WAVE facilitates polarized E-cadherin transport.

Cadherin dynamics drive morphogenesis, while defects in cadherin polarity contribute to diseases, including cancers. However, the forces polarizing cadherin membrane distribution are not well understood. We previously showed that WAVE-dependent branched actin polarizes cadherin distribution and suggested that one mechanism is protein transport. While previous studies suggested that WAVE is enriched at various endocytic organelles, the role of WAVE in protein traffic is understudied. Here we test the model that WAVE regulates cadherin by polarizing its transport. In support of this model we show that 1) endogenously tagged WAVE accumulates in vivo at several endocytic organelles, including recycling endosomes and at the Golgi; 2) likewise, cadherin protein accumulates at recycling endosomes and the Golgi; 3) loss of WAVE components reduces cadherin accumulation at apically directed RAB-11-positive recycling endosomes and increases accumulation at the Golgi. In addition, live imaging illustrates that dynamics and velocity of recycling endosomes enriched for RAB-11::GFP and RFP::RME-1 are reduced in animals depleted of WAVE components and RAB-11::GFP movements are misdirected, suggesting that WAVE powers and directs their movements. This in vivo study demonstrates the importance of WAVE in promoting polarized transport in epithelia and supports a model that WAVE promotes cell-cell adhesion and polarity by promoting cadherin transport.

RhoGAP RGA-8 supports morphogenesis in C. elegans by polarizing epithelia.

CDC-42 regulation of non-muscle myosin/NMY-2 is required for polarity maintenance in the one-cell embryo of Caenorhabditis elegans CDC-42 and NMY-2 regulate polarity throughout embryogenesis, but their contribution to later events of morphogenesis are less understood. We have shown that epidermal enclosure requires the GTPase CED-10/Rac1 and WAVE/Scar complex, its effector, to promote protrusions that drive enclosure through the branch actin regulator Arp2/3. Our analysis here of RGA-8, a homolog of SH3BP1/Rich1/ARHGAP17/Nadrin, with BAR and RhoGAP motifs, suggests it regulates CDC-42, so that actin and myosin/NMY-2 promote ventral enclosure during embryonic morphogenesis. Genetic and molecular data suggest RGA-8 regulates CDC-42, and phenocopies the CDC-42 pathway regulators WASP-1/WSP-1 and the F-BAR proteins TOCA-1 and TOCA-2. Live imaging shows RGA-8 and WSP-1 enrich myosin and regulate F-actin in migrating epidermal cells during ventral enclosure. Loss of RGA-8 alters membrane recruitment of active CDC-42. We propose TOCA proteins and RGA-8 use BAR domains to localize and regenerate CDC-42 activity, thus regulating F-actin levels, through the branched actin regulator WSP-1, and myosin enrichment. RhoGAP RGA-8 thus polarizes epithelia, to promote cell migrations and cell shape changes of embryonic morphogenesis.

WAVE regulates Cadherin junction assembly and turnover during epithelial polarization.

Actin is an integral component of epithelial apical junctions, yet the interactions of branched actin regulators with apical junction components are still not clear. Biochemical data have shown that α-catenin inhibits Arp2/3-dependent branched actin. These results suggested that branched actin is only needed at earliest stages of apical junction development. We use live imaging in developing C. elegans embryos to test models for how WAVE-induced branched actin collaborates with other apical junction proteins during the essential process of junction formation and maturation. We uncover both early and late essential roles for WAVE in apical junction formation. Early, as the C. elegans intestinal epithelium becomes polarized, we find that WAVE components become enriched concurrently with the Cadherin components and before the DLG-1 apical accumulation. Live imaging of F-actin accumulation in polarizing intestine supports that the Cadherin complex components and branched actin regulators work together for apical actin enrichment. Later in junction development, the apical accumulation of WAVE and Cadherin components is shown to be interdependent: Cadherin complex loss alters WAVE accumulation, and WAVE complex loss increases Cadherin accumulation. To determine why Cadherin levels rise when WVE-1 is depleted, we use FRAP to analyze Cadherin dynamics and find that loss of WAVE as well as of the trafficking protein EHD-1/RME-1 increases Cadherin dynamics. EM studies in adults depleted of branched actin regulators support that WVE-1 maintains established junctions, presumably through its trafficking effect on Cadherin. Thus we propose a developmental model for junction formation where branched actin regulators are tightly interconnected with Cadherin junctions through their previously unappreciated role in Cadherin transport.