Orientador: Marcelo Carnier Dornelas / Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-27T11:18:53Z (GMT). No. of bitstreams: 1 Rocha_DiegoIsmael_D.pdf: 4893065 bytes, checksum: 1f8621283157c9a9ebce9de36cac067a (MD5) Previous issue date: 2015 / Resumo: Em Passiflora, a grande variabilidade das estruturas florais e cores permitiu adaptação dessas espécies a diversos tipos de polinizadores. Em geral, espécies polinizadas pelo mesmo grupo de animais exibem um conjunto de caracteres que determinam esses sistemas de polinização. O presente estudo teve como objetivo caracterizar o desenvolvimento de estruturas florais peculiares de espécies do gênero (androginóforo e corona), bem como, os aspectos bioquímicos (cor, aroma e néctar) de espécies com diferentes sistemas de polinização (ornitofilia e quiropterofilia) e nos híbridos interespecíficos obtidos entre as mesmas. Nossas análises evidenciaram que a espécie ornitófila (P. coccinea - flor vermelha) e a espécie quiropterófila (P. setacea - flor branca) apresentaram caracteres morfológicos, bem como, a composição química do néctar contrastantes estando essas características florais de acordo com os seus sistemas de polinização. Nos híbridos, nem todas as características foram intermediárias, em relação às espécies parentais. O androginóforo curvo, comumente, observado em espécies do gênero polinizadas por morcegos foi atribuído como resposta à barreira física exercida pelo perianto e a redistribuição de auxina nos tecidos dessa estrutura. Com relação à cor, os híbridos apresentaram tonalidades de vermelho diferentes da coloração escarlate observada em P. coccinea. Porém, a diversidade de pigmentação caracterizada nestes genótipos de Passiflora pôde ser explicada pela combinação de moléculas específicas de antocianinas acumuladas em suas pétalas. Quanto à composição química do aroma, os híbridos apresentaram uma composição qualitativamente similar ao de P. coccinea, não sendo observados a herdabilidade de compostos característicos do aroma de P. setacea (benzenoides). Possível correlação entre cor e aroma foi sugerida. Acredita-se que esses resultados possam contribuir para a compreensão da evolução destes sistemas de polinização dentro do gênero Passiflora / Abstract: In Passiflora, the great variability of floral structures and colours allowed adaptation of these species to different types of pollinators. In general, species pollinated by the same group of animals exhibit a set of characters that determine these pollination systems. This study aimed to characterize the development of unique floral structures of the genus (androgynophore and corona), as well as the biochemical aspects (colour, scent and nectar) of species with different pollination systems (ornithophily and chiropterophily) and in interspecific hybrids obtained between of them. Our analyzes have shown that both, morphology and nectar composition from ornithophilous (P. coccinea - red flower) and chiropterophilous species (P. setacea - white flower) are contracting, and these resources according to their pollination systems. In hybrids, not all features were intermediate in relation to the parental species. The bend androgynophore, commonly observed in the Passiflora species pollinated by bats was assigned in response to the physical barrier exerted by the perianth and the redistribution of auxin in tissues of that structure. Regarding colour, the hybrids showed different shades of red staining in comparison to the scarlet red observed in P. coccinea. However, the diversity of pigmentation characterized in these Passiflora genotypes could be explained by the combination of specific molecules anthocyanins accumulated in their petals. Regarding scent composition, hybrids showed a similar composition to the P. coccinea and, it was not observed heritability of characteristic P. setacea compounds (benzenoids). Possible correlation between colour and scent has been suggested. It is believed that these results may contribute to understanding the evolution of these pollination systems within the genus Passiflora / Doutorado / Biologia Vegetal / Doutor em Biologia Vegetal

Comprendre comment la coordination des cellules entre elles permet l’émergence d’une forme est une des questions les plus fascinantes en biologie du développement. Au cours de cette thèse, nous avons utilisé les premiers stades de développement des feuilles dentelées d'Arabidopsis thaliana comme modèle pour étudier la relation entre les évènements cellulaires et la morphogenèse. Pendant le développement des feuilles d'Arabidopsis thaliana, le contrôle fin de la prolifération et de l'expansion cellulaire permet la croissance différentielle au niveau de la marge foliaire, nécessaire à la formation des indentations. Dans ce modèle, la croissance différentielle est le résultat de l'interaction entre la signalisation de l’auxine et l’activité des facteurs de transcription CUP SHAPED COTYLEDONS impliqués dans le maintien de l'identité des domaines frontières. Pour affiner la compréhension des relations complexes entre les facteurs de transcriptions CUC, les réponses auxiniques et les événements cellulaires à l'origine des indentations foliaires, nous avons utilisé des expériences d’imagerie en temps réel sur des primordia foliaires de lignées exprimant des rapporteurs de développement et/ou de réponse auxinique. Nos résultats ont révélé un contrôle dynamique de la croissance différentielle à la marge des feuilles et l'implication critique de CUC3 dans la répression locale de la croissance cellulaire. / How a shape arises from the coordinated behavior of cells is one of the most fascinating questions in developmental biology. Here we used the early stages of development of serrated leaves in Arabidopsis thaliana as a model to study the tight relation between cellular behaviour and morphogenesis. During Arabidopsis thaliana leaf development the fine control of cell proliferation and cell expansion sustains differential growth at the margin required for the formation of leaf outgrowth named teeth. In this model, differential growth is the result of interplay between auxin signaling and CUC transcription factors that are involved in the maintenance of boundary domain identity. To clarify the interconnected relations between patterns of CUC TFs and auxin responses as well as the cellular events behind serrations we used time-lapse experiments on vegetative primordia of lines expressing developmental and/or auxin response reporters. Our results revealed a tight and dynamic control of differential growth at the leaf margin and the critical involvement of CUC3 in the local repression of cell growth in combination with low auxin responses.

La symbiose endomycorhizienne à arbuscules (SMA) a une importance écologique et agronomique majeure car elle permet à la majorité des plantes terrestres, via une association avec des champignons du phylum des Glomeromycota, une meilleure acquisition de nutriments du sol. Les champignons grâce à un large réseau mycélien dans le sol, collectent et echangent des nutriments avec les plantes qui leurs fournissent des substrats carbonés issus de la photosynthèse. Le champignon mycorhizienne à arbuscules (CMA) Rhizophagus irregularis, capable de coloniser les racines de la plupart des plantes terrestres, sécrète des lipo-chitooligosaccharides (Myc-LCOs) et des chitooligosaccharides à chaîne courte (Myc-COs). L’ajout exogène de Myc-LCO stimule la colonisation de plusieurs espèces par des CMAs. Ces Myc-LCOs et les Myc-COs sont capables d’activer une voie de signalisation symbiotique requise pour l’établissement de la SMA. Cependant, le rôle de ces molécules et leur importance dans l’établissement de la symbiose restent à ce jour inconnu. Au cours de ma thèsej’ao étudié la SMA chez la plante modèle pour les monocotylédones tempérées, Brachypodium distachyon. Je me suis intéressé, tout d’abord, à la perception des signaux fongiques (Myc-LCOs et Myc-COs). Un mécanisme possible par lequel les Myc-LCOs pourraient stimuler la SMA, est leur capacité à induire une augmentation du nombre de racines latérales (RL), montrée chez la légumineuse modèle Medicago truncatula ainsi que chez la monocotylédone Oriza sativa (riz). Les réponses transcriptionnelles aux Myc-LCOs chez M. truncatula suggèrent une interaction entre la perception de Myc-LCOs et la signalisation auxine. Par ailleurs, l’ajout d’auxine stimule la SMA chez M. truncatula et le riz. Pendant mes travaux de thèse, nous avons pu montrer que des LCOs induisent aussi la formation de RLs chez B. distachyon. De plus, nous avons pu montrer un effet des LCOs sur l’homéostasie de l’auxine. Finalement, nous avons pu confirmer un rôle positif de l’auxine dans la mise en place de la SMA grâce à l’étude d’un mutant surproduisant de l’auxine. Par ailleurs, des travaux dans l’équipe ont permis d’identifier BdLYR1, un récepteur kinase contenant des motifs lysin (LysM-RLK) capable de lier les LCOs à haute affinité. Cependant, un mutant dans ce gène ne présente aucun phénotype mycorhizien suggérant une redondance fonctionnelle au niveau des récepteurs et/ou des signaux pour l’activation de la voie de signalisation symbiotique. J’ai initié la caractérisation du LysM-RLK BdLYR4, un récepteur potentiel de COs. Enfin, il a été observé que la réponse de croissance des plantes à la colonisation par les CMA est à la fois dépendantes de l’environnement, de la souche de champignon utilisée ainsi que du génotype de plante. Très peu d’études ont été réalisées dans le but d’identifier les mécanismes moléculaires qui régissent ces réponses de croissance. Pour pouvoir le faire, j’ai initié la caractérisation de la variabilité génétique pour les réponses de croissance aux CMAs chez B. distachyon dans le but d’identifier des génotypes contrastés ainsi que les variables phénotypiques les plus pertinentes à analyser. Ces travaux ont permis de trouver des conditions de culture qui permettent l’analyse de la réponse croissance chez B. distachyon. / The endomycorrhizal symbiosis with arbuscules (AM) is of major ecological and agronomic importance because it allows the majority of terrestrial plants, via an association with phylum fungi of Glomeromycota, to facilitate their acquisition of nutrients and water. Fungi through a large soil exchange surface collect and provide nutrients to plants that provide fungi with carbonaceous substrates derived from photosynthesis. The fungus AM Rhizophagus irregularis, capable of colonizing the roots of most terrestrial plants, secretes lipo-chitooligosaccharides (Myc-LCOs) and short-chain chitooligosaccharides (Myc-COs). These molecules are capable of activating a signaling pathway required for the establishment of the AM symbiosis. However, their role and relevance for AM symbiosis is completely unkown. During my phD, I studied perception of fungal signals (Myc-LCOs and Myc-COs) in B.distachyon. Myc-LCOs stimulate AM symbiosis in several plants and induce an increase in the number of lateral roots (RL) in legume plant Medicago truncatula, as well as in Oriza sativa (rice).Transcriptional responses to LCO in M.truncatula suggest a crosstalk between auxin and LCO. Moreover, the addition of auxin also stimulates AM symbiosis in M. truncatula and rice. During my phD, we showed that LCOs stimulates also lateral root formation in B.distachyon. In addition, we showed that these developmental effects are linked to a regulation of auxin biosyntheses and homeostasis. Finally, I could confirm the positive role of auxin in AM thanks to an auxin-overproducer mutant. In the team, BdLYR1, a high-affinity LCO binding protein has been identified. However, the mutant Bdlyr1 is not affected in mycorrhization, as it was also shown for its ortholog in rice. This can be explained by functional redundancy of the receptors but also by redundancy of fungal signals (Myc-CO). During my phD, I focused on BdLYR4, which is a good candidate for Myc-COs receptor. It has been reported mycorrhizal growth responses (MGR) in colonized plants. These growth responses depend on environmental conditions, fungal strain and plant genotype. To date, very few studies have been done in order to identify molecular mechanisms controlling MGRs. To do so, I initiate the characterization of natural variability for MGRs in B.distachyon in order to identify contrasted genotypes and the more convenient phenotipical and physiological traits to analyze. All this work leads us to identify good conditions that maximize differences between genotypes.

PhD thesis focuses on soil biochemistry, namely the effects of organic and inorganic substances on proteolytic ktivity in forest soils. The thesis is composed of several impacted journal articles and two manuscripts. Selected organic compounds feature L- and D-tartaric acid, phenolics, proteinogenic and non-proteinogenic amino acids, auxins and cytokinins, whilst inorganic substances are represented by salts. Furthermore, an article dealing with the effects of different forest management practises on soil enzymatic ktivity (including proteases) is adjoined. Its aim is to monitor the effect of forest regenerative elements (forest stand gaps) on enzymatic ktivity, whereas the intervention into forest ktivity changes the moisture and thermal conditions of the site. The results show that out of the organic substances tested L- and D-tartaric acid, proteinogenic and non-proteinogenic amino acids and auxins have a positive effect on the proteolytic ktivity. Conversely, phenols and cytokinins acted as inhibitors. Soil salinization also had a negative effect on proteolytic ktivity of forest soils. The Influence of forest management through forest regenerative elements on soil protease ktivity was not detected. Potential urease ktivity was the only enzyme that reacted to the change of climatic conditions of forest habitat; however, ktiv not a principal topic of this work.

The diploma thesis was aimed to monitor the expression profiles of selected genes in the growing of cotyledonary buds in pea (Pisum sativum L.). The apical stem was decapitated and 0.5% auxin paste was applied to the stump. From the treated plants cotyledonary buds were collected in intervals 0, 1, 3, 6, 12, 24, 48 and 120 hours. The biological material was analyzed for expression of genes PsDRM1, PsBRC1, PsAUX1, PsPIN1, PsYUC1. A gene expression normalization factor was calculated based on constitutive genes PsEF1-alpha, PsActin and Ps beta-tubulin. Application of auxin to the stem stump prevented the growth of cotyledonary buds. This inhibition of cotyledonary buds was accompanied by increased expression of dormancy markers PsDRM1, PsBRC1 and auxin biosynthesis gene PsYUC1 and decreased expression of the auxin transporter genes PsPIN1 and PsAUX1.

京都大学 / 0048 / 新制・課程博士 / 博士(理学) / 甲第21250号 / 理博第4420号 / 新制||理||1634(附属図書館) / 京都大学大学院理学研究科生物科学専攻 / (主査)教授 長谷 あきら, 教授 鹿内 利治, 准教授 小山 時隆 / 学位規則第4条第1項該当 / Doctor of Science / Kyoto University / DGAM

Tomato is a major horticulture crop grown across the globe. Unfortunately, its yield is reduced by 25% because of auxin herbicides and glyphosate drift. In this present study, wild germplasm of tomato was screened for herbicide tolerance. From the greenhouse study nine accessions for glyphosate and 2,4-D, eleven accessions for dicamba, five accessions for quinclorac, eight accessions for aminocyclopyrachlor, and two accessions for picloram and aminopyralid were identified to be tolerant. A few accessions were selected from each herbicide tolerant group for field trials at two locations in Mississippi in 2016 and 2017. Results indicated that TOM18 was most tolerant to dicamba herbicide, while TOM87 and TOM129 to glyphosate and quinclorac herbicide, respectively, on the basis of yield and injury. Molecular experiments were conducted to measure the genetic diversity among diverse germplasm. Genetic diversity analysis showed wild accessions to be highly diverse as compared to cultivated tomato.

No description available.

Cell dedifferentiation is a cell fate regression process in which the cell fate memory of a differentiated cell is erased, leading to regain stem cell characteristics. Auxin regulates both cell dedifferentiation and differentiation in plants. It is unknown how auxin controls the two opposite processes. Here the minimal auxin requirements for cell dedifferentiation were found, molecular markers associated with the cell dedifferentiation event were identified. When cellular auxin concentration exceeds the level of meristem cell, most differentiated cells undergo dedifferentiation. In differentiated cells, the polar auxin efflux system prevents cell dedifferentiation by reducing auxin accumulation, particularly in the presence of exogenous auxin. Classic plant tissue culture experiments have shown that exposure of cell culture to a high auxin to cytokinin ratio promotes root formation and a low auxin to cytokinin ratio leads to shoot regeneration. Since the auxin level is highly elevated in the shoot meristem tissues, it is unclear how a low auxin to cytokinin ratio promotes the regeneration of shoots. To identify genes mediating the cytokinin and auxin interaction during organogenesis in vitro, three allelic mutants that display root instead of shoot regeneration in response to a low auxin to cytokinin ratio are identified using a forward genetic approach in Arabidopsis. Molecular characterization shows that the mutations disrupt the AUX1 gene, which has been reported to regulate auxin influx in plants. Meanwhile, it was found that cytokinin substantially stimulates auxin accumulation and redistribution in calli and some specific tissues of Arabidopsis seedlings. In the aux1 mutants, the cytokinin regulated auxin accumulation and redistribution is substantially reduced. These results suggest that auxin elevation and other changes stimulated by cytokinin, instead of low auxin or exogenous auxin directly applied, is essential for shoot regeneration. In this study, as a part of interaction between auxin and cytokinin it was identified that the induction of ARR5 and ARR6 expression by cytokinin is subjected to the regulation of auxin. The expression of ARR5 and ARR6 follows a mutual exclusive pattern in response to the induction of exogenous auxin in Arabidopsis seedlings and calli. The results suggest that auxin interacts with the cytokinin via a gene and tissue specific induction of the negative regulators in the cytokinin signaling pathway.

Plants and pathogenic microorganisms are in constant conflict with each other. Understanding the molecular networks that trigger resistance, along with the molecular networks that pathogens might co-opt to infect susceptible plants, is important for developing the integrated, holistic perspective that is necessary for innovative development of engineered resistance to current and emerging pathogens. The first objective of the dissertation was to increase the understanding of mechanisms by which plants recognize pathogen attack and mount an appropriate defense response. These experiments focused on resistance triggered by the Arabidopsis thaliana R gene, RPP7, which encodes a coiled-coil nucleotide binding-leucine-rich repeat (CC-NB-LRR) protein that activates race-specific resistance to the downy mildew pathogen, Hyaloperonospora parasitica (Hpa). Previously-published genetic epistasis tests have established that RPP7 activates defense responses through a signaling mechanism that does not require accumulation of salicylic acid (SA), or components of the ethylene and jasmonate response pathways. Furthermore, RPP7 is not strongly compromised by mutations in genes associated with defense signal transduction (PAD4, NDR1, NPR1, RAR1). Double mutant combinations of these signal transduction components were analyzed to detect additive or functionally-redundant contributions to RPP7-dependent resistance. Most of the double mutants support an enhanced level of asexual sporulation compared to the single mutant parental lines. Time-course experiments with histochemical stains revealed that these double mutants delay, but do not suppress, the oxidative burst and the hypersensitive response. These results suggest that RPP7 activates multiple signaling pathways, each of which makes incremental contributions to the timing of defense activation. The second objective of the dissertation was to investigate the role that auxin plays in enabling virulent H. parasitica to colonize Arabidopsis. Transcript profiling revealed induction of auxin-associated genes in response to infection of Arabidopsis thaliana by virulent strains of the oömycete pathogen, H. parasitica. Experiments with the DR5 / Ph. D.