Collaborative Research: Regulation of Cellular Mechanics by Crosslinked Actin Networks - Role of Palladin and Alpha-actinin

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Intellectual Merit Cells in the body have a remarkable ability to sense "stiffness" in their environment - that is, a cell can distinguish between a hard substrate such as bone, and a softer substrate such as brain. This ability is critical for many aspects of cell function such as migration, wound healing and the proper formation of tissues and organs. In order to sense the mechanical properties of their environment, cells adjust their internal stiffness to match that of the external surface by reorganizing their internal cytoskeleton, which is made of a dynamic network of microscopic filaments. The main component of these filaments is a protein called actin. Various actin-binding proteins crosslink the actin filaments and organize them into multifilament bundles. Palladin is a newly identified actin cross-linking protein that is important in organizing these filament networks. Previous experiments have shown that palladin plays an essential role in embryonic development: when the palladin gene is silenced in mice, it results in lethal development defects that are characterized by a failure of cells to adhere properly to a substrate and to migrate appropriately. These deficits may arise because the cell's ability to adjust its internal stiffness is compromised in the absence of palladin. This proposal will test the hypothesis that palladin, by its ability to cross-link actin and its interaction with another actin cross-linker, alpha-actinin, determines the structure and mechanical properties of actin networks and enables the cell to sense its physical environment. Two types of approaches will be used to address this question. First, the structural and mechanical properties of actin networks assembled on a glass slide will be measured to elucidate how actin cross-linking by palladin contributes to actin organization. In addition, palladin levels will be genetically manipulated in living cells to study how altered actin organization, cellular stiffness and force generation impacts cellular mechano-sensitivity. Broader Impact This collaborative proposal will enhance the understanding of essential biological processes that underlie cell movement and tissue formation. The training of graduate and undergraduate students in interdisciplinary approaches from Physics and Cell Biology will be an integral part of the work. The cell lines that will be developed as part of this project will be made freely available to other investigators following their publication. A graduate course in Cell Mechanics will be developed based on the conceptual framework of this proposal. The PI and co-PI will also encourage minority students as well as high school students from the area to participate in research as part of the Louis Stokes Alliance for Minority Participation program at the University of Maryland and the University of North Carolina Research Apprenticeship Program. The PI will organize a one week biophysics laboratory demonstration as part of the Summer Girls Program in the Department of Physics to encourage participation of female students in science and technology fields.