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Coordinators: Mathieu Rousseau-Gueutin, Cyril Falentin et Anne-Marie Chèvre

Contacts: mathieu.rousseau-gueutin@inrae.fr; cyril.falentin@inrae.fr; anne-marie.chevre@inrae.fr

Research

Context and Issues

Polyploidy or Whole Genome Duplication (WGD) has and still plays a major role in the evolution, diversification, and phenotypic plasticity of eukaryotes, especially in flowering plants (Angiosperms). One of the best plant systems to study this phenomenon corresponds to the Brassica genus as (i) it has been subjected to both recent and ancient polyploidization events, (ii) it is possible to resynthesize polyploids in the lab, (iii) assembled genomes exist for both diploid and polyploid species. Within the laboratory, we are interested in understanding the immediate impact of allopolyploidy on genome structure (i.e. identification of structural variations using SNP array or DNA Seq data, and validation using molecular cytogenetics: Rousseau-Gueutin et al. 2017; Stein et al. 2017, Ferreira de Carvalho et al. 2020), on the epigenome (BS-Seq: G. Richard post-doc), transcription of duplicated genes or cytonuclear interactions (Ferreira de Carvalho et al. 2019). More precisely, we try to identify regions or molecular mechanisms that have facilitated its genome stabilization (by improving its meiotic stability: Pelé et al; 2018; Ferreira de Carvalho et al. 2020) and that plays a role in fertility (including evolution of the self-incompatibility system: Azibi et al.2020), thereafter facilitating its putative speciation success.

We are also working on the long-term evolutionary fate of genes duplicated by old polyploidization events. Indeed, Brassica species have been subjected to a whole triplication event since their divergence with Arabidopsis (about 20 millions of years ago), so that up to 3 copies of each Arabidopsis gene may be encountered for each diploid Brassica species. However, since these paleoploidization events in diploid species, important structural variations (i.e. reduction of chromosome number or structural variations: Boutte et al. 2020) and losses of most duplicated copies occurred. Additionally, we observed that the different oilseed rape varieties carry different translocations between A and C genomes (Rousseau-Gueutin et al. 2020) as synthetics but of smaller size. We are interested in understanding which and why some genes were retained in multiple copies (synteny and phylogenomic approaches), and if they retained the same function (using Crispr-Cas9 technology: Maud Facon post-doc). 

 Methodology 

• Comparative and evolutionary genomics (DNA-Seq, RNA-Seq and BS-Seq)
• Synteny analyses between fully assembled genomes
• Molecular phylogenetics
• Molecular cytogenetic (BAC-FISH and Oligo-Fish)
• Functional genetics (Crispr-Cas9)

Main results

Brassica species have been subjected to both ancient and recent polyploidization events. Taking advantage of various Brassica assembled genomes (for which we are deeply involved: see also project ‘Brassica Genomics and New Sequencing Technologies’ in close collaboration with Genoscope), we studied the fractionation process in Brassica diploid species and observed that most duplicated genes returned to single copy. Interestingly, we observed that genes involved in chloroplast protein complexes show a higher retention of duplicated copies compared to other protein complexes. These old duplicated copies are functional and under purifying section, indicating that they retained the same function and act synergetically (Ferreira et al. 2019). To verify unambiguously such hypothesis, we currently use the Crispr-Cas9 technology to K-O one to several duplicated copies of a gene and study its impact on the phenotype (ANR PIA Genius project ‘Duplex’: postdoc M. Facon). Similarly, we studied the impact of paleopolyploidy on the self-incompatibility system and observed the diversity of the evolutionary fate of duplicated genes, with cases of pseudogeneization, sub-functionalization and even a case of neo-functionalization (Azibi et al. 2020). In addition to this process of loss/retention of duplicated genes (via fractionation), we also observed that different diploid B. rapa morphotypes (which diverged less than 2 mya) presented important structural variations, resulting from recent insertions (mostly repeat elements) or inversions. Interestingly, these SVs may represent up to 15-20% of its genome and presumably played a role in the currently existing important B. rapa morphotypic diversity (Boutte et al. 2020). We recently provided a new oilseed rape reference genome (Rousseau-Gueutin et al. 2020) with which all regions carrying repeated elements are well assembled even pericentromeric regions. It will be the support of new studies on the dynamics of genome at the allotetraploid level.

To decipher the immediate impact of allopolyploidy on structural and functional genome evolution, we also created resynthesized B. napus population in the laboratory. We notably observed that the mating system type in the first generations following allopolyploidy played a major role in the putative speciation success of a poyploid. Indeed, progenies deriving from outcrossing between resynthesized B. napus rather than by selfing show a much lower level of aneuploidy, despite being subjected to numerous homoeologous exchanges (Rousseau-Gueutin et al. 2017). Using such populations, we also identified genomic regions that may improve genome stability (more bivalent pairing) or fertility of newly formed polyploids (Ferreira de Carvalho et al. 2020).

Parters

• CEA, Genoscope, Evry, France (teams ‘Research and Development Bioinformatics and Sequencing’ and ‘Eukaryotes genomes analyses’)
• INRAE UMR ‘Genetics, Diversity, and Ecophysiology of Cereals’, Clermont-Ferrand, France (J. Salse)
• INRAE UMR GQE (Quantitative Genetics and Evolution), Le Moulon, France (K. Alix)
• University of Lille UMR’ Evolution, Ecology and Paleotonlogy’, Lille, France (X. Vekemans)
• University of Rennes 1, UMR Ecobio (M.L. Aïnouche, A. Salmon)
• University of Science and Technology Houari Boumedienne, Bab Ezzouar, Algeria (H. Hadj-Arab)
• University of York, England (I. Bancroft)
• University of Giessen, Germany (R. Snowdon)
• Czech Academy of Science, Brno, Czech Republic (A. Kovarik) 

Funding and Support (last 5 years)

• Brittany State ‘Epitrans’: Post-doc salary of Gautier Richard (12/2019-06/2021): Facilitate oilseed rape breeding via epigenomic and transcriptomic changes (supervisors: A-M. Chevre and M. Rousseau-Gueutin)
• Marie-Slodowska Curie Actions – H2020 ‘Surfing’: Post-doc salary and bench fees of Julie Ferreira de Carvalho (06/2018-06/2020): StructURal and Functional dynamics of BrassIca napus polyploid Genome.
• Agreenskills plus ‘Dynamic of a polyploid species’: Post-doc salary of Julie Ferreira de Carvalho (05/2017-05/2018): Immediate impact of polyploidy and of human selection on evolutionary dynamic of B. napus.
• ANR PIA ‘Genius’ project Duplex (2017-2020): ’ Tuning a ménage à 4: how to deal with DUPLicated gene Expression (P.I.: M. Rousseau-Gueutin).
• INRAE Plant Breeding and Biology department ‘Allogen’ (2016-2017): Impact of interspecific hybridization and polyploidy on the evolutionary dynamic of B. napus genome. (P.I. M. Rousseau-Gueutin).
• France Génomique ‘Polysucess’ (2014-2020). How a polyploid becomes a new species: Brassica model (P.I.: A.M. Chèvre).
• ERANET project ‘ERACAPS’ (2014-2016): Evolution of genomes: structure-function relationships in the polyploid crop species (P.I.: I. Bancroft).

Publications (last 5 years)

• Azibi, T ; Hadj-Arab H., Lodé M., Ferreira de Caravalho J., Trotoux G., Nègre S., Gilet M.M., Boutte J., Lucas J., Vekemans, X., Chèvre A.M., Rousseau-Gueutin, M. (2020). Impact of whole genome triplication on the evolutionary history and the functional dynamics of regulatory genes involved in Brassica self-incompatibility signalling pathway. Plant Reproduction 33, 43-58. https://doi.org/10.1007/s00497-020-00385-x.
• Belser C., Istace B., Denis E., Dubarry M., Baurens F.-C., Falentin C., Genete M., Berrabah W., Chèvre A. M., Delourme R., Deniot G., Denoeud F., Duffé P., Engelen S., Lemainque A., Manzanares-Dauleux M. J., Martin G., Morice J., Noel B., Vekemans X., D’Hont A., Rousseau-Gueutin M., Barbe V., Cruaud C., Wincker P. & Aury J.-M. (2018). Chromosome-scale assemblies of plant genomes using nanopore long reads and optical maps. Nature Plants, 4(11), 879-887.  https://doi.org/10.1038/s41477-018-0289-4
• Bouchet A. S., Laperche A., Bissuel-Belaygue C., Baron C., Morice J., Rousseau-Gueutin M., Dheu J. E., George P., Pinochet X., Foubert T., Maes O., Dugué D., Guinot F. & Nesi N. (2016). Genetic basis of nitrogen use efficiency and yield stability across environments in winter rapeseed. BMC Genetics, 17(1), 1-21.  https://doi.org/10.1186/s12863-016-0432-z
• Boutte J., Maillet L., Chaussepied T., Letort S., Aury J.M., Belser C., Boideau F., Brunet A., Coriton C., Deniot G., Falentin C., Huteau V., Lodé M., Morice J., Trotoux G., Chèvre A.M., Rousseau-Gueutin M., Ferreria de Carvalho J. (2020)  Large genomic variants reveal unexplored intraspecific diversity in Brassica rapa genomes. Frontiers in Plant Sciences. DOI: 10.3389/fpls.2020.577536
• Ferreira De Carvalho J., Lucas J., Deniot G., Falentin C., Filangi O., Gilet M-M., Legeai F., Lodé M., Morice J., Trotoux G., Aury J-M., Barbe V., Keller J., Snowdon R., He Z., Denoeud F., Wincker P., Bancroft I., Chèvre A-M., Rousseau-Gueutin M. (2019). Cytonuclear interactions remain stable during allopolyploid evolution despite repeated whole-genome duplications in Brassica. Plant Journal, 98 (3), 434-447. DOI: 10.1111/tpj.14228
• Ferreira de Carvalho J., Stoeckel S., Eber F., Lodé-Taburel M., Gilet M-M, Trotoux, G., Morice J., Falentin C., Chèvre A-M., Rousseau-Gueutin M. (2020). Untangling structural factors and evolutionary drivers in nascent polyploids. New Phytologist 230(5):2072-2084.
• Giraud D., Lima O., Huteau V., Coriton O., Boutte J., Kovarik A., Leitch A., Leitch L., Ainouche M. & Salmon A. (2021). Evolutionary dynamics of transposable elements and satellite DNAs in polyploid Spartina species. Plant Science 302. https://doi.org/10.1016/j.plantsci.2020.110671
• Giraud D., Lima O., Rousseau-Gueutin M., Salmon A., Aïnouche, M.L. (2021) Gene and transposable element expression evolution following recent and past polyploidy events in Spartina species. Frontiers in Genetics.
• Martinez Palacios P., Jacquemot M.-P., Tapie M., Rousselet A., Diop M., Remoue C., Falque M., Lloyd A., Jenczewski E., Lassalle G., Chèvre A. M., Lelandais C., Crespi M., Brabant P., Joets J. & Alix K. (2019). Assessing the response of small RNA populations to allopolyploidy using resynthesized Brassica napus allotetraploids. Molecular Biology and Evolution, 36(4), 709-726.  https://doi.org/10.1093/molbev/msz007
• Pelé A., Rousseau-Gueutin M., Chèvre A. M. (2018). Speciation Success of Polyploid Plants Closely Relates to the Regulation of Meiotic Recombination. Frontiers in Plant Science, 9, 907.  https://doi.org/10.3389/fpls.2018.00907
• Pelé A., Trotoux G., Eber F., Lodé M., Gilet M., Deniot G., Falentin C., Nègre S., Morice J., Rousseau-Gueutin M., Chèvre
 A-M. (2016) “The poor lonesome A subgenome of Brassica napus var. Darmor (AACC) may not survive without its mate”. New Phytologist 213, 1886-1897.
• Rousseau-Gueutin M., Belser C., Da Silva C., Richard G., Istace B., Cruaud C., Falentin C., Boideau F., Boutte J., Delourme R., Deniot G., Engelen S., Ferreira de Carvalho J., Lemainque A., Maillet L., Morice J., Wincker P., Denoeud F., Chèvre A.M., Jean-Marc Aury J.M., 2020. Long-reads assembly of the Brassica napus reference genome, Darmor-bzh. GigaScience 9: 1-16 https://doi: 10.1093/gigascience/giaa137
• Rousseau-Gueutin M., Morice J., Coriton O., Huteau V., Trotoux G., Nègre S., Falentin C., Deniot G., Gilet M., Eber F., Pelé A., Vautrin S., Fourment J., Lodé M., Bergès H. & Chèvre A. M. (2017). The Impact of Open Pollination on the Structural Evolutionary Dynamics, Meiotic Behavior and Fertility of Resynthesized Allotetraploid Brassica napus L. G3: Genes|Genomes|Genetics, 7(2), 705-717.  https://doi.org/10.1534/g3.116.036517
• Rousseau-Gueutin, M., Keller, J., Ferreira De Carvalho, J., Aïnouche, Martin, G. (2018). The Intertwined Chloroplast and Nuclear Genome Coevolution in Plants. In: Diah Ratnadewi, dir., Dr. Hamim, Plant Growth and Regulation - Alterations to Sustain Unfavorable Conditions (p. 61-84). Londres, INT: InTech Open, DOI : 10.5772/intechopen.75673
• Sochorová J., Coriton O., Kuderová A., Lunerová J., Chèvre A. M. & Kovařík A. (2017). Gene conversion events and variable degree of homogenization of rDNA loci in cultivars of Brassica napus. Annals of Botany, 119(1), 13-26.  https://doi.org/10.1093/aob/mcw187
• Stein A., Coriton O., Rousseau-Gueutin M., Samans B., Schiessl S.V., Obermeier C., Parkin I.A., Chèvre A.M., Snowdon R.J.  (2017) Mapping of homoeologous chromosome exchanges influencing quantitative trait variation in Brassica napus. Plant Biotechnology Journal. doi: 10.1111/pbi.12732