Double-strand-break-induced homologous recombination in mammalian cells

2001 ◽  
Vol 29 (2) ◽  
pp. 196-201 ◽  
Author(s):  
R. D. Johnson ◽  
M. Jasin
Keyword(s):  
Homologous Recombination ◽  
Sister Chromatid ◽  
Mammalian Cells ◽  
Indirect Evidence ◽  
Strand Break ◽  
Double Strand Breaks ◽  
Crossing Over ◽  
Dna Double Strand Breaks ◽  
Dsb Repair ◽  
Strand Breaks

In mammalian cells, the repair of DNA double-strand breaks (DSBs) occurs by both homologous and non-homologous mechanisms. Indirect evidence, including that from gene targeting and random integration experiments, had suggested that non-homologous mechanisms were significantly more frequent than homologous ones. However, more recent experiments indicate that homologous recombination is also a prominent DSB repair pathway. These experiments show that mammalian cells use homologous sequences located at multiple positions throughout the genome to repair a DSB. However, template preference appears to be biased, with the sister chromatid being preferred by 2–3 orders of magnitude over a homologous or heterologous chromosome. The outcome of homologous recombination in mammalian cells is predominantly gene conversion that is not associated with crossing-over. The preference for the sister chromatid and the bias against crossing-over seen in mitotic mammalian cells may have developed in order to reduce the potential for genome alterations that could occur when other homologous repair templates are utilized. In attempts to understand further the mechanism of homologous recombination, the proteins that promote this process are beginning to be identified. To date, four mammalian proteins have been demonstrated conclusively to be involved in DSB repair by homologous recombination: Rad54, XRCC2, XRCC3 and BRCAI. This paper summarizes results from a number of recent studies.

scholarly journals SPO16 binds SHOC1 to promote homologous recombination and crossing-over in meiotic prophase I

2019 ◽  
Vol 5 (1) ◽  
pp. eaau9780 ◽  
Author(s):  
Qianting Zhang ◽  
Shu-Yan Ji ◽  
Kiran Busayavalasa ◽  
Chao Yu
Keyword(s):  
Homologous Recombination ◽  
Meiotic Recombination ◽  
Double Strand Breaks ◽  
Crossing Over ◽  
Dna Double Strand Breaks ◽  
Dsb Repair ◽  
Strand Breaks ◽  
Homologous Chromosomes ◽  
Recombination Nodules ◽  
Evolutionarily Conserved

Segregation of homologous chromosomes in meiosis I is tightly regulated by their physical links, or crossovers (COs), generated from DNA double-strand breaks (DSBs) through meiotic homologous recombination. In budding yeast, three ZMM (Zip1/2/3/4, Mer3, Msh4/5) proteins, Zip2, Zip4, and Spo16, form a “ZZS” complex, functioning to promote meiotic recombination via a DSB repair pathway. Here, we identified the mammalian ortholog of Spo16, termed SPO16, which interacts with the mammalian ortholog of Zip2 (SHOC1/MZIP2), and whose functions are evolutionarily conserved to promote the formation of COs. SPO16 localizes to the recombination nodules, as SHOC1 and TEX11 do. SPO16 is required for stabilization of SHOC1 and proper localization of other ZMM proteins. The DSBs formed in SPO16-deleted meiocytes were repaired without COs formation, although synapsis is less affected. Therefore, formation of SPO16-SHOC1 complex–associated recombination intermediates is a key step facilitating meiotic recombination that produces COs from yeast to mammals.

scholarly journals Resection is responsible for loss of transcription around a double-strand break in Saccharomyces cerevisiae

eLife ◽  
2015 ◽  
Vol 4 ◽  
Author(s):  
Nicola Manfrini ◽  
Michela Clerici ◽  
Maxime Wery ◽  
Chiara Vittoria Colombo ◽  
Marc Descrimes ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Homologous Recombination ◽  
Transcriptional Silencing ◽  
Strand Break ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
Dsb Repair ◽  
Strand Breaks ◽  
Transcriptional Inhibition ◽  
In Cis

Emerging evidence indicate that the mammalian checkpoint kinase ATM induces transcriptional silencing in cis to DNA double-strand breaks (DSBs) through a poorly understood mechanism. Here we show that in Saccharomyces cerevisiae a single DSB causes transcriptional inhibition of proximal genes independently of Tel1/ATM and Mec1/ATR. Since the DSB ends undergo nucleolytic degradation (resection) of their 5′-ending strands, we investigated the contribution of resection in this DSB-induced transcriptional inhibition. We discovered that resection-defective mutants fail to stop transcription around a DSB, and the extent of this failure correlates with the severity of the resection defect. Furthermore, Rad9 and generation of γH2A reduce this DSB-induced transcriptional inhibition by counteracting DSB resection. Therefore, the conversion of the DSB ends from double-stranded to single-stranded DNA, which is necessary to initiate DSB repair by homologous recombination, is responsible for loss of transcription around a DSB in S. cerevisiae.

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 RAP80 suppresses the vulnerability of R-loops during DNA double-strand break repair

2021 ◽  
Author(s):  
Takaaki Yasuhara ◽  
Reona Kato ◽  
Motohiro Yamauchi ◽  
Yuki Uchihara ◽  
Lee Zou ◽  
...  
Keyword(s):  
Active Regions ◽  
Strand Break ◽  
Double Strand Breaks ◽  
Critical Component ◽  
Dna Double Strand Breaks ◽  
End Joining ◽  
Dsb Repair ◽  
Strand Breaks ◽  
Chromosome Translocations ◽  
Dna Double Strand Break

AbstractR-loops, consisting of ssDNA and DNA-RNA hybrids, are potentially vulnerable unless they are appropriately processed. Recent evidence suggests that R-loops can form in the proximity of DNA double-strand breaks (DSBs) within transcriptionally active regions. Yet, how the vulnerability of R-loops is overcome during DSB repair remains unclear. Here, we identify RAP80 as a factor suppressing the vulnerability of ssDNA in R-loops and chromosome translocations and deletions during DSB repair. Mechanistically, RAP80 prevents unscheduled nucleolytic processing of ssDNA in R-loops by CtIP. This mechanism promotes efficient DSB repair via transcription-associated end-joining dependent on BRCA1, Polθ, and LIG1/3. Thus, RAP80 suppresses the vulnerability of R-loops during DSB repair, thereby precluding genomic abnormalities in a critical component of the genome caused by deleterious R-loop processing.

scholarly journals Hedgehog signaling enables repair of ribosomal DNA double-strand breaks

2020 ◽  
Vol 48 (18) ◽  
pp. 10342-10352
Author(s):  
Tshering D Lama-Sherpa ◽  
Victor T G Lin ◽  
Brandon J Metge ◽  
Shannon E Weeks ◽  
Dongquan Chen ◽  
...  
Keyword(s):  
Cancer Cells ◽  
Ribosomal Dna ◽  
Hedgehog Signaling ◽  
Strand Break ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
Dsb Repair ◽  
Strand Breaks ◽  
Rdna Sequences ◽  
Polymerase I

Abstract Ribosomal DNA (rDNA) consists of highly repeated sequences that are prone to incurring damage. Delays or failure of rDNA double-strand break (DSB) repair are deleterious, and can lead to rDNA transcriptional arrest, chromosomal translocations, genomic losses, and cell death. Here, we show that the zinc-finger transcription factor GLI1, a terminal effector of the Hedgehog (Hh) pathway, is required for the repair of rDNA DSBs. We found that GLI1 is activated in triple-negative breast cancer cells in response to ionizing radiation (IR) and localizes to rDNA sequences in response to both global DSBs generated by IR and site-specific DSBs in rDNA. Inhibiting GLI1 interferes with rDNA DSB repair and impacts RNA polymerase I activity and cell viability. Our findings tie Hh signaling to rDNA repair and this heretofore unknown function may be critically important in proliferating cancer cells.

scholarly journals Homologous recombination preferentially repairs heat-induced DNA double-strand breaks in mammalian cells

2016 ◽  
Vol 33 (3) ◽  
pp. 336-342 ◽  
Author(s):  
Akihisa Takahashi ◽  
Eiichiro Mori ◽  
Yosuke Nakagawa ◽  
Atsuhisa Kajihara ◽  
Tadaaki Kirita ◽  
...  
Keyword(s):  
Homologous Recombination ◽  
Mammalian Cells ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
Strand Breaks

scholarly journals RAD51 Is Required for the Repair of Plasmid Double-Stranded DNA Gaps from Either Plasmid or Chromosomal Templates

2000 ◽  
Vol 20 (4) ◽  
pp. 1194-1205 ◽  
Author(s):  
Stephan Bärtsch ◽  
Leslie E. Kang ◽  
Lorraine S. Symington
Keyword(s):  
Homologous Recombination ◽  
Double Strand Breaks ◽  
Crossing Over ◽  
Dna Double Strand Breaks ◽  
End Joining ◽  
Radiation Chemical ◽  
Strand Breaks ◽  
Independent Events ◽  
In The Wild ◽  
A Minor

ABSTRACT DNA double-strand breaks may be induced by endonucleases, ionizing radiation, chemical agents, and mechanical forces or by replication of single-stranded nicked chromosomes. Repair of double-strand breaks can occur by homologous recombination or by nonhomologous end joining. A system was developed to measure the efficiency of plasmid gap repair by homologous recombination using either chromosomal or plasmid templates. Gap repair was biased toward gene conversion events unassociated with crossing over using either donor sequence. The dependence of recombinational gap repair on genes belonging to the RAD52epistasis group was tested in this system. RAD51,RAD52, RAD57, and RAD59 were required for efficient gap repair using either chromosomal or plasmid donors. No homologous recombination products were recovered fromrad52 mutants, whereas a low level of repair occurred in the absence of RAD51, RAD57, orRAD59. These results suggest a minor pathway of strand invasion that is dependent on RAD52 but not onRAD51. The residual repair events in rad51mutants were more frequently associated with crossing over than was observed in the wild-type strain, suggesting that the mechanisms forRAD51-dependent and RAD51-independent events are different. Plasmid gap repair was reduced synergistically inrad51 rad59 double mutants, indicating an important role for RAD59 in RAD51-independent repair.

Detecting, signalling and repairing DNA double-strand breaks

2001 ◽  
Vol 29 (6) ◽  
pp. 655-661 ◽  
Author(s):  
S. P. Jackson
Keyword(s):  
Mammalian Cells ◽  
Molecular Mechanisms ◽  
Double Strand Breaks ◽  
Model Organisms ◽  
Dna Double Strand Breaks ◽  
Dsb Repair ◽  
Site Specific ◽  
Strand Breaks ◽  
Dna Dsbs ◽  
Recombination Processes

DNA double-strand breaks (DSBs) can be generated by a variety of genotoxic agents, including ionizing radiation and radiomimetic chemicals. They can also occur when DNA replication complexes encounter other forms of DNA damage, and are produced as intermediates during certain site-specific recombination processes. It is crucial that cells recognize DSBs and bring about their efficient repair, because a single unrepaired cellular DSB can induce cell death, and defective DSB repair can lead to mutations or the loss of significant segments of chromosomal material. Eukaryotic cells have evolved a variety of systems to detect DNA DSBs, repair them, and signal their presence to the transcription, cell cycle and apoptotic machineries. In this review, I describe how work on mammalian cells and also on model organisms such as yeasts has revelaed that such systems are highly conserved throughout evolution, and has provided insights into the molecular mechanisms by which DNA DSBs are recognized, signalled and repaired. I also explain how defects in the proteins that function in these pathways are associated with a variety of human pathological states.

Regulation of the cellular DNA double-strand break response

2007 ◽  
Vol 85 (6) ◽  
pp. 663-674 ◽  
Author(s):  
Kendra L. Cann ◽  
Geoffrey G. Hicks
Keyword(s):  
Mammalian Cells ◽  
Strand Break ◽  
Double Strand Break ◽  
Double Strand Breaks ◽  
Cell Cycle Checkpoints ◽  
Dna Double Strand Breaks ◽  
End Joining ◽  
Strand Breaks ◽  
Dna Double Strand Break ◽  
Cellular Dna

DNA double-strand breaks occur frequently in cycling cells, and are also induced by exogenous sources, including ionizing radiation. Cells have developed integrated double-strand break response pathways to cope with these lesions, including pathways that initiate DNA repair (either via homologous recombination or nonhomologous end joining), the cell-cycle checkpoints (G1–S, intra-S phase, and G2–M) that provide time for repair, and apoptosis. However, before any of these pathways can be activated, the damage must first be recognized. In this review, we will discuss how the response of mammalian cells to DNA double-strand breaks is regulated, beginning with the activation of ATM, the pinnacle kinase of the double-strand break signalling cascade.

scholarly journals Repair of site-specific double-strand breaks in a mammalian chromosome by homologous and illegitimate recombination.

1997 ◽  
Vol 17 (1) ◽  
pp. 267-277 ◽  
Author(s):  
R G Sargent ◽  
M A Brenneman ◽  
J H Wilson
Keyword(s):  
Homologous Recombination ◽  
Mammalian Cells ◽  
Repair Process ◽  
Strand Break ◽  
Double Strand Breaks ◽  
Illegitimate Recombination ◽  
Loss Of Function ◽  
Yeast Saccharomyces Cerevisiae ◽  
Site Specific ◽  
Strand Breaks

In mammalian cells, chromosomal double-strand breaks are efficiently repaired, yet little is known about the relative contributions of homologous recombination and illegitimate recombination in the repair process. In this study, we used a loss-of-function assay to assess the repair of double-strand breaks by homologous and illegitimate recombination. We have used a hamster cell line engineered by gene targeting to contain a tandem duplication of the native adenine phosphoribosyltransferase (APRT) gene with an I-SceI recognition site in the otherwise wild-type APRT+ copy of the gene. Site-specific double-strand breaks were induced by intracellular expression of I-SceI, a rare-cutting endonuclease from the yeast Saccharomyces cerevisiae. I-SceI cleavage stimulated homologous recombination about 100-fold; however, illegitimate recombination was stimulated more than 1,000-fold. These results suggest that illegitimate recombination is an important competing pathway with homologous recombination for chromosomal double-strand break repair in mammalian cells.

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