The Origin Recognition Complex, SIR1 , and the S Phase Requirement for Silencing

Abstract

Silencing of transcription in Saccharomyces cerevisiae has several links to DNA replication, including a role for the origin recognition complex (ORC), the DNA replication initiator, in both processes. In addition, the establishment of silencing at the HML and HMR loci requires cells to pass through the S phase of the cell cycle. Passage through S phase was required for silencing of HMR even under conditions in which ORC itself was no longer required. The requirement for ORC in silencing of HMR could be bypassed by tethering the Sir1 protein to the HMR-E silencer. However, ORC had a Sir1-independent role in transcriptional silencing at telomeres. Thus, the role of ORC in silencing was separable from its role in initiation, and the role of S phase in silencing was independent of replication initiation at the silencers.

Differential Requirement of DNA Replication Factors for Subtelomeric ARS Consensus Sequence Protosilencers in Saccharomyces cerevisiae

“…The establishment of silent chromatin requires a passage through S-phase (Miller and Nasmyth 1984), but the requirement for DNA replication per se has been ruled out (Dubey et al 1991;Kirchmaier and Rine 2001;Li et al 2001;Sharma et al 2001). Nevertheless, the list of DNA replication factors that affect silencing has been steadily growing (EhrenhoferMurray et al 1995(EhrenhoferMurray et al , 1999Dillin and Rine 1997;Fox et al 1997;Dziak et al 2003;Suter et al 2004;Liachko and Tye 2005). In many instances it is not clear if mutations in these factors affect silencing directly or indirectly through effects on DNA replication or cell cycle.…”

“…Furthermore, robust silencing at HMR does not take place until M-phase (Lau et al 2002). Nevertheless, many of the genes that affect silencing both at telomeres and at HM loci encode for bona fide DNA replication factors such as ORC2, ORC5, minichromosome maintenance (MCM)5, MCM10, CDC44(RF-C), CDC45, and POL30(PCNA) (Ehrenhofer-Murray et al 1995Dillin and Rine 1997;Fox et al 1997;Dziak et al 2003;Suter et al 2004;Liachko and Tye 2005).…”

Mating-Type Genes andMATSwitching inSaccharomyces cerevisiae

“…Yet when one examines these same sequences on the chromosome by two-dimensional gel electrophoresis, to detect structures characteristic of the presence of an origin of DNA replication, neither HML-E nor HML-I is active, while HMR-E does appear to act as a chromosomal origin, although only in a fraction of cell cycles (Dubey et al 1991;Rivier and Rine 1992;Collins and Newlon 1994). Whether silencing depends on origin activity has been the subject of much debate; current evidence suggests that the binding of the ORC proteins to silencer regions is a key step in establishing silencing, but it is not necessary that replication be initiated at that site (Fox et al 1993;Ehrenhofer-Murray et al 1995;Fox et al 1997;Li et al 2001). Indeed mutations of Orc5 were isolated that were defective in silencing but not in replication (Fox et al 1995).…”

Transcriptional silencing functions of the yeast protein Orc1/Sir3 subfunctionalized after gene duplication

“…Orc1, as part of ORC, binds silencers (16), and the BAH domain of Orc1 interacts with Sir1 (26,27) to stabilize other Sir proteins at silencers. This recruitment of Sir1 is thought to be the only function of ORC in silencing because ORC can be bypassed by tethering Sir1 to the silencer (27,35). Interestingly, the BAH domain of Orc1 is not essential for DNA replication in S. cerevisiae (31), although it is conserved in most eukaryotic orthologs of Orc1.…”