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Molecular insight into ribosome functions and translation regulation

English title Molecular insight into ribosome functions and translation regulation
Applicant Polacek Norbert
Number 143388
Funding scheme Project funding (Div. I-III)
Research institution Departement für Chemie und Biochemie Universität Bern
Institution of higher education University of Berne - BE
Main discipline Molecular Biology
Start/End 01.01.2013 - 30.06.2016
Approved amount 701'173.00
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All Disciplines (2)

Discipline
Molecular Biology
Organic Chemistry

Keywords (4)

non-coding RNAs; Ribosome; Translation regulation; nucleotide analog interference

Lay Summary (German)

Lead
Das Ribosom, aufgebaut aus ribosomaler RNA und ribosomalen Proteinen, ist für die Proteinsynthese in allen lebenden Zellen zuständig. Da das Ribosom den Hauptangriffspunkt vieler klinisch relevanter Antibiotika darstellt, ist dessen funktionelles Verständnis von potentieller Bedeutung für die Bekämpfung von Antibiotika Resistenzen und für die Erzeugung neuer anti-mikrobieller Substanzen.
Lay summary

Molekulare Einsichten in das Ribosom und die Regulation der Proteinsynthese

Inhalt und Ziel des Forschungsprojektes

Ziel dieses Forschungsprojektes ist es, funktionelle Zentren des Ribosoms chemisch zu modifizieren, und dadurch molekulare Details der Proteinsynthese aufzuklären. Dabei wird eine kürzlich von uns entwickelte Methode angewendet werden, die es erlaubt, stellenspezifisch chemisch modifizierte RNA Bausteine in die ribosomale RNA einzubauen. Diese Vorgangsweise vergrössert signifikant den Pool an chemisch unterschiedlichen Gruppen, die spezifisch in das Ribosom eingebracht werden können.

Regulation der Proteinsynthese ermöglicht Zellen, schnell und effizient auf geänderte Umweltbedingungen reagieren zu können. Im zweiten Hauptziel dieses Forschungsvorhabens wollen wir das Potenzial von kleinen nicht-Protein-kodierenden RNAs (ncRNAs) der Bäckerhefe untersuchen, direkt ans Ribosom zu binden und dadurch die Effizienz der Proteinsynthese zu regulieren.

Wissenschaftlicher und gesellschaftlicher Kontext des Forschungsprojekts

Die zentrale Rolle des Ribosoms während der Genexpression spiegelt sich auch in der Tatsache wider, dass mehr als die Hälfte aller bekannten Antibiotika die Translationsmaschinerie als Angriffspunkt haben. Molekulare Einsichten in die funktionsweisen des Ribosoms sind daher nicht nur für das bessere Verständnis der Proteinbiosynthese wesentlich, sondern eröffnen auch neue Möglichkeiten zur Bekämpfung von Antibiotika-resistenten Bakterien.

Direct link to Lay Summary Last update: 28.11.2012

Responsible applicant and co-applicants

Employees

Publications

Publication
cDNA library generation for the analysis of small RNAs by high-throughput sequencing.
Gebetsberger Jennifer, Fricker Roger, Polacek Norbert (2015), cDNA library generation for the analysis of small RNAs by high-throughput sequencing., in Methods in Molecular Biology, 139.
Expression of the vault RNA protects cells from undergoing apoptosis.
Amort Melanie, Nachbauer Birgit, Tuzlak Selma, Kieser Arnd, Schepers Aloys, Villunger Andreas, Polacek Norbert (2015), Expression of the vault RNA protects cells from undergoing apoptosis., in Nature communications, 6, 7030-7030.
Role of a ribosomal RNA phosphate oxygen during the EF-G-triggered GTP hydrolysis.
Koch Miriam, Flür Sara, Kreutz Christoph, Ennifar Eric, Micura Ronald, Polacek Norbert (2015), Role of a ribosomal RNA phosphate oxygen during the EF-G-triggered GTP hydrolysis., in Proceedings of the National Academy of Sciences of the United States of America, 112(20), 2561-8.
The integrity of the G2421-C2395 base pair in the ribosomal E-site is crucial for protein synthesis.
Koch Miriam, Clementi Nina, Rusca Nicola, Vögele Paul, Erlacher Matthias, Polacek Norbert (2015), The integrity of the G2421-C2395 base pair in the ribosomal E-site is crucial for protein synthesis., in RNA biology, 12(1), 70-81.
An mRNA-derived noncoding RNA targets and regulates the ribosome.
Pircher Andreas, Bakowska-Zywicka Kamilla, Schneider Lukas, Zywicki Marek, Polacek Norbert (2014), An mRNA-derived noncoding RNA targets and regulates the ribosome., in Molecular Cell, 147.
Ribosome-associated ncRNAs: an emerging class of translation regulators.
Pircher Andreas, Gebetsberger Jennifer, Polacek Norbert (2014), Ribosome-associated ncRNAs: an emerging class of translation regulators., in RNA Biology, 1335.
Slicing tRNAs to boost functional ncRNA diversity.
Gebetsberger Jennifer, Polacek Norbert (2013), Slicing tRNAs to boost functional ncRNA diversity., in RNA Biology, 1798.

Collaboration

Group / person Country
Types of collaboration
Prof. Alexander Mankin/University of Illinois at Chicago United States of America (North America)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
- Exchange of personnel
Dr. Marek Zywicki/Adam Mickiewicz University Poland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
Prof. Brice Felden/University of Rennes France (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Exchange of personnel
Prof. Ronald Micura/Inst. für Organische Chemie; Universität Innsbruck Austria (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
Dr. David Teis/Innsbruck Biocenter, Medizinische Universität Innsbruck Austria (Europe)
- in-depth/constructive exchanges on approaches, methods or results
Prof. Christian Leumann/Department für Chemie und Biochemie, Universität Bern Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication

Scientific events

Active participation

Title Type of contribution Title of article or contribution Date Place Persons involved
Swiss RNA worshop 2016 Poster The muman vault RNA confers apoptosis resistance to malignantly transformed cells 22.01.2016 Bern, Switzerland Nachbauer Birgit; Bracher Lisamaria; Polacek Norbert;
Swiss RNA workshop 2016 Poster Biology of tRNAs in Trypanosoma brucei during stress 22.01.2016 Bern, Switzerland Polacek Norbert; Cristodero Marina; Fricker Roger;
Swiss RNA workshop 2016 Poster Oxidized bases in the peptidyl transferase center and their effects on translation 22.01.2016 Bern, Switzerland Willi Jessica; Polacek Norbert;
Swiss RNA workshop 2016 Poster Mechanistic insight into function and processing of a ribosome-associated ncRNA (rancRNA) 22.01.2016 Bern, Switzerland Schneider Lukas; Polacek Norbert;
Translation control meeting 2015 Poster Ribosome-associated ncRNAs (rancRNAs) An emerging class of translation regulators 09.09.2015 Heidelberg, Germany Gonskikh Yulia; Polacek Norbert; Fricker Roger; Schneider Lukas;
Protein synthesis and translational control Poster An intergenic non-coding RNA targets and regulates the ribosome in H. volcanii. 09.09.2015 Heidelberg, Germany Polacek Norbert; Wyss Leander;
RNA meeting 2015 Poster Oxidized bases in the ribosome's peptidyl transferase center and their effects on translation 26.05.2015 Madison, United States of America Willi Jessica; Polacek Norbert;
Gordon Research Conference: Translation Machinery in Health & Disease Talk given at a conference The ribosome as novel target for stress-induced small regulatory RNAs 22.02.2015 Ventura, United States of America Polacek Norbert;
Swiss RNA workshop 2015 Poster Oxidized bases in the peptidyl transferase center and their effects on translation 23.01.2015 Bern, Switzerland Willi Jessica; Polacek Norbert;
Swiss RNA workshop 2015 Poster Ribosome associated non-coding RNAs and their regulatory role during translation 23.01.2015 Bern, Switzerland Schneider Lukas; Polacek Norbert;
Swiss RNA workshop 2015 Poster Genomic knock-out of rancRNAs in the archaeon H. volcanii 23.01.2015 Bern, Switzerland Polacek Norbert; Wyss Leander;
Swiss RNA workshop 2015 Poster Biology of tRNA halves in Trypanosoma brucei 23.01.2015 Bern, Switzerland Fricker Roger; Gonskikh Yulia; Polacek Norbert;
Swiss RNA workshop 2016 Poster An intergenic non-coding RNA targets and regulates the ribosome in H. volcanii. 22.01.2015 Bern, Switzerland Polacek Norbert; Wyss Leander;
Annual UK Archaea Workshop Poster Genomic knock-out of rancRNAs in the archaeon H. volcanii 08.01.2015 Nottingham, Great Britain and Northern Ireland Wyss Leander; Polacek Norbert;
RNA Biochemistry 2014 Talk given at a conference Stress-induced translation regulation by ribosome-bound ncRNAs 09.10.2014 Bonn, Germany Polacek Norbert;
DNA habitats and its RNA inhabitants Poster Screening for ncRNAs in Trypanosoma brucei 24.07.2014 Salzburg, Austria Polacek Norbert; Fricker Roger;
Ribosomes & Antibiotics meeting Talk given at a conference Stress-mediated translation regulation by ribosome-bound ncRNAs 09.06.2014 Tartu, Estonia Polacek Norbert;
RNA meeting 2014 Poster Expression of the vault RNA protects cells from undergoing apoptosis 03.06.2014 Quebec City, Canada Nachbauer Birgit; Polacek Norbert;
RNA meeting 2014 Poster tRNA-derived fragments target the small ribosomal subunit to fine-tune translation 03.06.2014 Quebec City, Canada Polacek Norbert;
Swiss RNA workshop 2014 Poster Genomic knock-out of ribosome-associated RNAs in the archaeon H. volcanii 24.01.2014 Bern, Switzerland Polacek Norbert; Wyss Leander;
Swiss RNA workshop 2014 Poster Ribosome-associated non-coding RNAs and their regulatory role during translation 24.01.2014 Bern, Switzerland Polacek Norbert; Schneider Lukas;
Microsymposium on small RNAs Poster Genomic knock-out of ribosome-associated ncRNAs in the archaeon H. volcanii 13.10.2013 Wien, Austria Polacek Norbert; Wyss Leander;
Ribosome Conference 2013 Talk given at a conference The ribosome as novel target for small regulatory RNAs 09.07.2013 Napa Valley, United States of America Polacek Norbert;
Regulating with RNA in Bacteria Talk given at a conference The ribosome as novel target for small regulatory RNAs 04.06.2013 Würzburg, Germany Polacek Norbert;
Swiss RNA workshop 2013 Poster Genomic knock-out of ribosome-associated ncRNAs in the archaeon H. volcanii 25.01.2013 Bern, Switzerland Polacek Norbert; Wyss Leander;


Associated projects

Number Title Start Funding scheme
166527 Stress-mediated effects on ribosome functions and translation control 01.07.2016 Project funding (Div. I-III)
157704 Direct electron detector and phase plate for cryo-transmission electron microscopy of biological samples 01.12.2014 R'EQUIP

Abstract

Molecular insight into ribosome functions and translation regulation1.SUMMARYThe ribosome is the largest known RNA enzyme and is regarded as one of the most ancient catalysts in biology. The ribosome, a multifunctional RNA-protein complex, translates the genome’s message into all proteins needed for life. Its crucial role in gene expression is mirrored by the fact that the ribosome represents the main target for clinical relevant antibiotics. Therefore the knowledge of its functioning is crucial for our understanding of antibiotic resistance and for the future design of new anti-microbial compounds. In this proposal we want to address two fundamental questions: (i)How does the ribosome function at the molecular and atomic level?(ii)How is the ribosome regulated during protein biosynthesis?To gain insight into these topics we will apply an experimental tool that has been established recently in my laboratory that allows manipulating individual functional groups or even single atoms of the large subunit ribosomal RNA. This technique significantly expands the chemical repertoire of residues that can be placed at specific sites within the ribosome and thus clearly exceeds the potential of standard mutagenesis. Here we want to investigate the role of two functional centers of the large ribosomal subunit, namely the ribosomal E-site as well as the peptidyl transferase center. Both of these sites have been shown to be targeted by antibiotics thus highlight their biological relevance. By establishing a novel in vitro selection regime we further want to ‘retro-evolve’ the large ribosomal subunit RNA in order to gain insight into the evolutionary history of this ancient RNA enzyme. Pivotal for cellular life is a well-controlled and coordinated gene expression system. Given the central position of translation in gene expression it is not surprising that protein synthesis is amply targeted by regulatory molecules, mostly regulatory proteins. More recently also small non-protein-coding RNAs (ncRNAs) have been added to the list of regulatory entities (e.g. microRNAs, anti-sense RNAs). However these ncRNAs regulating translation almost exclusively target the messenger RNAs rather than the ribosome directly. In this proposal we want to test the hypothesis, that small ncRNA do exist that directly bind to the ribosome and are capable of fine-tuning its activity. A recently started genomic screen in my group for small ncRNA ribo-regulators in the yeast Saccharomyces cerevisiae revealed hundreds of potential ribosome-associated ncRNAs. With the proposed research we now want to functionally characterize some of the most promising candidates to reveal their regulatory potential.Cumulatively, the proposed research will likely expand our current knowledge of ribosome functions and more precisely will shed new light on the catalytic and regulatory repertoire of non-protein-coding RNAs in biology.
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