Figure 5.

E2F DNA-binding activity in primary keratinocytes. Whole-cell extracts were prepared from primary keratinocytes derived from line 1.1 transgenic and nontransgenic newborn mice. E2F electrophoretic mobility-shift assay reactions contained 10 μg of either nontransgenic (Negative, lanes 1–4) or transgenic (K5 E2F1, lanes 5–11) extract and an end-labeled DNA fragment derived from the adenovirus E2 promoter. Excess (20 ng) double-stranded oligonucleotide containing either wild-type E2F sites (E2 wt, lanes 2 and 6) or mutated E2F sites (E2 mut, lanes 3 and 7) were added to the binding reaction to distinguish specific complexes from nonspecific (N.S.) complexes. Antibody specific for either E2F1 (lanes 4 and 8), Rb (lane 9), p107 (lane 10), or p130 (lane 11) was added to the binding reactions where indicated to identify proteins in specific complexes.