Glucose transporters in the 21st Century.

The ability to take up and metabolize glucose at the cellular level is a property shared by the vast majority of existing organisms. Most mammalian cells import glucose by a process of facilitative diffusion mediated by members of the Glut (SLC2A) family of membrane transport proteins. Fourteen Glut proteins are expressed in the human and they include transporters for substrates other than glucose, including fructose, myoinositol, and urate. The primary physiological substrates for at least half of the 14 Glut proteins are either uncertain or unknown. The well-established glucose transporter isoforms, Gluts 1-4, are known to have distinct regulatory and/or kinetic properties that reflect their specific roles in cellular and whole body glucose homeostasis. Separate review articles on many of the Glut proteins have recently appeared in this journal. Here, we provide a very brief summary of the known properties of the 14 Glut proteins and suggest some avenues of future investigation in this area.

Towards a theory of the credit-risk balance sheet

This article designs what it calls a Credit-Risk Balance Sheet (the risk being that of default by customers), a tool which, in principle, can contribute to revealing, controlling and managing the bad debt risk arising from a company¿s commercial credit, whose amount can represent a significant proportion of both its current and total assets.To construct it, we start from the duality observed in any credit transaction of this nature, whose basic identity can be summed up as Credit = Risk. ¿Credit¿ is granted by a company to its customer, and can be ranked by quality (we suggest the credit scoring system) and ¿risk¿ can either be assumed (interiorised) by the company itself or transferred to third parties (exteriorised).What provides the approach that leads to us being able to talk with confidence of a real Credit-Risk Balance Sheet with its methodological robustness is that the dual vision of the credit transaction is not, as we demonstrate, merely a classificatory duality (a double risk-credit classification of reality) but rather a true causal relationship, that is, a risk-credit causal duality.Once said Credit-Risk Balance Sheet (which bears a certain structural similarity with the classic net asset balance sheet) has been built, and its methodological coherence demonstrated, its properties ¿static and dynamic¿ are studied.Analysis of the temporal evolution of the Credit-Risk Balance Sheet and of its applications will be the object of subsequent works.

"In vitro" study of the molecular mechanism of the "E.Coli" Hsp 70/Hsp40/NEF chaperone system and its interactions with ?-sinuclein

SUMMARY Under stressful conditions, mutant or post-translationally modified proteins may spontaneously misfold and form toxie species, which may further assemble into a continuum of increasingly large and insoluble toxic oligomers that may further condense into less toxic, compact amyloids in the cell Intracellular accumulation of aggregated proteins is a common denominator of several neurodegenerative diseases. To cope with the cytotoxicity induced by abnormal, aggregated proteins, cells have evolved various defence mechanisms among which, the molecular chaperones Hsp70. Hsp70 (DnaK in E. coii) is an ATPase chaperone involved in many physiological processes in the cell, such as assisting de novo protein folding, dissociating native protein oligomers and serving as pulling motors in the import of polypeptides into organelles. In addition, Hsp70 chaperones can actively solubilize and reactivate stable protein aggregates, such as heat- or mutation-induced aggregates. Hsp70 requires the cooperation of two other co-chaperones: Hsp40 and NEF (Nucleotide exchange factor) to fulfil its unfolding activity. In the first experimental section of this thesis (Chapter II), we studied by biochemical analysis the in vitro interaction between recombinant human aggregated α-synuclein (a-Syn oligomers) mimicking toxic a-Syn oligomers species in PD brains, with a model Hsp70/Hsp40 chaperone system (the E. coii DnaK/DnaJ/GrpE). We found that chaperone-mediated unfolding of two denatured model enzymes were strongly affected by α-Syn oligomers but, remarkably, not by monomers. This in vitro observed dysfunction of the Hsp70 chaperone system resulted from the sequestration of the Hsp40 proteins by the oligomeric α-synuclein species. In the second experimental part (Chapter III), we performed in vitro biochemical analysis of the co-chaperone function of three E. coii Hsp40s proteins (DnaJ, CbpA and DjlA) in the ATP-fuelled DnaK-mediated refolding of a model DnaK chaperone substrate into its native state. Hsp40s activities were compared using dose-response approaches in two types of in vitro assays: refolding of heat-denatured G6PDH and DnaK-mediated ATPase activity. We also observed that the disaggregation efficiency of Hsp70 does not directly correlate with Hsp40 binding affinity. Besides, we found that these E. coii Hsp40s confer substrate specificity to DnaK, CbpA being more effective in the DnaK-mediated disaggregation of large G6PDH aggregates than DnaJ under certain conditions. Sensibilisées par différents stress ou mutations, certaines protéines fonctionnelles de la cellule peuvent spontanément se convertir en formes inactives, mal pliées, enrichies en feuillets bêta, et exposant des surfaces hydrophobes favorisant l'agrégation. Cherchant à se stabiliser, les surfaces hydrophobes peuvent s'associer aux régions hydrophobes d'autres protéines mal pliées, formant des agrégats protéiques stables: les amyloïdes. Le dépôt intracellulaire de protéines agrégées est un dénominateur commun à de nombreuses maladies neurodégénératives. Afin de contrer la cytotoxicité induite par les protéines agrégées, les cellules ont développé plusieurs mécanismes de défense, parmi lesquels, les chaperonnes moléculaires Hsp70. Hsp70 nécessite la collaboration de deux autres co-chaperonnes : Hsp40 et NEF pour accomplir son activité de désagrégation. Hsp70 (DnaK, chez E. coli) est impliquée par ailleurs dans d'autres fonctions physiologiques telles que l'assistanat de protéines néosynthétisées à la sortie du ribosome, ou le transport transmembranaire de polypeptides. Par ailleurs, les chaperonnes Hsp70 peuvent également solubiliser et réactiver des protéines agrégées à la suite d'un stress ou d'une mutation. Dans la première partie expérimentale de cette thèse (Chapter II), nous avons étudié in vitro l'interaction entre les oligomères d'a-synucleine, responsables entre autres, de la maladie de Parkinson, et le système chaperon Hsp70/Hsp40 (système Escherichia coli DnaK/DnaJ/GrpE). Nous avons démontré que contrairement aux monomères, les oligomères d'a-synucleine inhibaient le système chaperon lors du repliement de protéines agrégées. Cette dysfonction du système chaperon résulte de la séquestration des chaperonnes Hsp40 par les oligomères d'a-synucleine. La deuxième partie expérimentale (Chapitre III) est consacrée à une étude in vitro de la fonction co-chaperonne de trois Hsp40 d'is. coli (DnaJ, CbpA, et DjlA) lors de la désagrégation par DnaK d'une protéine pré-agrégée. Leurs activités ont été comparées par le biais d'une approche dose-réponse au niveau de deux analyses enzymatiques: le repliement de la protéine agrégée et l'activité ATPase de DnaK. Par ailleurs, nous avons mis en évidence que l'efficacité de désagrégation d'Hsp70 et l'affinité des chaperonnes Hsp40 vis-à-vis de leur substrat n'étaient pas corrélées positivement. Nous avons également montré que ces trois chaperonnes Hsp40 étaient directement impliquées dans la spécificité des fonctions accomplies par les chaperonnes Hsp70. En effet, DnaK en présence de CbpA assure la désagrégation de large agrégats protéiques avec une efficacité nettement plus accrue qu'en présence de DnaJ.

Affaires de patrons, usages de la frontière : géographie des réseaux marchands entre Niger, Bénin et Nigéria

Résumé L'objectif de la thèse est de comprendre le mode d'organisation économique spécifique aux petits centres urbains qui composent les espaces frontaliers sahéliens, en s'interrogeant sur leur concurrence ou leur complémentarité éventuelle à l'intérieur d'un régime de spatialité particulier. En s'appuyant sur l'exemple du carrefour économique de Gaya-Malanville-Kamba situé à la frontière entre le Niger, le Bénin et le Nigeria, il questionne le rôle de la ville-frontière ainsi que le jeu des acteurs marchands localement dominants, à partir de quatre grandes interrogations : Quelles sont les spécificités de l'Afrique sahélienne qui obligent à renouveler les approches géographiques de l'espace marchand? Quels sont les facteurs déterminants de l'activité économique frontalière? Les formes d'organisation de l'espace qui concourent à la structuration de l'économie sont-elles concurrentes ou coopératives? Les logiques économiques frontalières sont-elles compatibles avec l'orientation des programmes de développement adoptés par les pays sahéliens et leurs partenaires bi- ou multilatéraux? Dans une première partie, un modèle territorial de l'Afrique sahélienne permet de rendre compte de la prédominance des logiques circulatoires sur les logiques productives, une propriété essentielle de toute organisation économique confrontée à l'instabilité climatique. Dans une seconde partie, l'étude considère les facteurs déterminants de l'activité économique frontalière que sont le degré d'enclavement des territoires, la libre circulation des biens et des personnes, les relations concurrentielles ou coopératives qui lient les marchés ainsi que les liens clientélistes qui unissent patron et obligés. Une troisième partie est consacrée aux productions agricoles de tente organisées sous forme de coopératives paysannes ou d'initiatives privées. Une quatrième partie s'intéresse aux réseaux de l'import-export et du commerce de détail qui bénéficient de l'augmentation des besoins engendrée par l'urbanisation sahélienne. L'économie spatiale qui résulte de ces flux est organisée selon deux logiques distinctes : d'une part, les opportunités relatives à la production agricole conduisent certains investisseurs à intensifier l'irrigation pour satisfaire la demande des marchés urbains, d'autre part, les acteurs du capitalisme marchand, actifs dans l'import-export et la vente de détail, développent des réseaux informels et mobiles qui se jouent des différentiels nationaux. Les activités commerciales des villes-marchés connaissent alors des fluctuations liées aux entreprises productives et circulatoires de ces patrons, lesquelles concourent à l'organisation territoriale générale de l>Afrique sahélienne. Ces logiques évoluent dans un contexte fortement marqué par les politiques des institutions financières internationales, des agences bilatérales de coopération et des ONGs. Celles-ci se donnent pour ambition de transformer les économies, les systèmes politiques et les organisations sociales sahéliennes, en faisant la promotion du libéralisme, de la bonne gouvernance et de la société civile. Ces axes directeurs, qui constituent le champ de bataille contemporain du développement, forment un ensemble dans lequel la spécificité sahélienne notamment frontalière est rarement prise en compte. C'est pourquoi l'étude conclut en faveur d'un renouvellement des politiques de développement appliquées aux espaces frontaliers. Trois grands axes d'intervention peuvent alors être dégagés, lesquels permettent de réconcilier des acteurs et des logiques longtemps dissociés: ceux des espaces séparés par une limite administrative, ceux de la sphère urbaine et rurale et ceux du capitalisme marchand et de l'investissement agricole, en renforçant la coopération économique transfrontalière, en prenant en considération les interactions croissantes entre villes et campagnes et en appuyant les activités marchandes. Abstract: Urbanisation in West Africa is recent and fast. If only 10 % of the total population was living in urban areas in 1950, this proportion reached 40 % in 2000 and will be estimated to 60 % in 2025. Small and intermediate cities, located between the countryside and large metropolis, are particularly concerned with this process. They are nowadays considered as efficient vectors of local economic development because of fiscal or monetary disparities between states, which enable businessmen to develop particular skills based on local urban networks. The majority of theses networks are informal and extremely flexible, like in the Gaya - Malanville - Kamba region, located between Niger, Benin and Nigeria. Evidence show that this economic space is characterised by high potentialities (climatic and hydrological conditions, location on main economic West African axis) and few constraints (remoteness of some potentially high productive areas). In this context, this PhD deals with the economic relationships between the three market cities. Focusing on the links that unite the businessmen of the local markets - called patron; - it reveals the extreme flexibility of their strategies as well as the deeply informal nature of their activities. Through the analysis of examples taken from the commerce of agricultural products, import and export flows and detail activities, it studies the changes that have taken place in the city centres of Gaya, Malanville and Kamba. Meanwhile, this research shows how these cities represent a border economical area based on rival and complementary connections. In the first Part, it was necessary to reconsider the usual spatial analysis devoted to the question of economic centrality. As a matter of fact, the organisation of West African economic spaces is very flexible and mobile. Centrality is always precarious because of seasonal or temporary reasons. This is why the first chapters are devoted to the study of the specificity of the Sahelian territoriality. Two main elements are relevant: first the population diversity and second, the urban-rural linkages. In the second part, the study considers three main factors on which the cross-border economic networks are dependent: enclosure that prevents goods to reach the markets, administrative constraints that limit free trade between states and cities and the concurrent or complementary relationships between markets. A third part deals with the clientelist ties engaged between the patrons and their clients with the hypothesis that these relationships are based on reciprocity and inequality. A fourth part is devoted to' the study of the spatial organisation of commercial goods across the borders, as far as the agriculture commercial products, the import-export merchandises and the retail products are concerned. This leads to the conclusion that the economic activity is directly linked to urban growth. However, the study notices that there is a lack of efficient policies dealing with strengthening the business sector and improving the cross-border cooperation. This particularity allows us to favour new local development approaches, which would take into account the important potential of private economical actors. In the same time, the commercial flows should be regulated with the help of public policies, as long as they are specifically adapted to the problems that these areas have to deal with.

ATRIBUTOS FÍSICOS DE UM LATOSSOLO VERMELHO CULTIVADO COM PLANTAS DE COBERTURA, EM SEMEADURA DIRETA

A utilização do sistema de semeadura direta associado ao uso de plantas de cobertura e à rotação de culturas altera alguns atributos físicos do solo e pode indicar mudança em sua qualidade. Este estudo objetivou utilizar o índice de estabilidade de agregados (IEA) e o índice de sensibilidade (IS) para avaliar alterações nos atributos físicos do solo com o uso de diferentes plantas de cobertura após 12 anos de semeadura direta. Com delineamento de blocos ao acaso, utilizaram-se as coberturas: crotalária, milheto, sorgo, braquiária, pousio (vegetação espontânea), amostradas no ano de 2012, mais uma testemunha sem cobertura (semeadura convencional) desde o ano de 2000, em que foram coletadas três amostras por parcela, perfazendo um total de 12 amostras por tratamento, com quatro repetições. Foram avaliadas a densidade do solo (Ds), macroporosidade (Ma), microporosidade (Mi), porosidade total (PT), estabilidade dos agregados (EA), diâmetro médio geométrico (DMG) e ponderado (DMP), índice de sensibilidade (Is) e índice da porcentagem de agregados com diâmetro superior a 2 mm (AGRI). A utilização das diferentes coberturas e a introdução do sistema de semeadura direta após 12 anos causaram alterações positivas nos atributos físicos na camada superficial do solo. As correlações positivas e negativas significativas entre densidade do solo e os outros atributos físicos avaliados evidenciaram a sua importância como bom indicador da qualidade do solo. Por meio do índice de sensibilidade, constatou-se que microporosidade aumentou mais do que os outros atributos físicos após a implantação do sistema de semeadura direta na área.

EFICIÊNCIA DE SOLUÇÕES DENSAS NO FRACIONAMENTO FÍSICO DA MATÉRIA ORGÂNICA DO SOLO

Os mecanismos de estabilização da matéria orgânica (MO) têm sido estudados em solos tropicais e subtropicais brasileiros; no entanto, poucos trabalhos avaliaram a influência da parte metodológica do uso das soluções nos resultados obtidos. Objetivou-se avaliar a eficiência de duas soluções salinas (iodeto de sódio - NaI 1,8 kg dm-3 e politungstato de sódio - PTS 2,0 kg dm-3) na separação de frações densimétricas da MO em dois solos (Argissolo Vermelho do sul do Brasil - 220 g kg-1 argila e Latossolo Vermelho do Cerrado - 630 g kg-1 argila) e o reflexo dessa eficiência na magnitude e importância do mecanismo de proteção física por oclusão em agregados no acúmulo de MO em solos brasileiros. Amostras de solo coletadas nas camadas de 0,00-0,05; 0,05-0,10; e 0,10-0,20 m foram submetidas ao fracionamento físico densimétrico e separadas as frações leve livre (FLL), leve-oclusa (FLO) e pesada (FP) da MO do solo. O uso de PTS 2,0 kg dm-3 aumentou o rendimento de carbono orgânico (CO) das FLL e FLO em ambos os solos em relação à solução de NaI 1,8 kg dm-3, sendo o efeito mais pronunciado na FLO. A utilização do sistema plantio direto (PD) aumentou os estoques de CO total na camada de 0,00-0,05 m do Argissolo em relação ao solo em preparo convencional (PC). O mecanismo de proteção física da MO por oclusão em agregados foi efetivo no aumento dos estoques de C do solo, sendo responsável por aproximadamente ⅓ do acúmulo na camada superficial do Argissolo (0,00-0,05 m). Para o Latossolo, não houve diferenças entre o estoque de CO total do PD e do PC, porém o solo sob PD apresentou acúmulo de aproximadamente ⅔ do estoque de C como FLO na camada superficial do solo. A proteção física por oclusão em agregados é um mecanismo expressivo na estabilização e sequestro de C em solos tropicais e subtropicais, cuja importância pode ser mascarada pela baixa eficiência da solução de NaI em estudos de fracionamento densimétrico da MO do solo.

PRODUÇÃO DE MATÉRIA SECA E ACÚMULO DE MACRONUTRIENTES NA PARTE AÉREA DAS PLANTAS DE NIGER

O estudo da marcha de absorção de nutrientes e da produção de matéria seca em razão do estádio de desenvolvimento das culturas é de fundamental importância para subsidiar estratégias de adubação. Este trabalho objetivou determinar a produção de matéria seca e o acúmulo de macronutrientes na parte aérea das plantas de Guizotia abyssinica (niger), em diferentes estádios de crescimento e desenvolvimento. O delineamento experimental foi em blocos ao acaso, com cinco repetições. Amostras da parte aérea das plantas foram coletadas aos 15, 30, 45, 60, 75 e 90 dias após a emergência (DAE). Em cada coleta, as plantas foram divididas em folha, caule + ramos, capítulo e aquênios (grãos), secas até peso constante para posterior determinação da massa de matéria seca e teor de N, P, K, Ca, Mg e S. Os teores dos macronutrientes na parte aérea das plantas de niger, seguiram a seguinte ordem de exportação de nutrientes: N > Ca > Mg > P > K > S. Em contrapartida, quanto aos teores dos nutrientes nos grãos por ocasião da colheita do experimento, a quantidade dos elementos exportados seguiu a ordem: N > P > Ca > Mg > S > K. Atenção especial deve ser dada à manutenção da adequada disponibilidade de N e P nas fases inicias por causa do ciclo curto e do rápido surgimento das estruturas reprodutivas. A exportação de nutrientes e o índice de colheita são baixos para as plantas de niger, resultando maior oferta de nutrientes para o cultivo em rotação.

MÉTODOS DE INDEXAÇÃO DE INDICADORES NA AVALIAÇÃO DA QUALIDADE DO SOLO EM RELAÇÃO À EROSÃO HÍDRICA

A avaliação da qualidade dos solos agrícolas é importante para definição e adoção de práticas de manejo que garantam a sustentabilidade socioeconômica e ambiental. Os métodos de indexação dos indicadores de qualidade denominados Índice de Qualidade Integrado (IQI) e Índice de Qualidade Nemoro (IQN) foram utilizados neste estudo para avaliar a qualidade de solo em áreas experimentais de plantio de eucalipto. A seleção dos indicadores foi feita a partir de nove indicadores de qualidade do solo: diâmetro médio geométrico, permeabilidade à água, matéria orgânica, macro e microporosidade, volume total de poros, densidade do solo, resistência à penetração e índice de floculação, que estão relacionados à erosão hídrica. Os tratamentos constituíram de eucalipto plantado em nível, com e sem a manutenção dos resíduos, em desnível e solo descoberto, em dois biomas distintos, cujas vegetações nativas são Cerrado e Floresta. Os índices de qualidade do solo (IQS) apresentaram alta correlação com a erosão hídrica. Entre os sistemas manejados, o Eucalipto com manutenção do resíduo evidenciou valores mais elevados em ambos os índices, ressaltando-se a importância da cobertura vegetal e manutenção da matéria orgânica para conservação do solo e da água em sistemas florestais. Os IQS demonstraram alto coeficiente de correlação inversa com as perdas de solo e água. Em locais com as maiores taxas de erosão hídrica manifestaram também os menores valores de IQI e IQN. Assim, os índices testados permitiram avaliar com eficácia os efeitos dos manejos adotados sobre a qualidade do solo em relação à erosão hídrica.

Efeitos Tóxicos de Arsênio em Eisenia Andrei em Exposição a Solos do Entorno de Minerações de Ouro

Os produtos das atividades de mineração têm grande relevância econômica no Estado de Minas Gerais, Brasil. No entanto, seus inúmeros danos à biota, incluindo a fauna edáfica, resultam na necessidade de recuperação e biomonitoramento dessas áreas e do seu entorno. Considerando a importância ecológica e o potencial bioindicador das minhocas, o objetivo deste estudo foi avaliar a toxicidade de arsênio presente em solos do entorno de minerações de ouro, nas bacias dos rios Doce e São Francisco, a oligoquetas da espécie Eisenia andrei Bouché. Amostras de solos dessas áreas foram caracterizadas em relação às suas propriedades físicas e químicas, incluindo os metais Fe, Al, Cu, Ni, Zn e Mn e do metaloide As; com elas, foram conduzidos bioensaios, que avaliaram a sobrevivência, reprodução e variação da biomassa dos organismos. Também foram realizados testes de sensibilidade dessa espécie ao As adicionado a solo artificial tropical e em solo natural. Houve evidente efeito tóxico sobre a reprodução dessa espécie, que esteve altamente correlacionado com as concentrações de As nas amostras (r = -0,80). Amostras de solos naturais de locais avaliados nas duas áreas apresentaram concentrações de As acima do limite estipulado pela Resolução Conama nº 420 de 2009, chegando a 2.388,2 mg kg-1. A CL50 do As em solo artificial tropical (207,4 mg kg-1) foi seis vezes menor que a do solo natural (1.248,1 mg kg-1). A menor biodisponibilidade do As nos solos naturais foi atribuída a sua associação com o minério de Fe presente nas amostras. Concluiu-se que os solos do entorno de áreas de mineração causaram toxicidade crônica a E. andrei, em termos de sua reprodução, o que demonstrou o potencial de danos dessas atividades sobre a fauna edáfica dessas áreas. O estudo também forneceu informações ecotoxicológicas que podem ser úteis em avaliações de risco ambiental em solos tropicais.

Managing the crises : How do Swiss municipalities react to the crisis and what explains the differences ?

Swiss municipalities are to an import ant extent responsible for their own resources. Since these resources primarily depend on income and property tax from individuals and enterprises, their budgets are likely to be directly affected by the actual crisis of the financial sector and the economy. This paper investigates how the municipalities perceive this threat and how they reacted to it or plan to do so. In a nationwide survey conducted at the end of 2009 in all 2596 Swiss municipalities, we asked the local secretaries which measures they launch in order to cope with expected losses in tax income and a possible increase of welfare spending. Do the municipalities rather rely on Keynesian measures increasing public spending and accepting greater deficits, or do they try to avoid further deficits by austerity measures and a withdrawal of planned investments? The paper shows that only a few municipalities - mainly the bigger ones - expect to be strongly hit by the crisis. Their reactions, however, do not reveal the clear patterns theory lets to expect. Preferences for austerity measures and deficit spending become visible but many municipalities take measures from both theories. The strongest explaining factor whether municipalities react is the affectedness by the crisis followed by the fact that the municipality belongs to the French speaking part of the country. Size also has an effect whereas the strength of the Social Democrats is negligible. More difficult is it, to explain what kind of measures municipalities are likely to take.

Molecular mechanisms of penicillin-induced killing and penicillin tolerance in Streptococcus gordonii

Abstract The main thesis topic relates to the 'molecular mechanisms of penicillin-induced bacterial death. Indeed, bacteria have developed two principal mechanisms to escape the killing effect of ß-lactam antibiotics: resistance and tolerance. Resistant bacteria are characterized by their ability to grow in the presence of drug concentrations higher than the one inhibiting the growth of susceptible members of the same species. Hence, resistant bacteria have an increased minimal inhibitory concentration (MIC) of the drug. Nevertheless, when exposed to antibiotic concentrations exceeding their new MIC, resistant bacteria remain sensitive to the antibiotic killing effect. In contrast, tolerant bacteria have an unchanged MIC. However, they have a considerably increased ability to survive drug-induced killing, even at concentrations exceeding their MIC by several orders of magnitude. In other words, in the presence of the antibiotic, tolerant bacteria become persister cells which stop growing but are not killed. In the present thesis, it is shown that the survival phenotype of a tolerant Streptococcus gordonii strain depends on two components belonging to sugar metabolism pathways. First, the transcription factor CcpA which mediates a global regulatory mechanism allowing bacteria to utilize the most efficient sugar source for their growth. We show that the inactivation of the ccpA gene leads to a partial loss of penicillin tolerance both in vitro and in a rat model of experimental endocarditis. Second, the Enzyme I of the phosphotransferase system which is involved in the uptake and phosphorylation of sugars. Here, we -show that a single nucleotide mutation in ptsI, the gene encoding the Enzyme I, is sufficient to confer a fully tolerant phenotype in S. gordonii both in vivo and in vivo. The mutation results in a radical proline to arginine substitution in the C-terminal domain of the protein, probably leading to a decrease in its homodimerization and subsequent activity. Taken together our results prove that tolerance is a global survival mechanism linked to sugar metabolism. We hypothesize that, in the presence of the antibiotic, the already altered metabolic processes of the tolerant strain are completely inactivated. Hence, bacteria may enter in a dormant state and become insensitive to the bactericidal effect of ß-lactams, which depends on actively dividing cells. This thesis manuscript also contains two other side-projects. The first one establishes that the ability to form a biofilm is not a requisite for the successful establishment of endocarditis due to S. gordonii. The second one characterizes the S. gordonii a-phosphoglucomutase gene, and shows that its inactivation results in a loss of in vitro fitness and in vivo virulence. Résumé Le sujet principal de cette thèse concerne les mécanismes moléculaires de la mort bactérienne induite par la pénicilline. En effet, les bactéries ont développé deux mécanismes principaux pour échapper à l'effet bactéricide des ß-lactamines : la résistance et la tolérance. Les bactéries résistantes sont caractérisées par leur capacité de croître en présence de concentration d'antibiotiques plus élevées que celles inhibant la croissance des organismes sensibles de la même espèce. Les bactéries résistantes ont donc une augmentation de leur concentration minimale inhibitrice (CMI) à l'antibiotique. Néanmoins, quand elles sont exposées à des concentrations dépassant leur nouvelle CMI, elles restent sensibles à l'effet bactéricide. Au contraire, les bactéries tolérantes ont une CMI inchangée. Toutefois, elles ont une très importante capacité à survivre à l'effet bactéricide des ß-lactamines, ceci même à des concentrations excédant leur CMI de plusieurs ordres de grandeur. En d'autres termes, en présence de l'antibiotique, les bactéries tolérantes deviennent des cellules persistantes qui arrêtent leur croissance mais ne sont pas tuées. Dans la présente thèse, il est montré que le phénotype de survie d'un Streptococcus gordonii tolérant dépend de deux composants appartenant aux voies du métabolisme des sucres. Premièrement, le facteur de transcription CcpA qui contrôle un système global de régulation permettant à la bactérie d'utiliser les sources de sucre les plus efficaces pour sa croissance. Il est montré que l'inactivation du gène ccpA résulte en la perte partielle de la tolérance à la pénicilline aussi bien in vitro que dans un modèle d'endocardite expérimentale chez le rat. Deuxièmement, l'Enzyme I du système de phosphotransfert impliqué dans l'import et la phosphorylation des sucres. Nous montrons qu'une mutation ponctuelle d'un nucléotide dans ptsl, le gène codant pour l'Enzyme I, suffit à complètement conférer un phénotype tolérant chez S. gordonii aussi bien in vitro qu'in vivo. La mutation induit la substitution radicale d'une proline en une arginine dans le domaine C-terminal de la protéine, résultant probablement en une diminution de sa capacité d'homodimérisation et donc d'activité. Dans leur ensemble, nos résultats prouvent que la tolérance est un mécanisme global de survie lié au métabolisme des sucres. Nous présentons l'hypothèse que, en présence de l'antibiotique, les processus métaboliques déjà altérés de la souche tolérante deviennent complètement inactifs. En conséquence, les bactéries entreraient dans un état dormant nonréplicatif, devenant ainsi insensibles à l'effet bactéricide des ß-lactamines qui nécessite des cellules en cours de division active. Le manuscrit de cette thèse contient également deux projets secondaires. Le premier montre que la capacité de former un biofilm n'est pas un prérequis pour le succès de l'initiation de l'endocardite à S. gordonii. Le second caractérise le gène de l'a-phosphoglucomutase de S. gordonii et montre que son inactivation résulte en une perte de fitness in vitro et de virulence in vivo.

Hsp70 chaperones accelerate protein translocation and the unfolding of stable protein aggregates by entropic pulling.

Hsp70s are highly conserved ATPase molecular chaperones mediating the correct folding of de novo synthesized proteins, the translocation of proteins across membranes, the disassembly of some native protein oligomers, and the active unfolding and disassembly of stress-induced protein aggregates. Here, we bring thermodynamic arguments and biochemical evidences for a unifying mechanism named entropic pulling, based on entropy loss due to excluded-volume effects, by which Hsp70 molecules can convert the energy of ATP hydrolysis into a force capable of accelerating the local unfolding of various protein substrates and, thus, perform disparate cellular functions. By means of entropic pulling, individual Hsp70 molecules can accelerate unfolding and pulling of translocating polypeptides into mitochondria in the absence of a molecular fulcrum, thus settling former contradictions between the power-stroke and the Brownian ratchet models for Hsp70-mediated protein translocation across membranes. Moreover, in a very different context devoid of membrane and components of the import pore, the same physical principles apply to the forceful unfolding, solubilization, and assisted native refolding of stable protein aggregates by individual Hsp70 molecules, thus providing a mechanism for Hsp70-mediated protein disaggregation.

Proteomic data from human cell cultures refine mechanisms of chaperone-mediated protein homeostasis.

In the crowded environment of human cells, folding of nascent polypeptides and refolding of stress-unfolded proteins is error prone. Accumulation of cytotoxic misfolded and aggregated species may cause cell death, tissue loss, degenerative conformational diseases, and aging. Nevertheless, young cells effectively express a network of molecular chaperones and folding enzymes, termed here "the chaperome," which can prevent formation of potentially harmful misfolded protein conformers and use the energy of adenosine triphosphate (ATP) to rehabilitate already formed toxic aggregates into native functional proteins. In an attempt to extend knowledge of chaperome mechanisms in cellular proteostasis, we performed a meta-analysis of human chaperome using high-throughput proteomic data from 11 immortalized human cell lines. Chaperome polypeptides were about 10 % of total protein mass of human cells, half of which were Hsp90s and Hsp70s. Knowledge of cellular concentrations and ratios among chaperome polypeptides provided a novel basis to understand mechanisms by which the Hsp60, Hsp70, Hsp90, and small heat shock proteins (HSPs), in collaboration with cochaperones and folding enzymes, assist de novo protein folding, import polypeptides into organelles, unfold stress-destabilized toxic conformers, and control the conformal activity of native proteins in the crowded environment of the cell. Proteomic data also provided means to distinguish between stable components of chaperone core machineries and dynamic regulatory cochaperones.

Deficiência múltipla: formação de professores e processos de ensino-aprendizagem

O artigo discute diferentes dimensões do processo educacional de alunos com deficiência múltipla matriculados no atendimento educacional especializado - AEE - oferecido numa classe multifuncional em uma escola localizada na Baixada Fluminense, estado do Rio de Janeiro, no período letivo de 2013. Participaram da investigação quatro alunos com deficiência múltipla não oralizados e duas professoras do AEE. Metodologicamente, empregaram-se os referenciais da pesquisa-ação; na coleta de dados, a observação participante e entrevistas semiestruturadas; como base teórica, a matriz histórico-cultural de Vigotski. Os resultados evidenciaram, entre outros pontos, a importância da pesquisa-ação na formação continuada dos professores participantes. Igualmente, ilustraram as possibilidades de construção dos processos psicológicos superiores nos alunos com essa deficiência com base em intervenções pedagógicas sistemáticas com uso de recursos da comunicação alternativa e das tecnologias assistivas.

Problemáticas psicossociais da abordagem da noção de sujeito

O presente artigo trata da importância da questão do sujeito nas representações sociais. Para tanto, repassa alguns modelos da psicologia social e alude às novas perspectivas sobre o sujeito desenvolvidas nas ciências sociais e humanas. Feito um breve panorama do uso da categoria sujeito na história dessa disciplina, com destaque a seu retorno às ciências sociais nos anos 1990 e a noções correlatas, como "ator" e "agente", aponta-se a necessidade de trabalhar com a noção de "subjetivação" em pesquisas que considerem a subjetividade sem perder a dimensão social no estudo das representações, consideradas em dois níveis: o subjetivo e o transubjetivo.