Abstract
Bromodomain-containing protein 4 plays a central role in coordinating the complex epigenetic component of the innate immune response. Previous studies implicated BRD4 as a component of a chromatin-modifying complex that is dynamically recruited to a network of protective cytokines by binding activated transcription factors, polymerases, and histones to trigger their rapid expression via transcriptional elongation. Our previous study extended our understanding of the airway epithelial BRD4 interactome by identifying over 100 functionally important coactivators and transcription factors, whose association is induced by respiratory syncytial virus (RSV) infection. RSV is an etiological agent of recurrent respiratory tract infections associated with exacerbations of chronic obstructive pulmonary disease. Using a highly selective small-molecule BRD4 inhibitor (ZL0454) developed by us, we extend these findings to identify the gene regulatory network dependent on BRD4 bromodomain (BD) interactions. Human small airway epithelial cells were infected in the absence or presence of ZL0454, and gene expression profiling was performed. A highly reproducible dataset was obtained which indicated that BRD4 mediates both activation and repression of RSV-inducible gene regulatory networks controlling cytokine expression, interferon (IFN) production, and extracellular matrix remodeling. Index genes of functionally significant clusters were validated independently. We discover that BRD4 regulates the expression of its own gene during the innate immune response. Interestingly, BRD4 activates the expression of NFκB/RelA, a coactivator that binds to BRD4 in a BD-dependent manner. We extend this finding to show that BRD4 also regulates other components of its functional interactome, including the Mediator (Med) coactivator complex and the SWI/SNF-related, matrix-associated, actin-dependent regulator of chromatin (SMARC) subunits. To provide further insight into mechanisms for BRD4 in RSV expression, we mapped 7,845 RSV-inducible Tn5 transposase peaks onto the BRD4-dependent gene bodies. These were located in promoters and introns of cytostructural and extracellular matrix (ECM) formation genes. These data indicate that BRD4 mediates the dynamic response of airway epithelial cells to RNA infection by modulating the expression of its coactivators, controlling the expression of host defense mechanisms and remodeling genes through changes in promoter accessibility.
Highlights
Small-molecule Bromodomain-containing 4 (BRD4) inhibitor, we demonstrate the dynamic role that BRD4 plays in the expression of its interacting coactivators
BRD4 reshapes the protein-interacting complexes formed during the innate response to RSV
Integrating the results of the RNA-Seq and ATAC-Seq data, we identify the subset of BRD4dependent genes that undergo RSV-inducible nucleosomal repositioning, resulting in enhanced transcription factor accessibility
Summary
Bromodomain-containing 4 (BRD4) is a member of the bromodomain and extra-terminal domain (BET) family of proteins that plays pleiotropic roles in epigenetic control of inflammation-inducible gene expression (Nowak et al, 2008; Tian et al, 2016), maintenance of cellular identity (Loven et al, 2013), DNA damage signaling (Floyd et al, 2013), chromatin compaction/conformation (Devaiah et al, 2016a), cell cycle regulation, and so on [Reviewed by Devaiah et al (2016b)].In this study, we focus on the activity of BRD4 in controlling inducible innate antiviral genomic response. Recent studies applying native affinity isolation coupled with high-resolution mass spectrometry have substantially expanded the known protein interactions of BRD4 to include RNA polymerases, coactivators, transcription factors, histone acetyltransferases, and actin motors (Zhang et al, 2017; Mann et al, 2021). In this way, BRD4 forms highly dynamic complexes in a stimulus- and cell-type– dependent manner
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