An Arf-GEF Pathway for Regulating Actomyosin at Adherens Junctions: the Role of an Adaptor Protein

Cytoskeletal regulation is essential for morphogenesis during development. One down-regulator of actomyosin in the Drosophila embryo is the Arf-GEF Steppke. I investigated how Steppke is recruited to myosin-enriched adherens junctions. Its coiled-coil (CC) domain was shown to be necessary and sufficient for Steppke localization, while its PH domain was found to be dispensable. Homodimerization of Steppke’s CC domain also had minimal effect. Thus, I pursued Steppke’s adaptor, Stepping stone (Sstn). The proteins co-localized in vivo, Sstn was required for Step localization, and the proteins interacted directly through hydrophobic residues of their respective CC domains in vitro. A separate conserved region (CR) in Sstn was necessary and sufficient for Sstn localization, while its CC domain was dispensable. Remarkably, replacing the CC domain of Steppke with the CR of Sstn conveyed proper localization to Steppke. Thus, Steppke is recruited to actomyosin-rich adherens junctions through direct interaction with the adaptor protein Sstn.M.Sc.2020-11-15 00:00:0