Some spontaneous gross chromosomal rearrangements (GCRs) seem to result from DNA-replication errors. The chromatin-assembly factor I (CAF-I) and replication-coupling assembly factor (RCAF) complexes function in chromatin assembly during DNA replication and repair and could play a role in maintaining genome stability. Inactivation of CAF-I or RCAF increased the rate of accumulating different types of GCRs including translocations and deletion of chromosome arms with associated de novo telomere addition. Inactivation of CAF-I seems to cause damage that activates the DNA-damage checkpoints, whereas inactivation of RCAF seems to cause damage that activates the DNA-damage and replication checkpoints. Both defects result in increased genome instability that is normally suppressed by these checkpoints, RAD52-dependent recombination, and PIF1-dependent inhibition of de novo telomere addition. Treatment of CAF-I-or RCAF-defective cells with methyl methanesulfonate increased the induction of GCRs compared with that seen for a wild-type strain. These results indicate that coupling of chromatin assembly to DNA replication and DNA repair is critical to maintaining genome stability.

SummaryThe repair of spontaneous or induced DNA damage by homologous recombination (HR) in Saccharomyces cerevisiae will suppress chromosome rearrangements. Alternative chromosome healing pathways can result in chromosomal instability. One of these pathways is de novo telomere addition where the end of a broken chromosome is stabilized by telomerasedependent addition of telomeres at non-telomeric sites. De novo telomere addition requires the recruitment of telomerase to chromosomal targets. Subsequently, annealing of the telomerase reverse transcriptase RNA-template (guide RNA) at short regions of homology is followed by extension of the nascent 3 ¢ -end of the broken chromosome to copy a short region of the telomerase guide RNA; multiple cycles of this process yield the new telomere. Proteins including Pif1 helicase, the single-stranded DNA-binding protein Cdc13 and the Ku heterocomplex are known to participate in native telomere functions and also regulate the de novo telomere addition reaction. Studies of the sequences added at de novo telomeres have lead to a detailed description of the annealing-extension-dissociation cycles that copy the telomerase guide RNA, which can explain the heterogeneity of telomeric repeats at de novo and native telomeres in S. cerevisiae .

In telomerase-deficient Saccharomyces cerevisiae, telomeres are maintained by recombination. Here we used a S. cerevisiae assay for characterizing gross chromosomal rearrangements (GCRs) to analyze genome instability in post-senescent telomerase-deficient cells. Telomerase-deficient tlc1 and est2 mutants did not have increased GCR rates, but their telomeres could be joined to other DNAs resulting in chromosome fusions. Inactivation of Tel1 or either the Rad51 or Rad59 recombination pathways in telomerase-deficient cells increased the GCR rate, even though telomeres were maintained. The GCRs were translocations and chromosome fusions formed by nonhomologous end joining. We observed chromosome fusions only in mutant strains expressing Rad51 and Rad55 or when Tel1 was inactivated. In contrast, inactivation of Mec1 resulted in more inversion translocations such as the isochromosomes seen in human tumors. These inversion translocations seemed to be formed by recombination after replication of broken chromosomes.Telomeres function in replication and maintenance of chromosome ends, to prevent DNA ends from being inappropriately joined to each other and to prevent chromosome ends from activating checkpoints 1,2 . Telomeres are maintained by telomerase, which consists of the Est2 catalytic subunit, the Tlc1 RNA and other subunits 2 .Telomere maintenance also requires other proteins. These include the Tel1 protein kinase that functions in telomere protection and length regulation and proteins such as Cdc13 and Ku that target telomerase to telomeres and protect telomeres from degradation 2 . Proteins such as Pif1 help regulate telomere length 3 and prevent telomerase from adding telomeres to broken DNAs 3,4 . In telomerase-deficient S. cerevisiae cells telomeres are maintained by recombination 5,6 . Most mammalian cells lack telomerase 7 and have a limited lifespan. Immortalization and cancer progression require increased telomere maintenance capacity, either through upregulation of telomerase activity 7 or through the alternative lengthening of telomere pathway 8 .Recombination and the Tel1 and Mec1 checkpoints differentially effect genome rearrangements driven by telomere dysfunction in yeast L E T T E R S 612VOLUME 36 | NUMBER 6 | JUNE 2004 NATURE GENETICS RDKY5233 is a tlc1∆ type II strain. Additional relevant GCR rates include the tlc1∆ type I strain, RDKY5232 (3.1 × 10 -10 (0.9)); lig4∆ strain, RDKY3641 (1.6 × 10 -9 (9); ref. 10); tel1∆ lig4∆ strain, RDKY5238 (4.2 × 10 -9 (12)); and tel1∆ lig4∆ est2∆ strain, RDKY5240 (3.5 × 10 -9 (10)). ND, not determined.