5′-Flanking Sequences Mediate Butyrate Stimulation of Embryonic Globin Gene Expression in Adult Erythroid Cells

Abstract

A stable transfection assay was used to test the mechanism by which embryonic globin gene transcription is stimulated in adult erythroid cells exposed to butyric acid and its analogs. To test the appropriate expression and inducibility of chicken globin genes in murine erythroleukemia (MEL) cells, an adult chicken β-globin gene construct was stably transfected. The chicken β-globin gene was found to be coregulated with the endogenous adult mouse α-globin gene following induction of erythroid differentiation of the transfected MEL cells by incubation with either 2% dimethyl sulfoxide (DMSO) or 1 mM sodium butyrate (NaB). In contrast, a stably transfected embryonic chicken β-type globin gene, ρ, was downregulated during DMSO-induced MEL cell differentiation. However, incubation with NaB, which induces MEL cell differentiation, or α-amino butyrate, which does not induce differentiation of MEL cells, resulted in markedly increased levels of transcription from the stably transfected ρ gene. Analysis of histone modification showed that induction of ρ gene expression was not correlated with increased bulk histone acetylation. A region of 5′-flanking sequence extending from -569 to -725 bp upstream of the ρ gene cap site was found to be required for both downregulation of ρ gene expression during DMSO-induced differentiation and upregulation by treatment with NaB or α-amino butyrate. These data are support for a novel mechanism by which butyrate compounds can alter cellular gene expression through specific DNA sequences. The results reported here are also evidence that 5′-flanking sequences are involved in the suppression of embryonic globin gene expression in terminally differentiated adult erythroid cells.