Butyric acid and its various analogues are known to stimulate fetal hemoglobin synthesis in certain mammalian erythroleukemia cell lines and in patients affected with hemoglobinopathy disorders. Accelerated erythropoiesis and/or increased transcription are two possible mechanisms to explain induction of the Aγ gene by butyrate compounds. Increased transcription may be mediated by inhibition of histone deacetylase with resulting nucleosome deslabilization, and/or the binding of a butyrate induced trans-factor to the Aγ promoter. Previous truncational analysis of the Aγ promoter, using transgenic mice and mouse erythroleukemia cells, demonstrated that extension of the promoter from -822 to -893 resulted in a significant increase in Aγ mRNA production following treatment with α-amino butyric acid. This DNA sequence may serve as a binding site for butyrate induced nuclear trans-factors. The protein binding pattern in this region was analyzed by gel mobility shift assay. Synthetic oligonucleotides representing the 3′ and 5′ halves of this 71 base pair region were end labeled with [γ-32P]dATP and incubated with nuclear extracts from HeLa cells and butyrate induced and uninduced K562 cells. Following gel electrophoresis, three major retarded bands were observed. The lower two bands represented specific DNA-protein interactions between the 3′ (-822 to -856) probe and K562 extracts. The middle band showed decreased binding in induced K562 cells. The lower band showed increased binding in K562 cells induced with butyrate; comparable binding with HeLa nuclear extract suggests this protein is non-erythroid specific. No significant change in protein binding to the 5′ (-857 to -893) probe was observed. These data suggest that induction of Aγ gene expression by butyrate may involve altered DNA-protein interactions in the -822 to -856 region of the Aγ promoter.
|Original language||English (US)|
|Journal||Journal of Investigative Medicine|
|State||Published - Jan 1 1996|
ASJC Scopus subject areas
- Biochemistry, Genetics and Molecular Biology(all)