Rho‐kinase mediates diphosphorylation of myosin regulatory light chain in cultured uterine, but not vascular smooth muscle cells

Phosphorylation of myosin regulatory light chain ( RLC ) triggers contraction in smooth muscle myocytes. Dephosphorylation of phosphorylated RLC (p RLC ) is mediated by myosin RLC phosphatase ( MLCP ), which is negatively regulated by rho‐associated kinase ( ROK ). We have compared basal and stimulated concentrations of p RLC in myocytes from human coronary artery (h VM ), which has a tonic contractile pattern to myocytes from human uterus (h UM ), which has a phasic contractile pattern. Our studies reveal fundamental differences between h VM and h UM regarding the mechanisms regulating phosphorylation RLC . Whereas h VM responded to stimulation by phosphorylation of RLC at S19, h UM responded by forming diphosphorylated RLC (at T18 and S19; pp RLC ), which, compared to p RLC , causes two to threefold greater activation of myosin ATP ase that provides energy to power the contraction. Importantly, the conversion of p RLC to pp RLC is mediated by ROK . In h UM , MLCP has high activity for pp RLC and this is inhibited by ROK through phosphorylation of the substrate targeting subunit ( MYPT 1) at T853. Inhibitors of ROK significantly reduce contractility in both h VM and h UM . We demonstrated that inhibition of pp RLC in phasic myocytes (h UM ) is 100‐fold more sensitive to ROK inhibitors than is p RLC in tonic myocytes (h VM ). We speculate that these differences in phosphorylation of RLC might reflect evolution of different contractile patterns to perform distinct physiological functions. Furthermore, our data suggest that low concentrations of ROK inhibitors might inhibit uterine contractions with minimal effects on vascular tone, thus posing a novel strategy for prevention or treatment of conditions such as preterm birth.