Phosphorylation of caldesmon by p21-activated kinase - Implications for the Ca2+ sensitivity of smooth muscle contraction

We have previously shown that pal-activated kinase, PAK, induces Ca2+-indep endent contraction of Triton-skinned smooth muscle with concomitant increas e in phosphorylation of caldesmon and desmin but not myosin-regulatory ligh t chain (Van Eyk, J, E,, Arrell, D, K,, Foster, D, B., Strauss, J, D,, Hein onen, T, Y,, Furmaniak-Kazmierczak, E,, Cote, G. P,, and Mak, A. S, (1998) J, Biol, Chem. 273, 23433-23439), In this study, we provide biochemical evi dence implicating a role for PAK in Ca2+-independent contraction of smooth muscle via phosphorylation of caldesmon, Mass spectroscopy data show that s toichiometric phosphorylation occurs at Ser(657) and Ser(687) abutting the calmodulin-binding sites A and B of chicken gizzard caldesmon, respectively . Phosphorylation of Ser(657) and Ser(687) has an important functional impa ct on caldesmon, PAK-phosphorylation reduces binding of caldesmon to calmod ulin by about 10-fold whereas binding of calmodulin to caldesmon partially inhibits PAK phosphorylation, Phosphorylated caldesmon displays a modest re duction in affinity for actin-tropomyosin but is significantly less effecti ve in inhibiting actin-activated S1 ATPase activity in the presence of trop omyosin, We conclude that PAK-phosphorylation of caldesmon at the calmoduli n-binding sites modulates caldesmon inhibition of actin-myosin ATPase activ ity and may, in concert with the actions of Rho-kinase, contribute to the r egulation of Ca2+ sensitivity of smooth muscle contraction.