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Identification of Essential and Virulence Genes in Mycoplasma Pneumoniae

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Identification of Essential and Virulence Genes in Mycoplasma Pneumoniae Identification of essential and virulence genes in Mycoplasma pneumoniae Dissertation for the award of the degree “Doctor rerum naturalium” of the Georg-August-Universität Göttingen within the doctoral program “Microbiology & Biochemistry” of the Georg-August-University School of Science (GAUSS) submitted by Cedric Blötz from Northeim Göttingen 2019 THESIS COMMITTEE Prof. Dr. Jörg Stülke (Supervisor and 1st Reviewer) Institute of Microbiology and Genetics Department of General Microbiology, University of Göttingen Prof. Dr. Carsten Lüder (2nd Reviewer) Institute for Medical Microbiology Department for Medical Microbiology, University Medical Center of Göttingen PD Dr. Michael Hoppert (3rd Reviewer) Institute of Microbiology and Genetics Department of General Microbiology, University of Göttingen Further Members of the Examination Board Prof. Dr. Stefanie Pöggeler Institute of Microbiology and Genetics Department of Genetics of Eukaryotic Microorganism Prof. Dr. Fabian M. Commichau Institute of Microbiology and Genetics Department of General Microbiology, University of Göttingen Prof. Dr. Henning Urlaub Max-Planck-Institute for Biophysical Chemistry Bioanalytical Mass Spectrometry Group, Göttingen Date of oral examination: 02.04.2019 I STATEMENT OF AUTHORSHIP I hereby declare that the doctoral thesis entitled, “Identification of essential and virulence genes in Mycoplasma pneumoniae” has been written independently and with no other sources and aids than quoted. Cedric Blötz II DANKSAGUNG Lieber Jörg, gute 20.000 km haben wir mal mehr mal weniger stressfrei bereist. Unglaublich, wie viel ich dank dir und der Arbeit reisen konnte. Ich bin dir für all die Möglichkeiten und Freiheiten, die du mir gegeben hast wirklich dankbar. Schon als Bachelorstudent und auch im Master hatte ich das Gefühl, dass du ein Professor/ Doktorvater bist, dem seine Studenten und Doktoranden und auch ihre Themen wichtig sind. Ich find es toll, dass ich sogut wie immer zu dir konnte, um mir wissenschaftlichen, organisatorischen oder persönlichen Rat zu holen. Darüber hinaus ist es großartig, einen Chef zu haben, mit dem man bei Bier und Sektflöten immer Spaß haben kann. Christina, ohne dich wäre ich im Labor-“handwerklichen“ wie auch in meiner Ideenvielfalt was Versuche betrifft, heute nicht da wo ich bin. Was hätte ich ohne deine Weisheiten aus der Tageszeitung manchmal nur getan. Es schmerzt mich nicht mehr dein Student und Kollege zu sein. Wie gern habe ich mit dir gelacht. Auch dir Katrin möchte ich an dieser Stelle danken. Wie so viele habe ich unglaublich viel von dir gelernt und durch unsere Zusammenarbeit, Diskussionen und Anregungen profitiert. Wie aus euch beiden Lehrmeister, Kollegen und schließlich Freunde wurden ist wunderbar. Freunde. Anika - Larissa, das seid ihr, tolle Freundinnen. Anika, unglaublich wie die Zeit angefangen hat zu rasen seit wir uns im Bachelor kennengelernt haben. Wie oft du mich unterstützt hast. Wie oft ich dich mit Fragen und meiner „Überorganisation“ genervt habe. Wie oft wir zusammen Tränen gelacht haben. Larissa, du warst wohl die beste Studentin, die ich hatte und je haben konnte. Was haben wir gelacht, was hatten wir für Spaß. Du bist immer da, bei allem. Vielen Dank! Hannes du warst ein toller Labornachbar und Kollege. Danke, für so viele anregende Diskussionen und überdachte Versuchsplanungen, toll wie oft wir uns gegenseitig geholfen haben. Es war eine schöne Zeit. Auch allen anderen Kollegen und meinen Studenten (Neil, Alex, Erik, Anika, Tenzin) möchte ich hier danken. Ihr habt mir hier seit 2010 (Bachelor bis Doktor) die Arbeit zu einer unvergesslichen Zeit gemacht. Besonders betonen möchte ich die unermüdliche Arbeit und wirklich immer erquickende Art von Silvia, die nicht nur für mich, sondern für dieser Abteilung eine große Bereicherung ist. Miri, was soll ich sagen? Ich wurde aufgenommen, fast rausgeworfen und doch haben wir in den letzten 4 Jahren unglaubliche Momente erlebt. Eine unglaubliche Zeit endet, aber das ist kein Ende! Mit dir kann man Pferde stehlen, Weinkeltereien eröffnen oder HAIdefinition-Filme gucken. Es ist schön wie wir für einander da sind. Danke für alles. Vielen Dank dass du meine Arbeit so gründlich gelesen hast. „s“! Auf viele weitere schöne Momente! III HoMaMaDa oder auch Mama, Papa, Marc, Darleen - selbst wenn ihr fachlich weniger zu der Arbeit beigetragen habt, hätte ich all das nicht ohne euer Zutun geschafft. Ihr habt mir so viel Mut, Stärke und Liebe gegeben und mich immer unterstützt und oft mehr an mich geglaubt als ich selbst. Vielen Dank Marc, dass du meine Launen und meine fehlende Zeit einfach weggesteckt hast und immer zu mir stehst. Es ist schön eine solch unglaublich tolle Familie zu haben. An dieser Stelle möchte ich mich auch bei meinen Prüfern Carsten und Michael bedanken, die immer ein offenes Ohr hatten. Danke auch dir Fabian für immer volle Sektgläser und interessante Diskussionen. Ein Dank gilt auch den weiteren Mitgliedern meines Prüfungskomitees, Prof. Pöggeler und Prof. Urlaub. Vielen Dank auch all den tollen Kooperationspartnern in fast der ganzen Welt - Francis, Ludwig, Juri, Carole, Dan, Carlos, Roger, Maria, und Luis. IV From a few peaks rising above the fog we try to imagine what the hidden landscape underneath might look like. Pieter W. Postma BIOCHEMIST V VI LIST OF PUBLICATIONS Publications part of the dissertation: Blötz, C., Treffon, K., Kaever, V., Schwede, F., Hammer, E., and Stülke, J. (2017) Identification of the components involved in cyclic di-AMP signaling in Mycoplasma pneumoniae. Front Microbiol 8: 1328. Blötz, C., Lartigue, C., Valverde Timana, Y., Ruiz, E., Paetzold, B., Busse, J., and Stülke, J. (2018) Development of a replicating plasmid based on the native oriC in Mycoplasma pneumoniae. Microbiology 164: 1372–1382. Blötz, C., Krüger, L., Kahle, A., Singh, N., Dickmanns, A., Stülke, J., (2019) How to get rid of peroxides? The detoxification system and its regulation in Mycoplasma pneumoniae. (unpublished) Blötz, C., Singh, N., Dumbke, R., Stülke, J., (2019) Characterization of the immunoglobulin binding protein (IbpM) from Mycoplasma pneumoniae. (unpublished) Other publications: Blötz, C., and Stülke, J. (2017) Glycerol metabolism and its implication in virulence in Mycoplasma. FEMS Microbiol Rev 41: 640–652. Yus, E., Lloréns-Rico. V., Martínez, S., Gallo, C., Eilers, H., Blötz, C., Stülke, J., Lluch- Senar, M., Serrano, L. (2019) Reconstruction of regulatory network in a minimal bacterium reveals extensive non-transcription factor dependent regulation. (submitted) O’Reilly, F., Sinn, L., Blötz, C., Liang, X., Lenz, S., Mahamid, J., Stülke, J., and Rappsilber, J. (2019) In situ structural analysis reveals NusA as a physical link between transcription and translation in Mycoplasma pneumoniae. (unpublished) Other contribution: Bingyao, Z., Blötz, C., and Stülke, J. (2017) MycoWiki-database: mycowiki.uni- goettingen.de VII VIII TABLE OF CONTENT LIST OF ABBREVIATIONS ........................................................................................... XI SUMMARY ................................................................................................................... XIII CHAPTER 1 | Introduction ............................................................................................ 1 AIM OF THE THESIS ...................................................................................... 11 CHAPTER 2 | Cyclic di-AMP in a minimal organism ..................................................... 13 ABSTRACT ..................................................................................................... 14 MATERIALS AND METHODS ......................................................................... 17 RESULTS ....................................................................................................... 22 DISCUSSION .................................................................................................. 26 CHAPTER 3 | New tools for genetic manipulation of Mycoplasmas .............................. 29 ABSRTACT ..................................................................................................... 30 METHODS ...................................................................................................... 33 RESULTS ....................................................................................................... 37 DISCUSSION .................................................................................................. 44 CHAPTER 4 | Immunoglobulin binding protein IbpM ..................................................... 47 ABSTRACT ..................................................................................................... 48 MATERIALS AND METHODS ......................................................................... 51 RESULTS ....................................................................................................... 55 DISCUSSION .................................................................................................. 62 CHAPTER 5 | Peroxide detoxification in Mycoplasmas ................................................. 67 ABSTRACT ..................................................................................................... 68 MATERIALS AND METHODS ......................................................................... 71 RESULTS ....................................................................................................... 78 DISCUSSION .................................................................................................
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