African swine fever virus pI73R hijacks host mRNA to suppress interferon beta and reveals a druggable target for antiviral intervention

African swine fever (ASF) is an acute transmissible disease caused by the African swine fever virus (ASFV), which has evolved multiple mechanisms to circumvent the host immune response. This research reveals that ASFV pI73R (I73R protein) downregulates cGAS-STING-mediated interferon beta (IFN-β) activation. ASFV pI73R binds to <i>TBK1</i> and <i>IRF3</i> mRNAs, promoting their nuclear retention and attenuating IFN-β production. Furthermore, silencing <i>I73R</i> enhances <i>IFN-β</i> mRNA levels and increases TBK1 and IRF3 protein levels in primary porcine alveolar macrophages (PAMs) exposed to ASFV. Unlike other reported mechanisms of inhibition of IFN-β by other ASFV proteins, pI73R inhibits IFN-β through its RNA binding function. Using structure-based virtual screening, we identified a small-molecule compound, STL527159, which disrupts the pI73R-RNA interaction, thereby restoring nuclear export and translation of the target <i>TBK1</i> and <i>IRF3</i> mRNAs. This compound markedly reduced ASFV infection <i>in vitro</i>. These findings provide new perspectives on the immune evasion strategies used by ASFV and offer a novel theoretical foundation for the development of antiviral drugs and novel vaccine candidates against ASFV.