Comprehensive Analysis of Allelic, Cellular and Molecular Heterogeneity in Human 3-Dimensional Cardiac Microtissues with MYH7 Mutations

具有 MYH7 突变的人三维心脏微组织等位基因、细胞和分子异质性的综合分析

• 批准号: 9983170
• 负责人: John Travis Hinson
• 金额: $ 53.99万
• 依托单位: UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-07-01 至 2022-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9983170
• 关键词: 3-Dimensional; ATAC-seq; Actins; Alleles; Architecture; Arginine; Binding; Biological Assay; Biological Markers; Biomimetics; CRISPR/Cas technology; Cardiac; Cardiac Myocytes; Cell Line; Cells; Chromatin; Computing Methodologies; Consumption; Data; Disease; Disease model; Engineering; Epigenetic Process; Future; Gene Expression; Gene Expression Profile; Gene Mutation; Gene Targeting; Genes; Genetic; Genetic Transcription; Genotype-Tissue Expression Project; Glutamine; Goals; Heart failure; Heterogeneity; High-Throughput Nucleotide Sequencing; Human; Human Engineering; Hypertrophic Cardiomyopathy; Image; Immunofluorescence Immunologic; Individual; Inherited; Kinetics; Knock-out; Knowledge; Laboratories; Lead; Link; Location; Mechanics; Metabolic; Methionine; Mitochondria; Mitochondrial Proteins; Modeling; Molecular; Molecular Motors; Mutation; Myocardium; Myofibrils; Myosin Heavy Chains; Nuclear; Pathogenesis; Pathologic; Pathology; Pathway Analysis; Patients; Pattern; Phenotype; Prevalence; Proteins; RNA analysis; Regulatory Element; Research; Resolution; Risk; Role; Sarcomeres; Secondary to; Stress; Structure; Symptoms; Therapeutic; Three-dimensional analysis; Tissues; Transcript; Transposase; Treatment Efficacy; Valine; Variant; beta-Myosin; cell type; cohort; disease heterogeneity; genetic variant; genomic tools; heart dimension/size; heart function; in vivo; induced pluripotent stem cell; insight; new therapeutic target; novel; programs; response; single-cell RNA sequencing; stem cell differentiation; stem cell model; sudden cardiac death; therapeutic development; therapeutic target; transcriptome; transcriptome sequencing; treatment response

PROJECT SUMMARY/ABSTRACT Hypertrophic cardiomyopathy (HCM) patients are at risk for sudden cardiac death and progressive heart failure (HF), and there are no effective therapeutics, due in part to our limited genetic understanding of HCM pathogenesis. There are critical gaps in our current knowledge of the molecular mechanisms that link specific mutations in the beta myosin heavy chain gene (MYH7) to pathological thickening of the heart muscle that is associated with HCM. Our long-term goals are to utilize genomic tools combined with 3-dimensional cardiac microtissues derived from human induced pluripotent stem cells to interrogate mechanisms of HCM secondary to specific MYH7 variants, and to utilize these insights to identify new disease biomarkers and therapeutic targets for specific HCM patients. Our previous studies identified that two HCM-associated MYH7 variants, arginine 403 to glutamine and valine 606 to methionine that are located in the actin-binding domain of beta myosin heavy chain protein (MHC- β), generate increased microtissue contraction force with associated abnormalities in contraction kinetics. Studies by others have indicated that MYH7 variants located in distinct structural domains of MHC-β cause distinct phenotypes. These results lead to our central hypothesis that HCM is a heterogeneous disorder, in which patient symptoms and therapeutic responses are dependent on the location of the causative MYH7 variant(s) within the gene and on cell-type specific transcriptional and epigenetic programs, which initiate from abnormalities in contractile function. Guided by our comprehensive preliminary data, we propose to pursue three Specific Aims to determine multi-scale insights into HCM pathogenesis: (1) to characterize functional consequences of MYH7 variants localized to the actin-binding, ATP-binding and converter domains of MHC- β, (2) to identify cell type-specific transcriptional and epigenetic mechanisms of HCM in microtissues using paired single-cell RNA-seq and ATAC-seq and (3) to interrogate the function of C1ORF105, a nuclear-encoded mitochondrial protein that is associated with HCM and is specifically expressed in human cardiomyocytes. In summary, the execution of these aims will provide a more precise understanding of the functional role of MYH7 variant localization, generate novel cell-type specific and molecular mechanisms of HCM and identify critical molecular linkages between sarcomere and mitochondrial function that will broadly impact the field of HCM and heart failure.

项目总结/摘要 肥厚型心肌病（HCM）患者存在心源性猝死和进行性心脏病的风险。 失败（HF），并且没有有效的治疗方法，部分原因是我们对HCM的遗传理解有限 发病机制目前，我们对特定基因间相互作用的分子机制的认识存在重大空白。 β-肌球蛋白重链基因（MYH 7）突变导致心肌病理性增厚， 与HCM有关。我们的长期目标是利用基因组工具结合三维心脏 来源于人诱导多能干细胞的微组织，以询问HCM继发性机制 特异性MYH 7变体，并利用这些见解来确定新的疾病生物标志物和治疗靶点 针对特定的HCM患者。 我们先前的研究确定了两种HCM相关的MYH 7变体，精氨酸403到谷氨酰胺和 缬氨酸606至甲硫氨酸，其位于β肌球蛋白重链蛋白（MHC-1）的肌动蛋白结合结构域中。 β），产生增加的微组织收缩力，伴随收缩动力学的相关异常。 其他人的研究表明，位于MHC-β不同结构域的MYH 7变异体导致 不同的表型这些结果导致了我们的中心假设，即HCM是一种异质性疾病，其中 患者症状和治疗反应取决于致病MYH 7变体的位置 在基因内和细胞类型特异性转录和表观遗传程序上， 收缩功能异常。根据我们全面的初步数据，我们建议追求三个 具体目的是确定HCM发病机制的多尺度见解：（1）表征功能性 MYH 7变异体定位于MHC的肌动蛋白结合、ATP结合和转换结构域的后果， β，（2）使用配对的基因组DNA， 单细胞RNA-seq和ATAC-seq和（3）询问C1 ORF 105的功能，C1 ORF 105是一个核编码的 一种与HCM相关的线粒体蛋白，在人心肌细胞中特异性表达。 总之，这些目标的执行将提供对职能作用的更准确的理解 MYH 7变体定位，产生HCM的新细胞类型特异性和分子机制，并鉴定 肌节和线粒体功能之间的关键分子联系，将广泛影响 HCM和心力衰竭