L’objet de cette recherche porte sur l’évolution des dispositifs de formation avec l’introduction de technologies de réalité virtuelle dans des contextes industrialisés. Cette thèse Cifre s’appuie sur une intégration au sein du département « training » d’Airbus Helicopters. La démarche de recherche-action mise en œuvre permet d’observer et de participer à l’élaboration d’une technologie immersive pour la formation de pilotes. Cet outil est mis en situation avec des clients en formation et plusieurs démonstrations sont faites auprès d’acteurs de différents métiers. Nous mettons en avant des configurations hybrides dans le sens ou les situations de formation révèlent des aspects de la pratique et de la théorie telles qu’elles sont pensées dans la « langue de l’organisation ». Nous montrons alors que la mise en œuvre de telles technologies pourrait remettre en question les modèles rationalisés dans l’organisation. Ces éléments nous permettent d’envisager qu’une « approche par l’hybridation des dispositifs » amène l’organisation à interroger à chaque étape du projet ses propres modèles (objets structurés de partage d’information, organigramme, compétences et métiers des acteurs). C’est alors une remise en question en « profondeur » du projet de l’organisation apprenante qui est rendue possible, à partir de l’expérience des sujets. / The purpose of this research is to better understand how training devices evolve, with the introduction of virtual reality technology, in industrialized training situations. This work I based on an immersion in the "training" department of Airbus Helicopters. The research-action approach allows us to observe and participate in the emergence of a technology of reality for pilot’s training. The tool is put in situation with customers as part of their training and several demonstrations are made to actors of different trades around the training. We identify hybrid configurations in the sense that training situations are both practical and at the same time theoretical in the language of the organization. Then we show that the implementation of these technologies requires to rethink a traditional rationalization. These elements allow to suggest an "approach by the hybridization of systems" gives the organization the ability to reconsider its own project (structured objects of information, organization chart, skills and trades of actors). It is a depth questioning of the learning organization that is possible, from the experience of subjects.

O desenvolvimento de novos materiais tem tido papel fundamental nos recentes avanços tecnológicos. Esse progresso depende muito de fundamentos teóricos que abordem mecanismos microscópicos da matéria, ou seja, como átomos e moléculas interagem e geram configurações especiais, responsáveis pelo seu comportamento macroscópico. Dentre os materiais de interesse na atualidade estão os sistemas contendo impurezas magnéticas diluídas, isto é, átomos com camadas d ou f incompletas imersos, por exemplo, em metais não magnéticos, como átomos de ferro em uma matriz de cobre. Tradicionalmente, estes sistemas tem sido tratados através dos modelos de Kondo ou Anderson, os quais, desde os primeiros estudos na década de 1960, estão entre os mais importantes em física da matéria condensada. Neste trabalho, estudamos especificamente o modelo de Anderson de uma impureza. Ele se caracteriza por considerar uma correlação quando dois elétrons de spins opostos ocupam o nível localizado que representa a impureza. Além de, por outro termo no Hamiltoniano, contabilizar a hibridização eletrônica entre a banda de condução e a impureza, devido à superposição das funções de onda dos elétrons localizados e itinerantes. Em acréscimo ao modelo tradicional, incluímos um termo de hibridização adicional, que depende explicitamente do número de ocupação do nível localizado. Este termo de interação que acopla diretamente no Hamiltoniano o processo de hibridização e os efeitos de correlação é denominado de hibridização correlacionada. Através da estruturação e da consequente aplicação da técnica do Grupo de Renormalização Numérico - a qual estabelece uma transformação no Hamiltoniano, que a cada passo acrescenta uma escala de energia ao problema e constrói um método iterativo, no qual um Hamiltoniano é diagonalizado numericamente a cada iteração -, analisamos os efeitos de hibridização correlacionada sobre parte da física do modelo de Anderson de uma impureza. Em particular, isso é feito por meio de dados numéricos para a dependência da contribuição da impureza a três propriedades termodinâmicas - são elas: suscetibilidade magnética, calor específico e entropia - em função da temperatura, desde o topo da banda de condução até o nível de Fermi. / The development of new materials has been playing a fundamental role in the currently technological advances. This improvement is strongly dependent on the theoretical foundations which study the microscopic matter engine, i.e., the way atoms and molecules interact and create distinct configurations, responsible for their macroscopic behavior. Among the interesting materials, there are the dilute magnetic impurity systems. They are constituted by partially filled d or f orbital atoms immersed, for exemple, in nonmagnetic metals; like iron atoms in a copper background. Traditionally, such system has been described by the Kondo and Anderson models, which are, since the sixties, two of the most important models in condensed matter physics. In the present work, we specifically study the single-impurity Anderson model. It is characterized by taking correlation into account when two particles with opposite spins fill the impurity localized energy level. Beyond, by another term in the Hamiltonian, it considers the eletronic hybridization between impurity and conduction band, due their wave functions overlap. In addition to the usual model, we include a different hybridization term, which explicitly depends on localized level occupation number. This new interaction term, which couples hybridization process and correlation effects directy in the Hamiltonian, is named correlated hybridization. Through the exposition of Numerical Renormalization Group technique and its consequent enforcement - the procedure states a transformation in the Hamiltonian, where each step adds an energy scale to the problem and set up an iterative scheme, where a Hamiltonian is numerically diagonalized at each iteration - we analyse effects of correlated hybridization on part of the single-impurity Anderson model physics. In particular, this is done by numerical renormalization group data for the temperature dependence of the impurity contribution to three thermodynamical properties - they are: magnetic susceptibility, specific heat and entropy -, from the top of the conduction band until the Fermi level.

O presente estudo teve o objetivo de verificar o padrão de expressão dos genes HOXB13 e HHEX em carcinomas epidermóides de boca (CEB) através das técnicas de Transcriptase Reversa em Reação de Cadeia Polimerase (RT- PCR) e Hibridização In situ (ISH). Fragmentos de tecido tumoral e de tecido não tumoral adjacente à lesão foram obtidos de 30 pacientes portadores de CEB no Serviço de Cirurgia de Cabeça e Pescoço do HC - FMUSP. As amostras tiveram seus cDNAs extraí dos e submetidos à amplificação por PCR. Os “ amplicons" foram visualizados sob luz UV por eletroforese em gel de agarose a 1% contendo brometo de etí dio. A amplificação dos genes foram correlacionadas com a classificação UICC, TNM, graduação histológica, localização e espessura tumoral, invasão de tecidos adjacentes, perineural e vascular. Após seqüenciamento dos “ amplicons" e confirmação dos genes foram confecionadas as sondas de mRNA para realização da técnica de hibridização in situ. Os resultados obtidos através da técnica de RT-PCR mostraram que: a amplificação dos transcritos dos genes HOXB13 e HHEX podem ser detectados tanto no carcinoma epidermóide quanto no tecido não tumoral adjacente à lesão; não existindo diferença na amplificação dos transcritos de ambos genes para os dois grupos de tecido estudados; a amplificação do transcrito do gene HOXB13 mostrou relação estatí stica com os fatores prognósticos: espessura tumoral, invasão perineural e invasão vascular; a amplificação do transcrito do gene HHEX mostrou relação estatí stica com os fatores prognósticos: invasão vascular,envolvimento com os tecidos adjacentes e uma relação inversa com a idade do paciente. A expressão dos transcritos dos genes HOXB13 e HHEX, detectados pela técnica de ISH, mostrou um padrão de marcação consistente e invariável para os tecidos analisados, estando expressos tanto em carcinoma epidermóide quanto no tecido não tumoral adjacentes à lesão. Os resultados apontam para uma correlação entre a expressão do HHEX e do HOXB13 e alguns fatores prognósticos importantes podendo representar um indicador prognóstico valoroso para o entendimento do comportamento biológico do CEB. / The aim of this study was to verify the HOXB13 and HHEX genes expression in oral squamous cell carcinoma (OSCC) using Reverse Transcriptase Polymerase Chain Reaction (RT-PCR) and in situ Hybridization techniques. Tumoral tissues and adjacent non-tumoral oral mucosa specimens were obtained from 30 patients with OSCC at the Head and Neck Surgery Service HC (FMUSP). The samples were cDNA extracted and submitted to the RT-PCR technique. The amplicons were visualized in electrophoresis on a 1% agarose gel with ethidium bromide. Genes expressions were correlated with UICC staging, TNM stage, tumor location, tumor thickness, adjacent tissues involvement, vascular and perineural invasion, and cellular differentiation. Finally, direct sequence analysis was performed on PCR products to confirm cDNA sequence. Riboprobes were confectioned for in situ hybridization analysis. RT-PCR results showed HOXB13 and HHEX transcripts in both tumoral and non-tumoral tissue samples; no statistical correlation was verified between HOXB13/HHEX expressions and tumoral or non-tumoral tissues; there was a positive correlation between HOXB13 tumoral expression and tumor thickness, neural invasion, and vascular invasion; there was a positive correlation between HHEX tumoral expression and adjacent tissue involvement, vascular invasion, and a inverse relationship with patients age; ISH technique exhibited a consistent and invariable pattern of expression for both genes on both tumoral and non-tumoral tissue samples. Present results points out to a correlation between HOXB13 and HHEX expressions, and some important prognostic indicators, and this may represent a valuable tool to understand the biological behavior of OSCC.

A Síndrome de Rubinstein-Taybi (RTSs) é uma doença rara de herança autossômica dominante, caracterizada por dismorfismos craniofaciais, polegares e háluces alargados, deficiência intelectual e de crescimento. RTSs tem sido associada com mutações no gene CREBBP (CBP) e mutações menos frequentes no gene EP300 que foram descritas em oito indivíduos. CBP e p300 possuem alta homologia e são extremamente importantes em várias vias de sinalização, principalmente como coativadores de transcrição e na acetilação das histonas. Nosso estudo baseou-se na análise de alterações moleculares por sequenciamento direto do CBP, FISH e array-CGH em 20 pacientes com RTSs. Dos 20 pacientes avaliados por sequenciamento direto foram identificadas oito alterações moleculares, dentre estas, seis são alterações moleculares novas as quais não foram descritas na literatura, são elas: i) duas deleções (p.M747fs STOP830 e p.G1011fs STOP1021) ii) duas alterações do tipo nonsense (p.Arg1341X, p.Arg1498X) iii) três do tipo missense (p.Arg1907Trp, p.Leu604Pro e p.His1291Arg). Também identificamos um polimorfismo de único nucleotídeo (SNP) (rs115594471/ c.5874CT). Dois pacientes apresentaram deleção do gene CBP em um dos alelos, identificado pelo método array-CGH. Outro, apresentou uma translocação aparentemente equilibrada t(2;16), cuja análise subsequente com FISH revelou uma quebra na região do CBP. Neste trabalho, a taxa de detecção de alteração molecular no CBP por sequenciamento direto foi de 40% (08/20). Porém, a taxa de detecção das alterações moleculares no CBP foi de 55% (11/20), considerando a combinação das diferentes técnicas utilizadas (FISH, sequenciamento direto e array-CGH). Não houve correlação genótipo-fenótipo, exceto por uma maior frequência da presença de epicanto nos pacientes com alteração no CBP. Os resultados obtidos neste trabalho servem como o diagnóstico molecular para os pacientes com RTSs atendidos no Ambulatório do Laboratório de investigação Médica 001 (ALIM 001) do Instituto da Criança - FMUSP, contribuindo para uma melhor orientação médica, como também para realização do aconselhamento genético às famílias / Rubinstein-Taybi syndrome (RTSs) is a rare autosomal dominant disease characterized by craniofacial dysmorphisms, broad thumbs and toes, mental and growth deficiency. RTS has been associated with CREBBP (CBP) gene mutations and less frequently with mutations in EP300 gene, which have been reported in eight individuals. CBP and p300 have high homology and are extremely important in many signaling pathways especially as transcriptional coactivators and histone acetylation. Our study was based on the alteration analysis by direct sequencing of the CBP, by FISH and array-CGH in 20 RTSs patients. We identified eight molecular alterations in 20 RTSs patients evaluated by direct sequencing: i) two deletions (p.M747fs STOP830 and p.G1011fs STOP1021) ii) two nonsense alterations (p.Arg1341X and p.Arg1498X) iii) Three missense alteration (p.Arg1907Trp, p.Leu604Pro and p.His1291Arg). Single-nucleotide polymorphism were also identified (rs115594471 / c.5874CT), and six of these are new molecular alterations, not described in literature. Two RTSs patients studied had CBP gene deletion in one allele, identified by array-CGH method. Other patient, presented with apparent balanced translocation t(2;16) in which the subsequent analysis using FISH, showed a break in region of CBP. In this work, the rate of detection of molecular alteration in CBP by direct sequencing in RTSs patient was 40.0% (08/20). However, the rate of detection of molecular alteration in CBP was 55.0% (11/20), considering the combination of different techniques (FISH, direct sequencing and array-CGH. No significant correlation could be established in this study between the different types of mutations and genotype-phenotype of RTSs patients, except a higher frequency of the presence of epicanthus in the RTS patients with alteration in the CBP. The results of this study serve as a molecular diagnosis for RTSs patients treated at the Ambulatory of the Medical Investigation Laboratory 001 (ALIM 001) of the Instituto da Criança - FMUSP, and this contributes to better clinical management, such as making an appropriate genetic counseling for families

Michael Moore est une icône culturelle aux États-Unis. Son nom évoque la polémique, la contestation politique mais aussi le film documentaire à succès (Bowling For Columbine, primé aux Oscars, Fahrenheit 9/11, Palme d’Or à Cannes). Les six films du corpus, —Roger & Me (1989), The Big One (1998), Bowling For Columbine (2002), Fahrenheit 9/11 (2004), Sicko (2007) et Captain Mike Across America (Slacker Uprising) (2008) —, posent une triple problématique : la tension entre documentaire classique et documentaire hybride, la présence/absence du documentariste dans son film, et la réception du spectateur organisée par le divertissement et/ou la politique. Trois axes majeurs émergent, autour desquels l’articulation de la thèse peut s’effectuer : le divertissement, la politique et l’autosuffisance. La méthodologie pour rendre compte de cette triple problématique résulte d’un montage quali-quanti. Il s’agit de faire appel à l’analyse de discours mais aussi à l’analyse de contenu (pour mesurer la présence de manière quantitative). L’histoire américaine et l’histoire du documentaire sont également nécessaires pour situer Moore à la fois dans la lignée de certains de ses ainés, mais aussi pour mesurer sa propre influence sur le genre avec ses propres héritiers. Au bout du processus, il est possible de constater que Moore a insufflé un nouvel élan dans le genre du film documentaire en rejetant le modèle classique de type info-argument, rendant ses productions aussi populaires que des blockbusters hollywoodiens. Cette transformation vient de la combinaison inédite des finalités de divertissement et de politique pour créer un documentaire à la Michael Moore du type info-tainment. Avec l’ajout d’une troisième finalité d’autosuffisance, le documentaire moorien se signale aussi par une augmentation considérable de la présence du documentariste à l’écran et une densification de ses interventions au montage. En modifiant le documentaire classique, Moore a mis en place plusieurs stratégies de regard et de récit/storytelling afin de favoriser une assimilation efficace de ces trois finalités dans l’esprit du spectateur. / Michael Moore is a cultural icon in the United States. The mere mention of his name evokes polemics, political protest but also blockbuster documentary films (The Academy award winning Bowling For Columbine and Fahrenheit 9/11 which won the Palme d’Or at Cannes). The six films of this corpus—Roger & Me (1989), The Big One (1998), Bowling For Columbine (2002), Fahrenheit 9/11 (2004), Sicko (2007) and Captain Mike Across America (Slacker Uprising) (2008)—, are problematic in three ways: the discordant relationship between classic and hybrid documentary, the presence/absence of the documentary filmmaker in his own film, and the spectator’s reception of the film along the lines of entertainment and/or politics. Three main areas, thus, emerge around which the thesis can be constructed: entertainment, politics and self-sufficiency. The methodology used to attend to these three matters of discussion comes directly from quali-quanti research. This implies calling on discourse analysis but also content analysis (to measure Moore’s presence quantitatively).The history of the United States as well as that of the documentary film are also necessary to situate Moore not only in the footsteps of some of his predecessors but also to measure his own influence on the genre as evidenced by his very own successors. Ultimately, it can be noted that Moore has breathed new life into the documentary film genre by rejecting the traditional info-argument model, thereby making his films as successful as Hollywood blockbusters.This transformation arises from the unique combination of entertainment and politics to make a Michael Moore info-tainment style documentary. With the added third documentary finality of self-sufficiency, the Michael Moore documentary film is marked by a considerable increase of the filmmakers on-screen presence as well as density of the documentarian’s interventions in the editing room. By modifying the traditional documentary film form, Moore has established several strategies of gaze and narrative/storytelling in order to stimulate the effective absorption of his three main documentary finalities into the mind of the spectator.

De nombreux problèmes d'optimisation propres à différents secteurs industriels et académiques (énergie, chimie, transport, etc.) nécessitent de concevoir des méthodes de plus en plus efficaces pour les résoudre. Afin de répondre à ces besoins, l'objectif de cette thèse est de développer une bibliothèque composée de plusieurs métaheuristiques hybrides distribuées et massivement parallèles. Dans un premier temps, nous avons étudié le problème du voyageur de commerce et sa résolution par la méthode colonie de fourmis afin de mettre en place les techniques d'hybridation et de parallélisation. Ensuite, deux autres problèmes d'optimisation ont été traités, à savoir, le problème d'affectation quadratique (QAP) et le problème de la résolution structurale des zéolithes (ZSP). Pour le QAP, plusieurs variantes basées sur une recherche taboue itérative avec des diversifications adaptatives ont été proposées. Le but de ces propositions est d'étudier l'impact de : l'échange des données, des stratégies de diversification et des méthodes de coopération. Notre meilleure variante est comparée à six des meilleurs travaux de la littérature. En ce qui concerne le ZSP, deux nouvelles formulations de la fonction objective sont proposées pour évaluer le potentiel des structures zéolitiques trouvées. Ces formulations sont basées sur le principe de pénalisation et de récompense. Deux algorithmes génétiques hybrides et parallèles sont proposés pour générer des structures zéolitiques stables. Nos algorithmes ont généré actuellement six topologies stables, parmi lesquelles trois ne sont pas répertoriées sur le site Web du SC-IZA ou dans l'Atlas of Prospective Zeolite Structures. / Many optimization problems specific to different industrial and academic sectors (energy, chemicals, transportation, etc.) require the development of more effective methods in resolving. To meet these needs, the aim of this thesis is to develop a library of several hybrid metaheuristics distributed and massively parallel. First, we studied the traveling salesman problem and its resolution by the ant colony method to establish hybridization and parallelization techniques. Two other optimization problems have been dealt, which are, the quadratic assignment problem (QAP) and the zeolite structure problem (ZSP). For the QAP, several variants based on an iterative tabu search with adaptive diversification have been proposed. The aim of these proposals is to study the impact of: the data exchange, the diversification strategies and the methods of cooperation. Our best variant is compared with six from the leading works of the literature. For the ZSP two new formulations of the objective function are proposed to evaluate the potential of the zeolites structures founded. These formulations are based on reward and penalty evaluation. Two hybrid and parallel genetic algorithms are proposed to generate stable zeolites structures. Our algorithms have now generated six stable topologies, three of them are not listed in the SC-JZA website or in the Atlas of Prospective Zeolite Structures.

Spiroplasma citri est une bactérie phytopathogène transmise par la cicadelle Circuliferhaematoceps. L'absence de symptômes malgré la multiplication de S. citri dans l'hémolymphe, suggèreque le système immunitaire joue un rôle important dans la tolérance de la cicadelle vis-à-vis duspiroplasme.Le but de cette thèse a donc été d'étudier la réponse immunitaire de C. haematoceps aucours de l'infection par S. citri.Notre étude sur le système immunitaire de la cicadelle a montré la présence dans le plasma d'uneactivité antibactérienne et d'une activité phénoloxidase. Parmi les principaux types d'hémocytes unephagocytose des bactéries par les granulocytes et les plasmatocytes a été observée. Les gènessusceptibles d'être impliqués dans ces processus ont été recherchés par une approche par hybridationsoustractive. De manière étonnante, aucun gènes codant des récepteurs ni d'effecteurs connus del'immunité n'ont été identifiés. En revanche certains gènes (23 en tout) codent des protéines ayantpotentiellement un rôle immunitaire. Six de ces 23 gènes ont été retenus pour suivre leur expressionau temps précoce d'une infection bactérienne. Les résultats ont montré que les gènes codantI'Hexamérine, la DDBPl et la Thiorédoxine peroxydase étaient surexprimés lors de l'infection par 5.citri. Une approche fonctionnelle d'interférence par ARN a montré d'une part que I'Hexamérine étaitimpliquée dans l'activité phénoloxidase et d'autre part qu'elle jouait un rôle important dans la surviede C. haematococeps au cours de l'infection par 5. citri. En parallèle, le suivi de l'activité phénoloxidaseet de la phagocytose au cours de l'infection a montré que 5. citri était capable de s'adapter à laréponse immunitaire de l'insecte et d'y échapper. Ces résultats rejoignent ceux obtenus chez ladrosophile concernant S. poulsonii. / Spirop/asma citri is phytopathogenic bacteria transmitted by the leafhopper Circuliferhaematoceps. The absence of symptoms despite the multiplication of S. citri in the hemolymph,suggests that the immune system plays an important role in the tolerance of the leafhopper towardsthe spiroplasma infection. The purpose of this thesis was to study the immune response of C.haematoceps during the infection by 5. citri.The characterization of the immune system of the leafhopper showed that an antibacterial activity anda phenoloxidase activity were present in the plasma. The main types of hemocytes were identified.Among them, granulocytes and plasmatocytes are capable to phagocyte bacteria. The genes involvedin these immune processes were searched using subtractive hybridization method. lnterestingly, noneof the genes known to encode receptors or effectors of the immune system were identified. On theother hand 23 putative immune genes were identified. Six of these genes were retained to follow theirexpression in the early time of a bacterial infection. The results showed that the genes encodingHexamerin, DDBPl and Thioredoxin peroxidase were up-regulated during the infection by 5. citri. Afunctional approach by gene silencing showed that Hexamerin was involved in the phenoloxidaseactivity and played an important role in the survival of C. haematoceps during the infection by S. citri.Finally, the follow-up of the phenoloxidase activity and phagocytosis by hemocytes showed anadaptation and an evasion of S. citri from the immune response of the insect, according to the resultsobtained for 5. pou/sonii-infected drosophila.Keywords : 5piroplasma citri, phenoloxidase, phagocytosis, hemocytes, gene silencing, Hexamerine,subtractive hybridization.

Genetic changes are hallmarks of cancer development involving the activation and/or inactivation of oncogenes and tumour suppressor genes, respectively. In non-melanoma skin cancer (NMSC) development, the initiation of genetic mutations results from exposure to solar ultraviolet radiation. Non-melanoma skin cancers are comprised of basal cell carcinoma (BCC) and squamous cell carcinoma (SCC). Several related cutaneous lesions also exist, of which solar keratoses (SK) are widely accepted as a precursor dysplasia to SCC development. The study of recurrent genetic changes present within NMSC and SK should help reveal causative mutations in skin cancer development. Such analysis could also elucidate links in the genetic similarity of these dysplasia. The rapid screening of numerical changes in DNA sequence copy number throughout the entire genome has been made possible by the advent of comparative genomic hybridisation (CGH). This technique enables the identification of net gains and loss of genetic material within a tumour DNA sample. Chromosomal regions of recurrent gain or loss identify loci containing putative oncogenes and tumour suppressor genes, respectively with potential roles in NMSC tumourigenesis. Used in conjunction with tissue microdissection and universal degenerate PCR techniques this can enable the elucidation of aberrations in small histologically distinct regions of tumour. Such a technique can utilize archival material such as paraffin embedded tissue, which is the major source of neoplastic material available for cancer research. This study used the CGH technique to investigate aberrations in BCC, SCC and SK samples. The screening of copy number abnormalities (CNAs) in BCC revealed that although these tumours were close to diploid and generally genetically stable, they did contain several recurrent aberrations. The loss of genetic material at 9q was identified in a third of BCC tumours studied. This is characteristic of inactivation of the PTCH tumour suppressor gene, a known attribute in some sporadic BCC development. Validation of this loss was performed via loss of heterozygosity, demonstrating good concordance with the CGH data. In addition the over-representation of the 6p chromosome arm was revealed in 47% of biopsies. This novel CNA is also commonly observed in other cutaneous neoplasias, including Merkel cell carcinoma and malignant melanoma. This suggests a possible common mechanism in development and or promotion in these cutaneous dysplasias, the mechanisms of which have yet to be clearly defined. In contrast to BCC, numerical genetic aberrations in SCC and SK were much more frequent. Several regions of recurrent gain were commonly shared between both dysplasias including gain of 3q, 4p, 5p, 8q, 9q, 14q, 17p, 17q and 20q. Common chromosomal regions of loss included 3p, 8p, 9p, 11p, 13q and 17p. In addition loss of chromosome 18 was significantly observed in SCC in comparison to SK, a possible defining event in SK progression to SCC. The identification of shared genetic aberrations suggests a clonal and genetic relationship between the two lesions. This information further supports the notion for re-classification of SK to an SCC in situ or superficial SCC. Finally, the CNAs detected have been similarly observed in other squamous cell-derived tumours, for example cervical and head and neck SCC. This provides further evidence to common mechanisms involved in the initiation, development and progression of SCC neoplasia. This study has identified a number of recurrent chromosomal regions, some of which are novel in NMSC development. The further delineation of these loci should provide additional evidence of their significance and degree of involvement in NMSC tumourigenesis. The identification of the cancer-causing genes mapped to these loci will further demarcate the genetic mechanisms of BCC and SCC progression. An understanding of the events involved in skin cancer formation and progression should shed additional light on molecular targets for diagnostics, management and therapeutic treatment.

The occurrence of Boron (B) toxicity in Australian soils is recognised as a limiting factor for cereal productivity. A number of loci conferring tolerance to B toxicity have been identified in barley and chromosomally mapped. However, a lack of knowledge relating to the physiological and molecular events that occur under B toxicity and the molecular basis for B stress tolerance has been a bottleneck in harnessing available genetic diversity in barley and wheat. The recent advances in functional genomics provided an opportunity to examine B stress in barley in more detail. The aim of this project was to analyse genes differentially expressed under B stress in tolerant and intolerant barley to identify candidate genes involved in B toxicity tolerance. Two experimental approaches, Suppression Subtractive Hybridization (SSH) and microarray were adopted. Firstly, SSH was performed to examine gene expression in roots of selected tolerant and intolerant doubled haploid lines from a Clipper (B intolerant) X Sahara 3771 (B tolerant) mapping population, grown under moderate B stress. The SSH experiment aimed to investigate the early transcriptional response of B tolerant barley lines to B stress in order to identify the basis for B toxicity tolerance in roots. Differential screening of the subtracted library generated from B treated plants identified a total of 111 non-redundant clones up-regulated in bulked tolerant lines. On the other hand 94 clones were differentially expressed under non-treated conditions. Among the clones identified from subtracted library generated from B treated plants, metabolism was the largest functional category, representing 21% of the clones. The largest functional category in the subtracted library generated from non treated plants was cellular transport, representing 19% of the clones. Based on sequence similarity, about 170 transcripts identified in this experiment were assigned to chromosomal segments (bins) on the three homoeologous genomes of bread wheat. In total, 36 clones from the subtracted library generated from B treated plants were analysed as candidates. Nine were genetically mapped within the region of B tolerance QTL on three chromosomes (2H, 4H and 6H). The genes mapped to 4H and 6H QTL have the highest association with these loci in the Clipper X Sahara 3771 doubled haploid mapping population. A 4H B tolerance QTL candidate gene was identified as a B transporter gene with similarity to the Arabidopsis BOR1 gene. Genes identified to be differentially expressed in the tolerant lines from SSH suggest activation of a diverse defence response in the roots of barley plants under B stress. Data from SSH experiment indicate that cell wall-plasma membrane cytoskeleton continuum constitute the first action site against B toxicity and the influence of toxic B on K+ uptake could be the key initiating factor. In the second approach, the Affymetrix 22K Barley1 GeneChip(TM) was used to investigate B stress adaptation processes in barley. Gene expression was profiled in leaves of Sahara 3771 and Clipper plants grown under various B concentrations. The results show that the two genotypes respond differently to B toxicity. The B intolerance of Clipper is expressed through the induction of a high number of probe sets (2310) even at a low B concentration of 100 µM. In contrast, Sahara 3771 responded to a high B concentration (2000 µM) through the induction of only a few hundred (266) probe sets. In Sahara 3771 no change in the expression level of any probe sets was observed at 100 µM B. Altogether 286 probe sets showed differential expression in Sahara 3771 under three levels of B treatment (500, 1000 and 2000 µM). About 30% of these were down-regulated and about 70% were up-regulated in Sahara 3771 in response to B treatment. Most of the probe sets (59%) up-regulated in Sahara 3771 did not respond to B treatment in Clipper. These genes are either salt stress responsive or related to plant defense and thus could play a key role in protecting barley plants from the toxic effects of B. Two differentially expressed probe sets annotated as B transporters were identified between Sahara 3771 and Clipper under control condition. These two B transporter probe sets did not respond to B treatment but showed opposing expression patterns in the two varieties. One of these probe sets (Contig21126_at) is similar to the B transporter gene isolated from the SSH experiment that maps to the 4H tolerance locus. The map location and expression of this B transporter gene suggest that it could be the borate anion efflux transporter predicted by the proposed efflux model of B tolerance in Sahara 3771 barley. The other B transporter gene (Contig14139_at) showed over expression in Clipper under control condition and could be contributing to high B accumulation in Clipper which needs further investigation. Data from both experiments have indicated that B toxicity triggers oxidative stress and that jasmonate-based signaling plays a key role in B toxicity tolerance. SSH data indicate that Sahara 3771 which evolved in the harsh environment of Africa is more efficient in osmoregulation and ROS scavenging than Clipper. This trait is likely to give Sahara 3771 an edge over Clipper in tolerating toxic the effect of B. In addition to the efflux mechanism, which becomes less efficient with increasing B supply, Sahara 3771 appears to apply a number of other mechanisms for alleviating or withstanding toxic B induced stress to sustain growth. Some of these mechanisms are already known to be used by plants to cope with a number of stresses. / Thesis (Ph.D.) -- University of Adelaide, School of Agriculture, Food and Wine, 2008

<p>The telomeres are specialized structures at the end of the chromosomes composed of the repeated DNA sequence (TTAGGG)n and specific proteins bound to the DNA. The telomeres protect the chromosomes from degradation and end to end fusions. Due to the end-replication problem, the telomeric DNA shortens every cell division, forcing the cells into senescence at a critical telomere length. This process can be counteracted by activating a specialized enzyme, telomerase, which adds telomeric repeats to the chromosome ends leading to an extended or infinite cellular life span. Telomerase activity is absent in most somatic tissues but is found in germ cells, stem cells, activated lymphocytes and the vast majority of tumor cells and permanent cell lines. Hence, telomerase has been suggested as a target for cancer treatment as malignant cells almost exclusively express the enzyme and in that context telomere length measurements will be of great importance.</p><p>Telomere length is traditionally measured with a Southern blot based technique. A new method for telomere analysis of cells in suspension, called flow-FISH, was developed based on fluorescence in situ hybridization using a telomeric peptide nucleic acid (PNA) probe,</p><p>DNA staining with propidium iodide and quantification by flow cytometry. Flow-FISH had high reproducibility and the telomere length measurements showed good correlation with Southern blotting results. The flow-FISH technique also allows studies of cells in specific phases of the cell cycle and the replication timing of telomeric, centromeric and other repetitive sequences were analyzed in a number of cells. Like previous studies, centromeres were shown to replicate late in S phase while the telomere repeats were found to replicate early in S phase or concomitant with the bulk DNA, which is opposite to the patterns described in yeast.</p><p>In benign immunopurified lymphocytes from tonsils, high telomerase activity was found in germinal center (GC) B cells. This population also had high hTERT mRNA levels and displayed a telomere elongation as shown by flow-FISH and Southern blotting. Combined immunophenotyping and flow-FISH on unpurified tonsil cells confirmed the results.</p><p>Chronic lymphocytic leukemia (CLL), the most common leukemia in adults, can be divided into pre-GC CLL, characterized by unmutated immunoglobulin VH genes and worse prognosis, and post-GC CLL, with mutated VH genes and better prognosis. In 61 cases of CLL, telomere length was measured with Southern blotting and VH gene mutation status was analyzed. A new association was found between VH mutation status and telomere length, where cases with longer telomeres and mutated VH genes (post-GC CLL) had better prognosis</p><p>than CLL with short telomeres and unmutated VH genes (pre-GC CLL). A larger study of 112 CLL cases was performed using flow-FISH. The same correlation between telomere length and VH mutation status was found but gender seemed to be of importance as telomere length was a significant prognostic factor for the male CLL patients but not in the female group. Age of the patients and spread of disease seemed to affect the prognostic value of VH gene mutation status.</p>