A plant-like mechanism coupling m6A reading to polyadenylation safeguards transcriptome integrity and developmental gene partitioning in Toxoplasma

  1. Dayana C Farhat
  2. Matthew W Bowler
  3. Guillaume Communie
  4. Dominique Pontier
  5. Lucid Belmudes
  6. Caroline Mas
  7. Charlotte Corrao
  8. Yohann Couté
  9. Alexandre Bougdour
  10. Thierry Lagrange
  11. Mohamed-ali Hakimi  Is a corresponding author
  12. Christopher Swale  Is a corresponding author
  1. Université Grenoble Alpes, France
  2. European Molecular Biology Laboratory, France
  3. Institut Laue-Langevin, France
  4. CNRS, France

Abstract

Correct 3'end processing of mRNAs is one of the regulatory cornerstones of gene expression. In a parasite that must adapt to the regulatory requirements of its multi-host life style, there is a need to adopt additional means to partition the distinct transcriptional signatures of the closely and tandemly-arranged stage specific genes. In this study, we report our findings in T. gondii of an m6A-dependent 3'end polyadenylation serving as a transcriptional barrier at these loci. We identify the core polyadenylation complex within T. gondii and establish CPSF4 as a reader for m6A-modified mRNAs, via a YTH domain within its C-terminus, a feature which is shared with plants. We bring evidence of the specificity of this interaction both biochemically, and by determining the crystal structure at high resolution of the T. gondii CPSF4-YTH in complex with an m6A modified RNA. We show that the loss of m6A, both at the level of its deposition or its recognition was associated with an increase in aberrantly elongated chimeric mRNAs emanating from impaired transcriptional termination, a phenotype previously noticed in the plant model Arabidopsis thaliana. Nanopore direct RNA sequencing shows the occurrence of transcriptional read-through breaching into downstream repressed stage-specific genes, in the absence of either CPSF4 or the m6A RNA methylase components in both T. gondii and A. thaliana. Taken together, our results shed light on an essential regulatory mechanism coupling the pathways of m6A metabolism directly to the cleavage and polyadenylation processes, one that interestingly seem to serve, in both T. gondii and A. thaliana, as a guardian against aberrant transcriptional read-throughs.

Data availability

The Nanopore RNAseq data have been deposited in NCBI's SRA data PRJNA705300. The MS proteomics data have been deposited to the ProteomeXchange Consortium through the PRIDE partner repository with the dataset identifier PXD024326. Sequencing data have been deposited in GEO under accession code GSE168155:

The following data sets were generated

Article and author information

Author details

  1. Dayana C Farhat

    Team Host-pathogen interactions and Immunity to Infection, Université Grenoble Alpes, Grenoble, France
    Competing interests
    The authors declare that no competing interests exist.
  2. Matthew W Bowler

    European Molecular Biology Laboratory, Grenoble, France
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-0465-3351
  3. Guillaume Communie

    Institut Laue-Langevin, Institut Laue-Langevin, Grenoble, France
    Competing interests
    The authors declare that no competing interests exist.
  4. Dominique Pontier

    UMR5096, CNRS, Perpignan, France
    Competing interests
    The authors declare that no competing interests exist.
  5. Lucid Belmudes

    BIG-BGE, Université Grenoble Alpes, Grenoble, France
    Competing interests
    The authors declare that no competing interests exist.
  6. Caroline Mas

    Integrated Structural Biology Grenoble (ISBG), Université Grenoble Alpes, Grenoble, France
    Competing interests
    The authors declare that no competing interests exist.
  7. Charlotte Corrao

    Team Host-pathogen interactions and Immunity to Infection, Université Grenoble Alpes, Grenoble, France
    Competing interests
    The authors declare that no competing interests exist.
  8. Yohann Couté

    Institut de Biosciences et Biotechnologies de Grenoble, Université Grenoble Alpes, Grenoble, France
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-3896-6196
  9. Alexandre Bougdour

    Institut de Biosciences et Biotechnologies de Grenoble, Université Grenoble Alpes, Grenoble, France
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-5895-0020
  10. Thierry Lagrange

    UMR5096, CNRS, Grenoble, France
    Competing interests
    The authors declare that no competing interests exist.
  11. Mohamed-ali Hakimi

    Team Host-pathogen interactions and Immunity to Infection, Université Grenoble Alpes, Grenoble, France
    For correspondence
    mohamed-ali.hakimi@inserm.fr
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-2547-8233
  12. Christopher Swale

    Team Host-pathogen interactions and Immunity to Infection, Université Grenoble Alpes, Grenoble, France
    For correspondence
    christopher.swale@univ-grenoble-alpes.fr
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-9739-7774

Funding

Agence Nationale de la Recherche (Laboratoire d'Excellence (LabEx) ParaFrap [ANR-11-LABX-0024])

  • Dayana C Farhat
  • Mohamed-ali Hakimi
  • Christopher Swale

Agence Nationale de la Recherche (Project HostQuest,ANR-18-CE15-0023)

  • Charlotte Corrao
  • Alexandre Bougdour
  • Mohamed-ali Hakimi
  • Christopher Swale

European Research Council (ERC Consolidator Grant N{degree sign}614880 Hosting TOXO)

  • Mohamed-ali Hakimi

Fondation pour la Recherche Médicale (FRM FDT201904008364)

  • Dayana C Farhat
  • Mohamed-ali Hakimi

Agence Nationale de la Recherche (Proteomics French Infrastructure,Infrastructure Nationale en Biologie et Santé,ANR-10-INBS-08)

  • Lucid Belmudes
  • Yohann Couté

Agence Nationale de la Recherche (Laboratoires d'Excellences (LABEX) TULIP (ANR-10-LABX-41)")

  • Dominique Pontier
  • Thierry Lagrange

Agence Nationale de la Recherche (École Universitaire de Recherche (EUR)" TULIP-GS (ANR-18-EURE-0019)")

  • Dominique Pontier
  • Thierry Lagrange

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

Copyright

© 2021, Farhat 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.

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. Dayana C Farhat
  2. Matthew W Bowler
  3. Guillaume Communie
  4. Dominique Pontier
  5. Lucid Belmudes
  6. Caroline Mas
  7. Charlotte Corrao
  8. Yohann Couté
  9. Alexandre Bougdour
  10. Thierry Lagrange
  11. Mohamed-ali Hakimi
  12. Christopher Swale
(2021)
A plant-like mechanism coupling m6A reading to polyadenylation safeguards transcriptome integrity and developmental gene partitioning in Toxoplasma
eLife 10:e68312.
https://doi.org/10.7554/eLife.68312

Share this article

https://doi.org/10.7554/eLife.68312

Further reading

    1. Microbiology and Infectious Disease
    Dhaval Ghone, Edward L Evans ... Aussie Suzuki
    Research Article

    Virion Infectivity Factor (Vif) of the Human Immunodeficiency Virus type 1 (HIV-1) targets and degrades cellular APOBEC3 proteins, key regulators of intrinsic and innate antiretroviral immune responses, thereby facilitating HIV-1 infection. While Vif’s role in degrading APOBEC3G is well-studied, Vif is also known to cause cell cycle arrest, but the detailed nature of Vif’s effects on the cell cycle has yet to be delineated. In this study, we employed high-temporal resolution single-cell live imaging and super-resolution microscopy to monitor individual cells during Vif-induced cell cycle arrest. Our findings reveal that Vif does not affect the G2/M boundary as previously thought. Instead, Vif triggers a unique and robust pseudo-metaphase arrest, distinct from the mild prometaphase arrest induced by Vpr. During this arrest, chromosomes align properly and form the metaphase plate, but later lose alignment, resulting in polar chromosomes. Notably, Vif, unlike Vpr, significantly reduces the levels of both Protein Phosphatase 1 (PP1) and 2 A (PP2A) at kinetochores, which regulate chromosome-microtubule interactions. These results unveil a novel role for Vif in kinetochore regulation that governs the spatial organization of chromosomes during mitosis.

    1. Computational and Systems Biology
    2. Microbiology and Infectious Disease
    Ruihan Dong, Rongrong Liu ... Cheng Zhu
    Research Article

    Antimicrobial peptides (AMPs) are attractive candidates to combat antibiotic resistance for their capability to target biomembranes and restrict a wide range of pathogens. It is a daunting challenge to discover novel AMPs due to their sparse distributions in a vast peptide universe, especially for peptides that demonstrate potencies for both bacterial membranes and viral envelopes. Here, we establish a de novo AMP design framework by bridging a deep generative module and a graph-encoding activity regressor. The generative module learns hidden ‘grammars’ of AMP features and produces candidates sequentially pass antimicrobial predictor and antiviral classifiers. We discovered 16 bifunctional AMPs and experimentally validated their abilities to inhibit a spectrum of pathogens in vitro and in animal models. Notably, P076 is a highly potent bactericide with the minimal inhibitory concentration of 0.21 μM against multidrug-resistant Acinetobacter baumannii, while P002 broadly inhibits five enveloped viruses. Our study provides feasible means to uncover the sequences that simultaneously encode antimicrobial and antiviral activities, thus bolstering the function spectra of AMPs to combat a wide range of drug-resistant infections.