A stable phage lysin (Cpl-1) dimer with increased antipneumococcal activity and decreased plasma clearance.

Bacteriophages (phages) produce endolysins (lysins) as part of their lytic cycle in order to degrade the peptidoglycan layer of the infected bacteria for subsequent release of phage progeny. Because these enzymes maintain their lytic and lethal activity against Gram-positive bacteria when added extrinsically to the cells, they have been actively exploited as novel anti-infectives, sometimes termed enzybiotics. As with other relatively small peptides, one issue in their clinical development is their rapid inactivation through proteolytic degradation, immunological blockage and renal clearance. The antipneumococcal lysin Cpl-1 was shown to escape both proteolysis and immunological blockage. However, its short plasma half-life (20.5 min in mice) may represent a shortcoming for clinical usefulness. Here we report the construction of a Cpl-1 dimer with a view to increasing both the antipneumococcal specific activity and plasma half-life of Cpl-1. Dimerisation was achieved by introducing specific cysteine residues at the C-terminal end of the enzyme, thus favouring disulphide bonding. Compared with the native monomer, the constructed dimer demonstrated a two-fold increase in specific antipneumococcal activity and a ca. ten-fold decrease in plasma clearance. As several lysins are suspected to dimerise on contact with their cell wall substrate to be fully active, stable pre-dimerised enzymes may represent a more efficient alternative to the native monomer.

A novel derivative of the chelon desferrioxamine for site-specific conjugation to antibodies.

We describe the preparation of the modified chelator aminooxyacetyl-ferrioxamine, and the replacement of its iron atom by 67Ga at high specific activity. The aminooxy function of this compound was allowed to react with the aldehyde groups generated by the periodate oxidation of the oligosaccharide of a mouse IgG1 monoclonal antibody (MAb) directed against carcino-embryonic antigen (CEA). The use of the aminooxy group allowed a stable bond to be formed between the chelon and the antibody with no need for reduction. Iron was removed from the ferrioxamine moiety and replaced by 67Ga either before or after conjugation of the chelon to the antibody. In either case the labelled antibody was injected into nude mice bearing a human colon carcinoma having the appropriate antigenicity. Unoxidized antibody, labelled with 125I by conventional methods, was co-injected as an internal control. Additional control experiments were carried out with a non-immune IgG using the same 67Ga-labelled modified chelon as above. The in vivo distribution of the modified antibodies was evaluated at various times between 24 and 96 hr after injection. The methods used were gamma-camera imaging and, more quantitatively, gamma-counting of the various organs after dissection. Interestingly, with the metal-chelon-labelled antibody, the intensity and specificity of tumor labelling was comparable and in some cases superior to the results obtained with radio-iodinated antibody. In particular, there was almost no increase in liver and spleen uptake of radioactive metal relative to radio-iodine, contrary to what has been observed with most antibodies labelled with 111In after conjugation with DTPA.

Role of glutathione in macrophage activation: effect of cellular glutathione depletion on nitrite production and leishmanicidal activity.

We have examined the effects of two agents depleting the intracellular pool of glutathione (GSH) on macrophage activation induced by IFN-gamma + LPS, as measured by nitrite production and leishmanicidal activity. Diethylmaleate (DEM), which depletes intracellular GSH by conjugation via a reaction catalyzed by the GSH-S-transferase, strongly inhibited nitrite secretion and leishmanicidal activity when added before or at the time of addition of IFN-gamma + LPS; this inhibition was progressively lost when addition of DEM was delayed up to 10 hr. A close correlation was observed between levels of intracellular soluble GSH during activation and nitrite secretion. Inhibition was partially reversed by the addition of glutathione ethyl ester (GSH-Et). Buthionine sulfoximine (BSO), a specific inhibitor of gamma-glutamylcysteine synthetase, also inhibited macrophage activation, although to a lesser extent than DEM despite a more pronounced soluble GSH depletion. This inhibition was completely reversed by the addition of GSH-Et. DEM and BSO did not alter cell viability or PMA-triggered O2- production by activated macrophages, suggesting that the inhibitory effects observed on nitrite secretion and leishmanicidal activity were not related to a general impairment of macrophage function. DEM and BSO treatment reduced iNOS specific activity and iNOS protein in cytosolic extracts. DEM also decreased iNOS mRNA expression while BSO had no effect. Although commonly used as a GSH-depleting agent, DEM may have additional effects because it can also act as a sulhydryl reagent; BSO, on the other hand, which depletes GSH by enzymatic inhibition, has no effect on protein-bound GSH. Our results suggest that both soluble and protein-bound GSH may be important for the induction of NO synthase in IFN-gamma + LPS-activated macrophages.

Demyelination in brain cell aggregate cultures, induced by a monoclonal antibody against the myelin/oligodendrocyte glycoprotein (MOG).

Previous work has shown that aggregate cultures prepared from fetal rat telencephalon and grown in a chemically defined medium offer a useful model to study developmental processes such as myelin synthesis. Since compact myelin is formed in these cultures, we investigated the possibility to use this culture system to study demyelinating mechanisms. In particular, we examined the effect of a monoclonal antibody (8-18C5) directed against the myelin/oligodendrocyte glycoprotein (MOG). We found that addition of anti-MOG antibodies and complement to aggregate cultures led to a highly significant decrease in myelin basic protein (MBP) content and 2',3'-cyclic nucleotide 3'-phosphohydrolase (CNP) specific activity. These results indicate that, in our culture system, anti-MOG antibodies have a strong demyelinating effect.

Cardiac expression of ms1/STARS, a novel gene involved in cardiac development and disease, is regulated by GATA4.

Ms1/STARS is a novel muscle-specific actin-binding protein that specifically modulates the myocardin-related transcription factor (MRTF)-serum response factor (SRF) regulatory axis within striated muscle. This ms1/STARS-dependent regulatory axis is of central importance within the cardiac gene regulatory network and has been implicated in cardiac development and postnatal cardiac function/homeostasis. The dysregulation of ms1/STARS is associated with and causative of pathological cardiac phenotypes, including cardiac hypertrophy and cardiomyopathy. In order to gain an understanding of the mechanisms governing ms1/STARS expression in the heart, we have coupled a comparative genomic in silico analysis with reporter, gain-of-function, and loss-of-function approaches. Through this integrated analysis, we have identified three evolutionarily conserved regions (ECRs), α, SINA, and DINA, that act as cis-regulatory modules and confer differential cardiac cell-specific activity. Two of these ECRs, α and DINA, displayed distinct regulatory sensitivity to the core cardiac transcription factor GATA4. Overall, our results demonstrate that within embryonic, neonatal, and adult hearts, GATA4 represses ms1/STARS expression with the pathologically associated depletion of GATA4 (type 1/type 2 diabetic models), resulting in ms1/STARS upregulation. This GATA4-dependent repression of ms1/STARS expression has major implications for MRTF-SRF signaling in the context of cardiac development and disease.

Differentiation of rat striatal embryonic stem cells in vitro: monolayer culture vs. three-dimensional coculture with differentiated brain cells.

Several groups have demonstrated the existence of self-renewing stem cells in embryonic and adult mouse brain. In vitro, these cells proliferate in response to epidermal growth factor, forming clusters of nestin-positive cells that may be dissociated and subcultured repetitively. Here we show that, in stem cell clusters derived from rat embryonic striatum, cell proliferation decreased with increasing number of passages and in response to elevated concentrations of potassium (30 mM KCl). In monolayer culture, the appearance of microtubule-associated protein type-5-immunoreactive (MAP-5(+)) cells (presumptive neurons) in response to basic fibroblast growth factor (bFGF) was reduced at low cell density and with increasing number of passages. In the presence of bFGF, elevated potassium caused a more differentiated neuronal phenotype, characterized by an increased proportion of MAP-5(+) cells, extensive neuritic branching, and higher specific activity of glutamic acid decarboxylase. Dissociated stem cells were able to invade cultured brain cell aggregates containing different proportions of neurons and glial cells, whereas they required the presence of a considerable proportion of glial cells in the host cultures to become neurofilament H-positive. The latter observation supports the view that astrocyte-derived factors influence early differentiation of the neuronal cell lineage.

How unethical actions can be learned : the analysis of the sporting life courses of doping athletes

The aim of the present article was to present a specific activity approach called the 'life course of a practice' that could be used for the analysis of the long-term dynamic of learning within diverse social practices. The cases of unethical actions (doping) within a population of high-level athletes were examined, considering they could be fruitful exam- 10 ples for this type of analysis. After describing the theoretical construct 'life course of a practice', the places of these unethical actions in the sporting life courses and elements that characterize their emergence in the long-term dynamics are described. Last, targeted prevention measures of unethical actions over a lifetime are presented.

Reconstitution into liposomes of the tonoplast ATP-and pyrophosphate-dependent proton pumps from Rubus hispidus cell cultures.

The ATP- and pyrophosphate-dependent proton pumps from tonoplast-enriched vesicles prepared from Rubus hispidus cell cultures were solubilized in the presence of polyoxyethylene(9,10)p-t-octylphenol (Triton X-100) and reconstituted into liposomes of soybean phospholipids, using Bio-Beads SM-2 to remove the detergent. The specific activity of the two pumps was greatly increased by the solubilization-reconstitution procedure. Identical characteristics were found for pyrophosphate-dependent proton transport in native and reconstituted vesicles. On the other hand, the ATP-dependent proton transport of the reconstituted vesicles was no longer inhibited by KNO3.

Molecular basis of coagulation factor V deficiency caused by the R1698W inter-domain mutation.

Coagulation factor V (FV) deficiency is characterised by variable bleeding phenotypes and heterogeneous mutations. To add new insights into the FV genotype-phenotype relationship, we characterised the R1698W change in the A3 domain, at the poorly investigated interface with the A2 domain. The FV R1698W mutation was responsible for a markedly reduced expression level (10% of FV-WT) and specific activity in thrombin generation (0.39). Interestingly, the FVa1698W showed rapid activity decay upon activation due to increased dissociation rate between the heavy and light chains. The importance of the size and charge of the residue at position 1698 was investigated by three additional recombinant mutants, FVR1698A, FVR1698Q, and FVR1698E. FVR1698A and FVR1698Q expression (30 and 45% of FV-WT), specific activity (both 0.57) and stability were all reduced. Noticeably, FVR1698E showed normal activity and stability despite poor expression (10% of FV-WT). These data indicate the essential role of R1698 for normal biosynthetic process and support local flexibility for positively or negatively charged residues to produce stable and functional A3-A2 domain interactions. Their experimental alteration produces a gradient of FV defects, which help to interpret the wide spectrum of phenotypes in FV-deficient patients.

Demyelination induced in aggregating brain cell cultures by a monoclonal antibody against myelin/oligodendrocyte glycoprotein.

A monoclonal antibody (8-18C5) directed against myelin/oligodendrocyte glycoprotein (MOG) induced demyelination in aggregating brain cell cultures. With increasing doses of anti-MOG antibody in the presence of complement, myelin basic protein (MBP) concentration decreased in a dose-related manner. A similar, albeit less pronounced, effect was observed on specific activity of 2',3'-cyclic nucleotide 3'-phosphohydrolase. In the absence of complement, anti-MOG antibody did not induce detectable demyelination. In contrast to the effect of anti-MOG antibody and as expected, anti-MBP antibody did not demyelinate aggregating brain cell cultures in the presence of complement. These results provide additional support to the suggestion that MOG, a quantitatively minor myelin component located on the external side of the myelin membrane, is a good target antigen for antibody-induced demyelination. Indeed, they show that a purified anti-MOG antibody directed against a single epitope on the glycoprotein can produce demyelination, not only in vivo as previously shown, but also in cultures. Such an observation has not been made with polyclonal antisera raised against purified myelin proteins like MBP and proteolipid protein, the major protein components of the myelin membrane, or myelin-associated glycoprotein. These observations may have important implications regarding the possible role of anti-MOG antibodies in demyelinating diseases.

Na(+)-K(+)-ATPase gene expression during in vitro development of rat fetal forebrain.

The existence of at least three isoforms of Na(+)-K(+)-ATPase in adult brain tissues [alpha 1, kidney type; alpha 2 [or alpha(+)]; alpha 3] suggests that these genes might be regulated in a cell-specific and time-dependent manner during development. We have studied this question in serum-free aggregating cell cultures of mechanically dissociated rat fetal telencephalon. At the protein level, the relative rate of synthesis of the pool of alpha 1-, alpha 2-, and alpha 3-subunits increased approximately twofold over 15 days of culture, leading to a marked increase in the immunochemical pool of alpha-subunits as measured by a panspecific polyclonal antibody. Concomitantly, Na(+)-K(+)-ATPase enzyme-specific activity increased three- (lower forebrain) to sixfold (upper forebrain). The transcripts of all three alpha-isoforms and beta-subunit were detected in vitro in similar proportion to the level observed in vivo. alpha 3-mRNA (3.7 kb) was more abundant than alpha 1 (3.7 kb) or alpha 2 (5.3 and 3.4 kb). Cytosine arabinoside (0.4 microM) and cholera toxin (0.1 microM) were used to selectively eliminate glial cells or neurons, respectively. It was found that alpha 2-mRNA is predominantly transcribed in glial cell cultures, whereas alpha 3- and beta 1-mRNA (2.7, 2.3, and 1.8 kb) are predominant in neuronal cultures.

Biochemical characterization of a myelin fraction isolated from rat brain aggregating cell cultures.

Subcellular fractions isolated from rat brain aggregating cell cultures were studied by electron microscopy and showed the presence of typical myelin membranes. The chemical composition of purified culture myelin was similar to the fraction isolated from rat brain in terms of CNP specific activity, protein and lipid composition. The ratio of small to large components of myelin basic protein was comparable in culture and in vivo. These two proteins incorporated radioactive phosphorus. The major myelin glycoprotein was present and during development in culture its apparent molecular weight decreased although it never reached the position observed in myelin isolated from adult rats. In culture, the yield of myelin did not increase substantially between 33 and 50 days and was comparable to that of 15-day-old rat brain. The ratio basic protein to proteolipid protein resembled immature myelin and the cerebroside content was very low. A 'floating fraction' was isolated from the cultures and contained some myelin but mostly single membranes. Although these results indicate that myelin maturation is delayed in vitro this culture system provides substantial amounts of purified myelin to allow a complete biochemical analysis and metabolic studies during development.

La pertinence en science forensique. Une (en)quête épistémologique et empirique

A criminal investigation requires to search and to interpret vestiges of a criminal act that happened in a past time. The forensic investigator arises in this context as a critical reader of the investigation scene, in search of physical traces that should enable her to tell a story of the offence/crime which allegedly occurred. The challenge of any investigator is to detect and recognise relevant physical traces in order to provide forensic clues for investigation and intelligence purposes. Inspired by this obser- vation, the current research focuses on the following questions : What is a relevant physical trace? And, how does the forensic investigator know she is facing one ? The interest of such questions is to provide a definition of a dimension often used in forensic science but never studied in its implications and operations. This doctoral research investigates scientific paths that are not often explored in forensic science, by using semiotic and sociological tools combined with statistical data analysis. The results are shown following a semiotic path, strongly influenced by Peir- ce's studies, and a second track, called empirical, where investigations data were analysed and forensic investigators interviewed about their work practices in the field. By the semiotic track, a macroscopic view is given of a signification process running from the discove- red physical trace at the scene to what is evaluated as being relevant for the investigator. The physical trace is perceived in the form of several signs, whose meaning is culturally codified. The reasoning should consist of three main steps : 1- What kind of source does the discovered physical trace refer to ? 2- What cause/activity is at the origin of this source in the specific context of the case ? 3- What story can be told from these observations ? The stage 3 requires to reason in creating hypotheses that should explain the presence of the discovered trace coming from an activity ; the specific activity that is related to the investigated case. To validate this assumption, it would depend on their ability to respond to a rule of relevancy. The last step is the symbolisation of the relevancy. The rule would consist of two points : the recognition of the factual/circumstantial relevancy (Is the link between the trace and the case recognised with the formulated hypothesis ? ) and appropriate relevancy (What investment is required to collect and analyse the discovered trace considering the expected outcome at the investigation/intelligence level?). This process of meaning is based on observations and a conjectural reasoning subject to many influences. In this study, relevancy in forensic science is presented as a conventional dimension that is symbolised and conditioned by the context, the forensic investigator's practice and her workplace environment (culture of the place). In short, the current research states relevancy results of the interactions between parameters from situational, structural (or organisational) and individual orders. The detection, collection and analysis of relevant physical traces at scenes depends on the knowledge and culture mastered by the forensic investigator. In the study of the relation relevant trace-forensic investigator, this research introduces the KEE model as a conceptual map that illustrates three major areas of forensic knowledge and culture acquisition, involved in the research and evaluation of the relevant physical trace. Through the analysis of the investigation data and interviews, the relationship between those three parameters and the relevancy was highlighted. K, for knowing, embodies a rela- tionship to the immediate knowledge allowing to give an overview of the reality at a specific moment ; an important point since relevancy is signified in a context. E, for education, is considered through its relationship with relevancy via a culture that tends to become institutionalised ; it represents the theoretical knowledge. As for the parameter E, for experience, it exists in its relation to relevancy in the adjustments of the strategies of intervention (i.e a practical knowledge) of each practitioner having modulated her work in the light of success and setbacks case after case. The two E parameters constitute the library resources for the semiotic recognition process and the K parameter ensures the contextualisation required to set up the reasoning and to formulate explanatory hypotheses for the discovered physical traces, questioned in their relevancy. This research demonstrates that the relevancy is not absolute. It is temporal and contextual; it is a conventional and relative dimension that must be discussed. This is where the whole issue of the meaning of what is relevant to each stakeholder of the investigation process rests. By proposing a step by step approach to the meaning process from the physical trace to the forensic clue, this study aims to provide a more advanced understanding of the reasoning and its operation, in order to streng- then forensic investigators' training. This doctoral research presents a set of tools critical to both pedagogical and practical aspects for crime scene management while identifying key-influences with individual, structural and situational dimensions. - Une enquête criminelle consiste à rechercher et à faire parler les vestiges d'un acte incriminé passé. L'investigateur forensique se pose dans ce cadre comme un lecteur critique des lieux à la recherche de traces devant lui permettre de former son récit, soit l'histoire du délit/crime censé s'être produit. Le challenge de tout investigateur est de pouvoir détecter et reconnaître les traces dites pertinentes pour fournir des indices forensiques à buts d'enquête et de renseignement. Inspirée par un tel constat, la présente recherche pose au coeur de ses réflexions les questions suivantes : Qu'est-ce qu'une trace pertinente ? Et, comment fait le forensicien pour déterminer qu'il y fait face ? L'intérêt de tels questionnements se comprend dans la volonté de définir une dimension souvent utili- sée en science forensique, mais encore jamais étudiée dans ses implications et fonctionnements. Cette recherche se lance dans des voies d'étude encore peu explorées en usant d'outils sémiotiques et des pratiques d'enquêtes sociologiques combinés à des traitements statistiques de données. Les résultats sont représentés en suivant une piste sémiotique fortement influencée par les écrits de Peirce et une seconde piste dite empirique où des données d'interventions ont été analysées et des investigateurs forensiques interviewés sur leurs pratiques de travail sur le terrain. Par la piste sémiotique, une vision macroscopique du processus de signification de la trace en élément pertinent est représentée. La trace est perçue sous la forme de plusieurs signes dont la signification est codifiée culturellement. Le raisonnement se formaliserait en trois principales étapes : 1- Quel type de source évoque la trace détectée? 2- Quelle cause/activité est à l'origine de cette source dans le contexte précis du cas ? 3- Quelle histoire peut être racontée à partir de ces observations ? Cette dernière étape consiste à raisonner en créant des hypothèses devant expliquer la présence de la trace détectée suite à une activité posée comme étant en lien avec le cas investigué. Pour valider ces hypothèses, cela dépendrait de leur capacité à répondre à une règle, celle de la pertinence. Cette dernière étape consiste en la symbolisation de la pertinence. La règle se composerait de deux points : la reconnaissance de la pertinence factuelle (le lien entre la trace et le cas est-il reconnu dans l'hypothèse fournie?) et la pertinence appropriée (quel est l'investissement à fournir dans la collecte et l'exploitation de la trace pour le bénéfice attendu au niveau de l'investigation/renseignement?). Tout ce processus de signification se base sur des observations et un raisonnement conjectural soumis à de nombreuses influences. Dans cette étude, la pertinence en science forensique se formalise sous les traits d'une dimension conventionnelle, symbolisée, conditionnée par le contexte, la pratique de l'investigateur forensique et la culture du milieu ; en somme cette recherche avance que la pertinence est le fruit d'une interaction entre des paramètres d'ordre situationnel, structurel (ou organisationnel) et individuel. Garantir la détection, la collecte et l'exploitation des traces pertinentes sur les lieux dépend de la connaissance et d'une culture maîtrisées par le forensicien. Dans l'étude du rapport trace pertinente-investigateur forensique, la présente recherche pose le modèle SFE comme une carte conceptuelle illustrant trois grands axes d'acquisition de la connaissance et de la culture forensiques intervenant dans la recherche et l'évaluation de la trace pertinente. Par l'analyse des données d'in- terventions et des entretiens, le rapport entre ces trois paramètres et la pertinence a été mis en évidence. S, pour savoir, incarne un rapport à la connaissance immédiate pour se faire une représentation d'une réalité à un instant donné, un point important pour une pertinence qui se comprend dans un contexte. F, pour formation, se conçoit dans son rapport à la pertinence via cette culture qui tend à s'institutionnaliser (soit une connaissance théorique). Quant au paramètre E, pour expérience, il se comprend dans son rapport à la pertinence dans cet ajustement des stratégies d'intervention (soit une connaissance pratique) de chaque praticien ayant modulé leur travail au regard des succès et échecs enregistrés cas après cas. F et E formeraient la bibliothèque de ressources permettant le processus de reconnaissance sémiotique et S assurerait la contextualisation nécessaire pour poser le raisonnement et formuler les hypothèses explicatives pour les traces détectées et questionnées dans leur pertinence. Ce travail démontre que la pertinence n'est pas absolue. Elle est temporelle et contextuelle, c'est une dimension conventionnelle relative et interprétée qui se doit d'être discutée. C'est là que repose toute la problématique de la signification de ce qui est pertinent pour chaque participant du processus d'investigation. En proposant une lecture par étapes du processus de signification depuis la trace à l'indice, l'étude vise à offrir une compréhension plus poussée du raisonnement et de son fonctionnement pour renforcer la formation des intervenants forensiques. Cette recherche présente ainsi un ensemble d'outils critiques à portée tant pédagogiques que pratiques pour la gestion des lieux tout en identifiant des influences-clé jouées par des dimensions individuelles, structurelles et situationnelles.

Triodothyronine enhancement of neuronal differentiation in aggregating fetal rat brain cells cultured in a chemically defined medium.

Triiodothyronine (30 nM) added to serum-free cultures of mechanically dissociated re-aggregating fetal (15-16 days gestation) rat brain cells greatly increased the enzymatic activity of choline acetyltransferase and acetylcholinesterase throughout the entire culture period (33 days), and markedly accelerated the developmental rise of glutamic acid decarboxylase specific activity. The enhancement of choline acetyltransferase and acetylcholinesterase specific activities in the presence of triiodothyronine was even more pronouned in cultures of telencephalic cells. If triiodothyronine treatment was restricted to the first 17 culture days, the level of choline acetyltransferase specific activity at day 33 was 84% of that in chronically treated cultures and 270% of that in cultures receiving triiodothyronine between days 17 and 33, indicating that relatively undifferentiated cells were more responsive to the hormone. Triiodothyronine had no apparent effect on the incorporation of [3H]thymidine at day 5 or on the total DNA content of cultures, suggesting that cellular differentiation, rather than proliferation was affected by the hormone. Our findings in vitro are in good agreement with many observations in vivo, suggesting that rotation-mediated aggregating cell cultures of fetal rat brain provide a useful model to study thyroid hormone action in the developing brain.

Functional impact of genetic variation on gene expression

Numerous links between genetic variants and phenotypes are known and genome-wide association studies dramatically increased the number of genetic variants associated with traits during the last decade. However, how changes in the DNA perturb the molecular mechanisms and impact on the phenotype of an organism remains elusive. Studies suggest that many traitassociated variants are in the non-coding region of the genome and probably act through regulation of gene expression. During my thesis I investigated how genetic variants affect gene expression through gene regulatory mechanisms. The first chapter was a collaborative project with a pharmaceutical company, where we investigated genome-wide copy number variation (CNVs) among Cynomolgus monkeys (Macaca fascicularis) used in pharmaceutical studies, and associated them to changes in gene expression. We found substantial copy number variation and identified CNVs linked to tissue-specific expression changes of proximal genes. The second and third chapters focus on genetic variation in humans and its effects on gene regulatory mechanisms and gene expression. The second chapter studies two human trios, where the allelic effects of genetic variation on genome-wide gene expression, protein-DNA binding and chromatin modifications were investigated. We found abundant allele specific activity across all measured molecular phenotypes and show extended coordinated behavior among them. In the third chapter, we investigated the impact of genetic variation on these phenotypes in 47 unrelated individuals. We found that chromatin phenotypes are organized into local variable modules, often linked to genetic variation and gene expression. Our results suggest that chromatin variation emerges as a result of perturbations of cis-regulatory elements by genetic variants, leading to gene expression changes. The work of this thesis provides novel insights into how genetic variation impacts gene expression by perturbing regulatory mechanisms. -- De nombreux liens entre variations génétiques et phénotypes sont connus. Les études d'association pangénomique ont considérablement permis d'augmenter le nombre de variations génétiques associées à des phénotypes au cours de la dernière décennie. Cependant, comprendre comment ces changements perturbent les mécanismes moléculaires et affectent le phénotype d'un organisme nous échappe encore. Des études suggèrent que de nombreuses variations, associées à des phénotypes, sont situées dans les régions non codantes du génome et sont susceptibles d'agir en modifiant la régulation d'expression des gènes. Au cours de ma thèse, j'ai étudié comment les variations génétiques affectent les niveaux d'expression des gènes en perturbant les mécanismes de régulation de leur expression. Le travail présenté dans le premier chapitre est un projet en collaboration avec une société pharmaceutique. Nous avons étudié les variations en nombre de copies (CNV) présentes chez le macaque crabier (Macaca fascicularis) qui est utilisé dans les études pharmaceutiques, et nous les avons associées avec des changements d'expression des gènes. Nous avons découvert qu'il existe une variabilité substantielle du nombre de copies et nous avons identifié des CNVs liées aux changements d'expression des gènes situés dans leur voisinage. Ces associations sont présentes ou absentes de manière spécifique dans certains tissus. Les deuxième et troisième chapitres se concentrent sur les variations génétiques dans les populations humaines et leurs effets sur les mécanismes de régulation des gènes et leur expression. Le premier se penche sur deux trios humains, père, mère, enfant, au sein duquel nous avons étudié les effets alléliques des variations génétiques sur l'expression des gènes, les liaisons protéine-ADN et les modifications de la chromatine. Nous avons découvert que l'activité spécifique des allèles est abondante abonde dans tous ces phénotypes moléculaires et nous avons démontré que ces derniers ont un comportement coordonné entre eux. Dans le second, nous avons examiné l'impact des variations génétiques de ces phénotypes moléculaires chez 47 individus, sans lien de parenté. Nous avons observé que les phénotypes de la chromatine sont organisés en modules locaux, qui sont liés aux variations génétiques et à l'expression des gènes. Nos résultats suggèrent que la variabilité de la chromatine est due à des variations génétiques qui perturbent des éléments cis-régulateurs, et peut conduire à des changements dans l'expression des gènes. Le travail présenté dans cette thèse fournit de nouvelles pistes pour comprendre l'impact des différentes variations génétiques sur l'expression des gènes à travers les mécanismes de régulation.