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Molekulargenetische Analysen an Dem Mykophenolsäure Produzierenden Pilz Penicillium Brevicompactum

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Molekulargenetische Analysen an Dem Mykophenolsäure Produzierenden Pilz Penicillium Brevicompactum Molekulargenetische Analysen an dem Mykophenolsäure produzierenden Pilz Penicillium brevicompactum Dissertation zur Erlangung des Grades eines Doktors der Naturwissenschaften an der Fakultät für Biologie und Biotechnologie der Ruhr-Universität Bochum Internationale Graduiertenschule Biowissenschaften Ruhr-Universität Bochum Lehrstuhl für Allgemeine und Molekulare Botanik vorgelegt von Yasaman Mahmoudjanlou aus Teheran, Iran Bochum Mai, 2020 Referent: Prof. Dr. Ulrich Kück Korreferent: Prof. Dr. Dominik Begerow Molecular genetics of the biotechnologically relevant Penicillium brevicompactum Dissertation to obtain the degree Doctor Rerum Naturalium (Dr. rer. nat.) Submitted at the Faculty of Biology and Biotechnology Ruhr-University Bochum International Graduate School of Biosciences Ruhr University Bochum Department of General and Molecular Botany submitted by Yasaman Mahmoudjanlou from Tehran, Iran Bochum May, 2020 1st Supervisor: Prof. Dr. Ulrich Kück 2nd Supervisor: Prof. Dr. Dominik Begerow Danksagung Mein Dank gilt zu allererst meinem Doktorvater Herrn Prof. Dr. Ulrich Kück, für die außerordentlich interessante Themenstellung, die zahlreichen wissenschaftlichen Anregungen und Diskussionen sowie die verantwortungsvollen Aufgaben, welche ich in meiner Zeit als wissenschaftlicher Mitarbeiter am Lehrstuhl wahrnehmen durfte. Aus unserer gemeinsamen Arbeit konnte ich sehr viele Erkenntnisse schöpfen, welche mich auch über das wissenschaftliche Arbeiten hinaus geprägt haben. Zu jeder Zeit konnte ich mich auf Ihre Unterstützung und Ihr herausragendes Engagement verlassen. Bei Herrn Prof. Dr. Dominik Begerow bedanke ich mich für die Übernahme des Korreferats und seine stets offene Tür. Besonderer Dank gilt auch Herrn Prof. Dr. Christian Frisvad, dass er mich so herzlich in der Technical University of Denmark in Lyngby aufgenommen hat und es mir ermöglichte, mein methodisches Repertoire enorm zu erweitern. Auch bedanke ich mich bei allen aktuellen und ehemaligen Mitarbeitern des Lehrstuhls für Allgemeine und Molekulare Botanik. Allen voran Ingeborg Godehardt und Susanne Schlewinski für die Unterstützung bei meinen Experimenten. Dr. Tim Dahlmann für seine guten Ratschläge, gehaltvolle Diskussionen und seine enorme Hilfsbereitschaft, nicht zuletzt bei der Korrektur der vorliegenden Arbeit. Weiterhin gilt mein Dank PD Dr. Ines Teichert für das äußerst gewissenhafte Korrekturlesen und ihre überaus wertvollen Anregungen. PD Dr. Minou Nowrousian für anregende Diskussionen und ihre Hilfsbereitschaft in allen Belangen. Allen meinen Doktorschwestern und Doktorbrüdern möchte ich für die tolle gemeinsam verbrachte Zeit danken! Barbara Ramšak, für gute Ratschläge, welche mir halfen einige herausfordernde Probleme zu lösen und ein immer offenes Ohr – „These Penicillium!“. Dr. Sarah Schmidt, für ihr herzliches, freundliches Wesen und die gemeinsam verbrachte Zeit in Bibliothek und Cafeteria. Valentina Stein, für die regelmäßige Unterstützung bei der Durchführung meiner Experimente und ihr außergewöhnliches Engagement. Ramona Lütkenhaus, für die kontinuierliche Hilfsbereitschaft im Zusammenhang mit Experimenten und/oder diversen Laborgeräten. Nicht zuletzt gilt mein Dank Dr. Daria Radchenko, Ramona Märker, Xuemei Lin, Maria Shariatnasery und Hendrik Strotmeier für die stets inspirierende Zusammenarbeit. Meiner Familie und insbesondere Fabian Jasper-Möller danke ich, für die großartige und fortwährende Unterstützung, welche die vorliegende Arbeit erst ermöglicht haben. Table of Contents I Table of Contents Abbreviations ......................................................................................................................... II I. Introduction ............................................................................................................................................. 1 1. Biology of biotechnologically relevant Penicillium species ........................................................................ 1 1.1 Asexual versus sexual reproduction systems in Penicillia ................................................................. 1 1.1.1 Asexual reproduction .................................................................................................................... 1 1.1.2 Sexual reproduction ...................................................................................................................... 3 1.1.3 Parasexuality ................................................................................................................................. 5 1.2 Taxonomy .......................................................................................................................................... 6 1.2.1 Classification and phylogeny ......................................................................................................... 7 1.2.1.1 Classical classification ............................................................................................................. 7 1.2.1.2 Generic classification.............................................................................................................. 7 1.2.1.3 Infrageneric classification ....................................................................................................... 8 1.2.2 Nomenclature ............................................................................................................................... 9 1.2.3 Identification ............................................................................................................................... 10 1.3 Penicillia as producers of industrially relevant secondary metabolites and elaborator of food products ....................................................................................................................................................... 10 2. Genetic modification of Penicillium species as a tool for functional analysis and strain improvement .. 15 2.1 Conventional generation of recombinant strains ........................................................................... 15 2.2 Directed genetic modification ......................................................................................................... 17 3. Summary .................................................................................................................................................. 23 II. Scope of Thesis ...................................................................................................................................... 24 III. Mahmoudjanlou et al. 2019 ................................................................................................................... 28 IV. Mahmoudjanlou et al. 2020 ............................................................................................................... 29 V. Discussion .............................................................................................................................................. 30 1. Improvement of a transformation system for P. brevicompactum ......................................................... 30 2. Evidence of a heterothallic sexual state and cryptic sexuality in P. brevicompactum ............................. 36 3. Different attempts for induction of a sexual cycle in P. brevicompactum were unsuccessful ................ 39 4. Mating type genes are a suitable molecular marker for identification and phylogeny of Penicillium species .............................................................................................................................................................. 42 5. MAT1-1-1 controls asexual development of P. brevicompactum ............................................................ 45 6. P. brevicompactum MAT1-1-1 transcription factor is associated with germ tube formation and pellet morphology ...................................................................................................................................................... 47 VI. Summary ........................................................................................................................................... 50 VII. Zusammenfassung ............................................................................................................................. 51 VIII. References ......................................................................................................................................... 53 IX. Supplementary Data .............................................................................................................................. 75 X. Eigenanteil an Publikationen ................................................................................................................. 78 XI. Curriculum Vitae .................................................................................................................................... 79 XII. Erklärung ........................................................................................................................................... 81 Abbreviations II Abbreviations Δ deletion AMH anti-Mullerian hormone BiFC bimolecular fluorescence complementation bp base pair CAI codon adaptation index cDNA complementary DNA ChIP-seq chromatin Immunpräzipitation DNA-Sequenzierung CRISPR clustered regularly interspaced palindromic repeats CYA Czapek yeast agar DNA deoxyribonucleic acid dsRNA double-stranded EMSA electrophoretic mobility shift essays EPS Exopolysaccharides flbA fluffy low brlA expression GC3 third position of codons GFP
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