IFI16 restricts HSV-1 replication by accumulating on the hsv-1 genome, repressing HSV-1 gene expression, and directly or indirectly modulating histone modifications.
IFI16 restricts HSV-1 replication by accumulating on the hsv-1 genome, repressing HSV-1 gene expression, and directly or indirectly modulating histone modifications.
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DOI：
10.1371/journal.ppat.1004503
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发表时间：
2014-11
期刊：
影响因子：
6.7
通讯作者：
Chandran B
中科院分区：
文献类型：
--
作者：
Johnson KE;Bottero V;Flaherty S;Dutta S;Singh VV;Chandran B
干扰素-γ诱导因子16（IFN-γ inducible factor 16，IFI 16）是一种多功能的核蛋白，参与转录调控、干扰素-β（IFN-β）的诱导和炎性反应的激活。它与dsDNA的糖-磷酸骨架相互作用，并通过很大程度上未确定的机制调节病毒和细胞转录。IFI 16是人巨细胞病毒（HCMV）和单纯疱疹病毒（HSV-1）的限制因子，尽管HSV-1限制的机制尚不清楚。在这里，我们表明，IFI 16对HSV-1在人包皮成纤维细胞，骨肉瘤细胞和乳腺上皮癌细胞中的复制有深远的影响。IFI 16敲低使HSV-1产量增加6倍，而IFI 16过表达使病毒产量降低超过5倍。重要的是，在IFI 16的存在下，包括立即早期蛋白ICP 0和ICP 4、早期蛋白ICP 8和TK以及晚期蛋白gB和Us 11在内的HSV-1基因表达减少。消耗炎性小体衔接蛋白，ASC，或IFN诱导转录因子，IRF-3，不影响病毒产量。ChIP研究证实了在骨肉瘤（U2 OS）细胞和成纤维细胞中存在与HSV-1启动子结合的IFI 16。使用CRISPR基因编辑技术，我们产生了IFI 16蛋白表达永久缺失的U2 OS细胞。这些细胞和野生型（wt）U2 OS的ChIP分析表明，在感染后的不同时间，在IFI 16不存在的情况下，RNA聚合酶II、TATA结合蛋白（TBP）和Oct 1转录因子与病毒启动子的关联增加。虽然IFI 16没有改变病毒或细胞启动子的总组蛋白占有率，但它的缺失促进了活性染色质的标记，并减少了具有病毒和细胞基因启动子的抑制性染色质的标记。总的来说，这些研究首次证明了IFI 16阻止重要的转录激活因子与wt HSV-1启动子的结合，并提示了IFI 16限制wt HSV-1复制的潜在机制以及IFI 16在组蛋白修饰中的直接或间接作用。HSV-1是一种普遍存在的人类病原体，可终身感染，已进化出几种机制来逃避宿主免疫检测和应答。然而，它仍然受到细胞因子的调节。最近，宿主核蛋白IFI 16被证明参与对HSV-1感染的先天防御反应。在这里，我们提供的第一个证据表明，IFI 16抑制野生型HSV-1复制抑制病毒基因的表达，独立于其在免疫反应中的作用。我们发现，IFI 16结合HSV-1基因组在几个HSV-1基因的转录起始位点。使用我们产生的永久IFI 16阴性细胞系，我们证明IFI 16减少了重要转录因子的关联。IFI 16还通过增加抑制性染色质的标记物和减少用于激活病毒和细胞基因上的染色质的标记物来促进全局组蛋白修饰。对IFI 16在HSV-1生物学中的作用的这些见解表明，IFI 16的稳定化是抗病毒药物开发的有吸引力的途径。
Interferon-γ inducible factor 16 (IFI16) is a multifunctional nuclear protein involved in transcriptional regulation, induction of interferon-β (IFN-β), and activation of the inflammasome response. It interacts with the sugar-phosphate backbone of dsDNA and modulates viral and cellular transcription through largely undetermined mechanisms. IFI16 is a restriction factor for human cytomegalovirus (HCMV) and herpes simplex virus (HSV-1), though the mechanisms of HSV-1 restriction are not yet understood. Here, we show that IFI16 has a profound effect on HSV-1 replication in human foreskin fibroblasts, osteosarcoma cells, and breast epithelial cancer cells. IFI16 knockdown increased HSV-1 yield 6-fold and IFI16 overexpression reduced viral yield by over 5-fold. Importantly, HSV-1 gene expression, including the immediate early proteins, ICP0 and ICP4, the early proteins, ICP8 and TK, and the late proteins gB and Us11, was reduced in the presence of IFI16. Depletion of the inflammasome adaptor protein, ASC, or the IFN-inducing transcription factor, IRF-3, did not affect viral yield. ChIP studies demonstrated the presence of IFI16 bound to HSV-1 promoters in osteosarcoma (U2OS) cells and fibroblasts. Using CRISPR gene editing technology, we generated U2OS cells with permanent deletion of IFI16 protein expression. ChIP analysis of these cells and wild-type (wt) U2OS demonstrated increased association of RNA polymerase II, TATA binding protein (TBP) and Oct1 transcription factors with viral promoters in the absence of IFI16 at different times post infection. Although IFI16 did not alter the total histone occupancy at viral or cellular promoters, its absence promoted markers of active chromatin and decreased those of repressive chromatin with viral and cellular gene promoters. Collectively, these studies for the first time demonstrate that IFI16 prevents association of important transcriptional activators with wt HSV-1 promoters and suggest potential mechanisms of IFI16 restriction of wt HSV-1 replication and a direct or indirect role for IFI16 in histone modification. HSV-1, a ubiquitous human pathogen that establishes a life-long infection, has evolved several mechanisms to evade host immune detection and responses. However, it is still subject to regulation by cellular factors. Recently, a host nuclear protein, IFI16, was shown to be involved in the innate defense response to HSV-1 infection. Here, we provide the first evidence that IFI16 inhibits wild-type HSV-1 replication by repressing viral gene expression independent of its roles in the immune response. We show that IFI16 binds the HSV-1 genome at the transcription start sites of several HSV-1 genes. Using a permanently IFI16-negative cell line that we generated, we demonstrate that IFI16 reduces the association of important transcription factors. IFI16 also promotes global histone modifications by increasing the markers of repressive chromatin and decreasing the markers for activating chromatin on viral and cellular genes. These insights into the role of IFI16 in HSV-1 biology suggest that stabilization of IFI16 is an attractive avenue for antiviral drug development.
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