Novel functions of E-C coupling structural protein junctophilin-2 in the heart

E-C耦合结构蛋白junctophilin-2在心脏中的新功能

• 批准号: 9199431
• 负责人: Long-Sheng Song
• 金额: $ 48.85万
• 依托单位: UNIVERSITY OF IOWA
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-01-15 至 2019-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9199431
• 关键词: Animal Model; Binding; Biochemistry; CRISPR/Cas technology; Calpain; Cardiac; Cardiac Myocytes; Cardiac development; Cardiomyopathies; Cell Nucleus; Cell Proliferation; Cleaved cell; Clustered Regularly Interspaced Short Palindromic Repeats; Communication; Complex; Coupling; Data; Development; Down-Regulation; Echocardiography; Electrophysiology (science); Event; Future; Gene Expression; Gene Expression Profile; Genes; Genetic Transcription; Goals; Healthcare Systems; Heart; Heart Diseases; Heart Hypertrophy; Heart failure; Hospitalization; Human; Imaging Techniques; In Situ; Knock-in Mouse; Length; Light; Mediating; Membrane; Messenger RNA; Modeling; Molecular; Molecular Biology; Molecular Chaperones; Muscle Cells; N-terminal; Nuclear; Nuclear Localization Signal; Pathogenesis; Pathologic; Patients; Phenotype; Physiological; Pilot Projects; Property; Proteins; Publishing; Regulation; Reporter; Sarcoplasmic Reticulum; Signal Transduction; Stress; Structural Protein; System; Technology; Testing; Therapeutic; Transcriptional Regulation; Transducers; Transgenic Mice; Transgenic Organisms; Transmembrane Domain; Ventricular; base; chromatin modification; chromatin remodeling; in situ imaging; insight; interdisciplinary approach; junctophilin; mouse model; mutant; novel; novel therapeutic intervention; promoter; public health relevance; response; scaffold; transcriptome

 DESCRIPTION (provided by applicant): Junctophilin 2 (JP2) is a structural protein required for the formation of junctional couplings (i.e., cardiac dyads) between transverse (T)-tubule membrane and sarcoplasmic reticulum (SR) and is fundamental for local control of Ca2+-induced Ca2+ release and efficient contraction in ventricular myocytes during cardiac excitation- contraction (E-C) coupling. In animal models of heart failure, JP2 protein levels progressively decline, leading to T-tubule remodeling and loss of E-C coupling function. JP2 levels are also markedly reduced in human failing hearts. Our recently published and preliminary data demonstrate that JP2 is posttranslational regulated by calpain, a Ca2+-activated protease implicated in a variety of heart diseases. JP2 cleavage by calpain was originally hypothesized to underlie JP2 downregulation in failing hearts. In pilot studies, we made the novel and unexpected observation that an N-terminal JP2 cleavage fragment, termed "JP2NT," unlike full length JP2, is imported into nucleus, where it has a profound function in transcriptional reprogramming in cardiomyocytes. However, the (patho) physiological consequences of JP2NT nuclear localization and related molecular mechanisms are completely unknown. Based on these novel exciting preliminary findings, we hypothesize that, in response to stress, calpain-mediated cleavage of JP2 transforms JP2 from a structural protein into a transcriptional regulator that reprograms the transcriptional profile of damaged cardiomyocytes and represses the development of cardiac hypertrophy. This molecular event may be an important trigger for transcriptional remodeling, serving as an unappreciated protective mechanism under pathological conditions. We will test our hypothesis using a multidisciplinary approach, including multiple novels transgenic and CRISPR-based mouse models, molecular biology, biochemistry and a novel in situ confocal imaging technique developed by our lab. In Aim 1, we will explore the molecular mechanisms underlying JP2NT transcriptional regulation in cardiomyocytes, whereas Aim 2 will examine the patho(physiological) consequences of JP2NT in the heart. We anticipate the proposed studies will provide significant insights into the novel functions of JP2NT in membrane-to-nucleus signal transduction in the setting of cardiac stress and new insights into the cross- communication between cardiomyocyte structural remodeling and transcriptional remodeling in heart disease. This information will serve as a platform for the future development of novel heart failure therapeutics.

 描述（由申请人提供）：Junctophilin 2 (JP2) 是横 (T) 管膜和肌浆网 (SR) 之间形成连接耦合（即心脏二元组）所需的结构蛋白，对于心脏兴奋-收缩期间心室肌细胞中 Ca2+ 诱导的 Ca2+ 释放和有效收缩的局部控制至关重要 (E-C)耦合。在心力衰竭动物模型中，JP2 蛋白水平逐渐下降，导致 T 管重塑和 E-C 耦合功能丧失。人类衰竭心脏中的 JP2 水平也显着降低。我们最近发表的初步数据表明，JP2 受到钙蛋白酶（一种与多种心脏病有关的 Ca2+ 激活蛋白酶）的翻译后调节。最初假设钙蛋白酶对 JP2 的裂解是衰竭心脏中 JP2 下调的基础。在初步研究中，我们进行了新颖且意想不到的观察，即与全长 JP2 不同，N 端 JP2 裂解片段（称为“JP2NT”）被导入细胞核，在心肌细胞的转录重编程中具有深远的功能。然而，JP2NT 核定位的（病理）生理后果和相关分子机制尚不清楚。基于这些新颖的、令人兴奋的初步发现，我们假设，为了应对应激，钙蛋白酶介导的 JP2 裂解将 JP2 从结构蛋白转变为转录调节因子，重新编程受损心肌细胞的转录谱并抑制心脏肥大的发展。这种分子事件可能是转录重塑的重要触发因素，在病理条件下作为一种未被重视的保护机制。我们将使用多学科方法来检验我们的假设，包括多种新型转基因和基于 CRISPR 的小鼠模型、分子生物学、生物化学以及我们实验室开发的新型原位共聚焦成像技术。在目标 1 中，我们将探索心肌细胞中 JP2NT 转录调控的分子机制，而目标 2 将研究 JP2NT 在心脏中的病理（生理）后果。我们预计拟议的研究将为 JP2NT 的新颖功能提供重要见解 心脏应激环境下膜到核信号转导的研究，以及对心脏病中心肌细胞结构重塑和转录重塑之间交叉交流的新见解。该信息将作为未来开发新型心力衰竭疗法的平台。