IRF3 is Required for G10-Dependent Transcription and Anti-Alphaviral Activity.

(A) Immunoblot showing IRF3, STAT1, and GAPDH in THF-ISRE stably transduced with Cas9 and CRISPR gRNA directed against either STAT1 (THF-ISRE-ΔSTAT1) or IRF3 (THF-ISRE-ΔIRF3) as indicated. (B) Induction of IRF3/IFN-dependent LUC in THF lacking STAT1 following 7h exposure to 100μM G10, UV-inactivated CMV, or 1000U/mL IFNβ. Values displayed are average fold changes ±SD of quadruplicate measurements relative to cells exposed only to 1% DMSO. (C) Induction of IRF3/IFN-dependent LUC in THF lacking IRF3 following 7h exposure to 100μM G10, UV-inactivated CMV, or 1000U/mL IFNβ. Values displayed are as in B. (D) Immunoblot of lysates from THF-ISRE following 6h exposure to DMSO, UV-CMV, SeV or 100uM G10 as indicated showing phosphorylation status of IRF3 S386, total IRF3, and GAPDH. (E) Average media titers +SD of CHIKV, VEEV, and SINV at 24 (VEEV) or 48hpi (CHIKV, SINV) obtained from THF-ISRE-ΔIRF3 cells treated with 1% DMSO, 100μM G10, or 1000U/mL IFNβ as indicated. Infections were performed in triplicate.