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Phallusia Mammillata Embryos Marianne Roca

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Phallusia Mammillata Embryos Marianne Roca The spindle assembly checkpoint in Phallusia mammillata embryos Marianne Roca To cite this version: Marianne Roca. The spindle assembly checkpoint in Phallusia mammillata embryos. Development Biology. Sorbonne Université, 2019. English. NNT : 2019SORUS500. tel-03001896 HAL Id: tel-03001896 https://tel.archives-ouvertes.fr/tel-03001896 Submitted on 12 Nov 2020 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. Thèse de doctorat en biologie Sorbonne Université - Ecole doctorale complexité du vivant – ED515 Laboratoire de biologie du développement de Villefranche sur mer (LBDV) – UMR 7009 Equipe : mitose et contrôle du fuseau The spindle assembly checkpoint in Phallusia mammillata embryos Par Marianne Roca Dirigée par Elisabeth Christians and Stefania Castagnetti Présentée et soutenue publiquement le 29 novembre 2019 à Villefranche sur mer Devant un jury composé de : Dr. Roger Karess, rapporteur Pr. Eric Thompson, rapporteur Dr. Aude Dupré, examinatrice Pr. Elisabeth Chrisitians, directrice de thèse Dr. Stefania Castagnetti, encadrante (invité) Abstract During mitosis, progression through anaphase must take place only when all chromosomes are correctly attached to spindle microtubules to avoid chromosome mis-segregation and the generation of aneuploid cells (i.e. with an abnormal chromosome number). Embryos containing aneuploid cells can exhibit developmental defects and lethality. Furthermore, cancer cells are often aneuploid. To prevent such deleterious aneuploidy, a control mechanism, the spindle assembly checkpoint (SAC), delays metaphase- anaphase transition until all chromosomes are properly attached to spindle microtubules. However, the SAC is not efficient during early development in some species. During my thesis, I analyzed the activity of the SAC during the development of the marine chordate P. mammillata. I showed that in P. mammillata embryos, the SAC becomes efficient at the 8th cell cycle and its efficiency increases progressively in the following cell cycles. Although, I demonstrated that patterning of the embryo along the anteroposterior axis influences SAC efficiency, my experiments suggest that additional parameters modulate SAC efficiency. I searched the molecular mechanisms, which control SAC efficiency during development. I collected evidence showing that SAC components are present in oocytes and all post-fertilization stages. I found that SAC proteins localize at kinetochores during meiosis and at later stages when there is an efficient SAC while they do not accumulate on unattached kinetochores in early SAC deficient embryos. My thesis work establishes P. mammillata as a valuable experimental organism to study SAC regulation during embryogenesis. Keyword: Mitosis, Spindle assembly checkpoint, Aneuploidy, Chordates, Nocodazole, Embryo Résumé Le point de contrôle du fuseau mitotique (Spindle Assembly Checkpoint : SAC) retarde l’anaphase jusqu’à ce que tous les chromosomes soient attachés correctement aux microtubules. Le SAC permet ainsi d’éviter des erreurs de ségrégation des chromosomes aboutissant à des cellules filles aneuploïdes (i.e. avec un nombre anormal de chromosomes). L’aneuploïdie, délétère pour les cellules, peut entrainer des problèmes de développement et est observée dans les cancers. Cependant, chez certaines espèces, le SAC n’est pas efficace au cours de la phase précoce du développement embryonnaire. J’ai mis en évidence que chez l’ascidie P. mammillata, un organisme marin du groupe des chordés, le SAC devient efficace au 8ème cycle cellulaire et son efficacité augmente dans les cycles suivants. J’ai démontré qu’en partie ventrale l’identité des cellules antérieures induisait la présence d’un SAC plus efficace mais que d’autres facteurs modulaient aussi l’efficacité du SAC. J’ai étudié les mécanismes moléculaires impliqués dans les variations de l’efficacité du SAC au cours du développement. Mes expériences ont révélé la présence des composants du SAC tout au long de l’embryogenèse. Cependant, j’ai pu montrer que les protéines du SAC ne se localisent pas au niveau des kinétochores lorsque le SAC est inefficace au début du développement mais qu’elles s’y localisent bien dans l’ovocyte en méiose et dans l’embryon plus tardif, lequel se caractérise par un SAC actif. Ma thèse a permis de montrer que P. mammillata est un organisme expérimental de grand intérêt pour l’étude du SAC au cours de l’embryogenèse. Mot-clé : Mitose, Point de contrôle du fuseau mitotique, Aneuploïdie, Chordé, Nocodazole, Embryon Acknowledgments / Remerciement First of all, I would like to thank all my jury members for agreeing to look through my work and evaluate it. Aude Dupré merci pour vos conseils au cours des comités de thèse, c’est un plaisir de vous montrer jusqu’où ma thèse m’a menée. Roger Karess, I discovered the SAC in your team and it’s an honor to have you to judge my work. Eric Thompson thank you for agreeing to be a reporter bringing a tunicate eye on the project. After 3 years and more than 5207 mitosis analyzed, I have so many people to thank for helping me pursue my thesis and I hope I won’t forget too many. Stefania and Elisabeth thank you for taking me in your team and for teaching me so much during these 3 years. The time spent in the team was like a fairy tail with thousand and one species. Stefania you were always available and listening thanks a lot. Thank you for taking so much of your time to improve this manuscript, I learned a lot in scientific English writing thanks to you. Thank you for giving me the possibility to go to so many meetings where I heard and met incredibly smart and nice scientist. Elisabeth, merci de m’avoir accompagnée, soutenue et d’avoir enduré mon français. Lydia, je n’aurais pas pu faire cette thèse sans toi. Merci pour toutes les techniques que tu m’as enseignées et merci d’avoir pris la relève quand j’étais sur le point de devenir Mad avec mes clonages. Bravo pour ta persévérance avec Bub1. Merci aussi pour les expériences que tu as faites pour moi. Merci à l’ensemble de l’équipe ABC pour m’avoir tout enseigné, depuis les bases, sur le travail avec ce joli mdm. Sans vous, P. mammillata serait resté une illustre inconnue. Janet, there is so much to say even if you think you did nothing!! Thank you for having been a wonderful tutrice aussi bien en sciences qu’en franglais. Merci pour tes 42 idées nouvelles par semaine (pour ne pas dire jour), il ne me reste plus qu’à trouver la question. Merci à Gérard et Céline pour la bonne humeur matinale et l’ambiance musicale du matin (je n’oublierai jamais la fois où ce fut Mélanie). Céline, merci aussi pour tes conseils, tu m’as montré le chemin plus d’une fois. Merci pour tes discussions légères et ton rire quand j’étais prête à toute envoyer en l’air. Remi, j’aurais bien eu du mal à réaliser ce projet si tu n’avais pas eu l’idée d’utiliser le NLS et si tu ne nous avais pas passé la construction, merci beaucoup ! Et merci pour l’image du tadpole (Fig. 12D). Alex, you tought me a lot about ascidian and cell cycle, you showed me that there was a difference in SAC along the anteroposterior axis in my movies, thanks a lot. Thank you also for showing me how “easy” it is to perform pole ablation of P. mammillata eggs. After several months, yoga and diving lessons to help me relax and control my breath, I almost agree! Genia, where would I be without you? You literally drive me so much thanks a lot! Anne cela a fait du bien d’avoir quelqu’un à la mezza qui puisse à la fois s’amuser avec les lignes de commandes et s’émerveiller devant la structure d’une cellule ! Isa tu as été un professeur, un mentor, une amie… tu nous as montré à nous autres thésardes qu’il était possible d’arriver au bout and break free. 1 Mon temps passé au LBDV aurait été bien différent sans toi Sophie. Je n’aurais pas risqué de mourir de froid en février mais aurais sûrement écopé d’une piqure de méduse s’y elles ne fuyaient pas par peur que tu les découpes. Je n’aurais pas été fanny 2 ans de suite mais plutôt trois ! Je n’aurai probablement pas une aussi bonne capacité à suivre un protocole/recette. Tout se monnaye…. en chocolat ou gâteau ! Et je n’aurais pas su glisser aussi bien entre les embuches de l’administration. Merci d’avoir galéré juste avant moi. David, Anna now it is your turn to face the depth of French administration but I have no doubt that you will come back on the surface of this sea of paper. Julia ton passage fut bref mais intense, merci d’avoir ouvert mon esprit sur de nouveaux sujets. Maciek, you made me discover how pretty the ctenophors are, thanks for that. Isabella it was so great to meet you. Adrien tu as rejoint la mezza au pire moment, merci pour ton aide pour la gène ontologie. Angelica each time you come it is an event as big as Christmas! Manon, merci de m’avoir fait découvrir un coin d’Irlande en PACA �, désolée, mais sérieusement ce fut un chouette moment ! Merci à tous les membres de la mezza passé, présent et futur.
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