Inhibition of DNA synthesis in synchronized Chinese hamster cells treated in G1 with cycloheximide

The delay in the initiation of DNA synthesis after treatments during G1 with cycloheximide (CHM), an inhibitor of protein synthesis, has been studied in synchronous Chinese hamster ovary cells. Cells treated in mitosis for a duration of 1–5 h were delayed in the initiation of DNA synthesis by the length of the CHM treatment. However, cells treated after mitosis were delayed in the initiation of DNA synthesis by a length of time greater than the duration of the CHM treatment. The difference between the length of the delay and the duration of CHM treatment reached a maximum of about 4 h, and was roughly proportional to both the duration of the CHM treatment and the time in G1 when CHM was added. Treatment of cells as they entered S phase stopped DNA synthesis; however, after removal of CHM, DNA synthesis immediately resumed.These results suggest the following, G1 events requiring RNA and/or protein synthesis are involved in the initiation and continuation of DNA synthesis. Inhibition of protein synthesis during G1 results in the decay of these events (mean lifetime of 2.9 h), and the events which have decayed away must be repeated before DNA synthesis can be initiated. Events which occurred in the previous cycle, or in G1 more than 3.5 h prior to the addition of CHM, do not have to be repeated. In cells in very late G1 or early S phase, all the events leading to the initiation of DNA synthesis have been completed and do not have to be repeated following CHM treatments up to 5 h in duration.