All of the above findings demonstrated that eosinophil activation in allergic inflammation could be sensitively regulated by diverse stimuli. Besides, highly redundant functional effects and underlying signaling mechanisms were observed among different mediators. Elucidation of the mechanisms of eosinophil activation could improve our understanding of its complex and active role in the pathogenesis of allergic diseases, thereby providing biochemical basis for the development of more effective therapeutic strategies for treating the diseases. (Abstract shortened by UMI.) / Apart from the cytokines produced by T lymphoctyes, leptin, a cytokine produced by adipocytes, was also demonstrated to activate eosinophils. It was reported that the plasma levels of leptin were elevated in both obese and allergic patients. We found that leptin could activate eosinophils for survival enhancement, adhesion and migration, and secretion of cytokines and chemokines. Besides, we showed that the MAPKs and NF-kappaB pathways were involved in eosinophil adhesion, migration and mediator release induced by leptin, while Janus kinases (JAK)-signal transducers and activators of transcription (STAT) pathway was responsible for leptin-induced eosinophil survival. Our study indicated a potential correlation between obesity and exacerbation of allergic inflammation. / Eosinophilia is a hallmark pathological feature of allergic diseases and it has been targeted as a novel therapeutic strategy for allergic diseases. / In atopic dermatitis (AD), dermal infiltration of eosinophils is one of the pathological features of this disease. IL-31 is a novel Th2 cytokine reported to induce pruritus and skin dermatitis resembling human AD. Our study on the co-culture system of eosinophils and keratinocyte cell line HaCaT illustrated the production of pro-inflammatory cytokines and chemokines from the co-culture system of eosinophils and HaCaT cells, under the stimulation of IL-31. In co-culture system, surface expression of CD18 and intercellular adhesion molecule (ICAM)-1 on eosinophils and HaCaT cells was also up-regulated respectively, implying a direct interaction between the two cell types through their cell surface adhesion molecules. The interaction of eosinophils and HaCaT cells under IL-31 stimulation was shown to be mediated through MAPKs, NF-kappaB and PI3K pathways. These findings therefore elucidate the immunological roles of IL-31, eosinophils and keratinocytes in AD. / In the present study, we investigated the mechanisms of eosinophil activation induced by various stimuli including novel T helper type 2 (Th2) cytokines, adipokine, microbial products and direct interaction with tissue cells. The activation of eosinophils was studied in terms of survival enhancement, modulation of adhesion and migration, and the release of inflammatory mediators including cytokines, chemokines, granular proteins and superoxide. Using pharmacological and molecular approaches, we further investigated the intracellular signaling mechanisms regulating the eosinophil activation mediated by various stimuli. / Increasing evidence has indicated that bacterial and viral infections could intensify allergic responses. Our findings demonstrated that eosinophil activation could be elicited by microbes through toll-like receptors (TLRs), the recently discovered receptors for the recognition of conserved motifs in pathogens. We found that eosinophils could be activated by the ligands of TLR2, 5 and 7 in enhancing survival, adhesion and migration, release of pro-inflammatory cytokines, chemokines, granular proteins and superoxides. These stimulatory effects, mediated by TLR2, 5 and 7, were differentially regulated by MAPKs, NF-kappaB and phosphatidylinositol 3-kinase (PI3K) pathways. Moreover, an important finding of our study is the common involvement of focal adhesion kinase (FAK)-dependent extracellular-regulated protein kinase (ERK) phosphorylation in the signaling of TLR 2, 5 and 7, implying a special role of FAK in linking TLR signaling with MAPKs cascade in human eosinophils. Our study on microbe-induced eosinophil activation provided a potential explanation for linking infection with exacerbation of allergic diseases. / The interleukin (IL)-17 family is a newly discovered group of cytokines which was reported to be important in allergic inflammation. We studied the roles of two IL-17 family members, IL-175E/IL-25 and IL-17F on eosinophil activation. Both cytokines were found to induce the secretion of inflammatory cytokines and chemokines from eosinophils, in which IL-25 could also enhance eosinophil survival and adhesion. Besides, we found that the stimulatory effects induced by both IL-25 and IL-17F were mediated through mitogen-activated protein kinases (MAPKs) and nuclear factor kappa B (NF-kappaB) pathways. The main source of IL-17F has recently been reported to be a novel T lymphocyte population, Th17, which is specifically driven by IL-23. An important finding of our present study was the synergistic effects of IL-17F and its potent inducer, IL-23, on cytokine release from eosinophils. Since IL-23 was produced by macrophages and dendritic cells upon microbial stimulation, the synergistic effect of IL-17F and IL-23 on eosinophil activation might imply a potential role in linking microbial infection and allergic inflammation. Our findings also provide further support to the crucial role of the IL-17 family and Th17 lymphocytes in the amplification of allergic diseases. / Cheung, Fung Yi. / Source: Dissertation Abstracts International, Volume: 69-03, Section: B, page: 1552. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2007. / Includes bibliographical references (leaves 205-221). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English and Chinese. / School code: 1307.

Le système scolaire chilien a été parmi les premiers à instaurer un « pilotage par la qualité ». La Fondation Chili a été l’un des principaux acteurs de cette institutionnalisation. Le but de cette recherche doctorale est de découvrir comment les consultants en qualité de la Fondation Chili ont utilisé les objets techniques (Simondon, 1989) de leur boîte à outils pour influer sur l’implication des chefs d’établissements et de leurs équipes dans le processus d’institutionnalisation du pilotage par la qualité. Le champ de cohérence scientifique de cette recherche est celui de l’analyse institutionnelle (Lamihi & Monceau, 2002). Sept des dix anciens collègues du chercheur, toujours actifs comme consultants ont participé à cette recherche. Le dispositif de recherche a eu pour objectif prioritaire de permettre l’expression individuelle et collective des consultants pour faire émerger leurs implications en provocant des transductions dans leurs discours. La collecte des données empiriques a été effectuée à travers des entretiens individuels et collectifs, enregistrées en vidéo (Pesce, 2017). Le dispositif filmique a facilité la mise en réflexivité (Lallier, 2009 ; Rouch, 1979) des consultants sur leurs parcours et leurs pratiques au sein de la Fondation Chili. Pour élargir le champ d’analyse, quelques outils propres à l’analyse institutionnelle ont été intégrés au dispositif. Il s’agit tout d’abord de l’utilisation des analyseurs qui ont surgi des entretiens, ce que Lourau (1978) nommait des analyseurs naturels. J’ai également effectué une restitution des résultats avec celui des participants qui a été mon informateur privilégié, ceci afin d’affiner mes analyses. Enfin, l’écriture d’un journal de recherche a été essentielle pour l’analyse de mes propres implications primaires et secondaires en tant que praticien-chercheur. / The chilean school system has been among the first to introduce a « pilotage by the quality ». The Foundation Chile was one of the main actors of this institutionalization. The purpose of this doctoral research is to discover how the consultants in quality of the Foundation Chile have used the technical objects (Simondon, 1989) of their toolbox to influence the implication of school principals and their teams in the process of institutionalization of the pilotage by the quality. The field of scientific coherence of this research is that of the institutional analysis (Lamihi & Monceau, 2002). Seven of the ten former colleagues of the researcher are participated in this research. The research apparatus had for priority objective to allow the individual and collective expression of consultants, to make emerge their implications and by provoking transductions in their discourse. The collection of empirical data has been carried out through individual and collective interviews, all recorded in video (Pesce, 2017). The film apparatus facilitated the reflexivity (Lallier, 2009 ; Rouch, 1979) of consultants on their backgrounds and their practices within the Foundation Chile. To broaden the field of analysis, a few specific tools of the institutional analysis have been integrated to the research apparatus. The use of the analyzers which arose from interviews, that Lourau (1978) appointed of natural analyzers. I have also carried out a restitution of the results with that of participants who was my informant privileged, this in order to refine my analyzes. Finally, the writing of a diary study has been essential for the analysis of my own primary and secondary implications as a practitioner-researcher.

Liang, Fengting. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2010. / Includes bibliographical references (leaves 87-101). / Abstracts in English and Chinese. / DECLARATION --- p.I / ACKNOWLEDGEMENT --- p.II / ABBREVIATIONS --- p.III / ABSTRACT IN ENGLISH --- p.IV / ABSTRACT IN CHINESE --- p.VI / TABLE OF CONTENTS --- p.VIII / LIST OF FIGURES --- p.XI / LIST OF TABLES --- p.XIII / Chapter CHAPTER I- --- INTRODUCTION / Chapter 1.1 --- Roles of human bronchial epithelial cells --- p.1 / Chapter 1.2 --- Role of epithelium in airway inflammation --- p.3 / Chapter 1.3 --- Pathology of asthma --- p.5 / Chapter 1.4 --- The role of eosinophils in asthma --- p.7 / Chapter 1.5 --- "Effects of poly-L-arginine, a MBP analogue, on airway epithelium" --- p.10 / Chapter 1.6 --- Inflammatory pathways involved in epithelial cytokine production --- p.12 / Chapter 1.7 --- Roles and function of IL-6 and IL-8 in epithelial cells --- p.16 / Chapter 1.8 --- P2 receptors and inflammation --- p.18 / Chapter 1.9 --- Objectives --- p.20 / Chapter CHAPTER II- --- MATERIALS AND METHODS / Chapter 2.1 --- Materials and regents --- p.21 / Chapter 2.2 --- Cell culture --- p.22 / Chapter 2.3 --- RNA extraction and Real-time PCR --- p.23 / Chapter 2.4 --- Measurement of cytokine secretion by antibody array --- p.24 / Chapter 2.5 --- Quantification of IL-6 and IL-8 secretion --- p.27 / Chapter 2.6 --- Western Blotting --- p.28 / Chapter 2.7 --- NF-kB translocation assay --- p.29 / Chapter 2.8 --- Data analysis --- p.30 / Chapter CHAPTER III- --- RESULTS / Chapter 3.1 --- Poly-L-arginine-induced IL-6 and IL-8 release from 16HBE 14o- --- p.31 / Chapter 3.2 --- Signaling pathways involved in poly-L-arginine-induced IL-6 and IL-8 release --- p.34 / Chapter 3.2.1 --- "Effects of p38 MAPK, ERK1/2 and NF-kB inhibitors on poly-L-arginine-induced IL-6 and IL-8 release" --- p.35 / Chapter 3.2.2 --- Poly-L-arginine induces p38 MAPK and ERK1/2 phosphorylation --- p.43 / Chapter 3.2.3 --- Poly-L-arginine activates NF-kB translocation from cytoplasm to nucleus --- p.49 / Chapter 3.3 --- Effects of MAPK and NF-kB inhibitors on IL-6 and IL-8 mRNA expression on poly-L-arginine-challenged 16HBE14o- cells --- p.52 / Chapter 3.4 --- P2 receptors modulate poly-L-arginine-induced IL-6 and IL-8 ^ production --- p.55 / Chapter 3.4.1 --- Extracellular nucleotides modulate IL-6 and IL-8 production --- p.56 / Chapter 3.4.2 --- Effects of P2Y6 antagonist on poly-L-arginine-induced IL-6 and IL-8 production --- p.61 / Chapter 3.4.3 --- Effects of MAPKs inhibitors on UDP-induced IL-6 and IL-8 secretion --- p.64 / Chapter 3.4.4 --- UDP induces NF-kB translocation in 16HBE14o- cells --- p.67 / Chapter CHAPTER IV- --- DISCUSSION / Chapter 4.1 --- Involvement of p3 8 MAPK and NF-kB in poly-L-arginine-induced IL-6 and IL-8 secretion --- p.70 / Chapter 4.2 --- Involvement of p38 MAPK and NF-kB in poly-L-arginine-induced IL-6 and IL-8 mRNA elevation --- p.72 / Chapter 4.2.1 --- Regulation of NF-kB on IL-6 and IL-8 mRNA --- p.73 / Chapter 4.2.2 --- Regulation of p38 MAPK on IL-6 and IL-8 mRNA --- p.75 / Chapter 4.2.3 --- Crosstalk between NF-kB and p38 MAPK --- p.77 / Chapter 4.3 --- Extracellular nucleotides mediate IL-6 and IL-8 production in 16HBE14o- --- p.79 / Chapter 4.3.1 --- P2Y6 receptor is linked to poly-L-arginine-induced IL-6 and IL-8 release --- p.80 / Chapter 4.3.2 --- P2Y6 receptor regulates IL-6 and IL-8 secretion via p38 MAPK and NF-kB --- p.83 / Chapter 4.4 --- Summary --- p.86 / Chapter CHAPTER V- --- References --- p.87 / Publications --- p.102

Zhang, Siwei. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2008. / Includes bibliographical references (leaves 87-98). / Abstracts in English and Chinese. / Chapter 1. --- General introduction --- p.1 / Chapter 1.1. --- "Types, structure and function of human hemoglobin" --- p.1 / Chapter 1.1.1. --- Structure and functions of human hemoglobin --- p.1 / Chapter 1.1.2. --- Types of human hemoglobin --- p.2 / Chapter 1.2. --- Regulatory mechanism of human hemoglobin expression --- p.3 / Chapter 1.2.1. --- The human a and β locus --- p.3 / Chapter 1.2.2. --- Development of globin genes switching concept --- p.4 / Chapter 1.2.3. --- Factors that regulate globin gene expression --- p.5 / Chapter 1.2.3.1. --- The locus control region (LCR) --- p.5 / Chapter 1.2.3.2. --- The cis-regulatory elements --- p.5 / Chapter 1.2.3.3. --- The trans-acting factors --- p.6 / Chapter 1.3. --- The human hemoglobinopathies --- p.8 / Chapter 1.3.1. --- α-thalassemia --- p.8 / Chapter 1.3.2. --- β-thalassemia --- p.9 / Chapter 1.3.3. --- Sickle cell anemia --- p.10 / Chapter 1.4. --- Current approaches towards β-thalassemia treatment --- p.11 / Chapter 1.4.1. --- Blood transfusion --- p.11 / Chapter 1.4.2. --- Bone marrow transplantation --- p.12 / Chapter 1.4.3. --- Drug-induced activation of fetal hemoglobin production --- p.12 / Chapter 1.4.3.1. --- Hydroxyurea --- p.12 / Chapter 1.4.3.2. --- Butyrate and short-chain fatty acids --- p.13 / Chapter 1.4.3.3. --- "Mutagens, DNA methyltransferase inhibitors and other HbF inducible agents" --- p.13 / Chapter 1.4.3.4. --- Cucurbitacin D --- p.14 / Chapter 1.4.4. --- Gene therapy --- p.14 / Chapter 1.5. --- Research Objectives --- p.15 / Chapter 2. --- "Analysis of CuD, Hydroxyurea and other inducers on the induction of α, β, γ, δ， ε，ζ BP-1 genes and fetal hemoglobin induction" --- p.16 / Chapter 2.1. --- Introduction --- p.16 / Chapter 2.1.1. --- Properties of human K562 cell line --- p.16 / Chapter 2.1.2. --- Induction and measurement of fetal hemoglobin --- p.16 / Chapter 2.1.3. --- "Induction of α, β, γ, δ, ε , ζ and BP-1 gene and Real-time RT-PCR analysis" --- p.17 / Chapter 2.2. --- Materials --- p.18 / Chapter 2.2.1. --- Chemicals and reagents --- p.18 / Chapter 2.2.2. --- Kits --- p.19 / Chapter 2.2.3. --- Buffers and solutions --- p.19 / Chapter 2.2.4. --- Cell lines --- p.20 / Chapter 2.3. --- Experimental procedures --- p.20 / Chapter 2.3.1. --- Hemoglobin quantity measurement by HbF ELISA --- p.20 / Chapter 2.3.1.1. --- MTT assay --- p.21 / Chapter 2.3.1.2. --- Preparation of capture-antibody coated ELISA plates --- p.21 / Chapter 2.3.1.3. --- Plate blocking --- p.22 / Chapter 2.3.1.4. --- Sample and standard preparation --- p.22 / Chapter 2.3.1.5. --- HRP antibody and colorimetric detection --- p.23 / Chapter 2.3.1.6. --- Statistical analysis --- p.23 / Chapter 2.3.2. --- Preparation of mRNA extract from K562 cells --- p.23 / Chapter 2.3.3. --- Reverse transcription and Real-time PCR analysis --- p.24 / Chapter 2.4. --- Results --- p.25 / Chapter 2.4.1. --- CuD significantly upregulates HbF expression in K562 cells --- p.25 / Chapter 2.4.2. --- "CuD augments α, β, γ, δ, ε , ζ and BP-1 genes at different level in K562 cells" --- p.28 / Chapter 2.4.3. --- Cucurbitacin D-induced γ-globin gene activation requires12-24 hours in K562 cells --- p.31 / Chapter 2.5. --- Discussion --- p.33 / Chapter 2.5.1. --- Enhancement of fetal hemoglobin production using different chemical compounds --- p.33 / Chapter 2.5.2. --- CuD increased HbF synthesis by increasing γ-globin mRNA amount --- p.35 / Chapter 2.5.3. --- CuD and HU down-regulated the BP-1 gene expression --- p.36 / Chapter 3. --- Determination of potential signal transduction pathways during CuD and HU-mediated fetal hemoglobin production --- p.36 / Chapter 3.1. --- Introductions --- p.36 / Chapter 3.1.1. --- The p38 MAPK family --- p.37 / Chapter 3.1.2. --- The JAK2-STAT3 pathway --- p.38 / Chapter 3.1.3. --- Fundamentals on inhibition assay of p38 MAPK and JAK2-STAT3 pathway --- p.39 / Chapter 3.1.4. --- Fundamentals on nuclear translocation of STAT3 --- p.41 / Chapter 3.2. --- Materials --- p.41 / Chapter 3.2.1. --- Chemicals and reagents --- p.41 / Chapter 3.2.2. --- Kits --- p.44 / Chapter 3.2.3. --- Buffers and solutions --- p.44 / Chapter 3.3. --- Experimental procedures --- p.45 / Chapter 3.3.1. --- Detection of p3 8 MAPK phosphorylation status --- p.46 / Chapter 3.3.1.1. --- Preparation of cytosolic protein extracts --- p.46 / Chapter 3.3.1.2. --- Quantitative measurement of phospho-p38 and pan-p38 by ELIS A method --- p.46 / Chapter 3.3.1.2.1. --- Antigen adsorption and establishment of standard curves --- p.46 / Chapter 3.3.1.2.2. --- Plate washing and application of detection antibody --- p.47 / Chapter 3.3.1.2.3. --- Plate washing and application of secondary antibody --- p.47 / Chapter 3.3.1.2.4. --- Plate washing and chromogen detection --- p.48 / Chapter 3.3.2. --- Detection of signal cascade on JAK2-STAT3 pathway --- p.48 / Chapter 3.3.2.1. --- Preparation of cytosolic protein extracts for Western Blot detection --- p.48 / Chapter 3.3.2.2. --- Gel running and Western Blot detection --- p.48 / Chapter 3.3.3. --- Quantitative measurement of phospho-STAT3-Tyr705 using ELISA method --- p.50 / Chapter 3.3.3.1. --- Preparation of cytosolic protein extracts --- p.50 / Chapter 3.3.3.2. --- Reconstitution and Dilution of STAT3 [pY705] Standard --- p.50 / Chapter 3.3.3.3. --- Measurement of STAT3 [pY705] concentration in cell lysates --- p.51 / Chapter 3.3.4. --- Inhibitor assay of JAK2-STAT3 and p38 MAPK pathway --- p.52 / Chapter 3.3.4.1. --- Establishment of inhibitor assay --- p.52 / Chapter 3.3.4.2. --- HbF ELISA detection --- p.53 / Chapter 3.3.5. --- Detection of STAT3 nuclear translocation and DNA binding affinity --- p.53 / Chapter 3.3.5.1. --- Preparation of nuclear extract from K562 cells --- p.53 / Chapter 3.3.5.2. --- EMS A detection of transcriptional factors binding to γ-promoter region --- p.54 / Chapter 3.3.5.2.1. --- 3´ة end-labeling of EMS A probes --- p.54 / Chapter 3.3.5.2.2. --- Dot blotting for labeling efficiency estimation --- p.56 / Chapter 3.3.5.2.3. --- EMSA binding reaction and non-denaturing gel electrophoresis --- p.57 / Chapter 3.3.5.2.4. --- Membrane development and chemiluminescence detection --- p.58 / Chapter 3.3.5.3. --- Preparation of K562 samples for immunofluorescence detection --- p.60 / Chapter 3.3.5.3.1. --- Slide coating for cell capture --- p.60 / Chapter 3.3.5.3.2. --- Preparation of cell slide --- p.60 / Chapter 3.3.5.3.3. --- Sample fixation and antibody probing treatment --- p.60 / Chapter 3.3.5.3.4. --- Sample imaging and immunofluorescence detection --- p.61 / Chapter 3.4 --- Results --- p.62 / Chapter 3.4.1. --- Activation of p38 MAPK pathway and STAT3 phosphorylation by hydroxyurea --- p.62 / Chapter 3.4.1.1. --- "The p38 MAPK pathway is activated by hydroxyurea, but not activated by Cucurbitacin D" --- p.62 / Chapter 3.4.1.2. --- Increased p38 phosphorylation level elicits STAT3 phosphorylation at Ser727 site --- p.64 / Chapter 3.4.2. --- Activation of JAK2 and STAT3 phosphorylation by Cucurbitacin D --- p.66 / Chapter 3.4.2.1. --- Cucurbitacin D promotes JAK2 activation --- p.66 / Chapter 3.4.2.2. --- Cucurbitacin D and hydroxyurea promote STAT3 phosphorylation at Tyr705 site --- p.66 / Chapter 3.4.3. --- Basal activity of signal transduction pathways is essential for HbF induction --- p.69 / Chapter 3.4.3.1. --- Activation of γ-globin gene requires presence of basal phosphorylation level of p38 MAPK --- p.69 / Chapter 3.4.3.2. --- Inhibition on JAK2-STAT3 pathway results in reduced fetal hemoglobin production --- p.71 / Chapter 3.4.4. --- Translocation and DNA binding of STAT under Cucurbitacin D induction --- p.72 / Chapter 3.4.4.1. --- Cucurbitacin D and hydroxyurea both enhance binding affinity of transcriptional factors to the Gγ/Aγ promoter --- p.72 / Chapter 3.4.4.2. --- Cucurbitacin D and hydroxyurea induces nuclear translocation of STAT3 --- p.75 / Chapter 3.5. --- Discussion --- p.77 / Chapter 3.5.1. --- The role of p38 MAPK activation during γ-globin gene activation --- p.77 / Chapter 3.5.2. --- STAT3 phosphorylation at Ser727 site promotes transcription factor activity and γ-globin gene expression --- p.77 / Chapter 3.5.3. --- The role of JAK2-STAT3 activation during γ-globin gene activation --- p.78 / Chapter 3.5.4. --- Inhibitor assay --- p.79 / Chapter 3.5.5. --- Relations between STAT3 nuclear translocation and enhanced fetal hemoglobin production --- p.82 / Chapter 4. --- Summery and Prospect --- p.83 / Chapter 5. --- References --- p.87

Zhong, Lan. / Thesis Ph.D. Chinese University of Hong Kong 2014. / Includes bibliographical references (leaves 122-154). / Abstracts also in Chinese. / Title from PDF title page (viewed on 24, October, 2016). / Zhong, Lan.

Lam, Kwok Ho. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2011. / Includes bibliographical references (leaves 149-159). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.

Thesis (Ph.D.)--University of Minnesota, 1999. / Includes bibliographical references (leaves 74-98). Also available on the World Wide Web as a PDF file.

Manfouo Foutsop Germain (2010). Etude de lexpression des gènes cellulaires dérégulés dans des cellules T CD4+ exprimant Tat de HIV-1 : Focalisation sur la voie de signalisation TCR/CD3 (thèse de doctorat). Université de Liège-Gembloux Agro-Bio Tech, 157 pages, 11 tables, 29 figures. Résumé Nos travaux précédents ont suggéré que les gènes nef (negative factor) et tat (transactivateur de la transcription) du virus de limmunodéficience humaine de type 1 (HIV-1) pourraient induire la perte du récepteur des cellules T (TCR)/CD3 à la surface des lymphocytes T CD4+ infectés par le HIV-1, mais la contribution de tat dans cette perte navait pas été prouvée. Dans la première partie de ce travail nous avons utilisé un vecteur lentiviral exprimant les formes natives [1exon (72aa) ou 2exons (101aa)] ou des formes mutées du gène tat de HIV-1 pour transduire les cellules T CD4+. Ceci nous a permis de montrer que la production de la protéine Tat72aa ou Tat101aa entraîne une perte progressive du complexe TCR/CD3 en surface des cellules T CD4+. Nous avons constaté que cette perte de TCR/CD3 serait due à une diminution du taux des transcrits de CD3γ. Par ailleurs, nous avons montré que certains mutants du gène tat ont perdu leur activité transactivatrice et sont incapables dinduire la perte de TCR/CD3. Dans la deuxième partie des travaux, nous avons identifié par microarray, les gènes cellulaires dont les expressions sont communément altérées dans les cellules T CD4+ infectées par le HIV-1 ou transduites avec le gène tat-1exon ou tat-2exons. Plusieurs de ces gènes sont impliqués dans la voie de signalisation TCR/CD3. Nos travaux nous ont donc permis de montrer limplication du gène tat dans la dérégulation de la voie de signalisation TCR/CD3. Par conséquent, Tat pourrait induire une altération de la réponse immunitaire favorisant ainsi la progression de linfection par le HIV-1/SIDA. Manfouo Foutsop Germain (2010). A study of altered cellular gene expression in CD4+ T cells expressing HIV-1 Tat: Focus on the T cell receptor/CD3-directed pathway (PhD thesis). Université de Liège-Gembloux Agro-Bio Tech, 157 pages, 11 tables, 29 figures. Summary Our previous work has suggested that nef (negative factor) and tat (transactivator of transcription) are responsible for T cell receptor (TCR)/CD3 downmodulation on CD4+ T cells after infection with human immunodeficiency virus type 1 (HIV-1). However, the contribution of tat to this phenomenon has not been investigated. In the first part of this study we used lentiviral vector expressing the native tat gene [1exon (72aa) or two exon (101aa)] and various mutants to transduce CD4+ T cells. We found that Tat expression alone could provoke the progressive downmodulation of surface TCR/CD3 complexes due to the decrease of CD3γ transcripts. We also demonstrated that the majority of mutants lost their transactivation capability and their ability to downregulate the TCR/CD3 complex. In the second part of this work, we used gene expression arrays to identify cellular genes whose expression was altered in CD4+ T cells infected with HIV-1 or transduced with tat at different stages of receptor downregulation. We found progressive alteration of cellular gene expression in parallel with TCR/CD3 downregulation in the tat-transduced cells, which represented a subset of the changes detected in the HIV-1 infected cells, with significant number genes known to play a role in the TCR/CD3 signaling pathway. Our data demonstrate that tat deregulates the TCR/CD3 expression and signaling leading to altered immune function that likely favor HIV-1/AIDS disease progression. Copyright: Aux termes de la loi belge du 30 juin 1994, sur le droit dauteur et les droits voisins, seul lauteur a le droit de reproduire partiellement ou complètement cet ouvrage de quelque façon que ce soit ou dautoriser la reproduction partielle ou complète de quelque manière et sous quelque forme que ce soit. Toute photocopie ou reproduction sous autre forme est donc faite en violation de ladite loi et des modifications ultérieures.

High-throughput genome processing, sophisticated protein sequence analysis, programming, and information management were used to achieve two major advances in the comparative genomics of microbial signal transduction. First, an integrated and flexible bioinformatics platform and the Microbial Signal Transduction database (MiST) were developed, which facilitated the genome-wide analysis of bacterial signal transduction. This platform was used successfully for the high-throughput identification and classification of signal transduction proteins in more than 300 archaeal and bacterial organisms. Second, analysis of information encoded in prokaryotic genomes revealed that the majority of signal transduction systems consist of one-component systems a single protein containing both input and output domains but lacking phosphotransfer domains typical of two-component systems. The prevalence of one-component systems is a paradigm-shifting discovery because two-component systems are currently viewed as the primary mode of signal transduction in prokaryotes. One-component systems are more widely distributed among bacteria and archaea and display a greater diversity of domains than two-component systems. Additionally, in-depth bioinformatic analyses were performed that further characterized the function of two, input, signaling domains. In summary, this systematic, high-throughput delineation of microbial signal transduction is another step forward in our understanding of the genomic basis of life.

The general goal of this project was to use computational biology to understand signal transduction mechanisms in prokaryotes. Its specific focus was to characterize the cytoplasmic domain of methyl-accepting chemotaxis proteins (MCP_CD), a protein domain central to the function of chemotaxis, the most complex signaling network in prokaryotes. Chemotaxis enables cells to sense and respond to multiple external and internal stimuli by actively navigating to an optimal environment. MCP_CD is a central part of this circuit, but its coiled coil structure is difficult to analyze using traditional tools of computational biology. In this project, a new method for analysis of the domain was developed and used to gain insight into its function and evolution. Research advance 1: Characterization of the MCP_CD protein domain. Before this work, MCP_CD was known to have two distinct functional regions: the signaling region that activates the histidine kinase CheA and the methylation region where adaptation enzymes CheB and CheR store information about recent stimuli. The result of this project is classification of ~2000 MCP_CDs into twelve subfamilies. The unique mechanism of evolution of the domain has been clarified and precise boundaries of the adaptation and signaling regions determined. A new functional region, the flexible bundle subdomain, was identified and its contribution to the signaling mechanism elucidated by analysis of conserved sequence features. Conserved and variable sequence features in the adaptation and signaling subdomains led to a better understanding of the evolutionary history of the adaptation mechanism and of alternative higher-order arrangements of receptors within the membrane. Research advance 2: Development of a sensor / kinase correlation algorithm to couple diverse MCP_CD and kinase subfamilies. The receptor diversity discovered in this work is complemented by diversity in the kinases with which they interact. In this work, an algorithm was developed to associate receptor / kinase pairs which facilitated understanding of the function and evolution of chemotaxis. Research advance 3: Development of Cheops, a database of chemotaxis pathways. The Cheops (Chemotaxis operons) database presents the results of the sensor / kinase correlation algorithm and the information about receptor and kinase diversity in an integrated and intuitive way.