Transitions Between Front–Rear and Apical – Basal Polarity

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Remodeling Epithelial Cell Organization:
Transitions Between Front–Rear and
Apical–Basal Polarity
W.JamesNelson
DepartmentsofBiologyandMolecularandCellularPhysiology,StanfordUniversity,
Stanford,California94305
Correspondence:[email protected]
Polarized epithelial cells have a distinctive apical–basal axis of polarity for vectorial
transport of ions and solutes across the epithelium. In contrast, migratory mesenchymal
cells have afront–rearaxis of polarity. During development, mesenchymal cells convert
toepitheliabycoalescingintoaggregatesthatundergoepithelialdifferentiation.Signaling
networks and protein complexes comprising Rho family GTPases, polarity complexes
(Crumbs, PAR, and Scribble), and their downstream effectors, including the cytoskeleton
and the endocytic and exocytic vesicle trafficking pathways, together regulate the distri-
butions of plasma membrane and cytoskeletal proteins between front–rear and apical–
basal polarity. The challenge is to understand how these regulators and effectors are
adaptedtoregulatesymmetrybreakingprocessesthatgeneratecellpolaritiesthatarespe-
cializedfordifferentcellularactivitiesandfunctions.
THEBASICDESIGNOFPOLARIZED This conserved function of polarized epi-
EPITHELIALCELLS thelial cells requires that membrane proteins
aresortedandretainedinthecorrectapicalor
Epithelial cells are distinguished from other
basolateral membrane domain (Fig. 1B). A
celltypesbytheirorganizationintoadherent
majorsortingsitefornewlysynthesizedplasma-
groupsofcellsthatpartitiontheorganisminto
membraneproteinsisthetrans-Golgicomplex
discrete compartments—generally, a special-
(TGN) (the exocytic pathway) (reviewed in
ized internal compartment separated and
Folsch 2008; Mellman and Nelson 2008), but
protected from the external environment by
additional sorting events between the TGN
the epithelium. This organization provides a
and plasma membrane domains (Gravotta
number of unique physiological properties.
et al. 2007) and different membrane domains
Most importantly, aclosed epithelium enables
(transcytosis) (Casanova et al. 1990) occur in
the regulated exchange of nutrients and waste
the endocytic pathway (Fig. 1B). Some vesicle
betweentheinternalandexternalenvironments
trafficking between the TGN and the plasma
(Fig.1A)(reviewedinCereijidoetal.2004).
Editors:BruceBowermanandRongLi
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W.J.Nelson
A Functional apical–basal polarity B Structural apical–basal polarity
Primary
2Cl– Na+ cilium Apical
Crab gill APICAL membrane
cuticle
Mammalian Na+ 2Cl– Na+ K+ Apical Syn3 Bboadsyal
kidney AJC Endo
tubule lumen
K+ AJC Golgi
(Tight junction) Syn4
exocyst
Na+ K+ N
ATP
Basal
Na+ Cl– K+ Microtubules + Bmaesmoblartaenreal
+
+ + + +
BASAL
ECM
Figure1.Functionalandstructuralorganizationofpolarizedepithelia.(A)Functionalapical–basalpolarity.
Physiological studies of transporting epithelia across the phyla (e.g., crab gill and mammalian kidney
nephron)hasrevealedaremarkableconservationinthedistributionofionchannels(Clchannel,Kchannel)
transporters (Na,K,2Cl transporter) and pumps (Na/K-ATPase) between the apical and basolateral plasma
membranedomains.Thepolarizeddistributionoftheseproteinsgeneratesanapical–basalsodiumgradient
that is used to move other ions and solutes across the epithelium. (Redrawn and adapted from Cereijido
et al. 2004.) (B) Structural apical–basal polarity. Polarized epithelial cells have a distinctive apical–basal
polarity in the orientation of cell–cell and cell–extracellular matrix (ECM). Major structures of these cells
are also organized in the apical–basal axis: The organization of plasma membrane domains (apical and
basolateral), junctional complexes (APC, apical junctional complex), the centrosome (basal body),
microtubulesandprimarycilium,andthesecretorypathway(Golgi).Fordetails,seetext.
membrane occurs along microtubules (Jaulin solutes (Fig. 1A) (reviewed in Shin et al.
etal.2007;Lafontetal.1994),which,compared 2006).Second,severalclassesofmembranepro-
with migratory cells (see Fig. 2A), have an teinsbindtoacytoplasmicscaffoldcomplexof
unusual organization in polarized epithelial ankyrin–spectrinonthebasolateralmembrane,
cells (Bacallao et al. 1989) (Fig. 1B). Finally, including ion transporters and channels (re-
the docking and fusion of vesicles with the viewedinBennettandHealy2008).Theseinter-
correct membrane domain requires specific actions are important in regulating membrane
vesicle tethering (exocyst) and SNARE com- protein trafficking, and retention in different
plexes (Fig. 1B) (reviewed in Mellman and membrane domains (reviewed in Bennett and
Nelson2008). Healy2008).
Several mechanisms maintain the distri-
butions of proteinstothe apical or basolateral
EvolutionandDevelopmentalOriginsof
membrane domain. First, the tight junction
PolarizedEpithelia
acts as a molecular fence at the boundary
between the apical and basolateral membrane Epitheliaaroseatthetimeoftheemergenceof
domains (apical junctional complex [AJC]) primitive metazoans in Precambrian times,
(Fig. 1B) to prevent the free diffusion of pro- approximately 600 million years ago. The
teins from one domain to the other, and a evolution of mesenchymal cells in bilaterians
gate to the paracellular diffusion of ions and enabled a further diversification in the
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RemodelingEpithelialCellOrganization
A Front–rear polarity overview B Front–rear polarity signaling pathways
Rac1/Cdc42
Extracellular cues
RhoA Actin assembly
Leading edge Actin Scribble
Actomyosin
assembly
contraction + end MT Cdc42
Uropod Centrosome orientation Scribble/DLG
PAR complex Exocyst
Integrin N aPKC Rac1
PAR3/6
Golgi GSK3b aPKC
Ptdlns(3,4)P Integrin Exocytosis
2
TIAM-1 Numb
Ptdlns(3,4,5)P
3 bPIX1/GT1 GSK3 b
Rear Front
Scribble
LGL Integrin
endocytosis
APC
DLG
MT/centrosome
polarity
Figure 2. Functional and structural organization of migratory mesenchymal cells. (A) Front–rear polarity
overview.Migratorycellshaveadistinctiveorganizationbetweenthefrontofthecell(leadingedge)andrear
(uropod). Rho family GTPase activities have different functions in inducing actin polymerization at the
leadingedge(Cdc42,Rac1),andactomyosincontractionattherear(RhoA).Thecentrosome,microtubules,
andthesecretorypathway(Golgi)areorientedtowardthefrontofthecell.Endocyticandexocyticpathways
are also oriented in the rear–front orientation, which allows internalization of integrins from the rear, and
exocytosis and endocytosis of integrins in the front, all of which are required for cell migration.
Phosphatidylinositides have a polarized distribution in the front–rear polarity. Polarity protein complexes
are localized to the front of the cell (Scribble/DLG and PAR/aPKC). (B) Front–rear polarity signaling
pathways.Polarityproteincomplexes(blue)areupstreamanddownstreamofRhosmallGTPases(Rac1and
Cdc42)andtheirdownstreameffectors(yellow)andend-pointeffects(white).Fordetails,seetext.
distribution and organization of epithelial front–rear polarity designed for directional
structures(Baumetal.2008).Unlikeepithelial migration, to an epithelium with an apical–
cells, mesenchymal cells show weak cell–cell basal polarity designed for vectorial ion and
adhesion and are motile, allowing them to solutetransportiscomplex.Signalingpathways
migrate to different sites in the body cavity that initiate this mesenchymal-to-epithelial
where they convert to epithelial cells and con- transition (MET) involve soluble growth
tributetotheformationofsecondaryepithelia. factors and their receptors, and downstream
This provided additional complexity in com- signaling pathways(Baum etal.2008;Dressler
partmentalizationinmetazoansthatleadtothe 2006), and changes in patterns of gene
formation of physiologically different tissues expression(BoyleanddeCaestecker2006;van
andorgans(MagieandMartindale2008). derFlierandClevers2008).However,remodel-
The process of formation of polarized ing from front–rear to apical–basal polarity
epithelia during development is initiated by also involves the reorganization of generic
the coalescence of migratory (mesenchymal structures and cellular processes common to
and epithelial) cells into cell aggregates that allcelltypes,includingthecytoskeleton,endo-
undergo epithelial differentiation. The cellular cytic and exocytic vesicle trafficking pathways,
transformation from migratory cells with a and mechanisms for localizing and retaining
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W.J.Nelson
membrane proteins to different plasma of intracellular pathways required for mem-
membrane domains. Here, I focus on master brane protrusions in axonal growth cones
regulators and signaling pathways that control (Arimura and Kaibuchi 2005), migrating
front–rearpolarityofmigratory mesenchymal neutrophils (Stephens et al. 2008), and other
cells, establishing the basic design of the cells(Ridleyetal.2003).
developmental precursors to epithelia. I then
discuss how pathways and regulators common
Front–RearPolarityofRhoGTPases
to both mesenchymal and epithelial cells are
reused and adapted to regulate the symmetry Thegenerationoffront–rearpolarity ismedi-
breaking processes that lead to the conversion ated by the localized activation of members of
betweenfront–rearandapical–basalpolarity. the Rho family of small GTPases (Li et al.
2008; reviewed in Iden and Collard 2008). In
general, Cdc42 and Rac1 are activated at the
FRONT–REARPOLARITYIN
frontofthecell,whichresultsintherapidand
MIGRATORYCELLS
dynamic assemblyof actin filaments mediated
Migratory mesenchymal cells show front–rear by the Arp2/3 complex and formins (Pollard
polarityrequiredforpersistent,directionalcell 2007), and the capture (stabilization) and
migration (Fig. 2A) (reviewed in Ridley et al. polarized orientation of microtubules by
2003). Many different external cues initiate mDia (Palazzo et al. 2001) and microtubule
front–rear polarity, including growth factors plus-end binding proteins (Akhmanova et al.
and the extracellular matrix. Conversion of 2001; Wen et al. 2004) (reviewed in Li et al.
those cues into directional migration requires 2008)(Fig.2A).Theselocalizedchangesinthe
global changes in cell organization by protein actin and microtubule cytoskeletons generate
complexesandsignaling pathwaysthatcontrol membrane protrusions, dynamic assembly
thecytoskeletonandproteintrafficking. and disassembly of integrin-based contacts
with the extracellular matrix, and forward
movement of the leading edge (Li et al. 2008;
Front–RearPolarityofPhosphatidylinositides
Ridley et al. 2003). RhoA, on the other hand,
A central feature of polarized migrating cells, isactivatedattherearofthecellandpromotes
best studied in neutrophils and Dictyostelium, the assembly and activation of contractile
is the front–rear polarity of different phos- actomyosin networks that are necessary for
phatidylinositides in the plasma membrane detachmentoftherearofthecellfromtheextra-
(Funamoto et al. 2002; Iijima and Devreotes cellular matrix (Fig. 2A) (Ridley et al. 2003).
2002; Wang et al. 2002). Generally, phos- Local activation of these small GTPases affects
phatidylinositide-3,4,5-triphosphate (PtdIns cellorganizationgloballybymaintaininglocal-
(3,4,5)P )isenrichedintheplasmamembrane ized sites of actin polymerization (front) and
3
at the front of the cell, whereas phos- contractions (rear). Inhibition of local Rac1/
phatidylinositide-3,4-bisphosphate (PtdIns Cdc42 or RhoA activity, or disruption of
(3,4)P )isenrichedeverywhereelse,including the polarized organization of the actin and
2
the rear (Fig. 2A). The different distributions microtubule cytoskeletons results in a global
of phosphatidylinositides was thought to be a loss of directional migration by neutrophils
consequence of the spatial activation of phos- (Xuetal.2003).
phoinositide3-kinase(PI3-kinase)atthefront
of the cell, and phosphatase and tensin
RolesofPolarityComplexesinFront–Rear
homolog (PTEN) elsewhere. Although this
FunctionsofRhoGTPaseandCytoskeleton
picture has become more complex (Stephens
et al. 2008), the front–rear distribution of Polarity complexes regulate the cytoskeleton
PtdIns(3,4,5)P and PtdIns(3,4)P are impor- either upstream or downstream of Rho family
3 2
tantingeneratingasymmetry intheactivation GTPases (Fig. 2B) (reviewed in Iden and
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RemodelingEpithelialCellOrganization
Collard2008).Polarityproteinswereoriginally maintainingactiveCdc42atthefront,including
identified in epithelia as protein complexes PI3-kinase,whichconcentratesPtdIns(3,4,5)P
3
involvedinregulatingapical–basalpolarity,but atthefront (RaftopoulouandHall2004),and
theyarealsofound in migrating mesenchymal PAR4/LKB1 (Zhang et al. 2008). Interestingly,
cells. They include three protein complexes, ectopicexpressionofPAR4/LKB1insingleepi-
each defined by its core interacting proteins: thelial cellsresults in a dramatic globalpolari-
the Scribble complex (Scribble, DLG, and zation of the cytoskeleton and the formation
LGL), the PARtition (PAR) complex (PAR3/ of distinct plasma membrane domains in the
PAR6/atypical protein kinase C [aPKC]) and absence of eithercell–cell orcell–extracellular
PAR4/LKB1, and the Crumbs complex matrix adhesion (Baas et al. 2004). Thus, at
(Crumbs, PALS1, and PATJ) (see Fig. 2B) (see least in the context of ectopic expression,
McCaffrey and Macara 2009; Prehoda 2009). PAR4/LKB1 appears to be an example of a
These proteins are generally classified as regulator of autonomous symmetry breaking.
tumor suppressors (reviewed in Bilder 2004), PAR4/LKB1 substrates include another
but how they control cell proliferation is serine/threonine kinase PAR1 and the family
poorly understood (see the end of the article of ELKL-motif kinases (EMKs), also known
forfurtherdiscussion).Nevertheless,itisclear as microtubule-affinity-regulating kinases
thattheyplayessentialrolesincellorganization. (MARKs)(Illenbergeretal.1996).Avarietyof
They are generally localized at the front of studies in fibroblasts, epithelial cells, and
migrating cells (Fig. 2B), although Scribble Drosophila indicate that PAR1 and MARKs
and DLG have also been found at the rear of regulate microtubule remodeling and organi-
migrating T cells (Ludford-Menting et al. zation,andthedeliveryoftransportvesiclesto
2005).Littleisknownaboutplasmamembrane the plasma membrane (Cohen et al. 2004a;
binding sites for Scribble, or other Polarity Cohenetal.2004b;Elbertetal.2005).
complexesatthefront (orrear)ofcells. aPKC, a component of the PAR3/PAR6
Several mechanisms involving Polarity Polarity complex, is a downstream effector of
complexes locally activate Cdc42 and Rac1 at activated Cdc42 (Fig. 2B), although it may be
the leading edge plasma membrane (Fig. 2B). localizedtothefrontofthecellindependently
Scribble, in a complex with bPIX (PAK- of Cdc42 by PATJ (Shin et al. 2007), a com-
interacting exchange factor)—a guanine ponentoftheCrumbsPolaritycomplex.aPKC
exchange factor for Cdc42/Rac1—and GIT1 has a number of substrates critical for the
(G-protein coupled receptor interacting polarization of migrating cells (Fig. 2B). One
protein 1) (Audebert et al. 2004), controls substrate is T-cell-lymphoma invasion and
Cdc42 activation and localization (Osmani metastasis1 (TIAM1) (Nishimura et al. 2005),
et al. 2006). Knockdown of Scribble levels in a GEF that activates Rac1 (Pegtel et al. 2007).
mammalian tissue culture cells also causes a Another substrate is glycogen synthase
decrease in activation of Rac1, indicating that kinase-3b(GSK3b).GSK3bisinactivatedupon
the Scribble complex regulates Rac1 activity aPKC phosphorylation, one result of which is
directly or indirectly (Zhan et al. 2008). that nonphosphorylated APC, normally a
Additional mechanisms regulate Cdc42 and target of active GSK3b, bindsto and stabilizes
Rac1 activation at the leading edge (reviewed microtubules at the plasma membrane
in Arimura and Kaibuchi 2005), including (Zumbrunn et al. 2001). Interestingly, DLG, a
integrin engagement with the extracellular component of the Scribble Polarity complex,
matrix (Etienne-Manneville and Hall 2001), binds APC, thereby localizing DLG to the
Rap1, another Ras family GTPase (Bos 2005), front of migrating cells (Etienne-Manneville
which activates integrins and Cdc42 et al. 2005) (Fig. 2B). Finally, orientation of
(Takahashi et al. 2008), and Rac1 (Gerard microtubules in the front–rear axis affectsthe
et al. 2007) at the front of migrating cells. orientation of the centrosome, also called the
Several feedback loops may be involved in microtubule organization center (Fig. 2A),
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W.J.Nelson
which is critical for maintaining overall cell substrate accessibility) or location may fine
polarity and directional migration (Etienne- tunecellpolarizationandmigration.
MannevilleandHall2003). Several protein complexes that control
vesicle delivery to the plasma membrane are
localized to the front of migrating cells
(Fig. 2B). The complexof Scribble,bPIX, and
Front–RearPolarizationofEndocyticand
GIT1locallyregulatesCaþþ-dependentexocy-
ExocyticMembraneTraffickingPathways
tosis at synapses (Audebert et al. 2004), and
Membrane protein distributions in migratory thedistributionofthiscomplextothefrontof
cellsareorganizedinthefront–rearorientation the cell (Fig. 2B) could play a similar role to
by the exocytic and endocytic pathways promote local transport and vesicle fusion
(Fig. 2A). Local exocytosis of proteins occurs with the leading edge. Another protein in the
atthefrontofmigrationscells(fordiscussion, Scribble Polarity complex, LGL, may also play
see Bretscher 2008). It is unclear, however, if aroleinvesiclefusionattheplasmamembrane.
newly synthesized proteins are also targeted BothmammalianLGL(Muschetal.2002)and
specifically to the front of migrating cells. theyeasthomologSro7/77(Lehmanetal.1999)
Although specific sorting of membrane pro- bindt-SNAREs,anddeletionofSro7/77inyeast
teinsintheexocyticpathwaytoapicalandbaso- (Lehman et al. 1999) or LGL in neuronal cells
lateralmembranedomainsisgenerallythought (Klezovitchetal.2004)resultsininhibitionof
tobeaspecializationofepithelialcellspolarized someproteindeliverytotheplasmamembrane.
in the apical–basal axis (Fig. 1B), fibroblastic The Exocyst complex, which is thought to
cells appear to have a similar capacity tether transport vesiclesto sites of fusionwith
(Yoshimori et al. 1996). Further studies are theplasmamembrane(Wuetal.2008),isalso
required, however, to determine whether localized to the front of migratory cells
proteinsortingintheTGNandseparatedeliv- (Fig.2B)(SpiczkaandYeaman2008;Zuoetal.
ery of different classes of proteins to different 2006). The exocyst interacts with a number of
membranedomainsisimportantforestablish- proteins that could localize it to the leading
ing or maintaining front–rear polarity in edge,includingpaxillin(astructuralandsignal-
responsetoanextracellularcue. ing proteinassociatedwithintegrinfocaladhe-
The redistribution of membrane proteins sions), the bPIX-GIT1 complex (Spiczka and
after delivery to the plasma membrane occurs Yeaman 2008), and Apr1 (a protein in the
via the endocytic pathway. Integrins are an Arp2/3complex)(Zuoetal.2006).Importantly,
important class of proteins that are redistrib- knockdown of exocyst proteins results in
uted from the rear to the front of the cell by decreased delivery of integrins to the front of
the endocytic pathway (Hopkins et al. 1994; the cell and a decreased rate of cell migration
Lawson and Maxfield 1995) (Fig. 2A). The (Spiczka and Yeaman 2008; Zuo et al. 2006),
endocytic adaptor protein Numb (Salcini furthersupportingtheroleofpolarizedtraffick-
et al. 1997), another substrate for aPKC ingofproteinstothefrontofmigratorycells.
(Nishimura and Kaibuchi 2007), is involved In summary, regulation of front–rear
(Fig. 2B). In its nonphosphorylated state, polarity in migratory cells requires localized
Numb binds to and induces the endocytosis activation of different Rho family small
of integrins, and promotes cell migration GTPases at the front (Cdc42, Rac1) and rear
aPKC phosphorylation of Numb inhibits (Rho) of the cell that results in the polarized
Numb binding to integrins, and reduces cell orientation and assembly of the actin and
migration (Nishimura and Kaibuchi 2007). It microtubulecytoskeletons.Polaritycomplexes,
is important to recall that aPKC phos- particularly the Scribble complex and com-
phorylation of other substrates also promotes ponents of the PAR complex, play multiple
cell migration (see the previous discussion), roles in controlling the local distribution and
indicating that control of aPKC activity (or activation of Rac1 and Cdc42 at the front of
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RemodelingEpithelialCellOrganization
thecell,downstreamregulationofcytoskeleton space mediated by polarized distributions of
organizationandassembly,andlocalregulation ion channels and pumps between the apical
oftheendocyticandterminalexocyticmachin- (e.g., CFTR) and basolateral membrane
ery that are required for remodeling of mem- (e.g., Na/K-ATPase) (Ferrari et al. 2008;
braneproteindistributions. Houghton et al. 2003; Jaffe et al. 2008).
Coupledwiththelossofcell–celladhesionon
the forming apical membrane (see #2 above),
APICAL–BASALPOLARITYIN
vectorial pumping of fluid would help to fill
EPITHELIALCELLS
the luminal space; and (4) Repulsion of
Thetransitionfrommigratorycellstotheestab- E-cadherin cell–cell adhesion (e.g., during
lishment of a polarized simple epithelium formation of the heart tube in the Drosophila
requirestheaggregationofcellsthroughspecific embryo) (Santiago-Martinez et al. 2008). In
cell–cell interactions, followed by epithelial this case, inhibition of E-cadherin-mediated
differentiation. In general, the adhesive sur- cell–cell adhesion by Slit and Robo pathways
faces,formedbyinteractionsbetweencellsand inducesformationofaluminalspace.
withtheextracellularmatrix,formthebasolat- Cell adhesion to the extracellular matrix
eral plasma membrane domain. Characteristi- (ECM) is critical for signaling the formation
cally, the apical plasma membrane forms on of an apical membrane and luminal space
the“nonadhesive”surface.Inafewexceptional (reviewed in Bryant and Mostov 2008). In its
cases,however,anonadhesivesurfaceispresent simplest role, ECM adhesion enables cells to
immediately:Forexample, theopensurfaceof distinguish an adhesive surface (attached to
epithelial cell monolayers grown in tissue the ECM) and a nonadhesive surface. For
culture, the outside surface of the preimplan- example, single mammary epithelial cells
tationmammalianembryo,orsyncytialblasto- attached to an ECM (i.e., in the absence of
dermofDrosophila.Theformationofanapical cell–cell adhesion) secrete b-casein from the
surfacewithincellaggregatesismorecomplex. apical surface specifically (Streuli et al. 1995),
and RNA envelope viruses, which normally
bud from the apical (Influenza virus) and
TheFirstOvertEvidenceofApical–Basal
basolateral membrane (Vesicular Stomatitis
Polarity—Orientationofthe
virus, VSV) of polarizedmonolayers of MDCK
Apical/LuminalDomain
cells,butbudfromdifferentsitesontheplasma
Withinanaggregateofcells,aluminalnonad- membrane of single MDCK cells attached
hesive surface is not provided and, therefore, to a substratum (Rodriguez-Boulan and
hastobeformeddenovo.Severalmechanisms Pendergast1980).
appear to generate a nonadhesive surface and The specialized role of the ECM in the de
a lumenwithin an aggregate of cells (reviewed novoformationofthelumen/apicalmembrane
inBryantandMostov2008):(1)Selectiveapo- is important within cell aggregates such as
ptosis of cells in the center of the aggregate three-dimensional MDCK epithelial cysts
(Mailleux et al. 2008) removes cells, leaving (reviewed in Bryant and Mostov 2008).
behindaluminalspace;(2)Insertionofapical Although details of the mechanisms involved
proteins from intracellular stores into the are not fully understood, it appears that the
cell–cell contact, coupled with exclusion of initial step is Rac1-dependent adhesion of
E-cadherin and other basolateral proteins integrinstotheECMproteinlaminin(O’Brien
from that site (Ferrari et al. 2008; Jaffe et al. et al. 2001) (Fig. 3). Subsequently, the asym-
2008; Ojakian et al. 1997). In this case, the metric distribution of PtdIns(3,4,5)P to the
3
lackofcell–celladhesionproteinswouldallow basolateral membrane and PtdIns(3,4)P to
2
formation of a nonadhesive, luminal surface the apical membrane initiates the localization
on cells containing apical membrane proteins; of Cdc42 by annexin II (Martin-Belmonte
(3) Vectorial fluid secretion into the luminal et al. 2007), and the PAR complex to the
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W.J.Nelson
Endocytosis
PALS1 Primary cilium
Crumbs Rich1 Cdc42
AMOT Plasma membrane identity
PATJ PAR
Apical
Crumbs WASP Crumbs3 membrane Crumbs
PALS1 Arp2/3
PAR3 Dynamin Ptdlns(3,4)P
2
E-cadherin PAR3/6
Cdc42 Rac1 aPKC
APC Rho
Exocytosis Exocyst
Basolateral Scribble
Cargo tSNARE Golgi Dynein membrane DLG
ScribbleLGL 5)P3 mRNA Apoptosis ? Hippo
3,4, crusmdtbs bPIX/GIT1
ns( PAR3 Myc Warts
dl max
Pt
Rac1 APC Rac1 DIAP1-apoptosis
Transcription miR-200 DLG +
+ +
Integrin
JNK-apoptosis
E-cadherin
Laminin
Crumbs Snail TGFb
Extracellular cues
LGL/PATJ ZEB HGF
Laminin5
Figure3. Functionaland structuralorganization of polarizedepithelialcells.Center:Phosphatidylinositides
have a distinctive apical–basal polarity; PtdIns(3,4)P is localized in the apical membrane, and
2
PtdIns(3,4,5)P is localized in the basolateral membrane. Signals from the extracellular matrix (laminin)
3
through integrins and Rac1 activity orient cells, and are required for formation of the apical/luminal
domain. Microtubules, which are polarized in the apical–basal axis, are anchored at the basal and lateral
membranes(AJC)byAPCandotherproteincomplexes;microtubule/dynein-dependentdeliveryofmRNAs
totheapicalregionisrequiredforproperlocalizationofCrumbs,Sdt (PALS1),andPAR3.Modules:Polarity
protein complexes regulate several pathways critical for the establishment and maintenance of apical–basal
polarity.Generally,expressionofE-cadherin,Polarityproteincomplexes,andECMproteinsarerequiredto
establish and maintain apical–basal polarity (epithelial program). Extracellular cues, including growth
factors/cytokines (e.g., TGF-b or HGF) and the transcriptional repressors Snail and ZEB1 down-regulate
expressionofthisepithelialprogramcausingthelossofepithelialdifferentiationandcell–celladhesion,and
resulting in a front–rear polarization and cell migration (the transcriptional module). Plasma membrane
module involves mutually antagonistic regulation of the Crumbs (apical domain) and Scribble (basolateral
domain)complexes,andthePARcomplex.TheParcomplexalsolocallyregulatesRac1andRhoattheAJC
(see text for details). Components of the Crumbs (Crumbs3) and PAR complexes are localized to the
primary cilium (the primary cilium module) and regulate global cell polarity. The Crumbs complex (the
endocytosis module) and Scribble complex (the exocytosis module) control membrane protein organization
andstabilityattheAJC(seetextfordetails).Finally,theScribblecomplexappearstoplayaroleinregulating
apoptosis(theapoptosismodule).
apicalregionofthecell(seethefollowing).Itis theapicaldomain.Localizationofthesedifferent
unclear how PtdIns(3,4,5)P and PtdIns(3,4)P phosphatidylinositidestotheapicalandbasolat-
3 2
arelocallysynthesizedandlocalizedtodifferent eral membranes is important in apical–basal
membrane domains. One mechanism may polarityasectopicadditionofeitherphosphati-
involve localization of PTEN, which converts dylinositide to the “wrong” plasma membrane
PtdIns(3,4,5)P to PtdIns(3,4)P , to the PAR domain results in reversal of the apical–basal
3 2
complexattheAJC(vonSteinetal.2005).This polarity of membrane proteins (Martin-
could prevent PtdIns(3,4,5)P accumulation in Belmonteetal.2007).
3
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RemodelingEpithelialCellOrganization
Apical–BasalPolarizationofEndocyticand a template for binding the basal network of
ExocyticMembraneTraffickingPathways microtubules (Reilein and Nelson 2005), and
is associated with microtubules and the adhe-
As the cell generates apical–basal polarity, the
rens junction in some cell types (Hamada
exocytic and endocytic pathways establish a
and Bienz 2002) (Fig. 3). Microtubules may
complexinterrelationshipthatregulatescorrect
also attach indirectly to the cadherin–catenin
sorting of newly synthesized proteins to and
complex (adherens junction) through inter-
between the apical and basolateral membrane
actions with the dynactin complex (Lien et al.
domains.
2008; Ligon and Holzbaur 2007), and
A major sorting site for newly synthesized
p120catenin and kinesin 1 (Chen et al. 2003).
proteins in the exocytic pathway is the TGN
A population of microtubules is also localized
(Fig. 1B). Two pathways have been identified
to the adherens junction through their minus
that sort apical proteins by clustering them
endsbyNezhaandPLEKHA7thatbindtothe
into a microdomain of the TGN from which
cadherin–catenincomplex(Mengetal.2008).
apical transport vesicles form (reviewed in
Docking and fusion of vesicles with the
SchuckandSimons2004):clusteringbyglyco-
correctmembranedomainrequiresspecificves-
lipidraftsenrichedinsphingolipidsandcholes-
icletetheringandSNAREcomplexes(Fig.1B).
terol (Paladino et al. 2007), or by association
The exocyst vesicle-tethering complex is lo-
with galectin-3 oligomers (Delacour et al.
calized to the apex of the lateral membrane
2005). Sorting in the basolateral pathway
domainandspecifiesbasolateralvesicledelivery
involves recognition of diverse cytoplasmic
there(Grindstaffetal.1998).However,therole
sortingsignalsbycytosolicfactors,whichcluster
of the exocyst in vesicle trafficking in apical–
theseproteinsintoaTGNmicro-domainsepa-
basalpolarizedcellsmaybemoremultifaceted.
ratefromthatcontainingapicalproteins.Many
The exocyst has also been localized to apical
of these basolateral signals are either tyrosine-
endosomes where it regulates transcytosis of
ordileucine-basedandarerecognizedbycyto-
membraneproteinsbetweenthebasolateraland
solic adaptor proteins (AP) and clathrin
apicalmembranes(Oztanetal.2007),andatthe
(reviewed in Folsch 2008; Mellman and
apically localized primary cilium (Overgaard
Nelson2008).APsandclathrinarealsoinvolved
et al. 2009). In both cases, the exocyst likely
in sorting proteins in the endocytic pathway
acts as a vesicle-tethering complex, similar to
(Edeling et al. 2006), and several studies have
itsfunctionattheplasmamembrane.
showedtheintersectionofthebasolateralexo-
Different t-SNAREs are localized to the
cytic and endocytic pathways in basolateral
apical (syntaxin 3) and basolateral (syntaxin
proteinsorting(Gravottaetal.2007)andtrans-
4) membranes (Low et al. 1996) (Fig. 3) and
cytosisofproteinsbetweendifferentmembrane
specifythedeliveryofcognatetransportvesicles
domains(Casanovaetal.1990).
withthecorrectmembranedomain.Thisdiffer-
Some of the vesicletrafficking betweenthe
entiallocalizationisrequiredforcorrectvesicle
TGN and plasma membrane, and transcytosis
deliveryasmislocalizationof syntaxin 3tothe
between membrane domains occurs along
basolateral membrane causes apical proteins
microtubules (Jaulin et al. 2007; Lafont et al.
tobemistargetedtothebasolateralmembrane
1994).Microtubulesareorganizedintobundles
(Sharmaetal.2006).
aligned in the apical–basal axis of the cell,
with plus ends at the basal membrane and
Apical–BasalOrganizationandFunctionof
minus ends in the apical cytoplasm (Fig. 1B)
PolarityProteins
(Bacallao et al. 1989). How microtubules are
maintained in this apical–basal organization Adistinctivefeatureofpolarizedepitheliaisthe
is poorly understood. Adenomatous polyposis spatial distribution of intercellular junctional
coli (APC), a microtubule binding protein complexes and Polarity protein complexes
associated with the plasma membrane, forms (Crumbs,PAR,andScribble).Invertebrateand
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W.J.Nelson
invertebrate epithelial cells, the adherensjunc- 2005) and Stardust (Sdt, the Drosophila
tionandthetightjunctionarelocalizedtothe homolog of mammalian PALS1) (Fig. 3)
apexof the lateral membrane (the AJC) at the (Horne-Badovinac and Bilder 2008). Mecha-
boundary between the apical and basolateral nisms involved in localization of the Scribble
plasma membrane domains. In invertebrates, complex to the lateral membrane below the
the functional homolog of the tight junction, AJCareunknown.
the septate junction, is localized on the basal
sideoftheadherensjunction.
DownstreamEffectorsofPolarityProteinsin
Polarityproteinshavedistinctdistributions
Apical–BasalPolarity
attheAJC(Fig.3;theplasmamembraneidentity
module).TheCrumbscomplexislocatedtothe The different distributions of Polarity protein
apical side of the AJC (Tanentzapf and Tepass complexesaroundtheAJCarecriticaltomain-
2003), and at the primary cilium (Fan et al. tainingtheidentityoftheapicalandbasolateral
2004); the PAR complex is localized close to membrane domains (Fig.3; plasmamembrane
the AJC (Bilder et al. 2003; Izumi et al. 1998) identitymodule).GeneticstudiesinDrosophila
and at the primary cilium (Sfakianos et al. showed that the Crumbs complex is required
2007); and the Scribble complex is localized to regulate the formation of the apical mem-
on the lateral membrane below the AJC brane, whereas the Scribble complex regulates
(Bilderetal.2003;TanentzapfandTepass2003). the basolateral membrane; loss-of-function
Several mechanisms regulate the different mutations in the Crumbs complexor Scribble
distributions of these Polarity protein com- complex result in defects in apical–basal
plexes. In mammalian cells, the PAR complex polaritybecauseofalossoftheapicalandbaso-
binds through PAR3 to two cell–cell adhe- lateral membrane domain, respectively (Bilder
sion proteins, junction-associated molecule-A et al. 2003; Tanentzapf and Tepass 2003).
(JAM-A) (Ebnet et al. 2003) and nectin Importantly,lossofeither Crumbs orScribble
(Takekuni et al. 2003), that colocalize with signaling can be compensated by decreased
E-cadherinintheAJC.PAR4/LKB1alsoclosely expressionofScribbleorCrumbs,respectively,
colocalizes with E-cadherin (Sebbagh et al. demonstrating that functions of these com-
2009),whichmaylocallyregulateitsactivation plexes are mutually antagonistic (Bilder et al.
of PAR1 and MARKs, and thereby the organi- 2003;TanentzapfandTepass2003).
zationofmicrotubulesandthedeliveryofves- Despite strong genetic evidence for the
icles to the plasma membrane (Cohen et al. importance of these Polarity complexes in
2004a;Cohenetal.2004b;Elbertetal.2005). apical–basal polarity of epithelial cells, their
In vitro studies indicate that binding be- functions remain poorly understood. This is
tween different PDZ domain-containing pro- in contrast to roles for the PAR complex, par-
teins in the PAR complex (PAR3 and PAR6) ticularly aPKC, and some proteins in the
andCrumbscomplex(PALS1andPATJ)could Scribble complex in front–rear polarity in
be involved in positioning the Crumbs migrating cells (localization and activation of
complex close to the PAR complex in the Rho family small GTPases, orientation of the
region of the AJC (Fig. 3) (Lemmers et al. actin and microtubule cytoskeleton, and
2004; Roh and Margolis 2003). Other studies vesicle trafficking) (see Fig. 2). Are any of
in Drosophila have shown that restriction of these functions important for apical–basal
Crumbs to the apical domain of epithelial polarityinepithelialcells?
cells also requires dynein-dependent basal-to- RhofamilyGTPasesplayanumberofroles
apical transport of Crumbs mRNA along in polarized epithelial cells (Fig. 3; plasma
microtubules and localized translation in the membrane identity module). At the adherens
apical cytoplasm (Li et al. 2008). A similar junction, the activities of Rac and Rho antag-
mechanism maybeimportantin apical locali- onize each other as Rac stabilizes junctions,
zation of PAR3/bazooka (Harris and Peifer whereas Rho induces actomyosin contraction
10 CitethisarticleasColdSpringHarbPerspectBiol2009;1:a000513