Tian Bing, Yang Jun, Zhao Yingxin, Ivanciuc Teodora, Sun Hong, Garofalo Roberto P, Brasier Allan R
Department of Internal Medicine, University of Texas Medical Branch, Galveston, Texas, USA.
Sealy Center for Molecular Medicine, University of Texas Medical Branch, Galveston, Texas, USA.
J Virol. 2017 Feb 28;91(6). doi: 10.1128/JVI.00007-17. Print 2017 Mar 15.
The airway mucosa expresses protective interferon (IFN) and inflammatory cytokines in response to respiratory syncytial virus (RSV) infection. In this study, we examine the role of bromodomain containing 4 (BRD4) in mediating this innate immune response in human small airway epithelial cells. We observe that RSV induces BRD4 to complex with NF-κB/RelA. BRD4 is functionally required for expression of the NF-κB-dependent inflammatory gene regulatory network (GRN), including the IFN response factor 1 (IRF1) and IRF7, which mediate a cross talk pathway for RIG-I upregulation. Mechanistically, BRD4 is required for cyclin-dependent kinase 9 (CDK9) recruitment and phospho-Ser 2 carboxy-terminal domain (CTD) RNA polymerase (Pol) II formation on the promoters of , , and , producing their enhanced expression by transcriptional elongation. We also find that BRD4 independently regulates CDK9/phospho-Ser 2 CTD RNA Pol II recruitment to the IRF3-dependent IFN-stimulated genes (ISGs). , poly(I·C)-induced neutrophilia and mucosal chemokine production are blocked by a small-molecule BRD4 bromodomain inhibitor. Similarly, BRD4 inhibition reduces RSV-induced neutrophilia, mucosal CXC chemokine expression, activation of the IRF7-RIG-I autoamplification loop, mucosal IFN expression, and airway obstruction. RSV infection activates BRD4 acetyltransferase activity on histone H3 Lys (K) 122, demonstrating that RSV infection activates BRD4 These data validate BRD4 as a major effector of RSV-induced inflammation and disease. BRD4 is required for coupling NF-κB to expression of inflammatory genes and the IRF-RIG-I autoamplification pathway and independently facilitates antiviral ISG expression. BRD4 inhibition may be a strategy to reduce exuberant virus-induced mucosal airway inflammation. In the United States, 2.1 million children annually require medical attention for RSV infections. A first line of defense is the expression of the innate gene network by infected epithelial cells. Expression of the innate response requires the recruitment of transcriptional elongation factors to rapidly induce innate response genes through an unknown mechanism. We discovered that RSV infection induces a complex of bromodomain containing 4 (BRD4) with NF-κB and cyclin-dependent kinase 9 (CDK9). BRD4 is required for stable CDK9 binding, phospho-Ser 2 RNA Pol II formation, and histone acetyltransferase activity. Inhibition of BRD4 blocks Toll-like receptor 3 (TLR3)-dependent neutrophilia and RSV-induced inflammation, demonstrating its importance in the mucosal innate response Our study shows that BRD4 plays a central role in inflammation and activation of the IRF7-RIG-I amplification loop vital for mucosal interferon expression. BRD4 inhibition may be a strategy for modulating exuberant mucosal airway inflammation.
气道黏膜在对呼吸道合胞病毒（RSV）感染作出反应时会表达保护性干扰素（IFN）和炎性细胞因子。在本研究中，我们检测了含溴结构域4（BRD4）在介导人小气道上皮细胞这种先天性免疫反应中的作用。我们观察到RSV诱导BRD4与NF-κB/RelA形成复合物。BRD4在功能上是NF-κB依赖性炎性基因调控网络（GRN）表达所必需的，该网络包括干扰素反应因子1（IRF1）和IRF7，它们介导了RIG-I上调的串扰途径。从机制上讲，BRD4是细胞周期蛋白依赖性激酶9（CDK9）募集以及在、和启动子上磷酸化丝氨酸2羧基末端结构域（CTD）RNA聚合酶（Pol）II形成所必需的，通过转录延伸产生它们的增强表达。我们还发现BRD4独立调节CDK9/磷酸化丝氨酸2 CTD RNA Pol II募集至IRF3依赖性干扰素刺激基因（ISG）。，小分子BRD4溴结构域抑制剂可阻断聚肌苷酸-聚胞苷酸（poly(I·C)）诱导的嗜中性粒细胞增多和黏膜趋化因子产生。同样，抑制BRD4可减少RSV诱导的嗜中性粒细胞增多、黏膜CXC趋化因子表达、IRF7-RIG-I自动放大环的激活、黏膜IFN表达和气道阻塞。RSV感染激活了组蛋白H3赖氨酸（K）122上的BRD乙酰转移酶活性，表明RSV感染激活了BRD4。这些数据证实BRD4是RSV诱导的炎症和疾病的主要效应因子。BRD4是将NF-κB与炎性基因表达以及IRF-RIG-I自动放大途径偶联所必需的，并且独立促进抗病毒ISG表达。抑制BRD4可能是减少过度的病毒诱导的黏膜气道炎症的一种策略。在美国，每年有210万儿童因RSV感染需要医疗护理。第一道防线是受感染上皮细胞表达先天性基因网络。先天性反应的表达需要募集转录延伸因子，通过未知机制快速诱导先天性反应基因。我们发现RSV感染诱导含溴结构域4（BRD4）与NF-κB和细胞周期蛋白依赖性激酶9（CDK9）形成复合物。BRD4是稳定的CDK9结合、磷酸化丝氨酸2 RNA Pol II形成和组蛋白乙酰转移酶活性所必需的。抑制BRD4可阻断Toll样受体3（TLR3）依赖性嗜中性粒细胞增多和RSV诱导的炎症，证明其在黏膜先天性反应中的重要性。我们的研究表明BRD4在炎症以及对黏膜干扰素表达至关重要的IRF7-RIG-I放大环的激活中起核心作用。抑制BRD4可能是调节过度的黏膜气道炎症的一种策略。
