Phosphorylation and activation of myosin by Rho-associated kinase (Rho-kinase)

J Biol Chem. 1996 Aug 23;271(34):20246-9. doi: 10.1074/jbc.271.34.20246.

Abstract

The small GTPase Rho is implicated in physiological functions associated with actin-myosin filaments such as cytokinesis, cell motility, and smooth muscle contraction. We have recently identified and molecularly cloned Rho-associated serine/threonine kinase (Rho-kinase), which is activated by GTP Rho (Matsui, T., Amano, M., Yamamoto, T., Chihara, K., Nakafuku, M., Ito, M., Nakano, T., Okawa, K., Iwamatsu, A., and Kaibuchi, K. (1996) EMBO J. 15, 2208-2216). Here we found that Rho-kinase stoichiometrically phosphorylated myosin light chain (MLC). Peptide mapping and phosphoamino acid analyses revealed that the primary phosphorylation site of MLC by Rho-kinase was Ser-19, which is the site phosphorylated by MLC kinase. Rho-kinase phosphorylated recombinant MLC, whereas it failed to phosphorylate recombinant MLC, which contained Ala substituted for both Thr-18 and Ser-19. We also found that the phosphorylation of MLC by Rho-kinase resulted in the facilitation of the actin activation of myosin ATPase. Thus, it is likely that once Rho is activated, then it can interact with Rho-kinase and activate it. The activated Rho-kinase subsequently phosphorylates MLC. This may partly account for the mechanism by which Rho regulates cytokinesis, cell motility, or smooth muscle contraction.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Base Sequence
  • Cattle
  • Cell-Free System
  • Chickens
  • DNA Primers / chemistry
  • Enzyme Activation
  • Intracellular Signaling Peptides and Proteins
  • Molecular Sequence Data
  • Myosin Light Chains / metabolism*
  • Myosins / metabolism*
  • Peptide Mapping
  • Phosphorylation
  • Protein-Serine-Threonine Kinases / metabolism*
  • Rabbits
  • rho-Associated Kinases

Substances

  • DNA Primers
  • Intracellular Signaling Peptides and Proteins
  • Myosin Light Chains
  • Protein-Serine-Threonine Kinases
  • rho-Associated Kinases
  • Myosins