title: Enhancement of mDia2 activity by Rho-kinase-dependent phosphorylation of the diaphanous autoregulatory domain.
abstract: It is clear that RhoA activates the DRF (diaphanous-related formin) mDia2 by disrupting the molecular interaction between the DAD (diaphanous autoregulatory domain) and the DID (diaphanous inhibitory domain). Previous studies indicate that a basic motif within the DAD contributes to mDia2 auto-inhibition, and results shown in the present study suggest these residues bind a conserved acidic region within the DID. Furthermore, we demonstrate that mDia2 is phosphorylated by ROCK (Rho-kinase) at two conserved residues (Thr(1061) and Ser(1070)) just C-terminal to the DAD basic region. Phosphomimetic mutations to these residues in the context of the full-length molecule enhanced mDia2 activity as measured by increased actin polymerization, SRF (serum response factor)-dependent smooth muscle-specific gene transcription, and nuclear localization of myocardin-related transcription factor B. Biochemical and functional data indicate that the T1061E/S1070E mutation significantly inhibited the ability of DAD to interact with DID and enhanced mDia2 activation by RhoA. Taken together, the results of the present study indicate that ROCK-dependent phosphorylation of the mDia2 DAD is an important determinant of mDia2 activity and that this signalling mechanism affects actin polymerization and smooth muscle cell-specific gene expression.
authors.affiliation: Department of Pathology and Laboratory Medicine, University of North Carolina, Chapel Hill, NC 27599, USA.
authors.forename: Dean P
authors.initials: DP
authors.lastname: Staus
authors.forename: Joan M
authors.initials: JM
authors.lastname: Taylor
authors.forename: Christopher P
authors.initials: CP
authors.lastname: Mack
mesh_terms: D000818:Animals; D002460:Cell Line; D005455:Fluorescent Antibody Technique; D047468:Immunoprecipitation; D051379:Mice; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism*; D009252:NADPH Dehydrogenase / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D017434:Protein Structure, Tertiary; D054460:rho-Associated Kinases / Q000235:genetics / Q000378:metabolism*
chemical_list: D008869:Microtubule-Associated Proteins; C508552:Dia2 protein, mouse; D009252:NADPH Dehydrogenase; D054460:rho-Associated Kinases