A role for a replicator dominance mechanism in silencing

The role of the natural HMR ‐E silencer in modulating replication initiation and silencing by the origin recognition complex (ORC) was examined. When natural HMR ‐E was the only silencer controlling HMR , the silencer's ORC‐binding site (ACS) was dispensable for replication initiation but essential for silencing, indicating that a non‐silencer chromosomal replicator(s) existed in close proximity to the silencer. Further analysis revealed that regions flanking both sides of HMR ‐E contained replicators. In contrast to replication initiation by the intact silencer, initiation by the non‐silencer replicator(s) was abolished in an orc2‐1 mutant, indicating that these replicators were extremely sensitive to defects in ORC. Remarkably, the activity of one of the non‐silencer replicators correlated with reduced silencing; inactivation of these replicators caused by either the orc2‐1 mutation or the deletion of flanking sequences enhanced silencing. These data were consistent with a role for the ORC bound to the HMR ‐E silencer ACS in suppressing the function of neighboring ORC molecules capable of inhibiting silencing, and indicated that differences in ORC‐binding sites within HMR itself had profound effects on ORC function. Moreover, replication initiation by natural HMR ‐E was inefficient, suggesting that closely spaced replicators within HMR contributed to an inhibition of replication initiation.