Investigating the F-role of actin disassembly during Drosophila gastrulation

Massachusetts Institute Of Technology, Cambridge MA

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Abstract

? DESCRIPTION (provided by applicant): Tissue morphogenesis, the process that alters the shape of a tissue, is essential for embryonic development. Defects in tissue morphogenesis result in failed organ function and/or formation, as well as the progression of epithelial tumors, thus understanding the mechanism that promotes/regulates morphogenesis is critical. Some morphogenetic events are facilitated by apical constriction, a process that alters a columnar- shaped cell to a wedged-shaped cell. Defects in F-actin inhibit apical constriction, though an understanding of how F-actin dynamics regulate this process is lacking. Current models suggest that stable F-actin promotes apical constriction. During Drosophila gastrulation, apical constriction promotes epithelial bending, facilitating epithelial internalization and tube formatio within the embryo. Available tools and easily visualized morphological events by live imaging, makes Drosophila gastrulation an ideal system for investigating the role(s) for F-actin dynamics during apical constriction and tissue morphogenesis. To investigate how F-actin regulates morphogenesis, our lab has performed an RNAi screen that targeted 66 Drosophila genes that regulate F-actin during gastrulation. This screen identified essential roles for F-actin disassembly in mediating tissue morphogenesis. Loss of F-actin disassembly factors resulted in apical constriction, cell internalization, and/or tissue fusion defect, suggesting that dynamic actn is essential for these processes. F-actin and Myosin II generate apical constrictions in a pulsatile manner; each constriction is stabilized prior to the next. During constrictions, an increase of apical F-actin is observed. During stabilization, this F-actin concentration is lost an apical F-actin cables are present. I hypothesize that apically localized F-actin is disassembled and remodeled into F-actin cables to promote apical constriction. To test my hypothesis, I will determine if F-actin cable formation and apical constriction occurs properly in mutants with hindered F-actin disassembly using high-resolution, live imaging. Additionally, I will further elucidate the role for F-actin during morphogenesis by evaluating how F-actin organization promotes apical constriction. Drosophila abelson (abl) mutants have over abundant, disorganized apical F-actin. abl mutants also exhibit defects in apical constriction, thus I hypothesize that proper F-actin organization are essential for this processes. To test this hypothesis, I will elucidate the mechanism by which Abl promotes F-actin organization via live imaging and quantitative analysis techniques, gaining insight into how F-actin organization promotes apical constriction. Finally, the spatial and temporal regulation of F-actin dynamics during a dynamic process, such as apical constriction, is poorly understood. To further extend analysis of F-actin disassembly to apical constriction and later developmental stages, I will gain in vivo temporal control of the F-actin severing protein cofilin by acute inactivation through relocalization and sequestration. Cofilin inhibition during distinct developmental stages will gain a further understanding of how F- actin disassembly promotes morphogenesis.