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Séquençage Du Génome Du Parasite Intestinal Blastocystis Sp. (ST7)

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Séquençage Du Génome Du Parasite Intestinal Blastocystis Sp. (ST7) Séquençage du génome du parasite intestinal Blastocystis sp. (ST7) : vers une meilleure compréhension des capacités métaboliques d’organites apparentés aux mitochondries chez ce microorganisme anaérobie Michaël Roussel To cite this version: Michaël Roussel. Séquençage du génome du parasite intestinal Blastocystis sp. (ST7) : vers une meilleure compréhension des capacités métaboliques d’organites apparentés aux mitochondries chez ce microorganisme anaérobie. Sciences agricoles. Université Blaise Pascal - Clermont-Ferrand II, 2011. Français. NNT : 2011CLF22164. tel-00678598 HAL Id: tel-00678598 https://tel.archives-ouvertes.fr/tel-00678598 Submitted on 13 Mar 2012 HAL is a multi-disciplinary open access L’archive ouverte pluridisciplinaire HAL, est archive for the deposit and dissemination of sci- destinée au dépôt et à la diffusion de documents entific research documents, whether they are pub- scientifiques de niveau recherche, publiés ou non, lished or not. The documents may come from émanant des établissements d’enseignement et de teaching and research institutions in France or recherche français ou étrangers, des laboratoires abroad, or from public or private research centers. publics ou privés. N°D.U. : 2164 Université Blaise Pascal ECOLE DOCTORALE DES SCIENCES DE LA VIE, SANTE, AGRONOMIE, ENVIRONNEMENT N° d’ordre : 557 Thèse pour obtenir le grade de DOCTEUR D’UNIVERSITE Spécialité : Microbiologie Présentée et soutenue publiquement par ROUSSEL Michaël le 27 septembre 2011 Séquençage du génome du parasite intestinal Blastocystis sp. (ST7) : vers une meilleure compréhension des capacités métaboliques d’organites apparentés aux mitochondries chez ce microorganisme anaérobie Directeur de la thèse : M. DELBAC Frédéric, Professeur, Université Blaise Pascal Membres du jury : M. BRINGAUD Frédéric, DR CNRS, RMSB, Université Bordeaux Segalen (Bordeaux) M. HIRT Robert, Senior Lecturer, ICAMB, Université de Newcastle (Newcastle) Mme. DARFEUILLE-MICHAUD Arlette, Professeur, Université d’Auvergne (Clermont-Ferrand) M. VIVARES Christian, Professeur émérite, Université Blaise Pascal (Clermont-Ferrand) M. VISCOGLIOSI Eric, CR CNRS, Institut Pasteur Lille, Université Lille Nord, (Lille) LMGE, UMR-CNRS 6023 Equipe Interactions hôtes-parasites Université Blaise Pascal 24 Avenue des Landais, 63177 Aubière cedex A mes parents, Pour m’avoir encouragé et soutenu pendant toutes ces années, Je vous dédie cette thèse car c’est grâce à vous que je la soutiens aujourd’hui. Soyez certains de toute ma reconnaissance, admiration et affection A tous ceux qui me sont chers Remerciements Je remercie sincèrement, Messieurs les Docteurs Frédéric Bringaud et Robert Hirt d’avoir aimablement accepté d’être les rapporteurs de cette thèse. Je suis fier de vous compter parmi mes juges et vous assure ma reconnaissance et mon respect. Madame le Professeur Arlette Darfeuille-Michaud, d’avoir aimablement accepté de participer à ce jury, et de m’avoir permis de terminer ma thèse en m’acceptant sur un poste d’ATER. Monsieur Christian Vivares, pour m’avoir accompagné et soutenu en Master 2 et durant le concours de l’attribution des bourses de thèse, pour ses conseils avisés. Merci ! Monsieur Eric Viscogliosi, pour sa collaboration et son engagement dans nos travaux communs sur Blastocystis. Nous ne sommes pas au bout de nos surprises Eric ! Monsieur le professeur Frédéric Delbac, pour m’avoir suivi tout au long de cette thèse, pour sa disponibilité, sa confiance, son écoute, son soutien et ses conseils. Cela a été un plaisir de travailler dans ton équipe et sous ta direction Fred. Trouve ici toute ma gratitude et mon respect sincère. Je tiens à remercier, Tous ceux de l’équipe « Interactions hôtes-parasites » et du LMGE, pour leur accueil, leurs conseils, et leur gentillesse. Je tiens également à remercier tous ceux qui ont participé à ce travail. ROUSSEL Michaël Séquençage du génome du parasite intestinal Blastocystis sp. (ST7) : vers une meilleure compréhension des capacités métaboliques d’organites apparentés aux mitochondries chez ce microorganisme anaérobie Résumé Blastocystis sp., est un straménopile parasite anaérobie fréquemment rencontré dans le tractus gastro-intestinal de l’homme et de divers animaux. Ce microorganisme, parfois responsable de désordres digestifs aigus, pourrait conduire à des troubles fonctionnels intestinaux tels que le syndrome de l’intestin irritable (IBS). Le génome de Blastocystis sp., qui a fait l'objet d'un projet de séquençage en collaboration avec le Génoscope d’Evry, nous a permis de caractériser le plus petit génome de straménopile séquencé à ce jour (18,8 Mpb), avec une capacité codante de 6020 gènes. L’acquisition de nombreux gènes par transferts horizontaux est une caractéristique majeure de ce génome, qui montre d’abondants réarrangements génomiques. Bien qu’évoluant en anaérobiose, Blastocystis sp. possède des organites morphologiquement proches des mitochondries, appelés mitochondrion-like organelles (MLOs). Nous avons montré que ces organites comportaient un génome circulaire de type mitochondrial de 29,27 kpb, mais dépourvu des gènes codant pour les cytochromes. Des analyses in silico nous ont permis de caractériser le protéome des MLOs (365 protéines), conduisant à l’établissement d’un modèle prédictif des voies métaboliques associées à ces organites, avec notamment une chaine respiratoire limitée aux complexes I et II. Nous avons ainsi montré que les MLOs présentent des caractères communs aux mitochondries anaérobies et aux hydrogénosomes (présence d’une PFOR et d’une hydrogénase à fer), suggérant que Blastocystis sp. comporte des mitochondries anaérobies modifiées, qui résulteraient d’une adaptation du parasite à son environnement. Par ailleurs, la prédiction du sécrétome de Blastocystis sp. révèle la présence de facteurs de virulence potentiels, pouvant être impliqués dans l’altération de l’épithélium intestinal et le contournement du système immunitaire de l’hôte. Mots-clefs : génome, analyse in silico, mitochondrie anaérobie, hydrogénosome, sécrétome Abstract Blastocystis sp. is a highly prevalent anaerobic eukaryotic stramenopile parasite found in the intestinal tract of humans and various animals. This microorganism, sometimes associated with acute intestinal disorders, could be responsible for functional intestinal disorders such as the irritable bowel syndrom (IBS). As part of a collaborative sequencing project with the Genoscope (CEA Evry, France), we were able to caracterize the smallest stramenopile genome sequenced to date (18.8 Mbp) with a 6020 genes coding capacity. The gain of many genes through horizontal gene transfer is a major characteristic of this genome, which shows extensive genomic rearrangements. Despite the anaerobic nature of Blastocytists sp., this eukaryote harbours nevertheless mitochondrion-like organelles (MLOs). We have shown that these organelles have a 29.27 kbp mitochondrial-type circular genome that lacks cytochrome coding genes. In silico analysis allowed us to predict the MLOs proteome (365 proteins), with the subsequent predictive model of the metabolic pathways associated with these organelles, including an electron transport chain (ETC) restricted to complex I and II. We have shown that MLOs shared common characteristics with anaerobic mitochondrion and hydrogenosomes (presence of a PFOR and an iron-hydrogenase), which could mean that Blastocystis sp. harbours modifiedSOMMAIRE anaerobic mitochondrion that resulted from the parasite adaptation to its anaerobic environment. In addition, Blastocytis sp. secretome prediction reveals the presence of potential virulence factors, which could be involved in the degradation of the intestinal epithelium as well as the host immune system bypass. Key-words : genome, in silico analysis, anaerobic mitochondria, hydrogenosome, secretome SOMMAIRE LISTE DES ABREVIATIONS ..................................................................................... 1 INTRODUCTION .................................................................................................... 2 CHAPITRE I - REVUE BIBLIOGRAPHIQUE I. BLASTOCYSTIS SP., UN PARASITE INTESTINAL POLYMORPHE A LA DIVERSITE GENETIQUE COMPLEXE .................................................................................... 5 A. TAXONOMIE ET DIVERSITE GENETIQUE DE BLASTOCYSTIS SP. ........................................ 5 A.1 UNE IDENTIFICATION LABORIEUSE ............................................................................ 5 A.2 UN STRAMENOPILE PEU COMMUN ........................................................................... 6 A.3 DIVERSITE GENETIQUE ET CLASSIFICATION .................................................................. 8 A.3.1 Une diversité génétique complexe ................................................................... 9 A.3.2 Spéciation et classification .............................................................................. 11 B. LA BIOLOGIE DE BLASTOCYSTIS SP. ......................................................................... 14 B.1 UN ORGANISME POLYMORPHE .............................................................................. 14 B.1.1 La forme vacuolaire ......................................................................................... 15 B.1.2 La forme granulaire ......................................................................................... 17 B.1.3 La forme amiboïde .......................................................................................... 17 B.1.4 La forme kystique ...........................................................................................
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