Genome-wide RNAi Screen Reveals a New Role of a WNT/CTNNB1 Signaling Pathway as Negative Regulator of Virus-induced Innate Immune Responses
Figure 5
Pharmacological inhibition of GSK3 stabilizes an active CTNNB1 pool and negatively regulates antiviral innate immunity.
(A–B) Relative fold induction of IFNB1-driven reporter activity (A) and fold induction of CTNNB1-TCF/LEF dependent reporter activity TOP-flash (B) in HEK 293T cells transduced with lentivirus-expressing shRNA NT (control) or shRNA 45 targeting CTNNB1 for four days and subjected to GSK3 inhibition with BIO (5 µM) or BIO-acetoxime (10 µM) and SeV infection for 16 hours. (C) Immunoblot analysis of autophosphorylated GSK3α/β at Tyr279/216, dephosphorylated active CTNNB1 at Ser37/Thr41 and IFIT1 in HEK 293T cells treated with GSK3 inhibitors BIO (5 µM) or BIO-acetoxime (10 µM) and infected with SeV for 16 hours. (D) Immunoblot analysis of dephosphorylated active CTNNB1 at Ser37/Thr41, IFIT1 and phosphorylated NFKBIA (IκBα) at Ser32 in SeV-infected HEK 293T cells treated as described in (A–B) and treated with BIO (5 µM) or BIO-acetoxime (10 µM). (E) Relative fold induction of TOP-flash, IFNB1, ISG56, NF-κB and EF1α-driven reporter activity in HEK 293T cells subjected to GSK3 inhibition with BIO (5 µM), TCF/LEF interaction inhibition with PNU74654 (6 µM) or both inhibitors prior to SeV infection for 16 hours.