Binding of recombinant PACC to the gsn promoter.
(A) Upper panel, schematic representation of the pacC probe and the specific competitor oligo pacC. Lower panels, gel shift analysis using increasing amounts of recombinant PACC in the presence of specific competitors and polyclonal anti-PACC antibody. Lanes 1, 5 and 9, pacC probe, no protein added. Lanes 2, 6 and 10, gel shift analysis using 1.0, 2.0 and 5.0 µg of recombinant PACC. Lanes 3 and 7, gel shift analysis in the presence of the 146 bp specific competitor. Lanes 4 and 8, gel shift analysis using the specific competitor oligo pacC. Lanes 11 to 13, supershift assay using 5.0, 10.0 and 20.0 µL of anti-PACC antibody (1∶500). (B) Upper panel, schematic representation of the mutated mpacC probe. Lower panel, gel shift with wild-type and mutated probes. Analysis using different concentrations of recombinant PACC in the presence and in the absence of competitors. Lane 1 and 3, pacC and mpacC probes, respectively, no protein added. Lanes O, gel origin; SC, specific competitor; FP, free probe.