Mediator complex in cardiac development and function

心脏发育和功能的介导复合物

• 批准号: 10199010
• 负责人: Xi Fang
• 金额: $ 24.52万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-07-01 至 2023-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10199010
• 关键词: Address; Adult; Affect; Binding; Bioinformatics; Birth; Cardiac; Cardiac Myocytes; Cardiac development; Cardiology; Cardiomyopathies; Cardiovascular Diseases; Career Mobility; ChIP-seq; Complex; Congenital Heart Defects; DNA Binding; DNA Polymerase II; Development; Dilated Cardiomyopathy; Elements; Enhancers; Ensure; Event; Exhibits; Family; Gene Expression; Genes; Genetic Transcription; Genetic study; Goals; Head; Heart; Heart Abnormalities; Heart Diseases; Heart failure; Homeostasis; Human; Hypertrophy; Individual; Knock-out; Knockout Mice; Lead; Light; MED25 gene; Maintenance; Mediator of activation protein; Mentors; Mentorship; Missense Mutation; Modeling; Molecular; Morphogenesis; Morphology; Mus; Mutation; Oranges; Participant; Pathologic; Patients; Phase; Phenotype; Phosphotransferases; Physiological; Physiology; Play; RNA Polymerase II; Regulation; Regulator Genes; Research; Research Project Grants; Role; Shapes; Shprintzen syndrome; Signal Transduction; Structure; Tail; Time; Training; Transcript; Transcription Coactivator; Transcription Initiation; Transcriptional Regulation; cardiogenesis; career; cell type; congenital heart disorder; genomic locus; heart function; insight; mouse genetics; mouse model; novel; programs; promoter; skills; transcription factor; transcriptome sequencing

Mediator (MED) is a multi-subunit complex that plays a central role in transcription initiation by integrating regulatory signals from gene-specific transcriptional activators to RNA polymerase II. Genetic studies have identified mutations in specific MED subunits to be associated with human cardiac diseases. Specifically, deletions in MED15 are common in patients with DiGeorge/velocardiofacial syndrome, which typically includes congenital heart defects. A missense mutation in MED30 causes cardiomyopathy in mice. Emerging evidence suggests a critical role for “specialized” MED subunits in the regulation of temporal- and/or spatial-specific gene expression. However, little is known as to the subunit composition of MED in cardiomyocytes (CMs) at distinct developmental stages, and no studies have examined enrichment of MED complexes harboring individual “specialized” MED subunits at distinct genomic loci, as well as the specific transcription factors (TFs) they directly interact with to anchor them to functionally critical enhancer/promoter elements during heart development, or during the progression of cardiac disease. We found that transcript levels of MED subunits fluctuate within CMs during development. To gain further understanding into MED subunit specific roles in CMs, I generated novel mouse models with cardiac-specific knockout (cKO) of selected MED subunits. MED15, MED25, and MED30 cKO mice showed distinct phenotypes. MED15 cKO mice died immediately after birth, with cardiac defects observed from E12.5, whereas MED30 cKO mice died at E10-10.5 with dilated hearts. Conversely, MED25 cKO mice survived to adulthood and displayed no cardiac defects, demonstrating that not all MED subunits are indispensable for cardiac function. Accordingly, my hypothesis is that MED15 and MED30 subunits exhibit unique functional activities that shape key events in CM transcriptional regulation, morphogenesis, and heart function. My Specific Aims are: Aim 1. (K99) To elucidate stage-specific and locus- specific roles of MED15 or MED30 subunits in developing CMs by analyzing cardiac phenotypes of MED15 and MED30 cKO mice, identifying unique transcriptional regulatory elements enriched for MED15 or MED30 during early heart development, and identifying TFs that specifically interact with MED15 or MED30; and Aim 2. (R00) To understand the pathophysiological functions of MED15 and MED30 subunits during the progression of adult heart disease, including DCM and pathological hypertrophy, by utilizing inducible MED15 and MED30 CM-specific knockout (icKO) mouse models. I have a strong background and training record in mouse genetics and molecular cardiology. My training in the K99 phase will consist of structured mentorship by my primary mentor and complementary co-mentors/consultants, formal coursework, additional training in experimental skills (RNAseq, ChIP-seq and bioinformatics) and a program of career transition. This career plan and research project will ensure my successful transition to independent research, to fulfill my ultimate career goal of understanding the development, progression, and molecular basis of human cardiovascular diseases.

介体（Mediator，MED）是一种多亚基复合物，通过整合在转录起始过程中起重要作用 从基因特异性转录激活因子到RNA聚合酶II的调节信号。遗传学研究 在特定MED亚基中鉴定的突变与人类心脏疾病相关。具体地说， MED 15缺失在DiGeorge/腭心面综合征患者中很常见，通常包括 先天性心脏病MED 30的错义突变导致小鼠心肌病。新出现的证据 表明了“专门的”MED亚基在时间和/或空间特异性的调节中的关键作用。 基因表达。然而，对于心肌细胞（CM）中MED的亚基组成知之甚少， 不同的发展阶段，没有研究检查富集MED复合物， 不同基因组位点的单个“特化”MED亚基，以及特异性转录因子（TF） 它们直接相互作用以将它们锚到心脏过程中功能关键的增强子/启动子元件， 发展，或在心脏病的进展过程中。我们发现MED亚基的转录水平 在发展过程中在CM中波动。为了进一步了解MED亚基在 CMs，我产生了具有选定MED亚基的心脏特异性敲除（cKO）的新型小鼠模型。 MED 15、MED 25和MED 30 cKO小鼠表现出不同的表型。MED 15 cKO小鼠在注射后立即死亡。 出生，从E12.5观察到心脏缺陷，而MED 30 cKO小鼠在E10-10.5死亡， 心中相反，MED 25 cKO小鼠存活至成年，没有显示心脏缺陷，表明 并非所有MED亚基都是心脏功能不可或缺的。因此，我的假设是，MED 15 和MED 30亚基表现出独特的功能活性，形成CM转录调控中的关键事件， 形态发生和心脏功能。我的目标是：目标1。(K99)为了阐明阶段特异性和位点- 通过分析MED 15的心脏表型，探讨MED 15或MED 30亚基在发生CM中的特定作用 和MED 30 cKO小鼠，鉴定富集MED 15或MED 30的独特转录调控元件 在早期心脏发育过程中，并鉴定与MED 15或MED 30特异性相互作用的TF; 2. (R00)了解MED 15和MED 30亚基在糖尿病发病过程中的病理生理功能， 利用诱导型MED 15治疗成人心脏病进展，包括DCM和病理性肥大 和MED 30 CM特异性敲除（icKO）小鼠模型。我在以下领域拥有深厚的背景和培训记录 小鼠遗传学和分子心脏病学。我在K99阶段的培训将包括结构化的指导， 我的主要导师和辅助导师/顾问，正式课程，额外的培训， 实验技能（RNAseq，ChIP-seq和生物信息学）和职业过渡计划。这个职业规划 和研究项目将确保我成功地过渡到独立研究，以实现我的最终职业生涯 目的是了解人类心血管疾病的发生、发展和分子基础。