Tall fescue is the most common cool-season grass in the eastern USA, with broad economic and ecological importance to the region. Tall fescue is known to associate with a fungal endophyte, Neotyphodium coenophialum, whose presence can decrease biotic and abiotic stress experienced by the plant. This thesis evaluates the response of tall fescue and the fungal endophyte symbiosis to predicted climate change. I participated in two multi-factor climate change projects where I investigated the response of tall fescue tissue chemistry and growth to various climate change factors. Endophyte-infected (E+) tall fescue had decreased alkaloid production under elevated CO2 but increased alkaloid production under elevated temperatures. Significant differences between E+ and E- (endophyte-free) tall fescue tissue chemistry were also found, suggesting the endophyte interacts with the plant response to abiotic stress. Although several studies have reported benefits of endophyte infection for tall fescue growing under drought stress, my research found no differences between E+ and E- total growth and surprisingly showed increased mortality of E+ individuals under elevated temperature. Taken together, my research indicates that this grass-fungal relationship will respond to climate change, and may produce dramatic and unforeseen results that question the widely believed mutualistic nature of the symbiosis.

The Leaf Economics Spectrum (LES) describes the continuous range of leaf ecophysiological strategies that plants across the globe utilize to achieve a net-positive return on their resource investment. This spectrum is measured by traits such as leaf mass per area, leaf nitrogen, and leaf lifespan. Unsuitable leaf trait combinations are selected against in nature due to a net-loss on the return of initial resource investment, while some potentially extremely effective trait combinations may not be possible due to genetic constraints. Existing genetic variation in LES traits, however, suggests the spectrum may be less rigid than previously expected. To test this, we exploit phenotypic plasticity by subjecting cultivated sunflower, Helianthus annuus, to environmental stress to generate variability in LES traits. In a greenhouse experiment, treatment groups of Helianthus were exposed to six varying intensities each of four acute stress applications: nutrient limitation, high soil salinity, heat, and simulated herbivory. Leaves formed during the onset of stress and post-stress were sampled to measure leaf traits; changes in plant fitness were estimated through total plant biomass and the time of first flowering. Results from statistical analyses show a variety of significant effects that stress applications had on plant growth and LES traits. Our findings explore the possibility of achieving off-axis LES trait combinations through salt-induced trait plasticity, and if further studies are needed to reassess the rigidity of the spectrum.

Plants are continuously exposed to different abiotic and biotic stresses in their natural environment. Their capacity to survive depends on the capacity to perceive external signal and quality amount a defence response for protection from the stress perceived. The purpose of this project was to study the impact of combined abiotic stress and biotic stress on the outcome of the disease inducing Arabidopsis thaliana – Pseudomonas syringae interaction. This study included a focus on the role of ABA in these interactions and also whether 3´-O-β D- ribofuranosyl adenosine (hereafter it called ‘400’ compound), a novel adenosine derived compound induced during compatible interactions, was involved. The later involved the targetted disruption of a putative 400 biosynthetic pathway involving analysis of knockout mutants of enzymes; APD-ribose diphosphatase NAD binding / hydrolases of the NUDIX class, glucosyl transferases, ribosyltransferases, a ribose-phosphate pyrophosphokinase3 and galactosyltransferases. Unfortunately, none of these targeted interventions modified the host response to Pseudomonas infection, nor altered levels of 400 in challenged leaves. The primary research investigated the interaction between abiotic and biotic stresses in Arabidopsis plants focussing on the modulation of plant defence against multiple, and possibly antagonistic, stress responses and the role plant hormones play in this process. We showed that high light caused enhanced susceptibility to the already virulent Pseudomonas syringae DC3000pvsp61. The pathways contributing to this enhanced susceptibility were largely ABA independent. Subsequent characterization of transgenic lines expressing the soluble Arabidopsis abscisic acid receptors, PYRABACTIN RESISTANCE1-LIKE4-6 provided compelling evidence for a role for these receptors in DC3000 virulence strategies, but they contribute to a lesser extent to the enhanced susceptibility under high light. This was corroborated genetically by using mutants of the immediately downstream targets of PYLs, the type two protein phosphatase, specifically the triple mutant hab1-1/abi2-1/abi1-2. A number of epitope and fluorescent constructs were generated to facilitate future studies of the role of ABA signaling. Targetted profiling suggested that SA dynamics were altered under DC3000 challenged Arabidopsis grown under high light. Furthermore, differential accumulation of flavonoids suggested these may also play a role in attenuating host defences under high light. Finally we provide evidence based on comparative analysis of that the photoreceptors phytochrome double mutant phyA-211/phyB-9 and cry1/cry2 behave antagonistically in Arabidopsis response to DC3000. Overall our studies support the conclusion that plants abiotic stress (HL) response takes precedence over biotic stress (DC3000) responses and that abiotic stress is detrimental to plant immunity. The luciferase transgenic PYL lines showed high level of expression of ClucP::PYL5 plant tissues challenged 2hpi of DC3000 (OD600: 0.15) in comparison with C1lucP::PYL6. This result opposes to what RT-PCR reported; which was that three PYLs genes display similar expression level at 6hpi of hrpA or 18hpi of DC3000. The epitope tags of CaMV::HA transgenic plants showed HA-tagged signal with stunted phenotype in a range of PYL4, 5 and 6 plants but none of the plants displayed any differences in susceptibility to DC3000. Although, RT-PCR assay showed high levels of expression in the three PYLs, 6hpi of hrpA but no signal was detected in B8eGFP::PYL5 transgenic line either followed the DC3000 and hrpA infection or by examined plant seedlings at early stages under confocal microscopy.

Genotypes adapted to contrasting environments are expected to behave differently when placed in common controlled conditions, if their sensitivity to environmental cues or intrinsic growth behaviour are set to different thresholds, or are limited at distinct levels. This allows natural variation to be exploited as an unlimited source of new alleles or genes for the study of the genetic basis of quantitative trait variation. My doctoral work focuses on analysing natural variation for shoot growth and response to the environment in A. thaliana. Natural variation analyses aim at understanding how molecular genetic or epigenetic diversity controls phenotypic variation at different scales and times of plant development and under different environmental conditions, and how selection or demographic processes influence the frequency of those molecular variants in populations for them to get adapted to their local environment. As such, the analysis of A. thaliana natural variation can be addressed using a variety of approaches, from genetics and molecular methods to ecology and evolutionary questions. During my PhD, I got the chance to tackle several of those aspects through my contributions to three independent projects which have in common to exploit A. thaliana natural variation. The first one is the analysis of the pattern of polymorphism from a set of 102 A. thaliana accessions at the MOT1 gene coding for a molybdate transporter (an essential micronutrient) and responsible for contrasted growth and fitness among accessions in response to Mo availability in the soil. I showed at different geographical scales that MOT1 pattern of polymorphisms is not consistent with neutral evolution and shows signs of diversifying selection. This work helped reinforce the hypothesis that in some populations, mutations in MOT1 have been selected to face soils rich in Mo and potentially deleterious despite their negative effect on Mo-limiting soils. The second project consists in the characterisation and functional analysis of two putative receptor-like kinases (RLKs) identified from their effect on shoot growth specifically under mannitol-supplemented media and not in response to other osmotic constraints. The function of such RLKs in A. thaliana, which is not known to synthesize mannitol was intriguing at first but, through different experiments, we built the hypothesis that those RLKs could be activated by the mannitol produced by some pathogens such as fungi and participate to plant defensive response. The third project, in collaboration with Michel Vincentz's team from CBMEG (Brasil) and Vincent Colot (IBENS, Paris), consists in the analysis of the occurrence of natural epigenetic variants of the QQS gene in different populations from Central Asia and their possible phenotypic and adaptive consequences. Overall, these analyses of the genetic and epigenetic molecular variation leading to the biomass phenotype(s) in interaction with the environment provide clues as to how and where in the pathways adaptation is shaping natural variation.

Des génotypes adaptés à des environnements contrastés ont de grandes chances de se comporter différemment lorsqu’ils sont placés dans des conditions similaires et contrôlées, notamment si leur sensibilité aux signaux environnementaux et/ou leur croissance intrinsèque sont limitées à différents niveaux. De ce fait, la variabilité observée dans les populations naturelles peut être utilisée comme une source illimitée de nouveaux allèles ou gènes pour l’étude des bases génétiques de la variation des traits quantitatifs. Mon travail de doctorat a consisté en l’analyse de la variabilité naturelle pour la croissance et la réponse à l’environnement chez Arabidopsis thaliana. Le but des approches de génétique quantitative est de comprendre comment la diversité génétique et épigénétique contrôle la variabilité phénotypique observée dans les populations à différentes échelles, au cours du développement et sous différentes contraintes environnementales. De plus, ces analyses ont pour objectif de comprendre comment les processus adaptatifs et démographiques influencent la fréquence de ces variants dans les populations en fonction de leur environnement local. Ainsi, l’étude de la variabilité naturelle peut être appréhendée en utilisant diverses approches, de la génétique et des méthodes de biologie moléculaire aux études écologiques et évolutives. Au cours de mon doctorat, j’ai eu la chance de travailler sur plusieurs de ces aspects au travers de trois projets indépendants qui exploitent tous la variabilité naturelle d’A. thaliana.Le premier projet a consisté en l’analyse du pattern de polymorphisme observé dans des populations d’A. thaliana au gène MOT1 qui code pour un transporteur de molybdate (la forme assimilable du molybdène (Mo), un micro-élément essentiel) et qui est responsable d’une partie des variations de croissance et de fitness observées à l’échelle de l’espèce en fonction de la disponibilité en Mo des sols. J’ai montré à différentes échelles géographiques que le pattern de polymorphisme à MOT1 ne reflète pas une évolution neutre mais présente plutôt des traces de sélection diversifiante. Ce travail a contribué à renforcer l’hypothèse selon laquelle des mutations au niveau du gène MOT1 pourraient avoir été sélectionnées dans certaines populations pour faire face aux niveaux élevés de Mo observés dans certains sols et potentiellement délétères malgré leur effet négatif sur des milieux pauvres en Mo.Le deuxième projet portait sur la caractérisation et l’analyse fonctionnelle de deux récepteur-kinase putatifs (RLK) identifiés de part leurs effets sur la croissance foliaire spécifiquement en réponse à un stress induit par du mannitol mais pas sous d’autres contraintes osmotiques. La fonction de ces récepteurs chez A. thaliana -qui n’est pas connu pour produire du mannitol- peut paraître intrigante. Les différentes expériences réalisées au cours de cette thèse nous ont cependant permis de construire un modèle selon lequel ces récepteurs pourraient être activés par le mannitol produit par certains pathogènes tel que les champignons et participer aux réponses de défense de la plante.Le troisième projet a été réalisé en collaboration avec l’équipe de Michel Vincentz (CBMEG, Brésil) et de Vincent Colot (IBENS, Paris) et consiste en l’analyse de l’occurrence de variants épigénétiques naturels au gène QQS dans différentes populations d’Asie Centrale et de leurs possibles conséquences phénotypique et adaptative.En conclusion, l’analyse des variants génétiques et épigénétiques naturels à l’origine des variations de biomasse en interaction avec l’environnement permet de comprendre comment l’évolution façonne la variabilité naturelle. / Genotypes adapted to contrasting environments are expected to behave differently when placed in common controlled conditions, if their sensitivity to environmental cues or intrinsic growth behaviour are set to different thresholds, or are limited at distinct levels. This allows natural variation to be exploited as an unlimited source of new alleles or genes for the study of the genetic basis of quantitative trait variation. My doctoral work focuses on analysing natural variation for shoot growth and response to the environment in A. thaliana. Natural variation analyses aim at understanding how molecular genetic or epigenetic diversity controls phenotypic variation at different scales and times of plant development and under different environmental conditions, and how selection or demographic processes influence the frequency of those molecular variants in populations for them to get adapted to their local environment. As such, the analysis of A. thaliana natural variation can be addressed using a variety of approaches, from genetics and molecular methods to ecology and evolutionary questions. During my PhD, I got the chance to tackle several of those aspects through my contributions to three independent projects which have in common to exploit A. thaliana natural variation. The first one is the analysis of the pattern of polymorphism from a set of 102 A. thaliana accessions at the MOT1 gene coding for a molybdate transporter (an essential micronutrient) and responsible for contrasted growth and fitness among accessions in response to Mo availability in the soil. I showed at different geographical scales that MOT1 pattern of polymorphisms is not consistent with neutral evolution and shows signs of diversifying selection. This work helped reinforce the hypothesis that in some populations, mutations in MOT1 have been selected to face soils rich in Mo and potentially deleterious despite their negative effect on Mo-limiting soils. The second project consists in the characterisation and functional analysis of two putative receptor-like kinases (RLKs) identified from their effect on shoot growth specifically under mannitol-supplemented media and not in response to other osmotic constraints. The function of such RLKs in A. thaliana, which is not known to synthesize mannitol was intriguing at first but, through different experiments, we built the hypothesis that those RLKs could be activated by the mannitol produced by some pathogens such as fungi and participate to plant defensive response. The third project, in collaboration with Michel Vincentz’s team from CBMEG (Brasil) and Vincent Colot (IBENS, Paris), consists in the analysis of the occurrence of natural epigenetic variants of the QQS gene in different populations from Central Asia and their possible phenotypic and adaptive consequences. Overall, these analyses of the genetic and epigenetic molecular variation leading to the biomass phenotype(s) in interaction with the environment provide clues as to how and where in the pathways adaptation is shaping natural variation.