DEGRADATION OF E12/E47 IN SMOOTH MUSCLE CELLS

描述 (改编自申请者摘要)增殖和去分化 血管平滑肌细胞(VSMC)是血管闭塞的主要成分 血管疾病，这是发达国家的主要死亡原因 国家。作为预防心脏病专家，候选人的主要关注点是 一直致力于识别和治疗动脉粥样硬化的根本原因 患有既往心血管疾病的患者。作为一名调查员，他 旨在加深对这种疾病的了解。最基本的 螺旋-环-螺旋蛋白E12和E47促进细胞分化和 延缓细胞增殖。它们在VSMC中表达。申请人 克隆了一种新的泛素结合酶UBC-E2A，它针对 E12和E47。泛素-蛋白酶体系统可以显著改变 通过调节反式作用因子的降解来表达细胞表型。 通过瞄准E12和E47进行营业额，UBC-E2A可能会促进VSMC 增殖和去分化。这里的项目将描述 E12和E47蛋白在VSMC中的降解。首先，他将绘制UBC-E2A的地图 在E2a蛋白上的结合位点，然后他将确定E2a的作用 蛋白质磷酸化对其降解的影响。此分析将允许 设计一种与UBC-E2A竞争结合E2A蛋白和 抑制E2a蛋白降解。稳定的本构关系和 四环素诱导的全长义和全长平滑肌细胞克隆 将产生反义UBC-E2a和UBC-E2a抑制剂。他将使用 这些克隆用于研究UBC-E2A对细胞周期的限制作用 点，它调节细胞的增殖。此外，他还将学习 UBC-E2A对VSMC迁移、黏附和基质蛋白的影响 制作。哈佛大学心血管生物学实验室 公共卫生将作为这些研究的地点。实验室 占地12,000平方英尺，配备了专为 细胞和分子生物学研究。(摘要结束)

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)