LncRNA mechanism of heart failure

LncRNA心力衰竭机制

• 批准号: 9246562
• 负责人: CHING-PIN CHANG
• 金额: $ 40.22万
• 依托单位: INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-04-01 至 2019-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9246562
• 关键词: Adult; Alternative Splicing; Animals; Antisense RNA; Biochemical; Biological Assay; Cardiac; Cardiac Myocytes; Cardiomyopathies; Cell Culture Techniques; Chemicals; Chromatin; Chromatin Remodeling Factor; Code; Complex; Data; Electrophoretic Mobility Shift Assay; Episome; FDA approved; Failure; Feedback; Fetal Heart; Fibrosis; Foundations; Gene Expression; Gene Expression Regulation; Gene Targeting; Genes; Genetic; Genetic Models; Genetic Transcription; Heart; Heart Diseases; Heart Hypertrophy; Heart failure; Homeostasis; Human; Hypertrophy; Immunoprecipitation; Knock-out; Methods; Modeling; Molecular; Morbidity - disease rate; Mus; Myocardium; Myopathy; Myosin Heavy Chains; Neonatal; Nucleosomes; Nucleotides; Outcome; Pathogenesis; Pathologic; Pharmacotherapy; Proteins; RNA; Regulation; Reporter; Repression; Role; SMARCA4 gene; Societies; Stress; Switch Genes; Technology; Tissues; Transcript; Transgenic Organisms; Treatment Failure; Untranslated RNA; Western Blotting; Yeasts; arm; base; cardiogenesis; chromatin immunoprecipitation; design; epigenetic regulation; heart cell; heart function; human disease; in vivo; induced pluripotent stem cell; insight; mortality; pressure; prevent; programs; promoter; protein complex; public health relevance; restoration; success

DESCRIPTION (provided by applicant): Pathological stress induces transcriptional gene reprogramming in the heart muscle, leading to cardiac myopathy and heart failure. Such stress-induced gene reprogramming requires regulation at the chromatin level by chromatin-regulating factors. The activity of chromatin regulators can be modulated by an emerging class of RNAs-the long noncoding RNAs (lncRNAs), which are long RNA transcripts with low/no protein-coding potential. The role of lncRNAs in hypertrophy and heart failure, however, is essentially unknown. Most studies of lncRNAs are conducted in cell cultures or in yeast, and mouse genetic models are lacking. This program will focus on a new mouse genetic model to define the molecular function of a newly identified lncRNA in controlling cardiac gene expression, hypertrophy, and failure. Because RNAs can be chemically modified and delivered as a drug for therapy, the success of this program will lay down a foundation for designing new mechanism-based therapy for heart failure. Aim 1: Defining the in vivo regulation of an lncRNA in the heart. We will use transgenic and knockout technology of mouse genetics to define the genetic and molecular mechanisms by which this lncRNA expression is regulated in the heart. Methods also include immunostaining, western blot, quantitative PCR, chromatin immunoprecipitation (ChIP), and RNA immunoprecipitation (RNA-IP) analyses. Aim 2: Determining how this lncRNA interacts with chromatin-regulating factor in the heart. We will use molecular, cellular, and biochemical methods to determine the molecular mechanism by which the lncRNA controls the molecular functions of chromatin regulators in the hearts. The methods include RNA-IP, ChIP, quantitative PCR, electric mobility shift assays, episome-based reporter assays, and nucleosome assembly. Aim 3: Defining the function of human lncRNA-protein complex. We will use human heart tissues and iPS-derived heart cells to demonstrate the presence of lncRNA complex. Also, we will use molecular, cellular, and biochemical methods to define the function of human lncRNA complex. The methods include RNA-IP, ChIP analysis of small amount of tissues, quantitative PCR, electric mobility shift assays, episome-based reporter assays, and iPS-based technology.

描述(申请人提供)：病理性应激诱导心肌转录基因重编程，导致心肌病和心力衰竭。这种应激诱导的基因重编程需要染色质调节因子在染色质水平上进行调节。染色质调节剂的活性可以被一类新兴的RNA--长非编码RNAs(Long Non Coding RNAs，LncRNAs)调节，LncRNAs是一种具有低/无蛋白质编码潜力的长RNA转录体。然而，lncRNAs在肥厚和心力衰竭中的作用基本上是未知的。大多数对lncRNAs的研究都是在细胞培养或酵母中进行的，缺乏小鼠遗传模型。这个项目将集中在一个新的小鼠遗传模型上，以确定新发现的lncRNA在控制心脏基因表达、肥大和衰竭方面的分子功能。由于RNA可以被化学修饰并作为药物用于治疗，该计划的成功将为设计新的基于机制的心力衰竭治疗奠定基础。目的1：明确在心脏中对lncRNA的体内调控。我们将使用小鼠遗传学的转基因和基因敲除技术来确定这种lncRNA在心脏中表达的遗传和分子机制。方法还包括免疫染色、免疫印迹、定量PCR、染色质免疫沉淀(CHIP)和RNA免疫沉淀(RNA-IP)分析。目的2：确定该lncRNA如何与心脏中的染色质调节因子相互作用。我们将使用分子、细胞和生化方法来确定lncRNA控制心脏染色质调节分子功能的分子机制。这些方法包括RNA-IP、芯片、定量聚合酶链式反应、电子迁移率移位分析、基于Episome的报告分子分析和核小体组装。目的3：明确人lncRNA-蛋白质复合体的功能。我们将使用人类心脏组织和iPS来源的心脏细胞来证明lncRNA复合体的存在。此外，我们还将使用分子、细胞和生化方法来定义人类lncRNA复合体的功能。这些方法包括RNA-IP、少量组织的芯片分析、定量聚合酶链式反应、电迁移率分析、基于Episome的报告分析和基于iPS的技术。