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CAF-1 p150 and Ki-67 Regulate Nuclear Structure Throughout the Human Cell CycleMatheson, Timothy D. 09 January 2017 (has links)
The three-dimensional organization of the human genome is non-random in interphase cells. Heterochromatin is highly clustered at the nuclear periphery, adjacent to nucleoli, and near centromeres. These localizations are reshuffled during mitosis when the chromosomes are condensed, nucleoli disassembled, and the nuclear envelope broken down. After cytokinesis, heterochromatin is re-localized to the domains described above. However, the mechanisms by which this localization is coordinated are not well understood. This dissertation will present evidence showing that both CAF-1 p150 and Ki-67 regulate nuclear structure throughout the human cell cycle.
Chromatin Assembly Factor 1 (CAF-1) is a highly conserved three-subunit protein complex which deposits histones (H3/H4)2 heterotetramers onto replicating DNA during S-phase of the cell cycle. The N-terminal domain of the largest subunit of CAF-1 (p150N) is dispensable for histone deposition, and instead regulates the localization of specific loci (Nucleolar-Associated Domains, or “NADs”) and several proteins to the nucleolus during interphase. One of the proteins regulated by p150N is Ki-67, a protein widely used as a clinical marker of cellular proliferation. Depletion of Ki-67 decreases the association of NADs to the nucleolus in a manner similar to that of p150. Ki-67 is also a fundamental component of the perichromosomal layer (PCL), a sheath of proteins that surrounds all condensed chromosomes during mitosis. A subset of p150 localizes to the PCL during mitosis, and depletion of p150 disrupts Ki-67 localization to the PCL. This activity was mapped to the Sumoylation Interacting Motif (SIM) within p150N, which is also required for the localization of NADs and Ki-67 to the nucleolus during interphase. Together, these studies indicate that p150N coordinates the three-dimensional arrangement of both interphase and mitotic chromosomes via Ki-67.
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Market Uptake of Sustainable Aviation Fuel : An Investigation of how Swedavia can Contribute to Market Uptake of SAFNordström, Elin January 2021 (has links)
Previous studies have claimed a deadlock between fuel producers, airlines and policymakers when it comes to the market uptake of Sustainable Aviation Fuel (SAF). This report investigates the market from a stakeholder perspective, including producers, distributors, purchasers and logistic holders. Obstacles in terms of market maturity are identified as mainly the price gap between Conventional Aviation Fuel (CAF) and SAF. This is due to many things, whereas one being the detachment between travellers and the usage of SAF. It is also the current blend restriction of a maximum of 50 %, as well as extensive and expensive certification processes. Currently, there are few, but a lack of policies and incentives directed towards SAF and a lack of coherency between those that exist and are being implemented. Stakeholders agree that there is a need for developing these, as well as a need for additional financial support during stages of developing production facilities. When it comes to procurement of SAF, stakeholders agree upon that flexibility within different areas are advantageous for the market uptake of SAF, and in most regards, the procurement made by Swedavia is indeed flexible. An important aspect identified in the analysis of Swedavia’s procurement is the use of a business model which reduces the price gap between CAF and SAF for airlines and enabling Swedavia to claim the CO2 reduction of corporate travel. Multiple corporates joined the previous procurement of SAF for corporate travel, and Swedavia holds great opportunities to continue to contribute to the market uptake of SAF by expanding the concept. Multiple opportunities also lie in expanding the business concept to reach all travellers. Concluding, it is established that the current route, mainly referring to the procedure of procurement, is effective and carries great opportunity for further implementation and development. / <p>2021-06-04</p>
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Mass spectrometry as a tool to dissect the role of chromatin assembly factors in regulating nucleosome assemblyGharib, Marlène 12 1900 (has links)
L'assemblage des nucléosomes est étroitement couplée à la synthèse des histones ainsi qu’à la réplication et la réparation de l’ADN durant la phase S. Ce processus implique un mécanisme de contrôle qui contribue soigneusement et de manière régulée à l’assemblage de l’ADN en chromatine. L'assemblage des nucléosomes durant la synthèse de l’ADN est crucial et contribue ainsi au maintien de la stabilité génomique. Cette thèse décrit la caractérisation par spectrométrie de masse(SM) des protéines jouant un rôle critique dans l’assemblage et le maintien de la structure chromatinienne. Plus précisément,
la phosphorylation de deux facteurs d’assemblage des nucléosome, le facteur CAF-1, une chaperone d’histone qui participe à l'assemblage de la chromatine spécifiquement couplée à la réplication de l'ADN, ainsi que le complexe protéique Hir, jouant de plus un rôle important dans la régulation transcriptionelle des gènes d’histones lors de la progression normale du cycle cellulaire et en réponse aux dommages de l'ADN, a été examiné.
La caractérisation des sites de phosphorylation par SM nécéssite la séparation des protéines par éléctrophorèse suivi d’une coloration a l’argent. Dans le chapitre 2, nous demontrons que la coloration à l’argent induit un artéfact de sulfatation. Plus précisément, cet artéfact est causé par un réactif spécifiquement utilisé lors de la coloration. La sulfatation présente de fortes similitudes avec la phosphorylation. Ainsi, l’incrément de masse observé sur les peptides sulfatés et phosphorylés (+80 Da) nécéssite des instruments offrant une haute résolution et haute précision de masse pour différencier ces deux modifications.
Dans les chapitres 3 et 4, nous avons d’abord démontré par SM que Cac1, la plus
grande sous-unité du facteur CAF-1, est cible de plusieurs sites de phosphorylation. Fait intéréssant, certains de ces sites contiennent des séquences consensus pour les kinases Cdc7-Dbf4 et CDKs. Ainsi, ces résultats fournissent les premières évidences que CAF-1 est potentiellement régulé par ces deux kinases in vivo. La fonction de tous les sites de phosphorylation identifiés a ensuite été évaluée. Nous avons démontré que la phosphorylation de la Ser-503, un site consensus de la DDK, est essentielle à la répréssion transcriptionelle des gènes au niveau des télomères. Cependant, cette phosphorylation ne semble pas être nécéssaire pour d’autres fonctions connues de CAF-1, indiquant que le blocage de la phsophorylation de Cac1 Ser-503 affecte spécifiquement la fonction de CAF-1 aux structures hétérochromatiques des télomères. Ensuite, nous avons identifiés une intéraction physique entre CAF-1 et Cdc7-Dbf4. Des études in vitro ont également
demontré que cette kinase phosphoryle spécifiquement Cac1 Ser-503, suggérant un rôle potential pour la kinase Cdc7-Dbf4 dans l’assemblage et la stabilité de la structure
hétérochromatique aux télomères. Finalement, les analyses par SM nous ont également permi de montrer que la sous-unité Hpc2 du complexe Hir est phosphorylée sur plusieurs
sites consensus des CDKs et de Cdc7-Dbf4. De plus, la quantification par SM d’un site
spécifique de phosphorylation de Hpc2, la Ser-330, s’est révélée être fortement induite suite à l’activation du point de contrôle de réplication (le “checkpoint”) suite au dommage a l’ADN. Nous montrons que la Ser-330 de Hpc2 est phopshorylée par les kinases de point de contrôle de manière Mec1/Tel1- et Rad53-dépendante. Nos données préliminaires suggèrent ainsi que la capacité du complex Hir de réguler la répréssion transcriptionelle des gènes d'histones lors de la progression du cycle cellulaire normal et en réponse au dommage de l'ADN est médiée par la phosphorylation de Hpc2 par ces deux kinases.
Enfin, ces deux études mettent en évidence l'importance de la spectrométrie de masse dans la caractérisation des sites de phosphorylation des protéines, nous permettant ainsi de comprendre plus précisement les mécanismes de régulation de l'assemblage de la chromatine et de la synthèse des histones. / Nucleosome assembly entails a controlled mechanism that is tightly coupled to DNA and histone synthesis during DNA replication and repair in S-phase. Importantly, this contributes to the prompt and carefully orchestrated assembly of newly replicated DNA
into chromatin, which is essential for the maintenance of genomic integrity. This thesis
describes the mass spectrometric characterization of proteins critical in the regulation of nucleosome assembly behind the replication fork and chromatin structure. More specifically, the phosphorylation of Chromatin Assembly Factor 1 (CAF-1), a nucleosome assembly factor that uniquely functions during replication-coupled de novo nucleosome assembly in S-phase and the Hir protein complex, a second nucleosome assembly factor that also contributes to the transcriptional regulation of histone genes during normal cell cycle progression and in response to DNA damage, was examined.
We first demonstrated that characterization of protein phosphorylation by mass
spectrometry (MS), which often relies on the separation of proteins by gel electrophoresis
followed by silver staining for visualization, should be given careful considerations. In chapter 2, we report a potential pitfall in the interpretation of phosphorylation modifications due to the artifactual sulfation of serine, threonine and tyrosine residues caused by a specific reagent used during silver staining. Sulfation and phosphorylation both impart an
80 Da addition of these residues making them distinguishable only with MS systems offering high resolution and high mass accuracy capabilities.
Chapter 3 and 4 present the MS characterization of in vivo phosphorylation
occurring on CAF-1 and Hir proteins, respectively. We first demonstrated that Cac1, the largest subunit of CAF-1, is phosphorylated on several novel residues containing the consensus sequences recognized by either Cdc7-Dbf4 (DDK) or cyclin-dependent kinases(CDKs). These results have provided the first evidence that CAF-1 is regulated by these two kinases in vivo. The function of all identified Cac1 phosphorylation sites was then assessed. In vivo phenotypic studies showed that the specific phosphorylation of Ser-503, a Cac1 DDK-like site identified in our study, is essential for heterochromatin-mediated
telomeric silencing. Cac1-Ser-503 did not appear to be required for other known functions of CAF-1, including DNA damage resistance and mitotic chromosom segregation,
indicating that blocking Cac1 phosphorylation on Ser-503 sepcifically cripples CAF-1 function at telomeres. Next, biochemical purifications identified a physical interaction between CAF-1 and Cdc7-Dbf4. Consistent with this physical interaction data, in vitro kinase assay studies showed that Cdc7-Dbf4 specifically phosphorylates Cac1 Ser-503 thereby uncovering a novel role for Cdc7-Dbf4 in heterochromatin assembly and/or stability that is potentially mediated through CAF-1. Finally, MS analysis also showed that the Hpc2 subunit of the Hir protein complex is phosphorylated on several CDK- and DDK-like consensus sites. Furthermore, MS quantification of a specific phosphorylation site,Hpc2 Ser-330, was shown to be highly induced following the activation of the DNA
damage checkpoint in response to DNA damage. We show that Hpc2 Ser-330 is phopshorylated by checkpoint kinases in a Mec1/Tel1- and Rad53-dependent manner. Our preliminary data suggest that the ability of the Hir protein complex to regulate the transcriptional repression of histone genes during normal cell cycle progression and in response to DNA damage is mediated through the regulated phosphorylation of Hpc2 by these kinases.
Finally, these two studies highlight the importance of mass spectrometry in
characterizing protein phosphorylation events, which has yielded novel insights into the regulation of chromatin assembly by CAF-1 and histone synthesis mediated by Hir
proteins.
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Využití systémů řízení kvality při autoevaluaci středních škol ve Středočeském kraji / The Use of Quality Management Systems for Self evaluation of Secondary Schools in the Central Bohemian RegionVelflová, Romana January 2010 (has links)
The final issue deals with the introduction of Quality Management Systems in Central Bohemian high schools. The theoretical part describes principles of the Quality Management and more closely presents ISO 9001 standard, EFQM Excellence Model, CAF Model and CQAF Model. The research part determines opinion of school headmasters in Decree No 15/2005 Coll., that specifies requirements for the obligatory self-evaluation of schools. In next chapters through a survey and an analysis of school web pages is examined practice of Quality Management Systems implementation in high schools in Central Bohemia -- current status, awareness of head masters, demand and expectations. The last chapter offers a definite experience with the introduction of Quality Management Systems through additional interviews with school headmasters.
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Mass spectrometry as a tool to dissect the role of chromatin assembly factors in regulating nucleosome assemblyGharib, Marlène 12 1900 (has links)
L'assemblage des nucléosomes est étroitement couplée à la synthèse des histones ainsi qu’à la réplication et la réparation de l’ADN durant la phase S. Ce processus implique un mécanisme de contrôle qui contribue soigneusement et de manière régulée à l’assemblage de l’ADN en chromatine. L'assemblage des nucléosomes durant la synthèse de l’ADN est crucial et contribue ainsi au maintien de la stabilité génomique. Cette thèse décrit la caractérisation par spectrométrie de masse(SM) des protéines jouant un rôle critique dans l’assemblage et le maintien de la structure chromatinienne. Plus précisément,
la phosphorylation de deux facteurs d’assemblage des nucléosome, le facteur CAF-1, une chaperone d’histone qui participe à l'assemblage de la chromatine spécifiquement couplée à la réplication de l'ADN, ainsi que le complexe protéique Hir, jouant de plus un rôle important dans la régulation transcriptionelle des gènes d’histones lors de la progression normale du cycle cellulaire et en réponse aux dommages de l'ADN, a été examiné.
La caractérisation des sites de phosphorylation par SM nécéssite la séparation des protéines par éléctrophorèse suivi d’une coloration a l’argent. Dans le chapitre 2, nous demontrons que la coloration à l’argent induit un artéfact de sulfatation. Plus précisément, cet artéfact est causé par un réactif spécifiquement utilisé lors de la coloration. La sulfatation présente de fortes similitudes avec la phosphorylation. Ainsi, l’incrément de masse observé sur les peptides sulfatés et phosphorylés (+80 Da) nécéssite des instruments offrant une haute résolution et haute précision de masse pour différencier ces deux modifications.
Dans les chapitres 3 et 4, nous avons d’abord démontré par SM que Cac1, la plus
grande sous-unité du facteur CAF-1, est cible de plusieurs sites de phosphorylation. Fait intéréssant, certains de ces sites contiennent des séquences consensus pour les kinases Cdc7-Dbf4 et CDKs. Ainsi, ces résultats fournissent les premières évidences que CAF-1 est potentiellement régulé par ces deux kinases in vivo. La fonction de tous les sites de phosphorylation identifiés a ensuite été évaluée. Nous avons démontré que la phosphorylation de la Ser-503, un site consensus de la DDK, est essentielle à la répréssion transcriptionelle des gènes au niveau des télomères. Cependant, cette phosphorylation ne semble pas être nécéssaire pour d’autres fonctions connues de CAF-1, indiquant que le blocage de la phsophorylation de Cac1 Ser-503 affecte spécifiquement la fonction de CAF-1 aux structures hétérochromatiques des télomères. Ensuite, nous avons identifiés une intéraction physique entre CAF-1 et Cdc7-Dbf4. Des études in vitro ont également
demontré que cette kinase phosphoryle spécifiquement Cac1 Ser-503, suggérant un rôle potential pour la kinase Cdc7-Dbf4 dans l’assemblage et la stabilité de la structure
hétérochromatique aux télomères. Finalement, les analyses par SM nous ont également permi de montrer que la sous-unité Hpc2 du complexe Hir est phosphorylée sur plusieurs
sites consensus des CDKs et de Cdc7-Dbf4. De plus, la quantification par SM d’un site
spécifique de phosphorylation de Hpc2, la Ser-330, s’est révélée être fortement induite suite à l’activation du point de contrôle de réplication (le “checkpoint”) suite au dommage a l’ADN. Nous montrons que la Ser-330 de Hpc2 est phopshorylée par les kinases de point de contrôle de manière Mec1/Tel1- et Rad53-dépendante. Nos données préliminaires suggèrent ainsi que la capacité du complex Hir de réguler la répréssion transcriptionelle des gènes d'histones lors de la progression du cycle cellulaire normal et en réponse au dommage de l'ADN est médiée par la phosphorylation de Hpc2 par ces deux kinases.
Enfin, ces deux études mettent en évidence l'importance de la spectrométrie de masse dans la caractérisation des sites de phosphorylation des protéines, nous permettant ainsi de comprendre plus précisement les mécanismes de régulation de l'assemblage de la chromatine et de la synthèse des histones. / Nucleosome assembly entails a controlled mechanism that is tightly coupled to DNA and histone synthesis during DNA replication and repair in S-phase. Importantly, this contributes to the prompt and carefully orchestrated assembly of newly replicated DNA
into chromatin, which is essential for the maintenance of genomic integrity. This thesis
describes the mass spectrometric characterization of proteins critical in the regulation of nucleosome assembly behind the replication fork and chromatin structure. More specifically, the phosphorylation of Chromatin Assembly Factor 1 (CAF-1), a nucleosome assembly factor that uniquely functions during replication-coupled de novo nucleosome assembly in S-phase and the Hir protein complex, a second nucleosome assembly factor that also contributes to the transcriptional regulation of histone genes during normal cell cycle progression and in response to DNA damage, was examined.
We first demonstrated that characterization of protein phosphorylation by mass
spectrometry (MS), which often relies on the separation of proteins by gel electrophoresis
followed by silver staining for visualization, should be given careful considerations. In chapter 2, we report a potential pitfall in the interpretation of phosphorylation modifications due to the artifactual sulfation of serine, threonine and tyrosine residues caused by a specific reagent used during silver staining. Sulfation and phosphorylation both impart an
80 Da addition of these residues making them distinguishable only with MS systems offering high resolution and high mass accuracy capabilities.
Chapter 3 and 4 present the MS characterization of in vivo phosphorylation
occurring on CAF-1 and Hir proteins, respectively. We first demonstrated that Cac1, the largest subunit of CAF-1, is phosphorylated on several novel residues containing the consensus sequences recognized by either Cdc7-Dbf4 (DDK) or cyclin-dependent kinases(CDKs). These results have provided the first evidence that CAF-1 is regulated by these two kinases in vivo. The function of all identified Cac1 phosphorylation sites was then assessed. In vivo phenotypic studies showed that the specific phosphorylation of Ser-503, a Cac1 DDK-like site identified in our study, is essential for heterochromatin-mediated
telomeric silencing. Cac1-Ser-503 did not appear to be required for other known functions of CAF-1, including DNA damage resistance and mitotic chromosom segregation,
indicating that blocking Cac1 phosphorylation on Ser-503 sepcifically cripples CAF-1 function at telomeres. Next, biochemical purifications identified a physical interaction between CAF-1 and Cdc7-Dbf4. Consistent with this physical interaction data, in vitro kinase assay studies showed that Cdc7-Dbf4 specifically phosphorylates Cac1 Ser-503 thereby uncovering a novel role for Cdc7-Dbf4 in heterochromatin assembly and/or stability that is potentially mediated through CAF-1. Finally, MS analysis also showed that the Hpc2 subunit of the Hir protein complex is phosphorylated on several CDK- and DDK-like consensus sites. Furthermore, MS quantification of a specific phosphorylation site,Hpc2 Ser-330, was shown to be highly induced following the activation of the DNA
damage checkpoint in response to DNA damage. We show that Hpc2 Ser-330 is phopshorylated by checkpoint kinases in a Mec1/Tel1- and Rad53-dependent manner. Our preliminary data suggest that the ability of the Hir protein complex to regulate the transcriptional repression of histone genes during normal cell cycle progression and in response to DNA damage is mediated through the regulated phosphorylation of Hpc2 by these kinases.
Finally, these two studies highlight the importance of mass spectrometry in
characterizing protein phosphorylation events, which has yielded novel insights into the regulation of chromatin assembly by CAF-1 and histone synthesis mediated by Hir
proteins.
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AVALIAÇÃO INSTITUCIONAL DE IES: REFLEXÕES SOBRE COMPATIBILIDADE DE TRÊS MODELOS DE AUTO AVALIAÇÃO SINAES, FNQ e CAF / INSTITUTIONAL EVALUATION OF IES: REFLECTIONS ON COMPATIBILITY OF THREE MODELS SELF ASSESSMENT - SINAES, FNQ and CAFNascimento, Fabio Redin do 21 February 2013 (has links)
The Common Assessment Framework (CAF or Commom Assessment Framework), used in the education sector in European countries and the assessment tool of the National Quality Foundation (FNQ) named Model Management Excellence (MEG) used in public and private organizations in Brazil are assessment models of organizational performance, specially developed to help organizations apply the techniques of Total Quality Management (TQM) in order to better their performance levels and their management. However, the study lies in the general assessment of the quality management of Higher Education Institutions (HEIs) in Brazil based on the external evaluation component of the National Higher Education Evaluation (SINAES). Had the motivation to empirical perception about the difficulties that, in general, undergraduate courses face in adopting a system of quality management and implementation of a system with features evaluative management and continuous improvement. This thesis aims to propose the integration of Model Management Excellence Foundation National Quality - FNQ with the Common Assessment Framework - CAF, making a conversãoBrasil / Europe. Using the method of action research as directed qualitative research strategy, the study sought to reconcile the two models and processes your criteria with indicators of the National Assessment of Higher Education SINAES, generating a guidance booklet called SIMECA. This work will serve as support to managers of undergraduate seeking quality assurance and support for the IES rumen toward excellence, fulfilling the main objective of this work. For future research, it is recommended to deepen, through case studies, the study demonstrated the application. / A Estrutura Comum de Avaliação (Commom Assessment Framework ou CAF), usada no setor da educação em países europeus e o instrumento de avaliação da Fundação Nacional da Qualidade (FNQ) denominado Modelo de Excelência da Gestão (MEG) utilizada em organizações públicas e privadas do Brasil são modelos de avaliação de desempenho organizacional, especialmente desenvolvidos para ajudar as organizações aplicarem as técnicas da Gestão da Qualidade Total (TQM) a fim de melhorar os seus níveis de desempenho e da sua gestão. Contudo, o estudo situa-se no quadro geral da avaliação da gestão da qualidade das Instituições de Educação Superior Privada (IESP) no Brasil tendo como base o componente de avaliação externa do Sistema Nacional de Avaliação da Educação Superior (SINAES). Teve como motivação a percepção empírica acerca das dificuldades que, de modo geral, as IES enfrentam na adoção de um sistema de gestão da qualidade e na implementação de um sistema avaliativo com características gerenciais e de melhoria continua. A presente dissertação visa propor a integração do Modelo de Excelência da Gestão da Fundação Nacional da Qualidade FNQ com a Estrutura Comum de Avaliação CAF, fazendo uma conversão Brasil / Europa. Com a utilização do método de pesquisa qualitativa direcionada como estratégia de pesquisa, o trabalho buscou a compatibilização dos dois modelos e seus critérios processos com os indicadores do Sistema Nacional de Avaliação da Educação Superior SINAES, gerando uma cartilha de orientação denominada SIMECA. O presente trabalho servirá como apoio aos gestores educacionais que buscam a garantia da qualidade além do suporte para que as IES rumem em direção da excelência, cumprindo o objetivo principal deste trabalho. Para futuras pesquisas, recomenda-se aprofundar, por meio de estudo de casos, a aplicação do estudo demonstrado.
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El uso del valenciano, la actitud hacia la lengua y la destreza escrita : Un estudio sociolingüístico sobre el dominio del valenciano de jóvenes alicantinos / The use of Valencian language, the attitude towards the language and the written proficiency : A sociolinguistic study of the domain of Valencian of young AlicantinosHarnafi, Amina January 2016 (has links)
Los jóvenes de Alicante viven en una sociedad donde coexisten dos lenguas, el castellano y el valenciano. La lengua valenciana es utilizada por jóvenes de Alicante tanto en ámbitos formales como informales. Los jóvenes hablantes de valenciano sienten un lazo identificativo con la lengua que se rige por las actitudes de éstos. La actitud lingüística conlleva a una identificación lingüística del hablante, la cual es una construcción social que se rige por las normas que rodean al hablante. La identificación bilingüe podría, por lo tanto, ser influenciada e influenciar a su vez a la competencia lingüística. El propósito del presente estudio es investigar en qué situaciones y contextos es usado el valenciano; con el objetivo de estudiar la relación entre el grado de bilingüismo, la actitud lingüística y el nivel de complejidad, corrección y fluidez (CAF) en la expresión escrita del valenciano. Partimos de la hipótesis de que el valenciano tiene, para la generación joven de Alicante, una función social y que la actitud hacia la lengua es positiva, lo que a su vez se refleja en la destreza escrita. Para comprobar nuestra hipótesis nos hemos basado en un cuestionario de hábitos sociales, un test de nivel de valenciano y dos redacciones, una escrita en castellano y una en valenciano. Con la participación de 59 jóvenes alicantinos, llegamos a la conclusión de que el valenciano es usado tanto en ámbitos formales como informales y que la actitud hacia la lengua es positiva, y que consideran importante dominar y preservar el valenciano. Además, los participantes han presentado un nivel alto de destreza en el valenciano escrito, mostrando complejidad y fluidez en la expresión escrita. Por otro lado, la corrección es más elevada en el castellano. También se ha podido ver que la identificación de los participantes está relativamente correlacionada con la competencia lingüística. / The young people of Alicante live in society where two languages, Castilian and Valencian, coexist. The Valencian language is used by young people from Alicante in both formal and informal settings. Young speakers of Valencian feel a linguistic identification with the language, which is governed by social norms. Bilingual identification could, therefore, be influenced by and influence linguistic competence. The purpose of this study is to examine in which situations and contexts Valencian is used with the aim of evaluating the relationship between the degree of bilingualism, language attitude and level complexity, accuracy and fluency (CAF) in the written Valencian of young people of Alicante. We hypothesized that Valencian has, for the young generation of Alicante, a social function and the attitude towards the language is positive, which in turn is reflected in writing skills. To test our hypothesis we have used a questionnaire of social habits, a diagnostic test in Valencian and two essays, written in Castilian and Valencian. With the participation of 59 young Alicantinos, we conclude that Valencian is used in both formal and informal settings, that the attitude towards the language is positive and that it is considered important to master and preserve the Valencian language. In addition, the participants have proficiency in the Valencian written language, showing complexity and fluency in written expression. On the other hand, their accuracy is higher in Castilian. It has also been seen that the linguistic identification of the participants is relatively correlated to linguistic competence.
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Regulation of replication dependent nucleosome assemblyGopinathan Nair, Amogh 04 1900 (has links)
Chez les cellules humaines, environ 2 mètres d'ADN est compacté dans le noyau cellulaire par la formation d'une structure nucléoprotéique appelée chromatine. La chromatine est composée d'ADN enroulé à la surface d'un octamère de core histones pour former une structure appelée nucléosome. La structure de la chromatine doit être altérée afin d'accéder à l'information génétique pour sa réplication, sa réparation et sa transcription. La duplication de la chromatine lors de la phase S est cruciale pour la prolifération et la survie des cellules. Cette duplication de la chromatine requière une ségrégation des histones parentales, mais aussi une déposition d'histones néo-synthétisées sur l'ADN. Ces deux réactions résultent en formation de chromatine dès qu'une quantité suffisante d'ADNest générée par la machinerie de réplication. De plus, en raison de conditions intrinsèques et extrinsèques, la machinerie de réplication est souvent confrontée à de nombreux obstacles, sous la forme de lésions à l'ADN qui interfèrent avec la réplication de l'ADN. Sous ces conditions, l'assemblage de nucléosomes et la synthèse d'histones sont étroitement régulées afin d'éviter la production d'un excès d'histones et leurs nombreuses conséquences nuisibles à la cellule.
"Chromatin Assembly Factor 1" (CAF-1) est responsable de la déposition initiale des molécules d'H3 et H4 derrière les fourches de réplication. Pour permettre sa fonction d'assemblage de chromatine, CAF-1 est localisée aux fourches de réplication en vertue de sa liaison à une protéine appelée Proliferating Cell Nuclear Antigen (PCNA). Cependant, le mécanisme moléculaire par lequel CAF-1 exerce sa function demeure mal compris.
Dans le deuxième chapitre de ma thèse, j'ai exploré comment CAF-1 se lie à PCNA d'une manière distincte des nombreux autres partenaires de PCNA. Grâce à nos collaborateurs, des études de crystallographie ont démontré que CAF-1 se lie à PCNA grâce à une interaction non-canonique entre le "PCNA Interaction Peptide" (PIP) de CAF-1 et une interaction de type cation-pi (π). Nous avons aussi montré qu'une substitution d'un seul acide aminé, unique au PIP de CAF-1, abolit son interaction avec PCNA et sa capacité d'assemblage de nuclésomes. Nous avons aussi montré que le PIP de CAF-1 est situé à l'extrémité C-terminale d'une très longue hélice alpha qui est conservée à travers l'évolution parmi de nombreux homologues de CAF-1. Nos études biophysiques ontmontré que cette longue hélice alpha forme des structures oligomériques de type "coiled-coil", ce qui suggère certains mécanismes pour dédier un anneau de PCNA à l'assemblage de chromatine et ce, en dépit des nombreux intéracteurs de PCNA présents aux fourches de réplication.
Dans le troisième chapitre de ma thèse, nos collaborateurs et moi-même avons étudié les mécanismes moléculaires par lesquels les cellules parviennent à maintenir un équilibre délicat entre la synthèse d'ADN et la synthèse d'histones et ce, même en présence de lésions à l'ADN qui interfèrent avec la réplication. Chez Saccharomyces cerevisiae, nous avons montré que les kinases de réponse au dommage à l'ADN, Mec1/Tel1 et Rad53, inhibent la transcription des gènes d'histones en réponse aux liaisons à l'ADN qui interfèrent avec la réplication. Nous avons montré que la répression des gènes d'histones induite par le dommage à l'ADN est médiée par une phosphorylation extensive de Hpc2, l'une des sous-unités du complexe "Histone Gene Repressor" (HIR). Hpc2 contient un domaine qui se lie à l'histone H3. À partir de la structure d'Hpc2, nous avons généré des mutants qui, d'après la structure, sont incapables de se lier à l'histone H3. Nos résultats montrent que l'accumulation d'histones en excès provoquée par le dommage à l'ADN entraîne la phosphorylation d'Hpc2 and la liaison de l'excès d'histone H3 à Hpc2. Ces résultats suggèrent que la répression transcriptionnelle des gènes d'histones induite par le dommage à l'ADN est médiée, du moins en partie, par une simple rétroaction négative impliquant la liaison des histones en excès à la sous-unité Hpc2 du complexe HIR. / In human cells, roughly 2 meters of DNA is compacted into the cell nucleus by the formation of a nucleoprotein complex called chromatin. Chromatin is composed of DNA wrapped around an octamer of core histones to form so-called nucleosomes. Chromatin structure needs to be altered to access genetic information for processes like replication, repair and transcription. Duplication of chromatin during S phase is vital for cell proliferation and viability. Chromatin duplication requires segregation of parental histones, but also deposition of newly synthesized histones onto DNA. This process results in packaging all of the synthesized DNA with histones to form nucleosomes as soon as enough nascent DNA has emerged from the replication machinery. Moreover, as a result of intrinsic and extrinsic conditions, the replication machinery often encounters DNA lesions that impede the continuous synthesis of DNA. Under these conditions, nucleosome assembly and histone synthesis are tightly regulated to prevent the production of an excess of histone proteins and their deleterious consequences.
Chromatin Assembly Factor-1 (CAF-1) performs the initial step in chromatin assembly by depositing newly synthesized histone H3-H4 molecules behind replication forks. In order to perform its chromatin assembly function, CAF-1 localizes to DNA replication forks by binding directly to a protein known as the Proliferating Cell Nuclear Antigen (PCNA). However, the exact molecular mechanism by which this is achieved remains poorly understood.
Through the second chapter of my thesis, I have explored how CAF-1 binds PCNA in a manner that is distinct from the numerous other binding partners of PCNA. With the help of our collaborators, crystallographic studies demonstrated that CAF-1 binds to PCNA by virtue of a non-canonical PCNA interaction peptide (PIP) and a cation-pi (π) interaction. We have also shown that a single amino acid substitution, unique to the PIP of CAF-1, disrupts its binding to PCNA and chromatin assembly activity. We found that the CAF-1 p150 PIP resides at the extreme C-terminus of a long alpha helix that is evolutionarily conserved among numerous homologues of CAF-1. Our biophysical studies showed that this long alpha-helix is capable of forming higher-order coiled coils, which suggests mechanisms to dedicate one PCNA ring for chromatin assembly despite the presence of multiple PCNA interactors at replication forks.
In the third chapter of this thesis, our collaborators and I have addressed the crucial molecular mechanisms by which cells maintain a delicate balance between DNA and histone synthesis despite the presence of DNA lesions that interfere with replication. In Saccharomyces cerevisiae, we showed that the DNA damage response kinases Mec1/Tel1 and Rad53 inhibit histone gene transcription when DNA lesions block DNA replication. We also showed that this repression is mediated by phosphorylation of the Hpc2 subunit of the Histone Gene Repressor complex (HIR). Hpc2 contains a domain that directly binds to histone H3. Interestingly, structure-based mutants of Hpc2 predicted to be incapable of binding H3 are defective in DNA damage-induced transcriptional repression of histone genes in response to DNA damage during replication. Our results indicate that the accumulation of excess histones caused by DNA damage during S phase triggers extensive phosphorylation of Hpc2 and binding of excess H3 to Hpc2. This suggests that DNA damage-induced repression of histone genes is mediated, at least in part, by a simple negative feedback triggered by binding of excess histones to the Hpc2 subunit of the HIR complex.
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A cidade de S?o Paulo de 1870 a 1930: caf?, imigrantes, ferrovia, ind?stria / The city of S?o Paulo from 1870 to 1930: coffee, immigrants, railroads and industryMota, Paula de Brito 16 March 2007 (has links)
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Previous issue date: 2007-03-16 / This study approaches the processes of S?o Paulo urban evolution from 1870 to 1930, since its initial phase as villa until the city that later became into the current metropolis. For this, it analyzes in a concatenated manner, formulating casual connections, about the importance of coffee culture; the deployment and development of the railroads, decisive for the urban push registered on this period; the great immigrants contingent came from Europe in search of better enrichment opportunities and the influences caused by them in the social bodies at this time; the industrial development and the urban passage of Br?s quarter as stage of transformations occurred in the urgency of the industrial metropolis at the city of S?o Paulo from 1870 to 1930. / Este trabalho aborda os condicionantes da evolu??o urbana de S?o Paulo no per?odo de 1870 a 1930, desde o seu est?gio de vila at? a cidade que posteriormente se transfigurou na atual metr?pole. Para tanto, discorre de forma concatenada, formulando nexos causais, sobre a import?ncia da cultura do caf?; a implanta??o e desenvolvimento das estradas de ferro, decisivas para o grande impulso urbano registrado no per?odo em an?lise; o grande contingente de imigrantes vindos da Europa em busca de melhores oportunidades de enriquecimento e as influ?ncias por eles plasmadas nos corpos sociais da ?poca; o desenvolvimento industrial e o percurso urbano do bairro do Br?s como palco das transforma??es ocorridas na emerg?ncia da metr?pole industrial na cidade de S?o Paulo no per?odo de 1870 a 1930.
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Quatre essais sur les effets des rentes des ressources naturelles dans les pays en développement / Four essays on how incomes from natural ressources impact developing countriesMabali, Aristide 07 November 2016 (has links)
Cette thèse présente un ensemble de travaux de recherche en économie politique qui s’inscrivent dans le champ contemporain de la littérature sur la malédiction des ressources naturelles dans les pays en développement. Elle s’intéresse spécifiquement aux liens entre les rentes des ressources naturelles et les institutions politiques et les conflits, puis entre les rentes et la pauvreté sous ses différentes formes. Son champ géographique se situe à plusieurs niveaux, international pour un groupe de pays en développement, monographique et infranational pour le Tchad et ses régions. La première partie, en s’inspirant des théories des changements institutionnels, teste l’hypothèse que les rentes issues de différentes ressources naturelles ont des effets différenciés sur la qualité des institutions dans les pays en développement. Par la suite, elle teste l’hypothèse d’un « système de conflit régional » dans le Triangle République Centrafricaine, Tchad et Soudan. La deuxième partie évalue le dispositif « original » de gestion des revenus pétroliers sur différentes dimensions de la pauvreté au Tchad sur des données d’enquêtes de terrain. Le premier chapitre de cette partie teste l’hypothèse d’un biais urbain dans l’allocation de temps des enfants dans un contexte pétrolier au Tchad. Alors que le second est dédié à l’évaluation de l’impact de l’exploitation pétrolière sur le profil de la pauvreté dans la région productrice en utilisant les méthodes quasi-expérimentales d’évaluation d’impact. / This thesis is a collection of political economy research topics fitting into the contemporary field of literature on the natural resource curse in developing countries. The thesis focuses primarily on the links between natural resource rents and political institutions, before studying the impact of oil resource rents on poverty under its different forms. The geographical scope of the thesis is both macro for a group of developing countries, and micro in the case of Chad and its regions. The first part, drawing on institutional change theories, tests the hypothesis on whether rents generated by natural resources have a detrimental effect on institutions in developing countries, by considering different types of natural resources characterized by their different degree of appropriability. Besides, we evaluate the hypothesis of a "regional conflict system" in the “tormented Triangle” which includes Central African Republic, Chad and Sudan. The second part assesses the impact of the “unusual” system for allocating and managing the expected oil income from the Doba Oil project on many aspects of poverty in Chad by using field survey data. As such, the first chapter of this part tests the hypothesis of an urban bias in children's time allocation in the oil context in Chad. Regarding the second chapter, it investigates the impact of oil extraction on poverty profile in the oil producing region by using quasi-experimental evaluation methods.
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