African swine fever (ASF) is a devastating infectious disease caused by African swine fever virus (ASFV). The ASFV genome encodes multiple structural and non-structural proteins that contribute to evasion of host immunity. In this study, we determined that the viral non-structural protein MGF360-14L inhibits interferon-β (IFN-β) promoter activity induced by cGAS-STING signaling. MGF360-14L was also found to downregulate expression of the IRF3 protein and promote its degradation through ubiquitin-meditated proteolysis. Moreover, MGF360-14L was shown to interact with and destabilize IRF3 by facilitating E3 ligase TRIM21-mediated K63-linked ubiquitination of IRF3. Overall, our study revealed that MGF360-14L promotes degradation of IRF3 through TRIM21, thereby inhibiting type I interferon production. These findings provide new insights into the mechanisms underlying ASFV immune evasion.

Cyclic GMP-AMP synthase (cGAS) is a major DNA sensor. The recognition of cytosolic DNA by cGAS triggers a robust innate immune response that restricts the replication of diverse viral pathogens through the type I interferon (IFN) and nuclear factor-κB (NF-κB) pathways. African swine fever virus (ASFV) is a large and complex DNA virus reported to strongly inhibit the cGAS-STING signaling pathway. Herein, 12 ASFV structural proteins were screened to determine their effects on the cGAS-STING pathway. Ectopic expression of the ASFV caspid protein M1249L significantly inhibited the IFN-β promoter activity induced by the cGAS-STING pathway in a dose-dependent manner. And it could also downregulate the levels of IFN-β and several interferon-stimulating genes (ISGs) induced by cGAS-STING and 2'3'-cGAMP. Moreover, ASFV M1249L also suppressed phosphorylation of TBK1 by cGAS and STING overexpression. Further study showed that M1249L co-localized and interacted with interferon regulatory factor 3 (IRF3), which led to induce IRF3 degradation by lysosomal pathway. Taken together, our study revealed a novel strategy utilized by ASFV for cGAS-STING-related immune evasion.

African swine fever (ASF) is a devastating infectious disease in domestic pigs caused by African swine fever virus (ASFV) with a mortality rate of about 100%. However, the understanding of the interaction between ASFV and host is still not clear. In this study, the expression differences and functional analysis of microRNA (miRNA) in porcine peripheral blood lymphocytes of ASFV infected pigs and healthy pigs were compared based on Illumina high-throughput sequencing, then the GO and KEGG signal pathways were analyzed. The miRNA related to immunity and inflammation were screened, and the regulatory network of miRNA-mRNA was drawn. A total of 70 differentially expressed miRNAs were found (p ≤ 0.05). Of these, 45 were upregulated and 25 were downregulated in ASFV-infected pigs vs. healthy pigs. A total of 8179 mRNA genes targeted by these 70 differentially expressed miRNA were predicted, of which 1447 mRNA genes were targeted by ssc-miR-2320-5p. Five differentially expressed miRNA were validated by RT-qPCR, which were consistent with the RNA-Seq results. The GO analysis revealed that a total of 30 gene functions were significantly enriched, including 7 molecular functions (MF), 13 cellular components (CC), and 10 biological processes (BP). The KEGG enrichment analysis revealed that the differentially expressed genes were significantly enriched in pathways related to immunity, inflammation, and various metabolic processes, in which a total of two downregulated miRNAs after infection and eight upregulated miRNAs related to immunity and inflammation were screened in ASFV-infected pigs vs. healthy pigs. The network of miRNA-mRNA showed that the mRNA target genes were strongly regulated by ssc-miR-214, ssc-miR-199b-3p, and ssc-miR-199a-3p. The mRNA target genes were enriched into the MAPK signaling pathway, Toll-like receptor signaling pathway, TNF signaling pathway, and IL-17 signaling pathway by using a KEGG enrichment analysis. Therefore, ASFV could regulate immunity and metabolism-related pathways in infected pigs by inducing differential expression of miRNAs. These results provided a new basis for further elucidating the interactions between ASFV and the host as well as the immunity regulation mechanisms of ASFV, which will be conducive to better controlling ASF.