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: 10210431
• 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 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10210431
• 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; 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; therapeutically effective; 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.

项目概要/摘要 肥厚型心肌病 (HCM) 患者面临心源性猝死和进展性心脏病的风险 衰竭 (HF)，并且没有有效的治疗方法，部分原因是我们对 HCM 的遗传了解有限 发病。我们目前对连接特定分子机制的了解存在严重差距 β 肌球蛋白重链基因 (MYH7) 突变导致心肌病理性增厚，即 与 HCM 相关。我们的长期目标是利用基因组工具结合 3 维心脏 源自人类诱导多能干细胞的微组织用于探究 HCM 继发性机制 特定 MYH7 变体，并利用这些见解来识别新的疾病生物标志物和治疗靶点 针对特定 HCM 患者。 我们之前的研究发现了两种与 HCM 相关的 MYH7 变异，即精氨酸 403 至谷氨酰胺和 缬氨酸 606 变为甲硫氨酸，位于 β 肌球蛋白重链蛋白 (MHC- β），产生增加的微组织收缩力，并伴有收缩动力学异常。 其他人的研究表明，位于 MHC-β 不同结构域的 MYH7 变异导致 不同的表型。这些结果引出了我们的中心假设：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 MYH7 变异的位置 在基因内以及细胞类型特异性转录和表观遗传程序上，这些程序起始于 收缩功能异常。在我们全面的初步数据的指导下，我们建议采取三项行动 具体目标是确定 HCM 发病机制的多尺度见解：(1) 表征功能性 MYH7 变异的后果定位于 MHC 的肌动蛋白结合、ATP 结合和转换器结构域 β, (2) 使用配对鉴定微组织中 HCM 的细胞类型特异性转录和表观遗传机制 单细胞 RNA-seq 和 ATAC-seq 以及 (3) 询问 C1ORF105（核编码的）的功能 与 HCM 相关且在人类心肌细胞中特异性表达的线粒体蛋白。 总之，这些目标的执行将使人们更准确地理解职能作用 MYH7 变异定位，产生新的 HCM 细胞类型特异性和分子机制并识别 肌节和线粒体功能之间的关键分子联系将广泛影响以下领域 HCM 和心力衰竭。

项目成果

Reading Frame Repair of TTN Truncation Variants Restores Titin Quantity and Functions.

• DOI: 10.1161/circulationaha.120.049997
• 发表时间: 2022-01-18
• 期刊: Circulation
• 影响因子: 37.8
• 作者: Romano R;Ghahremani S;Zimmerman T;Legere N;Thakar K;Ladha FA;Pettinato AM;Hinson JT
• 通讯作者: Hinson JT