α1-Adrenoceptor-Mediated Phosphorylation of MYPT1 and CPI-17 in the Uterine Artery: Role of ERK/PKC By

We have previously demonstrated that ERK/PKC signaling pathways play a key role in the regulation of Ca 2+ sensitivity and contractility of the uterine artery. The present study tested the hypothesis that ERK and PKC differentially regulated myosin light chain phosphatase activity by phosphorylation of MYPT1 and CPI-17. Agonist-induced contractions and phosphorylation of MYPT1/Thr 696 , MYPT1/Thr 850 , and CPI-17/Thr 38 were measured simultaneously in the same tissues of isolated near-term pregnant ovine uterine arteries. Phenylephrine produced time-dependent concurrent increases in the phosphorylation of ERK 44/42 and MYPT1/Thr 850 , which preceded contractions. In addition, phenylephrine induced phosphorylation of CPI-17/Thr 38 that was concurrent with the contractions. In contrast, phenylephrine did not induce phosphorylation of MYPT1/Thr 696 in the uterine artery. PD098059 inhibited phosphorylation of ERK 44/42 and the initial peak phosphorylation of MYPT1/Thr 850 , but did not affect CPI-17/Thr 38 phosphorylation. Activation of PKC by PDBu induced a time-dependent phosphorylation of CPI-17/Thr 38 that preceded the contractions of the uterine artery. In addition, PDBu activated PKCα, and induced a co-immunoprecipitation of PKCα with caldesmon. The results suggest that phosphorylation of MYPT1/Thr 850 and CPI-17/Thr 38 play important roles in the regulation of agonist-mediated Ca 2+ sensitivity in the uterine artery, which are regulated in part by ERK and PKC, respectively. In addition, phosphorylated CPI-17 may regulate the Ca 2+ sensitivity by interacting with caldesmon and reversing its inhibitory effect on myosin ATPase.