Glicose e oxigênio são moléculas essenciais para a maioria dos organismos vivos. Além de sua importância nos processos de produção de energia - glicose como fonte de carbono e energia e oxigênio como aceptor dos elétrons doados por NADH e FADH2 - estes dois compostos funcionam como efetuadores, modulando vários processos metabólicos e fisiológicos nas células. Visto que a mitocôndria é um dos alvos afetados pelas disponibilidades destas duas moléculas, nós isolamos e seqüenciamos o genoma mitocondrial de Trichoderma reesei, um fungo multicelular empregado neste trabalho como sistema modelo. Foi estudado o efeito da variação de concentração de glicose e oxigênio sobre a expressão de transcritos do genoma mitocondrial, bem como sua implicação no metabolismo de glicose. São apresentadas análises da expressão gênica de aproximadamente 2000 transcritos de T. reesei submetido a concentrações limitantes de oxigênio dissolvido, realizadas com o emprego da técnica de microarrays de cDNA. Pelo menos 330 transcritos foram diferencialmente expressos em função da disponibilidade de oxigênio. Aqueles envolvidos nos processos de síntese protéica e divisão celular foram regulados negativamente, enquanto transcritos relacionados com funções de defesa celular e síntese de RNA foram positivamente regulados. Uma fração substancial de outros genes afetados pela baixa disponibilidade de oxigênio não possui, atualmente, funções celulares conhecidas. Esta observação deve contribuir para a posterior anotação funcional do genoma de T. reesei. Também foram identificados reguladores transcricionais diferencialmente expressos em baixas tensões de oxigênio. O perfil de expressão destes reguladores aponta-os como potenciais candidatos ao envolvimento com a expressão de genes afetados pela disponibilidade de oxigênio. / Glucose and oxygen are essential molecules in most of living organisms. In addition to their importance in production of energy - glucose as a carbon and energy source and oxygen as an acceptor of electrons donated by NADH and FADH2 - both molecules function as effectors modulating various metabolic and physiological processes in the cell. Because one of the targets affected by both molecules is the mitochondrion, we isolated and sequenced the mitochondrial genome of Trichoderma reesei, a multicellular fungus that is used in this study as a model system. The effect of varying the concentration of glucose and oxygen on the expression of the transcripts of the mitochondrial genome, and its implication on the metabolism of glucose, was studied. Gene-wide expression analyses of nearly 2000 transcripts of T. reesei under limited concentration of dissolved oxygen, using cDNA microarry technique, are presented. At least 330 transcripts were differentially expressed with respect to oxygen availability. Those involved in protein synthesis and cell division processes were downregulated, while transcripts involved in cell defense and RNA synthesis were upregulated. A substantive fraction of other anaerobically affected genes have currently unknown cellular roles, and these results should therefore contribute to further functional annotation of the genome. ln addition, we have identified transcriptional regulators that are differentially expressed at a low oxygen tensions. The expression profile of these regulators points them out as potential candidates involved in the expression of genes affected by oxygen availability.

Indiana University-Purdue University Indianapolis (IUPUI) / SETMAR is a chimeric protein that originates from the fusion of a SET domain to the mariner Hsmar1 transposase. This fusion event occurred approximately 50 million years ago, after the split of an anthropoid primate ancestor from the prosimians. Thus, SETMAR is only expressed in anthropoid primates, such as humans, apes, and New World monkeys. Evolutionary sequence analyses have revealed that the DNA-binding domain, one of the two functional domains in the Hsmar1 transposase, has been subjected to a strong purifying selection. Consistent with these analyses, SETMAR retains robust binding specificity to its ancestral terminal inverted repeat (TIR) DNA. In the human genome, this TIR sequence is dispersed in over 1500 perfect or nearly perfect sites. Given that many DNA-binding domains of transcriptional regulators are derived from transposases, we hypothesized that SETMAR may play a role in gene regulation. In this thesis, we determined the crystal structures of the DNA-binding domain bound to both its ancestral TIR DNA and a variant TIR DNA sequence at 2.37 and 3.07 Å, respectively. Overall, the DNA-binding domain contains two helix-turn-helix (HTH) motifs linked by two AT-hook motifs and dimerizes through its HTH1 motif. In both complexes, minor groove interactions with the AT-hook motifs are similar, and major groove interactions with HTH1 involve a single residue. However, four residues from HTH2 participate in nucleobase-specific interactions with the TIR and only two with the variant DNA sequence. Despite these differences in nucleobase-specific interactions, the DNA-binding affinities of SETMAR to TIR or variant TIR differ by less than two-fold. From cell-based studies, we found that SETMAR represses firefly luciferase gene expression while the DNA-binding deficient mutant does not. A chromatin immunoprecipitation assay further confirms that SETMAR binds the TIR sequence in cells. Collectively, our studies suggest that SETMAR functions in gene regulation.

On TFBS motif discovery, three novel GA based algorithms are developed, namely GALF-P with focus on optimization, GALF-G for modeling, and GASMEN for spaced motifs. Novel memetic operators are introduced, namely local filtering and probabilistic refinement, to significantly improve effectiveness (e.g. 73% better than MEME) and efficiency (e.g. 4.49 times speedup) in search. The GA based algorithms have been extensively tested on comprehensive synthetic, real and benchmark datasets, and shown outstanding performances compared with state-of-the-art approaches. Our algorithms also "evolve" to handle more and more relaxed cases, namely from fixed motif widths to most flexible widths, from single motifs to multiple motifs with overlapping control, from stringent motif instance assumption to very relaxed ones, and from contiguous motifs to generic spaced motifs with arbitrary spacers. / TF-TFBS associated sequence pattern (rule) discovery is further investigated for better deciphering protein-DNA interactions in regulation. We for the first time generalize previous exact TF-TFBS rules to approximate ones using a progressive approach. A customized algorithm is developed, outperforming MEME by over 73%. The approximate TF-TFBS rules, compared with the exact ones, have significantly more verified rules and better verification ratios. Detailed analysis on PDB cases and conservation verification on NCBI protein records illustrate that the approximate rules reveal the flexible and specific protein-DNA interactions with much greater generalized capability. / The comprehensive pattern discovery algorithms developed will be further verified, improved and extended to further deciphering transcriptionial regulation, such as inferring whole gene regulatory networks by applying TFBS and TF-TFBS patterns discovered and incorporating expression data. / Transcription Factor (TF) and Transcription Factor Binding Site (TFBS) bindings are fundamental protein-DNA interactions in transcriptional regulation. TFs and TFBSs are conserved to form patterns (motifs) due to their important roles for controlling gene expressions and finally affecting functions and appearances. Pattern discovery is thus important for deciphering gene regulation, which has tremendous impacts on the understanding of life, bio-engineering and therapeutic applications. This thesis contributes to pattern discovery involving TFBS motifs and TF-TFBS associated sequence patterns based on Evolutionary Computation (EC), especially Genetic Algorithms (GAs), which are promising for bioinformatics problems with huge and noisy search space. / Chan, Tak Ming. / Advisers: Kwong-Sak Leung; Kin-Hong Lee. / Source: Dissertation Abstracts International, Volume: 73-03, Section: B, page: . / Thesis (Ph.D.)--Chinese University of Hong Kong, 2010. / Includes bibliographical references (leaves 147-153). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [201-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.

Abstract not available

Vegetative propagation allows the amplification of selected genotypes for research, breeding, and commercial planting. However, efficient in vitro regeneration and genetic transformation remains a major obstacle to research and commercial application in many plant species. Our aims are to improve knowledge of gene regulatory circuits important to meristem organization, and to identify genes that might be useful for improving the efficiency of in vitro regeneration. In this thesis, we have approached these goals in two ways. First, we analyzed gene expression during poplar (Populus) regeneration using an AffymetrixGeneChip® array representing over 56,000 poplar transcripts. We have produced a catalog of regulated genes that can be used to inform studies of gene function and biotechnology. Second, we developed a GUS reporter system for monitoring meristem initiation using promoters of poplar homologs to the meristem-active regulatory genes WUSCHEL (WUS) and SHOOTMERISTEMLESS (STM). This provides plant materials whose developmental state can be assayed with improved speed and sensitivity. For the microarray study, we hybridized cDNAs derived from tissues of a female hybrid poplar clone (INRA 717-1 B4, Populus tremula x P. alba) at five sequential time points during organogenesis. Samples were taken from stems prior to callus induction, at 3 days and 5 days after callus induction, and at 3 and 8 days after the start of shoot induction. Approximately 15% of the monitored genes were significantly up-or down-regulated based on both Extraction and Analysis of Differentially Expressed Gene Expression (EDGE) and Linear Models for Microarray Data (LIMMA, FDR<0.01). Of these, over 3,000 genes had a 5-fold or greater change in expression. We found a very strong and rapid change in gene expression at the first time point after callus induction, prior to detectable morphological changes. Subsequent changes in gene expression at later regeneration stages were more than an order of magnitude smaller. A total of 588 transcription factors that were distributed in 45 gene families were differentially regulated. Genes that showed strong differential expression encoded proteins active in auxin and cytokinin signaling, cell division, and plastid development. When compared with data on in vitro callogenesis from root explants in Arabidopsis, 25% (1,260) of up-regulated and 22% (748) of down- regulated genes were in common with the genes that we found regulated in poplar during callus induction. When ~3kb of the 5' flanking regions of close homologs were used to drive expression of the GUSPlus gene, 50 to 60% of the transgenic events showed expression in apical and axillary meristems. However, expression was also common in other organs, including in leaf veins (40% and 46% of WUS and STM transgenic events, respectively) and hydathodes (56% of WUS transgenic events). Histochemical GUS staining of explants during callogenesis and shoot regeneration using in vitro stems as explants showed that expression was detectable prior to visible shoot development, starting 3 to 15 days after explants were placed onto callus inducing medium. Based on microarray gene expression data, a paralog of poplar WUS was detectably up-regulated during shoot initiation, but the other paralog was not. Surprisingly, both paralogs of poplar STM were down-regulated 3- to 6-fold during early callus initiation, a possible consequence of its stronger expression in the secondary meristem (cambium) than in shoot tissues. We identified 15 to 35 copies of cytokinin response regulator binding motifs (ARR1AT) and one copy of the auxin response element (AuxRE) in both promoters. Several of the WUS and STM transgenic events produced should be useful for monitoring the timing and location of meristem development during natural and in vitro shoot regeneration. / Graduation date: 2008

The entire dissertation/thesis text is included in the research.pdf file; the official abstract appears in the short.pdf file (which also appears in the research.pdf); a non-technical general description, or public abstract, appears in the public.pdf file. Title from PDF of title page (University of Missouri--Columbia, viewed on March 10, 2010). Vita. Thesis advisor: Lorson, Christian L. "December 2008" Includes bibliographical references

Thesis (Ph. D.)--Ohio State University, 2007. / Full text release at OhioLINK's ETD Center delayed at author's request

The entire dissertation/thesis text is included in the research.pdf file; the official abstract appears in the short.pdf file (which also appears in the research.pdf); a non-technical general description, or public abstract, appears in the public.pdf file. Title from PDF of title page (University of Missouri--Columbia, viewed on April 1, 2010). Vita. Thesis advisor: Habib Zaghouani. "May 2008" Includes bibliographical references.

In angiosperms, specialized reproductive structures are borne in flowers to ensure their reproductive success. After the vegetative growth, plants undergo reproductive phase change to produce flowers. Floral meristems (FMs) are generated on the flanks of inflorescence and groups of specialized stem cells in the FM differentiate into four whorls of organs of a flower. In dicots, floral meristem successively gives rise to sepals, petals, stamens and carpels; after which it terminates. The fate of organs formed on FM is under the control of genetic regulators, key among which are members of MADS box transcription factor family. Their individual and combined act confers distinct identities to floral organs. Grass flowers are highly modified in structure. Rice flower, a model for grasses, is borne on a short branch called spikelet and they together from the basic structural units of the rice infloresences known as panicle. The outer whorl organs of a grass floret are bract-like structures known as lemma and palea to dicot sepals is highly dibated (see Chapter 1). In grass florets, petal homologs are a pair of highly reduced, fleshy bracts known as lodicules, while stamen and carpel homologs occupy the same position and share the same functions as their dicot counterparts. Aside from these distinct outer whorl organs, the florets are subtended by two pairs of bracts known as empty glumes and rudimentary glumes. The genetic regulators that control their unique identities and those that perform conserved functions are very intriguing and central questions in plant developmental biology. Using various contemporary and complementary technologies, we have analysed the molecular functions and downstream pathways of a MADS box transcription factor, OsMADSI during the rice floret meristem specification and organ development. Further by reverse genetics and overexpression studies, we have also functionally characterized two target genes of OsMADSI, OsETTINI and OsETTINI2 to understand their roles downstream to OsMADSI during the rice floret development.

Cette thèse de doctorat s’inscrit dans le développement et l’utilisation de méthodes mathématiques et informatiques qui exploitent les données temporelles d’expression des gènes issues de puces à ADN afin de rationaliser et de modéliser les réseaux de régulation génique. Dans cette optique, nous nous sommes principalement intéressés aux données d’expression des gènes de la drosophile (Drosophila melanogaster) pendant son développement, du stade embryonnaire au stade adulte. Nous avons également étudié des données concernant le développement d’autres eucaryotes supérieurs, la réponse d’une bactérie soumises à différents stress et le cycle cellulaire d’une levure. Ce travail a été réalisé selon trois volets principaux :la détection des stades de développement et des perturbations, les classifications de profils d’expression et la modélisation de réseaux de régulation.<p><p>Premièrement, l’observation des données d’expression utilisées nous a conduits à approfondir l’étude des phénomènes survenant lors des changements de stades de développement de la drosophile. Dans ce but, deux méthodes de détection automatique de ces changements ont été développées et appliquées aux données temporelles disponibles sur le développement d’eucaryotes supérieurs. Elles ont également été appliquées à des données temporelles relatives à des perturbations externes de bactéries. Cette étude à montré qu’une formulation mathématique simple permettait de retrouver les instants expérimentaux où une perturbation ou un changement de stade de développement est observé, à partir uniquement des profils d’expression. Par ailleurs, la réponse à une perturbation externe s’avère non distinguable d’une succession de stades de développement, sur la base des seuls profils temporels d’expression.<p><p>Deuxièmement, en raison des dimensions du problème constitué par les données d’expression de plusieurs milliers de gènes et de l’impossibilité de distinguer le rôle dans la régulation des gènes qui présentent des profils d’expression similaires, il s’est avéré nécessaire de classifier les gènes selon leurs profils d’expression. En nous basant sur les résultats obtenus lors de la détection des stades de développement, la démarche suivie est de regrouper les gènes qui présentent des profils temporels d’expression aux comportements similaires non seulement au cours de la série temporelle complète, mais également dans chacun des stades de développement. Dans cette optique, trois distances ont été proposées et utilisées dans une classification hiérarchique des données d’expression de la drosophile.<p><p>Troisièmement, des structures de modèles linéaires et non linéaires ainsi que des méthodes d’estimation et de réduction paramétriques ont été développées et utilisées pour reproduire les données d’expression du développement de la drosophile. Les résultats de ce travail ont montré qu’avec une structure de modèle linéaire simple, la reproduction des profils expérimentaux était excellente et que, dans ce cas, le réseau de régulation génique de la drosophile pouvait se contenter d’une faible connectivité (en moyenne 3 connexions par classe de gènes) et ce, sans hypothèse a priori. Toutefois, les modèles linéaires ont ensuite sérieusement été remis en question par des analyses de robustesse aux perturbations paramétriques et de stabilité des profils après extrapolation dans le temps. Dès lors, quatre structures de modèles non linéaires et cinq méthodes de réduction paramétrique ont été proposées et utilisées pour concilier les critères de reproduction des données, de robustesse et de stabilité des réseaux identifiés. En outre, ces méthodes de modélisation ont été appliquées à un sous-ensemble de 20 gènes impliqués dans le développement musculaire de la drosophile et pour lesquels 36 interactions ont été validées expérimentalement, ainsi qu’à des profils synthétiques bruités. Nous avons pu constater que plus de la moitié des connexions et non-connexions sont retrouvées par trois modèles non linéaires. Les résultats de cette étude ont permis d’éliminer certaines structures de modèle et méthodes de réduction et ont mis en lumière plusieurs directions futures à suivre dans la démarche de modélisation des réseaux de régulation génique. / Doctorat en Sciences de l'ingénieur / info:eu-repo/semantics/nonPublished