THE ORIGIN RECOGNITION COMPLEX HAS ESSENTIAL FUNCTIONS IN TRANSCRIPTIONAL SILENCING AND CHROMOSOMAL REPLICATION

The role of the origin recognition complex (ORC) was investigated in r eplication initiation and in silencing. Temperature-sensitive mutation s in ORC genes caused defects in replication initiation at chromosomal origins of replication, as measured by two-dimensional (2-D) origin-m apping gels, fork migration analysis, and plasmid replication studies. These data were consistent with ORC functioning as a eukaryotic repli cation initiator. Some origins displayed greater replication initiatio n deficiencies in ore mutants than did others, revealing functional di fferences between origins. Alleles of ORC5 were isolated that were def ective for silencing but not replication, indicating that ORC's role i n silencing could be separated from its role in replication. In temper ature-sensitive ore mutants arrested in mitosis, temperature-shift exp eriments caused a loss of silencing, indicating both that ORC had func tions outside of the S phase of the cell cycle and that ORC was requir ed for the maintenance of the silenced state.