GENETIC ANALYSIS OF CARDIAC TERMINAL DIFFERENTIATION

心脏终末分化的遗传分析

• 批准号: 6390317
• 负责人: William Robb MacLellan
• 金额: $ 22.95万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-09-01 至 2004-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6390317
• 关键词: binding proteins; cardiac myocytes; cell differentiation; inhibitor /antagonist; laboratory rat; protein protein interaction; protein structure function; retinoblastoma protein; tissue /cell culture

Terminal differentiation is a process whereby highly specialized cells commit irreversibly to a cellular fate and undergo growth arrest. Recent studies in cardiac muscle have identified two major families of proteins involved in cardiac terminal differentiation, pocket proteins (retinoblastoma gene product (Rb), p107, p130) and the transcriptional coactivators, p300/CBP. One explanation for the dependence of normal differentiation on pocket proteins, is the requirement of some tissue-specific transcription factors on the coexpression of Rb for transcriptional activity. This led us and others to hypothesize that Rb might be the key to terminal differentiation in cardiac myocytes. However, the mechanism underlying this dependence or the factors interacting with Rb are unknown. Therefore, to determine the factors interacting with Rb in cardiac muscle, we utilized the yeast two-hybrid system to identify cardiac Rb- binding proteins. One novel protein, designated p300- and Rb- binding Inhibitor of differentiation-1 (PRI-1) was the predominant Rb-binding clone. PRI-1 is preferentially expressed in cardiac and skeletal muscle in a developmentally regulated manner. Initial studies in skeletal muscle demonstrated that overexpression of PRI-1 inhibited muscle-specific transcription. Repression of skeletal muscle-restricted genes was mediated by a block to transactivation by MyoD, independent of G1 exit, and, surprisingly, was potentiated by a mutation that prevents PRI-1 binding to Rb. Inhibition of MyoD may be explained by PRI-1's ability, like adenoviral protein E1A, to bind p300, an essential MyoD coactivator. Thus, PRI-1 binds both Rb and p300, was inhibited by Rb in differentiated muscle, and was a novel repressor of MyoD function. Studies in cardiac muscle demonstrate that PRI-1 also inhibits cardiac-specific gene expression although the mechanism or potential targets of PRI-1's inhibitory effects are unknown. Thus PRI-1 represents one of the first endogenous, mammalian inhibitors of cardiac differentiation to be identified. This first RO1 proposes to further define the role of PRI-1 in cardiac myocyte differentiation and elucidate its biochemical properties.

终末分化是高度特化的细胞不可逆地决定细胞命运并经历生长停滞的过程。 最近对心肌的研究已经确定了参与心脏终末分化的两个主要蛋白质家族：口袋蛋白（视网膜母细胞瘤基因产物（Rb）、p107、p130）和转录共激活因子p300/CBP。正常分化依赖于口袋蛋白的一种解释是，一些组织特异性转录因子需要 Rb 共表达来实现转录活性。 这导致我们和其他人推测 Rb 可能是心肌细胞终末分化的关键。 然而，这种依赖性的机制或与 Rb 相互作用的因素尚不清楚。 因此，为了确定与心肌中 Rb 相互作用的因素，我们利用酵母双杂交系统来鉴定心脏 Rb 结合蛋白。 一种称为 p300 和 Rb 结合分化抑制剂 (PRI-1) 的新型蛋白质是主要的 Rb 结合克隆。 PRI-1 以发育调节方式优先在心肌和骨骼肌中表达。 对骨骼肌的初步研究表明，PRI-1 的过度表达会抑制肌肉特异性转录。骨骼肌限制性基因的抑制是通过 MyoD 反式激活的阻断介导的，与 G1 退出无关，而且令人惊讶的是，抑制 PRI-1 与 Rb 结合的突变会增强这种抑制。 对 MyoD 的抑制可能是由于 PRI-1 与腺病毒蛋白 E1A 一样能够结合 p300（一种重要的 MyoD 共激活因子）的能力。 因此，PRI-1 结合 Rb 和 p300，在分化的肌肉中被 Rb 抑制，并且是一种新型的 MyoD 功能抑制因子。 心肌研究表明，PRI-1 还可以抑制心脏特异性基因表达，尽管 PRI-1 抑制作用的机制或潜在目标尚不清楚。 因此，PRI-1 代表了第一个被鉴定的内源性哺乳动物心脏分化抑制剂之一。 第一个 RO1 提议进一步定义 PRI-1 在心肌细胞分化中的作用并阐明其生化特性。