Figure 7

At least one A3B domain with catalytic site integrity is necessary for ZfL2-2 inhibition. HeLa cells were co-transfected with 1 μg of target ZfL2-2 plasmid and (A) 1 μg of effector plasmid coding for the wild-type A3B protein or its mutants, or (B) 2, 0.5 and 0.01 μg of plasmids encoding the A3B protein and its C-terminal deletion mutant, NA3B. Relative retrotransposition efficiency was determined by counting the fixed and stained neomycin resistant colonies, formed after 12 days of G418 selection, and calculated by setting the value obtained for control cells co-transfected with retrotransposon plasmid and an empty vector at 100%. The reported cytidine deaminase activity (CDA), or lack thereof, of each protein is designated with + or – (1, ref. [51]; 2, ref. [71]). Western blotting was performed using extracts from 293T cells transiently expressing HA and Myc epitope-tagged proteins. GAPDH protein levels were used as loading controls. Data are the means ± SD of at least three independent experiments. ***P < 0.001, **P < 0.01, *P < 0.05, t-test.