DEGRADATION OF E12/E47 IN SMOOTH MUSCLE CELLS

平滑肌细胞中 E12/E47 的降解

• 批准号: 2857551
• 负责人: GORDON S HUGGINS
• 金额: $ 11.5万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-01-01 至 2002-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2857551
• 关键词: active sites; antisense nucleic acid; cell adhesion; cell differentiation; cell growth regulation; cell migration; cell proliferation; chemical conjugate; enzyme inhibitors; enzyme mechanism; extracellular matrix proteins; laboratory rat; molecular cloning; phosphorylation; proteasome; protein binding; protein degradation; protein metabolism; protein structure; ubiquitin; vascular smooth muscle

DESCRIPTION (Adapted from applicants' abstract) The proliferation and dedifferentiation of vascular smooth muscle cells (VSMC) is a principal component of occlusive vascular disease, which is the leading cause of death in developed countries. As a preventive cardiologist, the candidate's principle focus has been to identify and treat the root causes of atherosclerosis in patients with established cardiovascular disease. As an investigator, he seeks to further the understanding of this disease. The basic helix-loop-helix proteins E12 and E47 promote cellular differentiation and retard cellular proliferation. They are expressed in VSMC. The applicant cloned a novel ubiquitin conjugating enzyme, UBC-E2A, directed against the E12 and E47. The ubiquitin-proteasome system can dramatically alter the cellular phenotype by regulating the degradation of trans-acting factors. By targeting E12 and E47 for turnover, UBC-E2A may promote VSMC proliferation and dedifferentiation. The project herein will characterize E12 and E47 protein degradation in VSMC. First he will map the UBC-E2A binding site on the E2A proteins, then he will determine the role of E2A protein phosphorylation on their degradation. This analysis will permit the design of a peptide to compete with UBC-E2A for binding the E2A proteins and inhibit E2A protein degradation. Stable constitutive and tetracycline-inducible smooth muscle cell clones of full-length sense and antisense UBC-E2A and UBC-E2A inhibitor will be generated. He will use these clones to study the effect of UBC-E2A on the cell cycle restriction point, which regulates cellular proliferation. In addition, he will study the effects of UBC-E2A on VSMC migration, adhesion, and matrix protein production. The Cardiovascular Biology Laboratory of the Harvard School of Public Health will serve as the site for these studies. The laboratory encompasses 12,000 square feet and is equipped with instruments designed for research in cellular and molecular biology. (End of Abstract)

描述 （改编自申请人的摘要）扩散和推导 血管平滑肌细胞（VSMC）是闭塞的主要成分 血管疾病，这是发达的主要死亡原因 国家。 作为预防性心脏病专家，候选人的主要重点 一直在识别和治疗动脉粥样硬化的根本原因 患有既定心血管疾病的患者。 作为调查员，他 试图进一步理解这种疾病。 基本 螺旋环螺旋蛋白E12和E47促进细胞分化和 延迟细胞增殖。 它们在VSMC中表示。 申请人 克隆了一种新颖的泛素结合酶，UBC-E2A，针对 E12和E47。 泛素 - 蛋白酶体系统可以极大地改变 通过调节跨作用因子的降解，细胞表型。 通过针对E12和E47进行营业额，UBC-E2A可以促进VSMC 扩散和去分化。 这里的项目将描述 VSMC中的E12和E47蛋白质降解。 首先，他将映射UBC-E2A E2A蛋白上的结合位点，然后他将确定E2A的作用 蛋白质磷酸化的降解。 该分析将允许 设计肽以与UBC-E2A竞争以结合E2A蛋白和 抑制E2A蛋白降解。 稳定的本构和 四环素诱导的平滑肌细胞克隆具有全长感和 将产生反义UBC-E2A和UBC-E2A抑制剂。 他会使用 这些克隆研究UBC-E2A对细胞周期限制的影响 点，调节细胞增殖。 此外，他还将学习 UBC-E2A对VSMC迁移，粘附和基质蛋白的影响 生产。 哈佛大学的心血管生物学实验室 公共卫生将作为这些研究的网站。 实验室 包括12,000平方英尺 细胞和分子生物学研究。 （抽象的结尾）