Figure 6.

RBM10 binds RIG-I and DENV genome and promotes RIG-I ubiquitination. (A) RBM10 and dsRNA (a marker of viral RNA) localization was visualized by indirect immunofluorescence and nuclei-cytoplasmic ratio was quantified. (B) A549 cells were either treated with IFN α or DENV infected and subjected to co-immunoprecipitation assays with anti-RBM10. RNase A (0.1 mg/ml) was added to lysates when indicated. Input and immunoprecipitated fractions were analyzed by western blot with the corresponding antibodies. (C) RNA-immunoprecipitation (RIP) assays with anti-RBM10 of lysates from A549 infected cells at 48 hpi. RT-qPCRs were performed with primers for SAT1 pre-mRNA and viral RNA and represented as RIP/input RNA. Average values from triplicates are shown with standard deviation and p-values, determined using a paired two-tailed t-test. Significant P-values are indicated by the asterisks above the graphs (***P< 0.001; **P< 0.01; *P< 0.05). (D) HEK 293T were transfected with the indicated expression vectors and after 48 h subjected to Ni-NTA affinity purification. Eluates and input fractions were analyzed by western blot with the corresponding antibodies.