Rap1 regulates apical contractility to allow embryonic morphogenesis without tissue disruption and acts in part via Canoe-independent mechanisms.

Embryonic morphogenesis is powered by dramatic changes in cell shape and arrangement driven by the cytoskeleton and its connections to adherens junctions. This requires robust linkage allowing morphogenesis without disrupting tissue integrity. The small GTPase Rap1 is a key regulator of cell adhesion, controlling both cadherin-mediated and integrin-mediated processes. We have defined multiple roles in morphogenesis for one Rap1 effector, Canoe/Afadin, which ensures robust junction-cytoskeletal linkage. We now ask what mechanisms regulate Canoe and other junction-cytoskeletal linkers during Drosophila morphogenesis, defining roles for Rap1 and one of its guanine nucleotide exchange factor (GEF) regulators, Dizzy. Rap1 uses Canoe as one effector, regulating junctional planar polarity. However, Rap1 has additional roles in junctional protein localization and balanced apical constriction-in its absence, Bazooka/Par3 localization is fragmented, and cells next to mitotic cells apically constrict and invaginate, disrupting epidermal integrity. In contrast, the GEF Dizzy has phenotypes similar to but slightly less severe than Canoe loss, suggesting that this GEF regulates Rap1 action via Canoe. Taken together, these data reveal that Rap1 is a crucial regulator of morphogenesis, likely acting in parallel via Canoe and other effectors, and that different Rap1 GEFs regulate distinct functions of Rap1.

Multivalent interactions make adherens junction-cytoskeletal linkage robust during morphogenesis.

Embryogenesis requires cells to change shape and move without disrupting epithelial integrity. This requires robust, responsive linkage between adherens junctions and the actomyosin cytoskeleton. Using Drosophila morphogenesis, we define molecular mechanisms mediating junction-cytoskeletal linkage and explore the role of mechanosensing. We focus on the junction-cytoskeletal linker Canoe, a multidomain protein. We engineered the canoe locus to define how its domains mediate its mechanism of action. To our surprise, the PDZ and FAB domains, which we thought connected junctions and F-actin, are not required for viability or mechanosensitive recruitment to junctions under tension. The FAB domain stabilizes junctions experiencing elevated force, but in its absence, most cells recover, suggesting redundant interactions. In contrast, the Rap1-binding RA domains are critical for all Cno functions and enrichment at junctions under tension. This supports a model in which junctional robustness derives from a large protein network assembled via multivalent interactions, with proteins at network nodes and some node connections more critical than others.