Restoration of the CCAAT box or insertion of the CACCC motif activate δ-globin gene expression

Delia C. Tang, David Ebb, Ross C. Hardison, Griffin P. Rodgers

Research output: Contribution to journalArticlepeer-review

23 Scopus citations

Abstract

Hemoglobin A2 (HbA2), which contains δ-globin as its non-α-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 δ-globin gene in adult erythroid cells, we first compared promoter sequences and found that the δ-globin gene differs from the β-globin gene in the absence of an erythroid Kroppel-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 δ- 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 δ 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 δ 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 δ-globin gene expression, with potential future clinical benefit.

Original languageEnglish (US)
Pages (from-to)421-427
Number of pages7
JournalBlood
Volume90
Issue number1
DOIs
StatePublished - Jul 1 1997

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Immunology
  • Hematology
  • Cell Biology

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