A plant-like mechanism coupling m6A reading to polyadenylation safeguards transcriptome integrity and developmental gene partitioning in Toxoplasma
- Université Grenoble Alpes, France
- European Molecular Biology Laboratory, France
- Institut Laue-Langevin, France
- 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:
Article and author information
Author details
Alexandre Bougdour
Mohamed-ali Hakimi
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.
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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
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