Fractionation of Rat Liver Chromatins: Effects of Cations,Hepatectomy and Actinomycin D 1

Abstract

Summary

Rat liver chromatin was fractionated by differential centrifugation of nuclear sonicates in a 0.25–2.5 M linear sucrose gradient. Diffuse, transitional, and dense chromatin fractions were resolved which contained 22 ± 4, 10 ± 4, and 68 ± 5% of the total DNA, respectively. The diffuse chromatin incorporated RNA precursors in vivo 10–15 times more actively, and contained 2–3 times more RNA, phospholipid and nonhistone protein per unit DNA than dense chromatin. Mg²⁺ and Ca²⁺ (0.001-0.003 M) caused the diffuse chromatin to sediment with the dense chromatin apparently due to aggregation. Na⁺ and K⁺ (0.05 M) rendered the dense inactive chromatin more buoyant, possibly by dispersing it. During liver regeneration (12-hr posthepatectomy), the diffuse chromatin content increased 50% and incorporation of ¹⁴C-orotate into its RNA trebled. Actinomycin D (a single dose of 1 mg/kg administered 16 hr before sacrifice) reduced the size of the diffuse chromatin fraction by 65% and the specific activity of its RNA by 70%.