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1. Exon junction complex dependent mRNA localization is linked to centrosome organization during ciliogenesis

Author

Oh Sung Kwon, Rahul Mishra, Adham Safieddine, Emeline Coleno, Quentin Alasseur, Marion Faucourt, Isabelle Barbosa, Edouard Bertrand, Nathalie Spassky, and Hervé Le Hir

Subjects
Science
Abstract

Exon junction complexes (EJCs) that mark untranslated mRNA are involved in transport, translation and nonsense-mediated mRNA decay. Here the authors show centrosomal localization of EJCs which appears to be required for both the localization of NIN mRNA around centrosomes and ciliogenesis.

Published
2021
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2. A choreography of centrosomal mRNAs reveals a conserved localization mechanism involving active polysome transport

Author

Adham Safieddine, Emeline Coleno, Soha Salloum, Arthur Imbert, Abdel-Meneem Traboulsi, Oh Sung Kwon, Frederic Lionneton, Virginie Georget, Marie-Cécile Robert, Thierry Gostan, Charles-Henri Lecellier, Racha Chouaib, Xavier Pichon, Hervé Le Hir, Kazem Zibara, Florian Mueller, Thomas Walter, Marion Peter, and Edouard Bertrand

Subjects
Science
Abstract

Centrosomes function as microtubule organizing centers where several mRNAs accumulate. By employing high-throughput single molecule FISH screening, the authors discover that 8 human mRNAs localize to centrosomes with unique cell cycle dependent patterns using an active polysome targeting mechanism.

Published
2021
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3. The Arabidopsis SWI/SNF protein BAF60 mediates seedling growth control by modulating DNA accessibility

Author

Teddy Jégu, Alaguraj Veluchamy, Juan S. Ramirez-Prado, Charley Rizzi-Paillet, Magalie Perez, Anaïs Lhomme, David Latrasse, Emeline Coleno, Serge Vicaire, Stéphanie Legras, Bernard Jost, Martin Rougée, Fredy Barneche, Catherine Bergounioux, Martin Crespi, Magdy M. Mahfouz, Heribert Hirt, Cécile Raynaud, and Moussa Benhamed

Subjects
SWI/SNF, Chromatin, Morphogenesis, Phytochrome Interacting Factor, G-box, DNA accessibility, Biology (General), QH301-705.5, Genetics, QH426-470
Abstract

Abstract Background Plant adaptive responses to changing environments involve complex molecular interplays between intrinsic and external signals. Whilst much is known on the signaling components mediating diurnal, light, and temperature controls on plant development, their influence on chromatin-based transcriptional controls remains poorly explored. Results In this study we show that a SWI/SNF chromatin remodeler subunit, BAF60, represses seedling growth by modulating DNA accessibility of hypocotyl cell size regulatory genes. BAF60 binds nucleosome-free regions of multiple G box-containing genes, opposing in cis the promoting effect of the photomorphogenic and thermomorphogenic regulator Phytochrome Interacting Factor 4 (PIF4) on hypocotyl elongation. Furthermore, BAF60 expression level is regulated in response to light and daily rhythms. Conclusions These results unveil a short path between a chromatin remodeler and a signaling component to fine-tune plant morphogenesis in response to environmental conditions.

Published
2017
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6. HT-smFISH: a cost effective and flexible workflow for high-throughput single molecule RNA imaging

Author

Adham Safieddine, Emeline Coleno, Frederic Lionneton, Abdel-Meneem Traboulsi, Soha Salloum, Charles-Henri Lecellier, Thierry Gostan, Virginie Georget, Cédric Hassen-Khodja, Arthur Imbert, Florian Mueller, Thomas Walter, Marion Peter, Edouard Bertrand, Institut de Génétique Moléculaire de Montpellier (IGMM), Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM), Institut de Biologie Paris Seine (IBPS), Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Institut de génétique humaine (IGH), BioCampus (BCM), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM), Méthodes et Algorithmes pour la Bioinformatique (MAB), Laboratoire d'Informatique de Robotique et de Microélectronique de Montpellier (LIRMM), Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM), Centre de Bioinformatique (CBIO), Mines Paris - PSL (École nationale supérieure des mines de Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL), Institut Curie [Paris], Imagerie et Modélisation - Imaging and Modeling, Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Cité (UPCité), Cancer et génome: Bioinformatique, biostatistiques et épidémiologie d'un système complexe, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut Curie [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut de Recherche en Cancérologie de Montpellier (IRCM - U1194 Inserm - UM), CRLCC Val d'Aurelle - Paul Lamarque-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Montpellier (UM), and ANR-19-P3IA-0001,PRAIRIE,PaRis Artificial Intelligence Research InstitutE(2019)

Subjects
RNA metabolism, Molecular imaging probes, High-throughput screening HTS, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, [SDV.BC.BC]Life Sciences [q-bio]/Cellular Biology/Subcellular Processes [q-bio.SC], General Biochemistry, Genetics and Molecular Biology
Abstract

International audience; The ability to visualize RNA in its native subcellular environment by using single-molecule fluorescence in situ hybridization (smFISH) has reshaped our understanding of gene expression and cellular functions. A major hindrance of smFISH is the difficulty to perform systematic experiments in medium- or high-throughput formats, principally because of the high cost of generating the individual fluorescent probe sets. Here, we present high-throughput smFISH (HT-smFISH), a simple and cost-efficient method for imaging hundreds to thousands of single endogenous RNA molecules in 96-well plates. HT-smFISH uses RNA probes transcribed in vitro from a large pool of unlabeled oligonucleotides. This allows the generation of individual probes for many RNA species, replacing commercial DNA probe sets. HT-smFISH thus reduces costs per targeted RNA compared with many smFISH methods and is easily scalable and flexible in design. We provide a protocol that combines oligo pool design, probe set generation, optimized hybridization conditions and guidelines for image acquisition and analysis. The pipeline requires knowledge of standard molecular biology tools, cell culture and fluorescence microscopy. It is achievable in ~20 d. In brief, HT-smFISH is tailored for medium- to high-throughput screens that image RNAs at single-molecule sensitivity.

Published
2023
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7. FISH-quant v2: a scalable and modular tool for smFISH image analysis

Author

Arthur Imbert, Wei Ouyang, Adham Safieddine, Emeline Coleno, Christophe Zimmer, Edouard Bertrand, Thomas Walter, Florian Mueller, Centre de Bioinformatique (CBIO), Mines Paris - PSL (École nationale supérieure des mines de Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL), Cancer et génome: Bioinformatique, biostatistiques et épidémiologie d'un système complexe, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut Curie [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Royal Institute of Technology [Stockholm] (KTH ), Institut de Biologie Paris Seine (IBPS), Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Institut de génétique humaine (IGH), Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM), Imagerie et Modélisation - Imaging and Modeling, Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Cité (UPCité), Centre de Bioinformatique, Biostatistique et Biologie Intégrative (C3BI), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), ANR-19-P3IA-0001,PRAIRIE,PaRis Artificial Intelligence Research InstitutE(2019), ANR-19-CE12-0007,TRANSFACT,Caracterisation des foyers de traduction: de nouvelles structures qui organisement finement le cytoplasme(2019), ANR-10-INBS-0004,France-BioImaging,Développment d'une infrastructure française distribuée coordonnée(2010), Walter, Thomas, PaRis Artificial Intelligence Research InstitutE - - PRAIRIE2019 - ANR-19-P3IA-0001 - P3IA - VALID, Caracterisation des foyers de traduction: de nouvelles structures qui organisement finement le cytoplasme - - TRANSFACT2019 - ANR-19-CE12-0007 - AAPG2019 - VALID, and Développment d'une infrastructure française distribuée coordonnée - - France-BioImaging2010 - ANR-10-INBS-0004 - INBS - VALID

Subjects
[INFO.INFO-CV]Computer Science [cs]/Computer Vision and Pattern Recognition [cs.CV], RNA localization, smFISH, Single Molecule Imaging, [INFO.INFO-BT] Computer Science [cs]/Biotechnology, [INFO.INFO-CV] Computer Science [cs]/Computer Vision and Pattern Recognition [cs.CV], image analysis, image analysis RNA localization transcription smFISH, Nanotechnology, RNA, RNA, Messenger, [INFO.INFO-BT]Computer Science [cs]/Biotechnology, transcription, Molecular Biology, In Situ Hybridization, Fluorescence
Abstract

International audience; Regulation of RNA abundance and localization is a key step in gene expression control. Single-molecule RNA fluorescence in situ hybridization (smFISH) is a widely used single-cell-single-molecule imaging technique enabling quantitative studies of gene expression and its regulatory mechanisms. Today, these methods are applicable at a large scale, which in turn come with a need for adequate tools for data analysis and exploration. Here, we present FISH-quant v2, a highly modular tool accessible for both experts and non-experts. Our user-friendly package allows the user to segment nuclei and cells, detect isolated RNAs, decompose dense RNA clusters, quantify RNA localization patterns and visualize these results both at the single-cell level and variations within the cell population. This tool was validated and applied on large-scale smFISH image data sets, revealing diverse subcellular RNA localization patterns and a surprisingly high degree of cell-to-cell heterogeneity.

Published
2021
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8. The kinesin KIF1C transports APC-dependent mRNAs to cell protrusions

Author

Emeline Coleno, Marion Peter, Marie-Cécile Robert, Tianhong Wang, Xavier Pichon, Kazem Zibara, Arthur Imbert, Edouard Bertrand, Florian Mueller, Thomas Walter, Stavroula Mili, Racha Chouaib, Konstadinos Moissoglu, Institut de Génétique Moléculaire de Montpellier (IGMM), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS), Franche-Comté Électronique Mécanique, Thermique et Optique - Sciences et Technologies (UMR 6174) (FEMTO-ST), Université de Technologie de Belfort-Montbeliard (UTBM)-Ecole Nationale Supérieure de Mécanique et des Microtechniques (ENSMM)-Université de Franche-Comté (UFC), Université Bourgogne Franche-Comté [COMUE] (UBFC)-Université Bourgogne Franche-Comté [COMUE] (UBFC)-Centre National de la Recherche Scientifique (CNRS), Laboratoire d'économie et de sociologie du travail (LEST), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Hospices Civils de Lyon (HCL), Imagerie et Modélisation - Imaging and Modeling, Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM), Equipe labellisée Ligue contre le Cancer, National Cancer Institute [Bethesda] (NCI-NIH), National Institutes of Health [Bethesda] (NIH), Institut de génétique humaine (IGH), Cancer et génome: Bioinformatique, biostatistiques et épidémiologie d'un système complexe, Institut Curie [Paris]-MINES ParisTech - École nationale supérieure des mines de Paris, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM), Lebanese University [Beirut] (LU), Centre de Bioinformatique, Biostatistique et Biologie Intégrative (C3BI), This project was supported by France BioImaging (ANR-10-INBS-04), the Agence Nationale de la Recherche (ANR-11-BSV8-018-02, ANR-14-CE10-0018-01 and ANR-19-CE12-0007-03), and the Ligue Nationale Contre le Cancer and the Fondation pour la Recherche Médicale. This work was supported by the Labex EpiGenMed, from the framework 'Investissements d’avenir'. This work was supported in part by the Intramural Research Program of the National Cancer Institute, NIH., ANR-10-INBS-0004,France-BioImaging,Développment d'une infrastructure française distribuée coordonnée(2010), ANR-11-BSV8-0018,EJCbirth,Impacts de la transcription par l'ARN polymérase II sur l'assemblage de l'EJC(2011), ANR-14-CE10-0018,HI-FISH,Etude systématique de l'expression des gènes à l'échelle des molécules uniques d'ARN(2014), ANR-19-CE12-0007,TRANSFACT,Caracterisation des foyers de traduction: de nouvelles structures qui organisement finement le cytoplasme(2019), ANR-10-LABX-0012,EpiGenMed,From Genome and Epigenome to Molecular Medicine: turning new paradigms in biology into the therapeutic strategies of tomorrow(2010), Mines Paris - PSL (École nationale supérieure des mines de Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut Curie [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Cité (UPCité), and Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS)

Subjects
Cytoplasm, RNA localization, [SDV]Life Sciences [q-bio], Adenomatous Polyposis Coli Protein, RNA transport, Kinesins, Biology, Article, RNA Transport, 03 medical and health sciences, local translation, 0302 clinical medicine, Live cell imaging, MRNA transport, Animals, Humans, RNA, Messenger, Molecular Biology, cytoplasmic protrusions, 030304 developmental biology, 0303 health sciences, Messenger RNA, RNA, Cell biology, Kinesin, Cell Surface Extensions, 030217 neurology & neurosurgery, HeLa Cells
Abstract

International audience; RNA localization and local translation are important for numerous cellular functions. In mammals, a class of mRNAs localize to cytoplasmic protrusions in an APC-dependent manner, with roles during cell migration. Here, we investigated this localization mechanism. We found that the KIF1C motor interacts with APC-dependent mRNAs and is required for their localization. Live cell imaging revealed rapid, active transport of single mRNAs over long distances that requires both microtubules and KIF1C. Two color imaging directly revealed single mRNAs transported by single KIF1C motors, with the 3’UTR being sufficient to trigger KIF1C-dependent RNA transport and localization. Moreover, KIF1C remained associated with peripheral, multimeric RNA clusters and was required for their formation. These results reveal a widespread RNA transport pathway in mammalian cells, in which the KIF1C motor has a dual role in transporting RNAs and clustering them within cytoplasmic protrusions. Interestingly, KIF1C also transports its own mRNA suggesting a possible feedback loop acting at the level of mRNA transport.

Published
2021
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9. Exon junction complex dependent mRNA localization is linked to centrosome organization during ciliogenesis

Author

Marion Faucourt, Adham Safieddine, Edouard Bertrand, Isabelle Barbosa, Quentin Alasseur, Rahul Mishra, Hervé Le Hir, Oh Sung Kwon, Nathalie Spassky, Emeline Coleno, Institut de biologie de l'ENS Paris (UMR 8197/1024) (IBENS), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Département de Biologie - ENS Paris, École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Institut de Génétique Moléculaire de Montpellier (IGMM), Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM), Département de Biologie - ENS Paris, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS), Institut de biologie de l'ENS Paris (IBENS), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Département de Biologie - ENS Paris, Institut de biologie de l'Ecole Normale Supérieure (IBENS), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL), École normale supérieure - Paris (ENS-PSL), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure - Paris (ENS-PSL), and bertrand, edouard

Subjects
0301 basic medicine, Translation, Centriole, [SDV]Life Sciences [q-bio], General Physics and Astronomy, Cell Cycle Proteins, Autoantigens, Microtubules, RNA Transport, DEAD-box RNA Helicases, Mice, Exon, 0302 clinical medicine, Neural Stem Cells, Basal body, Pericentriolar material, 0303 health sciences, Multidisciplinary, Cell Cycle, 030302 biochemistry & molecular biology, High-throughput screening, Nuclear Proteins, RNA-Binding Proteins, Exons, Cell biology, [SDV] Life Sciences [q-bio], Microtubule-Associated Proteins, Science, Molecular imaging, Biology, Article, General Biochemistry, Genetics and Molecular Biology, Cell-cycle exit, 03 medical and health sciences, Ciliogenesis, Animals, Humans, Cilia, RNA, Messenger, Cell Proliferation, 030304 developmental biology, Microtubule nucleation, Centrosome, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, General Chemistry, Cytoskeletal Proteins, 030104 developmental biology, Protein Biosynthesis, Eukaryotic Initiation Factor-4A, Exon junction complex, 030217 neurology & neurosurgery
Abstract

Exon junction complexes (EJCs) mark untranslated spliced mRNAs and are crucial for the mRNA lifecycle. An imbalance in EJC dosage alters mouse neural stem cell (mNSC) division and is linked to human neurodevelopmental disorders. In quiescent mNSC and immortalized human retinal pigment epithelial (RPE1) cells, centrioles form a basal body for ciliogenesis. Here, we report that EJCs accumulate at basal bodies of mNSC or RPE1 cells and decline when these cells differentiate or resume growth. A high-throughput smFISH screen identifies two transcripts accumulating at centrosomes in quiescent cells, NIN and BICD2. In contrast to BICD2, the localization of NIN transcripts is EJC-dependent. NIN mRNA encodes a core component of centrosomes required for microtubule nucleation and anchoring. We find that EJC down-regulation impairs both pericentriolar material organization and ciliogenesis. An EJC-dependent mRNA trafficking towards centrosome and basal bodies might contribute to proper mNSC division and brain development., Exon junction complexes (EJCs) that mark untranslated mRNA are involved in transport, translation and nonsense-mediated mRNA decay. Here the authors show centrosomal localization of EJCs which appears to be required for both the localization of NIN mRNA around centrosomes and ciliogenesis.

Published
2021
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10. FISH-quant v2: a scalable and modular analysis tool for smFISH image analysis

Author

Edouard Bertrand, Christophe Zimmer, Thomas Walter, Arthur Imbert, Adham Safieddine, Wei Ouyang, Emeline Coleno, Florian Mueller, Institut de génétique humaine (IGH), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS), Royal Institute of Technology [Stockholm] (KTH ), Institut de Biologie Paris Seine (IBPS), Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Imagerie et Modélisation - Imaging and Modeling, Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Cité (UPCité), Centre de Bioinformatique (CBIO), Mines Paris - PSL (École nationale supérieure des mines de Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL), Cancer et génome: Bioinformatique, biostatistiques et épidémiologie d'un système complexe, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut Curie [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Université Paris sciences et lettres (PSL), Centre de Bioinformatique, Biostatistique et Biologie Intégrative (C3BI), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), ANR-19-CE12-0007,TRANSFACT,Caracterisation des foyers de traduction: de nouvelles structures qui organisement finement le cytoplasme(2019), ANR-19-P3IA-0001,PRAIRIE,PaRis Artificial Intelligence Research InstitutE(2019), MINES ParisTech - École nationale supérieure des mines de Paris, Institut Curie [Paris]-MINES ParisTech - École nationale supérieure des mines de Paris, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM), Sorbonne Université (SU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Biologie mitochondriale – Mitochondrial biology, and Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS)-Université Paris Cité (UPC)

Subjects
Messenger RNA, RNA localization, business.industry, Computer science, [INFO.INFO-CV]Computer Science [cs]/Computer Vision and Pattern Recognition [cs.CV], RNA, Computational biology, Modular design, [SDV.BIBS]Life Sciences [q-bio]/Quantitative Methods [q-bio.QM], Image (mathematics), [INFO.INFO-TI]Computer Science [cs]/Image Processing [eess.IV], Scalability, Gene expression, %22">Fish , [INFO.INFO-BI]Computer Science [cs]/Bioinformatics [q-bio.QM], business
Abstract

Regulation of RNA abundance and localization is a key step in gene expression control. Single-molecule RNA fluorescence in-situ hybridization (smFISH) is a widely used single-cell-single-molecule imaging technique enabling a quantitative understanding of gene expression and its regulatory mechanisms. Recent progress in experimental techniques provides larger data-sets, requiring adequate tools for data analysis and exploration. Here, we present FISH-quant v2, a highly modular analysis tool accessible both for non-experts and experts, which we validated and applied on large-scale smFISH image datasets. Our package allows the user to detect isolated and clustered mRNA spots, segment nuclei and cells, quantify RNA localization patterns and visualize these results at the single-cell level.

Published
2021
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11. The kinesin KIF1C transports APC-dependent mRNAs to cell protrusions

Author

Thomas Walter, Xavier Pichon, Edouard Bertrand, Racha Chouaib, Florian Mueller, Emeline Coleno, Tianhong Wang, Marion Peter, Kazem Zibara, Arthur Imbert, Stavroula Mili, Konstadinos Moissoglu, Centre de Bioinformatique (CBIO), MINES ParisTech - École nationale supérieure des mines de Paris, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL), Cancer et génome: Bioinformatique, biostatistiques et épidémiologie d'un système complexe, Institut Curie [Paris]-MINES ParisTech - École nationale supérieure des mines de Paris, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM), and Université Paris sciences et lettres (PSL)

Subjects
0303 health sciences, Messenger RNA, RNA localization, Chemistry, RNA, RNA transport, [INFO.INFO-CV]Computer Science [cs]/Computer Vision and Pattern Recognition [cs.CV], [SDV.BIBS]Life Sciences [q-bio]/Quantitative Methods [q-bio.QM], Cell biology, 03 medical and health sciences, 0302 clinical medicine, Cytoplasm, Live cell imaging, [INFO.INFO-TI]Computer Science [cs]/Image Processing [eess.IV], Kinesin, MRNA transport, [INFO.INFO-BI]Computer Science [cs]/Bioinformatics [q-bio.QM], 030217 neurology & neurosurgery, 030304 developmental biology
Abstract

RNA localization and local translation are important for numerous cellular functions. In mammals, a class of mRNAs localize to cytoplasmic protrusions in an APC-dependent manner, with roles during cell migration. Here, we investigated this localization mechanism. We found that the KIF1C motor interacts with APC-dependent mRNAs and is required for their localization. Live cell imaging revealed rapid, active transport of single mRNAs over long distances that requires both microtubules and KIF1C. Two color imaging directly showed single mRNAs transported by single KIF1C motors, with the 3’UTR being sufficient to trigger KIF1C-dependent RNA transport and localization. Moreover, KIF1C remained associated with peripheral, multimeric RNA clusters and was required for their formation. These results reveal an RNA transport pathway in mammalian cells, in which the KIF1C motor has a dual role both in transporting RNAs and in promoting their clustering within cytoplasmic protrusions. Interestingly, KIF1C also transports its own mRNA suggesting a possible feedback loop acting at the level of mRNA transport.

Published
2021

12. A choreography of centrosomal mRNAs reveals a conserved localization mechanism involving active polysome transport

Author

Arthur Imbert, Marie-Cécile Robert, Hervé Le Hir, Emeline Coleno, Thierry Gostan, Virginie Georget, Adham Safieddine, Xavier Pichon, Frédéric Lionneton, Florian Mueller, Thomas Walter, Charles-Henri Lecellier, Racha Chouaib, Edouard Bertrand, Soha Salloum, Abdel-Meneem Traboulsi, Kazem Zibara, Marion Peter, Oh Sung Kwon, Institut de Génétique Moléculaire de Montpellier (IGMM), Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM), Equipe labellisée Ligue contre le Cancer, Centre de Bioinformatique (CBIO), MINES ParisTech - École nationale supérieure des mines de Paris, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL), Institut Curie [Paris], Institut de biologie de l'ENS Paris (UMR 8197/1024) (IBENS), Département de Biologie - ENS Paris, École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), BioCampus Montpellier (BCM), Université Montpellier 1 (UM1)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS), Laboratory of Stem Cells [Lebanese, Beirut] (ER045-PRASE), Lebanese University [Beirut] (LU), Imagerie et Modélisation - Imaging and Modeling, Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), This project was supported by France BioImaging (ANR-10-INBS-04), by the Agence Nationale de la Recherche (ANR-11-BSV8-018-02 and ANR-14-CE10-0018-01), the Fondation pour la Recherche Médicale ('Bioinformatics' grant), the Institut Pasteur, the Ligue Nationale Contre le Cancer, and the Labex EpiGenMed from the framework 'Investissements d’avenir'. This work was also funded in part by the French government under the management of Agence Nationale de la Recherche as part of the 'Investissements d’avenir' program, reference ANR-19-P3IA-0001 (PRAIRIE 3IA Institute)., ANR-19-P3IA-0001,PRAIRIE,PaRis Artificial Intelligence Research InstitutE(2019), ANR-10-INBS-0004,France-BioImaging,Développment d'une infrastructure française distribuée coordonnée(2010), ANR-11-BSV8-0018,EJCbirth,Impacts de la transcription par l'ARN polymérase II sur l'assemblage de l'EJC(2011), ANR-14-CE10-0018,HI-FISH,Etude systématique de l'expression des gènes à l'échelle des molécules uniques d'ARN(2014), Institut National de la Santé et de la Recherche Médicale (INSERM), Centre National de la Recherche Scientifique (CNRS)-Institut Pasteur [Paris], Université Paris sciences et lettres (PSL), Institut de génétique humaine (IGH), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS), Mines Paris - PSL (École nationale supérieure des mines de Paris), Institut de biologie de l'ENS Paris (IBENS), École normale supérieure - Paris (ENS-PSL), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure - Paris (ENS-PSL), BioCampus (BCM), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), LE HIR, Hervé, PaRis Artificial Intelligence Research InstitutE - - PRAIRIE2019 - ANR-19-P3IA-0001 - P3IA - VALID, Développment d'une infrastructure française distribuée coordonnée - - France-BioImaging2010 - ANR-10-INBS-0004 - INBS - VALID, BLANC - Impacts de la transcription par l'ARN polymérase II sur l'assemblage de l'EJC - - EJCbirth2011 - ANR-11-BSV8-0018 - BLANC - VALID, Appel à projets générique - Etude systématique de l'expression des gènes à l'échelle des molécules uniques d'ARN - - HI-FISH2014 - ANR-14-CE10-0018 - Appel à projets générique - VALID, Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Département de Biologie - ENS Paris, and Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)

Subjects
0301 basic medicine, General Physics and Astronomy, ASPM, Cell Cycle Proteins, smFISH, RNA Transport, 0302 clinical medicine, local translation, Gene expression, [SDV.BC.IC]Life Sciences [q-bio]/Cellular Biology/Cell Behavior [q-bio.CB], centrosomes, Cycloheximide, Multidisciplinary, High-throughput screening, Translation (biology), Translocon, Cell biology, [SDV.BBM.GTP] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], Drosophila, Puromycin, Science, Mitosis, Molecular imaging, Spindle Apparatus, Biology, General Biochemistry, Genetics and Molecular Biology, Article, 03 medical and health sciences, Open Reading Frames, Microtubule, Polysome, [SDV.BBM.GTP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], conserved polysome choreography at centrosomes mRNA localization, Animals, Humans, RNA, Messenger, Centrosome, RNA metabolism, polysome imaging, cotranslational targeting, General Chemistry, NUMA1, 030104 developmental biology, Polyribosomes, 030217 neurology & neurosurgery, HeLa Cells
Abstract

Local translation allows for a spatial control of gene expression. Here, we use high-throughput smFISH to screen centrosomal protein-coding genes, and we describe 8 human mRNAs accumulating at centrosomes. These mRNAs localize at different stages during cell cycle with a remarkable choreography, indicating a finely regulated translational program at centrosomes. Interestingly, drug treatments and reporter analyses reveal a common translation-dependent localization mechanism requiring the nascent protein. Using ASPM and NUMA1 as models, single mRNA and polysome imaging reveals active movements of endogenous polysomes towards the centrosome at the onset of mitosis, when these mRNAs start localizing. ASPM polysomes associate with microtubules and localize by either motor-driven transport or microtubule pulling. Remarkably, the Drosophila orthologs of the human centrosomal mRNAs also localize to centrosomes and also require translation. These data identify a conserved family of centrosomal mRNAs that localize by active polysome transport mediated by nascent proteins., Centrosomes function as microtubule organizing centers where several mRNAs accumulate. By employing high-throughput single molecule FISH screening, the authors discover that 8 human mRNAs localize to centrosomes with unique cell cycle dependent patterns using an active polysome targeting mechanism.

Published
2021
Full Text
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13. A Dual Protein-mRNA Localization Screen Reveals Compartmentalized Translation and Widespread Co-translational RNA Targeting

Author

Thomas Walter, Emeline Coleno, Florian Mueller, Adham Safieddine, Xavier Pichon, Nikolay Tsanov, Arthur Imbert, Marie-Cécile Robert, Aubin Samacoits, Edouard Bertrand, Racha Chouaib, Anthony A. Hyman, Kazem Zibara, Christophe Zimmer, Matthias Peter, Ina Poser, Oh Sung Kwon, Hervé Le Hir, Abdel-Meneem Traboulsi, Institut de Génétique Moléculaire de Montpellier (IGMM), Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM), Laboratory of Stem Cells [Lebanese, Beirut] (ER045-PRASE), Lebanese University [Beirut] (LU), Centre de Bioinformatique (CBIO), MINES ParisTech - École nationale supérieure des mines de Paris, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL), Cancer et génome: Bioinformatique, biostatistiques et épidémiologie d'un système complexe, Institut Curie [Paris]-MINES ParisTech - École nationale supérieure des mines de Paris, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut de biologie de l'ENS Paris (IBENS), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Département de Biologie - ENS Paris, École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Imagerie et Modélisation - Imaging and Modeling, Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), Centre de Bioinformatique, Biostatistique et Biologie Intégrative (C3BI), Max Planck Institute of Molecular Cell Biology and Genetics (MPI-CBG), Max-Planck-Gesellschaft, Equipe labellisée Ligue contre le Cancer, This project was supported by France BioImaging (ANR-10-INBS-04), the Agence Nationale de la Recherche (ANR-11-BSV8-018-02 and ANR-14-CE10-0018-01), the Institut Pasteur, the Ligue Nationale Contre le Cancer, and the Fondation pour la Recherche Médicale (FRM grant to E.B.). Furthermore, this work was supported by the French government under management of Agence Nationale de la Recherche as part of the 'Investissements d'avenir' program, reference ANR-19-P3IA-0001 (PRAIRIE 3IA Institute) and by the Labex EpiGenMed from the framework 'Investissements d’avenir.', ANR-10-INBS-0004,France-BioImaging,Développment d'une infrastructure française distribuée coordonnée(2010), ANR-11-BSV8-0018,EJCbirth,Impacts de la transcription par l'ARN polymérase II sur l'assemblage de l'EJC(2011), ANR-14-CE10-0018,HI-FISH,Etude systématique de l'expression des gènes à l'échelle des molécules uniques d'ARN(2014), ANR-19-P3IA-0001,PRAIRIE,PaRis Artificial Intelligence Research InstitutE(2019), ANR-10-LABX-0012,EpiGenMed,From Genome and Epigenome to Molecular Medicine: turning new paradigms in biology into the therapeutic strategies of tomorrow(2010), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS), Mines Paris - PSL (École nationale supérieure des mines de Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut Curie [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Département de Biologie - ENS Paris, École normale supérieure - Paris (ENS-PSL), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure - Paris (ENS-PSL), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), Institut de biologie de l'ENS Paris (UMR 8197/1024) (IBENS), Centre National de la Recherche Scientifique (CNRS)-Institut Pasteur [Paris], Zimmer, Christophe, Développment d'une infrastructure française distribuée coordonnée - - France-BioImaging2010 - ANR-10-INBS-0004 - INBS - VALID, BLANC - Impacts de la transcription par l'ARN polymérase II sur l'assemblage de l'EJC - - EJCbirth2011 - ANR-11-BSV8-0018 - BLANC - VALID, Appel à projets générique - Etude systématique de l'expression des gènes à l'échelle des molécules uniques d'ARN - - HI-FISH2014 - ANR-14-CE10-0018 - Appel à projets générique - VALID, PaRis Artificial Intelligence Research InstitutE - - PRAIRIE2019 - ANR-19-P3IA-0001 - P3IA - VALID, and Laboratoires d'excellence - From Genome and Epigenome to Molecular Medicine: turning new paradigms in biology into the therapeutic strategies of tomorrow - - EpiGenMed2010 - ANR-10-LABX-0012 - LABX - VALID

Subjects
RNA localization, Endosome, ASPM, translation factories, [SDV.BC.BC]Life Sciences [q-bio]/Cellular Biology/Subcellular Processes [q-bio.SC], Biology, General Biochemistry, Genetics and Molecular Biology, smFISH, Cell Line, 03 medical and health sciences, 0302 clinical medicine, local translation, Polysome, Gene expression, [SDV.BC.BC] Life Sciences [q-bio]/Cellular Biology/Subcellular Processes [q-bio.SC], Humans, RNA, Messenger, Molecular Biology, Gene, 030304 developmental biology, Centrosome, 0303 health sciences, Messenger RNA, co-translational targeting Manuscript, Translation (biology), Cell Biology, Translocon, Cell biology, Protein Transport, Gene Expression Regulation, Polyribosomes, Protein Biosynthesis, Beta-catenin, RNA, RNA transport, co-translational targeting, 030217 neurology & neurosurgery, Developmental Biology
Abstract

International audience; Local translation allows spatial control of gene expression. Here, we performed a dual protein-mRNA localization screen, using smFISH on 523 human cell lines expressing GFP-tagged genes. 32 mRNAs displayed specific cytoplasmic localizations with local translation at unexpected locations, including cytoplasmic protrusions, cell edges, endosomes, Golgi, the nuclear envelope, and centrosomes, the latter being cell-cycle-dependent. Automated classification of mRNA localization patterns revealed a high degree of intercellular heterogeneity. Surprisingly, mRNA localization frequently required ongoing translation, indicating widespread co-translational RNA targeting. Interestingly, while P-body accumulation was frequent (15 mRNAs), four mRNAs accumulated in foci that were distinct structures. These foci lacked the mature protein, but nascent polypeptide imaging showed that they were specialized translation factories. For β-catenin, foci formation was regulated by Wnt, relied on APC-dependent polysome aggregation, and led to nascent protein degradation. Thus, translation factories uniquely regulate nascent protein metabolism and create a fine granular compartmentalization of translation.

Published
2020
Full Text
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14. A conserved choreography of mRNAs at centrosomes reveals a localization mechanism involving active polysome transport

Author

Hervé Le Hir, Abdel-Meneem Traboulsi, Virginie Georget, Emeline Coleno, Xavier Pichon, Kazem Zibara, Thierry Gostan, Racha Chouaib, Oh Sung Kwon, Edouard Bertrand, Soha Salloum, Charles Lecellier, Frédéric Lionneton, Marion Peter, Marie-Cécile Robert, Adham Safieddine, Institut de Génétique Moléculaire de Montpellier (IGMM), Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM), Institut de biologie de l'ENS Paris (UMR 8197/1024) (IBENS), Département de Biologie - ENS Paris, École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), BioCampus Montpellier (BCM), Université Montpellier 1 (UM1)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS), Lebanese University [Beirut] (LU), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Université Montpellier 1 (UM1)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Montpellier (UM), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Département de Biologie - ENS Paris, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure - Paris (ENS Paris), and Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)

Subjects
polysome imaging, 0303 health sciences, [SDV]Life Sciences [q-bio], cotranslational targeting, ASPM, Translation (biology), Biology, NUMA1, Translocon, high-throughput screening, smFISH, Cell biology, 03 medical and health sciences, local translation, 0302 clinical medicine, Microtubule, Centrosome, Polysome, Gene expression, conserved polysome choreography at centrosomes mRNA localization, centrosomes, Mitosis, 030217 neurology & neurosurgery, 030304 developmental biology
Abstract

Local translation allows for a spatial control of gene expression. Here, we used high-throughput smFISH to screen centrosomal protein-coding genes, and we describe 8 human mRNAs accumulating at centrosomes. These mRNAs localize at different stages during cell cycle with a remarkable choreography, indicating a finely regulated translational program at centrosomes. Interestingly, drug treatments and reporter analyses revealed a common translation-dependent localization mechanism requiring the nascent protein. Using ASPM and NUMA1 as models, single mRNA and polysome imaging revealed active movements of endogenous polysomes towards the centrosome at the onset of mitosis, when these mRNAs start localizing. ASPM polysomes associate with microtubules and localize by either motor-driven transport or microtubule pulling. Remarkably, the Drosophila orthologs of the human centrosomal mRNAs also localize to centrosomes and also require translation. These data identify a conserved family of centrosomal mRNAs that localize by active polysomes transport mediated by nascent proteins.; Local translation allows for a spatial control of gene expression. Here, we used high-throughput smFISH to screen centrosomal protein-coding genes, and we describe 8 human mRNAs accumulating at centrosomes. These mRNAs localize at different stages during cell cycle with a remarkable choreography, indicating a finely regulated translational program at centrosomes. Interestingly, drug treatments and reporter analyses revealed a common translation-dependent localization mechanism requiring the nascent protein. Using ASPM and NUMA1 as models, single mRNA and polysome imaging revealed active movements of endogenous polysomes towards the centrosome at the onset of mitosis, when these mRNAs start localizing. ASPM polysomes associate with microtubules and localize by either motor-driven transport or microtubule pulling. Remarkably, the Drosophila orthologs of the human centrosomal mRNAs also localize to centrosomes and also require translation. These data identify a conserved family of centrosomal mRNAs that localize by active polysomes transport mediated by nascent proteins.

Published
2020
Full Text
View/download PDF

15. A localization screen reveals translation factories and widespread co-translational RNA targeting

Author

Ina Poser, Xavier Pichon, Edouard Bertrand, Nikolay Tsanov, Christophe Zimmer, Emeline Coleno, Hervé Le Hir, Oh Sung Kwon, Adham Safieddine, Thomas Walter, Marie-Cécile Robert, Marion Peter, Abdel-Meneem Traboulsi, Arthur Imbert, Kazem Zibara, Florian Mueller, Aubin Samacoits, Racha Chouaib, Anthony A. Hyman, Institut de Génétique Moléculaire de Montpellier (IGMM), Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM), Equipe labelisée Ligue Contre le Cancer, Laboratory of Stem Cells [Lebanese, Beirut] (ER045-PRASE), Lebanese University [Beirut] (LU), Faculty of Sciences [Lebanese University], Equipe labellisée Ligue contre le Cancer, Centre de Bioinformatique (CBIO), MINES ParisTech - École nationale supérieure des mines de Paris, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL), Cancer et génome: Bioinformatique, biostatistiques et épidémiologie d'un système complexe, Institut Curie [Paris]-MINES ParisTech - École nationale supérieure des mines de Paris, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut de biologie de l'ENS Paris (UMR 8197/1024) (IBENS), Département de Biologie - ENS Paris, École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Imagerie et Modélisation - Imaging and Modeling, Centre National de la Recherche Scientifique (CNRS)-Institut Pasteur [Paris], Centre de Bioinformatique, Biostatistique et Biologie Intégrative (C3BI), Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), Max Planck Institute of Molecular Cell Biology and Genetics (MPI-CBG), Max-Planck-Gesellschaft, Université Paris sciences et lettres (PSL), This project was supported by France BioImaging (ANR-10-INBS-04), the Agence Nationale de la Recherche (ANR-11-BSV8-018-02 and ANR-14-CE10-0018-01), the Institut Pasteur, the Ligue Nationale Contre le Cancer and the Fondation pour la Recherche Médicale (FRM grant to EB). This work was supported by the Labex EpiGenMed, from the framework 'Investissements d’avenir'., ANR-10-INBS-0004,France-BioImaging,Développment d'une infrastructure française distribuée coordonnée(2010), ANR-14-CE10-0018,HI-FISH,Etude systématique de l'expression des gènes à l'échelle des molécules uniques d'ARN(2014), Lemesle, Marie, Développment d'une infrastructure française distribuée coordonnée - - France-BioImaging2010 - ANR-10-INBS-0004 - INBS - VALID, Appel à projets générique - Etude systématique de l'expression des gènes à l'échelle des molécules uniques d'ARN - - HI-FISH2014 - ANR-14-CE10-0018 - Appel à projets générique - VALID, Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS), Faculty of Sciences [Lebanese University] | Faculté des Sciences [Université Libanaise], Mines Paris - PSL (École nationale supérieure des mines de Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut Curie [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut de biologie de l'ENS Paris (IBENS), École normale supérieure - Paris (ENS-PSL), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure - Paris (ENS-PSL), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Département de Biologie - ENS Paris, and Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)

Subjects
0303 health sciences, Messenger RNA, RNA localization, Translation (biology), translation factories, Biology, Translocon, smFISH, Cell biology, 03 medical and health sciences, 0302 clinical medicine, local translation, Cytoplasm, [SDV.SPEE] Life Sciences [q-bio]/Santé publique et épidémiologie, Polysome, co-translational targeting +Equal contributions, Gene expression, [SDV.SPEE]Life Sciences [q-bio]/Santé publique et épidémiologie, RNA transport, Gene, 030217 neurology & neurosurgery, 030304 developmental biology
Abstract

Publié sur BioRxiv le 21 mai 2020 : https://www.biorxiv.org/content/10.1101/2020.05.20.106989v1; Local translation allows a spatial control of gene expression. Here, we performed a dual protein/mRNA localization screen, using smFISH on 523 human cell lines expressing GFP-tagged genes. A total of 32 mRNAs displayed specific cytoplasmic localizations, and we observed local translation at unexpected locations, including cytoplasmic protrusions, cell edges, endosomes, Golgi, the nuclear envelope and centrosomes, the latter being cell cycle dependent. Quantitation of mRNA distribution and automatic pattern classification revealed a high degree of localization heterogeneity between cells. Surprisingly, mRNA localization frequently required ongoing translation, indicating widespread co-translational RNA targeting. Interestingly, while P-body accumulation was frequent (15 mRNAs), four mRNAs accumulated in foci that were distinct structures. These foci lacked the mature protein, but nascent polypeptide imaging showed that they were specialized translation factories. For β-catenin, foci formation was regulated by Wnt, relied on APC-dependent polysome aggregation, and led to nascent protein degradation. Thus, translation factories uniquely regulate nascent protein metabolism and create a fine granular compartmentalization of translation.

Published
2020

16. The Arabidopsis SWI/SNF protein BAF60 mediates seedling growth control by modulating DNA accessibility

Author

Juan S. Ramirez-Prado, Charley Rizzi-Paillet, Heribert Hirt, Teddy Jégu, Fredy Barneche, Stephanie Legras, Catherine Bergounioux, Emeline Coleno, Martin Rougée, Alaguraj Veluchamy, Bernard Jost, Cécile Raynaud, Moussa Benhamed, Serge Vicaire, David Latrasse, Martin Crespi, Anaïs Lhomme, Magdy M. Mahfouz, Magalie Perez, Institut des Sciences des Plantes de Paris-Saclay (IPS2 (UMR_9213 / UMR_1403)), Institut National de la Recherche Agronomique (INRA)-Université Paris-Sud - Paris 11 (UP11)-Université Paris Diderot - Paris 7 (UPD7)-Université d'Évry-Val-d'Essonne (UEVE)-Centre National de la Recherche Scientifique (CNRS), Université Sorbonne Paris Cité (COMUE) (USPC), Massachusetts General Hospital, Massachusetts General Hospital [Boston], Department of Genetics [Boston], Harvard Medical School [Boston] (HMS), Division of Biological and Environmental Sciences and Engineering, King Abdullah University of Science and Technology (KAUST), Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA), Institut de biologie de l'ENS Paris (IBENS), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Département de Biologie - ENS Paris, École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Universite Paris Sud, LabExSaclay Plant Sciences-SPS [ANR-10-LABX-0040-SPS], Université de Strasbourg (UNISTRA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Institut de biologie de l'ENS Paris (UMR 8197/1024) (IBENS), Département de Biologie - ENS Paris, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), École normale supérieure - Paris (ENS-PSL), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure - Paris (ENS-PSL), and Benhamed, Moussa

Subjects
0301 basic medicine, Phytochrome Interacting Factor, lcsh:QH426-470, DNA, Plant, Chromosomal Proteins, Non-Histone, [SDV]Life Sciences [q-bio], DNA accessibility, Morphogenesis, Regulator, PIF4, Arabidopsis, Biology, 03 medical and health sciences, Gene Expression Regulation, Plant, Basic Helix-Loop-Helix Transcription Factors, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, SWI/SNF, Chromatin, G-box, Gene, lcsh:QH301-705.5, Regulator gene, Genetics, Phytochrome, Arabidopsis Proteins, Research, fungi, biology.organism_classification, Chromatin Assembly and Disassembly, Hypocotyl, Cell biology, Nucleosomes, DNA-Binding Proteins, lcsh:Genetics, 030104 developmental biology, lcsh:Biology (General), Seedlings, Transcription Factors
Abstract

Background Plant adaptive responses to changing environments involve complex molecular interplays between intrinsic and external signals. Whilst much is known on the signaling components mediating diurnal, light, and temperature controls on plant development, their influence on chromatin-based transcriptional controls remains poorly explored. Results In this study we show that a SWI/SNF chromatin remodeler subunit, BAF60, represses seedling growth by modulating DNA accessibility of hypocotyl cell size regulatory genes. BAF60 binds nucleosome-free regions of multiple G box-containing genes, opposing in cis the promoting effect of the photomorphogenic and thermomorphogenic regulator Phytochrome Interacting Factor 4 (PIF4) on hypocotyl elongation. Furthermore, BAF60 expression level is regulated in response to light and daily rhythms. Conclusions These results unveil a short path between a chromatin remodeler and a signaling component to fine-tune plant morphogenesis in response to environmental conditions. Electronic supplementary material The online version of this article (doi:10.1186/s13059-017-1246-7) contains supplementary material, which is available to authorized users.

Published
2017
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17. Introns Protect Eukaryotic Genomes from Transcription-Associated Genetic Instability

Author

Vincent Géli, Ana Rita Grosso, Guilhem Janbon, Emeline Coleno, Sérgio F. de Almeida, Amandine Bonnet, Adrien Presle, Sree Rama Chaitanya Sridhara, Abdessamad El-Kaoutari, Benoit Palancade, Institut Jacques Monod (IJM (UMR_7592)), Université Paris Diderot - Paris 7 (UPD7)-Centre National de la Recherche Scientifique (CNRS), Faculdade de Medicina [Lisboa], Universidade de Lisboa = University of Lisbon (ULISBOA), Centre de Recherche en Cancérologie de Marseille (CRCM), Aix Marseille Université (AMU)-Institut Paoli-Calmettes, Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Biologie des ARN des Pathogènes fongiques - RNA Biology of Fungal Pathogens, Institut Pasteur [Paris] (IP), This work was supported by CNRS (to B.P.), Fondation ARC pour la Recherche sur le Cancer (to B.P.), Ligue Nationale contre le Cancer (to B.P., fellowship to A.B. and Equipe Labellisée 2014 to V.G.), EMBO (short-term fellowship to A.B.), Cancéropole PACA (fellowship to A.E.), and Fundaçao para a Ciencia e Tecnologia, Portugal (PTDC/BIM-ONC/0016-2014 to S.F.d.A. and IF/00510/2014 to A.R.G.). Bioinformatic and computing support at CRCM was provided by the CRCM Integrative Bioinformatics and Datacentre IT and Scientific Computing platforms., Universidade de Lisboa (ULISBOA), Institut Pasteur [Paris], Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Paoli-Calmettes, Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Aix Marseille Université (AMU), and Biologie des ARN des Pathogènes fongiques

Subjects
0301 basic medicine, Transcription, Genetic, MESH: Introns, [SDV]Life Sciences [q-bio], Candida glabrata, mRNA splicing, MESH: Genotype, 0302 clinical medicine, MESH: Structure-Activity Relationship, Minor spliceosome, Gene Expression Regulation, Fungal, Databases, Genetic, DNA, Fungal, MESH: Candida glabrata, MESH: Databases, Genetic, Genetics, Splice site mutation, MESH: Genomic Instability, Nucleic Acid Heteroduplexes, genetic instability, MESH: Cryptococcus neoformans, Group II intron, R-loops, MESH: Saccharomyces cerevisiae, messenger ribonucleoparticle, Phenotype, MESH: Nucleic Acid Conformation, MESH: Schizosaccharomyces, Ribonucleoproteins, RNA splicing, MESH: Fungal Proteins, transcription, MESH: Spliceosomes, MESH: Gene Expression Regulation, Fungal, MESH: Computational Biology, Spliceosome, Genotype, intron, RNA Splicing, Saccharomyces cerevisiae, Biology, MESH: Phenotype, Genomic Instability, Cell Line, Fungal Proteins, Structure-Activity Relationship, 03 medical and health sciences, mRNP, MESH: Nucleic Acid Heteroduplexes, Schizosaccharomyces, Humans, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Molecular Biology, Gene, MESH: DNA Damage, MESH: Humans, MESH: RNA, Fungal, MESH: Transcription, Genetic, Intron, Computational Biology, RNA, RNA, Fungal, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, Cell Biology, Introns, recombination, MESH: Cell Line, MESH: DNA, Fungal, MESH: Ribonucleoproteins, 030104 developmental biology, Cryptococcus neoformans, Spliceosomes, Nucleic Acid Conformation, MESH: RNA Splicing, 030217 neurology & neurosurgery, DNA Damage
Abstract

International audience; Transcription is a source of genetic instability that can notably result from the formation of genotoxic DNA:RNA hybrids, or R-loops, between the nascent mRNA and its template. Here we report an unexpected function for introns in counteracting R-loop accumulation in eukaryotic genomes. Deletion of endogenous introns increases R-loop formation, while insertion of an intron into an intronless gene suppresses R-loop accumulation and its deleterious impact on transcription and recombination in yeast. Recruitment of the spliceosome onto the mRNA, but not splicing per se, is shown to be critical to attenuate R-loop formation and transcription-associated genetic instability. Genome-wide analyses in a number of distant species differing in their intron content, including human, further revealed that intron-containing genes and the intron-richest genomes are best protected against R-loop accumulation and subsequent genetic instability. Our results thereby provide a possible rationale for the conservation of introns throughout the eukaryotic lineage.

Published
2017
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