View Program - 63rd Annual Drosophila Research Conference

During embryonic development, epithelial cells must change shape and move while maintaining a robust linkage between one another. In Drosophila, multiple molecular mechanisms are involved in maintaining this linkage between adherens junctions and the cytoskeleton. Our lab has focused on the role of the junction-cytoskeletal linker Canoe, which regulates diverse events in embryonic morphogenesis, from apical-basal polarity establishment to apical constriction to convergent elongation and collective cell migration. Canoe is activated by the small GTPase Rap1, which in turn is regulated by diverse Guanine nucleotide exchange factors (GEFs). We and others have defined the roles Rap1 plays during cellurization and mesoderm invagination, but little is known about its role in later embryonic morphogenesis. We are thus exploring the roles of Rap1, and those of one of its GEF regulators, Dizzy, contrasting these with roles of the Rap1 effector Canoe. We used maternally driven shRNAi to deplete both maternal and zygotic Dizzy. Our data reveal Dizzy is required to reinforce junctions under tension during germband extension, thus making Dizzy loss comparable to canoe maternal/zygotic mutants. Dizzy knockdown also altered junctional planar polarity, though not to the same extent as Canoe. Later in development the dizzy mutant epidermis remains relatively intact, with the most severe defects localized to the ventral epidermis, as seen previously in canoe mutants. Together these data suggest that Dizzy is the predominant GEF regulating Canoe via Rap1. We’re now extending this analysis to explore the role of Rap1 in morphogenetic movements. Our preliminary data suggest that Rap1 mutants have substantially more severe defects in epithelial integrity. Cell shapes are altered beginning at the onset of germband extension. The balance of apical contractility between different cells appears to be lost, leading to epithelial folding, and by the end of germband extension epithelial junctions become fragmented in many cells. These data suggest Rap1 must have other effectors in addition to Canoe—intriguingly, the Rap1 phenotypes are more similar to those of embryos lacking both Canoe and Pyd (fly ZO-1) function. These data also suggest additional GEFs must regulate Rap1 during these stages. Together, our data provide new insights into the mechanisms regulating cell shape change and ensuring tissue integrity.

Defining the roles of the small GTPase Rap1 and its regulator Dizzy in embryonic morphogenesis