Binding of nuclear proteins to the upstream element.

(A) The end-labeled, double-stranded oligonucleotide had a sense strand sequence 5′-GCTCGCCAGGGGTCGCTGCTCTCCCATG-3′, which includes the most upstream 24 bp of the upstream element (Figure 7A). Lanes 1-3 and lanes 4-6 represent complexes formed by extracts from mouse splenic erythroblasts (FVA cells) cultured for 0 hours (freshly isolated cells) or 24 hours, respectively. Lanes 1 and 4, no unlabeled competitor oligonucleotide; lanes 2 and 5, a 100-fold excess of a heterologous competitor oligonucleotide bearing a binding sequence for the transcription factor TAL1 (5′-CTCCCAGCAGCTGGCCTAGGAGATAGCAGCAG-3′); lanes 3 and 6, a 100-fold excess of unlabeled homologous competitor. (B) The end-labeled, double-stranded oligonucleotide had a sense strand sequence 5′-GTCCCTTTCTGGCGCGCACTCCTTTTGC-3′, which includes the sequence of the upstream element deleted in mutant 6 depicted in Figure 7A-B. Lanes 1-3 represent complexes formed with nuclear extracts of 0-hour splenic erythroblasts (FVA cells), and lanes 4-6 represent complexes formed with extracts from FVA cells cultured for 24 hours. Lanes 1 and 4, no unlabeled competitor oligonucleotide; lanes 2 and 5, a 100-fold excess of unlabeled homologous competitor oligonucleotide; lanes 3 and 6, a 100-fold excess of unlabeled heterologous competitor (TAL1 binding sequence as in Figure 8A).