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All Three Subunits of RecBCD Enzyme Are Essential for DNA Repair and Low-Temperature Growth in the Antarctic Pseudomonas syringae Lz4W

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All Three Subunits of RecBCD Enzyme Are Essential for DNA Repair and Low-Temperature Growth in the Antarctic Pseudomonas syringae Lz4W. / Pavankumar, Theetha L.; Sinha, Anurag K.; Ray, Malay K.

I: PLoS ONE, Bind 5, Nr. 2, e9412, 2010.

Publikation: Bidrag til tidsskriftTidsskriftartikelForskningfagfællebedømt

Harvard

Pavankumar, TL, Sinha, AK & Ray, MK 2010, 'All Three Subunits of RecBCD Enzyme Are Essential for DNA Repair and Low-Temperature Growth in the Antarctic Pseudomonas syringae Lz4W', PLoS ONE, bind 5, nr. 2, e9412. https://doi.org/10.1371/journal.pone.0009412

APA

Pavankumar, T. L., Sinha, A. K., & Ray, M. K. (2010). All Three Subunits of RecBCD Enzyme Are Essential for DNA Repair and Low-Temperature Growth in the Antarctic Pseudomonas syringae Lz4W. PLoS ONE, 5(2), [e9412]. https://doi.org/10.1371/journal.pone.0009412

Vancouver

Pavankumar TL, Sinha AK, Ray MK. All Three Subunits of RecBCD Enzyme Are Essential for DNA Repair and Low-Temperature Growth in the Antarctic Pseudomonas syringae Lz4W. PLoS ONE. 2010;5(2). e9412. https://doi.org/10.1371/journal.pone.0009412

Author

Pavankumar, Theetha L. ; Sinha, Anurag K. ; Ray, Malay K. / All Three Subunits of RecBCD Enzyme Are Essential for DNA Repair and Low-Temperature Growth in the Antarctic Pseudomonas syringae Lz4W. I: PLoS ONE. 2010 ; Bind 5, Nr. 2.

Bibtex

@article{58cdcf5783d942ed9908b2f579eb9cea,
title = "All Three Subunits of RecBCD Enzyme Are Essential for DNA Repair and Low-Temperature Growth in the Antarctic Pseudomonas syringae Lz4W",
abstract = "BACKGROUND: The recD mutants of the Antarctic Pseudomonas syringae Lz4W are sensitive to DNA-damaging agents and fail to grow at 4 degrees C. Generally, RecD associates with two other proteins (RecB and RecC) to produce RecBCD enzyme, which is involved in homologous recombination and DNA repair in many bacteria, including Escherichia coli. However, RecD is not essential for DNA repair, nor does its deletion cause any growth defects in E. coli. Hence, the assessment of the P. syringae RecBCD pathway was imperative.METHODOLOGY/PRINCIPAL FINDINGS: Mutational analysis and genetic complementation studies were used to establish that the individual null-mutations of all three genes, recC, recB, and recD, or the deletion of whole recCBD operon of P. syringae, lead to growth inhibition at low temperature, and sensitivity to UV and mitomycin C. Viability of the mutant cells dropped drastically at 4 degrees C, and the mutants accumulated linear chromosomal DNA and shorter DNA fragments in higher amounts compared to 22 degrees C. Additional genetic data using the mutant RecBCD enzymes that were inactivated either in the ATPase active site of RecB (RecB(K29Q)) or RecD (RecD(K229Q)), or in the nuclease center of RecB (RecB(D1118A) and RecB(Delta nuc)) suggested that, while the nuclease activity of RecB is not so critical in vivo, the ATP-dependent functions of both RecB and RecD are essential. Surprisingly, E. coli recBCD or recBC alone on plasmid could complement the defects of the Delta recCBD strain of P. syringae.CONCLUSIONS/SIGNIFICANCE: All three subunits of the RecBCD(Ps) enzyme are essential for DNA repair and growth of P. syringae at low temperatures (4 degrees C). The RecD requirement is only a function of the RecBCD complex in the bacterium. The RecBCD pathway protects the Antarctic bacterium from cold-induced DNA damages, and is critically dependent on the helicase activities of both RecB and RecD subunits, but not on the nuclease of RecBCD(Ps) enzyme.",
keywords = "Antarctic Regions, Bacterial Proteins/genetics, Blotting, Western, Cell Division/drug effects, Cold Temperature, DNA Repair, Exodeoxyribonuclease V/genetics, Genetic Complementation Test, Microbial Viability/drug effects, Mitomycin/pharmacology, Mutation, Nucleic Acid Synthesis Inhibitors/pharmacology, Operon/genetics, Protein Subunits/genetics, Pseudomonas syringae/enzymology, Recombination, Genetic, Ultraviolet Rays",
author = "Pavankumar, {Theetha L.} and Sinha, {Anurag K.} and Ray, {Malay K.}",
year = "2010",
doi = "10.1371/journal.pone.0009412",
language = "English",
volume = "5",
journal = "P L o S One",
issn = "1932-6203",
publisher = "Public Library of Science",
number = "2",

}

RIS

TY - JOUR

T1 - All Three Subunits of RecBCD Enzyme Are Essential for DNA Repair and Low-Temperature Growth in the Antarctic Pseudomonas syringae Lz4W

AU - Pavankumar, Theetha L.

AU - Sinha, Anurag K.

AU - Ray, Malay K.

PY - 2010

Y1 - 2010

N2 - BACKGROUND: The recD mutants of the Antarctic Pseudomonas syringae Lz4W are sensitive to DNA-damaging agents and fail to grow at 4 degrees C. Generally, RecD associates with two other proteins (RecB and RecC) to produce RecBCD enzyme, which is involved in homologous recombination and DNA repair in many bacteria, including Escherichia coli. However, RecD is not essential for DNA repair, nor does its deletion cause any growth defects in E. coli. Hence, the assessment of the P. syringae RecBCD pathway was imperative.METHODOLOGY/PRINCIPAL FINDINGS: Mutational analysis and genetic complementation studies were used to establish that the individual null-mutations of all three genes, recC, recB, and recD, or the deletion of whole recCBD operon of P. syringae, lead to growth inhibition at low temperature, and sensitivity to UV and mitomycin C. Viability of the mutant cells dropped drastically at 4 degrees C, and the mutants accumulated linear chromosomal DNA and shorter DNA fragments in higher amounts compared to 22 degrees C. Additional genetic data using the mutant RecBCD enzymes that were inactivated either in the ATPase active site of RecB (RecB(K29Q)) or RecD (RecD(K229Q)), or in the nuclease center of RecB (RecB(D1118A) and RecB(Delta nuc)) suggested that, while the nuclease activity of RecB is not so critical in vivo, the ATP-dependent functions of both RecB and RecD are essential. Surprisingly, E. coli recBCD or recBC alone on plasmid could complement the defects of the Delta recCBD strain of P. syringae.CONCLUSIONS/SIGNIFICANCE: All three subunits of the RecBCD(Ps) enzyme are essential for DNA repair and growth of P. syringae at low temperatures (4 degrees C). The RecD requirement is only a function of the RecBCD complex in the bacterium. The RecBCD pathway protects the Antarctic bacterium from cold-induced DNA damages, and is critically dependent on the helicase activities of both RecB and RecD subunits, but not on the nuclease of RecBCD(Ps) enzyme.

AB - BACKGROUND: The recD mutants of the Antarctic Pseudomonas syringae Lz4W are sensitive to DNA-damaging agents and fail to grow at 4 degrees C. Generally, RecD associates with two other proteins (RecB and RecC) to produce RecBCD enzyme, which is involved in homologous recombination and DNA repair in many bacteria, including Escherichia coli. However, RecD is not essential for DNA repair, nor does its deletion cause any growth defects in E. coli. Hence, the assessment of the P. syringae RecBCD pathway was imperative.METHODOLOGY/PRINCIPAL FINDINGS: Mutational analysis and genetic complementation studies were used to establish that the individual null-mutations of all three genes, recC, recB, and recD, or the deletion of whole recCBD operon of P. syringae, lead to growth inhibition at low temperature, and sensitivity to UV and mitomycin C. Viability of the mutant cells dropped drastically at 4 degrees C, and the mutants accumulated linear chromosomal DNA and shorter DNA fragments in higher amounts compared to 22 degrees C. Additional genetic data using the mutant RecBCD enzymes that were inactivated either in the ATPase active site of RecB (RecB(K29Q)) or RecD (RecD(K229Q)), or in the nuclease center of RecB (RecB(D1118A) and RecB(Delta nuc)) suggested that, while the nuclease activity of RecB is not so critical in vivo, the ATP-dependent functions of both RecB and RecD are essential. Surprisingly, E. coli recBCD or recBC alone on plasmid could complement the defects of the Delta recCBD strain of P. syringae.CONCLUSIONS/SIGNIFICANCE: All three subunits of the RecBCD(Ps) enzyme are essential for DNA repair and growth of P. syringae at low temperatures (4 degrees C). The RecD requirement is only a function of the RecBCD complex in the bacterium. The RecBCD pathway protects the Antarctic bacterium from cold-induced DNA damages, and is critically dependent on the helicase activities of both RecB and RecD subunits, but not on the nuclease of RecBCD(Ps) enzyme.

KW - Antarctic Regions

KW - Bacterial Proteins/genetics

KW - Blotting, Western

KW - Cell Division/drug effects

KW - Cold Temperature

KW - DNA Repair

KW - Exodeoxyribonuclease V/genetics

KW - Genetic Complementation Test

KW - Microbial Viability/drug effects

KW - Mitomycin/pharmacology

KW - Mutation

KW - Nucleic Acid Synthesis Inhibitors/pharmacology

KW - Operon/genetics

KW - Protein Subunits/genetics

KW - Pseudomonas syringae/enzymology

KW - Recombination, Genetic

KW - Ultraviolet Rays

U2 - 10.1371/journal.pone.0009412

DO - 10.1371/journal.pone.0009412

M3 - Journal article

C2 - 20195537

VL - 5

JO - P L o S One

JF - P L o S One

SN - 1932-6203

IS - 2

M1 - e9412

ER -

ID: 234877543