Generation and maintenance of epithelial cell polarity

Generation

and maintenance

of epithelial

cell polarity

B. Gumbiner Department

of Pharmacology,

University Current

Epithelial

of California

Opinion

in Cell Biology

polarity

Epithelia are tissues that form selective permeability barriers and regulate the exchange of substances between the external milieu and the internal environment of an organism. In order to perform these functions, epithelial cells are highly polarized. The expression of the polarized state depends not only on the autonomous properties of the individual epithelial cell, but also on interactions between the cells. This review will focus on Endings published in the past year that have begun to reveal the ways in which cell-cell and cell-substrate interactions contribute to the development of the polarized epithelium. The polarity of epithelial cells is manifest at many levels of organization (Fig. 1). Functionally, the most important attribute of epithelial polarity is the division of the plasma membrane into biochemically distinct apical and basolateral domains. The asymmetric distribution of surface enzymes, ion channels and solute transporter proteins endows the epithelium with its primary function: the vectorial transport of substances between luminal and serosal compartments of a tissue. This asymmetry must be maintained despite the continuity of the plasma membrane and the continual internalization, recycling, and turnover of membrane proteins. The process of polarization begins within the intraceUular membrane compartments, as the epithelial cells require a mechanism to sort continually plasma membrane proteins destined to one or other of the two membrane domains [ 11. Epithelial cells are also polarized morphologically. Typically, the apical ceU surface faces an open space or lumen and is frequently covered with microvilli. The basolateral membrane makes extensive contacts, having attachments to the basal lamina at the basal surface and to a junctional complex at the most apical edge of the lateral surface. Normally, all these features of the epithelial ceU operate in concert to give rise to the functional polarized state, although they can be manipulated and observed independently, allowing an experimental dissection of the mechanisms that contribute to the generation and maintenance of the final polarized state.

Generation

versus

maintenance

San Francisco,

of cell polarity

Many epithelial tissues develop from pre-existing polarized sheets of epithelial cells. Imagination and evagina-

San Francisco,

California,

USA

1990, 2:881487

tion of epithelia and symmetrical ceU division occur without disrupting the pre-existing pattern of polarity (Barties and Hubbard, Da, Biol1986,118:286295; Hatta and Takeichi, Nature 1986, 320447-449). There are, however, important events in development in which epithelial polarity seems to arise de nouo from non-polarized ceU types. For example, the renal tubules arise from induced mesenchymal cells that condense and polarize into epithelia [ 21. Perhaps the most significant de nova generation of an epithelium occurs at the earliest stages of mammalian development, when loosely associated blastomeres undergo compaction to form the trophectodermaI epithelium of the blastocyst (Fleming and Johnson, Annu Rev Cell Biol 1988, 4:459-485). In these cases, mechanisms must exist for generating the initial pattern of polarity. Even preformed epithelia must maintain the pattern of polarity in the face of dynamic membrane recycling and protein turnover. This pattern is plastic, however, because it can be reversed by manipulating extracellular cues. Madin-Darby canine kidney (MDCK) cells and thy roid epithelial cells, when grown in suspension, form polarized cysts with their apical membrane domain facing outwards [3,4]. When these cysts are placed in collagen gels they completely reverse polarity within 24 h so that their apical domains face the internal lumena. During reversal, components of the apical membrane do not simply translocate from one surface to the other. Rather, the entire apical domain seems first to be internalized and degraded and later to be resynthesized and assembled de nova at its new position. Therefore, even the long-term maintenance of polarity may involve mechanisms similar to those required for the de novo generation of polarity.

The organization domains

of epithelial

membrane

To maintain compositional differences between the apical and basolateral domains, a mechanism must exist to prevent mixing and randomization of plasma membrane proteins by diffusion in the plane of the membrane. Two mechanisms have been considered: restriction of membrane protein mobility by anchoring to the underlying membrane cytoskeleton [5,6], and the existence of a localized barrier to diffusion, or ‘fence’, at the tight junc-

Abbreviations CPtglycosyl-phosphatidylinositol;

MDCK-Madine-Darby

@ Current

Biology

canine

kidney;

Ltd ISSN 0955-0674

VAC-vacuolar

apical

compartment.

881

882

Cell-to-cell

contact

and extracellular

matrix

.

a)

Apical

(luminal)

(b) Microvilli

Na+

Na

Basolateral

Na

(serosal)

Fig. 1. Three important aspects of epithelial cell polarity. (a) The organization of the plasma membrane into two compositionally distinct domains. In the example shown, the presence of passive Na + channels in the apical domain and an active Na+ pump in the basolateral domain is responsible for the net luminal-to-serosal transport of Na+. (b) Sorting and pathways of intracellular transport of membrane proteins destined for either the apical or the basolateral domain. Sorting can occur either in the Golgi apparatus (left) or in the endosome after initial basolateral delivery and subsequent transcvtosis fright); solid arrow, pathway for basolateral proteins; broken arrow, pathway for apical proteins. (c) Morphological polarization of the cell. -

tion (van Meer and Simons, EMBO J 1986,5:1455-1464). These two mechanisms make different predictions about the mobility of surface proteins. Many polarized membrane constituents appear to diffuse freely and rapidly in the plane of the membrane. Measurements of protein mobility using fluorescence recovery after photobleaching (FRAP) indicate that the major fractions of several apical and basolateral membrane glycoproteins are free to diffuse (Salas et al, J Cell Biol1988, 107:23652376). Their diffusion coefficients of approximately 10 - 9 cm2 set - l are consistent with relatively free diffusion in a lipid bilayer. There are also several examples of specitic lipid molecules, that diffuse rapidly and are restricted to the apical membrane domain (van Meer and Simons, 1986). Tbo other classes of epithelial surface proteins are unlikely to be immobilized by attachment to the cytoskeleton, although their mobilities have not yet been determined. Several apical membrane proteins are anchored solely in the outer leaflet of the membrane via linkage to a glycosyl-phosphatidylinositol (GPI) moiety [7]. Also, many basolateral receptors and some apical proteins must behave dynamically in the membrane because they are continually endocytosed and recycled back to the surface (Fuller and Simons, J Cell Biol 1986, 103:1767-1n9;Matter et al, J Biol c3em 1990, 265:350%3512).

These dynamic properties suggest that

cytoskeletal anchoring cannot be the universal mechanism for restraining proteins to the apical or basolateral domain. Cytoskeletal anchoring probably occurs for only a subset of membrane proteins (see below). The pres-

ence of a barrier to diffusion at the boundary between membrane domains must be invoked to account for the maintenance of distinct membrane compositions. It appears that the tight junction, or zonufu occludens, forms the fence that separates the apical and basolatet-al plasma membrane domains. It co-localizes precisely with the boundaries of the distributions of several polarized membrane proteins (Gumbiner, Am J P&i01 1987, 253:C749-C758). The molecular structure of the tight junction is not well understood, but a model has been proposed for the tight junction fence (Fig. 2). Two tight junction proteins of unknown function have been identified. ZO-1 is a peripheral membrane protein closely associated with the cytoplasmic surface at the junction contact sites, and cingulin may be a cytoskeleton-associated protein localized near the tight junction [S] . The apical and the basolateral membranes are further organized into subdomains (Fig. 3). For example, cell-substratum adhesion receptors and components of hemidesmosomes are concentrated at the basal surface (Rapraeger et al, J Cell Bioll986, 103:2683-2696; Klatte et al, J Cell Bioi 1989, 109:3377-33901, whereas cell-cell adhesion molecules and cell junction proteins, including E-cadherin, desmosomal proteins, and the connexin protein of the gap junction, are localized to specilic regions of the lateral surface (91. Presumably, they are sublocalized as a result of their interactions with extracellular ligands in the basal lamina or in the neighboring cell. Certain proteins that are not involved in cell in teractions, such as ion transporters, are also localized

Generation

and maintenance

of epithelial

cell polarity

Cumbiner

Fig. 2. Model for the tight junction fence (modified from van Meer and Simons, EMBO 1 1986, 5:1455-W%). A putative integral membrane protein (X) forms a physical barrier to the movement of macromolecules. Integral membrane proteins (IMP), glycosylphosphatidylinositol-anchored proteins (GPO, and glycolipids (CL) all have large extracellular moieties that cannot diffuse through the occluding barrier in the intercellular space. The tight junction fence also extends into the outer leaflet of the lipid bilayer, because it restricts the diffusion of lipids in the external leaflet, but not the diffusion of lipids in the inner leaflet. In fact, the formation of an actual intercellular contact at the tight junction may not be required for the function of the fence, because functionally similar fences occur at contact-free sites in other cell types, such as the domain boundaries in the sperm plasma membrane (Myles et a/., / Cell &o/1984, 8:1905-1909), at the axon hillock in neurons (K Simons, personal communication), and at the foot processes of renal glomerular podocytes (Schnabel et al., Eur 1 Cell Viol 1989, 48:313-3262 Double headed arrows represent free diffusion past the tight junction.

in subdomains. In kidney epithelial cells, for example, both the band 3like anion exchanger and the Na+ -K+ ATPase are concentrated at highly infolded regions of the basolateral plasma membrane, but are relatively excluded from flat regions of the basal membrane in contact with the substratum and from the region of the junctional complex (Drenckhahn and Merte, EurJ Cell Bioll987, 45:107-l 15; Koob et al, Eur J Cell Bioll987, 45:230-237). The apical surface may be similarly subdivided for certain proteins into microvillus-associated and microvillus-free membrane surfaces (Kerjaschki ef al, J Cell Bioll984,98:1505-1513). The explanation for these non-homogeneous distributions of certain membrane proteins within the apical or the basolateral domains requires mechanisms in addition to the tight junction barrier. Some transporter proteins do interact directly with the underlying spectrin-based cytoskeleton. Linkage of the band 3 anion exchanger to the membrane cytoskeleton of the erythrocyte is mediated by its well characterized interaction with the protein ankyrin. In intercalated cells of the kidney, ankyrin co-localizes with band 3, and probably links it to the cytoskeleton in a similar way (Drenckhahn et al, Science 1985,230:1287-1289). The Na+-K+ATPase has been shown to interact directly with ankyrin in vitro [5,6]. It also co-localizes with ankyrin in the basolateral domain of kidney cells. The purpose of these membrane-cytoskeletal interactions may be to restrict the distributions of these transporter proteins to sub-

domains of the basolateral membrane. Membrane infoldings occur in epithelial cells to increase the surface area for transport functions, and it would not be surprising if the membrane cytoskeleton played an important role in their organization and stabilization.

Role of cell-substrate generation of polarity

adhesion

in the

The establishment of the apical-to-basal axis of polarity depends upon the adhesion of epithelial cells to the basal lamina in viva or to the culture substratum in vitro. In the example of polarity reversal in cultured epithelial cysts mentioned earlier [4], the cells respond to collagen in their environment and re-establish a new axis of polarity. The re-organization of the apical membrane domain is the most conspicuous response. In early mouse embryos, the eight-cell-stage blastomeres develop a morphologically discemable axis in response to contact with an extracellular substrate or another blastomere well before the development of a functional epithelium at compaction (Ziomek and Johnson, Cell 1980, 21935-942). An apical pole opposite the substrate contact site is evident as a prominent cluster of microvilli, which can also be detected as a surface ‘cap’ of concanavalin A receptors. These represent the majority of the cell surface glycoproteins. Their apparent localization is simply due to the increased membrane surface area asso-

883

884

Cell-to-cell

contact

and extracellular

matrix Surface

1

Coated

pits

enzyme

1

Fig. 3. Subdomain organization the apical and the basolateral membrane domains of epithelial

within plasma cells.

ciated with the microvilli, and is not equivalent to the presence of a compositionally distinct apical membrane domain. However, the formation of this polar structure is probably an early stage in the establishment of the apical-to-basal axis of polarity. Individual MOCK cells also seem to develop a distinct apical pole when they attach to a substrate (Ojakian and Schwimmer, J Cell Biof 1988, 107:2377-2387). Together, these lindings demonstrate that the extracellular matrix provides the cell with a cue to organize the axis of epithelial polarity and to create a presumptive apical pole.

sion in the kidney is restricted to the time of epithelial polarization, whereas the B chain is expressed throughout kidney development [ 101. An important challenge for future research will be to determine how laminin and/or collagen influence the axis of epithelial cell polarity.

The extracellular molecular signals for the polarization of the epithelial axis probably reside in the basal lamina. Although type I collagen has a striking inIluence on the polarity of cultured cells, it is not present in basal laminae. Type I collagen may exert its effects either indirectly by interacting with other matrix proteins or by substituting for type IV collagen, which is present in basal laminae and at the basal surface of cultured epithelial cells [ 31. Iaminin has been shown to play an essential role in epithelial polarization during the development of kidney tubules. Antibodies to the laminin A-chain block the morphological polarization of the tubules without interfering with the induction and condensation of the kidney mesenchyme (Klein et al., Cell 1988,55:331-341). The signal for polarization seems to reside in the laminin A chain, because antibodies to the B chain do not inhibit polarization. The A chain is localized at the basal cell surface, unlike the B chain which has a more widespread distribution, supporting the proposal that the A chain has a role in polarization. Moreover, the period of A chain expres-

Certain steps in cell polarization, such as the formation of a presumptive apical pole, do not require the establishment of cell-cell contact. Also, single epithelial cells are able to sort apical and basolateral membrane proteins intracellularly. Apical proteins can be sorted away from the plasma membrane into an unusual intracellular compartment called a vacuolar apical compartment (VAC; Vega Was et al., J Cell Biol 1988, 107:1717-1728). The physiological significance of this compartment is uncertain, but its existence demonstrates that the sorting machinery can function even when disengaged from the overt morphological polarization of the cell.

Role of cell-cell polarity

contacts

in the generation

of

Epithelial cells must form intimate cell-to-cell contacts, however, to express the entire spectrum of polarized features characteristic of mature epithelia. When cells are plated without Ca2+ to prevent cell-cell contact, many normally polarized plasma membrane proteins are distributed over the entire cell surface (Nelson and Veshneck, J Cell BioZl986, 103:1751-1765) [11,12]. Aher the initiation of cell-cell contact by the addition of Ca2+, the

Generation

cells develop their final state of polarization with distinct apical and basolateral domains. The formation of intimate cell-cell contacts has numerous effects on epithelial cells, many of which participate in the linal stages of polarization (Table 1). One of the most rapid events is the assembly of the tight junctions (Siliciano and Goodenough, J Cell Biol 1988, 107:238+2399), which immediately establishes the boundary between membrane domains. Initiation of cell-cell contact also stimulates rapid exocytosis of the VACs near the newly formed tight junctions (VegaSalas et al, 1988). Their addition to the surface forms an overt apical membrane domain almost instantaneously. Even the distribution of internal organelles, such as the Golgi apparatus, the centrioles, and microtubules, are inlluenced by the establishment of cell-cell contacts [ 13,141. These changes have important implications for the generation and maintenance of cell polarity, because microtubules are known to participate in the delivery of transport vesicles to the apical domain [15,16].

Table

1. Initiation

of ceil-ceil

epithelial

cell organization.

Response

to cefkell

contacts

has multiple

on

Dependence

contact

on E-cadherin

Formation Right

effects

of junctional

junctions,

zonula

complex

Yes’

desmosomes,

of gap junctions

Yestt

Increased

area of membrane

Yed

apposition Assembly

of fodrin-based

membrane

cytoskeleton

Exocytosis

of vacuolar

yes5 1181

apical

I ”

compartment Wistribution

of centrioles,

nicrotubules,

and Colgi

t 113,141

apparatus

‘Cumbiner

et al., / CeN Biol 1988, 107:1575-1587;

/ Cell Biol 1985, 101:1307-1315; JSA 85:7274-7278; 104:1527-1537;

BNelson //Vega-Sales

tMege

and Veshnock,

t8ehrens

et a/., Proc Nat/ Acad

et al., Sci

/ Cell Biol 1987,

er a/., / Cell Biol 1988,

maintenance

of epithelial

cell

polarity

Gumbiner

brane proteins [6,17]. In support of this hypothesis, the membrane cytoskeleton has been found to form complexes with the Na+ -K+ -ATP-ase in MDCK cells before the initiation of cell contacts [6], and its assembly at the basolateral membrane correlates in time with the localization of the Na+ -K+ -ATEase to the basolateral domain (Nelson and Veshnock, 1986). Although this mechanism does provide an explanation for the basolateral localization of the Na+-K+ATPase, it cannot account for the polarization of proteins that do not interact directly with the fodrin-based membrane cytoskeleton [ 181. Whether the membrane cytoskeleton is involved in localizing other molecules which indirectly establish the pattern of the basolateral domain, is not yet known. The fodrin-based cytoskeleton does not seem to polarize to regions of cell-cell contact in all epithelia Its organization is more complex in many other epithelial cell types, including hepatocytes and many cells of the renal tubules [ 191. Only the erythrocyte isoform of ankyrin is restricted to the basolateral domain of epithelial cells, but it is not expressed in all epithelia In contrast, the brain isoforms of ankyrin and fodrin, which are expressed in virtually all epithelia, are associated with both the apical and the basolateral membranes. To better understand the role of the membrane cytoskeleton in the generation of epithelial polarity, we will have to learn more about the identity of the receptors and modes of attachment for the different fodrin and ankyrin isoforms at each membrane domain. The protein primarily responsible for controlling the state of contact between epithelial cells is the Ca2+ dependent adhesion molecule E-cadherin, also called uvomomlin or LCAh4. It mediates several, if not all, of the contact-dependent events that participate in the generation of surface polarity (Table 1). The formation of the entire junctional complex, including the tight junction, depends on the function of E-cadherin (Gumbiner et al, J Cell Bioll988, 107:157>1587). E-cadherin also seems to induce fodrin and the Na+ -K+ -ATPase to accumulate in regions of cell-cell contact when it is expressed in non-polar fibroblasts by cDNA transfection [ 181. Whether E-cadherin is involved in VAC exocytosis or the rearrangements of internal cytoskeletal components and organelles has not been determined.

adhaerens)

Formation

and

107:1717-1728.

Cell-cell contact also initiates the assembly of the fodrin-based cytoskeleton at the basolateral surface (Nelson and Veshnock, J Cell Biol 1987, 104:1527-1537). It has been proposed that localized assembly of the membrane cytoskeleton at regions of cell-cell contact mediates the polarization of basolateral mem-

In order to mediate cell adhesion and influence multiple cellular events, E-cadherin needs to interact with components in the cytoplasm. The highly conserved cytoplasmic tail region of E-cadherin, which is required for cell adhesion, binds to a set of three polypeptides called the catenins [20,21]. These in turn may interact with the cytoskeleton. Two types of cytoskeletal interactions, both having implications for cell polarity, have been proposed. First, because E-cadherin localizes to the zonulu &eren.s junction in some epithelia, it may interact with the contractile actin lilament bundle associated with the ronula uu8aeren.s near the apical pole of the cell (Boiler et al, J Cell Biol 1985, 100:327-332). Second, a small fraction of the cellular E-cadherin has been reported to form complexes with the fodrin membrane cytoskeleton

885

886

Cell-to+ell

/contact

and extracellular

matrix

[17]. Further characterization of the catenins will probably reveal much about the nature of the interaction of Ecadherin with cytoskeletal structures.

Conclusion

and perspective

The generation qnd maintenance of polarity depends on multiple cellular processes, including intracellular protein sorting, the establishment of an apical pole opposite the site of contact with the extracellular matrix, the assembly of intercellular junctions, and the polarization of the cytoskeleton. One important challenge for the future will be to determine which molecules specify the establishment of pattern in the epithelial plasma membrane rather than respond to it. For example, what are the molecules responsible for targeting sorted transport vesicles to specific membrane domains? How are the distributions of these molecules influenced by cell-cell and cell-substrate contacts? Ultimately, we will be able to understand how all the components of the polarization process are integrated to create a polarized multicellular epithelium.

ceII polarity in multicellular epithelial (MDCK) cysts. / Cell Sci 1990, 95:153165. Exposure of the apical surface of MtXIK cysts (see 131) to collagen gels causes them to reverse polarity completely within 24 h. This paper demonstrates for the first time that reversal occurs by the degradation of the apical domain and the resynthesis of a new apical domain at the opposite pole of the cell. Thus, the maintenance of the apical pole, in addition to its initial formation, depends on a cue from the extracellular matrix. MORROWJS, CLUW CD, ARDITO T, MANN AS, k5HGARL4N M: Ankyrln links fodtin to the alpha subunit of Na+,K+ATpase in Madin-Darby canine kidney cells and in intact renaI tubule cells. / Ceu Bid 1989. 108:455465. A very nice study characterizing the interaction between the a-subunit of the Na+-K+-ATPase and the cytoskeletal protein ankyrin, and demonstrating their co-localization in the basolateral domain of epithelial cells. 5. o

l

6. a*

NELSON WJ, HAMMERTON RW: A membrane-cytoskeletal complex containing Na+,K+ -ATPase, at&pin, and fodrin in Madin-Darby canine kidney (MDCK) cells: implications for biogenesis of epithelial ceU polarity. J cell Biol 1989, 108:893902. The membrane cytoskeleton does not assemble in MDCK cells that are plated in low Ca2 + medium, preventing the formation of cell-cell contacts. This paper demonstrates for the first rime that protein complexes containing fodrin, ankyrin, and the Na+-K+-ATPase can be extracted from the cells with mild non-ionic detergents. It is proposed that these complexes represent normal precursors for the biogenesis of the basolateral plasma membrane.

7. me

Acknowledgements I wish to thank P McCrea, B Stevenson, M Symons and J White for their helpful comments on this manuscript.

Uswn MP, RODRIGUEZ-BOULW E: Glycophospholipid membrane anchoring provides clues to the mechanism of protein sorting in polarized epithelial cells. Trends Biockm .Sci 1990, 15:113118. An excellent review of recent papers by the authors and others demonstrating that GPI linkage of proreins to the membrane functions as a signal for targeting proteins to the apical domain. It also contains a good discussion of the nature of the various signals involved in targeting membrane proteins to either the apical or the basolateral domain.

8.

Annotated reading 0

00

references

and recommended

Of interest Of outstanding interest

1. SIMONSK, WAND~NCER-NESS A: Polarized sorting in epithelia. me Cell 1990, 2:207-210. An excellent recent review that discusses the intracellular transport pathways and sites of sorting for apical and basolateral membrane pro. teins in epithelia. A model for the mechanism of protein and Upid sorting Is proposed that accounts for the different types of sorting signals found in various polarized membrane proteins. 2. EKELOM P: Developmentally regulated conversion of mes0 enchyme to epithelium. FASESJ 1989. 332141-2150. A clear and concise review of the adhesion mechanisms involved in the de now development of polarized kidney tubules from induced mesenchymal tissue. 3. 0

WANG AZ, OJAKUN GK, NEWN WJ: Steps in the morphogenesis of a polarized epithelium I. Uncoupling the roles of ceU-ceIl and cellsubstratum contact in establishing plasma membrane polarity in multicellular epithelial (MDCK) cysts. / cf?u sci 1990, 95:137-151. This paper analyses the process of polarization of the MDCK epithelial cdl line when it grows as a cyst in suspension culture, using immunotluorescence staining for domain-specilic markers. The authors attempt to define the relative contributions of cell-cell and cell-substrate contact to the polarization of these markers. 4. a*

WANG AZ, OJAKIAN GK, NELSON WJ: Steps in the morphogenesis of a polarized epithelium Il. Disassembly and assembly of plasma membrane domains during reversal of epithelial

STEVENSONBR, HEINIZELMAN MB, ANDERXIN JM, Cm S, M~~SEKER MS: ZO-1 and cingulin: tight junction proteins with distinct identities and locahzations. Am / Pbysiol (C&II Pbysioll) 1989, 257:C621-C628. Two polypeptides, ZO-1 and cingulin, are localized to the region of the tight junction in a variety of epithelial ceU types. This paper demonsuates by imrnunochemical criteria that these two molecules are not closely related. Moreover, electron microscopic immunocytochemistry is used to show that ZO-1 is localized to the cytoplasmic membrane surface at the tight junction contact site, while cingulin is located further from the membranes. 0

STEVENSON BR, PAUL DL The molecular constituents of in9. 0 tercellular junctions. Curr opin cell Biol 1989, l&34-891. A clear, recent review of the molecular structures of several kinds of intercellular junctions, especially the epithelial junctions and the gap junctions.

10. a

EKBWM M, KLEIN G, MUGRAUER G, FECKER I DEUIZ~WNN R, TIMPL R, EKBLOM P: Transient and locally restricted expres-

sion of Iaminin A chain mRNA by developing epithelial ceUs during kidney organogenesis. Cell 1990, 60:337-346. A very thorough and careful study of the time and location of the expression of the laminin A chain during the morphogenesis of the kidney tubules. Iaminin A-chain expression by the developing epithelial cells coincides with the polarization of the tubular epithelia and then diminishes. 11. RODRIGUEZ-BOUIAN E, NELSON WJ: Morphogenesis of the po 00 larized epithelial celI phenotype. Science 1989, 245:71+725. A recent concise, but comprehensive, review on the subject of epitheliai ceU polarity. It includes some of the tindings covered in the present review, described from a somewhat different viewpoint, 12. @

CONITCERAS RG, Avll~ G, GUTIERREZC, BOUVARg, GONZALEZMARKAL

I,

DARZON

A,

CEREIJLDOM: Repolarization

BEAYY

G,

RODRIGUEZ-BOULW

E,

of Na+ -K+ pumps during es-

Generation tablishment of epithelial monolayers. Am J P&siol 1989, 257:6896-C905. MDCK cells cultured without Ca*+ are induced to form ceULcell contacts by the addition of Ca*+ The rapid development OF functional tight junctions traps some of the preexisting Na+ -K+-ATPase molecules in the apical domain. More slowly with time, the Na+ -K+ -ATPase be comes polarized to the basolateral surface. This Study shows that the formation of the tight junction fence is one of the earliest events to occur during the development of polarity following the initiation of ceU contacts. 13. a

BACAUAO R, ANI’ONY C, Darn C, KARF~ENII E, STEIZER EHK, StMONS K: The subcellular organization of Madin-Darby canine kidney cells during the formation of a polarized epithelium. / Cell Biol 1989, 109:2817-2832. The authors used confocal immunofluorescence microscopy and threedimensional image reconstruction to follow the organization of several di@erent qtoplasmic organelks during the development of a polarized epithelium from individual MIXK cells. Two days after plating, functional tight junctions form at the base of the cell. At this time, the Go@ apparatus surrounds the nucleus and the centrioles split and no longer nucleate microrubules. Subsequently, the junctions move upward to the apical end of the cell, along with the Golgi and the centrioles. Also, the microtubules become organized into an apical web and into polar long itudinal bundles parallel to the apical-basal axis with minus ends fowards the apical pole. These data indicate that extensive reorganization of microtubules and associated organeUes occurs during the formation of cell-cell contacts, resulting in the polarization of the cytoplasm. 14. *a

BUEM)IA B, Bt& M-H, G ttmrr~s G, KARSENTI E: Cytoskeletal control of centrioles movement during the establishment of polarity in Madin-Darby canine kidney cells. / cell Biol 1990, 110:1123-l 135. This paper demonstrates that the state of intercellular contacts, manipulated by the concentration of Ca*+ in the CUlNre medium, controls the separation of centrioles in MXK cells (see [ 131). Moreover, centriole separation depends on the integrity of actin filaments, and the authors propose that actin lilaments linked to cell junctions exert a force on the centrioles to bring about their movements.

H-P: Nocodazole, a EIUX U, KLUMPERMAh’ J. HAM microtubule-active drug, interferes with apical protein delivery in cultured intestinal epithelial cells (CaCo-2). J Cell Bid 1989, 108:13-22. Disruption of the microNbules in an epithelial ceU line causes apical proteins to be misdirected to the basolateral surface, but has no effect on the delivery of basolateral proteins. This paper and the next 1161 demonstrate that microtubules are involved in targeting apical transport vesicles to the apical domain. 15.

l

ACHLER C, FXMER D, MEKIX C, DRENCKHAHN D: Role of microtubules in polarized delivery of apical membrane proteins to the brush border of the intestinal epithelium. / Cell Biol 1989, 109:17+189. Disruption of microtubules in the intestinal epithelium causes brushborder enzymes (apical) to be misdirected to the basolateral surface. The cells even form miniature brush-border structures at the basolateral surface.

16. *

and maintenance

of epithelial

cell wlarity

Cumbiner

NELSON WJ, SHORE EM, WANG AZ, HAMMERTON RW: Identilication of a membrane-cytoskeletal complex containing the ceU adhesion molecule uvomorulin (E-cadherin), ankyrin. and fodrin in Madin-Darby canine kidney epithelial cells. J Cdl Bioi 1990, 110:349-357. Protein complexes containing fodrin. ankyrin, and the Na+-K+-ATPase can be extracted from MDCK cells CUlNKd in low Ca*+ medium (see lb]). Although most of the E-cadherin separates away from the fodrin-containing complexes, a small fraction seems to co-fractionate with fodrin. E.cadherin does not, however, copurify with the complexes containing the Na+-K+ -ATPase. The authors propose that E-cadherin forms its own complex with the fodrin-based membrane cytoskeleton. 17. l

18. 0

MCNEIU H, OZAWA M, KEMLER R NELSON WJ: Novel function of the ceU adhesion molecule uvomorulin as an inducer of cell surface polarity. Cell 1990, 62:30’+316. The immunofluorescence distributions of several plasma membrane markers was compared in normal fibroblasts and in fibroblasts transfected to express E-cadherin. The Na+-K+ -ATPase and fodrin accumulated in the region of cell-cell contact in E-cadherin-expressing cells, but not in the normal fibroblasts. However, another basolateral membrane protein, the H2 histocompatibility antigen, remained diFusely distributed, whether or not the cells expressed E-cadherin. Curiously, the bulk of the ceU surface glycoproteins, detected by wheat germ agglutinin staining, accumulated at regions of cell-cell contact even in normal non-transfected fibroblasts. DAVIS J, DAVIS I BENNET V: Diversity in membrane binding 19. 00 sites of a&yrins. / Biol Gem 1989, 2646417-6426. This paper demonstrates that epithelial tissues contain diverse membrane binding sites for various isoforms of ankyrin. It also shows by immunoIluorescence microscopy that most, if not all, epithelial ceU types express the brain isoforms of ankyrin and fodrin at both the apical and the basolateral plasma membranes. The erythroqte form of ankyrin is expressed in fewer ceU types, but is localized to the basolateral domain. OU\WA M. BARIBAULT H, KEM~ER R: The cytoplasmic domain of the ceU adhesion molecule uvomotulin associates with three independent proteins structurally related in tierent species. EMBO / 1989, 8:1711-1717. This excellent paper shows that a set of three distinct polypeptides, called ‘catenins’, co-immunoprecipitate with E-cadherin extracted from epithelial cells or from non-epithelial ceU lines nansfected with the Ecadherin cDNA Moreover, these polypeptides do not co-immunopre cipitate with mutant forms of E-cadherin that have deletions in the cy toplasmic tail domain. Therefore, the catenins interact with the cytoplasmic tail of E-cadherin, and are excellent candidates to mediate the linkage of E-cadherin to the cytoskeleton.

20. 00

21.

omWA

M, WGWAU)

M, f&MER

fk UVOmOditbCatet’h

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plex formation is regulated by a specific domain in the cy toplasmic region of the ceU adhesion molecule. Proc Null Acud Sci USA 1990, 87:424ti250. The terminal 70.amino-acid half of the E-cadherin cytoplasmic tail is responsible for the E-cadherin-catenin interaction (see [201>, because it induces catenin binding when fused to a marker protein. The paper also provides preliminary evidence that the b-catenin polypeptide interacts with E-cadherin, while a-catenin may interact with actin. l

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