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abstract: Engineered nanomaterials (ENMs) are added to numerous consumer products to enhance their effectiveness, whether it be for environmental remediation, mechanical properties, or as dietary supplements. Uses of ENMs include adding to enhance products, carbon for strength or dielectric properties, silver for antimicrobial properties, zinc oxide for UV sun-blocking properties, titanium dioxide for photocatalysis, or silica for desiccant properties. However, concerns arise from ENM functional properties that can impact the environment and a lack of regulation regarding ENMs leads to potential public exposure to ENMs and results in ill-informed public or manufacturer perceptions of ENMs. My dissertation evaluates the environmental, human health, and societal impacts of using ENMs, with a focus on ionic silver and nanosilver, in consumer and industrial products. Reproducible experiments served as functional assays to assess ENM distributions among various environmental matrices. Functional assay results were visualized using radar plots and aid in a framework to estimate likely ENM disposition in the environment. To assess beneficial uses of ENMs, bromide ion removal from drinking waters to limit disinfection by-product formation was studied. Silver-enabled graphene oxide materials were capable of removing bromide from water, and exhibited less competition from background solutes (e.g. natural organic matter) when compared against solely ionic silver addition to water for bromide removal. To assess complex interactions of ENMs with the microbiome, batch experiments were performed using fecal samples spiked with ionic silver or commercial dietary silver nanoparticles. Dietary nanosilver and ionic silver exposures to the fecal microbiome for 24 hours reduce short chain fatty acid (SCFA) production and changes the relative abundance of the microbiota. To understand the social perceptions of ENMS, statistically rigorous surveys were conducted to assess related perceptions related to the use of ENMs in drinking water treatment devices the general public and, separately, industrial manufacturers. These stakeholders are influenced by costs and efficiency of the technologies, consumer concerns of the safety of technologies, and environmental health and safety of the technologies. This dissertation represents novel research that took an interdisciplinary approach, spanning from wet-lab engineering bench scale testing to social science survey assessments to better understand the environmental, human health, and societal impacts of using ENMs such as nanosilver and ionic silver in industrial processes and consumer products. / Dissertation/Thesis / Doctoral Dissertation Civil, Environmental and Sustainable Engineering 2020

Le passage sélectif de molécules à travers la membrane interne des mitochondries est essentiel aux processus métaboliques des cellules eucaryotes. Cette communication cellulaire est assurée par des protéines transmembranaires de la famille des transporteurs mitochondriaux (MCF). Le transporteur ADP/ATP (AAC) est le membre le plus connu et le mieux caractérisé de cette famille. Il est responsable de l'import d'ADP dans la matrice mitochondriale et de l'export d'ATP après synthèse vers le cytosol. La structure d'AAC est connue mais plusieurs questions restent ouvertes concernant le mécanisme du transport, la sélectivité et l'état oligomérique, controversé, de la protéine. Pendant plusieurs années des études biochimiques réalisées sur la protéine solubilisée en détergent étaient en faveur d'une organisation dimérique du transporteur, mais la structure d'AAC, monomérique a remis en cause ce dogme. Afin de caractériser l'organisation oligomérique d'AAC in vivo, nous avons combiné plusieurs approches. Nous avons réalisé des expériences de FRET (Fluorescence Resonance Energy Transfer) directement sur des cellules mammifères ou bactériennes (E. coli) surexprimant la protéine AAC fusionnée avec des sondes FRET. En parallèle, nous avons mis au point des tests fonctionnels afin de contrôler l'état des mitochondries et l'activité du transporteur dans ces cellules. Enfin nous avons étudié la stoechiométrie de liaison de l'inhibiteur carboxyatractyloside grâce à des mesures de respiration sur des mitochondries extraites de foie de rat et placées dans différents états métaboliques. L'ensemble des résultats présentés dans ce manuscrit ont permis de montrer que 1) l'unité fonctionnelle d'AAC est monomérique 2) l'organisation structurale d'AAC dans les membranes natives dépend de l'état métabolique des mitochondries et peut être associée à des phénomènes de régulation. / The transport of small molecules through the inner mitochondrial membrane is essential in eukaryotic metabolism and is selectively controlled by a family of integral membrane proteins, the Mitochondrial Carrier Family (MCF). The ADP/ATP carrier (AAC), which is responsible for the import of ADP to the matrix of mitochondria and the export of newly synthesized ATP toward the cytosol, is the best-known and characterized MCF member. Although its structure sheds light on several aspects of the carrier activity, additional investigations are still required to decipher the whole transport mechanism, to understand the specificity and to characterize the controversial oligomeric state of the protein. For many years, based on studies mainly carried on detergent solubilized AAC the general consensus has been in favor of a dimeric organization of the carrier. The AAC three-dimensional structure, monomeric, broke this dogma. In order to get a precise insight into the in vivo oligomeric organization of AAC we combined several approaches. Fluorescence resonance energy transfer (FRET) measurements were performed directly on mammalian and E.coli cells expressing AAC labeled with several types of FRET probes. In parallel, different functional assays were established to control the state of the mitochondria in these cells and the transport activity of these AAC fusions. Lastly, measurements of the respiration rate coupled to the titration of the inhibitory effect of carboxyatractyloside on isolated rat liver mitochondria were used to investigate the organization of AAC in native mitochondria within two regimes of oxidative phosphorylation. Taken together the results described herein revealed that 1) AAC can function mechanistically as a monomer, 2) the organization of AAC in native membranes might be related to the state of the mitochondria and be involved in regulation.

L’identification des mutations constitutionnelles à l’origine d’une prédisposition génétique au cancer est essentielle à la prise en charge médicale des patients et de leurs familles. Depuis l’implémentation des technologies de séquençage à haut-débit dans les laboratoires diagnostiques, le principal défi n’est plus la détection des variations génétiques mais leur interprétation et leur classification. La question de l’interprétation de la variation est particulièrement cruciale lorsqu’elle conditionne la stratégie thérapeutique. Ainsi, il est essentiel de disposer de tests simples adaptables en routine diagnostique pour faciliter l’interprétation des variations génétiques. Dans ce contexte, nous avons utilisé un test fonctionnel développé par notre équipe pour classer des variations dans le gène TP53 à l’origine du syndrome de Li-Fraumeni et pour appréhender la corrélation génotype - phénotype chez les patients LFS. Dans un deuxième temps, nous avons évalué la pertinence d’une approche multi-omique (RNA-Seq et métabolomique) pour discriminer les cellules sauvages des cellules avec mutation hétérozygote du gène TP53 ou des gènes BRCA impliqués dans la prédisposition génétique aux cancers du sein et de l’ovaire. Sur la base des données de transcriptome, un modèle mathématique a été développé pour détecter les variants correspondant à des mutations délétères. Nous avons ensuite sélectionné les biomarqueurs les plus discriminants pour les intégrer dans un test fonctionnel de RT-MLPA dédié à la voie p53. Nous avons enfin adapté cet essai pour qu’il soit réalisable sur une simple prise de sang, sans immortalisation des lymphocytes du patient. / The identification of the constitutional mutation responsible for a genetic predisposition to cancer is essential to the clinical management of the patient and its relatives. With the implementation of high-throughput sequencing to the diagnostic routine of these pathologies, the challenge no longer lies within the detection of alterations but in their biological and clinical interpretation. While specific treatments are emerging, simple functional assays to help with the interpretation of the detected variants are needed. In this context, we used a functional test developed by our team to classify variations in the TP53 gene responsible for Li-Fraumeni syndrome and to understand the genotype-phenotype correlation in LFS patients. On the other hand, we assessed the relevance of a multi-omic approach (RNA-Seq and metabolomics) to discriminate wild-type cells from cells with a deleterious heterozygous mutation in TP53 or in the BRCA genes implicated in genetic predisposition to breast and ovarian cancers. Based on the transcriptomic data, a mathematical model has been developed to detect variants corresponding to deleterious mutations. Then we selected the most discriminating biomarkers and integrated them into a RT-MLPA functional assay dedicated to the p53 pathway. We finally adapted this test to be feasible on a simple blood test, without immortalization of the patient's lymphocytes.