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1. Virus Evolution Faced to Multiple Host Targets: The Potyvirus—Pepper Case Study

Author

Tamisier, Lucie, Lacombe, Séverine, Caranta, Carole, Gallois, Jean-Luc, Moury, Benoît, Ahmed, Rafi, Series Editor, Akira, Shizuo, Series Editor, Casadevall, Arturo, Series Editor, Galan, Jorge E., Series Editor, Garcia-Sastre, Adolfo, Series Editor, Malissen, Bernard, Series Editor, Rappuoli, Rino, Series Editor, Domingo, Esteban, editor, Schuster, Peter, editor, Elena, Santiago F., editor, and Perales, Celia, editor

Published
2023
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4. Genome Editing Tools for Potato Improvement

Author

Ravet, Karl, Sevestre, François, Chauvin, Laura, Chauvin, Jean-Eric, Lairy-Joly, Gisèle, Katz, Andrew, Devaux, Pierre, Szydlowski, Nicolas, Gallois, Jean-Luc, Pearce, Stephen, Veillet, Florian, Zhao, Kaijun, editor, Mishra, Rukmini, editor, and Joshi, Raj Kumar, editor

Published
2022
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6. AtHVA22a, a plant‐specific homologue of Reep/DP1/Yop1 family proteins is involved in turnip mosaic virus propagation.

Author

Xue, Mingshuo, Sofer, Luc, Simon, Vincent, Arvy, Nathalie, Diop, Mamoudou, Lion, Roxane, Beucher, Guillaume, Bordat, Amandine, Tilsner, Jens, Gallois, Jean‐Luc, and German‐Retana, Sylvie

Subjects
TURNIP mosaic virus, MEMBRANE proteins, PLANT proteins, BARLEY, PROTEINS
Abstract

The movement of potyviruses, the largest genus of single‐stranded, positive‐sense RNA viruses responsible for serious diseases in crops, is very complex. As potyviruses developed strategies to hijack the host secretory pathway and plasmodesmata (PD) for their transport, the goal of this study was to identify membrane and/or PD‐proteins that interact with the 6K2 protein, a potyviral protein involved in replication and cell‐to‐cell movement of turnip mosaic virus (TuMV). Using split‐ubiquitin membrane yeast two‐hybrid assays, we screened an Arabidopsis cDNA library for interactors of TuMV6K2. We isolated AtHVA22a (Hordeum vulgare abscisic acid responsive gene 22), which belongs to a multigenic family of transmembrane proteins, homologous to Receptor expression‐enhancing protein (Reep)/Deleted in polyposis (DP1)/Yop1 family proteins in animal and yeast. HVA22/DP1/Yop1 family genes are widely distributed in eukaryotes, but the role of HVA22 proteins in plants is still not well known, although proteomics analysis of PD fractions purified from Arabidopsis suspension cells showed that AtHVA22a is highly enriched in a PD proteome. We confirmed the interaction between TuMV6K2 and AtHVA22a in yeast, as well as in planta by using bimolecular fluorescence complementation and showed that TuMV6K2/AtHVA22a interaction occurs at the level of the viral replication compartment during TuMV infection. Finally, we showed that the propagation of TuMV is increased when AtHVA22a is overexpressed in planta but slowed down upon mutagenesis of AtHVA22a by CRISPR‐Cas9. Altogether, our results indicate that AtHVA22a plays an agonistic effect on TuMV propagation and that the C‐terminal tail of the protein is important in this process. [ABSTRACT FROM AUTHOR]

Published
2024
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9. Interdependence of a kinase and its cognate substrate plasma membrane nanoscale dynamics underlies Arabidopsis response to viral infection

Author

Jolivet, Marie-Dominique, primary, Deroubaix, Anne-Flore, additional, Boudsocq, Marie, additional, Abel, Nikolaj B., additional, Rocher, Marion, additional, Robbe, Terezinha, additional, Wattelet-Boyer, Valérie, additional, Huard, Jennifer, additional, Lefebvre, Dorian, additional, Lu, Yi-Ju, additional, Day, Brad, additional, Saias, Grégoire, additional, Ahmed, Jahed, additional, Cotelle, Valérie, additional, Giovinazzo, Nathalie, additional, Gallois, Jean-Luc, additional, Yamaji, Yasuyuki, additional, German-Retana, Sylvie, additional, Gronnier, Julien, additional, Ott, Thomas, additional, Mongrand, Sébastien, additional, and Germain, Véronique, additional

Published
2023
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11. Interdependence of a kinase and its cognate substrate plasma membrane nanoscale dynamics underliesArabidopsisresponse to viral infection

Author

Jolivet, Marie-Dominique, primary, Deroubaix, Anne-Flore, additional, Boudsocq, Marie, additional, Abel, Nikolaj B., additional, Rocher, Marion, additional, Robbe, Terezinha, additional, Wattelet-Boyer, Valérie, additional, Huard, Jennifer, additional, Lefebvre, Dorian, additional, Lu, Yi-Ju, additional, Day, Brad, additional, Saias, Grégoire, additional, Ahmed, Jahed, additional, Cotelle, Valérie, additional, Giovinazzo, Nathalie, additional, Gallois, Jean-Luc, additional, Yamaji, Yasuyuki, additional, German-Retana, Sylvie, additional, Gronnier, Julien, additional, Ott, Thomas, additional, Mongrand, Sébastien, additional, and Germain, Véronique, additional

Published
2023
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15. An iterative gene‐editing strategy broadens eIF4E1 genetic diversity in Solanum lycopersicum and generates resistance to multiple potyvirus isolates

Author

Kuroiwa, Kyoka, primary, Danilo, Benoit, additional, Perrot, Laura, additional, Thenault, Christina, additional, Veillet, Florian, additional, Delacote, Fabien, additional, Duchateau, Philippe, additional, Nogué, Fabien, additional, Mazier, Marianne, additional, and Gallois, Jean‐Luc, additional

Published
2023
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17. Rôle des facteurs d'initiation de la traduction 4E dans la résistance des plantes aux potyvirus : de la découverte des résistances naturelles à l'édition des gènes.

Author

Gallois, Jean-Luc and German-Retana, Sylvie

Subjects
*PLANT breeding, *PLANT viruses, *GENOME editing, *PLANT species, *CROPS, *PEPPERS
Abstract

Resistance to viruses is an important aspect of plant breeding. One way to achieve it is to select genetic resistances based on the susceptibility factors hijacked by the virus to infect the plants. Here, we recount work done on genes encoding translation initiation factors eIF4E, some of the most successful targets for obtaining resistance to potyviruses, starting from their characterization 20 years ago. With examples from different plant species, pepper, tomato, tobacco and arabidopsis, we present the basis of this type of resistances and their characteristics, highlighting the role of gene redundancy among 4E factors, their specificity for the virus and the need for the plant of a trade-off between resistance and development. Finally, we show how the new genome editing techniques could be used in plant breeding to develop eIF4E-based resistances in crops, mimicking the functional alleles that have been selected during evolution in many crops. [ABSTRACT FROM AUTHOR]

Published
2023
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18. An iterative gene editing strategy broadens eIF4E1 genetic diversity in Solanum lycopersicum and generates resistance to several potyvirus isolates

Author

Kuroiwa, Kyoka, primary, Danilo, Benoit, additional, Perrot, Laura, additional, Thenault, Christina, additional, Veillet, Florian, additional, Delacote, Fabien, additional, Duchateau, Philippe, additional, Nogué, Fabien, additional, Mazier, Marianne, additional, and Gallois, Jean-Luc, additional

Published
2022
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21. Monitoring systemic infection by cucumber mosaic virus using a small fluorescent protein iLOV in plants

Author

Kawakubo, Ayaka, Gallois, Jean-Luc, 1000090315606, Nakahara, Kenji S., Kawakubo, Ayaka, Gallois, Jean-Luc, 1000090315606, and Nakahara, Kenji S.

Abstract

Tracking virus infection in plants is useful to assess plant susceptibility or resistance. The recombinant cucumber mosaic virus (CMV) expressing a 2b-fused green fluorescent protein (GFP, 25 kDa) gradually loses (within weeks) the GFP gene in plants. Here, we constructed CMV expressing a flavin-based small fluorescent protein (iLOV, 10 kDa). CMV-iLOV still retains iLOV after 28 dpi in upper leaves of inoculated Nicotiana benthamiana and allows long-term monitoring of its distribution in plants. Using CMV-iLOV, we showed that pot1, a recessive allele for resistance to potyviruses, does not confer CMV resistance in tomato.

Published
2022

25. Gene Editing in Potato Using Technology

Author

Chauvin, Laura, Sevestre, François, Lukan, Tjaša, Nogué, Fabien, Gallois, Jean-Luc, Chauvin, Jean-Eric, Veillet, Florian, Institut de Génétique, Environnement et Protection des Plantes (IGEPP), Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-INSTITUT AGRO Agrocampus Ouest, Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro), Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS), Miniaturisation pour la Synthèse, l’Analyse et la Protéomique - UAR 3290 (MSAP), Department of Biotechnology and Systems Biology [Ljubljana, Slovenia], National Institute of Biology [Ljubljana] (NIB), Institut Jean-Pierre Bourgin (IJPB), AgroParisTech-Université Paris-Saclay-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Université de Rennes (UR)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-INSTITUT AGRO Agrocampus Ouest, Génétique et Amélioration des Fruits et Légumes (GAFL), and Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)

Subjects
[SDV.GEN.GPL]Life Sciences [q-bio]/Genetics/Plants genetics, Single-guide RNA, Agrobacterium tumefaciens, Protoplasts, [SDV]Life Sciences [q-bio], fungi, food and beverages, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, CRISPR-Cas9, Gene editing, HRM analysis, Potato, Plant regeneration
Abstract

International audience; Genome editing in the cultivated potato (Solanum tuberosum), a vegetatively propagated and highly heterozygous species, constitutes a promising trail to directly improve traits into elite cultivars. With the recent and successful development of the clustered regularly interspaced short palindromic repeat (CRISPR)-Cas9 system in eukaryotic cells, the plant science community has gained access to a powerful, inexpensive, and easy-to-use toolbox to target and inactivate/modify specific genes. The specificity and versatility of the CRISPR-Cas9 system rely on a variable 20 bp spacer sequence at the 5' end of a single-guide RNA (sgRNA), which directs the SpCas9 (Streptococcus pyogenes) nuclease to cut the target DNA at a precise locus with no or low off-target events. Using this system, we and other teams were able to knock out specific genes in potato through the error-prone non-homologous end-joining (NHEJ) DNA repair mechanism. In this chapter, we describe strategies to design and clone spacer sequences into CRISPR-SpCas9 plasmids. We show how these constructs can be used for Agrobacterium-mediated stable transformation or transient transfection of protoplasts, and we describe the optimization of these two delivery methods, as well as of the plant regeneration processes. Finally, the molecular screening and characterization of edited potato plants are also described, mainly relying on PCR-based methods such as high-resolution melt (HRM) analysis.

Published
2021

27. Comment les plantes se défendent-elles face aux virus ?

Author

German-Retana, Sylvie, PooggiN, Mikhail M., Gallois, Jean-Luc, ORTIZ, Diana, Moury, Benoît, Biologie du fruit et pathologie (BFP), Université de Bordeaux (UB)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Plant Health Institute of Montpellier (UMR PHIM), Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad)-Institut de Recherche pour le Développement (IRD)-Université de Montpellier (UM)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Génétique et Amélioration des Fruits et Légumes (GAFL), Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Unité de Pathologie Végétale (PV), Christian Lannou, Dominique Roby, Virginie Ravigné, Mourad Hannachi, and Benoit Moury

Subjects
[SDV.GEN.GPL]Life Sciences [q-bio]/Genetics/Plants genetics, résistance aux maladies, virus phytopathogène, variété résistante, Résistance quantitative, QTL, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, résistance qualitative, tolérance, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, pathologie végétale, ComputingMilieux_MISCELLANEOUS, [SDV.BV.PEP]Life Sciences [q-bio]/Vegetal Biology/Phytopathology and phytopharmacy
Abstract

International audience

Published
2021

28. A blue-print for gene function analysis through Base Editing in the model plant Physcomitrium (Physcomitrella) patens

Author

Guyon-Debast, Anouchka, Alboresi, Alessandro, Terret, Zoé, Charlot, Florence, Berthier, Floriane, Vendrell-Mir, Pol, Casacuberta, Josep M., Veillet, Florian, Morosinotto, Tomas, Gallois, Jean-Luc, Nogue, Fabien, Institut Jean-Pierre Bourgin (IJPB), AgroParisTech-Université Paris-Saclay-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Universita degli Studi di Padova, Génétique et Amélioration des Fruits et Légumes (GAFL), Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Spanish National Research Council (CSIC), Institut de Génétique, Environnement et Protection des Plantes (IGEPP), Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-AGROCAMPUS OUEST, Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), French National Research Agency French National Research Agency (ANR) [ANR11-BTBR-0001-GENIUS], LabEx Saclay Plant Sciences-SPS [ANR-10-LABX-0040-SPS], ANR-11-BTBR-0001,GENIUS,Ingénierie cellulaire : amélioration et innovation technologiques pour les plantes d'une agriculture(2011), University of Padova [Padova, Italy], Centre for Research in Agricultural Genomics (CRAG), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-INSTITUT AGRO Agrocampus Ouest, Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro), ANR-10-LABX-0040,SPS,Saclay Plant Sciences(2010), Agence Nationale de la Recherche (France), Institute of Plant Sciences Paris-Saclay, Università degli Studi di Padova = University of Padua (Unipd), and Université de Rennes (UR)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-INSTITUT AGRO Agrocampus Ouest

Subjects
Gene Editing, Physcomitrella patens, Adenine deaminase, Physcomitrium patens, APRT, Base editing, CRISPR, Cas9, Cytosine deaminase, Research, cytosine deaminase, [SDV]Life Sciences [q-bio], food and beverages, Bryopsida, Methods, Mutagenesis, Site-Directed, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, Clustered Regularly Interspaced Short Palindromic Repeats, CRISPR-Cas Systems
Abstract

CRISPR-Cas9 has proven to be highly valuable for genome editing in plants, including the model plant Physcomitrium patens. However, the fact that most of the editing events produced using the native Cas9 nuclease correspond to small insertions and deletions is a limitation., CRISPR-Cas9 base editors enable targeted mutation of single nucleotides in eukaryotic genomes and therefore overcome this limitation. Here, we report two programmable base-editing systems to induce precise cytosine or adenine conversions in P. patens., Using cytosine or adenine base editors, site-specific single-base mutations can be achieved with an efficiency up to 55%, without off-target mutations. Using the APT gene as a reporter of editing, we could show that both base editors can be used in simplex or multiplex, allowing for the production of protein variants with multiple amino-acid changes. Finally, we set up a co-editing selection system, named selecting modification of APRT to report gene targeting (SMART), allowing up to 90% efficiency site-specific base editing in P. patens., These two base editors will facilitate gene functional analysis in P. patens, allowing for site-specific editing of a given base through single sgRNA base editing or for in planta evolution of a given gene through the production of randomly mutagenised variants using multiple sgRNA base editing., The work, including study design, data collection, analysis and interpretation and manuscript writing, was supported by the French National Research Agency (ANR11-BTBR-0001-GENIUS). The IJPB benefits from the support of the LabEx Saclay Plant Sciences-SPS (ANR-10-LABX-0040-SPS).

Published
2021

29. A blueprint for gene function analysis through Base Editing in the model plant Physcomitrium (Physcomitrella) patens

Author

Guyon-Debast, Anouchka, Alboresi, Alessandro, Terret, Zoé, Charlot, Florence, Berthier, Floriane, Vendrell-Mir, Pol, Casacuberta i Suñer, Josep M., Veillet, Florian, Morosinotto, Tomas, Gallois, Jean-Luc, Nogué, Fabien, Guyon-Debast, Anouchka, Alboresi, Alessandro, Terret, Zoé, Charlot, Florence, Berthier, Floriane, Vendrell-Mir, Pol, Casacuberta i Suñer, Josep M., Veillet, Florian, Morosinotto, Tomas, Gallois, Jean-Luc, and Nogué, Fabien

Abstract

CRISPR-Cas9 has proven to be highly valuable for genome editing in plants, including the model plant Physcomitrium patens. However, the fact that most of the editing events produced using the native Cas9 nuclease correspond to small insertions and deletions is a limitation. CRISPR-Cas9 base editors enable targeted mutation of single nucleotides in eukaryotic genomes and therefore overcome this limitation. Here, we report two programmable base-editing systems to induce precise cytosine or adenine conversions in P. patens. Using cytosine or adenine base editors, site-specific single-base mutations can be achieved with an efficiency up to 55%, without off-target mutations. Using the APT gene as a reporter of editing, we could show that both base editors can be used in simplex or multiplex, allowing for the production of protein variants with multiple amino-acid changes. Finally, we set up a co-editing selection system, named selecting modification of APRT to report gene targeting (SMART), allowing up to 90% efficiency site-specific base editing in P. patens. These two base editors will facilitate gene functional analysis in P. patens, allowing for site-specific editing of a given base through single sgRNA base editing or for in planta evolution of a given gene through the production of randomly mutagenised variants using multiple sgRNA base editing.

Published
2021

30. A blueprint for gene function analysis through Base Editing in the model plant Physcomitrium (Physcomitrella) patens

Author

Agence Nationale de la Recherche (France), Institute of Plant Sciences Paris-Saclay, Guyon-Debast, Anouchka, Alboresi, Alessandro, Terret, Zoé, Charlot, Florence, Berthier, Floriane, Vendrell-Mir, Pol, Casacuberta, Josep M., Veillet, Florian, Morosinotto, Tomas, Gallois, Jean-Luc, Nogué, Fabien, Agence Nationale de la Recherche (France), Institute of Plant Sciences Paris-Saclay, Guyon-Debast, Anouchka, Alboresi, Alessandro, Terret, Zoé, Charlot, Florence, Berthier, Floriane, Vendrell-Mir, Pol, Casacuberta, Josep M., Veillet, Florian, Morosinotto, Tomas, Gallois, Jean-Luc, and Nogué, Fabien

Abstract

CRISPR-Cas9 has proven to be highly valuable for genome editing in plants, including the model plant Physcomitrium patens. However, the fact that most of the editing events produced using the native Cas9 nuclease correspond to small insertions and deletions is a limitation., CRISPR-Cas9 base editors enable targeted mutation of single nucleotides in eukaryotic genomes and therefore overcome this limitation. Here, we report two programmable base-editing systems to induce precise cytosine or adenine conversions in P. patens., Using cytosine or adenine base editors, site-specific single-base mutations can be achieved with an efficiency up to 55%, without off-target mutations. Using the APT gene as a reporter of editing, we could show that both base editors can be used in simplex or multiplex, allowing for the production of protein variants with multiple amino-acid changes. Finally, we set up a co-editing selection system, named selecting modification of APRT to report gene targeting (SMART), allowing up to 90% efficiency site-specific base editing in P. patens., These two base editors will facilitate gene functional analysis in P. patens, allowing for site-specific editing of a given base through single sgRNA base editing or for in planta evolution of a given gene through the production of randomly mutagenised variants using multiple sgRNA base editing.

Published
2021

32. A blueprint for gene function analysis through Base Editing in the model plantPhyscomitrium (Physcomitrella) patens

Author

Guyon‐Debast, Anouchka, primary, Alboresi, Alessandro, additional, Terret, Zoé, additional, Charlot, Florence, additional, Berthier, Floriane, additional, Vendrell‐Mir, Pol, additional, Casacuberta, Josep M., additional, Veillet, Florian, additional, Morosinotto, Tomas, additional, Gallois, Jean‐Luc, additional, and Nogué, Fabien, additional

Published
2021
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34. eIF4E-based resistance to potyviruses from natural variation to edited genes

Author

Gallois, Jean-Luc, Génétique et Amélioration des Fruits et Légumes (GAFL), and Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)

Subjects
[SDV.BIO]Life Sciences [q-bio]/Biotechnology, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, ComputingMilieux_MISCELLANEOUS
Abstract

International audience

Published
2020

35. Expanding the CRISPR Toolbox in P. patens Using SpCas9-NG Variant and Application for Gene and Base Editing in Solanaceae Crops

Author

Veillet, Florian, Perrot, Laura, Guyon-Debast, Anouchka, Kermarrec, Marie-Paule, Chauvin, Laura, Chauvin, Jean-Eric, Gallois, Jean-Luc, Mazier, Marianne, Nogué, Fabien, Institut de Génétique, Environnement et Protection des Plantes (IGEPP), Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-AGROCAMPUS OUEST, Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Génétique et Amélioration des Fruits et Légumes (GAFL), Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Institut Jean-Pierre Bourgin (IJPB), AgroParisTech-Université Paris-Saclay-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), ANR-10-LABX-0040-SPS, Agence Nationale de la Recherche, ANR-11-BTBR-0001,GENIUS,Ingénierie cellulaire : amélioration et innovation technologiques pour les plantes d'une agriculture(2011), ANR-17-EURE-0007,SPS-GSR,Ecole Universitaire de Recherche de Sciences des Plantes de Paris-Saclay(2017), Université de Rennes (UR)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-INSTITUT AGRO Agrocampus Ouest, and Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)

Subjects
SpCas9-NG, Physcomitrella patens, base editing, physcomitrella patens, tomato, lcsh:Chemistry, [SDV.GEN.GPL]Life Sciences [q-bio]/Genetics/Plants genetics, [SDV.BV.AP]Life Sciences [q-bio]/Vegetal Biology/Plant breeding, lcsh:Biology (General), lcsh:QD1-999, potato, CBE, xCas9, CRISPR-Cas9, lcsh:QH301-705.5, alternative PAM
Abstract

Genome editing has become a major tool for both functional studies and plant breeding in several species. Besides generating knockouts through the classical CRISPR-Cas9 system, recent development of CRISPR base editing holds great and exciting opportunities for the production of gain-of-function mutants. The PAM requirement is a strong limitation for CRISPR technologies such as base editing, because the base substitution mainly occurs in a small edition window. As precise single amino-acid substitution can be responsible for functions associated to some domains or agronomic traits, development of Cas9 variants with relaxed PAM recognition is of upmost importance for gene function analysis and plant breeding. Recently, the SpCas9-NG variant that recognizes the NGN PAM has been successfully tested in plants, mainly in monocotyledon species. In this work, we studied the efficiency of SpCas9-NG in the model moss Physcomitrella patens and two Solanaceae crops (Solanum lycopersicum and Solanum tuberosum) for both classical CRISPR-generated gene knock-out and cytosine base editing. We showed that the SpCas9-NG greatly expands the scope of genome editing by allowing the targeting of non-canonical NGT and NGA PAMs. The CRISPR toolbox developed in our study opens up new gene function analysis and plant breeding perspectives for model and crop plants.

Published
2020

43. Résistance aux virus: Quand la variabilité naturelle guide l'édition de gènes

Author

Gallois, Jean-Luc, Génétique et Amélioration des Fruits et Légumes (GAFL), Institut National de la Recherche Agronomique (INRA), and Gœthe Institut. Paris, FRA.

Subjects
[SDV]Life Sciences [q-bio], [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, ComputingMilieux_MISCELLANEOUS
Abstract

National audience

Published
2019

44. AllMine, a flexible pipeline for allele mining

Author

Bersez, Thomas, Gallois, Jean-Luc, Lagnel, Jacques, Génétique et Amélioration des Fruits et Légumes (GAFL), Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), and ANR-18-LEAP-0004,CASSANDRA,CASsava Sustainable Advancement & Nurturing by discovery of Disease Resistance Alleles(2018)

Subjects
[INFO.INFO-BI]Computer Science [cs]/Bioinformatics [q-bio.QM], ComputingMilieux_MISCELLANEOUS
Abstract

International audience

Published
2019

45. Mimicking natural polymorphism in eIF4E by CRISPR-Cas9 base editing is associated with resistance to potyviruses

Author

Bastet, Anna, Zafirov, Delyan, Giovinazzo, Nathalie, Guyon‐Debast, Anouchka, Nogué, Fabien, Robaglia, Christophe, Gallois, Jean‐Luc, Unité de recherche Génétique et amélioration des fruits et légumes (GALF), Institut National de la Recherche Agronomique (INRA), Luminy Génétique et Biophysique des Plantes (LGBP), Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Institut Jean-Pierre Bourgin (IJPB), Institut National de la Recherche Agronomique (INRA)-AgroParisTech, ANR-13-KBBE-0006,COBRA,A COmbination of systems Biology and experimental high-throughput approaches to engIneer durable Resistance against plAnt viruses in crops(2013), ANR-16-CE20-0008,PotyMove,Facteurs cellulaires recrutés par les potyvirus pour leur transport intercellulaire : de nouvelles sources de résistance des plantes?(2016), ANR-11-IDEX-02/10-LABX-0040,SPS,Saclay Plant Sciences(2011), Génétique et Amélioration des Fruits et Légumes (GAFL), Laboratoire de Génétique et Biophysique des Plantes -Faculté des Sciences de Luminy, Commissariat à l'énergie atomique et aux énergies alternatives (CEA), ANR-10-LABX-0040,SPS,Saclay Plant Sciences(2010), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), (AAP MARASME) ANR-KBBE COBRA (ANR-13-KBBE-0006), ANR-POTYMOVE (ANR-16-CE20-000803)Plant2Pro project ‘POTATOCRISP’. LabEx Saclay Plant Sciences-SPS (ANR-10-LABX- 0040-SPS)., ANR-11-IDEX-0002,UNITI,Université Fédérale de Toulouse(2011), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), and Inra Bap : AAP MARASME

Subjects
Arabidopsis thaliana, base editing, [SDV]Life Sciences [q-bio], Arabidopsis, Biotechnologies, polymorphisme, résistance aux agents pathogènes, potyvirus, Research Articles, Alleles, pisum sativum, Disease Resistance, Plant Diseases, Plant Proteins, cytidine deaminase, Gene Editing, Peas, food and beverages, résistance au virus, Plants, Genetically Modified, CRISPR-Cas9, eIF4E, Eukaryotic Initiation Factor-4E, CRISPR-Cas Systems, résistance génétique, CRISPR‐Cas9, Research Article
Abstract

International audience; In many crop species, natural variation in eIF4E proteins confers resistance to potyviruses. Gene editing offers new opportunities to transfer genetic resistance to crops that seem to lack natural eIF4E alleles. However, because eIF4E are physiologically important proteins, any introduced modification for virus resistance must not bring adverse phenotype effects. In this study, we assessed the role of amino acid substitutions encoded by a Pisum sativum eIF4E virus-resistance allele (W69L, T80D S81D, S84A, G114R and N176K) by introducing them independently into the Arabidopsis thaliana eIF4E1 gene, a susceptibility factor to the Clover yellow vein virus (ClYVV). Results show that most mutations were sufficient to prevent ClYVV accumulation in plants without affecting plant growth. In addition, two of these engineered resistance alleles can be combined with a loss-of-function eIFiso4E to expand the resistance spectrum to other potyviruses. Finally, we use CRISPR-nCas9-cytidine deaminase technology to convert the Arabidopsis eIF4E1 susceptibility allele into a resistance allele by introducing the N176K mutation with a single-point mutation through C-to-G base editing to generate resistant plants. This study shows how combining knowledge on pathogen susceptibility factors with precise genome-editing technologies offers a feasible solution for engineering transgene-free genetic resistance in plants, even across species barriers.

Published
2019

46. Analysis of the Tomato spotted wilt virus RNA-dependent RNA polymerase adaptative evolution and constrained domains using homology protein structure modeling

Author

Terret, Zoé, Bonnet, Grégori, Moury, Benoît, Gallois, Jean-Luc, Génétique et Amélioration des Fruits et Légumes (GAFL), Institut National de la Recherche Agronomique (INRA), Syngenta France, and Unité de Pathologie Végétale (PV)

Subjects
[SDV.SA]Life Sciences [q-bio]/Agricultural sciences, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, ComputingMilieux_MISCELLANEOUS, [SDV.BV.PEP]Life Sciences [q-bio]/Vegetal Biology/Phytopathology and phytopharmacy
Abstract

National audience

Published
2019

47. eIF4E-mediated resistance to viruses: from natural variation to synthetic alleles

Author

Gallois, Jean-Luc, Bastet, Anna, Gauffier, Camille, Robaglia, Christophe, Caranta, Carole, Génétique et Amélioration des Fruits et Légumes (GAFL), Institut National de la Recherche Agronomique (INRA), Aix Marseille Université (AMU), EMBO. Heidelberg, DEU., and University of Helsinki [Helsinki]. Helsinki, FIN.

Subjects
physiologie végétale, Vegetal Biology, approche génétique, variation génétique naturelle, food and beverages, facteur initiation eif4e, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, résistance au virus, Biologie végétale, ComputingMilieux_MISCELLANEOUS
Abstract

International audience; eIF4E translation initiation factors have emerged as major susceptibility factors for RNA viruses. Natural eIF4E-based resistance alleles are found in many species and are mostly variants that maintain the translation function of the protein. eIF4E genes represent major targets for engineering viral resistance, and gene-editing technologies can be used to make up for the lack of natural resistance alleles in some crops, often by knocking out eIF4E susceptibility factors. However, we report here how redundancy among eIF4E genes can restrict the efficient use of knockout alleles in breeding. We therefore discuss how gene-editing technologies can be used to design de novo functional alleles, using knowledge about the natural evolution of eIF4E genes in different species, to drive resistance to viruses without affecting plant physiology. As a proof of concept, we show in Arabidopsis how such trans-species synthetic gene design allows resistance pyramiding and broad spectrum engineering of virus resistance in plants at no yield loss.

Published
2018

48. Trans-species synthetic gene design allows resistance pyramiding and broad-spectrum engineering of virus resistance

Author

Bastet, Anna, Lederer, Baptiste, Giovinazzo, Nathalie, Arnoux, Xavier, German-Retana, Sylvie, Reinbold, Catherine, Brault, Veronique, Garcia, Damien, Djennane, Samia, Gersch, Sophie, Lemaire, Olivier, Robaglia, Christophe, Gallois, Jean-Luc, Génétique et Amélioration des Fruits et Légumes (GAFL), Institut National de la Recherche Agronomique (INRA), Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Biologie du fruit et pathologie (BFP), Université Bordeaux Segalen - Bordeaux 2-Institut National de la Recherche Agronomique (INRA)-Université Sciences et Technologies - Bordeaux 1, Santé de la vigne et qualité du vin (SVQV), Institut National de la Recherche Agronomique (INRA)-Université de Strasbourg (UNISTRA), Institut de biologie moléculaire des plantes (IBMP), Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA), Société Française de Virologie (SFV). FRA., Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Université Bordeaux Segalen - Bordeaux 2-Institut National de la Recherche Agronomique (INRA)-Université Sciences et Technologies - Bordeaux 1 (UB), and Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS)

Subjects
ComputingMilieux_MISCELLANEOUS, [SDV.BV.PEP]Life Sciences [q-bio]/Vegetal Biology/Phytopathology and phytopharmacy
Abstract

National audience

Published
2018

49. Trans-species synthetic gene design allows resistance pyramiding and broad-spectrum engineering of virus resistance in plants

Author

Bastet, Anna, Lederer, Baptiste, Giovinazzo, Nathalie, Arnoux, Xavier, German-Retana, Sylvie, Reinbold, Catherine, Brault, Veronique, Garcia, Damien, Djennane, Samia, Gersch, Sophie, Lemaire, Olivier, Robaglia, Christophe, Gallois, Jean-Luc, Génétique et Amélioration des Fruits et Légumes (GAFL), Institut National de la Recherche Agronomique (INRA), Biologie du fruit et pathologie (BFP), Université Bordeaux Segalen - Bordeaux 2-Institut National de la Recherche Agronomique (INRA)-Université Sciences et Technologies - Bordeaux 1 (UB), Santé de la vigne et qualité du vin (SVQV), Institut National de la Recherche Agronomique (INRA)-Université de Strasbourg (UNISTRA), Institut de biologie moléculaire des plantes (IBMP), Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS), Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Luminy Génétique et Biophysique des Plantes (LGBP), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), ANR-KBBE COBRA ANR-13-KBBE-0006, ANR-POTYMOVE ANR-16-CE20-000803, ANR-16-CE20-0008,PotyMove,Facteurs cellulaires recrutés par les potyvirus pour leur transport intercellulaire : de nouvelles sources de résistance des plantes?(2016), Unité de recherche Génétique et amélioration des fruits et légumes (GALF), Université Bordeaux Segalen - Bordeaux 2-Institut National de la Recherche Agronomique (INRA)-Université Sciences et Technologies - Bordeaux 1, Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Université de Strasbourg (UNISTRA)-Institut National de la Recherche Agronomique (INRA), Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), and Université Sciences et Technologies - Bordeaux 1-Université Bordeaux Segalen - Bordeaux 2-Institut National de la Recherche Agronomique (INRA)

Subjects
resistance, [SDV.BV.AP]Life Sciences [q-bio]/Vegetal Biology/Plant breeding, Arabidopsis thaliana, synthetic allele, translational research, potyvirus, eIF4E, fungi, food and beverages, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, Research Articles, Research Article
Abstract

UMR BFP - Equipe Virologie; International audience; To infect plants, viruses rely heavily on their host's machinery. Plant genetic resistances based on host factor modifications can be found among existing natural variability and are widely used for some but not all crops. While biotechnology can supply for the lack of natural resistance alleles, new strategies need to be developed to increase resistance spectra and durability without impairing plant development. Here, we assess how the targeted allele modification of the Arabidopsis thaliana translation initiation factor eIF4E1 can lead to broad and efficient resistance to the major group of potyviruses. A synthetic Arabidopsis thaliana eIF4E1 allele was designed by introducing multiple amino acid changes associated with resistance to potyvirus in naturally occurring Pisum sativum alleles. This new allele encodes a functional protein while maintaining plant resistance to a potyvirus isolate that usually hijacks eIF4E1. Due to its biological functionality, this synthetic allele allows, at no developmental cost, the pyramiding of resistances to potyviruses that selectively use the two major translation initiation factors, eIF4E1 or its isoform eIFiso4E. Moreover, this combination extends the resistance spectrum to potyvirus isolates for which no efficient resistance has so far been found, including resistance‐breaking isolates and an unrelated virus belonging to the Luteoviridae family. This study is a proof‐of‐concept for the efficiency of gene engineering combined with knowledge of natural variation to generate trans‐species virus resistance at no developmental cost to the plant. This has implications for breeding of crops with broad‐spectrum and high durability resistance using recent genome editing techniques.

Published
2018

50. A complex eIF4E locus impacts the durability of va resistance to Potato virus Y in tobacco

Author

Michel, Vincent, primary, Julio, Emilie, additional, Candresse, Thierry, additional, Cotucheau, Julien, additional, Decorps, Christophe, additional, Volpatti, Roxane, additional, Moury, Benoît, additional, Glais, Laurent, additional, Jacquot, Emmanuel, additional, de Borne, François Dorlhac, additional, Decroocq, Véronique, additional, Gallois, Jean‐Luc, additional, and German-Retana, Sylvie, additional

Published
2019
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