Viruses employ multiple strategies to inhibit host mRNA nuclear export. Distinct to the generally nonselective inhibition mechanisms, ORF10 from gammaherpesviruses inhibits mRNA export in a transcript-selective manner by interacting with Rae1 (RNA export 1) and Nup98 (nucleoporin 98). We now report the structure of ORF10 from MHV-68 (murine gammaherpesvirus 68) bound to the Rae1–Nup98 heterodimer, thereby revealing detailed intermolecular interactions. Structural and functional assays highlight that two highly conserved residues of ORF10, L60 and M413, play critical roles in both complex assembly and mRNA export inhibition. Interestingly, although ORF10 occupies the RNA-binding groove of Rae1–Nup98, the ORF10–Rae1–Nup98 ternary complex still maintains a comparable RNA-binding ability due to the ORF10–RNA direct interaction. Moreover, mutations on the RNA-binding surface of ORF10 disrupt its function of mRNA export inhibition. Our work demonstrates the molecular mechanism of ORF10-mediated selective inhibition and provides insights into the functions of Rae1–Nup98 in regulating host mRNA export.

病毒采用多种策略来抑制宿主mRNA的核输出。与通常的非选择性抑制机制不同，γ疱疹病毒的ORF10通过与Rae1（RNA出口1）和Nup98（核孔蛋白98）相互作用，以转录本选择性的方式抑制mRNA的输出。现在，我们报告从Rav1-Nup98异二聚体结合的MHV-68（鼠丙种疱疹病毒68）ORF10的结构，从而揭示详细的分子间相互作用。结构和功能分析强调，ORF10的两个高度保守的残基L60和M413在复杂装配和mRNA输出抑制中都起着关键作用。有趣的是，尽管ORF10占据Rae1-Nup98的RNA结合沟，但由于ORF10-RNA直接相互作用，ORF10-Rae1-Nup98三元复合物仍保持了相当的RNA结合能力。此外，ORF10的RNA结合表面的突变破坏了其mRNA输出抑制的功能。我们的工作证明了ORF10介导的选择性抑制的分子机制，并提供了对Rae1-Nup98在调节宿主mRNA输出中的功能的见解。