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PMID: 12563012 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Phosphorylation of the myosin phosphatase targeting subunit and CPI-17 during Ca2+ sensitization in rabbit smooth muscle.

The Journal of physiology ·Vol. 546 ·No. Pt 3 ·2003-02-01 ·Pages 879-89

Kitazawa T, Eto M, Woodsome TP, Khalequzzaman M

Abstract

Myosin phosphatase (MLCP) plays a critical regulatory role in the Ca(2+) sensitivity of myosin phosphorylation and smooth muscle contraction. It has been suggested that phosphorylation at Thr(695) of the MLCP regulatory subunit (MYPT1) and at Thr(38) of the MLCP inhibitor protein CPI-17 results in inhibition of MLCP activity. We have previously demonstrated that CPI-17 Thr(38) phosphorylation plays an important role in G-protein-mediated inhibition of MLCP in tonic arterial smooth muscle. Here, we attempted to evaluate the function of MYPT1 in phasic rabbit portal vein (PV) and vas deferens (VD) smooth muscles. Using site- and phospho-specific antibodies, phosphorylation of MYPT1 Thr(695) and CPI-17 Thr(38) was examined along with MYPT1 Thr(850), which is a non-inhibitory Rho-kinase site. We found that both CPI-17 Thr(38) and MYPT1 Thr(850) were phosphorylated in response to agonists or GTPgammaS concurrently with contraction and myosin phosphorylation in alpha-toxin-permeabilized PV tissues. In contrast, phosphorylation of MYPT1 Thr(695) did not increase. Comparable results were also obtained in both permeabilized and intact VD. The Rho-kinase inhibitor Y-27632 and the protein kinase C (PKC) inhibitor GF109203X suppressed phosphorylation of MYPT1 Thr(850) and CPI-17 Thr(38), respectively, in intact VD while MYPT1 Thr(695) phosphorylation was insensitive to both inhibitors. These results indicate that phosphorylation of MYPT1 Thr(695) is independent of stimulation of G-proteins, Rho-kinase or PKC. In the phasic PV, phosphorylation of CPI-17 Thr(38) may contribute towards inhibition of MLCP while the phasic visceral VD, which has a low CPI-17 concentration, probably utilizes other Ca(2+) sensitizing mechanisms for inhibiting MLCP besides phosphorylation of MYPT1 and CPI-17.

MeSH Terms
Amides/pharmacology Animals Calcium/metabolism Electrophysiology Endothelin-1/pharmacology Enzyme Inhibitors/pharmacology Indoles/pharmacology Isoenzymes/metabolism Male Maleimides/pharmacology Muscle Proteins/metabolism Muscle, Smooth/metabolism Muscle, Smooth, Vascular/physiology Myosin Light Chains/metabolism Myosin-Light-Chain Phosphatase Permeability Phenylephrine/pharmacology Phosphoprotein Phosphatases/metabolism Phosphoproteins/metabolism Phosphorylation Portal Vein/physiology Pyridines/pharmacology Rabbits Type C Phospholipases/pharmacology Vas Deferens/physiology Vasoconstriction
Chemicals
Amides CPI-17 protein, Oryctolagus cuniculus Endothelin-1 Enzyme Inhibitors Indoles Isoenzymes Maleimides Muscle Proteins Myosin Light Chains Phosphoproteins Pyridines Y 27632 Phenylephrine Phosphoprotein Phosphatases Myosin-Light-Chain Phosphatase Type C Phospholipases bisindolylmaleimide I Calcium
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Kitazawa Toshio
Department of Physiology and Biophysics, Georgetown University Medical Center, Washington, DC 20007, USA. [email protected]
Eto Masumi
Woodsome Terence P
Khalequzzaman Md
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44 references, click to expand
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Article Info
Journal
The Journal of physiology
Abbr.
J Physiol
ISSN
0022-3751
DOI
10.1113/jphysiol.2002.029306
Email
[email protected]
Published
2003-02-01
Pages
879-89
Language
English
Region
England
NLM ID
0266262
PMCID
PMC2342583
Subset
IM
Grants
NHLBI NIH HHS · R01 HL070881 · United States
NHLBI NIH HHS · HL70881 · United States
NHLBI NIH HHS · R01HL51824 · United States
External Links
PubMed source DOI: 10.1113/jphysiol.2002.029306 PMC Full Text View journal details
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