Pathogenic Role of Selected Cardiac Myocyte- and Fibroblast-Specific Epigenetic Changes in Laminopathies

选定的心肌细胞和成纤维细胞特异性表观遗传变化在核纤层蛋白病中的致病作用

• 批准号: 9119644
• 负责人: Ali J Marian
• 金额: $ 59.41万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-03-15 至 2020-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9119644
• 关键词: ARHGEF5 gene; Adult; Affect; Arrhythmia; Attenuated; CRISPR interference; Cardiac; Cardiac Myocytes; Cardiac development; Cause of Death; Cessation of life; Chromatin; DNA; DNA Methylation; Data; Defect; Dilated Cardiomyopathy; Disease; Embryo; Emerging Technologies; Employee Strikes; Enzymes; Epigenetic Process; Event; Exhibits; Fibroblasts; Fibrosis; Gene Expression; Gene Mutation; Genes; Genetic Transcription; Genomics; HDAC3 gene; Heart; Heart failure; Heterochromatin; Histone Deacetylation; Histones; Human; Injection of therapeutic agent; Knockout Mice; Lamin Type A; Lead; Mammals; Mediating; Membrane Proteins; Methylation; Methyltransferase; Modification; Molecular; Morbidity - disease rate; Mus; Mutation; Myocardial dysfunction; Neonatal; Nuclear; Nuclear Envelope; Organ; Pathogenicity; Phenotype; Precipitation; Prevention; RNA; Role; Staging; Striated Muscles; Structure; Testing; Transgenes; Virus; base; bisulfite sequencing; cell type; chromatin modification; clinical phenotype; coronary fibrosis; differential expression; disease-causing mutation; epigenetic regulation; gene therapy; genome-wide; improved; insight; mammalian genome; methylome; mortality; mutant; novel; premature; prevent; promoter; public health relevance; sudden cardiac death; transcriptome; transcriptomics

 DESCRIPTION (provided by applicant): The objective is to determine the pathogenic role of specific epigenetic modifications in dilated cardiomyopathy (DCM) in laminopathies, which encompass a wide range of diseases caused by mutations in LMNA gene, encoding lamin A/C (LMNA). Cardiac involvement is the primary cause of death in a subset that affects striated muscles. It manifests as DCM, conduction defects, arrhythmias, and sudden cardiac death. Chromatin state regulates gene expression in a cell type-specific manner. Dynamic chromatin modifications are critical for cardiac development and important for adult heart remodeling. In the mammalian genomes, LMNA interacts with chromatin, in a cell-type specific manner, at ~ 1,100 - 1,400 LMNA-associated domains (LADs). LMNA is implicated in H3K9 trimethylation by SUV39H1 and EHMT2 (G9a), histone deacetylation by HDACs and DNA methylation by DNMTs. LADs, mostly located in the heterochromatin loci, are enriched in H3K9me3 and H3K27me3 repressive and to a lesser extent activation histone marks. Preliminary data show marked epigenetic alterations in the heart of 2-week old Lmna-/- mice, preceding cardiac dysfunction. LMNA and several chromatin marks are differentially expressed in neonatal vs. adult cardiac myocytes (CMs) and in CMs vs. cardiac fibroblasts (CFs). Switching on expression of LMNAD300N in neonatal CMs leads to DCM and premature death, while its expression in adult CM induces a mild phenotype, findings in accord with the prominent role of epigenetics in neonatal CMs and parallel the phenotypic effects of Hdac3 deletion. Finally, AAV9 mediated expression of LMNA improves cardiac function and survival. Thus, we posit LMNA has distinct cell-type specific epigenetic functions in neonatal and adult CMs and CFs, which are pathogenic in DCM caused by LMNA mutations. To test this hypothesis, Lmna will be conditionally deleted in neonatal (P3) and adult (P60) CMs and CFs, two common cell types in the heart, using specific Cre deleter mice. Epigenetic basis of both cell type-dependent and -autonomous effects on DCM phenotype will be determined in isolated CMs and CFs by characterizing genome-wide promoter occupancy by specific histone marks, CpG methylation, and the whole transcriptome, prior to and after the onset of cardiac dysfunction and fibrosis (aim 1). In aim 2, epigenetic determinants of differential phenotypic effects upon expression of LMNAD300A, responsible for human DCM, in neonatal and adult CMs will be identified (as in aim 1), along with their induction and reversal upon switching on and off LMNAD300N expression, respectively. In aim 3, prevention and reversal of the epigenetic modifications (as in am 1) and the ensuing DCM will be determined upon AAV9- mediated expression the LMNAWT in the heart of CM-specific and systemic Lmna-/- mice. The findings will provide insight into the role of LMNA in cell type-specific epigenetic regulation of cardiac structure and function, the pathogenic role of the epigenetic modifications in DCM, and set the stage for targeted manipulation of selected epigenetic marks for the precise control, prevention, and reversion of DCM due to LMNA mutations.

 描述（由申请人提供）：目的是确定核纤层蛋白病中扩张型心肌病 (DCM) 中特定表观遗传修饰的致病作用，核纤层蛋白病是由编码核纤层蛋白 A/C (LMNA) 的 LMNA 基因突变引起的多种疾病。受累是影响横纹肌的子集的主要原因，其表现为 DCM、传导缺陷、心律失常和心源性猝死。以细胞类型特异性方式调节基因表达。动态染色质修饰对于心脏发育至关重要，对于成年心脏重塑也很重要。LMNA 以细胞类型特异性方式与染色质相互作用，约为 1,100 - 1,400 LMNA。 -相关结构域 (LAD) 涉及 SUV39H1 和 EHMT2 (G9a) 的 H3K9 三甲基化， HDAC 的组蛋白脱乙酰化和 DNMT 的 DNA 甲基化主要位于异染色质位点，富含 H3K9me3 和 H3K27me3 抑制性组蛋白标记，并在较小程度上激活组蛋白标记，初步数据显示 2 周龄 Lmna 心脏中存在明显的表观遗传改变。 -/- 小鼠，早期的 LMNA 和一些染色质标记在新生儿与成年小鼠中存在差异表达。成人心肌细胞 (CM) 以及 CM 与心脏成纤维细胞 (CF) 的比较 在新生儿 CM 中开启 LMNAD300N 的表达会导致 DCM 和过早死亡，而其在成人 CM 中的表达会诱导轻度表型，这些结果与显着作用一致。新生儿 CM 的表观遗传学作用与 Hdac3 缺失的表型效应相似。最后，AAV9 介导的 LMNA 表达可改善心脏功能和存活率。我们假设 LMNA 在新生儿和成人 CM 和 CF 中具有不同的细胞类型特异性表观遗传功能，这些功能在 LMNA 突变引起的 DCM 中具有致病性。为了检验这一假设，Lmna 将在新生儿 (P3) 和成人 (P60) CM 中被有条件地删除。和 CF，心脏中的两种常见细胞类型，将使用特定的 Cre 删除小鼠来确定对 DCM 表型的细胞类型依赖性和自主效应的表观遗传基础。通过特定组蛋白标记、CpG 甲基化和整个转录组来表征心脏功能障碍和纤维化发生之前和之后的 CM 和 CF（目标 2），差异表型影响的表观遗传决定因素。将鉴定新生儿和成人 CM 中导致人类 DCM 的 LMNAD300A 的表达（如目标 1），以及它们的诱导和逆转分别打开和关闭 LMNAD300N 表达 在目标 3 中，表观遗传修饰（如 am 1）和随后的 DCM 的预防和逆转将取决于 CM 特异性和全身性 Lmna 心脏中 LMNAWT 的表达。 -/- 小鼠。这些发现将深入了解 LMNA 在心脏结构和功能的细胞类型特异性表观遗传调节中的作用、表观遗传修饰在 DCM 中的致病作用，以及为有针对性地操纵选定的表观遗传标记奠定了基础，以精确控制、预防和逆转 LMNA 突变引起的 DCM。