Characterisation of an Escherichia coli line that completely lacks ribonucleotide reduction yields insights into the evolution of parasitism and endosymbiosis

Abstract

All life requires ribonucleotide reduction for de novo synthesis of deoxyribonucleotides. A handful of obligate intracellular species are known to lack ribonucleotide reduction and are instead dependent on their host for deoxyribonucleotide synthesis. As ribonucleotide reduction has on occasion been lost in obligate intracellular parasites and endosymbionts, we reasoned that it should in principle be possible to knock this process out entirely under conditions where deoxyribonucleosides are present in the growth media. We report here the creation of a strain of E. coli where all three ribonucleotide reductase operons have been fully deleted following introduction of a broad spectrum deoxyribonucleoside kinase from Mycoplasma mycoides. Our strain is able to grow in the presence of deoxyribonucleosides and shows slowed but substantial growth. Under limiting deoxyribonucleoside levels, we observe a distinctive filamentous cell morphology, where cells grow but do not appear to divide regularly. Finally, we examined whether our lines are able to adapt to limited supplies of deoxyribonucleosides, as might occur in the evolutionary switch from de novo synthesis to dependence on host production during the evolution of parasitism or endosymbiosis. Over the course of an evolution experiment, we observe a 25-fold reduction in the minimum concentration of exogenous deoxyribonucleosides necessary for growth. Genome analysis of replicate lines reveals that several lines carry mutations in deoB and cdd. deoB codes for phosphopentomutase, a key part of the deoxyriboaldolase pathway, which has been hypothesised as an alternative to ribonucleotide reduction for deoxyribonucleotide synthesis. Rather than synthesis via this pathway complementing the loss of ribonucleotide reduction, our experiments reveal that mutations appear that reduce or eliminate the capacity for this pathway to catabolise deoxyribonucleotides, thus preventing their loss via central metabolism. Mutational inactivation of both deoB and cdd is also observed in a number of obligate intracellular bacteria that have lost ribonucleotide reduction. We conclude that our experiments recapitulate key evolutionary steps in the adaptation to intracellular life without ribonucleotide reduction.

Data availability

All genome data have been deposited to the Single Read Archive (https://www.ncbi.nlm.nih.gov/sra) under accessions SRX17677859-SRX17677886. The following biosamples are associated with this project: SAMN30957267-SAMN30957273. The data have been deposited with links to BioProject accession number PRJNA882995 in the NCBI BioProject database (https://www.ncbi.nlm.nih.gov/bioproject/).

The following data sets were generated

Article and author information

Author details

  1. Samantha D Arras

    School of Biological Sciences, University of Auckland, Auckland, New Zealand
    Competing interests
    The authors declare that no competing interests exist.
  2. Nellie Sibaeva

    School of Biological Sciences, University of Auckland, Auckland, New Zealand
    Competing interests
    The authors declare that no competing interests exist.
  3. Ryan J Catchpole

    School of Biological Sciences, University of Canterbury, Christchurch, New Zealand
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-6641-647X
  4. Nobuyuki Horinouchi

    Division of Applied Life Sciences, Kyoto University, Kyoto, Japan
    Competing interests
    The authors declare that no competing interests exist.
  5. Dayong Si

    Division of Applied Life Sciences, Kyoto University, Kyoto, Japan
    Competing interests
    The authors declare that no competing interests exist.
  6. Alannah M Rickerby

    School of Biological Sciences, University of Auckland, Auckland, New Zealand
    Competing interests
    The authors declare that no competing interests exist.
  7. Kengo Deguchi

    Division of Applied Life Sciences, Kyoto University, Kyoto, Japan
    Competing interests
    The authors declare that no competing interests exist.
  8. Makoto Hibi

    Division of Applied Life Sciences, Kyoto University, Kyoto, Japan
    Competing interests
    The authors declare that no competing interests exist.
  9. Koichi Tanaka

    Division of Applied Life Sciences, Kyoto University, Kyoto, Japan
    Competing interests
    The authors declare that no competing interests exist.
  10. Michiki Takeuchi

    Division of Applied Life Sciences, Kyoto University, Kyoto, Japan
    Competing interests
    The authors declare that no competing interests exist.
  11. Jun Ogawa

    Division of Applied Life Sciences, Kyoto University, Kyoto, Japan
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-2741-621X
  12. Anthony M Poole

    School of Biological Sciences, University of Auckland, Auckland, New Zealand
    For correspondence
    a.poole@auckland.ac.nz
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-9940-2824

Funding

Royal Society Te Apārangi (17-UOA-257)

  • Jun Ogawa
  • Anthony M Poole

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Reviewing Editor

  1. Amie K Boal, Pennsylvania State University, United States

Version history

  1. Preprint posted: September 30, 2022 (view preprint)
  2. Received: September 30, 2022
  3. Accepted: April 5, 2023
  4. Accepted Manuscript published: April 6, 2023 (version 1)
  5. Version of Record published: April 21, 2023 (version 2)

Copyright

© 2023, Arras et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

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  1. Samantha D Arras
  2. Nellie Sibaeva
  3. Ryan J Catchpole
  4. Nobuyuki Horinouchi
  5. Dayong Si
  6. Alannah M Rickerby
  7. Kengo Deguchi
  8. Makoto Hibi
  9. Koichi Tanaka
  10. Michiki Takeuchi
  11. Jun Ogawa
  12. Anthony M Poole
(2023)
Characterisation of an Escherichia coli line that completely lacks ribonucleotide reduction yields insights into the evolution of parasitism and endosymbiosis
eLife 12:e83845.
https://doi.org/10.7554/eLife.83845

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https://doi.org/10.7554/eLife.83845

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