Chromatin fine-structure mapping of the goat beta F gene in fetal erythroid tissue.

Abstract

Using a restriction enzyme accessibility assay, we have previously demonstrated that the chromatin structure immediately proximal to the goat beta F-, beta C-, and beta A-globin genes changes in a manner which parallels their developmentally regulated expression. More specifically, the PvuII recognition sequence, located 9 nucleotides upstream from the transcriptional start site in each of the three genes, is accessible to digestion only in nuclei prepared from erythroid tissue in which the respective gene product is expressed. Here we describe two restriction enzyme sites further upstream from the transcription start sites (HindIII at -700 and SacI at -480) which were not accessible to digestion in fetal erythroid nuclei. Conversely, two sites within the second coding block of the beta F gene (AccI at +276 and BamHI at +470) were accessible in fetal erythroid tissue. The corresponding sites in the beta C and beta A genes were not available for digestion in the same fetal tissue. Processive exonuclease III digestion in situ from the three accessible restriction enzyme sites in the beta F gene allowed us to define more closely the limits of these open regions. Resistance to exonuclease III digestion was encountered at or near both intron-exon junctions flanking the first intervening sequence of the beta F gene. Conversely, no resistance to exonuclease III digestion was evident in either the first or second coding blocks or the 5' untranslated region. Digestion upstream from the PvuII site of the beta F gene was negligible. High-resolution mapping by S1 nuclease analysis indicated that the endpoint of exonuclease III digestion from this site lay immediately downstream of the ATA box.