Although coronavirus disease 2019 (COVID-19) causes cardiac dysfunction in up to 25% of patients, its pathogenesis remains unclear. Exposure of human induced pluripotent stem cell (iPSC)-derived heart cells to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) revealed productive infection and robust transcriptomic and morphological signatures of damage, particularly in cardiomyocytes. Transcriptomic disruption of structural genes corroborates adverse morphologic features, which included a distinct pattern of myofibrillar fragmentation and nuclear disruption. Human autopsy specimens from patients with COVID-19 reflected similar alterations, particularly sarcomeric fragmentation. These striking cytopathic features in cardiomyocytes provide insights into SARS-CoV-2-induced cardiac damage, offer a platform for discovery of potential therapeutics, and raise concerns about the long-term consequences of COVID-19 in asymptomatic as well as severe cases.
COVID-19 causes cardiac dysfunction in up to 50% of patients, but the pathogenesis remains unclear. Infection of human iPSC-derived cardiomyocytes with SARS-CoV-2 revealed robust transcriptomic and morphological signatures of damage in cardiomyocytes. These morphological signatures include a distinct pattern of sarcomere fragmentation, with specific cleavage of thick filaments, and numerous iPSC-cardiomyocytes that lacked nuclear DNA. Human autopsy specimens from COVID-19 patients also displayed marked sarcomeric disruption and similar fragmentation, as well as prevalently enucleated cardiomyocytes. These striking transcriptomic and cytopathic changes provide a roadmap to understand the mechanisms of COVID-19 cardiac damage, search for potential treatments, and determine the basis for prolonged cardiac morbidity observed in this pandemic.
Mutations in the cardiac splicing factor RBM20 lead to malignant dilated cardiomyopathy (DCM). To understand the mechanism of RBM20-associated DCM, we engineered isogenic iPSCs with DCM-associated missense mutations in RBM20 as well as RBM20 knockout (KO) iPSCs. iPSC-derived engineered heart tissues made from these cell lines recapitulate contractile dysfunction of RBM20-associated DCM and reveal greater dysfunction with missense mutations than KO. Analysis of RBM20 RNA binding by eCLIP reveals a gain-of-function preference of mutant RBM20 for 3′ UTR sequences that are shared with amyotrophic lateral sclerosis (ALS) and processing-body associated RNA binding proteins (FUS, DDX6). Deep RNA sequencing reveals that the RBM20 R636S mutant has unique gene, splicing, polyadenylation and circular RNA defects that differ from RBM20 KO. Super-resolution microscopy verifies that mutant RBM20 maintains very limited nuclear localization potential; rather, the mutant protein associates with cytoplasmic processing bodies (DDX6) under basal conditions, and with stress granules (G3BP1) following acute stress. Taken together, our results highlight a pathogenic mechanism in cardiac disease through splicing-dependent and -independent pathways.
Background:
Gene regulatory networks control tissue homeostasis and disease progression in a cell-type specific manner. Ubiquitously expressed chromatin regulators modulate these networks, yet the mechanisms governing how tissue-specificity of their function is achieved are poorly understood. BRD4, a member of the BET (Bromo- and Extra-Terminal domain) family of ubiquitously expressed acetyl-lysine reader proteins, plays a pivotal role as a coactivator of enhancer signaling across diverse tissue types in both health and disease, and has been implicated as a pharmacologic target in heart failure. However, the cell-specific role of BRD4 in adult cardiomyocytes remains unknown.
Methods:
We combined conditional mouse genetics, unbiased transcriptomic and epigenomic analyses, and classical molecular biology and biochemical approaches to understand the role of BRD4 in adult cardiomyocyte homeostasis.
Results:
Here, we show that cardiomyocyte-specific deletion of
Brd4
in adult mice leads to acute deterioration of cardiac contractile function with mutant animals demonstrating a transcriptomic signature enriched for decreased expression of genes critical for mitochondrial energy production. Genome-wide occupancy data show that BRD4 enriches at many downregulated genes (including the master co-activators
Ppargc1a
,
Ppargc1b
, and their downstream targets) and preferentially co-localizes with GATA4, a lineage determining cardiac transcription factor not previously implicated in regulation of adult cardiac metabolism. BRD4 and GATA4 form an endogenous complex in cardiomyocytes and interact in a bromodomainindependent manner, revealing a new functional interaction partner for BRD4 that can direct its locus and tissue specificity.
Conclusions:
These results highlight a novel role for a BRD4-GATA4 module in cooperative regulation of a cardiomyocyte specific gene program governing bioenergetic homeostasis in the adult heart.
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