Replication control is fundamental to genomic stability as aberrant replication within a single cell cycle can induce high rates of segmental amplification, chromosomal aneuploidy, and possibly other genomic instabilities. Current models for how eukaryotic cells prevent such re-initiation focus on the global cell-wide inhibition of replication proteins involved in loading the Mcm2-7 replicative helicase at origins (e.g. cyclin dependent kinase, CDK, inhibition of ORC, Cdc6, Cdt1, Mcm2-7). By preventing this initial step of initiation from reoccurring once S phase begins, re-initiation can be effectively prevented. Such models, however, treat origins as generic interchangeable elements and cannot account for the diverse efficiencies with which origins re-initiate when global control mechanisms are disrupted. These varied re-initiation efficiencies also cannot be explained by the well-documented diversity in origin timing and efficiency observed during normal S phase initiation. Instead, we now have evidence of a novel mechanism that contributes to the diversity in origin re-initiation efficiency.

Chapter 2 of this dissertation details the identification and characterization of genetic elements near ARS317 and ARS1238 that confer preferential re-replication on these and other origins when cell cycle control of MCM2-7 and Cdc6 is disrupted. These elements do not confer any detectable change on the replication efficiency or timing of adjacent origins, suggesting that their regulatory effect is specific to origin re-initiation. Hence, we refer to these elements as Re-Initiation Promoters (RIPs). The two RIPs mapped are AT rich sequences 40-50bp in size and exert their effects on adjacent origins in an orientation and distance dependent manner. Analysis of Mcm2-7 association with origins suggests that RIP elements allow local escape from the residual CDK inhibition of helicase loading when global CDK inhibition of Mcm2-7 and Cdc6 is disrupted.

Such local modulation of origin control suggests that there is a complex genomic landscape of re-replication potential, particularly when mechanisms preventing re-replication are partially or sporadically disrupted. Hence, if re-replication does contribute to genomic alterations, as has been speculated for cancer cells, some regions of the genome may be more susceptible to these alterations than others.