SARS-CoV-2 ORF6 protein does not antagonize interferon signaling in respiratory epithelial Calu-3 cells during infection.
SARS-CoV-2 ORF6 protein does not antagonize interferon signaling in respiratory epithelial Calu-3 cells during infection.
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严重急性呼吸综合征冠状病毒2（SARS-CoV-2）已造成数百万人死亡，对全球公共卫生构成重大威胁。病毒进化出不同的策略来对抗或逃避宿主的免疫反应。虽然SARS-CoV-2辅助蛋白ORF6的异位表达阻断了干扰素（IFN）的产生和下游IFN信号传导，但ORF6在呼吸道细胞真正病毒感染期间IFN信号传导中的作用尚不清楚。通过比较野生型（WT）和ORF6缺失（Δ ORF6）的SARS-CoV-2感染和呼吸道细胞中的IFN信号，我们发现Δ ORF6 SARS-CoV-2比WT病毒更有效地复制，从而刺激更强大的免疫信号。ORF6的丢失不会改变感染细胞中的先天信号传导：WT和Δ ORF6病毒均仅在旁观者细胞中诱导延迟的IFN应答。此外，在SARS-CoV-2感染的情况下，ORF6的表达对仙台病毒刺激的IFN诱导没有影响：在SARS-CoV-2感染的细胞和旁观者细胞中都观察到了IRF3的稳健易位。此外，IFN预处理类似地有效阻断WT和Δ ORF 6病毒复制，并且两种病毒在IFN-β处理后均不能抑制干扰素刺激基因（ISG）的诱导。然而，在用IFN-β处理后，在用WT病毒感染期间，仅旁观者细胞诱导STAT1易位，而Δ ORF6病毒感染的细胞现在显示易位。这表明在高IFN活化的条件下，ORF6可以减弱STAT1活化。这些数据提供了证据表明，ORF6不足以拮抗SARS-CoV-2感染的呼吸道细胞中的IFN产生或IFN信号传导，但可能影响刺激先天免疫途径的治疗剂的功效。先前的研究确定了几种SARS-CoV-2蛋白，包括ORF6，在非呼吸细胞中病毒蛋白过表达的情况下拮抗宿主先天免疫应答。我们着手确定ORF6在SARS-CoV-2感染呼吸道细胞期间IFN应答中的作用。使用缺失菌株，我们观察到感染没有减少，IFN信号转导的逃避没有差异，反应仅限于旁观者细胞。此外，仙台病毒诱导的IFN产生或IFN-β刺激的ISG表达的刺激在SARS-CoV-2病毒和缺乏ORF6的SARS-CoV-2病毒之间是相当的，这表明ORF6不足以抵消病毒感染期间的IFN诱导或IFN信号传导。
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused millions of deaths, posing a substantial threat to global public health. Viruses evolve different strategies to antagonize or evade host immune responses. While ectopic expression of SARS-CoV-2 accessory protein ORF6 blocks interferon (IFN) production and downstream IFN signaling, the role of ORF6 in IFN signaling during bona fide viral infection of respiratory cells is unclear. By comparing wild-type (WT) and ORF6-deleted (ΔORF6) SARS-CoV-2 infection and IFN signaling in respiratory cells, we found that ΔORF6 SARS-CoV-2 replicates more efficiently than WT virus and, thus, stimulates more robust immune signaling. Loss of ORF6 does not alter innate signaling in infected cells: both WT and ΔORF6 virus induce delayed IFN responses only in bystander cells. Moreover, expression of ORF6 in the context of SARS-CoV-2 infection has no effect on Sendai virus-stimulated IFN induction: robust translocation of IRF3 is observed in both SARS-CoV-2 infected and bystander cells. Furthermore, IFN pretreatment potently blocks WT and ΔORF6 virus replication similarly, and both viruses fail to suppress the induction of interferon-stimulated genes (ISGs) upon IFN-β treatment. However, upon treatment with IFN-β, only bystander cells induce STAT1 translocation during infection with WT virus, whereas ΔORF6 virus-infected cells now show translocation. This suggests that under conditions of high IFN activation, ORF6 can attenuate STAT1 activation. These data provide evidence that ORF6 is not sufficient to antagonize IFN production or IFN signaling in SARS-CoV-2-infected respiratory cells but may impact the efficacy of therapeutics that stimulate innate immune pathways. Previous studies identified several SARS-CoV-2 proteins, including ORF6, that antagonize host innate immune responses in the context of overexpression of viral proteins in non-respiratory cells. We set out to determine the role of ORF6 in IFN responses during SARS-CoV-2 infection of respiratory cells. Using a deletion strain, we observed no reduction of infection and no difference in evasion of IFN signaling, with responses limited to bystander cells. Moreover, stimulation of Sendai virus-induced IFN production or IFN-β-stimulated ISG expression was comparable between SARS-CoV-2 virus and SARS-CoV-2 lacking ORF6 virus, suggesting that ORF6 is not sufficient to counteract IFN induction or IFN signaling during viral infection.
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影响因子：
8.8
作者：
Dittmar M;Lee JS;Whig K;Segrist E;Li M;Kamalia B;Castellana L;Ayyanathan K;Cardenas-Diaz FL;Morrisey EE;Truitt R;Yang W;Jurado K;Samby K;Ramage H;Schultz DC;Cherry S
通讯作者：
Cherry S
影响因子：
10.5
作者：
Iwasaki A
通讯作者：
Iwasaki A
DOI：
10.1074/jbc.ac120.013788
发表时间：
2020-10-09
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
Felgenhauer U;Schoen A;Gad HH;Hartmann R;Schaubmar AR;Failing K;Drosten C;Weber F
通讯作者：
Weber F
影响因子：
8.8
作者：
Kimura I;Konno Y;Uriu K;Hopfensperger K;Sauter D;Nakagawa S;Sato K
通讯作者：
Sato K
影响因子：
5.4
作者：
Kopecky-Bromberg, Sarah A.;Martinez-Sobrido, Luis;Palese, Peter
通讯作者：
Palese, Peter