Function of Giant Sarcomere Matrix Proteins in Muscle

肌肉中巨肌节基质蛋白的功能

• 批准号: 6780842
• 负责人: Henk L. GRANZIER
• 金额: $ 33.03万
• 依托单位: WASHINGTON STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-08-01 至 2007-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6780842
• 关键词: atomic force microscopy; beta adrenergic agent; calcium ion; cardiac myocytes; elasticity; fluorescence spectrometry; genetically modified animals; heart function; immunoelectron microscopy; laboratory mouse; microarray technology; muscle proteins; muscle tension; myocardium; phosphorylation; polymerase chain reaction; posttranslational modifications; protein isoforms; protein kinase A; protein kinase C; protein structure function; recombinant proteins; sarcomeres; striated muscles

DESCRIPTION (provided by applicant): Our long-term objective is to understand the roles of titin in cardiac function and disease. Titin is the third myofilament of striated muscle and contains an extensible region, located in the I-band of the sarcomere, which functions as a molecular spring that, when extended, develops force. This force underlies a large fraction of the force developed by myocardium during the filling phase of the heart. Human myocardium expresses both stiff and compliant titin isoforms. Altered isoform expression profiles have been found in patients with dilated cardiomyopathy (DCM) and coronary artery disease. These are accompanied by altered passive stiffness. Furthermore, recent studies have discovered several titin mutations that lead to DCM. We propose to study the molecular basis of titin's extensibility, including the effects of posttranslational modifications on extensibility. We will use recombinant proteins representing titin's various spring elements and study their mechanical and structural properties, using atomic force microscopy and spectroscopic techniques. The effects of calcium and phosphorylation on mechanics and structure will be examined. Because titin isoform expression may regulate calcium sensitivity and beta-adrenergic control of titin-based elasticity, we will develop a titin exon microarray to characterize exon-splicing patterns. We will also develop and characterize several mouse knockout models in which spring elements have been excised in order to study the following hypothesized functions: 1) glutamate rich PEVK elements are calcium binding sites that can alter titin-based passive force in a calcium-dependent manner; 2) the cardiac unique N2B sequence can alter titin-based passive force via phosphorylation. This research will provide important and novel insights into the molecular basis of titin's elasticity and its adjustments via calcium and phosphorylation. Considering the multitude of 'titinopathies' that are rapidly being discovered, our findings will be important for understanding human cardiac function and disease.

描述（由申请人提供）：我们的长期目标是了解肌联蛋白在心脏功能和疾病中的作用。肌联蛋白是横纹肌的第三肌丝，并且包含位于肌节的I带中的可伸展区域，其充当分子弹簧，当伸展时，产生力。该力是在心脏的充盈阶段期间由心肌产生的力的大部分的基础。人心肌表达刚性和顺应性肌联蛋白亚型。在扩张型心肌病（DCM）和冠状动脉疾病患者中发现了异构体表达谱的改变。这些都伴随着改变被动刚度。此外，最近的研究发现了几种导致DCM的肌联蛋白突变。 我们建议研究肌联蛋白的可延伸性的分子基础，包括翻译后修饰对可延伸性的影响。我们将使用重组蛋白代表肌联蛋白的各种弹簧元件，并研究其机械和结构特性，使用原子力显微镜和光谱技术。将检查钙和磷酸化对力学和结构的影响。由于肌联蛋白亚型表达可能调节钙敏感性和β-肾上腺素能控制肌联蛋白为基础的弹性，我们将开发一个肌联蛋白外显子微阵列来表征外显子剪接模式。我们还将开发和表征几种小鼠敲除模型，其中弹簧元件已被切除，以研究以下假设功能：1）富含谷氨酸的PEVK元件是钙结合位点，可以以钙依赖性方式改变基于肌联蛋白的被动力; 2）心脏独特的N2 B序列可以通过磷酸化改变基于肌联蛋白的被动力。 这项研究将为肌联蛋白弹性的分子基础及其通过钙和磷酸化的调节提供重要和新颖的见解。考虑到正在迅速发现的众多“肌钙蛋白病”，我们的发现对于了解人类心脏功能和疾病将非常重要。