Chemical footprinting of 5S RNA chromatin in embryos of Drosophila melanogaster

Abstract

We have used the footprinting reagent (methidiumpropyl-EDTA) iron(II) [MPE X Fe(II)] to investigate the chromatin structure of the tandemly repeated 5S RNA genes of Drosophila melanogaster embryos. Indirect end-labelling analysis of the products of mild MPE X Fe(II) digestion of nuclei reveals one extended region of accessibility of the DNA-protein complex per 375-bp gene-spacer unit. Within this region, which spans the 135-bp gene itself, a segment of particularly high sensitivity (covering 40-60 bp) is located in the distal (3') portion of each gene. The majority of the nontranscribed spacer between genes is in a highly inaccessible chromatin conformation. This pattern is repeated for all copies of the gene-spacer unit examined, approximately 15 at each end of the cluster. Salt extraction experiments show a diminution of pattern intensity after treatment with 0.5 M KCl, but the pattern is not lost under conditions which permit nucleosome sliding. The results indicate a specific, periodic chromatin structure across these transcriptionally competent 5S genes, but one which is generated and maintained by factors other than simple nucleosome arrays. Presumably protein elements of the transcription complex play a dominant role in the structure observed.