Cardiac Muscle: Functional Role of Myosin - Binding Protein C in Contraction

心肌：肌球蛋白 - 结合蛋白 C 在收缩中的功能作用

• 批准号: 7880944
• 负责人: Robert W. Kensler
• 金额: $ 18.75万
• 依托单位: UNIVERSITY OF PUERTO RICO MED SCIENCES
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-08-15 至 2012-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7880944
• 关键词: Actins; Adrenergic Agents; Affect; Animals; Binding; Binding Sites; Biological Models; Caliber; Cardiac; Cardiac Myosins; Cardiovascular Physiology; Complement component C1s; Data; Electron Microscopy; England; Familial Hypertrophic Cardiomyopathy; Filament; Genetic; Goals; Head; Heart; Human; Hypertrophic Cardiomyopathy; Incubated; Knockout Mice; Location; Methods; Modeling; Mus; Mutation; Myocardial; Myocardial Contraction; Myocardium; Myosin ATPase; Myosin Heavy Chains; N-terminal; Oryctolagus cuniculus; Pathology; Phosphorylation; Play; Positioning Attribute; Property; Protein Isoforms; Proteins; Rattus; Reporting; Research; Resolution; Rodent; Role; Site; Structural Models; Structure; Surface; Testing; Thick; Thick Filament; Vertebral column; adrenergic; base; clinically relevant; interest; myosin-binding protein C; non-muscle myosin heavy chain-B; particle; reconstruction; research study; response

DESCRIPTION (provided by applicant): Mutations in the thick filament associated protein cMyBP-C are a major cause of familial hypertrophic cardiomyopathy. The pathological effects of mutations in cardiac myosin binding protein-C are consistent with increasing evidence that it plays both a structural and a regulatory role in myocardial contraction. Although the evidence suggesting a regulatory role for cMyBP-C in myocardial contraction appears compelling, the mechanism of action is far from clear. Most of the research on the functional role of cMyBPC and the effect of its phosphorylation has been done in rat or mouse myocardium which contains primarily the a-myosin heavy chain, despite evidence that thick filaments containing the (3-myosin heavy chain (such as in humans) may differ in the changes shown upon phosphorylation of cMyBP-C. This suggests the importance of investigating in more detail the effect of phosphorylation of cMyBP-C on the structure of the myocardial thick filament in species other than the rat; including species with the (3-myosin heavy chain such as the rabbit. In addition, increasing evidence suggests that N-terminus fragments of cMyBP-C may be able effect myocardial contractility independent of a tethering of the myosin S2 region to the backbone of the filament by cMyBP-C, and may also be able to directly bind actin. These studies emphasize the need to look at the effects of binding of these N-terminus fragments on the cross bridge arrangement and structure of the isolated cardiac thick filament; and further assess by electron microscopy the binding potential of these Nterminus fragments of cMyBP-C to actin. In addition in order to fully understand how cMyBP-C functions in the thick filament, it is vital to have higher resolution 30-reconstructions of the cardiac thick filaments than are currently available. The specfic aims of the research are to: 1) Determine the effect of phosphorylation of cMyBP-C on the crossbridge arrangement of isolated mouse, rat, and rabbit cardiac thick filaments; 2) Determine the effect of the C1-C2 and CO-C1 N-terminus fragments of cMyBP-C on the crossbridge arrangement of the isolated thick filaments from rabbit heart, and both wildtype and cMyBP-C"'" knockout mouse hearts; 3) Assess the of binding of C1-C2 and CO-C2 N-terminus fragments of cMyBP-C to actin; and 4) Compute higher resolution 3D-reconstructions of the cardiac thick filaments from both mouse and rabbit thick filaments by single particle analysis to precisely locate the myosin heads and cMyBP-C.

描述（由申请人提供）：粗丝相关蛋白cMyBP-C的突变是家族性肥厚型心肌病的主要原因。心肌肌球蛋白结合蛋白-C 突变的病理效应与越来越多的证据相一致，即它在心肌收缩中发挥结构和调节作用。尽管表明 cMyBP-C 在心肌收缩中发挥调节作用的证据似乎令人信服，但其作用机制尚不清楚。大多数关于 cMyBPC 的功能作用及其磷酸化影响的研究都是在主要含有 α-肌球蛋白重链的大鼠或小鼠心肌中进行的，尽管有证据表明含有 3-肌球蛋白重链（例如在人类中）的粗丝在 cMyBP-C 磷酸化时显示的变化可能有所不同。这表明更详细地研究磷酸化作用的重要性。 cMyBP-C 对大鼠以外物种心肌粗丝结构的影响；包括具有 3-肌球蛋白重链的物种，例如兔子。此外，越来越多的证据表明，cMyBP-C 的 N 末端片段可能能够影响心肌收缩力，而不依赖于 cMyBP-C 将肌球蛋白 S2 区域束缚在肌丝主干上，并且还可能能够直接结合肌动蛋白。这些研究 强调需要研究这些 N 末端片段的结合对离体心脏粗丝的跨桥排列和结构的影响；并通过电子显微镜进一步评估这些cMyBP-C N末端片段与肌动蛋白的结合潜力。此外，为了充分了解 cMyBP-C 在粗丝中的功能，对心脏粗丝进行更高分辨率的 30 重建至关重要。 可用。本研究的具体目的是： 1）确定cMyBP-C磷酸化对离体小鼠、大鼠和兔心脏粗丝桥排列的影响； 2) 确定cMyBP-C的C1-C2和CO-C1 N端片段对兔心脏分离的粗丝的横桥排列的影响，以及野生型和cMyBP-C"'" 敲除小鼠心脏； 3) 评估cMyBP-C的C1-C2和CO-C2 N端片段与肌动蛋白的结合； 4) 通过单粒子分析计算小鼠和兔子粗丝的心脏粗丝的更高分辨率 3D 重建，以精确定位肌球蛋白头和 cMyBP-C。