Specific pathways prevent duplication-mediated genome rearrangements

Ludwig Institute for Cancer Research, Department of Medicine, University of California School of Medicine, San Diego, 9500 Gilman Drive, La Jolla, California 92093-0669, USA.
Nature (Impact Factor: 41.46). 08/2009; 460(7258):984-9. DOI: 10.1038/nature08217
Source: PubMed


We have investigated the ability of different regions of the left arm of Saccharomyces cerevisiae chromosome V to participate in the formation of gross chromosomal rearrangements (GCRs). We found that the 4.2-kilobase HXT13-DSF1 region sharing divergent homology with chromosomes IV, X and XIV, similar to mammalian segmental duplications, was 'at risk' for participating in duplication-mediated GCRs generated by homologous recombination. Numerous genes and pathways, including SGS1, TOP3, RMI1, SRS2, RAD6, SLX1, SLX4, SLX5, MSH2, MSH6, RAD10 and the DNA replication stress checkpoint requiring MRC1 and TOF1, were highly specific for suppressing these GCRs compared to GCRs mediated by single-copy sequences. These results indicate that the mechanisms for formation and suppression of rearrangements occurring in regions containing at-risk sequences differ from those occurring in regions of single-copy sequence. This explains how extensive genome instability is prevented in eukaryotic cells whose genomes contain numerous divergent repeated sequences.

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    • "Slx8 shows an E3 ubiquitin ligase activity and interacts with an E2 ubiquitin conjugating enzyme Ubc4, while Slx5 binds to polysumoylated substrates via its multiple SIMs [11] [12]. Strains lacking SLX5/8 show many features of genomic instability, such as slow growth, increased sensitivity to genotoxins, gross chromosomal rearrangements, increased rates of chromosome loss and recombination [16] [17] [18] [19] [20] [21]. Consequently, the Slx5/8 STUbL is recruited to the sites of replication stress [18] and DNA double strand breaks (DSBs) [21], mediating relocation of irreparable DSBs to the nuclear pore [19]. "
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    ABSTRACT: The Saccharomyces cerevisiae Uls1 belongs to the Swi2/Snf2 family of DNA-dependent ATPases and a new protein family of SUMO-targeted ubiquitin ligases. Here we show that Uls1 is implicated in DNA repair independently of the replication stress response pathways mediated by the endonucleases Mus81 and Yen1 and the helicases Mph1 and Srs2. Uls1 works together with Sgs1 and we demonstrate that the attenuation of replication stress-related defects in sgs1Δ by deletion of ULS1 depends on a functional of Rad51 recombinase and post-replication repair pathway mediated by Rad18 and Rad5, but not on the translesion polymerase, Rev3. The higher resistance of sgs1Δ uls1Δ mutants to genotoxic stress compared to single sgs1Δ cells is not the result of decreased formation or accelerated resolution of recombination-dependent DNA structures. Instead, deletion of ULS1 restores stability of the rDNA region in sgs1Δ cells. Our data suggest that Uls1 may contribute to genomic stability during DNA synthesis and channel the repair of replication lesions into the Sgs1-dependent pathway, with DNA translocase and SUMO binding activities of Uls1 as well as a RING domain being essential for its functions in replication stress response.
    DNA Repair 09/2014; 21:24–35. DOI:10.1016/j.dnarep.2014.05.008 · 3.11 Impact Factor
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    • "Remarkably , deletion of SLX5 and SLX8 also cause increased rates of duplication - mediated rearrangements ( Table 1 , Putnam et al . , 2009 ) , despite the fact that deletion of SLX5 causes an increase in the sumoylation of Mms21 - targets . Moreover , the deletion of SIZ2 , which also increases the sumoylation of Mms21 - targets , causes a greater defect in suppressing duplication - mediated rearrangements alone and in combination with esc2D , slx5D , mms21 - 11 , and mms2"
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    ABSTRACT: Author Summary The human genome contains many “at-risk” sequences that are prone to mutations, including diverse repeated sequences, segmental duplications and regions of copy number variations. Such repetitive sequence elements can cause genome rearrangements through non-allelic homologous recombination (NAHR) and many human diseases are caused by chromosomal rearrangements mediated by NAHR. Here we discovered that Mms21 dependent sumoylation has a major role in suppressing duplication-mediated GCRs. In addition, we showed that mutations of additional genes in sumoylation pathway cause the highest GCR defect known to date, demonstrating a critical role of sumoylation pathway in preventing duplication-mediated GCRs. We further developed a new quantitative proteomics technology to measure sumoylation levels of individual sumoylated proteins on a proteome-wide scale and applied this approach to uncover distinct and overlapping activities of SUMO ligases for substrate sumoylation. We further established the roles of Esc2 and Slx5 in regulating the SUMO proteome. Taken together these findings suggest a model in which a fine balance of Mms21 activity towards its substrates is critical for the suppression of chromosomal rearrangements. Our findings thus have important implications for cancer genetics as well as new insights into the regulation and substrate specificity of protein sumoylation enzymes.
    PLoS Genetics 08/2013; 9(8):e1003670. DOI:10.1371/journal.pgen.1003670 · 7.53 Impact Factor
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    • "Therefore, we conclude that the BIR pathway responsible to complete the bridge and to produce aneuploidies in BIT has a different genetic requirement than the HO-induced BIR, which is characterized by breaks with homeologous or heterologous ends. This conclusion is perfectly in agreement with the recent observations that many HR-GCRs are Pol32-independent and that Pol32 is not required for allelic BIR [54]. Moreover, our data suggest that this HR-induced BIR is suppressed in absence of Sgs1. "
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    ABSTRACT: While in mammalian cells the genetic determinism of chromosomal translocation remains unclear, the yeast Saccharomyces cerevisiae has become an ideal model system to generate ad hoc translocations and analyze their cellular and molecular outcome. A linear DNA cassette carrying a selectable marker flanked by perfect homologies to two chromosomes triggers a bridge-induced translocation (BIT) in budding yeast, with variable efficiency. A postulated two-step process to produce BIT translocants is based on the cooperation between the Homologous Recombination System (HRS) and Break-Induced Replication (BIR); however, a clear indication of the molecular factors underlying the genetic mechanism is still missing. In this work we provide evidence that BIT translocation is elicited by the Rad54 helicase and completed by a Pol32-independent replication pathway. Our results demonstrate also that Rdh54 is involved in the stability of the translocants, suggesting a mitotic role in chromosome pairing and segregation. Moreover, when RAD54 is over-expressed, an ensemble of secondary rearrangements between repeated DNA tracts arise after the initial translocation event, leading to severe aneuploidy with loss of genetic material, which prompts the identification of fragile sites within the yeast genome.
    PLoS ONE 04/2013; 8(4):e60926. DOI:10.1371/journal.pone.0060926 · 3.23 Impact Factor
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