SYNCHRONIZED MAMMALIAN CELL CULTURES .3. VARIATION OF RIBONUCLEOTIDE REDUCTASE ACTIVITY DURING REPLICATION CYCLE OF CHINESE HAMSTER FIBROBLASTS

Ribonucleotide reductase activity was studied during the replication cycle of Colcemid synchronized Chinese hamster fibroblasts in culture. Enzyme activity dropped to a minimum value during the G1 phase, returned to the level observed in the metaphase cells as the cells entered the S phase, and continued to increase during the S and G2 phases. Actinomycin D, cycloheximide, and hydroxyurea were added to the cell cultures at the time of lowest activity (1.5 h after synchronization). With over 90 % inhibition of RNA synthesis by actinomycin D, enzyme activity returned by 4.5 h to the level observed in the metaphase cells, but a further increase was not seen. With inhibition of over 90 % of protein synthesis by cycloheximide, enzyme activity did not rise from the low level observed in the G1 phase. The addition of hydroxyurea produced an elevated level of reductase activity which rose during the S-phase to more than twice that of the control. Experiments in which extracts from cultures at different time points were mixed and assayed indicated that an inhibitor was not reponsible for the low levels of activity during the G1 phase. © 1969.