Objective Supra-molecular protein machineries control diverse cellular processes. Knowing their structural organization is crucial for understanding their function. As classical structural biology techniques are limited in studying such assemblies in their natural cellular environment, there is a critical methodological gap inhibiting a direct link between structure and function. Consequently, the structural intermediates underlying a full activity cycle of a large multi-protein complex have been impossible to visualize. Recent advances in fluorescence microscopy, in particular the development of groundbreaking superresolution microscopy (SRM) methods, can now help bridge this gap. With this interdisciplinary proposal, my group will develop unique and innovative optical, biological and computational imaging technologies to determine the structural organization of multi-protein assemblies in their functional cellular context.We will reach this goal by developing a method to robustly measure the precise 3D arrangements of proteins in supra-molecular assemblies in situ with nanometer isotropic resolution based on supercritical-angle detection and by measuring their absolute stoichiometries with engineered counting standards. We will also develop new data analysis tools to statistically analyze such data, taking into account the functional cellular context measured with correlative superresolution and electron microscopy, multi-color SRM and molecular biology tools. We will apply these new methods to address key questions on endocytosis, a fundamental membrane trafficking process. Our aim is to determine a time-resolved 3D superresolution localization map of the yeast endocytic proteins during the major functional transitions and to integrate these data into a mechanistic model of endocytosis. Importantly, the methods we develop here can be applied to many other large protein-based machines, and thus have the potential to have high impact in other key areas of cell biology. Fields of science natural sciencesphysical sciencesopticsmicroscopysuper resolution microscopynatural sciencesbiological sciencesbiochemistrybiomoleculesproteinsnatural sciencesbiological sciencescell biologynatural sciencesphysical sciencesopticsmicroscopyelectron microscopynatural sciencesbiological sciencesmolecular biologystructural biology Keywords superresolution microscopy endocytosis Programme(s) H2020-EU.1.1. - EXCELLENT SCIENCE - European Research Council (ERC) Main Programme Topic(s) ERC-2016-COG - ERC Consolidator Grant Call for proposal ERC-2016-COG See other projects for this call Funding Scheme ERC-COG - Consolidator Grant Coordinator UNIVERSITAT WIEN Net EU contribution € 143 750,00 Address Universitatsring 1 1010 Wien Austria See on map Region Ostösterreich Wien Wien Activity type Higher or Secondary Education Establishments Links Contact the organisation Opens in new window Website Opens in new window Participation in EU R&I programmes Opens in new window HORIZON collaboration network Opens in new window Other funding € 0,00 Beneficiaries (2) Sort alphabetically Sort by Net EU contribution Expand all Collapse all UNIVERSITAT WIEN Austria Net EU contribution € 143 750,00 Address Universitatsring 1 1010 Wien See on map Region Ostösterreich Wien Wien Activity type Higher or Secondary Education Establishments Links Contact the organisation Opens in new window Website Opens in new window Participation in EU R&I programmes Opens in new window HORIZON collaboration network Opens in new window Other funding € 0,00 EUROPEAN MOLECULAR BIOLOGY LABORATORY Germany Net EU contribution € 1 542 719,38 Address Meyerhofstrasse 1 69117 Heidelberg See on map Region Baden-Württemberg Karlsruhe Heidelberg, Stadtkreis Activity type Research Organisations Links Contact the organisation Opens in new window Website Opens in new window Participation in EU R&I programmes Opens in new window HORIZON collaboration network Opens in new window Other funding € 0,00
