title: Synchronous phosphorylation of CPI-17 and MYPT1 is essential for inducing Ca(2+)  sensitization in intestinal smooth muscle.
abstract: BACKGROUND
  Myosin phosphatase activity is regulated by mechanisms involving the phosphorylation of CPI-17 and MYPT1, primarily based on studies with tonic-type vascular smooth muscles. This study examined how these mechanisms contribute to the regulation of contraction of a phasic-type intestinal smooth muscle.
  
  
  METHODS
  Phosphorylation levels, tension, and Ca(2+) sensitization was detected in rat ileal smooth muscle. Key Results  In rat ileal smooth muscle, phosphorylation level of CPI-17 at Thr(38) and MYPT1 at Thr(853) , but not MYPT1 at Thr(696) , were increased with carbachol (1μmolL(-1) ) accompanied with muscle contraction. The PKC inhibitor Go6976 (1μmol L(-1) ) inhibited the carbachol-induced phosphorylation of CPI-17, whereas the Rho-associated kinase (ROCK) inhibitor, Y-27632 (10μmol L(-1) ) inhibited the carbachol-induced phosphorylation of both CPI-17 and MYPT1. Application of Go6976 or Y-27632 alone inhibited the carbachol-induced contraction; however, the combined application of these inhibitors did not inhibit the contraction in an additive manner. In β-escin-permeabilized ileal strip, treatment with antiphosphorylated antibodies for CPI-17 at Thr(38) and MYPT1 at Thr(853) and Thr(696) alone almost completely abolished the Ca(2+) sensitization due to carbachol with GTP.
  
  
  CONCLUSIONS
  In conclusion, receptor stimulation increases the Ca(2+) sensitivity of contractile elements through CPI-17 phosphorylation via the PKC/ROCK pathways and MYPT1 phosphorylation via the ROCK pathway, when these mechanisms operate cooperatively and/or synchronously in intestinal smooth muscle.
authors: Department of Veterinary Pharmacology, Graduate School of Agriculture and Life Sciences, the University of Tokyo, Tokyo, Japan.
authors: D
authors: D
authors: Mori
authors: M
authors: M
authors: Hori
authors: T
authors: T
authors: Murata
authors: T
authors: T
authors: Ohama
authors: H
authors: H
authors: Kishi
authors: S
authors: S
authors: Kobayashi
authors: H
authors: H
authors: Ozaki
mesh_terms: D000577:Amides / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002217:Carbachol / Q000494:pharmacology; D002227:Carbazoles / Q000494:pharmacology; D018679:Cholinergic Agonists / Q000494:pharmacology; D006160:Guanosine Triphosphate / Q000378:metabolism; D007422:Intestines / Q000033:anatomy & histology / Q000502:physiology*; D008297:Male; D009119:Muscle Contraction / Q000187:drug effects / Q000502:physiology; D009124:Muscle Proteins / Q000378:metabolism*; D009130:Muscle, Smooth / Q000187:drug effects / Q000502:physiology*; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D011188:Potassium / Q000378:metabolism; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D054645:Protein Phosphatase 1 / Q000378:metabolism*; D011725:Pyridines / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors
chemical_list: D000577:Amides; D002227:Carbazoles; D018679:Cholinergic Agonists; D009124:Muscle Proteins; D010750:Phosphoproteins; C491519:Ppp1r14a protein, rat; D011725:Pyridines; C081021:Go 6976; C108830:Y 27632; D006160:Guanosine Triphosphate; D002217:Carbachol; D054460:rho-Associated Kinases; D011493:Protein Kinase C; C499850:Ppp1r12a protein, rat; D054645:Protein Phosphatase 1; D011188:Potassium; D002118:Calcium