Interaction of Rho-kinase with myosin II at stress fibres.
Rho-kinase and myosin phosphatase cooperatively regulate the phosphorylation level of myosin light chain and are involved in the formation of stress fibres and smooth muscle contraction. Rho-kinase has been known to be localized at stress fibres, but little is known about the mechanism of its localization. Here we identified non-muscle myosin heavy chain IIA and IIB as the pleckstrin homology domain-interacting molecules by affinity column chromatography. The pleckstrin homology domain of Rho-kinase binds to myosin II directly in in vitro cosedimentation assay. The C-terminal region of the pleckstrin homology domain was important for this interaction, and the point mutations in the pleckstrin homology domain mutant (W1170A, W1340L) resulted in a decrease in the binding. We also found that the pleckstrin homology domain, but not the pleckstrin homology domain mutant (W1170A, W1340L), was localized at stress fibres in fibroblasts. These results indicate that Rho-kinase is localized at stress fibres through binding of the pleckstrin homology domain to myosin II.
Kawabata
Saeko
S
Department of Cell Pharmacology, Graduate School of Medicine, Nagoya University, Nagoya, Aichi 466-8550, Japan.
Usukura
Jiro
J
Morone
Nobuhiro
N
Ito
Masaaki
M
Iwamatsu
Akihiro
A
Kaibuchi
Kozo
K
Amano
Mutsuki
M
D000818:Animals; D001483:Base Sequence; D002846:Chromatography, Affinity; D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D005455:Fluorescent Antibody Technique; D047908:Intracellular Signaling Peptides and Proteins; D051379:Mice; D008854:Microscopy, Electron; D024462:Myosin Type II / Q000378:metabolism*; D041681:NIH 3T3 Cells; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D054460:rho-Associated Kinases
D017931:DNA Primers; D047908:Intracellular Signaling Peptides and Proteins; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; D024462:Myosin Type II