Les interférons (IFN) de type I et type III sont des cytokines induites par des pathogènes. L'IFN de type I (IFN α/β)se fixe à un récepteur constitué des chaînes IFNAR1 et IFNAR2. L'IFN de type III (3 λs) se fixe à un récepteur constitué des chaines IFNLR1 et IL-10R2. La liaison de ces IFNs à leur récepteur active la voie Jak/Stat, induit les mêmes gènes et des réponses cellulaires communes essentielles à la protection antivirale. L'IFN de type I joue un rôle pléiotropique et de ce fait la réponse cellulaire aux IFNs doit être contrôlée dans le temps et dans l'espace. Certains régulateurs négatifs tels que les SOCS ou les ubiquitine ligases ciblant la sous-unité IFNAR1 vont agir rapidement après la stimulation, alors que d'autres agissent à des temps plus tardifs, tels qu'USP18. USP18 est une cystéine protéase induite par l'IFN, elle clive ISG15, une molécule semblable à l'ubiquitine, à partir de protéines ISGylées. J'ai étudié comment une stimulation prolongée avec de l'IFN de type I ou III interfère avec la capacité de ces cellules à répondre à une re-stimulation par les IFN α, tout en maintenant leur sensibilité à l'IFN β et λ . Ce phénomène de désensibilisation différentielle n'est pas dû à une diminution des récepteurs à la surface des cellules mais à l'induction de la forme catalytiquement active d'USP18. Lors de traitements prolongés à l'IFN, l'accumulation d'USP18, dont l'expression est régulée par ISG15, inhibe progressivement la signalisation induite par l'IFN α. En conclusion, ces études montrent qu'USP18 fait partie intégrante des signaux transmis lors d'une stimulation par les IFN de type I et III et définit le seuil d'activité des différents sous-types α/β.

L'importance des activités protéolytiques associées à la membrane plasmique dans divers processus biologiques fondamentaux est de mieux en mieux définie. Les protéases de la famille ADAM (A Disintegrin and Metalloprotease), et ADAM10 en particulier, ont suscité un intérêt tout particulier du fait de l'importance de leurs substrats (récepteur de l'EGF, TNFα, Notch, APP...). Néanmoins, peu d'études se sont intéressées aux mécanismes régulant le trafic d'ADAM10.Les tétraspanines sont une super-famille de protéines de surface impliquées dans de nombreux processus biologiques fondamentaux parmi lesquels la migration, les interactions intercellulaires, la réponse immunitaire, la fusion des gamètes... L'une des caractéristiques majeure des tétraspanines est leur capacité à organiser un réseau d'interactions moléculaires appelé le " tetraspanin web ". De précédentes études menées dans le laboratoire ont montré qu'ADAM10 est associé au " tetraspanin web ". Néanmoins, la tétraspanine en interaction directe avec ADAM10 permettant son association au réseau n'est pas encore connue. Dans cette étude nous nous sommes intéressés à la régulation d'ADAM10 par les tétraspanines. Nous avons ainsi pu identifier une sous-famille de tétraspanines à 8 cystéines, les TspanC8 (Tspan5, Tspan10, Tspan14, Tspan15, Tspan17 et Tspan33), comme étant capables d'interagir directement avec ADAM10 et de réguler sa sortie du réticulum endoplasmique. Nous avons montré que Tspan5, Tspan14, Tspan15 et Tspan33 sont capables de réguler l'expression de surface d'ADAM10 et que Tspan10 et Tspan17 entrainent l'accumulation d'ADAM10 dans un compartiment endosomal tardif. Les TspanC8 pourraient également contribuer à la régulation de la spécificité de substrat d'ADAM10 puisque nous avons montré que l'expression des TspanC8 humaines Tspan5 et Tspan14 augmente l'activation de la voie Notch alors que Tspan15 n'a pas d'effet. Par ailleurs, les TspanC8 de Drosophile sont capables d'interagir directement avec Kuzbanian (l'orthologue d'ADAM10), permettent son accumulation à la surface cellulaire et régulent l'activation de la voie Notch dans différents contextes développementaux. Nous proposons que les TspanC8 soient une nouvelle famille de protéines ayant une fonction très conservée dans la régulation de l'activité et du trafic d'ADAM10, capables de réguler l'activation de la voie Notch.

Statement of problem: We have shown that the C677T 5,10 methylenetetrahydrofolate reductase (MTHFR) variant is associated with elevated maternal serum α-fetoprotein (MSAFP), the most common screening test for neural tube defects (NTD). Therefore, past contradictory studies of NTDs and C677T MTHFR may have been biased because of changes in case populations after prenatal diagnosis and termination of pregnancy. Further, an unexplained elevation of MSAFP is known to increase the risk for later pregnancy complications. Is the C677T MTHFR variant a predisposing genetic variant for both NTDs and later complications of pregnancy? Methods: A retrospective study of women with pregnancies resulting in NTD outcome and women with unexplained elevations of MSAFP was undertaken. Women and their partners were genotyped for the C677T MTHFR allele. Couples with a pregnancy resulting in a NTD outcome were compared to couples whose pregnancy outcome did not involve. Couples with unexplained elevations of MSAFP who did and did not have later complications of pregnancy were also compared. Allele frequencies for all groups were then compared against the previously established Manitoba population allele frequency (based on 977 consecutive newborn metabolic screening bloodspots). A review of all studies of NTDs and association with the C677T MTHFR variant was undertaken to determine if the association between the variant and MSAFP is a source of bias. NTD incidence was examined before and after folic acid food fortification introduced in Canada in 1999. Results: There is an increase in the allele frequency of the C677T MTHFR variant in parents with an unexplained elevated MSAFP followed by later complications of pregnancy. The C677T MTHFR variant is also a contributing genetic factor to NTDs worldwide. The incidence of NTDs in Manitoba has decreased by 37% since food fortification with folic acid was introduced. Conclusions: The C677T MTHFR variant is a contributing genetic factor to both later complications of pregnancy after an unexplained elevation of MSAFP and to NTDs. This variant is folate sensitive and folic acid fortification has reduced the incidence of NTDs. / February 2005

No description available.

Various α,β-unsaturated carbonyl compounds and derivatives were explored in order to expand the range of substrates for the 1,4-addition of alkenylboronates using 3,3′-disubstituted binaphthols. Enones 2.60 were examined and found to be suitable for conjugate addition under our proposed reaction conditions. The asymmetric 1,4-additions of alkenylboronates to enones 2.60 using catalytic amounts of 3,3′-disubstituted binaphthols was shown to occur with moderate to good yields and high enantioselectivities. The chiral products could serve as enantioenriched substrates for further transformation such as asymmetric reduction, which was performed with good yield and selectivity. The absolute configuration for the alkenylation of enones was also confirmed to be the (R) enantiomer using (S)-3,3′-disubstituted binaphthols via X-ray crystallographic analysis. Investigations into selective Baeyer-Villiger oxidation of 1,4-addition products of enones was also examined. Although the desire ester products were not obtained, intriguing informative findings were still obtained from the investigation.

Various α,β-unsaturated carbonyl compounds and derivatives were explored in order to expand the range of substrates for the 1,4-addition of alkenylboronates using 3,3′-disubstituted binaphthols. Enones 2.60 were examined and found to be suitable for conjugate addition under our proposed reaction conditions. The asymmetric 1,4-additions of alkenylboronates to enones 2.60 using catalytic amounts of 3,3′-disubstituted binaphthols was shown to occur with moderate to good yields and high enantioselectivities. The chiral products could serve as enantioenriched substrates for further transformation such as asymmetric reduction, which was performed with good yield and selectivity. The absolute configuration for the alkenylation of enones was also confirmed to be the (R) enantiomer using (S)-3,3′-disubstituted binaphthols via X-ray crystallographic analysis. Investigations into selective Baeyer-Villiger oxidation of 1,4-addition products of enones was also examined. Although the desire ester products were not obtained, intriguing informative findings were still obtained from the investigation.

A new analytical technique to measure the chemical composition distribution (CCD) of polyolefins, crystallization elution fractionation (CEF), was introduced in 2006 during the First International Conference on Polyolefin Characterization. CEF is a faster and higher resolution alternative to the previous polyolefin CCD analytical techniques such as temperature rising elution fractionation (TREF) and crystallization elution fractionation (CRYSTAF) (Monrabal et al., 2007). Crystallization elution fractionation is a liquid chromatography technique used to determine the CCD of polyolefins by combining a new separation procedure, dynamic crystallization, and TREF. In a typical CEF experiment, a polymer solution is loaded in the CEF column at high temperature, the polymer is allowed to crystallize by lowering the solution temperature, and then the precipitated polymer is eluted by a solvent flowing through the column as the temperature is raised. CEF needs to be calibrated to provide quantitative CCD results. A CEF calibration curve consists of a mathematical relationship between elution temperature determined by CEF and comonomer fraction in the copolymer that could be estimated by Fourier transform infrared spectroscopy (FTIR) and carbon-13 nuclear magnetic resonance (13C NMR). Different comonomer types in ethylene/α-olefin copolymers will have distinct calibration curves. The main objective of this thesis is to obtain CEF calibration curves for several different ethylene/α-olefin copolymers and to investigate which factors influence these calibration curves. A series of homogeneous ethylene/α-olefin copolymers (1-hexene, 1-octene and 1-dodecene) with different comonomer fractions were synthesized under controlled conditions to create CEF calibration standards. Their average chemical compositions were determined by 13C NMR and FTIR and then used to establish CEF calibration curves relating elution temperature and comonomer molar fraction in the copolymer.

Seasonally high nighttime temperatures (HNT) along the United States Gulf Coast and in regions of similar climate, during the critical stages of development, could reduce rice yield and quality. To study the effects of HNT on plant physiology, a method for applying a controlled heating treatment to plant canopies was developed using overhead infrared heaters, which are relatively inexpensive and are accurate, precise and reliable in rapidly controlling the temperature. The apparatus successfully maintained air temperatures within the set points plus/minus 0.5 degrees C, and was used for all the experiments. Several experiments were conducted to determine the response of various physiological parameters during and following exposure of rice plants to HNT (32 degrees C) or ambient nighttime temperature (ANT) (27 degrees C) starting from 2000 h until 0600 h, and with or without plant growth regulator treatments. The plant growth regulator treatments included alpha-tocopherol (vitamin E), glycine betaine (GB), and salicylic acid (SA), which play different roles in inducing thermo-tolerance in plants. High nighttime temperature had no effect on plant height, number of tillers and panicles, or rice net leaf photosynthetic rates. However, HNT increased leaf respiration (dark respiration in the night) (21%) and decreased membrane thermo-stability (60%), pollen germination (20%), spikelet fertility (18% as a % of total spikelets), grain length (2%), and grain width (2%). The HNT also hastened plant development. The combinations of these effects decreased rice yield by 90%. Moreover, under HNT, there were decreases in leaf chlorophyll concentration (7%) and nitrogen concentration (18%). Application of GB and SA increased total antioxidant capacity of the rice plants by 17%, thereby decreasing the leaf respiration rates, increasing membrane thermo-stability, pollen germination, and spikelet fertility, thus increasing the yield. High nighttime temperature decreased leaf starch concentration (14%), grain total nonstructural carbohydrate (TNC) concentration (9%), and grain extractable invertase activity (20%). Vitamin E- or GB-treated plants had greater grain soluble-sugar concentrations, whereas SA-treated plants had greater leaf soluble-sugar concentrations and lower grain TNC concentrations. Invertase activity was shown to be not rate limiting or required for sucrose degradation for starch synthesis in grain of 'Cocodrie' rice under short-term high nighttime temperatures exposures during grain filling. In conclusion, HNT decreased rice yield by increasing plant respiration, rate of crop development, and decreasing membrane thermo-stability, pollen germination, spikelet fertility and grain dimensions. Exogenous application of GB and SA increased yields under HNT, possibly acting through increased antioxidant levels, which might have protected the membranes and enzymes against heat-induced ROS-mediated degradation.

<p>Plasmacytoid dendritic cells (PDCs) are one cellular target of HIV-1 and respond to the virus by producing type I interferons and chemokines. PDCs exposed to HIV-1 strongly upregulate the expression of maturation markers such as CD83, CD80, CD86 and CCR7, which will turn them into professional antigen presenting cells with the ability to stimulate naïve CD4+T cells. When HIV-1 binds to the CD4 receptor and a co-receptor (CCR5 or CXCR4) on PDCs, the cell takes up the virus by endocytosis. In response to this, PDCs will become activated and express maturation markers on their surface that make them able to stimulate T cells to trigger an immune response. In this thesis, studies have been performed with different forms of HIV-1, i.e. opsonized virions covered in complement and antibodies since these forms are supposed to be more similar to how HIV appears in the body. According to our results there is no significant difference in PDC maturation between the free and opsonized HIV-1.</p>

The p53 protein promotes tumor eradication upon activation, making it an attractive target in cancer therapies. A reported 50% of all human cancers display aberrant activation of the MDM2 oncoprotein, which directly promotes tumorgenesis by inactivating the transcriptional activity of wild type p53, and is commonly associated with drug, chemo, and radio therapy resistance. Previously reported crystallographic analysis of the p53/MDM2 complex infers that the p53 protein forms a 2.5 turn amphipathic alpha helix whose hydrophobic face interacts within a deep hydrophobic cleft in the NH2-terminal domain of the globular MDM2. This suggests that the synthesis of small molecular antagonists of p53/MDM2 binding interactions, capable of reactivating wild type p53 function, show a promising therapeutic strategy in pharmaceutical discovery. The use of alpha helix mimics for the disruption of p53/MDM2 binding interactions has been amply documented in the literature; however, these compounds contain hydrophobic scaffolds that limit their usefulness as potential drug candidates. Presented is the design, synthesis, and biological evaluation of novel non-peptidic, drug-like, small molecule inhibitors to target p53/MDM2 binding interactions. The mimetics are designed to bind to the NH2-terminal domain of MDM2 protein leaving p53 unbound and capable of activation. The inhibitor design is based on an alpha helix mimetic scaffold derived from functionalized piperazines, diketopiperazines, and/or pyrimides. The mimetics are designed to have a comparably higher degree of solubility and notably facile synthesis yet still maintain the desired spacial arrangements of hydrophobic side chains in the ith, ith+4, and ith+7 positions of a natural alpha helix. The small molecules are designed to act as antagonists of protein/protein interactions, tumor inhibitors, and potent p53 activators.