Nucleosomes from normal and regenerating rat liver

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Abstract
Micrococcal-nuclease digestion of rat liver nuclei selectively released mononucleosomes associated with ADP-ribosylated [Caplan, Ord & Stocken (1978) Biochem. J. 174, 475–483] histone H1. Two classes of mononucleosome were detected, those that leaked out during digestion and those that were subsequently released by 5mm-sodium phosphate buffer (pH6.8)/0.2mm-NaEDTA. The former, from which histone H1 had been dissociated, contained 140-base-pair-length DNA and core histones;the latter contained core particles and mononucleosomes with histone H1 and 200-base-pair-length DNA. When normal liver nuclei were phosphorylated with [γ-32P]ATP, dissociated histone H1, which could be separated from core particles with Sephadex G-200, showed 32P uptake. 32P uptake into histones H2A and poly(ADP-ribosyl)ated H3 was appreciable in core particles, but was less evident in nucleosomes still containing histone H1. When [3H]-thymidine was given to partially hepatectomized rats in S-phase, 5–10min pulses in animals of over 300g body wt. showed the presence of high-specific-radioactivity DNA in released core particles and mononucleosomes compared with DNA retained in the nuclear pellets. Mononucleosomes from rat livers in S-phase with new, [3H]lysine-containing histones, had higher 32P incorporation in histones H1 and their core histones, than for di- or tri-nucleosomes. Thermal-denaturation properties of control and phosphorylated mononucleosomes and core particles were very similar; removal of histone H1 and non-histone chromosomal proteins in 0.5m-NaCl markedly increased the proportion of DNA ‘melting’ below 70°C.