Cadherin adhesome at a glance

This article is part of a Minifocus on Adhesion. For further reading, please see related articles: ‘Cycling around cell–cell adhesion with Rho GTPase regulators’ by Jessica McCormack et al. (J. Cell Sci. 126, 379-391). ‘E-cadherin–integrin crosstalk in cancer invasion and metastasis’ by Marta Canel et al. (J. Cell Sci. 126, 393-401). ‘Mechanosensitive systems at the cadherin–F-actin interface’ by Stephan Huveneers and Johan de Rooij (J. Cell Sci. 126, 403-413).Multicellular life hinges on molecular ensembles that are able to mediate adhesion between cells (Abedin and King, 2010). One such conserved structure is the adherens junction (AJ); here, transmembrane glycoproteins, the cadherins, interact homophilically with cadherins that are located on apposing cells, on the outside, and are anchored through a conglomerate of adaptor proteins to the actomyosin cytoskeleton on the inside (Harris and Tepass, 2010; Niessen et al., 2011).This Cell Science at a Glance article makes an inventory of the structural and regulatory components of AJs, and the interactions between them, collectively referred to as the cadherin adhesome, or cadhesome. Assembling this blueprint of building blocks and connections serves to highlight the molecular complexity and potential diversity of AJs, and offers an opportunity to address some important questions, such as: how do cadherins connect with the cytoskeleton? How are actin dynamics regulated at AJs? Which molecular switches regulate AJ dynamics? And what is the role of tension in the maturation of AJs?Cell–cell adhesion in AJs is primarily mediated by a twofold symmetric interaction between the first extracellular (EC1) domains of two cadherins on juxtapositioned cells (reviewed by Brasch et al., 2012). This trans-interaction has been defined by crystallography as a ‘strand swap’, in which a conserved tryptophan side chain of one EC1 inserts into a hydrophobic pocket in the EC1 of the opposing cadherin (Harrison et al., 2011). In addition to binding in trans, cadherins can also form cis-interactions through an interaction between the EC1 and EC2 domains of two cadherins that are located on the same cell (Harrison et al., 2011). Despite our detailed understanding of the interactions at the structural level, we are still missing many details regarding the organization of cadherins on the surface of cells. Whether cis-interactions precede or follow trans-interactions, for example, is still an open question. Chemical cross-linking experiments suggested that cis-dimers exist on the cell surface (Takeda et al., 1999), and some in vitro experiments suggested that lateral dimerization is required for adhesion (Brieher et al., 1996). However, other experiments and models indicate that trans-dimers, with their reduced molecular flexibility, are a prerequisite for the formation of cis-dimers and higher order assemblies (Brasch et al., 2012; Wu et al., 2010; Zhang et al., 2009).Before contacting another cell, cadherin is present all over the cell surface, including at the tips of filopodia and along the leading edge of lamellipodia. Shortly after a cell makes contact with another cell, the concentration of cadherin at the adhesion site increases as they cluster and form higher order structures (Adams et al., 1998; Raich et al., 1999; Vasioukhin et al., 2000). The clustering of cadherin, which is driven by trans- and cis-interactions on the outside of the cell, has profound effects on the inside of the cell. The intracellular tail of cadherin binds β-catenin before arrival at the membrane (Hinck et al., 1994), but after cadherins cluster, they recruit a multitude of additional structural and regulatory proteins that make up the AJ ‘plaque’ (Baum and Georgiou, 2011; Hartsock and Nelson, 2008; Troyanovsky, 1999). By far the most studied and best understood of these ‘plaque’ proteins are the catenins (α-, β-catenin and p120), which have important structural and regulatory roles (Gumbiner and McCrea, 1993; Nelson, 2008; Shapiro and Weis, 2009), but over 160 proteins have been reported to associate with AJs (see below). Concomitant with the clustering of cadherins and plaque proteins, F-actin is also recruited to and/or polymerized at these sites (Adams et al., 1996; Ivanov et al., 2005; Kovacs et al., 2002; Vasioukhin et al., 2000).Once formed, AJs constitute a physical link between the actin cytoskeleton of neighboring cells, providing the mechanical coordination needed for morphogenetic processes, such as gastrulation, neurulation, convergent extension, epithelial sheet-sealing and cell migration (Gumbiner, 2005; Halbleib and Nelson, 2006; Kardash et al., 2010; Lecuit, 2005; Lien et al., 2006). AJs also ‘sense’ their chemical and mechanical environment and serve as signaling centers in pathways controlling cell growth, differentiation and fate (Cavallaro and Dejana, 2011; Halbleib and Nelson, 2006; Lechler, 2012; Leckband et al., 2011). Importantly, AJs also regulate the homeostasis of adult tissues, and AJ dysfunction is involved in epithelial–mesenchymal transformation, a hallmark of cancer metastasis (Jeanes et al., 2008; Takeichi, 1993). Equally important as their assembly is the disassembly of AJs, a process that is tightly regulated by various signaling pathways (Baum and Georgiou, 2011; D'Souza-Schorey, 2005; Green et al., 2010); this way cells can modulate their adhesive properties in response to intrinsic or external cues in a temporally and spatially controlled manner.This short review and the accompanying poster are based on my compilation of information from primary literature from the last 30 years regarding AJ components and the direct interactions between them. The criteria used for assembling this dataset (the ‘Cadhesome’) are detailed in Box 1, and the complete annotated and referenced lists of components and interactions are available as supplementary material Tables S1 and S2.Over 170 proteins have been reported to colocalize with cadherin or catenins in AJs, and either directly interact with them or affect AJ dynamics (see supplementary material Table S1). Broadly speaking, the cadhesome is inhabited by two types of proteins: structural and regulatory proteins. Structural proteins consist of other transmembrane receptors in addition to cadherin, including Ca2+-independent immunoglobulin-like adhesion receptors of the nectin family (Sakisaka et al., 2007), cytoskeletal filaments, motors and over sixty adaptor proteins, many of which bind to cadherin or the cytoskeleton directly. For most of these adaptor proteins, it is not known whether they have a structural role or serve as a scaffold for signalling, or both. The presence of >70 regulatory proteins at AJs is not surprising, as a variety of cellular pathways regulate AJ dynamics, and signaling from AJs, in turn, controls important cellular behaviors (Braga, 2002; Cavallaro and Dejana, 2011; Lechler, 2012; McCrea et al., 2009; Stepniak et al., 2009). The largest groups of regulators are tyrosine kinases and phosphatases, and GTPases and their activators and inhibitors. These and other regulatory switches of the cadhesome will be discussed later.Although a few cadhesome components appear to be cell-type specific (e.g. neurojungin in neurons and KRIT1 in endothelia), the majority of cadhesome components have been observed in more than one cell type. This, however, does not necessarily mean that most components are present in every AJ. It has been shown that the composition of AJs can change over time (Green et al., 2010) in response to changes in tension (Taguchi et al., 2011), and varies between different cell types (Borrmann et al., 2000) and even between different AJs within the same cell (Peitsch et al., 1999). However, considerably more work remains before we can confidently classify subtypes of AJs according to their molecular composition and function.The complexity of AJs is further compounded by the large number of possible interactions between their constituents. Close to 390 pairwise interactions have been reported among the 174 cadhesome components (see supplementary material Table S2), a subset of which are illustrated in the accompanying poster. Close to two thirds of these interactions are classified as binding, whereas just over one third are classified as either activating or deactivating, depending on the respective enzymatic reaction. The dataset and the poster distinguish between 230 interactions that have been shown within the context of AJs, and 158 interactions that were found to occur outside of AJs, and for which it is not yet known whether they also take place in AJs.It is important to emphasize that although the poster depicts all of the possible interactions among cadhesome components, it is highly likely that only a subset of these interactions occur at the same time in the same AJ. Conceivably, every interaction is regulated and can be turned ‘on’ or ‘off’ under different conditions, thus allowing the AJ to respond to changes in the environment and signaling.Electron micrographs show a dense plaque of protein ∼25 nm wide between the plasma membrane and F-actin at AJs (for example see Yonemura et al., 1995). So far, we know very little about the three-dimensional organization of proteins within this plaque. Super-resolution microscopy and tagging of proteins in electron tomograms has recently been performed on integrin adhesion sites (Kanchanawong et al., 2010; Patla et al., 2010), and applying similar techniques to AJs should substantially enhance our understanding of their ultrastructure. Until then, the map of protein interactions might provide clues on the structural organization of AJs.The interaction between cadherins and the cytoskeleton has been the focus of intense research for the past 20 years, and yet the molecular details remain poorly understood (Yonemura, 2011). Two seemingly contradicting features characterize this interaction; (1) it is dynamic, as shown by fluorescence recovery after photobleaching (FRAP) experiments of cadherins, plaque proteins and actin (Yamada et al., 2005), and (2) it transmits tensile stresses, as observed in morphogenetic processes, such as apical constriction (Martin et al., 2009), and as directly demonstrated by cadherin adhesions that apply force on pillars (Ladoux et al., 2010), as well as by an intra-molecular stress sensor (Borghi et al., 2012). One model that is consistent with both characteristics hypothesises that cadherins are connected to F-actin by multiple transient links, with each link transmitting tension for the duration of its existence (Gates and Peifer, 2005).For many years it was thought that the cadherin–F-actin connection is solely mediated by cadherin-bound β-catenin that binds to F-actin-bound α-catenin (Gates and Peifer, 2005), but in 2005 the Nelson and Weiss groups contradicted this notion by showing that α-catenin that is bound to β-catenin cannot bind F-actin directly (Drees et al., 2005; Yamada et al., 2005). Nevertheless, it still appears to be the case that α-catenin and β-catenin are central to the binding of cadherin to F-actin, if not directly, then through additional adaptors, such as vinculin, eplin or zona occludens protein 1 (ZO1, also known as TJP1) (reviewed by Yonemura, 2011). Other transmembrane cadhesome proteins, such as nectin and vezatin, can connect to F-actin through adaptors, such as afadin, or motor proteins, such as myo7A (Küssel-Andermann et al., 2000; Sakisaka et al., 2007). In addition, AJs contain a number of membrane-bound actin-binding proteins, such as the so-called ezrin, radixin and moesin (ERM) proteins, which link cortical actin to the plasma membrane (Neisch and Fehon, 2011).There are also several lines of evidence to suggest an interaction between AJs and microtubules. The interaction between p120-catenin and the AJ components PLEKHA7 and Nezha (also known as CAMSAP3) anchors the minus ends of non-centrosomal microtubules to AJ, which could direct kinesin-driven vesicles towards AJ (Meng et al., 2008). Moreover, the interaction between β-catenin and dynein serves to capture the plus ends of microtubules at AJs and is important for the establishment of apico–basal cell polarity (Bellett et al., 2009; Ligon et al., 2001). In some specific cell types, such as in the lens or in dermal endothelial cells, there is evidence for a link between classical cadherins in AJ and the intermediate filament vimentin, mediated by γ-catenin and desmoplakin (Kowalczyk et al., 1998; Leonard et al., 2008).Cadherin adhesions are not simply tethered to a pre-formed actin network, but rather they have an active role in shaping the F-actin landscape that surrounds them (reviewed by Ratheesh and Yap, 2012). They do so by recruiting different actin nucleators and elongation factors, and a variety of actin dynamics regulators, cross-linkers and motors. At least three different actin structures can be found in association with AJs: F-actin bundles oriented parallel to and slightly separated from the plasma membrane; stress fibers oriented orthogonally to the plasma membrane with one end terminating at AJs; and a structurally unresolved pool of F-actin in close proximity to the membrane alongside cadherin clusters (reviewed by Niessen et al., 2011). The relationship between these different actin assemblies is not well defined. However, it has been shown that active actin polymerization that occurs adjacent to the membrane in AJs can contribute to the structure of circumferential actin bundles (Kovacs et al., 2011).Actin structures at AJs can appear to be stable for many minutes, but FRAP experiments suggest that the actin filaments themselves turn over within seconds, indicating that actin at AJs is continuously being polymerized and depolymerized (Kovacs et al., 2002; Kovacs et al., 2011; Yamada et al., 2005).In light of the variety of actin networks observed in AJs, it is not surprising to find in the cadhesome Arp2/3, which is responsible for nucleating and polymerizing a dendritically branched network, as well as the formin diaphanous and Ena/VASP proteins, which nucleate and elongate long unbranched actin filaments (Carramusa et al., 2007; Kovacs et al., 2002; Scott et al., 2006). Branched networks push membranes and drive protrusions as well as endocytosis, and unbranched filaments can coalesce with myosin to form contractile bundles (reviewed by Chesarone and Goode, 2009). How the activity of the different factors is coordinated in space and time at AJs is not known. The cadhesome also contains the activators of Arp2/3, mDia1 (also known as DIAPH1) and VASP, which themselves need to be activated by Rho GTPases. Thus, the type of F-actin network that is polymerized is likely to be determined by which of the activators are recruited into AJs, and by the balance between active Rho GTPases. Once several F-actin filaments are polymerized, they can be arranged into different higher-level structures, and connect with the AJs through different actin-binding adaptors. The cadhesome contains several actin cross-linking proteins, such as α-actinin and filamin, and several myosin motors that can serve as cross linkers as well as force-generating machines (Knudsen et al., 1995; Maddugoda et al., 2007; Smutny et al., 2010; Wakamatsu et al., 2011).During development and also in adults, cells regulate their AJs in response to changing needs (reviewed by Baum and Georgiou, 2011; Green et al., 2010; Lecuit, 2005). Long-term modulation of AJs, such as in epithelial–mesenchymal transition, is controlled at the levels of transcription, translation and trafficking, which determine the availability of cadhesome components at the membrane (Delva and Kowalczyk, 2009; van Roy and Berx, 2008). More rapid control of AJ dynamics is mediated by post-translational regulation of cadhesome proteins, which affects their activity and interactions (Bertocchi et al., 2012; van Roy and Berx, 2008). The types of post-translational regulation found within the cadhesome are: serine or threonine phosphorylation, tyrosine phosphorylation, GTPase binding, lipid binding and proteolysis.I refer to the activity of regulatory proteins as ‘switches’, because in many cases their action results in switching ‘on’ or ‘off’ an interaction of a protein or its activity (see also Zaidel-Bar and Geiger, 2010). For example, tyrosine phosphorylation of E-cadherin creates a binding site for the cell polarity protein NUMB (Wang et al., 2009); phosphorylation of a threonine residue on ERM proteins keeps them in an active, actin-binding state (Yonemura et al., 2002); binding of RAP1 to afadin activates its capacity to stabilize AJs (Hoshino et al., 2005); and cleavage of E-cadherin by ADAM10 downregulates cell–cell adhesion (Maretzky et al., 2005).Such regulatory switches are likely a general principle, although for most of the enzyme-substrate interactions in the cadhesome we do not know the functional implications of the modification for the target protein or for AJ dynamics. Nevertheless, grouping together all the known interactions between proteins from the same regulatory group helps to identify some design principles of AJ regulation. For example, it can be seen that regulators of actin dynamics are controlled by Rho GTPases, tyrosine phosphorylation and phosphatidylinositol 4,5-bisphosphate PtdIns(4,5)P2, whereas cadherins are regulated by serine/threonine phosphorylation and proteases.In a similar manner to integrin-mediated adhesions (Zaidel-Bar et al., 2004), cadherin-mediated adhesions undergo a process of maturation, during which their composition and associated actin structures evolve (Baum and Georgiou, 2011; Green et al., 2010; Troyanovsky, 1999). In years, it has that tension has an important role in this process et al., 2011; and 2010). However, it remains to determine which of the between and cadherin adhesions are driven by an in and clustering is thought to the GTPases and which and protrusions et al., 2000; Kovacs et al., These protrusions to the contact with the neighboring cell and thus on cadherin and At some during AJ maturation, to the of and (e.g. see et al., 2012; Ratheesh et al., there appears to be a from and to activity (Yamada and Nelson, 2007). the activity activates the formin mDia1 and Rho activity to the formation of contractile actomyosin bundles et al., 1999; Smutny et al., 2010). driven by myosin creates tension in the actin network, which on the AJ plaque proteins (Borghi et al., 2012; et al., 2010; et al., 2006; et al., 2005). tension appears to the of additional actin polymerization factors, which unbranched actin filaments et al., 2008). Ena/VASP might be recruited by an adaptor that has been shown to be in AJs, adhesions and stress fibers et al., 2012; et al., 2010; Zaidel-Bar et al., adaptor α-catenin is thought to contribute to AJ maturation in two In it is to be primarily bound to cadherin but as the of cadherin over a a subset of α-catenin could and bind to F-actin in a way that from binding et al., 2010; et al., 2005), thus the from a activity to In the contractile α-catenin that remains bound to cadherin has been to be a binding site for (Yonemura et al., 2010). The of the actin-binding protein serves to the link between cadherin and F-actin et al., 2010). Other proteins, such as myosin and have been shown to be recruited to AJs under tension et al., 2007; et al., 2011), a between and maturation of AJs.The model of AJ depicts cadherin only with the three catenins and By the complexity of the cadherin from the literature is It will be to as many cadhesome components as possible under various in order to the diversity of AJs, and to different subtypes of AJs in different cells, and also in different of the same cell and under different what we know about AJs also the in our For example, the number of phosphorylation in the cadhesome literature is only a of the phosphorylation by on cadhesome proteins (see at integrin has close to components (Zaidel-Bar and Geiger, 2010), and of adhesions suggest that the number of components is higher and 2012). However, assembling lists of proteins will not enhance our understanding it is with information on their molecular interactions and some cadhesome components have been the majority has been Thus, the cadhesome should be as a in of and cell will be needed to a cadherin that the features of AJs well for it to be into a model that can the dynamics of AJ assembly and disassembly in response to cellular that the most and important features of AJs, such as do not from the activity of one type of but rather are properties of the network of interactions between cadhesome proteins. to such as a will to be of my to the of the cadhesome and for the on the poster. to work could not be to space