Review
TRENDS in Cell Biology
Vol.15 No.4 April 2005
The building blocks for basolateral
vesicles in polarized epithelial cells
Heike Fölsch
Dept of Biochemistry, Molecular Biology and Cell Biology, Northwestern University, 2205 Tech Drive, Evanston, IL, 60208-3500, USA
After the discovery of basolateral sorting signals for
polarized delivery in epithelial cells in the early 1990s, it
was only about a decade later that the epithelial-cellspecific sorting adaptor AP-1B was discovered. AP-1B
decodes a subclass of basolateral sorting signals and
localizes to the recycling endosomes as opposed to the
trans-Golgi network, suggesting that this is its major
site of action. Furthermore, AP-1B does not simply select
its cargo but also facilitates the recruitment of the
exocyst complex needed for subsequent fusion with
the plasma membrane. This review discusses our
current knowledge of AP-1B function in cargo sorting
to the basolateral membrane and its impact on our
understanding of the similarities and differences
between AP-1B-minus fibroblasts and AP-1B-positive
epithelial cells.
Epithelial cells have polarized plasma membranes divided
into biochemically distinct apical and basolateral
domains. The apical domain is enriched in glycolipids
and cholesterol and constitutes the lumenal wall of
organs, whereas the basolateral membrane is in contact
with connective tissues. Tight junctions seal epithelial
monolayers and provide diffusion barriers that prevent a
mixing of apical and basolateral membrane components.
Established during embryogenesis, this polarity has to be
maintained throughout the life cycle of every individual
cell in an epithelial monolayer to ensure proper function.
To accomplish this task, epithelial cells developed
mechanisms to either specifically retain or to selectively
deliver proteins to the target membrane [1,2]. Newly
synthesized proteins (i.e. biosynthetic cargo) destined for
apical or basolateral plasma membrane domains, as well
as for endosomes and lysosomes, are transported together
from the endoplasmic reticulum (ER) through the Golgi
complex. Upon leaving the Golgi apparatus, however,
these proteins are sorted away from each other, according
to their final destination [3] (Figure 1). It is believed that
this sorting occurs in the trans-Golgi network (TGN), a
vesicular-tubular compartment adjacent to the transGolgi cisternae [3]. However, some sorting might also
occur in recycling endosomes, as discussed below. Recycling endosomes are defined as a perinuclear compartment
in which internalized transferrin (Tfn) receptors accumulate with time. Furthermore, they seem to be the primary
Corresponding author: Fölsch, H. ([email protected]).
Available online 3 March 2005
site of signal-mediated sorting of internalized transmembrane receptors during recycling [4].
Sorting is controlled by signals encoded in the cargo
proteins themselves and facilitated by cytosolic machineries that recognize these signals [1]. Apical targeting
signals often comprise N- and O-linked carbohydrate
chains attached to the ectodomain of proteins, and some
apical sorting signals might be localized in transmembrane domains. Another class of apical targeting determinants are glycosylphosphatidylinositol (GPI) anchors,
which direct the modified proteins to the membranes. In
general, it is believed that apical cargo proteins partition
into glycolipid membrane rafts that facilitate apical
delivery either through lipid–lipid or lipid–protein interactions [5]. The exact mechanism of this pathway,
Apical PM
TJ
Apical path
Endosome
or
lysosome
BL path
gi
Gol
Basolateral PM
TRENDS in Cell Biology
Figure 1. Schematic representation of the different sorting pathways that exist in
polarized epithelial cells. Newly synthesized transmembrane proteins move
together from the endoplasmic reticulum to the Golgi and through the Golgi
apparatus. Upon leaving the Golgi they are sorted away from each other and
delivered to endosomes or lysosomes (blue pathway), the apical plasma
membrane (purple) or the basolateral plasma membrane (green). Sorting to
endosomes, lysosomes or the basolateral membrane involves cytosolic heterotetrameric complexes of the clathrin adaptor protein family (Table 1). Abbreviations: BL, basolateral; PM, plasma membrane; TJ, tight junction.
www.sciencedirect.com 0962-8924/$ - see front matter Q 2005 Elsevier Ltd. All rights reserved. doi:10.1016/j.tcb.2005.02.006
Review
TRENDS in Cell Biology
Vol.15 No.4 April 2005
223
Table 1. Subunit composition of heterotetrameric clathrin adaptor complexes
Adaptor
complex
AP-1A
Subunits
Function
g, b1, m1A, s1
Clathrin binding
motif in b-subunit
Yes
AP-1B
Endosomal sorting
Ubiquitous
g, b1, m1B, s1
Yes
Basolateral sorting
Polarized epithelial cells
AP-2
a, b2, m2, s2
Yes
Endocytosis
Ubiquitous
AP-3A
d, b3A, m3A, s3
Yes
Endosomal sorting
Ubiquitous
AP-3B
d, b3B, m3B, s3
Yes
Synaptic vesicle formation
Neurons
AP-4
3, b4, m4, s4
No
Endosomal and/or basolateral sorting
Ubiquitous
however, is still poorly understood, and putative apical
sorting adaptors remain elusive.
In contrast to apical targeting determinants, targeting
signals for basolateral sorting are in most cases encoded in
the cytoplasmic tails of proteins and frequently contain
either crucial tyrosine or dileucine residues. Basolateral
targeting signals therefore resemble those that direct
proteins to lysosomes, and in general are cis-dominant
over apical targeting information (i.e. if the crucial
tyrosine residue in a basolateral targeting motif is
mutated, the resulting protein is often sorted to the apical
membrane) [6]. Another class of signals decoded in the
cytoplasmic tails of proteins are PDZ-interacting motifs.
These motifs interact with PDZ domains in other proteins,
are involved in apical and basolateral targeting and
serve most likely as retention signals at the respective
membrane [5].
Recently, much progress has been made furthering our
understanding of basolateral sorting. Elucidation of the
molecules that regulate apical membrane trafficking has
been less progressive. Therefore, this review mainly
focuses on recent advances in understanding the basolateral sorting of transmembrane proteins. This will
include a discussion of how cargo proteins might be
selected into nascent basolateral vesicles and how the
appropriate adaptor complex in turn might help to
orchestrate the recruitment of regulatory factors needed
for vesicle formation and subsequent vesicle fusion. The
sorting site of biosynthetic cargo – the TGN or recycling
endosomes – is currently a matter of debate, and we will
discuss recent evidence perhaps leading to a shift in favor
of recycling endosomes. Because a major sorting adaptor
for basolateral secretion, AP-1B, is only expressed in
epithelial cells and not in nonpolarized tissues, insights
into this unique pathway will help us not only to understand cell polarization in greater detail but also will
allow us to appreciate how fibroblasts differ from epithelial cells – this will ultimately help us to elucidate how
individual cells and tissues in the body can perform their
unique tasks.
Adaptors for basolateral sorting
To ensure proper sorting, the respective proteins need to
provide an address tag (in this case, a tyrosine- or
dileucine-based sorting signal), and these signals need to
be recognized by specific cytosolic sorting adaptors.
Typically, linear peptide signals are recognized by cytosolic complexes of the clathrin adaptor protein (AP) family
[7]. These complexes are heterotetrameric and comprise
two large subunits (g, a, d or 3 and b1–b4), one medium
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Expression pattern
(m1–m4) and one small subunit (s1–s2). There are four
major species, AP-1 (g, b1, m1, s1), AP-2 (a, b2, m2,
s2), AP-3 (d, b3, m3, s3) and AP-4 (3, b4, m4, s4) (Table 1).
AP-1, AP-3 and AP-4 facilitate sorting at the TGN or
endosomes, and AP-2 acts at the plasma membrane to
mediate internalization [7]. Whereas the large subunits
interact with various accessory proteins (discussed below),
the medium subunits interact directly with tyrosine-based
sorting signals and therefore are responsible for cargo
recognition [8].
In many cases, tyrosine-based basolateral sorting
signals are recognized by an epithelial-specific variant of
AP-1, AP-1B, which is not expressed in fibroblasts and
other nonpolarized cells types [9]. The ubiquitously
expressed AP-1 (now also called AP-1A) and AP-1B are
identical except for the medium subunits m1A or m1B,
respectively. m1A and m1B are nearly 80% identical [9] and
are coexpressed in all m1B-positive epithelial cells. Despite
their close homology, AP-1A and AP-1B form distinct
vesicle populations and have separate functions [10]. The
role of AP-1B first became evident from studies with
m1B-deficient LLC-PK1 kidney cells. In these cells,
basolateral proteins with tyrosine-based sorting signals,
such as low-density lipoprotein (LDL) or Tfn receptors
that, normally, interact with AP-1B, are missorted to
the apical domain. This sorting phenotype could be
‘corrected’ by exogenous expression of m1B [11]. Although
the importance of AP-1B in basolateral targeting is
clear, it is also evident that there are alternative
AP-1B-independent pathways to the basolateral surface
(Table 2). For example, FcII-B2 receptors, which recognize
antibodies for internalization, have dileucine signals and
are sorted basolaterally even in the absence of AP-1B [12].
Recently, AP-4 has been implicated in basolateral sorting
as well, because a knockdown of the medium subunit of
the AP-4 complex in MDCK cells resulted in a moderate
missorting phenotype of LDL receptors [13]. At present
Table 2. Confirmed AP-1B-dependent or AP-1B-independent
basolateral proteins
AP-1B
dependency
AP-1B-dependent
AP-1B-independent
Signal
Protein
Refs
Y-based
LDL receptor
Tfn receptor
AGPR-HI
VSVG
ErbB2
FcII B2 receptor
E-cadherin
CD147 (EMMPRIN)
H/K-ATPase b-subunit
NaC/KC-ATPase
[11]
[11]
[58]
[10]
[59]
[12]
[60]
[61]
[62]
[11]
Other
LL-based
L-based
Y-based
Other
224
Review
TRENDS in Cell Biology
it is not clear how AP-1B and AP-4 might work together
in basolateral secretion. Another aspect that needs
clarification is exactly where – at the TGN or at recycling
endosomes – in the basolateral pathway these different
adaptors might exert their functions.
Putative involvement of recycling endosomes in
basolateral sorting of biosynthetic cargo
Plasma membrane receptors are typically sorted multiple
times during their lifetime. It is believed that the initial
sorting occurs in the TGN for apical or basolateral
membrane delivery. This hypothesis was based on numerous observations consistent with the segregation of cargo
upon exit from the TGN [14–18]. By contrast, in a recent
study it was claimed that apical (GPI-anchored proteins)
and basolateral (VSVG) cargos can leave the TGN in the
same transport carriers in MDCK cells [19], thus questioning the sorting function of the TGN for plasma
membrane cargo. Nevertheless, none of these experiments
could truly discriminate between a direct pathway from
the TGN to the plasma membrane and pathways that
involve travel through endosomal populations, and the
conclusion that biosynthetic cargo did not travel through
endosomes was based mainly on negative evidence
obtained in nonpolarized cells [17,20].
After arrival at the plasma membrane, many receptors
Vol.15 No.4 April 2005
are readily internalized and reach a perinuclear compartment defined as recycling endosomes, from which they are
resorted for membrane delivery. Often, the same signals
used for targeted insertion of plasma membrane proteins
along the biosynthetic pathway are reused to faithfully
redeliver internalized receptors to the correct target
membrane [21]. As AP-1B is localized primarily at the
recycling endosomes [10,22], the question arises whether
AP-1B is only involved in recycling of internalized cargo or
possibly has an additional function in sorting of newly
synthesized cargo proteins [23]. This might be the case if
AP-1B cargo could traverse the recycling endosomes
during transport from the Golgi to the plasma membrane.
This might indeed be the case for at least some basolateral
membrane proteins. For example, although localized at
recycling endosomes, mutant alleles of Rab8 lead to
missorting of biosynthetic AP-1B cargo proteins [24].
Furthermore, recycling endosomes can be a sorting station
for newly synthesized VSVG [25], and it has been
suggested that biosynthetic polymeric IgA receptor can
travel through endosomes to reach the surface as well [26].
Therefore AP-1B positioned at the recycling endosomes
might be involved in sorting of both internalized and
biosynthetic cargo (Figure 2).
Alternatively, some cargo proteins might move directly
from the TGN to the basolateral plasma membrane if they
Apical path
AP-1B
Exocyst
fusion site
Sec6/8
Exo70
PKD?
Motor
protein?
AP-3/4?
TGN
4?
-3/
AP
Regulators:
Rab8, Cdc42,
RalA
Recycling endosomes
Apical cargo
Basolateral cargo
TRENDS in Cell Biology
Figure 2. Schematic illustration of events that are involved in building AP-1B vesicles at the recycling endosomes. Upon exit from the trans-Golgi network (TGN), proteins
destined for the plasma membrane can move directly or indirectly via recycling endosomes to the apical or basolateral membrane. TGN exit might be regulated by protein
kinase D (PKD) and facilitated by AP-3 or AP-4. Exit of basolateral cargo from the recycling endosomes might be dependent on AP-1B and the small GTPases Rab8 and Cdc42.
Transport of the vesicles towards the basolateral membrane might be aided by actin or microtubule-based motors. Fusion is dependent on the exocyst and probably the
small GTPase RalA (see main text for details). Proteins involved mainly in basolateral secretion are shown in green, and proteins that might control both apical and
basolateral pathways are shown in black. Finally, AP-3 and AP-4 are depicted in blue because they also have a function in lysosomal targeting in addition to their putative
involvement in basolateral secretion.
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TRENDS in Cell Biology
are able to interact with other adaptor proteins. Indeed,
such a scenario has been suggested for LDL receptors [22].
In this case, AP-4 might sort LDL receptors at the TGN for
delivery to the basolateral membrane and, only after
internalization, might this protein become dependent on
AP-1B for recycling back to the basolateral membrane
(Figure 2) [11,13,22]. Another alternative sorting adaptor
acting at the TGN for direct basolateral delivery might be
AP-3. This might or might not involve clathrin. It should
be noted that, although the b-subunit of AP-3 has a
clathrin-binding motif, the involvement of clathrin in
forming AP-3 vesicles is still a matter for debate [7]. So far,
AP-3 has only been shown to play a role in surface delivery
of VSVG in fibroblasts [27], and its involvement in
basolateral sorting is unclear. Furthermore, it is well
established that AP-3 is involved in sorting of cargos to
endosomes or lysosomes [7]. So perhaps AP-3 sorts cargo
proteins such as VSVG from the TGN to recycling
endosomes from where they transit to the surface,
perhaps in clathrin-coated vesicles [10]. As the clathrin
adaptor complexes have some overlapping signalrecognition capabilities [28], the fate of any given receptor
leaving the TGN destined for the plasma membrane might
be decided by which adaptors are available and how
strongly they interact with a given cargo protein. Clearly,
for a better definition of the role of AP-1B in basolateral
delivery, we need to elucidate how AP-1B differs from
AP-1A and the other adaptor complexes.
AP-1A and AP-1B – similarities and differences
The fact that AP-1A and AP-1B have largely nonoverlapping functions raises the question of how cells
discriminate between the two adaptors. As AP-1A and
AP-1B differ only in the composition of the medium
subunits, physiological differences between both adaptor
complexes must be due to m1A or m1B function. Recently,
the crystal structures of the AP-2 and AP-1A core
complexes, composed of the small, the medium and the
two large subunits without their C-terminal extensions,
were solved [29,30]. It is now apparent that the C-termini
of the medium subunits extend away from the adaptor
complexes to accommodate cargo recognition and binding
during vesicle formation (Figure 3) [29,30]. In addition to
cargo binding, the C-terminus of m2 binds to phosphatidylinositol (4,5)-bisphosphate [PtdIns(4,5)P2] at the plasma
membrane. Helen Yin and colleagues showed recently
that the recruitment of AP-1A to the TGN was dependent
on phosphatidylinositol (4)-phosphate [PtdIns(4)P] [31],
and m1A is predicted to bind to this lipid [30]. This might
be true for AP-1B as well.
The conformational change in the medium subunits
leading to their extension away from the core complex
seems to be facilitated by a phosphorylation event on the
medium subunit itself (Figure 3). It is thought that, upon
recruitment of AP-1A to TGN membranes through
activated Arf1, AP-1A is phosphorylated and thereby
becomes able to bind to its cargo molecules [32]. After
vesicle formation, it is believed that m1A is dephosphorylated to facilitate uncoating of the vesicles before their
fusion with the target membrane [32]. Since the postulated phosphorylation sites are conserved between m1A
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Vol.15 No.4 April 2005
225
Putative phosphatase
AP-1B
β1
AP-1B
γ µ1B
γ
σ1
σ1
Cytosolic form
β1
P?
µ1B
*
Cargo Lipid binding?
Putative kinase
TRENDS in Cell Biology
Figure 3. Schematic illustration of the conformational changes in m1B during
membrane recruitment of the adaptor complex AP-1B. For the cytosolic form of
AP-1B, it is predicted that the C-terminus of m1B is folded back onto the complex.
Upon membrane recruitment, the C-terminus might become phosphorylated by an
unknown kinase. This phosphorylation is predicted to lead to an extension of the
C-terminus away from the complex, thus enabling it to bind to its target cargo
protein and perhaps to lipids. Release of AP-1B from the membrane might be
facilitated by dephosphorylation of m1B, upon which the C-terminus folds back
against the complex.
and m1B [30,32], a similar mechanism probably regulates
AP-1B.
Another open question is whether AP-1B vesicles
incorporate any of the accessory proteins known to be
associated with AP-1A vesicles. For example, accessory
proteins that bind to the large g subunit of AP-1 have been
discovered. Among these factors are g-synergin and
EpsinR (also named enthoprotin or CLINT) (see [33] for
references). The function of g-synergin is still elusive,
whereas EpsinR is known to bind to PtdIns(4)P, just like
AP-1, and has a suggested function in AP-1A-mediated
vesicle formation [34]. Members of another protein family
– the GGAs – have been shown to interact with g-adaptin
as well. Initially, GGAs were described as monomeric
adaptor proteins that bind to acidic dileucine motifs in the
cytoplasmic tail of mannose 6-phosphate receptors (MPRs)
[35]. However, recently it has been suggested that they
also interact with g-adaptin of AP-1 [36]. Therefore,
perhaps AP-1A, g-synergin, EpsinR and the GGAs act
together at the TGN to build clathrin-coated vesicles.
As m1B is only expressed in epithelial cells, and most
studies with AP-1 effector proteins or co-adaptors thus far
have been performed in fibroblasts, it remains to be shown
whether they can also work together with AP-1B, or
whether AP-1B might have its own unique set of accessory
proteins. One possible way to ensure different sets of
accessory proteins for AP-1B might be to spatially
separate the location of AP-1B in cells from that of AP-1A
and other adaptor complexes. Indeed, as discussed above,
AP-1A and AP-1B do not colocalize. Instead, AP-1A
localizes to the TGN, whereas AP-1B primarily localizes
to the recycling endosomes [10,22,37]. Presently, it is
unclear why AP-1A and AP-1B show these different
localizations. Perhaps specific ‘docking’ factors of the
synaptotagmin family are involved. For example, m2, the
medium subunit of AP-2, has been implicated in binding to
synaptotagmin at the plasma membrane [38]. However, a
similar role for synaptotagmin family members in the
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TRENDS in Cell Biology
recruitment of adaptor complexes to internal membranes
has not been demonstrated. Alternatively, proteins
involved in AP-1B recruitment or AP-1B vesicle formation
might determine localization, perhaps by stabilizing AP-1B
on specific membranes. Indeed, proteins known to regulate basolateral targeting often colocalize with AP-1B at
the recycling endosomes [10,24].
Regulation of basolateral sorting
Like all membrane trafficking events, basolateral
secretion has to be tightly regulated, spatially as well as
temporally. For example, fission of vesicles should only
occur after the selection of cargo into nascent vesicles is
complete. Recently, Yeaman et al. reported that this step
might be regulated by the serine/threonine protein
kinase D (PKD) [39]. Subsequently, vesicles destined for
the basolateral membrane might move efficiently to the
cell surface on cytoskeletal tracks. Indeed, the actin-based
motor myosin IIA has been implicated in basolateral
secretion [40]. In addition, Kif13A, a plus-end-directed
kinesin motor protein, has been implicated in moving
MPR-containing AP-1 vesicles along microtubules in
m1B-positive MDCK cells [41]. However, so far there is no
direct evidence that Kif13A moves MPR–AP-1B vesicles in
addition to MPR–AP-1A. At the basolateral membrane,
vesicle fusion might depend on the basolateral t-SNARE
syntaxin 4 [2], a still elusive v-SNARE and the exocyst
complex (Figure 2).
The exocyst, an eight-subunit complex, was first discovered in yeast and later shown to be important for
basolateral targeting of LDL receptors [14]. As Sec6 and
Sec8 are localized laterally below tight junctions, it has
been suggested that the exocyst specifies the location at
the plasma membrane to which basolateral vesicles are
delivered [42,43]. Indeed, fusion activity is observed only
in the upper third of the basolateral membrane domain,
immediately below the tight junctions [44]. Interestingly,
membrane recruitment of the exocyst seems to be
enhanced by AP-1B expression in epithelial cells, and
AP-1B and exocyst subunits colocalize on membranes [10].
These data implicate AP-1B vesicles in utilizing the
exocyst for fusion with the basolateral membrane (Figure 2).
Assembly of the exocyst complex at the fusion site
might be regulated by RalA, a small GTPase that seems to
directly interact with the exocyst subunits Sec5 and Exo84
[45,46] (see Box 1 for explanation of the GTPase cycle).
Other small GTPases involved in basolateral secretion are
Rab8 and Cdc42 [24,47,48]. Both of these specifically
Box 1. Activation of small GTPases
Rho, Cdc42, Arf and Rab proteins belong to the superfamily of
Ras-like GTPases. They all cycle between an inactive GDP-bound
form and an active GTP-bound form. In the GDP form, the GTPases
are typically bound to a GDP-dissociation inhibitor (GDI) in the
cytosol. Membrane recruitment involves GDI-displacement factors
(GDFs) at the target membrane and GDP–GTP exchange factors
(GEFs) that activate the GTPase. In the GTP-bound form, the
GTPases interact with various effector molecules at the membranes.
The cycle closes when a GTPase-activating protein (GAP) binds and
stimulates GTP hydrolysis. Subsequently, the GDP form is extracted
from the membrane by GDI [63].
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Vol.15 No.4 April 2005
regulate basolateral delivery of AP-1B cargo [24]. Despite
their clear involvement in basolateral secretion, the exact
mechanisms by which Rab8 and Cdc42 control the AP-1B
pathway are still elusive. Rab8 might be involved in
exocyst or AP-1B recruitment [24], whereas Cdc42 might
act through alterations of the actin cytoskeleton [47,48] or
other activities. For example, in yeast, Cdc42 seems to
interact directly with the exocyst [49]. A challenge for the
future will be to elucidate the sequential action of the
GTPases involved in basolateral secretion and how they
work together to ensure correct protein delivery.
In summary, after selecting its cargo proteins, AP-1B
might facilitate the recruitment of exocyst subunits
and clathrin onto the nascent vesicles for the generation
of clathrin-coated vesicles at the recycling endosomes
[10,22,24,50]. This might be regulated by any of the
aforementioned GTPases (Figure 2).
‘Polarized’ sorting in epithelial cells and fibroblasts
The discovery of AP-1B as an adaptor complex for
basolateral protein sorting in epithelial cells raises two
important questions. First, how is it possible for regulators of membrane trafficking that are ubiquitously
expressed to specifically regulate the AP-1B pathway in
epithelial cells, and, second, what happens to AP-1B cargo
in nonpolarized cells? For example, many cell biologists
use a temperature-sensitive mutant of VSVG to analyze
membrane trafficking events. Over the years, it was
generally assumed that the sorting of VSVG (and other
basolateral transmembrane proteins) to the plasma
membrane is more or less the same in fibroblasts and
epithelial cells. These assumptions were based on early
studies showing that apical cargo was sorted into vesicles
after Golgi exit that are different to those used by basolateral
proteins [14–18]. However, it is now also clear that
basolateral sorting of VSVG is dependent on AP-1B,
and this cargo is packaged into clathrin-coated vesicles
after exit from the Golgi only when AP-1B is present [10].
Therefore, although fibroblasts have some sorting function
for plasma membrane proteins, m1B-positive epithelial cells
have in addition the AP-1B sorting system.
As AP-1B works together with some effector proteins
otherwise known for their involvement in cell migration, it
seems as if AP-1B might ‘hitchhike’ on a secretory
pathway that m1B-minus cells might use for polarized
exocytosis events at their leading edges, a process that
might involve recycling of membrane and proteins from
recycling endosomes [51]. Interestingly, two GTPases,
Rab11 and Arf6, involved in recycling can interact with
exocyst subunits [52,53]. This raises the possibility that
the exocyst is involved in polarized secretion during cell
migration [54]. So it appears that polarized secretion at
the leading edge of fibroblasts and basolateral protein
sorting in epithelial cells use similar proteins, with the
major exception of AP-1B. How then did AP-1B learn to
utilize preexisting machineries? Perhaps this is mediated
by regulators such as Rab8 that are more abundant in
epithelial cells as well [55]. Another possibility is that
some GTPase effector proteins are more abundant in
epithelial cells than in fibroblasts. Indeed, members of a
family of Rab11-interacting proteins, FIPs, show
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TRENDS in Cell Biology
differential expression patterns, with some of them being
more abundant in kidney than in other tissues, and this
might determine which downstream actions take place
(Prekeris, R., pers. commun.; [56,57]). For example,
although Rab11 has been implicated in Sec15 binding
[52], this GTPase seems to regulate apical, rather than
basolateral, sorting [56]. Perhaps a kidney-specific FIP
with a high affinity for Rab11 successfully competes with
Sec15. Alternatively, Rab11 might play a dual role in both
apical and basolateral sorting.
Taken together, it is possible that fibroblasts might
form ‘AP-1B-like’ vesicles upon stimulation of, for
example, Cdc42, and the same might be true for other
polarized cells such as neurons or hepatocytes that are
m1B-minus [9]. However, we have to be cautious in our
assumption that anything that might be true for these
types of vesicles will also apply to AP-1B vesicles as well.
Concluding remarks
Even though there has been much progress in furthering
our understanding of basolateral sorting of cargo that
interacts with AP-1B, we still know little about the
basolateral sorting of AP-1B-independent cargo. Perhaps
AP-1A, AP-3 or AP-4 plays some role in sorting this cargo
to endosomes and thus takes it away from apical sorting at
the TGN, but at present this is mere speculation. Different
from AP-1A, AP-3 and AP-4, AP-1B clearly localizes to
recycling endosomes as opposed to the TGN, and there is
some dispute regarding whether AP-1B plays a role in
sorting biosynthetic cargo. However, the role of AP-1B in
basolateral recycling of internalized receptors is undisputed. The knowledge that AP-1B vesicles incorporate
exocyst subunits, and the indication that Rab8, RalA and
Cdc42 might be important regulators of AP-1B vesicle
formation, is leading us towards a more molecular understanding of an important aspect of basolateral sorting. In
the future, it will be important to learn how these proteins
interact with one another to orchestrate vesicle formation,
transport to the fusion site and vesicle fusion. Do we know
all the proteins involved in AP-1B-dependent sorting?
Most certainly not; for example it is presently unknown
which v-SNARE might be incorporated into AP-1B
vesicles and how this is achieved. Nevertheless, with our
current knowledge of AP-1B, we are already starting to
appreciate the molecular differences between distinct
basolateral sorting pathways, perhaps acting at distinct
intracellular sites. Furthermore, similarities and differences between exocytosis in fibroblasts and polarized
epithelial cells are becoming more apparent. Our ability to
discriminate most notably the differences will ultimately
lead to a deeper understanding of how various cells and
tissues in the body fulfil their unique tasks.
Acknowledgements
I apologize to investigators whose work is not cited owing to space
constraints.
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