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1. Bread wheat TaSPO11‐1 exhibits evolutionarily conserved function in meiotic recombination across distant plant species

Author

Alain Nicolas, Giacomo Bastianelli, Emmanuel Guiderdoni, Robin Michard, Ian Fayos, Olivier Da Ines, Charles I. White, Pierre Sourdille, Génétique, Reproduction et Développement - Clermont Auvergne (GReD), Centre National de la Recherche Scientifique (CNRS)-Université Clermont Auvergne (UCA)-Institut National de la Santé et de la Recherche Médicale (INSERM), Génétique, Reproduction et Développement (GReD), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Clermont Auvergne [2017-2020] (UCA [2017-2020])-Centre National de la Recherche Scientifique (CNRS), Génétique Diversité et Ecophysiologie des Céréales (GDEC), Université Clermont Auvergne [2017-2020] (UCA [2017-2020])-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), SAS Meiogenix France, Amélioration génétique et adaptation des plantes méditerranéennes et tropicales (UMR AGAP), Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-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), 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), French government IDEX-ISITE initiative 16-IDEX-0001 (CAP20-25), French National Research Agency (ANR) 2014/1020, CNRS, INSERM, INRAE, Université Clermont Auvergne, Centre National de la Recherche Scientifique (CNRS)-Université Clermont Auvergne [2017-2020] (UCA [2017-2020])-Institut National de la Santé et de la Recherche Médicale (INSERM), Meiogenix, Dynamique de l'information génétique : bases fondamentales et cancer (DIG CANCER), Centre National de la Recherche Scientifique (CNRS)-Institut Curie [Paris]-Sorbonne Université (SU), Centre National de la Recherche Scientifique (CNRS), Institut National de la Santé et de la Recherche Médicale (Inserm), ANRT CIFRE grant no. 2014/1020, ANR-16-IDEX-0001,CAP 20-25,CAP 20-25(2016), Sourdille, Pierre, CAP 20-25 - - CAP 20-252016 - ANR-16-IDEX-0001 - IDEX - VALID, Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Institut Agro - 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), and Institut Curie [Paris]-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS)

Subjects
0106 biological sciences, 0301 basic medicine, [SDV]Life Sciences [q-bio], Plant Science, [SDV.GEN] Life Sciences [q-bio]/Genetics, 01 natural sciences, F30 - Génétique et amélioration des plantes, Arabidopsis, [SDV.GEN.GPL] Life Sciences [q-bio]/Genetics/Plants genetics, wheat, Conserved Sequence, Triticum, ComputingMilieux_MISCELLANEOUS, Plant Proteins, 2. Zero hunger, Genetics, meiotic recombination, biology, phytogénétique, food and beverages, Protéine, Complementation, [SDV] Life Sciences [q-bio], Meiosis, Ploidy, Méiose, Spo11, Gene Transfer, Horizontal, Aegilops, SPO11-1, Triticum aestivum, [SDV.CAN]Life Sciences [q-bio]/Cancer, Genes, Plant, Evolution, Molecular, [SDV.GEN.GPL]Life Sciences [q-bio]/Genetics/Plants genetics, 03 medical and health sciences, Aegilops tauschii, protein evolution, Gene, [SDV.GEN]Life Sciences [q-bio]/Genetics, Génome, Arabidopsis Proteins, fungi, Oryza, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, Cell Biology, biology.organism_classification, [SDV.BV.AP]Life Sciences [q-bio]/Vegetal Biology/Plant breeding, 030104 developmental biology, Triticum urartu, grasses, biology.protein, Homologous recombination, Sequence Alignment, 010606 plant biology & botany
Abstract

International audience; The manipulation of meiotic recombination in crops is essential to develop new plant varieties rapidly, helping to produce more cultivars in a sustainable manner. One option is to control the formation and repair of the meiosis-specific DNA double-strand breaks (DSBs) that initiate recombination between the homologous chromosomes and ultimately lead to crossovers. These DSBs are introduced by the evolutionarily conserved topoisomerase-like protein SPO11 and associated proteins. Here, we characterized the homoeologous copies of the SPO11-1 protein in hexaploid bread wheat (Triticum aestivum). The genome contains three SPO11-1 gene copies that exhibit 93-95% identity at the nucleotide level, and clearly the A and D copies originated from the diploid ancestors Triticum urartu and Aegilops tauschii, respectively. Furthermore, phylogenetic analysis of 105 plant genomes revealed a clear partitioning between monocots and dicots, with the seven main motifs being almost fully conserved, even between clades. The functional similarity of the proteins among monocots was confirmed through complementation analysis of the Oryza sativa (rice) spo11-1 mutant by the wheat TaSPO11-1-5D coding sequence. Also, remarkably, although the wheat and Arabidopsis SPO11-1 proteins share only 55% identity and the partner proteins also differ, the TaSPO11-1-5D cDNA significantly restored the fertility of the Arabidopsis spo11-1 mutant, indicating a robust functional conservation of the SPO11-1 protein activity across distant plants. These successful heterologous complementation assays, using both Arabidopsis and rice hosts, are good surrogates to validate the functionality of candidate genes and cDNA, as well as variant constructs, when the transformation and mutant production in wheat is much longer and more tedious.

Published
2020
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2. Engineering meiotic recombination pathways in rice

Author

Fayos, Ian, Mieulet, Delphine, Petit, Julie, Meunier, Anne Cecile, Perin, Christophe, Nicolas, Alain, Guiderdoni, Emmanuel, Amélioration génétique et adaptation des plantes méditerranéennes et tropicales (UMR AGAP), Institut National de la Recherche Agronomique (INRA)-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)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad), Département Systèmes Biologiques (Cirad-BIOS), Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad), Université Paris sciences et lettres (PSL), Meiogenix, Meiogenix Company, French National Research Agency (ANR), CGIAR research program RICE (CRP RICE), and Agropolis Foundation (RicE FUnctional GEnomics (REFUGE) platform project)

Subjects
Crossing over, Arabidopsis, Reviews, Oryza sativa, Review, Genes, Plant, F30 - Génétique et amélioration des plantes, crossovers, meiosis, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, Homologous Recombination, Vegetal Biology, rice, fungi, food and beverages, Oryza, recombination, Apomixie, Gène, apomixis, Méiose, Genetic Engineering, Biologie végétale
Abstract

International audience; In the last 15 years, outstanding progress has been made in understanding the function of meiotic genes in the model dicot and monocot plants Arabidopsis and rice (Oryza sativa L.), respectively. This knowledge allowed to modulate meiotic recombination in Arabidopsis and, more recently, in rice. For instance, the overall frequency of crossovers (COs) has been stimulated 2.3- and 3.2-fold through the inactivation of the rice FANCM and RECQ4 DNA helicases, respectively, two genes involved in the repair of DNA double-strand breaks (DSBs) as noncrossovers (NCOs) of the Class II crossover pathway. Differently, the programmed induction of DSBs and COs at desired sites is currently explored by guiding the SPO11-1 topoisomerase-like transesterase, initiating meiotic recombination in all eukaryotes, to specific target regions of the rice genome. Furthermore, the inactivation of 3 meiosis-specific genes, namely PAIR1, OsREC8 and OsOSD1, in the Mitosis instead of Meiosis (MiMe) mutant turned rice meiosis into mitosis, thereby abolishing recombination and achieving the first component of apomixis, apomeiosis. The successful translation of Arabidopsis results into a crop further allowed the implementation of two breakthrough strategies that triggered parthenogenesis from the MiMe unreduced clonal egg cell and completed the second component of diplosporous apomixis. Here, we review the most recent advances in and future prospects of the manipulation of meiotic recombination in rice and potentially other major crops, all essential for global food security.

Published
2019
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3. Aborting meiosis allows recombination in sterile diploid yeast hybrids

Author

Jean Luc Legras, Angela Lutazi, Melania D’Angiolo, Karl Persson, Sylvie Dequin, Benoit Albaud, Matteo De Chiara, Souhir Marsit, Agnès Llored, Lorenzo Tattini, Benjamin Barré, Simone Mozzachiodi, Jonas Warringer, Jia-Xing Yue, Alain Nicolas, Sophie Loeillet, Agurtzane Irizar, Neža Škofljanc, Raphaelle Laureau, Simon Stenberg, Gianni Liti, Institut de Recherche sur le Cancer et le Vieillissement (IRCAN), Université Nice Sophia Antipolis (... - 2019) (UNS), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Côte d'Azur (UCA), Centre de recherche de l'Institut Curie [Paris], Institut Curie [Paris], Sciences Pour l'Oenologie (SPO), Université Montpellier 1 (UM1)-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), Department of Chemistry and Molecular Biology, University of Gothenburg, Gothenburg, Sweden, Department of Chemistry and Molecular Biology, University of Gothenburg, Department of Chemistry and Molecular Biology, University Gothenburg, Department of chemistry, University of Gothenburg, Sweden, Fondation pour la Recherche MedicaleFRM EQU202003010413, CEFIPRA, Canceropole PACA (AAP Equipment 2018), ANR-11-LABX-0028,SIGNALIFE,Réseau d'Innovation sur les Voies de Signalisation en Sciences de la Vie(2011), ANR-13-BSV6-0006,AcrossTrait,Génétique des traits quantitatifs chez les levures: au delà des croisements habituels(2013), ANR-15-IDEX-0001,UCA JEDI,Idex UCA JEDI(2015), ANR-16-CE12-0019,PhenoVar,Comprendre les intéractions fonctionnelles entre Variations Structurelles des chromosomes et la diversité Phénotypes en utilisant le modèle levure(2016), ANR-18-CE12-0004,GoNG,Innovation évolutive par acquisition de nouveaux gènes(2018), Meiogenix, Sun Yat-Sen University [Guangzhou] (SYSU), Dynamique de l'information génétique : bases fondamentales et cancer (DIG CANCER), Centre National de la Recherche Scientifique (CNRS)-Institut Curie [Paris]-Sorbonne Université (SU), Columbia University Medical Center (CUMC), Columbia University [New York], Université du Québec à Rimouski (UQAR), Université de Montpellier (UM)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Department of Chemistry and Molecular Biology [Gothenburg], University of Gothenburg (GU), Plateforme de génomique [Institut Curie], Université Nice Sophia Antipolis (1965 - 2019) (UNS), Institut Curie [Paris]-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Université de Montpellier (UM)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Institut Agro - 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), Gestionnaire, Hal Sorbonne Université, Centres d'excellences - Réseau d'Innovation sur les Voies de Signalisation en Sciences de la Vie - - SIGNALIFE2011 - ANR-11-LABX-0028 - LABX - VALID, Blanc 2013 - Génétique des traits quantitatifs chez les levures: au delà des croisements habituels - - AcrossTrait2013 - ANR-13-BSV6-0006 - Blanc 2013 - VALID, Idex UCA JEDI - - UCA JEDI2015 - ANR-15-IDEX-0001 - IDEX - VALID, Comprendre les intéractions fonctionnelles entre Variations Structurelles des chromosomes et la diversité Phénotypes en utilisant le modèle levure - - PhenoVar2016 - ANR-16-CE12-0019 - AAPG2016 - VALID, and APPEL À PROJETS GÉNÉRIQUE 2018 - Innovation évolutive par acquisition de nouveaux gènes - - GoNG2018 - ANR-18-CE12-0004 - AAPG2018 - VALID

Subjects
Saccharomyces cerevisiae Proteins, Science, [SDV]Life Sciences [q-bio], Saccharomyces cerevisiae, General Physics and Astronomy, Evolutionary biology, Biology, Genome, Article, Evolutionary genetics, General Biochemistry, Genetics and Molecular Biology, Evolution, Molecular, 03 medical and health sciences, 0302 clinical medicine, Fungal genetics, Meiosis, Gene mapping, Genetic linkage, [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Homologous Recombination, 030304 developmental biology, Genetics, 0303 health sciences, Multidisciplinary, Chromosome Mapping, General Chemistry, biology.organism_classification, Diploidy, Genome evolution, Phenotype, Infertility, Hybridization, Genetic, Genome, Fungal, Ploidy, Homologous recombination, 030217 neurology & neurosurgery, Recombination
Abstract

Hybrids between diverged lineages contain novel genetic combinations but an impaired meiosis often makes them evolutionary dead ends. Here, we explore to what extent an aborted meiosis followed by a return-to-growth (RTG) promotes recombination across a panel of 20 Saccharomyces cerevisiae and S. paradoxus diploid hybrids with different genomic structures and levels of sterility. Genome analyses of 275 clones reveal that RTG promotes recombination and generates extensive regions of loss-of-heterozygosity in sterile hybrids with either a defective meiosis or a heavily rearranged karyotype, whereas RTG recombination is reduced by high sequence divergence between parental subgenomes. The RTG recombination preferentially arises in regions with low local heterozygosity and near meiotic recombination hotspots. The loss-of-heterozygosity has a profound impact on sexual and asexual fitness, and enables genetic mapping of phenotypic differences in sterile lineages where linkage analysis would fail. We propose that RTG gives sterile yeast hybrids access to a natural route for genome recombination and adaptation., Hybrids are often considered evolutionary dead ends because they do not generate viable offspring. Here, the authors show that sterile yeast hybrids generate genetic diversity through meiotic-like recombination by aborting meiosis and return to asexual growth.

Published
2021
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4. Massive crossover elevation via combination of HEI10 and recq4a recq4b during Arabidopsis meiosis

Author

Heïdi Serra, Catherine H Griffin, Raphael Mercier, Ian R. Henderson, Divyashree C Nageswaran, Mathilde Seguela-Arnaud, Christophe Lambing, Piotr Ziółkowski, Charles J Underwood, Stephanie D. Topp, Joiselle Blanche Fernandes, Department of Plant Sciences, University of Cambridge [UK] (CAM), Institut Jean-Pierre Bourgin (IJPB), Institut National de la Recherche Agronomique (INRA)-AgroParisTech, Université Paris Saclay (COmUE), Université Paris-Sud - Paris 11 (UP11), Biotechnology and Biological Sciences Research Council (BBSRC) [BB/L006847/1], BBSRC-Meiogenix IPA [BB/N007557/1], European Research Area Network for Coodinating Action in Plant Sciences/BBSRC 'DeCOP' [BB/M004937/1], European Research Council (SynthHotspot CoG), Gatsby Charitable Foundation [GAT2962], Royal Society University Research Fellowship, Bettencourt Schueller Foundation, and European Project: 606956,EC:FP7:PEOPLE,FP7-PEOPLE-2013-ITN,COMREC(2013)

Subjects
0301 basic medicine, Chromosomal Proteins, Non-Histone, Heterochromatin, Arabidopsis, Genetic recombination, 03 medical and health sciences, Meiosis, Centromere, Homologous chromosome, meiosis, RECQ4, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, Crossing Over, Genetic, Homologous Recombination, 2. Zero hunger, Genetics, crossover, Multidisciplinary, biology, Arabidopsis Proteins, fungi, DNA Helicases, HEI10, Helicase, Biological Sciences, DNA Methylation, Subtelomere, recombination, 030104 developmental biology, biology.protein, Homologous recombination
Abstract

International audience; During meiosis, homologous chromosomes undergo reciprocal crossovers, which generate genetic diversity and underpin classical crop improvement. Meiotic recombination initiates from DNA double-strand breaks (DSBs), which are processed into single-stranded DNA that can invade a homologous chromosome. The resulting joint molecules can ultimately be resolved as crossovers. In Arabidopsis, competing pathways balance the repair of ∼100-200 meiotic DSBs into ∼10 crossovers per meiosis, with the excess DSBs repaired as noncrossovers. To bias DSB repair toward crossovers, we simultaneously increased dosage of the procrossover E3 ligase gene HEI10 and introduced mutations in the anticrossovers helicase genes RECQ4A and RECQ4B. As HEI10 and recq4a recq4b increase interfering and noninterfering crossover pathways, respectively, they combine additively to yield a massive meiotic recombination increase. Interestingly, we also show that increased HEI10 dosage increases crossover coincidence, which indicates an effect on interference. We also show that patterns of interhomolog polymorphism and heterochromatin drive recombination increases distally towards the sub-telomeres in both HEI10 and recq4a recq4b backgrounds, while the centromeres remain crossover suppressed. These results provide a genetic framework for engineering meiotic recombination landscapes in plant genomes.

Published
2018
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5. Cold-conserved hybrid immature embryos for efficient wheat transformation

Author

Caroline Tassy, Giacomo Bastianelli, Pierre Sourdille, Robin Michard, Pierre Barret, Marie-Claire Debote, Alain Loussert, Alain Nicolas, Manon Batista, Génétique Diversité et Ecophysiologie des Céréales (GDEC), Institut National de la Recherche Agronomique (INRA)-Université Blaise Pascal - Clermont-Ferrand 2 (UBP), Meiogenix, Dynamique de l'information génétique : bases fondamentales et cancer (DIG CANCER), Institut Curie-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), PSL Research University, Centre National de la Recherche Scientifique (CNRS), Institut Curie, ANRT CIFRE [2014/1020], Institut National de la Recherche Agronomique (INRA)-Université Clermont Auvergne [2017-2020] (UCA [2017-2020]), Laboratoire de microbiologie et génétique moléculaires (LMGM), Centre de Biologie Intégrative (CBI), Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS)-Centre National de la Recherche Scientifique (CNRS), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Institut Curie [Paris]-Sorbonne Université (SU), Université Paris sciences et lettres (PSL), Université Blaise Pascal - Clermont-Ferrand 2 (UBP)-Institut National de la Recherche Agronomique (INRA), Laboratoire de microbiologie et génétique moléculaires - UMR5100 (LMGM), Université de Toulouse (UT)-Université de Toulouse (UT)-Centre National de la Recherche Scientifique (CNRS)-Université Toulouse III - Paul Sabatier (UT3), Université de Toulouse (UT)-Université de Toulouse (UT)-Centre National de la Recherche Scientifique (CNRS), and Institut Curie [Paris]-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS)

Subjects
0106 biological sciences, ble tendre, [SDV.BIO]Life Sciences [q-bio]/Biotechnology, Cold treatment, biolistic, Horticulture, Biology, 01 natural sciences, cold treatment, wheat, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, Hybrid, 2. Zero hunger, hybrids, Chinese spring, transformation, Plant physiology, food and beverages, Embryo, traitement au froid, Transgenesis, Transformation (genetics), microprojectile bombardment, soft wheat, Callus, embryonic structures, biolistique, 010606 plant biology & botany, transgénèse végétale
Abstract

Transgenesis through biolistic of immature embryos is the most convenient way to introduce artificially new genes in bread wheat (Triticum aestivum L.). However, only a few genotypes can be efficiently transformed. To improve the transformation of wheat varieties, we stored immature seeds at room temperature or 4 degrees C during 4 or 7 days and extracted immature embryos prior to transformation. Shelling stops the embryo's growth and almost all the embryos formed a callus on selective media when stored at 4 degrees C for 4 or 7 days (respectively 87% and 99%). We also used hybrid immature embryos derived from a cross between a transformable line (Courtot) and a non-transformable line (Chinese Spring) for biolistic transformation. Hybrid embryos showed the same response to biolistic than the responsive parent. All together, these results improve significantly the biolistic protocol for wheat transformation by reducing the number of mother plants in the greenhouse, and improve the transformation of additional genotypes through hybrid transformation.

Published
2019
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6. The Enhancement of Plant Disease Resistance Using CRISPR/Cas9 Technology

Author

Adriano Marocco, Vittoria Brambilla, Peter M. Rogowsky, Alessandra Lanubile, Virginia Maria Grazia Borrelli, Università cattolica del Sacro Cuore [Roma] (Unicatt), Università degli Studi di Milano [Milano] (UNIMI), Reproduction et développement des plantes (RDP), Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National de la Recherche Agronomique (INRA)-École normale supérieure - Lyon (ENS Lyon), Doctoral School on the Agro-Food System (Agrisystem) of Universita Cattolica del Sacro Cuore (Italy), biotechnology company Meiogenix, seed company Limagrain, Università cattolica del Sacro Cuore = Catholic University of the Sacred Heart [Roma] (Unicatt), Università degli Studi di Milano = University of Milan (UNIMI), École normale supérieure de Lyon (ENS de Lyon)-Institut National de la Recherche Agronomique (INRA)-Université Claude Bernard Lyon 1 (UCBL), and Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)

Subjects
0301 basic medicine, résistance aux maladies, disease resistance, [SDV]Life Sciences [q-bio], champignon, Review, Computational biology, Plant Science, virus, Plant disease resistance, Biology, lcsh:Plant culture, Genome, 03 medical and health sciences, CRISPR/Cas9, crop improvement, genome editing, fungus, bacteria, Genome editing, CRISPR, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, lcsh:SB1-1110, 2. Zero hunger, bactérie, Transcription activator-like effector nuclease, Bacterial disease, Bacteria, Cas9, génome, fungi, food and beverages, Zinc finger nuclease, Settore AGR/07 - GENETICA AGRARIA, 030104 developmental biology, plante, Crop improvement, Disease resistance, Fungus, Virus
Abstract

International audience; Genome editing technologies have progressed rapidly and become one of the most important genetic tools in the implementation of pathogen resistance in plants. Recent years have witnessed the emergence of site directed modification methods using meganucleases, zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and clustered regularly interspaced short palindrome repeats (CRISPR)/CRISPR-associated protein 9 (Cas9). Recently, CRISPR/Cas9 has largely overtaken the other genome editing technologies due to the fact that it is easier to design and implement, has a higher success rate, and is more versatile and less expensive. This review focuses on the recent advances in plant protection using CRISPR/Cas9 technology in model plants and crops in response to viral, fungal and bacterial diseases. As regards the achievement of viral disease resistance, the main strategies employed in model species such as Arabidopsis and Nicotiana benthamiana, which include the integration of CRISPR-encoding sequences that target and interfere with the viral genome and the induction of a CRISPR-mediated targeted mutation in the host plant genome, will be discussed. Furthermore, as regards fungal and bacterial disease resistance, the strategies based on CRISPR/Cas9 targeted modification of susceptibility genes in crop species such as rice, tomato, wheat, and citrus will be reviewed. After spending years deciphering and reading genomes, researchers are now editing and rewriting them to develop crop plants resistant to specific pests and pathogens.

Published
2018
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7. Efficient wheat transformation can be performed on cold-conserved immature hybrid embryos

Author

Michard, Robin, Debote, Marie-Claire, Loussert, Alain, Tassy, Caroline, Bastianelli, Giacomo, Nicolas, Alain, Barret, Pierre, Sourdille, Pierre, Génétique Diversité et Ecophysiologie des Céréales (GDEC), Université Blaise Pascal - Clermont-Ferrand 2 (UBP)-Institut National de la Recherche Agronomique (INRA), SAS Meiogenix France, UMR 3244, Univ Paris 06 (PSL), Centre National de la Recherche Scientifique (CNRS), Institut Curie, Institut National de la Recherche Agronomique (INRA)-Université Clermont Auvergne [2017-2020] (UCA [2017-2020]), Dynamique de l'information génétique : bases fondamentales et cancer (DIG CANCER), Institut Curie [Paris]-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Institut National de la Recherche Agronomique (INRA)-Université Blaise Pascal - Clermont-Ferrand 2 (UBP), and Centre National de la Recherche Scientifique (CNRS)-Institut Curie [Paris]-Sorbonne Université (SU)

Subjects
protéine de stockage, embryon, [SDV]Life Sciences [q-bio], education, fungi, food and beverages, embryo, qualité du blé
Abstract

Poster P0218Poster P0218; Efficient wheat transformation can be performed on cold-conserved immature hybrid embryos. Plant and Animals Genomics Conference XXV

Published
2018

8. Interhomolog polymorphism shapes meiotic crossover within the Arabidopsis RAC1 and RPP13 disease resistance genes

Author

Serra, Heïdi, Choi, Kyuha, Zhao, Xiaohui, Blackwell, Alexander R., Kim, Juhyun, Henderson, Ian R., Génétique Diversité et Ecophysiologie des Céréales (GDEC), Institut National de la Recherche Agronomique (INRA)-Université Clermont Auvergne [2017-2020] (UCA [2017-2020]), University of Cambridge [UK] (CAM), Pohang University of Science and Technology (POSTECH), Pohang Univ Sci & Technol, Dept Life Sci, Pohang, Gyeongbuk, South Korea, Partenaires INRAE, Royal Society University Research Fellowship, Gatsby Charitable Foundation GAT2962, BBSRC BB/L006847/1, BBSRC-Meiogenix IPA grant BB/N007557/1, ERC 'SynthHotSpot' Consolidator Grant, National Natural Science Foundation of China 61403318, EMBO long-term postdoctoral fellowship ALT 807-2009, RDA Next-Generation BioGreen 21 Program PJ01337001, NRF Basic Science Research Program NRF-2017R1D1AB03028374, Bettencourt Schueller Foundation, Gatsby Foundation Sainsbury studentship GAT3401, Choi, Kyuha [0000-0002-4072-3807], Blackwell, Alexander R [0000-0002-0369-4584], Kim, Juhyun [0000-0001-5462-3391], Henderson, Ian R [0000-0001-5066-1489], and Apollo - University of Cambridge Repository

Subjects
[SDV]Life Sciences [q-bio], Artificial Gene Amplification and Extension, Plant Science, polymorphisme, QH426-470, Biochemistry, Polymerase Chain Reaction, Database and Informatics Methods, DNA Breaks, Double-Stranded, Cell Cycle and Cell Division, Crossing Over, Genetic, Homologous Recombination, Disease Resistance, Chromosome Biology, Plant Anatomy, Eukaryota, High-Throughput Nucleotide Sequencing, Plants, Plants, Genetically Modified, Chromatin, Nucleic acids, Meiosis, Experimental Organism Systems, Cell Processes, [SDE]Environmental Sciences, Pollen, génétique de la résistance, Sequence Analysis, Research Article, DNA recombination, Bioinformatics, Arabidopsis Thaliana, Brassica, Research and Analysis Methods, Genes, Plant, Chromosomes, Plant, Model Organisms, Plant and Algal Models, Genetics, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, Molecular Biology Techniques, Molecular Biology, Plant Diseases, Evolutionary Biology, Polymorphism, Genetic, Population Biology, Arabidopsis Proteins, Organisms, Biology and Life Sciences, DNA, Cell Biology, arabidopsis, Mutation, Animal Studies, Sequence Alignment, Population Genetics
Abstract

During meiosis, chromosomes undergo DNA double-strand breaks (DSBs), which can be repaired using a homologous chromosome to produce crossovers. Meiotic recombination frequency is variable along chromosomes and tends to concentrate in narrow hotspots. We mapped crossover hotspots located in the Arabidopsis thaliana RAC1 and RPP13 disease resistance genes, using varying haplotypic combinations. We observed a negative non-linear relationship between interhomolog divergence and crossover frequency within the hotspots, consistent with polymorphism locally suppressing crossover repair of DSBs. The fancm, recq4a recq4b, figl1 and msh2 mutants, or lines with increased HEI10 dosage, are known to show increased crossovers throughout the genome. Surprisingly, RAC1 crossovers were either unchanged or decreased in these genetic backgrounds, showing that chromosome location and local chromatin environment are important for regulation of crossover activity. We employed deep sequencing of crossovers to examine recombination topology within RAC1, in wild type, fancm, recq4a recq4b and fancm recq4a recq4b backgrounds. The RAC1 recombination landscape was broadly conserved in the anti-crossover mutants and showed a negative relationship with interhomolog divergence. However, crossovers at the RAC1 5′-end were relatively suppressed in recq4a recq4b backgrounds, further indicating that local context may influence recombination outcomes. Our results demonstrate the importance of interhomolog divergence in shaping recombination within plant disease resistance genes and crossover hotspots., Author summary Sexually reproducing plants and animals produce gametes with half the number of chromosomes, which can participate in fertilization. A specialized cell division called meiosis generates gametes, where the chromosomes are copied once and segregated twice. A further key feature of meiosis is that chromosomes physically pair and undergo reciprocal exchanges, called crossovers. Due to independent chromosome segregation and crossovers, meiosis creates gametes that are genetically diverse, which has a major effect on patterns of sequence variation in populations. Interestingly, the frequency of crossover is also highly variable along the lengths of chromosomes and tends to be concentrated in narrow hotspots. Here we studied two crossover hotspots in detail that are located within disease resistance genes, using the model plant Arabidopsis. We show that within these hotspots, greater levels of genetic difference between the recombining chromosomes locally inhibits crossover formation. We also show that hotspots within one of these resistance genes are surprisingly resistant to genetic backgrounds that increase crossovers elsewhere in the genome. This indicates that patterns of polymorphism and hotspot location along the chromosome are both important for control of recombination activity.

Published
2018
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9. Identification of determining factors for meiotic recombination targeting in bread wheat

Author

Michard, Robin, Debote, Marie-Claire, Tassy, Caroline, Bastianelli, Giacomo, Nicolas, Alain, Barret, Pierre, Sourdille, Pierre, Génétique Diversité et Ecophysiologie des Céréales (GDEC), Institut National de la Recherche Agronomique (INRA)-Université Blaise Pascal - Clermont-Ferrand 2 (UBP), SAS Meiogenix France, Université Pierre et Marie Curie (Paris 6), Centre National de la Recherche Scientifique (CNRS), Institut Curie, Institut National de la Recherche Agronomique (INRA)-Université Clermont Auvergne [2017-2020] (UCA [2017-2020]), and Université Pierre et Marie Curie - Paris 6 (UPMC)

Subjects
ble tendre, recombinaison méiotique, soft wheat, urogenital system, [SDV]Life Sciences [q-bio], digestive, oral, and skin physiology, food and beverages
Abstract

Identification of determining factors for meiotic recombination targeting in bread wheat. 12. French 3R Meeting

Published
2017

10. Ciblage de la recombinaison méiotique chez le blé tendre : Comment ça marche ?

Author

Michard, Robin, Bastianelli, Giacomo, Nicolas, Alain, Sourdille, Pierre, Génétique Diversité et Ecophysiologie des Céréales (GDEC), Institut National de la Recherche Agronomique (INRA)-Université Clermont Auvergne [2017-2020] (UCA [2017-2020]), SAS Meiogenix France, Université Pierre et Marie Curie - Paris 6 (UPMC), Centre National de la Recherche Scientifique (CNRS), Institut Curie, Institut National de la Recherche Agronomique (INRA)-Université Blaise Pascal - Clermont-Ferrand 2 (UBP), Université Pierre et Marie Curie (Paris 6), and Université Blaise Pascal - Clermont-Ferrand 2 (UBP)-Institut National de la Recherche Agronomique (INRA)

Subjects
ble tendre, recombinaison méiotique, soft wheat, [SDV]Life Sciences [q-bio]
Abstract

Ciblage de la recombinaison méiotique chez le blé tendre : Comment ça marche ?. Journée de l'Ecole Doctorale SVSAE, les 20 ans

Published
2017

11. UGDR: a generic pipeline to detect recombined regions in polyploid and complex hybrid yeast genomes.

Author

Bedrat A

Subjects
Ploidies, Diploidy, Polymorphism, Genetic, Yeasts, Genome, Polyploidy
Abstract

Motivation: In eukaryotes, homologous recombination between the parental genomes frequently occurs during the evolutionary conserved process of meiosis, generating the genetic diversity transmitted by the gametes. The genome-wide determination of the frequency and location of the recombination events can now be efficiently performed by genotyping the offspring's polymorphic markers. However, genotyping recombination in complex hybrid genomes with existing methods remains challenging because of their strain and ploidy specificity and the degree of diversity and complexity of the parental genomes, especially in [Formula: see text] polyploids., Results: We present UGDR, a pipeline to genotype the polymorphisms of complex hybrid yeast genomes. It is based on optimal mapping strategies of NGS reads, comparative analyses of the allelic ratio variation and read depth coverage. We tested the UGDR pipeline with sequencing reads from recombined hybrid diploid yeast strains and various clinical strains exhibiting different degrees of ploidy. UGDR allows to plot the markers distribution and recombination profile per chromosome., Conclusion: UGDR detects and plots recombination events in haploids and polyploid yeasts, which facilitates the discovery and understanding of the yeast genetic recombination map and identify new out-performing recombinants., (© 2022. The Author(s).)

Published
2022
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12. Unlocking the functional potential of polyploid yeasts.

Author

Mozzachiodi S, Krogerus K, Gibson B, Nicolas A, and Liti G

Subjects
Meiosis, Polyploidy, Saccharomyces cerevisiae genetics, Plant Breeding, Saccharomyces cerevisiae Proteins genetics
Abstract

Breeding and domestication have generated widely exploited crops, animals and microbes. However, many Saccharomyces cerevisiae industrial strains have complex polyploid genomes and are sterile, preventing genetic improvement strategies based on breeding. Here, we present a strain improvement approach based on the budding yeasts' property to promote genetic recombination when meiosis is interrupted and cells return-to-mitotic-growth (RTG). We demonstrate that two unrelated sterile industrial strains with complex triploid and tetraploid genomes are RTG-competent and develop a visual screening for easy and high-throughput identification of recombined RTG clones based on colony phenotypes. Sequencing of the evolved clones reveal unprecedented levels of RTG-induced genome-wide recombination. We generate and extensively phenotype a RTG library and identify clones with superior biotechnological traits. Thus, we propose the RTG-framework as a fully non-GMO workflow to rapidly improve industrial yeasts that can be easily brought to the market., (© 2022. The Author(s).)

Published
2022
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13. Manipulation of Meiotic Recombination to Hasten Crop Improvement.

Author

Fayos I, Frouin J, Meynard D, Vernet A, Herbert L, and Guiderdoni E

Abstract

Reciprocal (cross-overs = COs) and non-reciprocal (gene conversion) DNA exchanges between the parental chromosomes (the homologs) during meiotic recombination are, together with mutation, the drivers for the evolution and adaptation of species. In plant breeding, recombination combines alleles from genetically diverse accessions to generate new haplotypes on which selection can act. In recent years, a spectacular progress has been accomplished in the understanding of the mechanisms underlying meiotic recombination in both model and crop plants as well as in the modulation of meiotic recombination using different strategies. The latter includes the stimulation and redistribution of COs by either modifying environmental conditions (e.g., T°), harnessing particular genomic situations (e.g., triploidy in Brassicaceae), or inactivating/over-expressing meiotic genes, notably some involved in the DNA double-strand break (DSB) repair pathways. These tools could be particularly useful for shuffling diversity in pre-breeding generations. Furthermore, thanks to the site-specific properties of genome editing technologies the targeting of meiotic recombination at specific chromosomal regions nowadays appears an attainable goal. Directing COs at desired chromosomal positions would allow breaking linkage situations existing between favorable and unfavorable alleles, the so-called linkage drag, and accelerate genetic gain. This review surveys the recent achievements in the manipulation of meiotic recombination in plants that could be integrated into breeding schemes to meet the challenges of deploying crops that are more resilient to climate instability, resistant to pathogens and pests, and sparing in their input requirements.

Published
2022
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14. Recombination in a sterile polyploid hybrid yeast upon meiotic Return-To-Growth.

Author

Serero A, Bedrat A, Baulande S, Bastianelli G, Colavizza D, Desfougères T, Pignède G, Quipourt-Isnard AD, and Nicolas A

Subjects
Genome, Fungal, Phenotype, Polyploidy, Homologous Recombination, Meiosis genetics, Saccharomyces cerevisiae genetics, Saccharomyces cerevisiae growth & development
Abstract

The sustainable future of food industry and consumer demands meet the need to generate out-performing new yeast variants. This is addressed by using the natural yeast diversity and breeding via sexual reproduction but the recovery of recombined spores in many industrial strains is limited. To circumvent this drawback, we examined whether or not the process of meiotic Return to Growth (RTG) that allows S. cerevisiae diploid cells to initiate meiotic recombination genome-wide and then re-enter into mitosis, will be effective to generate recombinants in a sterile and polyploid baking yeast strain (CNCM). We proceeded in four steps. First, whole genome sequencing of the CNCM strain revealed that it was an unbalanced polymorphic triploid. Second, we annotated a panel of genes likely involved in the success of the RTG process. Third, we examined the strain progression into sporulation and fourth, we developed an elutriation and reiterative RTG protocol that allowed to generate extensive libraries of recombinant RTGs, enriched up to 70 %. Altogether, the genome analysis of 122 RTG cells demonstrated that they were bona fide RTG recombinants since the vast majority retained the parental ploidy and exhibited allelic variations involving 1-60 recombined regions per cell with a length of ∼0.4-400 kb. Thus, beyond diploid laboratory strains, we demonstrated the proficiency of this natural non-GM and marker-free process to recombine a sterile and polyploid hybrid yeast, thus providing an unprecedented resource to screen improved traits., (Copyright © 2021 The Authors. Published by Elsevier GmbH.. All rights reserved.)

Published
2021
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15. Assessment of the roles of SPO11-2 and SPO11-4 in meiosis in rice using CRISPR/Cas9 mutagenesis.

Author

Fayos I, Meunier AC, Vernet A, Navarro-Sanz S, Portefaix M, Lartaud M, Bastianelli G, Périn C, Nicolas A, and Guiderdoni E

Subjects
CRISPR-Cas Systems, Meiosis, Mutagenesis, Arabidopsis genetics, Oryza genetics
Abstract

In Arabidopsis, chromosomal double-strand breaks at meiosis are presumably catalyzed by two distinct SPO11 transesterases, AtSPO11-1 and AtSPO11-2, together with M-TOPVIB. To clarify the roles of the SPO11 paralogs in rice, we used CRISPR/Cas9 mutagenesis to produce null biallelic mutants in OsSPO11-1, OsSPO11-2, and OsSPO11-4. Similar to Osspo11-1, biallelic mutations in the first exon of OsSPO11-2 led to complete panicle sterility. Conversely, all Osspo11-4 biallelic mutants were fertile. To generate segregating Osspo11-2 mutant lines, we developed a strategy based on dual intron targeting. Similar to Osspo11-1, the pollen mother cells of Osspo11-2 progeny plants showed an absence of bivalent formation at metaphase I, aberrant segregation of homologous chromosomes, and formation of non-viable tetrads. In contrast, the chromosome behavior in Osspo11-4 male meiocytes was indistinguishable from that in the wild type. While similar numbers of OsDMC1 foci were revealed by immunostaining in wild-type and Osspo11-4 prophase pollen mother cells (114 and 101, respectively), a surprisingly high number (85) of foci was observed in the sterile Osspo11-2 mutant, indicative of a divergent function between OsSPO11-1 and OsSPO11-2. This study demonstrates that whereas OsSPO11-1 and OsSPO11-2 are the likely orthologs of AtSPO11-1 and AtSPO11-2, OsSPO11-4 has no major role in wild-type rice meiosis., (© The Author(s) 2020. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Published
2020
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16. Bread wheat TaSPO11-1 exhibits evolutionarily conserved function in meiotic recombination across distant plant species.

Author

Da Ines O, Michard R, Fayos I, Bastianelli G, Nicolas A, Guiderdoni E, White C, and Sourdille P

Subjects
Aegilops genetics, Arabidopsis Proteins genetics, Evolution, Molecular, Meiosis genetics, Oryza genetics, Sequence Alignment, Conserved Sequence genetics, Gene Transfer, Horizontal genetics, Genes, Plant genetics, Plant Proteins genetics, Triticum genetics
Abstract

The manipulation of meiotic recombination in crops is essential to develop new plant varieties rapidly, helping to produce more cultivars in a sustainable manner. One option is to control the formation and repair of the meiosis-specific DNA double-strand breaks (DSBs) that initiate recombination between the homologous chromosomes and ultimately lead to crossovers. These DSBs are introduced by the evolutionarily conserved topoisomerase-like protein SPO11 and associated proteins. Here, we characterized the homoeologous copies of the SPO11-1 protein in hexaploid bread wheat (Triticum aestivum). The genome contains three SPO11-1 gene copies that exhibit 93-95% identity at the nucleotide level, and clearly the A and D copies originated from the diploid ancestors Triticum urartu and Aegilops tauschii, respectively. Furthermore, phylogenetic analysis of 105 plant genomes revealed a clear partitioning between monocots and dicots, with the seven main motifs being almost fully conserved, even between clades. The functional similarity of the proteins among monocots was confirmed through complementation analysis of the Oryza sativa (rice) spo11-1 mutant by the wheat TaSPO11-1-5D coding sequence. Also, remarkably, although the wheat and Arabidopsis SPO11-1 proteins share only 55% identity and the partner proteins also differ, the TaSPO11-1-5D cDNA significantly restored the fertility of the Arabidopsis spo11-1 mutant, indicating a robust functional conservation of the SPO11-1 protein activity across distant plants. These successful heterologous complementation assays, using both Arabidopsis and rice hosts, are good surrogates to validate the functionality of candidate genes and cDNA, as well as variant constructs, when the transformation and mutant production in wheat is much longer and more tedious., (© 2020 Society for Experimental Biology and John Wiley & Sons Ltd.)

Published
2020
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17. Engineering meiotic recombination pathways in rice.

Author

Fayos I, Mieulet D, Petit J, Meunier AC, Périn C, Nicolas A, and Guiderdoni E

Subjects
Arabidopsis, Genes, Plant, Genetic Engineering, Homologous Recombination, Meiosis, Oryza genetics
Abstract

In the last 15 years, outstanding progress has been made in understanding the function of meiotic genes in the model dicot and monocot plants Arabidopsis and rice (Oryza sativa L.), respectively. This knowledge allowed to modulate meiotic recombination in Arabidopsis and, more recently, in rice. For instance, the overall frequency of crossovers (COs) has been stimulated 2.3- and 3.2-fold through the inactivation of the rice FANCM and RECQ4 DNA helicases, respectively, two genes involved in the repair of DNA double-strand breaks (DSBs) as noncrossovers (NCOs) of the Class II crossover pathway. Differently, the programmed induction of DSBs and COs at desired sites is currently explored by guiding the SPO11-1 topoisomerase-like transesterase, initiating meiotic recombination in all eukaryotes, to specific target regions of the rice genome. Furthermore, the inactivation of 3 meiosis-specific genes, namely PAIR1, OsREC8 and OsOSD1, in the Mitosis instead of Meiosis (MiMe) mutant turned rice meiosis into mitosis, thereby abolishing recombination and achieving the first component of apomixis, apomeiosis. The successful translation of Arabidopsis results into a crop further allowed the implementation of two breakthrough strategies that triggered parthenogenesis from the MiMe unreduced clonal egg cell and completed the second component of diplosporous apomixis. Here, we review the most recent advances in and future prospects of the manipulation of meiotic recombination in rice and potentially other major crops, all essential for global food security., (© 2019 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.)

Published
2019
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18. Programming sites of meiotic crossovers using Spo11 fusion proteins.

Author

Sarno R, Vicq Y, Uematsu N, Luka M, Lapierre C, Carroll D, Bastianelli G, Serero A, and Nicolas A

Subjects
CRISPR-Cas Systems, Chromosomes, Fungal genetics, DNA Breaks, Double-Stranded, Gene Fusion, Gene Targeting methods, Recombination, Genetic, Reproducibility of Results, Transcription Activator-Like Effectors genetics, Transcription Activator-Like Effectors metabolism, Crossing Over, Genetic, Endodeoxyribonucleases genetics, Meiosis genetics, Saccharomyces cerevisiae genetics, Saccharomyces cerevisiae Proteins genetics
Abstract

Meiotic recombination shapes the genetic diversity transmitted upon sexual reproduction. However, its non-random distribution along the chromosomes constrains the landscape of potential genetic combinations. For a variety of purposes, it is desirable to expand the natural repertoire by changing the distribution of crossovers in a wide range of eukaryotes. Toward this end, we report the local stimulation of meiotic recombination at a number of chromosomal sites by tethering the natural Spo11 protein to various DNA-binding modules: full-length DNA binding proteins, zinc fingers (ZFs), transcription activator-like effector (TALE) modules, and the CRISPR-Cas9 system. In the yeast Saccharomyces cerevisiae, each strategy is able to stimulate crossover frequencies in naturally recombination-cold regions. The binding and cleavage efficiency of the targeting Spo11 fusions (TSF) are variable, being dependent on the chromosomal regions and potential competition with endogenous factors. TSF-mediated genome interrogation distinguishes naturally recombination-cold regions that are flexible and can be warmed-up (gene promoters and coding sequences), from those that remain refractory (gene terminators and centromeres). These results describe new generic experimental strategies to increase the genetic diversity of gametes, which should prove useful in plant breeding and other applications., (© The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Published
2017
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