Rozprawy doktorskie na temat „Spatial pathways”

This thesis concentrates on the induction of the immediate early genes zif268 and c-fos in a functional and dysfunctional brain network. Initial studies focused on the creation of a task to allow the study of immediate early gene activation after working memory. Previous studies using such a paradigm have compared the animals performing the experimental task with poorly matched control groups. Experiments in this thesis attempted to rectify this. A group of experiments using the water maze found decreases in Zif268 activation in the experimental group as compared to control groups, mainly in the hippocampus and some parahippocampal areas. This was believed to have arisen because of a streamlining of the brain network in the Working Memory group. An increase in c-Fos immunoreactivity was seen in the Working Memory group as compared to controls in prefrontal regions. Structural equation modelling analysis was performed, which allows immediate early gene counts to be used to analyse networks of brain regions. In the Working Memory group connections were seen between the parahippocampal regions and the subiculum that progressed via the hippocampus, indicating that the hippocampus was still engaged by the task. In the control group analysed no such hippocampal pathway was found. This water maze task was then used to study zif268-EGFP activation in a novel transgenic rodent model, where the Zif268 promoter drives the expression of a fluorescent protein EGFP. Activation of both EGFP and Zif268 immunoreactivity was seen in the CA1 region of the animals performing the control task. No EGFP activation was seen in this region in the Working Memory group even though EGFP expression was seen in other regions. The GFP protein was also able to be seen under direct visualisation in the CA1 and dentate gyrus region of control animals. Concerning the dysfunctional brain, gene expression was analysed in the retrosplenial cortex after NMDA lesions of the anterior thalamic nuclei. Previous research has shown that lesions produce a dramatic hypoactivity in the protein products of the immediate early genes c-fos and zif268. Microarray analysis of retrosplenial tissue revealed that as well as decreases in expression of genes related to repair and cell adhesion/neurogenesis, an increase in c-fos mRNA was seen in the lesion hemisphere of the brain. This pattern of expression is opposite to that of the protein. Possible reasons for this are discussed.

Thesis advisor: Rudolph Hon
Groundwater in the Shepley's Hill Landfill (SHL) area had a high arsenic concentration for at least 20 years. This study is aimed at understanding the migration pathways of arsenic in the SHL area and postulating the possible sources and mechanisms for the mobilization of arsenic. A direct-push sampling technique (DPT) was applied in the summer of 2010 within the SHL and its downgradient area, which provided groundwater samples from various depths and locations. A long-term monitoring dataset from 1998 to 2009 was utilized as a subsidiary source for temporal analysis. Spatial distributions of arsenic and other major dissolved compositions were analyzed. Extremely high arsenic concentrations (up to ~ 15000 µg/L) were detected in the deep glacial sand deposits close to a peat layer within the SHL. Arsenic concentrations decrease dramatically in the downgradient area north of the SHL. The transport of arsenic in the SHL area is similar to that of iron. The source of arsenic is likely to been within the boundary of the SHL. The glacial sand overburden within the SHL provides enough source for the arsenic mobilization. A possible mechanism of arsenic mobilization in the SHL area is that the reductive dissolution/desorption of arsenic from iron bearing minerals under a lasting reducing environment created by decompositions of organic matter in waste and peat
Thesis (MS) — Boston College, 2013
Submitted to: Boston College. Graduate School of Arts and Sciences
Discipline: Earth and Environmental Sciences

The dramatic global trend of population growth has led to a rapid urbanisation, resulting in unprecedented land cover change. The incarnation of accompanying developed has typified impermeable surfaces. These surfaces have disconnected the stormwater component of the natural hydrological cycle, disregarding it as a nuisance and designing it to be rapidly removed from urban areas. Utilising Sustainable Urban Drainage Systems (SUDS) offers opportunities in urban areas to recycle the water and challenge the perception that stormwater is a nuisance and of no value. The current context of drought experienced by Cape Town has highlighted the need for less reliance on surface water resources; implementing SUDS could be a way of reconnecting the hydrological urban water cycle. It could also help to repair the human disconnect from nature that is prevalent in urban areas. The research question explored the role of spatial planning in enabling the implementation of SUDS in the City Bowl, Cape Town. While conceptual and technical frameworks have been developed for SUDS in South Africa, at present there is no spatial guide as to how these interventions could be realised in a specific context and area. This research utilise s the tools of spatial planning to re-imagine the City Bowl in relation to water. The case study methods used, enabling a detailed understanding of the site. This was complemented by interviews with various planning professionals in order to understand the current role spatial planning plays in terms of implementing SUDS. The research suggest is that whilst SUDS has many constraints, the opportunities that they provide for improving water quality and quantity, and surrounding amenities, suggests that this is one which has to be embraced if the City Bowl is going to respond innovatively and sustainably to the drought. It also highlights the need to improve coordination across different spheres and departments of governance, and emphasises the need to value local community knowledge. A prevalent silo approach to complex problems is no longer acceptable. The implications of the research are that implementing SUDS in the City Bowl requires planners to embrace a water literacy approach to spatial plans, and in doing so, return the focus to water

Thesis (M.A.)--Kent State University, 2009.
Title from PDF t.p. (viewed Mar. 16, 2010). Advisor: Marilyn Norconk. Keywords: White-faced sakis; goal-directed foraging; spatial memory; mental maps; arboreal pathways; distance-minimization; topographical mental mapping; preference. Includes bibliographical references (p. 183-194).

Data visualization is an essential methodology for bioinformatics studies. Spatial Transcriptomics(ST) is a method that aims at measuring the transcriptome of tissue sections while maintaining its spacial information. Finally, the study of biological pathway focuses on a series of biochemical reactions that take place in organisms. As these studies generate a large number of datasets, this thesis attempts to combine the ST’s data with pathwayinformation and visualize it in an intuitive way to assist user comprehension and insight.In this thesis, Python was used for integrating the dataset and JavaScript libraries wereused for building the visualization. The processing of ST pathway data together with the data visualization interface are the outcomes of this thesis. The data visualization can show the regulation of pathways in the ST data and can be accessed by modern browsers. These outcomes can help users navigate the ST and pathway datasets more effectively.
Datavisualisering är en viktig del av bioinformatik. Spatial transkriptomik (ST) är en metod som mäter transkriptom, samtidigt som den behåller spatial information. Biologiskapathways å andrasidan fokuserar på biokemiska reaktioner som sker inom organismer. Dessa studier genererar mycket data, och denna avhandling försöker att kombinera ST-data med pathway information och få en intuitiv visualisering av det integrerade datat.I avhandlingen användes Python för att integrera datat och JavaScript bibliotek för attbygga visualiseringsverktyget. Avhandlingen resulterade i en metod för att integrera STdata och pathway information, samt ett visualiseringsverktyg för ovan nämnda information.Verktyget kan visa pathway regulationer i ST data och kan användas i moderna webbläsare.Forskningen resulterade i ett verktyg som kan hjälpa forskare att förstå ST och pathwaydata.

This dissertation contributes to the area of Applied Linguistics and foreign language teaching and learning by investigating whether formal instruction, as opposed to no specific instruction, plays a significant role in the acquisition of Spanish spatial prepositions: "en", "sobre", "de," and "a." The study investigates the effect of formal instruction for spatial concepts for which English native speakers could use the L1 to generate correct responses in the L2 and for spatial concepts that created confusion between the L1 and the L2. It also investigates the effect of formal instruction when prepositions are taught by rules. The study introduces a visual/spatial/kinesthetic methodology based on the Davis Symbol MasteryRTM program, originally designed to be used with juvenile and adult dyslexics. The study investigates whether 3-D clay modeling can create new mental representations of spatial concepts not existent in the L1 or resolve overlapping spatial concepts between the L1 and the L2. Advanced university learners of Spanish as a Second Language participated in this study. Results show that (a) formal instruction has a significant general effect for the mixed spatial concepts chosen for this study; (b) formal instruction has no significant effect over no specific instruction for concepts for which English native speakers could use the L1 to generate correct responses in the L2; (c) formal instruction, specifically 3-D clay intervention, can either help resolve the confusion in case of overlapping of spatial concepts between the L1 and L2 or create new mental representations not existent in the L1; and (d) formal instruction has no significant effect in the case of instruction by rules. The dissertation offers a novel theoretical explanation for why 3-D clay modeling may help resolve confusion in the case of overlapping of spatial concepts between the L1 and L2 or create new mental representations not existent in the L1. Vygotsky's Tools for Cognitive Development are extended: 3-D clay modeling provides a tool that is both concretely grounded and consciously systematically accessible. The dissertation also discusses motivation in learning based on Csikszentmihalyi's "Flow Theory".

South African cities are still largely unequal where the urban poor’s right to the city is not adequately addressed. As a result of the legacy of apartheid and the various housing policies that have been adopted to address housing segregation, Johannesburg remains spatially fragmented with the urban poor remaining isolated in the periphery of the city far from places of employment and opportunity. With the decentralization and deindustrialization of employment, commuting for the urban poor has become more time consuming and costly with an average monthly commute costing R1 500.00. The City of Johannesburg has committed to tackling these structural and spatial inequalities that exist in Johannesburg through its flagship programme called Corridors of Freedom. Corridors of Freedom aims to transform the spatial inequalities in Johannesburg through the creation of geographies of inclusion using transit oriented development (TOD). This is through investing in public transport with the development of the Rea Vaya Bus Rapid Transit System (BRT) and inclusionary housing as the backbone to eradicate spatial. Corridors of Freedom aims to address housing segregation through the extension of the Rea Vaya BRT system in peripheral townships and through inclusionary housing provision to promote inclusive development in Johannesburg. This study will evaluate whether Corridors of Freedom and its associated developments have the potential to improve the lives of potential beneficiaries of two townships, Kagiso in the West Rand and Soweto. The study seeks to understand how the presence of the Rea Vaya BRT system can assist the urban poor’s access to the city and places of employment and opportunity. A comparison is made between Soweto residents who directly benefit from using the Rea Vaya buses and the Kagiso residents who do not have access to the Rea Vaya buses. Several stakeholders were interviewed including both the residents of Kagiso and Soweto. It was concluded that the Rea Vaya BRT system is a poverty-reduction strategy for the urban poor and an attempt from the City of Johannesburg to correct the existing spatial inequalities.The Rea Vaya Bus Rapid Transit system along with well-located social housing is an important step towards addressing inclusivity and bringing people close to places of employment and opportunity.

Après le vieillissement, le diabète Mellitus de type 2 (DMT2) est le facteur de risque le plus important pour développer la maladie d'Alzheimer (MA). Le DMT2 est une maladie métabolique caractérisée par une hyperglycémie et une résistance à l’insuline qui se développe vers la cinquantaine et est fortement favorisée par l’obésité. Nous avons exploré l’impact potentiel du DMT2 sur le développement de la MA chez le Rat. Pour cela, nous avons utilisé un régime alimentaire cafétéria (RC) couplé à des injections de faibles doses de Streptozotocine (STZ) (STZ-CD). Les rats STZ-CD montrent des signes classiques de DMT2 et des déficits légers de consolidation en mémoire de reconnaissance spatiale. Afin d'imiter le développement des stades précoces de la MA, la moitié des rats reçoivent une infusion intracérébrale de peptides β-amyloïdes solubles (Aβ) qui ne conduisent pas à des déficits mnésiques durables. Par contre, le phénotype DMT2 chez les rats STZ-CD exacerbe les déficits mnésiques observés avec le peptide Aβ en les prolongeant dans le temps. L’enrichissement environnemental pendant une période critique de 2 semaines après l’infusion d’Aβ est capable de compenser les déficits mnésiques induits par le peptide Aβ et/ou le traitement STZ-CD ; mais d’une manière limitée dans le temps. Des analyses biochimiques dans l’aire CA1 de l’hippocampe ont été effectuées pour explorer de possibles altérations de la voie PI3K, de marqueurs de la cascade amyloïde et du DMT2. Le peptide Aβ seul induit peu de changements durables ; le phénotype DMT2 seul est associé à des changements pour quelques protéines-clé, largement en liaison avec le régime cafétéria. Par contre, la majorité des modifications dysfonctionnelles de protéines est observée chez les rats montrant un phénotype de type DMT2 et recevant le peptide Aβ. Ces altérations, similaires à celles rapportées chez des patients atteints de la MA et chez des modèles animaux de la MA, concernent notamment des protéines de la voie PI3K-Akt impliquée dans des fonctions comme l’autophagie et l’inflammation et des marqueurs de la MA. L’altération de ces protéines pourrait contribuer aux déficits mnésiques durables observés et mettre en lumière des mécanismes moléculaires induits par le DMT2 et promouvant un milieu neuronal favorisant le développement d’un stade précoce de la maladie d'Alzheimer
Following aging, type 2 Diabetes Mellitus (T2DM) is the most important risk factor of developing Alzheimer’s disease (AD). It is a metabolic disorder characterised by hyperglycemia and insulin resistance that develops in middle age and is promoted largely by obesity. In this study, we used a T2DM rat model to assess the potential impact T2DM may have on the development of AD. Rats were fed cafeteria-style diet (CD) coupled with low dose injections of Streptozotocin (STZ)(STZ-CD). We found that STZ-CD treated rats showed classic signs of T2DM and a modest deficit in consolidation of spatial recognition memory. In order to mimic the development of early stage AD, half of the rats were infused with a soluble oligomeric amyloid beta (Aβ), which alone was not sufficient to induce long-lasting memory deficits. Interestingly, the T2DM phenotype exacerbated the memory deficits induced by Aβ infusion by prolonging these deficits. Environmental enrichment during a critical two-week period following infusion of Aβ rescued memory deficits induced by Aβ and/or STZ-CD treatment; however, this was time-limited. Biochemical analyses were conducted mainly in proteins involved in the PI3K-Akt signalling pathway and markers of AD and T2DM in CA1 of the hippocampus. Aβ alone induced few long-lasting changes; T2DM phenotype alone induced some changes that were largely mediated by CD treatment alone; however, the majority of dysfunctional regulation of proteins was observed in rats showing a T2DM phenotype that were infused with Aβ. More importantly, many of these changes are similar to those reported in brains of AD patients or rodent models of the disease; notably key proteins in the PI3K-Akt signaling pathway that mediate functions such as autophagy, inflammation and markers of AD. Dysregulation of these proteins may contribute to the long-lasting memory deficits seen in this model, which may provide evidence of molecular mechanisms induced by T2DM that could promote a dysfunctional neuronal environment favouring the development of early stages of Alzheimer’s disease

Frequent use of phenylurea herbicide isoproturon (IPU) in agricultural fields has resulted not only in the contamination of the natural resources including soil and water but also in the adaptation of the soil microflora to its rapid degradation. However, up to now, the mechanisms underlying this microbial adaptation are not well elucidated. The aim of this study was to explore the processes and factors implicated in IPU degradation from the agricultural field to the genes coding for catabolic genes. The study carried out at the experimental field of Epoisses cropped with a winter wheat / barley / rape seed crop rotation indicated that as a result of its periodically repeated use, the soil microflora adapted to IPU mineralization activity. Further analysis using exploratory and geostatistical tools demonstrated the existence of spatial variability in IPU mineralization activity at the field scale which was correlated not only with several soil physico-chemical parameters like organic matter content, CEC and C/N ratio but also with the pesticide application plan over a three year crop rotation. In order to get further insight into underlying mechanisms, an IPU mineralizing bacterial culture and strain Sphingomonas sp. SH were isolated through enrichment cultures performed from two different adapted soils. Both had the catabolic activities highly specific for the mineralization of IPU and its metabolites but none of other structurally related phenylurea herbicides. IPU metabolic activity of both the mixed culture and the strain SH was found to be affected by pH with optimal activity taking place at pH 7.5. Based on the accumulation of different known metabolites during mineralization kinetics, IPU metabolic pathway was proposed to be initiated by two successive demethylations, followed by cleavage of the urea side chain resulting in the accumulation of 4-isopropylaniline, and ultimately the mineralization of the phenyl ring. In order to identify the genes involved in IPU degradation, BAC clone library was established from the genomic DNA of the bacterial culture. Although, the functional screening did not yield in identifying any BAC clone able to degrade IPU or its known metabolites, the PCR based screening led us to identify a cat gene cluster involved in ortho-cleavage of the phenyl ring of catechol through beta-ketoadipate pathway. Based on this finding, it was hypothesized that phenyl ring of 4-isopropylaniline formed during IPU transformation might be mineralized through ortho-cleavage of catechol. This finding allowed us to propose the lower IPU metabolic pathway which was not yet described.

Sound localisation is one of the most representative function of the auditory system and, as such, it has been extensively investigated across species. Spatial hearing can be dramatically altered across the life span, yet research in humans have highlighted the remarkable capacity of the brain to adapt to changes of listening conditions, such as temporary ear plugging or long lasting hearing impairments. Although several investigations have examined accommodation to altered auditory cues (Chapter 1), a common theoretical framework seems to lack and a number of questions remain open. This limits the possibility to translate our current knowledge into concrete clinical applications for individuals who experience spatial hearing difficulties after hearing loss. The current dissertation reflects the attempt to answer specific questions regarding the process of sound localisation. The first study (Chapter 2) aimed to investigate the relation between different reference frames in spatial hearing, namely egocentric and allocentric sound representation. We studies this topic in the context of a learning paradigm, assessing to what extent localisation of single sounds in simulated monaural hearing (unilateral ear plugging) can improve following an audio-visual spatial hearing training focused on egocentric sound processing vs allocentric sound processing. An untrained group was also included in the study. We found that localisation performance in the horizontal plane improved specifically in the side ipsilateral to the ear-plug for all groups. Yet, the trained groups showed a qualitatively different change of performance after four days of multisensory ego/allocentric training compared to the untrained group, providing initial evidence of the possible role of allocentric coding in acoustic space re-learning. These results further highlight the importance of including a test-retest group in paradigms of sound localisation training. The second study (Chapter 3) focused on a specific aspect of the phenomenological experience of spatial hearing, namely the subjective confidence about the perceived sound position. We examined the relation between objective localisation accuracy and subjective certainty while participants localised sounds in two different listening conditions – binaural or simulated monaural hearing. Results showed that overall subjective certainty on sound position decreased in the altered listening condition (unilateral ear-plugging). In simulated monaural hearing, localisation accuracy and spatial confidence dissociated. For instance, there were trials in which participants were accurate, but felt uncertain, and trials in which they were less accurate but expressed higher ratings of spatial confidence on sound position. Furthermore, subjective confidence increased as a function of time within the testing block, and it was related to the spatial distribution of the perceived sound-source position. The third study (Chapter 4) exploited magnetoencephalography (MEG) to study the dynamics of the cortical network implied in active sound localisation. We implemented a novel apparatus to study sound localisation in MEG with real sounds in external space, and collected behavioural and subjective responses (i.e., accuracy and confidence, as in Study 2) during this altered listening condition. Results showed that participants were able to perceive the spatial difference between the positions of stimulation, thus proving the reliability of our novel setting for the study of spatial hearing in MEG. MEG data highlight a distributed bilateral cortical network involved in active sound localisation, which emerged shortly after stimulus presentation (100—125 ms). The network comprise the classical dorsal auditory pathway plus other cortical regions usually underestimated in previous literature – most notably, regions in the central sulcus/precentral gyrus possibly involved in head movements. Connectivity analysis revealed different patterns of neural coupling, as a function of frequency band. In particular, coherence in high gamma revealed significant connections involving the parietal cortex and the posterior superior temporal cortex. In the final chapter (Chapter 5), I summarise the main findings of the three studies, discuss their implications and outline potential future directions.

Sound localisation is one of the most representative function of the auditory system and, as such, it has been extensively investigated across species. Spatial hearing can be dramatically altered across the life span, yet research in humans have highlighted the remarkable capacity of the brain to adapt to changes of listening conditions, such as temporary ear plugging or long lasting hearing impairments. Although several investigations have examined accommodation to altered auditory cues (Chapter 1), a common theoretical framework seems to lack and a number of questions remain open. This limits the possibility to translate our current knowledge into concrete clinical applications for individuals who experience spatial hearing difficulties after hearing loss. The current dissertation reflects the attempt to answer specific questions regarding the process of sound localisation. The first study (Chapter 2) aimed to investigate the relation between different reference frames in spatial hearing, namely egocentric and allocentric sound representation. We studies this topic in the context of a learning paradigm, assessing to what extent localisation of single sounds in simulated monaural hearing (unilateral ear plugging) can improve following an audio-visual spatial hearing training focused on egocentric sound processing vs allocentric sound processing. An untrained group was also included in the study. We found that localisation performance in the horizontal plane improved specifically in the side ipsilateral to the ear-plug for all groups. Yet, the trained groups showed a qualitatively different change of performance after four days of multisensory ego/allocentric training compared to the untrained group, providing initial evidence of the possible role of allocentric coding in acoustic space re-learning. These results further highlight the importance of including a test-retest group in paradigms of sound localisation training. The second study (Chapter 3) focused on a specific aspect of the phenomenological experience of spatial hearing, namely the subjective confidence about the perceived sound position. We examined the relation between objective localisation accuracy and subjective certainty while participants localised sounds in two different listening conditions – binaural or simulated monaural hearing. Results showed that overall subjective certainty on sound position decreased in the altered listening condition (unilateral ear-plugging). In simulated monaural hearing, localisation accuracy and spatial confidence dissociated. For instance, there were trials in which participants were accurate, but felt uncertain, and trials in which they were less accurate but expressed higher ratings of spatial confidence on sound position. Furthermore, subjective confidence increased as a function of time within the testing block, and it was related to the spatial distribution of the perceived sound-source position. The third study (Chapter 4) exploited magnetoencephalography (MEG) to study the dynamics of the cortical network implied in active sound localisation. We implemented a novel apparatus to study sound localisation in MEG with real sounds in external space, and collected behavioural and subjective responses (i.e., accuracy and confidence, as in Study 2) during this altered listening condition. Results showed that participants were able to perceive the spatial difference between the positions of stimulation, thus proving the reliability of our novel setting for the study of spatial hearing in MEG. MEG data highlight a distributed bilateral cortical network involved in active sound localisation, which emerged shortly after stimulus presentation (100—125 ms). The network comprise the classical dorsal auditory pathway plus other cortical regions usually underestimated in previous literature – most notably, regions in the central sulcus/precentral gyrus possibly involved in head movements. Connectivity analysis revealed different patterns of neural coupling, as a function of frequency band. In particular, coherence in high gamma revealed significant connections involving the parietal cortex and the posterior superior temporal cortex. In the final chapter (Chapter 5), I summarise the main findings of the three studies, discuss their implications and outline potential future directions.

No presente modelo em NetLogo, implementou-se um código onde patches genotipicamente homogêneos, reproduzem-se no mapa composto de 64 x 64 células. Buscam parceiros entre si, seguindo algumas orientações. O par reprodutivo deve estar dentro de uma determinada distância genética (G) e espacial (S). Estes parâmetros definem qual a máxima divergência genotípica permitida para a reprodução (G) e qual a distância espacial máxima entre dois possíveis parceiros reprodutivos (S). Além destes, o sliderM determina a probabilidade de ocorrer mutação nos genótipos resultantes das reproduções e A a amplitude, i.e., a quantidade de mudança sofrida pelo genótipo do agente. A princípio, geneticamente homogêneos, todos os indivíduos podem potencialmente formar pares. Contudo, com ocorrência de trocas genéticas e mutações, na formação da prole, aumenta-se a diversidade genética e há isolamento reprodutivo entre indivíduos. Obteu-se especiação dos agentes, ocorrência de corredor de fluxo gênico e mapa robusto de combinação de parâmetros.
In the present model in NetLogo, we implemented a code where genotypically homogeneous patches, reproduce in a map consisting of 64 x 64 cells. Seek partners among themselves by following some guidelines. The breeding pair must be within a certain genetic (G) and spatial (S) distance. These parameters define the maximum genotypic divergence which allowed for reproduction (G) and that maximum spatial distance between two potential reproductive partners (S). In addition, the slider M determines the probability of mutation in resulting genotypes and A the amplitude, i.e., the amount of change experienced by the genotype of the agent. Primarily, genetically homogeneous, all individuals can potentially form pairs. However, with the occurrence of genetic changes and mutations in the offspring formation, the genetic diversity increases and there is reproductive isolation between individuals. There were agents speciation, occurrence of genic flow pathway and robust map of matching parameters.

Dans le domaine de la biologie développementale, un des principaux défis est de comprendre comment des tissus multicellulaires, à l'origine indifférenciés, peuvent engendrer des formes aussi complexes que celles d'une fleur. De part son implication dans l'organogenèse florale, l'auxine est une phytohormone majeure. Nous avons donc déterminé son réseau binaire potentiel, puis y avons appliqué des modèles de clustering de graphes s'appuyant sur les profils de connexion présentés par ces 52 facteurs de transcription (FT). Nous avons ainsi pu identifier trois groupes, proches des groupes biologiques putatifs: les facteurs de réponse à l'auxine activateurs (ARF+), répresseurs (ARF-) et les Aux/IAAs. Nous avons détecté l'auto-interaction des ARF+ et des Aux/IAA, ainsi que leur interaction, alors que les ARF- en présentent un nombre restreint. Ainsi, nous proposons un mode de compétition auxine indépendent entre ARF+ et ARF- pour la régulation transcriptionelle. Deuxièmement, nous avons modélisé l'influence des séquences de dimérisation des FT sur la structure de l'interactome en utilisant des modèles de mélange Gaussien pour graphes aléatoires. Les groupes obtenus sont proches des précédents, et les paramètres estimés nous on conduit à conclure que chaque sous-domaine peut jouer un rôle différent en fonction de leur proximité phylogénétique.Enfin, nous sommes passés à l'échelle multi-cellulaire ou, par un graphe spatio-temporel, nous avons modélisé les premiers stades du développement floral d'A. thaliana. Nous avons pu extraire des caractéristiques cellulaires (3D+t) de reconstruction d'imagerie confocale, et avons démontré la possibilité de caractériser l'identité cellulaire en utilisant des méthodes de classification hiérarchique et des arbres de Markov cachés
A striking aspect of flowering plants is that, although they seem to display a great diversity of size and shape, they are made of the same basics constituents, that is the cells. The major challenge is then to understand how multicellular tissues, originally undifferentiated, can give rise to such complex shapes. We first investigated the uncharacterised signalling network of auxin since it is a major phytohormone involved in flower organogenesis.We started by determining the potential binary network, then applied model-based graph clustering methods relying on connectivity profiles. We demonstrated that it could be summarise in three groups, closely related to putative biological groups. The characterisation of the network function was made using ordinary differential equation modelling, which was later confirmed by experimental observations.In a second time, we modelled the influence of the protein dimerisation sequences on the auxin interactome structure using mixture of linear models for random graphs. This model lead us to conclude that these groups behave differently, depending on their dimerisation sequence similarities, and that each dimerisation domains might play different roles.Finally, we changed scale to represent the observed early stages of A. thaliana flower development as a spatio-temporal property graph. Using recent improvements in imaging techniques, we could extract 3D+t cellular features, and demonstrated the possibility of identifying and characterising cellular identity on this basis. In that respect, hierarchical clustering methods and hidden Markov tree have proven successful in grouping cell depending on their feature similarities

Les végétaux, contrairement aux animaux, génèrent la plupart de leurs organes et tissus au cours de leur développement post-embryonnaire et ce, grâce à des tissus contenant de petits amas de cellules souches appelés méristèmes. Le méristème apical caulinaire (MAC), situé à l’extrémité de la tige, génère toute la partie aérienne de la plante. A sa périphérie, les organes latéraux (fleurs ou feuilles) sont générés selon un patron spatio-temporel précis appelé phyllotaxie. De nombreuses données accumulées ces 20 dernières années ont démontré qu’une hormone végétale, l’auxine, joue un rôle prépondérant dans le contrôle du devenir des cellules dans le MAC. Un ensemble de données expérimentales couplées à des modèles mathématiques suggère que l’auxine s’accumule successivement dans les sites d’organogenèse grâce à l’auto-organisation de ses transporteurs membranaires et instruit les cellules à se différencier en organes.Fautes d’outils appropriés, il était impossible jusqu’alors de visualiser l’auxine in vivo et d’étudier sa dynamique temporelle. Nous avons généré un nouveau senseur de la signalisation de l’auxine, appelé DII-Venus, qui permet de visualiser de manière indirecte mais spécifique les niveaux relatifs d’auxine in planta avec une excellente résolution spatio-temporelle. Cet outil a permis de mettre en évidence pour la première fois des oscillations circadiennes d’auxine au niveau du MAC. Une analyse complète de la structure de la voie de réponse transcriptionelle à l’auxine, couplée à des approches de modélisation, a permis de mettre en évidence des propriétés « tampon » de la voie transcriptionnelle qui la rendent relativement insensible aux fluctuations d’auxine, et contribuent à la robustesse du programme organogénétique. En revanche, la voie non-transriptionnelle de réponse à l’auxine, sensible à ces oscillations, génère des rythmicités de croissance au niveau du MAC qui contribuent à déterminer la temporalité de l’émergence de nouveaux organes. Ces résultats démontrent ainsi pour la première fois que la rythmicité de l’émergence de nouveaux organes au niveau du MAC n’est pas uniquement une conséquence des capacités d’auto-organisation du tissu mais est aussi contrôlée, au moins partiellement, par une horloge biologique
Plants, contrarily to animals, are able to generate new organs and tissues throughout their lives thanks to the activity of specialized tissues containing stem cells called meristems. The shoot apical meristem (SAM), located at the shoot tip, generates all the aerial parts of the plant that arise after germination. At its periphery, organ production occurs following precise spatio-temporal patterns also known as phyllotaxis. During the past twenty years, the phytohormone auxin has been demonstrated to play a major role in this process. Indeed, both experimental and theoretical studies strongly suggest that auxin accumulates successively in sites of organogenesis thanks to its efflux carriers, and instructs cells to differentiate into organs.However, so far, very little is known about the actual temporal dynamics of auxin in tissues, because of the lack of appropriate tool to visualize auxin in vivo. We developed a new auxin signaling sensor, called DII-VENUS, that allows for monitoring auxin levels in planta with a good spatio-temporal resolution. Using this new tool, we were able to demonstrate that for the first time that the SAM is subjected to circadian oscillations of auxin levels. Our data suggest that these oscillations are not perceived by the auxin transcriptional pathway, which is predicted, according to our mathematical models, to exhibit buffering properties. However, they are perceived by the non-transcriptional putative receptor ABP1 and translated into rhythmic growth patterns at the SAM. These growth oscillations seem to regulate organ initiation in the meristem thus demonstrating for the first time the rhythmic emergence of organs at the SAM does not only result from the self-organizing properties of the tissue but is also controlled, at least partially, by a biological clock

Dans les organismes multicellulaires, plusieurs millier d’origines initient la réplication de l'ADN. Elles sont regroupées en domaines qui se répliquent tôt ou tard au cours de la phase S (origines précoces ou tardives). L'un des mécanismes régulant le programme de réplication est un point de contrôle intra phase S qui dépend des kinases ATR et Chk1. Cette voie est activée par un stress réplicatif engendré par le blocage des fourches de réplication aux origines précoces, en retour elle inhibe l’activation des origines tardives. Il a été proposé, que la protéine Polo-Like-Kinase 1 (Plk1) soit responsable du déclenchement des origines situées à proximité des fourches bloquées en cas de stress réplicatif. Cependant, aucune analyse de l’activation des origines n’a jamais été réalisée au cours d’une phase S non perturbée lorsque Plk1 est absente. Pour avoir une vue globale et unifiée du processus de réplication de l'ADN, des modèles numériques et analytiques ont été construits dans le passé, mais aucun d'eux n'intègrent le rôle de Chk1 et Plk1. L'objectif de ma thèse était d’étudier expérimentalement et analytiquement de quelle manière Chk1 peut réguler le déclenchement des origines dans l'espace et dans le temps. En particulier, de comprendre si Plk1 pouvait être impliquée dans cette régulation pendant une phase S non perturbée, à cette fin, j'ai utilisé le système réplicatif des extraits d’œuf de Xénopes. En premier lieu, j'ai intégré dans un modèle numérique l’action de Chk1 pour reproduire le programme de réplication du système Xénope. J'ai testé différents scénarios puis j’ai utilisé des données de peignage d'ADN obtenues précédemment dans des conditions d'inhibition de la kinase Chk1. Les simulations Monte Carlo obtenues ont été ajustées aux données expérimentales en optimisant les valeurs des paramètres libres des modèles. J'ai trouvé qu'il fallait ajouter deux hypothèses aux modèles de réplication développés précédemment: 1) la présence d’une forte inhibition du déclenchement des origines par Chk1 à partir du début de la phase S 2) la présence de domaines génomiques répliquant précocement et qui échappent à cette inhibition. Deuxièmement, j'ai montré expérimentalement que, Plk1 actif est recrutée sur la chromatine avant le début de la phase S non perturbée et qu'en l'absence de Plk1, la réplication de l'ADN est ralentie. De plus, l’absence de Plk1 entraîne une augmentation de la phosphorylation de Chk1 et une diminution de l’activité de la kinase Cdk2, ce qui suggère que Plk1 inhibe Chk1. En réalisant des expériences de peignage d’ADN, j'ai démontré qu’en l’absence de Plk1 on observe une baisse du niveau d’activation des origines. L'analyse de ces données par mon modèle numérique suggère que Plk1 régule négativement l’action de Chk1 levant ainsi son action inhibitrice sur l’activation globale des origines. Cet effet concorde avec mes observations expérimentales. Il semble cependant que Plk1 n’agisse pas à proximité directe des fourches de réplication, comme cela avait été proposé précédemment. Enfin, en assimilant le processus de réplication à un processus de nucléation et de croissance unidimensionnel, j'ai développé une nouvelle approche quantitative pour étudier la régulation du programme de réplication. Cette approche lie la similarité entre les profils spatiaux de réplication d'une molécule unique et les processus de régulation de la réplication de l'ADN. En analysant les données de peignage d'ADN, j'ai montré que le programme de réplication de l'ADN des embryons précoces de Xénope est régulé par deux processus exclusifs dans l'espace et dans le temps. L’un avec une fréquence faible d’activation des origines et une vitesse apparente de fourches élevée et un second, régulé par Plk1, présentant une fréquence d’activation élevée des origines avec une vitesse apparente de fourches faible
The initiation of DNA replication in multicellular organisms starts from several thousand genomic loci called replication origins. They are grouped into domains which replicate early or late during S phase. The firing of a replication origin creates two diverging replication forks that replicate flanking DNA. One of the mechanisms regulating DNA replication program is the ATR/Chk1 dependent intra-S phase checkpoint. This pathway is activated by replicative stress due to stalled replication forks at early firing origins and in turn, inhibits the late firing of origins. It has been proposed that the checkpoint recovery kinase Plk1 (Polo-Like-Kinase 1) could be responsible for allowing origin firing close to stalled forks in replication stress conditions. However, origin firing has not been analysed after Plk1 inhibition or depletion during unperturbed S phase. To assemble a comprehensive and unified view of the DNA replication process numerical and analytical models have been built in the past, but none of them integrates the role of checkpoint pathways. The goal of my thesis was to investigate experimentally and analytically how the checkpoint regulates the firing of origins in space and time and, in particular, whether the Plk1 is implicated in the regulation of origin firing during unperturbed S phase. To this end, I used the Xenopus in vitro system. First, I integrated in a numerical model the checkpoint pathway to describe the replication program in the Xenopus in vitro system. I tested different scenarios and used DNA combing data previously obtained by the laboratory after the inhibition of the checkpoint kinase Chk1. Monte Carlo simulated data were fitted to experimental data by optimizing the values of free parameters of models using a genetic algorithm. I found that two new hypothesis should be added to formerly built replication models: 1) a strong inhibition of origin firing by Chk1 from the beginning of S phase 2) the presence of early replicating genomic domains that evade the origin firing inhibition. Second, I experimentally showed that during unperturbed S phase active Plk1 is recruited to chromatin before the start of S phase and that in the absence of Plk1, DNA replication is slowed down. Moreover, Plk1 depletion led to an increase in Chk1 phosphorylation (p-Chk1) and a decrease of Cdk2 activity, suggesting that Plk1 inhibits the intra-S phase checkpoint. Performing DNA combing, I demonstrated that Plk1 depletion leads to a decrease in origin firing level. Analysis of the combing data by the developed numerical model suggested that during unchallenged S phase Plk1 down regulates the global origin firing inhibitory action of Chk1, consistent with the experimental observation of increased level of p-Chk1 in Plk1 depleted Xenopus egg extract. However, Plk1 does not seem to act close to replication forks as was proposed earlier. Finally, by considering replication process as a one-dimensional nucleation and growth process and using statistical methods, I developed a new quantitative approach to study the regulation of replication program. This approach links the similarity between single molecule replication patterns to DNA replication regulating processes. By analyzing DNA combing data, I showed that DNA replication program in Xenopus early embryos is regulated by two spatially and temporally exclusive processes. One with low frequency of origin firing and high apparent fork speed and a second, controlled by PlK1, with a high frequency of origin firing and a low apparent fork speed. Altogether my results demonstrate that Plk1 positively regulates replication origin firing during normal S phase by down regulating the replication checkpoint. The numerical model predicts the existence of replication timing domains in the Xenopus model system. Future work will show whether Plk1 regulates the replication program at the level of genomic domains

Tese de doutoramento, Geografia (Geografia Humana), Universidade de Lisboa, Instituto de Geografia e Ordenamento do Território, 2018
This doctoral thesis explores the notion of migrant spatial integration drawing from qualitative fieldwork carried out with migrants in Lisbon, Portugal. Departing from a critique of the scientific scholarship’s recurrent focus toward either the residential level of migrants’ urban integration or the affective/identity ties linking migrants to new territories, this thesis provides an alternative frame for the study of migrants’ relationships with urban space. It looks at spatial integration in terms of the construction and maintenance of relations of use and knowledge about the city and its resources. From this perspective, to ‘integrate’ urban space has to do with being able to navigate it and about being aware of its resources. The author argues that such a practical definition of spatial integration helps overcoming some of the scholarship’s perceived limitations, such as the residential bias, methodological nationalism, and the normativity often implicit in some integration accounts. At the same time, the proposed approach sheds light on migrants’ practical knowledges about urban space, acknowledging their active role as urban dwellers and their skills to manage life locally. The thesis develops its argument by recourse to two fundamental working-concepts: ‘becoming local’ and spatial apprenticeship. Through the first working-concept, the author invokes the figure of the ‘local’, understood as someone who inhabits a particular city and knows ‘how it works’, and raises the potentialities of thinking about migrants as ‘becoming locals’. By doing that, we are given a window into the practical strategies migrants resort to in order to manipulate the urban form to their own purposes and needs. As for spatial apprenticeship, the second working-concept, the author argues that it captures the diversity of urban expertise accumulated by migrant urbanites resulting from both the embodied and intersectional nature of migrants’ spatial practices and the dynamics of urban change and the specificities of cities. Finally, the thesis sustains that looking at the articulation between learning a city and putting that practical savoir-faire into practice is fundamental if we are to understand the ways migrants have managed the making of urban livelihoods together with the urgency for settlement, finding work, and making personal connections.
Esta tese de doutoramento tem como objetivo explorar os desdobramentos e potencialidades do conceito de integração espacial a partir de uma investigação qualitativa junto a diversos imigrantes em Lisboa, Portugal. Nas últimas décadas, a apreciação crítica dos estudos migratórios com relação à ideia de integração enquanto processo único, genérico e normativo deu lugar à consideração pormenorizada das várias dimensões constitutivas do processo de integração. Dessa forma, a literatura especializada tem investigado, entre outras, a integração de imigrantes no mercado laboral; na vida comunitária; nos sistemas de saúde e de bem-estar social etc. Dentro deste rol dos diferentes aspectos da integração de imigrantes, destaca-se, para efeito desta tese, sua dimensão espacial. Tradicionalmente, a investigação das espacialidades do processo de integração de imigrantes tem se debruçado, sobretudo, na sua esfera residencial. Nessa perspectiva, importantes trabalhos lançaram luz sobre o acesso dos imigrantes à habitação, sobre os padrões residenciais, sobre a concentração/dispersão residencial, entre outros tópicos. Para além dessa perspectiva, a literatura especializada tem também analisado a formação de laços de pertença e de identidade entre os imigrantes e os locais em que se estabeleceram. Grosso modo, e apesar de comprovadas exceções, subsiste a compreensão de que a concentração étnica residencial de imigrantes, bem como a falta de laços de identidade entre imigrantes e o espaço local, apontariam para um problema de integração espacial. Esta tese não busca chegar a uma conclusão a respeito da ideia de integração espacial de imigrantes a partir de seu aspecto residencial ou de pertença. Ao contrário, a investigação conduzida no presente trabalho procura repensar a integração espacial à luz de outro paradigma. O conceito de integração espacial aqui proposto sugere que ‘integrar-se’ ao espaço urbano implica na capacidade de navegar a cidade e de conhecer os seus recursos disponíveis. Em outras palavras, integração espacial é concebida nesta tese como o desenvolvimento e a manutenção de relações de uso e de conhecimento sobre o espaço praticado. A tese demonstra que tal definição possibilita caracterizar o processo de integração de imigrantes no espaço urbano sem reiterar algumas das limitações identificadas na literatura sobre o assunto, como o viés residencial, o nacionalismo metodológico, e mesmo a normatividade implícita na ideia de identidade/pertença. Além disso, defende-se que essa definição de integração espacial faz vir à tona os diversos conhecimentos práticos dos imigrantes sobre a cidade, uma vez que se reconhece seu papel ativo como residentes citadinos e suas estratégias para conduzir suas vidas e responder às necessidades que envolvem o uso de recursos urbanos. A tese explora os potenciais e as limitações da definição proposta de integração espacial por meio de dois conceitos fundamentais: ‘becoming local’ (‘tornando-se um local’) e spatial apprenticeship (aprendizagem espacial). No artigo intitulado Using the city: migrant spatial integration as urban practice (‘Usando a cidade: integração espacial de imigrantes e prática urbana’), a figura do ‘local’, entendido como aquele que habita numa determinada cidade e que entende intimamente seu funcionamento, é invocada como ferramenta para pensar o processo pelo qual imigrantes tornam-se ‘locais’. O artigo sustenta que a figura do ‘local’ é constituída essencialmente por experiência e conhecimentos práticos sobre a cidade, seus recursos, e como os utilizar. Pensar os imigrantes como ‘becoming locals’ é, nesse sentido, buscar compreender como são construídos os conhecimentos práticos sobre a cidade, pelos imigrantes, e como esses lançam mão desses conhecimentos em seus afazeres cotidianos. O artigo é construído com base em contributos de diversos imigrantes em Lisboa, que ilustram a multiplicidade do ‘ser-local’ e expõem diversos fatores que moldam suas relações com o espaço da cidade. O segundo conceito-chave está desenvolvido com mais detalhe no segundo artigo que compõe a tese. Em A user’s guide to Lisbon: mobilities, spatial apprenticeship and migrant urban integration (‘Um guia de utilizador de Lisboa: mobilidades, aprendizagem espacial e integração espacial de imigrantes’), a noção de spatial apprenticeship (aprendizagem espacial) é introduzida para descrever a complexidade da interação entre a materialidade do espaço urbano e a corporalidade das práticas descritas pelos imigrantes na cidade. A própria cidade, em sua materialidade, assimetria, distribuição e fraturas, revela, por meio do conceito de spatial apprenticeship, seu papel também ativo na cristalização dos diversos modos de uso do espaço urbano. Apoiando-se na contribuição da literatura sobre mobilidades, este artigo aponta para o aspecto aprendido da mobilidade urbana e para o valor desta aprendizagem enquanto reveladora de diferentes características, necessidades, posicionamentos e valores apresentados pelos imigrantes-utilizadores da cidade. O último artigo que integra esta tese chama-se Navigating urban life in Lisbon: a study of migrants’ mobilities and use of space (‘Navegando a vida urbana lisboeta: um estudo das mobilidades de imigrantes e de seu uso do espaço’). Neste texto, a ideia de integração espacial é vista de maneira mais abrangente, na sua intersecção com temáticas ligadas à mobilidade, ao bem-estar, e à participação na vida urbana. A definição de integração espacial é aqui aprofundada e sublinha o processo através do qual indivíduos imigrantes passam a ver como seus os recursos urbanos, isto é, reduzem o distanciamento prático-subjetivo entre si e a materialidade urbana. O artigo ilustra este processo a partir de diversos exemplos em que o uso da cidade responde às necessidades materiais do cotidiano e revela, analiticamente, geografias urbanas de exclusão, de racismo, ou de solidariedade e de fé. O material empírico principal* desta investigação foi produzido entre 2015 e 2016 em Lisboa. Dado o caráter exploratório e qualitativo do projeto, os participantes foram recrutados segundo uma lógica de amostragem intencional, de maneira a garantir a diversidade de perfis, seja em termos de nacionalidade (15 nacionalidades), status socioeconômico, gênero (11 mulheres; 14 homens), período de permanência em Portugal, religião, ou bairro de residência, etc. No total, 25 imigrantes contribuíram neste projeto, dos quais 11 podem ser classificados de imigrantes pós-coloniais (vindo de Angola, Brasil, Cabo Verde, Guiné Bissau, Moçambique, e São Tomé e Príncipe). A participação decorreu sempre de maneira voluntária e a partir de três métodos distintos: entrevistas em profundidade (cuja duração variou entre 1 e 2h), representações gráficas dos espaços frequentados feitas pelos participantes (mapas mentais), e escrita de um diário (time-space journal) onde os participantes detalharam os lugares frequentados ao longo de alguns dias (2 a 5 dias) e suas relações com os mesmos. Quando necessárias, entrevistas follow-up foram também realizadas. A conclusão da tese elenca as potencialidades e limitações da abordagem proposta, bem como situa-a no contexto da literatura recente sobre integração de imigrantes. Finalmente, são apontados alguns caminhos de investigação que podem beneficiar-se do conceito de integração espacial tal qual é aqui definido.

博士
國立清華大學
生命科學系
94
The endoplasmic reticulum (ER) is a multifunctional organelle controlling important cellular processes, including Ca2+ homeostasis, protein synthesis, protein trafficking, and apoptosis. Under physiologically or pharmacologically adverse conditions that perturb the calcium homeostasis, accumulation of unfolded or malfolded proteins in the ER lumen occurs, referred to as ER stress, and the cells will activate a series of signal transduction cascades collectively termed the unfolded protein response (UPR). One characteristic of the UPR is the induction of the ER resident stress proteins referred to as the glucose-regulated proteins (GRPs). The best characterized GRP78, also known as the immunoglobulin heavy chain binding protein or BiP, is thought to function in Ca2+ sequestration or as a molecular chaperone in the folding and assembly of membrane or secreted proteins. Previous reports of GRP78 in different cellular compartments prompted us to examine and compare the changes of GRP78 in intracellular distribution patterns in response to ER stress, specifically under calcium disturbance. Treatment with calcium ionophore A23187 and sarcoplasmic/endoplasmic reticulum Ca2+ ATPase inhibitor thapsigargin (TG) results in a decrease of [Ca2+]er with a concurrent increase of [Ca2+]c. The immunostaining of GRP78 coupled with confocal microscopy demonstrated the granular and perinuclear expression as normal ER distribution in cells under normal growing conditions. When the cells were exposed to A23187 or TG, the greater proportion of GRP78 displayed a diffused distribution throughout the cytoplasm at a slightly higher intensity. Overlapping of GRP78 and mitochondria marker apparently depicted worm-shaped strings, suggesting that there is an increase in the level of colocalization and that this might occur from an increase in the level of targeting. Cellular fractionation and protease digestion of isolated mitochondria from ER-stressed cells suggested that a significant portion of GRP78 is localized to the mitochondria and is protease-resistant. Localizations of GRP78 in ER and mitochondria were confirmed by immunoelectron microscopy. In ER-stressed cells, GRP78 mainly localized within the mitochondria and decorated the mitochondrial membrane compartment. Submitochondrial fractionation studies further indicated that the mitochondrial resided GRP78 is mainly located in the intermembrane space, inner membrane, and matrix. Furthermore, radioactive labeling followed by subcellular fractionation showed that a significant portion of the newly synthesized GRP78 is localized to the mitochondria in cells under UPR. The results in this study indicate that, at least under certain circumstances, the ER resided chaperone GRP78 can be retargeted to mitochondria and thereby may be involved in correlating UPR signaling between these two organelles. We also investigate the signaling pathway involved in the induction of GRP78 in cells under the treatment of geldanamycin (GA), which is a potent inducer of ER stress response. By using calcium monitoring, inhibitors screening and direct examination of the ROS contents rendered by GA, we show that GA exerts GRP78 inductive expression through a causative pathways connecting phospholipase C to intracellular calcium increase, PKC activation as well as ROS generation. Furthermore, the calcium influx from extracellular space, intracellular calcium store oscillations and mitochondrial calcium influx are important for the calcium mobilization and GRP78 expression induced by GA. The GA-induced GRP78 expression signaling cascades may represent regulation of the ER-stress response in cell survival or apoptotic program.

Bistable arches have two force-free stable equilibrium configurations. They also show multimodality by switching between their stable states in multiple deformation pathways. These two attributes and their nonlinear force-displacement characteristic are desirable in a range of engineering applications. The analytical and semi-analytical methods developed in this work enable faster analysis than finite element analysis and also facilitate closed-form relationships for insightful design of arches. We show that arch profiles composed using the basis set of buckling mode shapes of the straight column with the corresponding boundary conditions exhibit bistability. We analyze such arches by expressing their deformed profiles also in the same basis set. We assume that the arches are slender and shallow to derive their potential energy comprising bending and compression strain energies as well as the work potential. We solve the equilibrium equations obtained by minimizing potential energy using a semi-analytical method for analytically intractable general boundary conditions. In this method, we obtain the critical points on the force-displacement curve corresponding to switching and switch back forces and travel of the mid-point of the arch. We use this method to analyze and optimize arches of varying as-fabricated stress-free shapes and boundary conditions. We obtain an analytical relationship between the arch-profiles in the force-free states of the arch by equating the force to zero in the aforementioned equilibrium equations for both fixed-fixed and pinned-pinned boundary conditions. This relationship is bilateral, i.e., in one form it can be a used for analysis and in another for design. We derive necessary and sufficient conditions as well as corollaries from the bilateral relationship pertaining to the shapes of bistable arches. Deformation pathways in bistable arches can also be three-dimensional . These spatial deformation pathways can help reduce the switching and switch-back forces and might also, at times, adversely affect bistability. We model spatial pathways by incorporating additional energy terms due to out-of-plane bending and torsion into analysis of planar arches. We use a geometric relation by St. Venant and Michell to capture the coupling amongst the in-plane and out-of-plane deformations and rotation of the cross-sections. Furthermore, we show that non-planar arches, i.e., spatial arches, can be bistable too. Our analysis is extended to spatial arches by modifying the geometric relation to consider the additional out-of-plane curvature. We also present design of two applications based on bistable arches: an RF-MEMS switch and a mechanical OR gate. RF-MEMS switch utilizes bimodality and a novel initially-retracting electrothermal actuator to realize ON and OFF states with only two electrical terminals. The mechanical OR gate uses the bilateral relationship to design arch-profiles that achieve the OR gate logic. Additionally, we present two studies on bistability in axisymmetric shallow thin shells. In the first study, we optimize the shell-profile for maximum travel by numerical and semi-analytical approaches and compare the results with the shell obtained by revolving the optimal arch for maximum travel. In the second study, we discuss the design of a passive universal gripper based on a bistable shell that can grasp objects of varying shape.

To survive, many biological organisms need to accurately infer which features of their environment predict future rewards and punishments. In machine learning terms, this is the problem of spatial credit assignment, for which many supervised learning algorithms have been developed. In this thesis, I mainly propose that a dual-pathway, regression-like strategy and associated biological implementations may be used to solve this problem. Using David Marr's (1982) three-level philosophy of computational neuroscience, the thesis and its contributions are organized as follows: - Computational Level: Here, the spatial credit assignment problem is formally defined and modeled using probability density functions. The specific challenges of the problem faced by organisms and machine learning algorithms alike are also identified. - Algorithmic Level: I present and evaluate the novel hypothesis that the general strategy used by animals is to perform a regression over past experiences. I also introduce an extension of a probabilistic model for regression that substantially improves generalization without resorting to regularization. This approach subdues residual associations to irrelevant features, as does regularization. - Physical Level: Here, the neuroscience of classical conditioning and of the basal ganglia is briefly reviewed. Then, two novel models of the basal ganglia are put forward: 1) an online-learning model that supports the regression hypothesis and 2) a biological implementation of the probabilistic model previously introduced. Finally, we compare these models to others in the literature. In short, this thesis establishes a theoretical framework for studying the spatial credit assignment problem, offers a simple hypothesis for how biological systems solve it, and implements basal ganglia-based algorithms in support. The thesis brings to light novel approaches for machine learning and several explanations for biological structures and classical conditioning phenomena.
Note: While the thesis contains content from two articles (one journal, one conference), their publishers do not require special permission for their use in dissertations (information confirming this is in an appendix of the thesis itself).

Bone Morphogenetic Proteins (BMPs) are key regulators for a lot of diverse cellular processes. During embryonic development these proteins act as morphogens and play a crucial role particularly in organogenesis. BMPs have a direct impact on distinct cellular fates by means of concentration-gradients in the developing embryos. Using the diverse signaling input information within the embryo due to the gradient, the cells transduce the varying extracellular information into distinct gene expression profiles and cell fate decisions. Furthermore, BMP proteins bear important functions in adult organisms like tissue homeostasis or regeneration. In contrast to TGF-ß signaling, currently only little is known about how cells decode and quantify incoming BMP signals. There is poor knowledge about the quantitative relationships between signal input, transducing molecules, their states and location, and finally their ability to incorporate graded systemic inputs and produce qualitative responses. A key requirement for efficient pathway modulation is the complete comprehension of this signaling network on a quantitative level as the BMP signaling pathway, just like many other signaling pathways, is a major target for medicative interference. I therefore at first studied the subcellular distribution of Smad1, which is the main signal transducing protein of the BMP signaling pathway, in a quantitative manner and in response to various types and levels of stimuli in murine c2c12 cells. Results indicate that the subcellular localization of Smad1 is not dependent on the initial BMP input. Surprisingly, only the phospho-Smad1 level is proportionally associated to ligand concentration. Furthermore, the activated transducer proteins were entirely located in the nucleus. Besides the subcellular localization of Smad1, I have analyzed the gene expression profile induced by BMP signaling. Therefore, I examined two endogenous immediate early BMP targets as well as the expression of the stably transgenic Gaussia Luciferase. Interestingly, the results of these independent experimental setups and read-outs suggest oscillating target gene expression. The amplitudes of the oscillations showed a precise concentration-dependence for continuous and transient stimulation. Additionally, even short-time stimulation of 15’ activates oscillating gene-expression pulses that are detectable for at least 30h post-stimulation. Only treatment with a BMP type I receptor kinase inhibitor leads to the complete abolishment of the target gene expression. This indicated that target gene expression oscillations depend directly on BMP type I receptor kinase activity
Bone Morphogenetic Proteins (BMPs) stellen wichtige Regulatoren für eine Vielzahl von verschiedenen zellulären Prozessen dar. Während der Embryonalentwicklung agieren diese Proteine als Morphogene und spielen daher eine entscheidende Rolle für diesen Prozess, vor allem in der Organogenese. Durch Konzentrationsgradienten üben BMPs einen direkten Einfluss auf verschiedene zelluläre Schicksale im entwickelnden Embryo aus. Aufgrund dieser Gradienten gelangen vielfältige Signalinformationen zu den verschiedenen Zellen, welche die extrazelluläre Information in verschiedene Genexpressionsprofile und Zellschicksalsentscheidungen umwandeln. Darüber hinaus tragen BMPs wichtige Funktionen im erwachsenen Organismus, wie z.B. Gewebshomöostase oder -regeneration. Im Gegensatz zu dem verwandten TGF-ß Signaltransduktionsweg ist derzeit nur wenig über die zelluläre Übersetzung und Quantifizierung eingehender BMP-Signale bekannt. Es gibt wenige Kenntnisse über die quantitative Beziehung zwischen Signaleingang, Überträgerproteinen, ihren Zuständen sowie intrazellulären Positionen, und schließlich ihre Fähigkeit Signaleingänge systemisch zu integrieren und qualitative Antworten der Zelle zu produzieren. Eine wesentliche Voraussetzung für die effiziente Signaltransduktions-modulierung ist das vollständige Verständnis des Signalnetzwerkes auf einer quantitativen Ebene, da der BMP-Signalweg, wie auch viele andere Signalwege, ein wichtiges Ziel für medizinische Anwendungen und Medikamentenentwicklung ist. Daher untersuchte ich zunächst die subzelluläre Verteilung der wichtigsten Signalweiterleitungsproteine des BMP-Signalweges, der Smad1-Proteine, auf quantitativer Ebene und deren Reaktion auf verschiedene Stimulierungsarten und BMP-Konzentrationsstufen in murinen c2c12-Zellen. Die Ergebnisse zeigen, dass die subzelluläre Lokalisation von Smad1 unabhängig von der BMP-Konzentration ist und nur das phospho-Smad1 Level proportional zur Konzentration des Liganden steigt. Darüber hinaus befanden sich die aktiven Überträgerproteine nach Stimulierungvollständig im Zellkern. Neben der subzellulären Lokalisation von Smad1, habe ich das Genexpressionsprofil von BMP-Zielgenen analysiert. Ich untersuchte zwei endogene und frühe BMP-Zielgene sowie die Expression der stabil transgenen Gaussia Luciferase. Interessanterweise deuten die Ergebnisse dieser zwei unabhängigen Versuchsaufbauten und Detektionsmethoden auf eine oszillierende Expression der Zielgene hin. Die Amplituden der Schwingungen zeigten eine deutliche Konzentrationsabhängigkeit bei kontinuierlicher und transienter Stimulation. Außerdem aktiviert eine Kurzzeitstimulierung von 15 Minuten ebenfalls ein oszillierendes Genexpressionsprofil, welches für mindestens 30 Stunden nach der Stimulierung nachweisbar ist. Nur die Behandlung mit einem BMP Typ-I-Rezeptorkinaseinhibitor führt zur vollständigen Aufhebung der Zielgenexpression. Infolgedessen sind die Oszillationen der Zielgenexpression direkt von der Aktivität der BMP Typ-I-Rezeptorkinase abhängig

The Kerguelen Plateau is a biological hotspot and an important region for Southern Ocean fisheries. However, its remoteness and harsh environmental conditions make consistent sampling difficult. Therefore, significant knowledge gaps remain regarding regional-scale impacts of fishing and climate change. Very little is understood about the transfer of energy from prey to predators on the Kerguelen Plateau and even less is understood about how the food web is being impacted by fishing pressure and will be impacted by future climate change. Understanding the combined impacts of bottom-up and top-down processes on food web dynamics can help inform ecosystem-based management on the Kerguelen Plateau. The research in this thesis compiles current knowledge of the Kerguelen Plateau food web and uses Ecopath with Ecosim to explore ecosystem dynamics and identify the potential impacts of climate change and fishing over time and space. Previously published work to develop the first food web model with a balanced energy system for the Kerguelen Islands. This model forms the basis for the research in this thesis. Next, the entire plateau food web was incorporated by extending the domain of the model to include not only the waters surrounding the Kerguelen Islands but also the region further south surrounding Heard and McDonald Islands. This model was used to undertake a thorough exploration of food web structure and function, including answering questions regarding the transfer of energy through the food web and interactions between the fishery and the food web. Results from this work indicate that killer whales, cephalopods (squid) and myctophids (lanternfish) are key pathways for the transfer of energy from lower trophic levels to top predators while fishing activity (in the form of catch and by-catch) had little impact on overall food web dynamics. This work provides the first quantitative description of the food web on the Kerguelen Plateau. The Ecopath model was calibrated using time-series data, and the resulting Ecosim model was applied to explore climate and fishing impacts on the Kerguelen Plateau food web over the last 32 years (1986–2018). Model results again showed that fishing (fishing mortality) was not a driver of food web dynamics. Instead, analyses identified sea surface temperature, zonal wind speed and the Southern Annular Mode as drivers of biomass trends in the food web. Additionally, results indicate that Patagonian toothfish recruitment may be sensitive to surface warming. These results highlight the importance of considering environmental change in ecosystem-based management. Spatial differences in environmental processes, fishing pressure and marine protected areas (MPAs) on the Kerguelen Plateau could have local-scale impacts on the food web. The Ecosim model was spatially resolved by incorporating habitat maps in Ecospace. Food web dynamics were compared with fishing and without fishing present to evaluate spatial fishing impacts and food web distribution was evaluated in relation to the current locations of MPAs. Observed spatial distributions were replicated in the model and findings showed limited direct impacts of fishing on Patagonian toothfish and the food web. Modelled distributions showed that groups in the food web were well distributed within MPAs, indicating that current ecosystem-based management is representative of the food web. This thesis presents the first comprehensive analysis of food web dynamics on the Kerguelen Plateau. Results from this thesis identified key energy pathways, potential environmental drivers and evaluated the degree of protection provided by MPAs, all of which are important for conservation management and predicting Southern Ocean ecosystem response to change in the future. Findings suggest that current fishing for Patagonian toothfish is operating at sustainable levels and spatial management is representative of the food web. This work provides an important foundation for considering future ecosystem-based approaches, especially those that include climate change.

Estimating the risk associated with species and pathogen movements involves considerable uncertainty. One key uncertainty concerns the extent and frequency of human-mediated species and pathogen movements relative to the distribution of recipient ecosystems. Baitfish use in Ontario, Canada is one of many pathways with the potential to introduce and spread biota to beyond their current geographic range. To determine the biological risk associated with baitfish use, models were used to estimate the probability of species occurrences throughout pathway levels, from the commercial harvest level, to retail tank and angler purchases, to movement and release by the end-user (i.e., the angler). Vector activity, as the primary contributor of species movements and introductions associated with this pathway, was modeled within a spatial interaction framework that incorporated landscape structure (e.g., the distribution of angling populations, lake size and sportfish richness, and their physical separation via least-cost routing of transportation networks) to predict the extent of movement. Agent-based models of vector activity were used to relate vector movements to region-specific probability thresholds of risk activity. Model outputs were used to estimate the movement and introduction of species and pathogens to lake ecosystems resulting from a variety of infection scenarios. Species results identified a pronounced reduction in the probability of non-target species occurrences throughout pathway levels. However, the occurrence of biological invaders and other non-target fishes at retail levels implied incidental bycatch throughout the pathway. Relatively short, frequent vector movements associated with incidentally purchased species were very likely, yet would not contribute to species range expansions due to the homogeneity of biological communities at those levels. However, rarer, yet considerably lengthier, vector movements associated with key species and pathogens implied the potential for low-probability, long-distance species and pathogen movements resulting from human activities.

Intra-archipelago waterways, including tidal strait networks, present a complex set of barriers to, and conduits for sediment transport between marine basins. Tidal straits may also be the least well understood tide-dominated sedimentary environment. To address these issues, currents, sediment transport pathways, and seabed sedimentology & geomorphology were studied in the central Salish Sea (Gulf and San Juan Islands region) of British Columbia, Canada and Washington State, USA. A variety of data types were integrated: 3D & 2D tidal models, multibeam bathymetry & backscatter, seabed video, grab samples, cores and seismic reflection. This dissertation included the first regional sediment transport modelling study of the central Salish Sea. Lagrangian particle dispersal simulations were driven by 2D tidal hydrodynamics (~59-days). It was found that flood-tide dominance through narrow intra-archipelago connecting straits resulted in the transfer of sediment into the inland Strait of Georgia, an apparent sediment sink. The formative/maintenance processes at a variety of seabed landforms, including a banner bank with giant dunes, were explained with modelled tides and sediment transport. Deglacial history and modern lateral sedimentological and morphological transitions were also considered. Based on this modern environment, adjustments to the tidal strait facies model were identified. In addition, erosion and deposition patterns across the banner bank (dune complex) were monitored with 8-repeat multibeam sonar surveys (~10 years). With these data, spatially variable bathymetric change detection techniques were explored: A) a cell-by-cell probabilistic depth uncertainty-based threshold (t-test); and B) coherent clusters of change pixels identified with the local Moran's Ii spatial autocorrelation statistic. Uncertainty about volumetric change is a considerable challenge in seabed change research, compared to terrestrial studies. Consideration of volumetric change confidence intervals tempers interpretations and communicates metadata. Techniques A & B may both be used to restrict volumetric change calculations in area, to exclude low relative bathymetric change signal areas.
Graduate
2018-12-07

Signal transduction is a mode of cellular communication essential for an organism’s sustenance and survival. Cell communication in higher organisms is largely executed by two classes of cell surface receptors i.e. G-protein coupled Receptors (GPCRs) and receptor tyrosine kinases (RTKs). Though the activation of each of these receptors have shown to regulate cellular responses through a linear pathway the underlying signaling events are much more complicated. A number of studies have shown that the signaling regulators of the RTK mediated pathway can be cross-activated by GPCRs activation via a process termed as ‘transactivation.’ However, these studies are limited, as they have largely missed the spatio-temporal dynamics of the signaling regulators involved in the transactivation pathway. With this premise, the present research was structured to investigate the spatiotemporal dynamics of MAPK cascade transactivated by GPCR signalling pathways in live cells. Towards this broad objective, we subdivided the study into three part