The effect of 9-cis RA on Sp1 binding to the FR-β promoter by EMSA. (A) Lanes 1 to 10, 30 000 cpm 32P-labeled probe (– 88 nt to – 33 nt); lane 2, 2.5 μg KG-1 nuclear extracts; lanes 3 to 10, 5 μg KG-1 nuclear extracts; lane 4, 50-fold excess wild-type cold probe (– 88 nt to – 33 nt); lane 5, 100-fold excess wild-type unlabeled probe (– 88 nt to – 33 nt); lane 6, 100-fold unlabeled mutated probe ΔSp1 (– 88 nt, – 33 nt); lane 7, nuclear extracts mixed with vehicle; lane 8, nuclear extracts mixed with 0.1 μM 9-cis RA; lane 9, nuclear extracts mixed with 1.0 μM 9-cis RA; and lane 10, nuclear extract mixed with 10 μM 9-cis RA. (B) Lane 1, 5 μg nuclear extract from KG-1 cells that were treated with vehicle for 5 days; lane 2, 5 μg nuclear protein from KG-1 cells that were treated with ATRA (1 μM) for 5 days; and lane 3, 5 μg nuclear protein from KG-1 cells that were treated with TPA (0.05 μM) for 5 days.
