The silent
information regulators SIR2-4 negatively regulate chromosomal DNA
replication.
Donald Pappas,
Michael Weinreich
Chromosome Replication Lab, Van Andel Research Institute, 333 Bostwick Ave. NE,
Grand Rapids, MI 49503, USA (don.pappas@vai.org)
The Origin Recognition Complex (ORC) determines the sites of replication initiation by binding to origins and recruiting additional initiation factors to the chromatin prior to DNA synthesis. An essential step in initiation occurs when Cdc6p (ATP) binds to ORC and facilitates the recruitment of the MCM DNA helicase to origins. It is not known how Cdc6p promotes MCM binding to origins or what role Cdc6p ATP binding or hydrolysis has during initiation. To identify additional genes influencing this step we isolated extragenic suppressors of a cdc6-4 ts mutant that was compromised in the Walker A-box motif required for ATP binding. We recovered 7 independent mutants that were able to grow at high temperature but were simultaneously unable to mate with a cell of the opposite mating type due to loss-of-function mutations in the SIR2, SIR3, or SIR4 genes. A genetic analysis indicated that this negative regulation of initiation likely represents a novel role of the Sir2-4 proteins distinct from their well-known involvement in transcriptional silencing at the HM loci, at telomeres, or at the rDNA locus. We propose one model in which the Sir2-4 proteins negatively regulate the initiation of DNA replication through the generation of a repressive chromatin structure in the vicinity of some origins. Thus, when Cdc6p-ATP binding or hydrolysis is compromised, an 'open' chromatin structure becomes essential to origin function perhaps by allowing efficient recruitment of the MCM helicase.