Developmental Changes Affecting Cardiac Titin Function

影响心脏肌联蛋白功能的发育变化

• 批准号: 6808924
• 负责人: MARION Lewis GREASER
• 金额: $ 29.1万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-07-01 至 2008-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6808924
• 关键词: cardiac myocytes; chromatography; circular dichroism; developmental genetics; echocardiography; electrophoresis; embryo /fetus protein; gene mutation; genetic mapping; glutamates; heart circulation; heart function; hemodynamics; laboratory rabbit; laboratory rat; mature animal; molecular assembly /self assembly; muscle proteins; muscle tension; peptides; phosphorylation; protein binding; protein isoforms; protein sequence; protein structure function; surface plasmon resonance

DESCRIPTION (provided by applicant): Titin is a 3000-4000 kD protein found in heart and skeletal muscle, and it has been proposed to play a major role in controlling the resting or passive tension in these tissues. Major developmental changes have been found to occur in cardiac titin as a result of alternative splicing pathways. A unique rat strain has been discovered with an autosomal dominant mutation that leads to a delayed fetal-to-adult titin transition pattern. These animals will be used to test the role of titin in cardiac hemodynamics in the intact animal using echocardiography and pressure-volume relationship measurements. Mechanical experiments on single cardiomyocytes will test hypotheses regarding titin's role in rest tension and the Frank-Starling relationship. Interactions betweer_ positively charged PPAK peptides from titin's extensible PEVK region with negatively charged polyE peptides will be tested using chromatographic, circular dichroism, electrophoretic, and surface plasmon resonance techniques. Studies will be designed to determine if binding of the PPAK and polyE peptides is specific to adjacent regions in the sequence or if multiple types of interactions can occur. Nontitin peptides rich in glutamic acid will also be examined for binding. Previously proposed phosphorylation sites will be identified in expressed titin polypeptides, and these sites will also be assayed for )hosphorylation state in intact fetal and adult titins. These experiments will test the hypothesis that certain _ites must be phosphorylated for proper assembly and/or function. Gene mapping studies will be conducted to localize the mutation site leading to the delayed developmental titin isoform program. The proposed studies will provide new information regarding the structure and function of this giant protein and its relation to human health and cardiovascular disease. The work may also provide novel insights on mechanisms of alternative splicing, an area of increasing importance in understanding the proteome.

描述（由申请人提供）：TITIN是在心脏和骨骼肌中发现的3000-4000 kD蛋白，并已提议在控制这些组织中的静息或被动张力方面发挥重要作用。 由于替代剪接途径，已经发现在心脏滴定中发生了重大的发育变化。 已经发现了一种独特的大鼠菌株，它具有常染色体显性突变，从而导致胎儿到成年titin过渡模式延迟。 这些动物将用于使用超声心动图和压力量关系测量值来测试钛在完整动物中心脏血流动力学中的作用。 关于单个心肌细胞的机械实验将检验有关泰丁在静止张力和坦率宣传关系中的作用的假设。 从TITIN的可扩展PEVK区域带带负电荷Polye肽的阳性PPAK肽的相互作用将使用色谱，圆形二色性，电泳和表面等离子体共振技术进行测试。 将设计研究以确定PPAK和Polye肽的结合是否特定于序列中的相邻区域，还是可能发生多种类型的相互作用。 还将检查富含谷氨酸的非丁蛋白肽的结合。 先前提出的磷酸化位点将在表达的钛二肽中鉴定出来，并且这些位点还将在完整的胎儿和成人钛蛋白中分析）磷酸化状态。 这些实验将检验以下假设：某些_位必须磷酸化以进行适当的组装和/或功能。 将进行基因映射研究，以定位突变位点，导致延迟发育titin同工型程序。 拟议的研究将提供有关该巨型蛋白质及其与人类健康和心血管疾病的关系的新信息。 这项工作还可以提供有关替代剪接机制的新颖见解，这是对理解蛋白质组的重要性越来越重要的领域。