1. Biochemistry and Chemical Biology

Random-sequence genetic oligomer pools display an innate potential for ligation and recombination

  1. Hannes Mutschler  Is a corresponding author
  2. Alexander I Taylor
  3. Benjamin T Porebski
  4. Alice Lightowlers
  5. Gillian Houlihan
  6. Mikhail Abramov
  7. Piet Herdewijn
  8. Philipp Holliger  Is a corresponding author
  1. MRC Laboratory of Molecular Biology, United Kingdom
  2. Katholieke Universiteit Leuven, Belgium
https://doi.org/10.7554/eLife.43022
Share this article
Cite this article
  1. Hannes Mutschler
  2. Alexander I Taylor
  3. Benjamin T Porebski
  4. Alice Lightowlers
  5. Gillian Houlihan
  6. Mikhail Abramov
  7. Piet Herdewijn
  8. Philipp Holliger
(2018)
Random-sequence genetic oligomer pools display an innate potential for ligation and recombination
eLife 7:e43022.
https://doi.org/10.7554/eLife.43022
  2. Download BibTeX
  3. Download .RIS

Abstract

Recombination, the exchange of information between different genetic polymer strands, is of fundamental importance in biology for genome maintenance and genetic diversification mediated by dedicated recombinase enzymes. Here, we describe a pervasive non-enzymatic capacity for recombination (and ligation) in random-sequence genetic oligomer pools. Specifically, we examine random and semi-random eicosamer (N20) pools of RNA, DNA and the unnatural genetic polymers ANA (arabino-), HNA (hexitol-) and AtNA (altritol-nucleic acids). While DNA, ANA and HNA pools proved inert, RNA and AtNA pools displayed diverse modes of spontaneous intermolecular recombination, connecting recombination mechanistically to the vicinal ring cis-diol configuration shared by RNA and AtNA. Thus, the chemical constitution that renders both susceptible to hydrolysis emerges as the fundamental determinant of an innate capacity for recombination, which is shown to enable a concomitant increase in compositional, informational and structural pool complexity and hence evolutionary potential.

Data availability

All data generated or analysed during this study are included in the manuscript and supporting files. Source data files have been provided for: Figure 1, Figure 1-supplement 1, Figure 2, Figure 2-supplement 1, Figure 4, Figure 4-supplement 2, Figure 4-supplement 3, Figure 5, Figure 5-supplement 1, Figure 6 and Figure 6-supplement 2. Further source files are shell scripts for the motif search, extraction of secondary structure frequencies and size distributions and python scripts for the generation of random sequences using position-specific nucleotide frequencies and the simulations of the population level Shannon Index at different cleavage / ligation rates are available online (see Source Code Files 1-5).

The following data sets were generated

Article and author information

Author details

  1. Hannes Mutschler

    MRC Laboratory of Molecular Biology, Cambridge, United Kingdom
    For correspondence
    mutschler@biochem.mpg.de
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-8005-1657

  2. Alexander I Taylor

    MRC Laboratory of Molecular Biology, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-7684-1437

  3. Benjamin T Porebski

    MRC Laboratory of Molecular Biology, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.

  4. Alice Lightowlers

    MRC Laboratory of Molecular Biology, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.

  5. Gillian Houlihan

    MRC Laboratory of Molecular Biology, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.

  6. Mikhail Abramov

    REGA Institute, Katholieke Universiteit Leuven, Leuven, Belgium
    Competing interests
    The authors declare that no competing interests exist.

  7. Piet Herdewijn

    REGA Institute, Katholieke Universiteit Leuven, Leuven, Belgium
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-3589-8503

  8. Philipp Holliger

    MRC Laboratory of Molecular Biology, Cambridge, United Kingdom
    For correspondence
    ph1@mrc-lmb.cam.ac.uk
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-3440-9854

Funding

Medical Research Council (MC_U105178804)

  • Alexander I Taylor
  • Benjamin T Porebski
  • Gillian Houlihan
  • Philipp Holliger

Federation of European Biochemical Societies (FEBS Long-Term Fellowship)

  • Hannes Mutschler

KU Leuven (OT/1414/128)

  • Mikhail Abramov
  • Piet Herdewijn

FWO Vlaanderen (G078014N)

  • Mikhail Abramov
  • Piet Herdewijn

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

Reviewing Editor

  1. Ulrich Muller, UCSD, United States

Version history

  1. Received: October 21, 2018
  2. Accepted: November 16, 2018
  3. Accepted Manuscript published: November 21, 2018 (version 1)
  4. Version of Record published: December 11, 2018 (version 2)

Copyright

© 2018, Mutschler 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.

Metrics

  • 3,403
    views
  • 476
    downloads
  • 42
    citations

Views, downloads and citations are aggregated across all versions of this paper published by eLife.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Hannes Mutschler
  2. Alexander I Taylor
  3. Benjamin T Porebski
  4. Alice Lightowlers
  5. Gillian Houlihan
  6. Mikhail Abramov
  7. Piet Herdewijn
  8. Philipp Holliger
(2018)
Random-sequence genetic oligomer pools display an innate potential for ligation and recombination
eLife 7:e43022.
https://doi.org/10.7554/eLife.43022

Share this article

https://doi.org/10.7554/eLife.43022

Categories and tags

Further reading

    1. Biochemistry and Chemical Biology
    2. Computational and Systems Biology

    Enrichment of rare codons at 5' ends of genes is a spandrel caused by evolutionary sequence turnover and does not improve translation

    Richard Sejour, Janet Leatherwood ... Bruce Futcher
    Research Article

    Previously, Tuller et al. found that the first 30–50 codons of the genes of yeast and other eukaryotes are slightly enriched for rare codons. They argued that this slowed translation, and was adaptive because it queued ribosomes to prevent collisions. Today, the translational speeds of different codons are known, and indeed rare codons are translated slowly. We re-examined this 5’ slow translation ‘ramp.’ We confirm that 5’ regions are slightly enriched for rare codons; in addition, they are depleted for downstream Start codons (which are fast), with both effects contributing to slow 5’ translation. However, we also find that the 5’ (and 3’) ends of yeast genes are poorly conserved in evolution, suggesting that they are unstable and turnover relatively rapidly. When a new 5’ end forms de novo, it is likely to include codons that would otherwise be rare. Because evolution has had a relatively short time to select against these codons, 5’ ends are typically slightly enriched for rare, slow codons. Opposite to the expectation of Tuller et al., we show by direct experiment that genes with slowly translated codons at the 5’ end are expressed relatively poorly, and that substituting faster synonymous codons improves expression. Direct experiment shows that slow codons do not prevent downstream ribosome collisions. Further informatic studies suggest that for natural genes, slow 5’ ends are correlated with poor gene expression, opposite to the expectation of Tuller et al. Thus, we conclude that slow 5’ translation is a ‘spandrel’--a non-adaptive consequence of something else, in this case, the turnover of 5’ ends in evolution, and it does not improve translation.

    1. Biochemistry and Chemical Biology

    Uncovering the BIN1-SH3 interactome underpinning centronuclear myopathy

    Boglarka Zambo, Evelina Edelweiss ... Gergo Gogl
    Research Article

    Truncation of the protein-protein interaction SH3 domain of the membrane remodeling Bridging Integrator 1 (BIN1, Amphiphysin 2) protein leads to centronuclear myopathy. Here, we assessed the impact of a set of naturally observed, previously uncharacterized BIN1 SH3 domain variants using conventional in vitro and cell-based assays monitoring the BIN1 interaction with dynamin 2 (DNM2) and identified potentially harmful ones that can be also tentatively connected to neuromuscular disorders. However, SH3 domains are typically promiscuous and it is expected that other, so far unknown partners of BIN1 exist besides DNM2, that also participate in the development of centronuclear myopathy. In order to shed light on these other relevant interaction partners and to get a holistic picture of the pathomechanism behind BIN1 SH3 domain variants, we used affinity interactomics. We identified hundreds of new BIN1 interaction partners proteome-wide, among which many appear to participate in cell division, suggesting a critical role of BIN1 in the regulation of mitosis. Finally, we show that the identified BIN1 mutations indeed cause proteome-wide affinity perturbation, signifying the importance of employing unbiased affinity interactomic approaches.

    1. Biochemistry and Chemical Biology
    2. Chromosomes and Gene Expression

    Human DDX6 regulates translation and decay of inefficiently translated mRNAs

    Ramona Weber, Chung-Te Chang
    Research Article

    Recent findings indicate that the translation elongation rate influences mRNA stability. One of the factors that has been implicated in this link between mRNA decay and translation speed is the yeast DEAD-box helicase Dhh1p. Here, we demonstrated that the human ortholog of Dhh1p, DDX6, triggers the deadenylation-dependent decay of inefficiently translated mRNAs in human cells. DDX6 interacts with the ribosome through the Phe-Asp-Phe (FDF) motif in its RecA2 domain. Furthermore, RecA2-mediated interactions and ATPase activity are both required for DDX6 to destabilize inefficiently translated mRNAs. Using ribosome profiling and RNA sequencing, we identified two classes of endogenous mRNAs that are regulated in a DDX6-dependent manner. The identified targets are either translationally regulated or regulated at the steady-state-level and either exhibit signatures of poor overall translation or of locally reduced ribosome translocation rates. Transferring the identified sequence stretches into a reporter mRNA caused translation- and DDX6-dependent degradation of the reporter mRNA. In summary, these results identify DDX6 as a crucial regulator of mRNA translation and decay triggered by slow ribosome movement and provide insights into the mechanism by which DDX6 destabilizes inefficiently translated mRNAs.