Monkeypox virus (MPXV) is an orthopoxvirus (OPXV) that re-emerged in 2022 causing significant morbidity worldwide. Mpox disease presents with characteristic skin lesions and systemic inflammatory symptoms. Little is known about how MPXV modulates human innate immunity and cytokine responses, although significant evasion is expected from knowledge obtained from other OPXV. In this study I show that MPXV infection of human keratinocytes results in an inflammatory cytokine response defined by elevated IL-6, IL-8, IL-11 and LIF that was not shared with other OPXV. Genetic manipulation revealed that cytokine induction is dependent on nucleic acid sensing and multiple transcription factors. Activation of human fibroblasts with these cytokines did not restrict viral replication despite activating STAT3. Conversely, activation with IL-1β, TNFα and IFNβ resulted in a reduction of viral foci and significant restriction of MPXV and VACV replication. Furthermore, comparative analysis of cytokine responses highlighted that IL-1β and TNFα upregulate a subset of the ISG proteome and restriction by IL-1β and TNFα was partially alleviated by JAK1/2 inhibitor ruxolitinib, indicating ISG-dependent antiviral activity. Further analysis highlighted that IFN restriction was partly mediated by the transmembrane protein family IFITM, consistent with restriction at the level of establishing infection. Spread assays revealed that IFITMs impair VACV spread, mediated by EEV and restrict viral titres of a range of OPXV. Surprisingly, our study determined that IFITM antagonism is a conserved feature of OPXV infection. 

Collectively, this work reveals that MPXV has evolved a distinctive interplay with human innate responses characterized by selective modulation of inflammatory and chemotactic signalling whilst retaining effective suppression of antiviral cytokines. Furthermore, preactivating cells with IFN revealed selective antagonism of restrictive ISG by OPXV, indicating that OPXV interactions with the ISG proteome provide a source of novel antiviral factors to be exploited for IFN-based antivirals.