Evidence that the S.cerevisiae Sgs1 protein facilitates recombinational repair of telomeres during senescence

Department of Pathology and Laboratory Medicine, University of Pennsylvania School of Medicine, Philadelphia, PA, USA.
Nucleic Acids Research (Impact Factor: 9.11). 02/2006; 34(2):506-16. DOI: 10.1093/nar/gkj452
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RecQ DNA helicases, including yeast Sgs1p and the human Werner and Bloom syndrome proteins, participate in telomere biology, but the underlying mechanisms are not fully understood. Here, we explore the protein sequences and genetic interactors of Sgs1p that function to slow the senescence of telomerase (tlc1) mutants. We find that the S-phase checkpoint function of Sgs1p is dispensable for preventing rapid senescence, but that Sgs1p sequences required for homologous recombination, including the helicase domain and topoisomerase III interaction domain, are essential. sgs1 and rad52 mutations are epistatic during senescence, indicating that Sgs1p participates in a RAD52-dependent recombinational pathway of telomere maintenance. Several mutations that are synthetically lethal with sgs1 mutation and which individually lead to genome instability, including mus81, srs2, rrm3, slx1 and top1, do not speed the senescence of tlc1 mutants, indicating that the rapid senescence of sgs1 tlc1 mutants is not caused by generic genome instability. However, mutations in SLX5 or SLX8, which encode proteins that function together in a complex that is required for viability in sgs1 mutants, do speed the senescence of tlc1 mutants. These observations further define roles for RecQ helicases and related proteins in telomere maintenance.

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Available from: Julia Lee-Soety, Oct 09, 2015
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    • "This function may be necessary to allow alternative repair events, such as break-induced replication or microhomology-mediated recombination . Consistently, type II survivors of telomerase ablation, which survive thanks to imprecise recombination events between telomeric repeats, require Slx5/Slx8 activity [56] [57]. On the other hand, Slx5/ Slx8 was reported to inhibit Rad51-mediated (canonical ) recombination events [55]. "
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    ABSTRACT: The double membrane of the eukaryotic nucleus surrounds the genome, constraining it to a nuclear sphere. Proteins, RNA protein particles and artificial chromosome rings diffuse rapidly and freely throughout the nucleoplasm, while chromosomal loci show subdiffusive movement with varying degrees of constraint. In situ biochemical approaches and live imaging studies have revealed the existence of nuclear subcompartments that are enriched for specific chromatin states and/or enzymatic activities. This sequestration is thought to enhance the formation of heterochromatin, particularly when factors of limited abundance are involved. Implicit in the concept of compartmentation is the idea that chromatin is able to move from one compartment to another. Indeed, in budding yeast, gene activation, repression and the presence of persistent DNA double-strand breaks each has been shown to provoke subnuclear relocalization of chromatin. In some cases, movement has been linked to the action of ATP-dependent chromatin remodeling complexes, more specifically to the Snf2-related ATPase-containing complexes, SWR-C and INO80-C. Here we examine how these multi-subunit remodelers contribute to chromatin-based processes linked to the DNA damage response. We review recent evidence that supports a role for yeast SWR-C and INO80-C in determining the subnuclear position of damaged domains and finally, we recap the multiple ways in which these remodelers contribute to genomic integrity.
    Journal of Molecular Biology 10/2014; 427(3). DOI:10.1016/j.jmb.2014.10.015 · 4.33 Impact Factor
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    • "Accordingly, Mus81 was recently found at the telomeres of human cells that maintain telomeres through a recombination-based mechanism (92). Confirming previous results, deletion of MUS81 did not affect senescence in Control cells (39); (Figure 7A). In contrast, we found that mus81Δ VST cells displayed faster senescence compared with MUS81 VST cells. "
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    ABSTRACT: In the absence of telomerase, telomeres progressively shorten with every round of DNA replication, leading to replicative senescence. In telomerase-deficient Saccharomyces cerevisiae, the shortest telomere triggers the onset of senescence by activating the DNA damage checkpoint and recruiting homologous recombination (HR) factors. Yet, the molecular structures that trigger this checkpoint and the mechanisms of repair have remained elusive. By tracking individual telomeres, we show that telomeres are subjected to different pathways depending on their length. We first demonstrate a progressive accumulation of subtelomeric single-stranded DNA (ssDNA) through 5'-3' resection as telomeres shorten. Thus, exposure of subtelomeric ssDNA could be the signal for cell cycle arrest in senescence. Strikingly, early after loss of telomerase, HR counteracts subtelomeric ssDNA accumulation rather than elongates telomeres. We then asked whether replication repair pathways contribute to this mechanism. We uncovered that Rad5, a DNA helicase/Ubiquitin ligase of the error-free branch of the DNA damage tolerance (DDT) pathway, associates with native telomeres and cooperates with HR in senescent cells. We propose that DDT acts in a length-independent manner, whereas an HR-based repair using the sister chromatid as a template buffers precocious 5'-3' resection at the shortest telomeres.
    Nucleic Acids Research 01/2014; 42(6). DOI:10.1093/nar/gkt1328 · 9.11 Impact Factor
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    • "In fact, Sgs1 not only has a function in the resection of DSBs and telomeres but also has a critical function in processing recombination intermediates that occur at replication fork stalls, as detailed recently (Vanoli et al., 2010). In accordance with this, the deletion of Sgs1 has a deleterious effect on the growth of telomerase-negative cells (Cohen and Sinclair, 2001; Johnson et al., 2001; Azam et al., 2006). "
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    ABSTRACT: In many somatic human tissues, telomeres shorten progressively because of the DNA-end replication problem. Consequently, cells cease to proliferate and are maintained in a metabolically viable state called replicative senescence. These cells are characterized by an activation of DNA damage checkpoints stemming from eroded telomeres, which are bypassed in many cancer cells. Hence, replicative senescence has been considered one of the most potent tumor suppressor pathways. However, the mechanism through which short telomeres trigger this cellular response is far from being understood. When telomerase is removed experimentally in Saccharomyces cerevisiae, telomere shortening also results in a gradual arrest of population growth, suggesting that replicative senescence also occurs in this unicellular eukaryote. In this review, we present the key steps that have contributed to the understanding of the mechanisms underlying the establishment of replicative senescence in budding yeast. As in mammals, signals stemming from short telomeres activate the DNA damage checkpoints, suggesting that the early cellular response to the shortest telomere(s) is conserved in evolution. Yet closer analysis reveals a complex picture in which the apparent single checkpoint response may result from a variety of telomeric alterations expressed in the absence of telomerase. Accordingly, the DNA replication of eroding telomeres appears as a critical challenge for senescing budding yeast cells and the easy manipulation of S. cerevisiae is providing insights into the way short telomeres are integrated into their chromatin and nuclear environments. Finally, the loss of telomerase in budding yeast triggers a more general metabolic alteration that remains largely unexplored. Thus, telomerase-deficient S. cerevisiae cells may have more common points than anticipated with somatic cells, in which telomerase depletion is naturally programed, thus potentially inspiring investigations in mammalian cells.
    Frontiers in Oncology 04/2013; 3:101. DOI:10.3389/fonc.2013.00101
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