Texte remanié de: Thèse--Sci. nat.--Paris VI, 1986. / Bibliogr. p. 285-297. Index. Résumé en français, anglais, allemand, japonais.

Thesis-- University of Michigan, 1973.

Activity and transcription profiles of two classes of cysteine proteases, papain- and legumain-like cysteine proteases, as well as their potential inhibitors, cysteine protease inhibitors (cystatins), were investigated in soybean nodules during nodule development and after drought inducing premature senescence. During nodule development total protease activity increased with major activity bands detected protease zymography in older nodules. Expressed cysteine proteases during nodule development were detected by tagging proteases with the cysteine protease inhibitor DCG-04 with major DCG-04 tagged bands found in both young and old nodules. Increase in protease activity was associated with a significant decrease in nitrogenase activity of nodules measured as acetylene reduction. Semi-quantitative RT-PCR for cysteine protease and cystatin transcription profiling showed a decrease in transcription during development and also after drought treatment of several papain-like cysteine proteases (Glyma04g04400, Glyma17g05670, Glyma10g35100, and Glyma04g03090). In contrast, transcription of three legumain-like cysteine proteases (Glyma17g14680, Glyma05g04230 and Glyma14g10620) increased during nodule development and also after drought treatment. Transcription of two cystatins (Glyma13g27980 and Glyma05g28250) increased during nodule development with Glyma13g27980 strongly up-regulated after drought treatment and Glyma05g28250 constitutively strongly expressed in both well-watered and drought treated nodules. Overall, the study has contributed in establishing an expression profile of cysteine proteases and cystatins in soybean nodules. This knowledge provides a basis which can be used to determine the importance of the individual components of the cysteine protease – cystatin system, during soybean nodule development and during stress-induced premature nodule senescence. / Dissertation (MSc)--University of Pretoria, 2013. / gm2014 / Plant Science / unrestricted

No description available.

Quantitative multielemental analysis using Particle Induced X-ray Emission (PIXE) using 1 and 2 MeV proton bombardment on thin targets has been achieved. The method is based on the calculation of atomic ratios from experimentally determined relative x-ray efficiency curves. Sample preparation techniques involving digestion and homogenous deposition of samples and standards with a minimum of contamination have been investigated. The accuracy of the method has been evaluated using five standard reference materials obtained from the National Bureau of Standards. The elimination of bremsstrahlung associated with the charging effect of non-conducting samples in PIXE analysis has been accomplished using thin carbon foils in the beam path. Applications of the PIXE technique to studies on deep-sea ferromanganese nodules were performed. The utility of PIXE in the analysis of noduoles and in the following of the distribution of a large number of elements through the various stages of a processing scheme were demonstrated.

Graduation date: 1987

Étude moléculaire des étapes précoces de la symbiose actinorhizienne Casuarina-Frankia : analyse fonctionnelle des gènes de la plante hôte contrôlant l'infectionPlus de 80% des plantes peuvent établir une symbiose racinaire avec des champignons de l'ordre des Glomales et former des endomycorhizes à arbuscules (AM). En revanche, seules certaines espèces appartenant à dix familles d'angiospermes réunies dans le Clade des Eurosidées I peuvent établir une symbiose racinaire fixatrice d'azote. Il s'agit d'une part, des plantes de la famille des légumineuses (Fabacées) et de Parasponia associées à Rhizobium et d'autre part, des plantes actinorhiziennes associées à l'actinomycète Frankia. Comme chez les légumineuses, la symbiose actinorhizienne aboutit à la formation de nodosités (ou « nodules »), siège de la fixation d'azote par les bactéries. Cependant, contrairement aux nodules des légumineuses, le nodule actinorhizien présente une structure et un développement s'apparentant aux racines latérales. L'étude des nodosités actinorhiziennes est donc particulièrement intéressante tant pour rechercher les spécificités de cette symbiose, que pour déterminer quelles sont les caractéristiques communes avec les légumineuses. Nous avons étudié le rôle du gène CCaMK dans le processus symbiotique et l'organogenèse nodulaire chez l'arbre actinorhizien Casuarina glauca. CCaMK code pour une protéine kinase dépendante du calcium et de la calmoduline (« calcium and calmodulin dependent protein kinase »). Dans la cascade de signalisation conduisant à la nodulation et à la mycorhization chez les légumineuses, ce gène est positionné en aval des oscillations calciques (« calcium spiking ») qui ont lieu durant les premières étapes de l'interaction symbiotique. CCaMK jouerait un rôle dans la perception et le décodage des oscillations calciques, ainsi que leur transduction aux différents composants contrôlant les endosymbioses racinaires. Nous avons suivi l'expression spatio-temporelle de la fusion transcriptionnelle PromCgCCaMK::GUS au cours de la nodulation et montré que celle-ci était corrélée à la présence de Frankia tout au long du processus symbiotique, soulignant ainsi le rôle clé de CCaMK dans l'infection. Par ailleurs, nous avons cherché à déterminer l'importance du domaine autoinhibiteur de la protéine CCaMK dans l'activation du processus d'organogenèse du nodule. Pour cela, nous avons réalisé et introduit chez C. glauca des constructions géniques de CgCCaMK permettant l'expression de formes tronquées constitutivement actives, car dépourvues du domaine autoinhibiteur/CaM. Nous avons aussi utilisé des formes tronquées du gène MtCCaMK de Medicago truncatula. L'expression de ces formes tronquées de CCaMK a révélé que la levée de l'autoinhibition induit la formation de nodules spontanés indépendamment de l'actinobactérie Frankia. Les résultats obtenus suggèrent que la protéine dérégulée est capable de réactiver la voie de signalisation, ainsi que les gènes situés en aval de CCaMK, qui sont nécessaires à l'organogenèse nodulaire.Mots clés : Casuarina glauca, Frankia, CCaMK, infection, autoinhibition, nodules spontanés / Molecular study of the early stages of actinorhizal symbiosis Casuarina-Frankia: functional analysis of the host plant genes controlling the infectionMore than 80% of plant species are able to develop arbuscular mycorrhizal (AM) symbiosis in association with glomeromycete fungi. In contrast, only some species of the Eurosid I clade, confined to four orders and ten Angiosperm families, are able to form nitrogen-fixing root nodule symbioses with soil bacteria. This concerns plants of the legume family (Fabaceae) and Parasponia associated with Rhizobium bacteria and actinorhizal plants associated with the actinomycete Frankia. Similarly to Legumes, the actinorhizal symbiosis results in the formation of nitrogen-fixing root nodules. However, unlike legume nodule, the actinorhizal nodule has a same origin and structure than a lateral root. Thus, the study of actinorhizal nodules is of particular interest not only for investigating its specific properties but also, for determining common characteristics shared with legume nodules.We have studied the role of CgCCaMK gene during the symbiotic process and nodule organogenesis in the actinorhizal tree Casuarina glauca. CCaMK encodes a calcium and calmodulin dependent protein kinase. In the signalisation cascade leading to both nodulation and mycorrhization in legumes, this gene is acting downstream the calcium oscillations (« calcium spiking ») that occur during the early steps of the symbiotic interaction. It has been suggested that these calcium oscillations are decoded and transduced by the CCaMK protein.We have monitored the spatio-temporal expression of a PromCgCCaMK::GUS fusion during actinorhizal nodulation and have shown that reporter gene expression was correlated with the presence of Frankia along the symbiotic process. This data highlights the role of CgCCaMK during Frankia infection. In addition, we have investigated the role of the CCaMK autoinhibitory/CaM domain in actinorhizal nodule organogenesis. To achieve this goal, we have obtained truncated versions of CgCCaMK lacking the autoinhibitory/CaM domain, and then expressed them into C. glauca. We have also used truncated forms of MtCCaMK from Medicago truncatula. The expression of these CCaMK constructs from C. glauca and M. truncatula was found to induce spontaneous nodulation in the absence of Frankia bacteria. These results suggest that deregulation of the calcium and calmodulin dependent protein kinase is able to reactivate the symbiotic signalling pathway and genes acting downstream CCaMK that are needed for nodule organogenesis.Key words: Casuarina glauca, Frankia, CCaMK, infection, autoinhibition, spontaneous nodules

Deep sea polymetallic nodules are concretions of manganese and iron oxides formed around a nucleus. They accrete either hydrogenetically - metals precipitate from the seawater - or diagenetically - metals precipitate from the sediment pore water. The accretion process affects both the nodules morphology and geochemistry. In this study, fourteen polymetallic nodules from four ocean regions, namely the Clarion-Clippertone Zone (Northeast Pacific Ocean), the Central Indian Basin (Central Indian Ocean), the Mascarene Basin (West Indian Ocean), and the Rio Grande Rise (Southwest Atlantic Ocean), were used to compare morphological and geochemical aspects between the different oceanic regions. Computed Tomography (CT) was applied to study the nodules internal structure. Scanning Electron Microscopy (SEM) was used to describe the micro layers within the nodules. Chemical composition of growth layers and nuclei was determined by both Micro X-ray Fluoscence (μ-XRF) and Laser Ablation Inductively Coupled Plasma Mass Spectroscopy (LA-ICP-MS). Finally, X-ray Absorption Near Edge Spectroscopy (XANES) was performed in order to determine the speciation (i.e., the oxidation state) of Mn and Fe. Polymetallic nodules from the Central Indian Basin are diagenetic and the ones from the Mascarene Basin and the Rio Grande Rise are hydrogenetic, while nodules from the Clarion-Clippertone Zone are of mixed type. However, the dominant accretion process varies across the nodules resulting in inhomogeneous layer textures and chemical composition. Strong Mn and Fe fractionation occurs in the diagenetic and mixed type nodules accompanied by fractionation of the trace elements Ni, Cu, Co and Ti. Mn and Fe are present in the nodules mainly as oxidized species Mn4+ and Fe3+, independently of the degree of fractionation. Schematic models of the nodules environment of formation are proposed, in which and the fractionation of Mn and Fe is possibly the result of the variation of the redox front depth through time. / Nódulos polimetálicos de mar profundo são concreções de óxidos de manganês e de ferro ao redor de um núcleo. Os nódulos crescem através da precipitação hidrogenética - precipitação de metais da água do mar - ou diagenética - precipitação de metais da água intersticial do sedimento. O processo de acreção reflete na morfologia e geoquímica dos nódulos. Neste trabalho, quatorze nódulos polimetálicos provenientes de quatro regiões oceânicas - Clarion-Clippertone Zone (Oceano Pacífico Nordeste), Bacia Central do Índico (Oceano Índico Central), Bacia Mascarene (Oceano Índico Oeste) e Elevação de Rio Grande (Oceano Atlântico Sudoeste) - foram usados a fim de se comparar os aspectos morfológicos e geoquímicos dos nódulos entre regiões diferentes. A estrutura interna dos nódulos foi avaliada através da Tomografia Computadorizada por Raios-X (CT). Microscopia Eletrônica de Varredura (SEM) foi usada para descrever as micro camadas. A composição química foi determinada por Micro Fluorescência de Raios-X (μ-XRF) e por ablação a laser acoplada a espectrometria de massa com plasma indutivamente acoplado (LA-ICP-MS). Por fim, a Espectroscopia de Absorção de Raios-X próximo à borda (XANES) foi realizada a fim de se determinar a especiação (i.e., o número de oxidação) do Mn e do Fe. Os nódulos polimetálicos da Bacia Central do Índico são diagenéticos e os da Bacia Mascarene e Elevação do Rio Grande são hidrogênicos, enquanto que os da Clarion-Clippertone Zone são do tipo misto. Entretanto, o processo de acreção varia ao longo dos nódulos, resultando em textura das camadas e composição química heterogênea. Forte fracionamento entre Mn e Fe ocorre nos nódulos diagenéticos e do tipo misto, assim como entre os metais traço Ni, Cu, Co e Ti. O Mn e o Fe estão presentes nos nódulos principalmente na forma de espécies oxidadas Mn4+ e Fe3+, independentemente do efeito de fracionamento entre eles. Modelos esquemáticos do ambiente de formação dos nódulos são propostos e sugere-se que variações da profundidade da frente redox no sedimento ao longo do tempo são responsáveis pelo efeito de fracionamento entre o Mn e o Fe.

Deep sea polymetallic nodules are concretions of manganese and iron oxides formed around a nucleus. They accrete either hydrogenetically - metals precipitate from the seawater - or diagenetically - metals precipitate from the sediment pore water. The accretion process affects both the nodules morphology and geochemistry. In this study, fourteen polymetallic nodules from four ocean regions, namely the Clarion-Clippertone Zone (Northeast Pacific Ocean), the Central Indian Basin (Central Indian Ocean), the Mascarene Basin (West Indian Ocean), and the Rio Grande Rise (Southwest Atlantic Ocean), were used to compare morphological and geochemical aspects between the different oceanic regions. Computed Tomography (CT) was applied to study the nodules internal structure. Scanning Electron Microscopy (SEM) was used to describe the micro layers within the nodules. Chemical composition of growth layers and nuclei was determined by both Micro X-ray Fluoscence (μ-XRF) and Laser Ablation Inductively Coupled Plasma Mass Spectroscopy (LA-ICP-MS). Finally, X-ray Absorption Near Edge Spectroscopy (XANES) was performed in order to determine the speciation (i.e., the oxidation state) of Mn and Fe. Polymetallic nodules from the Central Indian Basin are diagenetic and the ones from the Mascarene Basin and the Rio Grande Rise are hydrogenetic, while nodules from the Clarion-Clippertone Zone are of mixed type. However, the dominant accretion process varies across the nodules resulting in inhomogeneous layer textures and chemical composition. Strong Mn and Fe fractionation occurs in the diagenetic and mixed type nodules accompanied by fractionation of the trace elements Ni, Cu, Co and Ti. Mn and Fe are present in the nodules mainly as oxidized species Mn4+ and Fe3+, independently of the degree of fractionation. Schematic models of the nodules environment of formation are proposed, in which and the fractionation of Mn and Fe is possibly the result of the variation of the redox front depth through time. / Nódulos polimetálicos de mar profundo são concreções de óxidos de manganês e de ferro ao redor de um núcleo. Os nódulos crescem através da precipitação hidrogenética - precipitação de metais da água do mar - ou diagenética - precipitação de metais da água intersticial do sedimento. O processo de acreção reflete na morfologia e geoquímica dos nódulos. Neste trabalho, quatorze nódulos polimetálicos provenientes de quatro regiões oceânicas - Clarion-Clippertone Zone (Oceano Pacífico Nordeste), Bacia Central do Índico (Oceano Índico Central), Bacia Mascarene (Oceano Índico Oeste) e Elevação de Rio Grande (Oceano Atlântico Sudoeste) - foram usados a fim de se comparar os aspectos morfológicos e geoquímicos dos nódulos entre regiões diferentes. A estrutura interna dos nódulos foi avaliada através da Tomografia Computadorizada por Raios-X (CT). Microscopia Eletrônica de Varredura (SEM) foi usada para descrever as micro camadas. A composição química foi determinada por Micro Fluorescência de Raios-X (μ-XRF) e por ablação a laser acoplada a espectrometria de massa com plasma indutivamente acoplado (LA-ICP-MS). Por fim, a Espectroscopia de Absorção de Raios-X próximo à borda (XANES) foi realizada a fim de se determinar a especiação (i.e., o número de oxidação) do Mn e do Fe. Os nódulos polimetálicos da Bacia Central do Índico são diagenéticos e os da Bacia Mascarene e Elevação do Rio Grande são hidrogênicos, enquanto que os da Clarion-Clippertone Zone são do tipo misto. Entretanto, o processo de acreção varia ao longo dos nódulos, resultando em textura das camadas e composição química heterogênea. Forte fracionamento entre Mn e Fe ocorre nos nódulos diagenéticos e do tipo misto, assim como entre os metais traço Ni, Cu, Co e Ti. O Mn e o Fe estão presentes nos nódulos principalmente na forma de espécies oxidadas Mn4+ e Fe3+, independentemente do efeito de fracionamento entre eles. Modelos esquemáticos do ambiente de formação dos nódulos são propostos e sugere-se que variações da profundidade da frente redox no sedimento ao longo do tempo são responsáveis pelo efeito de fracionamento entre o Mn e o Fe.

Heme Oxygenase (HO) is an enzyme universally found in animals, plants and microbes. In plants, the role of heme oxygenase in the synthesis of the phytochrome chromophore is well recognized and has been extensively studied; however its role in regulating reactive oxygen species (ROS) in plants is just beginning to be explored, particularly in legumes. Legumes interact with Rhizobium bacteria to form symbiotic nitrogen fixing nodules. ROS plays an important role in the development of roots as well as symbiotic nodules. In the model legume Medicago truncatula, ROS in the root is regulated in part by the LATD/NIP gene. The M. truncatula giraffe mutant has a deletion that removes the entire HO coding sequence. We have found that the M. truncatula GIRAFFE HO regulates expression of some of the LATD/NIP-regulated ROS genes such as RESPRATORY BURST OXIDASE HOMOLOG C (RBOHC) and a cell wall peroxidase (cwPRX2) in seedlings. This means that the wild-type function of GIRAFFE is to up-regulate expression of RBOHC and cwPRX2 in roots, in contrast to LATD/NIP, which down-regulates them. We also found that LATD/NIP and GIRAFFE do not regulate expression of each other in seedlings. Given that the highest expression of GIRAFFE HO is in a senescing nodule, we tested the expression of ROS-regulatory enzymes in senescing nodules. We found that GIRAFFE up-regulates expression of RBOHC during nitrate-induced nodule senescence. At present, with changing climatic conditions and exposure to various environmental stresses that can alter ROS homeostasis, characterizing the role of GIRAFFE in the antioxidant machinery of legumes can be useful in improving crop productivity and for enhancing soil fertility.