RNA viruses that replicate in the cytoplasm often disrupt nucleocytoplasmic transport to preferentially translate their own transcripts and prevent host antiviral responses. The Sarbecovirus accessory protein ORF6 has previously been shown to be the major inhibitor of interferon production in both SARS-CoV and SARS-CoV-2. SARS-CoV-2 ORF6 was recently shown to co-purify with the host mRNA export factors Rae1 and Nup98. Here, we demonstrate SARS-CoV-2 ORF6 strongly represses protein expression of co-transfected reporter constructs and imprisons host mRNA in the nucleus, which is associated with its ability to co-purify with Rae1 and Nup98. These protein-protein interactions map to the C-terminus of ORF6 and can be abolished by a single amino acid mutation in Met58. Overexpression of Rae1 restores reporter expression in the presence of SARS-CoV-2 ORF6. We further identify an ORF6 mutant containing a 9-amino acid deletion, ORF6 ∆22-30, in multiple SARS-CoV-2 clinical isolates that can still downregulate the expression of a co-transfected reporter and interact with Rae1 and Nup98. SARS-CoV ORF6 also interacts with Rae1 and Nup98. However, SARS-CoV-2 ORF6 more strongly co-purifies with Rae1 and Nup98 and results in significantly reduced expression of reporter proteins compared to SARS-CoV ORF6, a potential mechanism for the delayed symptom onset and pre-symptomatic transmission uniquely associated with the SARS-CoV-2 pandemic.

中文翻译：

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SARS-CoV-2 ORF6通过与Rae1和Nup98相互作用来破坏核质运输

在细胞质中复制的RNA病毒通常会破坏核质运输，从而优先翻译其自身的转录本并阻止宿主抗病毒反应。先前已证明Sarbecovirus辅助蛋白ORF6是SARS-CoV和SARS-CoV-2中干扰素产生的主要抑制剂。最近显示，SARS-CoV-2 ORF6与宿主mRNA输出因子Rae1和Nup98共纯化。在这里，我们证明了SARS-CoV-2 ORF6强烈抑制共转染的报道分子构建体的蛋白质表达，并监禁细胞核中的宿主mRNA，这与其与Rae1和Nup98共纯化的能力有关。这些蛋白质-蛋白质相互作用映射到ORF6的C末端，并且可以被Met58中的单个氨基酸突变所消除。在SARS-CoV-2 ORF6存在下，Rae1的过表达恢复了报告基因的表达。我们进一步在多个SARS-CoV-2临床分离株中鉴定出一个含有9个氨基酸缺失的ORF6突变体，ORF6Δ22-30，该突变体仍可以下调共转染的报告基因的表达并与Rae1和Nup98相互作用。SARS-CoV ORF6也与Rae1和Nup98相互作用。然而，SARS-CoV-2 ORF6与Rae1和Nup98更强烈地共纯化，与SARS-CoV ORF6相比，导致报告蛋白的表达显着降低，这是延迟症状发作和症状发生前传播的唯一潜在机制。 SARS-CoV-2大流行。在多个SARS-CoV-2临床分离株中仍然可以下调共同转染的报告基因的表达并与Rae1和Nup98相互作用。SARS-CoV ORF6也与Rae1和Nup98相互作用。然而，SARS-CoV-2 ORF6与Rae1和Nup98更强烈地共纯化，与SARS-CoV ORF6相比，导致报告蛋白的表达显着降低，这是延迟症状发作和症状发生前传播的唯一潜在机制。 SARS-CoV-2大流行。在多个SARS-CoV-2临床分离株中仍可下调共转染报告基因的表达并与Rae1和Nup98相互作用。SARS-CoV ORF6也与Rae1和Nup98相互作用。然而，SARS-CoV-2 ORF6与Rae1和Nup98更强烈地共纯化，与SARS-CoV ORF6相比，导致报告蛋白的表达显着降低，这是延迟症状发作和症状发生前传播的唯一潜在机制。 SARS-CoV-2大流行。