The primers used in this study.

<div><p>Cell-passage-adapted strains of African swine fever virus (ASFV) typically exhibit substantial genomic alterations and attenuated virulence in pigs. We have indicated that the HEK293T cells-adapted ASFV strain underwent genetic alterations and the <i>I7L</i> gene in the right variable region was deleted compared with the ASFV HLJ/2018 strain (ASFV-WT). A recent study has revealed that the deletion of the <i>I7L</i>-<i>I11L</i> genes results in attenuation of virulent ASFV <i>in vivo</i>, but the underlying mechanism remains largely unknown. Therefore, we hypothesized that the deletion of the <i>I7L</i> gene may be related to the pathogenicity of ASFV in pigs. We generated the <i>I7L</i> gene-deleted ASFV mutant (ASFV-ΔI7L) and found that the <i>I7L</i> gene deletion does not influence the replication of ASFV in primary porcine alveolar macrophages (PAMs). Using transcriptome sequencing analysis, we identified that the differentially expressed genes in the PAMs infected with ASFV-ΔI7L were mainly involved in antiviral immune responses induced by interferon gamma (IFN-<i>γ</i>) compared with those in the ASFV-WT-infected PAMs. Meanwhile, we further confirmed that the I7L protein (pI7L) suppressed the IFN-<i>γ</i>-triggered JAK-STAT signaling pathway. Mechanistically, pI7L interacts with STAT1 and inhibits its phosphorylation and homodimerization, which depends on the tyrosine at position 98 (Y98) of pI7L, thereby preventing the nuclear translocation of STAT1 and leading to the decreased production of IFN-<i>γ</i>-stimulated genes. Importantly, ASFV-ΔI7L exhibited reduced replication and virulence compared with ASFV-WT in pigs, likely due to the increased production of IFN-<i>γ</i>-stimulated genes, indicating that pI7L is involved in the virulence of ASFV. Taken together, our findings demonstrate that pI7L is associated with pathogenicity and antagonizes the IFN-<i>γ</i>-triggered JAK-STAT signaling pathway via inhibiting the phosphorylation and homodimerization of STAT1 depending on the Y98 residue of pI7L and the SH2 domain of STAT1, which provides more information for understanding the immunoevasion strategies and designing the live attenuated vaccines for ASF.</p></div>