Restoration of the CCAAT Box or Insertion of the CACCC Motif Activate δ-Globin Gene Expression
| Autor: | Griffin P. Rodgers, David H. Ebb, Ross C. Hardison, Delia C. Tang |
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| Rok vydání: | 1997 |
| Předmět: | |
| Zdroj: | Blood. 90:421-427 |
| ISSN: | 1528-0020 0006-4971 |
| DOI: | 10.1182/blood.v90.1.421 |
| Popis: | 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 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 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. |
| Databáze: | OpenAIRE |
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