TBK1 and IKKε protect target cells from IFNgamma-mediated inflammatory apoptosis

T cells produce interferon gamma (IFNγ), but it is unclear whether they can use IFNγ to directly kill target cells. Here, we report that tumor targets lacking TBK1 and IKKε are killed by IFNγ, which induces expression of TNFR1 and the Z-nucleic acid sensor, ZBP1 to trigger RIPK1-dependent apoptosis. Concurrent with apoptosis, IFNγ unexpectedly also induces hyperactivation of NFκB in TBK1 and IKKε double-deficient cells. The two kinases normally function to inhibit IKKa/b activity and in their absence, IFNγ induces elevated expression of inflammatory genes. Thus, IFNγ can induce an inflammatory form of apoptosis that is normally suppressed by TBK1 and IKKε. These kinases regulate three responses: (1) induction of type I IFN; (2) inhibition of RIPK1-dependent death; and (3) inhibition of NFκB-dependent inflammation. We propose that these kinases evolved these intertwined functions so that cells infected by pathogens encoding TBK1/IKKε antagonists to block type I IFN expression will be killed by IFNγ-secreting T cells to generate an alternate NFκB-dependent inflammatory response. To determine if target cells deficient in TBK1 and IKKε are sensitive to IFNγ-mediated cell death, we established B16-F1 melanoma cells with both kinases knocked-out by Crispr-Cas (DKO). B16 cells were also transduced with a non-targeting sgRNA as a control (NTNT). NTNT and DKO B16-F1 cells were treated with either media or IFNγ for 16 and 24 hours. We performed gene expression profiling and gene set enrichment analysis with the data obtained from RNA-seq comparing the 4 sample groups.