Cardiac Myosin-Binding Protein C: Molecular Mechanisms Governing Cardiac Contractility

心肌肌球蛋白结合蛋白 C：控制心脏收缩力的分子机制

基本信息

批准号：
10624275
负责人：
Sivaraj Sivaramakrishnan
金额：
$ 54.49万
依托单位：
UNIVERSITY OF VERMONT & ST AGRIC COLLEGE
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-06-01 至 2024-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10624275
关键词：
Ablation Actins Actomyosin Address Adrenergic Agents Affect Atomic Force Microscopy Binding Cardiac Cardiac Myosins Collaborations Contractile Proteins DNA DNA Sequence Alteration Defect Development Equilibrium Filament Generations Head Heart Heart failure Human Hypertrophic Cardiomyopathy Knowledge Lasers Location Mass Spectrum Analysis Measures Modeling Molecular Molecular Conformation Molecular Structure Mosses Motion Mutagenesis Mutation Myocardium Myofibrils Myosin ATPase N-terminal Nanotubes Performance Periodicity Phosphorylation Physiological Positioning Attribute Preparation Proteins Pump Regulation Relaxation Role Sarcomeres Structure Sudden Death Surface Techniques Testing Thick Thick Filament Thin Filament Thinness Time Transgenic Mice Vertebral column beta-Myosin biophysical techniques cell motility citrate carrier flexibility improved in vivo molecular mechanics mutant myosin-binding protein C nanometer nanoscale novel novel strategies operation recruit response single molecule spatial relationship superresolution imaging targeted treatment young adult

项目摘要

Cardiac myosin-binding protein C (cMyBP-C) is a sarcomeric thick filament associated protein that is essential to normal cardiac structure and function. The importance of cMyBP-C is emphasized by mutations to cMyBP-C being a leading cause of hypertrophic cardiomyopathy. Despite being a key regulator of cardiac contractility, the molecular mechanism by which cMyBP-C modulates actomyosin force and motion generation is far from certain. Although cMyBP-C's N-terminal domains can bind to actin and the myosin head region, it is not known which of these binding partners is physiologically relevant and whether these binding partner interactions modulate cardiac contractility by directly affecting actomyosin power generation or indirectly by altering Ca2+- dependent thin filament activation. With phosphorylation of cMyBP-C's N terminus occurring in response to β- adrenergic stimulation, phosphorylation may offer a measure of cMyBP-C functional tunability in order to enhance cardiac contractility. To address these questions, we propose two specific aims. Aim 1 will test the hypothesis that phosphorylation modulates cMyBP-C's N-terminal domain structure to influence its binding partner interactions (i.e. thin filament and myosin head region). We will use a novel mass-spectrometry technique and atomic force microscopy to characterize the molecular mechanics of cMyBP-C's N terminus that has been structurally altered due to phosphorylation or mutagenesis. The functional impact of these structural perturbations will be characterized in the context of cardiac myofibrils and native thick filaments to determine if cMyBP-C operates only where it exist in the thick filament and whether it can sequester cardiac myosin into a reserve pool of super-relaxed myosin heads. Thus, we will measure the location and time course of fluorescent-ATP turnover in single cardiac myofibrils and the force generated by native thick filaments in the laser trap in preparations from transgenic mice expressing phosphorylation and binding partner ablated mutant cMyBP-C. In Aim 2 we will create DNA-based “designer” thick filament nanotubes to define how the spatial relationships that normally exist in the thick filament between cMyBP-C and its myosin and actin binding partners are critical determinants of cMyBP-C's modes of operation. These DNA-nanotubes will allow exquisite nanometer spatial positioning of expressed cMyBP-C and human β-cardiac myosin on the nanotube surface relative to each other. By this novel approach we can assign cMyBP C's modulation of actomyosin motility to binding of the myosin head and/or thin filament, as assessed by both thin filament motility and force generation using the laser trap. With the knowledge and understanding of cMyBP-C function derived from these collective studies, targeted therapies directed at cMyBP-C binding partner interactions may be developed to help modulate and to improve cardiac performance in the failing heart.

心肌肌球蛋白结合蛋白 C (cMyBP-C) 是一种必需的肌节粗丝相关蛋白 cMyBP-C 的突变强调了 cMyBP-C 对正常心脏结构和功能的重要性。尽管是心肌收缩力的关键调节因子，但它是肥厚型心肌病的主要原因。 cMyBP-C 调节肌动球蛋白力和运动产生的分子机制远非如此。虽然 cMyBP-C 的 N 端结构域可以结合肌动蛋白和肌球蛋白头部区域，但尚不清楚。这些结合伙伴中的哪一个具有生理相关性以及这些结合伙伴是否相互作用通过直接影响肌动球蛋白发电或通过改变 Ca2+- 间接调节心肌收缩力依赖于 β- 的 cMyBP-C 的 N 末端发生磷酸化。肾上腺素能刺激、磷酸化可以提供 cMyBP-C 功能可调性的测量，以便为了解决这些问题，我们提出了两个具体目标：假设磷酸化调节 cMyBP-C 的 N 端结构域结构以影响其结合合作伙伴相互作用（即细丝和肌球蛋白头部区域）我们将使用一种新颖的质谱分析法。技术和原子力显微镜来表征 cMyBP-C N 末端的分子力学由于磷酸化或诱变而发生结构改变这些结构的功能影响。扰动将在心肌原纤维和天然粗丝的背景下进行表征，以确定是否 cMyBP-C 仅在粗丝中存在的地方起作用，以及它是否可以将心肌肌球蛋白隔离成因此，我们将测量超松弛肌球蛋白头的储备池的位置和时间过程。单个心肌原纤维中的荧光 ATP 周转以及心肌原纤维中天然粗丝产生的力表达磷酸化和结合伴侣消融突变体的转基因小鼠制剂中的激光陷阱在目标 2 中，我们将创建基于 DNA 的“设计师”粗丝纳米管来定义空间如何通常存在于粗丝中的 cMyBP-C 与其肌球蛋白和肌动蛋白结合之间的关系合作伙伴是 cMyBP-C 运行模式的关键决定因素，这些 DNA 纳米管将实现精致。表达的cMyBP-C和人β-心肌肌球蛋白在纳米管表面的纳米空间定位通过这种新颖的方法，我们可以将 cMyBP C 对肌动球蛋白运动的调节分配给肌球蛋白头和/或细丝的结合，通过细丝运动和力的产生来评估使用激光陷阱的知识和对 cMyBP-C 功能的理解源自于这些集体。研究表明，针对 cMyBP-C 结合伴侣相互作用的靶向疗法可能会被开发出来以帮助调节和改善衰竭心脏的心脏功能。

项目成果

期刊论文数量（7）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Dilated cardiomyopathy mutation in beta-cardiac myosin enhances actin activation of the power stroke and phosphate release.

扩张型心肌病β-心肌肌球蛋白突变增强了动力冲程的肌动蛋白激活和磷酸盐释放。

DOI：
10.1101/2023.11.10.566646
发表时间：
2023
期刊：
bioRxiv : the preprint server for biology
影响因子：
0
作者：
Bodt,SkylarML;Ge,Jinghua;Ma,Wen;Rasicci,DavidV;Desetty,Rohini;McCammon,JAndrew;Yengo,ChristopherM
通讯作者：
Yengo,ChristopherM

Cardiac myosin binding protein-C phosphorylation accelerates β-cardiac myosin detachment rate in mouse myocardium

心肌肌球蛋白结合蛋白-C 磷酸化加速小鼠心肌中β-心肌肌球蛋白脱离率

DOI：
10.1152/ajpheart.00673.2020
发表时间：
2021
期刊：
American Journal of Physiology-Heart and Circulatory Physiology
影响因子：
4.8
作者：
Tanner, Bertrand C.;Previs, Michael J.;Wang, Yuan;Robbins, Jeffrey;Palmer, Bradley M.
通讯作者：
Palmer, Bradley M.

Nanosurfer assay dissects β-cardiac myosin and cardiac myosin-binding protein C interactions.

DOI：
10.1016/j.bpj.2022.05.013
发表时间：
2022-06-21
期刊：
BIOPHYSICAL JOURNAL
影响因子：
3.4
作者：
Touma, Anja M.;Tang, Wanjian;Rasicci, David, V;Vang, Duha;Rai, Ashim;Previs, Samantha B.;Warshaw, David M.;Yengo, Christopher M.;Sivaramakrishnan, Sivaraj
通讯作者：
Sivaramakrishnan, Sivaraj

A dynamic Dab2 keeps myosin VI stably on track.

DOI：
10.1016/j.jbc.2021.100640
发表时间：
2021-01
期刊：
The Journal of biological chemistry
影响因子：
0
作者：
Cirilo JA Jr;Yengo CM
通讯作者：
Yengo CM

Dilated cardiomyopathy mutation E525K in human beta-cardiac myosin stabilizes the interacting-heads motif and super-relaxed state of myosin.

DOI：
10.7554/elife.77415
发表时间：
2022-11-24
期刊：
eLife
影响因子：
7.7
作者：
Rasicci DV;Tiwari P;Bodt SML;Desetty R;Sadler FR;Sivaramakrishnan S;Craig R;Yengo CM
通讯作者：
Yengo CM