Alff Peter J, Gavrilovskaya Irina N, Gorbunova Elena, Endriss Karen, Chong Yuson, Geimonen Erika, Sen Nandini, Reich Nancy C, Mackow Erich R
Molecular and Cellular Biology Graduate Program, SUNY at Stony Brook, Stony Brook, NY 11794, USA.
J Virol. 2006 Oct;80(19):9676-86. doi: 10.1128/JVI.00508-06.
Hantaviruses cause two diseases with prominent vascular permeability defects, hemorrhagic fever with renal syndrome and hantavirus pulmonary syndrome. All hantaviruses infect human endothelial cells, although it is unclear what differentiates pathogenic from nonpathogenic hantaviruses. We observed dramatic differences in interferon-specific transcriptional responses between pathogenic and nonpathogenic hantaviruses at 1 day postinfection, suggesting that hantavirus pathogenesis may in part be determined by viral regulation of cellular interferon responses. In contrast to pathogenic NY-1 virus (NY-1V) and Hantaan virus (HTNV), nonpathogenic Prospect Hill virus (PHV) elicits early interferon responses following infection of human endothelial cells. We determined that PHV replication is blocked in human endothelial cells and that RNA and protein synthesis by PHV, but not NY-1V or HTNV, is inhibited at 2 to 4 days postinfection. The addition of antibodies to beta interferon (IFN-beta) blocked interferon-directed MxA induction by >90% and demonstrated that hantavirus infection induces the secretion of IFN-beta from endothelial cells. Coinfecting endothelial cells with NY-1V and PHV resulted in a 60% decrease in the induction of interferon-responsive MxA transcripts by PHV and further suggested the potential for NY-1V to regulate early IFN responses. Expression of the NY-1V G1 cytoplasmic tail inhibited by >90% RIG-I- and downstream TBK-1-directed transcription from interferon-stimulated response elements or beta-interferon promoters in a dose-dependent manner. In contrast, expression of the NY-1V nucleocapsid or PHV G1 tail had no effect on RIG-I- or TBK-1-directed transcriptional responses. Further, neither the NY-1V nor PHV G1 tails inhibited transcriptional responses directed by a constitutively active form of interferon regulatory factor 3 (IRF-3 5D), and IRF-3 is a direct target of TBK-1 phosphorylation. These findings indicate that the pathogenic NY-1V G1 protein regulates cellular IFN responses upstream of IRF-3 phosphorylation at the level of the TBK-1 complex. These findings further suggest that the G1 cytoplasmic tail contains a virulence element which determines the ability of hantaviruses to bypass innate cellular immune responses and delineates a mechanism for pathogenic hantaviruses to successfully replicate within human endothelial cells.
汉坦病毒可引发两种具有显著血管通透性缺陷的疾病，即肾综合征出血热和汉坦病毒肺综合征。所有汉坦病毒都会感染人类内皮细胞，不过尚不清楚致病汉坦病毒与非致病汉坦病毒之间的差异所在。我们观察到，在感染后1天，致病和非致病汉坦病毒之间的干扰素特异性转录反应存在显著差异，这表明汉坦病毒的发病机制可能部分由病毒对细胞干扰素反应的调控所决定。与致病的纽约1型病毒（NY-1V）和汉滩病毒（HTNV）不同，非致病的普洛透斯山病毒（PHV）在感染人类内皮细胞后会引发早期干扰素反应。我们确定PHV在人类内皮细胞中的复制受阻，且在感染后2至4天，PHV的RNA和蛋白质合成受到抑制，而NY-1V或HTNV则不受影响。添加抗β干扰素（IFN-β）抗体可使干扰素介导的Mx A诱导减少90%以上，并证明汉坦病毒感染可诱导内皮细胞分泌IFN-β。用NY-1V和PHV共同感染内皮细胞，导致PHV诱导的干扰素反应性Mx A转录本减少60%，进一步表明NY-1V具有调控早期干扰素反应的潜力。NY-1V G1细胞质尾巴的表达以剂量依赖的方式抑制了超过90%的由RIG-I和下游TBK-1介导的来自干扰素刺激反应元件或β干扰素启动子的转录。相比之下，NY-1V核衣壳或PHV G1尾巴的表达对RIG-I或TBK-1介导的转录反应没有影响。此外，NY-1V和PHV的G1尾巴均未抑制由组成型活性形式的干扰素调节因子3（IRF-3 5D）介导的转录反应，而IRF-3是TBK-1磷酸化的直接靶点。这些发现表明，致病的NY-1V G1蛋白在TBK-1复合物水平上，在IRF-3磷酸化上游调节细胞干扰素反应。这些发现进一步表明，G1细胞质尾巴包含一个毒力元件，该元件决定了汉坦病毒绕过先天性细胞免疫反应的能力，并阐明了致病汉坦病毒在人类内皮细胞内成功复制的机制。
