Xia Yongyan, Nigos L Rayburn, Villalón-Letelier Fernando, Tessema Melkamu B, Brooks Andrew G, Bartok Eva, Behrendt Rayk, Londrigan Sarah L, Reading Patrick C, Farrukee Rubaiyea
Department of Microbiology and Immunology, The University of Melbourne, at the Peter Doherty Institute for Infection and Immunity, Victoria, Australia.
Institute of Experimental Haematology and Transfusion Medicine, University Hospital, University of Bonn, Bonn, Germany.
J Biol Chem. 2025 Aug 6;301(9):110567. doi: 10.1016/j.jbc.2025.110567.
Host cell restriction factors are intracellular proteins that target and inhibit virus replication. Membrane-associated RING-CH finger (MARCHF) proteins are a family of intracellular E3-ubiquitin ligases and some, including MARCHF8, have been implicated in restricting the replication of diverse RNA viruses. However, little is currently known as to whether MARCHF proteins mediate antiviral activity against DNA viruses. Herein, we used a doxycycline-inducible overexpression system to demonstrate that human MARCHF1 and MARCHF8 potently restrict productive HSV-1 replication and assessed MARCHF8-mediated restriction of HSV-1 in detail. A functional RING-CH domain and a tyrosine-based motif, located in the N- and C-terminal cytoplasmic tail of MARCHF8, respectively, were required for HSV-1 restriction. For many RNA viruses, MARCHF8-mediated restriction has been associated with downregulation of viral envelope glycoproteins from the cell surface, thereby limiting their subsequent incorporation into nascent virions. However, while MARCHF8 expression did not affect virus entry, translocation of viral genome to the nucleus and immediate early (IE) gene expression, it did inhibit HSV-1 genomic replication, and therefore subsequent late gene expression and release of infectious virions. MARCHF8-mediated restriction of HSV-1 occurred independent of other cellular factors known to impact genomic replication of HSV-1, namely SAMHD1 and CD81, and could also proceed efficiently in cells expressing a functional cGAS-STING pathway. To our knowledge, these studies are the first to demonstrate MARCHF8-mediated restriction against a human DNA virus. Moreover, inhibition of HSV-1 genomic replication represents a novel mechanism of MARCHF8-mediated virus restriction that is distinct to its reported antiviral activity against different RNA viruses.
宿主细胞限制因子是靶向并抑制病毒复制的细胞内蛋白质。膜相关的RING-CH指蛋白（MARCHF）是细胞内E3泛素连接酶家族，其中一些成员，包括MARCHF8，已被证明参与限制多种RNA病毒的复制。然而，目前对于MARCHF蛋白是否介导针对DNA病毒的抗病毒活性知之甚少。在此，我们使用强力霉素诱导的过表达系统证明人MARCHF1和MARCHF8能有效限制单纯疱疹病毒1型（HSV-1）的有效复制，并详细评估了MARCHF8介导的对HSV-1的限制作用。HSV-1限制需要分别位于MARCHF8 N端和C端细胞质尾巴中的功能性RING-CH结构域和基于酪氨酸的基序。对于许多RNA病毒，MARCHF8介导的限制与细胞表面病毒包膜糖蛋白的下调有关，从而限制它们随后掺入新生病毒颗粒中。然而，虽然MARCHF8的表达不影响病毒进入、病毒基因组向细胞核的转运以及立即早期（IE）基因的表达，但它确实抑制了HSV-1的基因组复制，因此也抑制了随后的晚期基因表达和感染性病毒颗粒的释放。MARCHF8介导的对HSV-1的限制独立于已知影响HSV-1基因组复制的其他细胞因子，即SAMHD1和CD81，并且在表达功能性cGAS-STING途径的细胞中也能有效进行。据我们所知，这些研究首次证明了MARCHF8介导的对人类DNA病毒的限制作用。此外，抑制HSV-1基因组复制代表了MARCHF8介导的病毒限制的一种新机制，这与其报道的针对不同RNA病毒的抗病毒活性不同。
