Thromboxane A2-induced contraction of rat caudal arterial smooth muscle involves activation of Ca2+ entry and Ca2+ sensitization:: Rho-associated kinase-mediated phosphorylation of MYPT1 at Thr-855, but not Thr-697
Thromboxane A2-induced contraction of rat caudal arterial smooth muscle involves activation of Ca2+ entry and Ca2+ sensitization:: Rho-associated kinase-mediated phosphorylation of MYPT1 at Thr-855, but not Thr-697
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DOI：
10.1042/bj20050237
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发表时间：
2005-08-01
影响因子：
4.1
通讯作者：
Walsh, MP
中科院分区：
文献类型：
--
作者：
Wilson, DP;Susnjar, M;Walsh, MP
在去内皮化的大鼠尾动脉中研究了TxA（2）（血栓烷A（2））模拟物U-46619激活血管平滑肌收缩的信号转导途径。U-46619引起的收缩可被TP受体（TxA（2）受体）拮抗剂SQ-29548、ROK（Rho相关激酶）抑制剂Y-27632和H-1152，MLCK（肌球蛋白轻链激酶）抑制剂ML-7、ML-9和渥曼青霉素，电压门控Ca 2+通道阻断剂尼卡地平，以及细胞外Ca 2+的清除;蛋白激酶C抑制剂GF 109203 x无作用。U-46619在α-毒素透化组织中引起Ca 2+敏化。U-46619诱导小GTdR RhoA的活化，与ROK的参与一致。研究了ROK的两个下游靶点：CPI-17 [蛋白激酶C增强的PP 1抑制蛋白（蛋白磷酸酶1型）17 kDa]，一种肌球蛋白轻链磷酸酶抑制剂，在功能位点（Thr-38）没有磷酸化; MYPT 1（肌球蛋白轻链磷酸酶的肌球蛋白靶向亚基）的磷酸化在Thr-855显著增加，但在Thr-697没有。U-46619诱发的收缩与肌球蛋白20 kDa轻链的磷酸化相关。我们的结论是：（i）U-46619通过激活Ca 2 +/钙调蛋白/MLCK途径和RhoA/ROK途径诱导收缩;（ii）MYPT 1的Thr-855在静息和响应于U-46619刺激时被ROK磷酸化;（iii）MYPT 1的Thr-697在静息条件下被除ROK之外的激酶磷酸化，并且响应于U-46619处理不增加;和（iv）ROK和蛋白激酶C均不磷酸化该血管平滑肌中的CPI-17以响应U-46619。
The signal transduction pathway whereby the TxA(2) (thromboxane A(2)) mimetic U-46619 activates vascular smooth muscle contraction was investigated in de-endothelialized rat caudal artery. U-46619-evoked contraction was inhibited by the TP receptor (TxA(2) receptor) antagonist SQ-29548, the ROK (Rho-associated kinase) inhibitors Y-27632 and H-1152, the MLCK (myosin light-chain kinase) inhibitors ML-7, ML-9 and wortmannin, the voltage-gated Ca2+-channel blocker nicardipine, and removal of extracellular Ca2+; the protein kinase C inhibitor GF109203x had no effect. U-46619 elicited Ca2+ sensitization in alpha-toxin-permeabilized tissue. U-46619 induced activation of the small GTPase RhoA, consistent with the involvement of ROK. Two downstream targets of ROK were investigated: CPI-17 [protein kinase C-potentiated inhibitory protein for PP1 (protein phosphatase type 1) of 17 kDa], a myosin light-chain phosphatase inhibitor, was not phosphorylated at the functional site (Thr-38); phosphorylation of MYPT1 (myosin-targeting subunit of myosin light-chain phosphatase) was significantly increased at Thr-855, but not Thr-697. U-46619-evoked contraction correlated with phosphorylation of the 20 kDa light chains of myosin. We conclude that: (i) U-46619 induces contraction via activation of the Ca2+/calmodulin/MLCK pathway and of the RhoA/ROK pathway; (ii) Thr-855 of MYPT1 is phosphorylated by ROK at rest and in response to U-46619 stimulation; (iii) Thr-697 of MYPT1 is phosphorylated by a kinase other than ROK under resting conditions, and is not increased in response to U-46619 treatment; and (iv) neither ROK nor protein kinase C phosphorylates CPI-17 in this vascular smooth muscle in response to U-46619.