New Tools for Old Organelles

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New Tools for Old Organelles New tools for old organelles Bioinformatics for comparative cell biology Marc Gouw Dissertation presented to obtain the Ph.D degree in Bioinformatics Instituto de Tecnologia Química e Biológica António Xavier | Universidade Nova de Lisboa Oeiras, July, 2015 New tools for old organelles Bioinformatics for comparative cell biology Marc Gouw Dissertation presented to obtain the Ph.D degree in Evolutionary Biology Instituto de Tecnologia Química e Biológica António Xavier | Universidade Nova de Lisboa Research work coordinated by: Oeiras July, 2015 Acknowledgments This thesis and the years of work and inspiration put into it would not have been possible without the help of many people. First of all, I just generally want to thank everyone for being so great. Especially those who in any way contributed to making the past few years in Portugal and at the IGC such a wonderful experience: without all of you, my life and this thesis would have turned out very different indeed. A special thanks to all of the past and present members of the Computational Genomics Laboratory, who have always been kind enough to be unforgivingly critical of my work, and equally enthusiastic to go enjoy a beer on a Friday afternoon. And especially Beatriz Ferreira Gomes and Yoan Diekmann, who were essential to my PhD project: You have been truly inspiring collogues and close friends. A lot of this work would not have been possible without you. The epic mtoc-explorer.org, and indeed this thesis would also not have been possible without Renato Alves. I blame you, your programming genius and incredible patience for teaching me so much. I also really want to thank my ever so important supervisors Jos´ePereira- Leal and M´onicaBettencourt-Dias. Your guidance, supervision and support over the past few years has been invaluable. Also Zita Carvalho-Santos, who's initial work laid the foundation of what turned into the major part of my PhD. The IGC is a wonderful place to study. I wanted to thank Prof. Howard and Prof. Coutinho for giving me the opportunity to be part of the IGC. I wanted to thank Thiago Carvalho, and Elio´ Sucena for providing a lot of practical and scientific support. Jorge Carneiro also provided incredibly useful feedback and advice throughout various parts of this project. Also my thesis committee Patr´ıciaBeldade and Nuno Moreno, you provided excellent advice (especially towards the end of my PhD), for which I will always be grateful. There are many at the IGC who I value incredibly as colleagues and friends, to whom I cannot express enough gratitude. Barbara Vreede, your never- ending excitement with science and life in general is truly inspiring (and highly contagious). I can't thank you enough for making the IGC a special place (and for being my gateway to coming to Portugal in the first place). And Dani Bodor, your vast knowledge, curiosity and excitement at the world of cell biology and beyond are incredible, just as your ability to catch a hammer. There are also a few people I would like to thank for helping me get this thesis finished. Jarek Surkont provided great feedback on chapter3. Renato Alves & Ricardo Leite for their comments and suggestions on chapter2. Lastly I wanted to thank those closest to me for always being there to support me, my work, thesis, or any other important (or meaningless) part in my life. Mom & Dad, thanks for you neverending support, for never doubting iii me, and always giving me an extra push to keep on going when I need it. Richard and Bernard, you are incredible brothers, and although you are far away, I know you guys always have my back. And the love of my life Iria, for her endless support while I was writing this thesis, but more generally for being such a wonderful person and part of my life. Thank you all !!! Financial Support Esta disserta¸c~aoteve o apoio financeiro da FCT e do FSE no ^ambito do Quadro Comunit´ariode Apoio, bolsa de doutoramento #SFRH/BD/51628/2011 e da Funda¸c~aoCalouste Gulbenkian. This dissertation had the financial support from FCT and FSE through the Quadro Comunitrio de Apoio, doctoral fellowship #SFRH/BD/51628/2011 and Funda¸c~ooCalouste Gulbenkian. Software This thesis was written entirely using the free and open source software programs LATEX, Inkscape & Python. Cover image Cover image is an adaptation of figure 2.1, which includes a modified version of a Chlamydomonas reinhardtii Transition Zone from Geimer and Melkonian (2004). iv Summary For hundreds of years biologists have studied the naturally occurring diversity in plant and animal species. The invention of the electron microscope in the first half of the 1900's reveled that cells also can be incredible complex (and often stunningly beautiful). However, despite the fact that the field of cell biology has existed for over 100 years we still lack a formal understanding of how cells evolve: It is unclear what the extents are in cell and organelle morphology, if and how diversity might be constrained, and how organelles change morphologically over time. The emergence of the eukaryotic cell over 1 billion years ago marks one of evolutions major transitions. In this branch of life the cellular architecture evolved from a relatively simple plan to a highly complex and compartmental- ized system of organelles. One of the most powerful ways to study evolution is to study diversity across a broad range of different species: The \compara- tive" approach to biology. In the context of eukaryotic evolution we call this \comparative cell biology", which we explore in this thesis. In this thesis we study two model systems for \comparative cell biology": Microtubule Organizing Centers (MTOCs) in chapters2 and3 and RabGTPases in chapter4. Each of these chapters explores a different angle of cellular evolution, and each chapter proposes new bioinformatics tools to enable a \comparative cell biology" approach. The first chapter addresses evolution of MTOCs from a purely morphological perspective. In order to achieve this we created mtoc-explorer.org, a community driven web-resource in which we collected ultrastructural data on MTOCs from over 100 species. Using this data we were able to determine some of the fundamental principles of the evolution of shape in organelles. We show that although diversity is a prominent theme in MTOC evolution, the total set of possible morphologies is constrained by functional requirements. In doing so we uncover a \spandrel" in cell biology: The requirement for microtubule based motility constraints the overall architecture of a cell's mitotic apparatus. Lastly we develop a model to measure ancestrality of organelles, and show convergent evolution of complex organelles in cells. One of the major goals in biology is determine the link between a species' v genome and its morphological and functional properties. In chapter3 we address this issue in a 32 species analysis using the eukaryotic cilium as a model organelle. Using a bioinformatics technique called \phylogenetic profiling” we ask how well we can use the presence and absence of a gene across multiple species to predict if a gene is functionally involved in the biogenesis or maintenance of the cilium. We found that the major improvements in \comparative cell biology" predictions are obtained by maximizing the taxonomic distribution of the species analyzed (representing as many eukaryotic lineages as possible). Lastly in chapter4 we explore the comparative approach using only sequence data. Rabs are a family of GTPases that are master regulators of intracellular trafficking, and are present in all major eukaryotic species. Each different family of Rabs is known to participate in different cellular processes. Therefore being able to identify which family a Rab belongs to allows one to make functional predictions about which processes can occur in a cell. In order to make these predictions possible, a bioinformatics pipeline the Rabifier and accompanying database RabDB.org were developed. This thesis marks the first application of \comparative cell biology" as a framework to study the evolution of the eukaryotic cell. From an evolutionary perspective, the most important finding of this work is that many of the principles we know from \organism" apply equally to the model systems studied in this thesis. Whether these principals hold for other organelles remains to be explored. Most importantly, in each of these chapters, this thesis provides bioinformatics tools for \comparative cell biology". vi Summ´ario Durante centenas de anos os bi´ologost^emestudado a diversidade natural que ocorre em todas as esp´eciesvegetais e animais. A inven¸c~aodo microsc´opio electr´onicona primeira metade do s´eculopassado, ajudou a revelar que as c´elulastamb´empodem ser incrivelmente complexas (e muitas vezes de uma beleza apaixonante). No entanto e apesar do facto de que o ramo da biologia celular existe h´amais de 100 anos, ainda n~aotemos um conhecimento formal de como as c´elulasevoluem: n~ao´eclaro a vastid~aodas c´elulase da morfologia dos organelos, e se e de que maneira, a diversidade pode ser limitante, e de que modo os organelos a mudam sua morfologia ao longo do tempo. O aparecimento da c´elulaeucari´oticah´amais de 1000 milh~oesde anos traduz-se numa das mais importantes transi¸c~oesevolutivas. Neste ramo a arquitetura celular evoluiu a partir de um esbo¸corelativamente simples para um sistema altamente complexo e compartimentalizado de organelos. Uma das formas mais poderosas para estudar a evolu¸c~ao´eestudar a di- versidade atrav´esde uma ampla gama de diferentes esp´ecies:a \abordagem comparativa" para a biologia. No contexto da evolu¸c~aoeucari´oticadesign´amos como \Biologia celular comparativa", a abordagem utilizada nesta tese. Dois sistemas modelo ser~aoestudados usando \biologia celular compar- ativa": o Centro organizador de microt´ubulos(MTOCs) nos captulos 2 e 3 e RabGTPases no cap´ıtulo4.
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