Cell cycle-dependent changes in conformation and composition of nucleosomes containing human histone gene sequences

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Abstract
Unfolding of the nucleosomes in transcriptionally active chromatin uncovers the sulfhydryl groups of histone H3 and permits the selective recovery of the unfolded nucleosomes by mercury-affinity chromatography. This new technique has been used to compare the nucleosomal proteins and their postsynthetic modifications in the unfolded and the compactly beaded nucleosomes of HeLa cells in logarithmic growth, and at different stages of the growth cycle. The Hg-bound nucleosomes are shown to be deficient in replicating DNA sequences, but to remain associated with fragments of nascent RNA chains (or RNP particles) during gradient centrifugations. Both nucleosome fractions contain a full complement of "core" histones but differ with respect to postsynthetic modifications. The Hg-bound nucleosomes contain high levels of the tri- and tetra-acetylated forms of histones H3 and H4. The unbound nucleosomes are deficient in acetylated histones but enriched in phosphorylated H2A. In synchronized HeLa cells, histone H2A and H4 gene sequences occur in the Hg-bound nucleosomes during the S-phase when their transcription takes place, but not in the G2-phase when the genes are repressed.