CHROMATIN ASSEMBLY FACTOR-I CONTRIBUTES TO THE MAINTENANCE, BUT NOT THE REESTABLISHMENT, OF SILENCING AT THE YEAST SILENT MATING LOCI

CAC1/RLF2 encodes the largest subunit of chromatin assembly factor I ( CAF-I), a complex that assembles newly synthesized histones onto recen tly replicated DNA in vitro. In vivo, cac1/rlf2 mutants are defective in telomeric silencing and mislocalize Rap1p, a telomere binding prote in. Here, we report that in cells lacking CAF-I the silent mating loci are derepressed partially. MATa cac1 cells exhibit an unusual respons e to or factor: They arrest and form mating projections (shmoos) initi ally, but are unable to sustain the arrest stare, giving rise to clust ers of shmooing cells. cac1 MATa HMLa HMRa strains do not form these s hmoo clusters, indicating that derepression of HML alpha causes the sh moo cluster phenotype in cac1 cells. When SIR3 is reintroduced into si r1 sir3 cells, HML remains derepressed indicating that SIR1 is require d for the re-establishment of silencing at HML. In contrast, when SIR3 is reintroduced into cac1 sir3 cells, silencing is restored to HML, i ndicating that CAF-I is not required for the re-establishment of silen cing. Loss of the other CAF-I subunits (Cac2p and Cac3p/Msi1p) also re sults in the shmoo cluster phenotype, implying that loss of CAF-I acti vity gives rise to this unstable repression of HML. Strains carrying c ertain mutations in the amino terminus of histone H4 and strains with limiting amounts of Sir2p or Sir3p also form shmoo clusters, implying that the shmoo cluster phenotype is indicative of defects in maintenan ce of the structural integrity of silent chromatin. MATa cac(-) sir1 d ouble mutants have a synergistic mating defect, suggesting that the tw o silencing mechanisms, establishment and maintenance, function cooper atively. We propose a model to explain the distinctions between the es tablishment and the maintenance of silent chromatin.