The GRP78/BIP promoter E2F1-like sequences in the distal GC-box and ERSE
1 element compete for nuclear binding with the specific Apaf-1 promoter E2F1
site. Biotin-labeled Apaf-1 E2F1 sequence (lanes 2-4) and
GRP78/BIP Sp1 binding sequences -324 to -311 (distal GC-box; lanes
5-7) and -126 to -108 (GC-rich element of ERSE 1; lanes 8-10)
were incubated with Saos-2ERE2F1 cell nuclear extracts and a 50-fold molar
excess of cold double-stranded specific competitor for the Apaf-1 E2F1 site
(lanes 3, 6, and 9) or an equimolar amount of mutant Apaf-1
E2F1 site competitor (lanes 4, 7, and 10). The
arrow indicates a specific band competed by double-stranded cognate
competitor. The faster migrating complexes B′ and C′ with the
GC-box E2F site are not E2F1-specific. Lane 1, binding without
nuclear extract.