Gene regulation lies at the heart of most biological processes and transcription factors are the key molecules that control tissues specific gene expression. In higher eukaryotes transcription factors control gene expression by binding regulatory DNA segments called cis-regulatory modules (CRMs). The increasing number of sequenced genomes of multicellular eukaryotes along with high-throughput methods such as whole genome microarray expression data allows for systematic characterization of the CRMs that control gene expression. A first step towards understanding gene regulation is the identification of the regulatory elements present in the genome. We take advantage of the large database of spatio-temporal patterns of gene expression in D. melanogaster embryogenesis to identify sets of developmentally co-expressed genes. We developed a computational method that identifies DNA binding sites for transcription factors from families of co-regulated genes that are expressed during Drosophila embryo development. This method discovers over-represented motifs in a set of co-regulated genes using the exhaustive motif enumeration technique. Clustering the predicted motifs identifies the CRMs, which assist in translating a combinatorial code of TF inputs into a specific gene expression output. The predicted CRMs were verified experimentally by searching the whole genome for the predicted CRMs and establishing expression pattern of the genes that are associated with these CRMs. It is well know that the gene expression is substantially controlled through CRMs and those key regulatory sequences are conserved in related species. The conservation of CRMs can be studied by comparing the related genomes and alignment methods are widely used computational tools for comparing the sequences. However, in distantly related species the CRM sequences are simply not align able. To identify the similar CRMs in distantly related species we developed a non-alignment based method for discovering similar CRMs in related species. This method is based on word frequencies where the given sequences are compared using Poisson based metric. When starting with a set of CRMs involved in Drosophila early embryo development, we show here that our non-alignment method successfully detects similar CRMs in distantly related species ( D. ananassae, D. pseudoobscura, D. willisoni, D. mojavensis, D. virilis, D. grimshawi ). This method proved efficient in discriminating the functional CRMs from the non-functional ones.

O Coup¬TF, αποτελεί ορφανό μέλος της υπεροικογένειας των υποδοχέων των στεροειδών/θυρεοειδών ορμονών και κατέχει κυρίαρχο ρόλο στην ανάπτυξη των εμβρύων όλων των μεταζώων. Στην παρούσα Διατριβή μελετήθηκε η cis¬ ρυθμιστική περιοχή του γονιδίου του, με σκοπό την ένταξή του στο γονιδιακό ρυθμιστικό δίκτυο του εμβρύου του αχινού. Με πειράματα in situ υβριδοποίησης βρέθηκε ότι το γονίδιο PlCoup¬TF εκφράζεται στο στοματικό εξώδερμα του γαστριδίου και στη βλεφαριδωτή ζώνη στον πλουτέα, στο είδος Paracentrotus lividus. Από παλαιότερα πειράματα είχε βρεθεί ότι το τμήμα της ανοδικής περιοχής που εκτείνεται από το -232 ως το ¬532 (τμήμα a), είναι απαραίτητο και επαρκές για την έκφραση του γονιδίου αναφοράς (gfp) στη βλεφαριδωτή ζώνη του πλουτέα. Εντός της περιοχής a ανευρέθησαν τρία πιθανά ρυθμιστικά στοιχεία (¬ 453, ¬432 και ¬377) του γονιδίου PlCoup¬TF, τα οποία αναγνωρίζονται από πρωτεΐνες εμβρυικού πυρηνικού εκχυλίσματος. Στοχευμένες μεταλλάξεις των στοιχείων αυτών, οδήγησαν σε μείωση της έκφρασης του γονιδίου αναφοράς (στοιχείο ¬453) και στην εκτοπική έκφρασή του (στοιχεία ¬432 και ¬377). Περαιτέρω μελέτη των παραγόντων που αναγνωρίζουν τα στοιχεία αυτά, οδήγησε στο συμπέρασμα ότι ο μεταγραφικός παράγοντας PlElk αναγνωρίζει το στοιχείο ¬ 453 και ρυθμίζει θετικά το γονιδίο του PlCoup¬TF και ο μεταγραφικός παράγοντας PlOtx αναγνωρίζει το στοιχείο -377 και καταστέλλει την έκφραση του PlCoup¬TF στο αντιστοματικό εξώδερμα. Τα αποτελέσματα της παρούσης εργασίας οδήγησαν στην ένταξη του γονιδίου PlCoup¬TF και των δύο ρυθμιστών του στο γονιδιακό ρυθμιστικό δίκτυο που καθορίζει τη διαφοροποίηση της βλεφαριδωτής ζώνης εντός του εμβρυικού εξωδέρματος. / Coup­TF, an orphan member of the nuclear receptor super family, has a fundamental role in the development of metazoan embryos. The study of the gene's regulatory circuit in the sea urchin embryo will facilitate the placement of this transcription factor in the well­studied embryonic Gene Regulatory Network (GRN). The Paracentrotus lividus Coup­TF gene (PlCoup­TF) is expressed throughout embryonic development preferentially in the oral ectoderm of the gastrula and the ciliary band of the pluteus stage. Two overlapping λ genomic clones, containing three exons and upstream sequences of PlCoup­TF, were isolated from a genomic library. The transcription initiation site was determined and 5′ deletions and individual segments of a 1930 bp upstream region were placed ahead of a GFP reporter cassette and injected into fertilized P.lividus eggs. Module a (−532 to −232), was necessary and sufficient to confer ciliary band expression to the reporter. Comparison of P.lividus and Strongylocentrotus purpuratusupstream Coup­TF sequences, revealed considerable conservation, but none within module a. 5′ and internal deletions into module a, defined a smaller region that confers ciliary band specific expression. Putative regulatory cis­acting elements (RE1, RE2 and RE3) within module a, were specifically bound by proteins in sea urchin embryonic nuclear extracts. Site­specific mutagenesis of these elements resulted in loss of reporter activity (RE1) or ectopic expression (RE2, RE3). It is proposed that sea urchin transcription factors, which bind these three regulatory sites, are necessary for spatial and quantitative regulation of the PlCoup­TF gene at pluteus stage sea urchin embryos. Additional experiments led us to the conclusion that transcription factor PlElk binds to the ­453 regulatory element and positively regulates PlCoup­TF gene in the ciliary band. Furthermore, PlOtx binds to the ­377 regulatory element and negatively regulates PlCoup­TF gene in the aboral ectoderm. These findings lead to the hierarchical positioning of PlCoup­TF within the embryonic GRN

Submitted by Gabriela Lopes (gmachadolopesufpel@gmail.com) on 2016-09-28T14:00:15Z No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) dissertação.pdf: 1451490 bytes, checksum: 8db111be8fc2a2a277ed6aa88f84a6e6 (MD5) / Approved for entry into archive by Aline Batista (alinehb.ufpel@gmail.com) on 2016-09-28T19:03:52Z (GMT) No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) dissertação.pdf: 1451490 bytes, checksum: 8db111be8fc2a2a277ed6aa88f84a6e6 (MD5) / Made available in DSpace on 2016-09-28T19:03:52Z (GMT). No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) dissertação.pdf: 1451490 bytes, checksum: 8db111be8fc2a2a277ed6aa88f84a6e6 (MD5) Previous issue date: 2013-06-28 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / As raízes possuem uma grande variedade de funções nas plantas, incluindo absorção de água, nutrientes e suporte estrutural. A combinação de métodos clássicos de genética e melhoramento com tecnologias moleculares de análise genômica abre uma nova perspectiva para a ampliação do conhecimento das bases genéticas e aceleração de programas de melhoramento. A maioria dos conhecimentos sobre as redes gênicas envolvidas no desenvolvimento radicular vem sendo acumuladas na espécie Arabidopsis thaliana, modelo de planta dicotiledônea. O entendimento dos mecanismos envolvidos na regulação da expressão dos genes é essencial para compreender a forma e a função dos sistemas. Os elementos cis-acting são regiões do DNA que atuam como interruptores moleculares envolvidos na regulação da transcrição de uma rede gênica dinâmica. Embora freqüentemente tenham somente cinco a 20 pb de tamanho, os elementos cis-acting são críticos para o entendimento da regulação gênica. O conhecimento destes elementos presentes na região promotora de famílias gênicas, poderá contribuir para a compreensão dos sistemas reguladores da expressão da rede gênica envolvida na formação do sistema radicular. O objetivo desse trabalho é identificar os elementos cis-acting presentes na região promotora de genes de arroz (Oryza sativa subsp japonica cv. Nipponbare) similares aos genes das famílias Argonauta, Cullin e Ara de Arabidopsis thaliana. A região promotora dos genes destas famílias no arroz foi investigada quanto à abundância destes elementos. As seqüências foram analisadas utilizando o programa “Signal Scan Search” do portal “Plant Cis-acting Regulatory DNA Elements” (PLACE) para a identificação dos diferentes elementos cis-acting. Foram detectados 96 diferentes elementos, sendo cinco destes (GAREAT (TAACAAR), TGACGTVMAMY (TGACGT), CCAATBOX1 (CCAAT), LECPLEACS2 (TAAAATAT) e SV40COREENHAN (GTGGWWHG), comuns as famílias gênicas Argonauta, Cullin e Ara. / The roots have a large range of functions in plants, including acquisition of water and nutrients, as well as structural support. The combination of classical methods of genetics and breeding with molecular technologies for genomic analysis opens a new perspective to expand the knowledge of the genetic basis and to accelerate breeding programs. The most advanced knowledge regarding gene networks involved in root development has been obtained in the model dicotyledon plant species Arabidopsis thaliana. Understanding the mechanisms involved in regulation of gene expression is essential to predict the form and function of systems. Cis-acting elements are DNA regions that act as molecular switches involved in the regulation of transcription of dynamic gene network. Although often having only five to 20 bp in size, cis-acting elements are critical to the understanding of gene regulation. Knowledge of the cis-acting elements present in the promoter region of gene families, can contribute to the understanding of the expression regulatory systems of these genes and others, involved with the root system. The objective of this study is to identify the cisacting elements present in the upstream region of rice (Oryza sativa subsp japonica cv. Nipponbare) genes, similar to gene families Argonauta, Cullin and Ara in Arabidopsis thaliana. The promoter region of these rice gene families were investigated for the abundance of cis-acting elements. The sequences were analyzed using the software “Signal Scan Search” of the website “Plant Cis-acting Regulatory DNA Elements” (PLACE) to the identification of different cis-acting elements. It were detected 96 different cis-acting elements, and five of these, (GAREAT (TAACAAR), TGACGTVMAMY (TGACGT), CCAATBOX1 (CCAAT), LECPLEACS2 (TAAAATAT) e SV40COREENHAN (GTGGWWHG) were common to the gene families Argonauta, Cullin and Ara.

Submitted by Fabio Sobreira Campos da Costa (fabio.sobreira@ufpe.br) on 2016-08-26T13:34:53Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) DISSERTAÇÃO VERSÃO DIGITAL FINAL.pdf: 1801824 bytes, checksum: fa9f05a820a2698b9f22a84423419651 (MD5) / Made available in DSpace on 2016-08-26T13:34:53Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) DISSERTAÇÃO VERSÃO DIGITAL FINAL.pdf: 1801824 bytes, checksum: fa9f05a820a2698b9f22a84423419651 (MD5) Previous issue date: 2016-02-29 / CAPEs / A presente investigação é instigada por dois pressupostos: 1) compreendendo a grande mídia como instituição integrada, e originada, no sistema capitalista, ela é foco de complexas lutas de/pelo poder para/de influenciar a produção de significados e representações sociais – ou seja, é espaço privilegiado no (re)conhecimento da (in)existência e (in)visibilização de vozes, temas e mundos, assim colaborando na (re)construção social da realidade; 2) na sociedade brasileira, pessoas trans constituem um dos grupos político-identitários que maior marginalização sofre diariamente – de ordem estrutural, muitas das violências transfóbicas/ cissexistas que enfrentam são resultado, e resultam, do/no apagamento de suas vozes, historicamente patologizadas/ criminalizadas pelos saberes ‘oficiais’ (principalmente os construídos pela medicina e ciências psi, reproduzidos nos mais variados espaços públicos e privados). Nossa hipótese é que a mídia é, também, espaço de (re)produção de violências e exclusão de pessoas trans – hipótese essa que ao final afirmamos, e justificamos, como verdadeira. Desta forma, o trabalho tem três objetivos centrais: 1) analisar os discursos produzidos (jan.2014 – jan.2015) por Aqui PE e Jornal de Commercio (versões impressas) acerca de transexualidade(s) e travestilidade(s); 2) através de entrevistas semi-estruturadas com alguns dos sujeitos envolvidos na produção noticiosa (editoras/es dos cadernos analisados), lançar pistas sobre algumas das concepções acerca de transexualidade e travestilidade que circulam nas redações dos jornais, bem como sobre outras variáveis que condicionem a construção noticiosa dos temas; 3) alinhavando teorizações provenientes, principalmente, das Teorias do Jornalismo, Estudos Críticos do Discurso e Estudos Queer, refletir sobre as possibilidades de mudança discursiva, portanto social, acerca da(s) transexualidade(s), travestilidade(s) e, necessária-simultaneamente, cisgeneridade(s). / The investigation is instigated by two assumptions: 1) understanding the mainstream media as an institution integrated, and originated, in the capitalist system, it is the focus of complex struggles of/for power to influence the production of meanings and social representations – i.e., its a privileged space for the recognition of (in)existence and (in)visibility of voices, themes and worlds, thus collaborating in the reconstruction of reality; 2) in Brazilian society, transgender people are one of the political/identity groups that, on daily bases, experience social marginalization – structural, many of the transphobic/ cissexists violence they face are the result, and result, from / in the deletion of their voices, historically pathologized/ criminalized by 'official' knowledge (mostly built by medicine and psychological sciences, reproduced in various public and private spaces). Our hypothesis is that the media is, also, space of (re)production of violence and exclusion of transgender people - a hypothesis that at the end we found to be true. Thus, the work has three main objectives: 1) to analyze the discourses produced (jan.2014 - jan.2015) on Aqui PE and Jornal do Commercio (printed versions) about transsexuality(s); 2) through semi-structured interviews with some of those involved in news production (publishers of the analyzed books), provide clues about some of the conceptions of transsexuality circulating in newsroom and newspapers, as well as other variables that condition the construction of news topics; 3) tacking theories derived mainly from Journalism Theories, Critical Discourse Studies and Queer Studies, reflect on the possibilities of discursive change, therefore social changes, about transgenderity and, necessary- simultaneously, cisgenderity.

Orientador: Lucia Elvira Alvares / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-13T02:44:23Z (GMT). No. of bitstreams: 1 Grade_CarlaVermeulenCarvalho_M.pdf: 54304678 bytes, checksum: 869f67d6a836e256bbd7ee9e15980659 (MD5) Previous issue date: 2009 / Resumo: A proteina Miostatina (tambem conhecida como GDF8) e um membro da superfamilia de crescimento e diferenciacao ß (TGF- ß) e e expressa quase que exclusivamente em musculatura esqueletica, tanto no embriao em desenvolvimento quanto no individuo adulto, onde circula livre pela corrente sanguinea. A Miostatina foi inicialmente identificada em 1997 por MCPHERRON et al. e, desde entao, muitos estudos tem demonstrado seu papel essencial na regulacao do desenvolvimento de musculatura esqueletica de aves e mamiferos. O nocaute genico da Miostatina causa hiperplasia e hipertrofia das fibras musculares, resultando em musculos individuais ate duas vezes maiores do que em animais selvagens. Isso demonstra que a Miostatina e um regulador negativo da deposicao de musculatura esqueletica. A estrutura e a funcao desta proteina sao conservadas em diversas especies, incluindo humanos, onde os niveis de Miostatina circulante no sangue se encontram aumentados durante condicoes de distrofia e na caquexia que acompanha alguns tipos de cancer e a AIDS. Um melhor entendimento dos mecanismos que regem a expressao da Miostatina e essencial para o desenvolvimento de estrategias que possam regular sua atividade durante tais condicoes. No presente trabalho, nos identificamos, com o uso de ferramentas de Bioinformatica, elementos cisregulatorios putativos (promotor e enhancers) que possivelmente regulam a transcricao do gene da Miostatina. Inicialmente foi realizada uma comparacao dos loci do GDF8, incluindo as regioes intergenicas adjacentes, provenientes dos genomas de Humano, Camundongo e Galinha. Essa analise revelou a presenca de diferentes regioes evolutivamente conservadas (RECs) adjacentes a sequencia codificadora desta proteina, sete downstream e uma upstream ao gene. Por terem sido mantidas relativamente conservadas ao longo da evolucao, essas regioes supostamente possuem um papel funcional, possivelmente como elementos cis-regulatorios do gene da Miostatina. Em seguida, com o intuito de entender as funcoes que cada uma dessas regioes possa estar exercendo sobre a regulacao da atividade transcricional do gene da Miostatina, foi realizada uma busca por sitios de ligacao para fatores transcricionais que tenham sido conservados evolutivamente nessas RECs. Muitos sitios conservados foram observados nas sete RECs downstream ao gene da Miostatina, entre eles estao sitios para fatores relacionados ao desenvolvimento de musculatura esqueletica (MyoD, Myogenin, E47, EN1), membros (Pax3, Tbx5) e coracao (Nkx2.5, Pitx2). Juntos, esses dados sugerem uma regulacao modular do gene da Miostatina durante a embriogenese dos vertebrados. A unica REC localizada upstream ao GDF8 representa o promotor minimo putativo deste gene. Essa hipotese e reforcada pela presenca de um sitio de ligacao conservado para a Proteina de Ligacao ao sitio TATA. Com o intuito de validar as hipoteses formuladas com base nas analises de Bioinformatica, no presente trabalho buscamos caracterizar funcionalmente o promotor minimo do gene da Miostatina. Para tanto, a regiao do promotor minimo foi inicialmente clonada em um vetor que nao contem promotor e possui como gene reporter o GFP. Essa construcao de expressao foi entao testada atraves de experimentos de eletroporacao em embrioes de galinha in ovo. A analise dos embrioes eletroporados revelou que a regiao de DNA elegida para as analises funcionais e capaz de dirigir a transcricao do gene reporter, indicando que ela corresponde ao promotor minimo do gene da Miostatina. Alem do sitio TATA, ha, na regiao do promotor, diversos sitios conservados para a ligacao de proteinas envolvidas na via de sinalizacao mediada por cAMP (CREB, ATF, NFY). Esse achado esta de acordo com estudos recentes que demonstram o envolvimento do cAMP na regulacao dos fatores miogenicos Myf5 e MyoD, bem como de Pax3, sugerindo que a atividade do gene da Miostatina tambem possa estar sendo regulada por essa via de sinalizacao. Outras regioes do genoma humano que possuem arquitetura semelhante a observada no promotor da Miostatina foram identificadas, demonstrando que outros genes podem estar sob influencia da mesma via de sinalizacao que regula a atividade do promotor da Miostatina, dentre eles genes envolvidos na miogenese e neurogenese. / Abstract: The Myostatin protein (also known as GDF8) is a member of the transforming growth factor-ß (TGF-ß) superfamily and is expressed almost exclusively in skeletal muscle, both in the embryo and in the adult, where the protein circulates in the blood flow. It was initially identified in 1997 by MCPHERRON et al., and since then many studies have been demonstrating its essential role in the regulation of the development of skeletal muscle from birds and mammals. The knockout of the Myostatin gene causes both hyperplasia and hypertrophy of the skeletal muscle fibers, resulting in muscles twice as big as the wildtype ones, thus showing that Myostatin is a negative regulator of skeletal muscle deposition. The GDF8 structure and function is conserved in many species, including humans where the Myostatin levels are increased during dystrophy conditions and in the cachexia that accompanies some types of cancer and AIDS. A better understanding of the mechanisms that rule the Myostatin expression is essential for the development of strategies that might regulate its activity during such conditions. In this research, we have identified, with the use of bioinformatic tools, the cis-regulatory elements (promoter and enhancers) that regulate the Myostatin gene transcription. We compared the GDF8 loci from human, chicken and mouse and found different evolutionary conserved regions (ECRs), adjacent to the GDF8 coding sequence. Because these intergenic sequences remained relatively conserved throughout evolution, they supposedly have a functional role, possibly as cis-regulatory elements for the Myostatin gene. Our analyses revealed the presence of seven possible enhancers downstream of the GDF8 gene and one conserved region upstream of it. In order to understand the role these regions might have in the regulation of Myostatin's transcription activity, we searched for binding sites that were also evolutionary conserved. Many conserved binding sites were observed in the RECs downstream to the Myostatin gene, and among them are sites for factors related to the development of the skeletal muscle (MyoD, Myogenin, E47, EN1), limbs (Pax3, Tbx5) and heart (Nkx2.5, AREB6, Pitx2). Together, these data suggest a modular regulation of the Myostatin gene during vertebrates' embryogenesis. The only REC observed upstream of the Myostatin locus represents the putative basal gene promoter. This hypothesis is strengthened by the presence of a binding site for the Tata Binding Protein conserved for the studied species. In this research, we aimed at functionally characterizing the Myostatin gene basal promoter. For that purpose, we cloned the studied region in a promoterless vector, which contains GFP as a reporter gene. This expression construct was then tested through in ovo electroporation assays. The analysis of the electroporated embryos revealed that the cloned DNA region is capable of driving the transcription of the reporter gene, which indicates that it truly corresponds to the basal promoter of the Myostatin gene. Moreover, there are conserved binding sites for the CREB and ATF1 transcription factors in the basal promoter, which are activated by the cAMP signaling path. This finding is in agreement with recent studies that demonstrate the involvement of cAMP in the regulation of the myogenic factors Myf5 and MyoD, as well as Pax3, thus suggesting that the activity of the Myostatin gene might be under the influence of this signaling path. Other regions of the human genome that have a similar architecture to the one observed in the Myostatin promoter were identified. This demonstrates that other genes are possibly under the influence of the same signaling path regulating the activity of the Myostatin promoter, among them genes involved in myogenesis and neurogenesis. / Mestrado / Histologia / Mestre em Biologia Celular e Estrutural

Submitted by Gabriela Lopes (gmachadolopesufpel@gmail.com) on 2017-02-08T14:22:01Z No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) dissertação.pdf: 1451490 bytes, checksum: 8db111be8fc2a2a277ed6aa88f84a6e6 (MD5) / Approved for entry into archive by Aline Batista (alinehb.ufpel@gmail.com) on 2017-02-15T16:51:40Z (GMT) No. of bitstreams: 2 dissertação.pdf: 1451490 bytes, checksum: 8db111be8fc2a2a277ed6aa88f84a6e6 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2017-02-15T16:51:40Z (GMT). No. of bitstreams: 2 dissertação.pdf: 1451490 bytes, checksum: 8db111be8fc2a2a277ed6aa88f84a6e6 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2013-06-28 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / As raízes possuem uma grande variedade de funções nas plantas, incluindo absorção de água, nutrientes e suporte estrutural. A combinação de métodos clássicos de genética e melhoramento com tecnologias moleculares de análise genômica abre uma nova perspectiva para a ampliação do conhecimento das bases genéticas e aceleração de programas de melhoramento. A maioria dos conhecimentos sobre as redes gênicas envolvidas no desenvolvimento radicular vem sendo acumuladas na espécie Arabidopsis thaliana, modelo de planta dicotiledônea. O entendimento dos mecanismos envolvidos na regulação da expressão dos genes é essencial para compreender a forma e a função dos sistemas. Os elementos cis-acting são regiões do DNA que atuam como interruptores moleculares envolvidos na regulação da transcrição de uma rede gênica dinâmica. Embora freqüentemente tenham somente cinco a 20 pb de tamanho, os elementos cis-acting são críticos para o entendimento da regulação gênica. O conhecimento destes elementos presentes na região promotora de famílias gênicas, poderá contribuir para a compreensão dos sistemas reguladores da expressão da rede gênica envolvida na formação do sistema radicular. O objetivo desse trabalho é identificar os elementos cis-acting presentes na região promotora de genes de arroz (Oryza sativa subsp japonica cv. Nipponbare) similares aos genes das famílias Argonauta, Cullin e Arade Arabidopsis thaliana. A região promotora dos genes destas famílias no arroz foi investigada quanto à abundância destes elementos. As seqüências foram analisadas utilizando o programa “Signal Scan Search” do portal “Plant Cis-acting Regulatory DNA Elements” (PLACE) para a identificação dos diferentes elementos cis-acting. Foram detectados 96 diferentes elementos, sendo cinco destes (GAREAT (TAACAAR), TGACGTVMAMY (TGACGT), CCAATBOX1 (CCAAT), LECPLEACS2 (TAAAATAT) e SV40COREENHAN (GTGGWWHG), comuns as famílias gênicas Argonauta, Cullin e Ara. / The roots have a large range of functions in plants, including acquisition of water and nutrients, as well as structural support. The combination of classical methods of genetics and breeding with molecular technologies for genomic analysis opens a new perspective to expand the knowledge of the genetic basis and to accelerate breeding programs. The most advanced knowledge regarding gene networks involved in root development has been obtained in the model dicotyledon plant species Arabidopsis thaliana. Understanding the mechanisms involved in regulation of gene expression is essential to predict the form and function of systems. Cis-actingelements are DNA regions that act as molecular switches involved in the regulation of transcription of dynamic gene network. Although often having only five to 20 bp in size, cis-actingelements are critical to the understanding of gene regulation. Knowledge of the cis-acting elements present in the promoter region of gene families, can contribute to the understanding of the expression regulatory systems of these genes and others, involved with the root system. The objective of this study is to identify the cisacting elements present in the upstream region of rice (Oryza sativa subsp japonica cv. Nipponbare) genes, similar to gene families Argonauta, Cullin and Arain Arabidopsis thaliana. The promoter region of these rice gene families were investigated for the abundance of cis-acting elements. The sequences were analyzed using the software “Signal Scan Search” ofthe website “Plant Cis-acting Regulatory DNA Elements” (PLACE) to the identification of different cis-acting elements. It were detected 96 different cis-acting elements, and five of these, (GAREAT (TAACAAR), TGACGTVMAMY (TGACGT), CCAATBOX1 (CCAAT), LECPLEACS2 (TAAAATAT) e SV40COREENHAN (GTGGWWHG) were common to the gene families Argonauta, Cullin andAra.

Industriellement, les problématiques d‘adhésion polymère/métal se rencontrent dans de nombreux secteurs tels que l'industrie automobile, ou les applications aéronautiques et électroniques. Les polyuréthanes (PU) sont fréquemment utilisés comme adhésifs structuraux, et sont obtenus à partir de polyols provenant de la pétrochimie (polyester et polyéther polyols). Cependant, ces produits ont des inconvénients notables sur le plan écologique car ils sont produits à partir de ressources non renouvelables, ils peuvent également générer une pollution de l'environnement, et leurs matières premières de départ sont d‘une part de plus en plus coûteuses et d‘autres part amenées à se raréfier dans les années à venir. Le caoutchouc naturel (NR) est une alternative intéressante aux polyols de synthèse car il est issu d‘une ressource végétale (hévéa), renouvelable et abondante, et également car il présente des propriétés mécaniques intéressantes. De plus, il peut être facilement modifié chimiquement, afin notamment d‘apporter des groupements hydroxyle capables de réagir ensuite avec des fonctions isocyanate pour former un polyuréthane. Dans ce travail, le polyisoprène hydroxytéléchélique (HTPI) ayant une fonctionnalité en hydroxyle de 2 a été synthétisé avec succès par époxydation contrôlée suivie de coupure oxydante de polyisoprène de hautes masses, puis réduction sélective des oligoisoprènes carbonyltéléchéliques obtenus. Ces HTPI de différentes masses molaires (1000-8000 g mol-1) ont été obtenus de façon reproductible. Des modifications chimiques ont été effectuées par époxydation à différents taux (10-60% EHTPI). Les différentes microstructures de ces oligomères ont été mises en évidence par FT-IR, RMN and SEC. Leurs propriétés thermiques ont été déterminées par ATG et DSC. Les propriétés de surface (énergie de surface, microscopie optique) et les propriétés d‘adhésion (test de clivage) de différents matériaux ont été caractérisées. Les échantillons à base de HTPI pur (sans époxyde) présentent un niveau d‘adhésion élevé. Des taux d‘époxydation proches de 30-40% permettent d‘obtenir des performances adhésives intéressantes. D‘autre part, l‘effet de la masse molaire est faible(cependant, une masse molaire plus élevée entraîne globalement une meilleure adhérence). Le niveau d‘adhérence observé est similaire à ceux mesurés pour des adhésifs structuraux utilisés dans l‘industrie automobile ou aéronautique. Le test de clivage est un test d‘adhérence sévère pour un joint adhésif, et les faibles propagations de fissures observées pour certaines formulations permettent d‘escompter des développements industriels prometteurs pour ces nouveaux polymères. / Industrially, metal/polymer adhesion is involved in a wide range of industries such as automotive industry, or aeronautic and electronic applications. Polyurethanes (PU) are frequently used as structural adhesives, and are based from polyols obtained from petrochemical products (polyester and polyether polyols). However, these products have some disadvantages as they are non-renewable resources, they may cause environmental pollution, and they tend to be exhausted in the near future. Natural rubber (NR) is an interesting choice to use as a starting material in PU synthesis, due to the fact that they are renewable source, abundant polymer and they have interesting mechanical properties and can be chemically modified. In this work, hydroxytelechelic polyisoprene (HTPI) having a hydroxyl functionality of 2 was successfully performed via controlled epoxidation and cleavage of high molecular weight polyisoprene, following by a selective reduction reaction of the obtained carbonyltelechelicoligoisoprenes. These HTPI with different molecular weights (1000-8000 g mol-1) were reproducible obtained. Chemical modifications on HTPI were performed by various percentage of epoxidation (10-60%, EHTPI). The different microstructures of these oligomers were evidenced by the characterization techniques FT-IR, NMR, SEC. Their thermal properties were also investigated by TGA and DSC.Surface properties (surface energy, optical microscopy) and adhesion properties (wedge test) of different materials have been characterized. To resume adherence results, pure HTPI samples (without any epoxy group) present a very high adhesion level. Epoxidation degrees close to 30-40% allow to obtain interesting adhesive performance. Elsewhere, the effect of molecular weight is slight (nevertheless, a higher Mn of HTPI induces globally a better adherence). The adherence level is similar to whose measured for structural adhesive used in car or aeronautic industry. The wedge test is a severe adherence test, and the low crack propagation observed for some formulations underlines promising industrial developments for this new polymers.

This dissertation explores the structural modification of indigo into its rare cis form by substitution of a short organic bridge between the two indole nitrogen atoms. The synthesized N,N′-tethered cis-indigoids are assessed for their physicochemical properties in order to understand the effect of the organic tether on cis-indigoid systems. Three literature compounds containing “simple” tether structures are synthesized and subjected to complete characterization. Among them, alkyl group based cis-indigos, 2.5 and 2.6 exhibit similar absorption wavelengths as the parent indigo while oxalyl-tethered indigo 2.7 shows a large hypsochromic shift due to the strong electron-withdrawing nature of the oxalyl group. The electronics of the tether also affected the HOMO and LUMO energy levels; the oxalyl tether lowered both energy levels in comparison to the alkyl tethers. However, the indigoid co-planarity was strongly affected by the ring size from the tether rather than electronics. A new tethered cis-indigo structure type was discovered in reactions involving quinones. This new tether type consists of a 2,2′-dihydroindigo unit connected to the cis-indigo backbone through a central C–C bond of the former. Incorporation of the quinone moiety was observed in 3.2 where the final structure is comprised of two molecules of indigo and one naphthoquinone, while structures of 3.3 and 3.4 contained two indigoid units only. Investigation by CV revealed the strength of the quinone altered the early reaction intermediates. These dimeric cis-indigos show a small hypsochromic shift in the absorption compared to the parent indigo and relatively planar cis-indigoid backbone. 3.2 demonstrates rich redox behaviours. 3.3 and 3.4 display dynamic behaviours observed by solution VT 1H NMR spectroscopy. Oxalyl-tethered cis-Nindigo 4.6a and cis-indigo monoimine 4.7a were synthesized and compared with the indigoid counterpart 2.7 to examine the influence of the arylimine groups. Absorption wavelengths of the imine group containing species depended strongly on the electronics of the tether, yet the number of imine groups affected redox potentials. Protonation of 4.6a (to give H+oxalyl Nindigo 4.6aH+) causes a bathochromic shift in the absorption and allows for much easier reduction. The acidity (pKa) of 4.6aH+ is estimated between 3.6 – 4.46 in DMSO. A protonated cis-Nindigo derivative 4.9aH+ was obtained via reduction of the oxalyl group and is compared with 4.6aH+. / Graduate / 2022-08-13

Gene regulation lies at the heart of most biological processes and transcription factors are the key molecules that control tissues specific gene expression. In higher eukaryotes transcription factors control gene expression by binding regulatory DNA segments called cis-regulatory modules (CRMs). The increasing number of sequenced genomes of multicellular eukaryotes along with high-throughput methods such as whole genome microarray expression data allows for systematic characterization of the CRMs that control gene expression. A first step towards understanding gene regulation is the identification of the regulatory elements present in the genome. We take advantage of the large database of spatio-temporal patterns of gene expression in D. melanogaster embryogenesis to identify sets of developmentally co-expressed genes. We developed a computational method that identifies DNA binding sites for transcription factors from families of co-regulated genes that are expressed during Drosophila embryo development. This method discovers over-represented motifs in a set of co-regulated genes using the exhaustive motif enumeration technique. Clustering the predicted motifs identifies the CRMs, which assist in translating a combinatorial code of TF inputs into a specific gene expression output. The predicted CRMs were verified experimentally by searching the whole genome for the predicted CRMs and establishing expression pattern of the genes that are associated with these CRMs. It is well know that the gene expression is substantially controlled through CRMs and those key regulatory sequences are conserved in related species. The conservation of CRMs can be studied by comparing the related genomes and alignment methods are widely used computational tools for comparing the sequences. However, in distantly related species the CRM sequences are simply not align able. To identify the similar CRMs in distantly related species we developed a non-alignment based method for discovering similar CRMs in related species. This method is based on word frequencies where the given sequences are compared using Poisson based metric. When starting with a set of CRMs involved in Drosophila early embryo development, we show here that our non-alignment method successfully detects similar CRMs in distantly related species ( D. ananassae, D. pseudoobscura, D. willisoni, D. mojavensis, D. virilis, D. grimshawi ). This method proved efficient in discriminating the functional CRMs from the non-functional ones.

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