Effect of vanadate on force and myosin light chain phosphorylation in skinned aortic smooth muscle.
The effects of vanadate were examined on Ca2+-activated force and phosphorylation of 20-kDa myosin light chain in membrane-permeabilized rabbit aortic smooth muscle strips. Addition of vanadate during maximum contraction reduced the force in a dose-dependent manner, and inhibited it almost completely at 1 mM. Two-dimensional polyacrylamide gel electrophoretic analyses revealed that vanadate also reduced the phosphorylation of 20- kDa myosin light chain in a dose-dependent manner from approximately 50% in the absence of vanadate to approximately 20% in the presence of 1 mM vanadate. The effects of 1 mM vanadate on purified myosin light chain kinase and phosphatase were then examined using purified myosin as substrate, and it was found that vanadate neither inhibited myosin light chain kinase nor activated myosin light chain phosphatase. These results indicate that the reduction in the 20-kDa myosin light chain phosphorylation level by vanadate may be effected through its inhibition of the force generation in skinned smooth muscle strip, as evidenced by the finding that vanadate eliminated the enhancement of myosin light chain kinase activity brought about by the interaction between purified myosin and actin.
Morimoto
S
S
Department of Pharmacology, Juntendo University School of Medicine, Hongo, Bunkyo-ku, Tokyo, 113-8421, Japan. morimoto@clipharm.med.kyushu-u.ac.jp
Sato
O
O
Ogawa
Y
Y
D000199:Actins / Q000302:isolation & purification / Q000378:metabolism; D000818:Animals; D001011:Aorta / Q000187:drug effects / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D011232:Chemical Precipitation; D004305:Dose-Response Relationship, Drug; D066298:In Vitro Techniques; D009119:Muscle Contraction / Q000187:drug effects; D009129:Muscle Tonus / Q000187:drug effects; D009131:Muscle, Smooth, Vascular / Q000187:drug effects* / Q000502:physiology*; D018994:Myosin Light Chains / Q000187:drug effects / Q000378:metabolism*; D009219:Myosin-Light-Chain Kinase / Q000187:drug effects / Q000302:isolation & purification / Q000378:metabolism; D043263:Myosin-Light-Chain Phosphatase; D009994:Osmolar Concentration; D010749:Phosphoprotein Phosphatases / Q000187:drug effects / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011817:Rabbits; D014638:Vanadates / Q000494:pharmacology*
D000199:Actins; D018994:Myosin Light Chains; D014638:Vanadates; D009219:Myosin-Light-Chain Kinase; D010749:Phosphoprotein Phosphatases; D043263:Myosin-Light-Chain Phosphatase; D002118:Calcium