Figure 2

RLR activation induces proteasomal degradation of the larger MAVS isoform after its polyubiquitination. (A) HeLa cells were infected with SeV H4 in the presence of MG132. At different times after infection, MAVS and its ubiquitination were analyzed in mitochondrial extracts by immunoblotting with a short and a long exposure respectively. (B) HeLa cells were infected with SeV H4 in the presence or the absence of MG132. Nine hours after infection, MAVS was analyzed in cell extracts by immunoblotting. (C) HEK293T cells were infected with SeV H4 in the presence or the absence of MG132, then at various times after infection, RIG-I, MAVS, p-IRF3, IRF3, p-IκBα and IκBα were analyzed in cell extracts by immunoblotting. (D) HeLa cells were infected or not with SeV H4 for 8 hr in the presence or the absence of MG132. Next, nuclear translocation of IRF3 was assessed by immunofluorescence. (E) HEK293T cells were transfected with either an IFN-β promoter reporter or with NF-κB reporter as well as with renilla luciferase as an internal control. 24 hr after transfection, cells were infected with SeV WT or SeV H4 or else left non-infected (-) and treated with different proteasome inhibitors. Luciferase assay was performed 8 hr after infection and was normalized using renilla luciferase activity. Data represent means ± SD (n = 3).