Identification of a stage selector element in the human gamma-globin gene promoter that fosters preferential interaction with the 5' HS2 enhancer when in competition with the beta-promoter

EMBO J. 1992 Aug;11(8):2961-9.

Abstract

The erythroid-specific enhancer within hypersensitivity site 2 (HS2) of the human beta-globin locus control region is required for high level globin gene expression. We investigated interaction between HS2 and the gamma- and beta-promoters using reporter constructs in transient assays in human erythroleukemia (K562) cells. The beta-promoter, usually silent in K562 cells, was activated by HS2. This activity was abolished when a gamma-promoter was linked in cis. Analysis of truncation mutants suggested that sequences conveying the competitive advantage of the gamma-promoter for HS2 included those between positions -53 and -35 relative to the transcriptional start site. This sequence, when used to replace the corresponding region of the beta-promoter, increased beta-promoter activity 10-fold when linked to HS2. The modified beta-promoter was also capable of competing with a gamma-promoter modified internally in the -53 to -35 region, when the two promoters were linked to HS2 in a single plasmid. The corresponding sequences from the Galago gamma-promoter, a species which lacks fetal gamma-gene expression, were inactive in analogous assays. We have identified and partially purified a nuclear protein found in human (fetal stage) erythroleukemia cells, but present in much lower concentration in murine (adult stage) erythroleukemia cells, that binds the -53 to -35 sequence of the gamma-promoter. We speculate that this region of the gamma-promoter functions as a stage selector element in the regulation of hemoglobin switching in humans.

MeSH terms

  • Animals
  • Base Sequence
  • Binding Sites
  • Binding, Competitive
  • Cell Line
  • Cell Nucleus / physiology
  • Chloramphenicol O-Acetyltransferase / genetics
  • Chloramphenicol O-Acetyltransferase / metabolism
  • Enhancer Elements, Genetic*
  • Gene Expression
  • Gene Expression Regulation, Neoplastic
  • Globins / genetics*
  • Humans
  • Kinetics
  • Leukemia, Experimental
  • Leukemia, Myelogenous, Chronic, BCR-ABL Positive
  • Luciferases / genetics
  • Luciferases / metabolism
  • Mice
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Promoter Regions, Genetic*
  • Recombinant Fusion Proteins / metabolism
  • Restriction Mapping
  • Transfection

Substances

  • Recombinant Fusion Proteins
  • Globins
  • Luciferases
  • Chloramphenicol O-Acetyltransferase