Functional roles of in vivo footprinted DNA motifs within an α-globin enhancer: Erythroid lineage and developmental stage specificities

Qingyi Zhang, Irene Rombel, G. Narender Reddy, Jong Back Gang, C. K.James Shen

Research output: Contribution to journalArticle

23 Citations (Scopus)

Abstract

Transcriptional regulation of the human α-like globin genes, embryonic ζ2 and adult α, during erythroid development is mediated by a distal enhancer, HS-40. Previous protein-DNA binding studies have shown that HS-40 consists of multiple nuclear factor binding motifs that are occupied in vivo in an erythroid lineage- and developmental stage-specific manner. We have systematically analyzed the functional roles of these factor binding motifs of HS-40 by site-directed mutagenesis and transient expression assay in erythroid cell cultures. Three of these HS-40 enhancer motifs, 5′NF-E2/ AP1, GT II, and GATA-1(c), positively regulate the ζ2-globin promoter activity in embryonic/fetal erythroid K562 cells and the adult a-globin promoter activity in adult erythroid MEL cells. On the other hand, the 3′NF-E2/AP1 motif is able to exert both positive and negative regulatory effects on the ζ2-globin promoter activity in K562 cells, and this dual function appears to be modulated through differential binding of the ubiquitous AP1 factors and the erythroid-enriched NF-E2 factor. Mutation in the GATA-1(d) motif, which exhibits an adult erythroid-specific genomic footprint, decreases the HS-40 enhancer function in dimethyl sulfoxide-induced MEL cells but not in K562 cells. These studies have defined the regulatory roles of the different HS-40 motifs. The remarkable correlation between genomic footprinting data and the mutagenesis results also suggests that the erythroid lineage- and developmental stage-specific regulation of human α-like globin promoters is indeed modulated by stable binding of specific nuclear factors in vivo.

Original languageEnglish
Pages (from-to)8501-8505
Number of pages5
JournalJournal of Biological Chemistry
Volume270
Issue number15
DOIs
Publication statusPublished - Apr 14 1995
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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