Light is a very important factor affecting every aspect of plant development. Plant developmental responses to light are sensitive to the direction, intensity, color, and duration of light. Light is perceived by an extensive set of photoreceptors that includes the red/far-red light–absorbing phytochromes and blue/UV-A light–absorbing cryptochromes. The Arabidopsis mutant seedling hyper-responsive to light 6 (shl6) has exaggerated developmental responses to available light. In the low light, shl6 seedlings have a phenotype similar to wild-type plants grown in high light, with short hypocotyls, expanded cotyledons, and well-developed first true leaves. In addition, the roots of shl6 are short and highly branched. The SHL6 gene was mapped to a position on chromosome 5 between simple sequence length polymorphism (SSLP) markers nga249 and nga151. Two cosmid clones from this interval (introduced by Agrobacterium-mediated transformation) complemented the shl6 mutant phenotype. One candidate gene identified by complementation is a member of the glycosyltransferase family. The sequence of shl6 mutant differs from wild type Columbia allele of this gene (At5g13000) by a single nucleotide substitution in the first exon. This putative SHL6 gene encodes a member of a glycan synthase-like (GSL12) gene family that includes callose synthase. The β-1,3-D-glucan callose is found in the cell plate of dividing cells, in pollen mother cell walls, and pollen tubes. Callose synthase and related genes have not been previously implicated in developmental responses to light. We also observed that 90% of Col-0 anthers showed high callose deposition, but shl6 mutant did not display callose deposition in the anthers. The pollen viability in the shl6 was lower than Col-0. The epidermal cell elongation in shl6 hypocotyls was reduced when compared with Col-0. Therefore, we conclude that the mutation in light-regulated SHL6/GSL12 was involved in the synthesis of callose as well as light signaling.

<p>Plant resistance hypothesis says that under a period of time when a plant is exposed to powerful herbivore attack it will prioritise defence as a major metabolic function. In theory, induced plant defence (resistance) will provide opportunities for this organism to “invest” in other functions, in example growth when attackers are absent.</p><p>One of the compounds taking part in plant defence is callose. This β-1,3-glucan is synthesised by callose synthase and broken down by β-1,3-glucanase. Deposition of callose occurs as a reaction to aphid attack an varies, depending on cultivars, and aphid species. In this experiment barley (Hordeum vulgare) cultivar Clipper is being infested with two types of aphids: Russian wheat aphid (RWA, Diuraphis noxia) and bird cherry-oat aphid (BCA, Rhopalosiphium padi) over a time period. Infestation by those two insects results in different callose formation and deposition level.</p><p>Six sequences encoding for putative callose synthase genes and nine sequences encoding for β-1,3-glucanase were examined by RT-PCR and Real – Time PCR methods for different expression patterns.</p><p>The results did not show any significant regulation of gene expression during RWA and BCA attack for any of these genes. Thus the pathway regulating aphid – induced callose deposition in barley reminds unresolved.</p>

Plant resistance hypothesis says that under a period of time when a plant is exposed to powerful herbivore attack it will prioritise defence as a major metabolic function. In theory, induced plant defence (resistance) will provide opportunities for this organism to “invest” in other functions, in example growth when attackers are absent. One of the compounds taking part in plant defence is callose. This β-1,3-glucan is synthesised by callose synthase and broken down by β-1,3-glucanase. Deposition of callose occurs as a reaction to aphid attack an varies, depending on cultivars, and aphid species. In this experiment barley (Hordeum vulgare) cultivar Clipper is being infested with two types of aphids: Russian wheat aphid (RWA, Diuraphis noxia) and bird cherry-oat aphid (BCA, Rhopalosiphium padi) over a time period. Infestation by those two insects results in different callose formation and deposition level. Six sequences encoding for putative callose synthase genes and nine sequences encoding for β-1,3-glucanase were examined by RT-PCR and Real – Time PCR methods for different expression patterns. The results did not show any significant regulation of gene expression during RWA and BCA attack for any of these genes. Thus the pathway regulating aphid – induced callose deposition in barley reminds unresolved.

Spores are fundamental to the reproductive success of all land plants. The success of a spore lies in its recalcitrant multi-layered spore wall commonly made of sporopollenin, cellulose, and pectin. However, other polysaccharides may be associated with the intine of spores, and their patterns of deposition vary across taxa. Callose, a plant 1-3-β-glucan polysaccharide, has unsubstantiated accounts of its presence and absence in association with the spore mother cell wall of hornworts for more than a century. To address this conundrum, I used aniline blue, a fluorochrome that has high specificity of binding to beta glucans and when excited with ultraviolet light, it will fluoresce yellow-green. However, due to the limited resolution power of that technique, I also used transmission electron microscopy (TEM) with immunogold labeling to observe the ultrastructural localization of callose using anticallose, a monoclonal antibody. I also bioinformatically probed the genomes and transcriptomes of hornworts to elucidate the callose synthase genes and enzymes which may be responsible for the putative callose deposition. Because of the asynchronous spore development, each sporophyte has a continuum of spores at various developmental stages. Subsequently, the sporophyte of many hornworts makes an ideal system to study all aspects of sporogenesis. For the first time, I provide unequivocal, correlative evidence for callose involvement in spore wall development in hornworts. Here we report on the spatiotemporal deposition of callose in sporogenesis of Phaeoceros carolinianus, and we show that callose is a common wall constituent of the spore intine in three other genera (Anthoceros, Notothylas, and Nothoceros). Callose deposition in hornworts is post-meiotic and begins during early spore wall development after a white lined lemella is formed and during expansion of the exine. As sporopollenin is deposited and the spore wall thickens, callose remains localized in the intine during the remainder of sporogenesis. The occurrence of callose in hornwort spores is a first record of this polysaccharide in the inner spore wall (intine or endospore) of any embryophyte. This suggests that callose may serve the same or similar roles in hornwort intine development and function as pectin-cellulose does in later diverging taxa. Bioinformatic tblastn techniques and molecular high through put Illumina genome sequencing combined with blast techniques for orthologs to callose synthase genes from Arabidopsis thaliana and Physcomitrella patens did not provide any evidence as to if callose synthase genes are present in hornworts This was due to database contamination, sample contamination, and sample quality.

Le grain de pollen, gamétophyte mâle, est le produit de la méiose mâle. Il présente une structure extrêmement diversifiée chez les Angiospermes. Afin d’étudier cette diversité, nous avons centré notre étude sur le type apertural. Défini par le nombre, la forme, et la disposition des apertures (zone amincie de la paroi), le type apertural joue un rôle majeur dans les processus de germination et de survie du pollen. L’étude de la méiose mâle a montré que les variations dans le type apertural peuvent être corrélées avec : le type de cytocinèse, la forme des tétrades, la formation des parois entre les futurs grains de pollens, et la position polaire ou groupée des apertures au sein de la tétrade. Récemment, chez un certain nombre d’espèces, il a été mis en évidence une corrélation entre la localisation des derniers dépôts additionnels de callose après la formation des parois intersporales et la position des futures apertures. Nous avons étudié la diversité et l’évolution des composants de la microsporogenèse ainsi que des dépôts de callose additionnels au sein des Angiospermes basales et des Monocotylédones. L’étude du développement de nombreuses morphologies polliniques variées chez ces groupes révèle la présence de dépôts additionnels de callose dans la grande majorité des cas. Nous avons montré qu’il existait un lien entre ces derniers dépôts et la position de la future aperture quelque soit la morphologie du grain de pollen. De plus, pour une même morphologie pollinique, nous avons mis en évidence des dépôts additionnels de callose différents. 7 types de microsporogenèse différents ont été observés pour les pollens monosulqués. Ils présentent les différences dans le type de cytocinèse, la forme des tétrades, la présence et la forme de dépôts de callose additionnels. Nous avons également étudié des espèces sélectionnées présentant différentes configurations d’apertures appartenant à la famille des Bromeliaceae, produits par divers types de dépôts de callose additionnels. On a reconstruit, par la méthode du maximum de parcimonie, l’évolution de différents caractères de la microsporogenèse et la présence ou l’absence des dépôts additionnels de callose intervenant dans la détermination du type apertural. Nos résultats confirment que la forme des tétrades tétragonales, en association avec le type de cytocinèse successive, le mode de formation des parois de façon centrifuge, et la présence des dépôts additionnels de callose, sont les états ancestraux chez les Monocotylédones. De plus, la forme des tétrades tétraédriques, en association avec le type de cytocinèse simultané, le mode de formation des parois de façon centripète, et la présence des dépôts additionnels de callose, sont les états ancestraux chez les Angiospermes basales. Les transitions observées des composants de la microsporogenèse ainsi que le type de dépôts additionnels de callose montrent la variabilité de ces caractères du développement chez les espèces étudiées. Les transitions observées des formes de la tétrade sont associées à des transitions au niveau du type de cytocinèse. Cela confirme un lien étroit entre ces deux caractères. Les transitions dans le mode de formation des parois, associées aux transitions dans tous les autres caractères, indiquent une corrélation entre les différents composants de la microsporogenèse chez les Angiospermes. En présence de dépôts de callose additionnels, la détermination du type apertural est déconnectée des composantes de la microsporogenèse. Des variations des dépôts additionnels conduisant à des pollens monosulqués montrent que le développement (les dépôts additionnels) n’est pas soumis à des contraintes et que la stase sur le type apertural monosulqué résulte de pressions de sélection. / The pollen grain, the male gametophyte of seed plants, is the product of microsporogenesis (male meiosis). It has a considerable structural diversity in flowering plants. To investigate this diversity, we have focused our study on the aperture pattern. The aperture pattern of pollen grains is defined by the shape and number of apertures, and their position when pollen grains are still assembled in tetrads at the end of meiosis. The outer pollen wall is punctuated with apertures, areas where the exine is thinner or even absent. The apertures are the places where the pollen tube emerges at germination. They also have an essential functional role for the survival of pollen grains. The study of microsporogenesis has shown that variations in the aperture pattern can be correlated with features of this developmental process: the type of cytokinesis, the tetrad shape, the way in which callose walls are formed among the four microspores and the position of the apertures within the tetrad (polar or grouped at the last point of contact among the microspores). Recently, in a certain number of species, a correlation has been evidenced between the location of the last points of additional callose deposition and aperture location after wall formation among the future microspores. We studied the diversity and evolution of the above-mentioned features of microsporogenesis and additional callose deposition at the late tetrad stage in a selection of species from basal angiosperms and monocots (with a particular sampling effort in Bromeliaceae) with various aperture patterns. The study of microsporogenesis in these groups revealed the presence of additional callose deposits in most cases. We show that there is a link between the last points of callose deposition and aperture location for several aperture types. In addition, for the same pollen morphology, we have observed different patterns of callose deposition. We described seven different microsporogenesis pathways associated to monosulcate pollen grain. They differ in the type of cytokinesis, tetrad shape, and presence and shape of additional callose deposition. Using Maximum Parsimony, we reconstructed the evolution of the features of microsporogenesis and the presence or absence of additional callose deposits likely to play a key role in aperture pattern determination. Our results confirm that the shape of tetragonal tetrads, in association with the successive cytokinesis, centrifugal cell wall formation, and the presence of additional callose deposits, is the ancestral states in monocots. Using our results on basal angiosperms, we confirm that tetrahedral tetrad, in association with simultaneous cytokinesis, centripetal cell wall formation, and the presence of additional callose deposits, are ancestral at the level of angiosperms. Our results highlight the large variability of the features of microsporogenesis in our study species. The transitions observed in the tetrad form character associated in the most cases with transitions in the cytokinesis character, confirm the relationship between these two characters. The transitions in intersporal wall formation are associated with transitions in all other characters. This suggests a correlation among the different components of microsporogenesis in angiosperms. On the opposite, the presence of additional callose deposits, which seems to be the key element in aperture pattern determination (contrary to previous hypotheses) is disconnected from the other features of microsporogenesis. The variations additional callose deposits leading to monosulcate pollen show that the development (additional callose deposits) is not due to developmental constraints and stasis on monosulcate apertural pattern is resulting from selective pressures.

The field of nanoecotoxicology has been pioneered in recent years as concern grows in response to the potential environmental hazards of engineered nanoparticle release. Silver nanoparticle (AgNP) release through induction into commercial products as an antimicrobiont is of particular interest. Plausible routes of AgNPs to reach aquatic systems and their biological impacts have been investigated, but none have addressed the potential remediation of these waters using the heavy metal accumulating fern Azolla caroliniana. This study employed biological staining techniques and fluorescence microscopy to identify oxidative stress, wounding responses of cell wall and membrane, and cell death of A. caroliniana roots to assess the capability of this plant to withstand AgNP exposure. Two concentrations series were applied, 0-1.0ppm and 0-10.0ppm for 1, 3, 5 days after transfer (DAT), 0ppm being a control. Oxidative stress, measured in production of non-specific ROS, increased in a dose-dependent manner with increasing AgNP concentration. Callose (1,3-β-glucan) was deposited in response to potential cell wall damage and was also observed to be elevated in a dose-dependent manner. Cell vitality appeared from a general decline in fluorescence of nucleic content to visual nuclei lysis. Statistically significant and severe responses to AgNPs was observed at 1 DAT but recovery could be seen at 3~5 DAT. In sum, these data suggest a toxicity threshold of 1.0ppm at which A. caroliniana roots can mediate exposure.

No description available.

Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal.

Peptídeos de sinalização ou hormonais desempenham papéis importantes nas plantas por serem determinantes no crescimento, desenvolvimento e defesa. RALF é um peptídeo ubíquo no reino vegetal e na planta modelo Arabidopsis thaliana os peptídeos RALF formam uma família multigênica de 37 membros alguns com expressão gênica tecido-específica e outros com expressão em toda a planta. A isoforma AtRALF1 é a mais estudada e é expressa principalmente na raiz e no hipocótilo. Uma das funções deste peptídeo é a regulação da expansão celular, um processo que também envolve auxina, giberelina, etileno, brassinosteróides e citocininas. O objetivo deste trabalho foi estudar a relação entre o AtRALF1 e o hormônio etileno, principalmente na expansão celular e alcalinização celular que são efeitos característicos do peptídeo. AtRALF1 inibe o crescimento da raiz primária porém plantas de arabidopsis tratadas simultaneamente com o peptídeo e inibidores da biossíntese ou percepção de etileno não tiveram o crescimento de suas raízes inibido. Etileno induz a deposição de calose e plantas selvagens expostas ao AtRALF1 e plantas que super-expressam AtRALF1 exibiram depósito de calose nas células da ponta da raiz. Curiosamente, quando a alcalinização do meio de células em suspensão induzida por AtRALF1 foi avaliada perante o aumento da produção de etileno ou perante o bloqueio de sua síntese ou percepção, não foi observada alteração na resposta. Os resultados aqui apresentados demonstram que a resposta de inibição da expansão celular ocasionada por AtRALF1 é dependente de etileno e sugere que os efeitos de alcalinização do meio extracelular e da inibição do crescimento da raiz primária estão dissociados. / Peptide hormones or signaling peptides play important roles that determine growth, development and defense in plants. RALF is a ubiquitous peptide in the plant kingdom and in model plant Arabidopsis thaliana comprises a multigene family of 37 members, some of them with tissue-specific gene expression and others that are expressed throughout the plant. AtRALF1 isoform is the most studied and is expressed in roots and hypocotyls. One of the peptide functions is the negative regulation of cell expansion, a process also controled by auxin, gibberelin, ethylene, brassinosteroids, cytokinin. The aim of this work was to study ethylene and AtRALF1 peptide relation, mainly in the cell alkalinization and inhibition of cell expansion responses that characteristic of the peptide. AtRALF1 inhibits root growth but simultaneous treatment with both AtRALF1 and inhibitors of the ethylene perception or biosynthesis show no inhibition on root growth. Ethylene increases callose deposition and wild-type plants treated with AtRALF1 or plants overexpressing the AtRALF1 gene show increased callose plate formation on root tips. Curiously, when the extracellular alkalinization induced by AtRALF1 was evaluated against ethylene production or against ethylene inhibitors, no alteration was observed. The data presented here reveal that the cell expansion inhibition caused by AtRALF1 is ethylene dependent and suggest that the extracellular alkalinization response and root growth inhibition are dissociated.

Spores are single-celled dispersal units surrounded by a wall of the highly resistant biopolymer sporopollenin. All land plants produce spores. Spore development is described in Physcomitrella patens, a moss with single-celled spores, and Dendroceros, a hornwort with multicellular spores. Correlated light, fluorescence and immuno-electron microscopy localizes callose in the aperture of developing spores in the model moss Physcomitrella. Twelve copies of callose synthase genes were annotated bioinformatically and compared with Arabidopsis callose synthase genes. This study identifies a suspect gene involved in moss spore exine development. Unicellular spores of Dendroceros following meiosis remain in tetrads, fill the intercapsular space, and are surrounded by a convoluted, homogeneous electron-opaque outer wall and narrow fibrillar inner wall. No precise pattern of cell division leads to multicellular spores of variable shape and cell number. Evolution of precocious endospory in epiphytic hornworts is a means to protect nascent spores while it develops biochemical and structural machinary to withstand drying. To advance knowledge of genetic control of spore wall development, the sequenced genome of Physcomitrella is probed using a bioinformatic approach to decipher the evolution of five selected genes putatively involved in spore wall formation. Those encoding for callose synthase provide the most complete results. Callose involvement in spore development is a plesiomorphic feature of land plants. Phylogenomic analysis of callose synthases in land plants with sequenced genomes revealed a single moss callose synthase basal in a clade containing the only Arabidopsis callose synthase implicated in exine development of pollen walls as well as two clades of moss specific callose synthase proteins. A predicted protein-protein interactome was constructed to investigate the protein landscape in Physcomitrella for proteins involved in sporogenesis. Orthologous genes were identified between Physcomitrellaand several other species to map orthologous interactions and predict the first bryophyte interactome. The Physcomitrella predicted protein-protein interactome contains 41,936 unique interactions for 4062 different proteins, none of which are associated with sporogenesis. Rather the most conserved interactions among proteins were those associated with metabolic processes. The utility of predicted protein interactions to infer biological roles, providing provisional molecular roadmaps is demonstrated to generate hypotheses for experimental approaches.