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.

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.