Hemoglobin A2 (HbA2), which contains
delta-globin as its non-
alpha-globin, represents a minor fraction of the Hb found in normal adults. It has been shown recently that HbA2 is as potent as HbF in inhibiting intracellular
deoxy-HbS polymerization, and its expression is therefore relevant to
sickle cell disease treatment strategies. To elucidate the mechanisms responsible for the low-level expression of the
delta-globin gene in adult erythroid cells, we first compared promoter sequences and found that the
delta-globin gene differs from the
beta-globin gene in the absence of an erythroid Krüppel-like factor (
EKLF) binding site, the alteration of the CCAAT box to CCAAC, and the presence of a GATA-1 binding site. Second, serial deletions of the human
delta-globin promoter sequence fused to a
luciferase (LUC) reporter gene were transfected into K562 cells. We identified both positive and negative regulatory regions in the 5' flanking sequence. Furthermore, a plasmid containing a single base pair (bp) mutation in the CCAAC box of the delta promoter, restoring the CCAAT box, caused a 5.6-fold and 2.4-fold (P < .05) increase of LUC activity in transfected K562 cells and MEL cells, respectively, in comparison to the wild-type delta promoter. A set of substitutions that create an
EKLF binding site centered at -85 bp increased the expression by 26.8-fold and 6.5-fold (P < .05) in K562 and MEL cells, respectively. These results clearly demonstrate that the restoration of either an
EKLF binding site or the CCAAT box can increase
delta-globin gene expression, with potential future clinical benefit.