A global view of meiotic double-strand break end resection

Mimitou, Eleni P.; Yamada, Shintaro; Keeney, Scott

doi:10.1126/science.aak9704

Cited by 169 publications

(242 citation statements)

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“…In this model, the size changes are a consequence of aberrant control of DSB numbers along with a change in nucleolytic DSB processing. This model is consistent with detection of a substantial subpopulation of unresected DSBs in Tel1-deficient cells (Mimitou et al 2017). This model also readily accounts for the apparent contribution of Rec114 phosphorylation and for the observation that reducing DSB numbers (by Spo11 +/− heterozygosity) in an Atm −/− background partially suppresses the changes in SPO11-oligo sizes (Lange et al 2011).…”

Section: Tel1 and Dsb Processingsupporting

confidence: 69%

“…One model to explain these findings is that Tel1 acts early in meiotic DSB processing. Indeed, Tel1 phosphorylates proteins necessary for Spo11 removal from DNA ends, such as Mre11 and Sae2 (Cartagena-Lirola et al 2006;Terasawa et al 2008), and Tel1 is required for both the efficient initiation of DSB resection and the generation of normal-length exonucleolytic resection tracts (Mimitou et al 2017). Thus, altered oligo sizes may reflect changes in the positions of Mre11-dependent endonucleolytic cleavage (Cannavo and Cejka 2014) and/or decreased 3 ′ →5 ′ exonuclease activity of Mre11 (Garcia et al 2011).…”

Section: Tel1 and Dsb Processingmentioning

confidence: 99%

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Numerical and spatial patterning of yeast meiotic DNA breaks by Tel1

2016

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The Spo11-generated double-strand breaks (DSBs) that initiate meiotic recombination are dangerous lesions that can disrupt genome integrity, so meiotic cells regulate their number, timing, and distribution. Mechanisms of this regulation remain poorly understood. Here, we use Spo11-oligonucleotide complexes, a byproduct of DSB formation, to reveal aspects of the contribution of the Saccharomyces cerevisiae DNA damage-responsive kinase Tel1 (ortholog of mammalian ATM). A tel1Δ mutant has globally increased amounts of Spo11-oligonucleotide complexes and altered Spo11-oligonucleotide lengths, consistent with conserved roles for Tel1 in control of DSB number and processing. A kinase-dead tel1 mutation similarly increases Spo11-oligonucleotide levels but mutating known Tel1 phosphotargets on Hop1 and Rec114 does not, implicating Tel1 kinase activity and clarifying roles of Tel1 phosphorylation substrates. Deep sequencing of Spo11 oligonucleotides demonstrates that Tel1 shapes the genome-wide DSB landscape in unexpected ways. Early in meiosis, Tel1 absence causes widespread changes in DSB distributions across large chromosomal domains. Many of these changes are erased as meiosis proceeds, however, illustrating homeostatic behavior of DSB regulatory systems. We further find that effects of Tel1 are distinct but partially overlapping with previously described contributions of the recombination regulator Cst9 (also known as Zip3). Finally, we provide evidence indicating that Tel1-dependent DSB interference influences the population-average DSB landscape but also demonstrate that locally inhibitory effects of an artificial hotspot insertion can be both Tel1-independent and chromosomal context-dependent. Our findings delineate Tel1 roles in regulating number and location of DSBs and illuminate the complex interplay between Tel1 and other pathways for DSB control.

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Section: Tel1 and Dsb Processingsupporting

confidence: 69%

Section: Tel1 and Dsb Processingmentioning

confidence: 99%

Numerical and spatial patterning of yeast meiotic DNA breaks by Tel1

2016

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“…The sensitivity of the 3′ ssDNA of resected DSBs to ssDNA-specific endonucleases provides a tool to map resection endpoints by sequencing the region where ssDNA transitions to dsDNA (Figure 1) (Mimitou et al, 2017). We isolate genomic DNA of meiotic cells in low-melting point agarose plugs to protect the DNA from random shearing.…”

Section: Overview Of S1-seqmentioning

confidence: 99%

“…We have shown that the sequential treatment with S1 nuclease and T4 DNA polymerase can degrade the 2-nt 5′ overhang of unresected DSBs to which Spo11 is still covalently bound (Mimitou et al, 2017). We therefore reasoned that S1-seq can provide a facile method for nucleotide-resolution mapping of the unresected Spo11 DSBs that accumulate in sae2Δ or rad50S mutants (Alani, Padmore, & Kleckner, 1990; McKee & Kleckner, 1997; Prinz, Amon, & Klein, 1997).…”

Section: Adaptations Of S1-seqmentioning

confidence: 99%

“…Mre11 then degrades the nicked strand towards the break via its 3′→5′ exonuclease activity, releasing Spo11 bound to short oligos (Garcia, Phelps, Gray, & Neale, 2011; Zakharyevich et al, 2010). The Mre11 nicks also constitute entry points for the 5′→3′ exonuclease, Exo1, which extends resection to an average of 822 nt (Mimitou, Yamada, & Keeney, 2017; Zakharyevich et al, 2010). During recombination in vegetative cells, a parallel pathway resects DSBs redundantly with Exo1, and depends on the concerted action of the Sgs1–Top3–Rmi1 complex andthe Dna2 endonuclease (Mimitou & Symington, 2008; Zhu, Chung, Shim, Lee, & Ira, 2008).…”

Section: Introductionmentioning

confidence: 99%

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S1-seq Assay for Mapping Processed DNA Ends

Mimitou

Keeney

2018

Methods in Enzymology

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During meiosis, the specialized cell division giving rise to gametes, numerous DNA double-strand breaks (DSBs) are introduced at multiple places throughout the genome by the topoisomerase-like protein Spo11. Homologous recombination, a highly-conserved DSB repair pathway, is employed for their repair and ensures the formation of chiasmata and the proper segregation of homologous chromosomes. In the initial steps of recombination, end resection takes place, wherein Spo11 is endonucleolytically released and the 5′-terminal strands of each DSB are exonucleolytically processed, exposing the ssDNA necessary to identify a homologous repair template. DNA removed by DSB processing is reconstituted by DNA synthesis, which copies genetic information from the intact homologous template. We developed a next-generation sequencing assay, termed S1-seq, to study DNA end resection genome-wide at high spatial resolution during yeast meiotic recombination. The assay relies on the fact that removal of the ssDNA tails of resected DSBs marks the position where resection stopped. Molecular features of resection are revealed by sequencing of these ssDNA-to-dsDNA junctions and comparison to high-resolution Spo11 DSB maps. We describe the experimental and computational methods for S1-seq as applied to meiosis in the SK1 strain of budding yeast Saccharomyces cerevisiae, and discuss how it can also be applied to map DSBs and recombination intermediates.

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Meiotic Recombination Pathways

Reitz¹

2020

Encyclopedia of Life Sciences

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During meiosis, one round of DNA replication is followed by two rounds of division to create the gametes that are obligatory for biparental reproduction. To achieve the reductional segregation that is required during the first meiotic division, most eukaryotes utilise a program that involves the deliberate formation of DNA double‐stranded breaks in their genomes, followed by regulated homologous recombination and reciprocal exchange of genetic material between homologous chromosomes. This reciprocal exchange of genetic information physically links the homologous chromosomes to one another and provides the tension that is necessary for their segregation. Meiotic recombination is highly regulated to ensure that at least one reciprocal crossover event occurs between each pair of homologous chromosomes. Key Concepts Meiotic recombination physically links the homologous chromosomes to one another, creating the tension that is required for their segregation. Meiosis I is a reductional segregation that halves the ploidy of the cell. This process requires bi‐orientation of the homologous chromosomes but mono‐orientation of the sister chromatids, a feat that is achieved through structural modification of the sister chromatid centromeres by monopolin. Directed homologous recombination physically links the homologous chromosomes to one another, through preferential use of the homologous chromosome as the repair template, rather than the sister chromatid, followed by resolution of the recombination intermediate into a crossover. DNA double‐stranded break fate is decided early in the meiotic recombination pathway, with crossovers exclusively arising through a double‐Holliday junction intermediate and non‐crossovers primarily being formed through the synthesis‐dependent strand annealing pathway. Formation of the synaptonemal complex, a proteinaceous structure that assembles between homologous chromosomes during meiosis, is interdependent with meiotic recombination.

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A global view of meiotic double-strand break end resection

Cited by 169 publications

References 56 publications

Numerical and spatial patterning of yeast meiotic DNA breaks by Tel1

Numerical and spatial patterning of yeast meiotic DNA breaks by Tel1

S1-seq Assay for Mapping Processed DNA Ends

Meiotic Recombination Pathways

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