scholarly journals A global view of meiotic double-strand break end resection

Science ◽  
2017 ◽  
Vol 355 (6320) ◽  
pp. 40-45 ◽  
Author(s):  
Eleni P. Mimitou ◽  
Shintaro Yamada ◽  
Scott Keeney
Keyword(s):  
Meiotic Recombination ◽  
Strand Break ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Naked Dna ◽  
Global View ◽  
Genome Wide ◽  
End Resection

DNA double-strand breaks that initiate meiotic recombination are exonucleolytically processed. This 5′→3′ resection is a central, conserved feature of recombination but remains poorly understood. To address this lack, we mapped resection endpoints genome-wide at high resolution inSaccharomyces cerevisiae. Full-length resection requires Exo1 exonuclease and the DSB-responsive kinase Tel1, but not Sgs1 helicase. Tel1 also promotes efficient and timely resection initiation. Resection endpoints display pronounced heterogeneity between genomic loci that reflects a tendency for nucleosomes to block Exo1, yet Exo1 also appears to digest chromatin with high processivity and at rates similar to naked DNA in vitro. This paradox points to nucleosome destabilization or eviction as a defining feature of the meiotic resection landscape.

scholarly journals A global view of meiotic double-strand break end resection

2016 ◽  
Author(s):  
Eleni P. Mimitou ◽  
Shintaro Yamada ◽  
Scott Keeney
Keyword(s):  
Strand Break ◽  
Dna Double Strand Breaks ◽  
Timely Initiation ◽  
Strand Breaks ◽  
Naked Dna ◽  
Genome Wide ◽  
Sequencing Method ◽  
End Resection

AbstractThe DNA double-strand breaks that initiate homologous recombination during meiosis are subject to extensive 5′→3′ exonucleolytic processing. This resection is a central and conserved feature of recombination, yet its mechanism is poorly understood. Using a purpose-made deep-sequencing method, we mapped meiotic resection endpoints genome-wide at high spatial resolution inSaccharomyces cerevisiae. Generating full-length resection tracts requires Exo1 exonuclease activity and the DNA-damage responsive kinase Tel1, but not the helicase Sgs1. Tel1 is also required for efficient and timely initiation of resection. We find that distributions of resection endpoints at individual genomic loci display pronounced heterogeneity that reflects a tendency for nucleosomes to block Exo1 in vivo, yet modeling experiments indicate that Exo1 digests chromatin with high apparent processivity and at rates approaching those for naked DNA in vitro. This paradox points to nucleosome destabilization or eviction as a determining feature of the meiotic resection landscape.

scholarly journals End-resection at DNA double-strand breaks in the three domains of life

2013 ◽  
Vol 41 (1) ◽  
pp. 314-320 ◽  
Author(s):  
John K. Blackwood ◽  
Neil J. Rzechorzek ◽  
Sian M. Bray ◽  
Joseph D. Maman ◽  
Luca Pellegrini ◽  
...  
Keyword(s):  
Dna Repair ◽  
Homologous Recombination ◽  
Strand Break ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Homologous Chromosomes ◽  
Domains Of Life ◽  
Strand Invasion ◽  
End Resection

During DNA repair by HR (homologous recombination), the ends of a DNA DSB (double-strand break) must be resected to generate single-stranded tails, which are required for strand invasion and exchange with homologous chromosomes. This 5′–3′ end-resection of the DNA duplex is an essential process, conserved across all three domains of life: the bacteria, eukaryota and archaea. In the present review, we examine the numerous and redundant helicase and nuclease systems that function as the enzymatic analogues for this crucial process in the three major phylogenetic divisions.

scholarly journals i-BLESS is an ultra-sensitive method for detection of DNA double-strand breaks

2018 ◽  
Vol 1 (1) ◽  
Author(s):  
Anna Biernacka ◽  
Yingjie Zhu ◽  
Magdalena Skrzypczak ◽  
Romain Forey ◽  
Benjamin Pardo ◽  
...  
Keyword(s):  
Cell Fate ◽  
Genome Stability ◽  
Strand Break ◽  
Double Strand Breaks ◽  
Universal Method ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Agarose Beads ◽  
Genome Wide ◽  
Dna Double Strand Break

AbstractMaintenance of genome stability is a key issue for cell fate that could be compromised by chromosome deletions and translocations caused by DNA double-strand breaks (DSBs). Thus development of precise and sensitive tools for DSBs labeling is of great importance for understanding mechanisms of DSB formation, their sensing and repair. Until now there has been no high resolution and specific DSB detection technique that would be applicable to any cells regardless of their size. Here, we present i-BLESS, a universal method for direct genome-wide DNA double-strand break labeling in cells immobilized in agarose beads. i-BLESS has three key advantages: it is the only unbiased method applicable to yeast, achieves a sensitivity of one break at a given position in 100,000 cells, and eliminates background noise while still allowing for fixation of samples. The method allows detection of ultra-rare breaks such as those forming spontaneously at G-quadruplexes.

scholarly journals Identification of regulators of poly-ADP-ribose polymerase (PARP) inhibitor response through complementary CRISPR knockout and activation screens

2019 ◽  
Author(s):  
Kristen E. Clements ◽  
Anastasia Hale ◽  
Nathanial J. Tolman ◽  
Claudia M. Nicolae ◽  
Anchal Sharma ◽  
...  
Keyword(s):  
Histone Acetyltransferase ◽  
Positive Response ◽  
Parp Inhibitor ◽  
Strand Break ◽  
Double Strand Breaks ◽  
Repair Pathway ◽  
Genetic Determinants ◽  
Strand Breaks ◽  
Genome Wide ◽  
End Resection

AbstractInhibitors of poly-ADP-ribose polymerase 1 (PARPi) are highly effective in killing cells deficient in the homologous recombination (HR) DNA repair pathway, such as those lacking BRCA2. In light of this, PARPi have been utilized in recent years to treat BRCA2-mutant tumors, with many patients deriving impressive clinical benefit. However, positive response to PARPi is not universal, even among patients with HR-deficient tumors. Here, we present the results of three genome-wide CRISPR knockout and activation screens which provide an unbiased look at genetic determinants of PARPi response in wildtype or BRCA2-knockout cells. Strikingly, we reveal that depletion of the histone acetyltransferase TIP60, a top hit from our screens, robustly reverses the PARPi sensitivity caused by BRCA2 deficiency. Mechanistically, we show that TIP60 depletion rewires double strand break repair in BRCA2-deficient cells by promoting 53BP1 binding to double strand breaks to suppress end resection. Our work provides a comprehensive set of putative biomarkers that serve to better understand and predict PARPi response, and identifies a novel pathway of PARPi resistance in BRCA2-deficient cells.

scholarly journals Genome-Wide Redistribution of Meiotic Double-Strand Breaks in Saccharomyces cerevisiae

2006 ◽  
Vol 27 (5) ◽  
pp. 1868-1880 ◽  
Author(s):  
Nicolas Robine ◽  
Norio Uematsu ◽  
Franck Amiot ◽  
Xavier Gidrol ◽  
Emmanuel Barillot ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Meiotic Recombination ◽  
Binding Sites ◽  
Chromosomal Region ◽  
Double Strand Breaks ◽  
Dna Binding Domain ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Genome Wide ◽  
Local Stimulation

ABSTRACT Meiotic recombination is initiated by the formation of programmed DNA double-strand breaks (DSBs) catalyzed by the Spo11 protein. DSBs are not randomly distributed along chromosomes. To better understand factors that control the distribution of DSBs in budding yeast, we have examined the genome-wide binding and cleavage properties of the Gal4 DNA binding domain (Gal4BD)-Spo11 fusion protein. We found that Gal4BD-Spo11 cleaves only a subset of its binding sites, indicating that the association of Spo11 with chromatin is not sufficient for DSB formation. In centromere-associated regions, the centromere itself prevents DSB cleavage by tethered Gal4BD-Spo11 since its displacement restores targeted DSB formation. In addition, we observed that new DSBs introduced by Gal4BD-Spo11 inhibit surrounding DSB formation over long distances (up to 60 kb), keeping constant the number of DSBs per chromosomal region. Together, these results demonstrate that the targeting of Spo11 to new chromosomal locations leads to both local stimulation and genome-wide redistribution of recombination initiation and that some chromosomal regions are inherently cold regardless of the presence of Spo11.

scholarly journals Mouse TEX15 is essential for DNA double-strand break repair and chromosomal synapsis during male meiosis

2008 ◽  
Vol 180 (4) ◽  
pp. 673-679 ◽  
Author(s):  
Fang Yang ◽  
Sigrid Eckardt ◽  
N. Adrian Leu ◽  
K. John McLaughlin ◽  
Peijing Jeremy Wang
Keyword(s):  
Meiotic Recombination ◽  
Strand Break ◽  
Male Meiosis ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Homologous Chromosomes ◽  
Recombination Proteins ◽  
Meiotic Chromosomes ◽  
Dna Double Strand Break

During meiosis, homologous chromosomes undergo synapsis and recombination. We identify TEX15 as a novel protein that is required for chromosomal synapsis and meiotic recombination. Loss of TEX15 function in mice causes early meiotic arrest in males but not in females. Specifically, TEX15-deficient spermatocytes exhibit a failure in chromosomal synapsis. In mutant spermatocytes, DNA double-strand breaks (DSBs) are formed, but localization of the recombination proteins RAD51 and DMC1 to meiotic chromosomes is severely impaired. Based on these data, we propose that TEX15 regulates the loading of DNA repair proteins onto sites of DSBs and, thus, its absence causes a failure in meiotic recombination.

scholarly journals Phytotherapeutics oridonin and ponicidin show additive effects combined with irradiation in pancreatic cancer in vitro

2017 ◽  
Vol 51 (4) ◽  
pp. 407-414 ◽  
Author(s):  
Jakob Liermann ◽  
Patrick Naumann ◽  
Franco Fortunato ◽  
Thomas E. Schmid ◽  
Klaus-Josef Weber ◽  
...  
Keyword(s):  
Pancreatic Cancer ◽  
Strand Break ◽  
Double Strand Break ◽  
Double Strand Breaks ◽  
Double Strand Break Repair ◽  
Additive Effects ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Dna Double Strand Break

Abstract Background Chemoradiation of locally advanced non-metastatic pancreatic cancer can lead to secondary operability by tumor mass reduction. Here, we analyzed radiomodulating effects of oridonin and ponicidin in pancreatic cancer in vitro. Both agents are ent-kaurane diterpenoids, extracted from Isodon rubescens, a plant that is well known in Traditional Chinese Medicine. Cytotoxic effects have recently been shown in different tumor entities for both agents. Materials and methods Pancreatic cancer cell lines AsPC-1, BxPC-3, Panc-1 and MIA PaCa-2 were pretreated with oridonin or ponicidin and irradiated with 2 Gy to 6 Gy. Long-term survival was determined by clonogenic assay. Cell cycle effects and intensity of γH2AX as indicator for DNA double-strand breaks were investigated by flow cytometry. Western blotting was used to study the DNA double-strand break repair proteins Ku70, Ku80 and XRCC4. Results Oridonin and ponicidin lead to a dose-dependent reduction of clonogenic survival and an increase in γH2AX. Combined with irradiation we observed additive effects and a prolonged G2/M-arrest. No relevant changes in the levels of the DNA double-strand break repair proteins were detected. Conclusions Pretreatment with oridonin or ponicidin followed by irradiation lead to an additional reduction in survival of pancreatic cancer cells in vitro, presumably explained by an induced prolonged G2/M-arrest. Both agents seem to induce DNA double-strand breaks but do not interact with the non-homologous end joining (NHEJ) pathway.

scholarly journals Factors influencing meiotic recombination revealed by whole-genome sequencing of single sperm

Science ◽  
2019 ◽  
Vol 363 (6433) ◽  
pp. eaau8861 ◽  
Author(s):  
Anjali Gupta Hinch ◽  
Gang Zhang ◽  
Philipp W. Becker ◽  
Daniela Moralli ◽  
Robert Hinch ◽  
...  
Keyword(s):  
Meiotic Recombination ◽  
Double Strand Breaks ◽  
Pseudoautosomal Region ◽  
Whole Genome ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Molecular Assays ◽  
Factors Influencing ◽  
Genome Wide ◽  
Crossover Probability

Recombination is critical to meiosis and evolution, yet many aspects of the physical exchange of DNA via crossovers remain poorly understood. We report an approach for single-cell whole-genome DNA sequencing by which we sequenced 217 individual hybrid mouse sperm, providing a kilobase-resolution genome-wide map of crossovers. Combining this map with molecular assays measuring stages of recombination, we identified factors that affect crossover probability, including PRDM9 binding on the non-initiating template homolog and telomere proximity. These factors also influence the time for sites of recombination-initiating DNA double-strand breaks to find and engage their homologs, with rapidly engaging sites more likely to form crossovers. We show that chromatin environment on the template homolog affects positioning of crossover breakpoints. Our results also offer insights into recombination in the pseudoautosomal region.

Repair pathway choice for double-strand breaks

2020 ◽  
Vol 64 (5) ◽  
pp. 765-777 ◽  
Author(s):  
Yixi Xu ◽  
Dongyi Xu
Keyword(s):  
Cell Cycle ◽  
Double Strand Breaks ◽  
Homologous Region ◽  
Gene Repair ◽  
Repair Pathway ◽  
Dna Double Strand Breaks ◽  
Environmental Radiation ◽  
Strand Breaks ◽  
Dna End Resection ◽  
End Resection

Abstract Deoxyribonucleic acid (DNA) is at a constant risk of damage from endogenous substances, environmental radiation, and chemical stressors. DNA double-strand breaks (DSBs) pose a significant threat to genomic integrity and cell survival. There are two major pathways for DSB repair: nonhomologous end-joining (NHEJ) and homologous recombination (HR). The extent of DNA end resection, which determines the length of the 3′ single-stranded DNA (ssDNA) overhang, is the primary factor that determines whether repair is carried out via NHEJ or HR. NHEJ, which does not require a 3′ ssDNA tail, occurs throughout the cell cycle. 53BP1 and the cofactors PTIP or RIF1-shieldin protect the broken DNA end, inhibit long-range end resection and thus promote NHEJ. In contrast, HR mainly occurs during the S/G2 phase and requires DNA end processing to create a 3′ tail that can invade a homologous region, ensuring faithful gene repair. BRCA1 and the cofactors CtIP, EXO1, BLM/DNA2, and the MRE11–RAD50–NBS1 (MRN) complex promote DNA end resection and thus HR. DNA resection is influenced by the cell cycle, the chromatin environment, and the complexity of the DNA end break. Herein, we summarize the key factors involved in repair pathway selection for DSBs and discuss recent related publications.

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