title: Ca2+-independent, inhibitory effects of cyclic adenosine 5'-monophosphate on Ca2+ regulation of phosphoinositide 3-kinase C2alpha, Rho, and myosin phosphatase in vascular smooth muscle.
abstract: We have recently demonstrated in vascular smooth muscle (VSM) that membrane depolarization by high KCl induces Ca(2+)-dependent Rho activation and myosin phosphatase (MLCP) inhibition (Ca(2+)-induced Ca(2+)-sensitization) through the mechanisms involving phosphorylation of myosin-targeting protein 1 (MYPT1) and 17-kDa protein kinase C (PKC)-potentiated inhibitory protein of PP1 (CPI-17). In the present study, we investigated whether and how cAMP affected Ca(2+)-dependent MLCP inhibition by examining the effects of forskolin, cell-permeable dibutyryl cAMP (dbcAMP), and isoproterenol. Forskolin, but not its inactive analog 1,9-dideoxyforskolin, inhibited KCl-induced contraction and the 20-kDa myosin light chain (MLC) phosphorylation without inhibiting Ca(2+) mobilization in rabbit aortic VSM. dbcAMP mimicked these forskolin effects. We recently suggested that Ca(2+)-mediated Rho activation is dependent on class II alpha-isoform of phosphoinositide 3-kinase (PI3K-C2alpha). Forskolin inhibited KCl-induced stimulation of PI3K-C2alpha activity. KCl-induced membrane depolarization stimulated Rho in a manner dependent on a PI3K but not PKC and stimulated phosphorylation of MYPT1 at Thr(850) and CPI-17 at Thr(38) in manners dependent on both PI3K and Rho kinase, but not PKC. Forskolin, dbcAMP, and isoproterenol inhibited KCl-induced Rho activation and phosphorylation of MYPT1 and CPI-17. Consistent with these data, forskolin, isoproterenol, a PI3K inhibitor, or a Rho kinase inhibitor, but not a PKC inhibitor, abolished KCl-induced diphosphorylation of MLC. These observations indicate that cAMP inhibits Ca(2+)-mediated activation of the MLCP-regulating signaling pathway comprising PI3K-C2alpha, Rho, and Rho kinase in a manner independent of Ca(2+) and point to the novel mechanism of the cAMP actions in the regulation of vascular smooth muscle contraction.
authors.affiliation: Department of Physiology, Kanazawa University Graduate School of Medicine, 13-1 Takara-machi, Kanazawa, Ishikawa 920-8640, Japan.
authors.forename: Mohammed Ali
authors.initials: MA
authors.lastname: Azam
authors.forename: Kazuaki
authors.initials: K
authors.lastname: Yoshioka
authors.forename: Shinsuke
authors.initials: S
authors.lastname: Ohkura
authors.forename: Noriko
authors.initials: N
authors.lastname: Takuwa
authors.forename: Naotoshi
authors.initials: N
authors.lastname: Sugimoto
authors.forename: Koichi
authors.initials: K
authors.lastname: Sato
authors.forename: Yoh
authors.initials: Y
authors.lastname: Takuwa
mesh_terms: D000818:Animals; D003994:Bucladesine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D058541:Class II Phosphatidylinositol 3-Kinases; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000502:physiology*; D007545:Isoproterenol / Q000494:pharmacology; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects; D009124:Muscle Proteins / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000201:enzymology*; D043263:Myosin-Light-Chain Phosphatase / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology*; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011817:Rabbits; D014661:Vasoconstriction
chemical_list: C491521:CPI-17 protein, Oryctolagus cuniculus; D009124:Muscle Proteins; D010750:Phosphoproteins; D005576:Colforsin; D003994:Bucladesine; D000242:Cyclic AMP; D019869:Phosphatidylinositol 3-Kinases; D058541:Class II Phosphatidylinositol 3-Kinases; D043263:Myosin-Light-Chain Phosphatase; D007545:Isoproterenol; D002118:Calcium