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Identification of new genes contributing to the extreme radioresistance of Deinococcus radiodurans using a Tn5-based transposon mutant library.

Dulermo R, Onodera T, Coste G, Passot F, Dutertre M, Porteron M, Confalonieri F, Sommer S, Pasternak C - PLoS ONE (2015)

Bottom Line: Here, we have developed an extremely efficient in vivo Tn5-based mutagenesis procedure to construct a Deinococcus radiodurans insertion mutant library subsequently screened for sensitivity to genotoxic agents such as γ and UV radiations or mitomycin C.Interestingly, preliminary characterization of previously undescribed radiosensitive mutants suggests the contribution of cyclic di-AMP signaling in the recovery of D. radiodurans cells from genotoxic stresses, probably by modulating several pathways involved in the overall cell response.Our analyses also point out a new transcriptional regulator belonging to the GntR family, encoded by DR0265, and a predicted RNase belonging to the newly described Y family, both contributing to the extreme radioresistance of D. radiodurans.

View Article: PubMed Central - PubMed

Affiliation: Univ. Paris-Sud, Institute for Integrative Biology of the Cell (I2BC), Université Paris Saclay, CEA, CNRS, Orsay, France.

ABSTRACT
Here, we have developed an extremely efficient in vivo Tn5-based mutagenesis procedure to construct a Deinococcus radiodurans insertion mutant library subsequently screened for sensitivity to genotoxic agents such as γ and UV radiations or mitomycin C. The genes inactivated in radiosensitive mutants belong to various functional categories, including DNA repair functions, stress responses, signal transduction, membrane transport, several metabolic pathways, and genes of unknown function. Interestingly, preliminary characterization of previously undescribed radiosensitive mutants suggests the contribution of cyclic di-AMP signaling in the recovery of D. radiodurans cells from genotoxic stresses, probably by modulating several pathways involved in the overall cell response. Our analyses also point out a new transcriptional regulator belonging to the GntR family, encoded by DR0265, and a predicted RNase belonging to the newly described Y family, both contributing to the extreme radioresistance of D. radiodurans. Altogether, this work has revealed new cell responses involved either directly or indirectly in repair of various cell damage and confirmed that D. radiodurans extreme radiation resistance is determined by a multiplicity of pathways acting as a complex network.

No MeSH data available.


Related in: MedlinePlus

Cell morphology in D. radiodurans deleted for DR2462 observed by microscopy.D. radiodurans cells from the wild type strain (top panels) and from ΔDR2462 strain (bottom panels) were grown to OD650 = 0.3. Left panels: Nomarski interference contrast (DIC). Middle panels: membrane staining (FM4-64). Right panels: DNA staining (DAPI). All pictures are at the same scale (bar = 2 μm).
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pone.0124358.g007: Cell morphology in D. radiodurans deleted for DR2462 observed by microscopy.D. radiodurans cells from the wild type strain (top panels) and from ΔDR2462 strain (bottom panels) were grown to OD650 = 0.3. Left panels: Nomarski interference contrast (DIC). Middle panels: membrane staining (FM4-64). Right panels: DNA staining (DAPI). All pictures are at the same scale (bar = 2 μm).

Mentions: In B. subtilis, depletion of RNase Y (YmdA) resulted in an aberrant distribution of cell lengths, with a few unusually longer cells and many short, almost spherical, cells reminiscent of minicells [81,82]. It was proposed that this phenotype might be related to increased concentration of dnaA transcript [80], since overexpression of DnaA has been linked to aberrant changes in cell shape [83]. D. radiodurans mutant bacteria devoid of DR2462 protein grew normally and did not show dramatic alterations of their morphologies when they were observed by microscopy after nucleoid and membrane staining, except that they are slightly smaller than wild type cells (Fig 7), suggesting a more modest role of RNase Y in D. radiodurans cell viability than those played by RNase Y in B. subtilis.


Identification of new genes contributing to the extreme radioresistance of Deinococcus radiodurans using a Tn5-based transposon mutant library.

Dulermo R, Onodera T, Coste G, Passot F, Dutertre M, Porteron M, Confalonieri F, Sommer S, Pasternak C - PLoS ONE (2015)

Cell morphology in D. radiodurans deleted for DR2462 observed by microscopy.D. radiodurans cells from the wild type strain (top panels) and from ΔDR2462 strain (bottom panels) were grown to OD650 = 0.3. Left panels: Nomarski interference contrast (DIC). Middle panels: membrane staining (FM4-64). Right panels: DNA staining (DAPI). All pictures are at the same scale (bar = 2 μm).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4401554&req=5

pone.0124358.g007: Cell morphology in D. radiodurans deleted for DR2462 observed by microscopy.D. radiodurans cells from the wild type strain (top panels) and from ΔDR2462 strain (bottom panels) were grown to OD650 = 0.3. Left panels: Nomarski interference contrast (DIC). Middle panels: membrane staining (FM4-64). Right panels: DNA staining (DAPI). All pictures are at the same scale (bar = 2 μm).
Mentions: In B. subtilis, depletion of RNase Y (YmdA) resulted in an aberrant distribution of cell lengths, with a few unusually longer cells and many short, almost spherical, cells reminiscent of minicells [81,82]. It was proposed that this phenotype might be related to increased concentration of dnaA transcript [80], since overexpression of DnaA has been linked to aberrant changes in cell shape [83]. D. radiodurans mutant bacteria devoid of DR2462 protein grew normally and did not show dramatic alterations of their morphologies when they were observed by microscopy after nucleoid and membrane staining, except that they are slightly smaller than wild type cells (Fig 7), suggesting a more modest role of RNase Y in D. radiodurans cell viability than those played by RNase Y in B. subtilis.

Bottom Line: Here, we have developed an extremely efficient in vivo Tn5-based mutagenesis procedure to construct a Deinococcus radiodurans insertion mutant library subsequently screened for sensitivity to genotoxic agents such as γ and UV radiations or mitomycin C.Interestingly, preliminary characterization of previously undescribed radiosensitive mutants suggests the contribution of cyclic di-AMP signaling in the recovery of D. radiodurans cells from genotoxic stresses, probably by modulating several pathways involved in the overall cell response.Our analyses also point out a new transcriptional regulator belonging to the GntR family, encoded by DR0265, and a predicted RNase belonging to the newly described Y family, both contributing to the extreme radioresistance of D. radiodurans.

View Article: PubMed Central - PubMed

Affiliation: Univ. Paris-Sud, Institute for Integrative Biology of the Cell (I2BC), Université Paris Saclay, CEA, CNRS, Orsay, France.

ABSTRACT
Here, we have developed an extremely efficient in vivo Tn5-based mutagenesis procedure to construct a Deinococcus radiodurans insertion mutant library subsequently screened for sensitivity to genotoxic agents such as γ and UV radiations or mitomycin C. The genes inactivated in radiosensitive mutants belong to various functional categories, including DNA repair functions, stress responses, signal transduction, membrane transport, several metabolic pathways, and genes of unknown function. Interestingly, preliminary characterization of previously undescribed radiosensitive mutants suggests the contribution of cyclic di-AMP signaling in the recovery of D. radiodurans cells from genotoxic stresses, probably by modulating several pathways involved in the overall cell response. Our analyses also point out a new transcriptional regulator belonging to the GntR family, encoded by DR0265, and a predicted RNase belonging to the newly described Y family, both contributing to the extreme radioresistance of D. radiodurans. Altogether, this work has revealed new cell responses involved either directly or indirectly in repair of various cell damage and confirmed that D. radiodurans extreme radiation resistance is determined by a multiplicity of pathways acting as a complex network.

No MeSH data available.


Related in: MedlinePlus