Nucleosomes and subnucleosomes: heterogeneity and composition

Abstract

Previous studies ( Varshavsky, Bakayev and Georgiev, 1976a) have shown that chromatin subunits (mononucleosomes) and their oligomers in a mild staphylococcal nuclease digest of chromatin display a heterogeneous content of histone H1. We now report that a mild staphylococcal nuclease digest of either chromatin or nuclei from mouse Ehrlich tumor cells contains mononucleosomes of three discrete kinds. The smallest mononucleosome (MN 1) contains all histones except H1 and a DNA fragment 140 base pairs (bp) long. The intermediate mononucleosome (MN 2) contains all five histones and a DNA fragment 170 bp long. The third mononucleosome (MN 3) also contains all five histones, but its DNA fragment is longer and more heterogeneous in size (180–200 bp). Most of the MN 3 particles are rapidly converted by nuclease into mononucleosomes MN 1 and MN 2 There exists, however, a relatively nuclease-resistant subpopulation of the MN 3 mononucleosomes. These 200 bp MN 1 particles contain not only histones but also nonhistone proteins, and are significantly more resistant to nuclease than the bulk of MN 3 particles and the smaller mononucleosomes MN 1 and MN 2. There are eight major kinds of staphylococcal nuclease-produced soluble subnucleosomes (SN). The SN 1 is a set of naked double-stranded DNA fragments ∼20 bp long. The SN 2 is a complex of a specific basic nonhistone protein (molecular weight ∼16,000 daltons) and a DNA fragment ∼27 bp long. The SN 3 contains histone H4, the above-mentioned specific nonhistone protein and a DNA fragment ∼27 bp long. The SN 4 contains histones H2a, H2b, H4 and a DNA fragment ∼45 bp long. The SN 5 contains histones H2a, H2b, H3 and a DNA fragment ∼55 bp long. The SN 6 is a complex of histone H1 and a DNA fragment ∼35 bp long. Subnucleosomes SN 7 and SN 8 each contain all the histones except H1, and DNA fragments ∼100 and ∼120 bp long, respectively. Nuclease digestion of isolated mono- or dinucleosomes does not produce some of the subnucleosomes. These and related findings indicate that the cleavage required to generate these subnucleosomes result from some aspect of chromatin structure which is lost upon digestion to mono- and dinucleosomes.