TRANSCRIPTION FACTOR E2F BINDS DNA AS A HETERODIMER

E2F is a mammalian transcription factor that appears to play an import ant role in cell cycle control. DNA affinity column-purified E2F from HeLa cells reproducibly exhibits multiple protein bands when analyzed by SDS/PAGE. After electrophoretic purification, electroelution, and r efolding of the individual protein components, the E2F DNA binding act ivity of the individual proteins was poor. However, upon mixing the in dividual components together, a dramatic (100- to 1000-fold) increase in specific DNA binding activity was observed. The five protein bands isolated can be separated into two groups based on apparent molecular mass. Optimal reconstitution of activity requires one of the two prote ins found in the group of larger molecular mass (almost-equal-to 60 kD a) and one of the three proteins in the smaller-sized group (almost-eq ual-to 50 kDa). The reconstituted heterodimer is identical to authenti c affinity-purified E2F by three criteria: DNA-binding specificity, DN A footprinting pattern, and binding to the retinoblastoma gene product . A recently cloned protein with E2F-like activity, RBP3/E2F-1, is rel ated to the protein components of the group of larger molecular mass, as determined by Western blot analysis and reconstitution experiments. These data suggest that E2F, like many other transcription factors, b inds DNA as an oligomeric complex composed of at least two distinct pr oteins.