Phosphorylation of CPI-17, an inhibitor of myosin phosphatase, by protein kinase N

Tetsuya Hamaguchi, Masaaki Ito, Jianhua Feng, Tetsuya Seko, Mutsumi Koyama, Hirofumi Machida, Koujiro Takase, Mutsuki Amano, Kozo Kaibuchi, David J. Hartshorne, Takeshi Nakano

Research output: Contribution to journalArticlepeer-review

78 Citations (Scopus)


CPI-17 Is a phosphorylation-dependent inhibitory protein for smooth muscle myosin phosphate. Phosphorylation at Thr38, in vitro, by protein kinase C or Rho-kinase enhances the inhibitory potency toward myosin phosphatase. Phosphorylation of CPI-17 by protein kinase N (PKN), a fatty acid- and Rho- activated serine/threonine kinase, and its effect on smooth muscle myosin phosphatase activity were investigated. CPI-17 was phosphorylated by GST-PKN-CAT, a constitutively active GST-fusion fragment of PKN, to 1.46 mol of P/mol of CPI-17, in vitro. The K(m) value of CPI-17 for PKN was 0.96 μM. Phosphorylation of PKN dramatically increased the inhibitory effect of CPI-17 on myosin phosphatase activity. The major and inhibitory phosphorylation site was identified as Thr38 using a point mutant of CPI-17 and a phosphorylation - state specific antibody. Thus, CPI-17 is a substrate of PKN and might be involved in the Ca2+ sensitization of smooth muscle contraction as a downstream effector of Rho and/or arachidonic acid. (C) 2000 Academic Press.

Original languageEnglish
Pages (from-to)825-830
Number of pages6
JournalBiochemical and Biophysical Research Communications
Issue number3
Publication statusPublished - 11-08-2000
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology


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