Protection of discrete DNA fragments by the complex H1-octamerhistones or H5-octamerhistones after micrococcal nuclease digestion

Abstract

Several authors, including ourselves, have reported the existence of chromatosomes with DNA size larger than 166 bp in bird erythrocyte chromatin. It was tempting to correlate this increased DNA size with the presence of histone H5. In order to substantiate this hypothesis, we performed a micrococcal nuclease digestion kinetic on: chicken erythrocyte chromatin, either native, selectively depleted from H1, or from H1 and H5; and rat liver chromatin, either native or partially H1 depleted. The comparative analysis of the lengths of DNA in the chromatosome size region led to the following conclusions: - denaturing gels clearly reveal a first discrete pause at 178 nucleotides in H1 depleted chicken erythrocyte chromatin as well as in partially H1-depleted rat liver chromatin, before the material accumulates at the next intermediate 166 nucleotide chromatosome pause. - the generation of all discrete chromatosome bands is critically dependent on low ionic strength conditions and low Ca++ concentrations during the digestion, suggesting it may result from the protection of DNA cleavage sites by histone H5 or H1, C or N terminal domains.