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ERC Implementing Arrangements Call for Expression of Interest 2017 ERC Implementing Arrangements Call for Expression of Interest 2017 Project ID: Project Acronym: Evaluation Panel: LS1 - Molecular and 637987 ChromArch Structural Biology and Biochemistry Principal Investigator: Dr Christof Gebhardt Host Institution: Universitaet Ulm - DE Single Molecule Mechanisms of Spatio-Temporal Chromatin Architecture Chromatin packaging into the nucleus of eukaryotic cells is highly sophisticated. It not only serves to condense the genomic content into restricted space, but mainly to encode epigenetic traits ensuring temporally controlled and balanced transcription of genes and coordinated DNA replication and repair. The non-random three-dimensional chromatin architecture including looped structures between genomic control elements relies on the action of architectural proteins. However, despite increasing interest in spatio-temporal chromatin organization, mechanistic details of their contributions are not well understood.With this proposal I aim at unveiling molecular mechanisms of protein–mediated chromatin organization by in vivo single molecule tracking and quantitative super-resolution imaging of architectural proteins using reflected light sheet microscopy (RLSM). I will measure the interaction dynamics, the spatial distribution and the stoichiometry of architectural proteins throughout the nucleus and at specific chromatin loci within single cells. In complement single molecule force spectroscopy experiments using magnetic tweezers (MT), I will study mechanisms of DNA loop formation in vitro by structure-mediating proteins. Integrating these spatio-temporal and mechanical single molecule information, I will in the third sup-project measure the dynamics of relative end-to-end movements and the forces acting within a looped chromatin structure in living cells.Taken together, my experiments will greatly enhance our mechanistic understanding of three-dimensional chromatin architecture and inspire future experiments on its regulatory effects on nuclear functions and potential therapeutic utility upon controlled modification. Keywords of the ERC project: chromatin topology, single molecule imaging, super-resolution imaging, force spectroscopy Keywords that characterize the scientific profile of the potential visiting researcher/s: physicist, experience in instrument development, interest in biological questions Index: -1 - Project ID: Project Acronym: Evaluation Panel: LS1 - Molecular and 309433 EPIPLURETRO Structural Biology and Biochemistry Principal Investigator: Dr Jose L. Garcia-Perez Host Institution: FUNDACION PUBLICA ANDALUZA PROGRESO Y SALUD - ES Epigenetic control and impact of mammalian retrotransposons in pluripotent genomes Almost half of the human genome is made of Transposable Elements (TEs), whose ongoing activity continually impacts our genome. However, little is known about how the host regulates TEs and their genomic and epigenomic impacts. EpiPluriRetro will advance research in a new groundbreaking concept: that TEs are active in our pluripotent genome, and that epigenetic regulation is employed therein to regulate TE activity. LINE-1 retrotransposons comprise approximately 20% of the mammalian genome, and L1 retrotransposition events can create genetic diversity by a variety of mechanisms. From acting as simple insertion mutagens to inducing other complex genomic alterations it is becoming increasingly evident that the activity of TEs is a major force driving human genome evolution. It has been demonstrated that the main mutagenic load associated with TE mobilization occurs during early human embryogenesis (i.e., our pluripotent genome). EpiPluriRetro will examine how epigenetic mechanisms influence LINE-1 retrotransposition in pluripotent cells. To do that, we will combine genetic, biochemical and genomics approaches to identify pluripotent host factors that influence the fate of LINE-1 retrotransposition. In addition, EpiPluriRetro will analyze the impact of LINE-1 insertions in our pluripotent genome and the Epimutagenic impact of new LINE-1 mobilization events in pluripotent cells. To do that, we have developed an innovative approach to analyze the effect of LINE-1 insertions within human genes without biases, including epigenetic alterations induced by a new L1 insertion. EpiPluriRetro will help to understand how the activity of TEs is controlled in our heritable genome, which will directly impact our knowledge in how new genetic diseases are generated in humans. In addition, EpiPluriRetro will allow us to describe a new concept in human biology, as we will analyze how new TE insertions can modify the chromatin status of flanking genomic regions where they insert. Keywords of the ERC project: Epigenetics Retrotransposons, pluripotent genome, mutataion, genomic instability, Stem cell biology Keywords that characterize the scientific profile of the potential visiting researcher/s: Transposable Elements, Stem cell biology, Epiogenetics Index: -2 - Project ID: Project Acronym: Evaluation Panel: LS1 - Molecular and 340551 Birtoaction Structural Biology and Biochemistry Principal Investigator: Dr Laszlo Tora Host Institution: Centre Europeen De Recherche En Biologie Et Medecine - FR From birth to action: regulation of gene expression through transcription complex biogenesis Transcriptional regulation of protein coding genes in eukaryotic cells requires a complex interplay of sequence- specific DNA-binding factors, co-activators, general transcription factors (GTFs), RNA polymerase II and the epigenetic status of target sequences. Nuclear transcription complexes function as large multiprotein assemblies and are often composed of functional modules. The regulated decision-making that exists in cells governing the assembly and the allocation of factors to different transcription complexes to regulate distinct gene expression pathways is not yet understood. To tackle this fundamental question, we will systematically analyse the regulated biogenesis of transcription complexes from their sites of translation in the cytoplasm, through their assembly intermediates and nuclear import, to their site of action in the nucleus. The project will have four main Aims to decipher the biogenesis of transcription complexes: I) Investigate their co-translation-driven assembly II) Determine their cytoplasmic intermediates and factors required for their assembly pathways III) Uncover their nuclear import IV) Understand at the single molecule level their nuclear assembly, dynamics and action at target genes To carry out these aims we propose a combination of multidisciplinary and cutting edge approaches, out of which some of them will be high-risk taking, while others will utilize methods routinely run by the group. The project builds on several complementary expertise and knowledge either already existing in the group or that will be implemented during the project. At the end of the proposed project we will obtain novel results extensively describing the different steps of the regulatory mechanisms that control the assembly and the consequent gene regulatory function of transcription complexes. Thus, we anticipate that the results of our research will have a major impact on the field and will lead to a new paradigm for contemporary metazoan transcription. Keywords of the ERC project: Gene regulation through transcription complex assembly regulation, Keywords that characterize the scientific profile of the potential visiting researcher/s: co-translational assembly pathways, selective ribosome profiling, bioinfo analyses, Index: -3 - Project ID: Project Acronym: Evaluation Panel: LS1 - Molecular and 637733 Pentabrain Structural Biology and Biochemistry Principal Investigator: Dr Hugues Nury Host Institution: Centre National De La Recherche Scientifique Cnrs - FR Structural studies of mammalian Cys-loop receptors In the brain, Cys-loop receptors mediate fast neurotransmission. They function as allosteric signal transducers across the plasma membrane: upon binding of one or more neurotransmitter molecules to an extracellular site, the receptors undergo complex conformational transitions that result in transient opening of an intrinsic ion channel. The Cys-loop family comprises receptors activated by serotonin, acetylcholine, glycine and GABA. Mammalian receptors are also the targets of a legion of psycho-active and therapeutic compounds (including nicotine, benzodiazepines, anti-emetics, general anaesthetics). Our structural knowledge is currently limited to invertebrate homologues. Atomic structures mammalian receptors are therefore acutely missing in order to understand their physiological role in molecular terms, and to be able to develop new drugs targeting them. The project proposes to decipher the operation mechanism, the pharmacology and conformational transitions of mammalian Cys-loop receptors. Starting with a solid body of preliminary results, we will obtain new high- resolution structures, taking advantage of antibody-based crystallization chaperones. We will try and record for the first time a ‘molecular movie’ of the gating conformational transition in cristallo. On the way, we will also investigate the potential of antibody-based modulators of Cys-loop receptors for biomedical applications.The applicant has solved in the past the structures of a bacterial Cys-loop receptor and of the mouse serotonin receptor. The proposed research will take place at the CNRS in Grenoble, France, in a very favourable environment for structural biology. Keywords of the ERC project: membrane protein, crystallography,
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