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Cell wall mannans of Candida albicans mask β-(1,3)-glucan from recognition by Dectin-1, contributing to innate immune evasion. Glucan exposures are predominantly single receptor-ligand interaction sites of nanoscale dimensions. Candida species vary in basal glucan exposure and molecular complexity of mannans. We used super-resolution fluorescence imaging and a series of protein mannosylation mutants in C. albicans and C. glabrata to investigate the role of specific N-mannan features in regulating the nanoscale geometry of glucan exposure. Decreasing acid labile mannan abundance and α-(1,6)-mannan backbone length correlated most strongly with increased density and nanoscopic size of glucan exposures in C. albicans and C. glabrata, respectively. Additionally, a C. albicans clinical isolate with high glucan exposure produced similarly perturbed N-mannan structures and elevated glucan exposure geometry. Thus, acid labile mannan structure influences the nanoscale features of glucan exposure, impacting the nature of the pathogenic surface that triggers immunoreceptor engagement, aggregation, and signaling. Graus et al. find that N-mannan structural features regulated by Candida mannosyltransfersases control glucan exposure. Loss of mannan increased the frequency and size of glucan exposures and changed multivalent receptor engagement. Changes to mannan structure in a bloodstream isolate are associated with elevated glucan exposure at the nanoscale.

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Mitochondria are the power plant of most non-green eukaryotic cells. In order to understand mitochondrial functions and their regulation, knowledge of the spatiotemporal organization of their proteins is important. Mitochondrial membrane proteins can diffuse within membranes. They are involved in diverse functions e.g. protein import, cell respiration, metabolism, metabolite transport, fusion, fission or formation of the mitochondrial architecture. Furthermore, mitochondria compose of different subcompartments with different tasks. Especially, the inner mitochondrial membrane (IM), where the oxidative phosphorylation (OXPHOS) takes places, has a complex architecture with cristae extending into the matrix. The present work revealed the restricted localization of some mitochondrial proteins to specific membrane sections and linked it to their function or bioenergetic circumstances in the living cell. 
Single particle tracking (SPT) techniques like tracking and localization microscopy (TALM) allow to localize proteins with a precision below 20 nm. Additionally, tracking single proteins provides information about their mobility, dynamic and their spatiotemporal organization. TALM uses proteins, which were genetically tagged either with the HaloTag® (HaloTag) or the fSnapTag® (fSnapTag). These tags can be orthogonally and posttranslationally stained with specific and self-marking dyes. If the dyes are conjugated to the respective substrate of the tag. Single molecule labeling of mitochondrial proteins was performed substoichiometrically using membrane permeable rhodamine dyes, either tetramethylrhodamine (TMR) or silicon rhodamine (SiR). TALM allowed to localize proteins in different mitochondrial subcompartments. The gained trajectories and trajectory maps of mitochondrial proteins revealed their spatiotemporal organization. In the case of IM proteins like F1FO ATP synthase (Complex V - CV) a restricted diffusion in the CM, which is part of the continuous IM, was determined. The unimpeded diffusion of mitochondrial proteins in the outer mitochondrial membrane (OM) was compared with the mobility of IM proteins. The diffusion of mitochondrial IM proteins was restricted by the IM architecture and their diffusion coefficients were lower. Furthermore, significant differences of different mitochondrial IM proteins were compared, showing different localizations in the IM often coupled to their function, accompanied by different spatiotemporal organization and diffusion coefficients. Furthermore, a distinction was made between diffusion of proteins in the inner boundary membrane (IBM) and proteins that preferentially diffuse in the cristae membrane (CM). Evaluating trajectory maps, the different subcompartments in the IM were revealed by trajectories and the trajectory directionality, allowing the identification of mitochondrial proteins, which mark these subcompartments.
The morphology of mitochondria / mitochondrial networks and their bioenergetic parameters are linked to the metabolic states of the cell. In this work, the connection of the spatiotemporal protein organization of CV and the IM architecture was uncovered on the micro- and nanoscopic level and linked to the metabolic state of the cell. It was determined that the spatiotemporal organization of the CV was altered, when CV was inhibited. In addition, the bioenergetic influence of cells on the spatiotemporal behavior of CV and the reorganization of the IM architecture was investigated by TALM and compared with results of electron microscopy images. It was shown that starvation of cells led to a loss of cristae and thus to an increased mobility and spatiotemporal reorganization of CV. Taken together, the results presented in this work showed that a correctly functioning and active CV helps to maintain the IM architecture and both, the spatiotemporal organization of CV and the IM architecture were coupled to the metabolic state.. 
In order to investigate putative protein-protein interactions by colocalization and co-locomotion studies on single molecule level, dual color SPT is needed. Therefore, posttranslational and substoichimetric labeling as performed in TALM was tested for its potential of protein-protein interaction studies of mitochondrial membrane proteins. Here, a genetically double tagged translocase of the outer membrane subunit-20 (Tom20) (Tom20:HaloTag:fSnapTag) acted as a positive control. It turned out that substoichimetric, posttranslational labeling of mitochondrial proteins was not suitable for protein-protein interaction studies on mitochondrial proteins, because it was restricted by the low labeling degrees needed for TALM. However, dual-color TALM still allowed to study effects of proteins influencing the IM architecture and to study their influence on the spatiotemporal organization of CV. The co-transfection of Mic10, as the central protein of the mitochondrial inner membrane organizing system / mitochondrial contact site complex / mitochondrial organizing structure (MINOS / MICOS / MitOS (MINOS/MICOS)), altered the regular and aligned organization of the cristae. This was measured by a changed spatiotemporal organization of the CV, such as the loss of the perpendicular oriented of CV subunit-γ (CV-SUγ) cristae trajectories. In contrast to this, co-transfection of CV subunit-e (CV-SUe), important for dimerization of CV, increased the number of cristae trajectories. 
Mitochondria are three-dimensional (3D) cell organelles. Consequently, subcompartments like the IBM and CM are a 3D space in which CV is localized and diffuses. Thus, the diffusion of mitochondrial proteins is underestimated by two-dimensional SPT e.g. lateral confined diffusion can result from mitochondrial proteins diffusing along the z-axis of the microscope. In order to reveal the 3D spatiotemporal organization of CV, the potential of TALM to be extended to a 3D-SPT technique was investigated. Therto a cylindrical lens was installed in the emission path of a total internal reflection fluorescence (TIRF) microscope. This leads to an astigmatically distorted point spread function (PSF) of the fluorescent single molecule signals. This distortion allowed the reconstruction of single molecule localizations of CV to a superresolved image of the IM, in living cells. In addition, 3D-TALM enabled to display the 3D architecture of the IM by 3D trajectories of CV. 3D-TALM was able to detect whether CV diffuses in the IBM or in the CM, and extended the information about its mobility in the CM that it takes place in a disc-like manner. In this way it could be shown that CV is mobile within the cristae in all directions. Finally, 3D-TALM revealed an altered IM architecture caused by the metabolic state of the cell. As performed in two-dimensional TALM, the cells were kept under starving conditions. Here the now tubular IM architecture was revealed by 3D-TALM. The reversed metabolic state under improved respiratory conditions unexpectedly led to a more diverse IM architecture. These ultrastructural changes were also revealed by electron microscopy. Consequently, 3D-TALM enabled the study of IM architecture by tracking CV under different metabolic conditions, allowing an ultrastructural analysis of mitochondria in living cells. In addition, 3D TALM provided the spatiotemporal organization of CV under different metabolic conditions, so that the diffusion coefficients of CV could be related to changes in IM architecture caused by the metabolic condition.

Les neutrophiles sont les leucocytes les plus nombreux et les premières cellules à arriver au site de l’infection où elles internalisent les pathogènes par phagocytose. Dès le début du processus, la NADPH oxydase s’assemble au phagosome où elle permet la production de formes réactives de l’oxygène contribuant ainsi à la destruction du pathogène. La sous-unité catalytique membranaire de la NADPH oxydase, Nox2, est donc présente à la coupe phagocytaire puis au phagosome. Le dessein de cette étude était de déterminer quelles sont les sources subcellulaires de la protéine Nox2, de savoir si la protéine s’accumule au phagosome et le cas échéant selon quelle cinétique. Dans le but de comprendre la dynamique de la protéine Nox2, la protéine d’échafaudage IQGAP1 qui est associée au cytosquelette a également été étudiée. Enfin l’étude de l’organisation spatiale de la protéine Nox2 à la synapse phagocytaire a également été abordé.En utilisant des cellules neutrophil-like (PLB-985) ainsi que des neutrophiles humains, notre étude a montré par immunofluorescence la présence de la protéine Nox2 dans des endosomes de recyclage ou dans des endosomes précoces. Lors de la phagocytose ils avoisinent le phagosome suggérant leur implication dans l’apport de la protéine Nox2 à la membrane de ce dernier. L’utilisation de cellules PLB-985 pour lesquelles l’expression de Nox2 a été supprimée puis réintroduite avec un transgène codant pour la protéine GFP-Nox2 montre que la sous-unité Nox2 s’accumule au phagosome pendant les vingt minutes suivant sa fermeture. Dans notre étude, la protéine IQGAP1 ne semble pas avoir d’effet sur la phagocytose ou sur la production de FRO par la NADPH oxydase. Enfin, grâce à une technique de microscopie super-résolution (le dSTORM) l’évolution du pattern de Nox2 dans la membrane a été évalué au cours du temps en phagocytose frustrée. En dix minutes, le nombre de clusters de protéine Nox2 augmente mais leur taille reste inchangée. / Neutrophils are the most numerous leukocytes and the first cells to arrive at the site of infection where they internalize pathogens by phagocytosis. From the beginning of the process, the NADPH oxidase is assembled at the phagosome, where it allows the production of reactive oxygen species (ROS), thus contributing to the destruction of the pathogen. The membrane bound catalytic subunit of the NADPH oxidase, Nox2, is therefore recruited at the phagocytic cup and then at the phagosome. The purpose of this study was to determine, which are the subcellular sources of the Nox2 protein, whether the protein accumulates at the phagosome and if so, according to which kinetics. In order to modify the dynamics of the Nox2 protein, the scaffold protein IQGAP1 that is associated with the cytoskeleton was also studied. Finally, the spatial organization of the Nox2 protein in the phagocytic synapse was also investigated.Using neutrophil-like cells (PLB-985) as well as human neutrophils, our study showed by immunofluorescence the presence of the Nox2 protein in recycled or early endosomes. During phagocytosis, they are close to the phagosome, suggesting their involvement in the contribution of the Nox2 protein to the phagosome membrane. The use of PLB-985 for which Nox2 expression has been suppressed and then reintroduced with a transgene encoding the GFP-Nox2 protein shows that the Nox2 subunit accumulates at the phagosome during the first twenty minutes after its closure. In our study, the protein IQGAP1 does not appear to have any effect on phagocytosis or on the production of ROS by NADPH oxidase. Finally, using super resolution microscopy (dSTORM) the evolution of the Nox2 pattern in the membrane has been evaluated over time in frustrated phagocytosis. Within ten minutes, the number of Nox2 protein clusters increases but their size remains unchanged.

Ce travail de thèse propose d’étudier les propriétés photophysiques de systèmes moléculaires à la fois photochromes et fluorescents. Sous l’effet de la lumière, ces molécules peuvent subir une désexcitation radiative, non radiative ou une photo-isomérisation. Dans la première partie de ce travail, la compétition entre ces différentes voies a été étudiée en détail grâce à deux photochromes proches sur le plan structurel et présentant une émission de fluorescence de couleur différente pour chaque forme du photochrome. Les efficacités relatives et les mécanismes de désexcitation ont été étudiés à l’échelle macroscopique. Cette comparaison a été accompagnée d’une étude sous microscope pour laquelle des nano-bâtonnets d’or ont été intégrés au système photochrome-fluorescent de manière à étudier l’influence d’un champ plasmon de surface localisé sur les différents phénomènes de désexcitation à une échelle proche de celle de la nanoparticule individuelle. Une deuxième partie de ce travail consiste en la préparation et l’étude des propriétés photophysiques de nanoparticules organiques constituées de dyades fluorescentes et photochromes. La fluorescence de ce type de nano-objets peut être efficacement contrôlée par la lumière : la conversion de quelques unités photochromes peut permettre, par transfert d’énergie intermoléculaire, d’éteindre la totalité de la fluorescence d’une nanoparticule. L’observation de cet effet pour des nanoparticules individuelles a été mise en application pour le développement d’une méthode de microscopie optique super-résolution. / The main purpose of this PhD work is the study of fluorescent and photochromic molecular systems. Under light illumination, such molecules may undergo radiative and non-radiative deactivation or photoisomerization. In the first part of this work, this competition has been investigated for structurally related photochromic compounds with different fluorescence colors corresponding to both isomers. Efficiency and mechanisms of the deactivation pathways were unraveled at the macroscopic scale. This competition was further studied under microscope with gold nanorods included in the sample, in order to study the effect of localized surface plasmon resonance on the different deactivation processes.The second part of this work consists in the preparation and the photophysical study of organic nanoparticles composed of fluorescent-photochromic dyads. Fluorescence of such nano-objects can be efficiently driven by light : the switching of a few photochromic units is enough to turn off the entire fluorescence of a nanoparticle by intermolecular energy transfer. Observation of this effect at the level of individual nanoparticles allowed the developement of a super-resolution method for optical microscopy.

Mapping of earth resources, environmental monitoring, and many other systems require high-resolution wide-area imaging. Since images often have to be captured at night or in inclement weather conditions, a capability is provided by Synthetic Aperture Radar (SAR). SAR systems exploit radar signal's long-range propagation and utilize digital electronics to process complex information, all of which enables high-resolution imagery. This gives SAR systems advantages over optical imaging systems, since, unlike optical imaging, SAR is effective at any time of day and in any weather conditions. Moreover, advanced technology called Interferometric Synthetic Aperture Radar (InSAR), has the potential to apply phase information from SAR images and to measure ground surface deformation. However, given the current state of technology, the quality of InSAR data can be distorted by several factors, such as image co-registration, interferogram generation, phase unwrapping, and geocoding. Image co-registration aligns two or more images so that the same pixel in each image corresponds to the same point of the target scene. Super-Resolution (SR), on the other hand, is the process of generating high-resolution (HR) images from a low-resolution (LR) one. SR influences the co-registration quality and therefore could potentially be used to enhance later stages of SAR image processing. Our research resulted in two major contributions towards the enhancement of SAR processing. The first one is a new learning-based SR model that can be applied with SAR, and similar applications. A second major contribution is utilizing the devised model for improving SAR co-registration and InSAR interferogram generation, together with methods for evaluating the quality of the resulting images. In the case of phase unwrapping, the process of recovering unambiguous phase values from a two-dimensional array of phase values known only modulo $2\pi$ rad, our research produced a third major contribution. This third major contribution is the finding that quantum annealers can resolve problems associated with phase unwrapping. Even though other potential solutions to this problem do currently exist - based on network programming for example - network programming techniques do not scale well to larger images. We were able to formulate the phase unwrapping problem as a quadratic unconstrained binary optimization (QUBO) problem, which can be solved using a quantum annealer. Since quantum annealers are limited in the number of qubits they can process, currently available quantum annealers do not have the capacity to process large SAR images. To resolve this limitation, we developed a novel method of recursively partitioning the image, then recursively unwrapping each partition, until the whole image becomes unwrapped. We tested our new approach with various software-based QUBO solvers and various images, both synthetic and real. We also experimented with a D-Wave Systems quantum annealer, the first and only commercial supplier of quantum annealers, and we developed an embedding method to map the problem to the D-Wave 2000Q_6, which improved the result images significantly. With our method, we were able to achieve high-quality solutions, comparable to state-of-the-art phase-unwrapping solvers. / Graduate

<p>High-throughput phenotyping has emerged as a powerful approach to expedite crop breeding programs. Modern remote sensing systems, including manned aircraft, unmanned aerial vehicles (UAVs), and terrestrial platforms equipped with multiple sensors, such as RGB cameras, multispectral, hyperspectral, and infrared thermal sensors, as well as light detection and ranging (LiDAR) scanners are now widely used technologies in advancing high throughput phenotyping. These systems can collect high spatial, spectral, and temporal resolution data on various phenotypic traits, such as plant height, canopy cover, and leaf area. Enhancing the capability of utilizing such remote sensing data for automated phenotyping is crucial in advancing crop breeding. This dissertation focuses on developing deep learning and transfer learning methodologies for crop phenotyping using multi-modality remote sensing and environmental data. The techniques address two main areas: multi-temporal/across-field biomass prediction and multi-scale remote sensing data fusion.</p> <p><br></p> <p>Biomass is a plant characteristic that strongly correlates with biofuel production, but is also influenced by genetic and environmental factors. Previous studies have shown that deep learning-based models are effective in predicting end-of-season biomass for a single year and field. This dissertation includes development of transfer learning methodologies for multiyear,</p> <p>across-field biomass prediction. Feature importance analysis was performed to identify and remove redundant features. The proposed model can incorporate high-dimensional genetic marker data, along with other features representing phenotypic information, environmental conditions, or management practices. It can also predict end-of-season biomass using mid-season remote sensing and environmental data to provide early rankings. The framework was evaluated using experimental trials conducted from 2017 to 2021 at the Agronomy Center for Research and Education (ACRE) at Purdue University. The proposed transfer learning techniques effectively selected the most informative training samples in the target domain, resulting in significant improvements in end-of-season yield prediction and ranking. Furthermore, the importance of input remote sensing features was assessed at different growth stages.</p> <p><br></p> <p>Remote sensing technology enables multi-scale, multi-temporal data acquisition. However, to fully exploit the potential of the acquired data, data fusion techniques that leverage the strengths of different sensors and platforms are necessary. In this dissertation, a generative adversarial network (GAN) based multiscale RGB-guided model and domain adaptation framework were developed to enhance the spatial resolution of multispectral images. The model was trained on limited high spatial resolution images from a wheel-based platform and then applied to low spatial resolution images acquired by UAV and airborne platforms.</p> <p>The strategy was tested in two distinct scenarios, sorghum plant breeding, and urban areas, to evaluate its effectiveness.</p>

[pt] A super resolução (SR) é um tema de suma importância em domínios de conhecimentos variados, como por exemplo a área médica, de monitoramento e de segurança. O uso de redes neurais profundas para a resolução desta tarefa é algo extremamente recente no universo da sísmica, tendo poucas referências, as quais começaram a ser divulgadas há menos de 2 anos. Todavia, a literatura apresenta uma vasta gama de métodos, que utilizam redes neurais para a super resolução de imagens naturais. Tendo isto em vista, o objetivo deste trabalho é explorar tais abordagens aplicadas em dados sísmicos sintéticos de reservatórios. Para isto, foram empregados modelos de importância cronológica na literatura e foram comparados com um método clássico de interpolação e com os modelos da literatura de super resolução de imagens sísmicas. São estes modelos: o SRCNN, o RDN, a abordagem do Deep Image Prior e o SAN. Por fim, os resultados apresentam que o PSNR obtido por arquiteturas de projetos no domínio da sísmica equivale a 38.23 e o melhor resultado das arquiteturas propostas 38.62, mostrando o avanço que tais modelos trazem ao campo da sísmica. / [en] Super resolution (SR) is a topic of notable importance in domains of assorted knowledge, such as the medical, monitoring, and security areas. The use of deep neural networks to solve this task is something extremely recent in the seismic field, with few references, which began to be published less than 2 years ago. However, the literature presents a wide range of methods, using neural networks for the super resolution of natural images. With this in mind, the objective of this work is to explore such approaches applied to synthetic seismic data from reservoirs. For this, models of chronological importance in the literature were used and compared with a classic interpolation method and with models of the literature of super resolution of seismic images. These models are: SRCNN, RDN, the Deep Image Prior approach and SAN. The results show that the PSNR obtained by architectures developed for the seismic domain is equivalent to 38.23 and the best result of the proposed architectures is 38.62, showing the progress that such models bring to the seismic domain.

No description available.