Erythroid Krüppel-like factor is recruited to the CACCC box in the β-globin promoter but not to the CACCC box in the γ-globin promoter: The role of the neighboring promoter elements

The programmed expression of the five β-like globin genes (ɛ, (A)γ, (G)γ, δ, and β) is characterized by a series of switches that are developmentally regulated. The (A)γ- and (G)γ- (fetus) to β-globin (adult) switch depends on transcription factor erythroid Krüppel-like factor (EKLF), which, like Sp1, binds to CACCC boxes. EKLF is essential for the expression of the β-globin but not the γ-globin gene. Because both γ-globin and β-globin promoters contain the CACCC box, and their promoter elements are similar, it is not known why the two promoters behave so differently. In this report, we searched for the functional differences between the two promoters by studying their ability to recruit EKLF. We used the in vivo PIN*POINT assay to show that EKLF is recruited to the β-globin promoter but not to the γ-globin promoter. We show that this selectivity is a result of differences in surrounding promoter elements and not CACCC box alone. One of the differences between the two promoters with a functional consequence is the CCTTG repeat that is present in the γ-globin promoter but not in the β-globin promoter. The repeat, when inserted in the β-globin promoter, decreases EKLF recruitment to and activity of the β-globin promoter, suggesting that the repeat functions as a suppressor element. The CCTTG repeat can also suppress the SV40 promoter in cis, and the suppressor factor binding to the repeat can be squelched with a plasmid containing a high copy number of the repeat. These findings may have implications in designing drug targets for treatment of β-globin disorders.