Signature Redacted Author

Total Page:16

File Type:pdf, Size:1020Kb

Signature Redacted Author Single-molecule studies of protein degradation ARCHE and kinesin-8 motility MASS^C I STTUTE by by L~oEAPR 15 2015 Yongdae Shin B.S., Seoul National University (2007) LIBRARIES S.M., Massachusetts Institute of Technology (2009) Submitted to the Department of Mechanical Engineering in partial fulfillment of the requirements for the degree of Doctor of Philosophy in Mechanical Engineering at the MASSACHUSETTS INSTITUTE OF TECHNOLOGY February 2015 Massachusetts Institute of Technology 2015. All rights reserved. Signature redacted Author ..................... Department of Mechanical Engineering October 6, 2014 Certified by.redacted Matthew J. Lang Associate Professor of Chemical and Biomolecular Engineering, Vanderbilt University Signature redacted Thesis Supervisor C ertified by ....... ................................. Roger D. Kamm Professor of Mechanical Engineering Signature redacted Chair, Thesis Committee A ccepted by ........... ......................... David E. Hardt Professor of Mechanical Engineering Chairman, Department Committee on Graduate Studies I Single-molecule studies of protein degradation and kinesin-8 motility by Yongdae Shin Submitted to the Department of Mechanical Engineering on October 6, 2014, in partial fulfillment of the requirements for the degree of Doctor of Philosophy in Mechanical Engineering Abstract Molecular machines drive living organisms out of equilibrium and perform critical functions in almost every aspect of cellular processes. They are mechanical enzymes transducing free energy stored in chemical forms to generate motions and forces useful in cell. How these tiny machines operate in the presence of thermal agitation has been studied for a few model systems but still largely elusive. Especially, mechanisms of how molecular machines that were evolved from common ancestors diversified their machine actions to fit specific cellular requirements need to be answered. This thesis explores mechanisms of molecular machines in an effort to reveal roles of thermal fluctuations on machine functions, using two biological systems: ClpXP proteases and kinesin-8 Kif18B. As members of bigger protein families, these two fascinating nanomachines perform important cellular tasks in protein quality control and spindle formation, respectively. AAA+ ClpX unfoldases use energy from ATP binding and hydrolysis to drive mechanical unfolding and translocation of target proteins to associated peptidase ClpP. Previous ensemble biochemical and structural studies uncovered many aspects of degradation activity of ClpXP but the mechanistic understanding of ClpXP func- tion is currently still lacking. We employed single-molecule biophysical techniques including optical trapping and single-molecule fluorescence to directly monitor un- folding and translocation activity of single ClpX hexamer as well as conformational dynamics of single subunit of ClpX. Statistical kinetic analyses on unfolding and translocation uncovered that unfolding kinetics were dominated by futile ATP hy- drolysis and translocation steps contained more than one rate-limiting process for each physical stepping. Single-molecule fluorescence resonance energy transfer (sm- FRET) assay revealed dynamic switching of ClpX subunit conformations between multiple states. In the absence of nucleotide, ClpX explored available conformational spaces thermally in an erratic manner. Nucleotide binding to ClpX hexamer leads to ring contraction as well as defined hexameric conformation arrangements. Con- formational transitions were not directly coupled to ATP hydrolysis, suggesting an important role of thermal fluctuation in ClpX machine function. 3 The Kinesin-8s are plus-end directed motors that negatively regulate microtubule length. The canonical members of this kinesin sub-family showed ultra-processivity which enables Kinesin-8s to enrich preferentially at the plus-ends of microtubules to alter microtubule dynamics. Kif18B is an understudied human Kinesin-8 that also limits MT growth during mitosis. Using single-molecule assays, we found that Kif18B was only modestly processive, and that the motor switched frequently between plus- end directed and diffusive modes of motility. Measurements with truncated motors showed diffusion was promoted by a second MT-binding site located in the Kifl8B tail. Our model accounting for motility switching is consistent with autoinhibition mechanism of Kinesin-1, implying that kinesins may share common regulatory mech- anisms to drive varying functional consequences. Thesis Supervisor: Matthew J. Lang Title: Associate Professor of Chemical and Biomolecular Engineering, Vanderbilt University 4 Acknowledgments Looking back on past years of graduate school, many things have happened in my life. It was not only an academic journey but also a time to look into myself. Facing the end of the graduate life, I am happy that I feel like I know what I want to do and what I should do better than I did 7 years ago. I would like to thank all people who gave me huge support and help and made my graduate life productive and enjoyable. I want to thank my advisor, Matt Lang for having me in his laboratory. When I first came to MIT, I was a naive college graduate and rather a theory-minded person. He trained and guided me to think how to precisely measure things and to build high-end instruments to accomplish that. Now I always think how better measurements can be made to tackle problems facing me. In addition to being a fantastic research advisor, Matt has been a mentor for personal stuffs as well as my career. I really appreciate his support and patience during my stay in his lab and it is just impossible to list here all his role and contribution during my graduate study. I also want to thank Bob Sauer, Tania Baker and Ryoma (Puck) Ohi for giving me a chance to work on ClpXP and Kif18B. Bob and Tania provided insightful advices on ClpXP, without them it would have been impossible to continuously make progress in my ClpX study. Puck introduced me a world of cell biology. His support and encouragement helped me to keep moving on. I am also grateful to my thesis com- mittee, Roger Kamm and Peter So, for their commitment and critical advice during my graduate study. I was very fortunate enough to work with many talented collaborators. Joey Davis and Andreas Martin taught me how to work with ClpXP and made critical contribu- tions to make the first single-molecule fluorescence assay for ClpXP. Adrian Olivares's invaluable contribution made it possible for us to propose the mechanochemical cy- cles of ClpX. I also want to thank Ben Stinson, Andrew Nager and Karl Schmitz for their significant help in developing the fluorescence assay for ClpXP conformational dynamics. Sun Taek Kim introduced me to the exciting world of immunology. I am also thankful to Yaqing Du who provided most kinesins I used in my thesis and taught 5 me biology of the cell division. It has been a great pleasure to interact and share memory with the former and current Lang Lab members; Carlos Castro, David Appleyard, Ricardo Brau, Mo Khalil, Jorge Ferrer, Hyungsuk Lee, Marie Aubin-Tam, Hoi Siew Kit, Bill Hesse, Ted Feldman, Juan Carlos Cordova, Sonia Brady, Harris Manning, Yinnian Feng, Nikki Reinemann, Taishi Zhang, James Smith and Dibyendu Das. Everyday life was enjoyable due to their presence in the lab and it has been great fun to exchange scientific idea with them. I also want to thank my friends in KGSAME for their invaluable support. Finally, and most importantly, I am thankful to my family for their countless encouragement and support; many thanks to Mom, Dad and Nuna. This work is dedicated to my wife, Mihee. It was only possible with her support and love. I cannot finish without mentioning my little boy and girl; Jeongmu and Jeongwon. Their smile made me to forget all hard works in the lab and to realize what is important in my life. 6 Contents 1 Introduction 15 1.1 Molecular Machines ............................ 16 1.1.1 ClpXP Protease ........ .................. 18 1.1.2 K inesin .............................. 21 1.2 Single-Molecule Techniques .............. .. 23 1.2.1 Single-Molecule Fluorescence ...... ............. 24 1.2.2 Optical Trapping ...... ........... ........ 28 2 Monitoring conformational dynamics of single ClpX subunits 31 2.1 Sum m ary ....... ............. ............ 31 2.2 Introduction ........... ............. ........ 32 2.3 R esults .................... ............... 33 2.3.1 Single-Molecule FRET assay for ClpXP ... .......... 33 2.3.2 Nucleotide dependence of ClpX conformation ......... 35 2.3.3 Substrate dependence ...... ............ ..... 39 2.3.4 K inetics ................. ............. 39 2.4 D iscussion .......... ............ ........... 44 2.5 Materials and Methods ...... ......... ........ ... 45 2.5.1 Single-molecule fluorescence assay . ...... ..... .... 45 2.5.2 Data analysis .... ..... ..... .... ..... .... 46 3 Single-molecule kinetics of ClpXP-mediated protein unfolding and translocation 47 7 3.1 Summary . ..... .... ..... ..... .... ..... .. 47 3.2 Introduction ... ...... ..... ...... ...... .... 48 3.3 Results and Discussion .... ..... ...... ...... ... 49 3.3.1 Single-molecule optical trapping assay of ClpXP ...... 49 3.3.2 Single-molecule kinetics ................... 51 3.3.3 Multiple ATP hydrolysis cycles yet single rate limiting step . 56 3.3.4 Translocation kinetics .............. ...... 59 3.4 Materials and Methods .................. ...... 64 3.4.1 Protein constructs . .......... ........... 64 3.4.2 Single-molecule
Recommended publications
  • Josephine Bay Paul Center for Comparative Molecular Biology And
    OCTOBER 2, 2014 CONTACT US DIRECTIONS TO MBL HOME ABOUT MBL EDUCATION RESEARCH GIVING MBL NEWS HOME Josephine Bay Paul Center for Comparative Molecular Biology and Evolution RESOURCES HIGHLIGHTS RESEARCH Josephine Bay Paul Center for Comparative Molecular Biology and Evolution Eugene Bell Center for Regenerative Biology & Tissue Engineering Cellular Dynamics Program The Ecosystems Center Program in Sensory Physiology and Behavior Whitman Center for Research and Discovery EDUCATION MBLWHOI LIBRARY GIFTS PEOPLE In the Josephine Bay Paul Center for Comparative Molecular Biology and Evolution, scientists explore the evolution and interaction of genomes of diverse organisms with significant roles in environmental biology and human health. Bay Paul Center scientists integrate the powerful tools of genome science, molecular phylogenetics, and molecular ecology to advance our understanding of how living organisms are related to each other, to provide the tools to quantify and assess biodiversity, and to identify genes and metabolic processes of ecological and biomedical importance. Publications | Staff List © 1996-2014, The Marine Biological Laboratory MARINE BIOLOGICAL LABORATORY, MBL, and the Join the MBL community: 1888 logo are registered trademarks and service marks of The Marine Biological Laboratory. OCTOBER 2, 2014 CONTACT US DIRECTIONS TO MBL HOME ABOUT MBL EDUCATION RESEARCH GIVING MBL NEWS HOME Bay Paul Center Publications RESOURCES Akerman, NH; Butterfield, DA; and Huber, JA. 2013. Phylogenetic Diversity and Functional Gene Patterns of Sulfur- oxidizing Subseafloor Epsilonproteobacteria in Diffuse Hydrothermal Vent Fluids. Front Microbiol. 4, 185. HIGHLIGHTS RESEARCH Alliegro, MC; and Alliegro, MA. 2013. Localization of rRNA Transcribed Spacer Domains in the Nucleolinus and Maternal Procentrosomes of Surf Clam (Spisula) Oocytes.” RNA Biol.
    [Show full text]
  • Machine-Learning Based Classification of Textual Stimuli To
    MACHINE-LEARNING BASED CLASSIFICATION OF TEXTUAL STIMULI TO PROMOTE IDEATION IN BIOINSPIRED DESIGN A Dissertation by MICHAEL W. GLIER Submitted to the Office of Graduate Studies of Texas A&M University in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY Chair of Committee, Daniel A. McAdams Co-Chair of Committee, Julie S. Linsey Committee Members, Richard J Malak Tracy A. Hammond Head of Department, Andreas Polycarpou August 2013 Major Subject: Mechanical Engineering Copyright 2013 Michael W. Glier ABSTRACT Bioinspired design uses biological systems to inspire engineering designs. One of bioinspired design’s challenges is identifying relevant information sources in biology for an engineering design task. Currently information can be retrieved by searching biology texts or journals using biology-focused keywords that map to engineering functions. However, this search technique can overwhelm designers with unusable results. This work explores the use of text classification tools to identify relevant biology passages for design. Further, this research examines the effects of using biology passages as stimuli during idea generation. Four human-subjects studies are examined in this work. Two surveys are performed in which participants evaluate sentences from a biology corpus and indicate whether each sentence prompts an idea for solving a specific design problem. The surveys are used to develop and evaluate text classification tools. Two idea generation studies are performed in which participants generate and record solutions for designing a corn shucker using either different sets of biology passages as design stimuli, or no stimuli. Based 286 sentences from the surveys, a k Nearest Neighbor classifier is developed that is able to identify helpful sentences relating to the function “separate” with a precision of 0.62 and recall of 0.48.
    [Show full text]
  • A Study of the Cell Biology of Motility in Eimeria Tenella Sporozoites
    A STUDY OF THE CELL BIOLOGY OF MOTILITY IN Eimeria tenella SPOROZOITES by David Robert Bruce Department of Biology University College London A thesis presented for the degree of Doctor of Philosophy in the University of London 2000 ProQuest Number: U643145 All rights reserved INFORMATION TO ALL USERS The quality of this reproduction is dependent upon the quality of the copy submitted. In the unlikely event that the author did not send a complete manuscript and there are missing pages, these will be noted. Also, if material had to be removed, a note will indicate the deletion. uest. ProQuest U643145 Published by ProQuest LLC(2016). Copyright of the Dissertation is held by the Author. All rights reserved. This work is protected against unauthorized copying under Title 17, United States Code. Microform Edition © ProQuest LLC. ProQuest LLC 789 East Eisenhower Parkway P.O. Box 1346 Ann Arbor, Ml 48106-1346 ABSTRACT A study on the cell biology of motility inEimeria tenella sporozoites Eimeria tenella is an obligate intracellular parasite within the phylum Apicomplexa. It is the causative agent of coccidiosis in domesticated chickens and under modem farming conditions can have a considerable economic impact. Motility is employed by the sporozoite to effect release from the sporocyst and enable invasion of appropriate host cells and occurs at an average speed of 16.7 ± 6 pms'\ Frame by frame video analysis of gliding motility shows it to be an erratic non­ substrate specific process and this observation was confirmed by studies of bead translocation across the cell surface occurring at an average speed of 16.9 ± 7.6 pms'^ Incubation with cytochalasin D, 2,3-butanedione monoxime and colchicine, known inhibitors of the motility associated proteins actin, myosin and tubulin respectively, indicated that it is an actomyosin complex which generates the force to power sporozoite motility.
    [Show full text]
  • Spatial Organization of Large Cells
    Spatial Organization of Large Cells A dissertation presented by Martin Helmut Wuehr to The Committee on Higher Degrees in Systems Biology in partial fulfillment of the requirements for the degree of Doctor of Philosophy in the subject of Systems Biology Harvard University Cambridge, Massachusetts March 2010 i © 2010 – Martin H. Wuehr All rights reserved. ii Timothy John Mitchison Martin Helmut Wuehr Spatial Organization of Large Cells Abstract The rationale for my research was to investigate unusually large cells, fertilized frog and fish eggs, to obtain a unique perspective on a cell’s spatial organization with a focus on cell division. First, we investigated how spindle size changes during cleavage stages while cell size changes by orders of magnitude. To do so we improved techniques for immunofluorescence in amphibian embryos and generated a transgenic fish line with fluorescently labeled microtubules. We show that in smaller cells spindle length scales with cell length, but in very large cells spindle length approaches an upper limit and seems uncoupled from cell size. Furthermore, we were able to assemble mitotic spindles in embryonic extract that had similar size as in vivo spindles. This indicates that spindle size is set by a mechanism that is intrinsic to the spindle and not downstream of cell size. Second we investigated how relatively small spindles in large cells are positioned, and oriented for symmetric cell division. We show that the localization and orientation of these spindles are determined by location and iii orientation of sister centrosomes set by the anaphase-telophase aster of the previous cycle. Third we researched the mechanism by which asters center and align centrosomes relative to the longest axis.
    [Show full text]
  • Cell Biology Annual Meeting Program
    Cell Biology Annual Meeting Program Shirley Tilghman, President Julie Theriot, Program Chair Karen Oegema, Local Organizer ascb.org/2015meeting Search Your Smartphone App Store for ASCB2015 Submit Your Research Committed to Excellence, Rigor, and Breadth eNeuro has joined JNeurosci in the PubMed Central database. Learn more at SfN.org DON’T MISS OUT ON ANOTHER ISSUE! SUBSCRIBE Sign up now to receive FOR FREE* the monthly print magazine. TODAY! Each issue contains feature articles on hot new trends in science, profiles of top-notch researchers, reviews of the latest tools and technologies, and much, much more. * Only available for a limited time. Not all requests qualify for a free subscription. www.the-scientist.com/subscribe The Company of Biologists is a UK based charity and not-for-profit publisher run by biologists for biologists. The Company aims to promote research and study across all branches of biology through the publication of its five journals. Development Advances in developmental biology and stem cells dev.biologists.org Journal of Cell Science The science of cells jcs.biologists.org The Journal of Experimental Biology At the forefront of comparative physiology and integrative biology jeb.biologists.org Disease Models & Mechanisms Basic research with translational impact dmm.biologists.org Biology Open Facilitating rapid peer review for accessible research bio.biologists.org In addition to publishing, The Company makes an important contribution to the scientific community, providing grants, travelling fellowships and sponsorship
    [Show full text]
  • Opposing Motor Activities Are Required for the Organization of the Mammalian Mitotic Spindle Pole Tirso Gaglio,* Alejandro Saredi,* James B
    Opposing Motor Activities Are Required for the Organization of the Mammalian Mitotic Spindle Pole Tirso Gaglio,* Alejandro Saredi,* James B. Bingham,* M. Josh Hasbani,* Steven R. Gill,* Trina A. Schroer,* and Duane A. Compton* *Department of Biochemistry, Dartmouth Medical School, Hanover, New Hampshire 03755; and*Department of Biology, 220A Mudd Hall, The Johns Hopkins University, Baltimore, Maryland 21218 Abstract. We use both in vitro and in vivo approaches in cultured cells results in the complete lack of organi- to examine the roles of Eg5 (kinesin-related protein), zation of microtubules and the failure to efficiently con- cytoplasmic dynein, and dynactin in the organization of centrate the NuMA protein despite its association with the microtubules and the localization of NuMA (Nu- the microtubules. Simultaneous immunodepletion of clear protein that associates with the Mitotic A__ppara- these proteins from the cell free system for mitotic aster tus) at the polar ends of the mammalian mitotic spindle. assembly indicates that the plus end--directed activity of Perturbation of the function of Eg5 through either im- Eg5 antagonizes the minus end--directed activity of cy- munodepletion from a cell free system for assembly of toplasmic dynein and a minus end--directed activity as- mitotic asters or antibody microinjection into cultured sociated with NuMA during the organization of the mi- cells leads to organized astral microtubule arrays with crotubules into a morphologic pole. Taken together, expanded polar regions in which the minus ends of the these results demonstrate that the unique organization microtubules emanate from a ring-like structure that of the minus ends of microtubules and the localization contains NuMA.
    [Show full text]
  • Measuring Mitotic Spindle Dynamics in Budding Yeast
    Measuring mitotic spindle dynamics in budding yeast Kemp Plumb B.Sc. Department of Physics McGill University Montreal,Qu´ ebec´ July, 2009 A thesis submitted to McGill University in partial fulfillment of the requirements of the degree of Master of Science c Kemp Plumb, 2009 Library and Archives Bibliothèque et Canada Archives Canada Published Heritage Direction du Branch Patrimoine de l’édition 395 Wellington Street 395, rue Wellington Ottawa ON K1A 0N4 Ottawa ON K1A 0N4 Canada Canada Your file Votre référence ISBN: 978-0-494-66191-8 Our file Notre référence ISBN: 978-0-494-66191-8 NOTICE: AVIS: The author has granted a non- L’auteur a accordé une licence non exclusive exclusive license allowing Library and permettant à la Bibliothèque et Archives Archives Canada to reproduce, Canada de reproduire, publier, archiver, publish, archive, preserve, conserve, sauvegarder, conserver, transmettre au public communicate to the public by par télécommunication ou par l’Internet, prêter, telecommunication or on the Internet, distribuer et vendre des thèses partout dans le loan, distribute and sell theses monde, à des fins commerciales ou autres, sur worldwide, for commercial or non- support microforme, papier, électronique et/ou commercial purposes, in microform, autres formats. paper, electronic and/or any other formats. The author retains copyright L’auteur conserve la propriété du droit d’auteur ownership and moral rights in this et des droits moraux qui protège cette thèse. Ni thesis. Neither the thesis nor la thèse ni des extraits substantiels de celle-ci substantial extracts from it may be ne doivent être imprimés ou autrement printed or otherwise reproduced reproduits sans son autorisation.
    [Show full text]
  • October- 2018 Journal.Pmd
    Current Trends in Biotechnology and Pharmacy ISSN 0973-8916 (Print), 2230-7303 (Online) Editors Prof.K.R.S. Sambasiva Rao, India Prof. Karnam S. Murthy, USA krssrao@abap.co.in skarnam@vcu.edu Editorial Board Prof. Anil Kumar, India Dr.P. Ananda Kumar, India Prof. P.Appa Rao, India Prof. Aswani Kumar, India Prof. Bhaskara R.Jasti, USA Prof. Carola Severi, Italy Prof. Chellu S. Chetty, USA Prof. K.P.R. Chowdary, India Dr. S.J.S. Flora, India Dr. Govinder S. Flora, USA Prof. H.M. Heise, Germany Prof. Huangxian Ju, China Prof. Jian-Jiang Zhong, China Dr. K.S.Jagannatha Rao, Panama Prof. Kanyaratt Supaibulwatana, Thailand Prof.Juergen Backhaus, Germany Prof. Jamila K. Adam, South Africa Prof. P.B.Kavi Kishor, India Prof. P.Kondaiah, India Prof. M.Krishnan, India Prof. Madhavan P.N. Nair, USA Prof. M.Lakshmi Narasu, India Prof. Mohammed Alzoghaibi, Saudi Arabia Prof.Mahendra Rai, India Prof. Milan Franek, Czech Republic Prof.T.V.Narayana, India Prof. Nelson Duran, Brazil Dr. Prasada Rao S.Kodavanti, USA Prof. Mulchand S. Patel, USA Dr. C.N.Ramchand, India Dr. R.K. Patel, India Prof. P.Reddanna, India Prof. G.Raja Rami Reddy, India Dr. Samuel J.K. Abraham, Japan Dr. Ramanjulu Sunkar, USA Dr. Shaji T. George, USA Prof. B.J. Rao, India Prof. Sehamuddin Galadari, UAE Prof. Roman R. Ganta, USA Prof. B.Srinivasulu, India Prof. Sham S. Kakar, USA Prof. B. Suresh, India Dr. N.Sreenivasulu, Germany Prof. Swami Mruthinti, USA Prof.Sung Soo Kim, Korea Prof. Urmila Kodavanti, USA Prof. N. Udupa, India Assistant Editors Dr.Giridhar Mudduluru, Germany Dr.
    [Show full text]
  • Philipp M. Niethammer Cell Biology Program, Memorial Sloan Kettering Cancer Center, 1275 York Avenue New York, NY 10065, USA
    Philipp M. Niethammer Cell Biology Program, Memorial Sloan Kettering Cancer Center, 1275 York Avenue New York, NY 10065, USA. Box: 278, Phone: 212-639-5332 (office), 617-910-7167 (cell), niethamp@mskcc.org ORCID: https://orcid.org/0000-0003-4175-4012 Curriculum Vitae PERSONAL STATEMENT I am an experimental biologist with a background in biochemistry, cell biology, and advanced imaging techniques. For my PhD and postdoc, I trained with Eric Karsenti, Philippe Bastiaens (EMBL Heidelberg, Germany), and Tim Mitchison (Harvard Medical School, Boston, USA), using frogs and zebrafish as experimental systems. My general biological interest is to understand the biochemical and biophysical basis of morphogenesis in cells, tissues, and organisms. I am interested in the question of how biochemical signaling circuits generate and process spatiotemporal information to spatially guide morphogenetic events and how this warrants the specificity of a morphogenetic response. One of my primary research directions, past and future, has been the development of new probes and microscopy-based methods to image these circuits inside living cells, tissues, and organisms. This approach was exemplified by my development of novel optical probes for microtubule regulation during my PhD and the first implementation of a ge- netically encoded H2O2 sensor in a vertebrate animal during my postdoc. My PhD and postdoctoral training in the Department of Cell Biology and Biophysics at EMBL Heidelberg and the Department of Systems Biology at Harvard Medical School have minted me with a physical perspective on biological problems. In my lab, I have further developed quantitative intravitral imaging approaches to decipher the early regulation of inflammation and healing after tissue damage in zebrafish.
    [Show full text]
  • The Role of GCP8 in Microtubule Nucleation and Cell Cicle Progression
    The role of GCP8 in microtubule nucleation and cell cicle progression Artur Ezquerra Gonzalez ADVERTIMENT. La consulta d’aquesta tesi queda condicionada a l’acceptació de les següents condicions d'ús: La difusió d’aquesta tesi per mitjà del servei TDX (www.tdx.cat) i a través del Dipòsit Digital de la UB (diposit.ub.edu) ha estat autoritzada pels titulars dels drets de propietat intel·lectual únicament per a usos privats emmarcats en activitats d’investigació i docència. No s’autoritza la seva reproducció amb finalitats de lucre ni la seva difusió i posada a disposició des d’un lloc aliè al servei TDX ni al Dipòsit Digital de la UB. No s’autoritza la presentació del seu contingut en una finestra o marc aliè a TDX o al Dipòsit Digital de la UB (framing). Aquesta reserva de drets afecta tant al resum de presentació de la tesi com als seus continguts. En la utilització o cita de parts de la tesi és obligat indicar el nom de la persona autora. ADVERTENCIA. La consulta de esta tesis queda condicionada a la aceptación de las siguientes condiciones de uso: La difusión de esta tesis por medio del servicio TDR (www.tdx.cat) y a través del Repositorio Digital de la UB (diposit.ub.edu) ha sido autorizada por los titulares de los derechos de propiedad intelectual únicamente para usos privados enmarcados en actividades de investigación y docencia. No se autoriza su reproducción con finalidades de lucro ni su difusión y puesta a disposición desde un sitio ajeno al servicio TDR o al Repositorio Digital de la UB.
    [Show full text]
  • A Decade of Molecular Cell Biology: Achievements and Challenges
    PERSPECTIVES a reaction–diffusion mechanism. I attribute 10-YEAR ANNIVERSARY SERIES — VIEWPOINT this concept to Eric Karsenti, who mooted the idea in the mid 1980s for signals dif- A decade of molecular cell biology: fusing away from DNA in eggs. However, it wasn’t proven until the development of fluorescence resonance energy transfer achievements and challenges (FRET)-based activity biosensors in the past decade2,3. In general, fluorescence sensors Asifa Akhtar, Elaine Fuchs, Tim Mitchison, Reuben J. Shaw, Daniel St Johnston, of biochemical activity are a very important Andreas Strasser, Susan Taylor, Claire Walczak and Marino Zerial development. Abstract | Nature Reviews Molecular Cell Biology celebrated its 10‑year anniversary Reuben J. Shaw. One area close to our own during this past year with a series of specially commissioned articles. work is the unexpected re-emergence of To complement this, here we have asked researchers from across the field for their metabolism and its relationship to growth insights into how molecular cell biology research has evolved during this past control and cancer. Advances in autophagy decade, the key concepts that have emerged and the most promising interfaces continue to amaze me in terms of how little basic information we actually have on how a that have developed. Their comments highlight the broad impact that particular cell works. Autophagy regulators are highly advances have had, some of the basic understanding that we still require, and the conserved proteins in a central cell biologi- collaborative approaches that will be essential for driving the field forward. cal process that is deregulated in common human diseases, yet much of the biochemi- cal framework for this process has been What do you feel have been the Takahashi were paradigm-shifting.
    [Show full text]
  • On the Role of Microtubules in Cell Polarity a Reconstituted Minimal System Vendel, K.J.A
    Delft University of Technology On the role of microtubules in cell polarity A reconstituted minimal system Vendel, K.J.A. DOI 10.4233/uuid:34f174d5-f8ad-4bf2-86f7-47660a84fe64 Publication date 2020 Document Version Final published version Citation (APA) Vendel, K. J. A. (2020). On the role of microtubules in cell polarity: A reconstituted minimal system. https://doi.org/10.4233/uuid:34f174d5-f8ad-4bf2-86f7-47660a84fe64 Important note To cite this publication, please use the final published version (if applicable). Please check the document version above. Copyright Other than for strictly personal use, it is not permitted to download, forward or distribute the text or part of it, without the consent of the author(s) and/or copyright holder(s), unless the work is under an open content license such as Creative Commons. Takedown policy Please contact us and provide details if you believe this document breaches copyrights. We will remove access to the work immediately and investigate your claim. This work is downloaded from Delft University of Technology. For technical reasons the number of authors shown on this cover page is limited to a maximum of 10. ON THE ROLE OF MICROTUBULES IN CELL POLARITY A RECONSTITUTED MINIMAL SYSTEM ON THE ROLE OF MICROTUBULES IN CELL POLARITY A RECONSTITUTED MINIMAL SYSTEM Proefschrift ter verkrijging van de graad van doctor aan de Technische Universiteit Delft, op gezag van de Rector Magnificus Prof. dr. ir. T. H. J. J. van der Hagen, voorzitter van het College voor Promoties, in het openbaar te verdedigen op 21 september 2020 om 12:30 uur door Kim Jikke Anna VENDEL Master in physics, Universiteit Leiden, Nederland geboren te Amsterdam, Nederland.
    [Show full text]