Dissertations / Theses on the topic 'Or 3-1762 or 3'

In this thesis I set out to characterise the interaction of 14-3-3 with lipid raft-like membrane domains. 14-3-3 associated with DRMs isolated from rate brain extract in a 14-3-3 isoform specific manner. Inhibition of 14-3-3 target protein interactions suggested that this association was reliant on binding of 14-3-3 to a membrane protein. The interaction of 14-3-3 with DRMs was further investigated in N2a and PC12 cells and also be detergent-free method. There are, however, some concerns over whether DRMs represent physiological membrane domains. Therefore I also investigated quantitatively the colocalisation of 14-3-3 with membrane domains in intact cells by confocal microscopy, using the lipid raft marker cholera toxin B subunit (CTXB). For comparison, other DRM-resident proteins, Thy-1, syntaxin-1a and SNAP-25, were also examined. SNAP-25 and Thy-1 showed a high degree of coincidence with CTXB in PC12 cells, 14-3-3 also colocalised with CTXB but to a much lower extent. As lipid rafts have been implicated in the control of regulated exocytosis the high coincidence of SNAP-25 and lack of coincidence of syntaxin-1 with CTXB is of interest. Cholesterol depletion, which affects the integrity of lipid raft-like domains, revealed some discrepancies between the association of 14-3-3., Thy-1, SNAP-25 and syntaxin-1a with DRMs and CTXB domains imaged in intact PC12 and N2a cells. CTXB clusters were partially disrupted in N2a cells and the coincidence of SNAP-25, but not 14-3-3, with CTXB was reduced. These findings indicate a role for lipid raft-like domains in controlling the spatial distribution of SNAP-25 on the plasma membrane. However, the lack of cholesterol dependent 14-3-3 localisation indicates that other membrane compartmentalisation mechanism may affect 14-3-3 distribution.

A P-3 Scheduling Support System (P-3 S3) is a Management Information System (MIS) that was designed using structured techniques. Structured analysis was used to determine the functionality and data requirements. Computer Assisted Systems Engineering (CASE) tools were used to document the analysis and design. The system was designed to be implemented in dBase III Plus, a data base management tool developed by Ashton Tate. P-3 S3 is designed to run on a microcomputer with the MS-DOS operating system. It provides real-time access to historical data and provides suggested personnel assignments to the user. The design provides for faster flight schedule generation and prevention of conflicting schedule events. Keywords: software development; patrol squadrons; theses

A series of conjugated [3+3] Schiff-base macrocycles containing both a central crown ether-like pocket and three tetradentate N[sub 2]O[sub 2] binding sites were prepared and investigated. The formation mechanism was investigated through the synthesis and study of macrocycle fragments. Further understanding of the macrocycle conformations and dynamics was obtained through computational studies. A monoreduced macrocycle where one of the six imines has been reduced was obtained as a by-product of macrocycle formation. Reactivity studies and deuterium labeling investigations revealed that the selective reducing agent is likely a benzimidazoline. This intermediate is generated in situ during the formation of the nonreduced macrocycle and with macrocycle reduction is converted to a stable benzimidazole unit. Upon addition of small cations, the conjugated Schiff-base macrocycles assemble into tubular structures. Spectroscopic and mass spectrometric studies have shown that the cations bind to the crown ether-like centre of the macrocycle and induce aggregation to form structures composed of alternating cations and macrocycles. With the addition of seven equivalents of Zn[sup 2+] or Cd[sup 2+] to these fully conjugated macrocycles surprising heptametallic complexes were obtained. Here, the trimetallated macrocycle is first formed (with metal ions bound to the three N[sub 2]O[sub 2] pockets) and then this templates the formation of a [M[sub 4]O][sub 6+] cluster that caps the cone-shaped macrocycle. NMR studies indicated that these zinc complexes dimerize under certain solvent conditions forming capsule-like structures resembling cavitands used in host-guest chemistry. Variations of these [3+3] Schiff-base macrocycles were prepared by modifying the substituents of the diformyl diol unit. In this way naphthalene-based macrocycles were prepared. Studies on a series of related model compounds revealed that the ketoenamine isomer is stabilized in these macrocycles rather than the enolimine isomer as observed in the analogous phenyl-based macrocycles.
Science, Faculty of
Chemistry, Department of
Graduate

Ce travail presente les proprietes spectrales d'orthoborates de formulation (lnbo#3 : ln = la, in, y) dopes par eu#3#+ ou tb#3#+, en correlation avec leurs structures cristallographiques. Deux procedes de synthese ont ete utilises pour preparer les composes: une voie solide classique et une voie sol-gel minerale dont les parametres tels que le ph, le rapport b/ln et les temperatures d'elaboration ont ete precises. La mise au point d'un protocole de synthese par voie liquide a permis de redefinir les domaines d'existence des trois formes polymorphiques aragonite, vaterite et calcite en fonction des rayons ioniques des cations trivalents et de mettre en evidence la possibilite tout a fait originale de preparer a basse temperature des representants de certaines de ces varietes. Tous les composes ont ete controles par analyse chimique, drx, ir et rmn du #1#1b ce qui a permis de caracteriser les formulations et les enchainements cristallographiques. Un laser accordable de largeur spectrale 0,1 cm#-#1 a ete utilise pour induire le fluorescence visible des composes. L'analyse des spectres optiques de l'ion eu#3#+ utilise comme sonde structurale a permis d'identifier les centres emetteurs de lumiere. Leurs environnements sont compatibles avec les descriptions structurales faites dans le cas de labo#3 et inbo#3. Par contre un desaccord a ete observe pour ybo#3. La redetermination structurale de ce compose menee sur monocristal dans le groupe d'espace p6#3/m correle les resultats optiques. Les mesures comparatives des intensites de photoluminescence de eu#3#+ et tb#3#+ realisees entre les echantillons prepares par voie solide et ceux prepares par technique sol-gel montrent des gains en efficacite tres appreciables, de l'ordre de 30 a 40% pour les phases preparees en solution. Une meilleure homogeneite des phases preparees par le procede sol-gel semble etre a la base de telles performances. D'autre part la haute absorption dans l'uv du vide des ions eu#3#+ et tb#3#+ dans ces composes, conferent a ces materiaux de reelles potentialites pour une application dans les ecrans a excitation plasma

Magister Scientiae - MSc (Biotechnology)
Lately, there has been serious contamination of soils and ground water caused by mineral mining and environmental pollution leading to heavy metal accumulation within soils and over time rendering the soil infertile. The contaminated soil affects crop; yield, germination percentage and leads to seed dormancy. Additionally, as plants are sessile organisms, exposure to environmental stress involves different defense cycles and signalling molecules. Oxidative stress, increases the production of reactive oxygen species (ROS) at greater rates than that of the metabolism. Moreover, oxidative damage leads to the loss in cellular function and eventually death. Nonetheless, plants have adapted ROS-scavenging systems driven by reduction-oxidation reactions as defense mechanisms. In this way, cellular homeostasis is an essential defending process and finding means to optimize these reactions would benefit in the development of plants. Hence, environmentally friendly solutions are being reported such as using glucosinolates, a secondary plant metabolite found in relatively high concentrations within crucifers such as Brassica napus L.
2020-04-30

Etude de la fluorescence. Une serie de composes isotopes de formule generale linaln::(2)f::(8) a ete synthetisee avec des lanthanides de faible rayon ionique (ln = ho -> lu). Un laser a impulsion accordable de largeur spectrale 0,5cm**(-1) a ete utilise pour exciter selectivement dans l'uv ou dans le visible un des niveaux absorbants au-dessus de **(5)d::(4) de l'ion tb**(3+) ou de **(5)d::(0) de l'ion eu**(3+). Mise en evidence d'un transfert d'energie tb**(3+) -> eu**(3+)

This study was designed to determine whether chis-quared based tests for independence give reliable estimates (as compared to the exact values provided by Fisher's exact probabilities test) of the probability of a relationship between the variables in 3 X 3, 3 X 4 , and 3 X 5 contingency tables when the sample size is 10, 20, or 30. In addition to the classical (uncorrected) chi-squared test, four methods for continuity correction were compared to Fisher's exact probabilities test. The four methods were Yates' correction, two corrections attributed to Cochran, and Mantel's correction. The study was modeled after a similar comparison conducted on 2 X 2 contingency tables and published by Michael Haber.

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Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)
Cintiladores são materiais capazes de absorver radiações de alta energia e convertê-las eficientemente em radiações ultravioleta ou visível. Avanços significativos na eletrônica de detecção dessas radiações motivaram as pesquisas visando à busca por novos materiais cintiladores de alta eficiência e à maior compreensão do mecanismo de cintilação. Nesse contexto, os compostos de matriz GdAlO3 e Gd1-xLuxAlO3 dopados com Eu3+, Tb3+ ou Pr3+ são materiais cintiladores promissores, pois apresentam alta densidade, elevados valores de coeficiente de atenuação de massa e propriedades espectroscópicas relevantes como emissão intensa, tempo de vida de mili ou microssegundos e transferência de energia a partir dos íons Gd3+ para os ativadores. Neste trabalho, foi investigada a preparação dos cintiladores GdAlO3 e Gd1-xLuxAlO3 nominalmente puros e dopados com Eu3+, Tb3+ e Pr3+ através do método dos precursores poliméricos (método Pechini) e suas propriedades estruturais e espectroscópicas (fotoluminescência e luminescência com excitação por raios X) foram investigadas. O perfil espectral de emissão das amostras de GdAlO3 dopadas com Eu3+ evidencia a ocupação de dois sítios distintos pelos íons dopantes quando em concentrações superiores a 1,0%-mol, provavelmente devido à distribuição não homogênea dos íons na matriz. A transferência de energia Gd3+→Ln3+ foi evidenciada pela espectroscopia de fotoluminescência e é um processo importante no mecanismo de cintilação desses materiais, como evidenciado pela comparação das medidas espectroscópicas de fotoluminescência e luminescência com excitação por raios X das amostras de GdAlO3:Pr3+. Com a incorporação de Lu3+ na matriz de GdAlO3, a intensidade de emissão aumenta até a concentração de 10%-mol de Lu3+, decrescendo...
Scintillators materials are compounds that can absorb ionizing radiations and efficiently convert them into UV-VIS radiations. Despite of impressive advances in the technology of direct detection, the search for new scintillators materials with high efficiency and for the precise understanding of the scintillation mechanism is still the focus of research in this area. In this context, the compounds GdAlO3 and Gd1-xLuxAlO3 doped with Eu3+, Tb3+ and Pr3+ are promising scintillators materials, since they have high density, high values of mass attenuation coefficient and relevant spectroscopic properties, such as bright emission, lifetime of milli or microseconds and the energy transfer from Gd3+ ions to the activators. This work describes the preparation of GdAlO3 and Gd1-xLuxAlO3 nominally pure and doped with Eu3+, Tb3+ e Pr3+ by the polymeric precursor method (Pechini method) and the study of their structural and spectroscopic (photoluminescence and X-rays excited optical luminescence) properties. Emission spectral profile of Eu3+-doped GdAlO3 indicates the occupation of two different sites by the dopant ions when the concentration is higher than 1.0 mol-%, probably due to inhomogeneous distribution of ions in the host. The energy transfer Gd3+→Ln3+ was evidenced by photoluminescence spectroscopy and is an important process in the scintillation mechanism of these materials, as showed by comparison of spectroscopic measurements of photoluminescence and X-rays excited optical luminescence of GdAlO3:Pr3+ sample. The emission intensity increases by incorporating Lu3+ in the GdAlO3 host until the Lu3+ concentration of 10 mol-%, decreasing at higher concentrations. This behavior is probably assigned to the balance between increasing the absorption of X-rays and the reduction of the transfer process from Gd3+ ions with increasing concentration of... (Complete abstract click electronic access below)

Orientador: Marian Rosaly Davolos
Banca: Carlos Frederico de Oliveira Graeff
Banca: Luiz Antonio de Oliveira Nunes
Resumo: Cintiladores são materiais capazes de absorver radiações de alta energia e convertê-las eficientemente em radiações ultravioleta ou visível. Avanços significativos na eletrônica de detecção dessas radiações motivaram as pesquisas visando à busca por novos materiais cintiladores de alta eficiência e à maior compreensão do mecanismo de cintilação. Nesse contexto, os compostos de matriz GdAlO3 e Gd1-xLuxAlO3 dopados com Eu3+, Tb3+ ou Pr3+ são materiais cintiladores promissores, pois apresentam alta densidade, elevados valores de coeficiente de atenuação de massa e propriedades espectroscópicas relevantes como emissão intensa, tempo de vida de mili ou microssegundos e transferência de energia a partir dos íons Gd3+ para os ativadores. Neste trabalho, foi investigada a preparação dos cintiladores GdAlO3 e Gd1-xLuxAlO3 nominalmente puros e dopados com Eu3+, Tb3+ e Pr3+ através do método dos precursores poliméricos (método Pechini) e suas propriedades estruturais e espectroscópicas (fotoluminescência e luminescência com excitação por raios X) foram investigadas. O perfil espectral de emissão das amostras de GdAlO3 dopadas com Eu3+ evidencia a ocupação de dois sítios distintos pelos íons dopantes quando em concentrações superiores a 1,0%-mol, provavelmente devido à distribuição não homogênea dos íons na matriz. A transferência de energia Gd3+→Ln3+ foi evidenciada pela espectroscopia de fotoluminescência e é um processo importante no mecanismo de cintilação desses materiais, como evidenciado pela comparação das medidas espectroscópicas de fotoluminescência e luminescência com excitação por raios X das amostras de GdAlO3:Pr3+. Com a incorporação de Lu3+ na matriz de GdAlO3, a intensidade de emissão aumenta até a concentração de 10%-mol de Lu3+, decrescendo... (Resumo completo, clicar acesso eletrônico abaixo)
Abstract: Scintillators materials are compounds that can absorb ionizing radiations and efficiently convert them into UV-VIS radiations. Despite of impressive advances in the technology of direct detection, the search for new scintillators materials with high efficiency and for the precise understanding of the scintillation mechanism is still the focus of research in this area. In this context, the compounds GdAlO3 and Gd1-xLuxAlO3 doped with Eu3+, Tb3+ and Pr3+ are promising scintillators materials, since they have high density, high values of mass attenuation coefficient and relevant spectroscopic properties, such as bright emission, lifetime of milli or microseconds and the energy transfer from Gd3+ ions to the activators. This work describes the preparation of GdAlO3 and Gd1-xLuxAlO3 nominally pure and doped with Eu3+, Tb3+ e Pr3+ by the polymeric precursor method (Pechini method) and the study of their structural and spectroscopic (photoluminescence and X-rays excited optical luminescence) properties. Emission spectral profile of Eu3+-doped GdAlO3 indicates the occupation of two different sites by the dopant ions when the concentration is higher than 1.0 mol-%, probably due to inhomogeneous distribution of ions in the host. The energy transfer Gd3+→Ln3+ was evidenced by photoluminescence spectroscopy and is an important process in the scintillation mechanism of these materials, as showed by comparison of spectroscopic measurements of photoluminescence and X-rays excited optical luminescence of GdAlO3:Pr3+ sample. The emission intensity increases by incorporating Lu3+ in the GdAlO3 host until the Lu3+ concentration of 10 mol-%, decreasing at higher concentrations. This behavior is probably assigned to the balance between increasing the absorption of X-rays and the reduction of the transfer process from Gd3+ ions with increasing concentration of... (Complete abstract click electronic access below)
Mestre

En el desarrollo de esta tesis hemos analizado distintos aspectos preliminares relacionados con la utilización de óxidos complejos, tales como las perovskitas de manganeso, en la implementación de dispositivos con potenciales aplicaciones en magnetoelectrónica. En concreto, hemos abordado a fondo distintos aspectos relacionados con la fabricación de uniones túnel magnéticas basadas en el sistema La2/3Ca1/3MnO3 (LCMO)/SrTiO3 (STO).
Nuestro trabajo ha puesto de manifiesto que la técnica de pulverización catódica de RF es adecuada para el crecimiento de capas epitaxiales de LCMO de alta calidad sobre sustratos monocristalinos diversos (STO, LaAlO3 (LAO)). Las heteroestructuras LCMO/STO y LCMO/LAO han sido objeto de un detallado análisis poniendo de manifiesto que el desacoplo de parámetros de red juega un papel muy importante tanto sobre el tipo de crecimiento de las heteroestructuras como sobre las propiedades físicas. El sistema LCMO/STO (δ=1.2%) crece tensionado en el plano y permite crecer capas de hasta unos 100 nm sin que haya relajación parcial de la tensión estructural. En cambio, el sistema LCMO/LAO (δ=-1.8%) crece comprimido en el plano y muestra señales de relajación parcial de la tensión estructural incluso para espesores en torno a los 10 nm.
El análisis de las propiedades magnéticas y de transporte muestran que ambos sistemas LCMO/STO y LCMO/LAO adolecen de una notable disminución tanto de su temperatura de transición magnética, TC, como de su magnetización de saturación, MS, al disminuir el espesor de las capas, siendo éste más acusado en el caso del LCMO/STO. Tratamientos térmicos al aire a alta temperatura permiten una mejora sustancial de las propiedades magnéticas y de transporte en ambos sistemas. La mejora es mucho más rápida en el sistema LCMO/LAO. Así mismo, el aumento del espesor de las capas también promueve una mejora de las propiedades, que se aproximan a las del material en forma masiva conforme el grosor aumenta. Esta evolución se ha relacionado con la aparición de una capa no magnética en la superficie de las capas de LCMO que se pone de manifiesto, predominantemente para las capas de LCMO/LAO ultra delgadas. Por otro lado, hemos visto como esta capa inerte puede ser reducida hasta un orden de magnitud mediante tratamientos térmicos de las muestras a alta temperatura.
Nuestro trabajo ha puesto de manifiesto que la tensión estructural juega un papel fundamental y que las mejoras de TC y MS con los tratamientos térmicos están fundamentalmente ligadas a una relajación parcial de la tensión estructural. Otras posibilidades, tales como la incorporación de oxigeno o la variación de la microestructura de las muestras han sido descartadas en base a medidas de efecto Hall y análisis de microscopía electrónica de alta resolución.
Para el estudio de las propiedades de transporte en las interfases hemos desarrollado un procedimiento experimental basado en la utilización de un microscopio de fuerzas atómicas trabajando en modo corriente permitiendo la caracterización de barreras aislantes e interfases entre un metal noble y un óxido complejo. Este método permite la caracterización de barreras y superficies soslayando los problemas derivados del desconocimiento preciso del contacto entre la punta de AFM y la superficie de la capa. Se realizaron medidas mediante la definición de nanocontactos de diferente geometría y tamaño realizados por distintas técnicas de nanofabricación. Mediante el estudio de las curvas I(V) en nanocontactos de LCMO/metal noble (Au, Pt) constatamos la existencia de una barrera superficial aislante (túnel) para capas finas vírgenes de LCMO/LAO. Mediante el ajuste de las curvas I(V) utilizando el modelo de Simmons hemos derivado valores para la capa aislante que concuerdan muy bien con otros datos reportados, además esta capa aislante puede ser reducida drásticamente mediante tratamientos térmicos en atmósfera ambiente.
In this Thesis we have analyzed different preliminary aspects related to the use of complex oxides, such as manganes perovskites, for implementation of functional devices with potential application on magnetoelectronics. To be precise, we have faced in deep the different aspects related with the fabrication of magnetic tunnel junctions based on the La2/3Ca1/3MnO3 (LCMO)/SrTiO3 (STO) system.
Our work makes evident the suitability of RF sputtering technique for growing high quality epitaxial layers of LCMO onto several single-crystalline substrates (STO, LaAlO3 (LAO)). The heterostructures LCMO/STO and LCMO/LAO have been object of a detailed analysis showing up that structural misfit plays a significant role both on the heterostructures growth process and on the physical properties. The system LCMO/STO (δ=1.2%) is under in-plane tensile strain but, thin films up to 100 nm thick can be grown without any partial relieve of the structural strain. In contrast, the system LCMO/LAO (δ=-1.8%) grows under in-plane compressive strain and shows clear traces of partial relaxation of structural strain even for thicknesses around 10nm.
The analysis of the magnetic and transport properties indicates that both systems have an outstanding depression on both the temperature transition, TC, and the saturation magnetization, MS, when decreasing the thin film thickness, being this phenomenon more pronounced in the case of LCMO/STO. High temperature annealing processes in air promote a remarkable improvement of the magnetic and transport properties in both systems. This improvement is faster and more evident in the LCMO/LAO system.
In addition, we have verified that the increment of the film thicknesses also promotes an improvement of the physical properties, approaching that of bulk samples. This evolution has been correlated with the existence of a non magnetic layer at the topmost layers of the LCMO thin films, whose effect becomes more relevant as the film thickness decreases and being dominant for ultra-thin films in the LCMO/LAO system. It has been observed that this dead layer can be reduced up to one order of magnitude by high temperature annealing processes.
Our work has demonstrated that structural strain plays a fundamental role on the magnetic and transport properties of these systems and the improvements of TC and MS with annealing processes is strongly linked to a partial relaxation of the structural strain. Other possibilities, such as oxygen uptake or modifications of the microstructure of the samples have been ruled out in base of Hall Effect measurements and high resolution transmission electron microscopy analysis.
To study transport properties at the interfaces we have developed an experimental procedure based on the use of an atomic forces microscope (AFM) working in current sensing mode, permiting the electrical characterization of insulating barriers and the interfaces between the precious metal and the complex oxide. This method allows characterizing transport properties of barriers and interfaces avoiding the problems related to the uncertainty of the exact nature of the contact between the AFM tip an the analyzed surface. Measurements were realized by defining nanocontacts, with different geometries and sizes, using different nanofabrication techinques.

Initiation of DNA replication is a cellular process whose regulation is not well understood yet. It certainly depends on cis-acting elements, or origins of DNA replication sequences, and trans-acting factors, or initiator proteins. Here, we have characterized the Cruciform Binding Protein (CBP) as an origin binding protein and its role in initiation of DNA replication.
CBP was previously shown to contain the β, ɣ, ε, and ζ isoforms of the 14-3-3 family, which is composed of seven mammalian isoforms (β, ɣ, ε, η, σ, τ, and ζ) that can form homo- and heterodimers, and plays a variety of roles in different cellular processes. In this thesis, I showed by Western blot analysis with anti-14-3-3σ antibody, which partially interfered with the CBP-cruciform DNA complex formation, that the isolated CBP-cruciform DNA complex contained also the σ isoform. The same antibody reduced the in vitro DNA replication efficiency of HeLa cell total extracts, in an assay that used p186, a plasmid bearing the 186-bp minimal origin of ors8, as template DNA. Similarly, I found that antibodies against 14-3-3 isoforms β, ɣ, ε, and ζ also interfered with the CBP-cruciform DNA complex formation, and reduced the in vitro p186 replication efficiency of HeLa cell total extracts. The five isoforms of 14-3-3 (β, ɣ, ε, σ, and ζ) were found to associate with the monkey cell (CV-1) origins of DNA replication ors8 and ors12 in a cell cycle-dependent manner, the association being higher at the G1/S phase. Furthermore, we found that 14-3-3 yeast homologues, Bmh1p and Bmh2p, were able to bind cruciform DNA in vitro, and to associate in vivo with the autonomous replication sequence 307 (ARS307) in a cell cycle-dependent manner, again the association being higher at G1/S. Finally, I showed that recombinant 14-3-3ζ, tagged with maltose-binding protein (r14-3-3ζ-MBP), could only bind cruciform DNA after pre-incubation with a CBP-enriched HeLa cell extract (FTH), in which it heterodimerized with endogenous 14-3-3 isoforms β and ε. Addition of r14-3-3ζ-MBP to HeLa cell total extracts increased the in vitro replication of p186, suggesting that increased CBP activity could lead to multiple rounds of initiation of DNA replication.

The extension of axons to their appropriate targets during development or regeneration relies on the growth cone, a specialized structure that integrates extracellular signals to guide axon growth. Achieving the many complex connections formed by the nervous system while relying on a limited number of cues, requires the growth cone to modulate its response to signals in both space and time. The mechanisms by which the growth cone is able to spatially and temporally regulate its underlying cytoskeletal response to guidance cues are only beginning to become understood. Proteomic analysis of isolated growth cones identified the 14-3-3 adaptor proteins as major constituents of the growth cone. Given their role in regulating spatial and temporal activity of multiple cell signalling events, 14-3-3s are excellent candidates to play a similar role in the growth cone. This thesis establishes the critical role of 14-3-3 proteins in the spatial and temporal regulation of growth cone response to axon guidance cues. In chapter 2, we describe how loss of 14-3-3 function in dorsal root ganglion (DRG) neurons, converts their response to gradients of nerve growth factor (NGF) and myelin associated glycoprotein (MAG) from repellent to attractive. This switch can be blocked by antagonizing cAMP-dependent kinase (PKA), implicating 14-3-3 in the regulation of PKA activity. Consistently, we find that 14-3-3s interact with PKA and that disrupting this interaction results in the release of active catalytic subunits of PKA. In Chapter 3, we describe the role of 14-3-3 proteins and PKA in establishing a cell intrinsic, time dependent switch in the turning response of commissural interneurons to gradients of Sonic Hedgehog (SHH). The change from an attractive to a repellent response to SHH at the midline is critical in the proper formation of the axonal projections of dorsal commissural neurons in the developing spinal cord, and interfering with this switch in vivo, either by prematurely overexpressing 14-3-3s or by blocking 14-3-3 function results in aberrant axon growth. In Chapter 4, we identify a role for 14-3-3 proteins in mediating the signalling of another guidance cue used by commissural neurons, Netrin-1. Together these studies demonstrate the 14-3-3s are a multifunctional family of adaptor proteins that provide a critical layer of regulation to axon guidance signals, allowing the growth cone to respond to cues appropriately during development and provide a potentially interesting target for modulating growth cone responses after axon injury.
La projection des axones vers leurs cibles appropriées au cours du développement ou de la régénération s'appuie sur le cône de croissance, une structure spécialisée qui intègre les signaux extracellulaires pour guider la croissance axonale. La mise en place des connexions multiples et complexes formées par le système nerveux, tout en s'appuyant sur un nombre limité de signaux, requière que le cône de croissance module sa réponse à ces signaux tant dans l'espace que dans le temps. Les mécanismes par lesquels le cône de croissance est en mesure réguler la réponse spacio-temporelle de son cytosquelette à des signaux de guidage commencent seulement à être compris. L'analyse protéomique de cônes de croissance isolés a identifié les protéines adaptatrices 14-3-3 en tant que constituants principaux du cône de croissance. Compte tenu de leur rôle dans la régulation de l'activité spatiale et temporelle de multiples événements de signalisation cellulaires, les protéines 14-3-3 sont d'excellents candidats pour jouer un rôle similaire dans le cône de croissance. Cette thèse établit le rôle essentiel des protéines 14-3-3 dans la régulation spatiale et temporelle de la réponse du cône de croissance à des signaux de guidage axonal. Dans le chapitre 2, nous décrivons comment la perte de fonction des protéines 14-3-3 dans les neurones du ganglion de la racine dorsale (DRG), convertit leur réponse à des gradients de facteur de croissance nerveuse (NGF) et glycoprotéine associée à la myéline (MAG) de la répulsion à l'attraction. Ce commutateur peut être bloqué par l'inactivation de la kinase dépendante à AMPc (PKA), ce qui implique 14-3-3 dans la régulation de l'activité de PKA. Dans ce sens, nous constatons que les protéines 14-3-3 interagissent avec PKA et que perturber cette interaction résulte en la libération de sous-unités catalytiques actives de la PKA. Dans le chapitre 3, nous décrivons le rôle des protéines 14-3-3 et de PKA dans l'établissement d'un commutateur intrinsèque à la cellule, et dépendant du temps lors de l'orientation des axones des interneurones commissuraux soumis à un gradient de Sonic Hedgehog (Shh). Le changement d'une réponse attractive à une réponse répulsive à SHH au niveau de la ligne médiane est essentiel à la formation des projections axonales appropriées des neurones commissuraux dorsaux de la moelle épinière lors du développement, et interférer avec ce commutateur in vivo, soit en surexprimant prématurément les 14-3 – 3, soit en bloquant la fonction des 14-3-3, entraîne une croissance axonale aberrante. Dans le chapitre 4, nous identifions un rôle pour les protéines 14-3-3 dans la médiation d'un autre signal de guidage utilisé par les neurones commissuraux, la nétrine-1. Ensemble, ces études montrent que les protéines 14-3-3 sont une famille multifonctionnelle de protéines adaptatrices qui fournissent un niveau supplémentaire de régulation du guidage axonal, permettant au cône de croissance de répondre à des signaux de façon appropriée au cours du développement et de fournir une cible potentiellement intéressante pour la modulation de cette croissance après blessure axonale.

[2+2+1] iron and cobalt mediated cycloadditions have been applied towards the synthesis of 3-amino and 3-oxycyclopentadienones. These methods have provided an efficient means of generating [3.3.0], [4.3.0], and [5.3.0] bicyclic-cyclopentadienone ring systems. In an effort to probe the reactivity of our metal complexed dienones, we proceeded to subject them to cycloaddition reactions. With this goal in mind, we applied oxidative demetallation conditions to provide the metal free bicyclic-cyclopentadienones. Cycloaddition of the bicyclic-dienones with a number of pi donors and acceptors provided chemo and regioselective cycloadducts in an efficient fashion. Final studies included investigations of the iron cyclopentadienones in diastereoselective reactions.