Teses / dissertações sobre o tema "Abiotic stress factors"

Anthropogenic disturbances can alter the abiotic composition of freshwater systems. These compositional changes can act as physiological stressors towards system inhabitants. However, little is known about how these altered abiotic factors influence stress hormones (corticosterone) in larval amphibians. Throughout the following chapters, I examined the effects of several abiotic factors on baseline and stress-induced corticosterone levels in the larvae of four amphibian species: Jefferson salamander (Ambystoma jeffersonianum), spotted salamander (A. maculatum), wood frog (Rana sylvatica), and grey treefrog (Hyla versicolor). Chapter II examined corticosterone level differences throughout development in A. jeffersonianum and R. sylvatica larvae under field, mesocosm, and laboratory venues. Baseline corticosterone levels in R. sylvatica increased near metamorphic climax in all venues, but not in A. jeffersonianum. Rather, baseline corticosterone levels differed with respect to venue throughout development in A. jeffersonianum. Chapter III examined corticosterone level differences among free-living A. jeffersonianum populations and possible abiotic factors underlying these hormone differences. Corticosterone levels significantly differed across populations. Increased baseline corticosterone levels significantly correlated to low pH. There was also a trend for increased baseline corticosterone levels to be positively correlated with chloride levels and negatively correlated with conductivity. Chapter IV examined the effects of laboratory manipulated pH on corticosterone levels in A. jeffersonianum, A. maculatum, R. sylvatica, and H. versicolor. There was a significant correlation between increased baseline corticosterone levels to low pH in all four species. Prey consumption (in both Ambystoma species) and survival (in A. jeffersonianum, A. maculatum, and R. sylvatica) were also negatively correlated to low pH. Chapter V examined the effects of increased conductivity on corticosterone levels in A. jeffersonianum, R. sylvatica, and H. versicolor. Increased conductivity exposure significantly correlated to increased baseline corticosterone levels in A. jeffersonianum and R. sylvatica. Prey consumption in A. jeffersonianum was also negatively correlated to increased conductivity. My dissertation shows that abiotic factors, such as pH and conductivity, can influence corticosterone levels in larval amphibians. These results suggest that corticosterone levels in larval amphibians may be a suitable biomarker reflective of altered freshwater habitat quality. However, my results also suggest that one should use a high degree of caution when using corticosterone levels in larval amphibians as a means to infer the health status of a population.
Ph. D.

Mestrado em Engenharia Agronómica - Protecção das Plantas - Instituto Superior de Agronomia - UL
O sobreiro (Quercus suber L.) é uma árvore autóctone do Mediterrâneo que desempenha um papel-chave no seu ecossistema. A relevância económica da espécie, aliada ao seu declínio populacional, tanto devido a ameaças biológicas (ex.: P. cinnamomi), como abióticas (ex.: calor e secura), levaram a um acrescido interesse nesta árvore. Num projecto anterior (SuberStress) do laboratório de acolhimento foram selecionados 10 genes diferencialmente expressos em diferentes stresses aplicados separadamente, entre eles QsUNK1, de função desconhecida. Para a caracterização de QsUNK1 foram inicialmente determinadas as sequências genómica (2264pb com três intrões de 819, 486 e 506pb) e do promotor (identificados 1220pb). Por “Yeast-1-Hybrid” (Y1H) identificaram-se cinco potenciais factores de transcrição (TFs) por rastreio de uma biblioteca de cDNAs de stresse abiótico (calor e secura) com “baits” de levedura contendo parte do promotor (-797 a -260pb). Para a validação funcional deste gene utilizaram-se duas linhas descritas como mutantes no gene homólogo em Arabidopsis (AT3G55646). Plântulas de Arabidopsis (selvagens e mutantes) e plantas de sobreiro foram sujeitas a ensaios de calor, por choque ou aclimatação. Por análise da expressão em sobreiro, verificou-se indução de QsUNK1 em ambas as condições de stresse. Em Arabidopsis, AtUNK1 não revelou variações de expressão claras, pelo que o gene não está a ser silenciado. Igualmente não se confirmou o “knockout” nas duas linhas analisadas. Para validar a ligação TF-promotor e efectuar estudos funcionais, foram desenvolvidas linhas embriogénicas de sobreiro
N/A

Wheat is the most important grain crop grown in our country providing greatest part of the daily nutritional requirement. Abiotic factors including salinity, drought, cold and heat stresses affect quality and yield of wheat varieties used for the production of both bread and pasta flour. NAC proteins form one of the widest families of plant specific transcription factors. Members of this family are related with development, defense and abiotic stress responses. TaNAC69-1 and TtNAM-B2 genes were isolated from T.aestivum and T.turgidum, respectively. Then they were cloned into different monocot and dicot expression vectors to be used for further wheat and tobacco genetic transformation studies. To understand effects of salinity, drought, cold and heat stresses on expression profiles of TaNAC69-1 and TtNAM-B2 genes, quantitative real time PCR was performed. The time series expression profiles of TaNAC69-1 show that it was signi

Many stress situations limit plant growth, resulting in crop production difficulties. Population growth, limited availability and over-utilization of arable land, and intolerant crop species have resulted in tremendous strain being placed on agriculturalists to produce enough to sustain the world's population. An understanding of the principles involved in plant resistance to environmental stress will enable scientists to harness these mechanisms to create stress-tolerant crop species, thus increasing crop production, and enabling the farming of previously unproductive land. This research project uses computational and bioinformatics techniques to explore the promoter regions of genes, encoding proteins that are up- or down-regulated in response to specific abiotic stresses, with the aim of identifying common patterns in the cis-elements governing the regulation of these abiotic stress responsive genes. An initial dataset of fifty known genes encoding for proteins reported to be up- or down-regulated in response to plant stresses that result in water-deficit at the cellular level viz. drought, low temperature, and salinity, were identified, and a postgreSQL database created to store relevant information pertaining to these genes and the proteins encoded by them. The genomic DNA was obtained where possible, and the promoter and intron regions identified. The Neural Network Promoter Prediction (NNPP) software package was used to predict the transcription start signal (TSS) and the promoter searching software tool, TESS (Transcription Element Search Software) used to identify known and user-defined cis-elements within the promoter regions of these genes. Currently available promoter prediction software analysis tools are reported to predict one promoter per kilobase of DNA, whilst functional promoters are thought to only occur one in 30-40 kilobases, which indicates that a large perccntage of predictions are likely to be false positives (pedersen et. al., 1999). NNPP was chosen as it was rated as the highest performing promoter prediction software tool by Fickett and Hatzigeorgiou (1997) in a thorough review of eukaryotic promoter prediction algorithms, however results were less than promising as very few predicted TSS were identified in the area 50 bps up- and downstream of the gene start site, where biologically functional TSSs are known to occur (Reese, 2000; Fickett and Hatzigeorgiou, 1997). TESS results seemed to support the hypothesis that drought, low-temperature and high salinity plant stress response proteins have similar as-elements in their promoter regions, and suggested links to various other gene regulation mechanisms viz. gibberellin-, light-, auxin- and development-regulated gene expression, highlighting the vast complexity of plant stress response processes. Although far from conclusive, results provide a valuable basis for future comparative promoter studies that will attempt to deduce possible common transcriptional initiation of abiotic stress response genes.
KMBT_363
Adobe Acrobat 9.54 Paper Capture Plug-in

Dissertação para obtenção do Grau de Mestre em Engenharia do Ambiente Perfil de Gestão de Sistemas Ambientais
Plants are emitting chemical-signals to the atmosphere in response to stress factors - Volatile Organic Compounds (VOCs). VOCs have higher influence on atmosphere chemistry: they are acting as photochemical precursors in tropospheric ozone formation. Present work studies VOCs emission released by rice (Oryza sativa L cv. Aríete) cycle in paddy fields, in aleatory schemes with three replicates, in two separate soil plots with different textures (silty clay and loamy sand), studying open field conditions and open top chambers (OTCs) under influence of treatments with induced abiotic stress (increase temperature and simultaneously temperature and CO2 atmospheric concentration enhancement). VOCs were extracted from plant by solid phase micro extraction (SPME) and stem distillation extraction (SDE), and analyzed by gas chromatography coupled to mass spectrometry (GC/MS) using two GC capillary columns with different polarities, one non-polar (DB-5) and other polar (DB-WAX). A total of 33 VOCs using a non-polar column and 22 VOCs using a polar column, in both set of results were identified the three main classes of compounds: green leaf volatiles (GLV), monoterpenes and sesquiterpenes. Between rice cycle VOCs vary their trend and on vegetative stage were observed more VOCs, followed by ripening and lesser on reproductive. Silty clay soil demonstrated higher amount of VOCs released if compared with loamy sand texture. Between OTCs, more compounds were released by increasing temperature than simultaneously temperature and CO2. In Intergovernmental Panel for Climate Change (IPCC) scenarios with emergent trend of increasing temperature and CO2 atmospheric concentration, two effects are inherent to rice VOCs emission, one negative with higher emission related with temperature and other positive with less emission associated CO2. Field data measurements addictions in air quality models will help achievements of realistic previsions and better understand the effect of climate change in air quality on a global scale.
Portuguese Foundation for Science and Technology; FCT-UNL and partners from INIAV and UTAD, on a project named PTDC/AGR-AAM/102529/2008

Stress responsive genes play a key role in shaping the manner in which plants process and respond to environmental stress. Their gene products are linked to DNA transcription and its consequent translation into a response product. However, whilst these genes play a significant role in manufacturing responses to stressful stimuli, transcription factors coordinate access to these genes, specifically by accessing a gene&rsquo
s promoter region which houses transcription factor binding sites. Here transcriptional elements play a key role in mediating responses to environmental stress where each transcription factor binding site may constitute a potential response to a stress signal. Arabidopsis thaliana, a model organism, can be used to identify the mechanism of how transcription factors shape a plant&rsquo
s survival in a stressful environment. Whilst there are numerous plant stress research groups, globally there is a shortage of publicly available stress responsive gene databases. In addition a number of previous databases such as the Generation Challenge Programme&rsquo
s comparative plant stressresponsive gene catalogue, Stresslink and DRASTIC have become defunct whilst others have stagnated. There is currently a single Arabidopsis thaliana stress response database called STIFDB which was launched in 2008 and only covers abiotic stresses as handled by major abiotic stress responsive transcription factor families. Its data was sourced from microarray expression databases, contains numerous omissions as well as numerous erroneous entries and has not been updated since its inception.The Dragon Arabidopsis Stress Transcription Factor database (DASTF) was developed in response to the current lack of stress response gene resources. A total of 2333 entries were downloaded from SWISSPROT, manually curated and imported into DASTF. The entries represent 424 transcription factor families. Each entry has a corresponding SWISSPROT, ENTREZ GENBANK and TAIR accession number. The 5&rsquo
untranslated regions (UTR) of 417 families were scanned against TRANSFAC&rsquo
s binding site catalogue to identify binding sites. The relational database consists of two tables, namely a transcription factor table and a transcription factor family table called DASTF_TF and TF_Family respectively. Using a two-tier client-server architecture, a webserver was built with PHP, APACHE and MYSQL and the data was loaded into these tables with a PYTHON script. The DASTF database contains 60 entries which correspond to biotic stress and 167 correspond to abiotic stress while 2106 respond to biotic and/or abiotic stress. Users can search the database using text, family, chromosome and stress type search options. Online tools have been integrated into the DASTF 
database, such as HMMER, CLUSTALW, BLAST and HYDROCALCULATOR. User&rsquo
s can upload sequences to identify which transcription factor family their sequences belong to by using HMMER. The website can be accessed at http://apps.sanbi.ac.za/dastf/ and two updates per year are envisaged.

Le blé tendre, Triticum aestivum, est une des céréales les plus cultivées dans le monde. Le changement climatique qui se développe actuellement contraint fortement les cultures et altère leur rendement. La compréhension des mécanismes de réponse du blé tendre aux stress abiotiques est donc une problématique d’actualité. Plusieurs grandes familles de facteurs de transcription, dont la famille NAC,interviennent dans le développement de la plante et dans sa réponse aux stress environnementaux. Cette thèse, structurée en 3 volets, est ciblée sur l’étude de la famille NAC chez le blé tendre : les TaNAC. Dans un premier temps, nous avons étudié la structuration génomique et phylogénétique des 488 membres de la famille TaNAC, recensés à partir de la base de données la plus récente du blé tendre.Nous avons aussi étudié l’histoire évolutive de cette famille, qui a été marquée par des événements de duplication et de rétroposition. Enfin, une analyse de sa diversité allélique a permis d’identifier des gènes qui présentent des SNP montrant une forte association avec des paramètres d’accumulation des protéines de réserve dans le grain. Le deuxième chapitre de cette thèse a porté sur l’étude de l’expression de ces 488 gènes TaNAC dans plusieurs organes et en réponse aux stress thermique et sécheresse. Une analyse globale a été réalisée à partir de données bio-informatiques, suivie d’une étude in planta de l’expression d’une sélection de 23 gènes. Les profils d’expression obtenus ont révélé l’existence de 4 gènes TaNAC, encore jamais décrits dans la littérature et qui interviennent dans le développement du grain de blé tendre mais aussi dans sa réponse adaptative à plusieurs stress abiotiques. Le troisième volet de cette thèse a donc porté sur la caractérisation génétique, moléculaire et physiologique de ces 4 facteurs de transcription TaNAC. Ils appartiennent à un clade rassemblant des séquences présentant des similitudes génomique et structurale. De plus, ils sont localisés dans le noyau et leurs profils d’expression sont similaires, avec toutefois un niveau variable entre gènes et entre homéologues pour chaque gène. En réponse à un stress thermique modéré, ce profil d’expression est accéléré au cours du développement du grain ; le stade 120°Cj étant le stade clé qui montre la plus grande différence d’expression de ces gènes entre les conditions contrôle et stressée. Pour des raisons techniques, la production de plantes transgéniques sur- et sous-exprimant ces gènes n’a pas permis de valider l’implication de ces 4 TaNAC dans le développement du grain et en réponse à la température. Une analyse de génétique d’association a toutefois permis de mettre en évidence un lien entre des marqueurs moléculaires situés dans ces gènes et l’accumulation des protéines de réserve.Globalement, les résultats obtenus ont montré que des membres de la famille TaNAC sont impliqués dans le développement du blé tendre et dans sa réponse aux stress abiotiques. Plus particulièrement, 4 facteurs de transcription TaNAC semblent jouer un rôle clé dans l’accumulation des protéines dans le grain en réponse à un stress thermique modéré
Bread wheat, Triticum aestivum, is one of the most cultivated cereal in the world. The climate change that is currently developing strongly constrains crops and impairs their yield. Understanding the wheat response mechanisms to abiotic stresses is therefore a current issue. Several major families of transcription factors, including the NAC family, are involved in the plant development and its response to environmental stresses. This thesis, structured in three parts, is focused on the study of the NAC family in bread wheat (TaNAC).First, we studied the genomic and phylogenetic structure of the 488 members of the TaNAC family identified from the latest database of bread wheat. We also studied the evolutionary history of this family, which was marked by duplication and retroposition events. Finally, an analysis of its allelic diversity allows us to identify genes with SNP showing a strong association with storage protein accumulation parameters in the grain. In a second part, we studied the expression of these 488 TaNAC genes in several organs and in response to heat and drought. An overall analysis was performed using bioinformatic data, followed by an in planta study of the expression of a selection of 23 genes. The expression profiles revealed that four TaNAC genes, never described in the literature, are involved in the wheat grain development but also in its adaptive response to several abiotic stresses. In a third part, we focused on the genetic, molecular and physiological characterization of these four TaNAC transcription factors. They belong to a clade gathering sequences with genomic and structural similarities. Moreover, they are localized in the nucleus and their expression profiles are similar, with a variable level between genes and between homeologs for each gene. In response to moderate heat stress, this expression profile is accelerated during grain development and a key stage at 120°Cj was identified, it shows the greatest difference in genes expression level between control and stressed conditions. For technical reasons, the production of transgenic plants over- and under-expressing these genes did not validate the involvement of these 4 TaNAC in grain development and in its temperature response. An association genetic analysis, however, showed a link between molecular markers located in these genes and the storage proteins accumulation. Overall, the results showed that members of the TaNAC family are involved in the bread wheat development and its response to abiotic stresses. In particular, four TaNAC transcription factors appear to play a key role in grain protein accumulation in response to a moderate heat stress

Du fait de sa croissance exceptionnelle combinée aux propriétés supérieures de son bois, l'Eucalyptus est devenu le feuillu le plus planté au monde et s'est imposé comme source de biomasse pour la production de papier et de biocarburants de seconde génération. Le bois est composé de parois secondaires lignifiées et sa formation est finement régulée par un réseau complexe, et globalement mal connu, de facteurs de transcription (FT). Les parois secondaires sont composées de 80% de polysaccharides, ciblés pour la plupart des bioproduits à haute valeur ajoutée, tandis que la lignine (20%) est responsable de la récalcitrance de la biomasse à la dégradation enzymatique mais augmente le potentiel énergétique du bois par combustion. Malgré son adaptabilité remarquable à différents sols et climats, la croissance de l'Eucalyptus varie fortement suivant ces facteurs. L'Eucalyptus est largement planté sur des sols lessivés dans les régions tropicales et subtropicales où les plantations industrielles font face à des épisodes de sécheresse de plus en plus fréquents, en combinaison avec des forts manques de nutriments, nécessitant de gros apports en fertilisants. Dans les région tempérées telles que l'Europe du Nord, la principale limitation à l'implantation de cet arbre dépourvu d'endodormance, est l'exposition au froid. Ces contraintes abiotiques sont aggravées par le changement climatique et leur impact sur la formation du bois et sa qualité restent peu documentés. Quelques données suggèrent que ces stress affectent le dépôt de la paroi secondaire ainsi que la structure du xylème. Cependant, ces résultats sont très hétérogènes entre différentes espèces et principalement focalisés sur des tissus différents du bois. La sélection de clones adaptés et le développement de pratiques culturales plus viables, sont essentiels pour améliorer la productivité et la qualité du bois, ce qui requiert une meilleure compréhension de la réponse des arbres au froid et au manque d'eau en interaction avec la nutrition. Dans le but de décrypter les régulations induites par le froid dans la différenciation du xylème, nous avons effectué une approche ciblée sur des Eucalyptus acclimatés au froid. Des analyses de biochimie, d'histochimie et de transcriptomique, ont révélé que le froid déclenche un dépôt de paroi secondaire précoce dans les cellules du xylème en développement, caractérisé par un fort dépôt de lignine. En parallèle, pour caractériser l'effet du manque d'eau combiné à différents régimes nutritifs, sur la formation et la qualité du bois, nous avons tiré profit d'un dispositif expérimental mis en place au champ avec un clone commercial d'Eucalyptus, soumis à une exclusion de pluie combinée à une fertilisation au potassium. Nous avons combiné des analyses globales du transcriptome et du métabolome, avec l'analyse des propriétés structurales et biochimiques du bois. L'approche intégrative de ces jeux de données a révélé que la fertilisation au potassium induit une répression de la biosynthèse de la paroi secondaire ainsi qu'une régulation de l'activité cambiale et la modification dans les propriétés du bois, avec une forte interaction avec l'exclusion d'eau. Ces deux approches ont permis l'identification de différents FT non caractérisés qui constituent des candidats prometteurs dans le contrôle de l'activité cambiale et du dépôt de paroi secondaire chez un ligneux. Leur caractérisation fonctionnelle chez le peuplier et l'Eucalyptus a révélé un nouveau régulateur clé de la biosynthèse de paroi secondaire, et plusieurs facteurs MYB potentiellement impliqués dans la balance entre formation de la paroi secondaire et croissance
Due to its outstanding growth combined to superior wood properties, Eucalyptus genus has become the most planted hardwood on earth and emerged as the most appealing sources of renewable biomass feedstock for paper and second-generation biofuels. Wood is composed of lignified secondary cell walls (SCWs) and its formation is tightly regulated by a complex, partially unknown, transcription factors (TFs) network. SCWs are composed by 80% of polysaccharides targeted for most of value-added bioproducts, whereas lignin (20%) is responsible for biomass recalcitrance to enzymatic degradation but increase wood energetic potential for combustion. Despite its remarkable adaptability to various soils and climate environment Eucalyptus growth varies strongly according to these factors. Eucalyptus is extensively grown in highly weathered soils in tropical and subtropical regions where plantations are facing more frequent drought episodes in combination to nutrient starvation, requiring high amounts of expensive fertilizers. In temperate regions such as North of Europe, the main limitation for the expansion of this non-dormant tree is cold exposure, which reduces dramatically its growth. The effects of these stresses are emphasized in the actual context of climate change which induces sharp contrasting periods, and their impacts on wood formation and quality remain unknown. Scarce data from literature suggest that these stresses affect secondary cell wall (SCW) deposition as well as xylem cell patterning. However these results are highly heterogeneous among different species and mainly focused on non-woody tissues. The selection of adapted clones and the development of more sustainable cultural practices are crucial to improve wood productivity and quality, which require a better understanding of tree response to cold and water stress in interaction with nutrition. In order to unravel the regulation of xylem differentiation by low temperature, we performed a targeted approach on cold-acclimated Eucalyptus trees. By biochemical, histochemical and transcriptomic analyses, we revealed that low temperature trigger a precocious SCW deposition in developing xylem cells, characterized by a strong lignin deposition. In parallel, we aimed to characterize the effect of water stress combined to different mineral nutrition regimes, on wood formation and quality. To this end, we took advantage of an experimental design set up on field with a highly productive Eucalyptus commercial clone submitted to both rainfall exclusion combined to potassium fertilization. We combined large scale analyses of transcriptome and metabolome, with wood structural and biochemical properties analyses. The integrative approach with these datasets revealed that potassium fertilization induces a repression of SCW biosynthesis, together with regulation of cambial activity and modifications in wood properties, with a strong interaction with water exclusion. Both approaches allowed to point out several uncharacterized yet TFs which are highly promising candidates in the control of cambial activity and SCW deposition in a woody perennial. Characterization of their function in poplar and Eucalyptus revealed a new key regulator of SCW biosynthesis in wood, and several MYB TFs potentially involved in the trade-off between SCW biosynthesis and growth

Submitted by Gabriela Lopes (gmachadolopesufpel@gmail.com) on 2016-09-14T17:13:51Z No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Mariana_Kruger.pdf: 976408 bytes, checksum: 6bfde814984b8524fba2bd3708136696 (MD5)
Approved for entry into archive by Aline Batista (alinehb.ufpel@gmail.com) on 2016-09-14T18:28:25Z (GMT) No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Mariana_Kruger.pdf: 976408 bytes, checksum: 6bfde814984b8524fba2bd3708136696 (MD5)
Made available in DSpace on 2016-09-14T18:28:25Z (GMT). No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Mariana_Kruger.pdf: 976408 bytes, checksum: 6bfde814984b8524fba2bd3708136696 (MD5) Previous issue date: 2013-12-19
Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES
O arroz (Oryza sativa L.) é uma cultura que possui grande importância alimentar e econômica, sendo largamente cultivado em ecossistemas de terras baixas, o qual apresenta condições de drenagem deficiente. Nessas condições, o baixo potencial redox do meio leva a redução do ferro presente na solução do solo o qual torna-se prontamente disponível, podendo se tornar tóxico à planta quando absorvido em excesso. A toxidez por ferro é um dos mais importantes estresses abióticos a limitar a produção de arroz irrigado em nível mundial. Em áreas de cultivo de arroz do Rio Grande do Sul, o ferro tem causado problemas de toxidez e o uso de cultivares tolerantes torna-se uma importante estratégia para solucionar o problema, sendo necessário a identificação de genes frente aos diferentes mecanismos que conferem essa tolerância às plantas para aplicação em programas de melhoramento. Estudos recentes descobriram a importância de alguns fatores de transcrição responsivos ao etileno (ERFs) na resposta vegetal a diferentes estresses. Dessa forma o objetivo deste trabalho foi avaliar o perfil de expressão diferencial de 32 ERFs em arroz submetidos a condições de estresse por ferro, em três cultivares, Nipponbare, Epagri 108 (tolerante à toxidez por ferro) e BR-IRGA 409 (sensível à toxidez por ferro) e identificar elementos cis na região promotora de genes ERFs em arroz. Os resultados obtidos relatam que em arroz, a maioria dos membros da família ERF demonstram perfil diferencial nas cultivares Nipponbare, BR -IRGA 409 e Epagri 108, em condições de excesso de ferro. Os ERFs expressos na cultivar tolerante (Epagri 108) podem ser potencialmente importantes na indução da tolerância, enquanto que os que foram altamente induzidos na cultivar sensível (BR – IRGA 409) pertencem a um mecanismo de resposta o qual não é eficiente. Análises de elementos cis demonstram que a regulação da expressão de todos os ERFs analisados se dá de forma complexa e que existe uma associação entre a presença de determinado elemento cis com o perfil de expressão.
Rice (Oryza sativa L.) is a crop of high nourishment and financial importance, largely grown in flat and flooded ecosystems which present deficient drainage conditions. Under such conditions, the low redox potential of the environment leads to the reduction of iron present in the soil solution, which becomes readily available and can become toxic to the plant when absorbed in excess. Iron toxicity is one of the most important types of abiotic stress, limiting the production of irrigated rice worldwide. In rice growth areas of Rio Grande do Sul, Brazil, iron has caused toxicity problems and the use of tolerant cultivars becomes an important strategy in solving the problem, where the identification of genes is necessary in face of the different mechanisms which confer this tolerance to the plants for application in improvement programs. Recent studies discovered the importance of some transcription factors responsive to ethylene (ERFs) in vegetable response to different kinds of stress. Thus, the aim of this study was to evaluate the differential expression profile of 32 ERFs in rice subjected to iron stress conditions in three cultivars; Nipponbare, Epagri 108 (tolerant to iron toxicity) and Br-Irga 409 (sensitive to iron toxicity) and to identify cis elements in the ERF gene promotion region in rice. The results obtained demonstrate that in rice, most members of the ERF family showed differential profiles in the cultivars Npponbare, Epagri 108 and Br-Irga 409, under iron stress conditions. ERFs expressed in the tolerant cultivar (Epagri 108) may be potentially important in the induction of tolerance while those which were more highly induced in the sensitive cultivar (Br-IRGA 409) belong to a response mechanism which is not efficient. Analyses of cis elements demonstrate that the regulation of expression of all analyzed ERFs takes place in complex form and that there exists an association between the presence of a specific cis element and the expression profile.

Doutoramento em Biologia
Rapid climatic changes are taking place in Arctic, subarctic and cold temperate regions, where predictions point to an increase in freeze-thaw events, changes in precipitation, evaporation and salinity patterns. Climate change may therefore result in large impacts in ecosystem functioning and dynamics, especially in the presence of contaminants due to intense anthropogenic activities. Even though multiple stress approaches have received increasing interest in the last decades, the number of such studies is limited. In particular, knowledge on the effect of freezethaw events and salinity fluctuations on ecotoxicology of soil invertebrates is lacking, especially important when considering supralittoral species. Therefore, the aim of this thesis was to investigate the effects of low temperature and salinity fluctuations, singly and in combination with contaminants, in the freeze-tolerant and euryhaline enchytraeid Enchytraeus albidus. The assessment of population level endpoints (survival and reproduction), along with physiological and biochemical parameters such as levels of cryoprotectants, ice/water content, oxidative stress biomarkers, cellular energy allocation, and tissue concentration of chemicals (when applied), provided new and valuable knowledge on the effects of selected physical and chemical stressors in E. albidus, and allowed the understanding of adjustments in the primary response mechanisms that enable worms to maintain homeostasis and survival in harsh environments such as polar and temperate-cold regions. The presence of moderate levels of salinity significantly increased freeze-tolerance (mainly evaluated as survival, cryoprotection and ice fraction) and reproduction of E. albidus. Moreover, it contributed to the readjustments of cryoprotectant levels, restoration of antioxidant levels and changed singnificantly the effect and uptake of chemicals (copper cadmium, carbendazim and 4-nonylphenol). Temperature fluctuations (simulated as daily freeze-thaw cycles, between -2ºC and -4ºC) caused substancial negative effect on survival of worms previsouly exposed to non-lethal concentrations of 4-nonylphenol, as compared with constant freezing (-4ºC) and control temperature (2ºC). The decrease in cryoprotectants, increase in energy consumption and the highest concentration of 4-nonylphenol in the tissues have highlighted the high energy requirements and level of toxicity experienced by worms exposed to the combined effect of contaminants and freezing-thawing events. The findings reported on this thesis demonstrate that natural (physical) and chemical stressors, singly or in combination, may alter the dynamics of E. albidus, affecting not only their survival and reproduction (and consequent presence/distribution) but also their physiological and biochemical adaptations. These alterations may lead to severe consequences for the functioning of the ecosystems along the Arctic, subarctic and cold temperate regions, where they play an important role for decomposition of dead organic matter. This thesis provides a scientific basis for improving the setting of safety factors for natural soil ecosystems, and to underline the integration of similar investigations in ecotoxicology, and eventually in risk assessment of contaminants.
As alterações climáticas estão a atingir rapidamente as regiões do Ártico, SubÁrtico e as regiões temperadas, apontando as previsões para um aumento de eventos de congelamento-descongelamento, bem como mudanças nos padrões de precipitação, evaporação e de salinidade. Estas alterações climáticas poderão resultar em impactos francamente negativos no funcionamento e dinâmica de ecossistemas, especialmente quando associados à presença de contaminantes resultantes da intensa atividade antropogénica. Embora a incorporação de stressores múltiplos em estudos de ecotoxicidade tenha recebido um crescente interesse pela comunidade científica, o seu número é ainda reduzido. Particularizando, o conhecimento dos efeitos de eventos de congelamento-descongelamento e de flutuações de salinidade permanecem desconhecidos, especialmente quando se consideram espécies supra-litorais. Neste contexto, o objetivo geral da presente tese consistiu em investigar os efeitos das flutuações de temperaturas e salinidade, individualmente ou em combinação com contaminantes, no enquitraídeo tolerante ao frio e eurialino - o Enchytraeus albidus. A avaliação de parâmetros populacionais (sobrevivência, reprodução e bioacumulação), fisiológicos (níveis de crioprotetores, conteúdo em gelo / água, temperatura de fusão e sobrecongelamento) e bioquímicos (biomarcadores de stress oxidativo, alocação de energia celular) permitiu compilar novas e valiosas informações sobre os efeitos dos stressores físicos e químicos selecionados no enquitraídeo e compreender quais os reajustes nos mecanismos de resposta primários que lhes permitem manter a homeostasia e sobrevivência em ambientes inóspitos como as regiões Polares e temperadas-frias. A presença de níveis moderados de salinidade aumentou significativamente a tolerância a temperaturas congelantes (essencialmente avaliada como sobrevivência, crioprotecção e fracção de gelo extracelular) e a reprodução do E. albidus. Além disso, contribuiu para a regulação de crioprotectores, restauração dos níveis de antioxidantes nestes organismos e alterou significativamente o efeito e a incorporação/absorção de substâncias químicas (cádmio, cobre carbendazim e 4-nonilfenol). As flutuações de temperatura (simuladas como ciclos diários de congelamento-descongelamento, com temperaturas entre 2ºC e -4ºC) causaram um efeito substancialmente negativo na sobrevivência de organismos previamente expostos a concentrações não letais de 4-nonilfenol, quando comparados com organismos expostos a uma temperatura congelante constante (-4ºC) ou à temperatura controlo (2ºC). A diminuição na crioproteção, o aumento no consumo de energia e a maior concentração de 4-nonilfenol nos tecidos vieram sublinhar o elevado gasto energético e o nível de toxicidade sofrido pelos organismos expostos à combinação de contaminantes e eventos de congelamento e descongelamento. Os resultados apresentados nesta tese demonstram, assim, que a presença de stressores naturais (físicos) e químicos, isoladamente ou em combinação, podem alterar a dinâmica do E. albidus, afetando não só a sua sobrevivência e reprodução (e consequente presença / distribuição), mas também as suas adaptações fisiológicas e bioquímicas. Essas alterações podem levar a consequências graves para o funcionamento dos ecossistemas do Ártico, subÁrtico e regiões temperadas-frias, uma vez que estes organismos desempenham um papel importante para a decomposição de matéria orgânica morta. Esta tese fornece ainda uma base científica para melhorar a atribuição de coeficientes de segurança para os ecossistemas naturais do solo, alertando para a integração de investigações semelhantes em ecotoxicologia, e, eventualmente, para a avaliação de risco ecológico de contaminantes.

Plants respond to environmental stress by altering their gene expression. Under stress conditions some genes are activated and some genes are repressed. Even though a lot of work has been done to understand mechanisms of gene activation under abiotic stress very little information is available on how stress responsive genes are kept repressed under normal growth conditions. Recent work has revealed that plants use transcriptional repression as common mechanism of gene repression. Transcriptional repression is achieved by recruitment co-repressor complexes to the target genes. Recent studies have revealed that the co-repressor LUH complexes with SLK1 and SLK2 to silence Arabidopsis thaliana stress responsive genes. However, the transcription factors involved in the recruitment of this complex to its target genes are not known. In this study, we identified SLK2INT1, as a novel transcription factor that is involved in silencing of select Arabidopsis thaliana stress responsive genes by recruiting the LUH-SLK2 complex.

Thesis (Ph.D.)--Kent State University, 2009.
Title from PDF t.p. (viewed Nov. 16, 2009). Advisor: Mark Kershner. Keywords: crayfish; stream; predation; current velocity; water depth; grain size; resource competition. Includes bibliographical references.

Le développement post-natal est une étape cruciale pour le reste de la vie d’un individu car c’est à ce moment que sont finalisées et maturées les fonctions physiologiques et comportementales individuelles et que la morphologie finale est acquise. Toute contrainte ou tout stress perçu par la progéniture pendant cette période peut avoir des conséquences morphologiques, physiologiques et/ou comportementales non négligeables à court mais aussi à long terme. In fine, c’est alors sa fitness qui peut être affectée par la qualité des conditions de développement subies. Ce doctorat vise à améliorer notre compréhension de l’importance des conditions environnementales abiotiques (climat, dérangement anthropique et exposition à un pesticide) et des soins parentaux rencontrés durant la croissance sur la qualité des poussins produits. Grâce à l’étude des marqueurs du stress et de l’allostasie (réponse au stress et longueur des télomères) chez des poussins d’Albatros à sourcils noirs et de Pétrel des neiges, nous avons pu démontrer, dans un premier temps, qu’à court terme, la qualité des soins parentaux – approximée par l’âge des individus reproducteurs – était un facteur très important pour la mise en place du phénotype du poussin. Dans un second temps, la manipulation des taux de corticostérone durant le développement chez des poussins de Moineau domestique (mimant une contrainte développementale) semble impacter à long terme les performances individuelles. En effet, à l’âge adulte, une réduction du métabolisme et des dépenses énergétiques, une réduction de l’attractivité sexuelle et une augmentation de l’investissement parental ont été mis en évidence. Au vu des résultats obtenus dans le cadre de ce doctorat, nous discutons de l’influence des conditions de développement sur la fitness des individus en s’appuyant sur les hypothèses évolutives détaillées dans la littérature scientifique
Post-natal development is a crucial step for the rest of life. Indeed, individual physiological and behavioral functions are set-up and matured during that life-stage and final morphology is acquired at that time. Any stress or constraint perceived by the offspring during this period can have significant morphological, physiological and/or behavioral consequences in the short but also in the long term. In fine, an individual’s fitness can be affected by the quality of its developmental conditions. This PhD aims to improve our understanding of the impact of abiotic developmental conditions (climate, human disturbance and exposure to a pesticide) and parental care on the quality of the produced chicks. Firstly, through the study of markers of stress and allostasis (stress response and telomere length) in Black-browed albatross and Snow petrel’s chicks, we demonstrated that in the short term, the quality of parental care - approximated by the age of the breeding individuals - was a major factor determining a chick’s phenotype. Secondly, the manipulation of corticosterone levels during development in House sparrow chicks (mimicking a developmental constraint) seems to have long-term impacts on individual performance. More precisely, in adulthood, I found that this experimental manipulation of developmental conditions was associated with a reduced metabolism, a reduced sexual attractiveness, and an increased parental investment during adulthood. Using the results obtained during this PhD, I discuss the influence of developmental conditions on individual fitness in an evolutionary context

Plants possess highly sensitive perception systems by which they recognize signal compounds originating from microbes. These molecular cues play an important role in both symbiotic and pathogenic relationships. Establishment of the soybean (Glycine max)-Bradyrhizobium symbiosis is orchestrated by specific signal molecules exchanged between appropriate plant and microbe partners: flavonoids as plant-to-bacteria signals, and Nod factor as bacteria-to-plant signals. How this signaling process interacts with stress conditions (abiotic and biotic) is the subject of this thesis. The abiotic stresses were suboptimal growth temperature, low pH, and salinity. Suboptimal growth temperatures affected the ability of the microsymbiont, Bradyrhizobim japonicum, to perceive nod gene inducers (genistein) and produce Nod factor. Nod Bj-V (C18:1, MeFuc) production by B. japonicum strains 523C and USDA110 was strongly affected by suboptimal growth temperature. Nod factor production declined with temperature, from 28 to 15°C. Strain USDA110 was more affected by decreased temperature than strain 532C. Decreased Nod factor production at low temperature was due to both decreased bacterial growth and lower production efficiency (Nod factor per cell). When a 1:1 mixture of Nod factor Nod Bj-V (C18:1, MeFuc) and Nod Bj-V (Ac, C16:0, MeFuc) was applied to soybean roots, root hair deformation increased as Nod factor concentration increased under stressfully low temperature and low pH conditions. High salinity stress strongly reduced the root hair deformation caused by Nod factor, and increasing the concentrations of added Nod factor did not over come this. Exogenous application of Nod Bj-V (C18:1, MeFuc), from strain 532C, to soybean root systems under two root zone temperatures (RZTs---17 and 25°C) reduced the progression of disease (powdery mildew---Microsphaera difussa) development on soybean leaves; this effect increased with Nod factor concentration and was gr

L'Eucalyptus, feuillu le plus planté dans le monde, est fortement exposé au froid en raison de l'absence de dormance. Les gènes DREB sont connus comme étant les principaux régulateurs de la réponse aux stress abiotiques. Un nombre élevé de gènes DREB1/CBF (C-Repeat Factor) a été identifié chez Eucalyptus grandis. Le but de l'étude est de mieux comprendre le rôle de la voie DREB chez Eucalyptus pour le contrôle de la tolérance au stress, du développement et de la formation du bois. La présente étude a permis une annotation des gènes CBF et DREB2 dans le cadre d'un projet de sequençage partiel du génome d'E. gunnii. Une analyse complète de l'expression des genes par qRT-PCR a été réalisée sur les différents organes des deux espèces d'Eucalyptus après les traitements au stress. L'existence d'une copie de CBF supplémentaire dans le génome E. gunnii par rapport à E. grandis suggère que ce groupe est encore en évolution contrairement au groupe DREB2. Un nombre élevé de transcrits CBF chez E. gunnii, tolérant au froid, et forte une vitesse d'induction ce ces facteurs chez E. grandis, à croissance rapide, suggère que les facteurs CBF sont impliqués à la fois dans la protection au stress et la limitation de croissance. Des facteurs de transcription des familles MYB, NAC, KNOX et AP2/ERF impliqués dans le contrôle de la croissance et de la formation de la paroi cellulaire ont été identifiés comme étant des gènes putatifs cibles de CBF. Ces résultats sont en accord avec le phénotype modifié de surexpresseurs CBF. Les deux approches suggèrent un rôle central de la voie de DREB dans le compromis entre la croissance et la résistance au stress chez cette espèce ligneuse
Eucalyptus, the most widely planted hardwood in the world, is highly exposed to the cold due to the lack of dormancy. DREB (Drought Responsive Element Binding) genes are known as master regulators of abiotic stress response. A high number of the DREB1/CBF (C-Repeat Factor) genes has been annotated in Eucalyptus grandis. The aim of the study was to better understand the role of DREB pathway in Eucalyptus for the control of stress tolerance, development and wood formation. The present study provides an annotation of the CBF and DREB2 genes from a partial draft of the E. gunnii genome sequence. A comprehensive transcriptional analysis through high-throughput qRT-PCR was carried out on different organs from the two Eucalyptus species after stress treatments. An additional CBF copy in the E. gunnii genome compared to E. grandis suggests that this group is still evolving unlike the DREB2 group. The higher CBF transcript amounts in the cold tolerant E. gunnii together with higher induction rates in the fast growing E. grandis suggest that CBF factors promote both stress protection and growth limitation. In addition, transcription factors from MYB, NAC, KNOX and AP2/ERF families involved in the control of growth and cell wall formation have been identified as putative CBF target genes. These results are in agreement with the modified phenotype of CBF overexpressors. Both approaches suggest a central role of DREB pathway in the trade-off between growth and stress resistance in this woody species

ABSTRACT TRANSFORMATION OF POTATO WITH MYB4 TRANSCRIPTION FACTOR AND EVALUATION OF ABIOTIC STRESS TOLERANCE AND GENE EXPRESSION PROFILES IN TRANSGENIC PLANTS Kalemtas, Gü
lsü
m Ph.D., Department of Biology Supervisor: Prof. Dr. Hü
seyin Avni Ö
ktem February 2011, 257 pages Potato (Solanum tuberosum L. cv. Kennebec) was transformed via Agrobacterium tumefaciens (EHA105) harbouring two different binary vectors containing Oryza sativa myb4 gene, which encodes MYB4 transcription factor
under the control of CaMV35S promoter or cold inducible COR15a promoter. The transgenic plants were not growth retarded and there was no significant difference (p<
0.05) in their tuber yield compared to wild-type plants. Wild-type and transgenic plants were subjected to abiotic stresses to compare their stress tolerances. There was no significant difference in boron, freezing and drought tolerances of wild-type and transgenic lines. Two of the transgenic lines were more salt tolerant than wild-type with respect to growth parameters. Transcriptomes of wild-type and these two lines, one expressing myb4 under the control of 35S promoter and the other COR15a promoter, were analyzed to elucidate the myb4-regulated processes and downstream target genes in potato. Differentially regulated genes in transgenic lines showed that myb4 controls a large and complex transcriptional network associated with diverse cellular processes, primarily defense and rescue, metabolism and development. Genes involved in sucrose synthesis, some peroxidases and CBF3 transcription factor were up-regulated in transgenic plants upon exposure to freezing stress. This suggested that myb4 may configure freezing response in potato primarily by oxidative stress defence mechanisms, osmotic adjustment or activation of CBF3 regulated genes that may confer cold tolerance. Despite up-regulation of these stress related genes, transgenic potato was not more drought or freezing tolerant compared to WT under the tested conditions. Further experiments should be conducted to better elucidate the involvement of these genes in regulation of stress response in transgenic potato expressing myb4.

La demande mondiale de nourriture et d’energie est en constante augmentation. L’accès à de nouvelles terres arables étant limité, les agriculteurs devront produire environ 70% de plus par hectare d'ici 2050. L'optimisation et la rationalisation des systèmes de production agricole sont essentielles pour assurer la sécurité alimentaire des populations dans des conditions durables. La communauté agricole serait en mesure d'optimiser les performances des cultures en ayant accès à une meilleure compréhension des systèmes de production, notamment concernant leur potentiel et principaux facteurs limitants. Dans ce contexte, la caractérisation des conditions de croissance des cultures prenant en compte leurs impacts sur le potentiel de production et sur les pertes de rendement est importante pour identifier les marges de progrès et proposer des systèmes agricoles améliorés.Dans cette étude une approche est proposée pour caractériser l'environnement de grandes zones géographiques prenant en compte les impacts des ressources clés au développement des plantes (température, rayonnement, eau et azote) sur la performance des cultures. Le niveau de disponibilité de ces ressources a été estimé en utilisant des approches de modélisation de culture, intégrant données météorologiques et caractéristiques de sol. L'efficacité d'utilisation des ressources disponibles a été caractérisée par les pratiques culturales ainsi que la stratégie et l’environnement technique des agriculteurs. Une procédure générique d’intégration de la donnée a été établie et utilisée pour décrire la production de maïs grain en Amérique du Nord de 1991 à 2013. Le jeu de données résultant de cette intégration couvre 84% des superficies plantées en maïs, décrites par 1 558 unités spatiales élémentaires, pour un total de 28 303 scénarios indépendants. Cette méthode combine et harmonise les observations de rendement issues des statistiques agricoles avec un large ensemble de descripteurs pertinents de conditions de croissance.Un sous-ensemble de 21 facteurs limitants a été identifié par une analyse de sélection de variables pour expliquer 66% de la variabilité des pertes de rendement observées. Compte tenu du nombre de dimensions du jeu de donnée, la méthode relaxed lasso a été choisie comme un compromis intéressant entre capacité de prédiction et d’interprétation. Les descripteurs sélectionnés montrent la contribution de la gestion des cultures à la variabilité des pertes de rendement, en particulier les niveaux d'intensification de l’agriculture et l’environnement technique des agriculteurs. Le troisième facteur principal est la disponibilité en eau et la sécheresse qui en résulte. L’utilisation des connaissances en physiologie dans la conception de descripteurs a considérablement amélioré la capacité d’interprétation de la proposition et la confiance des utilisateurs finaux dans l'approche.La structure des facteurs limitant le rendement a été utilisée pour identifier les Environnements Types les plus fréquents. Deux approches ont été proposes : l’une axée uniquement sur les facteurs physiologiques et l’autre intégrant également l’identification de marchés de taille homogène. 11 Environnements Types ont été identifiés en fonction des facteurs physiologiques et 8 en combinant facteurs physiologiques et contraintes de taille de marché. Les Environnements Types basés sur les facteurs physiologiques sont très informatifs sur l'évolution historique des pratiques culturales et les changements de stratégie des agriculteurs au cours du temps. Une telle catégorisation des conditions de croissance a montré des capacités à reproduire l'expertise de terrain et à soutenir l'évaluation de génotypes. Trois cas d'utilisation ont servi à illustrer l'intérêt de l'approche pour (i) décrire l'historique environnemental d'un marché, (ii) définir des populations d’Environnements cibles (TPE) et (iii) évaluer la pertinence de l'échantillonnage environnemental de réseaux multi-lieux (MET)
The global demand for food and energy is constantly increasing. As access to spare arable land is limited, growers will have to produce about 70% more per hectare by 2050. Optimization and rationalization of the agricultural production systems is then critical to ensure food security under sustainable conditions. The agricultural community would be in a position to optimize crop performance by better understanding the cropping systems, especially regarding potential production and limiting factors. In this context, the ability to characterize crop growing conditions in regards to their impacts on potential yield and yield gap is important in order to identify margins of progress and propose improved farming systems.An approach is proposed to characterize the crop environment of large geographical areas taking into consideration the impact on crop performance of the key resources for plant development (temperature, solar radiation, water and nitrogen). The level of availability of each resource was estimated by using crop modeling approaches integrating weather data and soil characteristics as inputs. The efficiency of use of the available resources was characterized by crop practices, grower strategy and grower technical environment. A generic integration procedure was established and used to describe corn production for grain in North America from 1991 to 2013. The resulting dataset covers 84% of the total corn planted area, deciphered in 1,558 elementary spatial units, for a total of 28,303 independent scenarios. Such a method combines and harmonizes, at scale, yield observations from agricultural statistics with a large set of relevant descriptors of growing conditions.A subset of 21 main limiting factors was identified through variable-selection analysis to explain 66% of the observed yield-gap variability. The relaxed lasso method resulted in an interesting compromise between interpretability and prediction ability. The selected descriptors highlighted the contribution of crop management in yield-gap variability, especially regarding levels of crop intensification and the technical environment of growers. The third main factor is water availability and resulting drought. The integration of knowledge in crop physiology into descriptor design significantly improved the interpretability of the proposal and the confidence of end-users in the approach.Yield-gap patterns were used to identify the set of most frequent Environment Types. Two approaches were proposed: one focused on crop physiology drivers and another also considered market-size homogeneity among Environment Types. This resulted in 11 Environment Types based on crop physiology drivers and 8 Environment Types when combining crop physiology drivers and market constraints. Crop-physiology Environment Types are very informative on the historical evolution of cultural practices and the changes in grower strategies over the studied period. Such categorization of growing conditions demonstrated the ability to reproduce field expertise and support genotype evaluation. Three business Use Cases were used to illustrate the interest of the approach in (i) describing the environmental history of a market, (ii) defining relevant Target Population Environments (TPE) and (iii) evaluating the environmental sampling relevance of Multi-Environment Trials (MET)

Symbiosis between bacteria and single celled eukaryotes, such as ciliates and amoebae, is a widespread phenomenon. In the past years, the number of species characterizations of symbiotic bacteria, has constantly grown. Especially members of the order Rickettsiales (Alphaproteobacteria), which comprises obligate intracellular human and animal pathogens as well as other non-pathogenic intracellular species, got into the focus. However, biodiversity of both, protists and associated bacterial symbionts, is still largely underestimated. Thus, I hypothesised that different kind of bacteria occur naturally in the ciliate Paramecium, that paramecia can serve as potential reservoir for human and animal pathogens, and that the species distribution of Paramecium and its endosymbionts shows geographical patterns. In the present thesis, I provide data on the distribution and diversity of Paramecium species and their endosymbionts isolated from freshwater and brackish habitats from North and South America. The most frequent species were Paramecium biaurelia, Paramecium caudatum and Paramecium multimicronucleatum for the North American samples as well as P. multimicronucleatum and Paramecium calkinsi for the South American samples indicating geographical differences in the species distribution of Paramecium between the two continents. Monoclonal cultures were screened for the presence of endosymbionts by fluorescence in situ hybridization (FISH) using universal and specific probes. I detected bacterial endosymbionts belonging to different taxa of Rickettsiales sensu lato in Paramecium-cultures isolated from eight different habitats. Moreover, I characterized the four novel endosymbiont species “Candidatus Fokinia solitaria”, “Candidatus Fokinia cryptica”, “Candidatus Bealeia paramacronuclearis” and “Candidatus Jurandia parameciophila”, respectively, presenting detailed descriptions. Thus, I did not only detect various bacterial symbionts in different host species from two continents, but I also identified bacteria related to human and animal pathogens. Moreover, phylogenetic information provided by the four novel bacterial species helped me to revise the taxonomic organisation of the order Rickettsiales, which I split into the two orders Rickettsiales sensu stricto and Holosporales ord. nov. The importance and severity of a symbiosis for its host can be estimated, among others, by studying the symbionts transmission modes. Most host-symbiont systems in nature show a mixed mode of transmission, even if one transmission type occurs only rarely. Moreover, evolution of exclusively vertically or horizontally transmitted symbionts is scarce. However, circumstances for horizontal transmission of many symbiont species so far described as vertically transmitted, are completely unknown and still need to be revealed. Therefore, I assumed that both, vector organisms and environmental stress, can trigger horizontal transmission. I performed different sets of infection experiments using vector organisms and environmental stress factors as driving forces to trigger horizontal transmission. In experiments focused on vector-induced transmission, I was not able to confirm vector properties of Holospora towards Caedibacter, as described towards food bacteria. However, I rarely observed vector-independent horizontal transmission of Caedibacter taeniospiralis towards symbiont-free Paramecium species. The here tested genotypes of C. taeniospiralis might still take advantage of additional horizontal transmission, even if it occurs rarely. In addition, I tested the effect of salinity and temperature stress on the transmission of the recently described bacterial symbionts “Candidatus Megaira polyxenophila” and “Ca. Bealeia paramacronuclearis” as potentially interesting representatives. The involved symbiont and host genotypes differed in their response to the induced environmental stress. Results of horizontal transmission turned out to be ambiguous. While temperature stress did not show any effect on transmission, salinity stress could possibly induce horizontal transmission of “Ca. Megaira polyxenophila” at slightly increased salinity concentrations in regard to normal conditions indicating the significance of environmental stress for the selection of the preferred transmission mode.
Symbiosen zwischen Bakterien und einzelligen Eukaryoten wie Ciliaten und Amöben sind weitverbreit. In den vergangenen Jahren stieg die Anzahl der Artbeschreibungen symbiotisch lebender Bakterien stetig an. Besonders Vertreter der Ordnung Rickettsiales (Alphaproteobacteria), welche obligat intrazelluläre Menschen- und Tierpathogene sowie andere nicht-pathogene, intrazelluläre Arten beinhaltet, rückten in den Fokus. Die Biodiversität von Protisten und mit ihnen assoziierte, bakterielle Symbionten wird jedoch weitestgehend unterschätzt. Daher untersuchte ich, welche verschiedenen Bakterienarten im Ciliaten Paramecium vorkommen, ob Paramecien als potentielles Reservoir für menschen- und tierpathogene Bakterien dienen können und ob die Artverbreitung von Paramecium und seiner Symbionten geographische Muster aufweist. In der vorliegenden Arbeit, präsentiere ich Verbreitungs- und Diversitätsdaten von Paramecium-Arten sowie deren Endosymbionten, welche aus nord- und südamerikanischen Süß- und Brackwasserproben isoliert wurden. Die am häufigsten erfassten Arten in nordamerikanischen Proben waren Paramecium biaurelia, Paramecium caudatum und Paramecium multimicronucleatum, sowie P. multimicronucleatum und Paramecium calkinsi in südamerikanischen Proben. Die identifizierten Arten weisen auf geographische Unterschiede in der Artverteilung der beiden Kontinente hin. Monoklonale Massenkulturen wurden unter Verwendung universeller und spezifischer Sonden mittels Fluoreszenz in situ Hybridisierung auf das Vorhandensein von Endosymbionten überprüft. Dabei entdeckte ich bakterielle Endosymbionten verschiedener Rickettsiales sensu lato-Vertreter in Paramecienkulturen aus acht verschiedenen Habitaten. Darüber hinaus charakterisierte ich die vier neuen Endosymbiontenarten “Candidatus Fokinia solitaria”, “Candidatus Fokinia cryptica”, “Candidatus Bealeia paramacronuclearis” und “Candidatus Jurandia parameciophila” und lieferte detaillierte Artbeschreibungen. Somit erfasste ich nicht nur verschiedene Symbionten in unterschiedlichen Paramecium-Arten der beiden Kontinente, sondern identifizierte auch Bakterien, welche nahe mit relevanten Pathogenen verwandt sind. Die phylogenetischen Informationen, die mir diese vier neuen Bakterienarten lieferten, halfen mir die taxonomische Organisation der Ordnung Rickettsiales zu revidieren, welche ich in die zwei Ordnungen Rickettsiales sensu stricto und Holosporales ord. nov. aufspaltete. Die Bedeutung einer Symbiose für seinen Wirt kann unter anderem durch die Erforschung der Übertragungswege abgeschätzt werden. Die meisten natürlichen Wirt-Symbionten-Systeme zeigen gemischte Übertragungswege, auch wenn eine der Übertragungsarten nur wenig genutzt wird. Ferner tritt eine Evolution hin zu sich ausschließlich vertikal oder horizontal übertragenden Symbionten selten auf. Die Bedingungen, unter denen eine horizontale Übertragung der meisten sich als vertikal übertragend beschriebenen Symbionten stattfindet, sind unbekannt und müssen nach wie vor untersucht werden. Deshalb legte ich meiner Arbeit die Annahme zugrunde, dass sowohl Vektororganisen als auch Umweltstress eine horizontale Übertragung begünstigen können. Ich führte verschiedene Infektionsexperimente durch, in denen ich die Effekte von Vektororganismen und verschiedener abiotischer Stressfaktoren als potenzielle Auslöser für eine horizontale Übertragung untersuchte. In den Experimenten, die sich mit einer Vektor-induzierten Übertragung beschäftigten, konnte ich keine möglichen Vektoreigenschaften von Holospora gegenüber Caedibacter nachweisen, wie sie gegenüber Futterbakterien bekannt sind. In seltenen Fällen beobachtete ich jedoch eine Vektor-unabhängige, horizontale Übertragung von Caedibacter taeniospiralis auf Symbionten-freie Paramecium-Arten. Die hier getesteten C. taeniospiralis-Genotypen sind möglicherweise immer noch dazu in der Lage, die Vorteile einer zusätzlichen horizontalen Übertragung zu nutzen. Zusätzlich testete ich die Effekte von Salz- und Temperaturstress auf die Übertragung der kürzlich beschriebenen, bakteriellen Symbionten “Candidatus Megaira polyxenophila” und “Ca. Bealeia paramacronuclearis” als potentiell interessante Vertreter. Die getesteten Symbionten- und Wirtsgenotypen unterschieden sich hinsichtlich ihrer Reaktion auf den ausgelösten abiotischen Stress. Die erzielten Ergebnisse zur horizontalen Übertragung erscheinen jedoch nicht eindeutig. Während Temperaturstress keine Effekte auf die Übertragung hatte, führte leichter Salzstress zu einer möglichen horizontalen Übertragung von “Ca. Megaira polyxenophila”, was auf die Bedeutung abiotischer Stressfaktoren bei der Wahl des bevorzugten Übertragungsweges hinweist.
Le simbiosi fra batteri ed eucarioti unicellulari, come i ciliati e le amebe, è un fenomeno molto diffuso. Negli anni passati, il numero di descrizioni di specie di batteri simbionti è cresciuto costantemente. In particolare, sono venuti alla ribalta i membri dell’ordine Rickettsiales (Alphaproteobacteria), che comprende patogeni intracellulari obbligati umani e di animali, oltre ad altre specie di batteri intracellulari non patogeni. Comunque, la biodiversità dei protisti e dei batteri simbionti a loro associati è ancora largamente sottostimata. Perciò, è stato ipotizzato che ci siano diversi tipi di batteri che vivono in associatione con il ciliato Paramecium in natura, che I parameci possano fungere da possibili serbatoi naturali per patogeni umani e animali, e che la distribuzione delle specie di Paramecium e dei suoi simbionti presenti dei pattern geografici. Nel presente lavoro di tesi, sono presentati dati sulla distribuzione e diversità di specie del genere Paramecium e dei loro batteri endosimbionti, isolate da ambienti dulciacquicoli e salmastri dal nordamerica e sudamerica. Le specie più frequenti sono state Paramecium biaurelia, Paramecium caudatum e Paramecium multimicronucleatum per i campioni nordamericani, e P. multimicronucleatum e Paramecium calkinsi per i campioni sudamericani, il che indica differenze geografiche nella distribuzione della specie di Paramecium fra I gue continenti. Colture monoclonali sono state saminate per la presenza di endosimbionti tramite ibridazione in situ a fluorescenza (FISH), impiegando sonde universali e specifiche. Sono stati rilevati batteri endosimbionti appartenenti a diversi taxa di Rickettsiales sensu lato in colture di paramecio isolate da otto diversi habitat. Inoltre, sono state caratterizzate quattro nuove specie di endosimbionti, rispettivamente, “Candidatus Fokinia solitaria”, “Candidatus Fokinia cryptica”, “Candidatus Bealeia paramacronuclearis” e “Candidatus Jurandia parameciophila”), fornendo descrizioni dettagliate. Perciò non solo sono stati caratterizzati diversi batteri simbionti in differenti specie di ospiti da due continenti, ma sono anche stati identificati batteri affini a patogeni umani e animali. I dati filogenetici forniti dalle quattro nuove specie mi hanno permesso di revisionare l’organizzazione tassonomica dell’ordine Rickettsiales, che è stato diviso nei due ordini Rickettsiales sensu stricto e Holosporales ord. nov. L’importanza e la severità di una simbiosi per l’ospite può essere stimata studiando le modalità di trasmissione del simbionte. La maggior parte dei sistemi ospite-simbionte in natura presentano una modalità mista di trasmissione, anche se una tipo di trasmissione avviene solo occasionalmente. Inoltre, l’evoluzione di simbionti trasmessi eclusivamente per via verticale o orizzontale è scarsa. Comunque, le circostanze per la trasmissione orizzontale di molti simbionti finora descritti come trasmessi orizzontalmente sono completamente sconosciute, e necessitano di essere rivelate. Perciò, è stato ipotizzato che sia organismi vettori che stress ambientali possano indurre la trasmissione orizzontali. Sono stati eseguiti diversi set di esperimenti di infezione, impiegando organismi vettori e fattori di stress ambientali come agenti che potessero indurre tramissione orizzontale. In esperimenti incentrati sulla trasmissione indotta da vettori, non è stato possibile confermare capacità di vettore da parte di Holospora nei confronti di Caedibacter, come invece descritto nei confronti di batteri a vita libera. Comunque, in rari casi è stata osservata trasmissione orizzontale di Caedibacter taeniospiralis verso specie di Paramecium indipendente dal vettore. I genotipi testati di C. taeniospiralis potrebbero comunque trarre un vantaggio dalla possibilità alternativa di trasmissione orizzontale, anche se questa avviene solo di rado. Inoltre, è stato testato l’effetto dello stress da salinità e temperatura sulla trasmissione dei batteri simbionti recentemente descritti “Candidatus Megaira polyxenophila” e “Ca. Bealeia paramacronuclearis” come rappresentanti potenzialmente interessanti. I genotipi coinvolti di ospiti e simbionti hanno risposto diversamente a fattori indotti di stress ambientale. I risultati sono stati ambigui in termini di trasmissione orizzontale. Mentre lo stress da temperatura non ha mostrato un effetto sulla trasmissione, è stata rilevata una possibile induzione della trasmissione di “Ca. Megaira polyxenophila” in presenza di basso stress salino rispetto alle condizioni normali, indicando l’importanza degli stress ambientali per la selezione della modalità di trasmissione.

Climate change is a growing concern for the scientific community. For the Iberian Peninsula it is predicted that climate change driven alterations will be more profound, as well as for mountain regions. The predictable alterations will have profound implications on biota, being amphibians suggested as one of the most vulnerable classes due to their unique characteristics and high susceptibility to environmental factors. With this in mind the aim of our study was to assess the effects of climate change related abiotic factors in amphibian larvae from two distinct altitudes (low and high). To achieve this aim two different approaches were made. The first consisted in using a full factorial experimental design to assess the effects of two abiotic factors (pH and temperature) in larvae from both altitudes. The second consisted in assessing the effect of several exposure volumes/water column heights in tadpoles from high altitude which will be more prone to be affected to desiccation effect. The endpoints measured for both approaches were both morphological (size) and physiological (antioxidant defense system and peroxidative damage). From our approach the main results showed that temperature has higher influence than pH in the development time and also in the size in tadpoles, being the effects dependent on the origin (low or high altitude) of the animals. Also, the physiological parameters suggest that animals from high altitudes present higher stress to high temperatures. Furthermore, the scenario of the lowest volume/water column height (desiccation) also revealed high stress for the animals. Overall, the most relevant results of the present study, indicate that the alteration in abiotic factors, mainly temperature has influence on the development of P. perezi and that high altitude populations may be more severely affected in a scenario of climate change were temperature increases and desiccation occurs.
As alterações climáticas são uma preocupação crescente na comunidade científica. As previsões apontam para que estas alterações sejam mais intensas na Península Ibérica, assim como nas regiões montanhosas. As alterações previstas irão ter implicações acentuadas na biota, sendo os anfíbios apontados como uma das classes mais vulneráveis devidos às suas características únicas e a sua alta suscetibilidade a fatores ambientais. Posto isto, o principal objetivo deste estudo foi avaliar os efeitos de fatores abióticos relacionados com as alterações climáticas em larvas de anfíbios oriundos de duas altitudes diferentes (elevada e baixa). Para alcançar este objetivo foram feitas duas abordagens diferentes. A primeira consistiu em avaliar os efeitos de dois fatores (pH e temperatura) em girinos de ambas as altitudes. O segundo compreendeu avaliar o efeito de diferentes volumes/colunas de água em girinos de altitude elevada, pois estes serão mais propensos a serem afetados pela dessecação. Os parâmetros medidos, para as duas abordagens experimentais, foram morfológicos (tamanho) e fisiológicos (sistema de defesa antioxidante e dano peroxidativo). A nossa abordagem mostrou, no geral, que a temperatura teve maior influência que o pH no tempo de desenvolvimento e no tamanho dos girinos, sendo que os efeitos dependem da origem dos animais (baixa ou elevada altitude). Também os parâmetros fisiológicos sugerem que os animais de altitudes elevadas apresentam um maior stress às temperaturas elevadas. Além disso, o menor volume de água (dessecação) também revelou maior stress para os animais. No geral, os resultados mais relevantes deste estudo, indicam que as alterações nos fatores abióticos, principalmente na temperatura, tiveram influência no desenvolvimento de P. perezi e as populações de altitudes elevadas podem ser mais severamente afetadas num cenário de alterações climáticas em que a temperatura aumente e a dessecação ocorra.
Mestrado em Ecologia Aplicada

Stress signaling pathway regulates proteins which are critical for reprogramming of metabolic synthesis and gene expression to achieve homeostasis and cellular stability under stress conditions. The understanding of stress signaling mechanism and response will increase the ability to improve plant‘s oralgal resistance to stress. The theoretical part summarizes the current knowledge of biosynthetic pathway and miRNA roles and presents selected model organisms (Chlamydomonas, Arabidopsis). As well, it introduces abiotic stress and seeks links between abiotic factors, secondary metabolites and miRNAs. The experimental part deals with selected abiotic factors [lycorine, UV-C radiation, ZnSe quantum dots (QDs), CO2] and their influence on Chlamydomonas reinhardtii and Arabidopsis thaliana. It has been shown that 100 and 250µM ZnSe QDs in the form of foliar feeding caused oxidative stress in the leaves without morphological changes in Arabidopsis thaliana. Additionaly, 250µM concentration inhibited the viability of Agrobacterium tumefaciens, a Gram-negative bacterium infecting plants, by 60 %. Furthermore, the regulatory biochemical, molecular and post-transcriptional pathways of Chlamydomonas reinhardtii response to UV-C radiation and lycorine were identified. Further, three specific miRNAs (Cre06.g281600, Cre06.g30900, Cre16.g662600) have been studied in connection with carbon concentrating mechanism (CCM) in Chlamydomonas reinhardtii. Expression of mentioned miRNAs had a positive correlation with target mRNA and revealed the potential regulatory role of miRNAs during CCM adaprion and possibility to increase the efficiency of photosynthesis. Despite, the role of epigenetics including miRNA interference is still unclear in the unicellular algae.

碩士
國立臺灣大學
植物科學研究所
102
WRKY transcription factors are a large protein family found in plants. The name of WRKY proteins is derived from their prominent DNA-binding region, designated the WRKY domain, a highly conserved region constituted by about 60 amino acid residues. It has been known that - WRKY transcription factors play important roles in the regulation of genes associated with plant defense responses - but the role of WRKY proteins in abiotic stress responses is still unclear in Arabidopsis thaliana. In this study, we showed that WRKY17, the member of WRKY IId subfamily, could interact with the HD2-type HDAC HD2C. Disruption of WRKY17 rendered plants sensitive to salt. Compared with wrky17.2 and hd2c-1 plants, wrky17.2/hd2c-1 double mutant plants displayed increased salt sensitivity. In addition, the expression of ABA-responsive genes was changed in these mutants. These data suggested that WRKY17 may play a role in abiotic stress responses.

博士
國立臺灣大學
植物科學研究所
101
AP2/ERF proteins play crucial roles in plant growth and development and in responses to biotic and abiotic stresses. ETHYLENE RESPONSE FACTOR 53 (AtERF53) is a transcription factor and belongs to group 1 in the ERF family which was induced in the early stage of dehydration and salt treatment, especially was highly induced by dehydration. The functional study of AtERF53 is hampered because its protein expression in Arabidopsis is vulnerable to degradation in overexpressed transgenic lines. In a previous study, we found that RING domain ligase2 (RGLG2) interacted with AtERF53 in the nucleus, and mediated AtERF53 ubiquitination for proteasome degradation. Taking advantage of the RING domain ligase1/ RING domain ligase2 (rglg1rglg2) double mutant in which AtERF53 can express stably, we investigated the physiological function of AtERF53. In this study, we demonstrate that expression of AtERF53 in wild-type Arabidopsis was responsive to heat and abscisic acid (ABA) treatment. Histological analysis showed that AtERF53 was expressed in stems, leaves, roots, and especially in the vasculature, but not in guard cells or inflorescence. From results of the cotransfection experiment, we concluded that AtERF53 has positive transactivation activity. Overexpression of AtERF53 in the rglg1rglg2 double mutant conferred better heat-stress tolerance and had resulted in higher endogenous ABA and proline levels compared to rglg1rglg2 double mutants. AtERF53 also has a function to regulate guard-cell movement because the stomatal apertutre of AtERF53 overexpressed in rglg1rglg2 double mutant was smaller than that in the rglg1rglg2 double mutant under ABA treatment. In a global gene expression study, we found higher expression of many stress-related genes, such as DREB1A, COR15A, COR15B , PLC, P5CS1, cpHSC70s, and proline and ABA metabolism-related genes. Furthermore, we identified several downstream target genes of AtERF53 by chromatin immunoprecipitation assay. In conclusion, these genetic, molecular and biochemical studies may explain how the AtERF53 transcription factor contributes to abiotic stress tolerance in Arabidopsis.

碩士
輔仁大學
生命科學系碩士班
101
Stress is defined as the external factor that exerts disadvantageous influences on plant growth. There are two types of stress, i.e. biotic and abiotic. The abiotic stress includes drought, salinity, heat, cold, ... etc. When plants encounter stresses, one plant hormone, i.e. abscisic acid (ABA), will activates several transcription factors and plays key role in plant stress response. The MYB gene family is reported to be among those involved in regulation of stress response pathway. However, the function and regulatory role of this gene family still remain mostly unclear. In order to obtain more insight about how these gene function in plant under stresses, in this study, we investigate one of the MYB gene family, i.e. AtMYB107 gene, which had showed three-fold magnitude of expression change under abiotic stresses using microarray analysis. After treated 10-day old Arabidopsis seedlings with various stress conditions, we have observed significant increments of AtMYB107 gene expression under ABA, cold, drought, and UV-C stresses. Unexpectedly, the AtMYB107 mutant line (mut), obtained from T-DNA insertion in promoter, showed increased AtMYB107 gene expression after cold treatment. It may imply a negatively regulatory role of the promoter insertion site. In addition, only root and chlorophyll content have significant differences between two AtMYB107 transgenic line and wild type plants in normal condition. The exogenous ABA test, stress tolerance assay, and detection of candidate genes prove the overexpression of AtMYB107 confers ABA sensitivity and UV-B, drought, cold tolerance. Finally, the UV-B stress and osmotic stress response is link up again in Our results.

Abiotic stress is one of the major factors that affect food production worldwide and, therefore understanding stress responsive proteins and engineering plants for abiotic stress tolerance is very important. In the present study, the biological role of pea pathogenesis-related 10.4 (PR-10.4; also known as abscisic acid responsive 17; ABR17) in abiotic stress tolerance has been investigated. Our investigation on ribonuclease (RNase) activity of ABR17 suggested that highly conserved histidine-69 and glutamic acid-148 are important for RNase activity. In order to further investigate the biological role(s) of ABR17, transcriptional profiling of pea ABR17-mediated gene expression changes in ABR17-transgenic Arabidopsis thaliana plants was carried out using microarrays. Our results indicated that pea ABR17 modulates many plant growth/development genes most of which are cytokinin (CK) responsive. These results agree very well with previously reported enhanced endogenous CKs in these transgenic plants. However, no significant changes in transcript abundance of CK biosynthetic genes were observed between transgenic and wild-type plants, suggesting an alternate source of CK in ABR17-transgenic plants. It is speculated that ABR17 may act as either a CK reservoir (through its reported CK binding property) or may be responsible for isopentenylated-tRNA degradation (through its demonstrated RNase activity) thereby increasing endogenous CK pools. Furthermore, microarray analysis of salinity stressed ABR17-Arabidopsis indicated that ABR17 modulates many stress responsive genes that included four putative AP2 family genes (RAP2.6-At1g43160, RAP2.6L-At5g13330, DREB26-At1g21910 and DREB19-At2g38340). Functional characterization of these genes suggested that they are transcription factors and they play very important roles in abiotic stress response in addition to growth and development. Moreover, overexpression of RAP2.6L and DREB19 genes enhanced salinity and drought tolerance in Arabidopsis. Taken together, our results suggest that pea ABR17 proteins are important in abiotic stress responses as they may act as source of enhanced CKs and they may also modulate expression of stress responsive genes to enhance stress tolerance in plants. However, additional research aimed at deciphering the links between ABR17 and CK biosynthesis as well as the mechanism of ABR17-mediated gene expression changes should be conducted in order to get more insights into the biological roles of PR10 proteins in planta.
Plant Science

碩士
輔仁大學
生命科學系碩士班
106
External environmental factors that limit plant survival, growth and reproduction are called stress and can be divided into biotic and abiotic stress. Abiotic stress includes low temperature, high temperature, drought stress, salt stress and UV stress, etc. Abscisic acid (ABA) plays an important regulatory role when plants encounter stress and activates transcription factors to initiate further anti-stress responses. Among all families of transcription factors, MYB family is considered to be the largest group and the members of this group are considered to be related with different types of stresses. In order to gain a further confirmation of the function and expression of the MYB gene when plants are under stress. In this study, microarray data provided by TAIR has been used for selection of stress-response AtMYB85 gene. Ten-day-old Arabidopsis thaliana wild type (WT) plants were treated with various kinds of stress conditions and analyzed by qRT-PCR. The results showed that AtMYB85 gene was expressed under ABA, drought and salt treatments, indicating that it may be involved in these stress regulatory pathways. Then we use forward primer and reverse primer to screen homozygous plants from transgenic lines established in the lab and mutant line obtained from SALK by PCR. RNA was extracted from each line and tested by RT-PCR. AtMYB85 gene expression was inhibited in mutant lines and significantly increased in over-expression lines. In the ABA test, the seeds of mutants were found to be significantly inhibited by exogenous ABA and delayed in their germination time. Under drought and salt stress, the physiological indexes of the mutants were obviously affected, which were more sensitive to stress. In the tolerance test, it was found that the tolerance of salt stress-treated mutants decreased while that of over-expression strains increased. After molecular testing of candidate genes related to salt stress, the expression of these candidate genes was considered to be associated with stress regulatory pathways. Therefore, we assumed that activation of AtMYB85 would affect the production of plant lignin and confer drought and salt tolerance to over-expression plants. Under salt stress, ABA-dependent and SOS-responsive pathways were also regulated by this gene.

碩士
國立臺灣大學
植物科學研究所
100
Plants face various environmental stresses, such as drought, salinity, heat and cold directly. In response to heat stress (HS), heat shock response (HSR) is a conserved mechanism of increasing heat shock protein (HSP) genes expression through a heat shock factors (HSFs)-dependent mechanism. According to microarray databases, AtHSFA6a is induced by salt and osmotic stress, while AtHSFA6b expression is responsive to cold, salt, osmotic and drought treatment; in addition both genes are upregulated by ABA treatment suggesting the AtHSFA6s may involve in ABA-mediated responses. This report describes our investigation of the role of AtHSFA6s in response to abiotic stress in Arabidopsis by genetic and molecular approaches. Both AtHSFA6s were induced by abscisic acid (ABA), salt and osmotic stress. Downregulated AtHSFA6b expression in ABA-deficient and ABA-insensitive mutants indicates that ABA signaling is required for AtHSFA6b expression. Notably, ABRE acts as the functional cis-element in the promoter region of AtHSFA6b, suggests AtHSFA6b is a member involved in ABA-dependent pathway. Analysis of cotyledon greening, root growth and salt tolerance in mutant lines and transgenic plants indicated that AtHSFA6b participates in ABA-mediated stress responses and acts as a positive regulator. In addition, thermotolerance tests also showed AtHSFA6b is involved in basal- and acquired-thermotolerance. These studies reveal AtHSFA6b is a positive regulator in ABA-mediated stresses, and add a new signaling pathway into the complex HSF network.

Effects of a clear-cut timber harvest of the riparian zone were studied on 18 stream systems in watersheds influenced by a continental climate regime in the central interior of British Columbia. Streams selected in the Stuart-Takla and Willow-Bowron watersheds were sampled between mid-September and late October 1995. Study streams were classified according to stages of clear-cut harvest in the riparian zone: streams within unharvested reaches, streams without canopy cover (timber harvest less than 5 years old) or streams with a deciduous canopy cover (timber harvest more than 20 years old). Measurements were taken of topographic and habitat attributes, of watersheds and productivity and the benthic macroinvertebrate community. 'Uncut' stream channels had higher gradients and narrower widths than were found in 'old cut' streams suggesting that stream channel dimensions of streams in logged areas may have changed over time. More specifically, 'recent cut' streams had the lowest canopy cover and the highest chlorophyll α biomass. However, in 'old cut' streams with dense canopy cover, chlorophyll α biomass was similar to levels in 'uncut' streams. In addition, although density of benthic macroinvertebrates did not appear to have changed over time, total biomass increased soon after timber harvest. After more than 20 years, biomass measurements of macroinvertebrates were similar to levels found in unharvested streams. Density and biomass of the scraper functional guild, in particular, were lowest in 'recent cut' streams. These streams also had the highest densities of predators and invertebrate parasites and the highest biomass of collector functional guilds and leeches. Higher biomass of the two collector guilds in 'recent cut' streams may have been linked to an increase in the quality of their food source, i.e., FPOM, although this was not measured in this study. Higher biomass of scrapers and collectors found soon after the timber harvest may have attracted resident fish into these streams, and so increased fish parasite biomass. After more than 20 years following timber harvest, biotic and abiotic factors may have returned to levels found prior to timber harvests. Future work is required to determine the factors that influence changes in benthic community composition and instream habitat in stream ecosystems following timber harvest in this region.

This thesis performs de novo predictions for functionally significant sequence motifs in the Arabidopsis genome under two separate contexts. Each study applies the use of genomic positional information, statistical over-representation and several biologically contextual filters to maximize the visibility of biological signal in prediction results. Numerous literature supported motifs are prevalent in the results of both studies and a number of novel motif patterns possess a strong potential for in planta significance. The first study examines the statistical over-representation of C-terminal tripeptides as a means for identifying eukaryotic conserved protein targetting signatures. Comparative genomics is applied to the analysis of tripeptide frequencies in the C-terminus of 7 eukaryotic proteomes. While biological signal is maximized through the filtering of both simple sequences and homologous sequences present across protein families. The second study introduces a methodology for the effective prediction of transcription factor binding sites in Arabidopsis. A collection of motif prediction algorithms and a novel enumerative strategy are applied to the prediction of cis-acting regulatory elements within the promoters of genes found coexpressed within distinct tissues and under specific abiotic stress treatments. Overall, the analysis identifies 4 known motifs in expected contexts, 5 known motifs in novel contexts and 7 novel motifs with a high potential for biological function.