The kinetics of non-histone chromosomal protein (NHCP) synthesis were studied in Chinese hamster ovary (CHO) plateau phase cells stimulated to proliferate and were compared to NHCP synthesis kinetics in two populations of synchronous G1 traversing cells. In all cases, NHCP synthesis rates increase 3to 5-fold as cells traversed G , and attained maximum values one hour before semi-conservative DNA replication began. Similar to results in synchronous GI cells, the molecular weight distributions of the NHCP fraction from stimulated plateau phase cells underwent only minor changes, measured by sodium dodecylsulfate (SDS) polyacrylamide gel electrophoresis, as these cells moved toward S phase. Yet, during this progression after plateau phase and in the transition from early G , to late G, in synchronous cells, the total NHCP fraction increased significantly (1.5-2-fold) in amount per cell. These data indicate that plateau phase cells are similar to early GI cells, both in terms of their amounts of non-histone per cell and in their subsequent NHCP synthesis kinetics as they move toward S phase. These results extend previous findings which suggested that NHCP synthesis was coupled to DNA replication and demonstrate that the increased NHCP synthesis and accumulation in chromatin may be a biochemical marker for G1 progression.
Previous studies have shown that NHCP synthesis rates increase significantly as cells traverse G1 phase (Stem and Borun, '72; Gerner and Humphrey, '73) and support the hypothesis that the synthesis of these chromatin associated proteins is involved in the processes controlling semi-conservative DNA replication in mammalian cells (see Stein and Baserga, '72, for review). However, before a cause/ effect relationship between NHCP synthesis and DNA replication can be seriously considered, a more complete understanding of the temporal relationship of these two biochemical components must be obtained. The possibility exists that the increased synthesis of non-histones, which are a heterogeneous class of proteins containing many species, may only be coincidental to DNA synthesis. Some data already exists to support a causal relationship. It has been shown that increased GI synthesis of non-histones is inhibited by the drug 5-azacytidine, as is subsequent DNA (Tsuboi and Baserga, '72). A previous report demonstrated that non-histones from late G1 cells, inhibited from entering S phase by hydroxyurea (HU), continue to be synthesized at elevated rates (5-fold higher than non-histones from early G1 cells) (Gerner and Humphrey, '73). In addition, data in that communication showed that non-histones synthesis rates decreased more than 4-fold during the first hour after cells left late GI and entered early S phase.
The purpose of the studies contained in this report was to further illucidate the temporal relation of NHCP synthesis in G1 traversing cells and DNA replication. If these two fractions are coupled in some manner, their relationship should be conserved when the length of GI is changed. This report describes the kinetics of NHCP