Angioplasty-induced epigenomic remodeling entails BRD4 and EZH2 hierarchical regulations

Özer

Molecular Therapy - Methods & Clinical Development

et al. 2021

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Bromodomain protein BRD4 reads histone acetylation (H3K27ac), an epigenomic mark of transcription enhancers. CCAAT enhancer binding protein delta (CEBPD) is a transcription factor typically studied in metabolism. While both are potent effectors and potential therapeutic targets, their relationship was previously unknown. Here we investigated their interplay in vascular smooth muscle cell (SMC) inflammation. Chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) revealed H3K27ac/BRD4 enrichment at Cebpd in injured rat carotid arteries. While genomic deletion of BRD4-associated enhancer in SMCs in vitro decreased Cebpd transcripts, BRD4 gene silencing also diminished Cebpd mRNA and protein, indicative of a BRD4 control over CEBPD expression. Bromodomain-1, but not bromodomain-2, accounted for this BRD4 function. Moreover, endogenous BRD4 protein co-immunoprecipitated with CEBPD, and both proteins co-immunoprecipitated the Cebpd promoter and enhancer DNA fragments. These co-immunoprecipitations (coIPs) were all abolished by the BRD4-bromodomain blocker JQ1, suggesting a BRD4/CEBPD /promoter/enhancer complex. While BRD4 and CEBPD were both upregulated upon tumor necrosis factor alpha (TNF-α) stimulation of SMC inflammation (increased interleukin [IL]-1b, IL-6, and MCP-1), they mediated this stimulation via preferentially elevated expression of platelet-derived growth factor receptor alpha (PDGFRα, versus PDGFRβ), as indicated by loss- and gain-of-function experiments. Taken together, our study unravels a hierarchical yet collaborative BRD4/CEBPD relationship, a previously unrecognized mechanism that prompts SMC inflammation and may underlie other pathophysiological processes as well.

Section: Introductionmentioning

confidence: 99%

A hierarchical and collaborative BRD4/CEBPD partnership governs vascular smooth muscle cell inflammation

Özer

Molecular Therapy - Methods & Clinical Development

et al. 2021

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“…Arteries were collected for ChIPseq at post-angioplasty day 7 as this is the peak time of pro-IH molecular and cellular events 14,15 . Epigenomic marks involved in transcriptional activation, including BRD4, H3K27ac, and H3K4me1 16 , were chosen for ChIP experiments, and the main body of data is available in our other report 17 . BRD4 and H3K27ac are associated with active enhancers, and H3K4me1 can be found with active, inactive, or poised enhancers.…”

Section: Angioplasty Injury Induces Brd4 and H3k27ac Enrichment At Thmentioning

confidence: 99%

Partnership between epigenetic reader BRD4 and transcription factor CEBPD

Zhang

Urabe³

et al. 2020

Preprint

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Vascular smooth muscle cell (SMC) state/phenotype transitions underlie neointimal hyperplasia (IH) predisposing to cardiovascular diseases. Bromodomain protein BRD4 is a histone acetylation reader and enhancer mark that co-activates transcription elongation. CCAAT enhancer binding protein delta (CEBPD) is a transcription factor typically studied in adipogenesis and immune cell differentiation. Here we investigated the association between BRD4 and CEBPD in SMC state transition.Chromatin immunoprecipitation sequencing (ChIPseq) showed enrichment of BRD4 and histone acetylation (H3K27ac) at Cebpd and enhancer in rat carotid arteries undergoing IH. In vitro, BRD4 silencing with siRNA reduced SMC expression of CEBPD. Bromodomain-1 but not bromodoamin-2 accounted for this BRD4 function. Endogenous BRD4 co-IP'ed with CEBPD; Cebpd promoter and enhancer DNA fragments co-IP'ed with CEBPD or endogenous BRD4 (ChIP-qPCR). These co-IPs were abolished by the BRD4 bromodomain blocker JQ1. TNFa upregulated both BRD4 and CEBPD. Silencing CEBPD averted TNFa-induced inflammatory SMC state transition (heightened IL-1b, IL6, and MCP-1 mRNA levels), so did JQ1. CEBPD overexpression increased PDGFRa preferentially over PDGFRb; so did TNFa, and JQ1 abolished TNFa's effect.Our data reveal a BRD4/CEBPD partnership that promotes CEBPD's own transcription and inflammatory SMC state transition, thus shedding new light on epigenetic reader and transcription factor cooperative actions in SMC pathobiology.

Angioplasty induces epigenomic remodeling in injured arteries

et al. 2022

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Neointimal hyperplasia/proliferation (IH) is the primary etiology of vascular stenosis. Epigenomic studies concerning IH have been largely confined to in vitro models, and IH-underlying epigenetic mechanisms remain poorly understood. This study integrates information from in vivo epigenomic mapping, conditional knockout, gene transfer and pharmacology in rodent models of IH. The data from injured (IH-prone) rat arteries revealed a surge of genome-wide occupancy by histone-3 lysine-27 trimethylation (H3K27me3), a gene-repression mark. This was unexpected in the traditional view of prevailing post-injury gene activation rather than repression. Further analysis illustrated a shift of H3K27me3 enrichment to anti-proliferative genes, from pro-proliferative genes where gene-activation mark H3K27ac(acetylation) accumulated instead. H3K27ac and its reader BRD4 (bromodomain protein) co-enriched at Ezh2; conditional BRD4 knockout in injured mouse arteries reduced H3K27me3 and its writer EZH2, which positively regulated another pro-IH chromatin modulator UHRF1. Thus, results uncover injury-induced loci-specific H3K27me3 redistribution in the epigenomic landscape entailing BRD4→EZH2→UHRF1 hierarchical regulations. Given that these players are pharmaceutical targets, further research may help improve treatments of IH.