Histone acetylation in replication and transcription: turnover at specific acetylation sites in histone H4 from Physarum polycephalum.

Histone H4 from growing cells is partially acetylated at lysines 5, 8, 12, and 16. The turnover rate at each of these sites was investigated by pulse-labeling plasmodia of Physarum polycephalum with [3H]acetate for 55 min in either S phase or G2 phase of the cell cycle. Labeled histone H4 was purified and digested with a protease which cleaves on the carboxyl side of arginine residues. The peptide containing the acetylation sites was purified by high-performance liquid chromatography. Subfractions of the peptide were obtained due to differences in acetyllysine content. Each subfraction was subjected to automated Edman degradation and the radioactivity released after each cycle was determined. Histone H4 was acetylated uniformly in vitro and acetylated peptide 1-23 was used as a control. The results show a very striking preference for turnover on lysine-5 in the "low acetyl" subfraction from cells in S phase; the "high acetyl" subfraction showed turnover at all four sites. The peptides labeled in G2 phase showed turnover mainly at positions -8, -12, and -16. The data imply that the patterns of histone acetyl turnover associated with replication and transcription are nonrandom and distinct. The results have implications for nucleosome structure particularly the possible role of lysine-5 in chromosome maturation and for the design of experiments to test chromatin function in vitro.