Coordinate Role of Vinculin and Metavinculin in Actin Organization

Abstract

Vinculin (Vcn) is an essential cytoskeletal protein that acts as a scaffold to link transmembrane receptors to actin filaments, thereby playing a crucial role in cell adhesion, motility, and force transmission between cells. While Vcn is ubiquitously expressed, metavinculin (MVcn), a larger isoform of Vcn, is selectively expressed in smooth and cardiac muscle cells. Similar to Vcn, MVcn can directly associate with actin and remodel the actin cytoskeleton. However, distinct from Vcn, MVcn contains an additional exon which encodes a 68-residue insert. Point mutations in the 68-residue insert have been associated with altered actin organization and heart disease, notably dilated cardiomyopathy (DCM) and hypertrophic cardiomyopathy (HCM). To better understand the coordinate role of MVcn and Vcn in actin filament assembly and the consequences of cardiomyopathy-related mutations, we conducted a series of actin co-sedimentation and negative stain EM experiments. Consistent with previous findings, the tail domain of MVcn (MVt) is unable to induce actin bundling. In the presence of sub-stoichiometric amounts MVt relative to Vcn tail domain (Vt), MVt inhibits the assembly of actin filaments into parallel bundles. In contrast to wild type MVt, MVt CM mutants induce the formation of disordered higher-order F-actin assemblies. To probe the molecular basis for differences in actin reorganization observed for MVt and MVt CM mutants, and negative regulation of Vt-mediated actin bunding by MVt, we experimentally examined computational models of actin-induced MVt conformational changes. Results from these studies will be presented.