USP2a enhances type-II and type-III IFNs-mediated signaling and antiviral efficacy.

(A, B) 293T cells transfected with empty vector or Flag-USP2a were treated with IFNγ (1,000 IU/ml) and IFNλ (50 ng/ml) as indicated. The immunoblotting was performed as indicated. (C) HepG2 cells were transfected with or without HA-USP2a, together with GAS-Luc and Renilla. The luciferase activity was measured 4 hrs after IFNγ (2,000 IU/ml) treatment. (D) HepG2 cells were transfected with or without Flag-USP2a, together with ISRE-Luc and Renilla. The luciferase activity was measured 4 hrs after IFNλ (50 ng/ml) treatment. (E, F) HepG2 cells transfected with shCON or shUSP2a were collected after IFNγ (1,000 IU/ml) or IFNλ (15 ng/ml) treatment for 6 hrs, and the mRNA levels of IFIT1 and Mx1 (E) or ISG15 and ISG54 (F) were analyzed by quantitative RT-PCR. (G) HepG2 cells transfected with empty vector or Flag-USP2a were treated with IFNγ (100 IU/ml) or IFNλ (0.5 ng/ml) overnight, and then cells were challenged by VSV-GFP (MOI = 0.5). After 24 hrs, VSV-GFP levels were detected by fluorescence. (H) KB cells were transfected with or without Flag-USP2a. KB cells were then challenged by VSV as indicated. pY701-STAT1 levels were determined by immunoblotting. (I) KB cells transfected with shCON and shUSP2a were infected with VSV for 2 hrs, the level of VSV viral RNA was analyzed by quantitative RT-PCR. (J) HUVEC cells transfected with shCON and shUSP2a were infected with DENV for 2 hrs, the level of DENV viral RNA was analyzed by quantitative RT-PCR. (K) A549 cells transfected with shCON and shUSP2a were infected with H1N1 for 2 hrs, the level of H1N1 viral RNA was analyzed by quantitative RT-PCR. *p<0.05, **p<0.01, ***p<0.001.