title: Rho-associated kinase ROCK activates LIM-kinase 1 by phosphorylation at threonine 508 within the activation loop.
abstract: LIM-kinase 1 (LIMK1) phosphorylates cofilin, an actin-depolymerizing factor, and regulates actin cytoskeletal reorganization. LIMK1 is activated by the small GTPase Rho and its downstream protein kinase ROCK. We now report the site of phosphorylation of LIMK1 by ROCK. In vitro kinase reaction revealed that the active forms of ROCK phosphorylated LIMK1 on the threonine residue and markedly increased its cofilin-phosphorylating activity. A LIMK1 mutant (T508A) with replacement of Thr-508 within the activation loop of the kinase domain by alanine was neither phosphorylated nor activated by ROCK. Replacement of Thr-508 by serine changed the ROCK-catalyzed phosphorylation residue from threonine to serine. A LIMK1 mutant with replacement of Thr-508 by two glutamates increased the kinase activity about 2-fold but was not further activated by ROCK. In addition, wild-type LIMK1, but not its T508A mutant, was activated by co-expression with ROCK in cultured cells. These results suggest that ROCK activates LIMK1 in vitro and in vivo by phosphorylation at Thr-508. Together with the recent finding that PAK1, a downstream effector of Rac, also activates LIMK1 by phosphorylation at Thr-508, these results suggest that activation of LIMK1 is one of the common targets for Rho and Rac to reorganize the actin cytoskeleton.
authors.affiliation: Biological Institute, Graduate School of Science, Tohoku University, Sendai 980-8578, Japan.
authors.forename: K
authors.initials: K
authors.lastname: Ohashi
authors.forename: K
authors.initials: K
authors.lastname: Nagata
authors.forename: M
authors.initials: M
authors.lastname: Maekawa
authors.forename: T
authors.initials: T
authors.lastname: Ishizaki
authors.forename: S
authors.initials: S
authors.lastname: Narumiya
authors.forename: K
authors.initials: K
authors.lastname: Mizuno
mesh_terms: D000818:Animals; D019556:COS Cells; D004789:Enzyme Activation; D047908:Intracellular Signaling Peptides and Proteins; D054461:Lim Kinases; D016297:Mutagenesis, Site-Directed; D009447:Neuroblastoma / Q000378:metabolism*; D010766:Phosphorylation; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D013912:Threonine; D020741:rho GTP-Binding Proteins / Q000378:metabolism*; D054460:rho-Associated Kinases
chemical_list: D047908:Intracellular Signaling Peptides and Proteins; D013912:Threonine; D011494:Protein Kinases; D054461:Lim Kinases; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; D020741:rho GTP-Binding Proteins