FIGURE 3.

A3G counteracts the interaction between MOV10 and AGO2. A and B, 293T cells were co-transfected with a plasmid expressing A3G and a plasmid containing the Renilla luciferase reporter gene with perfect/non-perfect binding sites for mir25 or mir16 in the 3′-UTR. This plasmid also contains the firefly luciferase gene as an intraplasmid transfection normalization reporter. pcDNA3.1 and psiCHECK-2 were also transfected as controls. At 48 h post-transfection, Renilla/firefly luciferase activity was measured. C, 293T cells were co-transfected with pcDNA3.1-A3G-HA, psiCHECK-2, and Renilla luciferase-specific siRNA. pcDNA3.1 and gfp-specific siRNA were also transfected as controls. At 48 h post-transfection, Renilla/firefly luciferase activity was measured. Data in A, B, and C represent mean ± S.D. (error bars). **, statistically significant, ≤0.01. D, 293T cells were co-transfected with HA-AGO1- (or HA-AGO2-), A3G-FLAG-, and MOV10-FLAG-expressing plasmids. E, 293T cells were co-transfected with HA-AGO2-, MOV10-FLAG- and different amounts of A3G-expressing plasmids. D and E, transfected-cells were lysed and immunoprecipitated with anti-HA antibody at 24 h post-transfection. The precipitated samples were then analyzed by immunoblotting using anti-FLAG and anti-HA antibodies. Values in D and E represent percentage of MOV10-FLAG normalized against HA-AGO1 (or HA-AGO2) and compared with control.