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Senecavirus A (SVA), an important emerging porcine virus, has outbreaks in different regions and countries each year, becoming a virus with global prevalence. SVA infection has been reported to induce macroautophagy/autophagy; however, the molecular mechanisms of autophagy induction and the effect of SVA on autophagy remain unknown. This study showed that SVA infection induced the autophagy process in the early stage of SVA infection, and the rapamycin-induced autophagy inhibited SVA replication by degrading virus 3 C protein. To counteract this, SVA utilized 2AB protein inhibiting the autophagy process from promoting viral replication in the late stage of SVA infection. Further study showed that SVA 2AB protein interacted with MARCHF8/MARCH8 and LC3 to degrade the latter and inhibit the autophagy process. In addition, we found that MARCHF8 was a positive regulator of type I IFN (IFN-I) signaling. During the autophagy process, the SVA 2AB protein targeted MARCHF8 and MAVS forming a large complex for degradation to deactivate IFN-I signaling. Together, our study reveals the molecular mechanisms of selective autophagy in the host against viruses and reveals potential viral strategies to evade the autophagic process and IFN-I signaling for successful pathogenesis. Abbreviations: Baf A1: bafilomycin A1; Co-IP: co-immunoprecipitation; CQ: chloroquine; DAPI: 4′,6-diamidino-2-phenylindole; hpi: hours post-infection; IFN: interferon; ISG: IFN-stimulated gene; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MARCHF8/MARCH8: membrane associated ring-CH-type finger 8; MAVS: mitochondrial antiviral signaling protein; MOI: multiplicity of infection; Rapa: rapamycin; RT: room temperature; siRNA: small interfering RNA; SVA: Senecavirus A; TCID50: 50% tissue culture infectious doses. [ABSTRACT FROM AUTHOR]

Interferon-induced BST2 (bone marrow stromal cell antigen 2) inhibits viral replication by tethering enveloped virions to the cell surface to restrict viral release and by inducing the NFKB-dependent antiviral immune response. However, the mechanism by which BST2 uses the selective autophagy pathway to inhibit viral replication is poorly understood. In this study, we showed that BST2 expression was significantly increased during porcine epidemic diarrhea virus (PEDV) infection of Vero cells by IRF1 targeting its promoter. We also showed that BST2 suppressed PEDV replication by binding and degrading the PEDV-encoded nucleocapsid (N) protein. The downregulation of N protein was blocked by macroautophagy/autophagy inhibitors but not a proteasome inhibitor, implying that the N protein was degraded via the selective autophagy pathway. Both the BST2 and N protein interacted with the E3 ubiquitin ligase MARCHF8/MARCH8 and the cargo receptor CALCOCO2/NDP52, and the ubiquitination of N protein was necessary for the degradation of N mediated by the BST2-MARCHF8 axis. The knockdown of MARCHF8 or ATG5 with small interfering RNAs blocked the selective autophagy pathway, rescued the protein abundance of PEDV N in 293T cells, and prevented the inhibition of PEDV replication by BST2 in Vero cells. Together, our data demonstrate the novel mechanism of BST2-mediated virus restriction, in which BST2 recruits MARCHF8 to catalyze the ubiquitination of the PEDV N protein. The ubiquitinated N protein is then recognized by CALCOCO2/NDP52, which delivers it to autolysosome for degradation through the selective autophagy pathway. Abbreviations: 3MA: 3-methyladenine; ATG: autophagy-related; Baf A1: bafilomycin A1; BST2: bone marrow stromal cell antigen 2; CALCOCO2/NDP52: calcium binding and coiled-coil domain 2; CC: coiled-coil; ChIP: chromatin immunoprecipitation; Co-IP: co-immunoprecipitation; CQ: chloroquine; CT: cytoplasmic tail; DAPI: 4′,6-diamidino-2-phenylindole; GPI: glycosyl-phosphatidylinositol; hpi: hours post infection; IRF1: interferon regulatory factor 1; ISG: IFN-stimulated gene; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MARCHF8/MARCH8: membrane-associated ring-CH-type finger 8; MOI: multiplicity of infection; N protein: nucleocapsid protein; PED: porcine epidemic diarrhea; PEDV: porcine epidemic diarrhea virus; RT: room temperature; siRNA: small interfering RNA; STAT: signal transducer and activator of transcription; TCID50: 50% tissue culture infectious doses; TM: transmembrane. [ABSTRACT FROM AUTHOR]