Visualizing the dynamics of chromosome structure formation coupled with DNA replication

Abstract

A basic question of cell biology is how DNA folds to chromosome. Numbers of examples have suggested the involvement of DNA replication in chromosome structure formation. To visualize and identify the dynamics of chromosome structure formation and to elucidate the involvement of DNA replication in chromosome construction, Cy3-2'-deoxyuridine-5'-triphosphate direct-labeled active replicating DNA was observed in prematurely condensed chromosomes (PCCs) under a confocal scanning microscope utilized with drug-induced premature chromosome condensation (PCC) technique that facilitates the visualization of interphase chromatin as condensed chromosome form. S-phase PCCs revealed clearly the drastic dynamics of chromosome formation that transits during S-phase from a 'cloudy nebula' to numerous numbers of 'beads on a string' and finally to 'striped arrays of banding structured chromosome' along with the progress of DNA replication. The number, distribution, and shape of replication foci were also measured in individual subphases of S-phase more precisely than reported previously; maximally, approximately 1,400 foci of 0.35 microm average radius size were scored at the beginning of the S-phase, and the number reduced to approximately 100 at the end of the S-phase. Drug-induced PCC clearly provided the new insight that eukaryote DNA replication is tightly coupled with the chromosome condensation/compaction for the construction of the higher-ordered structure of the eukaryote chromosome.