Crb3 stabilizes activated Ezrin-Radixin-Moesin to organize the apical domain of multiciliated cells

Cell shape changes mainly rely on the remodeling of the actin cytoskeleton. Multiciliated cells (MCCs) of the mucociliary epidermis of Xenopus laevis embryos, as they mature, dramatically reshape their apical domain to grow cilia, in coordination with the underlying actin cytoskeleton. Crumbs (Crb) proteins are multifaceted transmembrane apical polarity proteins known to recruit actin linkers and promote apical membrane growth. Here, we identify the homeolog Crb3.L as an important player for apical domain morphogenesis in differentiating Xenopus MCCs. We found that Crb3.L is initially present in cytoplasmic vesicles in the vicinity of ascending centrioles/basal bodies (BBs), then at the expanding apical membrane concomitantly with BB docking, and finally in the ciliary shaft of growing and mature cilia. Using morpholino-mediated knockdown, we show that Crb3.L-depleted MCCs display a complex phenotype associating reduction in the apical surface, disorganization of the apical actin meshwork, centriole/BB migration defects, as well as abnormal ciliary tuft formation. Based on prior studies, we hypothesized that Crb3.L could regulate Ezrin-Radixin Moesin (ERM) protein subcellular localization in MCCs. Strikingly, we observed that endogenous phospho-activated ERM (pERM) is recruited to the growing apical domain of inserting MCCs, in a Crb3.L-dependent manner. Our data suggest that Crb3.L recruits and/or stabilizes activated pERM at the emerging apical membrane to allow coordinated actin-dependent expansion of the apical membrane in MCCs.