Stinus Ruiz de Gauna, Sonia

English

DNA, the molecule that contains the genetic information encoding the characteristics of a given organism, is mostly confined within the nucleus of the cells that compose the organism. The telomeres are the physical ends of the DNA molecule and their function is to protect it. Telomerase is the protein in charge of the maintenance of telomeres. Telomere dysfunction has severe consequences for human health and is a hallmark of ageing and cancer. However, it is not yet well understood what minimally defines a telomere. Specifically, we used baker’s yeast to study how telomeres are distinguished from naturally occurring breaks within the DNA molecule. Telomeres and DNA breaks resemble each other but are processed differently: while telomeres are maintained by telomerase, DNA breaks are repaired by a set of specialized proteins. It is essential to distinguish between telomeres and DNA breaks, and failure to do so can have fatal consequences. We determined the minimum length of a telomere to still be sensed as such and the key protein, Cdc13, that sets this minimum length. We have also studied telomeric G-quadruplexes, structures that form within the telomere that are proposed to influence telomere biology. We have found that G-quadruplexes protect telomeres when other telomere protecting mechanisms (e.g. telomerase) are absent, but their role is not essential. Thus, we have determined that a minimum telomere length and the Cdc13 protein are essential features of telomeres, but G-quadruplexes are not

Proceedings - University of Groningen

   University of GroningenOn the molecular biology of telomeresStinus Ruiz de Gauna, SoniaIMPORTANT NOTE: You are advised to consult the publisher's version (publisher's PDF) if you wish to cite fromit. Please check the document version below.Document VersionPublisher's PDF, also known as Version of recordPublication date:2018Link to publication in University of Groningen/UMCG research databaseCitation for published version (APA):Stinus Ruiz de Gauna, S. (2018). On the molecular biology of telomeres: Lessons from budding yeast.University of Groningen.CopyrightOther than for strictly personal use, it is not permitted to download or to forward/distribute the text or part of it without the consent of theauthor(s) and/or copyright holder(s), unless the work is under an open content license (like Creative Commons).The publication may also be distributed here under the terms of Article 25fa of the Dutch Copyright Act, indicated by the “Taverne” license.More information can be found on the University of Groningen website: https://www.rug.nl/library/open-access/self-archiving-pure/taverne-amendment.Take-down policyIf you believe that this document breaches copyright please contact us providing details, and we will remove access to the work immediatelyand investigate your claim.Downloaded from the University of Groningen/UMCG research database (Pure): http://www.rug.nl/research/portal. For technical reasons thenumber of authors shown on this cover page is limited to 10 maximum.Download date: 03-06-2022On the molecular biologyof telomeresLessons from budding yeastSonia Stinus Ruiz de GaunaOn the molecular biology of  telomeres - Lessons from budding yeastThe work presented in this thesis was conducted at the European Research Institute for the Biology of  Ageing, University Medical Center Groningen, University of  Gron-ingen, Groningen, the Netherlands.Printed by: ProefschriftMaken | www.proefschriftmaken.nlISBN (print version): 978-94-6380-078-5ISBN (digital version): 978-94-6380-075-4Copyright © 2018 by Sonia Stinus Ruiz de GaunaAll rights reserved. No parts of  this book may be reproduced or transmitted in any form or by any means without prior permission of  the author.On the molecular biology of telomeresLessons from budding yeastPhD thesis to obtain the degree of PhD at theUniversity of Groningenon the authority of theRector Magnificus Prof. E. Sterkenand in accordance withthe decision by the College of Deans.This thesis will be defended in public on Monday 3 December 2018 at 11.00 hoursby Sonia Stinus Ruiz de Gauna born on 18 April 1989in Donostia - San Sebastián, SpainSupervisorProf. G. de Haan Co-supervisorDr. M. Chang Assessment CommitteeProf. M.A.T.M. van Vugt Prof. F. Foijer Prof. B. LukeTable of contentsChapter 1: Introduction  71. Telomeres 91.1. Structure of  telomeres 91.2. Protection of  Saccharomyces cerevisiae telomeres  101.2.1. Single-stranded telomeric DNA binding proteins 101.2.2. Double-stranded telomeric DNA binding proteins  111.2.3. Telomere-associated proteins 121.3. Regulation of  telomere length 141.3.1. Telomere length regulation by homologous recombination mechanisms  141.4. Transcription at telomeres: telomere position effect and TERRA 151.5. Biology of  mammalian telomeres 161.5.1. Telomere structure and composition of  the shelterin complex 161.5.2. Regulation of  telomere length 181.5.3. Transcription of  human telomeres: telomere position effect and TERRA 182. G-quadruplexes  192.1. Role of  G-quadruplexes in replication 202.2. Role of  G-quadruplexes at telomeres 212.2.1. Telomerase regulation by G-quadruplexes  222.2.2. Telomere protection by G-quadruplexes 223. Thesis overview 23Chapter 2: A sharp Pif1-dependent threshold separates DNA double-strand breaks from critically short telomeres  25Abstract 26Introduction 27Results 28Identification of  a Pif1-insensitivity threshold at DNA ends 28A DSB-telomere transition also exists at chromosome ends 30Pif1 is not inhibited by DNA damage kinases  33Artificial telomerase recruitment does not outcompete Pif1 34The DSB-telomere transition recapitulates the differential regulation of  Pif1 36Investigating the molecular trigger of  the DSB-telomere transition 37Cdc13 function influences the fate of  DNA ends 38Discussion 43Methods 45Supplementary data 50Table of contentsChapter 3: Telomerase regulation by the Pif1 helicase – a length-dependent effect?  61Chapter 4: Investigating the role of  G-quadruplexes at Saccharomyces cerevisiae telomeres  69Abstract 70Introduction 71Results 72G-quadruplexes mediate a non-essential telomere protection function 72tlc1-tm cells senesce very rapidly in the absence of  telomerase 74Telomerase-dependent telomere extension is dramatically increased at tlc1-tm telomeres 75Telomere binding proteins are affected in tlc1-tm telomeres 76Telomere homeostasis is altered in tlc1-tm cells 78tlc1-tm repeats are not counted as telomeric sequence in terms of  telomere length homeostasis 80Discussion 81Materials & methods 83Supplementary data 88Chapter 5: Discussion and future perspectives 91A ~40 nt length threshold separates telomeres from DSBs 92Development of  the inducible STEX assay 93Cdc13-independent telomerase recruitment to chromosome ends 94Non-essential G-quadruplex-mediated telomere protection 94Characterisation of  a Rap1-free telomere 95Bibliography 97Appendix 113Nederlandse samenvatting 114Summary in English 116Resumen en español 118Résumé en français 120Curriculum vitae 124List of  publications 125Acknowledgements 126

On the molecular biology of telomeres:Lessons from budding yeast

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On the molecular biology of telomeres:Lessons from budding yeast

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Proceedings - University of Groningen