Rab GTPases in vesicular
Marino
European
Zerial and Harald
Molecular
Biology
Laboratory,
transport
Stenmark
Heidelberg,
Germany
Specificity
and directionality
are two features shared by the numerous
steps of membrane
transport that connect cellular organelles. By shuttling
between
specific membrane
compartments
and the cytoplasm,
small
GTPases of the Rab family appear to regulate
membrane
traffic in a
cyclical manner. The restriction
of certain Rab proteins to differentiated
cell types supports a role for these GTPases in defining the specificity of
membrane
trafficking.
Current
Opinion
in Cell Biology
Introduction
Eukaryotic cells are divided into distinct membranebound organelles. While this functional compartmentalization has the advantage of both efficiency and regulation, it also requires controlled interactions between
the various organelles. Exchange of constituents between organefles is thought to occur through vesicular or
tubular intermediates that selectively deliver proteins and
lipids from one compartment to another [ 10-3.1. Specific
molecules control this complex trafficking network. Over
the past few years, a large number of small GTPases of
the Rab family have been described and localized to various intracellular compartments. By alternating between
two distinct conformations in a nucleotide-dependent
fashion, these proteins function as molecular switches.
Through this mechanism they have been proposed to
ensure fidelity in the process of docking and/or fusion
of vesicles with their correct acceptor compartment [4].
The cycle of GTP binding and hydrolysis would allow
multiple rounds of vesicular transport and confer vectorial@. This raiew focuses on the involvement of Rab
proteins in individual trafficking routes between intracellular compartments.
Function
of Rab proteins
in membrane
traffic
The role of Rab GTRases in membrane traffic was first
demonstrated by studies on Sec4p and Yptlp, which
function in the exocytic pathway of Saccbaromyces cere
~r&ae [2*]. Rab proteins, the mammalian counterparts of
yeast Sec4p and Yptlp [ 51, are localized to distinct intracellular compartments (Table 1) and a combination of
in vitro and in vivo studies has emphasized their role in
the regulation of different exocytic and endocytic transport processes.
1993, 5~613-620
In the exocytic pathway, in vitro studies have shown that
the Rablb protein is involved in endoplasmic reticulum
(ER) to Golgi transport [6] and work in intact cells has
indicated that this transport process requires also Rabla
and Rab2 proteins [ 71. The participation of three different Rab proteins in ERGolgi transport might reflect sequential steps of vesicular transport or, alternatively, the
concerted function of several Rab GTPases at the same
step. Rab proteins also function in regulated secretion.
Synthetic peptides corresponding to the effector region
of Rab3a stimulate regulated exocytosis [S-lo].
In the endocytic pathway, both Rab4 and Rab5 proteins
are associated with early endosomes (Rab5 is also on the
plasma membrane) [ 11,121. However, they have distinct
activities. An increase in the level of Rab5 stimulates endocytosis and expands the size of the early endosomes,
whereas expression of a Rab5 GTP-binding defective mutant has the opposite effect [13*]. These morphological
alterations are consistent with a role of Rab5 in the fusion between early endosomes in vitro [ 141. In contrast,
overexpression of Rab4 increases the re&x of endocytic
markers to the cell surface and induces the accumulation
of endocytic tubules and tubule clusters that could correspond either to structurally altered early endosomes or
even to a distinct recycling compartment [IS*].
Another example of distinct function of Rab proteins in
the same organelle is provided by Rab7 and Rab9. Both
proteins have been local@ed to late endosomes [12,16],
but only Rab9 spectically functions in transport from this
compartment to the tram&olgi network (TGN) in vitro
[16]. While the role of mammalian Rab7 remains unknown, functional studies on a structural homologue
in S. cerevzs&e, Ypt7p, suggest that this protein may
function at a late step of the endocytic pathway [17’].
Ypt7 null mutants are viable and exhibit normal exocytosis and u-factor internalization, but a-factor degradation
is inhibited and the vacuole is fragmented. By analogy
Abbreviations
CHM-choroideremia;
CDL-GDP
DC&-dominant
suppressor
of Set+ ER-endoplasmic
reticulum;
CAP-GTPase activating
dissociation
inhibitor;
W-mammalian
suppressor
of Sec4; NSF-N-ethylmaleimide-sensitive
fusion
SNAP-soluble
NSF attachment
protein;
SNARE-SNAP
receptor;
TCN-trans-Colgi
network.
@ Current
Biology
Ltd ISSN 0955+674
protein;
protein;
613
614
Membranes
.
Table 1. Summary
and
of the
cells.
mammalian
Protein
Localization
Yptl
(yeast)
Sec4
ER-Golgi
localization,
functional
ER-Golgi
ER-Golgi
intermediate
compartment
Synaptic
vesicles
Chromaffin
granules
Rab3a
RaMa
Early
Rab5a
Rab7
Rab8
Rab17
Apical
basolateral
‘Regulatory
most
clathrin-
Late endosomes
Post-Golgi
basolateral
secretory
vesicles
Late endosomes,
TGN
Rab9
factors
Rab proteins.
active
of Yptl,
on different
Sec4
and
Interacting
DSSC,
Rab proteins
in yeast
components
MSSC
Rabphilin-3A
Not determined
Golgi-plasma
membrane
transport
Transport
from late endosomes
to TGN
Not determined
dense tubules,
plasma
membrane
See text
components
Regulated
exocytosis
in pancreatic
acinar
cells, adrenal
chromaffin
cells
and mast cells
Early endosomes-plasma
membrane
recycling
pathway
Plasma membrane-early
endosome
transport
and homotypic
fusion
between
early endosomes
Not determined
Transport
in endocytic
pathway
endosomes
Plasma membrane,
coated
vesicles,
early endosomes
Middle
Colgi-TGN
Not determined
Rab6
Ypt7
interacting
Fusion of post-ER
vesicles
with plasma
membrane
Fusion of post-Golgi
vesicles
with plasma
membrane
ER-Golgi
transport
in yeast and mammalian
cells
ER-cis-Colgi
and c&medial
Colgi transport
ER-Goigi
transport
dabla
Rab2
and
Function
Secretory
vesiclesplasma
membrane
Rabl b
properties
Rab proteins.
Rab GDI
is not
listed
among
the
interacting
components,
as it interacts
with
for references.
with mammalian endocytic pathway organization and localization of Rab7 to late endosomes, Ypt7p is thought to
control transport between early and late endosomal compartments in yeast [ 17’1. This interpretation is consistent
with the existence of kinetically and biochemically distinct endqtic organelles in yeast [ 181. Certainly, these
observations suggest that the yeast endocytic machinery
shares features with the mammalian endocytic apparatus.
A vesicle transport process can be typically divided into
several steps: budding, uncoating, docking and fusion
[1~3*]. While Rab proteins are implicated in the regulation of distinct membrane traffic events, the exact step
controlled by each protein is not clear. Both Sedp and
Rab5 appear to control vesicle docking/fusion. However,
while yeast Yptlp is thought to function in the same step
[2*], its mammalian-related protein, Rablb, is thought
to be essential for vesicle formation [ 61. The interpretation of in vivo studies is complicated by the fact that
transport between compartments may occur in both directions, forwards and backwards, and affecting one direction is likely to influence the other. Overexpression
of Rab5 stimulates endocytosis but also accelerates recycling from the early endosomes to the plasma membrane [WI. Likewise, inhibition of retrograde transport
could, in principle, indirectly affect anterograde movement between the ER and the Golgi, and vice versa
Clearly, these studies have shown that changes in the
balance between vesicles coming from and returning to
a compartment have profound effects on organelle organization. Similar effects have also been observed with
other components of the traf3cking machinery. For example, the sbi6ire mutation in a dynamin-related protein
of Drosophila mehnogaster is thought to block coated
vesicle budding from the plasma membrane [W-21 1.
This causes an increase in its surface area, an accumulation of elongated coated pits and a decrease in the
number of endocytic tubular elements [ 191. Thus, similar to the overexpression of Rab4 and Rab5, the mutation
induces drastic morphological changes. Altogether, these
biochemical and morphological studies only represent a
first step towards a more detailed dissection of Rab protein function.
Rab CTPases in vesicular
Complexity
of the Rab protein
family
The large number of Rab GTPases identiiied in mammalian cells (30 different members) 122-241 led to the
hypothesis that each step of vesicular traffic is regulated
by at least one Rab protein [4]. Nevertheless, the implications of this complexity are not clear. First, the picture is
complicated by the existence of subgroups of Rab proteins sharing high sequence identity (e.g. Rabla, Rablb,
Rab3a, Rab3b, Rab3c, Rab3d). The high sequence conservation suggests that proteins belonging to one subgroup
may have a similar function, as supported by the finding
that both Rabla and Rablb regulate ER-Golgi transport
[7]. However, this simple interpretation is countered by
the fact that members of a subgroup may have different
biochemical properties: both the exocytlc Rabla and the
endocytlc RaMa proteins contain a phosphotylation site
for p34u
kinase [25,26*] whereas Rablb and Rab4b
do not [27]. Anti-Rab5a specific antibodies block early
endosome fusion in vitro, despite the presence of the
Rab5b and Rab5c proteins [28]. Different members of
the Rab3 subgroup carry out distinct functions (see below). Altogether, these differences question the idea that
Rab isofonns are functionally redundant. Instead, the diversity might be responsible for hne tuning intracellular
transport by the coordinated activity of several Rab isoforms.
Second, recent morphological data indicate that several
Rab proteins localize to the same organelle. Early endosomes provide a striking example: they contain at least
three additional Rab proteins besides the Rab4 and Rab5
subgroups (VM Olkkonen, P Dupree, 1 Killisch, A Lutcke,
M Zerial, et al. and A Liitcke, A Valencia, VM Olkkonen,
G Griffiths, RG Patton, et al., unpublished data). What
are the roles played by these different Rab proteins? One
possibility is that they function in either of the transport
routes that meet in this compartment: transport from and
to the cell surface, and from the TGN and to late endosomes [28]. Alternatively, several distinct Rab proteins
might be required in a given transport process. These
proteins might carry out either the same function (e.g.
vesicle docking) or different functions (e.g. budding, uncoating, docking or fusion).
transport
Zerial
and Stenmark
indicate that epithelial cells, which have distinct apical and basolateral transport pathways, and tmnscymtic
routes &vkewed in [33,34]), also express specific Rab
proteins once they become polarized. In the developing
kidney, Rab17 is absent from the mesenchymal precursors but is induced upon their differentiation into epithelial cells. In the adult organ, the protein is associated
with the basolateral plasma membrane and with apical
tubules [35*]. Both expression and localization suggest
that Rabl7 may function in transcytosis, an epithelial cell
specilic transport process [ 331.
Rab8, a ubiquitously expressed protein sharing high sequence homology with Sec4 [27], functions in tratfic
from the TGN to the basolateral surface in polarized
MDCK cells and to the somatodendrltic plasma membrane in hippocampal neurons [36,37]. These two domains are thought to correspond to tile plasma membrane of non-polarized cells [ 381. This suggests that Rab8
does not function in an epithelial cell speciIic pathway. In
contrast, another as yet unidenti6ed GTP-binding protein
is found on immuno-isolated apical exocytic vesicles and
is implicated in apical exocytosis, the epithelial cell specific transport step (361.
Collectively, these data suggest that specialized Rab proteins are part of the machinery regulating cell type specilic transport processes. The localization and function of
the Rab proteins must depend on other cell type specific
regulatory components, for example, factors that regulate
their GTP-GDP cycle.
Regulatory
factors
for Rab proteins
Rab proteins
The cycle of GTP binding and hydrolysis of Rab proteins has been postulated to ensure directionality in the
vesicle docking/fusion process [ 41. According to a common view [4,39], a speciiic Rab protein is recruited on
the donor membrane or on nascent transport vesicles in
its GTP-bound form. Hydrolysis of GTP by the Rab protein is thought to trigger fusion of the vesicle with the acceptor membrane [4,40*]. Subsequently, the GDP-bound
Rab protein is released into the cytosol and returned to
the donor membrane. There, a GDP-GTP exchange protein catalyzes GDP release and GTP binding and allows
the cycle to continue.
The notion that Rab proteins control distinct intracellular
transport processes suggests that cell type speciiic steps
in membrane traific also require specific Rab proteins.
Several Rab proteins that are cell type or tissue-specific
have been identiIied. For instance, neurons, exocrine
and endocrine cells, which display regulated secretion,
speciIically express the Rab3a protein [ 29-311.
The expression of cell type specific Rab proteins is
regulated during differentiation. Expression of Rab3d
increases during ditferentiation of 3T3-Ll cells into
adipocytes. This protein might control the insulindependent exocytosis of vesicles containing glucose
transporter in these specialized cells [32]. Recent data
Proteins that promote GDP-GTP exchange by members
of the Rab protein family have been characterized in
yeast and mammalian cells [41,42*,43m]. Do these factors regulate the activity of speciiic Rab proteins? In vitro,
yeast Dss4 stimulates GDP dissociation of both Sec4p and
Yptlp [42*]. Mammalian suppressor of Sec4, Mss4 protein, that acts on Sec4p, also stimulates GDP release from
Yptlp and mammalian Rab3a [43’]. Thus, exchange proteins appear to be active on multiple Rab proteins. If Rab
proteins associate with their respective compartments in
the GTP-bound form [ 391, then exchange factors might
function in association with organelle-specific components. In contrast, GTPase-activating proteins (GAPS) appear to be more selective as the recently identified GAP
Cell type specific
615
616
Membranes,
of Ypt6p (GYP6) does not act efficiently 0; other Ypt
proteins [44*].
Association of Rab proteins with membranes requires
gemnylgeranyl groups on the carboxyl-terminal cysteine
motif [45,46]. This process is reversed by a cytosolic
factor, Rab GDI, originally puriiied as a GDP dissociation inhllitor for Rab3a 1471. Rab GDI is able to
recognize several geranylgeranylated Rab proteins and
extract them from membranes in vitro [48-521. These
combined properties suggest that Rab GDI could function to remove Rab proteins from membranes after GTP
hydrolysis and to return them to the donor membrane.
As predicted from such a function, cytosolic Rab proteins
are found in complex with Rab GDI [50-521 and a small
but significant fraction of Rab GDI is associated with various organelles [50]. Thus, Rab GDI appears to be an
essential component that ensures the cyclical and vectorial function of Rab proteins.
Is the function of GDI regulated? Analysis of the quartet mutation in D. mekznogaster suggests that it might
be. Mutations in the quartet locus are associated with
pleiotropic developmental defects in the larvae and
distinguished by the presence of a basic shift in the
isoelectric point of three abundant proteins, one of
which has been identihed as a Drosopbilu homologue of
Rab GDI [ 531. This suggests that Drosophila Rab GDI is
post-translationally mod&d or demodiiied by the wildtype quartet gene product. Although there is no evidence
that quurtetencodes a kinase, it is interesting to note that
the cytosolic form of mammalian GDI seems to be highly
phosphorylated (J Gruenbetg, personal communication).
Phosphorylation or other post-translational modifications
of Rab GDI might be necessary to inhibit reassociation of
Rab proteins with the target membrane and direct them
to the donor compartment.
Other components may also be involved in the Rab protein cycle. Geranylgeranylation of Rab proteins is catalyzed by geranylgeranyl transferase II, an enzyme that,
in contmst to other known isoprenyl transferases [54],
requires sequences amino-terminal to the target cysteine
motif [55-571. This peculiarity may be due to a third enzyme subunit, component A [ 541. Perhaps component A
recognizes Rab proteins and presents them to component B, the catalytic part of the transferase.
Intriguingly, the gene products of human and mouse
choroideremia (CI-IM), a disease associated with degeneration of the retina and the underlying vasculature, the
choroid, share sequence homology with rat component
A 158,591. Two additional iindings support the possibility that CHM could be a member of a family of A
components. First, in cytosol of lymphoblasts from CHM
patients, geranylgeranylation activity is reduced and isoprenylation of Rab3a is more a&c&d than that of Rabla
[&I. Second, another CHM-like gene, CHML, has been
identiiied [ 611. Interestingly, CHM shares sequence similarity with Rab GDI [62]. This sequence conservation
might reflect the ability of both GDI and component A
to interact with geranylgeranylated Rab proteins. However, unlike Rab GDI, component A can also recognize
the non-prenylated form of Rab proteins, and it interacts
with both the GTP- and the GDP-bound forms [59’].
Therefore, it is thought that component A would be
more likely to act as a chaperone for Rab proteins during
the initial process of membrane association, than to remove F&b proteins from membranes. This function might
be necessary to prevent interactions of the hydrophobic
geranylgeranyl moiety with non-target membranes.
Rab proteins
machinery
and the vesicle
transport
If Rab proteins are implicated in the regulation of
vesicular transport, the real challenge now is to understand how they function. For this task it is important to study the relationship between Rab proteins
and other elements of the transport machinery. Different vesicle fusion processes in mammalian cells and
in yeast require N-ethylmaleimide-sensitive fusion protein (NSF) and the soluble NSF attachment proteins
(SNAPS) [ 1*,2*]. More recently, a search for SNAP receptors (SNARES) has yielded four different factors from
bovine brain, all associated with the synapse [63**,64*].
VAMP/synaptobrevin-2 is found on transport vesicles (vSNARE), while syntaxin A and B are on target membranes
(t-SNARES). It is not clear where the fourth protein,
SNAP-25 (synaptosome-associated protein of 25 kDa) is
localized. These findings have led to the hypothesis that,
in general, specificity in the process of vesicle fusion
with its target compartment might be mediated by the
interaction of a specific v-SNARE with its complementary
t-SNARE through the NSF-SNAPS complex (Fig. 1).
Where do Rab proteins fit into this scheme? The answer
is not there yet. However, recent progress suggests possible connections
between Rab proteins and the vesicle
fusion apparatus. Rabphilin-$4, a protein that interacts
with Rab3a, has recently been puriiied and characterized [65,66*]. Structural analysis of the deduced amino
acid sequence from the corresponding cDNA revealed
high homology with synaptotagmin, a synaptic vesicle
integral membrane protein [67]. However, unlike synaptotagmin, rabphilin3A contains neither a signal peptide
nor a putative transmembrane domain. The precise function of synaptotagmin is unknown, although it has been
postulated to be a regulatory component of calcium-dependent exocytosis in the presynaptic terminal [67,68].
The identification of rabphilin3A provides a potential
link between Pab proteins and t-SNARES.In fact, syntaxin
(a t-SNARE) can be co-immunoprecipitated with synaptotagmin suggesting an association of the two proteins
[67], and rabphilin3A is a synaptotagmin-like molecule.
In addition, the SLK’Z/SEC22 and SLY12IBETl genes,
which are multicopy suppressors of functional loss of
YPTl in S. cerevtie, encode two members of the synaptobrevin (v-SNARE) family [69-711. Therefore, it is possible to envisage interactions between a rabphilin and a
t-SNARE and between a Rab protein and a v-SNARE (Fig.
1). The speciiicity of interactions between V-SNARESand
t-SNARESin a complex with NSF and SNAPSmight thus
require further regulation by a Rab protein. This regula-
Rab
(a)
z
GTPases
in vesicular
transport
Zerial
and
Stenmark
617
lated by GAPS and exchange proteins) control the interactions between Rab proteins and their target molecules?
The synaptotagmin-like protein rabphilin-3A is certainly
an interesting candidate for a Rab target [66*]. Complicated molecular machineries are often regulated by a
number of different GTPases, and Rab proteins are not
the only GTPases implicated in intracellular transport.
Compelling evidence indicates that members of other
subfamilies of GTPases function in regulating different
aspects of membrane traffic, such as vesicle budding
and vesicle coat assembly and disassembly [28,39]. Future studies will reveal whether there are links between
Rab proteins and these other GTPases.
The identification of several tissue- and cell type specific
Rab proteins suggests that the complexity of this fanily of GTPases could still increase, when one considers the high heterogeneity of cell types from different
organs. Learning more about the distribution of Rab
proteins in various cell types will contribute to understanding the relationship between structural features of
organelles and their specialized functions, and between
sub-compartments and transport routes.
Fig. 1. Model for possible
interactions
between
Rab proteins
and
known
components
of the membrane
traffic
machinery.
Proper
docking
and fusion of a vesicle
with its correct
target
membrane
depends
on interactions
between
specific
components
present
on both
compartments.
(a) NSF and
SNAPS interact
with
vSNARE- and t-SNARE-type
molecules
1671. (b) interaction
between
the vesicle-specific
v-SNARE
and its target-specific
t-SNARE
is still
hypothetical.
Cc) The vesicle-specific,
CTP-bound
Rab protein
interacts
with its corresponding
rabphilin,
and Cd) possibly,
to a vSNARE. (e) Rabphilin
might interact
with a t-SNARE
as suggested
by the interaction
of syntaxin
with
synaptotagmin
[671. In this
chain
of connections,
the CTPase
activity
of Rab proteins
could
be utilized
to proof-read
I41 the interaction
between
the correct
pair of SNARES.
tory mechanism
might be used to proof-read the correct
interactions between these components. As rabphilin3A can only recognize the GTP-bound form of Rab3A
[66*], GTP hydrolysis would confer unidirectional@ to
the docking/fusion process.
Conclusions
A comprehensive analysis of the molecules implicated
in membrane trticking is beginning to establish which
components are common to many transport processes,
and which are specific. The data obtained so far suggest
that, whereas NSF and SNAPS belong to the first cathegory, Rab proteins belong to the latter one. Although a
number of studies have demonstrated the role of these
GTPases as regulators of membrane traffic, their precise function remains unknown. Proteins that regulate
the membrane association and nucleotide state of Rab
proteins have now been identified, and further mechanistic studies will undoubtedly focus on three key questions. First, what brings about the organelle-specific localization of Rab proteins? Second, which are their target
molecules? Third, how does the nucleotide state (regu-
Acknowledgements
We thank
Jean
phy, Rob Parton,
suggestions.
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BAILLY E, MCCAFFREVM, TOUCHOT N, ZAHRAOUIA, GOUD
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In this paper, it is shown that mutation of the consensus site for
p34dd kinase in the carboxyl terminus of RaMa prevents its phosphorylation during mitosis and blocks its appearance in the cytosol.
Post-translational modilication of the carboxyl-terminal cysteine motif
of RaMa was shown to be unafTected by phosphorylation, and membrane association was found to re-occur upon dephosphorylation after
mitosis.
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GRUENBERGJ, CIAGUE MJ: Regulation of Intracellular Membrane Transport. Curr @in Cell Biol 1992, 4:593-W.
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Whereas normal Chinese hamster ovaq cells contained only 20% of
their transferring receptors on their surface, this number was raised to
80 % In stable transfectants overexpressing Rab4. Overexpression also
prevented delivery of intemalized transferrin to acidic endosomes, and
a large fraction of internalized transferrin was associated with small,
tubular dusters incapable qf efficient acidI6cation. Overexpression of
Rab4 did not affect initial rates of transfenin internalization, but the
rate of transferrin recycling and a fluid phase marker to the plasma
membrane were increased. The results suggest that Rab4 controls the
rate of recycling, possibly by controlling the rate of transport between
the early endosome and a putative recycling compartment.
16.
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17.
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A S. cerevtiegene encoding Ypt7, a protein with 63 % sequence identity to mammalian Rab7, was cloned. Ypt7 and gab7 have identical effector regions. Gene disruption of Ypt7 had essentially no effect on the
growth properties of yeast ceUs at various temperatures. However, the
Ypt7 null mutants were characterized by a highly fragmented vacuole.
while uptake of the pheromone alpha factor occurred at a normal rate,
its degradation was severely inhibited in the null mutants.
18.
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Cloning of a mouSe cDNA for a novel Rab protein, Rabl7, is reported.
Northern blot analysis revealed that Rab17 is expressed exclusively in
polarized epitheiial cells and is thus the first example of an epitheiiai
ceii speciUc Rab protein. In situ hybridization on murine developing
kidneys showed that rabl7 expression was turned on concomittantIy
with the onset of epitheiiai differentiation. In the adult kidney, Rab17
was found to be associated with the basolaterai plasma membrane and
with apical tubules. Localization data suggest that Rabl7 might be involved in reguiating transcytosis.
36.
HUBER IA, PIMPUKAR S, PARTON RG, VIRTA H, ZERIAL M, SIMONS
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A mutation (Gin79-+Leu) in the Sec4 protein strongiy inhibited its in
trinsic GTPase activity. A partially purified GAP activity stimulated the
hydrolysis rate of the mutant to about 30% of the GAP-stimulated,
wild-type Sec4. The se&Leui’9 allele can function as the oniy copy
of sec4 in yeast cells, but the cells are cold-sensitive, exhibit slowed invertase secretion and accumulate secretory vesicles. It thus appears that
the mutation in Sec4 causes a partial loss of function. This is the best
evidence so far that GTP hydrolysis by a member of the Sec4/ypt/Rab
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41.
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with a molecular mass of 17kDa. It was found to stimulate GDP
dissociation from Sec4p as weU as from Yptlp in vitro. Post-translational modification of Sec4p was not required for its interaction with
Dss4-lp. The activity on Sec4p was about twofold higher than that on
Yptl. Dss4-lp was not active on Rab3a and Ra.52.Disruption of the
wild-type dss4 gene was not lethal. However, synthetic lethality was observed when ds4 disruption was combined with certain ret mutants
showing lethality in the presence of the se&-8 mutation. It is therefore
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Mammalian
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1993, 268: in press.
REGAZZIR, K~KUCHIA, TAXAI Y, WO~IM
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53.
Z.AHNER JE,
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CHENEY CM: A Dnxopbiiu
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A rat-brain cDNA library was expressed in temperature-sensitive se&8
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the mss4 cDNA was isolated. The Mss4 protein has a molecular mass
of 14kDa and has sequence homology with Dss4-lp [41]. Ms.&p stimulated GDP release from Sec4p, and to a lesser extent from Rab3a and
Yptl, but was inactive on Ras2. The mss4 gene is expressed in a variety of tissues, suggesting that its natural target is one or more widely
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Yeast cells were transformed with a yeast genomic library on a multicopy plasmid, and extracts from the individual transformants wem
assayed for their ability to stimulate the GTPase activity of Ypd. A
single, positive colony was isolated, and from this the Cl?6 (GAP of
Ypt6) gene was cloned. The gene product, Gyp6, has no sign&ant
homology to other known proteins, Including other GTPase-activating
proteins. Purified Gyp6 strongly stimuiated the GTPase activity of Ypt6,
and had a weak activity on Ypt7. It did not stimulate the GTPase activ
ity of Yptl, Sec4, YpUA, H-ras or Rab2. Whereas @tideletion mutants
are temperature-sensitive, disruption of gyp6resulted in no phenotypic
alterations.
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1992, 267:394&3945.
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KH~~RAVI-FARR, Ct&tx GJ, ABE K, COX AD, MCLUN T, LUtZ
RJ, S~NENSKYM, DER CJ: Ras (CXXX) and Rab (CCKXC)
Prenylation
Signal Sequences
Distinct.
J Biol C&m 1992.
Are
Unique
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FunctlonaIIy
267:2436324368.
CHAVR~ERP, NIGG EA, ZE~UAI. M: Isoprenylation
Rab Proteins
on StructuraIly
Distinct
Cysteine
Motifs.
Sci 1992, 102:857-865.
57.
P!ZTER M,
58
Cit!ZMERS
FPM, VAN DE POL DJR, VAN KERXHOFFLPM, WtElUNGA
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In
Patients
with Choroideremia
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59.
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SEABRA MC, BROWN MS, SIAUGHTER
(2% SUDHOF TC,
JL
A of Rab
PuriIicatIon
of
Component
of
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GOID!%IN
GeranyIgeranyI
620
Membranes
Transferase: Possible Identitv with the Choroideremia Gene
Product. Cell 1992, 70:104P-1057.
Partial peptide sequences of purllied rat component A of geranylgetanyl
transferase II showed high similarity to the human and mouse CHM
gene products.
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SEABRAMC, BROW MS, GOIDSTEINJL: Retinal Degeneration
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in Choroideremia: Deficiency of Rab Geranylgeranyl Transfemse.
Science
1992,
SNAPS.a- and Y-SNAPS are ubiquitously expressed and act synerglstially, whereas B-SNAP is bralnspeclftc. B-SNAP competes for the same
biding site on SNAP receptors as a-SNAP, but unlike a-SNAP it cannot
restore cell-free Golgl transport in cytosol from secl7 mutant yeast. It
is most abundant in areas of high neuron density and may be involved
in neurosecretion.
65.
259:377+X.
Cytosol prepared from lymphoblasts of patients suiTering from the Xlinked eye disease CHM exhibited reduced ability to geranylgeranylate
RabIa and Rab3a in c&n The ability to geranylgeranylate Rab3a was
more alfected than that of Rabla The results are consistent with the
idea that the CHM gene product may be member of a family of A components of Rab getanylgeranyl transferases.
61.
CREMERsFPM, MOUOY CM, VAN DE POL DJR, VAN DEN HURK
JAJM, BACH I, GEURT~VAN KERNELAHM, ROPERSH-H: An Autosomal Homologue of the Choroideremia Gene Colocalizes
with the Usher Syndrome ‘Qpe II Locus on the Distal Part
of Chromosome lq. Hum Mol Gene1 1992, 1:71-75.
62.
FODOR E, LEE RT, O’DONNELLJJ: Analysis of Choroderemia
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SOMR T, WHI?EHEARTSW, BRUNNERM, ERDJUMEN~-BROMAGE
H, GEROMANOS S, TEMPST P, ROTHMANJE: SNAP Receptors
Implicated in Vesicle Targeting and Fusion. Nahtre 1993,
362:318324.
SNAPSare soluble receptors for the general fusion protein NSF. In this
paper, SNAP receptors (SNARES) from bovine brain were isolated by
the use of al??nity columns containing NSF/SNAP complexes. Remarkably, of the four SNARESidentified in this manner, all are proteins that
have been previously found to be associated with the synapse. One
of them, VAMP/synaptobrevln 2, is a synaptic vesicle protein, whereas
two others, syntaxin A and syntaxln B, are found on the synaptic plasma
membrane. The fourth SNARE, SNAP-25, is also thought to be associated with synaptic vesicles or plasma membrane. The results thus indicate that SNAP/NSF can intemct with SNARESin the vesicular membrane
(V-SNARES)as well as with SNARESin the target membrane (t.SNARFs).
Whereas NSF and SNAPS appear to be universal components of the
vesicle fusion apparatus, it is possible that families of V-SNARESand tSNARESensure speciiic interactions between vesicles and their target
membranes.
64.
Ww
SW, Gatrr IC, BRUNNERM, CIAR( DO, MAYERT,
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B~JHROWSA, ROTHMANJE: SNAP Family of NSF Attachment
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SH~RATAJCI
H, KA~~UCHIK, YAMAGUCHIT, WA~A K, HO~UUCHI
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J
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1992,
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SHIRATAKIH, KAIBUCHI K, SAKODA T, KISHIDA S, YAMAGUCHI
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T, WADA K, MNM, TAKA~ Y: RabphiIin-3A, a Putative
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This paper reports the cloning of a cDNA encoding tabphilin3A
a protein isolated from bovine brain crude membranes by chemical crosslinking to Rab3a. The carboxyl-terminal part of rabphilln3A
shows significant sequence homology to the synaptic vesicle protein
synaptotagmln, and contains two Cz domains like those found in protein klnase C and synaptotagmin. Unlike synaptotagmin, rabphilin3A
does not contain any putative transmembrane region. Rabphliin3A was
found to form a complex with the GTPyS form of Rab3a, but not with
Rab3a-GDP and not with the GTPyS-form of Rabll.
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67.
BENNET MK, SCHELUR RI-I: The Molecular Machinery for
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Sci USA
1993, 90:25592563.
68.
SHOJ~-KAQIY, YOSH~DAA SATO K, HOSHINO T, OGURA A,
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NEWMANAP, SHIMJ, FERRO-NOVICK
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are Members of a Group of Interacting Yeast Genes Required for Transport from the Endoplasmic Reticulum to
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DASCHERC, Osstc R G~uwrtz D, SCHM~I-~HD: Identiiication
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Suppress the Functional Loss of KPTI, a Member of the
RAS Supedamily.
Mol Cell Biol 1991, 11:872+385.
71.
NEWMANAP, GIW J, MANctm P. ROSSI G, UAN JP, FEattoNO~~CKS: SEC22 and SLY2 are Identical. Mol Cell Biol 1992,
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This paper reports the cloning and sequencing of cDNAs encoding a.,
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M Zerial and H Stenmark, European laboratoty of Molecular Biology,
Postfach 10.229, Meyerhofstrasse 1,6900 Heidelberg, Germany.