Role of β-catenin and γ-catenin in haematopoiesis : Generation of floxed Delta like 4 gene targeted mice

Résumé La voie de signalisation de Wnt est extrêmement conservée au cours de l'évolution. Les protéines Wnt sont des molécules sécrétées qui se lient à la famille de récepteurs Frizzled. Cette interaction mène à la stabilisation de la protéine β-caténine, qui va s'accumuler dans le cytoplasme puis migrer dans le noyau où elle peut s'hétérodimériser avec les facteurs de transcription de la famille TCF/LEF. Il a été démontré que cette voie de signalisation joue un rôle important durant la lymphopoïèse et de récents résultats suggèrent un rôle clé de cette voie dans le renouvellement des Cellules Souches Hématopoïétique (CSH). Des études se basant sur un système de surexpression de protéines montrent clairement que la voie Wnt peut influencer l'hématopoïèse. Cependant, le rôle de la protéine β-caténine dans le système hématopoïétique n'a jamais été testé directement. Ce projet de thèse se propose d'étudier la fonction de la protéine β-caténine par sa délétion inductible via le système Cre-loxP. De façon surprenante, nous avons pu démontrer que les progéniteurs de la moelle osseuse, déficients en β-caténine, ne montrent aucune altération dans leur capacité à s'auto-renouveler et/ou à reconstituer toutes les lignées hématopoïétiques (myéloïde, érythroïde et lymphoïde) dans les souris-chimères. De plus, le développement, la survie des thymocytes ainsi que la prolifération des cellules T périphériques induite par un antigène, sont indépendants de β-caténine. Ces résultats suggèrent soit que la protéine β-caténine ne joue pas un rôle primordial dans le système hématopoiétique, soit que son absence pourrait être compensée par une autre protéine. Un candidat privilégié susceptible de se substituer à β-caténine, serait plakoglobine, aussi connu sous le nom de γ-caténine. En effet, ces deux protéines partagent de multiples caractéristiques structurelles. Afin de démontrer que la protéine γ-caténine peut compenser l'absence de β-caténine, nous avons généré des souris dans lesquelles, le système hématopoïétique est déficient pour ces deux protéines. Cette déficience combinée de β- caténine et γ-caténine ne perturbe pas la capacité des Cellules Souche Hématopoïétique-Long Terme (CSH-LT) de se renouveler, par contre elle agit sur un progéniteur précoce déjà différencié de la moelle osseuse. Ces résultats mettent en évidence que la protéine γ-caténine est capable de compenser l'absence de protéine β-caténine dans le système hématopoïétique. Par conséquent, ce travail contribue à une meilleure connaissance de la cascade Wnt dans l'hématopoïèse. Summary The canonical Wnt signal transduction pathway is a developmentally highly conserved. Wnts are secreted molecules which bind to the family of Frizzled receptors in a complex with the low density lipoprotein receptor related protein (LRP-5/6). This initial activation step leads to the stabilization and accumulation of β-catenin, first in the cytoplasm and subsequently in the nucleus where it forms heterodimers with TCF/LEF transcription factor family members. Wnt signalling has been shown to be important during early lymphopoiesis and has more recently, been suggested to be a key player in self-renewal of haematopoietic stem cells (HSCs). Although mostly gain of function studies indicate that components of the Wnt signalling pathway can influence the haematopoietic system, the role of β-catenin has never been directly investigated. The aim of this thesis project is to investigate the putatively critical role of β-catenin in vivo using the Cre-loxP mediated conditional loss of function approach. Surprisingly, β-catenin deficient bone marrow (BM) progenitors arc not impaired in their ability to self-renew and/or to reconstitute all haematopoietic lineages (myeloid, erythroid and lymphoid) in both mixed and straight bone marrow chimeras. In addition, both thymocyte development and survival, and antigen-induced proliferation of peripheral T cells are β- catenin independent. Our results do not necessarily exclude the possibility of an important function for β-catenin mediated Wnt signalling in the haematopoietic system, it rather raises the question that β-catenin is compensated for by another protein. A prime candidate that may take over the function of β-catenin in its absence, is the close relative plakoglobin, also know as γ-catenin. This protein shares multiple structural features with β-catenin. In order to investigate whether γ-catenin can compensate for the loss of β-catenin we have generated mice in which the haematopoietic compartment is deficient for both proteins. Combined deficiency of β-catenin and γ-catenin does not perturb Long Term-Haematopoietic Stem Cells (LT-HSC) self renewal, but affects an already lineage committed progenitor population within the BM. Our results demonstrate that y-catenin can indeed compensate for the loss of β-catenin within the haematopoietie system.

Analysis of experiments in square lattice with emphasis on variance components. i. Individual analysis

This paper focused on four alternatives of analysis of experiments in square lattice as far as the estimation of variance components and some genetic parameters are concerned: 1) intra-block analysis with adjusted treatment and blocks within unadjusted repetitions; 2) lattice analysis as complete randomized blocks; 3) intrablock analysis with unadjusted treatment and blocks within adjusted repetitions; 4) lattice analysis as complete randomized blocks, by utilizing the adjusted means of treatments, obtained from the analysis with recovery of interblock information, having as mean square of the error the mean effective variance of this same analysis with recovery of inter-block information. For the four alternatives of analysis, the estimators and estimates were obtained for the variance components and heritability coefficients. The classification of material was also studied. The present study suggests that for each experiment and depending of the objectives of the analysis, one should observe which alternative of analysis is preferable, mainly in cases where a negative estimate is obtained for the variance component due to effects of blocks within adjusted repetitions.

Variation in soil nutrient concentrations and bulk density within peatland forest sites

Abstract

Generation of biological association networks: A novel strategy to detect new targets in cancer therapy

The aim of this work was to design a novel strategy to detect new targets for anticancer treatments. The rationale was to build Biological Association Networks from differentially expressed genes in drug-resistant cells to identify important nodes within the Networks. These nodes may represent putative targets to attack in cancer therapy, as a way to destabilize the gene network developed by the resistant cells to escape from the drug pressure. As a model we used cells resistant to methotrexate (MTX), an inhibitor of DHFR. Selected node-genes were analyzed at the transcriptional level and from a genotypic point of view. In colon cancer cells, DHFR, the AKR1 family, PKC¿, S100A4, DKK1, and CAV1 were overexpressed while E-cadherin was lost. In breast cancer cells, the UGT1A family was overexpressed, whereas EEF1A1 was overexpressed in pancreatic cells. Interference RNAs directed against these targets sensitized cells towards MTX.

Evolutionarily stable learning schedules and cumulative culture in discrete generation models.

Individual learning (e.g., trial-and-error) and social learning (e.g., imitation) are alternative ways of acquiring and expressing the appropriate phenotype in an environment. The optimal choice between using individual learning and/or social learning may be dictated by the life-stage or age of an organism. Of special interest is a learning schedule in which social learning precedes individual learning, because such a schedule is apparently a necessary condition for cumulative culture. Assuming two obligatory learning stages per discrete generation, we obtain the evolutionarily stable learning schedules for the three situations where the environment is constant, fluctuates between generations, or fluctuates within generations. During each learning stage, we assume that an organism may target the optimal phenotype in the current environment by individual learning, and/or the mature phenotype of the previous generation by oblique social learning. In the absence of exogenous costs to learning, the evolutionarily stable learning schedules are predicted to be either pure social learning followed by pure individual learning ("bang-bang" control) or pure individual learning at both stages ("flat" control). Moreover, we find for each situation that the evolutionarily stable learning schedule is also the one that optimizes the learned phenotype at equilibrium.

Testing the effects of genetic crossing distance on embryo survival within a metapopulation of brown trout (Salmo trutta)

Predicting progeny performance from parental genetic divergence can potentially enhance the efficiency of supportive breeding programmes and facilitate risk assessment. Yet, experimental testing of the effects of breeding distance on offspring performance remains rare, especially in wild populations of vertebrates. Recent studies have demonstrated that embryos of salmonid fish are sensitive indicators of additive genetic variance for viability traits. We therefore used gametes of wild brown trout (Salmo trutta) from five genetically distinct populations of a river catchment in Switzerland, and used a full factorial design to produce over 2,000 embryos in 100 different crosses with varying genetic distances (FST range 0.005-0.035). Customized egg capsules allowed recording the survival of individual embryos until hatching under natural field conditions. Our breeding design enabled us to evaluate the role of the environment, of genetic and nongenetic parental contributions, and of interactions between these factors, on embryo viability. We found that embryo survival was strongly affected by maternal environmental (i.e. non-genetic) effects and by the microenvironment, i.e. by the location within the gravel. However, embryo survival was not predicted by population divergence, parental allelic dissimilarity, or heterozygosity, neither in the field nor under laboratory conditions. Our findings suggest that the genetic effects of inter-population hybridization within a genetically differentiated meta-population can be minor in comparison to environmental effects.

Chemical and isotopic equilibrium between CO2 and CH4 in fumarolic gas discharges: Generation of CH4 in arc magmatic-hydrothermal systems

The chemical and isotopic composition of fumarolic gases emitted from Nisyros Volcano, Greece, and of a single gas sample from Vesuvio, Italy, was investigated in order to determine the origin of methane (CH,) within two subduction-related magmatic-hydrothermal environments. Apparent temperatures derived from carbon isotope partitioning between CH4 and CO2 of around 340degreesC for Nisyros and 470degreesC for Vesuvio correlate well with aquifer temperatures as measured directly and/or inferred from compositional data using the H2O-H-2-CO2-CO-CH4 geothermometer. Thermodynamic modeling reveals chemical equilibrium between CH4, CO2 and H2O implying that carbon isotope partitioning between CO2 and CH, in both systems is controlled by aquifer temperature. N-2/(3) He and CH4/(3) He ratios of Nisyros fumarolic gases are unusually low for subduction zone gases and correspond to those of midoceanic ridge environments. Accordingly, CH4 may have been primarily generated through the reduction of CO, by H, in the absence of any organic matter following a Fischer-Tropsch-type reaction. However, primary occurrence of minor amounts of thermogenic CH4 and subsequent re-equilibration with co-existing CO2 cannot be ruled out entirely- CO2/He-3 ratios and delta(13)C(CO2) values imply that the evolved CO2 either derives from a metasomatized mantle or is a mixture between two components, one outgassing from an unaltered mantle and the other released by thermal breakdown of marine carbonates. The latter may contain traces of organic matter possibly decomposing to CH4 during thermometamorphism. Copyright (C) 2004 Elsevier Ltd.

The Architecture of a Fourth Generation Mobile Network

Neljännen sukupolven mobiiliverkot yhdistävät saumattomasti televerkot, Internetin ja niiden palvelut. Alkuperin Internetiä käytettiin vain paikallaan pysyviltä tietokoneilta perinteisten televerkkojen tarjotessa puhelin- ja datapalveluita. Neljännen sukupolven mobiiliverkkojen käyttäjät voivat käyttää sekä Internetiin perustuvia että perinteisten televerkkojen palveluita liikkuessaankin. Tämä diplomityö esittelee neljännen sukupolven mobiiliverkon yleisen arkkitehtuurin. Arkkitehtuurin perusosat kuvaillaan ja arkkitehtuuria verrataan toisen ja kolmannen sukupolven mobiiliverkkoihin. Aiheeseen liittyvät Internet-standardit esitellään ja niiden soveltuvuutta mobiiliverkkoihin pohditaan. Langattomia, lyhyen kantaman nopeita liitäntäverkkotekniikoita esitellään. Neljännen sukupolven mobiiliverkoissa tarvittavia päätelaitteiden ja käyttäjien liikkuvuuden hallintamenetelmiä esitellään. Esitelty arkkitehtuuri perustuu langattomiin, lyhyen kantaman nopeisiin liitäntäverkkotekniikoihin ja Internet-standardeihin. Arkkitehtuuri mahdollistaa yhteydet toisiin käyttäjiin ilman tietoa heidän senhetkisestä päätelaitteesta tai sijainnista. Internetin palveluitavoidaan käyttää missä tahansa neljännen sukupolven mobiiliverkon alueella. Yleiskäytöistä liikkuvuuden hallintamenetelmää yhden verkon alueelle ehdotetaan. Menetelmää voidaan käyttää yhdessä esitellyn arkkitehtuurin kanssa.

Transport Resource Management within UMTS Radio Network Subsystem

Tällä hetkellä haastavin telekommunikaatioteollisuuden tutkimus – ja kehitystoiminta on keskittynyt kolmannen sukupolven matkapuhelinjärjestelmien ympärille. Järjestelmien standardointityössä on saatu aikaiseksi ensimmäiset vakaat spesifikaatioversiot ja kaupallista toimintaa ollaan parhaillaan aloittelemassa Japanissa ja Euroopassa. Eräs kolmannen sukupolven järjestelmistä on UMTS (Universal Mobile Telecommunications System). Tämä diplomityö antaa yleiskuvan UMTS järjestelmästä ja sen eri verkkoelementtien toiminnallisuuksista. Päähuomio on kiinnitetty radioverkkojärjestelmään (UMTS Terrestrial Radio Access Network) ja erityisesti sen radioaliverkkojärjestelmään (Radio Network Subsystem), joka koostuu radioverkonohjaimesta (Radio Network Controller) ja joukosta siihen kuuluvia tukiasemia (Node B). Radioverkonohjain ja tukiasemat on yhdistetty avoimen rajapinnan kautta jota kutsutaan Iub -rajapinnaksi. Rajapinta tarjoaa radioverkonohjaimelle mahdollisuuden kontrolloida tukiasemia signalointiviestien avulla ja mahdollistaa tehokkaan ja luotettavan käyttäjätiedon siirron radioaliverkkojärjestelmän sisällä. Tämän diplomityön pääasiallinen sisältö on siirtoresurssien hallinta Iub -rajapinnan ylitse. Työssä esitellään ja selitetään siirtoverkon arkkitehtuuri. Myös kaikki Iub:ssä sijaitsevat protokollat ja toiminnalliset yksiköt jotka vaikuttavat siirtoresurssien hallintaan esitellään ja kuvataan yksityiskohtaisesti. Päähuomio on kiinnitetty sovellusprotokolliin sekä rajapinnan siirtoverkko- että radioverkkokerroksella sekä näiden protokollien väliseen vuorovaikutukseen. Kyseiset protokollat ovat Node B Application Part (NBAP) ja Access Link Control Application Part (ALCAP). Työn toteutusosassa käydään lävitse NBAP –protokollan prototyypin ja Node B Manager –toiminnallisen yksikön prototyypin implementaatio.

Standardisation process of Universal Mobile Telecommunication System in Third Generation Partnership Project

Third Generation Partnership Project (3GPP) on organisaatio, joka määrittelee ja ylläpitää kolmannen sukupolven matkapuhelinverkon standardeja. Organisaatio luotiin monien eri standardointielinten toimesta havaittaessa, ettei maailmanlaajuista kolmannen sukupolven matkapuhelinteknologiaa voitaisi määritellä ilman laajaa yhteistyötä. 3GPP:ssä standardointityö on jakautunut usealle tekniselle määrittelyryhmälle. Jokaisen ryhmän tehtävänä on kehittää määrittelyjä ja raportteja omalla vastuualueellaan. 3GPP:ssä määrittelytyötä tehdään samanaikaisesti teknillisten määrittelyryhmien välillä. Tämä vaatii tiukkoja sääntöjä määrittelyjen luonti-, hyväksyntä- ja ylläpitotehtäviin. Vain siten on mahdollista hallita määrittelyihin tulevia muutoksia ja tarvittavaa kokonaistyömäärää. Tämä diplomityö kuvaa 3GPP:n määrittelemän UMTS-teknologian. Työssä keskitytään tarkemmin 3GPP-organisaation rakenteeseen, määritysten tekemiseen ja työskentelytapoihin. Tämä diplomityö osoittaa millainen organisaatio ja säännöt vaaditaan maailmanlaajuisen matkapuhelinjärjestelmän kehittämiseen.

Performance Management of a Next Generation Network

Today's communication networks consist of numerous interdependent network components. To manage these networks and to ensure their reliable and efficient operation to meet the increasing customer usability demands, extensive network management tools are required from the service provider. The goal of this study was to adapt the Next Generation Network (NGN) providing VoIP services within a performance oriented network management system. This study focuses only on NGN network and the project was implemented as an assignment of the Network Operations Center of Elisa Corporation. The theoretical part of this study introduces the network environment of the Elisa NGN platform: its components and used signalling protocols as well as other exploitable communication protocols. In addition, the Simple Network Management Protocol (SNMP) is closely examined since it is commonly used as the basis of IP (Internet Protocol) network management. Also some primary applications enabled by the NGN technology are introduced. The empirical part of this study contains a short overview of the implemented network performance management system and its properties. The most crucial monitored MIB modules, SNMP parameters and implemented performance measurements are described. The trap topology and the role of the traps for management of the NGN platform are considered and finally, the conclusion based on the several disquisitions is made supported with suggestions for future improvements.

Ecological and physiological variance in T-cell mediated immune response in Cory's shearwaters

T-cell mediated immune response (CMI) hasbeen widely studied in relation to individual andfitness components in birds. However, few studieshave simultaneously examined individual and socialfactors and habitat-mediated variance in theimmunity of chicks and adults from the samepopulation and in the same breeding season. Weinvestigated ecological and physiological variancein CMI of male and female nestlings and adults in abreeding population of Cory's Shearwaters(Calonectrisdiomedea) in theMediterranean Sea. Explanatory variables includedindividual traits (body condition, carbon andnitrogen stable isotope ratios, plasma totalproteins, triglycerides, uric acid, osmolarity,β-hydroxy-butyrate, erythrocyte meancorpuscular diameter, hematocrit, andhemoglobin) and burrow traits(temperature, isolation, and physicalstructure). During incubation, immune responseof adult males was significantly greater than thatof females. Nestlings exhibited a lower immuneresponse than adults. Ecological and physiologicalfactors affecting immune response differed betweenadults and nestlings. General linear models showedthat immune response in adult males was positivelyassociated with burrow isolation, suggesting thatmales breeding at higher densities suffer immunesystem suppression. In contrast, immune response inchicks was positively associated with bodycondition and plasma triglyceride levels.Therefore, adult immune response appears to beassociated with social stress, whereas a trade-offbetween immune function and fasting capability mayexist for nestlings. Our results, and those fromprevious studies, provide support for anasymmetrical influence of ecological andphysiological factors on the health of differentage and sex groups within a population, and for theimportance of simultaneously considering individualand population characteristics in intraspecificstudies of immune response.

Ecological and physiological variance in T-cell mediated immune response in Cory's shearwaters

T-cell mediated immune response (CMI) hasbeen widely studied in relation to individual andfitness components in birds. However, few studieshave simultaneously examined individual and socialfactors and habitat-mediated variance in theimmunity of chicks and adults from the samepopulation and in the same breeding season. Weinvestigated ecological and physiological variancein CMI of male and female nestlings and adults in abreeding population of Cory's Shearwaters(Calonectrisdiomedea) in theMediterranean Sea. Explanatory variables includedindividual traits (body condition, carbon andnitrogen stable isotope ratios, plasma totalproteins, triglycerides, uric acid, osmolarity,β-hydroxy-butyrate, erythrocyte meancorpuscular diameter, hematocrit, andhemoglobin) and burrow traits(temperature, isolation, and physicalstructure). During incubation, immune responseof adult males was significantly greater than thatof females. Nestlings exhibited a lower immuneresponse than adults. Ecological and physiologicalfactors affecting immune response differed betweenadults and nestlings. General linear models showedthat immune response in adult males was positivelyassociated with burrow isolation, suggesting thatmales breeding at higher densities suffer immunesystem suppression. In contrast, immune response inchicks was positively associated with bodycondition and plasma triglyceride levels.Therefore, adult immune response appears to beassociated with social stress, whereas a trade-offbetween immune function and fasting capability mayexist for nestlings. Our results, and those fromprevious studies, provide support for anasymmetrical influence of ecological andphysiological factors on the health of differentage and sex groups within a population, and for theimportance of simultaneously considering individualand population characteristics in intraspecificstudies of immune response.

Decay of linkage disequilibrium within genes across HGDP-CEPH human samples: most population isolates do not show increased LD

Background It is well known that the pattern of linkage disequilibrium varies between human populations, with remarkable geographical stratification. Indirect association studies routinely exploit linkage disequilibrium around genes, particularly in isolated populations where it is assumed to be higher. Here, we explore both the amount and the decay of linkage disequilibrium with physical distance along 211 gene regions, most of them related to complex diseases, across 39 HGDP-CEPH population samples, focusing particularly on the populations defined as isolates. Within each gene region and population we use r2 between all possible single nucleotide polymorphism (SNP) pairs as a measure of linkage disequilibrium and focus on the proportion of SNP pairs with r2 greater than 0.8. Results Although the average r2 was found to be significantly different both between and within continental regions, a much higher proportion of r2 variance could be attributed to differences between continental regions (2.8% vs. 0.5%, respectively). Similarly, while the proportion of SNP pairs with r2 > 0.8 was significantly different across continents for all distance classes, it was generally much more homogenous within continents, except in the case of Africa and the Americas. The only isolated populations with consistently higher LD in all distance classes with respect to their continent are the Kalash (Central South Asia) and the Surui (America). Moreover, isolated populations showed only slightly higher proportions of SNP pairs with r2 > 0.8 per gene region than non-isolated populations in the same continent. Thus, the number of SNPs in isolated populations that need to be genotyped may be only slightly less than in non-isolates. Conclusion The 'isolated population' label by itself does not guarantee a greater genotyping efficiency in association studies, and properties other than increased linkage disequilibrium may make these populations interesting in genetic epidemiology.

INFLUENTIAL FACTORS IN SURFACE WATER QUALITY IN CATCHMENTS WITHIN THE PAMPA BIOME WITH DIFFERENT LAND USE1

The aim of this study was to identify, by multivariate statistical technique, the physic, chemical and biological variables that best characterize the quality of surface waters in two small rural catchments with different land uses (eucalyptus silviculture (SC) vs. pasture and extensive livestock (LC)) located in Rosário do Sul, RS - Brazil. Monitoring was conducted during the months of August 2011 to August 2012 and the following parameters were analyzed: Ca2+, Mg2+, K+, SO42-, Cl-, pH, electrical conductivity, turbidity, alkalinity, suspended and dissolved solids, biochemical oxygen demand , total coliforms, Escherichia coli and temperature, flow and rainfall. Through the use of FA/PCA, it was found that the model best fit to express water quality of in LC that was composed of five factors which account for 83.5% of the total variance, while for SC, four factors accounted for 85.12% of the variance. In LC, the five main factors were, respectively, soluble salts, diffuse pollution, solid, and both anthropogenic and organic factors. In SC, the four factors were namely: soluble salts, mineral, nutritional and diffuse pollution factors. The results of this study showed that by replacing the traditional soil usage (pasture and livestock) with planted forest, diffuse pollution was attenuated but, however, it did not result in major changes in the physical-chemical and biological characteristics of the water. Another point to note is that factorial analysis did not result in a large reduction in the number of variables, once the best model fit occurred with the addition of 15 of 18 analyzed variables (LC) and 17 of 18 analyzed variables (SC).