Binding of multiple factors to the MMTV promoter in crude and fractionated nuclear extracts

Abstract

Hormone activation of MMTV transcription results in the establishment of a tightly bound transcription factor complex at the promoter (Cordingley et al., Cell 48, 261–270, 1987). We have characterized two fractionable binding activities which participate in this complex. One factor, previously identified as the mouse homologue of NF-1 (or CTF), protects sequences −82 to −56 from exonuclease III digestion in vitro. Sequences protected by a second factor (−42 to −4) span the TATA box of the promoter, suggesting that the binding activity in this fraction is equivalent to the HeLa cell transcription factor TFIID (Sawadogo and Roeder, Cell 43, 165–175, 1986). The downstream boundary of exonuclease protection by the putative TATA-binding factor is −4; DNasel footprinting of this fraction, however, showed additional protection of discrete sites downstream of the cap site. The apparent concentration and promoter-specific binding activity of both factors is unaffected by hormone treatment of the cells.