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）：嗜连接蛋白2（JP 2）是形成连接偶联（即，在横（T）-小管膜和肌浆网（SR）之间的心脏二分体（cardiac dyads），并且是在心脏兴奋-收缩（E-C）偶联期间局部控制心室肌细胞中Ca 2+诱导的Ca 2+释放和有效收缩的基础。在心力衰竭的动物模型中，JP 2蛋白水平逐渐下降，导致T-小管重塑和E-C偶联功能丧失。JP 2水平在人类衰竭的心脏中也显著降低。我们最近发表的初步数据表明，JP 2是翻译后调节钙蛋白酶，钙激活蛋白酶涉及各种心脏疾病。最初假设钙蛋白酶对JP 2的切割是衰竭心脏中JP 2下调的基础。在初步研究中，我们进行了新的和意想不到的观察，N-末端JP 2切割片段，称为“JP 2NT”，不像全长JP 2，被导入到细胞核中，在那里它在心肌细胞的转录重编程中具有深远的功能。然而，（病理）生理后果的JP 2NT核定位和相关的分子机制是完全未知的。基于这些新的令人兴奋的初步研究结果，我们假设，在应激反应中，钙蛋白酶介导的JP 2裂解将JP 2从结构蛋白转化为转录调节因子，重新编程受损心肌细胞的转录谱并抑制心肌肥大的发展。这种分子事件可能是转录重塑的重要触发因素，在病理条件下充当未被认识到的保护机制。我们将使用多学科方法来测试我们的假设，包括多个新的转基因和基于CRISPR的小鼠模型，分子生物学，生物化学和我们实验室开发的一种新的原位共聚焦成像技术。在目标1中，我们将探索心肌细胞中JP 2NT转录调控的分子机制，而目标2将研究心脏中JP 2NT的病理（生理）后果。我们预计，拟议的研究将为JP 2NT的新功能提供重要的见解 在心脏应激环境中的膜-核信号转导，以及心脏病中心肌细胞结构重塑和转录重塑之间的交叉通讯的新见解。这些信息将作为未来开发新型心力衰竭治疗方法的平台。