Separation of basal histone synthesis from S-phase histone synthesis in dividing cells

Abstract

Histone synthesis in cycling tissue-culture cells can be separated into basal synthesis and S-phase synthesis. S-phase histone synthesis comprises about 90% of total histone synthesis, and in this regard our results support the generally held model that most histone synthesis is tightly linked to DNA synthesis. Basal histone synthesis is less than 10% of the S-phase synthesis but persists throughout the S, G2 and G1 phases of the cell cycle, while S-phase synthesis rises in parallel with the increase in DNA synthesis. In Chinese hamster ovary cells, the H2A variants Z and X, and the H3 variant 3, participate in basal histone synthesis while the H2A variants 1 and 2, and the H3 variant 2, participate in S-phase histone synthesis. H2B and H4 participate in both kinds of synthesis. Because specific H2A and H3 variants are involved, basal histone synthesis cannot be attributed to contamination by cells in S-phase. Whole cells and isolated nuclei yield the same results. Basal histone synthesis can be demonstrated in cells synchronized by mitotic shake-off either alone or in combination with isoleucine starvation and colcemid blockage. It is also differentiated from S-phase histone synthesis by its relative insensitivity to hydroxyurea, indicating that basal histone synthesis is not linked or is at least less tightly linked to DNA synthesis. The basal H2A variants X and Z of five other cell lines all show similar relative insensitivities to hydroxyurea treatment. These lines include attached cultures, suspension cultures, a normal line and transformed lines from human, mouse, rat and Chinese hamster, indicating that basal histone synthesis may be a property of all cycling cells. The integration of basal histones into chromatin seems to follow the same pathway as S-phase histones, but the kinetics are somewhat slower.