Ca2+-desensitizing hypoxic vasorelaxation: pivotal role for the myosin binding subunit of myosin phosphatase (MYPT1) in porcine coronary artery.
Acute hypoxia dilates most systemic arteries leading to increased tissue perfusion. We showed that at high stimulus conditions, porcine coronary artery was relaxed by hypoxia without a change in [Ca(2+)](i). This 'Ca(2+)-desensitizing hypoxic relaxation' was validated in permeabilized porcine coronary artery smooth muscle (PCASM) in which hypoxia decreased force and myosin regulatory light chain phosphorylation (p-MRLC) despite fixed [Ca(2+)]. Rho kinase-dependent phosphorylation of MYPT1 (p-MYPT1) is associated with decreased MRLC phosphatase (MLCP) activity, and increased Ca(2+) sensitivity of both p-MRLC and force. We tested the hypothesis that hypoxia induces Ca(2+)-desensitizing hypoxic relaxation via dephosphorylation of p-MYPT1, consequently increasing MLCP activity and thus decreasing p-MRLC. alpha-Toxin-permeabilized PCASM pretreated with ATPgammaS did not relax in response to hypoxia. Moreover, when MRLC but not MYPT1 was protected from ATPgammaS thiophosphorylation by the MRLC kinase inhibitor ML7 (300 mum), hypoxia remained ineffective. In contrast, hypoxic relaxation was preserved with further addition of the Rho kinase inhibitor Y27632 (1 mum), to attenuate thiophosphorylation of MYPT1. Importantly, measurements of p-MRLC, and p-MYPT1 at T696 and T853 (human sequence) paralleled that of force. We conclude that Ca(2+)-desensitizing hypoxic relaxation requires dephosphorylation of p-MYPT1. Moreover, no kinases, other then those inhibited by ML7 and Y27632, nor their associated phosphoproteins can be involved in Ca(2+)-desensitizing hypoxic relaxation.
Wardle
Robert L
RL
Department of Molecular and Cellular Physiology, University of Cincinnati College of Medicine, Cincinnati, OH, USA. wardlerl@uc.edu
Gu
Min
M
Ishida
Yukisato
Y
Paul
Richard J
RJ
D000818:Animals; D002118:Calcium / Q000494:pharmacology*; D015687:Cell Hypoxia / Q000187:drug effects / Q000502:physiology; D003331:Coronary Vessels / Q000187:drug effects / Q000502:physiology*; D066298:In Vitro Techniques; D009119:Muscle Contraction / Q000187:drug effects / Q000502:physiology*; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000502:physiology*; D043263:Myosin-Light-Chain Phosphatase / Q000378:metabolism*; D009218:Myosins / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D011485:Protein Binding; D013552:Swine; D014664:Vasodilation / Q000187:drug effects / Q000502:physiology*
D043263:Myosin-Light-Chain Phosphatase; D009218:Myosins; D002118:Calcium