ASSOCIATION OF AUTONOMOUS REPLICATION ACTIVITY WITH REPLICATION ORIGINS IN A HUMAN-CHROMOSOME

A systematic analysis of the correlation of autonomous replication act ivity with initiation of replication in a human chromosome was perform ed. The temporal order of replication of segments in a pericentric 320 -kb MEN203 locus on human chromosome 10 (10q11.2) was determined by pu lse-labeling of cells with Ei-bromodeoxyuridine after synchronization with aphidicolin. The entire MEN203 locus replicated during the late S phase. Two distinct segments replicated earlier than the others in th e locus, indicating that replication was initiated within or near thes e segments. Two other segments also showed an earlier response than th e respective neighboring regions. These results suggest that the MEN20 3 locus contains two distinct replication origins and two possible ori gins that may be used less frequently. The results were essentially co nfirmed by synchronization of the cell cycle with mimosine. Analysis o f autonomous replication activity of 10-kb long chromosome fragments c overing the 320-kb region showed that certain fragments replicated two or three times more efficiently than others. The results are consiste nt with our previous observations with randomly cloned human chromosom e fragments. The replication origins colocalized with fragments exhibi ting relatively high autonomous replication activity. Thus, the capaci ty for autonomous replication of chromosome fragments might be prerequ isite for the initiation of chromosomal replication. (C) 1998 Academic Press.