Dispersive initiation of replication in the Chinese hamster rhodopsin locus

Several higher eukaryotic replication origins appear to be composed of broa d zones of potential nascent strand start sites, while others are more circ umscribed, resembling those of yeast, bacteria, and viruses. The most deloc alized origin identified so far is similar to 55 kb in length and lies betw een the convergently transcribed dihydrofolate reductase (DHFR) and the 2BE 2121 genes on chromosome 2 in the Chinese hamster genome. In some of our st udies, we have utilized the rhodopsin origin as an early replicating intern al standard for assessing the effects of deleting various parts of the DHFR locus on DHFR origin activity. However, it had not been previously establi shed that the rhodopsin locus was located at a site far enough away to be i mmune to such deletions, nor had the mechanism of initiation at this origin been characterized. In the present study, are have localized the rhodopsin domain to a pair of small metacentric chromosomes and have used neutral/ne utral 2-D gel replicon mapping to show that initiation in this origin is al so highly delocalized, encompassing a region more than 50 kb in length that includes the nontranscribed rhodopsin gene itself. The initiation zone is flanked at least on one end by an actively transcribed gene that does not s upport initiation. Thus, the DHFR and rhodopsin origins belong to a class o f complex, polydisperse origins that appears to be unique to higher eukaryo tic cells. (C) 2000 Academic Press.