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Wildlife diseases often occur under quantifiable and consistent patterns, which can be understood to statistically predict their occurrence and spread across landscapes. Chronic wasting disease (CWD) is a neurodegenerative disease in the deer family Cervidae caused by a prion, a pathogenic and misfolded variant of a naturally occurring protein. Managing and controlling CWD is imperative for conservation of ecologically and economically important cervid species, but unclear transmission mechanisms within landscapes complicate evidence-based management. Gaps of information in the landscape ecology for CWD are particularly pronounced for areas with recent disease emergence and spread, such as within the CWD cluster in the Mid-Atlantic United States. Thus, I identified current gaps in information and sought to fill neglected areas of research, specifically focusing on landscape determinants for CWD occurrence and spread in the state of Virginia. In chapter 2, I conducted a scoping study that collected and synthesized decades of CWD research and identified trends with respect to statistical and mathematical modeling methods used, connectivity within the CWD research community, and the geographic areas from which studies were performed. In chapter 3, I investigated landscape determinants for CWD in Virginia using remote sensing landscape data and an epidemiological dataset from Virginia Department of Wildlife Resources (DWR) using diverse algorithms and model evaluation techniques. Finally, in chapter 4, I modeled landscape connectivity between confirmed CWD cases to examine potential paths and barriers to CWD spread across landscapes. My results indicate that landscape ecology was rarely incorporated throughout CWD's 50+ year history. I provide evidence that remotely-sensed landscape conditions can be used to predict the likelihood of CWD occurrence and connectivity in Virginia landscapes, suggesting plausible CWD spread. I suggest areas of future work by explicitly identifying gaps in CWD research and diagnostic methods from which models are based, and encourage further consideration of host's ecology in modeling. By integrating remotely-sensed data into my modeling framework, the workflow should be easily adaptable to new study areas or other wildlife diseases. / Master of Science / Understanding why diseases occur in some locations and not others can be a critical challenge for disease ecologists. One disease that has received significant attention from the media and scientific community is chronic wasting disease (CWD), which is caused by a misfolded protein called a prion. Virginia Department of Wildlife Resources (DWR) has identified a stark increase in the number of CWD cases since first discovered in 2009, which threatens white-tailed deer populations and a 500 million dollar industry used for conservation of Virginia wildlife species. Previous research found that CWD does not occur randomly on the landscape, but otherwise little is known about the landscape ecology of CWD. To provide insight on Virginia's CWD outbreak, I assessed methods used to investigate other CWD outbreaks in both space and time. Also, I used landscape data collected from satellites and data from CWD cases in Virginia, and applied statistical tools to identify patterns in the landscape that were linked with CWD cases. My results suggest that landscapes were rarely examined to understand CWD, and instead, researchers focused on understanding how populations will respond to the disease. I also provide evidence that, at least in Virginia, researchers can use satellite information with disease data to predict CWD on the landscape and estimate its spread. This information can be used by wildlife managers to control the disease. For example, disease surveillance can be increased in areas where CWD has been predicted, or herd sizes can be reduced in areas likely to promote disease spread. This information could also be used to tailor wildlife health regulations aimed to minimize the risk of other deer populations acquiring the disease. Ultimately, the landscape plays an important role in CWD, but research on this topic is limited; therefore, additional research is needed to understand and eventually control this disease affecting ecologically and culturally important game species.

Les prions sont des agrégats amyloïdes infectieux. Le prion [Het-s] de Podospora anserina est un des prions le mieux caractérisé. Le prion [Het-s] est impliqué dans l’incompatibilité végétative – un processus biologique qui a lieu au cours des anastomoses entre des souches génétiquement différentes. Quand une souche [Het-s] fusionne avec une souche exprimant l’allèle alternatif du gène het-s – l’allèle het-S – une réaction de mort cellulaire programmée est déclenchée. Les deux protéines diffèrent de 13 acides aminés et partagent une architecture en deux domaines ; un domaine globulaire en N-terminal nommé HeLo et un domaine PFD (Prion Forming Domain) en C-terminal. Il a été établi qu’en présence des fibres amyloïdes de [Het-s], la protéine HET-S agit en ‘pore-forming’ toxine : la transconformation du PFD de HET-S par les fibres amyloïdes du [Het-s] active le domaine HeLo de HET-S et entraîne la mort cellulaire. Afin de mieux caractériser les propriétés du repliement β-solénoïde du prion [Het-s], nous avons entrepris l’exploration in vivo des relations structure-fonction de ce repliement par une approche d’alanine scanning. Au cours de nos recherches pour des homologues de HET-S/s, nous avons identifié un partenaire fonctionnel de HET-S – une protéine appelée NWD2. NWD2 est une protéine STAND et partage une séquence homologue (3-23) au PFD de HET-S/s. Les protéines STAND, après la reconnaissance d’un ligand, forment des plateformes oligomériques pour transduire le signal. Des analyses génomiques in silico réalisées dans plusieurs génomes fongiques nous ont amené à proposer que la transduction du signal via une protéine STAND à repliement amyloïde est un mécanisme ancien et conservé chez les champignons. Dans ce contexte nous avons identifié deux nouveaux motifs PFD putatifs – σ et PP. En soumettant à l’épreuve notre hypothèse, nous avons d’abord démontré que NWD2 interagit avec HET-S/s en fonction d’un ligand spécifique in vivo et l’interaction est dépendante de la séquence NWD2(3-23) homologue au PFD de HET-S/s. Nous avons ensuite exploré le motif PP associé à un domaine HeLo-like (HELL) dans le génome de Chaetomium globosum. En démontrant la nature amyloïde et prion-like du motif PP ainsi que l’analogie fonctionnelle entre ce motif et le PFD de HET-S/s in vivo nous avons apporté des arguments supplémentaires en faveur de l’implication des repliements amyloïdes dans la transduction du signal chez les champignons filamenteux. / Prions are infectious amyloid aggregates. Podospora anserina’s [Het-s] is one of the best characterized fungal prions with a remarkably high prevalence in wild populations. [Het-s] functions in vegetative incompatibility - a biological process occurring during anastomosis between two genetically incompatible strains. When an [Het-s] prion infected strain fuses with a strain expressing the alternative allelic variant of the het-s locus – het-S – a cell death reaction of the heterokaryon occurs. Differing by 13 amino acids both proteins shares two domain architecture; a globular N-terminal domain called HeLo and a C-terminal Prion Forming Domain (PFD). It has been demonstrated that in presence of [Het-s] amyloid fibers HET-S turns into a pore-forming toxin: transconformation of the HET-S PFD by [Het-s] fibers triggers the refolding of the HET-S HeLo domain, inducing the cell death reaction. In an attempt to better characterize the conserved features of the [Het-s] β-solenoid fold we have used a mutational alanine scanning approach and explored in vivo the existing relations between structure and prion functions of [Het-s]. During our quest for new distant homologues of HET-S/s, we have identified a functional partner of HET-S toxin called NWD2. NWD2 is a STAND protein and shares a homology sequence (3-23) in the HET-S/s PFD. STAND proteins form signal transducing hubs through oligomerization upon ligand recognition. Several in silico analysis in various fungal genomes led us to propose that signal transduction via a STAND protein using an amyloid prion-like fold is a general widespread mechanism in fungi. In that context, we have proposed two novel putative PFD motifs called σ and PP. Testing experimentally our hypothesis, we have first demonstrated that NWD2 interacts with HET-S/s upon ligand recognition in vivo and the interaction is dependant of the NWD2(3-23) region. We have then explored the newly identified putative prion domain PP, associated to a Helo-like domain (termed HELL) from the filamentous fungus Chaetomium globosum. By demonstrating the amyloid, prion-like nature of the PP motif and the functional analogy between PP and HET-S/s PFD domain in vivo, we expose further evidences supporting the implication of amyloid folds in signal transduction in filamentous fungi.

Os príons são proteínas que causam um grupo de doenças neurodegenerativas invariavelmente fatais, sendo uma das mais conhecidas a encefalopatia espongiforme bovina (ou doença da vaca louca). A proteína príon celular (PrPc), rica em estrutura α-helicoidal, sofre uma mudança na sua estrutura secundária produzindo a proteína patológica (PrPSc; o príon) na qual prevalecem folhas-β. Devido a falta de dados de estruturais de alta resolução dos príons, simulações de DM podem ser particularmente úteis para estudar o redobramento de PrP. Estudos experimentais e computacionais, descritos na literatura, indicam que a utilização de pH ácido é capaz de criar alguma instabilidade estrutural, produzindo um ganho de estrutura-β na região N-terminal antes desestruturada. Este trabalho se propõe a investigar computacionalmente as mudanças estruturais na proteína príon celular do hamster Sírio induzidas por alteração de pH. Para isso, foi avaliada a influência do uso de diferentes campos de força (GROMOS, AMBER e OPLS), diferentes estados de protonação dos resíduos de histidina, diferentes condições iniciais e diferentes métodos de cálculo de interações eletrostáticas de longo alcance (GRF e SPME). A partir da evolução temporal das estruturas secundárias, foi observada uma forte dependência dos resultados com o uso de diferentes parâmetros de simulação. De fato, a tendência de pH descrita na literatura não foi claramente observada neste trabalho. Isso pode estar associado com a necessidade de se investir mais em múltiplas simulações de dinâmica molecular para quantificar com maior precisão o comportamento estrutural dos fragmentos protéicos em cada pH de estudo. / Prions are proteins that cause a group of invariably fatal neurodegenerative diseases, one of the most known being bovine spongiform encephalopathy (or mad cow disease). The cellular prion protein (PrPc), rich in α-helical structure, undergoes a change in its secondary structure producing the pathological protein (PrPSc, the prion) in which β-sheet structure prevails. Because of the lack of high-resolution prion structural data, MD simulations can be particularly useful to study PrP misfolding. Experimental and computational studies, described in literature, indicate that the use of low pH is capable to create some structural instability, producing a gain of β-structure content in the otherwise unstructured N-terminal region. This work aims to investigate computationally structural changes in the cellular prion protein of Syrian hamster induced by pH change. For this, we evaluated the influence of different force fields (GROMOS, AMBER and OPLS), different protonation states of histidine residues, different initial conditions and different methods for calculating long-range electrostatic interactions (GRF and SPME). From the time evolution of the secondary structures, we observed a strong dependence on the simulation parameters. In fact, the pH tendency described in literature was not clearly observed in this work. It may be associated with the need to invest more in multiple molecular dynamics simulations to quantify more accurately the structural behavior of the protein fragments in each pH study.

Os príons são proteínas que causam um grupo de doenças neurodegenerativas invariavelmente fatais, sendo uma das mais conhecidas a encefalopatia espongiforme bovina (ou doença da vaca louca). A proteína príon celular (PrPc), rica em estrutura α-helicoidal, sofre uma mudança na sua estrutura secundária produzindo a proteína patológica (PrPSc; o príon) na qual prevalecem folhas-β. Devido a falta de dados de estruturais de alta resolução dos príons, simulações de DM podem ser particularmente úteis para estudar o redobramento de PrP. Estudos experimentais e computacionais, descritos na literatura, indicam que a utilização de pH ácido é capaz de criar alguma instabilidade estrutural, produzindo um ganho de estrutura-β na região N-terminal antes desestruturada. Este trabalho se propõe a investigar computacionalmente as mudanças estruturais na proteína príon celular do hamster Sírio induzidas por alteração de pH. Para isso, foi avaliada a influência do uso de diferentes campos de força (GROMOS, AMBER e OPLS), diferentes estados de protonação dos resíduos de histidina, diferentes condições iniciais e diferentes métodos de cálculo de interações eletrostáticas de longo alcance (GRF e SPME). A partir da evolução temporal das estruturas secundárias, foi observada uma forte dependência dos resultados com o uso de diferentes parâmetros de simulação. De fato, a tendência de pH descrita na literatura não foi claramente observada neste trabalho. Isso pode estar associado com a necessidade de se investir mais em múltiplas simulações de dinâmica molecular para quantificar com maior precisão o comportamento estrutural dos fragmentos protéicos em cada pH de estudo. / Prions are proteins that cause a group of invariably fatal neurodegenerative diseases, one of the most known being bovine spongiform encephalopathy (or mad cow disease). The cellular prion protein (PrPc), rich in α-helical structure, undergoes a change in its secondary structure producing the pathological protein (PrPSc, the prion) in which β-sheet structure prevails. Because of the lack of high-resolution prion structural data, MD simulations can be particularly useful to study PrP misfolding. Experimental and computational studies, described in literature, indicate that the use of low pH is capable to create some structural instability, producing a gain of β-structure content in the otherwise unstructured N-terminal region. This work aims to investigate computationally structural changes in the cellular prion protein of Syrian hamster induced by pH change. For this, we evaluated the influence of different force fields (GROMOS, AMBER and OPLS), different protonation states of histidine residues, different initial conditions and different methods for calculating long-range electrostatic interactions (GRF and SPME). From the time evolution of the secondary structures, we observed a strong dependence on the simulation parameters. In fact, the pH tendency described in literature was not clearly observed in this work. It may be associated with the need to invest more in multiple molecular dynamics simulations to quantify more accurately the structural behavior of the protein fragments in each pH study.

Os príons são proteínas que causam um grupo de doenças neurodegenerativas invariavelmente fatais, sendo uma das mais conhecidas a encefalopatia espongiforme bovina (ou doença da vaca louca). A proteína príon celular (PrPc), rica em estrutura α-helicoidal, sofre uma mudança na sua estrutura secundária produzindo a proteína patológica (PrPSc; o príon) na qual prevalecem folhas-β. Devido a falta de dados de estruturais de alta resolução dos príons, simulações de DM podem ser particularmente úteis para estudar o redobramento de PrP. Estudos experimentais e computacionais, descritos na literatura, indicam que a utilização de pH ácido é capaz de criar alguma instabilidade estrutural, produzindo um ganho de estrutura-β na região N-terminal antes desestruturada. Este trabalho se propõe a investigar computacionalmente as mudanças estruturais na proteína príon celular do hamster Sírio induzidas por alteração de pH. Para isso, foi avaliada a influência do uso de diferentes campos de força (GROMOS, AMBER e OPLS), diferentes estados de protonação dos resíduos de histidina, diferentes condições iniciais e diferentes métodos de cálculo de interações eletrostáticas de longo alcance (GRF e SPME). A partir da evolução temporal das estruturas secundárias, foi observada uma forte dependência dos resultados com o uso de diferentes parâmetros de simulação. De fato, a tendência de pH descrita na literatura não foi claramente observada neste trabalho. Isso pode estar associado com a necessidade de se investir mais em múltiplas simulações de dinâmica molecular para quantificar com maior precisão o comportamento estrutural dos fragmentos protéicos em cada pH de estudo. / Prions are proteins that cause a group of invariably fatal neurodegenerative diseases, one of the most known being bovine spongiform encephalopathy (or mad cow disease). The cellular prion protein (PrPc), rich in α-helical structure, undergoes a change in its secondary structure producing the pathological protein (PrPSc, the prion) in which β-sheet structure prevails. Because of the lack of high-resolution prion structural data, MD simulations can be particularly useful to study PrP misfolding. Experimental and computational studies, described in literature, indicate that the use of low pH is capable to create some structural instability, producing a gain of β-structure content in the otherwise unstructured N-terminal region. This work aims to investigate computationally structural changes in the cellular prion protein of Syrian hamster induced by pH change. For this, we evaluated the influence of different force fields (GROMOS, AMBER and OPLS), different protonation states of histidine residues, different initial conditions and different methods for calculating long-range electrostatic interactions (GRF and SPME). From the time evolution of the secondary structures, we observed a strong dependence on the simulation parameters. In fact, the pH tendency described in literature was not clearly observed in this work. It may be associated with the need to invest more in multiple molecular dynamics simulations to quantify more accurately the structural behavior of the protein fragments in each pH study.

Les encéphalopathies spongiformes transmissibles sont des maladies neurodégénératives caractérisées par une vacuolisation intense et une perte neuronale associées à l’accumulation d’une protéine prion pathologique : la PrPSc. A cause de la longue période d’incubation silencieuse de ces maladies, les individus souffrant d’une maladie à prions peuvent exposer des patients qui recevraient leur sang ou un de leurs organes à un risque de contamination iatrogène. De plus, lorsque le diagnostic survient, les dommages cérébraux sont souvent massifs, et l’issue toujours fatale. A ce jour, aucun traitement n’est disponible, et l’ensemble des stratégies testées a échoué.L’alternative développée par le laboratoire est celle de la thérapie cellulaire couplée à la thérapie génique, en utilisant des cellules souches embryonnaires (ES) délivrant des molécules anti-prions. Concernant le choix des molécules anti-prions, nous avons choisi les mutants PrP-DN. Notre hypothèse repose sur des études montrant qu’une lysine au codon 219 de la PrP chez l’Homme ou une arginine au codon 171 de la PrP ovine, protègent du développement d’une ESST. L’étude de ces mutants, en cellules infectées ou dans des souris transgéniques, a permis de montrer que les PrP mutées n’étaient pas converties en PrPSc et qu’elles exerçaient un effet protecteur dit « dominant négatif » sur la conversion de la PrPC sauvage en PrPSc.Un des projets du laboratoire avait donc pour objectif d’utiliser les cellules souches exprimant les mutants PrP-DN pour développer une stratégie de thérapie génique et cellulaire des maladies à prions : l’hypothèse étant que les cellules greffées pourraient non seulement réparer le tissu lésé mais que ce dernier serait également protégé de l’infection par les prions.Une première approche de thérapie génique et cellulaire avait été initiée au laboratoire et montrait des résultats plutôt encourageants. En effet, la greffe de cellules souches neurales murines exprimant des PrP-DN et produites à partir de cellules souches embryonnaires murines, conduisait, pour une partie des souris, à un allongement du temps d’incubation de la maladie, ainsi qu’à une diminution de l’astrogliose et de la vacuolisation.Dans ce contexte, le premier objectif de ma thèse a été de valider l’approche thérapeutique en montrant que les cellules greffées délivraient des mutants PrP-DN capables d’inhiber la réplication du prion. Nous avons opté pour un modèle de culture organotypique infectée par des prions murins. En plus de répondre aux exigences éthiques de la directive 2010/63/UE, ce modèle offre l’avantage de pouvoir réaliser plus d’essais, des cinétiques d’accumulation de PrPSc et de visualiser le devenir des cellules greffées. Enfin, opter pour cette stratégie permettait de transposer dans un modèle humanisé des travaux précédemment réalisés et ayant montré des résultats encourageants. Pour cela, il a été nécessaire de mettre en place un modèle prion ex vivo de culture organotypique de tranches de cerveau, dans lequel il était possible de réaliser des greffes et permettant d’évaluer l’effet inhibiteur des mutant-PrP-DN sur la réplication du prion. Par ailleurs, notre groupe fait partie, avec d’autres groupes travaillant avec des cellules souches mésenchymateuses, de l’équipe « Biologie des cellules souches et médecine régénératrice », il nous est apparu pertinent d’évaluer l’effet des MSC sur la pathologie prions dans des modèles prions en culture organotypique et en particulier d’évaluer l’impact de greffes de MSC concomitantes aux greffes de NSC-PrP-DN. En effet, ces cellules sont décrites comme pouvant induire un microenvironnement neuroprotecteur en limitant la prolifération des cellules de la microglie et des astrocytes, et peuvent favoriser la différenciation des NSC. Enfin notre dernier objectif visait à transposer les outils murins vers des outils « humains » en produisant des NSC humaines issues d’ES et exprimant une PrP humaine portant les mutations DN. / Transmissible spongiform encephalopathies are neurodegenerative diseases characterized by a strong vacuolization, a neuronal lost and deposits of prion pathologic protein: PrPSc. This PrPSc accumulation is the result of the conformational conversion of the host encoded endogenous PrPC protein. Although the incidence of these diseases in humans remains low (about one to two cases per million inhabitants per year), these diseases remain a public health problem. Indeed, because of their long and silent incubation period, patients with prion disease may expose people through blood transfusion or organ transplantation with a risk of iatrogenic contamination. In addition, when the diagnosis occurs, brain damage is often massive, and the outcome is always fatal and rapidly occurs. Until now, there is no treatment that could be proposed to patients.The alternative developed by our laboratory for several years, is a strategy of cell therapy coupled with gene therapy. The general objective is to use pluripotent embryonic stem cells (ESC) and graft them as a “medicine” not only to orchestrate a functional recovery of the damaged zones and protect the grafted cells from prion propagation but also to deliver anti-prion molecules.For the anti-prion molecules, we have chosen dominant negative PrP mutants (PrP-DN). Our choice is based on studies showing that a lysine at codon 219 of the human PrP or an arginine at codon 171 of the ovine PrP protect against the development of a TSE. Study of these mutants in infected cells or in transgenic mice showed that the mutated PrPC were not converted into PrPSc. Moreover, they exibit a so-called "dominant negative" protective effect on the conformational conversion of their wild-type PrPC counterparts. A first approach of gene and cell therapy was initiated in the laboratory and has shown encouraging results. Indeed, the graft of murine neural stem cells (NSC) derived from murine embryonic stem cells and expressing anti-prion molecules, has allowed, for some of the mice, to an increase the incubation time of the diseaseas well as to a decrease of astrogliosis and vacuolization.In this context, the first objective of my thesis was to validate the therapeutic approach by showing that the grafted cells were able to inhibit prion replication trough the dominant negative effect of the PrP-DN. To address this point, we have chosen to use an organotypic culture model infected with murine prions (22L strain). In addition to fill the ethical requirements under the European Directive 2010/63 and the 3Rs, organotypic culture models offer the advantage to perform and repeat experiments, kinetic tests, and PrPres analysis. This model also allows to visualize the fate of the grafted cells. Finally, by choosing this strategy, it will be possible to transpose into a humanized model the work previously performed on mouse organotypic cultures.To achieve this task, it was necessary to establish an ex vivo prion model of organotypic culture brain slices, in which it was possible to perform grafts and to evaluate the inhibitory effect of PrP-DN mutants on the prion replication.In addition, as our group is included in the team "Stem cell biology and regenerative medicine" (led by Pr Jorgensen) in which several stem cells are studied (liver stem cells and mesenchymal stem cells (MSC)) and because MSC have been shown to provide protective effects when grafted in the brain of mice with neurological diseases, it was therefore relevant to evaluate the effect of MSC on prion pathology in our prion models and in particular to evaluate the impact of concomitant MSC grafts on NSC-PrP-DN.In a last step, our goal was to transpose the mouse tools (NSC from ES and expressing the PrP-DN mutants) to "human" tools by producing human NSCs derived from human ESC and expressing human PrP-DN, and to characterized the resulted cells.

Une protéine prion peut adopter deux conformations distinctes, l’une cellulaire et l’autre prion. La conformation prion est le résultat de son agrégation en fibre amyloïde. Cette fibre est le support de l’information prion à partir duquel les isoformes cellulaires sont convertis en forme prion de façon autocatalytique. La transmission de l’information prion repose donc sur la transmission de cette fibre au cours des divisions cellulaires, qui est réalisée par de petits polymères. Ceux-ci sont le résultat d’un équilibre entre la fragmentation et la polymérisation de la fibre. Une perturbation de cet équilibre provoque une agrégation massive de la protéine prion, menant à la perte de l’information prion.L’objectif de ma thèse était de comprendre ce qui définit in vivo la transmission du prion. Mon modèle d’étude est la protéine Ure2p propageant le prion [URE3] dans la levure S. cerevisiae. J’ai montré que la concentration cellulaire d’Ure2p détermine la vitesse d’agrégation de la protéine prion et donc son efficacité de transmission. En effet, de trop fortes concentrations cellulaires sont incompatibles avec la propagation du prion. La concentration cellulaire d’Ure2p définit également la diversité des souches prions. Un crible génétique m’a permit de mettre en évidence que la présence de séquences centromériques surnuméraires dans la cellule interfère avec la transmission du prion [URE3]. Le même phénomène est observé avec une augmentation du niveau de ploïdie de la cellule. Dans les deux cas, la surexpression du chaperon Hsp104 restaure une propagation normale du prion. / A prion protein can adopt two distinct conformations, one cellular and one prion. Prion conformation is the result of its aggregation into amyloid fibers. This fiber is the support of the prion information from which the cellular isoforms are converted into prion form by autocatalytic manner. The prion information transmission is therefore based on the transmission of this fiber during cell division, which is done by small polymers. These are the result of a balance between fragmentation and polymerization of the fiber. A disturbance of this balance causes a massive aggregation of the prion protein, leading to the prion information loss.The objective of my thesis was to understand what defined in vivo the prion transmission. My studying model was the Ure2p protein propagating the [URE3] prion in S. cerevisiae yeast. I showed that the Ure2p cellular concentration determined the aggregation speed of the prion protein and thus its transmission efficiency. Indeed, too high cellular concentrations are incompatible with the prion propagation. The cellular concentration of Ure2p also defines the prion strains diversity. A genetic screen allowed me to highlight that the presence of centrometric supernumerary sequences in the cell interferes with the [URE3] prion transmission. The same phenomenon is observed with an increase in the cell ploidy. In both cases, overexpression of the Hsp104 chaperone restores normal prion propagation.