THÈSE DE DOCTORAT de l’Université de recherche Paris Sciences et Lettres PSL Research University Préparée à l’Institut Curie Pyrimidine and NAD salvage pathways: unexpected allies in the maintenance of genome stability Ecole doctorale n°582 CBMS I Cancérologie : biologie - médecine - santé Spécialité : Aspects moléculaires et cellulaires de la biologie COMPOSITION DU JURY : Dr. Simon Saule Institut Curie, Président du jury Soutenue par Sandra Cunha Silveira Dr. Françoise Dantzer Ecole supérieure de biotechnologie de Le 9 Octobre 2018 Strasbourg (ESBS), Rapporteur h Dr. Alexander Ishchenko Gustave Roussy, Rapporteur Dirigée par Mounira Amor-Guéret Dr. Pierre Cordelier Centre de recherche en cancérologie de h Toulouse, Examinateur Dr. Fatima Mechta-Grigoriou Institut Curie, Examinateur Dr. Mounira Amor-Guéret Institut Curie, Directeur de thèse Pyrimidine and NAD salvage pathways: unexpected allies in the maintenance of genome stability Voies de sauvetage du pool des pyrimidines et du NAD: des alliés inattendus dans le maintien de la stabilité du génome Acknowledgements After four intensive years, my PhD came to an end. It is time to reflect about the people that accompanied me during this journey. My 4 years at Institut Curie had a major impact on my life as a researcher and as well as a person. Therefore, I would like to address a kind acknowledgment to those who contributed to the success of my PhD. First, I would like to thank PSL Research University for funding my first 3 years of PhD and La ligue contre le cancer and Institut Curie for funding my 4th and last year of PhD. I would like to thank all the jury members: Dr. Françoise Dantzer, Dr. Alexander Ishchenko, Dr. Pierre Cordelier, Dr. Fatima Mechta-Grigoriou and Dr. Simon Saule. Thank you very much for accepting to participate in my thesis defense. I would also like to thank my PhD supervisor, Mounira Amor-Guéret for accepting me as her PhD student in her team. Also, I would like to thank Mounira for her patience, advices and ideas that were very helpful during these four years. Her constant challenges allowed me to learn and further develop my skills. Now, I would like to thank all the former and current members of the team. Starting by the formers, I would like to thank my office neighborhoods Elias Bou Samra and Hamza Mameri for all the funny moments but also for the most serious ones (scientific discussions). Elias, thank you for your kind advices, help and scientific discussions. I hope that you are “enjoying” your new job at L’Oréal. Hamza, even though you are at Montpellier, starting your new career in a new institute and team, you always have time to discuss with me. Thank you very much for taking some of your precious time to help me. Finally, Hamza, I’m still waiting for the “blagues pourries” during lunch time. A thank to Alexis Fouquin. Although you came from another team to this one, thank you very much for your sympathy. You did an excellent PhD that allowed you to obtain a job at AstraZeneca. I hope you are loving your new career. Finally, I would like to thank Simon Gemble for teaching me all the techniques necessary to start my project. Thank you for your patience and time during my first year. Now, I would like to thank the current members. First my “professeure” Géraldine Buhagiar- Labarchède. Géraldine, a special and big thank you for your help during these four years. Thank you for always having a solution for a problem and a big thank you for your kindness and patience to correct every time I mispronounce a French word. As you said, I speak French exactly as your children…LOL…. Rosine, thank you very much for your “savoir-faire” that we need to learn in a laboratory. Also, thanks for your help during my project and for all the little discussions. I would like to thank Matthieu Grattia and Vincent Pennaneach. Matthieu, you are now working at Curie-Paris, but thank you for all the good moments, chocolate contraband and for your little jokes about my French. Vincent, thank you for your serenity and interesting discussions. I would like also to thank the formers and current members of your unit UMR3348 that contributed to a friendly environment during these 4 years. A special word goes to my Portuguese compatriot Ana Silva. Thank you for your friendship (for being there when I needed) and funny moments we share together. Also, thank you for the invitation to your special day that allowed me to discovery a beautiful place in your country. A special gratitude to Anissia Ait Saada, a good friend always with a kind word. Also, a thank to Julien Hardy for all the jokes about the Portuguese’s ruling the world xD. A special thanks to the girls from Stephan’s Vagner lab, Alina Chakraborty, Hélène Malka-Mahieu, Iris 5 Tanaka, Melissa Leriche, Sabrina Mennour and Rym Sfaxi from all the funny moments and company during lunchtime. I would also to thank Ahmed El-Marjou for his help during the construction of the NAMPT recombinant protein and Liliane Mouawad for teaching me and helping me during the molecular docking for NAMPT. I thank also the plateforme IBiSA d’imagerie cellulaire et tissulaire, Claire Lovo and Marie- Nöelle for the help with the microscopy experiments. I would also like to thank Patricia Duchambon for her patience, time and help during the construction of the NAMPT recombinant protein. It was complicated, but we did it. I would like to thank Jérôme Guitton and Christelle Machon for all the HPLC-MS experiments and time invested in my project. Lastly, I would like to thank another Portuguese compatriot, Sofia Ferreira, for all the conversations between us. For having accompanied the development of my thesis, I would like to thank the members of my thesis committee: Sarah Lambert, Martin Dutertre, Valérie Schreiber, Alexander Ishchenko and Yegor Vassetzky. In a more personal level, I would also like to thank Ana Luisa Ribeiro for your friendship. It has been a long time since we finished your Master degree and even though I am in Paris, we are always in touch. Thank you for your support and kind words during these years. I would like to thank both Alain and Marie Baron for accepting me with open arms in their home when I arrived at Paris. Thank you for all the kindness and help during the process to search a PhD in France. A special thanks to Manuel and Emilia Fernandes de Araujo for all the help during the search for a PhD and for letting me to stay in their home during my journeys at Gustave Roussy. A very big thank you for my parents-in-law for accepting me in the family as a new daughter. Thank you very much for all the help when I arrive at Paris. For all the help during my search for the PhD as well to make a little easier to overcome the challenges of being in a new country. A special thanks for my sister-in-law, Stacy Baron. Although they are very far, I would like to thank my parents, Serafim Silveira and Luisa Cunha, as well as my little sister, Diana Silveira, for supporting my choice to leave Portugal to pursuit my professional career. Finally, I would like to thank my husband (and boyfriend), Brian Baron, for all he did during these past years. Thank you very much for all your support, advice and friendship when I need. Also, thank for all the discussions about biology and physics, although I don’t have the same opinion (Biology rules!!!). I am very proud of you that you found your dream job as Medical Physicist. You deserve it. Thank you, everyone 6 Table of contents Acknowledgements ................................................................................................................... 5 Table of contents ....................................................................................................................... 7 List of figures ............................................................................................................................ 11 List of tables ............................................................................................................................. 13 Abbreviations ........................................................................................................................... 15 Abstract .................................................................................................................................... 19 Résumé ..................................................................................................................................... 21 Introduction ............................................................................................................................. 23 Bloom syndrome and the BLM protein ........................................................................ 25 1. Bloom syndrome ........................................................................................................... 27 1.1. Clinical features ...................................................................................................... 27 1.2. Cancer predisposition ............................................................................................ 28 1.3. Bloom syndrome cellular phenotypes ................................................................... 30 1.3.1. Cytogenetic features....................................................................................... 30 1.3.2. DNA Replication abnormalities....................................................................... 31 1.3.3. Mitotic abnormalities ..................................................................................... 31 1.3.4. Oxidative