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

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 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.