Coordinate remodeling of actin filaments and membranes occurs during many cellular processes. One protein that links the actin cytoskeleton and membranes is the GTPase dynamin. Dynamin is best characterized for its role during clathrin-dependent endocytosis where it participates in the formation of nascent endocytic vesicles by constricting membrane tubules and promoting fission or vesicle separation. However, whereas dynamin alone can promote formation of lipid tubules in vitro, it does not efficiently catalyze fission. Recent work suggested that factors that impart longitudinal tension on the membrane tubules assist in dynamin-induced fission. Actin filaments also participate in endocytosis, and may function, in part, to provide tension on membranes that facilitates efficient fission by dynamin. In addition, dynamin localizes with actin regulators at sites of actin filament remodeling. I propose that dynamin GTPase activity regulates actin filament dynamics, an activity that may also be coordinated with dynamin's ability to remodel membranes, and is directly involved in tumor cell growth and metastasis. The following specific aims will guide this research program: 1) Specific Aim 1. Determine the mechanism by which dynamin2 influences actin filament dynamics and filament organization. I will use biochemical and microscopy approaches to determine the mechanisms by which dynamin2 influences filament organization and filament turnover;2) Specific Aim 2. Determine the mechanism by which actin filaments influence dynamin2-dependent formation of membrane tubules and fission. I will use optical and electron microscopy, together with biochemical approaches, to determine how actin filament bundles formed by cortactin and Arp2/3 complex influence membrane fission by dynamin2;3) Specific Aim 3. Determine the effects of perturbing dynamin2 on cell migration and lamellipodia formation at the leading edge of migrating cells. I will overexpress dominant-negative dynamin2 mutants in migrating cells and determine the effects on overall cell motility and lamellipodia dynamics.
TO PUBLIC HEALTH: Tumor cells rapidly proliferate and migrate to other tissues. These two processes may be linked by the protein dynamin. The goal of the research outlined in this proposal is to determine the mechanism by which dynamin coordinately interacts with cellular membranes and the cell migration machinery, which may lead to the development of alternative therapies against tumor cell growth and metastasis.
| Mooren, Olivia L; Li, Jinmei; Nawas, Julie et al. (2014) Endothelial cells use dynamic actin to facilitate lymphocyte transendothelial migration and maintain the monolayer barrier. Mol Biol Cell 25:4115-29 |
| Mooren, Olivia L; Galletta, Brian J; Cooper, John A (2012) Roles for actin assembly in endocytosis. Annu Rev Biochem 81:661-86 |
| Galletta, Brian J; Mooren, Olivia L; Cooper, John A (2010) Actin dynamics and endocytosis in yeast and mammals. Curr Opin Biotechnol 21:604-10 |
| Mooren, Olivia L; Schafer, Dorothy A (2009) Constricting membranes at the nano and micro scale. Proc Natl Acad Sci U S A 106:20559-60 |
| Mooren, Olivia L; Kotova, Tatyana I; Moore, Andrew J et al. (2009) Dynamin2 GTPase and cortactin remodel actin filaments. J Biol Chem 284:23995-4005 |
