TY - JOUR
T1 - Rho-kinase phosphorylates eNOS at threonine 495 in endothelial cells
AU - Sugimoto, Masayuki
AU - Nakayama, Masanori
AU - Goto, Takaaki M.
AU - Amano, Mutsuki
AU - Komori, Kimihiro
AU - Kaibuchi, Kozo
N1 - Funding Information:
We thank Dr. Toshio Hayashi (Department of Geriatrics, Nagoya University, Japan) for the pBK-CMV-eNOS; Dr. Tsunehiko Higuchi (Graduate School of Pharmaceutical Sciences, Nagoya City University, Japan) for baculovirus transfer vectors for human eNOS; Dr. Yasuo Watanabe (Showa Pharmaceutical University, Tokyo, Japan) and Dr. Takeshi Urano (Department of Biochemistry Shimane University, Shimane, Japan) for helpful discussions; and Kaibuchi laboratory staff for technical aid. This research was supported in part by Grants-in-Aid for Scientific Research from the Ministry of Education, Culture, Sport, Science and Technology of Japan (MEXT), a Grants-in-Aid for Creative Scientific Research from MEXT, the 21st Century Center of Excellence Program, a Research Grant for Nervous and Mental Disorders from the Ministry of Health and Welfare; and the Program for Promotion of Fundamental Studies in Health Science of the National Institute of Biomedical Innovation.
PY - 2007/9/21
Y1 - 2007/9/21
N2 - Endothelial nitric oxide synthase (eNOS) produces nitric oxide (NO), which is involved in various physiological functions of the cardiovascular system. eNOS is activated by dephosphorylation at Thr495 and phosphorylation at Ser1177. Inhibition of Rho-kinase, an effector of the small GTPase RhoA, leads to activation of Akt/PKB, which phosphorylates eNOS at Ser1177 and thereby promotes NO production. However, little is known about the effects of Rho-kinase on phosphorylation of Thr495. We here found that the constitutively active form of Rho-kinase phosphorylated eNOS at Thr495 in vitro. Expression of the constitutively active form of RhoA or Rho-kinase increased this phosphorylation in COS-7 cells. Addition of thrombin to cultured human umbilical vein endothelial cells induced phosphorylation of eNOS at Thr495. Treatment with Y27632, a Rho-kinase inhibitor, suppressed thrombin-induced phosphorylation at Thr495. These results indicate that Rho-kinase can directly phosphorylate eNOS at Thr495 to suppress NO production in endothelium.
AB - Endothelial nitric oxide synthase (eNOS) produces nitric oxide (NO), which is involved in various physiological functions of the cardiovascular system. eNOS is activated by dephosphorylation at Thr495 and phosphorylation at Ser1177. Inhibition of Rho-kinase, an effector of the small GTPase RhoA, leads to activation of Akt/PKB, which phosphorylates eNOS at Ser1177 and thereby promotes NO production. However, little is known about the effects of Rho-kinase on phosphorylation of Thr495. We here found that the constitutively active form of Rho-kinase phosphorylated eNOS at Thr495 in vitro. Expression of the constitutively active form of RhoA or Rho-kinase increased this phosphorylation in COS-7 cells. Addition of thrombin to cultured human umbilical vein endothelial cells induced phosphorylation of eNOS at Thr495. Treatment with Y27632, a Rho-kinase inhibitor, suppressed thrombin-induced phosphorylation at Thr495. These results indicate that Rho-kinase can directly phosphorylate eNOS at Thr495 to suppress NO production in endothelium.
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U2 - 10.1016/j.bbrc.2007.07.030
DO - 10.1016/j.bbrc.2007.07.030
M3 - Article
C2 - 17651694
AN - SCOPUS:34547502405
SN - 0006-291X
VL - 361
SP - 462
EP - 467
JO - Biochemical and Biophysical Research Communications
JF - Biochemical and Biophysical Research Communications
IS - 2
ER -