Proton fluxes through plasma membranes are essential for regulating intracellular and extracellular pH and mediating co-transport of metabolites and ions. Although conventional electrical measurements are highly sensitive and precise for proton current detection, they provide limited specificity and spatial information. My thesis focuses on developing optical approaches to visualize proton fluxes from ion channels and transporters. It has been demonstrated that channel-mediated acid extrusion causes proton depletion at the inner surface of the plasma membrane. Yet, proton dynamics at the extracellular microenvironment are still unclear. In Chapter II, we developed an optical approach to directly measure pH change in this nanodomain by covalently attaching small-molecule, fluorescent proton sensors to the cell’s glycocalyx using glyco-engineering and copper free ‘click’ chemistry. The extracellularly facing sensors enable real-time detection of proton accumulation and depletion at the plasma membrane, providing an indirect readout of channel and transporter activity that correlated with whole-cell proton current. Moreover, the proton wavefront emanating from one cell was readily visible as it crossed over nearby cells. The transport of monocarboxylates, such as lactate and pyruvate is critical for energy metabolism and is mainly mediated by proton-coupled monocarboxylate transporters (MCT1-MCT4). Although pH electrodes and intracellular fluorescent pH sensors have been widely used for measuring the transport of proton-coupled MCTs, they are unable to monitor the subcellular activities and may underestimate the transport rate due to cell’s volume and intracellular buffering. In Chapter III, we used the Chapter II approach to visualize proton-coupled transport by MCT1-transfected HEK293T cells and observed proton depletion followed by a recovery upon extracellular perfusion of L-lactate or pyruvate. In addition, we identified a putative MCT, CG11665/Hrm that is essential for autophagy during cell death in Drosophila. The results demonstrate that Hrm is a bona fide proton-coupled monocarboxylate transporter that transports pyruvate faster than lactate. Although the approach developed in Chapter II enables visualization of proton fluxes from ion channels and transporters, it’s not applicable in some cell types which cannot incorporate unnatural sialic acid precursors into their glycocalyx, such as INS-1 cells and cardiomyocytes. To address this, in Chapter IV we developed a pH-sensitive, fluorescent WGA conjugate, WGA-pHRho that binds to endogenous glycocalyx. Compared to the results in Chapter II and III, cell surface-attached WGA-pHRho has similar fluorescent signals in response to proton fluxes from proton channel Hv1, omega mutant Shaker-IR R362H and MCT1. With WGA-pHRho, we were able to label the plasma membrane of INS-cells and cardiomyocytes and visualized the transport activity of MCT1 in these cells. Taken together, these findings provide news insights into proton dynamics at the extracellular environment and provide new optical tools to visualize proton fluxes from ion channels and transporters. Moreover, the modularity of the approaches makes them adaptable to study any transport events at the plasma membrane in cells, tissues, and organisms.

L'objectif initial de cette thèse est d'apporter une solution à ce problème en proposant, premièrement, une approche cognitive basée sur le paradigme appelé Belief-Desire-Intention (BDI) pour représenter les processus de prise de décision des acteurs humains, et deuxièmement, une validation de cette approche dans le contexte d'un modèle complet de changement d'usage des sols dans lequel la plupart des facteurs cités ci-dessus sont également simulés. Le résultat de ce travail est une approche générique qui a été validée sur un modèle intégrant le changement d'usage des sols d'une région située dans le Delta du Mékong au Vietnam. Nos contributions principales sont les suivantes : Intégration d'une architecture BDI au sein d'une plateforme de modélisation à base d'agents (GAMA) ; Conception d'un cadre générique baptisé " Multi-Agent Based Land-Use Change " (MAB-LUC) permettant de modéliser et de simuler les changements d'usage des sols en prenant en compte les décisions des agriculteurs ; Proposition d'une solution permettant d'intégrer et d'évaluer les facteurs socio-économiques et environnementaux dans le cadre de la planification agraire et d'intégrer MAB-LUC dans le processus existant proposé par la FAO. Ce travail, au-delà du cas d'étude concernant le Delta du Mékong, a enfin été conçu de façon générique afin que la méthodologie utilisée puisse être généralisée à la modélisation de systèmes socio-écologiques où les facteurs humains doivent être représentés avec précision. / Integrated modeling approaches (multi-simulation, multimodeling, etc.) have proven challenging in practice. The first challenge deals with the technical aspects of coupling different computational or mathematical components. The second challenge lies in the alignment of the semantics of these components so that their integration does make sense, which is particularly critical in pluridisciplinary models. A number of approaches have been proposed in the last 20 years but none of them is really suitable to our context . We propose in this thesis an alternate approach, called co-modeling, which borrows concepts and tools from agent-based modeling, agent-oriented software engineering and multimodel ecologies. Simply speaking, a co-model can be defined as a multi-agent system of models and datasets. Each model or dataset is represented by one or several agents interacting with one another within the context of a larger representation of their — potentially dynamic — environment. The proposed approach does not aim at providing a general solution to the two challenges above, but at providing a framework in which modelers can easily implement their solution or test different coupling solutions. The proposed approach is fully implemented within the GAMA agent-based modeling platform. Its advantages are shown in terms of flexibility, composability and reusability in a number of case studies. The first case study is the dynamic coupling of equation-based and agent-based models to obtain “switching” models dynamically. The second one is the design of a complex integrated model where three formalisms and four modeling approaches have been successfully coupled.

This thesis presents the characterization process of an imaging device for a satellite. The camera system was built by the Space Dynamics Laboratory (SDL) and will be used in the Michelson Interferometer for Global High-resolution Thermospheric Imaging (MIGHTI) instrument for National Aeronautics and Space Administration's (NASA) Ionospheric Con- nection Explorer (ICON) satellite. This mission will further scientists' understanding of the connection between the Earth's weather and ionospheric conditions. The ionosphere, a part of the atmosphere, interferes with satellite communications, causing disturbances and disruptions. By learning more about the ionosphere through the data collected by this instrument, scientists will better understand its effects on our communications.

Accurate energetic modeling of large molecular systems is always desired by chemists. For example, ligand-protein binding simulations and enzymatic catalysis studies all involve with a small energy difference. The energetic accuracy depends largely on a proper handling of electronic correlations. Molecular mechanics (MM) methods deliver a parameterized Newtonian treatment to these problems. They show great capability in handling large calculations but give only qualitatively good results. Quantum mechanics (QM) methods solve Schrödinger equations and exhibit much better energy accuracy, though the computational cost can be prohibitive if directly applied to very large systems. Fragment-based methods have been developed to decompose large QM calculations into fragment calculations. However, most current schemes use a self- consistent field (SCF) method on fragments, in which no electronic correlation is accounted for. The super-system energy is computed as a sum of fragment energies plus two-body corrections and, possibly, three-body corrections (a "body" is a fragment). Higher order corrections can be added. Nevertheless, many problems require the treatment of high order electronic correlations. The coupled-cluster (CC) theory is the state-of-the-art QM method for handling electronic correlations. The CC wavefunction contains correlated excitations up to a given truncated level and coincidental excitations for all possible electronic excitations. It is a brilliant way of including more electronic correlations while maintaining a low-order scaling. In the proposed excitonic coupled-cluster (X-CC) theory, substantial modifications have been made to allow CC algorithms to act on the collective coordinates of fragment fluctuations to obtain super-system energy. The X-CC theory is designed to achieve accurate energetic modeling results for large chemical systems with much improved affordability and systematic improvability. The test system used in this work is a chain of beryllium atoms. A 30-fragment X-CCSD(2) calculation delivered matching accuracy with traditional CCSD method. An X-CCSD(2) calculation on a chain of 100 bonded fragments finished in 7 hours on a single 2.2 GHz CPU core. The X-CC scheme also demonstrates the ability in handling charge transfer problems. Due to the use of fluctuation basis in the test cases, the excitonic algorithms can be easily generalized to inhomogeneous systems. This will be investigated in future work.

京都大学 / 0048 / 新制・課程博士 / 博士(工学) / 甲第18933号 / 工博第3975号 / 新制||工||1612(附属図書館) / 31884 / 京都大学大学院工学研究科社会基盤工学専攻 / (主査)教授 木村 亮, 教授 勝見 武, 准教授 木元 小百合 / 学位規則第4条第1項該当 / Doctor of Philosophy (Engineering) / Kyoto University / DFAM

京都大学 / 0048 / 新制・課程博士 / 博士(農学) / 甲第19048号 / 農博第2126号 / 新制||農||1032(附属図書館) / 学位論文||H27||N4930(農学部図書室) / 31999 / 京都大学大学院農学研究科応用生命科学専攻 / (主査)教授 植田 充美, 教授 植田 和光, 教授 小川 順 / 学位規則第4条第1項該当 / Doctor of Agricultural Science / Kyoto University / DFAM

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The electrochemical behavior of nickel (Ni) in different concentrations of sulfuric acid (H2SO4) was studied via cyclic voltammetry (CV) over a range of potentials (0.0 V– 3.0 V) at room temperature. The presented work displays novel experiments where external forcing by a platinum (Pt) electrode changed the proton concentration at a Ni electrode surface in order to control the frequency and magnitude of periodic oscillations produced. When studying unique phenomena such as the Ni phenomena in this thesis, efficient, durable, and inexpensive technology is always beneficial. A coupled microelectrode array sensor or CMAS which has been used for over four decades to study pitting corrosion, crevice corrosion, intergranular corrosion, galvanic corrosion, and other heterogeneous electrochemical processes were fabricated in a novel, systematic, inexpensive, and time efficient process. The presented work shows how to make the CMAS and proved that they functioned properly.

Cells dedicate tremendous amounts of energy to express essential genes for survival. During transcription, RNA polymerase (RNAP) actively scans the template strand of DNA, stalling when it meets DNA damages. Stalled RNAP prevents repair by the nucleotide excision repair pathway (NER); a sub-pathway of NER named transcription-coupled repair (TCR) resolve this problem by removing RNAP and recruiting repair proteins. In Escherichia coli, a TCR protein named “Mutation Frequency Decline” (Mfd) couples removal of RNAP through its motor activity with recruitment of the NER repair proteins. Mfd can be divided into two functional halves; the N-terminal region (MfdN, domains 1-3) is essential for NER protein recruitment, and the C-terminal region (MfdC, domains 4-7) is responsible for RNAP-interaction and motor activity. Data suggest Mfd undergoes large conformational movement to activate RNAP removal and repair protein recruitment. To study the activation mechanism of Mfd, we created several full-length “hyperactive” mutants by perturbing interactions between MfdN and MfdC. In all mutants, residue 79 in domain 1 is changed from aspartic acid to arginine (D79R), disrupting a key salt bridge interaction with arginine 804 in domain 6. The linker connecting MfdN and MfdC was made cleavable to allow separation of MfdN and MfdC, which enable us to study activities in equal molar concentration. We have studied the effect of the D79R mutation in vivo (cytotoxicity and UV sensitivity) and in vitro (enzyme activity and thermal stability), and demonstrate that this single residues change render the enzyme “hyperactive”. This confirms our model of activation: activation of Mfd results from breaking communication between MfdN and MfdC