FIG. 7.

SUMOylation of CTCF contributes to its repressive effect on the c-myc P2 promoter. HEK 293 cells were transfected with 500 ng of the c-myc P2 luciferase reporter construct either with no added CTCF-encoding plasmid (far left black bar) or with 10, 50, or 100 ng of plasmid DNAs encoding either wild-type CTCF or CTCF bearing mutations in the SUMO modification sites. The luciferase activities were measured as described in Materials and Methods and were normalized to the activity of a cotransfected plasmid encoding Renilla luciferase. The repressive effect of CTCF on the promoter is shown by the concentration-dependent four- to fivefold reduction in luciferase activity (black bars). This reduction was partially decreased by either the single VRKE or MRTE mutation or the double VRKE and MRTE mutations, which decrease the SUMOylation of CTCF. The asterisks indicate that the results were statistically significant (P < 0.05) when a pairwise Student t test was used to compare the CTCF mutants to the wild-type conditions.