Studies of mammalian chromosome replication. II. Evidence for the existence of defined chromosome replicating units
Lau, Y.F.; Arrighi, F.E.
Chromosoma 83(5): 721-741
1981
ISSN/ISBN: 0009-5915 PMID: 7028418 Document Number: 171185
Sister chromatids of metaphase chromosomes can be differentially stained if the cells have replicated their DNA semiconservatively for 2 cell cycles in a medium containing 5-bromodeoxyuridine (BrdU). When prematurely condensed chromosomes (PCC) are induced in cells during the 2nd S phase after BrdU is added to the medium, the replicated chromosome segments show sister chromatid differential (SCD) staining. This PCC-SCD system was used on synchronous and asynchronous Chinese hamster ovary (CHO) cells to show that the replication patterns of the CHO cells can be categorized into G1/S, early, early-mid, mid-late and late S phase patterns according to the amount of replicated chromosomes. During the first 4 h of the S phase, the replication patterns show SCD staining in chains of small chromosome segments. The amount of replicated chromosomes increase during the mid-late and late S categories (last 4 h). Small SCD segments are also present during these late intervals of the S phase. Measurements of these replicated segments indicate the presence of characteristic chromosome fragment sizes between 0.2-1.2 .mu.m in all S phase cells except those at G1/S which contain no SCD fragments. These small segments are operationally defined as chromosome replicating units or chromosomal replicons. They are interpreted to be composed of clusters of molecular DNA replicons. The larger SCD segments in the late S cells may arise by the joining of adjacent chromosomal replicons. Further application of this PCC-SCD method to study the chromosome replication process of 2 other rodents, Peromyscus eremicus and Microtus agrestis, with peculiar chromosomal locations of heterochromatin has demonstrated an ordered sequence of chromosome replication. The euchromatin and heterochromatin of the 2 spp. undergo 2 separate sequences of decondensation, replication and condensation during the early-mid and mid-late intervals, respectively, of the S phase. Similar-sized chromosomal replicons are present in both types of chromatin. Mammalian chromosomes evidently are replicated in groups of replicating units, or chromosomal replicons, along their lengths. The organization and structure of these chromosomal replicons with respect to those of the interphase nucleus and metaphase chromosomes are discussed.