0163-769X/98/$03.00/0
Endocrine Reviews 19(2): 173–202
Copyright © 1998 by The Endocrine Society
Printed in U.S.A.
Endocrinopathies in the Family of Endoplasmic
Reticulum (ER) Storage Diseases: Disorders of
Protein Trafficking and the Role of ER Molecular
Chaperones*
PAUL S. KIM
AND
PETER ARVAN
Division of Endocrinology (P.S.K.), University of Cincinnati College of Medicine, Cincinnati, Ohio
45267; and Division of Endocrinology (P.A.) and Department of Developmental and Molecular Biology,
Albert Einstein College of Medicine, Bronx, New York 10461
I. Introduction
I. Introduction
A. Overview
B. Protein folding in the ER
C. Supervised folding: the concept of molecular chaperones and folding catalysts
D. Co- and posttranslational modifications are factors
that can influence folding
II. ER Molecular Chaperones, Folding Catalysts, and Molecular Escorts
A. Binding protein (BiP)
B. GRP94
C. Calnexin and calreticulin
D. Disulfide isomerase and prolyl isomerase: families
of folding catalysts
E. ERp72 and ER60
F. HSP47
G. Molecular escorts: pro-peptides, transport subunits,
receptor-associated protein (RAP), and 7B2
III. Models of ER to Golgi Traffic Influence Models of Quality Control
A. Escape from ER retention as one hypothesis to explain anterograde protein traffic from the ER
B. Cargo receptors as another hypothesis to explain
anterograde protein traffic from the ER
C. What provides quality control of ER export?
D. ER-associated degradation
IV. Endocrinopathies as Models of Defective Protein Export
A. Congenital hypothyroid goiter with thyroglobulin
deficiency
B. Familial neurohypophyseal diabetes insipidus
C. Osteogenesis imperfecta and disorders of procollagen biosynthesis
D. ERSDs affecting lipoprotein metabolism
E. Other selected nonendocrine and endocrine ERSDs
V. Summary: A Proposed Classification of ERSDs
A. Overview
A
LL EUKARYOTIC cells secrete proteins. Higher eukaryotic tissues, in general, and many endocrine
glands, in particular, are differentiated to release abundant
quantities of specialized proteins. Most of the proteins released from cells are carried to the plasma membrane via the
biosynthetic transport pathway. The entire pathway is comprised of specific transport vesicles that shuttle their cargo
through a series of intracellular way-stations (1): at each
successive station, specific sorting decisions can be made on
the basis of transport signals (2, 3) and retention signals
(4 – 6). Exportable proteins enter at the endoplasmic reticulum (ER),1 the first membrane-bounded compartment of this
pathway (7, 8). Functions of the ER include the synthesis,
initial modification, and export of polypeptides destined for
secretion or to be located at the plasma membrane. One of the
most important jobs of the ER is to provide an environment
to facilitate the proper folding and assembly of newly synthesized exportable proteins. In addition, the ER contains
mechanisms to monitor the fidelity of these early biosynthetic events in the protein export pathway. This has been
called “ER quality control” (9), which involves machinery
1
The following abbreviations are used: ER, endoplasmic reticulum;
ERSD, ER storage disease; HSP, heat shock protein; GRP, glucoseregulated protein; BiP, immunoglobulin heavy chain-binding protein;
SREBP, sterol-regulatory element-binding protein; CHOP, C/EBP homologous protein; GADD153, growth arrest and damage-inducible gene
153; GPI, glycosylphosphatidylinositol; UGGT, UDP-glucose:glycoprotein-glucosyltransferase; GlcNac, N-acetyl glucosamine; Man, mannose;
PDI, protein disulfide isomerase; C-X-X-C, cysteine-X-X-cysteine; PPI,
peptidylprolyl isomerase; FKBP, FK506 binding protein; Erp72, endoplasmic reticulum protein 72; ER60, endoplasmic reticulum protein 60;
RAP, receptor-associated protein; LDL, low-density lipoprotein; LRP,
low-density lipoprotein-receptor related protein; PC, prohormone convertase; KDEL, lysine-aspartate-glutamate-leucine; COP, coat protein
complex; ERGIC53, a protein residing in the compartment intermediate
between ER and Golgi; MHC, major histocompatability complex; ERAD,
ER-associated degradation; Tg, thyroglobulin; FDI, familial diabetes
insipidus; DI, diabetes insipidus; AVP, vasopressin; NP, neurophysin;
OI, osteogenesis imperfecta; LDL-R, low-density lipoprotein receptor;
MTP, microsomal triglyceride transfer protein; LPL, lipoprotein lipase;
AAT, a-1-antitrypsin; CF, cystic fibrosis; CFTR, cystic fibrosis transmembrane conductance regulator.
Address correspondence to: Peter Arvan, M.D., Ph.D., Division of
Endocrinology, Albert Einstein College of Medicine, 1300 Morris Park
Avenue, Bronx, New York 10461.
*This work was supported by NIH Grants DK-40344 to (P.A.) and
DK-02113 (to P.S.K.), as well as support from Knoll Pharmaceuticals.
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KIM AND ARVAN
designed to try to prevent premature export of incompletely
or improperly folded proteins from the ER, as well as machinery intended to initiate the removal of misfolded, incompetent proteins (10). These features of the ER have
evolved to reduce potential harm posed by exportable proteins that are prone to aggregation and malfunction. Thus,
ER quality control machinery is designed to differentiate
normal and abnormal forms of a wide variety of exportable
proteins, presumably by recognizing structural signals that
are enriched in misfolded and incompletely folded molecules.
Recent years have witnessed the identification of numerous inborn errors of metabolism that affect secretory or
plasma membrane proteins. In many instances, mutations
causing minor changes in protein primary structure lead to
intracellular retention of the affected proteins, suggesting
that proper conformation is critical for protein transport as
well as biological activity. Nowadays, scientists are increasingly combining molecular and biochemical analyses in the
hopes of identifying precisely how mutations produce folding defects that lead to abnormal protein trafficking. In this
report, we review defective protein export as the cause of a
variety of endocrinopathies that fall into the category of
Endoplasmic Reticulum Storage Diseases (ERSDs) (11).
These disorders include certain forms of congenital hypothyroid goiter, osteogenesis imperfecta, diabetes insipidus,
familial hypercholesterolemia, and others. In each case, the
disease leads to accumulation in the ER of a critically important protein that is unable to reach its target site and
therefore is unable to perform its physiologically intended
function.
Morphological studies of cells affected by ERSDs routinely
reveal expansion and dilation of the ER compartment, which
may in part be due to accumulation of misfolded exportable
proteins. Moreover, the compensatory response in such cells
also frequently includes a selective induction of the synthesis
(and supranormal accumulation) of several ER resident proteins that are thought to participate in ER quality control.
Most of these proteins are considered to be molecular chaperones, a subtopic that is considered further in Section II.
This review will provide an endocrinologist’s perspective
of protein folding in the ER. From this vantage point, we
consider how subtle mutations in the coding sequences of
polypeptide hormone precursors and other exportable proteins, in conjunction with ER quality control, can lead to
defective protein trafficking, causing a disease phenotype.
Finally, we review specific representative endocrinopathies
in greater detail, as a means to highlight the underlying
similarities and differences in phenotypes and modes of
transmission of ERSDs, with an eye toward identifying future directions of endocrine investigation.
B. Protein folding in the ER
Until recently, the principle of protein biogenesis relied
entirely on the hypothesis that each peptide chain can selfassemble into a stable, low free-energy conformation, based
solely on information contained within the primary structure
(12, 13). The idea that few additional factors were required
for proper folding was supported by studies of the renatur-
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ation of small polypeptides in dilute solution at reduced
temperature. However, folding of proteins in living cells (14),
especially within the ER compartment, occurs under highly
restrictive conditions unique to this microenvironment (15,
16). For one thing, the protein concentration in the lumen of
the ER may be as high as 100 mg/ml (17). Second, secretory
proteins are translocated into the ER as they are being translated; thus, the NH2 termini of secretory proteins routinely
begin to fold in the ER before the COOH termini have even
been synthesized (18, 19). Third, the ER is the site for de novo
addition of N-linked core oligosaccharides, as well as initial
carbohydrate processing that may exert both direct and indirect effects on glycoprotein folding (see Section I.D). Fourth,
the composition of ions and small molecules in the intraluminal environment of the ER is highly regulated: this includes levels of calcium (20) and ATP (21). Finally, in mammalian cells, the ER is a more oxidizing environment than the
soluble cytoplasm, owing to the transport of oxidized glutathione (22), which fosters the formation of disulfide bridges
(23). All of these conditions, but particularly the oxidizing
environment and extraordinary concentration of nascent
(unfolded) polypeptides, lead to an increased possibility for
improper intra- and intermolecular associations.
In spite of these obstacles, a high fraction of newly synthesized secretory and plasma membrane proteins are successfully folded and exported from the ER. Indeed, while a
fraction of the initial cohort of newly made exportable proteins may enter novel misfolded states, for endogenous proteins in general, most of the cohort follows a “statisticallymost-probable” folding pathway, proceeding through a
predictable series of conformational intermediates en route
to the native state (24 –28). It is believed that this process has
evolved via cotranslational domain-dependent folding (19)
in conjunction with the actions of compartment-specific molecular chaperones.
C. Supervised folding: the concept of molecular chaperones
and folding catalysts
It is believed that the overall speed and efficiency of exportable protein folding is enhanced through a combination
of interactions with another group of proteins resident to the
ER. This group includes members of highly conserved families of molecular chaperones, as well as others to be described, some of which are viewed primarily as folding catalysts. Found in every living cell, molecular chaperones were
originally defined as families of proteins that assist in the
self-assembly of other chains but are generally not part of the
final functional unit (29). Thus, by definition, classical molecular chaperones interact only transiently with their “substrate” proteins (for further definition, see Section II and Fig.
1, below). Of the ER molecular chaperones, several are
known to be members of the larger family of heat shock
proteins (HSPs), conserved even to prokaryotes (which lack
ER and other cytoplasmic organelles). Heat shock is only one
of several different types of stress that can cause protein
denaturation, and HSP60, HSP70, and HSP90 classes of heat
shock proteins, so named for their approximate molecular
weights, are known to play crucial compensatory roles that
allow cell survival in the face of stress, by limiting and po-
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175
FIG. 1. Categories of ER helper proteins described in the text. The relative abundance of many ER resident proteins varies considerably with
cell type and is stimulated under conditions of cellular stress (see text), but many of these molecules are very abundant, perhaps on the order
of 10 mg/ml, even under resting conditions (92, 457). This schematic figure divides helper proteins in the ER into four general categories that
are defined in the text at the beginning of Section II. These groupings are not complete lists, e.g., the protein-specific ER chaperones do not
include BAP31, an ER membrane protein that interacts selectively with certain members of the cellubrevin/synaptobrevin family (459). Some
proteins listed are not discussed individually in the text. Under the enzymes that regulate folding are included molecules that may have both
direct and indirect effects on protein conformation, e.g., sugar processing in the ER may directly influence folding as well as by glycoprotein
association with calnexin and calreticulin (see Section II.C). Other molecules listed fall into “gray areas” with respect to category. For example,
PDI has been described as both a foldase and a classic ER chaperone (see Section II.D). Likewise, it is possible that ERp72 and/or ER60, listed
here as classic chaperones, might exhibit PDI-like catalytic activity in vivo. Further, prolyl hydroxylase and glutamyl carboxylase, although
widely expressed among different tissues, might be considered as protein-specific. Moreover, classic ER chaperones may also have the potential
to promote the folding and export of some secretory proteins and promote the degradation of others (see Section III). Finally, a new category
of proteins are listed here here under the name of molecular escorts (see Section II.G). In some cases, a secretory propeptide may be important
for nascent chain folding and may therefore be considered as a protein-specific chaperone, but the pro-region frequently escorts the rest of the
polypeptide through the secretory pathway, warranting inclusion in both categories. RAP is similarly considered in these two categories. The
common a-subunit of glycoprotein hormones is considered here as a molecular escort, especially in the case of FSH or TSH (246); however, aand b-subunits of glycoprotein hormones remain associated even after secretion, where the heterodimeric structure may be important for
biological activity (see Section II.G).
tentially reversing aggregation of misfolded proteins (30). In
the cells of higher eukaryotes, each intracellular compartment, including the ER, has its own subset of HSPs and other
chaperones (except for the absence of an identifiable HSP60
class member in the ER). In the HSP70 class, all members
share remarkable homology in their “substrate”-binding regions—and the same is true of the HSP90 class. Thus members within a given class tend to differ mostly within the
region of the chaperone that specifies targeting to a particular
subcellular compartment. However, certain metabolic insults may tend to trigger a chaperone stress response limited
more or less selectively to a specific compartment (31–33).
Indeed, glucose deprivation is a relatively selective stimulant
of the ER stress response; thus, certain ER chaperones also go
under the name of GRPs, for glucose-regulated proteins (34).
Generally speaking, HSPs are abundantly expressed under normal conditions but their synthesis is further induced
under stress conditions. The regulation of ER chaperone
levels is a complex process that has been reviewed elsewhere
(35), although recent papers have shed increasing light on the
signaling pathway responsible for induction of chaperone
synthesis in response to the presence of accumulating unfolded secretory proteins—now known as the unfolded protein
response (36). It had been suspected for some time that a
signaling cascade for increased synthesis of the ER-HSP70
family member (known as BiP) begins with an increase in the
ratio of bound/unbound chaperone (37). A reduction in the
“free” level of chaperones [as a consequence of increased
association with binding sites on available incompletely
folded proteins (38, 39)] leads to the activation of one (40 – 42)
or more (43) possible protein kinases, triggering a signal that
induces further synthesis of BiP and other ER chaperones (31,
44) through what is thought to be a predominantly transcriptional mechanism (45). The kinase IRE-1, which transmits its signal across the ER membrane, is a type 1 membrane
protein whose N terminus (in the ER lumen) has no homology to other proteins, while its C terminus (in the cytosol)
contains a predicted serine kinase. The activity of IRE-1 transcriptionally regulates the stability of a specific transcription
factor, HAC1 (46, 47). Other factors implicated in the unfolded protein response may include sterol-regulatory
element-binding proteins (48), induced by sterol depletion
(36), as well as CHOP (also known as GADD153) which is a
member of the C/EBP family of transcriptional factors that
can be markedly induced as a consequence of certain forms
of ER stress (45).
The accumulation of proteins in the ER membrane may
trigger a second distinct signal transduction pathway, recently termed the ER-overload response (36). In this case,
increasing presence of either misfolded or nascent membrane
proteins is capable of causing the activation of another transcriptional factor, nuclear factor-kB, which regulates a different cascade of gene expression (49). Some forms of cellular
stress can trigger both unfolded protein and ER-overload
responses, while others are selective for only one signaling
pathway. In addition to these transcriptional mechanisms,
cells appear to be able to posttranslationally regulate ER
chaperone activity, to a variable extent, via ADP ribosylation
(50), phosphorylation (51–55), as well as formation of oligomeric chaperone complexes (56).
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To the extent that HSP classes of ER chaperones monitor
protein folding, the transcriptional regulation described
above is a mechanism designed to ensure that adequate
availability of these supervisory molecules is maintained at
all times. The importance of available free chaperones is
underscored by the recent understanding that even after
dissociation from a given chaperone, incompletely folded
“substrate” proteins routinely rebind to the same or another
copy of that chaperone, contributing to an increase in the
bound chaperone fraction. Although the structural basis for
the binding of different chaperones appears to vary, the
property of cyclic association-dissociation is a common feature ranging from the bacterial chaperonins (57) to ER chaperones that are primarily involved in recognition of polypeptide (58) or carbohydrate moieties (9). Importantly, the fact
that molecular chaperones act on “substrate” proteins does
not violate the principle of self-assembly. As a rule, classic
chaperones interact with many different “substrates” without conferring steric information to influence the final folded
structure. However, by interacting with nascent chains,
chaperones prevent (and may even reverse) undesirable protein-protein interactions; this increases the chances that
newly made proteins will have the opportunity to achieve
their native structure. Because ER chaperone associations are
based on recognition of features enriched in incompletely
folded versions of exportable proteins, associations of ER
chaperones are usually (but not always) at their highest levels immediately upon nascent chain translocation into the
ER, and terminated before export of the “substrate” protein
from this compartment.
Not every interaction with individual ER resident proteins
will enhance folding speed or even folding efficiency (see
Section III), although some available data tend to suggest
enhancement of efficiency at the possible expense of delayed
protein folding (59). Promotion of productive folding is likely
to be the net effect of interactions with both chaperones and
folding catalysts. However, the extent to which roles played
by the binding of individual chaperones are overlapping vs.
unique (60), and how the different chaperones may cooperate
in the folding process for a wide variety of proteins (14),
remains largely unknown.
D. Co- and posttranslational modifications are factors that
can influence folding
Although the flow of genetic information ends when the
primary structure has been synthesized, co- and posttranslational modifications, under the influence of local environmental factors (discussed in Section I.B), can also affect the
folding outcome for many exportable proteins. Of course,
many important modification steps (e.g., terminal glycosylation, sulfation, phosphorylation, and dibasic proteolytic
cleavage events) take place as proteins are transported
through the Golgi complex, which can significantly alter
protein destination and biological function. However, with
the exception of proteolytic cleavage, Golgi processing activities generally have fewer effects on underlying protein
conformation than the processing activities of the ER, which
are the subject of this section.
One of the most important ER modifications is the pro-
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teolytic cleavage of the predominantly hydrophobic ;20
amino acid signal sequence (61) by the signal peptidase complex; the signal peptide is degraded after it has served to
target the nascent chain into the ER translocation channel
(62). Indeed, failure to remove the signal peptide generally
results in severe, irreversible misfolding of secretory proteins
(63).
Also, as the nascent chain enters the ER lumen, the rapid
collapse of its hydrophobic domains is accompanied by the
ordered formation of intramolecular disulfide bonds (26,
64), which stabilize secondary and tertiary structure and
can be critical for maintaining a biologically active conformation (65). Indeed, most of the cysteine residues of
exportable proteins eventually form disulfides (23), while
similar covalent bonds are not observed in cytosolic proteins or in the cytosolic domains of transmembrane proteins.
Within the oxidizing ER environment, reactive thiols
also can form mispaired disulfide bonds. Subsequent reshuffling and correction of aberrant disulfide bonds (see
Section II.D) may represent one of the rate-limiting steps
in protein folding (23). Studies of exportable proteins mutated to lack specific cysteine residues have provided additional evidence for the importance of correct disulfide
bond formation. Similarly, treatment of live cells with
dithiothreitol or other membrane-permeant reductants results in unfolding of many newly made proteins; upon
removal of dithiothreitol, reduced proteins begin to properly refold, reoxidize, and ultimately leave the ER, albeit
at a slower rate (66 –71). Intermolecular disulfide bonds
may also be important in the formation and maintenance
of quaternary structure (28, 72).
One of the next most important ER modifications is
the addition of N-linked carbohydrates to glycoproteins
(73). A large preassembled, oligosaccharide containing two
N-acetylglucosamines, nine mannoses, and three terminal
glucoses (74) is transferred cotranslationally from a dolichollinked intermediate to an asparagine residue (of the consensus sequence, Asn-X-Ser/Thr), as the nascent polypeptide
emerges through the translocation channel in the ER membrane (Fig. 2). ER membrane proteins known as ribophorins
(75), as part of a protein complex encoded by at least seven
genes (76), assist in the catalysis of this initial glycosylation
reaction. Further ER carbohydrate modifications then occur
through the actions of glucosidases and other processing
enzymes (Fig. 2, discussed in Section II.C). Although not
found on all exported proteins, carbohydrate moieties often
assist in the folding, stability, and solubility of nascent exportable polypeptides (77), and in some cases glycosylation
is required for the folding of subunits that occurs before
oligomeric assembly (78). Thus, it is not surprising that inhibition of N-linked glycosylation frequently leads to misfolding and aggregation of nascent chains. Once fully folded,
however, removal of sugar groups generally has little impact
on protein solubility and conformation.
Quaternary structural maturation is another important
modification, which can range from simple homodimers to
larger hetero-oligomeric complexes that are associated either covalently or noncovalently (79). Although there are
exceptions (79a), proper oligomeric assembly is frequently
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177
FIG. 2. N-Linked oligosaccharide processing of exportable glycoproteins in the ER. N-Linked carbohydrates may be added to Asn-residues of
exportable proteins containing the consensus acceptor sequence Asn-X-Ser/Thr. The N-linked carbohydrate is added en bloc via a dolicholphosphate intermediate. The two proximal N-acetyl glucosamine residues are represented as dark, filled ovals. The triantennary Man9 structure
is shown as ovals containing the letter M. The terminal glucose residues are indicated as small boxes containing the letter G. Shortly after
oligosaccharide addition, monosaccharide removal begins, first with glucosidase 1 removing the outermost glucose residue and glucosidase 2
removing the next outermost residue. These glucosidase reactions can be pharmacologically inhibited with deoxynorjirimicin or castanospermine
(see Section III.C). It is the remaining monoglucosylated carbohydrate form that has been shown to interact with calnexin and calreticulin (see
text). A single terminal glucose may be restored onto Man9 (or even Man8 or Man7) by the action of UGGT, which is thought to act in a cyclical,
repetitive fashion on deglucosylated glycoproteins that are sensed by the enzyme to be misfolded. Such reglucosylation stimulates reassociation
with calnexin and calreticulin (see text). Mannosidase action may begin before, but occurs largely after, the glucose cycling is completed;
importantly, trimming below the Man7 stage may eliminate the oligosaccharide from further suitability as a substrate for the UGGT enzyme.
required for ER to Golgi transport of secretory and plasma
membrane glycoproteins, whereas unassembled monomers are often retained in the ER until the protein is
degraded or an assembly partner becomes available (72).
Other covalent modifications that can also have profound effects on protein folding may be limited to certain
subgroups of exported proteins. For example, cotranslational hydroxylation of proline within the tripeptide GlyPro-Pro repeats of collagen, catalyzed by an ER enzyme
known as prolyl-4-hydroxylase, is essential for triple-helix
formation and stability (80, 81). Hydroxylation of lysine
side chains also plays an important role in collagen stability (82, 83). Similarly, carboxylation of glutamyl residues by a vitamin K-dependent mechanism plays an important structural role in the stability of a number of bloodclotting factors and calcium-binding proteins (84, 85).
Moreover, covalent attachment of a glycosylphosphatidylinositol anchor near the carboxy termini of a certain subset of exportable proteins is associated with the proteolytic
cleavage of the C-terminal ;20 amino acids (86, 87); failure
to remove these residues can lead to protein aggregation
and transport incompetence (88). Other roles for glycosylphosphatidylinositol anchors in protein folding and ER
export per se currently remain unknown.
II. ER Molecular Chaperones, Folding Catalysts, and
Molecular Escorts
For antibodies to identify antigenic proteins, an entire
system exists for gene rearrangement to produce Ig “variable
regions” intended to serve as high-affinity peptide-interaction sites for diverse recognition of foreign products. The
complexity of this situation can be loosely analogized to the
ER, where a wide range of small peptides are exposed, in the
context of nascent polypeptide folding, that are normally not
exposed in the respective native structures. Although generally hydrophobic, there is undoubtedly considerable diversity in the primary structures exposed in unfolded
patches of nascent exportable polypeptides. For this reason
in eukaryotic cells, a system has evolved to minimize exposure of these unfolded patches and thereby decrease the risk
of promoting improper intrachain and interchain peptide
interactions. However, instead of being based on gene rearrangement, the system of recognition of unfolded proteins in
the ER involves a finite series of genes producing proteins
whose peptide-interaction sites tend to be more promiscuous
than those of antibodies. Concomitant with this promiscuity
is a tendency toward lower affinity interaction with any
particular polypeptide. By differences in peptide interaction
specificities of different chaperones, and promiscuity of in-
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KIM AND ARVAN
teraction of each chaperone, as well as the sheer concentration of chaperones in the ER, dynamic interactions with a
wide range of “substrates” are produced. This idea underscores the basic mechanism of ER quality control, to be discussed further (Section II.C). For the present, we wish to
describe properties of a selected subset of known 1) ER chaperones, 2) folding catalysts (Section II.A–F), and 3) molecular
escorts (Section II.G) in the protein export pathway (Fig. 1).
In conjunction with the schema in Fig. 1, we define these
molecules as follows: 1) ER chaperones are simply viewed as
binding proteins whose association with exportable protein
“substrates” is regulated by the concentrations of the two
components and their binding affinity in a bimolecular interaction. 2) Folding catalysts are true enzymes, which also
physically interact with substrate proteins, but in so doing
they lower the activation energy required for a discrete conformational change within an exportable protein. In the secretory pathway, chaperones and folding catalysts tend to
reside predominantly in the ER, where they serve their primary biological functions, while 3) transport subunits and
molecular escorts routinely accompany their “substrate”
proteins out of the ER, and persistent interaction may even
be required for ER exit. Note that we have not attempted to
review each individual molecule listed in Fig. 1.
A. Binding protein (BiP)
BiP (also known as GRP78), the most studied ER molecular
chaperone, is a member of the HSP70 class (89 –91), a major
calcium-binding protein in the ER lumen (92), and an essential gene product that even the simplest eukaryotes cannot live without (93, 94). Like other members of the HSP70
class, BiP possesses a peptide-binding groove lined with
hydrophobic side chains, which is thought to interact optimally with hepta- or octapeptides enriched in aromatic/
hydrophobic residues in alternating positions — a feature
common to many naturally occurring peptide chains (95–97).
In properly folded globular proteins, such features are normally buried in the hydrophobic core or are used at the
interface between subunits for protein oligomerization. This
helps to account for the observation that although BiP interacts with a remarkably wide range of exportable proteins,
it tends to associate most strongly with unfolded, unassembled, or aberrant polypeptides (25, 98 –106).
In addition to the role of BiP in posttranslational folding,
this chaperone has been proposed to be involved in the
translocation of nascent polypeptides across the ER membrane (107–111). Further, BiP plays an indirect but key role
in the fusion of nuclear membranes during fertilization between haploid yeast cells (112, 113). Although the physiological relevance of these functions for mammalian cells in
vivo remains to be determined (114), it is presumed from
these studies that luminal BiP binding serves to stabilize
nascent polypeptide chains as well as the luminal domains
of certain endogenous ER membrane proteins.
Misfolded exportable proteins, such as those produced by
mutation or abnormal glycosylation, have been found to
interact with BiP for periods long after their synthesis (e.g.,
Refs. 115–117). Such prolonged interaction occurs via persistent reassociation of the chaperone (58) to BiP-binding
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sites that cannot be properly buried within the hydrophobic
core of misfolded polypeptides. Because each round of BiP
dissociation is coupled to a round of ATP hydrolysis (56,
118), ATP consumption in the ER of cells that have accumulated misfolded polypeptides is expected to increase. Further, misfolding of exportable proteins can be found in cells
depleted of ATP (66, 119); this observation has been used to
suggest that cyclical binding of BiP and other ATP-dependent chaperones facilitates protein maturation (15). However, it must be pointed out that because ATP hydrolysis is
required for polypeptide release from BiP (120), the depletion
of ATP— or the use of BiP mutants that cannot hydrolyze
ATP (121–124)—produces defective chaperone dissociation
that is likely to inhibit the maturation and ER egress of
exportable proteins (see Section III). Thus, the results of experiments using these approaches cannot and do not independently establish that BiP binding normally promotes folding or export from the ER under physiological conditions,
although such a conclusion has been suggested based on in
vivo protein refolding after heat shock (125).
En route to normal folding, assembly, and exit from the ER,
BiP binding to exportable proteins is typically observed only
transiently (25, 126, 127). Importantly, existence of such transient interaction also cannot distinguish models in which BiP
binding is thought to promote folding and export from those
where it is thought to delay folding and export (see Section
III). In addition, because individual BiP-binding sites are
represented by relatively small stretches of primary structure, incompletely folded versions of large polypeptides may
expose multiple potential BiP-binding sites during protein
folding, simultaneously or in series (71). For example, at a
moment in time in the steady state, the average stoichiometry
of association between BiP and nascent thyroglobulin (a
;330-kDa monomer) has been estimated at ;10:1 in the
thyroid ER (25). Potentially, the earliest folding intermediates of nascent secretory proteins may bind even more than
the average number of BiP molecules, while later, more
folded forms are likely to associate with progressively fewer
BiP molecules.
As described in an earlier section, BiP levels are transcriptionally regulated by the unfolded protein response. Importantly, ordinary physiological dynamics of the production of
exportable proteins is sufficient to regulate the synthesis of
BiP and other ER chaperones (128 –130).
Lastly, coupled with the recent identification of 14 hsp70
family members in the yeast genome (59), a second, novel
hsp70 member in the ER, LHS-1, has been described, which
is a nonessential gene that exhibits partial BiP-like function
(131). However, it is possible that in mammalian cells the
function of the yeast LHS-1 is subserved by its non-hsp70
homolog, GRP170 (132).
B. GRP94
GRP94 (also known as endoplasmin), the product of a
single gene (133), is a member of the HSP90 class (134) and
is also a major ER luminal calcium-binding protein (92, 135)
that is transcriptionally coregulated with BiP under most
conditions (34, 35). Considerably less is known about the
peptide-binding specificity of GRP94, but by analogy to other
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ENDOCRINE ERSDs
members of the HSP90 class, GRP94 is likely to act in a
cooperative way (136, 137) in associating with nascent
polypeptides that have exposed unfolded patches. For instance, GRP94 might be one of the luminal proteins whose
binding enhances completion of nascent chain translocation
into the ER lumen (110). Further, BiP and GRP94 are found
in ternary complexes in which they are simultaneously involved in direct interactions with exportable proteins (138,
139), although their precise association-dissociation kinetics
may not be identical (140).
Like BiP, GRP94 can also be found to interact with misfolded exportable proteins for prolonged periods after synthesis (117, 138, 141–143), as well as for transient periods with
normal proteins maturing in the export pathway (140, 143).
Individual polypeptides may expose more than one potential
GRP94 binding site during folding, either simultaneously or
in series (140). Sequence analysis has revealed two potential
ATP-binding sites in GRP94 (53), which exhibits increased
binding to polypeptide “substrates” under conditions of ATP
depletion (33, 144). This and other observations led to the
conclusion that GRP94 binds ATP and exhibits weak ATP
hydrolytic activity (21, 145). However, a recent study has
demonstrated that GRP94 interactions with peptides are nucleotide-independent, and that ATP binding and hydrolysis
are not inherent properties of this chaperone (146). Such an
interpretation is of particular interest because an adenine
nucleotide-binding site has recently been unambiguously
identified in the cytoplasmic homolog, hsp90 (147). Thus,
although the molecular mechanism remains poorly understood, it is presumed that GRP94, like BiP, can undergo cycles
of unbinding and rebinding to peptide “substrates” (15). It
should be noted that thus far (148), such binding of GRP94
has not been demonstrated to result in enhancement of folding or export of proteins from the ER (139).
As noted above, GRP94 levels are transcriptionally coregulated with those of BiP as part of the unfolded protein
response (55, 130, 149). However, there are notable examples
where disproportionate changes in one chaperone over the
other is observed (150), indicating subtleties in transcriptional and/or translational regulation that are not well understood.
C. Calnexin and calreticulin
Calnexin, the only major molecular chaperone of the ER
that is an integral membrane protein (151), is a single-spanning, calcium-binding protein of the ER membrane (152, 153).
The CNX1 gene, a homolog that is believed to encode a form
of calnexin from the yeast Saccharomyces pombe, is an essential
gene whose critical function is contained within its ER luminal domain (154, 155).
The binding/recognition function of mammalian calnexin
has been an area of intense interest; almost immediately it
was realized that treatment of cells with tunicamycin, a condition that causes severe misfolding of glycoproteins (and
generally causes their increased binding to BiP), interferes
with the binding of many such glycoproteins to calnexin
(153). Nevertheless, calnexin does indeed bind to misfolded,
mutant proteins and to folding intermediates of proteins en
route to export (156 –158). The carbohydrate dependence of
179
calnexin binding has led to the proposal that calnexin is a
lectin (159, 160) serving as part of a “chaperone apparatus”
that includes two independent enzyme activities: glucosidase II and UDP-glucose:glycoprotein-glucosyltransferase
(UGGT) (161). This proposal is based upon knowledge of the
processing pathway for N-linked carbohydrates in the ER
(Fig. 2).
A 14-saccharide unit is initially added en bloc to N-linked
consensus acceptor sites in glycoproteins. This oligosaccharide includes two N-acetyl glucosamine (GlcNac) residues
anchored in series to an asparagine in the exportable
polypeptide (Fig. 2). Attached to this disaccharide is a triantennary structure comprised of nine mannoses (called
Man9). At one antenna of the Man9 are a string of three
terminal glucose residues (74). Normally, the three terminal
glucoses are removed from N-linked oligosaccharides before
glycoprotein exit from the ER, by the sequential action of
glucosidase I (which removes the outermost glucose) and
glucosidase II (which sequentially removes the remaining
two glucose residues) (162). However, a single terminal glucose residue is restored onto Man9 if the UGGT enzyme
“senses” the glycoprotein to be unfolded (163, 164). This
sensing involves interaction of the UGGT enzyme both with
exposed hydrophobic residues as well as an exposed GlcNac
residue at the base of the peptide-bound oligosaccharide
(165). Calnexin in turn has been proposed to preferentially
bind to the Glucose1-Man9-GlcNac2 form of exportable glycoproteins in the ER (77, 166). All carbohydrate-binding activity of calnexin resides in its luminal domain (167). Recently, association of the calnexin luminal domain with
monoglucosylated RNase B in vitro was shown to be very
dynamic, suggesting that calnexin undergoes rapid on/off
binding to monoglucosylated glycoproteins (167). Glucosidase II removal of terminal glucoses is not likely to occur
while glycoproteins are bound to calnexin (167); however,
during the period when glycoproteins have been released
from calnexin, glucosidase II can remove the terminal glucose such that rebinding to calnexin cannot occur. In this
view, cycles of rebinding to calnexin (168, 169) are triggered
solely by the action of the UGGT enzyme.
In contrast with the view of calnexin acting solely as a
lectin in the ER lumen, others have suggested chaperone
function involving the transmembrane, nonlectin portion of
calnexin (170, 171) or chaperone binding to transmembrane
domains of exportable proteins (172). In addition it has been
shown that calnexin-“substrate” complexes are not dissociated even when the oligosaccharide is cleaved from the
polypeptide upon endoglycosidase digestion (166, 173, 174),
suggesting that some interactions with calnexin might occur
in a glycan-independent manner (175). Further, certain unglycosylated proteins have also been shown (71, 176, 177) to
associate with calnexin. Calnexin binding to unglycosylated
proteins occurs commonly with protein aggregates that have
been suggested to be separate from the productive maturation pathway (178). However, there is reason to believe that
protein aggregates may indeed participate in productive
maturation (25, 179); moreover, such aggregates bound to
calnexin are reversible in vivo, leading to successful export
from the ER (71). Thus, while the capability of calnexin to
interact with monoglucosylated glycoproteins is unequivo-
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cally established (167), whether this represents the complete
story of its role in physiological protein maturation continues
to be debated (180, 181).
Calnexin is thought to be in close proximity to nascent
chains upon their entry into the ER lumen; thus, it is not
surprising that calnexin is hypothesized to play a cotranslational as well as posttranslational role in the folding of
exportable proteins (71, 182). During the cotranslational period, calnexin binding may act to shield reactive free thiols
from forming incorrect disulfide bonds, although this improvement in folding efficiency may actually slow down
kinetic progression through the folding pathway (168, 183).
Of course, these features are not discordant with observations that posttranslational rebinding of calnexin to monoglucosylated side chains (184) is also correlated with productive folding of exportable glycoproteins (169).
Calreticulin (181, 185), a homolog of calnexin, is another
major calcium binding protein (92, 135), the topology of
which is entirely luminal. Although its function as part of an
ER chaperone apparatus has been studied far less than that
of calnexin (186), it shares with calnexin the capability of
recognizing monoglucosylated glycoproteins, with partially
overlapping specificity (187, 188). Calreticulin and calnexin
both go through cycles of binding and release, indicating
roles in monitoring and assisting protein folding (168, 169).
Importantly, however, calreticulin is a full participant in the
transcriptionally regulated, ER unfolded protein response
(189), a feature noted to be absent for calnexin (151). Calreticulin also plays a role in intracellular calcium signaling
(190, 191), in transcriptional regulation of steroid-sensitive
gene expression (192), and in a number of other important
cellular functions (reviewed separately in Ref. 185).
Calmegin is the newest member of the calnexin family,
found only in the testis, where it is thought to perform a
function in the folding of proteins that are necessary for the
ability of sperm to adhere to and fertilize eggs (193).
D. Disulfide isomerase and prolyl isomerase: families of
folding catalysts
Protein disulfide isomerase (PDI), another major component of the ER lumen, is a true foldase, in that it catalyzes
thiol-disulfide interchange with a broad substrate specificity,
and it shows strong homology with bacterial thioredoxin (23,
194, 195). The regulation of PDI synthesis overlaps only partially with those of BiP, GRP94, calreticulin, and other ER
residents that exhibit the unfolded protein response (189,
196). Instead, PDI expression seems to be proportional to the
flux of nascent chains into the ER [i.e., especially high in
secretory tissues (197–199)], and its activity may be further
regulated posttranslationally (200). Recent reports indicate
that PDI may undergo dimerization, autophosphorylation,
and ATP hydrolysis that is stimulated in the presence of
denatured polypeptides (201). PDI increases both the rate
and efficiency of proper folding and export of proteins that
contain disulfide bonds (195, 202).
PDI1 is a gene essential for viability in yeast (203). There
are different opinions regarding the significance of this observation. First, critical to the disulfide isomerase catalytic
activity are two -CXXC- motifs. However, polypeptide bind-
Vol. 19, No. 2
ing by PDI has been reported not to involve these motifs
(204), and PDI also has been shown to exhibit the potential
for assisting folding of proteins that do not possess disulfide
bonds (205). Further, certain deletions in PDI that leave residual disulfide isomerase activity are nevertheless lethal,
while cells carrying a variant PDI in which both-CGHCactive sites are disrupted (i.e., no measurable isomerase activity in vitro) remain viable (206) and able to assist in protein
folding (207). On the other hand, the ER lumen appears to
have other proteins (discussed further, below) that can function as disulfide isomerases in the absence of PDI1 enzyme
activity (208). With this in mind, it is interesting that Escherichia coli thioredoxin can complement null mutants of yeast
PDI only if thioredoxin is mutated to contain a reactive CXXC
motif (209). These and similar studies have led some to conclude that the essential function of PDI is in fact to unscramble nonnative disulfide bonds (210). Nevertheless, PDI is
recognized to be multifunctional; it is for instance a wellrecognized subunit of the prolyl-hydroxylase complex that is
involved in collagen synthesis (211), and it heterodimerizes
with the 97-kDa subunit of the microsomal triglyceride transfer protein complex (212).
Peptidylprolyl isomerase (PPI) catalyzes cis-trans isomerization of proline side chains, and enhances the rate of protein folding in vitro (213). PPIs are also termed immunophilins and are comprised of cyclophilins (which bind the
immunosuppressant cyclosporin A) and FKBPs (which bind
the immunosuppressant FK506). The ER luminal FKBPs are
transcriptionally regulated with other members of the unfolded protein response (149, 214).
E. ERp72 and ER60
ERp72, one of the more recently described luminal chaperones (215), is also a calcium-binding protein in the ER (135,
208) and is a member of the PDI superfamily (see above).
Although ERp72 is not an essential gene product, its overexpression can rescue nonviable cells deficient for PDI (216).
Indeed, ERp72 contains three copies of the -CXXC- active site
motif found in PDI (215). While it seems plausible that ERp72
may be able to exhibit limited PDI-like activity (208, 217), it
also may be that its ability to rescue cells lacking PDI could
be associated with the reported ability of ERp72 to assist in
the degradation of proteins that cannot fold in the ER (218).
ERp72 has been shown to interact with misfolded versions
of exportable proteins (33, 219, 220), but evidence for its
interaction with normal protein folding intermediates in vivo
is not well established (143). Nevertheless, ERp72 is clearly
regulated with other proteins exhibiting the ER unfolded
protein response (196, 221).
ER60/calregulin, which also contains thioredoxin-like sequences and thus shares homology with ERp72 and PDI, has
been implicated in the ER-associated degradation of misfolded proteins (222). Although initially hypothesized to be
a phosphoinositide-specific phospholipase C (223), this has
not been independently confirmed. By contrast, although
additional study is clearly needed, some evidence for a thioldependent protease activity of ER60 is accumulating (224).
Moreover, it has recently been reported that processing of
N-linked carbohydrates in exportable protein “substrates”
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ENDOCRINE ERSDs
may be required for their association with ER60 (225), which
has been proposed to act in conjunction with either calnexin
or calreticulin (226).
F. HSP47
Heat shock protein 47 (HSP47) is a collagen-specific stress
protein that performs chaperone function during folding and
assembly of newly synthesized procollagen molecules (227).
Because type I procollagen forms a triple helix beginning at
the carboxy terminus (see Section IV.C), much of its folding
must occur after translocation of the nascent chain has been
completed. Early association of HSP47 is facilitated by its
binding to the amino-terminal globular domain of the collagen propeptide (228), but the chaperone appears to remain
associated throughout most, if not all, subsequent stages of
tertiary structural maturation (229, 230). Although it may act
in concert with other, more ubiquitous ER chaperones (144,
231), the services of HSP47 appear to be unique to cells
secreting collagen and its homologs (232). Because of this,
HSP47 falls into a “gray area” of protein-specific ER chaperones, as is the case with the microsomal triglyceride transfer protein (see Fig. 1 and Section IV.D, below).
G. Molecular escorts: pro-peptides, transport subunits,
receptor-associated protein (RAP), and 7B2
Figure 1 provides a schematic to categorize a number of
additional polypeptides that provide helper function to the
protein export pathway, yet should nevertheless be distinguished from conventional ER chaperones. These include the
propeptides of many polypeptide hormones, transport subunits, and a subgroup of proteins for which we propose to
adopt the name “molecular escorts” (233). There are growing
numbers of examples of proteins in each of these subgroups,
but we mention only one or two representative examples. We
emphasize that these distinctions may change over time, as
molecular information about the roles of helper proteins in
the folding pathways of individual proteins becomes clearer.
Importantly, not all polypeptides that participate in the
folding and trafficking of exportable proteins are ER residents. Specifically, the propeptide regions of many exportable proteins themselves serve primarily a structural, rather
than a functional, role (234). Thus, folding in the absence of
the propeptide, or in the presence of a mutated propeptide,
may perturb the conformational maturation and ER export
of polypeptide hormone precursors (235). Such proregions
have been referred to as intramolecular chaperones, but a
distinction should be made between this idea and classic ER
chaperones (Fig. 1) which (except under unusual circumstances) are not transported down the secretory pathway,
and whose turnover in the cell is far slower, and which
routinely associate with folding intermediates of more than
one kind of “substrate” protein.
Not unlike the situation with the propeptides are subunits
of oligomeric proteins that function primarily in polypeptide
transport and stabilization, while playing a relatively minor
role in subsequent biological activity. A case in point is the
a-subunit of the glycoprotein hormones (LH, FSH, CG, TSH),
which combines noncovalently with b-subunit in the ER
181
(236). Both subunits must be partially folded before subunit
assembly (64, 237–239), and the specific regions used by aand b-subunits to combine have begun to be mapped (240 –
243). When expressed by themselves, b-subunits tend to be
relatively poorly secreted, whereas when coexpressed with
a, a much higher fraction of b is secreted, as heterodimers
(244). Although a heterodimeric structure (and therefore, the
presence of a subunit) is needed for biological activity (245),
the evidence indicates that the a-subunit, which is common
to all of these hormones and is therefore unlikely to provide
biological specificity, plays an important role in export of the
different glycoprotein hormones, in particular for FSH and
for TSH (246).
Another subgroup of molecules that should be distinguished from conventional ER chaperones is represented by
the “molecular escorts” (233). For illustrative purposes, we
review only two members of this subgroup.
RAP is a ;40 kDa polypeptide which interacts shortly after
the biosynthesis of low-density lipoprotein receptor (LDLR)related protein (LRP), and RAP travels with LRP molecules,
having the potential to remain associated on the cell surface
(247). By itself, RAP is a protein that is believed to be retained
within the ER (248). Further, LRP in the absence of RAP is
also defective for ER exit (233). However RAP association
with LRP allows both partners to exit the ER (249), presumably by maintaining a given LRP in a favorable conformation
(250) and by preventing premature association of LRP ligands in the ER (251), which could lead to receptor retention
and/or degradation. Instead, dissociation of RAP, when it
occurs, takes place after the proteins have reached the cell
surface, where LRP ligands can displace the escort.
7B2 exhibits restrictive expression in neuroendocrine tissues, where it is synthesized as a ;25-kDa precursor protein
and is released from the regulated secretory pathway as
processed proteolytic fragments. The carboxy-terminal domain of the pro7B2 protein is responsible for inhibiting the
prohormone processing activity of prohormone convertase 2
(PC2) (252, 253), while the larger N-terminal portion of processed 7B2 contains independent stimulatory actions on the
catalytic activity of PC2 (254). Evidently, several different
domains of the pro-7B2 protein may interact with the proform of PC2 (255). Recently, it was determined that pro-7B2
and pro-PC2 assemble (in a 1:1 stoichiometry) in the ER of
neuroendocrine secretory cells, and this assembly leaves the
ER together (256) before dissociation in the most distal portions of the Golgi complex (257, 258). Because pro-PC2 may
be transported through the secretory pathway more slowly
or less efficiently in the absence of pro-7B2 (259), pro-7B2 has
been called a chaperone (256) but may be better viewed as a
molecular escort. Such a distinction also represents a “gray
area” since recombinant 7B2 may be able to associate with at
least one other unrelated protein, based on in vitro studies
(260).
III. Models of ER to Golgi Traffic Influence Models of
Quality Control
Newly synthesized plasma membrane proteins and secretory proteins must enter or cross the ER membrane in a
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highly unfolded state (18). After folding is accomplished—
except for molecules that are permanent residents of the ER
itself—these proteins are transported by the budding of carrier vesicles destined for the Golgi complex. The budding of
ER export vesicles can be identified by the presence of coat
proteins that are recruited to the cytosolic side of the ER
membrane (261) (see Section III.B, below).
Although quality control monitoring of the secretory pathway may not occur exclusively in the ER, as described in the
Overview (Section I.A), the ER contains mechanisms that are
intended to help cells decide which proteins are ready for
export and which are not. Since some newly synthesized
proteins are destined to be permanent residents of the ER
while others are destined for forward transport, at some
point there must be a partitioning of proteins that remain in
the ER from those that advance further (262). There is currently considerable debate about the mechanisms by which
this sorting is achieved. We believe this may also bear on the
question of why many misfolded secretory proteins are not
exported. The following two sections propose two very different hypothetical mechanisms by which ER exit of exportable proteins might occur, and it remains possible that none,
all, or parts of both mechanisms may be correct, or that other
mechanisms not considered in this review (263) may come
into play. Nevertheless, for the present, such hypotheses are
necessary to begin to understand diseases of protein folding
and secretion (264).
A. Escape from ER retention as one hypothesis to explain
anterograde protein traffic from the ER
Over the past decade, a dominant view has been that no
specific signals are required for soluble proteins to undergo
forward transport, while luminal resident proteins are maintained in the ER by specific mechanisms, including both
prevention from forward trafficking (265) as well as retrieval
of ER residents that have escaped (262, 266). Retrieval via
retrograde vesicular transport to the ER is the better understood mechanism (267) and involves a receptor, Erd2p (268),
that binds to a C-terminal recognition motif comprised of the
sequence K-D-E-L or a close variant, which is common to
luminal ER resident proteins. A short cytosolically disposed
motif serves a similar retrieval function for ER transmembrane proteins (266, 269, 270). However, retrieval may not be
the major mechanism that retains ER resident proteins, because even in the absence of a KDEL tail, luminal ER molecular chaperones are very slow to exit the ER (271). This ER
retention has been attributed in some way to the calcium
binding and calcium levels in the ER lumen, possibly resulting in the formation of an insoluble protein matrix (265).
Indeed, mutation of the calcium-binding domain of calreticulin has a profound effect on the egress of this molecular
chaperone, even when the KDEL sequence, and receptor,
remain intact (272). Moreover, new genes are now being
described that play roles in KDEL-independent retention of
luminal ER resident proteins, although these mechanisms are
still not well understood (273).
Thus, it is currently thought that the primary mechanism maintaining the localization of ER luminal resident
proteins involves their direct retention within this com-
Vol. 19, No. 2
partment, with the KDEL signal serving as a back-up system for retrieving escaped ER chaperones. With this in
mind, secretory proteins must exhibit at least two properties for their successful exit from the ER. First, in general,
they must express neither an ER retention domain nor a
C-terminal retrieval motif used by the luminal ER resident
proteins. Second, they must not bind tightly or extensively
to ER chaperones that are themselves anchored in the ER.
In other words, in this model, for secretory proteins to exit
the ER, they must escape from the clutches of anchored ER
chaperones. Moreover, this model predicts that the residence time of newly synthesized secretory proteins will be
directly proportionate to the extent of their binding to ER
chaperones (139). This feature could explain why the average length of time that newly synthesized exportable
proteins spend in the ER differs for each protein species
(274, 275).
B. Cargo receptors as another hypothesis to explain
anterograde protein traffic from the ER
In the past 2 yr, an alternate sorting model of protein exit
from the ER has emerged (276). In this view, at or just before
the time of budding of a newly formed ER transport vesicle,
certain proteins are selectively extracted by receptors for
entry into these ER transport vesicles (277). Support for such
a model is based on the idea that cytosolic proteins, which
are known to coat the surface of budding membrane vesicles,
are likely to be recruited by cytoplasmically disposed domains of transmembrane receptors, while the luminal domains of these receptors in turn could recognize specific
ligands on the luminal side of the membrane (278, 279). This
kind of model has strong precedent in post-Golgi trafficking
(280), e.g., in the delivery of newly synthesized lysosomal
proenzymes as well as endocytic ligands via coated vesicles
(281–283). Between the ER and Golgi compartments, two
different kinds of multicomponent coat protein complexes,
known as COPI (277) and COPII (261), have been described
to be recruited to the cytosolic surface of membranes in
preparation for vesicle budding (284)—and neither class of
coated vesicle includes ER resident proteins (285–287). These
features have provided an incentive to search for anterograde sorting signals (3) and putative cargo receptors captured by transport vesicles (288).
Recently, in mammalian cells, ERGIC53, a recycling membrane protein that functions as a mannose-binding lectin
(289, 290), has been hypothesized to extract secretory proteins from the ER (262). In addition, in yeast, at least one ER
membrane component, EMP24, has been postulated as a
potential recycling receptor for selected exportable proteins
(291). Further, loss of the yp24A protein, homologous to
EMP24, is thought to inhibit the formation of transport vesicles (292). Moreover, the recent identification of other homologous proteins in higher organisms, and the demonstration that members of this p24 family of transmembrane
proteins express short cytosolically disposed motifs necessary to recruit coat protein complexes (288) has led to the
hypothesis that the p24 family represents one class of longsought cargo receptors.
However, it must be pointed out that the formation of
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ENDOCRINE ERSDs
coated transport vesicles does not even require the presence
of luminal contents (293). Certainly, lectin-like sorting interactions like those proposed for ERGIC53 (294) cannot solely
account for the ER exit rate of secretory proteins in mammalian cells, where many secreted proteins (e.g., proinsulin)
are unglycosylated. Moreover, in yeast, evidence indicates
that the phenotype associated with a loss of EMP24, one of
the p24 family, is not restricted to secretory protein transport
but includes altered cellular handling of luminal ER chaperones as well as protein disulfide isomerase (273). Thus, the
cargo receptor hypothesis, while gaining increasing interest,
still has a number of obstacles to overcome.
C. What provides quality control of ER export?
As noted in the Overview (Section I.A), the ER quality
control machinery is designed to try to prevent the export of
incompletely/improperly folded versions of exportable proteins. This might happen by one of several different mechanisms. For one, unfolded proteins may aggregate and become insoluble, thereby becoming unable to advance into the
lumen of ER transport vesicles (295). However, not all unfolded versions of exportable proteins are insoluble, or even
aggregated. In these cases, hypotheses designed to explain
how selective protein export is prevented depend largely on
which model is favored for the mechanism of normal protein
export (reviewed above). If anterograde traffic out of the ER
requires presentation of certain features to cargo receptors,
then it is possible that unfolded proteins fail to present the
required features and thus cannot be recognized and cannot
be carried forward into Golgi-bound transport vesicles. In
such a case, ER chaperones might serve a role in promoting
sufficient folding to assist in the presentation of exportable
proteins to cargo receptors. Alternatively, if anterograde traffic requires escape from ER chaperones, then it is possible
that unfolded proteins fail to bury chaperone recognition
sites. In that case, unfolded patches exposed on exportable
proteins serve as de facto ER retention signals, because they
either promote the formation of insoluble aggregates (which
cannot advance because of intrinsic biophysical properties)
or binding to ER chaperones (which prevent protein advance
because the chaperones are anchored in the ER—see Section
III.A). If the latter view is correct, a corollary is that for all
proteins retained in the ER that are not intrinsically insoluble,
the retained protein must be bound to one or more chaperones in order that its export be prevented. With these hypotheses in mind, it is worth reviewing what is known about
the binding of ER chaperones with respect to helping or
hindering protein export.
Based on current knowledge, there are two kinds of studies suggesting the promotion of protein export as a consequence of binding to ER chaperones. First is the case of
calnexin and calreticulin (169, 183), which associate with
monoglucosylated carbohydrate side chains of a wide variety of glycoproteins (167, 296) such that association is abrogated by pretreatment of cells with castanospermine or deoxynorjirimicin (inhibitors of carbohydrate processing that
prevent formation of monoglucosylated core sugars on glycoproteins, see Section II.C). In this case, drug treatment can
be clearly shown to diminish the folding and transit of a
183
subset of glycoproteins from the ER to the Golgi complex
(297, 298). A second method capitalizes on the fact that most
ER chaperones have a much longer half-life in the cell ($ 1
day) than the secretory proteins with which they interact
(minutes to hours). Thus, treatment of cells with cycloheximide, an inhibitor of protein synthesis, can allow for the
drainage of previously synthesized exportable proteins from
the ER, while the ER chaperones remain at normal concentrations. This causes the fraction of unoccupied chaperones
to increase. Because cycloheximide effects are reversible, the
drug can be washed away and the resumption of protein
synthesis allows new exportable proteins to be introduced
into an ER that now has an increased availability of chaperones. In such a case, aggregation of newly synthesized
secretory protein has been observed to be diminished (25). By
this means, increased availability of BiP and other chaperones might enhance protein maturation and traffic through
the anterograde transport pathway (299).
On the other hand, many of the ER chaperones described
in Section II have been specifically implicated in the retention
of exportable proteins within the ER. Indeed, association of
BiP with Ig heavy chains (90) and light chains (101) has been
directly correlated with their failure to undergo export from
the ER, and indeed, the loss of BiP-binding sites from surface
or secreted Igs restores their ability to undergo intracellular
transport (300, 301)— even if they are incompletely folded or
assembled. Similarly, soluble (truncated) forms of the T cell
receptor a-chain do not aggregate but exist as monomers,
and yet they are not secreted; instead, they coprecipitate with
BiP, and manipulations that cause BiP dissociation allow for
a-chain secretion in vivo (302). Also along this spectrum,
unassembled subunits of oligomeric membrane proteins are
typically retained in the ER bound to BiP, but they often
undergo high molecular weight aggregation that may prevent their entry into ER export vesicles (303, 304).
The situation with calnexin appears to be analogous: the
T cell receptor b-chain is prevented from ER export in the
absence of assembly with a-chain, and the retained b-chain
is bound to calnexin (305). For MHC class I heavy chains that
have not yet assembled with b2-microglobulin and antigenic
peptide, export from the ER is impeded by association with
calnexin (306), even while the assembly of heavy chains with
b2-microglobulin is promoted (183). Similarly, dissociation
from calnexin parallels the egress of MHC class II heterotrimers from the ER, suggesting that the chaperone is involved
in retention of unfolded/unassembled subunits (307). Moreover, in yeast cells, deletion of the calnexin homolog CNE1
does not lead to a loss of cell viability but, rather, inhibits ER
retention of selected proteins that normally cannot exit this
compartment (308).
Similar reports regarding retention of exportable proteins
within the ER have been attributed to GRP94 (138) and other
ER chaperones (232). These reports are especially common in
ERSDs (caused by the presence of mutant forms of exportable
proteins) where the ER unfolded protein response causes
remarkable induction of the synthesis of these ER chaperones
to levels $ 1 order of magnitude above the normal range.
Certainly, in ERSDs, a higher than normal number of these
chaperones are likely to be bound to exportable proteins
entrapped in the ER. However, ERSDs cannot be used to
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KIM AND ARVAN
provide definitive evidence regarding the effects of chaperone binding on the retention of exportable proteins, because
it is difficult to ascribe how much of the retention phenotype
could be due to underlying abnormalities associated with the
mutation of the exportable protein, rather than effects of
increased chaperone binding. For this reason, scientists have
attempted to independently express BiP or other ER chaperones via constitutive promoters, to increase the free level
of ER chaperones in the absence of an ERSD, with the intention of increasing chaperone binding to exportable proteins. When BiP is made increasingly available in the ER, this
is sufficient to blunt or block the ER unfolded protein response in Chinese hamster ovary (CHO) cells when an experimental stress is imposed (309). Equally importantly, increased availability of BiP inhibits export from the ER of
those secretory proteins known to bind BiP (309). Analogous
results have recently been reported upon increased expression of GRP94 (139).
Retention of exportable upon increased expression of BiP
or GRP94 might be explained by a saturation of the ER,
causing the diminished availability of folding promoters and
other key resident proteins in the ER (i.e., “chaperone imbalance”). As an alternative, increased chaperone expression
might simply cause increased complex formation with susceptible “substrate” proteins. The former idea has been rendered less likely by recent demonstrations that the levels of
a wide array of ER resident proteins are not changed as a
consequence of selective overexpression of an individual ER
chaperone (139, 309a). By contrast, experimental evidence
favoring the alternative view is the fact that decreased expression of BiP (using antisense methodology, which does
not diminish the levels of foldases or other ER resident proteins) actually increases the ER export of certain heterologous
proteins expressed in CHO cells (119, 310). Moreover, these
effects are selective, as overexpression of BiP, or even overexpression of mutant BiP, which cannot undergo ATP-dependent release of a “substrate” protein, fails to cause the
retention of secretory proteins that lack a demonstrable BiPbinding site (124).
The message transmitted by these results is that to execute
their role with respect to protein export, chaperones must not
only associate with unfolded secretory proteins for a period
of time—they must also dissociate during the folding process
(25). As described in Section I.C, repeated cycles of binding
and release to the same or different unfolded polypeptides
is a common function of molecular chaperones in all compartments. This has left open the hotly debated question of
what stage in the chaperone-binding cycle does the polypeptide folding actually takes place: during the period of chaperone association or during the period of release (311)? There
is good reason to believe that chaperone binding, at least in
some circumstances, may decrease the assembly of oligomeric subunits (168), decrease the folding of monomers, and
actually promote monomer unfolding (69). Although semantically inappropriate, such behavior has been termed an “anti-chaperone” function of ER chaperones and is dependent
upon the particular “substrate” and the availability and stoichiometry of chaperone binding (103).
Thus, to summarize this section, quality control of ER
export (9) may be provided by any of three different mech-
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anisms: 1) inability of an exportable protein to properly
present itself to cargo receptors mediating ER exit (an increasingly popular model but one for which direct evidence
is still lacking); 2) formation of protein aggregates that are
biophysically unable to enter transport vesicles (for which
indirect support has been obtained in only a few cases); and
3) repetitive, cyclical binding by ER chaperones, which mediates ER retention and does not necessarily facilitate folding
in all cases (for which there is the largest amount of available
evidence, albeit indirect). The latter model has obvious implications for ERSDs (see below), and suggests that successful export from the ER takes place only when all chaperonebinding sites on the exportable polypeptide are buried, or
when ER chaperones are otherwise disabled or overwhelmed. Thus, according to this model, one simple way to
think about the role of ER chaperones in the regulation of
protein flow out of the ER may be the dam concept, in which
the dam (comprising all available ER chaperones) serves as
a central regulator. The “height” and “tightness” of the dam,
represented by the levels of ER chaperones and their respective affinities for “substrate”-binding sites, regulates the escape of exportable proteins from the ER. The rigor of ER
quality control is related to both parameters. Ideally, ER
quality control should be sufficiently zealous to retain only
mutant proteins that may have lost their primary biological
activity or that may have other unwanted toxic effects. However, as we shall see further in Section IV, this quality control
is not perfectly efficient in all situations. Indeed, ER quality
control machinery may exhibit drastically different retention
properties for different mutant protein subunits whose primary structures differ from the native primary structure only
by the loss of a single free cysteine or a single disulfide bond,
in some cases even leading to export that is augmented over
that of the wild-type protein (79a, 312, 313), but in most cases
leading to a diminution of protein export.
D. ER-associated degradation
In cells, both wild-type and mutant versions of exportable
proteins are subject to misfolding under normal or stressed
conditions, although the fraction that is irreversibly misfolded may vary (315). The development of a system that is
not only able to recognize irreversibly misfolded proteins,
but to target them for degradation, is an essential function for
cell survival, because inexorable accumulation of undegraded misfolded proteins in the ER is likely to clog the
secretory pathway and become toxic to cells (316). A central
question to resolve for the coming decade is: although the
lysosomal compartment of cells is primarily designed for
macromolecular digestion, how do misfolded versions of
exportable proteins that cannot readily reach the lysosome
(because of ER quality control restraints) undergo degradation? Unlike autophagy (317–321), a phenomenon known as
ER-associated degradation, or ERAD (322), has been defined
by 1) insensitivity to classic inhibitors of lysosomal proteases
and intracellular transport blockers, 2) immunolocalization
of degradative substrates in the ER, as well as 3) lack of
Golgi-type sugar processing on the substrates (323, 324).
At this stage, most of the machinery responsible for ERAD
remains largely unidentified in higher eukaryotes. With few
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ENDOCRINE ERSDs
possible exceptions (143, 324), proteolytic fragments are generally not observed as intermediates in the ERAD process,
complicating its characterization. In some instances, the ER
compartment itself has been implicated as the site of degradation (325), suggesting that at least some of the ERAD
machinery is contained within this compartment. Further,
evidence has been presented that redox events, or the availability of free thiols, may strongly influence the activity of
ERAD machinery (326, 327). Indeed, ER60 and ERp72, which
contain copies of the -C-X-X-C- motif conserved in PDI (described above) have been postulated either to be potential
cysteine proteases or molecules that may target misfolded
proteins to ERAD (218, 222–224). Additional new genes are
being found that apparently influence this process (328).
One recently proposed mechanism of ERAD that is gaining great attention is the idea of dislocation of proteins from
the ER membrane (329 –331), or the ER lumen (322, 332),
presumably via reverse translocation through the ER translocon (333), i.e., back to the cytosol, for selective protein
degradation by the ubiquitin-proteasome proteolytic pathway (334 –336). The steps involed in this process are far from
being completely worked out, but may include initial partial
proteolysis within the ER, ubiquitination, extraction from the
ER, and the removal of all previously attached N-linked
carbohydrates, before complete proteasomal digestion (10,
337).
Interestingly, even though essentially all exportable proteins that fail to escape the ER are eventually degraded,
several studies have revealed that some proteins turn over
rapidly while others disappear more slowly (338). As molecular chaperones play major roles in assisting or preventing
protein degradation in other cellular compartments (339), it
seems likely that they may play similar diverse roles within
the ER. At this early stage in our knowledge, it should perhaps not be surprising that BiP and calnexin association have
been correlated with the destruction of misfolded exportable
proteins by ERAD (105, 322, 340, 341) just as others have
suggested that binding of these very chaperones helps to
protect from ERAD (168, 342, 343). In addition, inhibitors of
proteasomal proteolysis may cause the accumulation of undegraded, misfolded secretory proteins, leading to induction
of ER chaperone synthesis as part of the unfolded protein
response (344). More work is clearly needed to understand
how molecular chaperones help to distinguish misfolded
proteins targeted for ERAD from normal early folding intermediates of exported proteins and, more specifically, the
relationships between ERAD and the quality control of ER
export. Whatever the answers, it would appear that ER chaperones are likely to be intimately involved.
IV. Endocrinopathies as Models of Defective Protein
Export
A. Congenital hypothyroid goiter with thyroglobulin
deficiency
Thyroglobulin (Tg), a large secretory prohormone, provides the matrix for both iodine storage and thyroid hormone
synthesis. As a result, conditions that prevent Tg from reaching the thyroid follicle lumen, a key step in regulation of
185
thyroid hormone synthesis and storage, lead to clinically
significant hypothyroidism (345).
Export from the ER represents the rate-limiting step in the
overall process of secretion of Tg (139). Much of the time after
biosynthesis is used to convert Tg into a transport-competent
form, which involves multiple processing steps within the
ER (346). Cotranslationally, collapse of hydrophobic domains into the globular core, addition of N-linked sugars,
and formation of intramolecular, intradomain disulfide
bonds stabilize the nascent Tg molecule, as does protective
association with ER chaperones such as calnexin (71), BiP
(25), and GRP94 (143). During the posttranslational period,
while mispaired disulfide bonds are being corrected, these
ER chaperones begin to dissociate from Tg. The dissociation
is correlated with a period in the normal folding pathway
during which Tg matures through a series of discrete folding
intermediates, ultimately leading to compact monomers.
Moreover, immunopurified BiP-nascent Tg complexes have
been analyzed under native conditions, directly demonstrating BiP association only with early Tg folding intermediates
(25). These data indicate progressive BiP dissociation during
the conformational maturation of Tg. The compact monomers, no longer detectably bound to BiP, represent the first
structures in the Tg folding pathway that are competent to
assemble into homodimers—the form normally observed to
exit the ER (Fig. 3). Perturbations that interfere with the
pathway of Tg progression to homodimers invariably inhibit
ER export of Tg. On the other hand, chronic physiological
exposure to TSH, the dominant regulator of thyrocyte metabolism, accelerates the formation of compact monomers
and consequent dimer asssembly, even though the number
of new Tg molecules fluxing through the ER is greatly augmented. This hormonally stimulated increase in Tg export is
accompanied by modest elevations in the levels of PDI, BiP,
and GRP94 and accelerated dissociation of nascent Tg from
BiP (130) and perhaps other chaperones, as well.
In many human patients with congenital hypothyroid goiter, the disease is detected upon neonatal screening. The
prevalence of defective Tg synthesis or secretion from such
screening has been estimated to be 1/40,000 newborns (345).
However, the diagnosis can be difficult, especially when
there is an incomplete block in Tg metabolism. If the condition is not diagnosed during the neonatal period, the illness
is then detected upon progressive goiter growth in early life.
In such cases, asymptomatic parents are generally heterozygous carriers of Tg mutations, and there is frequently a history of consanguinity (347). In most human patients suffering
from congenital hypothyroid goiter with defective Tg, the
molecular defect has not been established. Although postulated defects in these cases have centered around abnormal
translation or altered posttranslational modifications such as
glycosylation, one of the more common defects may involve
Tg trafficking out of the ER. This, is supported by the frequent finding of markedly distended thyrocyte ER, which
immunostains positively with anti-Tg, while a relative absence of Tg is found in the follicle lumen and in the circulation. Moreover, a recent biochemical investigation of affected individuals from two independent kindreds with
congenital Tg deficiency revealed marked elevations in the
thyroidal levels of ER chaperones BiP and GRP94 (up to
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Vol. 19, No. 2
FIG. 3. Hypothesis for how luminal chaperones may influence productive and unproductive branches of the Tg folding pathway within the ER.
We propose that nascent Tg translocation into the ER lumen is completed with the assistance of calnexin and perhaps BiP and other chaperones.
The newly translated Tg monomers are highly unfolded, unstable, and excellent “substrates” for the binding of GRP94, BiP, and other
chaperones. In this model, the binding of GRP94 and BiP do not necessarily promote advancement to mature folded monomers; indeed, their
binding per se may kinetically inhibit such advancement (slight block shown in figure), which proceeds during the part of the chaperone
interaction cycle in which chaperones are released from a particular domain on Tg—the unbound domain then uses this period to properly bury
chaperone-binding sites, leading to progressive folding toward the native state. In the productive folding pathway, the appearance of compact
monomers is followed rapidly by Tg assembly into homodimers. It is not known what chaperones, if any, play a role in the dimerization process
(shown as question mark above the arrow signifying dimerization). Dimers are the predominant, if not exclusive, form of Tg that undergoes export
from the ER. An alternative, unproductive pathway taken by a significant fraction of unfolded Tg monomers involves the formation of protein
complexes (downward vertical arrow), which are likely to include one or more improper intrachain disulfide bonds as well as nonnative
intermolecular hydrophobic interactions. GRP94 and BiP bind extensively to molecules that either have formed or are prone to forming these
complexes, thereby inhibiting advancement down this potentially unproductive folding pathway. The extent of chaperone-mediated promotion
of productive folding is likely to be related to minimizing Tg entrapment in protein aggregates (block of downward vertical arrow), rather than
direct enhancement of Tg conformational maturation. Because of the oxidizing environment of the ER, some of the molecules in Tg complexes
form aberrant interchain disulfide bonds. Nevertheless, the cyclic association of numerous chaperones occurs in conjunction with some proper
monomer folding that proceeds even within the complexes. Thus, of those Tg aggregates that do occur, many are reversible in the folding pathway,
suggesting that unfolded monomers are in a dynamic equilibrium with Tg complexes. PDI is likely to play a major role in breaking mispaired
disulfide bonds within Tg complexes; PDI therefore acts as a true foldase for Tg (shown as upward vertical arrow). However, an indeterminate
portion of Tg does not recover from these misfolded states; this is especially true for mutant Tg, which is intrinsically defective in progression
along the normal folding pathway. In this case, it is possible that ERp72, and perhaps ER60, assist in initiating ER-associated degradation
of Tg, which may eventually occur in the cytosol (see text).
;10-fold), while intracellular Tg was synthesized but never
acquired complex carbohydrate modifications, indicating its
failure to reach the Golgi compartment (348). These and other
biological defects observed in the thyroids of patients with
hereditary goitrous hypothyroidsm appear very similar to
those found in animal models of this disease, in particular,
the cog/cog mouse.
The cog (congenital goiter) mutation, which arose spontaneously in the inbred AKR/J strain, is an autosomal recessive
gene tightly linked to the Tg locus in the central region of
mouse chromosome 15 (349), associated with the development of congenital hypothyroid goiter and its attendant
growth (350) and neurological (351) sequelae. Morphological
studies of cog/cog mice indicate that the thyrocytes have abnormally distended ER (352, 353) as do those of certain human patients suffering from congenital goiter due to Tg
deficiency (354, 355). As in the humans, Tg mRNA is abundant and apparently normal in size (356), but purified Tg
protein exhibits abnormal biophysical properties including
enhanced susceptibility to proteolysis (357, 358). An extensive analysis has recently shown that the mutant mice synthesize a full-length Tg protein, which undergoes N-linked
glycosylation and glucose trimming in the ER, indistinguish-
able from that in normal thyroid tissue; nevertheless, homodimerization of Tg is virtually undetectable. Consequently, the Tg protein is deficient for export, resulting in
diminished synthesis of thyroid hormone in the affected
mice. The underlying Tg folding defect appears to be due to
a temperature-sensitive mutation such that at 37 C, the quantity of Tg protein arriving in the Golgi complex is below the
limits of detection (143). The physiological response to this
illness includes the specific thyroidal induction of five ER
molecular chaperones: BiP, GRP94, ERp72, ER60, and calreticulin. Based on the logic described in Section III.C, Tg export
is probably prevented either because of the formation of
protein aggregates that are unable to advance into ER export
vesicles, or because of increased binding to ER chaperones
that function to retain misfolded proteins, or both. Indeed,
unlike normal mouse Tg, Tg from the cog/cog mice exhibits
near-quantitative, prolonged binding to GRP94 and BiP
(143), and the vast majority of newly synthesized Tg protein
is ultimately degraded without ever reaching the Golgi complex.
Remarkably, even without exogenous hormone replacement, cog/cog mice, as well as certain human patients with
congenital goiter due to Tg deficiency, become biochemically
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ENDOCRINE ERSDs
euthyroid with age. The reason seems to be that a very minor
fraction of Tg can in fact escape the ER and be exported to
the Golgi complex and beyond (143), where it becomes iodinated (358). Theoretically, if only 2% of all Tg reaches the
follicle lumen, as the goiter reaches 50 times normal size, total
thyroid hormone production could normalize even without
any improvement of Tg folding or special actions of molecular chaperones. In addition, as a consequence of chronic
hypothyroxinemia resulting in supranormal TSH stimulation, T3 may be preferentially formed (359). Of course, the
precise nature of the mutation in the encoded Tg is likely to
be a determining factor in the success of such compensatory
responses, both qualitatively and quantitatively. Recently, a
cDNA library has been generated from thyroids of the mutant cog/cog mice, and a full-length Tg cDNA has been successfully isolated. By comparing the Tg coding sequence to
that of the unaffected parental AKR/J strain, it is expected
that the precise molecular defect should soon be identified.
However, thus far there is no evidence that Tg mutations
leading to congenital goiter are conserved between species,
or even between different affected human kindreds (345).
Preliminary sequencing does indicate that the primary structures of all four hormonogenic domains within Tg (360) are
preserved in the Tg cDNA from the mutant mice.
Because Tg folding and export from the ER is normally a
slow step in the thyroid hormonogenic pathway, mutations
in Tg that by “sequence-gazing” may appear only minor in
nature, may be sufficient to further reduce Tg export to the
point where its transport becomes limiting for thyroid hormone synthesis. Thus, it is probable that many (but by no
means all) kindreds with genetically transmitted hypothyroid goiter, including those previously attributed to other
causes such as defective iodination machinery, may suffer
from a lack of available Tg substrate at the site where thyroid
hormone formation takes place (361). Assuming that each of
the two Tg alleles are expressed equally in heterozygous
carriers of this illness, Tg export in such cases might represent as much as 75% of normal (if wild-type/mutant dimers
are competent for transport) and as little as 25% of normal (if
only wild-type homodimers are competent for transport).
Thus, humans and animals heterozygous for Tg synthesis
defects are likely to have an increased predilection for hypothyroidism, especially in the setting of iodide deficiency or
comparable metabolic insult. Normally, there is sufficient
gain control by TSH stimulation of the thyroid gland, such
that frank hypothyroidism should not develop in heterozygotes, although chronic low-level TSH stimulation may predispose to goiter and autonomous thyroid growth in these
individuals (362).
B. Familial neurohypophyseal diabetes insipidus
Familial central diabetes insipidus (FDI) is an uncommon
form of diabetes insipidus (DI) caused by absence of circulating arginine vasopressin (AVP), a nonapeptide derived
from a larger single chain neurophysin-vasopressin precursor synthesized by magnocellular neurons of the hypothalamus (363, 364). A single copy of the AVP peptide is immediately preceded by a leader peptide that is cleaved from the
preprohormone by signal peptidase. Subsequent proteolytic
187
processing of the prohormone generates AVP as well as
neurophysin (NP) and a C-terminal glycoprotein fragment.
Accounting for more than two-thirds of the prohormone
mass, NP is thought to act as a molecular escort for AVP
along the intracellular secretory pathway (see Section II.G), by
forming a noncovalent heterodimer with AVP, which then
combines with other heterodimers and ultimately forms
even higher order complexes in the secretory pathway (364,
365).
Remarkably, genetic analyses have clearly established that
most FDI in humans is autosomal dominant (363, 366, 367).
An insight into this mode of transmission may be gained
from autopsy studies, in which patients with FDI are found
to exhibit marked degeneration of the neurons responsible
for AVP synthesis, storage, and secretion, for which an abnormal prohormone structure has been implicated as the root
cause. Molecular studies of affected families reveal assorted
point mutations in the AVP gene that tend to cluster in the
NP coding region, although they are not exclusive to this
region (368 –371). In rare cases, mutations affecting only the
leader sequence (372) or the translational initiation codon
(373) of the AVP-NP precursor lead to FDI. A delay in removal of the leader sequence is likely to result in severe
protein misfolding, which may already be irreversible by the
time the leader is, if ever, removed (63). In such a case the
mutants would not be expected to exit the ER as a result of
ER quality control. The more common mutations affecting
the NP-coding region may dramatically alter the structural
integrity of the AVP-NP precursor and/or the processed
complex. But why would such mutations in either the leader
sequence or the NP region lead to cell death? One possibility
supported by recent studies (460) is that intracellular accumulation of the mutant gene product results in cellular toxicity (ERSD type A-II, see Section V and Fig. 4, below). While
it is by no means established, it seems probable in human
patients that intracellular transport of AVP-NP from the ER
is affected; moreover, from the identification of signal sequence mutations leading to FDI, an intracellular transport
block can be suspected.
Interestingly, the Brattleboro rat, a remarkable animal
model of FDI, synthesizes a mutant precursor protein that
has been shown to accumulate in the ER (374, 375). In this
case, a frame shift due to a single base deletion in the NPcoding region results in elimination of the stop codon, causing translational readthrough of the poly-A tail of AVP-NP
mRNA. Thus, the mutant neurophysin-vasopressin precursor not only contains an altered NP domain, but a new
carboxy terminus that includes a polylysine tail that cannot
be fully translocated into the ER lumen. However, genetic
transmission in the Brattleboro rat is autosomal recessive,
which does not fit well with the idea that toxic protein accumulation causes complete neuronal cell death, at least
within heterozygous individuals (363). Such a finding is consistent with the possibility that in the Brattleboro rat, the
misfolded AVP-NP protein may be successfully degraded,
but this may not necessarily be true in many of the human
kindreds (see Section V). Evidently, studies are needed to
provide a cellular explanation for the discrepancy in the
mode of inheritance between most human kindreds with FDI
and the Brattleboro rat model (376, 377), which should in-
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KIM AND ARVAN
clude an analysis of the levels and functions of ER molecular
chaperones in different animals with this illness.
Finally, it should be pointed out that certain forms of
genetically inherited nephrogenic DI (378, 379) may also be
ERSDs. An autosomal recessive phenotype can apparently be
caused by point mutations within the AVP-regulated water
channel, known as aquaporin-2, which results in quantitative
retention of the mutant channel in the ER (380). Additional
mutants, such as those found in the V2 AVP receptor (381–
383), are also considered likely candidates as ERSDs.
C. Osteogenesis imperfecta and disorders of procollagen
biosynthesis
Collagen, secreted by connective tissues, exhibits a characteristic triple helical structure made possible by Gly-X-Y
repeats (in which X and Y are most often proline and hydroxyproline, respectively). The triple helix can further pack
to form the superhelix found in collagen fibrils (82). Of the
roughly 15 different types of collagen, type I procollagen is
by far the most studied (384). Type I is a hetero-trimer of two
pro-a-1 chains and one pro-a-2 chain. These chains are encoded by the COL1A1 and COL1A2 genes, respectively (83,
385, 386). As a prepro-a-chain enters the ER lumen, removal
of its lengthy signal peptide and glycosylation near the C
terminus takes place (387). Further, specific prolyl and lysyl
residues become hydroxylated, due to the actions of prolyl4-hydroxylase (211) and lysyl hydroxylase (388), two vitamin
C-dependent enzymes in the ER (389). Occasionally, hydroxylysines are further modified by glycosylation. Shortly
after completion of pro-a-chain synthesis, heterotrimer assembly is initiated by associations between the globular regions at the extreme C-termini of each pro-a-monomer (83,
390). The noncovalent triple helix is then propagated back
from the C- to the N terminus, while interchain disulfide
bonds begin to form, further stabilizing the helical structure.
Once completed, the trimers undergo intracellular transport,
whereupon the nonhelical N- and C terminal ends are proteolytically cleaved off and the trimers further assemble into
superhelical collagen fibrils (83, 386).
Many ER resident proteins, including HSP47 (227, 228,
230), participate along the folding and assembly pathway of
procollagens (144, 229, 391). In addition to its role as one of
the subunits of prolyl-4-hydroxylase, PDI may also act independently both as a molecular chaperone and folding catalyst during procollagen maturation (80, 81). Several other
ER chaperones and folding catalysts that have been reported
to assist in procollagen folding are PPI, BiP, and possibly
GRP94 (102, 144, 392).
Despite somewhat confusing nomenclature, great
progress has been made in recent years in elucidating the
molecular bases for osteogenesis imperfecta (OI). Mutations
in type I procollagen, the major protein component of bone
matrix, characterizes this heterogeneous group of “brittle
bone diseases” (83, 386). Interestingly, OI can be inherited
either as an autosomal dominant (the vast majority of cases),
or in a recessive mode. Because of the multiple and complex
steps of procollagen biosynthesis, genetic errors are manifested at numerous possible stages in the pathway; this, and
the ease of obtaining primary cultured disease fibroblasts,
Vol. 19, No. 2
may account for why OI is one of the most diverse and
extensively studied ERSDs. At present, more than 150 mutations in the two genes encoding type I procollagen are
reportedly responsible for various forms of OI, ranging from
death in utero to only mild bone fragility (393–395). Mutations
have been mapped all along the coding regions, but frequently represent point mutations in one of the Gly-X-Y
repeats, which disrupt the triple helical domain (396 – 400).
Indeed, it has been suggested that the severity of the disease
can be roughly correlated with the location of the mutation
along the length of pro-a: mutations in the C-terminal globular region generally have been associated with more severe
phenotypes (83, 401). Moreover, mutant chains can associate
with normal chains, creating an unstable or poorly functional
complex that accounts for the autosomal dominant transmission. This is further supported by the observation that
“null alleles” actually tend to cause relatively milder disease
(402, 403).
In one recent study, a Trp13123 Cys substitution near the
C terminus of pro-a-1 was found to be responsible for a
lethal form of OI (391). First, despite normal mRNA stability, primary fibroblast cell cultures revealed a greater
reduction in total collagen synthesis than would be expected from a null allele, due to a higher than normal rate
of intracellular degradation of pro-a-chains. Second, malfolded mutant subunits containing aberrant disulfide
bonds were found to accumulate in the ER in association
with BiP and were selectively degraded in a pre-Golgi
compartment. Third, BiP levels were further increased in
the mutant cells by ascorbate treatment, which causes a
rapid increase in procollagen synthesis. Nevertheless,
some of the procollagen trimers containing a mutant subunit were secreted, although they were abnormally posttranslationally modified, further perturbing the already
deficient extracellular matrix. Interestingly, the more common procollagen mutants that contain defects located
within the triple helical structure exhibited neither increased interaction with BiP nor induction of BiP synthesis
(102).
Considerable progress has also been made in delineating
the molecular mechanism of other procollagen disorders, for
which similarly numerous mutations exist that range in severity of phenotype (404 – 409). It is beyond the scope of this
paper to review all the mutations that cause both common
and rare diseases of collagen (83, 385). However, we note that
in a recently reported case, a single amino acid substitution
of Gly8533 Glu within the sequence encoding the triple helix
of type II procollagen (COL2A1) produces a lethal form of
hypochondrogenesis, in which the affected chondrocytes exhibit dilated ER that contains mutant procollagen molecules
impaired in their assembly and intracellular transport (404).
Finally, defective intracellular transport of other noncollagen
proteins of the extracellular matrix may also lead to varying
forms of OI and other skeletal dysplasias (405, 406, 408, 410,
411).
D. ERSDs affecting lipoprotein metabolism
Many clinically significant mutations have been identified
that affect lipoprotein metabolism (for review, see Refs. 412
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and 413); this section notes only a few examples, which are
apparent ERSDs. The LDLR, a 160-kDa cell surface glycoprotein, is responsible for the hepatic uptake of most LDL
particles from the circulation by endocytosis (414 – 416). Thus
far, several dozen mutations causing autosomal dominant
hypercholesterolemia have been reported, with molecular
defects ranging from lack of synthesis, altered intracellular
transport, or abnormal function (417). In one study, using
human fibroblasts isolated from homozygous patients derived from three different Lebanese families, a single base
substitution in the coding sequence near the C terminus of
the protein resulted in a truncated receptor that was quantitatively retained in the ER (418). The steady state level of the
mutant receptor was reduced ;10-fold, and additional evidence suggested rapid ER degradation of the mutant protein. Subsequent microscopic studies of skin fibroblasts obtained from similar patients have confirmed ER retention of
mutant LDLR (419).
Apolipoprotein B is synthesized in the ER of hepatocytes
and enterocytes, where it is lipid-loaded as a consequence of
interaction with the microsomal triglyceride transfer protein
(MTP) complex. Remarkably, abetalipoproteinemia is an
ERSD not due to mutations in the apolipoprotein B gene, but
is due to an absence of functional MTP (420). Several of the
reported mutations in such patients map to the region of the
coding sequence in MTP responsible for interaction with a
PDI subunit of MTP (421). Moreover, MTP has been proven
to physically associate with newly synthesized apolipoprotein B, and in the absence of lipid-loading by MTP, apolipoprotein B is subject to rapid intracellular degradation that
can be prevented by inhibitors of proteasomal proteolysis
(422). In the absence of such inhibitors, unique proteolytic
fragments of the apolipoprotein can be found in plasma
(423). At present it is unclear whether MTP assists as a cotranslocational chaperone for apolipoprotein B as it crosses
the ER membrane (424), or acts primarily during posttranslational folding (425).
Lipoprotein lipase (LPL), a 55-kDa glycoprotein secreted
by fat and muscle cells, is transported to the capillary endothelium where it catalyzes hydrolysis of triglycerides
within chylomicrons and very-low-density lipoprotein particles. LPL deficiency, inherited as an autosomal recessive
trait, accounts for most cases of familial hyperchylomicronemia (426). Regulation of LPL activity occurs primarily at the
level of its synthesis, N-linked glycosylation, trimming of
terminal glucose residues, and homodimerization that leads
to formation of the active enzyme (427– 429). Most of these
LPL processing steps occur in the ER (430, 431). Many LPL
mutations have been reported to be single amino acid substitutions in the coding sequence, but Pro2073 Leu accounts
for the majority of all cases of familial chylomicronemia due
to LPL deficiency (432). Biochemical comparisons of this
mutant and wild-type LPL expressed in COS-1 cells showed
similar levels of mRNA, but significantly reduced secretion
of the mutant LPL with its greater intracellular retention.
Moreover, the small, secreted amount of this LPL mutant is
functionally inactive (ERSD type B-I; see Section V and Fig.
4, below).
189
E. Other selected nonendocrine and endocrine ERSDs
a-1-Antitrypsin (AAT) deficiency is a well-known cause of
juvenile pulmonary emphysema, due to absence of the hepatically secreted serine protease inhibitor at its site of action
in the lungs. For review of the many forms of AAT deficiency,
see Ref. 433. Herein, we wish to mention only one antitrypsin
mutant in particular, the so-called PiZ, or Z-variant. Interestingly, extensive hepatic damage and early cirrhosis have
been directly linked with the expression of this particular
mutant AAT in some patients (11). Although a high fraction
of the Z-variant is degraded within the ER (434, 435), the
hepatic injury is thought to result from hepatocellular accumulation of an undegraded fraction of insoluble polymeric
Z-variant within the ER (436). It is tempting to speculate (see
Ref. 460) that this cellular toxicity could represent a similar
mechanism to that reported for familial central diabetes insipidus (see Section IV.B). It should be pointed out, however,
that AAT deficiency due to the Z-variant is an autosomal
recessive disease. This observation suggests that gene dosage
may affect not only the amount of antitrypsin that is secreted
but also the amount of antitrypsin that may contribute to
protein accumulation within the ER (ERSD type B-II, see
Section V and Fig. 4, below). Indeed, it is controversial
whether patients heterozygous for the Z-variant may be predisposed to develop liver cirrhosis without further metabolic
insult. However, it should be noted that expression of the
human Z-variant in transgenic mice does result in liver cirrhosis among a subpopulation of these animals.
Interestingly, unlike other AAT mutant alleles, the Z-variant of AAT is not thought to elevate the steady-state levels
of BiP (437); an inference from these data is that it may bind
BiP only poorly (see Section II.A). Since polymerization of the
Z-variant of AAT in the ER apparently represents an ordered,
protease-resistant assembly, it is possible that exposed domains on the misfolded mutant antitrypsin may be used
either for chaperone binding or polymeric assembly. Based
on the relative affinities of these competing interactions, as
well as the expression level of the “substrate,” a fraction of
the molecules might be favored to form an insoluble polymer
depending on chaperone concentrations and availability.
Failure to either export or completely degrade the molecule
may lead to a situation in which gradual, but inexorable
accumulation of this indigestible aggregate develops, ultimately leading to general cellular stress (438) and toxicity
(see Fig. 4, below).
Cystic fibrosis (CF) is one of the most common hereditary
disorders in the Caucasian population, and an allele resulting
in a single Phe deletion at position 508 in the CF transmembrane conductance regulator (CFTR), present in more than
70% of all cases, is responsible for protein misfolding (439).
In the homozygous state, abnormal chloride conductance
appears to be caused by functional absence of the CFTR at the
plasma membrane of epithelial cells, where this polytopic
membrane protein is normally expressed (440). Furthermore,
it has been well established that the mutant CFTRDF508
protein is synthesized but fails to exit the ER, wherein it is
retained by association with calnexin (441). Cytoplasmic
molecular chaperones also appear to interact with the
CFTRDF508 mutant (442), which is not surprising, since the
190
KIM AND ARVAN
Vol. 19, No. 2
FIG. 4. A generic classification of ERSDs. All ERSDs begin with ER retention of misfolded proteins (center of figure), which is merely the end
of the spectrum of ER retention that already exists for wild-type exportable proteins (315, 441). As described in Section IV, these mutants may
either be potentially functional (type A) or nonfunctional (type B). Nonfunctional mutants will be dominantly inherited if they can assemble
with the wild-type gene product, creating nonfunctional oligomers, or if 13 gene expression of the wild-type allele is insufficient for hormonal
homeostasis. The latter case is generally rare because endocrine feedback mechanisms allow for increased hormone production from the
wild-type gene product. In many dominant and most recessive ERSDs, even though the mutant proteins cannot escape the ER, they nevertheless
are efficiently degraded intracellularly (subtypes A-I and B-I). More rarely, mutant proteins may not be efficiently degraded, such that an
undegraded portion may accumulate (460), which can lead to cellular toxicity or cell death (subtypes A-II and B-II). If toxic accumulation occurs
with only a 13 level of gene expression, a dominant mode of inheritance will be apparent. If both mutant alleles must be expressed before toxic
accumulation is detected, such as for the Z-variant of a1-antitrypsin (see Section IV.E), then cellular toxicity will exhibit a recessive pattern
of transmission.
majority of each CFTR polypeptide chain is cytoplasmically
disposed. The ER-retained protein is then rapidly degraded
by a pathway thought to involve ubiquitination followed by
proteosomal proteolysis [(315, 329, 330), see Section III.D].
Remarkably, there is strong evidence that the CFTRDF508
protein has near-normal functional capability as a chlorideconducting channel, although the vast majority of the protein
never reaches its intended site of biological action (443). More
remarkably, an important fraction of the wild-type CFTR also
fails to be exported from the ER in association with calnexin
and is then rapidly degraded (441). Evidently, the difference
in cellular phenotype between the wild-type and mutant
DF508 protein is largely a matter of degree. In support of this,
an increased fraction of recombinant CFTRDF508 is detected
at the plasma membrane in cells grown at reduced temperature, and this cell surface CFTR is functionally competent
(440). Presumably, the slower rate of polypeptide chain folding at lower temperatures may increase the probability of
reaching a “more correct” final tertiary structure.
Table 1 summarizes a number of additional endocrinopathies in which ERSDs can be suspected or have been implicated. Although detailed cell and molecular studies are
still needed in many of these cases, the fundamental link
appears to be defective folding and intracellular transport of
important endocrine polypeptides (Table 1). This table is by
no means comprehensive; for example, not included therein
is an important new animal model that suffers from a global
polyendocrinopathy associated with notable obesity and infertility, known as the fat/fat mouse, which is caused by an
ERSD involving defective export of the prohormoneprocessing enzyme, carboxypeptidase E (455, 456).
V. Summary: A Proposed Classification of ERSDs
From the studies described in this review, it is clear that
structural information dictates not only the functional properties of exportable proteins, but also their ability to be transported in the intracellular secretory pathway. In ERSDs, the
precise nature of the defect determines both the severity of
the phenotype and the mode of inheritance. To our knowledge, all genetically inherited ERSDs are attributable to mutations in the coding sequence of exportable proteins; thus
far, with the exception of abetalipoproteinemia (see Section
IV.D), no mutations in ER chaperones (other than those that
scientists have genetically engineered) have been reported as
the cause of spontaneous disease.
The elevations of ER chaperones in ERSDs may differ
between mutations, between tissues, between individual patients, and between different physiological states (i.e., such as
before and after hormone replacement therapy) in the same
patient. Thus, measurement of ER chaperone levels plays an
important diagnostic role, but probably should not be used
as the sole basis to classify these illnesses. Moreover, because
mutant secretory proteins have been reported to occur in
virtually every organ system, ERSDs are more readily classified at the cell biological level, by the responses of the cells
that actually synthesize the secretory protein, rather than the
April, 1998
ENDOCRINE ERSDs
TABLE 1. Some endocrinopathies due to ERSDs
Congenital hypothyroidism and related disorders
Thyroglobulin deficiency
Thyroid peroxidase deficiency
Thyroxine binding globulin (TBG) deficiency
Diabetes insipidus and related disorders
Vasopressin precursor defect
Vasopressin V2-receptor defect
Aquaporin defect
Osteogenesis imperfecta and related disorders
Type I procollagen deficiency
Type II procollagen deficiency
Type IV procollagen deficiency
Lipid disorders
LDL receptor deficiency
Microsomal triglyceride transfer protein
deficiency
Lipoprotein lipase deficiency
Lipoprotein(a) defect
Diabetes mellitus
Insulin receptor defect
Growth disorders
Growth hormone receptor defect
Primary hypoparathyroidism
Preproparathormone defect
Refs.
See text
444 – 446
447, 448
See text
See text
See text
See text
See text
See text
See text
See text
449
450
191
teins to the cytosol for proteasomal proteolysis. Resistance of
untransported mutant protein to ER-associated degradation
will predispose to a dominant ERSD (460). In such a case,
although the mutant allele could could form oligomeric hybrids with the wild-type allele, complete nonmixing of the
normally exported wild-type allele and toxic accumulation of
the mutant allele is another distinct scenario that can also
produce a dominant mode of inheritance. For cells that rapidly and repetitively divide, it may be possible to escape
cellular lethality under conditions in which the ER is continuously being expanded in preparation for another round
of cell division. These cells, while still markedly abnormal,
may in effect be able to “outgrow” the toxic accumulation. By
contrast, highly differentiated cells that maintain only
steady-state quantities of ER are likely to be at greater risk of
cell injury or death in the face of perpetual accumulation of
misfolded proteins in the ER (460). Such considerations may
also be important factors in the age of onset for particular
manifestations that represent the natural history of ERSDs.
451– 453
454
hormone deficiency associated with the illness at the endorgan level.
With these ideas in mind, we present a schematic view in
Fig. 4. According to this schema, all ERSDs begin with ER
retention of the affected proteins or their subunits. Mutants
may then be divided into two groups: type A, where the
biological activity is preserved although the protein is transport-deficient; and type B, where the mutant has no potential
for functional activity. Both categories include both recessive
and dominant mutations. The primary clinical difference
between these two classes is that type A ERSDs may be
amenable to therapies designed to down-regulate the quality
control of ER export so that potentially functional molecules
can escape the ER and reach their intended intracellular
destination. In both types of ERSDs, in most cases, the retained mutant protein is efficiently degraded in the ER (subtypes A-I and B-I). In these cases, the predominant, global
phenotypes involve the symptoms and signs of hormone
deficiency. However, careful biochemical and cell biological
studies reveal various abnormalities in glandular function,
typically including the elevation of the levels of one or more
ER chaperones. As described in Section I.C, such elevations
are a consequence of chronic adaptation to the presence of
unfolded mutant secretory protein (the synthesis of which is
stimulated all the more by endocrine feedback loops). As
described in Section III, the elevated chaperones appear to be
integrally related to the ER retention as well as perhaps the
ERAD process that removes the misfolded proteins. In these
cases, the ER compartment may expand, but the secretory
cells are likely to survive.
In the more unusual subtype II (subtypes B-II and perhaps
A-II), the mutant protein exhibits an intrinsic tendency to
resist ERAD, creating a potentially dangerous accumulation
of indigestible material (Fig. 4). This may be due to the
unusual production of novel, protease-resistant protein complexes, or it may be due to the formation of protein assemblies that prevent the reverse translocation of mutant pro-
Acknowledgment
We thank Dr. I. Boime (Washington University, St. Louis, MO) for
helpful discussions.
References
1. Rothman JE 1994 Mechanisms of intracellular protein transport.
Nature 372:55– 63
2. Kornfeld S 1986 Trafficking of lysosomal enzymes in normal and
disease states. J Clin Invest 77:1– 6
3. Nishimura N, Balch WE 1997 A Di-acidic signal required for selective export from the endoplasmic reticulum. Science 277:556 –
558
4. Machamer CE 1993 Targeting and retention of Golgi membrane
proteins. Curr Opin Cell Biol 5:606 – 612
5. Kuliawat R, Arvan P 1994 Distinct molecular mechanisms for
protein sorting within immature secretory granules of pancreatic
b-cells. J Cell Biol 126:77– 86
6. Munro S 1995 An investigation of the role of transmembrane domains in Golgi protein retention. EMBO J 14:4695– 4704
7. Rapoport TA, Rolls MM, Jungnickel B 1996 Approaching the
mechanism of protein transport across the ER membrane. Curr
Opin Cell Biol 8:499 –504
8. Andrews DW, Johnson AE 1996 The translocon: more than a hole
in the ER membrane? Trends Biochem Sci 21:365–369
9. Hammond C, Helenius A 1995 Quality control in the secretory
pathway. Curr Opin Cell Biol 7:523–529
10. Kopito RR 1997 ER quality control: the cytoplasmic connection.
Cell 88:427– 430
11. Callea F, Brisigotti M, Fabbretti G, Bonino F, Desmet VJ 1992
Hepatic endoplasmic reticulum storage diseases. Liver 12:357–362
12. Anfinsen CB 1973 Principles that govern the folding of protein
chains. Science 181:223–230
13. Pace CN, Shirley BA, McNutt M, Gajiwala K 1996 Forces contributing to the conformational stability of proteins. FASEB J 10:
75– 83
14. Johnson JL, Craig EA 1997 Protein folding in vivo: unraveling
complex pathways. Cell 90:201–204
15. Rothman JE 1989 Polypeptide chain binding proteins: catalysts of
protein folding and related processes in cells. Cell 59:591– 601
16. Gething M-J, Sambrook J 1992 Protein folding in the cell. Nature
355:33– 45
17. Koch GLE 1987 Reticuloplasmins: a novel group of proteins in the
endoplasmic reticulum. J Cell Sci 87:491– 492
18. Helenius A, Marquardt T, Braakman I 1992 The endoplasmic
192
19.
20.
21.
22.
23.
24.
25.
26.
27.
28.
29.
30.
31.
32.
33.
34.
35.
36.
37.
38.
39.
40.
41.
42.
KIM AND ARVAN
reticulum as a protein folding compartment. Trends Cell Biol 2:227–
231
Netzer WJ, Hartl FU 1997 Recombination of protein domains facilitated by co-translational folding in eukaryotes. Nature 388:343–
349
Sambrook JF 1990 The involvement of calcium in transport of
secretory proteins from the endoplasmic reticulum. Cell 61:197–199
Clairmont CA, De Maio A, Hirschberg CB 1991 Translocation of
ATP into the lumen of rough endoplasmic reticulum-derived vesicles and its binding to luminal proteins including BiP (GRP 78) and
GRP 94. J Biol Chem 267:3983–3990
Hwang C, Sinskey AJ, Lodish HF 1992 Oxidized redox state of
glutathione in the endoplasmic reticulum. Science 257:1496 –1502
Freedman RB 1995 The formation of protein disulfide bonds. Curr
Opin Struct Biol 5:85–91
Gliniak BC, Kozak SL, Jones RT, Kabat D 1991 Disulfide bonding
controls the processing of retroviral envelope glycoproteins. J Biol
Chem 266:22991–22997
Kim P, Bole D, Arvan P 1992 Transient aggregation of nascent
thyroglobulin in the endoplasmic reticulum: relationship to the
molecular chaperone, BiP. J Cell Biol 118:541–549
Weissman JS, Kim PS 1991 Reexamination of the folding of BPTI:
predominance of native folding intermediates. Science 253:1386 –
1392
de Silva A, Braakman I, Helenius A 1993 Posttranslational folding
of vesicular stomatitis virus G protein in the ER: involvement of
noncovalent and covalent complexes. J Cell Biol 120:647– 655
Prabakaran D, Kim PS, Dixit VM, Arvan P 1996 Oligomeric assembly of thrombospondin in the endoplasmic reticulum of thyroid epithelial cells. Eur J Cell Biol 70:134 –141
Ellis RJ, Van Der Vies SM, Hemmingsen SM 1989 The molecular
chaperone concept. Biochem Soc Symp 55:145–153
Craig EA, Weissman JS, Horwich AL 1994 Heat shock proteins and
molecular chaperones: mediators of protein conformation and
turnover in the cells. Cell 78:365–372
Price BD, Mannheim-Rodman LA, Calderwood SK 1992 Brefeldin
A, thapsigargin, and AIF4- stimulate the accumulation of GRP78
mRNA in a cycloheximide dependent manner, whilst induction by
hypoxia is independent of protein synthesis. J Cell Physiol 152:
545–552
Goering PL, Fisher BR, Kish CL 1993 Stress protein synthesis
induced in rat liver by cadmium precedes hepatotoxicity. Toxicol
Appl Pharmacol 122:139 –148
Kuznetsov G, Bush KT, Zhang PL, Nigam SK 1996 Perturbations
in maturation of secretory proteins and their association with endoplasmic reticulum chaperones in a cell culture model for epithelial ischemia. Proc Natl Acad Sci USA 93:8584 – 8589
Lee AS 1992 Mammalian stress response: induction of the glucoseregulated protein family. Curr Opin Cell Biol 4:267–273
Little E, Ramakrishnan M, Roy B, Gazit G, Lee AS 1994 The
glucose-regulated proteins (GRP78 and GRP94): functions, gene
regulation, and applications. Crit Rev Eukaryot Gene Expr 4:1–18
Pahl HL, Baeuerle PA 1997 Endoplasmic-reticulum-induced signal
transduction and gene expression. Trends Cell Biol 7:50 –55
Kozutsumi Y, Segal M, Normington K, Gething MJ, Sambrook J
1988 The presence of malfolded proteins in the endoplasmic reticulum signals the induction of glucose regulated proteins. Nature
332:462– 464
Ng DTW, Watowich SS, Lamb RA 1992 Analysis in vivo of GRP78BiP/substrate interactions and their role in induction of the GRP78BiP gene. Mol Biol Cell 3:143–155
Kohno K, Normington K, Sambrook J, Gething MJ, Mori K 1993
The promoter region of the yeast KAR2 (BiP) gene contains a
regulatory domain that responds to the presence of unfolded proteins in the endoplasmic reticulum. Mol Cell Biol 13:877– 890
Cox JS, Shamu CE, Walter P 1993 Transcriptional induction of
genes encoding endoplasmic reticulum resident proteins requires
a transmembrane protein kinase. Cell 73:1197–1206
Mori K, Ma W, Gething MJ, Sambrook J 1993 A transmembrane
protein with a cdc21/CDC28-related kinase activity is required for
signaling from the ER to the nucleus. Cell 74:743–756
Welihinda AA, Kaufman RJ 1996 The unfolded protein response
pathway in Saccharomyces cerevisiae. Oligomerization and trans-
43.
44.
45.
46.
47.
48.
49.
50.
51.
52.
53.
54.
55.
56.
57.
58.
59.
60.
61.
62.
63.
64.
Vol. 19, No. 2
phosphorylation of Ire1p (Ern1p) are required for kinase activation.
J Biol Chem 271:18181–18187
Chavany C, Mimnaugh E, Miller P, Bitton R, Nguyen P, Trepel
J, Whitesell L, Schnur R, Moyer J, Neckers L 1996 p185erbB2 binds
to GRP94 in vivo. Dissociation of the p185erbB2/GRP94 heterocomplex by benzoquinone ansamycins precedes depletion of
p185erbB2. J Biol Chem 271:4974 – 4977
Cao X, Zhou Y, Lee AS 1995 Requirement of tyrosine- and serine/
threonine kinases in the transcriptional activation of the mammalian grp78/BiP promoter by thapsigargin. J Biol Chem 270:494 –502
Wang XZ, Lawson B, Brewer JW, Zinszner H, Sanjay A, Mi LJ,
Boorstein R, Kreibich G, Hendershot LM, Ron D 1996 Signals
from the stressed endoplasmic reticulum induce C/EBP-homologous protein (CHOP/GADD153). Mol Cell Biol 16:4273– 4280
Cox JS, Walter P 1996 A novel mechanism for regulating activity
of a transcription factor that controls the unfolded protein response.
Cell 87:391– 404
Sidrauski C, Cox JS, Walter P 1996 tRNA ligase is required for
regulated mRNA splicing in the unfolded protein response. Cell
87:405– 413
Brown MS, Goldstein JL 1997 The SREBP pathway: regulation of
cholesterol metabolism by proteolysis of a membrane-bound transcription factor. Cell 89:331–340
Pahl HL, Baeuerle PA 1995 A novel signal transduction pathway
from the endoplasmic reticulum to the nucleus is mediated by
transcription factor NF-kappa B. EMBO J 14:2580 –2588
Staddon JM, Bouzyk MM, Rozengurt E 1992 Interconversion of
GRP78/BiP. A novel event in the action of Pasteurella multocida
toxin, bombesin, and platelet-derived growth factor. J Biol Chem
267:25239 –25245
Freiden PJ, Gaut JR, Hendershot LM 1992 Interconversion of three
differentially modified and assembled forms of BiP. EMBO J 11:
63–70
Gaut JR, Hendershot LM 1993 The immunoglobulin-binding protein in vitro autophosphorylation site maps to a threonine within
the ATP binding cleft but is not a detectable site of in vivo phosphorylation. J Biol Chem 268:12691–12698
Csermely P, Miyata Y, Schnaider T, Yahara I 1995 Autophosphorylation of grp94 (endoplasmin). J Biol Chem 270:6381– 6388
Feldweg AM, Srivastava PK 1995 Molecular heterogeneity of tumor rejection antigen/heat shock protein GP96. Int Cancer 63:310 –
314
Nakai A, Kawatani T, Ohi S, Kawasaki H, Yoshimori T, Tashiro
Y, Miyata Y, Yahara I, Satoh M, Nagata K 1995 Expression and
phosphorylation of BiP/GRP78, a molecular chaperone in the endoplasmic reticulum, during the differentiation of a mouse myeloblastic cell line. Cell Struct Funct 20:33–39
Blond-Elguindi S, Fourie AM, Sambrook JF, Gething MJ 1993
Peptide-dependent stimulation of the ATPase activity of the molecular chaperone BiP is the result of conversion of oligomers to
active monomers. J Biol Chem 268:12730 –12735
Weissman JS, Kashi Y, Fenton WA, Horwich AL 1994 GroELmediated protein folding proceeds by multiple rounds of binding
and release of nonnative forms. Cell 78:693–702
Knittler MR, Haas IG 1992 Interaction of BiP with immunoglobulin light chain molecules: cycles of sequential binding and release.
EMBO J 11:1573–1581
Rassow J, von Ahsen O, Bomer U, Pfanner N 1997 Molecular
chaperones: towards a characterization of the heat-shock protein 70
family. Trends Cell Biol 7:129 –133
Tian G, Vainberg IE, Tap WD, Lewis SA, Cowan NJ 1995 Specificity in chaperonin-mediated protein folding. Nature 375:250 –253
Rapoport TA, Jungnickel B, Kutay U 1996 Protein transport across
the eukaryotic endoplasmic reticulum and bacterial inner membranes. Annu Rev Biochem 65:271–303
Lyko F, Martoglio B, Jungnickel B, Rapoport TA, Dobberstein B
1995 Signal sequence processing in rough microsomes. J Biol Chem
270:19873–19878
Scheele G, Jacoby R 1983 Proteolytic processing of presecretory
proteins is required for development of biological activities in
pancreatic exocrine proteins. J Biol Chem 258:2005–2009
Huth JR, Mountjoy K, Perini F, Bedows E, Ruddon RW 1992
Domain-dependent protein folding is indicated by the intracellular
April, 1998
ENDOCRINE ERSDs
kinetics of disulfide bond formation of human chorionic gonadotropin b subunit. J Biol Chem 267:21396 –21403
65. Srinivasan N, Sowdhamini R, Ramakrishnan C, Balaram P 1990
Conformations of disulfide bridges. Int J Pept Protein Res 36:147–
155
66. Braakman I, Helenius J, Helenius A 1992 Role of ATP and disulphide bonds during protein folding in the endoplasmic reticulum. Nature 356:260 –262
67. Braakman I, Helenius J, Helenius A 1992 Manipulating disulfide
bond formation and protein folding in the endoplasmic reticulum.
EMBO J 11:1717–1722
68. Lodish HF, Kong N, Wikstrom L 1992 Calcium is required for
folding of newly made subunits of the asialoglycoprotein receptor
within the endoplasmic reticulum. J Biol Chem 267:12753–12760
69. Kaji EH, Lodish HF 1993 Unfolding of newly made retinol-binding
protein by dithiothreitol. Sensitivity to retinoids. J Biol Chem 268:
22188 –22194
70. Kim PS, Kim K-R, Arvan P 1993 Disulfide-linked aggregation of
thyroglobulin normally occurs during nascent protein folding.
Am J Physiol 265:C704 –C711
71. Kim PS, Arvan P 1995 Calnexin and BiP act as sequential molecular
chaperones during thyroglobulin folding in the endoplasmic reticulum. J Cell Biol 128:29 –38
72. Hurtley SM, Helenius A 1989 Protein oligomerization in the endoplasmic reticulum. Annu Rev Cell Biol 5:277–307
73. Kornfeld R, Kornfeld S 1985 Assembly of asparagine-linked oligosaccharides. Annu Rev Biochem 54:631– 664
74. Petrescu AJ, Butters TD, Reinkensmeier G, Petrescu S, Platt FM,
Dwek RA, Wormald MR 1997 The solution NMR structure of
glucosylated N-glycans involved in the early stages of glycoprotein
biosynthesis and folding. EMBO J 16:4302– 4310
75. Kelleher DJ, Kreibich G, Gilmore R 1992 Oligosaccharyltransferase activity is associated with a protein complex composed of
ribophorins I and II and a 48 kd protein. Cell 69:55– 65
76. Reiss G, te Heesen S, Gilmore R, Zufferey R, Aebi M 1997 A
specific screen for oligosaccharyltransferase mutations identifies
the 9 kDa OST5 protein required for optimal activity in vivo and in
vitro. EMBO J 16:1164 –1172
77. Helenius A 1994 How N-linked oligosaccharides affect glycoprotein folding in the endoplasmic reticulum. Mol Biol Cell 5:253–265
78. Feng W, Huth JR, Norton SE, Ruddon RW 1995 Asparagine-linked
oligosaccharides facilitate human chorionic gonadotropin betasubunit folding but not assembly of prefolded beta with alpha.
Endocrinology 136:52– 61
79. Doms RW 1990 Oligomerization and protein transport. Methods
Enzymol 191:841– 854
79a.Bedows E, Norton SE, Huth JR, Suganuma N, Boime I, Ruddon
RW 1994 Misfolded human chorionic gonadotropin beta subunits
are secreted from transfected Chinese hamster ovary cells. J Biol
Chem 269:10574 –10580
80. Chessler SD, Byers PH 1992 Defective folding and stable association with protein disulfide isomerase/prolyl hydroxylase of type
I procollagen with a deletion in the pro alpha 2(I) chain that preserves the Gly-X-Y repeat pattern. J Biol Chem 267:7751–7757
81. Tasanen K, Parkkonen T, Chow LT, Kivirikko KI, Pihlajaniemi
T 1988 Characterization of the human gene for a polypeptide that
acts both as the beta subunit of prolyl 4-hydroxylase and as protein
disulfide isomerase. J Biol Chem 263:16218 –16224
82. Prockop DJ, Kivirikko KI 1984 Heritable diseases of collagen.
N Engl J Med 311:376 –386
83. Prockop DJ, Kivirikko KI 1995 Collagens: molecular biology, diseases, and potentials for therapy. Annu Rev Biochem 64:403– 434
84. Rowling PJE, Freedman RB 1993 Folding, assembly, and posttranslational modification of proteins within the lumen of the endoplasmic reticulum. In: Borgese N, Harris JR (eds) Subcellular
Biochemistry. Plenum Press, New York, pp 41– 80
85. Vermeer C 1990 Gamma-carboxyglutamate-containing proteins
and the vitamin K-dependent carboxylase. Biochem J 266:625– 636
86. Brown DA 1992 Interaction between GPI-anchored proteins and
membrane lipids. Trends Cell Biol 2:43– 45
87. Udenfriend S, Kodukula K 1995 How glycosylphosphatidylinositol-anchored membrane proteins are made. Annu Rev Biochem
64:563–591
193
88. Moran P, Caras IW 1992 Proteins containing an uncleaved signal
for glycophosphatidylinositol membrane anchor attachment are
retained in a post-ER compartment. J Cell Biol 119:763–772
89. Haas IG, Wabl M 1983 Immunoglobulin heavy chain binding
protein. Nature 306:387–389
90. Bole DG, Hendershot LM, Kearney JF 1986 Posttranslational association of immunoglobulin heavy chain binding protein with
nascent heavy chains in nonsecreting hybridomas. J Cell Biol 102:
1558 –1566
91. Munro S, Pelham HRB 1986 An hsp70-like protein in the ER:
identity with the 78 kd glucose-regulated protein and immunoglobulin heavy chain binding protein. Cell 46:291–300
92. Macer DR, Koch GL 1988 Identification of a set of calcium-binding
proteins in reticuloplasm, the luminal content of the endoplasmic
reticulum. J Cell Sci 91:61–70
93. Rose MD, Misra LM, Vogel JP 1989 KAR2, a karyogamy gene, is
the yeast homolog of the mammalian BiP/GRP78 gene. Cell 57:
1211–1221
94. Normington K, Kohno K, Kozutsumi Y, Gething MJ, Sambrook
J 1989 S. cerevisiae encodes an essential protein homologous in
sequence and function to mammalian BiP. Cell 57:1223–1236
95. Flynn GC, Pohl J, Flocco MT, Rothman JE 1991 Peptide-binding
specificity of the molecular chaperone BiP. Nature 353:726 –730
96. Blond-Elguindi S, Cwirla SE, Dower WJ, Lipshutz RJ, Sprang SR,
Sambrook JF, Gething MJ 1993 Affinity panning of a library of
peptides displayed on bacteriophages reveals the binding specificity of BiP. Cell 75:717–728
97. Fourie AM, Sambrook JF, Gething MJ 1994 Common and divergent peptide binding specificities of hsp70 molecular chaperones.
J Biol Chem 269:30470 –30478
98. Kassenbrock CK, Garcia PD, Walter P, Kelly RB 1988 Heavychain binding protein recognizes aberrant polypeptides translocated in vitro. Nature 333:90 –93
99. Hurtley SM, Bole DG, Hoover-Litty H, Helenius A, Copeland CS
1989 Interactions of misfolded influenza virus hemagglutinin with
binding protein (BiP). J Cell Biol 108:2117–2126
100. Machamer CE, Doms RW, Bole DG, Helenius A, Rose JK 1990
Heavy chain binding protein recognizes incompletely disulfidebonded forms of vesicular stomatitis virus G protein. J Biol Chem
265:6879 – 6883
101. Dul JL, Burrone OR, Argon Y 1992 A conditional secretory mutant
in an Ig L chain is caused by replacement of tyrosine/phenylalanine 87 with histidine. J Immunol 149:1927–1933
102. Chessler SD, Byers PH 1993 BiP binds type I procollagen pro alpha
chains with mutations in the carboxyl-terminal propeptide synthesized by cells from patients with osteogenesis imperfecta. J Biol
Chem 268:18226 –18233
103. Puig A, Gilbert HF 1994 Anti-chaperone behavior of BiP during the
protein disulfide isomerase-catalyzed refolding of reduced denatured lysosome. J Biol Chem 269:25889 –25896
104. Schmitz A, Maintz M, Kehle T, Herzog V 1995 In vivo iodination
of a misfolded proinsulin reveals co-localized signals for BiP binding and for degradation in the ER. EMBO J 14:1091–1098
105. Beggah A, Mathews P, Beguin P, Geering K 1996 Degradation and
endoplasmic reticulum retention of unassembled a- and b-subunits
of Na, K-ATPase correlate with interaction with BiP. J Biol Chem
271:20895–20902
106. Leitzgen K, Knittler MR, Haas IG 1997 Assembly of immunoglobulin light chains as a prerequisite for secretion. A model for
oligomerization-dependent subunit folding. J Biol Chem 272:3117–
3123
107. Vogel JP, Misra LM, Rose MD 1990 Loss of BiP/GRP78 function
blocks translocation of secretory proteins in yeast. J Cell Biol 110:
1885–1895
108. Nguyen TH, Law DTS, Williams DB 1991 Binding protein BiP is
required for translocation of secretory proteins into the endoplasmic reticulum in Saccharomyces cerevisiae. Proc Natl Acad Sci USA
88:1565–1569
109. Sanders SL, Whitfield KM, Vogel JP, Rose MD, Schekman RW
1992 Sec61p and BiP directly facilitate polypeptide translocation
into the ER. Cell 69:353–365
110. Nicchitta CV, Blobel G 1993 Lumenal proteins of the mammalian
194
111.
112.
113.
114.
115.
116.
117.
118.
119.
120.
121.
122.
123.
124.
125.
126.
127.
128.
129.
130.
131.
KIM AND ARVAN
endoplasmic reticulum are required to complete protein translocation. Cell 73:989 –998
Lyman SK, Schekman R 1997 Binding of secretory precursor
polypeptides to a translocon subcomplex is regulated by BiP. Cell
88:85–96
Latterich M, Schekman R 1994 The karyogamy gene KAR2 and
novel proteins are required for ER-membrane fusion. Cell 78:87–98
Ng DT, Walter P 1996 ER membrane protein complex required for
nuclear fusion. J Cell Biol 132:499 –509
Panzner S, Dreier L, Hartmann E, Kostka S, Rapoport TA 1995
Posttranslational protein transport in yeast reconstituted with a
purified complex of Sec proteins and Kar2p. Cell 81:561–570
Dorner AJ, Bole DG, Kaufman RJ 1987 The relationship of Nlinked glycosylation and heavy chain-binding protein association
with the secretion of glycoproteins. J Cell Biol 105:2665–2674
Ng DT, Hiebert SW, Lamb RA 1990 Different roles of individual
N-linked oligosaccharide chains in folding, assembly, and transport of the simian virus 5 hemagglutinin-neuraminidase. Mol Cell
Biol 10:1989 –2001
Katsumi A, Senda T, Yamashita Y, Yamazaki T, Hamaguchi M,
Kojima T, Kobayashi S, Saito H 1996 Protein C Nagoya, an elongated mutant of protein C, is retained within the endoplasmic
reticulum and is associated with GRP78 and GRP94. Blood 87:
4164 – 4175
Flynn GC, Chappell TG, Rothman JE 1989 Peptide binding and
release by proteins implicated as catalysts of protein assembly.
Science 245:385–390
Dorner AJ, Kaufman RJ 1994 The levels of endoplasmic reticulum
proteins and ATP affect folding and secretion of selective proteins.
Biologicals 22:103–112
Nigam SK, Goldberg AL, Ho S, Rohde MF, Bush KT, Sherman
MY 1994 A set of endoplasmic reticulum proteins possessing properties of molecular chaperones includes Ca21 binding proteins and
members of the thioredoxin superfamily. J Biol Chem 269:1744 –
1749
Te Heesen S, Aebi M 1994 The genetic interaction of kar2 and wbp1
mutations. Distinct functions of binding protein BiP and N-linked
glycosylation in the processing pathway of secreted proteins in
Saccharomyces cerevisiae. Eur J Biochem 222:631– 637
Simons JF, Ferro-Novick S, Rose MD, Helenius A 1995 BiP/Kar2p
serves as a molecular chaperone during carboxypeptidase Y folding in yeast. J Cell Biol 130:41– 49
Hendershot L, Wei J, Gaut J, Melnick J, Aviel S, Argon Y 1996
Inhibition of immunoglobulin folding and secretion by dominant
negative BiP ATPase mutants. Proc Natl Acad Sci USA 93:5269 –
5274
Morris JA, Dorner AJ, Edwards CA, Hendershot LM, Kaufman RJ
1997 Immunoglobulin binding protein (BiP) function is required to
protect cells from endoplasmic reticulum stress but is not required
for the secretion of selective proteins. J Biol Chem 272:4327– 4334
Jamsa E, Vakula N, Arffman A, Kilpelainen I, Makarow M 1995
In vivo reactivation of heat-denatured protein in the endoplasmic
reticulum of yeast. EMBO J 14:6028 – 6033
Ng DT, Randall RE, Lamb RA 1989 Intracellular maturation and
transport of the SV5 type II glycoprotein hemagglutinin-neuraminidase: specific and transient association with GRP78-BiP in the
endoplasmic reticulum and extensive internalization from the cell
surface. J Cell Biol 109:3273–3289
Hendershot LM 1990 Immunoglobulin heavy chain and binding
protein complexes are dissociated in vivo by light chain addition.
J Cell Biol 111:829 – 837
Wiest DL, Burkhardt JK, Hester S, Hortsch M, Meyer DI, Argon
Y 1990 Membrane biogenesis during B cell differentiation: most
endoplasmic reticulum proteins are expressed coordinately. J Cell
Biol 110:1501–1511
Watowich SS, Morimoto RI, Lamb RA 1991 Flux of the paramyxovirus hemagglutinin-neuraminidase glycoprotein through the endoplasmic reticulum activates transcription of the GRP78-BiP gene.
J Virol 65:3590 –3597
Kim PS, Arvan P 1993 Hormonal regulation of thyroglobulin export from the endoplasmic reticulum of cultured thyrocytes. J Biol
Chem 268:4873– 4879
Craven RA, Egerton M, Stirling CJ 1996 A novel Hsp70 of the yeast
132.
133.
134.
135.
136.
137.
138.
139.
140.
141.
142.
143.
144.
145.
146.
147.
148.
149.
150.
151.
152.
Vol. 19, No. 2
ER lumen is required for the efficient translocation of a number of
protein precursors. EMBO J 15:2640 –2650
Dierks T, Volkmer J, Schlenstedt G, Jung C, Sandholzer U, Zachmann K, Schlotterhose P, Neifer K, Schmidt B, Zimmermann R
1996 A microsomal ATP-binding protein involved in efficient protein transport into the mammalian endoplasmic reticulum. EMBO
J 15:6931– 6942
Maki RG, Eddy RLJ, Byers M, Shows TB, Srivastava PK 1993
Mapping of the genes for human endoplasmic reticular heat shock
protein gp96/grp94. Somat. Cell Mol Genet 19:73– 81
Mazzarella RA, Green M 1987 ERp99, an abundant conserved
glycoprotein of the endoplasmic reticulum, is homologous to the
90-kDa heat shock protein (hsp90) and the 94-kDa glucose regulated protein (GRP94). J Biol Chem 262:8875– 8883
Van PN, Peter F, Soling HD 1989 Four intracisternal calciumbinding glycoproteins from rat liver microsomes with high affinity
for calcium. No indication for calsequestrin-like proteins in inositol
1,4,5-trisphosphate-sensitive calcium sequestering rat liver vesicles. J Biol Chem 264:17494 –17501
Bose S, Weikl T, Bugl H, Buchner J 1996 Chaperone function of
Hsp90-associated proteins. Science 274:1715–1717
Schumacher RJ, Hansen WJ, Freeman BC, Alnemri E, Litwack G,
Toft DO 1996 Cooperative action of Hsp70, Hsp90, and DnaJ proteins in protein renaturation. Biochemistry 35:14889 –14898
Melnick J, Aviel S, Argon Y 1992 The endoplasmic reticulum stress
protein GRP94, in addition to BiP, associates with unassembled
immunoglobulin chains. J Biol Chem 267:21303–21306
Muresan Z, Arvan P 1997 Thyroglobulin transport along the secretory pathway. Investigation of the role of molecular chaperone,
GRP94, in protein export from the endoplasmic reticulum. J Biol
Chem 272:26095–26102
Melnick J, Dul JL, Argon Y 1994 Sequential interaction of the
chaperones BiP and GRP94 with immunoglobulin chains in the
endoplasmic reticulum. Nature 370:373–375
Navarro D, Qadri I, Pereira L 1991 A mutation in the ectodomain
of herpes simplex virus 1 glycoprotein B causes defective processing and retention in the endoplasmic reticulum. Virology 184:253–
264
Schaiff WT, Hruska Jr KA, McCourt DW, Green M, Schwartz BD
1992 HLA-DR associates with specific stress proteins and is retained in the endoplasmic reticulum in invariant chain negative
cells. J Exp Med 176:657– 666
Kim PS, Kwon O-Y, Arvan P 1996 An endoplasmic reticulum
storage disease causing congenital goiter with hypothyroidism.
J Cell Biol 133:517–527
Ferreira LR, Norris K, Smith T, Hebert C, Sauk JJ 1994 Association
of Hsp47, Grp78, and Grp94 with procollagen supports the successive or coupled action of molecular chaperones. J Cell Biochem
56:518 –526
Li Z, Srivastava PK 1993 Tumor rejection antigen gp96/grp94 is an
ATPase: implications for protein folding and antigen presentation.
EMBO J 12:3143–3151
Wearsch PA, Nicchitta CV 1997 Interaction of endoplasmic reticulum chaperone GRP94 with peptide substrates is adenine nucleotide-independent. J Biol Chem 272:5152–5156
Prodromou C, Roe SM, O’Brien R, Ladbury JE, Piper PW, Pearl
LH 1997 Identification and structural characterization of the ATP/
ADP-binding site in the Hsp90 molecular chaperone. Cell 90:65–75
Melnick J, Argon Y 1995 Molecular chaperones and the biosynthesis of antigen receptors. Immunol Today 16:243–250
Bush KT, Hendrickson BA, Nigam SK 1994 Induction of the
FK506-binding protein, FKBP13, under conditions which misfold
proteins in the endoplasmic reticulum. Biochem J 303:705–708
Miles MF, Wilke N, Elliot M, Tanner W, Shah S 1994 Ethanolresponsive genes in neural cells include the 78-kilodalton glucoseregulated protein (GRP78) and 94-kilodalton glucose-regulated
protein (GRP94) molecular chaperones. Mol Pharmacol 46:873– 879
Bergeron JJM, Brenner MB, Thomas DY, Williams DB 1994 Calnexin: a membrane-bound chaperone of the endoplasmic reticulum. Trends Biochem Sci 19:124 –128
Wada I, Rindress D, Cameron PH, Ou W-J, Doherty II JJ, Louvard
D, Bell AW, Dignard D, Thomas DY, Bergeron JJM 1991 SSRa and
April, 1998
153.
154.
155.
156.
157.
158.
159.
160.
161.
162.
163.
164.
165.
166.
167.
168.
169.
170.
171.
172.
173.
ENDOCRINE ERSDs
associated calnexin are major calcium binding proteins of the endoplasmic reticulum membrane. J Biol Chem 266:19599 –19610
Ou W-J, Cameron PH, Thomas DY, Bergeron JJM 1993 Association of folding intermediates of glycoproteins with calnexin during
protein maturation. Nature 364:771–776
Jannatipour M, Rokeach LA 1995 The Schizosaccharomyces pombe
homologue of the chaperone calnexin is essential for viability. J Biol
Chem 270:4845– 4853
Parlati F, Dignard D, Bergeron JJM, Thomas DY 1995 The calnexin
homologue cnx11 in Schizosaccharomyces pombe, is an essential gene
which can be complemented by its soluble ER domain. EMBO J
14:3064 –3072
Hochstenbach F, David V, Watkins S, Brenner MB 1992 Endoplasmic reticulum resident proteins of 90 kilodaltons associates
with the T- and B-cell antigen receptors and major histocompatibility complex antigens during their assembly. Proc Natl Acad Sci
USA 89:4734 – 4738
Le A, Steiner JL, Ferrell GA, Shaker JC, Sifers RN 1994 Association between calnexin and a secretion-incompetent variant of human alpha1-antitrypsin. J Biol Chem 269:7514 –7519
Tatu U, Hammond C, Helenius A 1995 Folding and oligomerization of influenza hemagglutinin in the ER and the intermediate
compartment. EMBO J 147:1340 –1348
Hammond C, Braakman I, Helenius A 1994 Role of N-linked
oligosaccharide recognition, glucose trimming, and calnexin in glycoprotein folding and quality control. Proc Natl Acad Sci USA
91:913–917
Ora A, Helenius A 1995 Calnexin fails to associate with substrate
proteins in glucosidase-deficient cell lines. J Biol Chem 270:26060 –
26062
Hebert DN, Foellmer B, Helenius A 1995 Glucose trimming and
reglucosylation determine glycoprotein association with calnexin
in the endoplasmic reticulum. Cell 81:425– 433
Trombetta ES, Simons JF, Helenius A 1996 Endoplasmic reticulum glucosidase II is composed of a catalytic subunit, conserved
from yeast to mammals, and a tightly bound noncatalytic HDELcontaining subunit. J Biol Chem 271:27509 –27516
Sousa MC, Ferrero-Garcia MA, Parodi AJ 1992 Recognition of the
oligosaccharide and protein moieties of glycoproteins by the UDPGlc:glycoprotein glucosyltransferase. Biochemistry 31:97–105
Parker CG, Fessler LI, Nelson RE, Fessler JH 1995 Drosophila
UDP-glucose:glycoprotein glucosyltransferase: sequence and characterization of an enzyme that distinguishes between denatured
and native proteins. EMBO J 14:1294 –1303
Sousa M, Parodi AJ 1995 The molecular basis for the recognition
of misfolded glycoproteins by the UDP-Glc:glycoprotein glucosyltransferase. EMBO J 14:4196 – 4203
Ware FE, Vassilakos A, Peterson P, Jackson M, Lehrman M, Williams D 1995 The molecular chaperone calnexin binds
Glc1Man9GlcNAc2 oligosaccharide as an initial step in recognizing
unfolded glycoproteins. J Biol Chem 270:4697– 4704
Zapun A, Petrescu SM, Rudd PM, Dwek RA, Thomas DY,
Bergeron JJM 1997 Conformation-independent binding of monoglucosylated ribonuclease B to calnexin. Cell 88:29 –38
Hebert DN, Foellmer B, Helenius A 1996 Calnexin and calreticulin
promote folding, delay oligomerization and suppress degradation
of influenza hemagglutinin in microsomes. EMBO J 15:2961–2968
Wada I, Kai M, Imai S, Sakane F, Kanoh H 1997 Promotion of
transferrin folding by cyclic interactions with calnexin and calreticulin. EMBO J 16:101–113
Margolese L, Waneck GL, Suzuki CK, Degen E, Flavell RA, Williams DB 1993 Identification of the region on the class I histocompatibility molecule that interacts with the molecular chaperone, p88
(calnexin, IP90). J Biol Chem 268:17959 –17966
Capps GG, Zuniga MC 1994 Class I histocompatibility molecule
association with phosphorylated calnexin. Implications for rates of
intracellular transport. J Biol Chem 269:11634 –11639
Loo TW, Clarke DM 1995 P-glycoprotein. Associations between
domains and between domains and molecular chaperones. J Biol
Chem 270:21839 –21844
Arunachalam B, Cresswell P 1995 Molecular requirements for the
interaction of class II major histocompatibility complex molecules
and invariant chain with calnexin. J Biol Chem 270:2784 –2790
195
174. Zhang Q, Tector M, Salter RD 1995 Calnexin recognizes carbohydrate and protein determinants of class I major histocompatibility complex molecules. J Biol Chem 270:3944 –3948
175. van Leeuwen JE, Kearse KP 1996 Calnexin associates exclusively
with individual CD3 delta and T cell antigen receptor (TCR) alpha
proteins containing incompletely trimmed glycans that are not
assembled into multisubunit TCR complexes. J Biol Chem 271:
9660 –9665
176. Rajagopalan S, Xu Y, Brenner MB 1994 Retention of unassembled
components of integral membrane proteins by calnexin. Science
263:387–390
177. Carreno BM, Schreiber KL, McKean DJ, Stroynowski I, Hansen
TH 1995 Aglycosylated and phosphatidylinositol-anchored MHC
class I molecules are associated with calnexin. Evidence implicating
the class I-connecting peptide segment in calnexin association.
J Immunol 154:5173–5180
178. Cannon KS, Hebert DN, Helenius A 1996 Glycan-dependent and
-independent association of vesicular stomatitis virus G protein
with calnexin. J Biol Chem 271:14280 –14284
179. Marks MS, Germain RN, Bonifacino JS 1995 Transient aggregation of major histocompatibility complex class II chains during
assembly in normal spleen cells. J Biol Chem 270:10475–10481
180. Williams DB 1995 The Merck Frosst Award Lecture 1994/La conference Merck Frosst 1994. Calnexin: a molecular chaperone with
a taste for carbohydrate. Biochem Cell Biol 73:123–132
181. Helenius A, Trombetta ES, Hebert DN, Simons JF 1997 Calnexin,
calreticulin and the folding of glycoproteins. Trends Cell Biol 7:193–
200
182. Chen W, Helenius J, Braakman I, Helenius A 1995 Cotranslational
folding and calnexin binding during glycoprotein synthesis. Proc
Natl Acad Sci USA 92:6229 – 6233
183. Vassilakos A, Cohen-Doyle MF, Peterson PA, Jackson MR, Williams DB 1996 The molecular chaperone calnexin facilitates folding
and assembly of class I histocompatibility molecules. EMBO J 15:
1495–1506
184. Van Leeuwen JE, Kearse KP 1997 Reglucosylation of N-linked
glycans is critical for calnexin assembly with T cell receptor (TCR)
alpha proteins but not TCRbeta proteins. J Biol Chem 272:4179 –
4186
185. Krause KH, Michalak M 1997 Calreticulin. Cell 88:439 – 443
186. Wada I, Imai S, Kai M, Sakane F, Kanoh H 1995 Chaperone
function of calreticulin when expressed in the endoplasmic reticulum as the membrane-anchored and soluble forms. J Biol Chem
270:20298 –20304
187. Peterson JR, Ora A, Van PN, Helenius A 1995 Transient, lectin-like
association of calreticulin with folding intermediates of cellular and
viral glycoproteins. Mol Biol Cell 6:1173–1184
188. van Leeuwen JE, Kearse KP 1996 Deglucosylation of N-linked
glycans is an important step in the dissociation of calreticulin-class
I-TAP complexes. Proc Natl Acad Sci USA 93:13997–14001
189. McCauliffe DP, Yang YS, Wilson J, Sontheimer RD, Capra JD
1992 The 59-flanking region of the human calreticulin gene shares
homology with the human GRP78, GRP94, and protein disulfide
isomerase promoters. J Biol Chem 267:2557–2562
190. Camacho P, Lechleiter JD 1995 Calreticulin inhibits repetitive intracellular Ca21 waves. Cell 82:765–771
191. Bastianutto C, Clementi E, Codazzi F, Podini P, De Giorgi F,
Rizzuto R, Meldolesi J, Pozzan T 1995 Overexpression of calreticulin increases the Ca21 capacity of rapidly exchanging Ca21
stores and reveals aspects of their lumenal microenvironment and
function. J Cell Biol 130:847– 855
192. Burns K, Duggan B, Atkinson EA, Famulski KS, Nemer M,
Bleackley RC, Michalak M 1994 Modulation of gene expression by
calreticulin binding to the glucocorticoid receptor. Nature 367:476 –
480
193. Ikawa M, Wada I, Kominami K, Watanabe D, Toshimori K, Nishimune Y, Okabe M 1997 The putative chaperone calmegin is required for sperm fertility. Nature 387:607– 611
194. Edman JC, Ellis L, Blacher RW, Roth RA, Rutter WJ 1985 Sequence
of protein disulfide isomerase and implications of its relationship
to thioredoxin. Nature 317:267–270
195. Weissman JS, Kim PS 1993 Efficient catalysis of disulphide bond
196
196.
197.
198.
199.
200.
201.
202.
203.
204.
205.
206.
207.
208.
209.
210.
211.
212.
213.
214.
215.
216.
217.
KIM AND ARVAN
rearrangements by protein disulfide isomerase. Nature 365:185–
188
Dorner AJ, Wasley LC, Raney P, Haugejorden S, Green M, Kaufman RJ 1990 The stress response in Chinese hamster ovary cells.
Regulation of ERp72 and protein disulfide isomerase expression
and secretion. J Biol Chem 265:22029 –22034
Freedman RB 1989 Protein disulfide isomerase: multiple roles in
the modification of nascent secretory proteins. Cell 57:1069 –1072
Marcus N, Shaffer D, Farrar P, Green M 1996 Tissue distribution
of three members of the murine protein disulfide isomerase (PDI)
family. Biochim Biophys Acta 1309:253–260
Iida KI, Miyaishi O, Iwata Y, Kozaki KI, Matsuyama M, Saga S
1996 Distinct distribution of protein disulfide isomerase family
proteins in rat tissues. J Histochem Cytochem 44:751–759
Tsibris JCM, Hunt LT, Ballejo G, Barker WC, Toney LJ, Spellacy
WN 1989 Selective inhibition of protein disulfide isomerase by
estrogens. J Biol Chem 264:13967–13970
Guthapfel R, Gueguen P, Quemeneur E 1996 ATP binding and
hydrolysis by the multifunctional protein disulfide isomerase.
J Biol Chem 271:2663–2666
Robinson AS, Hines V, Wittrup KD 1994 Protein disulfide isomerase overexpression increases secretion of foreign proteins in
Saccharomyces cerevisiae. Biotechnology 12:381–384
Farquhar R, Honey N, Murant SJ, Bossier P, Schultz L, Montgomery D, Ellis RW, Freedman RB, Tuite MF 1991 Protein disulfide isomerase is essential for viability in Saccharomyces cerevisiae.
Gene 108:81– 89
Noiva R, Freedman RB, Lennarz WJ 1993 Peptide binding to
protein disulfide isomerase occurs at a site distinct from the active
sites. J Biol Chem 268:19210 –19217
Cai H, Wang CC, Tsou CL 1994 Chaperone-like activity of protein
disulfide isomerase in the refolding of a protein with no disulfide
bonds. J Biol Chem 269:24550 –24552
LaMantia ML, Lennarz WJ 1993 The essential function of yeast
protein disulfide isomerase does not reside in its isomerase activity.
Cell 74:899 –908
Hayano T, Hirose M, Kikuchi M 1995 Protein disulfide isomerase
mutant lacking its isomerase activity accelerates protein folding in
the cell. FEBS Lett 377:505–511
Rupp K, Birnbach U, Lundstrom J, Van PN, Soling HD 1994
Effects of CaBP2, the rat analog of ERp72, and of CaBP1 on the
refolding of denatured reduced proteins. Comparison with protein
disulfide isomerase. J Biol Chem 269:2501–2507
Chivers PT, Laboissiere MCA, Raines RT 1996 The CXXC motif:
imperatives for the formation of native disulfide bonds in the cell.
EMBO J 15:2659 –2667
Laboissiere MC, Sturley SL, Raines RT 1995 The essential function
of protein-disulfide isomerase is to unscramble non-native disulfide bonds. J Biol Chem 270:28006 –28009
John DC, Grant ME, Bulleid NJ 1993 Cell-free synthesis and assembly of prolyl 4-hydroxylase: the role of the beta-subunit (PDI)
in preventing misfolding and aggregation of the alpha-subunit.
EMBO J 12:1587–1595
Wetterau JR, Combs KA, Spinner SN, Joiner BJ 1990 Protein
disulfide isomerase is a component of the microsomal triglyceride
transfer protein complex. J Biol Chem 265:9801–9807
Freskgard P-O, Bergenhem N, Jonsson B-H, Svensson M, Carlsson U 1992 Isomerase and chaperone activity of prolyl isomerase
in the folding of carbonic anhydrase. Science 258:466 – 468
Partaledis JA, Berlin V 1993 The FKB2 gene of Saccharomyces cerevisiae, encoding the immunosuppressant-binding protein FKBP13, is regulated in response to accumulation of unfolded proteins
in the endoplasmic reticulum. Proc Natl Acad Sci USA 90:5450 –
5454
Mazzarella RA, Srinivasan M, Haugejorden SM, Green M 1990
ERp72, an abundant luminal endoplasmic reticulum protein, contains three copies of the active site sequences of protein disulfide
isomerase. J Biol Chem 265:1094 –1101
Gunther R, Srinivasan M, Haugejorden S, Green M, Ehbrecht IM,
Kuntzel H 1993 Functional replacement of the Saccharomyces cerevisiae Trg1/Pdi1 protein by members of the mammalian protein
disulfide isomerase family. J Biol Chem 268:7728 –7732
Lundstrom-Ljung J, Birnbach U, Rupp K, Soling HD, Holmgren
218.
219.
220.
221.
222.
223.
224.
225.
226.
227.
228.
229.
230.
231.
232.
233.
234.
235.
236.
237.
238.
Vol. 19, No. 2
A 1995 Two resident ER-proteins, CaBP1 and CaBP2, with thioredoxin domains, are substrates for thioredoxin reductase: comparison with protein disulfide isomerase. FEBS Lett 357:305–308
Urade R, Takenaka Y, Kito M 1993 Protein degradation by ERp72
from rat and mouse liver endoplasmic reticulum. J Biol Chem
268:22004 –22009
Kuznetsov G, Chen LB, Nigam SK 1994 Several endoplasmic
reticulum stress proteins, including ERp72, interact with thyroglobulin during its maturation. J Biol Chem 269:22990 –22995
Feng W, Matzuk MM, Mountjoy K, Bedows E, Ruddon RW,
Boime I 1995 The asparagine-linked oligosaccharides of the human
chorionic gonadotropin beta subunit facilitate correct disulfide
bond pairing. J Biol Chem 270:11851–11859
Srinivasan M, Lenny N, Green M 1993 Identification of genomic
sequences that mediate the induction of the endoplasmic reticulum
stress protein, ERp72, by protein traffic. DNA Cell Biol 12:807– 822
Urade R, Kito M 1992 Inhibition by acidic phospholipids of protein
degradation by ER-60 protease, a novel cysteine protease, of the
endoplasmic reticulum. FEBS Lett 312:83– 86
Urade R, Nasu M, Moriyama T, Wada K, Kito M 1992 Protein
degradation by the phosphoinositide-specific phospholipase C-a
family from rat liver endoplasmic reticulum. J Biol Chem 267:
15152–15159
Otsu M, Urade R, Kito M, Omura F, Kikuchi M 1995 A possible
role of ER-60 protease in the degradation of misfolded proteins in
the endoplasmic reticulum. J Biol Chem 270:14958 –14961
Oliver JD, van der Wal FJ, Bulleid NJ, High S 1997 Interaction of
the thiol-dependent reductase ERp57 with nascent glycoproteins.
Science 275:86 – 88
Elliott JG, Oliver JD, High S 1997 The thiol-dependent reductase
ERp57 interacts specifically with N-glycosylated integral membrane proteins. J Biol Chem 272:13849 –13855
Nakai A, Satoh M, Hirayoshi K, Nagata K 1992 Involvment of the
stress protein HSP47 in procollagen processing in the endoplasmic
reticulum. J Cell Biol 117:903–914
Hu G, Gura T, Sabsay B, Sauk J, Dixit SN, Veis A 1995 Endoplasmic reticulum protein Hsp47 binds specifically to the Nterminal globular domain of the amino-propeptide of the procollagen I alpha 1 (I)-chain. J Cell Biochem 59:350 –367
Smith T, Ferreira LR, Hebert C, Norris K, Sauk JJ 1995 Hsp47 and
cyclophilin B traverse the endoplasmic reticulum with procollagen
into pre-Golgi intermediate vesicles. A role for Hsp47 and cyclophilin B in the export of procollagen from the endoplasmic reticulum. J Biol Chem 270:18323–18328
Satoh M, Hirayoshi K, Yokota S, Hosokawa N, Nagata K 1996
Intracellular interaction of collagen-specific stress protein HSP47
with newly synthesized procollagen. J Cell Biol 133:469 – 483
Ferreira LR, Norris K, Smith T, Hebert C, Sauk JJ 1996 Hsp47 and
other ER-resident molecular chaperones form heterocomplexes
with each other and with collagen type IV chains. Connect Tissue
Res 33:265–273
Nagata K 1996 Hsp47: a collagen-specific molecular chaperone.
Trends Biochem Sci 21:22–26
Willnow TE, Armstrong SA, Hammer RE, Herz J 1995 Functional
expression of low density lipoprotein receptor-related protein is
controlled by receptor-associated protein in vivo. Proc Natl Acad Sci
USA 92:4537– 4541
Weissman JS, Kim PS 1992 The pro region of BPTI facilitates
folding. Cell 71:841– 851
Mercure C, Thibault G, Lussier-Cacan S, Davignon J, Schiffrin
EL, Reudelhuber TL 1995 Molecular analysis of human prorenin
prosegment variants in vitro and in vivo. J Biol Chem 270:16355–
16359
Hoshina H, Boime I 1982 Combination of rat lutropin subunits
occurs early in the secretory pathway. Proc Natl Acad Sci USA
79:7649 –7653
Ruddon RW, Krzesichi RF, Norton SD, Beebe JS, Peters BP 1987
Detection of a glycosylated, incompletely folded form of human
chorionic gonadotropin b subunit that is a precursor of hormone
assembly in trophoblastic cells. J Biol Chem 262:12533–12540
Bedows E, Huth JR, Ruddon RW 1992 Kinetics of folding and
assembly of the human chorionic gonadotropin b subunit in transfected Chinese hamster ovary cells. J Biol Chem 267:8880 – 8886
April, 1998
ENDOCRINE ERSDs
239. Huth JR, Perini F, Lockridge O, Bedows E, Ruddon RW 1993
Protein folding and assembly in vitro parallel intracellular folding
and assembly. Catalysis of folding and assembly of the human
chorionic gonadotropin alpha beta dimer by protein disulfide
isomerase. J Biol Chem 268:16472–16482
240. Bielinska M, Boime I 1992 Site-directed mutagenesis defines a
domain in the gonadotropin alpha-subunit required for assembly
with the chorionic gonadotropin beta-subunit. Mol Endocrinol
6:261–271
241. Furuhashi M, Suzuki S, Suganuma N 1996 Disulfide bonds 7–31
and 59 – 87 of the alpha-subunit play a different role in assembly of
human chorionic gonadotropin and lutropin. Endocrinology 137:
4196 – 4200
242. Muyan M, Furuhashi M, Sugahara T, Boime I 1996 The carboxyterminal region of the beta-subunits of luteinizing hormone and
chorionic gonadotropin differentially influence secretion and assembly of the heterodimers. Mol Endocrinol 10:1678 –1687
243. Grossmann M, Szkudlinski MW, Dias JA, Xia H, Wong R, Puett
D, Weintraub BD 1996 Site-directed mutagenesis of amino acids
33– 44 of the common alpha-subunit reveals different structural
requirements for heterodimer expression among the glycoprotein
hormones and suggests that cyclic adenosine 39,59-monophosphate
production and growth promotion are potentially dissociable functions of human thyrotropin. Mol Endocrinol 10:769 –779
244. Corless CL, Matzuk MM, Ramabhadran TV, Krichevsky A,
Boime I 1987 Gonadotropin beta subunits determine the rate of
assembly and the oligosaccharide processing of hormone dimer
in transfected cells. J Cell Biol 104:1173–1181
245. Chen F, Wang Y, Puett D 1992 The carboxy-terminal region of the
glycoprotein hormone alpha-subunit: contributions to receptor
binding and signaling in human chorionic gonadotropin. Mol Endocrinol 6:914 –919
246. Matzuk MM, Kornmeier CM, Whitfield GK, Kourides IA, Boime
I 1988 The glycoprotein alpha-subunit is critical for secretion and
stability of the human thyrotropin beta-subunit. Mol Endocrinol
2:95–100
247. Williams SE, Ashcom JD, Argraves WS, Strickland DK 1992 A
novel mechanism for controlling the activity of alpha 2-macroglobulin receptor/low density lipoprotein receptor-related protein.
Multiple regulatory sites for 39-kDa receptor-associated protein.
J Biol Chem 267:9035–9040
248. Bu G, Geuze HJ, Strous GJ, Schwartz AL 1995 39 kDa receptorassociated protein is an ER resident protein and molecular chaperone for LDL receptor-related protein. EMBO J 14:2269 –2280
249. Farquhar MG, Saito A, Kerjaschki D, Orlando RA 1995 The Heymann nephritis antigenic complex: megalin (gp330) and RAP. J Am
Soc Nephrol 6:35– 47
250. Bu G, Rennke S 1996 Receptor-associated protein is a folding
chaperone for low density lipoprotein receptor-related protein.
J Biol Chem 271:22218 –22224
251. Willnow TE, Rohlmann A, Horton J, Otani H, Braun JR, Hammer
RE, Herz J 1996 RAP, a specialized chaperone, prevents ligandinduced ER retention and degradation of LDL receptor-related
endocytic receptors. EMBO J 15:2632–2639
252. Martens GJM, Braks JAM, Eib DW, Zhou Y, Lindberg I 1994 The
neuroendocrine polypeptide 7B2 is an endogenous inhibitor of
prohormone convertase PC2. Proc Natl Acad Sci USA 91:5784 –5787
253. van Horssen AM, van den Hurk WH, Bailyes EM, Hutton JC,
Martens GJ, Lindberg I 1995 Identification of the region within the
neuroendocrine polypeptide 7B2 responsible for the inhibition of
prohormone convertase PC2. J Biol Chem 270:14292–14296
254. Braks JA, Martens GJ 1995 The neuroendocrine chaperone 7B2 can
enhance in vitro POMC cleavage by prohormone convertase PC2.
FEBS Lett 371:154 –158
255. Zhu X, Lamango NS, Lindberg I 1996 Involvement of a polyproline
helix-like structure in the interaction of 7B2 with prohormone convertase 2. J Biol Chem 271:23582–23587
256. Braks JAM, Martens GJM 1994 7B2 is a neuroendocrine chaperone
that transiently interacts with prohormone convertase PC2 in the
secretory pathway. Cell 78:263–273
257. Benjannet S, Savaria D, Chretien M, Seidah NG 1995 7B2 is a
specific intracellular binding protein of the prohormone convertase
PC2. J Neurochem 64:2303–2311
197
258. Braks JA, Van Horssen AM, Martens GJ 1996 Dissociation of the
complex between the neuroendocrine chaperone 7B2 and prohormone convertase PC2 is not associated with proPC2 maturation.
Eur J Biochem 238:505–510
259. Zhu X, Lindberg I 1995 7B2 facilitates the maturation of proPC2 in
neuroendocrine cells and is required for the expression of enzymatic activity. J Cell Biol 129:1641–1650
260. Chaudhuri B, Stephan C, Huijbregts RP, Martens GJ 1995 The
neuroendocrine protein 7B2 acts as a molecular chaperone in the
in vitro folding of human insulin-like growth factor-1 secreted from
yeast. Biochem Biophys Res Commun 211:417– 425
261. Schekman R, Orci L 1996 Coat proteins and vesicle budding.
Science 271:1526 –1533
262. Pelham HRB 1995 Sorting and retrieval between the endoplasmic
reticulum and Golgi apparatus. Curr Opin Cell Biol 7:530 –535
263. Cole NB, Sciaky N, Marotta A, Song J, Lippincott-Schwartz J 1996
Golgi dispersal during microtubule disruption: regeneration of
Golgi stacks at peripheral endoplasmic reticulum exit sites. Mol
Biol Cell 7:631– 650
264. Thomas PJ, Qu BH, Pedersen PL 1995 Defective protein folding as
a basis of human disease. Trends Biochem Sci 20:456 – 459
265. Booth C, Koch GLE 1989 Perturbation of cellular calcium induces
secretion of luminal ER proteins. Cell 59:729 –737
266. Letourneur F, Gaynor EC, Hennecke S, Demolliere C, Duden R,
Emr SD, Riezman H, Cosson P 1994 Coatomer is essential for
retrieval of dilysine-tagged proteins to the endoplasmic reticulum.
Cell 79:1199 –1207
267. Lewis MJ, Pelham HR 1996 SNARE-mediated retrograde traffic
from the Golgi complex to the endoplasmic reticulum. Cell 85:205–
215
268. Semenza JC, Hardwick KG, Dean N, Pelham HR 1990 ERD2, a
yeast gene required for the receptor-mediated retrieval of luminal
ER proteins from the secretory pathway. Cell 61:1349 –1357
269. Teasdale RD, Jackson MR 1996 Signal-mediated sorting of membrane proteins between the endoplasmic reticulum and the Golgi
apparatus. Annu Rev Cell Dev Biol 12:27–54
270. Nilsson T, Warren G 1994 Retention and retrieval in the endoplasmic reticulum and the Golgi apparatus. Curr Opin Cell Biol
6:517–521
271. Ceriotti A, Colman A 1988 Binding to membrane proteins within
the endoplasmic reticulum cannot explain the retention of the glucose-regulated protein GRP78 in Xenopus oocytes. EMBO J 7:633–
638
272. Sonnichsen B, Fullekrug J, Nguyen Van P, Diekmann W, Robinson DG, Mieskes G 1994 Retention and retrieval: both mechanisms cooperate to maintain calreticulin in the endoplasmic reticulum. J Cell Sci 107:2705–2717
273. Elrod-Erickson MJ, Kaiser CA 1996 Genes that control the fidelity
of endoplasmic reticulum to Golgi transport identified as suppressors of vesicle budding mutations. Mol Biol Cell 7:1043–1058
274. Lodish HF, Kong N, Snider M, Strous GJAM 1983 Hepatoma
secretory proteins migrate from rough endoplasmic reticulum to
Golgi at characteristic rates. Nature 304:80 – 83
275. Scheele G, Tartakoff A 1985 Exit of nonglycosylated secretory
proteins from the rough endoplasmic reticulum is asynchronous in
the exocrine pancreas. J Biol Chem 260:926 –931
276. Bannykh SI, Balch WE 1997 Membrane dynamics at the endoplasmic reticulum-Golgi interface. J Cell Biol 138:1– 4
277. Rothman JE, Wieland FT 1996 Protein sorting by transport vesicles. Science 272:227–234
278. Balch WE, McCaffery JM, Plutner H, Farquhar MG 1994 Vesicular
stomatitis virus glycoprotein is sorted and concentrated during
export from the endoplasmic reticulum. Cell 76:841– 852
279. Aridor M, Bannykh SI, Rowe T, Balch WE 1995 Sequential coupling between COPII and COPI vesicle coats in endoplasmic reticulum to Golgi transport. J Cell Biol 131:875– 893
280. Schmid SL 1997 Clathrin-coated vesicle formation and protein
sorting: an integrated process. Annu Rev Biochem 66:511–548
281. Pearse BMF, Robinson MS 1990 Clathrin, adaptors, and sorting.
Annu Rev Cell Biol 6:151–171
282. Ohno H, Stewart J, Fournier MC, Bosshart H, Rhee I, Miyatake
S, Saito T, Gallusser A, Kirchhausen T, Bonifacino JS 1995 In-
198
283.
284.
285.
286.
287.
288.
289.
290.
291.
292.
293.
294.
295.
296.
297.
298.
299.
300.
301.
302.
KIM AND ARVAN
teraction of tyrosine-based sorting signals with clathrin-associated
proteins. Science 269:1872–1875
Mauxion F, Le Borgne R, Munier-Lehmann H, Hoflack B 1996 A
casein kinase II phosphorylation site in the cytoplasmic domain of
the cation-dependent mannose 6-phosphate receptor determines
the high affinity interaction of the AP-1 Golgi assembly proteins
with membranes. J Biol Chem 271:2171–2178
Tisdale EJ, Plutner H, Matteson J, Balch WE 1997 p53/58 binds
COPI and is required for selective transport through the early
secretory pathway. J Cell Biol 137:581–593
Bednarek SY, Ravazzola M, Hosobuchi M, Amherdt M, Perrelet
A, Schekman R, Orci L 1995 COPI- and COPII-coated vesicles bud
directly from the endoplasmic reticulum in yeast. Cell 83:1183–1196
Kuehn MJ, Schekman R, Ljungdahl PO 1996 Amino acid permeases require COPII components and the ER resident membrane
protein Shr3p for packaging into transport vesicles in vitro. J Cell
Biol 135:585–595
Rowe T, Aridor M, McCaffery JM, Plutner H, Nuoffer C, Balch
WE 1996 COPII vesicles derived from mammalian endoplasmic
reticulum microsomes recruit COPI. J Cell Biol 135:895–911
Fiedler K, Veit M, Stamnes MA, Rothman JE 1996 Bimodal interaction of coatomer with the p24 family of putative cargo receptors. Science 273:1396 –1399
Arar C, Carpentier V, Le Caer J-P, Monsigny M, Legrand A, Roche
A-C 1995 ERGIC-53, a membrane protein of the endoplasmic reticulum-Golgi intermediate compartment, is identical to MR60, an
intracellular mannose-specific lectin of myelomonocytic cells. J Biol
Chem 270:3551–3553
Itin C, Schindler R, Hauri HP 1995 Targeting of protein ERGIC-53
to the ER/ERGIC/cis-Golgi recycling pathway. J Cell Biol 131:
57– 67
Schimmoller F, Singer-Kruger B, Schroder S, Kruger U, Barlowe
C, Riezman H 1995 The absence of Emp24p, a component of ERderived COPII-coated vesicles, causes a defect in transport of selected proteins to the Golgi. EMBO J 14:1329 –1339
Stamnes MA, Craighead MW, Hoe MH, Lampen N, Geromanos
S, Tempst P, Rothman JE 1995 An integral membrane component
of coatomer-coated transport vesicles defines a family of proteins
involved in budding. Proc Natl Acad Sci USA 92:8011– 8015
Yeung T, Barlowe C, Schekman R 1995 Uncoupled packaging of
targeting and cargo molecules during transport vesicle budding
from the endoplasmic reticulum. J Biol Chem 270:30567–30570
Itin C, Roche AC, Monsigny M, Hauri HP 1996 ERGIC-53 is a
functional mannose-selective and calcium-dependent human homologue of leguminous lectins. Mol Biol Cell 7:483– 493
Pfeffer SR, Rothman JE 1987 Biosynthetic protein transport and
sorting by the endoplasmic reticulum and Golgi. Annu Rev Biochem 56:829 – 852
Rodan AR, Simons JF, Trombetta ES, Helenius A 1996 N-linked
oligosaccharides are necessary and sufficient for association of
glycosylated forms of bovine RNase with calnexin and calreticulin.
EMBO J 15:6921– 6930
Lodish HF, Kong N 1984 Glucose removal from N-linked oligosaccharides is required for efficient maturation of certain secretory
glycoproteins from the rough endoplasmic reticulum to the Golgi
complex. J Cell Biol 98:1720 –1729
Hammond C, Helenius A 1994 Folding of VSV G protein: sequential interactions with BiP and calnexin. Science 266:456 – 458
Harmsen MM, Bruyne MI, Raue HA, Maat J 1996 Overexpression
of binding protein and disruption of the PMR1 gene synergistically
stimulate secretion of bovine prochymosin but not plant thaumatin
in yeast. Appl Microbiol Biotechnol 46:365–370
Hendershot L, Bole D, Kohler G, Kearney JF 1987 Assembly and
secretion of heavy chains that do not associate posttranslationally
with immunoglobulin heavy chain-binding protein. J Cell Biol 104:
761–767
Pollok BA, Anker R, Eldridge P, Hendershot L, Levitt D 1987
Molecular basis of the cell-surface expression of immunoglobulin
mu chain without light chain in human B lymphocytes. Proc Natl
Acad Sci USA 84:9199 –9203
Suzuki CK, Bonifacino JS, Lin AY, Davis MM, Klausner RD 1991
Regulating the retention of T-cell receptor a chain variants within
Vol. 19, No. 2
the endoplasmic reticulum: Ca21-dependent association with BiP.
J Cell Biol 114:189 –205
303. de Silva AM, Balch WE, Helenius A 1990 Quality control in the
endoplasmic reticulum: folding and misfolding of vesicular stomatitis virus G protein in cells and in vitro. J Cell Biol 111:857– 866
304. Bonnerot C, Marks MS, Cosson P, Robertson EJ, Bikoff EK, Germain RN, Bonifacino JS 1994 Association with BiP and aggregation of class II MHC molecules synthesized in the absence of invariant chain. EMBO J 13:934 –944
305. David V, Hochstenbach F, Rajagopalan S, Brenner MB 1993 Interaction with newly synthesized and retained proteins in the endoplasmic reticulum suggests a chaperone function for human
integral membrane protein IP90 (calnexin). J Biol Chem 268:9585–
9592
306. Jackson MR, Cohen-Doyle MF, Peterson PA, Williams DB 1994
Regulation of MHC class I transport by the molecular chaperone,
calnexin (p88, IP90). Science 263:384 –387
307. Anderson KS, Cresswell P 1994 A role for calnexin (IP90) in the
assembly of class II MHC molecules. EMBO J 13:675– 682
308. Parlati F, Dominguez M, Bergeron JJM, Thomas DY 1995 Saccharomyces cerevisiae CNE1 encodes an endoplasmic reticulum (ER)
membrane protein with sequence similarity to calnexin and calreticulin and functions as a constituent of the ER quality control
apparatus. J Biol Chem 270:244 –253
309. Dorner AJ, Wasley LC, Kaufman RJ 1992 Overexpression of
GRP78 mitigates induction of glucose regulated proteins and
blocks secretion of selective proteins in Chinese hamster ovary
cells. EMBO J 11:1563–1571
309a.Muresan Z, Arvan P 1998 Enhanced binding of the molecular
chaperone, BiP, slows thyroglobulin export from the endoplasmic
reticulum. Mol Endocrinol, in press
310. Dorner AJ, Krane MG, Kaufman RJ 1988 Reduction of endogenous GRP78 levels improves secretion of a heterologous protein in
CHO cells. Mol Cell Biol 8:4063– 4070
311. Ellis RJ, Hartl FU 1996 Protein folding in the cell: competing
models of chaperonin function. FASEB J 10:20 –26
312. Sitia R, Neuberger M, Alberini C, Bet P, Fra A, Valetti C, Williams
G, Milstein C 1990 Developmental regulation of IgM secretion: the
role of the carboxy-terminal cysteine. Cell 60:781–790
313. Alberini CM, Bet P, Milstein C, Sitia R 1990 Secretion of immunoglobulin M assembly intermediates in the presence of reducing
agents. Nature 347:485– 487
314. Deleted in proof
315. Ward CL, Kopito RR 1994 Intracellular turnover of cystic fibrosis
transmembrane conductance regulator. J Biol Chem 269:25710 –
25718
316. Klausner RD, Sitia R 1990 Protein degradation in the endoplasmic
reticulum. Cell 62:611– 614
317. Dunn WAJ 1990 Studies on the mechanisms of autophagy: formation of the autophagic vacuole. J Cell Biol 110:1923–1933
318. Tooze J, Hollinshead M, Ludwig T, Howell K, Hoflack B, Kern
H 1990 In exocrine pancreas, the basolateral endocytic pathway
converges with the autophagic pathway immediately after the
early endosome. J Cell Biol 111:329 –345
319. Lawrence BP, Brown WJ 1992 Autophagic vacuoles rapidly fuse
with pre-existing lysosomes in cultured hepatocytes. J Cell Sci
102:515–526
320. Field MC, Moran P, Li W, Keller GA, Caras IW 1994 Retention and
degradation of proteins containing an uncleaved glycosylphosphatidylinositol signal. J Biol Chem 269:10830 –10837
321. Wainwright LJ, Field MC 1997 Quality control of glycosylphosphatidylinositol anchor attachment in mammalian cells: a biochemical study. Biochem J 321:655– 664
322. McCracken AA, Brodsky JL 1996 Assembly of ER-associated protein degradation in vitro: dependence on cytosol, calnexin, and
ATP. J Cell Biol 132:291–298
323. Chen C, Bonifacino JS, Yuan LC, Klausner RD 1988 Selective
degradation of T cell antigen receptor chains retained in a pre-Golgi
compartment. J Cell Biol 107:2149 –2161
324. Amara JF, Lederkermer G, Lodish HF 1989 Intracellular degradation of unassembled asialoglycoprotein receptor subunits: a preGolgi, nonlysosomal endoproteolytic cleavage. J Cell Biol 109:3315–
3324
April, 1998
ENDOCRINE ERSDs
325. Gardner AM, Aviel S, Argon Y 1993 Rapid degradation of an
unassembled immunoglobulin light chain is mediated by a serine
protease and occurs in a pre-Golgi compartment. J Biol Chem
268:25940 –25947
326. Fra AM, Fagioli C, Finazzi D, Sitia R, Alberini CM 1993 Quality
control of ER synthesized proteins: an exposed thiol group as a
three-way switch mediating assembly, retention and degradation.
EMBO J 12:4755– 4761
327. Young J, Kane LP, Exley M, Wileman T 1993 Regulation of selective protein degradation in the endoplasmic reticulum by redox
potential. J Biol Chem 268:19810 –19818
328. Knop M, Finger A, Braun T, Hellmuth K, Wolf DH 1996 Der1, a
novel protein specifically required for endoplasmic reticulum degradation in yeast. EMBO J 15:753–763
329. Ward CL, Omura S, Kopito RR 1995 Degradation of CFTR by the
ubiquitin-proteasome pathway. Cell 83:121–127
330. Jensen TJ, Loo MA, Pind S, Williams DB, Goldberg AL, Riordan
JR 1995 Multiple proteolytic systems, including the proteasome,
contribute to CFTR processing. Cell 83:129 –135
331. Wiertz EJHJ, Jones TR, Sun L, Bogyo M, Geuze HJ, Ploegh HL
1996 The human cytomegalovirus US11 gene product dislocates
MHC class I heavy chains from the endoplasmic reticulum to the
cytosol. Cell 84:769 –779
332. Hiller MM, Finger A, Schweiger M, Wolf DH 1996 ER degradation
of a misfolded luminal protein by the cytosolic ubiquitin-proteasome pathway. Science 273:1725–1728
333. Wiertz EJHJ, Tortorella D, Bogyo M, Yu J, Mothes W, Jones TR,
Rapoport TA, Ploegh HL 1996 Sec61-mediated transfer of a membrane protein from the endoplasmic reticulum to the proteasome
for destruction. Nature 384:432– 438
334. Ciechanover A 1994 The ubiquitin-proteasome proteolytic pathway. Cell 79:13–21
335. Goldberg AL 1995 Functions of the proteasome: the lysis at the end
of the tunnel. Science 268:522–523
336. Jentsch S, Schlenker S 1995 Selective protein degradation: a journey’s end within the proteasome. Cell 82:881– 884
337. Coux O, Tanaka K, Goldberg AL 1996 Structure and functions of
the 20S and 26S proteasomes. Annu Rev Biochem 65:801– 847
338. Bonifacino JS, Lippincott-Schwartz J 1991 Degradation of proteins
within the endoplasmic reticulum. Curr Opin Cell Biol 3:592– 600
339. Hayes SA, Dice JF 1996 Roles of molecular chaperones in protein
degradation. J Cell Biol 132:255–258
340. Knittler MR, Dirks S, Haas IG 1995 Molecular chaperones involved in protein degradation in the endoplasmic reticulum: quantitative interaction of the heat shock cognate protein BiP with partially folded immunoglobulin light chains that are degraded in the
endoplasmic reticulum. Proc Natl Acad Sci USA 92:1764 –1768
341. Qu D, Teckman JH, Omura S, Perlmutter DH 1996 Degradation
of a mutant secretory protein, alpha1-antitrypsin Z, in the endoplasmic reticulum requires proteasome activity. J Biol Chem 271:
22791–22795
342. Kearse KP, Williams DB, Singer A 1994 Persistence of glucose
residues on core oligosaccharides prevents association of TCRa and
TCRb proteins with calnexin and results specifically in accelerated
degradation of nascent TCRa proteins within the endoplasmic reticulum. EMBO J 13:3678 –3686
343. Liu Y, Choudhury P, Cabral CM, Sifers RN 1997 Intracellular
disposal of incompletely folded human a1-antitrypsin involves
release from calnexin and post-translational trimming of asparagine-linked oligosaccharides. J Biol Chem 272:7946 –7951
344. Bush KT, Goldberg AL, Nigam SK 1997 Proteasome inhibition
leads to a heat-shock response, induction of endoplasmic reticulum
chaperones, and thermotolerance. J Biol Chem 272:9086 –9092
345. Medeiros-Neto G, Stanbury JB 1994 Defects in Tg gene expression
and Tg secretion. In: Inherited Disorders of the Thyroid System.
Chapter 7. CRC Press, Boca Raton, FL, vol 221:107–138
346. Kim PS, Arvan P 1991 Folding and assembly of newly synthesized
thyroglobulin occurs in a pre-Golgi compartment. J Biol Chem
266:12412–12418
347. Medeiros-Neto G, Targovnik HM, Vassart G 1993 Defective thyroglobulin synthesis and secretion causing goiter and hypothyroidism. Endocr Rev 14:165–183
348. Medeiros-Neto G, Kim PS, Yoo SE, Vono J, Targovnik H, Ca-
349.
350.
351.
352.
353.
354.
355.
356.
357.
358.
359.
360.
361.
362.
363.
364.
365.
366.
367.
368.
369.
199
margo R, Hossain SA, Arvan P 1996 Congenital hypothyroid goiter
with deficient thyroglobulin. Identification of an endoplasmic reticulum storage disease (ERSD) with induction of molecular chaperones. J Clin Invest 98:2838 –2844
Taylor BA, Rowe L 1987 The congenital goiter mutation is linked
to the thyroglobulin gene in the mouse. Proc Natl Acad Sci USA
84:1986 –1990
Beamer WG, Maltais LJ, DeBaets MH, Eicher EM 1987 Inherited
congenital goiter in mice. Endocrinology 120:838 – 840
Sugisaki T, Beamer WG, Noguchi T 1992 Microcephalic cerebrum
with hypomyelination in the congenital goiter mouse (cog). Neurochem Res 17:1037–1040
Adkison LR, Taylor S, Beamer WG 1990 Mutant gene-induced
disorders of structure, function and thyroglobulin synthesis in
congenital goitre (cog/cog) in mice. J Endocrinol 126:51–58
Mayerhofer A, Amador AG, Beamer WG, Bartke A 1988 Ultrastructural aspects of the goiter in cog/cog mice. J Hered 79:200 –203
Cabrer B, Brocas H, Perez-Castillo A, Pohl V, Navas JJ, Targovnik
H, Centenera JA, Vassart G 1986 Normal level of thyroglobulin
messenger ribonucleic acid in a human congenital goiter with thyroglobulin deficiency. J Clin Endocrinol Metab 63:931–940
Ohyama Y, Hosoya T, Kameya T, Suzuki N, Nakamura S, Kazahari K, Shibayama K, Yokota Y, Matsura N 1994 Congenital
euthyroid goitre with impaired thyroglobulin transport. Clin Endocrinol (Oxf) 41:129 –135
Targovnick HM, Vono J, Billerbeck EC, Cerrone GE, Varela V,
Mendive F, Wajchenberg BL, Medeiros-Neto G 1995 A 138-nucleotide deletion in the thyroglobulin ribonucleic acid messenger in
a congenital goiter with defective thyroglobulin synthesis. J Clin
Endocrinol Metab 80:3356 –3360
Basche M, Beamer WG, Schneider AB 1989 Abnormal properties
of thyroglobulin in mice with inherited congenital goiter (cog/cog).
Endocrinology 124:1822–1829
Fogelfeld L, Harel G, Beamer WG, Schneider AB 1992 Low-molecular-weight iodoproteins in the congenital goiters of cog/cog
mice. Thyroid 2:329 –335
Dunn JT 1992 Thyroglobulin: chemistry and biosynthesis. In:
Braverman L, Utiger R (eds) Werner and Ingbar’s: The Thyroid,
Lippincott, Philadelphia, pp 98 –110
Malthiery Y, Marriq C, Berge-Lefranc J-L, Franc J-L, Henry M,
Lejeune P-J, Ruf J, Lissitzky S 1989 Thyroglobulin structure and
function: recent advances. Biochimie 71:195–210
Arvan P, Kim PS, Kuliawat R, Prabakaran D, Muresan Z, Yoo SE,
Hossain SA 1997 Intracellular protein transport to the thyrocyte
plasma membrane: potential implications for thyroid physiology.
Thyroid 7:89 –105
Yoshida S, Takamatsu J, Kuma K, Murakami Y, Sakane S, Katayama S, Tarutani O, Ohsawa N 1996 A variant of adenomatous
goiter with characteristic histology and possible hereditary thyroglobulin abnormality. J Clin Endocrinol Metab 81:1961–1966
Miller WL 1993 Molecular genetics of familial central diabetes
insipidus. J Clin Endocrinol Metab 77:592–595
Robertson GL 1995 Diabetes insipidus. Endocrinol Metab Clin
North Am 24:549 –572
Schmale H, Bahnsen U, Richter D 1993 Structure and expression
of the vasopressin precursor gene in central diabetes insipidus. Ann
NY Acad Sci 689:74 – 82
Repaske DR, Summar ML, Krishnamani MR, Gultekin EK, Arriazu MC, Roubicek ME, Blanco M, Isaac GB, Phillips JA3 1996
Recurrent mutations in the vasopressin-neurophysin II gene cause
autosomal dominant neurohypophyseal diabetes insipidus. J Clin
Endocrinol Metab 81:2328 –2334
Rittig S, Robertson GL, Siggaard C, Kovacs L, Gregersen N,
Nyborg J, Pedersen EB 1996 Identification of 13 new mutations in
the vasopressin-neurophysin II gene in 17 kindreds with familial
autosomal dominant neurohypophyseal diabetes insipidus. Am J
Hum Genet 58:107–117
Bahnsen U, Oosting P, Swaab DF, Nahke P, Richter D, Schmale
H 1992 A missense mutation in the vasopressin-neurophysin precursor gene cosegregates with human autosomal dominant neurohypophysial diabetes insipidus. EMBO J 11:19 –23
Nagasaki H, Ito M, Yuasa H, Saito H, Fukase M, Hamada K,
Ishikawa E, Katakami H, Oiso Y 1995 Two novel mutations in the
200
370.
371.
372.
373.
374.
375.
376.
377.
378.
379.
380.
381.
382.
383.
384.
385.
386.
387.
388.
KIM AND ARVAN
coding region for neurophysin-II associated with familial central
diabetes insipidus. J Clin Endocrinol Metab 80:1352–1356
Rauch F, Lenzner C, Nurnberg P, Frommel C, Vetter U 1996 A
novel mutation in the coding region for neurophysin-II is associated with autosomal dominant neurohypophyseal diabetes insipidus. Clin Endocrinol (Oxf) 44:45–51
Ueta Y, Taniguchi S, Yoshida A, Murakami I, Mitani Y, Hisatome
I, Manabe I, Sato R, Tsuboi M, Ohtahara A, Nanba E, Shigemasa
C 1996 A new type of familial central diabetes insipidus caused by
a single base substitution in the neurophysin II coding region of the
vasopressin gene. J Clin Endocrinol Metab 81:1787–1790
McLeod JF, Kovacs L, Gaskill MB, Rittig S, Bradley GS, Robertson GL 1993 Familial neurohypophyseal diabetes insipidus associated with a signal peptide mutation. J Clin Endocrinol Metab
77:599A–599G
Rutishauser J, Boni-Schnetzler M, Boni J, Wichmann W, Huisman T, Vallotton MB, Froesch ER 1996 A novel point mutation in
the translation initiation codon of the pre-pro-vasopressin-neurophysin II gene: cosegregation with morphological abnormalities
and clinical symptoms in autosomal dominant neurohypophyseal
diabetes insipidus. J Clin Endocrinol Metab 81:192–198
Schmale H, Borowiak B, Holtgreve-Grez H, Richter D 1989 Impact of altered protein structures on the intracellular traffic of a
mutated vasopressin precursor from Brattleboro rats. Eur J Biochem 182:621– 627
Morris JF, Pow DV, Mohr E, Richter D 1995 Production of hybrid
oxytocin/vasopressin precursors and accumulation of oxytocin
precursors in the rough endoplasmic reticulum of rat magnocellular neurons. Adv Exp Med Biol 395:49 –58
Pow DV, Morris JF, Ward AR 1992 Immuno-electron microscopic
evidence for two different types of partial somatic repair of the
mutant Brattleboro vasopressin gene. Neuroscience 50:503–512
van Leeuwen FW 1992 Mutant vasopressin precursor producing
cells of homozygous Brattleboro rat as a model for co-expression
of neuropeptides. Prog Brain Res 92:149 –155
Deen PM, Verdijk MA, Knoers NV, Wieringa B, Monnens LA,
van Os CH, van Oost BA 1994 Requirement of human renal water
channel aquaporin-2 for vasopressin-dependent concentration of
urine. Science 264:92–95
Rosenthal W, Seibold A, Bichet DG, Birnbaumer M 1995 Diabetes
insipidus: nephrogenic DI. In: Meyers RA (ed) Molecular Biology
and Biotechnology. VCH Publishers Inc., New York, pp 213–214
Deen PM, Croes H, van Aubel RA, Ginsel LA, van Os CH 1995
Water channels encoded by mutant aquaporin-2 genes in nephrogenic diabetes insipidus are impaired in their cellular routing. J Clin
Invest 95:2291–2296
Bichet DG, Arthus MF, Lonergan M, Hendy GN, Paradis AJ,
Fujiwara TM, Morgan K, Gregory MC, Rosenthal W, Didwania
A, Antaramian A, Birnbaumer M 1993 X-linked nephrogenic diabetes insipidus mutations in North America and the Hopewell
hypothesis. J Clin Invest 92:1262–1268
Rosenthal W, Seibold A, Antaramian A, Lonergan M, Arthus MF,
Hendy GN, Birnbaumer M, Bichet DG 1992 Molecular identification of the gene responsible for congenital nephrogenic diabetes
insipidus. Nature 359:233–235
Wenkert D, Merendino Jr JJ, Shenker A, Thambi N, Robertson
GL, Moses AM, Spiegel AM 1994 Novel mutations in the V2
vasopressin receptor gene of patients with X-linked nephrogenic
diabetes insipidus. Hum Mol Genet 3:1429 –1430
van der Rest M, Garrone R 1991 Collagen family of proteins.
FASEB J. 5:2814 –2823
Kuivaniemi H, Tromp G, Prockop DJ 1991 Mutations in collagen
genes: causes of rare and some common diseases in humans.
FASEB J 5:2052–2060
Aurbach GD, Marx SJ, Spiegel AM 1992 Chemistry of bone and
bone proteins: collagen biosynthesis. In: Wilson JD, Foster DW
(eds) Williams’ Textbook of Endocrinology. W.B. Saunders, Phildelphia, pp 1483–1484
Clark CC, Kefalides NA 1978 Localization and partial composition
of the oligosaccharide units on propeptide extensions of type I
procollagen. J Biol Chem 253:47–51
Kellokumpu S, Sormunen R, Heikkinen J, Myllyla R 1994 Lysyl
hydroxylase, a collagen processing enzyme, exemplifies a novel
389.
390.
391.
392.
393.
394.
395.
396.
397.
398.
399.
400.
401.
402.
403.
404.
405.
Vol. 19, No. 2
class of luminally-oriented peripheral membrane proteins in the
endoplasmic reticulum. J Biol Chem 269:30524 –30529
Pacifici M, Iozzo RV 1988 Remodeling of the rough endoplasmic
reticulum during stimulation of procollagen secretion by ascorbic
acid in cultured chondrocytes. A biochemical and morphological
study. J Biol Chem 263:2483–2492
Beck K, Boswell BA, Ridgway CC, Bachinger HP 1996 Triple helix
formation of procollagen type I can occur at the rough ER membrane. J Biol Chem 271:21566 –21573
Lamande SR, Chessler SD, Golub SB, Byers PH, Chan D, Cole
WG, Sillence DO, Bateman JF 1995 Endoplasmic reticulum-mediated quality control of type I collagen production by cells from
osteogenesis imperfecta patients with mutations in the pro alpha
1 (I) chain carboxyl-terminal propeptide which impair subunit
assembly. J Biol Chem 270:8642– 8649
Steinmann B, Bruckner P, Superti-Furga A 1991 Cyclosporin A
slows collagen triple-helix formation in vivo: indirect evidence for
a physiologic role of peptidyl-prolyl cis-trans-isomerase. J Biol
Chem 266:1299 –1303
Bonadio J, Byers PH 1985 Subtle structural alterations in the chains
of type I procollagen produce osteogenesis imperfecta type II. Nature 316:363–366
Prockop DJ, Chu M-L, De Wet W, Myers JC, Pihlajaniemi T,
Ramirez F, Sippola M 1985 Mutations in osteogenesis imperfecta
leading to the synthesis of abnormal type I procollagens. Ann NY
Acad Sci 460:289 –297
Wenstrup RJ, Willing MC, Starman BJ, Byers PH 1990 Distinct
biochemical phenotypes predict clinical severity in nonlethal variants of osteogenesis imperfecta. Am J Hum Genet 46:975–982
Cohen-Solal L, Zolezzi F, Pignatti PF, Mottes M 1996 Intrafamilial
variable expressivity of osteogenesis imperfecta due to mosaicism
for a lethal G382R substitution in the COL1A1 gene. Mol Cell
Probes 10:219 –225
Culbert AA, Wallis GA, Kadler KE 1996 Tracing the pathway
between mutation and phenotype in osteogenesis imperfecta: isolation of mineralization-specific genes. Am J Med Genet 63:167–174
Lund AM, Schwartz M, Raghunath M, Steinmann B, Skovby F
1996 Gly802Asp substitution in the pro alpha 2(I) collagen chain in
a family with recurrent osteogenesis imperfecta due to paternal
mosaicism. Eur J Hum Genet 4:39 – 45
Mundlos S, Chan D, Weng YM, Sillence DO, Cole WG, Bateman
JF 1996 Multiexon deletions in the type I collagen COL1A2 gene in
osteogenesis imperfecta type IB. Molecules containing the shortened alpha2(I) chains show differential incorporation into the bone
and skin extracellular matrix. J Biol Chem 271:21068 –21074
Zhuang J, Tromp G, Kuivaniemi H, Castells S, Prockop DJ 1996
Substitution of arginine for glycine at position 154 of the alpha 1
chain of type I collagen in a variant of osteogenesis imperfecta:
comparison to previous cases with the same mutation. Am J Med
Genet 61:111–116
Oliver JE, Thompson EM, Pope FM, Nicholls AC 1996 Mutation
in the carboxy-terminal propeptide of the pro alpha 1(I) chain of
type I collagen in a child with severe osteogenesis imperfecta (OI
type III): possible implications for protein folding. Hum Mutat
7:318 –326
Redford-Badwal DA, Stover ML, Valli M, McKinstry MB, Rowe
DW 1996 Nuclear retention of COL1A1 messenger RNA identifies
null alleles causing mild osteogenesis imperfecta. J Clin Invest
97:1035–1040
Willing MC, Deschenes SP, Slayton RL, Roberts EJ 1996 Premature chain termination is a unifying mechanism for COL1A1 null
alleles in osteogenesis imperfecta type I cell strains. Am J Hum
Genet 59:799 – 809
Bogaert R, Tiller GE, Weis MA, Gruber HE, Rimoin DL, Cohn
DH, Eyre DR 1992 An amino acid substitution (Gly853–.Glu) in
the collagen alpha 1(II) chain produces hypochondrogenesis. J Biol
Chem 267:22522–22526
Bonaventure J, Rousseau F, Legeai-Mallet L, Le Merrer M, Munnich A, Maroteaux P 1996 Common mutations in the fibroblast
growth factor receptor 3 (FGFR 3) gene account for achondroplasia,
hypochondroplasia, and thanatophoric dwarfism. Am J Med Genet
63:148 –154
April, 1998
ENDOCRINE ERSDs
406. Francomano CA, McIntosh I, Wilkin DJ 1996 Bone dysplasias in
man: molecular insights. Curr Opin Genet Dev 6:301–308
407. Gilbert-Barnes E, Langer LO, Opitz JM, Laxova R, Sotelo-Arila C
1996 Kniest dysplasia: radiologic, histopathological, and scanning
EM findings. Am J Med Genet 63:34 – 45
408. Horton WA 1996 Molecular genetic basis of the human chondrodysplasias. Endocrinol Metab Clin North Am 25:683– 697
409. Mundlos S, Chan D, McGill J, Bateman JF 1996 An alpha 1(II)
Gly913 to Cys substitution prevents the matrix incorporation of
type II collagen which is replaced with type I and III collagens in
cartilage from a patient with hypochondrogenesis. Am J Med Genet
63:129 –136
410. Dyne KM, Valli M, Forlino A, Mottes M, Kresse H, Cetta G 1996
Deficient expression of the small proteoglycan decorin in a case of
severe/lethal osteogenesis imperfecta. Am J Med Genet 63:161–166
411. Zabel B, Hilbert K, Stoss H, Superti-Furga A, Spranger J, Winterpacht A 1996 A specific collagen type II gene (COL2A1) mutation presenting as spondyloperipheral dysplasia. Am J Med Genet
63:123–128
412. Schaefer EJ, Genest Jr JJ, Ordovas JM, Salem DN, Wilson PW 1994
Familial lipoprotein disorders and premature coronary artery disease. Atherosclerosis 108:S41–S54
413. Pullinger CR, Kane JP 1995 Lipid metabolism and transport. In:
Meyers RA (ed) Molecular Biology and Biotechnology. VCH Publishers Inc., New York, pp 494 –509
414. Brown MS, Anderson RGW, Goldstein JL 1983 Recycling receptors: the round trip itinary of LDL recpetors. Cell 32:663– 667
415. Shite S, Seguchi T, Shimada T, Ono M, Kuwano M 1990 Rapid
turnover of low-density lipoprotein receptor by a non-lysosomal
pathway in mouse macrophage J774 cells and inhibitory effect of
brefeldin A. Eur J Biochem 191:491– 497
416. Foster JD, Strauss III JF, Paavola LG 1993 Cellular events involved
in hormonal control of receptor-mediated endocytosis: regulation
occurs at multiple sites in LDL pathway, including steps beyond
the receptor. Endocrinology 132:337–350
417. Goldstein JL, Hobbs HH, Brown MS 1995 Familial hypercholesterolemia. In: Scriver CR, Beaudet AL, Sly WS, Valle D (eds) The
Metabolic and Molecular Basis of Inherited Disease. McGraw Hill,
New York, pp 1981–2030
418. Lehrman MA, Schneider WJ, Brown MS, Davis CG, Elhammer A,
Russell DW, Goldstein JL 1987 The Lebanese allele at the low
density lipoprotein receptor locus. Nonsense mutation produces
truncated receptor that is retained in endoplasmic reticulum. J Biol
Chem 262:401– 410
419. Pathak RK, Merkle RK, Cummings RD, Goldstein JL, Brown MS,
Anderson RG 1988 Immunocytochemical localization of mutant
low density lipoprotein receptors that fail to reach the Golgi complex. J Cell Biol 106:1831–1841
420. Wetterau JR, Aggerbeck LP, Bouma ME, Eisenberg C, Munck A,
Hermier M, Schmitz J, Gay G, Rader DJ, Gregg RE 1992 Absence
of microsomal triglyceride transfer protein in individuals with
abetalipoproteinemia. Science 258:999 –1001
421. Ricci B, Sharp D, O’Rourke E, Kienzle B, Blinderman L, Gordon
D, Smith-Monroy C, Robinson G, Gregg RE, Rader DJ, Wetterau
JR 1995 A 30-amino acid truncation of the microsomal triglyceride
transfer protein large subunit disrupts its interaction with protein
disulfide-isomerase and causes abetalipoproteinemia. J Biol Chem
270:14281–14285
422. Wu X, Zhou M, Huang LS, Wetterau J, Ginsberg HN 1996 Demonstration of a physical interaction between microsomal triglyceride transfer protein and apolipoprotein B during the assembly of
ApoB-containing lipoproteins. J Biol Chem 271:10277–10281
423. Du EZ, Wang SL, Kayden HJ, Sokol R, Curtiss LK, Davis RA 1996
Translocation of apolipoprotein B across the endoplasmic reticulum is blocked in abetalipoproteinemia. J Lipid Res 37:1309 –1315
424. Wang S, McLeod RS, Gordon DA, Yao Z 1996 The microsomal
triglyceride transfer protein facilitates assembly and secretion of
apolipoprotein B-containing lipoproteins and decreases cotranslational degradation of apolipoprotein B in transfected COS-7 cells.
J Biol Chem 271:14124 –14133
425. Patel SB, Grundy SM 1996 Interactions between microsomal triglyceride transfer protein and apolipoprotein B within the endo-
426.
427.
428.
429.
430.
431.
432.
433.
434.
435.
436.
437.
438.
439.
440.
441.
442.
443.
444.
201
plasmic reticulum in a heterologous expression system. J Biol Chem
271:18686 –18694
Etienne J, Brault D 1992 Lipoprotein lipase deficiencies. Ann Biol
Clin (Paris) 50:299 –309
Ben-Zeev O, Doolittle MH, Davis RC, Elovson J, Schotz MC 1992
Maturation of lipoprotein lipase. Expression of full catalytic activity
requires glucose trimming but not translocation to the cis-Golgi
compartment. J Biol Chem 267:6219 – 6227
Masuno H, Blanchette-Mackie EJ, Schultz CJ, Spaeth AE, Scow
RO, Okuda H 1992 Retention of glucose by N-linked oligosaccharide chains impedes expression of lipoprotein lipase activity: effect
of castanospermine. J Lipid Res 33:1343–1349
Busca R, Pujana MA, Pognonec P, Auwerx J, Deeb SS, Reina M,
Vilaro S 1995 Absence of N-glycosylation at asparagine 43 in human lipoprotein lipase induces its accumulation in the rough endoplasmic reticulum and alters this cellular compartment. J Lipid
Res 36:939 –951
Carroll R, Ben-Zeev O, Doolittle MH, Severson DL 1992 Activation of lipoprotein lipase in cardiac myocytes by glycosylation
requires trimming of glucose residues in the endoplasmic reticulum. Biochem J 285:693– 696
Park JW, Blanchette-Mackie EJ, Scow RO 1996 Brefeldin A enables
synthesis of active lipoprotein lipase in cld/cld and castanospermine-treated mouse brown adipocytes via translocation of Golgi
components to endoplasmic reticulum. Biochem J 317:125–134
Ma Y, Henderson HE, Murthy V, Roederer G, Monsalve MV,
Clarke LA, Normand T, Julien P, Gagne C, Lambert M, Davignon
J, Lupien PJ, Brunzell J, Hayden MR 1991 A mutation in the
human lipoprotein lipase gene as the most common cause of familial chylomicronemia in French Canadians. N Engl J Med 324:
1761–1766
Sifers RN, Finegold MJ, Woo SL 1992 Molecular biology and
genetics of alpha1-antitrypsin deficiency. Semin Liver Dis 12:301–
310
Le A, Graham KS, Sifers RN 1990 Intracellular degradation of the
transport-impaired human PiZ a1-antitrypsin variant. Biochemical
mapping of the degradative event among compartments of the
secretory pathway. J Biol Chem 265:14001–14007
Le A, Ferrell GA, Dishon DS, Le Q-QA, Sifers RN 1992 Soluble
aggregates of the human PiZ a1-antitrypsin variant are degraded
within the endoplasmic reticulum by a mechanism sensitive to
inhibitors of protein synthesis. J Biol Chem 267:1072–1080
Carlson JA, Rogers BB, Sifers RN, Finegold MJ, Clift SM, DeMayo FJ 1989 Accumulation of PiZ a1-antitrypsin causes liver
damage in transgenic mice. J Clin Invest 83:1183–1190
Graham KS, Le A, Sifers RN 1990 Accumulation of the insoluble
PiZ variant human a1-antitrypsin within the hepatic endoplasmic
reticulum does not elevate the steaty-state level of grp78/BiP. J Biol
Chem 265:20463–20468
Perlmutter DH, Schlesinger MJ, Pierce JA, Punsal PI, Schwartz
AL 1989 Synthesis of stress proteins is increased in individuals with
homozygous PiZZ a1-antitrypsin deficiency and liver disease.
J Clin Invest 84:1555–1561
Sato S, Ward CL, Krouse ME, Wine JJ, Kopito RR 1996 Glycerol
reverses the misfolding phenotype of the most common cystic
fibrosis mutation. J Biol Chem 271:635– 638
Welsh MJ, Smith AE 1993 Molecular mechanism of CFTR chloride
channel dysfunction in cystic fibrosis. Cell 73:1251–1254
Pind S, Riordan JR, Williams DB 1994 Participation of the endoplasmic reticulum chaperone calnexin (p88, IP90) in the biogenesis
of the cystic fibrosis transmembrane conductance regulator. J Biol
Chem 269:12784 –12788
Yang Y, Janich S, Cohn JA, Wilson JM 1993 The common variant
of cystic fibrosis transmembrane conductance regulator is recognized by hsp 70 and degraded in a pre-Golgi nonlysosomal compartment. Proc Natl Acad Sci USA 90:9480 –9484
Pasyk EA, Foskett JK 1995 Mutant (DF508) cystic fibrosis transmembrane conductance regulator Cl2 channel is functional when
retained in the endoplasmic reticulum of mammalian cells. J Biol
Chem 270:12347–12350
Abramowicz MJ, Targovnik HM, Varela V, Cochaux P, Krawiec
L, Pisarev MA, Propato FV, Juvenal G, Chester HA, Vassart G
1992 Identification of a mutation in the coding sequence of the
202
445.
446.
447.
448.
449.
450.
451.
452.
KIM AND ARVAN
human thyroid peroxidase gene causing congenital goiter. J Clin
Invest 90:1200 –1204
Medeiros-Neto GA, Billerbeck AE, Wajchenberg BL, Targovnik
HM 1993 Defective organification of iodide causing hereditary
goitrous hypothyroidism. Thyroid 3:143–159
Carvalho DP, Rego KG, Rosenthal D 1994 Thyroid peroxidase in
dyshormonogenic goiters with organification and thyroglobulin
defects. Thyroid 4:421– 426
Miura Y, Mori Y, Kambe F, Tani Y, Oiso Y, Seo H 1994 Impaired
intracellular transport contributes to partial thyroxine-binding
globulin deficiency in a Japanese family. J Clin Endocrinol Metab
79:740 –744
Miura Y, Kambe F, Yamamori I, Mori Y, Tani Y, Murata Y, Oiso
Y, Seo H 1994 A truncated thyroxine-binding globulin due to a
frameshift mutation is retained within the rough endoplasmic reticulum: a possible mechanism of complete thyroxine-binding
globulin deficiency in Japanese. J Clin Endocrinol Metab 78:283–287
White AL, Hixson JE, Rainwater DL, Lanford RE 1994 Molecular
basis for “null” lipoprotein(a) phenotypes and the influence of
apolipoprotein(a) size on plasma lipoprotein(a) level in the baboon.
J Biol Chem 269:9060 –9066
Taylor SI 1992 Lilly Lecture: molecular mechanisms of insulin
resistance. Lessons from patients with mutations in the insulinreceptor gene. Diabetes 41:1473–1490
Duquesnoy P, Sobrier ML, Amselem S, Goossens M 1991 Defective membrane expression of human growth hormone (GH) receptor causes Laron-type GH insensitivity syndrome. Proc Natl
Acad Sci USA 88:10272–10276
Edery M, Rozakis-Adcock M, Goujon L, Finidori J, LeviMeyrueis C, Paly J, Djiane J, Postel-Vinay MC, Kelly PA 1993
453.
454.
455.
456.
457.
458.
459.
460.
Vol. 19, No. 2
Lack of hormone binding in COS-7 cells expressing a mutated
growth hormone receptor found in Laron dwarfism. J Clin Invest
91:838 – 844
Rosenfeld RG, Rosenbloom AL, Guevara-Aguirre J 1994 Growth
hormone (GH) insensitivity due to primary GH receptor deficiency.
Endocr Rev 15:369 –390
Arnold A, Horst SA, Gardella TJ, Baba H, Levine MA, Kronenberg HM 1990 Mutation of the signal peptide-encoding region of
the preproparathyroid hormone gene in familial isolated hypoparathyroidism. J Clin Invest 86:1084 –1087
Naggert JK, Fricker LD, Varlamov O, Nishina PM, Rouille Y,
Steiner DF, Carroll RJ, Paigen BJ, Leiter EH 1995 Hyperproinsulinaemia in obese fat/fat mice associated with carboxypeptidase E
mutation which reduces enzyme activity. Nat Genet 10:135–142
Varlamov O, Leiter EH, Fricker L 1996 Induced and spontaneous
mutations at Ser202 of carboxypeptidase E. Effect on enzyme expression, activity, and intracellular routing. J Biol Chem 271:13981–
13986
Rowling PJ, McLaughlin SH, Pollock GS, Freedman RB 1994 A
single purification procedure for the major resident proteins of the
ER lumen: endoplasmin, BiP, calreticulin and protein disulfide
isomerase. Protein Expr Purif 5:331–336
Deleted in proof
Annaert WG, Becker B, Kistner U, Reth M, Jahn R 1997 Export of
cellubrevin from the endoplasmic reticulum is controlled by
BAP31. J Cell Biol 139:1397–141
Ito M, Jameson JL, Ito M 1997 Molecular basis of autosomal dominant neurohypophyseal diabetes insipidus. Cellular toxicity
caused by the accumulation of mutant vasopressin precursors
within the endoplasmic reticulum. J Clin Invest 99:1897–1905