Conversion of a replication origin to a silencer through a pathway shared by a Forkhead transcription factor and an S phase cyclin.
Silencing of the mating-type locus HMR in Saccharomyces cerevisiae requires DNA elements called silencers. To establish HMR silencing, the origin recognition complex binds the HMR-E silencer and recruits the silent information regulator (Sir)1 protein. Sir1 in turn helps establish silencing by stabilizing binding of the other Sir proteins, Sir2-4. However, silencing is semistable even in sir1Delta cells, indicating that SIR1-independent establishment mechanisms exist. Furthermore, the requirement for SIR1 in silencing a sensitized version of HMR can be bypassed by high-copy expression of FKH1 (FKH1(hc)), a conserved forkhead transcription factor, or by deletion of the S phase cyclin CLB5 (clb5Delta). FKH1(hc) caused only a modest increase in Fkh1 levels but effectively reestablished Sir2-4 chromatin at HMR as determined by Sir3-directed chromatin immunoprecipitation. In addition, FKH1(hc) prolonged the cell cycle in a manner distinct from deletion of its close paralogue FKH2, and it created a cell cycle phenotype more reminiscent to that caused by a clb5Delta. Unexpectedly, and in contrast to SIR1, both FKH1(hc) and clb5Delta established silencing at HMR using the replication origins, ARS1 or ARSH4, as complete substitutes for HMR-E (HMRDeltaE::ARS). HMRDeltaE::ARS1 was a robust origin in CLB5 cells. However, initiation by HMRDeltaE::ARS1 was reduced by clb5Delta or FKH1(hc), whereas ARS1 at its native locus was unaffected. The CLB5-sensitivity of HMRDeltaE::ARS1 did not result from formation of Sir2-4 chromatin because sir2Delta did not rescue origin firing in clb5Delta cells. These and other data supported a model in which FKH1 and CLB5 modulated Sir2-4 chromatin and late-origin firing through opposing regulation of a common pathway.
Department of Biomolecular Chemistry, University of Wisconsin School of Medicine and Public Health, Madison, WI 53706, USA.
Laurieann
L
Casey
Erin E
EE
Patterson
Ulrika
U
Müller
Catherine A
CA
Fox
D002384:Catalysis; D018797:Cell Cycle Proteins / Q000378:metabolism*; D019926:Cyclin B / Q000378:metabolism*; D051858:Forkhead Transcription Factors / Q000378:metabolism*; D015966:Gene Expression Regulation, Fungal; D020868:Gene Silencing; D005800:Genes, Fungal; D005838:Genotype; D008954:Models, Biological; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D018741:Replication Origin / Q000235:genetics*; D016196:S Phase; D012441:Saccharomyces cerevisiae / Q000166:cytology* / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D037941:Silencer Elements, Transcriptional / Q000235:genetics*; D013489:Suppression, Genetic
C076558:CLB5 protein, S cerevisiae; D018797:Cell Cycle Proteins; D019926:Cyclin B; C464906:Fkh1 protein, S cerevisiae; D051858:Forkhead Transcription Factors; D012333:RNA, Messenger; D029701:Saccharomyces cerevisiae Proteins