scholarly journals Plasmid-mediated induction of recombination in yeast.

Genetics ◽  
1994 ◽  
Vol 137 (1) ◽  
pp. 41-48 ◽  
Author(s):  
R Silberman ◽  
M Kupiec
Keyword(s):  
Dna Damage ◽  
Recombination Frequency ◽  
Strand Break ◽  
Double Strand Break ◽  
Yeast Cells ◽  
Wild Type ◽  
Show Evidence ◽  
Type Information

Abstract Diploid yeast cells heteroallelic at the HIS3 locus were transformed with a minichromosome (centromeric plasmid) carrying homology to the HIS3 region and containing the same two mutations as were present in the chromosomes. When a double-strand break (DSB) was introduced in the region of homology, an increase in the recombination frequency between heteroalleles (leading to His+ cells) was observed, although the plasmid was unable to donate wild-type information. This induction of recombination was dependent on the presence of homology between the plasmid sequences and the chromosomes. We show evidence for the physical involvement of the plasmid in tripartite recombination events, and we propose models that can explain the interactions between the plasmid-borne and chromosomal-borne alleles. Our results suggest that the mitotic induction of recombination by DNA damage is due to localized initiation of recombination events, and not to a general induction of recombination enzymes in the cell.

scholarly journals Loss of BCL-3 sensitises colorectal cancer cells to DNA damage, revealing a role for BCL-3 in double strand break repair by homologous recombination

2021 ◽  
Author(s):  
Christopher Parker ◽  
Adam Christian Chambers ◽  
Dustin Flanagan ◽  
Tracey J Collard ◽  
Greg Ngo ◽  
...  
Keyword(s):  
Dna Damage ◽  
Homologous Recombination ◽  
Cellular Response ◽  
Parp Inhibitor ◽  
Strand Break ◽  
Double Strand Break ◽  
Parp Inhibition ◽  
Adjuvant Chemoradiotherapy ◽  
Wild Type ◽  
Γ Irradiation

Objective: The proto-oncogene BCL-3 is upregulated in a subset of colorectal cancers (CRC) and increased expression of the gene correlates with poor patient prognosis. The aim is to investigate whether inhibiting BCL-3 can increase the response to DNA damage in CRC.Design: The function of BCL-3 in DNA damage response was studied in vitro using siRNA and CRISPR-Cas9 genome editing and in vivo using Bcl3-/- mice. DNA damage induced by γ-irradiation and/or cisplatin was quantified using H2AX and RAD51 foci, repair pathways investigated using HR/NHEJ assays and treatment with the PARP inhibitor olaparib. Result: Suppression of BCL-3 increases double strand break number and decreases homologous recombination in CRC cells, supported by reduced RAD51 foci number and increased sensitivity to PARP inhibition. Importantly, a similar phenotype is seen in Bcl3-/-mice, where the intestinal crypts of these mice exhibit sensitivity to DNA damage and a greater number of double strand breaks compared to wild type mice. FurthermoreApc.Kras-mutant x Bcl3-/- mice exhibit increased DNA damage and reduced RAD51+ cells compared to their wild type counterparts when treated with cisplatin. Conclusion: This work identifies BCL-3 as a regulator of the cellular response to DNA damage and suggests that elevated BCL-3 expression could increase resistance of tumour cells to DNA damaging agents including radiotherapy. These findings offer a rationale for targeting BCL-3 in CRC as an adjuvant to conventional therapies and suggest that BCL-3 expression in tumours could be a useful biomarker in stratification of rectal cancer patients for neo-adjuvant chemoradiotherapy.

scholarly journals Rad51-Independent Interchromosomal Double-Strand Break Repair by Gene Conversion Requires Rad52 but Not Rad55, Rad57, or Dmc1

2007 ◽  
Vol 28 (3) ◽  
pp. 897-906 ◽  
Author(s):  
Thomas J. Pohl ◽  
Jac A. Nickoloff
Keyword(s):  
Gene Conversion ◽  
Strand Break ◽  
Double Strand Break ◽  
Single Strand ◽  
Homology Search ◽  
Wild Type ◽  
Dsb Repair ◽  
Single Strand Annealing ◽  
In The Wild ◽  
Break Induced Replication

ABSTRACT Homologous recombination (HR) is critical for DNA double-strand break (DSB) repair and genome stabilization. In yeast, HR is catalyzed by the Rad51 strand transferase and its “mediators,” including the Rad52 single-strand DNA-annealing protein, two Rad51 paralogs (Rad55 and Rad57), and Rad54. A Rad51 homolog, Dmc1, is important for meiotic HR. In wild-type cells, most DSB repair results in gene conversion, a conservative HR outcome. Because Rad51 plays a central role in the homology search and strand invasion steps, DSBs either are not repaired or are repaired by nonconservative single-strand annealing or break-induced replication mechanisms in rad51Δ mutants. Although DSB repair by gene conversion in the absence of Rad51 has been reported for ectopic HR events (e.g., inverted repeats or between plasmids), Rad51 has been thought to be essential for DSB repair by conservative interchromosomal (allelic) gene conversion. Here, we demonstrate that DSBs stimulate gene conversion between homologous chromosomes (allelic conversion) by >30-fold in a rad51Δ mutant. We show that Rad51-independent allelic conversion and break-induced replication occur independently of Rad55, Rad57, and Dmc1 but require Rad52. Unlike DSB-induced events, spontaneous allelic conversion was detected in both rad51Δ and rad52Δ mutants, but not in a rad51Δ rad52Δ double mutant. The frequencies of crossovers associated with DSB-induced gene conversion were similar in the wild type and the rad51Δ mutant, but discontinuous conversion tracts were fivefold more frequent and tract lengths were more widely distributed in the rad51Δ mutant, indicating that heteroduplex DNA has an altered structure, or is processed differently, in the absence of Rad51.

scholarly journals Schizosaccharomyces pombe KAT5 contributes to resection and repair of a DNA double strand break

Genetics ◽  
2021 ◽  
Author(s):  
Tingting Li ◽  
Ruben C Petreaca ◽  
Susan L Forsburg
Keyword(s):  
Dna Damage ◽  
Homologous Recombination ◽  
Chromatin Remodeling ◽  
Dna Damage Response ◽  
Histone Acetyltransferase ◽  
Strand Break ◽  
Double Strand Break ◽  
Dna Damage Checkpoint ◽  
Dna Double Strand Break ◽  
Damage Response

Abstract Chromatin remodeling is essential for effective repair of a DNA double strand break. KAT5 (S. pombe Mst1, human TIP60) is a MYST family histone acetyltransferase conserved from yeast to humans that coordinates various DNA damage response activities at a DNA double strand break (DSB), including histone remodeling and activation of the DNA damage checkpoint. In S. pombe, mutations in mst1+ causes sensitivity to DNA damaging drugs. Here we show that Mst1 is recruited to DSBs. Mutation of mst1+ disrupts recruitment of repair proteins and delays resection. These defects are partially rescued by deletion of pku70, which has been previously shown to antagonize repair by homologous recombination. These phenotypes of mst1 are similar to pht1-4KR, a non-acetylatable form of histone variant H2A.Z, which has been proposed to affect resection. Our data suggest that Mst1 functions to direct repair of DSBs towards homologous recombination pathways by modulating resection at the double strand break.

scholarly journals PARP-1–dependent recruitment of cold-inducible RNA-binding protein promotes double-strand break repair and genome stability

2018 ◽  
Vol 115 (8) ◽  
pp. E1759-E1768 ◽  
Author(s):  
Jung-Kuei Chen ◽  
Wen-Ling Lin ◽  
Zhang Chen ◽  
Hung-wen Liu
Keyword(s):  
Dna Damage ◽  
Genome Stability ◽  
Rna Binding ◽  
Binding Protein ◽  
Rna Binding Protein ◽  
Strand Break ◽  
Double Strand Break ◽  
Dsb Repair ◽  
Active Involvement ◽  
Parp 1

Maintenance of genome integrity is critical for both faithful propagation of genetic information and prevention of mutagenesis induced by various DNA damage events. Here we report cold-inducible RNA-binding protein (CIRBP) as a newly identified key regulator in DNA double-strand break (DSB) repair. On DNA damage, CIRBP temporarily accumulates at the damaged regions and is poly(ADP ribosyl)ated by poly(ADP ribose) polymerase-1 (PARP-1). Its dissociation from the sites of damage may depend on its phosphorylation status as mediated by phosphatidylinositol 3-kinase-related kinases. In the absence of CIRBP, cells showed reduced γH2AX, Rad51, and 53BP1 foci formation. Moreover, CIRBP-depleted cells exhibited impaired homologous recombination, impaired nonhomologous end-joining, increased micronuclei formation, and higher sensitivity to gamma irradiation, demonstrating the active involvement of CIRBP in DSB repair. Furthermore, CIRBP depleted cells exhibited defects in DNA damage-induced chromatin association of the MRN complex (Mre11, Rad50, and NBS1) and ATM kinase. CIRBP depletion also reduced phosphorylation of a variety of ATM substrate proteins and thus impaired the DNA damage response. Taken together, these results reveal a previously unrecognized role for CIRBP in DSB repair.

scholarly journals The nucleoporin 153, a novel factor in double-strand break repair and DNA damage response

Oncogene ◽  
2012 ◽  
Vol 31 (45) ◽  
pp. 4803-4809 ◽  
Author(s):  
C Lemaître ◽  
B Fischer ◽  
A Kalousi ◽  
A-S Hoffbeck ◽  
J Guirouilh-Barbat ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Damage Response ◽  
Strand Break ◽  
Double Strand Break ◽  
Double Strand Break Repair ◽  
Damage Response ◽  
Break Repair
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