Dynamics of adherens junctions in epithelial establishment, maintenance, and remodeling

Investigating the mechanisms controlling the asymmetric division of neocortical progenitors that generate neurones in the mammalian brain is crucial for understanding the abnormalities of cortical development. Partitioning of fate determinants is a key instructive step and components of the apical junctional complex (adherens junctions), including the polarity proteins PAR3 and aPKC as well as adhesion molecules such as N-cadherin, have been proposed to be candidate determinants. In this study, however, we found no correlation between the partitioning of N-cadherin and fate determination. Rather, we show that adherens junctions comprise three membrane domains, and that during asymmetrical division these are split such that both daughters retain the adhesive proteins that control cell position, but only one daughter inherits the polarity proteins along with the apical membrane. This provides a molecular explanation as to how both daughters remain anchored to the ventricular surface after mitosis, while adopting different fates.

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’

Asymmetric division is a fundamental mechanism for generating cellular diversity. In the central nervous system of Drosophila, neural progenitor cells called neuroblasts undergo asymmetric division along the apical-basal cellular axis. Neuroblasts originate from neuroepithelial cells, which are polarized along the apical-basal axis and divide symmetrically along the planar axis. The asymmetry of neuroblasts might arise from neuroblast-specific expression of the proteins required for asymmetric division. Alternatively, both neuroblasts and neuroepithelial cells could be capable of dividing asymmetrically, but in neuroepithelial cells other polarity cues might prevent asymmetric division. Here we show that by disrupting adherens junctions we can convert the symmetric epithelial division into asymmetric division. We further confirm that the adenomatous polyposis coli (APC) tumour suppressor protein is recruited to adherens junctions, and demonstrate that both APC and microtubule-associated EB1 homologues are required for the symmetric epithelial division along the planar axis. Our results indicate that neuroepithelial cells have all the necessary components to execute asymmetric division, but that this pathway is normally overridden by the planar polarity cue provided by adherens junctions.

Estrogens Determine Adherens Junction Organization and E-Cadherin Clustering in Breast Cancer Cells via Amphiregulin.

Caenorhabditis elegans provides a simplified, in vivo model system in which to study adherens junctions (AJs) and their role in morphogenesis. The core AJ components-HMR-1/E-cadherin, HMP-2/β-catenin and HMP-1/α-catenin-were initially identified through genetic screens for mutants with body axis elongation defects. In early embryos, AJ proteins are found at sites of contact between blastomeres, and in epithelial cells AJ proteins localize to the multifaceted apical junction (CeAJ)-a single structure that combines the adhesive and barrier functions of vertebrate adherens and tight junctions. The apically localized polarity proteins PAR-3 and PAR-6 mediate formation and maturation of junctions, while the basolaterally localized regulator LET-413/Scribble ensures that junctions remain apically positioned. AJs promote robust adhesion between epithelial cells and provide mechanical resistance for the physical strains of morphogenesis. However, in contrast to vertebrates, C. elegans AJ proteins are not essential for general cell adhesion or for epithelial cell polarization. A combination of conserved and novel proteins localizes to the CeAJ and works together with AJ proteins to mediate adhesion.

Epithelial tissues undergo extensive structural remodeling during embryonic development. Tissue remodeling is often enabled by oriented cell rearrangements that are linked with patterns of mechanical stresses in the tissue and with tissue mechanical properties. Cell rearrangements and their links to tissue structure have largely been studied at the apical side of tissues at the level of adherens junctions. Less is known about the involvement of basolateral domains in cell rearrangements. Here we use live confocal imaging to quantify cell rearrangements, cell packing structure, and cell morphology in 3D in the converging and extending Drosophila germband epithelium. We report gradients in cell shapes and tissue structure along the apical-basal axis of the germband, suggesting that the apical and basolateral domains display distinct behaviors. Cell rearrangements initiate at apical as well as basolateral positions, with initiation frequencies also displaying a gradient along the apical-basal axis. Analyses of cell contact angles and non-muscle myosin II localization patterns during cell rearrangements indicate that rearrangements can be actively driven in lateral as well as in apical regions of the tissue. Following initiation, rearrangements propagate across the apical-basal axis and lateral cell contacts remodel; these events involve scutoids and other complex 3D cell shapes as intermediate states. These findings uncover novel aspects of the cell rearrangements that drive dynamic remodeling of epithelia and reveal links between rearrangements and gradients in tissue structure along the apical-basal axis.

Notch-dependent Abl signaling regulates cell motility during ommatidial rotation in Drosophila.

The formation and maintenance of epithelia is critical for animal development and survival. Central to epithelial integrity are cadherin-based complexes called adherens junctions (AJs), which form physically robust but inherently dynamic cell-cell adhesions. How AJs function at the molecular level remains incompletely understood because techniques to study the central AJ proteins within the dynamic adhesion structure are scarce. Using a conformation sensitive probe that is amenable to fluorescence lifetime and anisotropy imaging, we demonstrate that the maturation of AJs is accompanied by a conformational change in the actin-binding domain of α-catenin. The structural transition depends on the degree of junctional maturation and requires actin polymerisation, but it is insensitive to vinculin binding to α-catenin. These different conformational states correlate with distinct α-catenin mobilities, with α-catenin unexpectedly showing an overall increased protein turnover in mature AJs. Collectively, the data reveal that α-catenin undergoes a previously proposed C-terminal conformational transition during epidermal differentiation to form mechanically stable yet dynamic cell-cell adhesions.

Based on recent data showing that overexpression of the Na,K-ATPase beta(1) subunit increased cell-cell adhesion of nonpolarized cells, we hypothesized that the beta(1) subunit can also be involved in the formation of cell-cell contacts in highly polarized epithelial cells. In support of this hypothesis, in Madin-Darby canine kidney (MDCK) cells, the Na,K-ATPase alpha(1) and beta(1) subunits were detected as precisely co-localized with adherens junctions in all stages of the monolayer formation starting from the initiation of cell-cell contact. The Na,K-ATPase and adherens junction protein, beta-catenin, stayed partially co-localized even after their internalization upon disruption of intercellular contacts by Ca(2+) depletion of the medium. The Na,K-ATPase subunits remained co-localized with the adherens junctions after detergent treatment of the cells. In contrast, the heterodimer formed by expressed unglycosylated Na,K-ATPase beta(1) subunit and the endogenous alpha(1) subunit was easily dissociated from the adherens junctions and cytoskeleton by the detergent extraction. The MDCK cell line in which half of the endogenous beta(1) subunits in the lateral membrane were substituted by unglycosylated beta(1) subunits displayed a decreased ability to form cell-to-cell contacts. Incubation of surface-attached MDCK cells with an antibody against the extracellular domain of the Na,K-ATPase beta(1) subunit specifically inhibited cell-cell contact formation. We conclude that the Na,K-ATPase beta(1) subunit is involved in the process of intercellular adhesion and is necessary for association of the heterodimeric Na,K-ATPase with the adherens junctions. Further, normal glycosylation of the Na,K-ATPase beta(1) subunit is essential for the stable association of the pump with the adherens junctions and plays an important role in cell-cell contact formation.

Delta-Notch signaling plays an essential role in cell fate determination in many tissue types, including the central nervous system. Although the signaling mechanism of Notch has been extensively studied, the behaviors of its ligands are not well understood. In the present study, we found that, in the developing neural tube, Dll1(Delta-like 1) was mainly localized on the processes extending from nascent neurons toward both the pia and the ventricle and accumulated at apical termini, where adherens junctions (AJs) were formed. To understand the mechanism of Dll1 localization, we searched for binding proteins for Dll1 and identified a scaffolding molecule, MAGI1. In the developing spinal cord, MAGI1 mRNA was highly expressed in the ventricular zone, where Dll1 mRNA was expressed. MAGI1 protein accumulated at the AJs formed around the termini of apically extending processes and was partially colocalized with Dll1. MAGI1 bound not only to Dll1 but also to N-cadherin-beta-catenin complexes. In cultured AJ-forming fibroblasts, MAGI1 was localized at AJs, and Dll1 was recruited to these AJs through binding to MAGI1. In addition, Dll1 was stabilized on the cell surface by MAGI1. Taken together, these results suggest that Dll1 is presented on the surface of AJs formed at the apical termini of processes through interaction with MAGI1 to activate Notch on neighboring cells in the developing central nervous system.

Chromatographic Assay for Determining the Effects of Microenvironment on Dimerization of Epithelial Cadherin K14E Mutant

Collective mechanical responses of cadherin-based adhesive junctions as predicted by simulations

Drosophila represents a paradigm for the analysis of the cellular, molecular and genetic mechanisms of development and is an ideal model system to study the contribution of Adherens Junctions (AJs) and their major components, cadherins, to morphogenesis. The combination of different techniques and approaches has allowed researchers to identify the requirements of these epithelial junctions in vivo in the context of a whole organism. The functional analysis of mutants for AJ core components, particularly for Drosophila DE-cadherin, has shown that AJs play critical roles in virtually all stages of development. For instance, AJs maintain tissue integrity while allowing the remodelling and homeostasis of many tissues. They control cell shape, contribute to cell polarity, facilitate cell-cell recognition during cell sorting, orient cell divisions, or regulate cell rearrangements, among other activities. Remarkably, these activities require a very fine control of the organisation and turnover of AJs during development. In addition, AJs engage in diverse and complex interactions with the cytoskeleton, signalling networks, intracellular trafficking machinery or polarity cues to perform these functions. Here, by summarising the requirements of AJs and cadherins during Drosophila morphogenesis, we illustrate the capital contribution of this model system to our knowledge of the mechanisms and biology of AJs.

Desmosomal Junctions Govern Tissue Integrity and Actomyosin Contractility in Apoptotic Cell Extrusion.

Complexities of Crumbs Function and Regulation in Tissue Morphogenesis