Protein kinase regulated by dsRNA downregulates the interferon production in dengue virus- and dsRNA-stimulated human lung epithelial cells.
Protein kinase regulated by dsRNA downregulates the interferon production in dengue virus- and dsRNA-stimulated human lung epithelial cells.
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dsRNA 调节的蛋白激酶下调登革热病毒和 dsRNA 刺激的人肺上皮细胞中干扰素的产生
DOI：
10.1371/journal.pone.0055108
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发表时间：
2013
期刊：
影响因子：
3.7
通讯作者：
Huang X
中科院分区：
文献类型：
--
作者：
Li Y;Xie J;Wu S;Xia J;Zhang P;Liu C;Zhang P;Huang X
背景登革热病毒（DENV）在热带和亚热带地区发现，每年影响数百万人。目前，没有针对登革热病毒的特异性疫苗或抗病毒治疗。先天免疫已被证明是重要的宿主抵抗登革病毒感染。虽然已经发现由双链RNA（PKR）调节的蛋白激酶促进响应于由几种病毒感染的先天性信号传导，但是其在响应于DENV感染的先天性反应中的作用仍然不清楚。本研究旨在探讨PKR在DENV诱导的先天性免疫应答中的作用。方法学/主要发现通过RNAi，PKR的沉默显著增强了DENV感染的人肺上皮A549细胞中干扰素（IFN）-β的表达。Western blot和免疫荧光显微镜数据显示，PKR敲低上调了DENV 2感染后先天性信号级联的激活，包括p38和JNK丝裂原活化蛋白激酶（MAPK）、干扰素调节因子-3和NF-κB。同样，在poly（IC）攻击的细胞中也观察到PKR对IFN产生的负调节作用。此外，PKR敲低介导的IFN诱导被RIG-I或IPS-1沉默减弱。最后，过表达无催化活性的PKR突变体（K296 R），但不表达缺乏dsRNA结合活性的突变体（K64 E）或双突变体（K64 EK 296 R），逆转了PKR敲低介导的IFN诱导，表明PKR下调IFN产生需要dsRNA结合活性。结论/意义PKR是DENV和poly（IC）诱导IFN产生的负调节因子，这种调节依赖于其dsRNA结合活性。这些发现揭示了PKR在先天免疫中的新的调节作用，表明PKR可能是抗DENV治疗的有希望的靶点。
Background Dengue virus (DENV) is found in the tropical and subtropical regions and affects millions of people annually. Currently, no specific vaccine or antiviral treatment against dengue virus is available. Innate immunity has been shown to be important for host resistance to DENV infection. Although protein kinase regulated by double-stranded RNA (PKR) has been found to promote the innate signaling in response to infection by several viruses, its role in the innate response to DENV infection is still unclear. Our study aimed to investigate the role of PKR in DENV-induced innate immune responses. Methodology/Principal Findings By RNAi, silencing of PKR significantly enhanced the expression of interferon (IFN)-β in DENV infected human lung epithelial A549 cells. Western blot and immunofluorescence microscopy data showed that PKR knockdown upregulated the activation of innate signaling cascades including p38 and JNK mitogen-activated protein kinases (MAPKs), interferon regulatory factor-3 and NF-κB, following DENV2 infection. Likewise, a negative regulatory effect of PKR on the IFN production was also observed in poly(IC) challenged cells. Moreover, the PKR knockdown-mediated IFN induction was attenuated by RIG-I or IPS-1 silencing. Finally, overexpression of a catalytically inactive PKR mutant (K296R), but not of a mutant lacking dsRNA binding activity (K64E) or the double mutant (K64EK296R), reversed the IFN induction mediated by PKR knockdown, suggesting that the dsRNA binding activity is required for PKR to downregulate IFN production. Conclusions/Significance PKR acts as a negative regulator of IFN induction triggered by DENVs and poly(IC), and this regulation relies on its dsRNA binding activity. These findings reveal a novel regulatory role for PKR in innate immunity, suggesting that PKR might be a promising target for anti-DENV treatments.
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