Regulation of Smooth Muscle Cell Differentiation by RhoA

RhoA 对平滑肌细胞分化的调节

• 批准号: 6747567
• 负责人: Christopher P. Mack
• 金额: $ 28.97万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-06-01 至 2007-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6747567
• 关键词: actin binding protein; angiotensin II; biological signal transduction; cell differentiation; cell growth regulation; fungal proteins; gel mobility shift assay; gene expression; genetic promoter element; genetically modified animals; guanine nucleotide binding protein; immunoprecipitation; integrins; laboratory mouse; muscle cells; phenotype; phosphorylation; protein protein interaction; reporter genes; thrombin; tissue /cell culture; transcription factor; vascular smooth muscle; yeast two hybrid system

DESCRIPTION (provided by applicant): Vascular smooth muscle cell (SMC) differentiation is a very important process during vasculogenesis and angiogenesis, and it is recognized that alterations in SMC phenotype play a role in the progression of several prominent cardiovascular diseases including atherosclerosis, hypertension, and restenosis. The overall goal of the proposed studies is to identify the mechanisms by which environmental signals regulate SMC differentiation marker gene expression. Serum response factor (SRF) is the only transcription factor that has been shown to regulate all of the SMC differentiation marker genes. Since RhoA regulates SRF, and because RhoA activity is affected by many environmental cues (matrix, contractile agonists, growth factors, mechanical stretch) we hypothesize that Rho signaling is important for regulating SMC-specific transcription, and ultimately, SMC phenotype. Our specific aims are as follows: Aim 1 - to identify the Rho-dependent signaling pathways that contribute to the regulation of SMC-specific gene expression. We will measure Rho activation in SMC, and we will use dominant negative approaches to determine the extent to which Rho and Rho effectors contribute to the regulation of SMC-specific promoter/reporter constructs. Aim 2 - to identify the mechanisms by which Rho affects SRF-dependent transcription in SMC. We will study the effects of Rho on the regulation of SRF phosphorylation and DNA binding, and on SRF's interaction with other transcription factors. Aim 3 - to determine the effects of RhoA signaling on SMC differentiation in vivo. Because SMC phenotype is regulated by many diverse environmental cues that cannot be accurately reproduced in culture models, it is important to study SMC differentiation in vivo. We will express dominant negative and constitutively active forms of Rho specifically in SMC in mice using the SM myosin heavy chain promoter. Measuring SMC differentiation marker gene expression, examining vessel and organ morphology, and determining proliferation indices will assess effects on SMC differentiation. Completion of these aims should lead to a better understanding of the signaling and transcription mechanisms that regulate SMC differentiation and perhaps to novel targets for cardiovascular therapeutic intervention.

描述（申请人提供）：血管平滑肌细胞（SMC）分化是血管发生和血管生成过程中非常重要的过程，并且人们认识到SMC表型的改变在包括动脉粥样硬化、高血压和再狭窄在内的几种重要心血管疾病的进展中发挥作用。拟议研究的总体目标是确定环境信号调节 SMC 分化标记基因表达的机制。血清反应因子 (SRF) 是唯一已被证明可以调节所有 SMC 分化标记基因的转录因子。由于 RhoA 调节 SRF，并且 RhoA 活性受到许多环境因素（基质、收缩激动剂、生长因子、机械拉伸）的影响，我们假设 Rho 信号对于调节 SMC 特异性转录以及最终调节 SMC 表型非常重要。我们的具体目标如下： 目标 1 - 确定有助于调节 SMC 特异性基因表达的 Rho 依赖性信号通路。我们将测量 SMC 中的 Rho 激活，并使用显性失活方法来确定 Rho 和 Rho 效应子对 SMC 特异性启动子/报告基因构建体调节的贡献程度。目标 2 - 确定 Rho 影响 SMC 中 SRF 依赖性转录的机制。我们将研究 Rho 对 SRF 磷酸化和 DNA 结合的调节以及 SRF 与其他转录因子相互作用的影响。目标 3 - 确定 RhoA 信号传导对 SMC 体内分化的影响。由于 SMC 表型受到许多不同环境因素的调节，而这些环境因素无法在培养模型中准确再现，因此研究 SMC 体内分化非常重要。我们将使用 SM 肌球蛋白重链启动子在小鼠 SMC 中特异性表达 Rho 的显性失活和组成型活性形式。测量 SMC 分化标志物基因表达、检查血管和器官形态并确定增殖指数将评估对 SMC 分化的影响。完成这些目标应该可以更好地理解调节 SMC 分化的信号传导和转录机制，并可能导致心血管治疗干预的新靶标。