Master of Science / Department of Mechanical and Nuclear Engineering / Douglas S. McGregor / Fission chambers are ideal neutron flux surveillance instruments to ensure nuclear reactor control and safety. They can provide online, in-core, real-time measurements covering the dynamic range of neutron flux including pulse, Campbell, and current mode over decades of reactor operation cycles. The first patented fission chamber was developed by Baer et al. in 1957. It was a cylindrical assembly thermal fission counter having sensitivity of 0.7 count/neutron cm⁻² for a background measurement of 5 counts/second with ability to operate at a temperature range of 20-80 ºC [3]. Since then, fission chamber technology was developed to come up with miniature and sub-miniature dimensions withstanding high irradiation and high temperature environment making them suitable for in-core online diagnosis. Since the introduction of high temperature fission chamber technology starting in the 1970’s, the need of the advancement in modeling of the fission chambers to improve their performance has become important. The development of modeling depends upon the understanding and consideration of underlying physics of these detectors. The validation of modeling of fission chambers will need the quantification of uncertainty introduced at every stage from neutron-deposit interaction to signal shaping. Based on this objective, a detailed review was performed on fission chamber modeling and simulation covering neutron flux self-shielding, fissile deposit evolution, fission product emission, auto-absorption, electron-ion pair creation, charge recombination and avalanche, space charge effect, charge transport, propagation of electronic pulse and pulse shaping. The analytical methods, algorithmic treatments, simulation, and computation codes used so far in case of modeling different aspects of fission chambers were reviewed. Along with the numerical methods and computer codes for simulating electron drift and charge transport for the usual gas chamber detectors, the use of several fissile material evolution techniques and computation codes were observed in case of fission chamber modeling. The use higher order statistics to handle fluctuation mode and to treat noisy data were observed. In recent years, fission chamber modeling made reasonable improvement in detail physics modeling. Several analytical methods like advanced statistics for Campbellng mode and electric field distortion due to space charge effect need to be incorporated in computation codes. More progress in the areas of evolution of gas behavior, consideration of Penning, recombination, and avalanche effect still needed.

For direct cycle gas cooled high temperature reactor designs, operating conditions may be limited as a result of excessive maintenance dose rates caused by the 110mAg source term on the turbine. The accurate prediction of silver fission and activation products’ release during normal operation is required to ensure regulatory compliance and economic viability of planned power plants. Fuel qualification programs should provide satisfactory results to ensure correct analyses, but will however not be available for many years. In the meantime data from the German fuel development program may be utilized. Traditionally diffusion models were used to derive transport parameters from limited irradiation testing of fuel materials and components. Best estimates for all applicable German fuel irradiation tests with defendable uncertainty ranges were never derived. However, diffusion theory and current parameters cannot account for all irradiation and heat-up test results, and for some tests, it appears unacceptably conservative. Other transport mechanisms have been suggested and alternative calculation models are being considered. In this thesis the relevant German material and irradiation tests were evaluated with the current PBMR metallic fission product release calculation model. Transport through all the fuel materials and components and from the sphere to the coolant gas was considered and best possible models and parameters were suggested. For the transport of silver through the SiC layer an alternative suggested model called the Molecular Vapour Transport Release (MVR) Model was evaluated against the traditional diffusion model. From this evaluation it was shown that classical diffusion modelling was still a viable model to predict silver transport in SiC. The MVR model was found to be a feasible model as well. However, due to the much larger verification and validation effort required, it was decided to use the diffusion model until such time that experimental results become available that might elucidate the exact physical transport model. The evaluation also showed that the diffusion model used must be quantified in a detailed evaluation of all applicable irradiation tests. A study of all German irradiation tests was previously performed and the applicable irradiation tests were identified. A detailed evaluation of these irradiation tests were performed with an updated diffusion model. New transport and material parameters were derived in this detailed evaluation and compared with existing values. An evaluation of some heat-up tests of irradiated fuel spheres was performed to assess the range for which the newly derived transport parameters are valid. The different models with their old and newly derived parameters were used to analyse sample PBMR cores. Recommendations were made to the suitability of the different models and parameters for future PBMR silver fission and activation product analyses. / Thesis (DPhil)--University of Pretoria, 2009. / Physics / unrestricted

Exonic sequences of eukaryotic genes are interspersed with introns which when accurately removed from the primary transcript (pre-mRNA) results in a functional transcript. These splicing reactions are carried out by the spliceosome, consisting of U1, U2, U4, U5, U6 snRNAs and 150 non-snRNP proteins, which assemble onto the pre-mRNA and catalyzes the two invariant transesterification reactions (Will and Luhrmann, 2006). The flexibility in choice of splice sites allows for alternative splicing which has immensely contributed to eukaryotic genome evolution and in diversifying the metazoan proteome (Nilesen and Graveley, 2010). Dynamic yet ordered interactions between U2, U5 and U6 snRNAs and Prp8, Prp16, Prp17, Prp18, Slu7 and Prp22 splicing factors are required in vitro for second-step of splicing of budding yeast and human model transcripts (Umen and Guthrie, 1995a; Horowitz, 2012). ScSlu7 aids 3’ss selection while its strongly associated partner ScPrp18 stabilises U5 snRNA-exonic interactions (James et al., 2002; Aronova et al., 2007). These factors are dispensable in vitro, for the splicing of introns with short branch nucleotide to 3’ss distances (Brys and Schwer, 1996; Zhang and Schwer, 1997). Nearly 43% of fission yeast genes have short introns, with degenerate splice-signals and unconventional Py(n) tracts (Kuhn and Kaufer, 2003). As these features differ extensively from budding yeast and are interestingly more representative of fungal and other eukaryotic introns, fission yeast is an attractive unicellular model to investigate alternate splice-site recognition and assembly mechanisms. Mechanistic details of the second catalytic step are poorly understood in fission yeast. Strikingly, mutations in 3’ss and Py(n) tract intronic cis elements, known to block second step splicing in budding yeast, cause pre-catalytic arrest with unspliced pre-mRNA accumulation in fission yeast (Romfo and Wise, 1997). Studies in our laboratory focussed on understanding the functions for fission yeast SpPrp18 and SpSlu7 predicted to be second-step factors, revealed remarkable differences as compared to their budding yeast counterparts. Unexpectedly, SpPrp18 and SpSlu7 were found by our lab to be required before catalysis and these proteins do not directly associate with each other. Genome-wide splicing studies in a missense slu7-2 mutant indicated widespread yet intron-specific splicing functions for SpSlu7 (Banerjee et al., 2013). Crucial functions were attributed to helix-5 and conserved region loop of SpPrp18 and in vivo splicing analysis in selected cellular transcripts in a missense mutant (V194R) also revealed intron-specific functions (Thesis, N Vijaykrishna). In this study, we have advanced our understanding of SpPrp18 functions by identifying its global substrates and correlating with its intron-specific roles. Through molecular and genetic approaches, we have probed its role in splicing/spliceosome assembly. We identified intronic features within substrates that increase the propensity for the requirement of SpSlu7 for efficient splicing. Further, using findings from the genome-wide alternative splicing patterns in SpSlu7 and SpPrp18 mutants, we have attempted to understand their role in splice-site choice and thus alternative splicing. Ia. Understanding global splicing functions and spliceosomal interactions of fission yeast splicing factor SpPrp18 Since SpPrp18 is an essential gene, our lab generated the strains (prp18-5int [V194R] and WTint), where the thiamine-repressible promoter allowed conditional expression of wild-type or mutant allele integrated at the heterologous leu1 locus. Splicing efficiency of certain cellular transcripts with differing intron characteristics was assessed by semi-quantitative RT-PCR studies and the data suggested intron-specific SpPrp18 roles (in collaboration with Vijaykrishna N). This prompted us to investigate the global splicing role for SpPrp18 for which we used splicing-sensitive microarrays having custom-designed probes to distinguish unspliced pre-mRNA and spliced mRNA for every individual pombe intron. RNA from prp18-5int (V194R) and WTint cells was used in these experiments. We derived a stringent dataset of 258 introns which were statistically significant and correlated in two biological replicate RNA samples, for various probes. Hierarchical clustering of this dataset showed that the depletion of wild-type SpPrp18 triggered a range of splicing phenotypes like (A) pre-mRNA accumulation with mRNA reduction (B) pre-mRNA accumulation (C) spliced mRNA reduction and (D) unchanged pre-mRNA and mRNA levels. Statistical analysis of cis motifs that may correlate with the substrate-specific SpPrp18 splicing functions was done, but the data showed a lack of a global discriminatory primary sequence feature. However, a subtle intron-specific role for Py(n) tracts located between 5’ss and BrP was deduced for SpPrp18. This lead was validated by examining the in vivo splicing efficiency of minitranscripts with wild-type or an altered Py tract length, carried out for a SpPrp18 dependent and an independent intron. To specifically address if SpPrp18 activity was required for second-step splicing we investigated, using primer extension analyses, for lariat intron-3’exon species, an intermediate formed after step 1. We observed that even in prp18-5int dbr1∆ double mutants (where lariat molecules are not degraded) the cells accumulate only unspliced pre-mRNA and not lariat intermediates, a signature of an early arrest prior to the first transesterification reaction. Strengthening these findings, positive genetic interactions were noted between prp18-5int and ts mutants in two factors (U2AF59 and SpPrp1) involved in precatalytic spliceosome assembly and activation. On the whole, our genome-wide studies indicate intron-specific pre-catalytic functions for SpPrp18 supported by genetic interactions with early acting splicing factors involved in spliceosomal assembly and activation. Ib. Identification of intronic features that determine substrate-specific splicing functions for SpSlu7 In vitro studies with ScSlu7 and hSlu7 show their influence in 3’ss selection when BrP to 3’ss distance is greater than 7 nts and 23 nts respectively; but the global substrates are not known in either species (Brys and Schwer, 1996; Chua and Reed, 1999b). Genome-wide analysis of the splicing efficiency changes in cells with the mis-sense spslu7+ mutant (slu7-2), previously carried out in our lab, revealed a spectrum of splicing defects (Banerjee et al., 2013). To further understand the intron context-specific roles for SpSlu7, we examined intronic cis features that may correlate with SpSlu7 dependence. Statistical analyses of the affected (422 introns) and unaffected categories (90 introns) revealed that intron length, BrP to 3’ss distance and AU content are multiple discriminatory cis features that govern SpSlu7 splicing functions. To assess the contribution of these intronic features we tested whether altering these cis elements changes a transcript’s dependency (or otherwise) on SpSlu7 by RT-PCR analyses. For these studies, we generated plasmid expressed mini-genes containing the respective wild-type intron or intron with altered BrP-3’ss distances. We used nab2+ I2 as a case of an intron spliced independent of SpSlu7 and rhb1+ I1 as a representative for SpSlu7 dependent intron. Experiments testing their in vivo splicing status proved that BrP-3’ss distance is a cis feature that dictates SpSlu7 splicing functions in a context-dependent manner. The intronic AU content particularly between the 5’ss and the BrP was assessed in minigene constructs where a chimeric intron was generated by swapping the low AU containing sequences in the 5’ss to BrP stretch of cdc2+ I2 with AU rich bpb1+ I1 5’ end sequences. The results reaffirmed that low intronic AU content particularly at the 5’ end co-relates with SpSlu7 dependency. Hence, we have deduced novel intronic elements, which perhaps in combination, create a contextual dependence for SpSlu7 to facilitate efficient splicing. II. Alternative splice-site selection in fission yeast and studies on the role of splicing factors SpSlu7 and SpPrp18 Budding yeast second-step splicing factors ScSlu7 and ScPrp18 mediate 3’ss choice in the single intron containing transcripts. Fission yeast genome encodes cis and trans factors that promote alternative splicing similar to higher eukaryotes. In this study, we have devised a data analysis pipeline to identify alternative splice events in multi-intronic transcripts of fission yeast. Further, we utilised this information to interrogate the global role for SpSlu7 and SpPrp18 in alternate splice site selection. We mapped the microarray probe sequences corresponding to all theoretically possible non-consecutive splice junctions of S. pombe transcripts onto two independent experimental next-generation (NGS) transcriptomes from wild-type samples and identified 104 exon skipping events with NGS reads more than 3 (Wilhelm et al., 2008; Rhind et al., 2011). We further generated a stringent list of ten exon skipping events having high sequence reads as well as raw intensity value in our microarray experiments with wild-type cells. Two representative events from this list, an abundant rps13+ exon 2 skipped alternative mRNA and less abundant ats1+ exon 3 skipped alternative mRNA were then taken up for experimental analyses by semi-quantitative RT-PCR assays. We confirmed these events and further noted that SpSlu7 and SpPrp18 were required for the constitutive splicing of ats1+ E2-I2-E3-I3-E4 cassette. On the other hand, SpSlu7, and not SpPrp18, exerted a subtle influence on the skipping of exon 3. In addition to exon 3 skipped mRNA, we detected an intron 3 retained ats1+ alternative mRNA (E2-E3-I3-E4) in wild-type cells. Assessment of this event in cells metabolically depleted of SpSlu7 and SpPrp18 showed a reduced abundance of this species in both instances. This suggests a role for functional SpSlu7 and SpPrp18 in retaining intron 3 in ats1+ transcripts in vivo. Among the ten microarray probes, custom-designed to detect specifically the mRNA isoforms arising from altered use of donor 5’ splice sites, we were able to detect in wild-type cells the utilisation of a downstream alternate 5’ss in intron 1 of D-Tyr-tRNA deacylase. Comparative assessment of this splicing event in prp18-5int and slu7-2 mutant cells revealed that SpPrp18 is preferentially required for the utilisation of its alternative 5’ss and such a role has not yet been attributed to its budding yeast and human homologs. On the other hand, SpSlu7 was required equally for utilisation of canonical and non-canonical 5’ss. Differential requirement for SpSlu7 for the utilisation of an upstream non-canonical 3’ss and the canonical 3’ss in DUF3074 intron 1, was noted. This role of SpSlu7 in 3’ss selection is similar to that known from in vitro studies of its budding yeast and human counterparts. Overall, we identified and experimentally validated novel alternate splice events in fission yeast and we infer an important role for SpSlu7 and SpPrp18 in both 5’ss and 3’ss selection.

Un dysfonctionnement mitochondrial est impliqué dans plusieurs maladies neurodégénératives, corrélé avec une augmentation des niveaux de stress oxydant. Les anomalies mitochondriales observées dans les tissus des patients, les modèles animaux et cellulaires des maladies de Huntington et de Parkinson, suggèrent l'implication de la mitochondrie dans leur pathogénie.Les deux projets discutés dans ce manuscrit se focalisent sur le rôle des aspects particuliers de la physiologie mitochondriale au cours des deux maladies. / Mitochondrial dysfunction has been implicated in several neurodegenerative diseases and is correlated with augmented levels of intracellular oxydant stress. The mitochondrial defects observed in tissues from patients, as well as in animal and cellular models of Huntington’s and Parkinson’s diseases, suggest the implication of mitochondria in the pathogenesis of these diseases. The two projects discussed in this manuscript focus on the role of particular aspects of mitochondrial physiology in these diseases. By the first project we show the role of defective mitochondrial respiratory chain compex II in several rodent models of Huntington’s disease. By using a lentivirus-based gene transfert strategy we highlight the neuroprotective potential of the striatal overexpression of the subunits of complex II. The second project focus on Parkin and PINK1, two proteins implicated in the autosomal recessive, hereditary forms of Parkinson’s disease and in mitochondrial quality control mechanisms, such as mitophagy. In a cellular model we show that the two proteins facilitate Drp1-dependent mitochondrial fission. We show that Parkin may facilitate the signaling pathways controlling the activity of the pro-fission protein Drp1. This effect is probably indirect and mostly PINK1-independent. On the contrary, in mitochondrial depolarization conditions, by FRET (Förster Resonance Energy Transfer) a direct spatial coordination of Parkin, PINK1 and Drp1 is observed, which seems to be determinant for the efficiency of mitophagy. My projects shed new light on pathogenic mechanisms and open new perspectives in the research on these diseases.

Les céramiques utilisées dans les Réacteurs à Eau Presurisée sont constituées de dioxyde d'uranium. Irradiée à fort taux de combustion en réacteur, elles présentent en périphérie de pastille une microstructure particulière, dénommée RIM, avec des pores de l'ordre du micromètre fortement pressurisés en gaz de fission. Lors des traitements thermiques (TT) simulant des situations incidentielles ou accidentelles de réacteur, un relâchement important de la zone de RIM est observé. Nous avons considéré que le mécanisme de relâchement du gaz contenu dans les bulles pressurisées est la fracturation mécanique des joints de grains du RIM. Puis nous avons comparé les différents types de sollicitations mécaniques auxquelles sont soumis un joint de grain à la contrainte à rupture de l'oxyde. La première sollicitation est induite par les bulles de gaz surpressurisées du RIM ; elle impose un champ de contrainte à un niveau microscopique i.e. à l'échelle d'une bulle de gaz et son environnement local. La seconde sollicitation est générée par l'interaction mécanique entre la pastille et la gaine. cette sollicitation impose un champ de contrainte à un niveau macroscopique i.e. à l'échelle de la zone de RIM et de son environnement global. La dernière sollicitation résulte de la déformation due à l'évolution structurale du RIM en TT. Les résultats expérimentaux de la thèse montrent que les champs de contraintes microscopiques et macroscopiques n'expliquent pas la fracturation des joints de grains du RIM en TT. Les sollicitations induites par l'évolution structurale du RIM en fonction de la température est un mécanisme possible pour expliquer le comportement mécanique global du RIM en TT. / The ceramics used in Power Water Reactors ar made of uranium dioxide. Irradiatd at high Burn-up, they present a characteristic zone in periphery called High Burn-Up Structure or RIM zone with micrometer pores containing over-pressurizzed gas bubbles. Annealing texts simulating incidental or accidental reactor situations, a strong release of the RIM zone is observed. We have considered that the fission gas release mechanism is the mechanical fracture of the RIM grain boundaries. The we have compared the diffrerent types of mechanical stress applied to a grain boundary with the fracture stress of the oxide. The first stress is due to RIM over-pressurized gas bubbles, these bubbles apply a stress field determined at a microscopic level i.e. at the gas bubbles scale and its local environment. The second stress is generated by the Pellet Cladding Mechanical Interaction (PCMI). This stress applies a stress field on a microscopic scale i.e. at the RIM zone and its overall environent. The last stress is occured by a strain due to the RIM structural evolution during annealing test. The experimental results show that microscopic and macroscopic stress fields to do not explain the RIM grain boundary fracture during annealing test. The stresses induced by the RIM structural evolution as a function of the temperature is a possible mechanism to explain the overal mechanical behavior of the RIM zone during annealing test.

No description available.

Epigenetics is an area of biology that studies heritable changes in gene ex- pression without any change in the DNA sequence. The most studied epige- netic mechanisms are DNA methylation, RNA interference and histone mod- ifications. There are over 130 different modifications that can be attached to histones, and the most commonly studied are methylation, acetylation, phos- phorylation, sumoylation and ubiquitination. The modifications, spread out through the genome, form the histone code, which recruits transcription fac- tors and modifies the accessibility of the DNA, which results in either active or silenced transcription. The silenced form of chromatin is known as heter- ochromatin and is usually found in regions of the chromosome that need to be highly regulated. To study epigenetics, the model organism Schizosac- charomyces pombe is used widely used. S. pombe is a single cell, rod shaped, fission yeast. The simplicity of S. pombe and its similarities to high- er eukaryotes makes it a good model organism for studying epigenetics. We find that, when mutating evolutionary conserved amino acids in the Clr2 protein, which is involved in heterochromatin formation in S. pombe, there is a change in silencing in different heterochromatic regions. When constructs of Clr2 with the BAH domain deleted are overexpressed, there is an increase in silencing in the central core centromere of chromosome II of S. pombe.

Group living is a social strategy adopted by many species, where individuals can exhibit long-term social affiliation with others, strengthened through cooperative behaviour and often kinship. For highly social mammals, changes in group membership may have significant consequences for the long-term viability and functioning of a population. Detecting significant social events is essential for monitoring the social dynamics of such populations and is crucial to determining the factors underlying these events. Detecting when changes in social organization occur, especially with incomplete data, poses significant analytical challenges. To resolve this issue, I developed and assessed a straightforward, multi-stage and generalizable method with broad utility for ecologists interested in detecting and subsequently investigating causes of changes in social organization. My approach illustrates the frequency and ecological relevance of group fission and fusion events in a population of fish-eating ‘Resident’ killer whales (Orcinus orca). Group fission is a process commonly found in social mammals, yet is poorly described in many taxa, and has never been formally described in killer whales. To address this gap, I provided the first description of matrilineal fission in killer whales, from a threatened but growing Northern Resident killer whale population in which matrilineal fission has been observed for the past three decades. I also undertook the first comprehensive assessment of how killer whale intragroup cohesion is influenced by group structure, demography and resource abundance. Fission in Northern Resident killer whales occurred both along and across maternal lines, where animals dispersed in parallel with their closest maternal kin. I show that fission in this population is driven primarily by population growth and the demographic conditions of groups, particularly those dictating the nutritional requirements of the group. I posit that intragroup food competition is the most likely explanation for group fission in this population, where prey abundance also has ancillary effects. As group fission can have a direct impact on the fitness of group members and the long-term viability of a population, my findings underscore the importance of incorporating studies of sociality into the management of threatened populations of social mammals. / Graduate / 0329 / 0472

京都大学 / 0048 / 新制・課程博士 / 博士(理学) / 甲第22876号 / 理博第4642号 / 新制||理||1667(附属図書館) / 京都大学大学院理学研究科化学専攻 / (主査)教授 寺西 利治, 教授 島川 祐一, 教授 長谷川 健 / 学位規則第4条第1項該当 / Doctor of Science / Kyoto University / DGAM

京都大学 / 新制・課程博士 / 博士(理学) / 甲第23018号 / 理博第4695号 / 新制||理||1673(附属図書館) / 京都大学大学院理学研究科地球惑星科学専攻 / (主査)准教授 河上 哲生, 教授 田上 高広, 教授 下林 典正 / 学位規則第4条第1項該当 / Doctor of Science / Kyoto University / DFAM