Double-strand break repair in bacteria: a view from<i>Bacillus subtilis</i>

Ayora, Silvia; Carrasco, Begoña; Cárdenas, Paula P.; César, Carolina Elvira; Cañas, Cristina; Yadav, Tribhuwan; Marchisone, Chiara; Alonso, Juan C.

doi:10.1111/j.1574-6976.2011.00272.x

Cited by 114 publications

(122 citation statements)

References 276 publications

(697 reference statements)

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“…Studies on RecF and RecR in E. coli (44,45), B. subtilis (17,46), and D. radiodurans (19,47) have led to the proposal that these two proteins function in concert with RecO as RecA mediators in the repair of DNA breaks and gaps. The M. smegmatis ⌬recO mutant is extremely sensitive to DNA damage and participates in two different pathways of recombination, recA-dependent HR and recA-independent SSA (10), implying that the function of RecO in the SSA pathway is distinct from its classic RecA mediator function.…”

Section: Resultsmentioning

confidence: 99%

RecF and RecR Play Critical Roles in the Homologous Recombination and Single-Strand Annealing Pathways of Mycobacteria

2015

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Mycobacteria encode three DNA double-strand break repair pathways: (i) RecA-dependent homologous recombination (HR), (ii) Ku-dependent nonhomologous end joining (NHEJ), and (iii) RecBCD-dependent single-strand annealing (SSA). Mycobacterial HR has two presynaptic pathway options that rely on the helicase-nuclease AdnAB and the strand annealing protein RecO, respectively. Ablation of adnAB or recO individually causes partial impairment of HR, but loss of adnAB and recO in combination abolishes HR. RecO, which can accelerate annealing of single-stranded DNA in vitro, also participates in the SSA pathway. The functions of RecF and RecR, which, in other model bacteria, function in concert with RecO as mediators of RecA loading, have not been examined in mycobacteria. Here, we present a genetic analysis of recF and recR in mycobacterial recombination. We find that RecF, like RecO, participates in the AdnAB-independent arm of the HR pathway and in SSA. In contrast, RecR is required for all HR in mycobacteria and for SSA. The essentiality of RecR as an agent of HR is yet another distinctive feature of mycobacterial DNA repair. N ature has evolved mechanisms to repair the structurally diverse DNA lesions that arise during chromosomal replication and from exogenous mutagens. Double-strand breaks (DSBs) are especially deleterious, insofar as a single chromosomal DSB can be lethal to a cell if not repaired prior to cell division. The major mechanisms of DSB repair are conserved, at least in their general principles, from bacteria to humans. This parallel is especially relevant for mycobacteria, which implement the same three distinct DSB repair pathways operative in eukarya: homologous recombination (HR), nonhomologous end joining (NHEJ), and singlestrand annealing (SSA). Mycobacterial NHEJ relies on the end-binding protein Ku, DNA ligase LigD (a multifunctional ligase-polymerase-3=-phosphoesterase), DNA ligase LigC1, and two additional polymerases (PolD1 and PolD2) (1-9). The SSA pathway of mycobacteria repairs DSBs that arise between repeats, with consequent deletion of the DNA between the repeats. SSA is independent of the strand exchange protein RecA but requires the helicase-nuclease RecBCD and the strand-annealing protein RecO (9,10). IMPORTANCE This study clarifies the molecular requirements for homologous recombination in mycobacteria. Specifically, we demonstrate that RecF and RecR play important roles in both theThe central agent of homology search in the HR pathways of all organisms is a strand exchange protein that polymerizes on the single-strand DNA (ssDNA) resulting from end resection at a DSB. In bacteria, the strand exchange protein is RecA; in eukarya, it is Rad51. RecA nucleation on ssDNA is regulated in order to prevent inappropriate nucleoprotein filament formation, which can be toxic (11). The presence of single-strand binding protein (SSB) on ssDNA prevents RecA loading unless a "mediator" protein or protein complex catalyzes the exchange of SSB for RecA to create the RecA nucleoprotein filam...

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Section: Resultsmentioning

confidence: 99%

RecF and RecR Play Critical Roles in the Homologous Recombination and Single-Strand Annealing Pathways of Mycobacteria

2015

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“…To further test if MutS2 promotes homologous recombination, we employed a transformation efficiency assay. Transformation of naturally competent B. subtilis cells with plasmid or chromosomal DNA is dependent on the function of several homologous recombination proteins, although the requirements for chromosomal DNA integration relative to plasmid assembly and maintenance are not identical (4,47). Thus, if MutS2 functions by promoting homologous recombination, as the MMC phenotype suggests, we would expect to observe decreased transformation efficiency using plasmid and/or chromosomal DNA.…”

Section: Figmentioning

confidence: 98%

MutS2 Promotes Homologous Recombination in Bacillus subtilis

Burby

Simmons

2017

J Bacteriol

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Bacterial MutS proteins are subdivided into two families, MutS1 and MutS2. MutS1 family members recognize DNA replication errors during their participation in the well-characterized mismatch repair (MMR) pathway. In contrast to the well-described function of MutS1, the function of MutS2 in bacteria has remained less clear. In Helicobacter pylori and Thermus thermophilus, MutS2 has been shown to suppress homologous recombination. The role of MutS2 is unknown in the Grampositive bacterium Bacillus subtilis. In this work, we investigated the contribution of MutS2 to maintaining genome integrity in B. subtilis. We found that deletion of mutS2 renders B. subtilis sensitive to the natural antibiotic mitomycin C (MMC), which requires homologous recombination for repair. We demonstrate that the C-terminal small MutS-related (Smr) domain is necessary but not sufficient for tolerance to MMC. Further, we developed a CRISPR/Cas9 genome editing system to test if the inducible prophage PBSX was the underlying cause of the observed MMC sensitivity. Genetic analysis revealed that MMC sensitivity was dependent on recombination and not on nucleotide excision repair or a symptom of prophage PBSX replication and cell lysis. We found that deletion of mutS2 resulted in decreased transformation efficiency using both plasmid and chromosomal DNA. Further, deletion of mutS2 in a strain lacking the Holliday junction endonuclease gene recU resulted in increased MMC sensitivity and decreased transformation efficiency, suggesting that MutS2 could function redundantly with RecU. Together, our results support a model where B. subtilis MutS2 helps to promote homologous recombination, demonstrating a new function for bacterial MutS2.IMPORTANCE Cells contain pathways that promote or inhibit recombination. MutS2 homologs are Smr-endonuclease domain-containing proteins that have been shown to function in antirecombination in some bacteria. We present evidence that B. subtilis MutS2 promotes recombination, providing a new function for MutS2. We found that cells lacking mutS2 are sensitive to DNA damage that requires homologous recombination for repair and have reduced transformation efficiency. Further analysis indicates that the C-terminal Smr domain requires the N-terminal portion of MutS2 for function in vivo. Moreover, we show that a mutS2 deletion is additive with a recU deletion, suggesting that these proteins have a redundant function in homologous recombination. Together, our study shows that MutS2 proteins have adapted different functions that impact recombination.

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“…3 and 4). Because ionizing radiation causes such a wide spectrum of direct and indirect DNA damage, it will be interesting to determine the contributions of additional error-free and error-prone DNA repair pathways (e.g., mismatch repair, translesion synthesis, and homologous recombination [29,[42][43][44]), to gain further detailed insights into spore resistance to ionizing radiation.…”

Section: Discussionmentioning

confidence: 99%

Resistance of Bacillus subtilis Spore DNA to Lethal Ionizing Radiation Damage Relies Primarily on Spore Core Components and DNA Repair, with Minor Effects of Oxygen Radical Detoxification

Moeller

Raguse

Reitz

et al. 2014

Appl Environ Microbiol

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eThe roles of various core components, including ␣/␤/␥-type small acid-soluble spore proteins (SASP), dipicolinic acid (DPA), core water content, and DNA repair by apurinic/apyrimidinic (AP) endonucleases or nonhomologous end joining (NHEJ), in Bacillus subtilis spore resistance to different types of ionizing radiation including X rays, protons, and high-energy charged iron ions have been studied. Spores deficient in DNA repair by NHEJ or AP endonucleases, the oxidative stress response, or protection by major ␣/␤-type SASP, DPA, and decreased core water content were significantly more sensitive to ionizing radiation than wild-type spores, with highest sensitivity to high-energy-charged iron ions. DNA repair via NHEJ and AP endonucleases appears to be the most important mechanism for spore resistance to ionizing radiation, whereas oxygen radical detoxification via the MrgA-mediated oxidative stress response or KatX catalase activity plays only a very minor role. Synergistic radioprotective effects of ␣/␤-type but not ␥-type SASP were also identified, indicating that ␣/␤-type SASP's binding to spore DNA is important in preventing DNA damage due to reactive oxygen species generated by ionizing radiation.

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Double-strand break repair in bacteria: a view fromBacillus subtilis

Cited by 114 publications

References 276 publications

RecF and RecR Play Critical Roles in the Homologous Recombination and Single-Strand Annealing Pathways of Mycobacteria

RecF and RecR Play Critical Roles in the Homologous Recombination and Single-Strand Annealing Pathways of Mycobacteria

MutS2 Promotes Homologous Recombination in Bacillus subtilis

Resistance of Bacillus subtilis Spore DNA to Lethal Ionizing Radiation Damage Relies Primarily on Spore Core Components and DNA Repair, with Minor Effects of Oxygen Radical Detoxification

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