MYOSIN VI FUNCTION AND MECHANISM

Washington University, Saint Louis MO

Investigators

Linked publications & trials

Abstract

DESCRIPTION (provided by applicant): This application for Competitive Revision Supplement is in response to NOT-OD-09-058, under the ARRA NIH announcement: NIH Announces the Availability of Recovery Act Funds for Competitive Revision Applications. In this application we propose to expand the scope of our currently funded project R01 GM060494 entitled "Myosin VI function and mechanism." Actin structures play many important roles in cells of multi-cellular organisms that go beyond participating in cell motility. They are crucial in generating differentiated cell form and mediating specialized cell function. Such structures are organized based on structural motifs found in motile cells, such as meshwork and parallel bundles, but often have very different kinetics of formation and turnover and perform very different functions, requiring different mechanical and physical properties. We are studying interesting example of a specialized actin structure important in cellular differentiation is the cellular remodeling that occurs during spermatid individualization in Drosophila. During this process a group of 64 spermatids interconnected by cytoplasmic bridges are separated into individual sperm. Cone-shaped actin structures move along the length of the highly elongated cell, remodeling the membrane and pushing out cellular contents and the bulk of the cytoplasm. This actin structure is extremely dense, so it can push the cytoplasmic contents out of the cell. Myosin VI, an actin-based motor protein, plays a key role in building the dense actin meshwork that forms the cone front. When it is absent, insufficient actin accumulates, the cones don't function properly, and the removal of cytoplasm and organelles is defective. Myosin VI's role is to stabilize the actin meshwork. In building this interesting actin structure, myosin VI cooperates with the arp 2/3 complex, which is important for meshwork formation. Profilin also is required, being important for assembling the bundles that form the rear of the cones, probably in collaboration with a formin. Other proteins are located on the actin cones, including fascin (singed), villin (quail), cortactin, and dynamin. In our currently funded work (R01GM060494), we focus on understanding myosin VI's mechanism of action in its actin stabilization role. However, we hypothesize that other proteins play important roles in generating this interesting structure and we propose to extend the scope of our work by screening an existing collection of male sterile mutants to identify these other proteins. We will visually screen for mutants with defects in actin cone structure and myosin VI localization. Once a collection of genes important in this process is identified, we will examine how the proteins work together to generate this structure and ask whether similar mechanisms are used in other cells and processes. Since in other situations dense actin structures are important for localizing cellular components or structural support, the information we obtain about this process may be relevant beyond Drosophila spermatogenesis. The work proposed here will extend the scope of our funded work, result in the hiring of additional personnel, the purchase of equipment and supplies, involve the contracting of services for generation and analysis of DNA sequence data, and accelerate the pace of scientific discovery. . PUBLIC HEALTH RELEVANCE: Actin structures play important roles in all cells and are the key cellular components that give differentiated cells their shape and organization. Defects in these structures and in actin organization and dynamics underlie many aspects of disease. Our studies will identify important regulators of actin assembly and organization in particular specialized cell types. By studying these regulators and how they help in formation of the correct structures at the correct time, in the correct place and with the correct form, a better understanding of the basic cellular processes underlying normal cell function and disease will emerge.