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This Article Full Text Full Text (PDF) All Versions of this Article: 289/6/L925    most recent 00054.2005v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in Web of Science Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via Web of Science (4) Citing Articles via Google Scholar Google Scholar Articles by Sakai, H. Articles by Misawa, M. Search for Related Content PubMed PubMed Citation Articles by Sakai, H. Articles by Misawa, M.

Acetylcholine-induced phosphorylation and membrane translocation of CPI-17 in bronchial smooth muscle of rats

Department of Pharmacology, School of Pharmacy, Hoshi University, Shinagawa-ku, Tokyo, Japan

Submitted 28 January 2005 ; accepted in final form 16 July 2005

A translocation of protein kinase C (PKC) from cytosol to plasma membrane has been reported as an association with agonist-induced Ca²⁺ sensitization in smooth muscle contraction. Therefore, it is possible that a downstream target of PKC, CPI-17 [PKC-potentiated inhibitory protein for heterotrimeric myosin light chain (MLC) phosphatase of 17 kDa], might also be translocated to membrane when activated. To confirm this hypothesis, cytosolic and membrane CPI-17 was measured in acetylcholine (ACh)- and high-K⁺ depolarization-stimulated bronchial smooth muscle of rats. An active form of CPI-17, i.e., Thr38-phosphorylated CPI-17, was also measured in cytosolic and membrane fractions. Immunoblot analyses demonstrated a translocation of CPI-17 from cytosolic to membrane fraction by ACh, but not high-K⁺ depolarization, stimulation in time- and concentration-dependent manners. Interestingly, phosphorylated CPI-17 was detected only in membrane fractions in the ACh-stimulated tissues. However, in the high-K⁺ depolarization-stimulated tissues, phosphorylated CPI-17 was not detected both in membrane and cytosolic fraction. To estimate downstream of activated CPI-17, immunoblotting for phosphorylated MLC was performed in ACh- or high-K⁺ depolarization-stimulated tissues. ACh- and high-K⁺ depolarization-induced phosphorylation of MLC was observed in its contraction-dependent manner. In conclusion, we, for the first time, suggested that CPI-17 is translocated and phosphorylated by ACh, but not high-K⁺ depolarization, in rat bronchial smooth muscle. ACh-induced translocation and phosphorylation of CPI-17 might be caused via the activation of muscarinic receptor.

high-potassium depolarization-stimulated tissues