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Restoration of the CCAAT box or insertion of the CACCC motif activates [corrected] delta-globin gene expression.

Abstract
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.
AuthorsD C Tang, D Ebb, R C Hardison, G P Rodgers
JournalBlood (Blood) Vol. 90 Issue 1 Pg. 421-7 (Jul 01 1997) ISSN: 0006-4971 [Print] United States
PMID9207479 (Publication Type: Journal Article, Research Support, U.S. Gov't, P.H.S.)
Chemical References
  • DNA Transposable Elements
  • Globins
  • Hemoglobin A2
Topics
  • Adult
  • Amino Acid Sequence
  • Cell Line
  • DNA Transposable Elements (genetics)
  • Gene Expression Regulation
  • Globins (genetics)
  • Hemoglobin A2 (genetics)
  • Humans
  • Molecular Sequence Data
  • Sequence Alignment
  • Sequence Analysis, DNA
  • Transfection

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