Aerospace Engineering / The aim of this thesis is to explore the high strain rate behavior of metallic specimens using electromagnetic inductive loading as the means to inflict the required high strain rate deformation on laboratory scale specimens, allowing for controlled, repeatable experiments to be performed. Three separate experiments were designed and performed, using helical and spiral coils as the sources of radial and unidirectional loading. The first experiment evaluated the effect of applying a polymer coating on 30.5 mm diameter, Al 6061- O tube samples, in two lengths, 18 and 36 mm. The expanding tube experiment was used to apply a radial loading on the specimens and record the event. Several optical techniques were then used to evaluate the behavior of the samples. Coatings of polyurea and polycarbonate were used. It was observed that the polycarbonate coating seemed to have a more profound effect on the behavior of the metal, by applying a larger restraining pressure on the tube surface during the expansion process, and thereby modifying the stress state of the specimen. The second experiment looked to design an experimental arrangement to test the plane strain, high strain rate behavior of Al 6061-O tubes of different lengths. A 112 mm long solenoid was designed and manufactured, and testing was performed on 30.5 mm diameter Al 6061-O tubes in lengths of 50, 70 and 90 mm. It was observed that the coil behaved similar to shorter ones at low voltages and that the longer the specimen used, the more its deformation path approached a plane strain condition. Finally, a third experiment was performed to develop an experiment to accelerate a plate to high linear velocities, as a means to evaluate the use of a flat spiral coil as the driver for future experiments based upon electromagnetic inductive loading. A prototype coil was manufactured and installed into a converted expanding tube experimental setup. Three samples were tested in several sizes, and materials: aluminum and steel. Speeds in the range of 45 to 251 m/s were obtained, validating the apparatus as a viable method to provide a unidirectional loading. / text

Nesse trabalho, são investigadas as propriedades dinâmicas de modelos da mecânica estatística na criticalidade. Inicialmente, trabalhando com modelos de spin e utilizando os conceitos de persistência global e de dimensão anômala da magnetização inicial, mostramos que o modelo de Baxter-Wu não está na mesma classe de universalidade dos modelos de Potts com quatro estados e de Ising com interação de três spins em uma direção, todos bidimensionais. Na segunda parte da tese, estudamos o fenômeno de crescimento da superfície gerada pela deposição segundo as regras que definem os autômatos celulares probabilísticos propostos por Grassberger (modelos A e B). Esses dois autômatos não pertencem à classe de universalidade de Domany-Kinzel e apresentam novos expoentes críticos, cuja origem se deve à conservação de paridade. Determinamos o expoente de crescimento beta w, válido em tempos curtos, assim como os outros expoentes críticos associados ao crescimento de superfície (alfa e z). Nossas estimativas se comparam bem com os resultados obtidos a partir de razões de inteiros propostas por Jensen para os expoentes beta, ni paralelo e ni perpendicular. Finalmente, investigamos a transição de fase entre o estado helicoidal e o estado desordenado (random coil) da polialanina e do fragmento peptídico PTH(1-34), que corresponde aos resíduos 1 a 34 da região aminoterminal do hormônio das paratireóides. Nosso cálculo, que leva em conta as interações entre todos os átomos da molécula, está baseado em uma abordagem de tempos curtos. Os resultados dessa análise indicam que a transição helix-coil das polialaninas e do PTH(1-34) é de segunda ordem e apontam para uma classe de universalidade para a transição helix-coil em homopolímeros e proteínas (partindo de um estado helicoidal). / In this work we investigated dynamic properties of statistical mechanical models at criticality. At first, using the concepts of global persistence and anomalous dimension of initial magnetization, we showed that the Baxter-Wu model does not belong to the same universality class as 4-state Potts model and Ising with multispin interaction in one direction. In the sequence, we studied the roughening behavior generated by deposition governed by rules defined by probabilistic cellular automata proposed by Grassberger (A and B models). Those models are known do not belong to the Domany-Kinzel universality class. They are characterized by different exponents which are related to the parity conserving (PC). We estimated the growth exponent beta w, in short-time regimen, such as, other critical exponents associated to the surface growth (alpha and z). Our results are in good agreement with those expected for parity conserving universality class. At last we studied the phase transition between the completely helical state and the random coil of the polyalanine, such as, for the 34-residue human parathyroid fragment PTH(1-34). Our short-time simulations of the helix-coil transition are based on a detailed all-atom representation of proteins. The results indicate that helix-coil transition in polyalanine and PTH(1-34) is a second-order phase transition and suggest a universality class to the helix-coil transition in homopolymer and (helical) proteins.

This thesis deals with measurement method pulsed quantities of electromagnetic field. This thesis was focused on methods, which make use of optic effect. Specially Faraday effect, Pockels effect and magneto-optic Kerr effect. Next it was focused on method makes use of Rogowski coil. At the end was shortly noticed about method makes use of calorimetric sensor. For experimentally realized has been chosen Faraday Magneto-optic Effect. In optic laboratory has been realized absolute methods and different methods making use of Wollaston polarizer on pneumatic optical desk. The main advantage of this method is the capability to measure the high frequency and high current signals. Next in this thesis has been written theory about Rogowski coil. This method has been experimentally realized in laboratory.

The main aim of this thesis is design air-conditioning system and optimization of indoor climate by solar shading. In theoretical part I focused on dividing and description off different types of solar shading. In the next part I designed suitable air-conditioning system for the open space office. The last part is first concentrating on the simulation of the indoor climate in software BSim2014. And then on evaulation of the appropriate solar shading system, whick I can use in the building.

Coiled-coil motifs are part of 5–10 % of the eukaryotic proteome and are involved in important cellular processes such as membrane trafficking, chromosome segregation or mechanosensing. Their canonical form is well understood and based on a heptad repeat with hydrophobic amino acids at positions 1 and 4. A sequence of these peptides folds into an α-helix and two, or more, of these helices bind together by winding around each other, covering up the hydrophobic residues and giving rise to the coiled-coil structure. Coiled-coil proteins appearing in nature do, however, deviate from this form by introducing discontinuities into the heptad repeat. The effect of these imperfections on the structure is only known for few cases and not generally understood or predictable. The additional impact of these discontinuities on the dynamic function of coiled-coil domains is unknown altogether. Here, in order to tackle these questions, the adhesive forces between the α-helices are studied in single-molecule experiments. To measure these small forces (∼ pN) with a high spatial and temporal resolution, a dual-trap optical tweezers setup was constructed. Special emphasis was put on realizing the required high resolution, a large degree of automation and versatility during the building process. The instrument’s performance was assessed by recording force-extension curves of DNA yielding results for the molecular parameters persistence length and stretch modulus in good agreement with those found in the literature. Additionally, the Allan deviation was computed for different configurations of beads and a high stability and resolution of the instrument was found with optimal performance on the time scale of 1–10 s. Optical tweezers require calibration to accurately measure forces. To this end, generally a scheme is used that leverages the Brownian motion of a trapped object in the harmonic potential, created by the laser focus, to determine the parameters required to convert the analog voltage signal to distances and forces. However, this approach requires prior knowledge of the bead’s drag coefficient. A method was suggested previously that allows to measure this parameter by exciting the trapped bead through an external fluid flow and observing its response. Yet, this scheme was proposed for single-trap devices only. The precision and versatility of the new instrument was increased by extending this technique to work with two traps and implementing it in the apparatus. To this aim, the underlying equations of a trapped bead’s motion were modified to include hydrodynamic interactions between the objects resulting from the external fluid flow. It was found that a single multiplicative factor is sufficient to correct the calibration results for the hydrodynamic effects and ensure precise calibration. The drag coefficient of several beads yielded the same result for a single and two traps within the measurement error thus confirming the validity of the method. The newly built instrument was then used to study the coiled-coil protein early endosome antigen 1 (EEA1). This 200 nm long homodimer was shown to undergo an entropic collapse upon binding a small GTPase at the N-terminus. For further investigations of this effect and the adhesives forces at play, an experiment was designed here to unzip the two α-helices of the protein. To this end, DNA handles were attached to each of the two helices using a sortase A based ligation reaction as force moderators and first optical tweezers experiments were performed with the protein-DNA chimera. Thus, the necessary tools for unzipping assays of EEA1 are now at hand to further research the entropic collapse process. To summarize, a dual-trap optical tweezers setup was built, the calibration routine extended and realized in a more precise way and the instrument was used to investigate binding energies of EEA1 α-helices. / Coiled-Coil Strukturmotive sind in 5–10 % aller Proteine von Eukaryoten vertreten und wichtiger Teil zellulärer Prozesse wie Membrantransport, Segregation von Chromosomen oder Mechanoperzeption. Ihre grundlegende Struktur besteht aus dem sogenannten Heptadenmuster, einer Sequenz aus sieben Aminosäuren mit hydrophoben Molekülen an Position eins und vier. Eine Reihe dieser Muster kann sich zu einer α-Helix falten und zwei, oder mehr, solcher Helices sich umeinander winden, um die hydrophoben Moleküle abzuschirmen. Das Ergebnis ist eine Coiled-Coil- oder Doppelwendelstruktur. Natürlich vorkommende Coiled-Coil Proteine weichen jedoch durch Fehlstellen im Heptadenmuster von dieser kanonischen Form ab. Die Auswirkung dieser Störstellen auf die Struktur des gesamten Moleküls ist bisher nur für einige wenige Fälle untersucht und nicht allgemein vorstanden oder vorhersagbar. Der zusätzliche Einfluss dieser Fehlstellen auf die Funktion und dynamischen Prozesse solcher Proteine ist gänzlich unbekannt. Um diesen Fragen nachzugehen werden hier die Bindungskräfte zwischen den α-Helices in Einzelmolekülstudien untersucht. Um diese winzigen Kräfte (∼ pN) mit hoher räumlicher und zeitlicher Auflösung untersuchen zu können, wurde im Rahmen der vorliegenden Arbeit eine optische Doppelfalle konstruiert. Besonderes Augenmerk lag dabei auf dem Erreichen der erforderlichen Auflösung, einem hohen Grad an Automatisierung und der vielfälting Einsatzfähigkeit des Instruments. Die Leistungsfähigkeit dieses Kraftmikroskops wurde besonders durch zwei Experimente überprüft und sichergestellt. Zum einen wurden DNA Moleküle gedehnt und die Polymerparameter Persistenzlänge und Zugmodul gemessen, welche sehr gut mit veröffentlichten Referenzwerten übereinstimmten. Zum anderen wurde die Allan Schwankung für verschiedene experimentelle Konfigurationen von mikroskopischen Kugeln ermittelt, was eine hohe Stabilität und Auflösung des Gerätes, mit optimaler Leistung bei Mittelung auf Zeitskalen von 1–10 s, bestätigte. Optische Fallen müssen kalibriert werden, um Kräfte exakt messen zu können. Im Allgemeinen kommt dafür ein Verfahren zum Einsatz, welches die brownsche Bewegung eines gefangenen Objektes im harmonischen Potential des Laserfokus ausnutzt. Aus diesen Fluktuationen werden die benötigten Parameter ermittelt, um das gemessene analoge Spannungssignal in Abstände und Kräfte umzuwandeln. Dieser Ansatz erfordert jedoch die Kenntnis des Reibungskoeffizienten des gehaltenen Objektes, meist einer mikroskopischen Kugel. Daher wurde eine Methode vorgeschlagen, die durch ein oszillierendes Flussfeld eine zusätzliche Bewegung der Kugel anregt aus welcher der Reibungskoeffizient bestimmt werden kann. Dieses Vorgehen reduziert die im vornherein benötigten Informationen, wurde jedoch nur für eine einzelne optische Falle entwickelt. Der Ansatz wurde in dieser Arbeit erweitert, indem die zu zugrundeliegenden Bewegungsgleichungen einer gefangenen Kugel um hydrodynamische Wechselwirkungen zwischen mehreren Objekten ergänzt und die Kalibrationparameter basierend darauf hergeleitet wurden. Im Ergebnis konnte gezeigt werden, dass ein einzelner multiplikativer Faktor ausreicht, um die Hydrodynamik zu berücksichtigen und die exakte Kalibration des Instruments sicherzustellen. Dieses Vorgehen wurde überprüft, indem der Reibungskoeffizient einer einzelnen oder mehrerer mikroskopischer Kugeln gleichzeitig durch Anlegen eines externen Flussfeldes gemessen wurde. Die Ergebnisse stimmen im Rahmen der Messgenauigkeit überein und bestätigen damit den gewählten Ansatz. Das neu implementierte Kraftmikroskop wurde im Folgenden eingesetzt, um das Coiled-Coil Protein Early Endosome Antigen 1 (EEA1) zu erforschen. Dieser 200 nm lange Homodimer kollabiert aufgrund entropischer Kräfte sobald eine kleine GTPase an seinen N-Terminus bindet. Um diesen Effekt und die wirkenden Bindungskräfte besser zu verstehen, wurde hier ein Experiment entwickelt bei dem die beiden α-Helicen auseinandergezogen werden. Dazu wurde mittels einer Sortase A basierten Ligationsreaktion an jede Helix ein DNA-Stück gebunden, über welches Kräfte auf das Molekül übertragen werden können. Erste Experimente wurden mit der optischen Doppelfalle und dieser Protein-DNA Chimäre durchgeführt. Somit sind alle benötigten Werkzeuge zum weiteren Studium des entropischen Kollapses von EEA1 verfügbar, indem die Bindungskräfte der α-Helicen untersucht werden. Zusammenfassend wurde eine hoch auflösende Doppelfalle konstruiert, die Kalibrationsmethode weiterentwickelt und verfeinert und das Kraftmikroskop zur Erforschung der Bindungskräfte der α-Helicen von EEA1 eingesetzt.

Specifically binding ions induce the transition of anionic polyacrylate coils from extended conformation to collapsed globules passing through a cascade of intermediate states when solution conditions approach the L-type precipitation threshold. It is the conformation of these intermediate states on surfaces and in solution which is at the focus of this thesis. In comparing the surface and solution conformations of intermediate states, we were able to qualitatively and quantitatively underline the effects of sample history. Two types of quantitative comparisons have been emphasized. In real space, the radius of gyration values of adsorbed molecules have been evaluated incorporating fully the x, y and z axes. These values have been compared with radius of gyration values of the very same sample solution obtained using SLS. In reciprocal space, a novel image processing protocol has been used to generate the 2D form factor curve wherein the correlation maxima have been compared with corresponding maxima obtained for the very same sample solution using small angle scattering techniques like the SANS. The influence of bivalent ions, respectively, Strontium, Lead and Calcium, on the shape of polyacrylate coils is studied. In the last case, temperature has been introduced as a secondary parameter to shed further light on the mechanism by which polyelectrolytebivalent ion complexation takes place. Both scattering and AFM experiments reveal formation of necklace-like structures as intermediates for NaPA-Sr2+ system. Since the mol. wt. of the NaPA coils used was relatively large in this case, adsorption on mica surfaces was strong. Under such conditions, the molecules undergo a z collapse upon flux drying but do not get altered in x and y directions. The ratio Rg(AFM)/Rg(SLS) was found to be in the range 0.7-0.9. The remaining (insignificant) differences in the Rg Abstract ii values arise due to the fact that AFM gives the square root of number averaged mean squared radius of gyration while SLS gives the square root of z-averaged mean squared radius of gyration. The differences in radius of gyration values observed in solution and on surfaces were more prominent for NaPA-Pb2+ system. Again, although both scattering and AFM reveal necklace-like structures as intermediates, the ratio Rg(AFM)/Rg(SLS) was now found to be nearly 0.6. The fact that Rg(AFM) is the square root of number averaged and Rg(SLS) is the square root of z-averaged mean squared radius of gyration, alone cannot explain this low value. Since the mol. wt. of NaPA coils used in this case was quite low, adsorption on mica surfaces was weak. Under such conditions, the molecule does not only undergo a z collapse upon flux drying, but also shrinks in the x and y directions due to capillary forces. Finally, with NaPA-Ca2+ system, the picture did not show a one-to-one correspondence between solution and surface conformations at all. In fact, it showed a one-step-ahead correspondence. As already stated, the coil to globule transition was induced by increasing the equilibration temperature from 15°C to 30°C in this case. SANS could not identify any necklace-like intermediates in solution at the equilibration temperature of 15°C while AFM scans at this temperature showed the beginning of formation of pearls. Likewise, at the equilibration temperature of 30°C, SANS could identify necklace-like intermediates in solution with a large majority of dumbbells while AFM scans at this temperature showed a mix of dumbbells, sausage-like structures and globules. Indeed, we were witnessing an accelerated coil to globule transition on surfaces as compared to the situation in solution resulting in a pre-emption in the formation of intermediate states on surfaces. Since the ratio Rg(AFM)/Rg(SLS) (given the square root of number averaged and the square root of z-averaged mean Abstract iii squared values respectively) at the equilibration temperature of 30°C showed a range of 0.7-0.9 indicating strong adsorption of the relatively high mol. wt. NaPA coil on mica surfaces, our suspect were the substrate-sample interaction forces. The AFM scans were therefore analysed with 2D form factor curves, a better protocol when no assumptions about the shape of adsorbed molecules are made a priori, to trace the effects of sample history. The thesis establishes the general utility of AFM to capture the essential features of a collapsing coil which the very coil exhibits in solution. The shape of the coil on surface and in solution may not be exactly the same, yet reveal the same characteristics. The comparative advantages and disadvantages of salt pre-treated mica surfaces and chemically modified mica surfaces have been brought out. Finally, a definitive new insight is gained as regards the mechanism of coil collapse induced by specifically binding ions. The entropic nature of the process as well as the visualized shape of the collapsing intermediates does not support a mechanism along an electrostatically driven shrinking with linear, rod-like arrays of pearls as intermediates. On a molecular level, it is the liberation of water molecules and Na+ ions which promotes binding of bivalent ions to COO- residues. This binding in turn increases the hydrophobicity of the polyacrylate chains. As a consequence, the chains shrink due to an increased propensity for polymerpolymer contacts (and finally precipitate).

Organic coatings are frequently applied to metals, often in circumstances where there is a need to protect the substrate from corrosion or to improve the aesthetic quality of a product. Increasingly, coatings are also expected to provide additional functionality, such as anti-bacterial properties. Concurrent with the need to satisfy ever more demanding specifications, coating formulators are being obliged to comply with stringent environmental legislation. A research area which may benefit the development of coating formulations is that of nanocomposite synthesis, in which the composite additives have at least one dimension in the nanometre size range. The current research has sought to modify a waterborne organic coating primer with two types of nano-sized additives: layered clays and titanium dioxide nanoparticles. Dispersion of the particles at loadings up to 5% w/w was achieved by ball milling and the modified coatings were subjected to numerous tests to ascertain the effect of the additives. Titanium dioxide was found to improve the tribological properties and corrosion resistance of the coatings, while the addition of magnesium aluminium hydroxycarbonate (hydrotalcite) layered clay had a deleterious effect. Some coatings exhibited a cracked morphology, although no correlation between particle loading and the presence of cracks could be determined. A preliminary investigation into substrate preparation was also conducted. This ensured that the degree of cleanliness of the hot-dip galvanized substrates was both satisfactory and consistent.

Experiments in high energy density (HED) plasma physics have become more accessible with the increasing availability of high-intensity pulsed lasers. Extending the experiment parameters to include magnetized HED plasmas requires a field source that can generate fields of order 100 tesla. This dissertation discusses the design and implementation of a pulsed field driver with a designed maximum of 2.2 MA from a 160 kJ capacitor bank. Faraday rotation measurement of 63 T for a 1.0 MA discharge supported Biot-Savart estimates for a single-turn coil with a 1 cm bore. After modification, the field driver generated up to 15 T to magnetize supernova-like spherical blast waves driven by the Texas Petawatt Laser. The presence of the high field suppressed blast wave expansion, and had the additional effect of revealing a cylindrical plasma along the laser axis. / text

Mechanics and biological function of nucleic acids are intimately coupled. The DNA double helix must be opened to allow base pairing of RNA during transcription; RNA must bend and fold in its many cellular functions. Presented in this dissertation are two investigations of mechanical deformations of nucleic acids, conducted with optical tweezers.In the introduction, the mechanical properties of DNA and RNA and their relevance to their cellular functions are introduced, to give the reader context for the results presented in the Chapters 2 and 3. This is followed by an introduction to the theory of semiflexible polymer elasticity. The optical tweezers instruments used in conducting these investigations are then presented, along with calibration procedures and a short introduction to optical trapping physics.Chapter 2 presents an investigation of the effect of downstream DNA tension on initiation by T7 RNA polymerase. A hidden Markov model is fit to force-dependent lifetimes obtained from optical tweezers experiments, allowing us to identify which steps in initiation are force-dependent and estimate rates and transition state distances. We find that 1-2 pN of tension is sufficient to turn o gene expression by causing transcription bubble collapse and destabilizing the bound state. Our force-dependence scheme and estimated transition distances provide independent supportfor the \scrunching" model of initiation.The effects of cation binding and screening on single-stranded helix formation in poly(A) RNA are presented in Chapter 3. Magnesium and calcium bind to poly(A), stabilize the helix, and change its mechanical properties. A new model of helix-coil transitions is presented and used to estimate energetics and mechanical properties.Chapter 4 presents the first fully objective algorithm for use in analyzing the noisy staircaselike data that is often produced by single-molecule fluorescence experiments. A test based on the SIC (BIC) statistic is used in conjunction with a progressive step-placement scheme to locate changepoints (steps) in noisy data. Its performance is compared to other step detection algorithms in use by biophysicists by repeating tests performed in a recent review.Experimental protocols and computer codes used in these investigations are presentedin detail in the appendices.

La polykystose rénale autosomique dominante (PKRAD) est la maladie génétique rénale la plus commune touchant 1/500 personnes. Elle se caractérise principalement par la formation de kystes rénaux dans tous les segments du néphron, entraînant l’insuffisance rénale, et par des manifestations extrarénales kystiques (foie, pancréas, rate) et non-kystiques (anomalies cardiaques, vasculaires et cérébrales). Deux gènes, PKD1 et PKD2, sont responsables de 85 et 15% des cas respectivement. Ces gènes encodent les polycystine-1 (PC-1) et -2 (PC-2) qui forment un complexe à la membrane plasmique et ciliaire des cellules épithéliales rénales. PC-1 est une protéine transmembranaire de 4302 acides aminés possédant un court domaine intracellulaire incluant un motif coiled-coil impliqué dans l’interaction entre PC-1 et PC-2 in-vitro. L’importance du coiled-coil est démontrée par des mutations affectant spécifiquement ce motif chez des patients PKRAD. Le mécanisme pathogénétique responsable de la PKRAD est indéterminé. Chez la souris, la PKRAD se développe suite à l’ablation (Pkd1-/-) ou lors de la surexpression (SBPkd1TAG) de Pkd1, ce qui suggère un effet de dosage. Des anomalies ciliaires sont aussi souvent associées à PKRAD. Mon objectif était de déterminer in-vivo le mécanisme pathogénétique de la polycystine-1 dans le développement des symptômes PKRAD rénaux et extrarénaux et plus spécifiquement, le rôle du motif coiled-coil dans le mécanisme de kystogenèse. Pour ce faire, nous avons généré deux constructions, Pkd1 sauvage (Pkd1TAG) et Pkd1 tronquée de son motif coiled-coil (Pkd1ΔCoiled-coil), par recombinaison homologue à partir du BAC-Pkd1 sauvage comprenant la séquence murine entière de Pkd1. Trois lignées de souris Pkd1TAG générées par microinjection démontrent un niveau d’expression de Pkd1 qui corrèle avec le nombre de copie du transgène (2, 5 et 15 copies). Les souris Pkd1TAG reproduisent la PKRAD en développant des kystes rénaux dans toutes les parties du néphron et des cils primaires plus longs que les contrôles non transgéniques. Les analyses physiologiques supportent que les souris Pkd1TAG développent une insuffisance rénale et démontrent une augmentation du volume urinaire de même qu’une diminution de l’osmolalité, de la créatinine et des protéines urinaires. De plus, les souris Pkd1TAG développent des kystes hépatiques, des anomalies cardiaques associées à des dépôts de calcium et des anévrismes cérébraux. La sévérité du phénotype augmente avec l’expression de Pkd1 appuyant l’hypothèse d’un mécanisme de dosage. Nous avons aussi déterminé que l’expression du transgène Pkd1TAG complémente le phénotype létal-embryonnaire des souris Pkd1-/-. D’autre part, nous avons générés 4 lignées de souris Pkd1ΔCoiled-coil (2 et 15 copies du transgène) dont le nombre de copies corrèle avec le niveau d’expression du transgène. Ces souris Pkd1ΔCoiled-coil, contrairement aux Pkd1TAG de même âge, ne développent pas de kystes et possèdent des cils primaires de longueur normale. Afin d’évaluer le rôle du motif coiled-coil en absence de polycystine-1 endogène, nous avons croisé les souris Pkd1ΔCoiled-coil avec les souris Pkd1-/-. Contrairement aux souris Pkd1-/- qui meurent in-utéro, les souris Pkd1ΔCoiled-coil; Pkd1-/- survivent ~10 à 14 jours après la naissance. Elles démontrent des kystes rénaux et pancréatiques sévères, un retard de croissance et des anomalies pulmonaires. Tous les segments du néphron sont affectés. Mon projet démontre que la surexpression de Pkd1 est un mécanisme pathogénique de la PKRAD tant au niveau rénal qu’extrarénal. De plus, il démontre que le motif coiled-coil est un élément déterminant dans la kystogenèse/PKRAD in-vivo. / Autosomal dominant polycystic kidney disease (ADPKD) is a common genetic disorder affecting 1:500 people worldwide, independently from sex and origin. ADPKD is characterized by formation of large bilateral kidney cysts affecting all segments of the nephron and increasing progressively in size and number leading to end stage renal failure by mid-fifty. Moreover, this systemic disease includes several extrarenal symptoms such as intracranial aneurysms, valvular defects and cysts formation in the liver and the pancreas. PKD1 and PKD2 genes mutations are involved in 85 and 15 % of the clinical cases. PKD genes encode polycystin-1 (PC-1) and -2 (PC-2), which both form a complex at the cell and ciliary membrane of renal epithelial cells. PC-1 is a large transmembrane protein with a small intracellular tail including a coiled-coil motif implicated in PC-1/PC-2 interaction in-vitro. Interestingly, specific mutations affecting the coiled-coil motif cause ADPKD in humans. The pathogenetic mechanism of ADPKD is unknown. In mice, both ablation (Pkd1-/-) or overexpression (SBPkd1TAG) of Pkd1 cause ADPKD, suggesting a dosage model. Ciliary anomalies are also linked to polycystic kidney disease. Herein, we evaluated in-vivo the role of Pkd1 in the development of renal and extrarenal manifestations of ADPKD and more specifically, the role of the coiled-coil motif in cystogenesis. We generated two constructions, wildtype Pkd1 (Pkd1TAG) and coiled-coil deleted Pkd1 (Pkd1ΔCoiled-coil), by homologous recombination from the wildtype Pkd1-BAC comprising the whole Pkd1 murine sequence. Three Pkd1TAG mice lines have been generated by microinjection and show expression patterns correlating with the copy number of the transgene (2, 5 and 15 copy). All Pkd1TAG mice develop renal cysts affecting all nephron segments as in ADPKD and longer primary cilia compared to wildtype mice. Physiologic analysis supports renal failure by increased urinary output and decreased of urinary proteins, osmolality and creatinin levels. Pkd1TAG mice also show cysts in the liver, cardiac and valvular anomalies associated with calcium deposition and cerebral aneurysms. The severity of the phenotype increased with Pkd1 expression suggesting a dosage model. Importantly, the Pkd1TAG transgene rescue embryonic lethality of Pkd1-/- mice. Furthermore, we generated 4 lines of Pkd1ΔCoiled-coil mice of 2 and 15 copies of the transgene correlating also to the level of expression. Compared to age-matched Pkd1TAG, Pkd1ΔCoiled-coil mice develop no cysts and show normal cilia length. To gain more insights on the role of coiled-coil motif in absence of endogenous Pc-1, we mated Pkd1ΔCoiled-coil with Pkd1-/- mice. Compared to the lethal embryonic Pkd1-/- mice, Pkd1ΔCoiled-coil; Pkd1-/- live ~ 10 to 14 days. They show severe renal and pancreatic cysts as well as growth retardation and pulmonary defects. My study demonstrates that Pkd1 overexpression is a pathogenic mechanism to induce ADPKD renal and extrarenal phenotype. Moreover, this work shows that the coiled-coil motif of polycystin-1 is a critical determinant in ADPKD cystogenesis.