Defining the mechanisms whereby MEK1/2 and ERK1/2 signaling control cardiomyocyte directional growth

定义 MEK1/2 和 ERK1/2 信号控制心肌细胞定向生长的机制

• 批准号: 9393390
• 负责人: Kelly Grimes
• 金额: $ 5.71万
• 依托单位: CINCINNATI CHILDRENS HOSP MED CTR
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-07-24 至 2020-07-23
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9393390
• 关键词: Adaptor Signaling Protein; Adenoviruses; Adult; Affect; Alpha Cell; Binding; Biological Assay; Cardiac; Cardiac Myocytes; Cardiovascular Diseases; Categories; Cell Nucleus; Cell Proliferation; Cells; Cytoplasm; Cytoskeletal Proteins; Cytosol; Data; Dilated Cardiomyopathy; Disease; Dissociation; Docking; Growth; Heart; Heart Hypertrophy; Heart failure; Hypertrophic Cardiomyopathy; Hypertrophy; Immunoprecipitation; Incidence; Individual; Knowledge; Laboratories; Length; Literature; Location; MAPK1 gene; MAPK3 gene; Mediating; Mediator of activation protein; Membrane; Mitogen-Activated Protein Kinases; Molecular; Movement; Mus; Muscle Cells; Nuclear; Pathologic; Pathway interactions; Patients; Pattern; Phosphorylation; Phosphotransferases; Play; Protein Kinase; Proteins; Proteomics; Publishing; Rattus; Regulation; Research; Role; Sarcomeres; Scaffolding Protein; Series; Signal Pathway; Signal Transduction; Stimulus; Stress; Time; Transgenic Mice; Ventricular; Width; Yeasts; arm; cell growth; cell type; exhaustion; experimental study; in vivo; inhibitor/antagonist; insight; nebulette; new therapeutic target; novel; novel therapeutics; phosphoproteomics; pressure; prevent; response; spatiotemporal; transcription factor; treatment strategy; yeast two hybrid system

Project Summary/Abstract Pathological growth of the heart can be separated into two categories: hypertrophic cardiomyopathy, whereby the ventricles undergo thickening and cardiomyocytes increase in width, and dilated cardiomyopathy, in which the ventricular walls become thinner and cardiomyocytes lengthen. New avenues of research to broaden our understanding of cardiac growth mechanisms are critical to treating the many patients worldwide suffering from these cardiovascular diseases. The experiments outlined in this proposal seek to increase the knowledge of such mechanisms by determining how cardiomyocytes grow preferentially in length or width. MEK1/2 and ERK1/2 have long been known to regulate cell growth and proliferation in many cell types. However, adult cardiomyocytes have very low proliferative capacities; therefore, the MEK1/2-ERK1/2 signaling pathway may be specialized in these cells to control growth. Indeed the sponsor's lab has previously found that these kinases regulate the switch between cardiomyocyte growth in different directions. When signaling through this pathway is increased, cardiomyocytes increase in width, adding sarcomeres in parallel. However, the inhibition of MEK1/2-ERK1/2 causes significant lengthening of cardiomyocytes and addition of sarcomeres in series. Canonical MEK1/2-ERK1/2 activation causes translocation of most ERK1/2 proteins to the nucleus to elicit cell growth; however, lengthening growth still occurs in cardiomyocytes when ERK1/2 do not translocate to the nucleus and remain in the cytoplasm. Therefore there are likely specific actions of MEK1/2 and ERK1/2 in the cytoplasm that regulate directional growth of cardiomyocytes, namely interactions with or regulation of proteins in the cytoplasm. This proposal seeks to understand the molecular underpinnings of these kinases' actions on directional growth by first determining the spatiotemporal changes in MEK1/2 and ERK1/2 during hypertrophic stimuli. The roles that these proteins' subcellular localizations play in directing cardiomyocyte lengthening or widening will also be assessed. Proteins which directly bind MEK1/2 and ERK1/2 will be identified through yeast two-hybrid and in vivo proteomics assays. These studies are underway and have already led to the identification of a novel interaction between ERK2 and the Z-disk protein nebulette. Finally a phosphoproteomic screen was conducted to determine proteins that may not necessarily bind MEK1/2 and ERK1/2 but may be downstream effectors. The results showed an array of targets, many of which are cytoskeletal or adaptor proteins, which displayed changes in phosphorylation upon the induction or inhibition of MEK1/2-ERK1/2 signaling. That so many cytoskeletal proteins were affected by changes in this signaling pathway supports the hypothesis that these kinases have specific actions in the cytoplasm to modulate directional growth. Overall, these studies will greatly expand our knowledge of the mechanisms that control cardiomyocyte growth in different directions, and thereby may provide us with new therapeutic targets to manipulate in conditions of pathological cardiac growth.

项目摘要/摘要 心脏的病理性生长可分为两类：肥厚性心肌病， 脑室增厚，心肌细胞增宽，扩张性心肌病， 室壁变薄，心肌细胞延长。拓宽研究的新途径 我们对心脏生长机制的了解对于治疗世界各地的许多患者来说至关重要 从这些心血管疾病中。这项提案中概述的实验试图增加人们对 通过确定心肌细胞如何在长度或宽度上优先生长来研究这种机制。MEK1/2及 长期以来，ERK1/2在许多细胞类型中调节细胞的生长和增殖。然而，成人 心肌细胞的增殖能力很低，因此，MEK1/2-ERK1/2信号通路可能是 专门研究这些细胞以控制生长。事实上，赞助商的实验室之前已经发现，这些激酶 调节心肌细胞向不同方向生长的切换。当通过这条通路发出信号时 是增加的，心肌细胞的宽度增加，平行增加肌节。然而，它的抑制作用 MEK1/2-ERK1/2导致心肌细胞显著延长和肌节串联增加。 典型的MEK1/2-ERK1/2激活导致大多数ERK1/2蛋白移位到细胞核以诱导细胞 然而，当ERK1/2不移位到心肌细胞中时，心肌细胞仍然发生延长的生长。 并留在细胞质中。因此，可能存在MEK1/2和ERK1/2在脑内的具体作用。 调节心肌细胞定向生长的细胞质，即与蛋白质的相互作用或调节 在细胞质中。这项提议试图了解这些激酶作用于 通过首先确定肥厚过程中MEK1/2和ERK1/2的时空变化来定向生长 刺激物。这些蛋白的亚细胞定位在指导心肌细胞延长或 还将对扩阔范围进行评估。将通过酵母鉴定与MEK1/2和ERK1/2直接结合的蛋白质 双杂交和体内蛋白质组学分析。这些研究正在进行中，并已导致识别 ERK2和Z-Disk蛋白星云之间的一种新的相互作用。最后进行了磷酸蛋白质组筛选。 进行以确定可能不一定结合MEK1/2和ERK1/2但可能位于下游的蛋白质 效应器。结果显示了一系列的靶点，其中许多是细胞骨架或接头蛋白，它们 在诱导或抑制MEK1/2-ERK1/2信号时，显示磷酸化的变化。是这样吗？ 许多细胞骨架蛋白受到这一信号通路变化的影响，支持这一假说 这些激酶在细胞质中有特定的作用来调节定向生长。总体而言，这些研究将 极大地扩展了我们对控制心肌细胞在不同方向生长的机制的了解，以及 从而可能为我们在病理性心脏生长条件下操作提供新的治疗靶点。