Saccharomyces cerevisiae as a Model System To Define the Chromosomal Instability Phenotype

Abstract

Translocations, deletions, and chromosome fusions are frequent events seen in cancers with genome instability. Here we analyzed 358 genome rearrangements generated in Saccharomyces cerevisiae selected by the loss of the nonessential terminal segment of chromosome V. The rearrangements appeared to be generated by both nonhomologous end joining and homologous recombination and targeted all chromosomes. Fifteen percent of the rearrangements occurred independently more than once. High levels of specific classes of rearrangements were isolated from strains with specific mutations: translocations to Ty elements were increased in telomerasedefective mutants, potential dicentric translocations and dicentric isochromosomes were associated with cell cycle checkpoint defects, chromosome fusions were frequent in strains with both telomerase and cell cycle checkpoint defects, and translocations to homolog genes were seen in strains with defects allowing homoeologous recombination. An analysis of human cancer-associated rearrangements revealed parallels to the effects that strain genotypes have on classes of rearrangement in S. cerevisiae.The development and progression of cancer are correlated with genetic instability. These genomic changes are associated with either a microsatellite instability (MSI) phenotype, involving defects in mismatch repair and a dramatic increase in base substitution and insertion/deletion mutation rates, or a chromosomal instability (CIN) phenotype, involving changes in chromosome number and structure (reviewed in reference 55); however, some tumors have been observed with both MSI and CIN. A causal role for CIN in tumorigenesis is still debated; however, most cancers are associated with dramatic changes to the chromosomal complement (68), and several hereditary cancer predisposition syndromes are closely linked to CIN (49). Using the mutator hypothesis (61), it has been argued that changes in gene dosage, a loss of heterozygosity, a deregulation of gene expression, and the generation of gain-of-function protein chimeras due to CIN are sufficient to drive tumorigenesis in many cases, even without mutations in oncogenes or tumor suppressor genes (29). Thus, understanding CIN is likely to provide some insight into the mechanisms of tumorigenesis.One of the major barriers to understanding the CIN phenotype, even for model organisms such as the yeast Saccharomyces cerevisiae, is that there are very few genetic systems capable of systematic characterizations of genome rearrangements (49, 105). In principle, rearrangements can arise through both homologous recombination (HR) and nonhomologous end-joining (NHEJ) pathways. In practice, the lack of a detailed understanding of the genetic and biochemical mechanisms that underlie these types of rearrangements in vivo has remained a bottleneck to understanding the generation of genome rearrangements in cancer and other human diseases.An assay designed to analyze the formation of translocations and other gross chromosomal rearrangements (GCRs) was developed wit...

Mapping genomic hotspots of DNA damage by a single-strand-DNA-compatible and strand-specific ChIP-seq method

“…One important method in this category is the gross chromosomal rearrangement (GCR) assay developed by the Kolodner laboratory, which is based on the selection against two markers near the left end of chromosome V in budding yeast (Chen and Kolodner 1999). The rearrangement breakpoints in hundreds of GCR-containing strains have been mapped, although doing such mapping one strain at a time is a time-and cost-consuming endeavor (Putnam et al 2005;Chan and Kolodner 2012).…”

Break-induced replication: What is it and what is it for?

“…Most of the translocation events described are mediated by no homology at the break point or very short sequence homologies and are dependent on DNL4, an essential component of the NHEJ pathway. 37 In the absence of both RAD52 and DNL4 the translocation class of GCRs is eliminated. An unusual class of translocations is detected between highly diverged sequences in the absence of the Sgs1 helicase and at least one other protein involved either in the DNA damage checkpoint response, chromatin assembly or the DNA unwinding.…”

Gene Copy-Number Variation in Haploid and Diploid Strains of the Yeast Saccharomyces cerevisiae

“…For both the translocations and interstitial deletions, the rate of events in a wild-type strain is very low (,10 29 /division); the breakpoints of the events occur between CAN1 and PCM1 and typically involved micro-or no sequence homology. Most of these rearrangements reflect nonhomologous end-joining (NHEJ) events, although homologous-recombination (HR) events involving very small homologies are also observed (Putnam et al 2005). More recent versions of the GCR assay showed that the introduction of homology either from a dispersed repeated gene family member (HXT13) or from a Ty element inserted between CAN1 and PCM1 elevated the rate of URA3-CAN1 codeletions by two to three orders of magnitude.…”