Hu Haidai, Dong Jiazhen, Liang Deguang, Gao Zengqiang, Bai Lei, Sun Rui, Hu Hao, Zhang Heng, Dong Yuhui, Lan Ke
Key Laboratory of Molecular Virology and Immunology, Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai, China.
Beijing Synchrotron Radiation Facility, Institute of High Energy Physics, Chinese Academy of Sciences, Beijing, China.
J Virol. 2015 Nov 4;90(3):1158-68. doi: 10.1128/JVI.01392-15. Print 2016 Feb 1.
The oncogenic herpesvirus Kaposi's sarcoma-associated herpesvirus (KSHV) is known to encode four viral interferon regulatory factors (vIRF1 to -4) to subvert the host antiviral immune response, but their detailed DNA-binding profiles as transcription factors in the host remain uncharacterized. Here, we first performed genome-wide vIRF2-binding site mapping in the human genome using chromatin immunoprecipitation coupled with high-throughput sequencing (ChIP-seq). vIRF2 was capable of binding to the promoter regions of 100 putative target genes. Importantly, we confirmed that vIRF2 can specifically interact with the promoters of the genes encoding PIK3C3, HMGCR, and HMGCL, which are associated with autophagosome formation or tumor progression and metastasis, and regulate their transcription in vivo. The crystal structure of the vIRF2 DNA-binding domain (DBD) (referred to here as vIRF2DBD) showed variable loop conformations and positive-charge distributions different from those of vIRF1 and cellular IRFs that are associated with DNA-binding specificities. Structure-based mutagenesis revealed that Arg82 and Arg85 are required for the in vitro DNA-binding activity of vIRF2DBD and can abolish the transcription regulation function of vIRF2 on the promoter reporter activity of PIK3C3, HMGCR, and HMGCL. Collectively, our study provided unique insights into the DNA-binding potency of vIRF2 and suggested that vIRF2 could act as a transcription factor of its target genes in the host antiviral immune response.
The oncogenic herpesvirus KSHV is the etiological agent of Kaposi's sarcoma, primary effusion lymphoma, and multicentric Castleman's disease. KSHV has developed a unique mechanism to subvert the host antiviral immune responses by encoding four homologues of cellular interferon regulatory factors (vIRF1 to -4). However, none of their DNA-binding profiles in the human genome have been characterized until now, and the structural basis for their diverse DNA-binding properties remain poorly understood. In this study, we performed the first genome-wide vIRF2-binding site mapping in the human genome and found vIRF2 can bind to the promoter regions of 100 target cellular genes. X-ray structure analysis and functional studies provided unique insights into its DNA-binding potency and regulation of target gene expression. Our study suggested that vIRF2 could act as a transcription factor of its target genes and contribute to KSHV infection and pathogenesis through versatile functions.
致癌性疱疹病毒卡波西肉瘤相关疱疹病毒（KSHV）已知编码四种病毒干扰素调节因子（vIRF1至-4）以颠覆宿主抗病毒免疫反应，但其作为宿主中转录因子的详细DNA结合谱仍未得到表征。在此，我们首先使用染色质免疫沉淀结合高通量测序（ChIP-seq）在人类基因组中进行全基因组vIRF2结合位点图谱绘制。vIRF2能够结合100个推定靶基因的启动子区域。重要的是，我们证实vIRF2可以与编码PIK3C3、HMGCR和HMGCL的基因的启动子特异性相互作用，这些基因与自噬体形成或肿瘤进展及转移相关，并在体内调节它们的转录。vIRF2 DNA结合结构域（DBD）（此处称为vIRF2DBD）的晶体结构显示出与vIRF1和与DNA结合特异性相关的细胞IRF不同的可变环构象和正电荷分布。基于结构的诱变表明，Arg82和Arg85是vIRF2DBD体外DNA结合活性所必需的，并且可以消除vIRF2对PIK3C3、HMGCR和HMGCL启动子报告基因活性的转录调节功能。总体而言，我们的研究为vIRF2的DNA结合能力提供了独特见解，并表明vIRF2可以在宿主抗病毒免疫反应中作为其靶基因的转录因子发挥作用。
致癌性疱疹病毒KSHV是卡波西肉瘤、原发性渗出性淋巴瘤和多中心Castleman病的病原体。KSHV通过编码细胞干扰素调节因子的四种同源物（vIRF1至-4），开发出一种独特的机制来颠覆宿主抗病毒免疫反应。然而，到目前为止，它们在人类基因组中的DNA结合谱均未得到表征，并且对其多样的DNA结合特性的结构基础仍知之甚少。在本研究中，我们在人类基因组中首次进行了全基因组vIRF2结合位点图谱绘制，发现vIRF2可以结合100个靶细胞基因的启动子区域。X射线结构分析和功能研究为其DNA结合能力和靶基因表达调控提供了独特见解。我们的研究表明，vIRF2可以作为其靶基因的转录因子，并通过多种功能促进KSHV感染和发病机制。
