Spatial and functional organization of cells in tissues is determined by cell-cell adhesion, thought to be initiated through trans-interactions between extracellular domains of the cadherin family of adhesion proteins, and strengthened by linkage to the actin cytoskeleton. Prevailing dogma is that cadherins are linked to the actin cytoskeleton through beta-catenin and alpha-catenin, although the quaternary complex has never been demonstrated. We test this hypothesis and find that alpha-catenin does not interact with actin filaments and the E-cadherin-beta-catenin complex simultaneously, even in the presence of the actin binding proteins vinculin and alpha-actinin, either in solution or on isolated cadherin-containing membranes. Direct analysis in polarized cells shows that mobilities of E-cadherin, beta-catenin, and alpha-catenin are similar, regardless of the dynamic state of actin assembly, whereas actin and several actin binding proteins have higher mobilities. These results suggest that the linkage between the cadherin-catenin complex and actin filaments is more dynamic than previously appreciated.

Epithelial cell-cell junctions, organized by adhesion proteins and the underlying actin cytoskeleton, are considered to be stable structures maintaining the structural integrity of tissues. Contrary to the idea that alpha-catenin links the adhesion protein E-cadherin through beta-catenin to the actin cytoskeleton, in the accompanying paper we report that alpha-catenin does not bind simultaneously to both E-cadherin-beta-catenin and actin filaments. Here we demonstrate that alpha-catenin exists as a monomer or a homodimer with different binding properties. Monomeric alpha-catenin binds more strongly to E-cadherin-beta-catenin, whereas the dimer preferentially binds actin filaments. Different molecular conformations are associated with these different binding states, indicating that alpha-catenin is an allosteric protein. Significantly, alpha-catenin directly regulates actin-filament organization by suppressing Arp2/3-mediated actin polymerization, likely by competing with the Arp2/3 complex for binding to actin filaments. These results indicate a new role for alpha-catenin in local regulation of actin assembly and organization at sites of cadherin-mediated cell-cell adhesion.

Tissue morphogenesis during development is dependent on activities of the cadherin family of cell-cell adhesion proteins that includes classical cadherins, protocadherins, and atypical cadherins (Fat, Dachsous, and Flamingo). The extracellular domain of cadherins contains characteristic repeats that regulate homophilic and heterophilic interactions during adhesion and cell sorting. Although cadherins may have originated to facilitate mechanical cell-cell adhesion, they have evolved to function in many other aspects of morphogenesis. These additional roles rely on cadherin interactions with a wide range of binding partners that modify their expression and adhesion activity by local regulation of the actin cytoskeleton and diverse signaling pathways. Here we examine how different members of the cadherin family act in different developmental contexts, and discuss the mechanisms involved.Cadherins were originally identified as cell surface glycoproteins responsible for Ca 2+ -dependent homophilic cell-cell adhesion during morula compaction in the preimplantation mouse embryo and during chick development (Yoshida and Takeichi 1982;Gallin et al. 1983;Peyrieras et al. 1983). Subsequently, >100 family members have been identified with diverse protein structures, but all with characteristic extracellular cadherin repeats (ECs) (Nollet et al. 2000). Cadherins are important in both simple and complex organisms. In addition to vertebrates, insects, and nematodes, members of the cadherin family are found in unicellular choanoflagellates (King et al. 2003), the diploblast Hydra (Hobmayer et al. 2000), and the sponge Oscarella carmela (Nichols et al. 2006).In the three decades since their discovery, it has become clear that the role of cadherins is not limited to mechanical adhesion between cells. Rather, cadherin function extends to multiple aspects of tissue morphogenesis, including cell recognition and sorting, boundary formation and maintenance, coordinated cell movements, and the induction and maintenance of structural and functional cell and tissue polarity. Cadherins have been implicated in the formation and maintenance of diverse tissues and organs ranging from polarization of simple epithelia, to mechanically linking hair cells in the cochlea, to providing an adhesion code for neural circuit formation during wiring of the brain (Yagi and Takeichi 2000;Gumbiner 2005). Given the breadth of their functions, it is not surprising that defective cadherin expression has also been linked directly to a wide variety of diseases including the archetypal disruption of normal tissue architecture, metastatic cancer .The large size of the cadherin family and the structural diversity of its members may have evolved to enable the many types of cell interactions required for tissue morphogenesis in complex organisms. The number of cadherins as well as distinct features of their gene structure permit precise temporal and spatial transcriptional regulation of cadherin subtypes, and variations in protein structure, particularly the cytopl...

complex containing at least two RAS-related GTP-binding proteins assembles at the site marked by the cue, *Department of Molecular and Cell Biology then a polarized actin and septin cytoskeleton assem-University of California, Berkeley bles, and the secretory apparatus becomes oriented Berkeley, California 94720 toward the spatial cue. Considerable information about † Department of Molecular and Cellular Physiology bud formation has been obtained during the past several Beckman Center for Molecular and Genetic Medicine years, and some advances in our understanding of mat-Stanford University School of Medicine ing projection formation and orientation have also been Stanford, California 94305-5426 made recently. Bud Site Selection Occurs in Response Cell polarity is the ultimate reflection of complex mechato Intrinsic Spatial Cues nisms that establish and maintain functionally special-Both genotype and nutritional conditions determine ized domains in the plasma membrane and cytoplasm. which of three budding patterns yeast cells will adopt. The spatial arrangement and protein composition of MATa and MAT␣ cells construct bud sites adjacent to these domains facilitate cellular processes as diverse the previous bud site (axial budding pattern), while as differentiation, localized membrane growth, activa-MATa/MAT␣ cells bud from sites that are either near tion of the immune response, directional cell migration, the previous bud site or at the opposite end of the cell and vectorial transport of molecules across cell layers. (bipolar budding pattern) (Chant and Pringle, 1995, and In this review, two phylogenetically distant eukaryotic references therein). A yeast cell undergoing pseudohycells, budding yeast and mammalian epithelial cells, are phal growth always buds from the same pole, namely examined to highlight advances in our understanding the pole opposite the original junction with its mother of how cell polarity is established. Both of these cells cell (unipolar budding pattern) (Kron et al., 1994). These are characterized by a high degree of cellular asymmetry three budding patterns presumably optimize the evolu-(see Figure 1) and have been used extensively to study tionary fitness of yeast growing in the haploid and diploid how cell polarity is developed. The specific focus here states, depending on the supply of nutrients (Gimeno is on the molecular nature of the intrinsic and extrinsic and Fink, 1992). Placement of cortical cues for each spatial cues that establish structural and molecular budding pattern is dependent upon one of two cytoskelasymmetry at the cell surface, the mechanisms that inetal proteins, septins and actin. terpret signals from these cues to generate new mem-Partial loss-of-function mutations in septins, cytoskelbrane domains, and the reorganization of the cell around etal proteins that are arranged in a ring at the bud neck these spatially defined sites (see Figure 2). The evidence and are required for cytokinesis (Sanders and Field, supports a model in which a hierarchy of three sequen-199...

Polarized epithelial cells play fundamental roles in the ontogeny and function of a variety of tissues and organs in mammals. The morphogenesis of a sheet of polarized epithelial cells (the trophectoderm) is the first overt sign of cellular differentiation in early embryonic development. In the adult, polarized epithelial cells line all body cavities and occur in tissues that carry out specialized vectorial transport functions of absorption and secretion. The generation of this phenotype is a multistage process requiring extracellular cues and the reorganization of proteins in the cytoplasm and on the plasma membrane; once established, the phenotype is maintained by the segregation and retention of specific proteins and lipids in distinct apical and basal-lateral plasma membrane domains.

Helicobacter pylori translocates the protein CagA into gastric epithelial cells and has been linked to peptic ulcer disease and gastric carcinoma. We show that injected CagA associates with the epithelial tight-junction scaffolding protein ZO-1 and the transmembrane protein junctional adhesion molecule, causing an ectopic assembly of tight-junction components at sites of bacterial attachment, and altering the composition and function of the apical-junctional complex. Long-term CagA delivery to polarized epithelia caused a disruption of the epithelial barrier function and dysplastic alterations in epithelial cell morphology. CagA appears to target H.pylori to host cell intercellular junctions and to disrupt junction-mediated functions.

Cell polarity is defined as asymmetry in cell shape, protein distributions and cell functions. It is characteristic of single-cell organisms, including yeast and bacteria, and cells in tissues of multi-cell organisms such as epithelia in worms, flies and mammals. This diversity raises several questions: do different cell types use different mechanisms to generate polarity, how is polarity signalled, how do cells react to that signal, and how is structural polarity translated into specialized functions? Analysis of evolutionarily diverse cell types reveals that cell-surface landmarks adapt core pathways for cytoskeleton assembly and protein transport to generate cell polarity.

In budding yeast, the Sec6/8p complex is essential for generating cell polarity by specifying vesicle delivery to the bud tip. We show that Sec6/8 homologs are components of a cytosolic, approximately 17S complex in nonpolarized MDCK epithelial cells. Upon initiation of calcium-dependent cell-cell adhesion, approximately 70% of Sec6/8 is rapidly (t(1/2) approximately 3-6 hr) recruited to sites of cell-cell contact. In streptolysin-O-permeabilized MDCK cells, Sec8 antibodies inhibit delivery of LDL receptor to the basal-lateral membrane, but not p75NTR to the apical membrane. These results indicate that lateral membrane recruitment of the Sec6/8 complex is a consequence of cell-cell adhesion and is essential for the biogenesis of epithelial cell surface polarity.