Cdc42 defines apical identity and regulates epithelial morphogenesis by promoting apical recruitment of Par6-aPKC and Crumbs

Abstract

Cdc42 regulates epithelial morphogenesis together with the Par complex (Baz/Par3-Par6-aPKC), Crumbs (Crb/CRB3) and Stardust (Sdt/PALS1). However, how these proteins work together and interact during epithelial morphogenesis is not well understood. To address this issue, we used the genetically amenable Drosophila pupal photoreceptor and follicular epithelium. We show that during epithelial morphogenesis active Cdc42 accumulates at the developing apical membrane and cell-cell contacts, independently of the Par complex and Crb. However, membrane localization of Baz, Par6-aPKC and Crb all depend on Cdc42. We find that although binding of Cdc42 to Par6 is not essential for the recruitment of Par6 and aPKC to the membrane, it is required for their apical localization and accumulation, which we find also depends on Par6 retention by Crb. In the pupal photoreceptor, membrane recruitment of Par6-aPKC also depends on Baz. Our work shows that Cdc42 is required for this recruitment and suggests that this factor promotes the handover of Par6-aPKC from Baz onto Crb. Altogether, we propose that Cdc42 drives morphogenesis by conferring apical identity, Par-complex assembly and apical accumulation of Crb.

Epidermal PAR-6 and PKC-3 are essential for postembryonic development ofCaenorhabditis elegansand control non-centrosomal microtubule organization

“…We also found that depletion of PAR-3 from the epidermis resulted in a partial loss of PAR-6. This is consistent with findings that multiple mechanisms can promote cortical recruitment of Par6-aPKC, including binding of Par6 to Par3 or Cdc42, and binding of aPKC to phospholipids (Dong et al, 2020;Hong et al, 2003;Hutterer et al, 2004;Joberty et al, 2000;Nagai-Tamai et al, 2002;Nunes de Almeida et al, 2019;Rodriguez et al, 2017;Wang et al, 2017;Wodarz et al, 2000). Conversely, the apical localization of PAR-3 was dependent upon PKC-3.…”

“…In mature epithelia, however, the bulk of Par3 segregates to the apical/lateral border, where it plays an essential role in the positioning and assembly of apical junctions (Achilleos et al, 2010;Georgiou et al, 2008;Harris and Peifer, 2004;Harris and Tepass, 2008;Izumi et al, 1998;Leibfried et al, 2008;Totong et al, 2007;Yamanaka et al, 2001). The release of Par3 in epithelia depends on phosphorylation of Par3 by aPKC, and involves handoff of Par6-aPKC to Cdc42 and the epithelial specific Crumbs polarity complex (Bilder et al, 2003;Harris and Peifer, 2005;Hong et al, 2003;Krahn et al, 2010;Morais-de-Sá et al, 2010;Nagai-Tamai et al, 2002;Nunes de Almeida et al, 2019;Walther and Pichaud, 2010).…”

The brachyceran de novo gene PIP82, a phosphorylation target of aPKC, is essential for proper formation and maintenance of the rhabdomeric photoreceptor apical domain in Drosophila

“…However, at 72 hrs APF, when two distinct apical domains are distinguishable, the intersection between PIP82 and Crumbs was greatly diminished; PIP82 localized to the base of the rhabdomere while Crumbs was concentrated to the stalk membrane with some Crumbs residual staining in the rhabdomere ( Fig 3E–3H ); PIP82 colocalization with Rhodopsin 1 (Rh1) at the base of the rhabdomere suggested PIP82 is not in the microvilli but rather found in the region of the rhabdomere terminal web [ 43 ] ( S7 Fig ). Moreover, utilizing an antibody that recognizes aPKC [ 33 ], we observed a non-overlapping spatial pattern of the PIP82 and aPKC domains at one day post eclosion ( Fig 3I–3L ). In this case, PIP82 demarcates the base of the rhabdomere and aPKC the stalk membrane of each photoreceptor.…”

“…Moreover, utilizing an antibody that recognizes aPKC [33], we observed a non-overlapping spatial pattern of the PIP82 and aPKC domains at one day post eclosion (Fig 3I-3L). In this case, PIP82 demarcates the base of the rhabdomere and aPKC the stalk membrane of each photoreceptor.…”

“…Moreover, the absence of aPKC results in photoreceptor cell death prior to differentiation [32]. The initial apical identity of photoreceptors is established by Cdc42 with the subsequent recruitment of Bazooka and the PAR6/aPKC complex [33]. In addition, the retention of aPKC by the transmembrane protein Crumbs at the apical surface reinforces the asymmetry between the apical and lateral membranes [34][35][36][37][38][39].…”