Regulation Vascular Smooth Muscle Myosin Phosphatase

调节血管平滑肌肌球蛋白磷酸酶

• 批准号: 7021245
• 负责人: MASUMI ETO
• 金额: $ 27.13万
• 依托单位: THOMAS JEFFERSON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-07-15 至 2011-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7021245
• 关键词: G protein; RNA interference; biological signal transduction; computer program /software; computer simulation; enzyme activity; fluorescence microscopy; intravital microscopy; kinase inhibitor; laboratory rat; molecular /cellular imaging; molecular site; muscle contraction; myosins; nuclear magnetic resonance spectroscopy; phosphatase inhibitor; phosphoprotein phosphatase; phosphorylation; protein kinase C; serine threonine protein kinase; tissue /cell culture; vascular smooth muscle

DESCRIPTION (provided by applicant): The long-term goal of this project is to understand regulatory mechanisms for vascular smooth muscle contraction. Vascular smooth muscle is a target for treatment of hypertension to reduce blood pressure. Agonist stimuli induce activation of G-proteins that results in phosphorylation of myosin and contraction of smooth muscle cells. This process is governed by the inhibition of myosin phosphatase, and causes the Ca2+-independent contraction. Thus, the activity of myosin phosphatase is a determinant of the Ca2+ sensitivity of the contraction. RhoA-dependent kinase (ROCK) is believed to phosphorylate a regulatory subunit of myosin phosphatase (MYPT1) to cause inactivation of the phosphatase. However, recent studies imply that phosphorylation of MYPT1 is unchanged during the contraction, so that a critical question still remains: How is myosin phosphatase inhibited in response to agonist stimulation of smooth muscle? A novel inhibitor protein for myosin phosphatase, called CPI-17, is a candidate and highly expressed in vascular smooth muscles. The kinases activated with agonist stimuli, such as protein kinase C (PKC) and ROCK, phosphorylate CPI-17 at Thr38, which converts it into a potent inhibitor. Phosphorylation and dephosphorylation of CPI-17 reversibly occur in parallel to contraction and relaxation of the muscle. Thus, CPI-17 is a key molecule to understand the regulation of myosin phosphatase in smooth muscle. Another myosin phosphatase inhibitor in the CPI-17 family, named PHI-1, is also expressed in smooth muscle. We will ask how two inhibitors control myosin phosphatase in smooth muscle. Aim 1 will test the hypothesis that CPI-17 and PHI-1 are phosphorylated by separate subsets of kinase signals in response to G-protein activation. Aim 2 is to define the role of phosphorylation of CPI-17 at Ser12 and Ser128, which are recently detected in cells. Aim 3 is to elucidate molecular mechanisms for specific inhibition of myosin phosphatase by CPI-17 by mutational/computational analyses. This project will uncover molecular mechanisms how multiple signals from G-protein activation converge onto myosin phosphatase inhibitors to control smooth muscle contraction, which is a major physiological importance in vascular biology. (Relevance) Drugs, blocking the contraction, are used for treatment of hypertension, asthma and erectile dysfunction. The results of this project will expose potential therapeutic targets for smooth muscle diseases.

描述（由申请人提供）：本项目的长期目标是了解血管平滑肌收缩的调节机制。血管平滑肌是治疗高血压以降低血压的靶点。激动剂刺激诱导G蛋白活化，导致肌球蛋白磷酸化和平滑肌细胞收缩。这一过程是由肌球蛋白磷酸酶的抑制，并导致Ca 2+非依赖性收缩。因此，肌球蛋白磷酸酶的活性是收缩的Ca 2+敏感性的决定因素。RhoA依赖性激酶（ROCK）被认为磷酸化肌球蛋白磷酸酶（MYPT 1）的调节亚基以引起磷酸酶的失活。然而，最近的研究表明，MYPT 1的磷酸化在收缩过程中是不变的，所以一个关键的问题仍然存在：肌球蛋白磷酸酶是如何抑制平滑肌激动剂刺激的反应？一种新的肌球蛋白磷酸酶抑制剂蛋白，称为CPI-17，是一种候选蛋白，在血管平滑肌中高度表达。激动剂刺激激活的激酶，如蛋白激酶C（PKC）和ROCK，在Thr 38磷酸化CPI-17，将其转化为有效的抑制剂。CPI-17的磷酸化和去磷酸化可逆地平行于肌肉的收缩和松弛发生。因此，CPI-17是了解平滑肌中肌球蛋白磷酸酶调节的关键分子。CPI-17家族中的另一种肌球蛋白磷酸酶抑制剂，命名为PHI-1，也在平滑肌中表达。我们将探讨两种抑制剂如何控制平滑肌中的肌球蛋白磷酸酶。目的1将测试CPI-17和PHI-1被响应于G蛋白活化的激酶信号的单独子集磷酸化的假设。目的2是确定最近在细胞中检测到的CPI-17在Ser 12和Ser 128处磷酸化的作用。目的3是通过突变/计算分析阐明CPI-17特异性抑制肌球蛋白磷酸酶的分子机制。该项目将揭示来自G蛋白激活的多种信号如何汇聚到肌球蛋白磷酸酶抑制剂上以控制平滑肌收缩的分子机制，这在血管生物学中具有重要的生理意义。（相关性）阻止收缩的药物用于治疗高血压、哮喘和勃起功能障碍。该项目的结果将揭示平滑肌疾病的潜在治疗靶点。