E2F-6 binds specifically to an E2F site. A labeled oligonucleotide corresponding to the E2F binding site in the DHFR promoter was incubated with in vitro translated myc-DP2 or HA-E2F-6 protein (lanes 1 and 2) or cotranslated myc-DP2 and HA-E2F-6 proteins (lanes 3–11). Unlabeled DHFR wild-type (wt) or mutated (mut) oligonucleotides were added in the amounts indicated. In supershift experiments, 1 μl of anti-HA antibody (HA) or anti-Myc antibody (9E10) was added. Filled arrow, position of the specific E2F-6 complex; open arrow, endogenous E2F binding activity in the reticulocyte lysate; ∗, HA supershifted band; solid circle, 9E10 supershifted band.