Letter | Published:

Rates and mechanisms of bacterial mutagenesis from maximum-depth sequencing

Nature volume 534, pages 693696 (30 June 2016) | Download Citation

Abstract

In 1943, Luria and Delbrück used a phage-resistance assay to establish spontaneous mutation as a driving force of microbial diversity1. Mutation rates are still studied using such assays, but these can only be used to examine the small minority of mutations conferring survival in a particular condition. Newer approaches, such as long-term evolution followed by whole-genome sequencing2,3, may be skewed by mutational ‘hot’ or ‘cold’ spots3,4. Both approaches are affected by numerous caveats5,6,7. Here we devise a method, maximum-depth sequencing (MDS), to detect extremely rare variants in a population of cells through error-corrected, high-throughput sequencing. We directly measure locus-specific mutation rates in Escherichia coli and show that they vary across the genome by at least an order of magnitude. Our data suggest that certain types of nucleotide misincorporation occur 104-fold more frequently than the basal rate of mutations, but are repaired in vivo. Our data also suggest specific mechanisms of antibiotic-induced mutagenesis, including downregulation of mismatch repair via oxidative stress, transcription–replication conflicts, and, in the case of fluoroquinolones, direct damage to DNA.

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Acknowledgements

We thank A. Heguy and the NYU Genome Technology Center, which is partially supported by the Cancer Center Support Grant, P30CA016087, at the Laura and Isaac Perlmutter Cancer Center. This work used computing resources at the High Performance Computing Facility of the Center for Health Informatics and Bioinformatics at the NYU Langone Medical Center. We thank D. Dwyer and K. Shankarling for materials, and T. Artemyev for his contribution. This work was supported by NIH grant R01GM107329 and HHMI (E.N.) and NCI PSOC grant U54 CA193313 (B.M.). J.J. was supported by the NYU Medical Scientist Training Program and a National Defense Science and Engineering Graduate Fellowship.

Author information

Author notes

    • Bud Mishra
    •  & Evgeny Nudler

    These authors jointly supervised this work.

Affiliations

  1. Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, New York, New York 10016, USA

    • Justin Jee
    • , Aviram Rasouly
    • , Ilya Shamovsky
    • , Yonatan Akivis
    • , Susan R. Steinman
    •  & Evgeny Nudler
  2. Courant Institute of Mathematical Sciences, New York University, New York, New York 10012, USA

    • Justin Jee
    •  & Bud Mishra
  3. Howard Hughes Medical Institute, New York University School of Medicine, New York, New York 10016, USA

    • Aviram Rasouly
    •  & Evgeny Nudler

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Contributions

J.J. and I.S. designed the MDS protocols. J.J., A.R., and E.N. designed the biological experiments. J.J., A.R., and Y.A. performed the experiments. J.J., B.M., S.S., and I.S. performed the data analysis. J.J. and E.N. wrote the manuscript with input from all co-authors. B.M. and E.N. supervised the research.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Bud Mishra or Evgeny Nudler.

Reviewer Information Nature thanks N. Luscombe, I. Martincorena, J. Wang and the other anonymous reviewer(s) for their contribution to the peer review of this work.

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    Supplementary Information

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DOI

https://doi.org/10.1038/nature18313

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