cGMP-dependent relaxation of smooth muscle is coupled with the change in the phosphorylation of myosin phosphatase.
Nitric oxide/cGMP pathway induces vasodilatation, yet the underlying mechanism is obscure. In the present study, we studied the mechanism of cGMP-induced relaxation of the smooth muscle contractile apparatus using permeabilized rabbit femoral arterial smooth muscle. 8-Br-cGMP-induced relaxation was accompanied with a decrease in myosin light chain (MLC) phosphorylation. MLC phosphatase (MLCP) activity, once decreased by agonist-stimulation, recovered to the resting level on addition of 8-Br-cGMP. Because MLCP activity is regulated by the phosphorylation of a MLCP-specific inhibitor, CPI17 at Thr38 and MBS (myosin binding subunit of MLCP) at Thr696, we examined the effect of 8-Br-cGMP on the phosphorylation of these MLCP modulators. Whereas CPI17 phosphorylation was unchanged after addition of 8-Br-cGMP, MBS phosphorylation at Thr696 was significantly decreased by 8-Br-cGMP. We found that 8-Br-cGMP markedly increased MBS phosphorylation at Ser695 in the fiber pretreated with phenylephrine. MBS phosphorylation of Thr696 phosphorylated MBS at Ser695 partially resumed MLCP activity inhibited by Thr696 phosphorylation. Whereas Ser695 phosphorylation was markedly increased, the extent of diphosphorylated MBS at Ser695 and Thr696 in fibers was unchanged after cGMP-stimulation. We found that MBS phosphatase activity in arteries for both diphosphorylated MBS and monophosphorylated MBS at Thr696 significantly increased by 8-Br-cGMP, whereas MBS kinase activity was unchanged. These results suggest that the phosphorylation at Ser640 induced by cGMP shifted the equilibrium of the Thr641 phosphorylation toward dephosphorylation, thus increasing MLCP activity. This results in the decrease in MLC phosphorylation and smooth muscle relaxation.
University of Massachusetts Medical School, Department of Physiology, 55 Lake Avenue North, Worcester, MA 01655, USA.
Kensei
K
Nakamura
Yasuhiko
Y
Koga
Hiroyasu
H
Sakai
Kazuaki
K
Homma
Mitsuo
M
Ikebe
D000818:Animals; D006152:Cyclic GMP / Q000031:analogs & derivatives* / Q000494:pharmacology / Q000502:physiology; D009126:Muscle Relaxation / Q000502:physiology*; D009130:Muscle, Smooth / Q000187:drug effects / Q000201:enzymology*; D043263:Myosin-Light-Chain Phosphatase / Q000378:metabolism*; D009924:Organ Culture Techniques; D010766:Phosphorylation / Q000187:drug effects; D011817:Rabbits
C016276:8-bromocyclic GMP; D043263:Myosin-Light-Chain Phosphatase; D006152:Cyclic GMP