Replication pattern of a large homogenously staining chromosome region in antifolate-resistant Chinese hamster cell lines.

We have investigated the replication pattern of a large, homogenously staining chromosome region (HSR) in two antifolate-resistant Chinese hamster cell lines. This region is believed to be the location of an amplified genetic sequence which includes at least the gene coding for dihydrofolate reductase and which may be present in as many as 200 copies. It is shown that the HSR in both cell lines is among the first chromosome regions to begin DNA synthesis after reversal of an early G1 block. In cells synchronized in the S period with hydroxyurea, it is also clear that the HSR in both cell lines begins replication at many sites within its length in early S. The replicons comprising the HSR therefore may respond to a common initiation signal in early S. In on cell line (A3), replication of the HSR requires, at most, 3 hours of a 7-hour S period; in a second line (MQ19), replication proceeds for approximately 5 hours. In neither line does replication of the HSR occur concomitantly with synthesis of characteristic late replicating regions. These results were confirmed in exponential cultures using a retroactive labeling technique. The significance of these findings is discussed with reference to the possible origin and arrangement of the amplified sequence in these two cell lines.