Studies on nonmuscle myosin-IIs: From subunit assembly to human disease; from contractility to cell sheet migration

Myosins play important cell biological roles throughout phylogeny. In D. melanogaster, nonmuscle myosin-II function, provided by zip/MyoII is necessary for dorsal closure, an essential cell sheet movement in embryogenesis. I investigated the tissue specific requirements of zip/MyoII function and show that this motor protein is responsible for generating the forces necessary for cell shape changes in both amnioserosa cells and in the dorsal-most cells of the lateral epidermis. Restoration of nonmuscle myosin-II function either ubiquitously, in just the amnioserosa, or in just the dorsal most cells of the lateral epidermis is sufficient to rescue dorsal closure. Additionally, uneven nonmuscle myosin-II contractility along the lateral epidermis during dorsal closure results in the misalignment of epidermal stripes.; Native, nonmuscle myosin-II is composed of two heavy chains, two regulatory light chains and two essential light chains. Little is known regarding the stability of the native nonmuscle myosin-II complex under genetic conditions that cause one of the three polypeptides to be absent. Here, I show that in D. melanogaster both the heavy chain and regulatory light chain polypeptide levels depend on the presence of each other. In contrast, essential light chain levels were unaffected when either heavy chain or regulatory light chain were reduced. Novel D. melanogaster essential light chain binding partners were identified, including: myosin VIIA, myosin VI, myosin V and a microtubule associating protein called abnormal spindle.; In humans, MYH9-related disorders are dominant hematological disorders resulting from mutations in a homologue of the fly zip/MyoII gene (nonmuscle myosin-IIA). We showed that four common rod mutations directly affect the structure and assembly properties of individual heavy chain molecules. Nevertheless, when four MYH9 rod mutations were engineered into zip/MyoII these failed to cause similar structural and assembly defects as observed for the human heavy chain. In rescue experiments, mutant zip/MyoII heavy chains have wild type activity.; To better understand how other myosin superfamily members function, I made antibodies, GFP labeled rescue and dominant negative constructs for ck/MyoVIIA; whose human homologue is essential for hearing. These constructs provided key reagents for studies on ck/MyoVIIA function in hairs and bristles (Kiehart et al., Genetics 2004) and for its essential contribution to fly hearing (Todi et al., Curr Biol 2005). Together my studies connect the power of fly genetics with in vivo imaging and biochemical analysis to provide novel insights into myosin function.