This thesis is focused on the biosystematic study of the Central-European endemic Spergularia echinosperma. With the combined use of morphometric analyses, genome size measurements and molecular tools, the taxonomic issues associated with this species have been clarified. The existence of S. kurkae, a stable allotetraploid hybrid between diploid S. echinosperma and tetraploid S. rubra, has been proven. Based on several lines of evidence, including distinct morphological separation and frequent occurrence in the absence of the parental species, treating S. kurkae as a separate species is proposed. In addition, two infraspecific taxa within S. echinospermaS. echinosperma subsp. echinosperma and S. echinosperma subsp. albensisdiffering in distributions and ecology have been described. A complete revision of the localities of S. echinosperma, S. kurkae and S. rubra in the Czech Republic is also presented. Furthermore, the development of 16 polymorphic microsatellite loci for S. echinosperma is reported.

Cette étude vise à explorer les mécanismes de tolérance des plantes aux xénobiotiques organiques de la famille des HAPs (phénanthrène), à travers l’analyse de l’impact des évènements de spéciation par hybridation et duplication génomique (allopolyploïdie). Nous avons pour cela mené une approche comparative sur un modèle de spéciation allopolyploïde récente, constitué des espèces parentales hexaploïdes S. alterniflora et S. maritima, et de l’allopolyploïde S. anglica qui résulte de la duplication du génome de leur hybride F1 S. x townsendii. Une approche intégrative basée sur des analyses physiologiques et moléculaires nous a permis de montrer que chez Spartina l’hybridation et le doublement du génome augmentent la tolérance aux xénobiotiques. Le parent paternel S. maritima se montre particulièrement sensible au phénanthrène par rapport au parent maternel S. alterniflora. Différentes analyses transcriptomiques ont permis l’identification de novo de transcrits spécifiquement exprimés en condition de stress, et l’annotation des petits ARNs (miARNs, leurs gènes cibles, et siARNs) agissant en tant que régulateurs de l’expression des gènes et la régulation des éléments transposables. Les analyses d’expression différentielle en réponse au stress ont permis de générer un modèle de régulation (miARN/gènes cibles) en réponse aux HAPs, testé par validation fonctionnelle en système hétérologue chez Arabidopsis. Un travail exploratoire de profilage du microbiome de la rhizosphère des spartines exposées au phénanthrène a été réalisé pour préciser les mécanismes de dégradation des xénobiotiques dans l’environnement en vue d’une application dans les stratégies de remédiation verte. / We explored mechanisms involved in tolerance to organic xenobiotics belonging to PAHs (phenanthrene), in the context of allopolyploid speciation (hybrid genome duplication). We developed a comparative approach, using a recent allopolyploidization model including the hexaploid parental species S. alterniflora and S. maritima, and the allopolyploid S. anglica, which resulted from genome doubling of the F1 hybrid S. x townsendii. Integrative approach based on physiological and molecular analyses highlights that hybridization and genome doubling enhance tolerance to xenobiotics in Spartina. The paternal parent S. maritima exhibits higher sensitivity compared to the maternal parent S. alterniflora. Various transcriptomic analyses were performed, to identify de novo stress responsive transcripts, and to annotate small RNAs (miRNAs, their target genes, and siRNAs) involved in gene expression and transposable element regulations. Differential expression analyses in response to stress allowed us to develop a putative miRNA regulatory network (miRNA/target genes) in response to PAH, functionally validated in Arabidopsis as heterologous system. An exploratory profiling of Spartina rhizosphere microbiome exposed to phenanthrene was also performed to characterize environmental degradation abilities, in the perspective of optimizing green remediation strategies.

Understanding the composition, evolution, and function of the cotton (Gossypium) genome is complicated by the joint presence of two genomes in its nucleus (AT and DT genomes). Specifically, read-mapping (a fundamental part of next-generation sequence analysis) cannot adequately differentiate reads as belonging to one genome or the other. These two genomes were derived from progenitor A-genome and D-genome diploids involved in ancestral allopolyploidization. To better understand the allopolyploid genome, we developed PolyCat to categorize reads according to their genome of origin based on homoeo-SNPs that differentiate the two genomes. We re-sequenced the genomes of extant diploid relatives of tetraploid cotton that contain the A1 (Gossypium herbaceum), A2 (Gossypium arboreum), or D5 (Gossypium raimondii) genomes. We identified 24 million SNPs between the A-diploid and D-diploid genomes. These analyses facilitated the construction of a robust index of conserved SNPs between the A-genomes and D-genomes at all detected polymorphic loci. This index can be used by PolyCat to assign reads from an allotetraploid to its genome-of-origin. Continued characterization of the Gossypium genomes will further enhance our ability to manipulate fiber and agronomic production of cotton. We re-sequenced 34 allotetraploid cotton lines, representing all 7 tetraploid cotton species. The analysis of these genomes-using PolyCat and PolyDog-provides us with the beginnings of a HapMap-like resource for cotton species, including indices of both homoeo-SNPs and allele-SNPs. With this information, we explore the phylogenetic relationships among cotton species, including the newly characterized species G. ekmanianum and G. stephensii. We examine gene conversion both recent and ancient, discovering that recent gene conversion is extremely rare, and ancient gene conversion is far less extensive than previously believed, with many previously identified conversion events being more probably due to autapamorphic SNPs in the descent of diploid relatives. In order to carry out these experiments, many tools for next-generation sequence analysis were developed. These tools, along with PolyCat and PolyDog, comprise the tool suite BamBam.

Family Poaceae is one of the most derived groups of monocots. Major mechanisms of grasses evolution and particularly of tribe Triticeae and genus Elymus are polyploidisation and hybridisation. Presented diploma thesis assessed frequency and direction of hybridisation between Elymus hispidus and E. repens in Central Europe. Thesis also evaluated evolutionary importance (significantly higher ratio of hybridisation and polyploidisation) of hybrid swarm in Nature Reserve Čertoryje (Bílé Karpaty Mountains/the White Carpathians, the Czech Republic). Putative hybridizing populations were 14,4 % of sampled localities, i.e. direct presence hybrids (7,9 %) or both parental species (8,4 %). Aneuploids and higher polyploids were found exclusively in Čertoryje hybrid swarm, except one putative DNA nonaploid plant. The genome in situ hybridisation confirmed and improved knowledge of particular species/cytotypes chromosome counts and further specified their genome composition. GISH newly characterized genome composition of interspecific hybrids and higher allopolyploids and revealed their heterogeneous origin. Flow cytometry revealed continuum of absolute genome sizes among parental species and their hybrid. The continuum was rather asymmetric in direction towards E. hispidus. Model hybrid swarm Čertoryje produce...

Traditional evolutionary models depict evolution as a bifurcating pattern with a single ancestor diverging to form two lineages. However, reticulate species resulting from hybridization and recombination have unique histories shared with two independent lineages, not one. Accounting for the genetic histories of reticulate species increases the power and ability to recover biologically meaningful relationships. The genus Collomia (Polemoniaceae) is used to explore issues of reticulation and the importance of accounting for gene histories in a phylogenetic analysis. The issue of reticulation within species trees is discussed with a multilabeled, network approach being explored to better represent the genus's evolutionary history. Wherry's hypotheses regarding the relationships that exist within Collomia are addressed and the need for a new intrageneric section is recognized based on support from multiple, independent genes and morphology. Sections Collomiastrum and Courtoisia remain as previously circumscribed. Section Collomia is modified with removal of two species, C. grandiflora and C. biflora from the section and by erecting a new section, Calyperona. A morphological key is included for each section and their corresponding species, followed by sectional discussions. Finally, the evolution of lifecycle duration, seed morphology, and pollen morphology are discussed based on the phylogeny of the genus.

This thesis comprises studies of the phylogenetic relationships in the flowering plant clade Potentilleae in Rosaceae. The relationships were elucidated by using DNA sequence data from the nuclear genome as well as from the plastid genome. In particular, the focus of the studies was the investigation of allopolyploidy, i.e. speciation as a result of hybridization and subsequent chromosome doubling. A phylogenetic method was used for identifying allopolyploidy through comparison of trees resulting from the analyses of different DNA sequences. Five sub-clades were investigated. First, both the sister clades that together contain all of Potentilleae: Fragariinae and Potentilla. Secondly, three subclades of Fragariinae, namely Alchemilla in wide sense, Sibbaldia and relatives, and Fragaria. The aim was to unravel the phylogenetic relationships, including instances of allopolyploidy. Classification issues were discussed in relation to the phylogenetic results. The split between Potentilla (=Potentillinae) and Fragariinae received better support than in previous studies. The phylogeny of Fragariinae was found to be consistent with classifying ten genera: Alchemilla in wide sense (incl. Aphanes and Lachemilla), Comarum, Sibbaldia, Sibbaldianthe, Sibbaldiopsis, Chamaerhodos, Drymocallis, Dasiphora, Potaninia, Fragaria, and also including a few orphan Potentilla species. The segregated genera Ivesia, Horkelia, Horkeliella and Duchesnea were found to be nested within Potentilla, corroborating earlier studies, while the segregated genus Argentina (P. anserina and close relatives) showed an ambiguous position. Plastid and nuclear (ribosomal) phylogenies were compared and incongruences were detected as potential instances of allopolyploid speciation. Five strongly supported incongruences were detected in Fragariinae and four of them were considered to be potentially caused by allopolyploidy. In addition, five supported incongruences were found in Potentilla. Alchemilla in the wide sense was found to contain four major clades, African Alchemilla, Eurasian Alchemilla, Lachemilla and Aphanes. Both Lachemilla and Aphanes were nested within Alchemilla and it was suggested that the name Alchemilla should be used in the wide sense, i.e. including both the genera Lachemilla and Aphanes. The genus Sibbaldia as commonly classified was shown to be polyphyletic in five different places in Potentilleae. Three Sibbaldia clades ended up in Fragariinae and two in Potentilla. A phylogeny of Fragaria, based on a nuclear low/single copy DNA region was estimated. The gene copy phylogeny was used to construct a reticulate tree hypothesizing allopolyploid speciation events. The evolution of Fragaria was shown to have been shaped by polyploidy. / At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 2: Manuscript. Paper 4: Manuscript. Paper 5: Manuscript.

No description available.

Ray-finned fishes (Actinopterygii) exhibit the greatest biodiversity among vertebrates. The vast majority of extant actinopterygian fish species belong to clade Teleostei - a lineage whose significant evolutionary success might have resulted from a teleost specific whole- genome duplication (TSGD) that occurred at the onset of this group, subsequent to its divergence from the rest of actinopterygian lineages. Despite the growing body of sequenced fish genomes and analyses of their transcriptomes, the largest contribution to understanding fish genomes comes from analyses of DNA content and from cytogenetics. Genomes of ray-finned fishes and especially those of Teleostei exhibit vast diversity and rapid dynamics of repetitive DNA sequences whose variability is reflected in a wide range of fish genome sizes and in the dynamics behind karyotype differentiation. Therefore, ray-finned fishes offer a unique opportunity to study genome variability as a driving force underlying morphological and ecological diversification, evolution and adaptation. Particularly, the mapping of repetitive DNA sequences by means of fluorescence in situ hybridization (FISH) has proven to be a very useful and informative approach during the last two decades and contributed greatly to our understanding of the fish genome...

Le succès évolutif de la polyploïdie, notamment de l’allopolyploïdie (où la duplication de génome complet est associée à une hybridation entre génomes différenciés) est en partie lié au fait que cet événement s’accompagne de nombreux changements dans l'organisation du génome et la régulation de l'expression des gènes. On parle du « choc génomique » de l’hybridation interspécifique et de l’allopolyploïdie. Ces sources de diversité génétique, à la fois structurale et fonctionnelle, apparaissent utiles et nécessaires à l'adaptation et l’évolution des espèces. Alors que de nombreuses études portant sur la compréhension des mécanismes moléculaires à l’origine du succès des allopolyploïdes ont concerné les modifications de l’expression des gènes, mes travaux de thèse ont porté sur les agents non codants du génome que sont les éléments transposables et les petits ARN non codants. Le modèle d'étude est le colza (Brassica napus, AACC), espèce allotétraploïde issue de l'hybridation entre les espèces diploïdes navette (B. rapa, AA) et chou (B. oleracea, CC). Nous disposions de colzas néo-synthétisés, étudiés à différentes générations d’autofécondation, permettant de caractériser les changements génomiques accompagnant la formation puis l’évolution du génome néo-allopolyploïde. Une étude a tout d’abord été menée sur un élément transposable (ET) spécifique du génome C, Bot1, en vue d’identifier de nouvelles transpositions survenant chez les colzas néo-synthétisés par rapport aux parents diploïdes, par une approche SSAP. Quelques rares événements de transposition ont été identifiés. Ces résultats, confrontés à ceux obtenus sur deux autres ET, ont permis de mettre en évidence un impact modéré de l’allopolyploïdie sur la transposition de ces différents ET. Par contre, il est apparu que des changements de méthylation auraient accompagné cette allopolyploïdisation, sans doute à l’origine de la réactivation et la transposition de quelques copies de Bot1. Les petits ARN non codants ont été suggérés comme impliqués dans les différents événements génomiques accompagnant la formation d’un génome allopolyploïde. Pour étudier la dynamique d’expression des petits ARN chez des colzas néo-synthétisés pris à deux générations d’autofécondation (S1, S5) en comparaison de leurs parents diploïdes, j’ai exploité des données de séquençage haut débit obtenues pour 11 banques construites à partir des tiges de ces différents génotypes. J’ai ainsi démontré, qu’à une échelle globale, les petits ARN présentaient une réponse immédiate mais transitoire à l’événement d’allopolyploïdie. Les fractions particulièrement affectées par l’allopolyploïdie se sont révélées correspondre (1) à des petits ARN interférents dérivés d’éléments transposables avec une baisse de leur abondance en génération précoce S1, et (2) à des populations de petits ARN de 21 nucléotides exprimées uniquement de manière très précoce, de l’hybride F1 à la génération S1. Nous avons notamment identifié des transcrits de type viral correspondant à ces petits ARN de 21-nt, et présentant les mêmes profils d’expression (de l’hybride F1 à la génération S1), suggérant une réactivation d’éléments viraux endogènes (EVE) en réponse à l’hybridation et l’allopolyploïdie. L’ensemble de mon étude a démontré la mise en place d’une succession des voies de régulation par petits ARN où ET et EVE, réactivés au niveau transcriptionnel, sont immédiatement soumis à une répression post-transcriptionnelle (PTGS), renforcée ensuite par une répression de leur transcription (TGS). L’hypothèse d’une absence de cette régulation par petits ARN lors des phénomènes de nécrose et létalité hybride, amène à envisager ces populations de petits ARN comme les clés de la réussite de la formation d’un génome hybride, où la répression immédiate et efficace des ET et autres endovirus, réactivés suite au choc génomique, se révèle être une nécessité. / The evolutionary success of polyploid species is partly due to the dynamic changes in genome organization and gene expression patterns that occur at the onset of the polyploid formation. These changes are promoted by the merging of divergent genomes into a single nucleus (i.e. allopolyploidy) that causes a “genomic shock”; they are thought to provide a rich source of new genetic material upon which selection can act to promote adaptation and evolution. Many studies have thus aimed to uncover molecular mechanisms that are responsible for the evolutionary success of allopolyploid species, most of them focusing on gene expression changes. In the present PhD thesis, my interest has been concentrated on the non-coding components of the genome: transposable elements and small non-coding RNAs. My study involves oilseed rape (Brassica napus, AACC), a relatively young allopolyploid species that originated from hybridizations between B. rapa (AA) and B. oleracea (CC). Specifically, I have used resynthesized B. napus polyploids advanced by self-pollination of single plants for several generations; I have analyzed these plants at different generations for genomic changes accompanying polyploid formation and subsequent evolution. In a first part, sequence-specific amplification polymorphism (SSAP) targeting the C genome-specific transposable element Bot1, was used to evaluate transposition rate of Bot1 in resynthesized B. napus in comparison with the diploid parents. Only a few transposition events were identified. When combined with the results obtained for two other TEs, this work suggests that allopolyploidy has only a moderate impact on TE transposition and restructuring. The changes observed in SSAP profiles led us to hypothesize that some of them resulted from changes in DNA methylation, resulting in rare but highly specific TE activation and transposition. In a second part, I have concentrated on small non-coding RNAs (sRNAs), which are thought to mediate different aspects of the response to the “genomic shock” induced by allopolyploid formation. Comprehensive analyses of sRNA expression in resynthesized B. napus allopolyploids have been carried out by deep sequencing sRNAs from 11 libraries prepared from stems of three allotetraploids (surveyed at the two generations S1 and S5) and the two diploid parents. Characterization of sRNA distributions in these plants indicates that sRNAs show an immediate but transient response to allopolyploidy. The sRNAs derived from transposable elements (down-regulated in the S1) or targeting unknown sequences (no Blast hit against any available public database) were particularly affected. The use of B. napus mRNAseq data revealed that these latest unknown candidates, which are 21-nt long and over-expressed in the earliest generations (F1, S0, S1) were derived from endogenous viral elements (EVE). We confirmed that these EVEs showed the same expression patterns as the 21-nt long sRNAs that specifically target them (over-expression in the F1, S0 and S1). These results suggest that (at least) some EVEs might be reactivated as a response to the merging of divergent genomes (in interspecific hybrids and newly formed allopolyploids). Altogether, our results have demonstrated a succession of sRNA pathways that counteract the reactivation of some specific TEs and/or EVEs at the onset of polyploid formation; reactivated TEs and/or EVEs being immediately repressed at the post-transcriptional level (PTGS), and then fully repressed by transcriptional gene silencing (TGS) in the subsequent generations. Such data lead to hypothesize that sRNAs are essential to overcome interspecific hybrid incompatibilities due to the uncontrolled and deleterious reactivation of TEs / EVEs. Therefore, sRNAs should be considered as the guardians of genome integrity even in newly-formed allopolyploids.

No description available.