Fig. 5

Analysis of the interaction between ZNF746 or ZNF777 protein fragments and endogenous TRIM28. Extracts from HeLa cells after transient expression of GST/GST-ZNF fusion constructs were subjected to immunoprecipitation using TRIM28 as bait. Input protein extracts (“X”) and eluted immunoprecipitates (“IP”) were analyzed by Western blotting. The blots were probed with monoclonal antibodies against TRIM28 and rabbit polyclonal antibodies against GST, followed by respective secondary antibodies with different fluorescent tags. Images represent black/white representations of cropped regions (see Methods) informative to evaluate TRIM28 precipitation (upper part) and GST/GST-fusion protein co-immunoprecipitations (lower parts) from the same lanes. White block arrows indicate GST/GST-fusion protein signals and the black block arrows point to TRIM28 bands in the input cell lysate lanes. GST alone was used as a negative control whereas the KRAB domain of ZNF10 served as positive control. Further controls were obtained by mock immunoprecipitation from non-transfected HeLa lysates (label “HeLa”). a Results for Z746a-derived GST fusions. b data of Z746b-derived GST fusions. c analysis of Z777-derived GST fusions. Asterisks indicate unspecific bands