Alpha-catenin-dependent recruitment of the centrosomal protein CAP350 to adherens junctions allows epithelial cells to acquire a columnar shape.

Maria P Gavilan,Marina Arjona,Michel Bornens,Rosa M Rios,Etienne Formstecher,Angel Zurbano,Juan R Martinez-Morales,Barry M Gumbiner

doi:10.1371/journal.pbio.1002087

Abstract

Epithelial morphogenesis involves a dramatic reorganisation of the microtubule cytoskeleton. How this complex process is controlled at the molecular level is still largely unknown. Here, we report that the centrosomal microtubule (MT)-binding protein CAP350 localises at adherens junctions in epithelial cells. By two-hybrid screening, we identified a direct interaction of CAP350 with the adhesion protein α-catenin that was further confirmed by co-immunoprecipitation experiments. Block of epithelial cadherin (E-cadherin)-mediated cell-cell adhesion or α-catenin depletion prevented CAP350 localisation at cell-cell junctions. Knocking down junction-located CAP350 inhibited the establishment of an apico-basal array of microtubules and impaired the acquisition of columnar shape in Madin-Darby canine kidney II (MDCKII) cells grown as polarised epithelia. Furthermore, MDCKII cystogenesis was also defective in junctional CAP350-depleted cells. CAP350-depleted MDCKII cysts were smaller and contained either multiple lumens or no lumen. Membrane polarity was not affected, but cortical microtubule bundles did not properly form. Our results indicate that CAP350 may act as an adaptor between adherens junctions and microtubules, thus regulating epithelial differentiation and contributing to the definition of cell architecture. We also uncover a central role of α-catenin in global cytoskeleton remodelling, in which it acts not only on actin but also on MT reorganisation during epithelial morphogenesis.

Highlights

Epithelial polarisation involves a coordinated series of events resulting in the asymmetric segregation of plasma membrane into apical and basolateral domains
Epithelial differentiation requires the coordination in space and time of several mechanisms that lead to the acquisition of distinctive epithelial features, including apical-basal polarity, specialised cell
By using a previously characterised goat polyclonal antibody (Fig. 1A, CAP350g), we found that in methanol- or paraformaldehyde (PFA)-fixed Madin-Darby canine kidney II (MDCKII) epithelial cells, CAP350 is essentially detected at the CTR (Fig. 1B, left panel)

Summary

Introduction

Epithelial polarisation involves a coordinated series of events resulting in the asymmetric segregation of plasma membrane into apical and basolateral domains. The establishment of apicobasal polarity is accompanied by asymmetric distribution of intracellular organelles, cytoskeleton reorganisation, and polarised membrane trafficking [1]. MTs are prominently organised in bundles aligned along the apicobasal axis with their minus ends oriented toward the apical membrane and plus ends toward the basal membrane and as networks of mixed polarity underneath the apical and basal membranes. The molecular events underlying MT reorganisation and their contribution to epithelial morphogenesis remain unclear. These mechanisms seem to be different depending on the tissue. Cadherins form an adhesive interface all along the lateral domain but are organised into special complexes known as zonula adherens (ZA) that are localised at the apical end. The molecular players underlying the AJ–MT connection remain, mostly unknown

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