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The Arabidopsis cyclic nucleotide-gated ion channels (AtCNGCs) form a large family consisting of 20 members. It has been suggested that CNGCs contribute to a wide array of biological functions such as pollen tube growth and pathogen defence signalling. However, the precise mechanisms by which AtCNGCs act, and the extent of their biological roles, have yet to be fully elucidated. AtCNGC11/12, the chimeric CNGC that resulted from the fusion of AtCNGC11 and 12, induces a number of pathogen defence related phenotypes in the Arabidopsis mutant cpr22. Spontaneous lesion formation is one such phenotype. Interestingly, when AtCNGC11/12 is transiently expressed in N. benthamiana it causes cell death which was characterized in this study. Also, AtCNGC11/12 was used to investigate the structural features responsible for the proper function and regulation of AtCNGCs. Electron microscopic analysis of the AtCNGC11/12-induced cell death showed similar characteristics to programmed cell death (PCD), such as plasma membrane shrinkage and vesicle formation. Interestingly caspase-1 inhibitors and the silencing of vacuolar processing enzyme, a plant enzyme with caspase-1 activity, suppressed the induction of cell death. Additionally, pharmacological analyses indicated that the AtCNGC11/12-indiced cell death was also dependent on Ca2+. Furthermore, 3 amino acid residues, R190, A225, and G287, were demonstrated to be essential for AtCNGC11/12-induce cell death. Taken together, these results indicate that the cell death that develops in the cpr22 mutant is indeed PCD and that AtCNGC11/12, is at the point of, or up-stream of, the Ca2+ signal necessary for the development of HR. Furthermore, the functionality of AtCNGC11/12 as a model for AtCNGC structure-function analyses was demonstrated by the identification of several amino acids necessary for cell death development. Yoshioka et al. (2006) demonstrated that the loss of AtCNGC11 or 12 results in decreased resistance to avirulent isolates of the oomycete pathogen, H. arabidopsidis. Thus, the present biological role suggested for AtCNGC11 and 12 is in pathogen defence, specifically within effector triggered immunity (ETI). Like AtCNGC11 and 12, AtCNGC2 has been demonstrated to contribute to pathogen defence signalling but has also been implicated in other physiological responses such as ion stress and senescence. To better understand the roles of AtCNGC11 and 12 in both pathogen defence and other Ca2+ dependent signalling processes, I have investigated promoter:GUS reporter lines, as well as, AtCNGC11 and 12 KO and RNAi silenced lines subjected to various treatments. From this work, I have demonstrated that AtCNGC11 and 12 have similar expression patterns during pathogen defence, development, and dark-induced senescence. Additionally, the findings presented here further characterize AtCNGC11 and 12 as contributors to ETI rather than PAMP triggered immunity. Furthermore, I demonstrated that AtCNGC11 and 12 are likely involved in the endogenous movement of Ca2+, contributing to a range of Ca2+ associated signalling pathways including gravitropism and senescence. Taken together, these results have greatly improved the characterization of AtCNGC11 and 12; significantly contributing to the understanding of a large and increasingly important channel family.

The Arabidopsis cyclic nucleotide-gated ion channels (AtCNGCs) form a large family consisting of 20 members. It has been suggested that CNGCs contribute to a wide array of biological functions such as pollen tube growth and pathogen defence signalling. However, the precise mechanisms by which AtCNGCs act, and the extent of their biological roles, have yet to be fully elucidated. AtCNGC11/12, the chimeric CNGC that resulted from the fusion of AtCNGC11 and 12, induces a number of pathogen defence related phenotypes in the Arabidopsis mutant cpr22. Spontaneous lesion formation is one such phenotype. Interestingly, when AtCNGC11/12 is transiently expressed in N. benthamiana it causes cell death which was characterized in this study. Also, AtCNGC11/12 was used to investigate the structural features responsible for the proper function and regulation of AtCNGCs. Electron microscopic analysis of the AtCNGC11/12-induced cell death showed similar characteristics to programmed cell death (PCD), such as plasma membrane shrinkage and vesicle formation. Interestingly caspase-1 inhibitors and the silencing of vacuolar processing enzyme, a plant enzyme with caspase-1 activity, suppressed the induction of cell death. Additionally, pharmacological analyses indicated that the AtCNGC11/12-indiced cell death was also dependent on Ca2+. Furthermore, 3 amino acid residues, R190, A225, and G287, were demonstrated to be essential for AtCNGC11/12-induce cell death. Taken together, these results indicate that the cell death that develops in the cpr22 mutant is indeed PCD and that AtCNGC11/12, is at the point of, or up-stream of, the Ca2+ signal necessary for the development of HR. Furthermore, the functionality of AtCNGC11/12 as a model for AtCNGC structure-function analyses was demonstrated by the identification of several amino acids necessary for cell death development. Yoshioka et al. (2006) demonstrated that the loss of AtCNGC11 or 12 results in decreased resistance to avirulent isolates of the oomycete pathogen, H. arabidopsidis. Thus, the present biological role suggested for AtCNGC11 and 12 is in pathogen defence, specifically within effector triggered immunity (ETI). Like AtCNGC11 and 12, AtCNGC2 has been demonstrated to contribute to pathogen defence signalling but has also been implicated in other physiological responses such as ion stress and senescence. To better understand the roles of AtCNGC11 and 12 in both pathogen defence and other Ca2+ dependent signalling processes, I have investigated promoter:GUS reporter lines, as well as, AtCNGC11 and 12 KO and RNAi silenced lines subjected to various treatments. From this work, I have demonstrated that AtCNGC11 and 12 have similar expression patterns during pathogen defence, development, and dark-induced senescence. Additionally, the findings presented here further characterize AtCNGC11 and 12 as contributors to ETI rather than PAMP triggered immunity. Furthermore, I demonstrated that AtCNGC11 and 12 are likely involved in the endogenous movement of Ca2+, contributing to a range of Ca2+ associated signalling pathways including gravitropism and senescence. Taken together, these results have greatly improved the characterization of AtCNGC11 and 12; significantly contributing to the understanding of a large and increasingly important channel family.

No description available.

Du point de vue écologique, les efforts de tension ou de compression développés par la maturation du bois au cours de la croissance permettent à l'arbre de maintenir sa posture verticale face à la gravité. Ces efforts, appelés autocontraintes de maturation, résultent de la mise en place d'un bois particulier, le bois de réaction. Sur le plan mécanique, c'est la dissymétrie des autocontraintes entre les deux faces du tronc de l'arbre qui joue un rôle moteur dans le redressement, qui s'exprime par une courbure. Ce travail vise à développer de nouveaux outils théoriques et métrologiques pour étudier le lien entre morphologie de l'arbre et autocontraintes dans un cadre biomécanique et écologique. Une première approche vise à mettre en relation la dynamique de croissance et la compétition du peuplement avec la morphologie et la réaction de contrôle de la posture des arbres. Elle utilise des données issues d'un essai sylvicole de long terme (plantations de hêtres de différentes densités initiales laissées en croissance pendant 26 ans). L'analyse rétrospective de la production du bois de réaction sur des rondelles prélevées sur ces arbres a permis d'évaluer les différentes composantes du mouvement gravitropique au cours du temps. Une loi d'échelle, établie entre la vitesse de courbure et la circonférence des tiges, montre (i) l'effet prépondérant du diamètre de la tige dans sa capacité de réaction, et (ii) l'absence d'effet additionnel de la compétition. En fin d'expérience, l'évaluation de la morphologie des arbres a permis de confirmer le lien entre l'inclinaison et l'élancement de la tige avec les indicateurs de contraintes de croissance. D'un point de vue mécanique, la morphologie de l'arbre s'interprète (i) par la forme de sa tige (inclinaison et courbures), et (ii) par la distribution spatiale de sa biomasse, qui peuvent fournir des variables biomécaniques candidates pour élaborer des modèles de stimulus-réponse. Ainsi, un important travail méthodologique couplant la technologie LiDAR terrestre (un instrument de numérisation laser 3D très prometteur pour les mesures forestières) à des techniques de modélisation géométrique a permis d'obtenir des maquettes 3D précises de la structure ligneuse d'arbres de différentes espèces. Les maquettes ont permis de modéliser la contrainte de flexion exercée par la biomasse aérienne des arbres en vue d'une mise en relation avec les indicateurs de contraintes de croissance mesurées sur leur grume. Les résultats montrent que la contrainte de flexion est une variable très prometteuse pour évaluer le degré de réaction des arbres. La méthodologie ouvre également des perspectives originales pour le suivi temporel de la morphologie de l'arbre en lien avec son interprétation biomécanique. / From an ecological point of view, the tension or compression stresses developed by wood maturation during growth allow the tree to maintain its vertical posture against gravity. These stresses, called growth stresses, results from the formation of a particular wood called reaction wood. From a mechanical point of view, the asymmetry of growth stresses between the two opposite faces of the tree trunk causes its reorientation, characterised by a curvature. This work aims at developing new theoretical and metrological tools to assess the link between tree morphology and growth stresses in a biomechanical and ecological framework. The first approach aimed at establishing the link between growth dynamics and competition of the stand with tree morphology and reaction of posture control. It is based on data taken in a long-term forestry experiment (beech plantations of different initial planting densities that grew during 26 years). The retrospective analysis of reaction wood production on wood discs taken on the trees allowed to assess the variation of their gravitropic performance over time. The scaling law established between curvature rate and stem circumference showed (i) the leading effect of the diameter of a stem on its reactivity, and (ii) the absence of additional effects of competition. At the end of the experimentation, the assessment of the tree morphology allowed to confirm the link between stem leaning and slenderness with growth stresses indicators. From a mechanical point of view, tree morphology can be assess by (i) the shape of its stem (leaning, curvatures), and (ii) the spatial distribution of its biomass, that can provide biomechanical variables for stimulus-response models. Therefore, an important methodological work was performed, based on terrestrial LiDAR technology (a promising tool for forest measurements based on 3D laser digitisation) coupled to geometrical modelling. It allowed to obtain accurate 3D mocks-up representing the woody structure of trees of variables species. The mocks-up allowed to model the bending stress exerted by the aerial biomass of the trees with the aim of linking it to the growth stresses indicators. The results show that bending stress is a promising variable for assessing the degree of reaction of trees. The developed methodology also gives many perspectives for monitoring tree morphology over time with the aim of biomechanical interpretation.

Les arbres sont capables de modifier l’orientation de leurs branches et de leur tronc par la production asymétrique de bois précontraint. Il existe des modèles biomécaniques développés pour simuler ces mouvements, mais ils ne simulent pas correctement le redressement (ou mouvement gravitropique) de jeunes arbres à l’échelle de temps intra-annuelle. La méconnaissance de la cinétique de maturation et des propriétés mécaniques du bois vert est responsable de ces résultats. Les travaux présentés dans ce mémoire ont pour objectifs de caractériser le comportement mécanique du bois vert au cours de sa maturation, et de développer un modèle biomécanique qui puisse simuler quantitativement le gravitropisme de jeunes peupliers. Des comportements mécaniques non-linéaires sont révélés par des essais de traction cycliques sur de fines lamelles de bois vert. Ils sont quantifiés par une grandeur mécanique liant rigidité et déformation. Des essais de flexion réalisés sur des planchettes renseignent quant à eux sur l’évolution intra-cerne du module élastique. Ces campagnes d’essais montrent une augmentation puis une diminution du module au cours de la maturation des cellules. Des essais de fluage indiquent que le comportement viscoélastique du bois vert se modélise par un modèle de Burgers. Les propriétés viscoélastiques du bois vert sont ainsi déterminées. Les propriétés mécaniques obtenues sont utilisées dans un modèle biomécanique développé pour modéliser l’évolution spatio-temporelle des propriétés. Le gravitropisme de jeunes peupliers est alors modélisé grâce à la prise en compte du comportement viscoélastique du bois vert, de la maturation continue des cellules, et de la variation des déformations de maturation au cours de la saison de végétation. / Trees are able to modify the orientation of their trunk and branches by asymmetrical production of prestressed wood. Biomechanical models designed to simulate these movements exist, but they cannot fit the righting-up movement (also called gravitropism) of young poplar trees at the intra-annual scale. The lack of knowledge of green wood maturation and mechanical properties is suspected to be responsible for this discrepancy. The aims of this study are to characterize mechanical properties of green wood during the maturation process, and to develop a biomechanical model that simulates quantitatively the gravitropism of young poplars. Nonlinear mechanical behavior is observed in cyclic tensile tests performed on thin lamellas of green wood. A relationship between stiffness and strain enables the characterization of this behavior. The intra-ring evolution of modulus of elasticity is measured using 3-points bending tests on small boards. Both of these experimental campaigns show that wood stiffness increases then decreases while cells are maturating. Creep tests show that green wood viscoelastic behaviour is described by a Burgers’ model. As a result, green wood viscoelastic properties are determined. These mechanical properties are used in a new biomechanical model designed for considering spatio-temporal evolutions of wood properties. Then the gravitropic movements of young poplars are simulated by considering viscoelastic behaviour of green wood, continuous maturation of cells, and variation of maturation strains along the growing season.

Auxin is phytohormone that regulates several developmental processes and environmental responses. One of the most well-described outcome of the auxin signalling pathway is regulation of gene transcription. Aux/IAA proteins play an important role in this process, acting as transcriptional repressors. Recent studies revealed that several root growth responses are too rapid to be explained by changes in the level of transcription. The correlation between the amount of Aux/IAAs and the root growth rate suggests that these proteins might be involved in root growth regulation, especially during rapid growth responses that are not associated with transcriptional reprogramming. This work is focused on one of the 29 Arabidopsis Aux/IAA proteins - the IAA17/AXR3 protein. First, we produced stable transgenic lines of Arabidopsis thaliana expressing different combinations of fluorescently labelled AXR3-1 proteins and/or fused to subcellular localization tags under the control of different tissue-specific promoters, in order to characterize the subcellular localization of the studied protein. Subsequent visualization by confocal microscopy methods confirmed information about the role of IAA17/AXR3 protein in root growth responses, its involvement in auxin signalling, and gravitropism. Next, we showed that the...

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