LEAFY (LFY) est un facteur de transcription central pour le développement des plantes, en particulier pour la floraison chez les angiospermes. LFY est très conservée, même chez les espèces ne portant pas de fleurs. On dispose de nombreuses données génétiques sur LFY et son réseau de régulation chez la plante modèle Arabidopsis thaliana, mais les mécanismes moléculaires impliqués dans son fonctionnement ne sont pas entièrement élucidés. LFY possède deux domaines conservés : un domaine de liaison à l'ADN et un domaine de fonction inconnue en position N-terminal. L'objectif a été de comprendre le rôle du domaine N-terminal et d'étudier l'évolution de la spécificité de liaison à l'ADN de LFY. Nous avons obtenu la structure cristallographique du domaine N-terminal de LFY et découvert qu'il s'agissait d'un domaine SAM (Sterile Alpha Motif) permettant l'oligomérisation de la protéine. Nous avons validé l'importance de cette propriété pour la fonction florale de LFY chez A. thaliana. Nous avons ensuite montré, par des analyses in vitro et in vivo en ChIP-seq que l'oligomérisation influençait la liaison à l'ADN en permettant une liaison coopérative sur plusieurs sites de liaison, en assurant la sélectivité de la protéine vis-à-vis de l'ADN et en permettant l'accès de la protéine à des régions génomiques où la conformation de la chromatine est normalement défavorable à la liaison. Cette étude intégrative a permis de mieux comprendre le fonctionnement de LFY. Des modifications dans les réseaux de régulation de l'expression des gènes sont source de nouveauté et d'évolution. LFY étant très conservée et ne faisant pas partie d'une famille multigénique, nous nous sommes demandé si sa spécificité de liaison à l'ADN avait évoluée. Nous avons montré que LFY était apparue chez les algues multicellulaires et que sa spécificité avait connue au moins deux changements majeurs au cours de l'évolution. Nous avons expliqué ces modifications au niveau moléculaire par des approches de biologie structurale et de biochimie. Nous avons identifié une espèce chez qui LFY a une spécificité relâchée et nous proposons qu'une telle forme ait pu permettre les transitions d'une spécificité à une autre. / LEAFY (LFY) is a key transcription factor for plant development, particularly for flowering in angiosperms. LFY is highly conserved in plants, including non-flowering species. Despite a wealth of genetic data about LFY and its regulatory network in the model plant Arabidopsis thaliana, how the protein works at the molecular level is not fully understood. It has two conserved domains: a DNA binding domain and a N-terminal domain of unknown function. My two main projects were to understand the role of the N-terminal domain and to study LFY DNA binding specificity evolution. We obtained LFY N-terminal domain crystal structure and discovered it was a Sterile-Alpha-Motif (SAM) mediating LFY oligomerisation. We validated the importance of that property for flower development in A. thaliana. Using both in vitro analyses and a ChIP-seq experiment, we pointed out that oligomerisation is required for proper DNA binding. It enables cooperative binding on several LFY binding sites, increases the protein selectivity towards DNA and allows LFY to access genomic regions where the chromatin conformation normally prevents binding. This integrative study provides a better understanding of how LFY works. The rewiring of transcriptional networks provides a rich source of evolutionary novelty. As LFY is highly conserved and single copy in most plant genomes, we asked whether its DNA binding specificity had evolved. We showed that LFY was present since multicellular algae and that it underwent at least two major shifts in DNA-binding specificity during plant evolution. We provided a structural explanation for the two newly identified DNA binding modes and we identified a LFY form with a relaxed specificity that could have served as an intermediate between evolutionary transitions.

Orientadores: Angelo Pires do Prado, Thomas Michael Lewinsohn / Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-09-11T21:19:37Z (GMT). No. of bitstreams: 1 Braun_MarinaReiter_D.pdf: 4685052 bytes, checksum: 6fca81032502f6931c9f8333e10b544b (MD5) Previous issue date: 2011 / Resumo: Na presente tese foram descritas dez novas espécies de Agromyzidae endófagos de capítulos de Asteraceae. Essas espécies estão distribuídas nas duas subfamílias representadas nos gêneros Melanagromyza, Ophyomyia (Agromyzinae), Calycomyza e Liriomyza (Phytomyzinae). No gênero Melanagromyza, segundo maior da família, foram descritas sete novas espécies. Pela primeira vez foi encontrada nos gêneros Calycomyza, Liriomyza e Ophyomyia a herbivoria de capítulos de asteráceas. Além de aumentar o conhecimento sobre a diversidade do grupo, outra contribuição desta tese é a avaliação da especialização alimentar do grupo, que mantém o padrão da família com uma baixa proporção de generalistas. A partir dos dados de interação desses endófagos construímos uma rede de interações entre quinze espécies de Melanagromyza e suas plantas hospedeiras com o objetivo de avaliar sua estrutura de interações. Esta rede foi testada para investigar a existência de aninhamento e compartimentalização dessas interações. Outras questões investigadas foram a relação entre afinidade morfológica dos herbívoros e sua utilização de recursos, além do parentesco das plantas hospedeiras utilizadas pelas espécies de herbívoros. A estrutura de interações dessa rede possui um padrão bastante compartimentalizado, o que era esperado em interações de herbivoria entre insetos que apresentam alta especialização alimentar. Quanto aos efeitos do parentesco na estruturação da rede de interações, verificamos resultados diferentes para os insetos e as plantas. A classificação das espécies de Melanagromyza conforme sua morfologia genital não teve relação com o grupamento dos herbívoros análise de agrupamento bidirecional. Por outro lado, para as plantas, observamos grande concordância entre a classificação taxonômica das espécies e os grupos formados a partir da análise de agrupamento bidirecional / Abstract: This study describes ten new species of endophagous Agromyzidae of Asteraceae flower-heads. Those species belong to genera of the two subfamilies, respectively Melanagromyza, Ophyomyia (Agromyzinae), Calycomyza and Liriomyza (Phytomyzinae). In Melanagromyza, the second largest genus in the family, seven new species were described. Herbivory of Asteraceae flower-heads was recorded for the first time in Calycomyza, Liriomyza and Ophyomyia. Besides increasing the knowledge of agromyzid diversity, this study contributes to the evaluation of host range patterns in the family, which confirms that most agromyzids are specialized on a plant genus or tribe, with a small percentage of generalistic species. From the feeding records between endophages and their host plants we built an interaction matrix, in order to evaluate their interaction structure. The network shows both a nested and a compartmented pattern, suggesting a compound structure. The structure of the analyzed network has both a nested and a compartmented pattern. Compartmentation is expected for interactions between herbivore insects with such specialized feeding habits. In addition, other issues were investigated in this study: Do morphologically similar herbivore species use the same plants? Are plants used by the same herbivore species phylogenetically related? Regarding the family relationship based on the interactions structure, we found divergent results for insects and for plants. The categorization of Melanagromyza species based on their male genital morphology showed no relation with the herbivore groups based on their host use in a bidirectional cluster analysis. On the other hand, plants showed strong agreement between the taxonomic affinities of the species and the groups based on their shared herbivores in the bidirectional analysis / Doutorado / Ecologia / Doutor em Ecologia

CAPES / Honey is generally known for its therapeutic value and the determination of its main floral source allows the certification to the consumer the properties related to its origin. Thus, the aim of this work was to evaluate the quality and characterize eleven honeys from Apis mellifera from the state of Rio de Janeiro according to physico-chemical parameters, to phenolics profile, to the antioxidant activity and also through the use of chemonetric analysis applied to 1H NMR data and HPLC-DAD. Two samples of assa peixe, six samples of cambara and three samples of morr?o-de-candeia honeys from diferente regions of Rio de Janeiro were analyzed. The physico-chemical parameters determined were: HMF content and color through a spectrophotometric method, free acidity and pH. The content of free amminoacids was also determined through the c?dmium-ninhydrine together with total proteins via the Bradford method. The antioxidant ability of honeys and their extracts was qualitatively determined through the total phenolics content using the Folin-Denis method. Total flavonoids were determined by the complexation method with aluminium chloride. The quantitative antioxidant activity of honeys was determined by the trapping of the 2,2-diphenyl-1-picryl-hydrazyl (DPPH) radical, by trapping of the ABTS.+ radical and also by the iron reduction method (FRAP). The identification and quantitation of polyphenols in the extracts were done by HPLC-PDA. The use of multivariate analysis for the 1H NMR data and HPLC enabled the distinction of the honeys analyzed in this work. Thus, the use of 1H NMR and HPLC data combined with multivariate analysis may be employed as a new strategy for the fast and non-destructive typification of Brazilian honeys. / No mercado, m?is s?o conhecidos pelo seu poder terap?utico e a designa??o de sua principal fonte floral permite atestar ao consumidor as propriedades de sua origem. Assim, o objetivo deste trabalho foi avaliar a qualidade e caracterizar onze m?is de Apis mellifera provenientes do Estado do Rio de Janeiro, por meio de par?metros f?sico-qu?micos, do perfil de subst?ncias fen?licas, de seus potenciais antioxidantes, al?m de usar m?todos quimiom?tricos aplicados aos dados de RMN de 1H e CLAE-DAD. Para tanto, utilizou-se duas amostras de mel de assa peixe, seis de cambar? e tr?s de morr?o de candeia de diferentes munic?pios do Rio de Janeiro. Os par?metros f?sico-qu?micos avaliados foram: teor de HMF e a cor utilizando m?todo espectrofotom?trico, acidez livre e pH. Al?m dessas determina??es, o conte?do de amino?cidos livre foi avaliado pelo m?todo de c?dmio-ninidrina, e prote?nas totais pelo m?todo de Bradford. A capacidade antioxidante dos m?is e de seus extratos foi avaliada qualitativamente atrav?s do conte?do de fen?licos total pelo m?todo de Folin-Denis, e de flavon?ides total pelo m?todo de complexa??o com cloreto de alum?nio. A quantifica??o do potencial antioxidante foi realizada pela captura do radical 2,2-difenil-1-picril-hidrazil (DPPH), captura do radical livre ABTS.+, al?m do m?todo de redu??o do ?on f?rrico (FRAP). A identifica??o e quantifica??o das subst?ncias polifen?licas dos extratos foi feita por cromatografia l?quida de alta efici?ncia com detector de arranjo de diodos (CLAE-DAD). A aplica??o da an?lise multivariada aos dados de RMN de H e de CLAE-DAD distinguiu os m?is de assa peixe, cambar? e morr?o de candeia produzidos no Estado do Rio de Janeiro. Assim, o uso de RMN de 1H e CLAE-DAD combinado com a quimiometria pode ser uma nova estrat?gia para tipifica??o de m?is brasileiros de forma r?pida e n?o destrutiva.

Brazil is the largest citrus producer in the world and has a large global citrus market share. However, several diseases affect the crop, being postbloom fruit drop (PFD) one of them. PFD has gained importance in São Paulo State for the displacement of citrus areas to regions with weather conditions more favorable for this disease. The accurate identification of the causal agent of the PFD has been performed and it was renamed as Colletotrichum abscissum. The origin of the initial inoculum is still an enigma for PFD epidemics and the hypotheses that the initial inoculum could be present in propagation material have been discussed but it has never been demonstrated. The objective of this work was to detect and quantify Colletotrichum abscissum from citrus leaves of budwood increase block and citrus nursery plants by qPCR. Four commercial citrus farms from São Paulo State, Brazil with budwood increase block and citrus nursery plants of Pera and Valencia sweet orange varieties were used for this work. C. abscissum was detected in budwood increase block and in nursery plant in both varieties (Valencia and Pera) at the four farms sampled. Out of 122 budwood increase block samples, 89 (73%) were positive for C. absicissum. From nursery plants, out of 175 samples, 129 (73%) were detected with the pathogen. The majority of the positive samples of budwood increase blocks and nursery plants contained 10 to 200 and 10 to 400 conidia of C. absicissum, respectively. With the methods used was not possible to isolate the fungus from vegetative material. This finding suggests a new long distances dispersion type of C. abscissum in the cycle of postbloom fruit drop by propagation material. Confirmation of C. abscissum in budwood increase block and nursery plants would lead to update regulations for the production of certified citrus nursery trees and searching for new control strategies of the pathogen. / O Brasil é o maior produtor de citros do mundo e possui uma grande participação no mercado global de citros. No entanto, várias doenças afetam a cultura, sendo uma delas a podridão floral dos citros (PFC). PFC ganhou importância no Estado de São Paulo pelo deslocamento de áreas de citros para regiões com condições climáticas mais favoráveis para a doença. A identificação precisa do agente causal do PFC foi realizada, tendo sido renomeado como Colletotrichum abscissum. A origem do inóculo inicial ainda é um enigma para as epidemias de PFC e as hipóteses do que o inóculo inicial poderia estar presente no material de propagação já foram discutidas, mas nunca foram demonstradas. O objetivo deste trabalho foi detectar e quantificar Colletotrichum abscissum em folhas de borbulheiras e mudas de citros por meio de qPCR. Neste trabalho, foram utilizadas quatro fazendas comerciais de citros do Estado de São Paulo, Brasil, com borbulheiras e viveiros de mudas de citros das variedades laranja Pera e Valência. C. abscissum foi detectado em borbulheiras e em mudas em ambas as variedades (Valência e Pêra) nas quatro fazendas amostradas. Das 122 amostras de folhas de borbulheiras, 89 (73%) foram positivas para C. absicissum. Das 175 amostras de folhas de mudas de citros, 129 (73%) foram detectadas com o patógeno. A maioria das amostras positivas de borbulheiras e mudas de citros continham 10 a 200 e 10 a 400 conídios de C. absicissum, respectivamente. Com os métodos utilizados, não foi possível isolar o fungo do material vegetativo. Esta descoberta sugere um novo tipo de dispersão a longas distâncias de C. abscissum no ciclo de podridão floral dos citros por meio do material de propagação. A confirmação de C. abscissum nas borbulheiras e mudas de citros levaria à atualização da regulamentação para a produção de mudas de citros certificadas e à busca de novas estratégias de controle do patógeno.

Submitted by Edisangela Bastos (edisangela@ufpa.br) on 2015-10-01T17:02:24Z No. of bitstreams: 2 license_rdf: 22974 bytes, checksum: 99c771d9f0b9c46790009b9874d49253 (MD5) Tese_EcologiaPolinizacaoBuriti.pdf: 1000900 bytes, checksum: 046bde2f9102fa9a27a027cd77e03c04 (MD5) / Approved for entry into archive by Ana Rosa Silva (arosa@ufpa.br) on 2015-10-06T15:18:21Z (GMT) No. of bitstreams: 2 license_rdf: 22974 bytes, checksum: 99c771d9f0b9c46790009b9874d49253 (MD5) Tese_EcologiaPolinizacaoBuriti.pdf: 1000900 bytes, checksum: 046bde2f9102fa9a27a027cd77e03c04 (MD5) / Made available in DSpace on 2015-10-06T15:18:21Z (GMT). No. of bitstreams: 2 license_rdf: 22974 bytes, checksum: 99c771d9f0b9c46790009b9874d49253 (MD5) Tese_EcologiaPolinizacaoBuriti.pdf: 1000900 bytes, checksum: 046bde2f9102fa9a27a027cd77e03c04 (MD5) Previous issue date: 2013 / O estudo das palmeiras nativas é importante por seu grande valor econômico e na manutenção das comunidades de várias espécies de vertebrados e invertebrados que se alimentam de seus frutos, sementes e folhas. A eficiência na produção dos frutos das palmeiras está diretamente relacionada com a presença de insetos polinizadores, principalmente besouros, abelhas e moscas. A palmeira Mauritia flexuosa, comumente conhecida como buriti, é a espécie mais abundante do Brasil e é também chamada de “árvore da vida”, por ser 100% utilizável. Este trabalho teve como objetivo contribuir para o conhecimento da ecologia da polinização do buriti em ambiente de restinga, no município de Barreirinhas, Maranhão, Brasil. Para tanto, obteve-se dados sobre fenologia reprodutiva, biologia floral, sistema reprodutivo e visitantes florais. Para o acompanhamento fenológico foram selecionados 25 indivíduos de cada sexo, os quais foram observados de agosto/2009 a outubro/2012. As fenofases de floração e frutificação foram relacionadas com as variáveis climáticas através de correlação de Spearman. O processo de abertura e longevidade floral foi acompanhado durante o pico de floração da espécie, verificando-se a viabilidade polínica, a receptividade estigmática, as regiões emissoras de odor e a ocorrência de termogênese. Para determinar o sistema reprodutivo foram feitos testes de polinização cruzada e apomixia. O transporte de grãos de pólen pelo vento foi observado, por meio de lâminas de vidros untadas com vaselina que permaneceram penduradas próximas às inflorescências pistiladas durante 24 horas. Os visitantes florais foram coletados através do ensacamento de 20 inflorescências de cada sexo, sendo classificados de acordo com a frequência e o comportamento. O buriti apresentou padrão fenológico anual, sincrônico e sazonal, com floração de agosto a novembro e pico de queda dos frutos em setembro, o que corresponde à estação seca, diferindo do observado na Amazônia, onde estes eventos fenológicos ocorreram na estação chuvosa. Esta diferença pode ser justificada pela grande disponibilidade de água na região, o que faz com que o buriti não necessariamente dependa das chuvas para florescer. Este fato foi evidenciado pela correlação significativa negativa das fenofases com a precipitação e com a umidade relativa. A forte incidência solar e a disponibilidade de água no ambiente contribuíram para o sucesso na floração e frutificação do buriti. Além disto, fatores bióticos podem ter exercido influência no comportamento fenológico, cuja estratégia reprodutiva adotada parece ser a sincronização da floração e da frutificação com a atividade dos polinizadores e dispersores. Dessa maneira a espécie garante a sua reprodução em um período ótimo para a germinação de sementes e estabelecimento de plântulas. O sistema reprodutivo do buriti é xenogâmico. O conjunto de características florais, aliado à abundância de pólen e ao forte odor leva a crer que essa palmeira tenha como principal estratégia de polinização a cantarofilia, porém o vento também possui grande importância na polinização. Além de apresentar polinização do tipo misto (ambofilia), as flores do buriti atraíram uma grande variedade de visitantes, cuja riqueza foi maior que a observada na Amazônia. / The study of native palms is important for its great value economic and in maintenance of communities of several species of vertebrates and invertebrates that feed on their fruits, seeds and leaves. The efficiency in the production of palm fruits is directly related to the presence of pollinating insects, especially beetles, bees and flies. The palm Mauritia flexuosa, commonly known as buriti, is the most abundant species in Brazil and is also called the "tree of life" because it is 100% usable. This study aimed to contribute to the knowledge of the pollination ecology of buriti in Restinga, in the municipality of Barreirinhas, Maranhão, Brazil. Therefore, we obtained data on reproductive phenology, floral biology, breeding system, and flower visitors. For phenological monitoring 25 individuals of each sex were selected, which were observed from August/2009 to October/2012. The stages of a flower and fruit were related to climatic variables through correlation Speaman. The process of anthesis and floral longevity was observed during peak flowering species, verifying the pollen viability, stigmatic receptivity, the regions emitting scent and occurrence of thermogenesis. Tests of cross-pollination and apomixis were made to check the reproductive system. The transport of pollen grains by wind was observed through glass slides greased with vaseline which have remained hanging near the pistillate inflorescences for 24 hours. The floral visitors were collected by bagging inflorescences of 20 of each sex and were classified according to the frequency and behavior. The buriti presented annual, synchronous and seasonal phenological pattern with flowering from August to November and peak fruit falling in September, which corresponds to the dry season, differing from that of the Amazon, where these phenological events occurred in rainy season. This difference can be explained by the greater availability of water in the region, which makes the buriti not necessarily depend on the rains to flowering. This was evidenced by the significant negative correlation of phenophases with precipitation and relative humidity. A strong solar irradiation and water availability in the environment contributed to the success in flowering and fruiting of buriti. In addition, biotic factors may have influenced the phenology, whose reproductive strategy adopted appears to be synchronization of flowering and fruiting with the activity of pollinators and seed dispersers. Thus the species warrants its reproduction in an optimal period for seed germination and seedling establishment. The reproductive system of buriti is xenogamic. The set of floral traits, combined with the abundance of pollen and the strong odor suggests that the buriti has as its main strategy pollination cantharophly, but the wind has a great importance in pollination. Besides presenting a mixed pollination (ambophily), flowers buriti attracted a wide variety of visitors, whose wealth was higher than those observed in the the Amazon.

There is considerable evidence that pollinator shifts can explain many differences in flower morphology between closely related plant species, but the extent to which pollinator shifts can explain the maintenance of among-population variation in floral traits within species is poorly known. In this thesis, I combined comparative and experimental approaches to examine the evolution of floral traits in the moth-pollinated orchid Platanthera bifolia. More specifically, I investigated (1) the relationship between flower and pollinator morphology, (2) phenotypic selection on morphology and phenology in populations in contrasting environments, (3) components of prezygotic reproductive isolation among divergent populations, and (4) the adaptive and functional significance of two correlated floral traits. A study of Scandinavian of P. bifolia populations revealed that spur length was positively correlated with proboscis length of local pollinators, which suggests that variation in spur length reflects adaptive evolution in response to geographically variable pollinator-mediated selection. A phenotypic selection study on Öland, SE Sweden, suggested that disruptive selection on spur length contributes to the maintenance of a bimodal distribution of spur length in mixed habitats, but provided very limited evidence of divergent selection on plant morphology and flowering phenology in grassland and woodland habitats. Field experiments revealed strong reproductive isolation between divergent populations on Öland, due to differences in spatial distribution, flowering phenology, and pollinators, and among-population incompatibility. The results suggest that prezygotic reproductive isolation contributes to the maintenance of population differentiation in floral traits in P. bifolia. A field manipulation experiment demonstrated that spur length but not perianth size affects pollination success and seed production. This suggests that among-population differentiation in perianth size may be the result of a genetic correlation with spur length. Taken together, the results of this thesis suggest that pollinator-mediated selection can shape the evolution of intraspecific floral variation.

The goals of this research project were to investigate aspects of pollination biology of two native boreal species: Vaccinium myrtilloides (Canadian blueberry) and Vaccinium vitis-idaea (lingonberry) in central Saskatchewan. Accordingly, surveys of insect taxa visiting the flowers were performed, and determination of the effectiveness of these insect taxa to serve as pollinators was measured through pollen deposition and pollen tube growth in the style. Accompanying fieldwork, and morphological and anatomical studies were done for the two food rewards offered by flowers of both Vaccinium species: pollen released through poricidal anthers and nectar secreted from the nectary atop the inferior ovary. Pollen-ovule (P/O) ratios were determined for the two study species in Saskatchewan (V. myrtilloides, V. vitis-idaea) as well as for five other Vaccinium species from eastern Canada (Nova Scotia – V. angustifolium, V. boreale, V. caespitosum, V. corymbosum, and V. uliginosum). Pollen, released at maturity as tetrads, were converted to total pollen grains per flower to yield P/O ratios ranging from 238 (V. caespitosum) to 2,008 (V. vitis-idaea), but 736 for the latter in Saskatchewan. These P/O ratios are indicative of a breeding system ranging from facultative autogamy to facultative xenogamy. Additionally, the structure of mature stamens and pollen tetrads was studied in V. myrtilloides and V. vitis-idaea. Each anther was functionally bilocular; had a single-cell thickness (i.e., epidermis) with regularly occurring papillae; lacked an endothecium; and possessed two distal, hollow tubules each terminating in a pore. Overall pollen grain viability was 76-97% (V. myrtilloides) and 51-93% (V. vitis-idaea), with about 20% of tetrads having only 1-3 grains viable, and 12% and 27% of tetrads entirely non-viable in V. myrtilloides and V. vitis-idaea, respectively. Pollen tetrads occasionally were connected by a sticky substance resembling pollenkitt, but viscin threads were absent. One instance of precocious (in situ) germination of tetrads was recorded within anthers of V. myrtilloides. The floral nectary was a disk of secretory tissue situated between the stamens and the style. The epidermis possessed solitary stomata that were variable in number, but not different between V. myrtilloides and V. vitis-idaea. The nectary was vascularized by phloem alone; many traces were found for V. myrtilloides throughout the nectary, whereas V. vitis-idaea had few traces at the nectary base, concentrated at the inner side of the disk closest to the style base. Young sclerenchyma cells were found throughout the nectary parenchyma. Nectar production started on the day of anthesis for both species, although many flowers of V. vitis-idaea appeared to have no measureable nectar at that time. V. myrtilloides produced a larger range of nectar solutes per flower (0 - 3684.1 μg), than V. vitis-idaea iii (1.29 to 1147.62 μg) over both years; nectar volumes per flower never exceeded 5 μL. Nectar was measured daily in flowers aged 1 – 4 days in 2010 and 1 – 6 days in 2011, however, over the two years at the same study site there was no clear pattern of secretion and reabsorption throughout flower life for V. myrtilloides, and only a gradual increase for V. vitis-idaea as flowers aged. Insect visitors to flowers surveyed in 2010 included a large proportion of honeybees (Apis mellifera) as visitors to both species, whereas in 2011 there were no honeybees present at the field site. There was a larger proportion of hoverflies (Syrphidae) found on the flowers of V. vitis-idaea than on V. myrtilloides. Other visitors to V. myrtilloides were bees (Bombus, Andrena, Osmia, Colletes) and wasps (Vespidae), whereas flowers of V. vitis-idaea were visited by bees (Bombus, Andrena, Osmia, Lasioglossum, Colletes, Hylaeus), an ant (Formicidae) and a butterfly (Lycaenidae). Bombus spp. were shown to be pollinators of V. myrtilloides. Andrena spp. were probable pollinators, whereas honeybees appeared to be poor pollinators. Bombus spp. seemed probable pollinators of V. vitis-idaea and hoverflies to be barely more than visitors, though small sample sizes did not allow for conclusive evidence. The time that an insect spent on a virgin flower had no relationship to the pollination result. Among individuals of various Bombus spp. that did or did not sonicate flowers of V. myrtilloides, the action of “buzz pollination” was shown to result in an increase in the number of pollen tetrads deposited and in pollen tube growth.

Comprendre la diversité morphologique des fleurs passe par l'étude de son origine moléculaire et développementale et de ses conséquences fonctionnelles et écologiques. Le périanthe est composé d'organes stériles, sépales et pétales, qui jouent un rôle majeur dans le succès reproducteur des plantes pollinisées par les animaux du fait de leur fonction d'attraction.Cette thèse propose une approche multidisciplinaire visant à comprendre l'origine génétique et moléculaire de la diversité morphologique du périanthe et sa signification évolutive, à l'aide du modèle Nigella damascena L. Cette Renonculacée présente un dimorphisme spontané. La forme probablement ancestrale, trouvée en populations naturelles, a un périanthe bipartite composé de cinq sépales pétaloïdes et huit pétales nectarifères. Dans la forme variante, cultivée à des fins d'horticulture, les pétales sont remplacés par un nombre élevé d'organes allant d'une forme proche des sépales à une forme proche des étamines.La première partie de cette thèse est consacrée à l'étude de l'origine développementale, génétique et moléculaire du dimorphisme, par la caractérisation détaillée de la morphologie florale et de son développement dans les deux morphes dans le cadre d'une approche gène candidat. Par analyse d'expression et validation fonctionnelle, nous avons montré que le gène NdAP3-3 est responsable de l'ensemble des aspects du dimorphisme floral de N. damascena, ce qui suggère que ce gène joue un rôle dans l'identité du pétale mais aussi dans l'architecture du méristème, potentiellement via la régulation du nombre d'organes et de la frontière entre périanthe et étamines.La seconde partie de cette thèse concerne l'impact du dimorphisme floral sur le mode de reproduction des deux morphes et leur maintien potentiel. Nous avons caractérisé les stratégies reproductives et la valeur sélective des deux morphes en conditions naturelles dans des populations expérimentales. Le variant sans pétale est peu visité par les pollinisateurs, et se reproduit majoritairement en autogamie. L'analyse de la vigueur de ses descendants suggère une dépression de consanguinité. Par ailleurs, dans notre matériel, il semble que l'allèle donnant le phénotype sans pétale soit lié à un allèle augmentant la valeur sélective. A la lumière de nos résultats, nous discutons les conditions du maintien de ce polymorphisme.

LEAFY (LFY) est un facteur de transcription central pour le développement des plantes, en particulier pour la floraison chez les angiospermes. LFY est très conservée, même chez les espèces ne portant pas de fleurs. On dispose de nombreuses données génétiques sur LFY et son réseau de régulation chez la plante modèle Arabidopsis thaliana, mais les mécanismes moléculaires impliqués dans son fonctionnement ne sont pas entièrement élucidés. LFY possède deux domaines conservés : un domaine de liaison à l'ADN et un domaine de fonction inconnue en position N-terminal. L'objectif a été de comprendre le rôle du domaine N-terminal et d'étudier l'évolution de la spécificité de liaison à l'ADN de LFY. Nous avons obtenu la structure cristallographique du domaine N-terminal de LFY et découvert qu'il s'agissait d'un domaine SAM (Sterile Alpha Motif) permettant l'oligomérisation de la protéine. Nous avons validé l'importance de cette propriété pour la fonction florale de LFY chez A. thaliana. Nous avons ensuite montré, par des analyses in vitro et in vivo en ChIP-seq que l'oligomérisation influençait la liaison à l'ADN en permettant une liaison coopérative sur plusieurs sites de liaison, en assurant la sélectivité de la protéine vis-à-vis de l'ADN et en permettant l'accès de la protéine à des régions génomiques où la conformation de la chromatine est normalement défavorable à la liaison. Cette étude intégrative a permis de mieux comprendre le fonctionnement de LFY. Des modifications dans les réseaux de régulation de l'expression des gènes sont source de nouveauté et d'évolution. LFY étant très conservée et ne faisant pas partie d'une famille multigénique, nous nous sommes demandé si sa spécificité de liaison à l'ADN avait évoluée. Nous avons montré que LFY était apparue chez les algues multicellulaires et que sa spécificité avait connue au moins deux changements majeurs au cours de l'évolution. Nous avons expliqué ces modifications au niveau moléculaire par des approches de biologie structurale et de biochimie. Nous avons identifié une espèce chez qui LFY a une spécificité relâchée et nous proposons qu'une telle forme ait pu permettre les transitions d'une spécificité à une autre.

Les plantes conservent la capacité à former de nouveaux organes tout au long de leur vie grâce au maintien de structures contenant des cellules souches, les méristèmes. La formation des fleurs, structures reproductives de la plante, est une étape essentielle de son cycle de vie. Afin d’assurer un développement floral complet, un méristème doit être formé de novo au sein du jeune bouton floral. Des données éparses de la littérature indiquent que le facteur de transcription LEAFY, en plus d’être un régulateur clé de l’identité florale, est aussi impliqué dans la mise en place du méristème floral.Dans la première partie de ce travail nous explorons le rôle de LEAFY dans l’initiation du méristème floral. Cette étude est concentrée sur un gène cible de LEAFY, le facteur de transcription REGULATOR OF AXILLARY MERISTEMS1 (RAX1). Nous montrons notamment que la voie régulée par LEAFY/RAX1 agit en parallèle du facteur de transcription REVOLUTA pour permettre la mise en place du méristème floral.Dans la deuxième partie de ce travail nous étudions les propriétés du domaine N-terminal de LEAFY. Ce domaine permet l’oligomérisation de LEAFY ainsi que potentiellement sa liaison aux régions fermées de la chromatine. Nous étudions également de manière plus exploratoire le rôle de ce domaine dans la régulation de l’expression du gène AGAMOUS, un important régulateur du développement floral. / Plants have the capacity to continuously produce organs throughout their life because they maintain stem cells containing structures called meristems. The formation of flowers is an essential step of the plant’s life-cycle. In order to ensure flower development, a new meristem must be formed within the young flower bud. Various data across the literature indicate that the transcription factor LEAFY is involved flower meristem formation in addition to its role as a master regulator of flower identity.In the first part of this work we explore the role of LEAFY in the initiation of flower meristem. This study focuses on a LEAFY target gene, the transcription factor REGULATOR OF AXILLARY MERISTEMS1 (RAX1). We show that the LEAFY/RAX1 pathway acts in parallel of the transcription factor REVOLUTA to allow flower meristem formation.In the second part of this work we study the properties of the N-terminal domain of LEAFY. This domain mediates LEAFY oligomerization and potentially its binding to closed chromatin regions. We also study in a more prospective manner the role of this domain in the transcriptional regulation of AGAMOUS, an important regulator of flower development.