Anna Bichler has joined the lab for her BSc thesis work. She will follow up on some novel candidate proteins that we found to be induced by the Unfolded Protein Response. A warm welcome to our new colleague!
In a collaborative effort spearheaded by the Ahrends lab at the ISAS (Leibniz-Institut für Analytische Wissenschaften) in Dortmund, we established a targeted proteomics approach aimed at analyzing components of the Unfolded Protein Response (UPR), an adaptive signal transduction pathway triggered by the accumulation of unfolded proteins in the endoplasmic reticulum. The UPR comprises an important cellular stress response that aims at re-instating cellular homoeostasis and it plays a key role in a variety of disorders (including diabetes, neurodegenerative disorders, and inflammatory processes). It has also emerged as an attractive target for therapeutic intervention in cancer due to its implication in tumor progression, malignancy and resistance to therapy. The newly developed high-resolution targeted proteomics strategy combines high specificity and sensitivity, allowing the accurate quantification of UPR proteins down to the lower attomol range in a straightforward way without any prior enrichment or fractionation approaches. This has allowed us to determine cellular protein copy numbers of UPR receptors, transducers and effectors, yielding novel insights into an important cellular stress response pathway.
Read the full manuscript at Scientific Reports: Nguyen et al. A sensitive and simple targeted proteomics approach to quantify transcription factor and membrane proteins of the unfolded protein response pathway in glioblastoma cells.
The 24th annual meeting of the RNA Society took place from June 11th to 16th in the beautiful city of Krakow, Poland. It was a scientifically very stimulating conference with great talks on virtually all aspects of RNA biology. I am very happy that our abstract on how Drosophila Sister-of-Sex-lethal antagonizes Sex-lethal auto-regulatory feedback to reinforce a male-specific gene expression pattern was selected for a talk.
The conference also was the perfect opportunity to catch up with colleagues many of which have become close friends over the years – the RNA society truly has become my scientific family!
An EMBO practical course on Measuring Translation Dynamics by Ribosome Profiling is coming up in May 2020. Fantastic speakers are supporting the course including Nicholas Ingolia (University of California, Berkeley, USA), Rachel Green (Johns Hopkins University School of Medicine, Baltimore, USA), Thomas Preiss (The Australian National University, Canberra, AU), Anne Willis (University of Cambridge, UK), Marina Rodnina (Max Planck Institute for Biophysical Chemistry, Göttingen, DE), and Gerben Menschaert (BIOBIX, University of Ghent, BE). The Couse is organized by Sebastian Leidel, Pavel Baranov and Jan Medenbach and will include numerous lectures as well as hands-on training on how to perform ribosome profiling experiments and how to analyze the data. More information will be available soon on the EMBL courses website.
We could successfully extend funding of the Collaborative Research Centre 960 (SFB960) ‘RNP biogenesis: assembly of ribosomal and non-ribosomal RNPs and control of their function’. To continue our ambitious research programs, we are now seeking highly motivated PhD students. We offer a highly competitive research environment and exciting research projects. For more information click here.
Mutations that alter the activity of RNA-binding proteins or their abundance have been implicated in numerous diseases such as neurodegenerative disorders and various types of cancer. This highlights the importance of RBP proteostasis and the necessity to tightly control the expression levels and activities of RBPs. In many cases, RBPs engage in an auto-regulatory feedback by directly binding to and influencing the fate of their own mRNAs, exerting control over their own expression.
Together with our colleagues Michaela Müller-McNicoll from the Institute of Cell Biology and Neuroscience at the Goethe University Frankfurt, Oliver Rossbach from the Institute of Biochemistry at the Justus-Liebig-University Giessen, and Jingyi Hiu at the State Key Laboratory of Molecular Biology (CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology), we have reviewed RBP-mediated autogenous feedback regulation in eukaryotic organisms. For this feedback control, RBPs employ a variety of mechanisms operating at all levels of post-transcriptional regulation of gene expression to either to maintain protein abundance within a physiological range (exerting negative feedback) or to enforce and stabilize cell fate decisions through generation of binary, genetic on/off switches.
The article has just been published in the Journal of Molecular Cell Biology – click here to read the full version.
Good news: the SFB 960 ‘RNP biogenesis: assembly of ribosomes and non-ribosomal RNPs and control of their function’ has been granted another funding period. We are grateful for the generous support by the Deutsche Forschungsgemeinschaft (DFG) that will allow us and our colleagues to continue and extend our scientific programs and to ensure further top-level education of PhD students within the Graduate Research Academy RNA Biology. We want to thank all reviewers that were involved in the selection process and we are looking forward to another four years of exciting science!
A news article on this topic can also be found on the homepage of the University of Regensburg (in German): DFG verlängert Regensburger Sonderforschungsbereich zur Ribosomen-Entstehung
New manuscript published – RhoA regulates translation of the Nogo-A decoy SPARC in white matter-invading glioblastomas
A collaborative effort lead by Björn Tews and supported by the research consortium ‘Systems Biology of the Unfolded Protein Response in Glioma’ (SUPR-G, generously funded by the BMBF in the framework of the e:med initiative) has resulted in a recent publication in Acta Neuropathologica that demonstrates a function of the peptide SPARC in migration and infiltrative growth of glioblastoma cells. SPARC production and secretion is enhanced via regulation of the UPR sensor IRE1 via AKT. SPARC secretion then prevents Nogo-A from inhibiting migration via RhoA. Advanced ultramicroscopy in undissected mouse brains reveals that gliomas require SPARC for invading into white matter structures and its depletion reduces tumor dissemination which significantly prolongs survival and improves response to cytostatic therapy. The discovery of a novel RhoA-IRE1 axis now provides a druggable target for interfering with SPARC production and underscores its therapeutic value. The full publiation can be accessed here.
Mark your calendars! On May 10th at 10a.m. in room VKL 5.1.31, Benedikt Beckmann will talk about novel methods to study RNA-protein interactions during bacterial infection (click here for the advert). Benedikt is a group leader at the Integrated Research Institute (IRI) for the Life Sciences in Berlin, an institute shared between the Humboldt University Berlin, the Charité University Clinic and the Max-Delbrück-Center for Molecular Medicine.
His research focuses on RNA-protein interactions in host cells during infection by pathogenic bacteria which he studies by employing classical molecular biology methods in combination with systems biology approaches and bioinformatics. Using Salmonella typhimurium as a model system, he investigates whether bacterial-derived non-coding RNAs and RNA-binding proteins can directly manipulate host cell gene expression on a post-transcriptional level e.g. to manipulate the immune repsponse of the host.
Recently, Bene established a novel methodological approach to generate RNA interactomes by employing organic extraction for the purification of UV cross-linked ribonucleoproteins (PTex). Previous methods relied on oligo-dT oligonucleotides to capture cross-linked RNPs by hybridization to the poly(A)-tail, limiting their applicability to eukaryotic organisms. Furthermore, Bene will talk about a recently developed machine learning-based tool for the de novo prediction of RNA-binding proteins across a wide variety of species (TriPepSVM).
New manuscript published – Purification of cross-linked RNA-protein complexes by phenol-toluol extraction (PTex)
We are happy that the collaborative effort spearheaded by Benedikt Beckmann at the Integrated Research Institute (IRI) for the Life Sciences has now resulted in a publication. We have described the approach earlier (see here) which, in a nutshell, allows the purification of cross-linked ribonucleoproteins by a series of organic extractions. Access the full article here at Nature Communications.
On February 14th and 15th, the second Regensburg – Canberra Symposium on RNP Biology took place at the John Curtin School of Medical Research at the Australian National University (ANU) in Canberra with the aim to further strengthen our collaborations and to pave the way for establishment of an international PhD program. It was great to meet down under and to hear about the exciting research on RNA biology at the ANU.
I would like to thank our colleagues in Canberra very much for organization of this wonderful meeting and for their generous hospitality.
New manuscript from the lab published in Nucleic Acids Research: Drosophila Sister-of-Sex-lethal reinforces a male-specific gene expression pattern by controlling Sex-lethal alternative splicing.
In a collboration with the labs of Stefan Schneuwly, Gunter Meister (both at the University of Regensburg), Michael Krahn (Westfälische Wilhelms-Universität Münster), and Oliver Rossbach (Justus-Liebig-University Giessen), we could demonstrate that the protein Sister-of-sex-lethal (Ssx) is required in male flies to suppress production of Sex-lethal (Sxl).
Genomic tagging of the Sex-lethal (Sxl) locus in flies to reveal Sxl protein mis-expression (arrowheads) in male flies mutant for Sister-of-Sex-lethal. Arrows mark expression of a Sxl isoform in neural cell bodies and projections.
Most higher eukaryotes reproduce sexually, increasing the variability in the offspring. This allows e.g. rapid adaption to a new (or changing) environment or the cleansing of harmful mutations from a population. Sexual reproduction in higher eukaryotes usually involves individuals of different sex: males and females. Not surprisingly, the genetic programs that determine sex and control sexual differentiation need to be particularly robust in order to ensure survival of the population.
In Drosophila, a single protein, the master regulator Sex-lethal (Sxl), governs female development by controlling the expression of key factors involved in female morphology and behaviour. Once expressed, it engages in an auto-regulatory, positive feedback loop to ensure its sustained expression. This stably ‘flips the switch’ and commits to female development.
In contrast, in males Sxl expression needs to be shut-off which is achieved by alternative splicing that generates RNA isoforms encoding truncated, non-functional Sxl protein. Fluctuations inherent to gene expression can, however, produce small amounts of functional Sxl protein. When left unchallenged, this protein can trigger a self-enforcing cascade resulting in Sxl protein expression snowballing out of control. Until now, however, it remained unclear how males completely shut off the Sxl expression cascade and protect themselves against runaway protein production to ensure robust sex-specific development.
We have discovered a safeguard mechanism that prevents Sxl production in adult male flies. We identified the protein Sister of Sex-lethal (Ssx) as the first antagonist of Sxl-mediated auto-regulatory splicing that defines a precise threshold level for activation of the auto-regulatory, positive feedback loop that controls Sxl expression. We could show that Ssx exerts function by competing with Sxl for the same RNA regulatory elements thus preventing Sxl from triggering the self-enforcing expression cascade in adult male animals.
From October 8th to 10th 2018, a hands-on methods course took place at the University of Regensburg, aimed at training PhD students in the latest methods of ribonucleoprotein (RNP) analysis. Topics included the quantification of protein-RNA interactions, single molecule analyses of RNPs by fluorescence resonance energy transfer (FRET) or fluorescence in situ hybridization (FISH), RNA structure probing, and comprehensive capture of RNA-binding proteins by interactome profiling from cultured cells. The course attracted PhD students from labs around the globe, some travelling from as far as Canberra, Australia, to get insight in and training on a variety of biophysical and biochemical methods. The design of the course was also aimed at fostering interactions between the participants, allowing them to build networks with peers working on related topics. Besides the lectures and the practical hands-on training, it was particularly the vivid and intense discussions among the participants that made this course a big success.
We are extremely grateful for the support of many expert speakers and trainers that delivered lectures on the latest methodological developments and that supervised the experimental work. In particular we want to thank the invited speakers from industry and academia that travelled to Regensburg to provide expert guidance: Rastislav Horos (European Molecular Biology Laboratory, EMBL, Heidelberg), Agatha Korytowski (Malvern Panalytical), Stefanie Nunes Rosa (Swedish University of Agricultural Sciences, Uppsala), Joel Perez-Perri (EMBL Heidelberg), Bruno Sargueil (CNRS/Paris Descartes University), and Ralf Seidel (Peter Debye Institute for Soft Matter Physics, Universität Leipzig).
The course was organized by the Graduate Research Academy RNA Biology of the Collaborative Research Centre 960 (SFB960, Ribosome formation: principles of RNP biogenesis and control of their function) that also provided financial support.
A particular highlight is coming up next week: Prof Thomas Cech will be awarded a honorary doctorate by the University of Regensburg. Tom Cech, now Professor at the University of Colorado at Boulder and director of the BioFrontiers Institute, was awarded the Nobel Prize in chemistry in 1989 jointly with Sidney Altman for the discovery of the catalytic activity of RNA. His revolutionary discovery of self-splicing in Tetrahymena thermophila radically changed the way we were thinking about RNA – what was previously considered a mere carrier of genetic information turned out to play the key function in fundamental biological processes. Exemplarily, protein synthesis is catalyzed by the RNA moiety of the large ribosomal subunit which promotes the peptidyl transfer reaction by which peptide bonds are formed between amino acids. His discovery also had a huge impact on my own scientific career and convinced me that RNA is the most interesting biomolecule to study. I am very much looking forward to the Symposium on Monday, October 22nd (starting 3:30p.m. in lecture hall H51) and to finally get the opportunity to meet Tom Cech in person!