[Organization of DNA replication domains in S-phase nuclei of human cells]

Each chromosome of eukaryotic cells contains multiple units of DNA replication that are activated during S-phase of cell cycle according to a definite program. It is considered at present that the main independent units of replication in mammalian cells represent groups of 20-25 adjacent synchronously activated small (with the average size 100 kbp) replicons. After labelling of nascent DNA with nonradioactive DNA precursors and immunofluorescent staining of incorporated label, discrete replication domains (RDs) are detected in S-phase nuclei. It is assumed that each RD is formed by a single group of synchronously activated small replicons. Since the average rate of replication fork movement is 2 kbp/min, a group of small replicons should finish DNA synthesis within 25 min, and only during this time one RD should incorporate the replicative label. We have studied the duration of DNA synthesis in individual RDs in S-phase human cells using double replicative labelling that can be detected in the nucleus by specific reagents. Our results indicate that in the main fraction of RDs DNA synthesis lasts more than 90 min, that contradicts the generally accepted model of organization of replication units in mammalian cells (Hand, 1978), but is in agreement with an alternative model, according to which the main replication units are single or clustered big replicons more than 300 kbp in size (Liapunova, 1994).