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Deciphering Additional Mechanisms of Mtorc1 Signaling in Skeletal

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Deciphering Additional Mechanisms of Mtorc1 Signaling in Skeletal Deciphering additional mechanisms of mTORC1 signaling in skeletal muscle Inauguraldissertation zur Erlangung der Würde eines Doktors der Philosophie vorgelegt der Philosophisch-Naturwissenschaftlichen Fakultät der Universität Basel von Kathrin Chojnowska aus Deutschland Basel, 2019 Originaldokument gespeichert auf dem Dokumentenserver der Universität Basel edoc.unibas.ch Genehmigt von der Philosophisch-Naturwissenschaftlichen Fakultät auf Antrag von Prof. Dr. Markus A. Rüegg Prof. Dr. Christoph Handschin Basel, den 19.02.2019 Prof. Dr. Martin Spiess Dekan der Philosophisch-Naturwissenschaftlichen Fakultät Table of Content 1 Acknowledgements ........................................................................................................... 7 2 List of Abbreviations .......................................................................................................... 9 3 Aim of the Study .............................................................................................................. 13 4 Outline of the Thesis ....................................................................................................... 15 5 Introduction ...................................................................................................................... 17 5.1 Skeletal muscle in health and disease- a metabolic organ ...................................... 17 5.2 mTORC1 and its role in skeletal muscle .................................................................. 19 6 Chapter 1: Sustained activation of mTORC1 in skeletal muscle causes impaired mitophagy with a subsequent increase in mitochondrial size but without altering mitochondrial homeostasis .............................................................................................. 25 6.1 Summary .................................................................................................................. 25 6.2 Introduction .............................................................................................................. 27 6.2.1 Mitochondria- the powerhouse of the cell ......................................................... 27 6.2.2 Mitochondria in skeletal muscle ........................................................................ 30 6.3 Results ..................................................................................................................... 33 6.4 Discussion ................................................................................................................ 45 7 Chapter 2: Long-term calorie restriction reduces skeletal muscle myopathy in TSCmKO mice ................................................................................................................................. 51 7.1 Summary .................................................................................................................. 51 7.2 Introduction .............................................................................................................. 53 7.2.1 Calorie restriction and its beneficial effects ...................................................... 53 7.2.2 Calorie restriction and mTORC1 ....................................................................... 54 7.2.3 Calorie restriction and its benefits on skeletal muscle ...................................... 55 7.3 Results ..................................................................................................................... 57 7.4 Discussion ................................................................................................................ 71 7.5 Appendix .................................................................................................................. 77 8 Chapter 3: Sustained activation of mTORC1 in skeletal muscle leads to disturbed calcium homeostasis and excitation-transcription coupling ............................................. 81 8.1 Summary .................................................................................................................. 81 8.2 Introduction .............................................................................................................. 83 8.2.1 Skeletal muscle composition- Differentiation of fiber types .............................. 83 8.2.2 Calcium signaling in skeletal muscle ................................................................ 85 8.2.3 Calcium signaling in disease and aging ............................................................ 90 8.3 Results ..................................................................................................................... 93 8.4 Discussion .............................................................................................................. 101 9 Material and Methods .................................................................................................... 107 10 Appendix ....................................................................................................................... 117 11 References .................................................................................................................... 133 Acknowledgements 1 Acknowledgements First and foremost, I would like to express my thanks to Prof. Dr. Markus Rüegg for giving me the opportunity to develop scientific expertise and subsequently obtaining my PhD in his laboratory. I am very grateful for your continuous support in my PhD studies and all the related research projects that I have been conducting. I very much cherished your guidance and immense knowledge, the freedom to develop my own ideas and to gain expertise but also to develop my professional, as well as personal skills. I am glad for your patience, in particular throughout our sometimes rocky path and that you accepted and appreciated me as an individual with all my strength and weaknesses in your laboratory. Besides my advisor, I would also like to thank the rest of my thesis committee: Prof. Dr. Christoph Handschin and Prof. Dr. Susan Treves for their insightful comments and encouragement, their critical questions throughout my committee meetings and other presentations. I also would like to thank both for their unique support when acquiring new techniques and utilizing their laboratory specialization. I appreciated the collaborations and I am very grateful for the teamwork. Frankly speaking, I enjoyed working with both of you and your lab members as a team a lot. Thanks to your support, I was able to widen my knowledge, and horizon on skeletal muscle research. Sincere gratitude goes to Dr. Perrine Castets and Dr. Nathalie Rion. You have not just been incredibly helpful within the first year of my PhD by teaching me various techniques to improve precision to a very high quality standard, but you also always had an open ear, be it for scientific discussions or giving me food for thought and expertise in general. I would also like to thank my fellow labmates for the stimulating discussions and for all the fun we have had throughout the four years. I had a wonderful time with you and I am grateful for the immense support. Special thanks to Dr. Daniel Jacob Ham for his support and expertise in lab animal handling and statistics. Thanks goes to Dr. Shuo Lin for the continuous support and knowhow be it force measurements or microscopy. I do not know whether I would still sit in front of the microscope, if it was not for your support throughout the years. Page | 7 Acknowledgements I would also like to mention the support of the “coffee gang”. You were a great sanction and I appreciated the fruitful discussions we always had. Special thanks goes to my lab-buddy, sitting next to me. You were the best support and friend I could hope for in a busy, professional work environment. I would also like to thank the members of Prof. Dr. Bernhard Bettler’s and Prof. Dr. Michael Hall’s laboratory for their continuous support, for any technical or theoretical questions. Special thanks go to Diana Weissenberger for all the emotional support during the countless nights that I have been working. Many thanks go to Dr. Martin Gassmann for answering all my questions regarding the world of academia — which is, frankly speaking, sometimes still a mystery to me. I hope you know how much I appreciated your help, especially during the last phase of my PhD. I am certain that there is no amount of chocolate, which can possibly compensate for what you have done for me. I thank all my friends, from school, from University, from my time at the Biozentrum and from life itself. I am proud to have an amazing chosen family that I can count on. I am in the lucky position of receiving great support from all of you. Thank you for always finding encouraging and motivating words to cheer me up when skies are grey. Special thanks go to my family for their tremendous support throughout my PhD and my professional career in general. I would not have been able to stand here today, if it was not for you. Thank you for always being supportive, no matter which ideas I have had, what I wanted to become or where I wanted to go next. I truly appreciate your never-ending support. Lastly, I cannot express how thankful I am for having Claude Chautems by my side. You were the best support and help I could wish for. Thank you for always being patient with me and for your ability of calming me down during this emotional and challenging chapter of my life. I very much appreciated your help, and I cannot wait to start our new chapter in Zurich and to see what the future holds for us.
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