Possible involvement of the novel CPI-17 protein in protein kinase C signal transduction of rabbit arterial smooth muscle.
1. CPI-17 has recently been identified as a novel protein in vascular smooth muscle. In vitro , its phosphorylation and thiophosphorylation by protein kinase C (PKC) specifically inhibits the type 1 class of protein phosphatases, including myosin light chain (MLC) phosphatase. 2. Both of the phosphorylated CPI-17 states dose-dependently potentiated submaximal contractions at constant [Ca2+] in beta-escin-permeabilized and Triton X-100-demembranated arterial smooth muscle, but produced no effect in intact and less intensely permeabilized (alpha-toxin) tissue. Thiophosphorylated CPI-17 (tp-CPI) induced large contractions even under Ca2+-free conditions and decreased Ca2+ EC50 by more than an order of magnitude. Unphosphorylated CPI-17 produced minimal but significant effects. 3. tp-CPI substantially increased the steady-state MLC phosphorylation to Ca2+ ratios in beta-escin preparations. 4. tp-CPI affected the kinetics of contraction and relaxation and of MLC phosphorylation and dephosphorylation in such a manner that indicates its major physiological effect is to inhibit MLC phosphatase. 5. Results from use of specific inhibitors in concurrence with tp-CPI repudiate the involvement of general G proteins, rho A or PKC itself in the Ca2+ sensitization by tp-CPI. 6. Our results indicate that phosphorylation of CPI-17 by PKC stimulates binding of CPI-17 to and subsequent inhibition of MLC phosphatase. This implies that CPI-17 accounts largely for the heretofore unknown signalling pathway between PKC and inhibited MLC phosphatase.
Li
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Department of Physiology and Biophysics, Georgetown University School of Medicine, Washington, DC 20007, USA.
Eto
M
M
Lee
M R
MR
Morita
F
F
Yazawa
M
M
Kitazawa
T
T
D000818:Animals; D001011:Aorta / Q000166:cytology; D002118:Calcium / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000378:metabolism*; D005263:Femoral Artery / Q000166:cytology; D006153:Guanosine Diphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D008297:Male; D009119:Muscle Contraction / Q000187:drug effects / Q000502:physiology; D009131:Muscle, Smooth, Vascular / Q000737:chemistry / Q000201:enzymology*; D043263:Myosin-Light-Chain Phosphatase; D010446:Peptide Fragments / Q000494:pharmacology; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism* / Q000494:pharmacology; D054648:Protein Phosphatase 2; D011506:Proteins / Q000378:metabolism*; D011817:Rabbits; D011994:Recombinant Proteins / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D013457:Sulfur Compounds / Q000378:metabolism; D013552:Swine; D013873:Thionucleotides / Q000494:pharmacology; D014661:Vasoconstriction / Q000187:drug effects / Q000502:physiology
D004791:Enzyme Inhibitors; D010446:Peptide Fragments; D011506:Proteins; D011994:Recombinant Proteins; D013457:Sulfur Compounds; D013873:Thionucleotides; C073882:phosphoprotein phosphatase inhibitor 1; C073885:protein kinase C (19-31); D006153:Guanosine Diphosphate; C023427:guanosine 5'-O-(2-thiodiphosphate); D011493:Protein Kinase C; D010749:Phosphoprotein Phosphatases; D054648:Protein Phosphatase 2; D043263:Myosin-Light-Chain Phosphatase; D002118:Calcium