Figure 3

Effects of mutations in the UP site on the binding of endogenous nuclear proteins. (a) The sequence of the BRCA1 proximal promoter is shown with previously characterized sites boxed. E2F sites characterized by Bindra and Glazer [27] (E2FA and E2FB) are shown by light grey boxes while the BRCA1 first exon is shown by the darker box and by the arrow showing the transcriptional start site. The arrows indicate the potential ets/GABP binding sites. (b) The sequence of the UPFR6 probe is indicated, which encompasses both the UP and E2F sites (dotted and grey box marked E2FB). Arrows identify putative GABP alpha/beta binding sites. The sequences of the various mutant probes are indicated along with their names. (c) The ³²P-labelled UPFR6 probes described in (b) were incubated in binding reactions with 5 μg of nuclear extract from the MCF-7 and T-47D cell lines and bandshift assays were performed. Only the DNA/protein complexes are shown, with the Upper, Lower and non-specific (NS) bands indicated as in Figure 1.