How the mammalian endoplasmic reticulum handles aggregation-prone β-sheet proteins

Misfolded proteins are prone to engage in aberrant intermolecular interactions that can lead to formation of large aggregate structures. Aggregation causes loss-of-function toxicity because the aggregating protein fails to reach its native fold and function. In addition, Protein aggregates may exert gain-of-toxicity, which is due to the shear presence of Aggregate conformations that sequester important cellular factors and disturb cell morphology. Protein aggregation is associated with a large number of human diseases. The endoplasmic reticulum (ER) is a membrane-bound cellular organelle andthe site of synthesis of one third of the eukaryotic proteome including secretory proteins and proteins destined for the endomembrane system. After co-translational translocation into the ER, nascent proteins are assisted to fold by molecular chaperones and are subject to post-translational modifications. Secretory proteins are retained in the ER lumen until they are correctly folded and are then delivered to the Golgi apparatus for further modifications. If a protein fails to fold properly after repeated folding cycles, it is instead targeted for degradation via the ER-associated degradation pathway (ERAD). The aim of the study presented in this thesis was to determine how the human ER quality control (ERQC) machinery deals with aggregation-prone proteins. This is of great interest because protein aggregates are differentially regulated by distinct cellular environments and many of the proteins that aggregate in diseases are in fact synthesised in the ER. To this end, we utilised de novo designed amyloidogenic β-proteins as generic models for protein aggregation. Due to their lack of evolved biological function, these model proteins allow the exclusive study of gain-of-function toxicity and enable us to dissect the effect of the ER environment on amyloidogenic proteins. We determined that ER-targeted versions of the model β-sheet proteins are significantly less toxic and more soluble than their non-targeted counterparts, which form toxic insoluble aggregates in the cytosol and nucleus. We found that the ER-targeted β-protein ER-β23 is recognised by ERQC machinery and efficiently retained in the ER lumen in a soluble polymeric state. Strikingly, ER-β23 interacted with factors of the ERAD pathway,even though it was not efficiently degraded. Instead, ER-β23 inhibited the degradation of other ERAD substrates by sequestering low-abundant ERAD factors. The presented results demonstrate a marked capacity of the ER to prevent the secretion of potentially toxic aggregation-prone proteins and to limit the formation of insoluble aggregates in the ER lumen. In addition, the data reveal a mechanism by which amyloidogenic proteins may disturb ER proteostasis. Another aim of this study was to analyse the effects of small molecule proteostasis modulators. We found that the anti-dopaminergic drugs fluphenazine and droperidolas well as the epidermal growth factor receptor (EGFR) inhibitors gefitinib and erlotinib improved proteostasis in the presence of Protein aggregates. In case of the former, this effect was most likely achieved via induction of the cytosol stress response. In summary, the work presented in this thesis provides novel insights into how aggregation-prone proteins behave in the environment of the ER and also demonstrates the potential of using small molecule modulators to improve cellular proteostasis in a disease context

How the mammalian endoplasmic reticulum handles aggregation-prone β-sheet proteins

Misfolded proteins are prone to engage in aberrant intermolecular interactions that can lead to formation of large aggregate structures. Aggregation causes loss-of-function toxicity because the aggregating protein fails to reach its native fold and function. In addition, Protein aggregates may exert gain-of-toxicity, which is due to the shear presence of Aggregate conformations that sequester important cellular factors and disturb cell morphology. Protein aggregation is associated with a large number of human diseases. The endoplasmic reticulum (ER) is a membrane-bound cellular organelle andthe site of synthesis of one third of the eukaryotic proteome including secretory proteins and proteins destined for the endomembrane system. After co-translational translocation into the ER, nascent proteins are assisted to fold by molecular chaperones and are subject to post-translational modifications. Secretory proteins are retained in the ER lumen until they are correctly folded and are then delivered to the Golgi apparatus for further modifications. If a protein fails to fold properly after repeated folding cycles, it is instead targeted for degradation via the ER-associated degradation pathway (ERAD). The aim of the study presented in this thesis was to determine how the human ER quality control (ERQC) machinery deals with aggregation-prone proteins. This is of great interest because protein aggregates are differentially regulated by distinct cellular environments and many of the proteins that aggregate in diseases are in fact synthesised in the ER. To this end, we utilised de novo designed amyloidogenic β-proteins as generic models for protein aggregation. Due to their lack of evolved biological function, these model proteins allow the exclusive study of gain-of-function toxicity and enable us to dissect the effect of the ER environment on amyloidogenic proteins. We determined that ER-targeted versions of the model β-sheet proteins are significantly less toxic and more soluble than their non-targeted counterparts, which form toxic insoluble aggregates in the cytosol and nucleus. We found that the ER-targeted β-protein ER-β23 is recognised by ERQC machinery and efficiently retained in the ER lumen in a soluble polymeric state. Strikingly, ER-β23 interacted with factors of the ERAD pathway,even though it was not efficiently degraded. Instead, ER-β23 inhibited the degradation of other ERAD substrates by sequestering low-abundant ERAD factors. The presented results demonstrate a marked capacity of the ER to prevent the secretion of potentially toxic aggregation-prone proteins and to limit the formation of insoluble aggregates in the ER lumen. In addition, the data reveal a mechanism by which amyloidogenic proteins may disturb ER proteostasis. Another aim of this study was to analyse the effects of small molecule proteostasis modulators. We found that the anti-dopaminergic drugs fluphenazine and droperidolas well as the epidermal growth factor receptor (EGFR) inhibitors gefitinib and erlotinib improved proteostasis in the presence of Protein aggregates. In case of the former, this effect was most likely achieved via induction of the cytosol stress response. In summary, the work presented in this thesis provides novel insights into how aggregation-prone proteins behave in the environment of the ER and also demonstrates the potential of using small molecule modulators to improve cellular proteostasis in a disease context

The coming decade of digital brain research: a vision for neuroscience at the intersection of technology and computing

In recent years, brain research has indisputably entered a new epoch, driven by substantial methodological advances and digitally enabled data integration and modelling at multiple scales— from molecules to the whole brain. Major advances are emerging at the intersection of neuroscience with technology and computing. This new science of the brain combines high-quality research, data integration across multiple scales, a new culture of multidisciplinary large-scale collaboration and translation into applications. As pioneered in Europe’s Human Brain Project (HBP), a systematic approach will be essential for meeting the coming decade’s pressing medical and technological challenges. The aims of this paper are to: develop a concept for the coming decade of digital brain research, discuss this new concept with the research community at large, to identify points of convergence, and derive therefrom scientific common goals; provide a scientific framework for the current and future development of EBRAINS, a research infrastructure resulting from the HBP’s work; inform and engage stakeholders, funding organisations and research institutions regarding future digital brain research; identify and address the transformational potential of comprehensive brain models for artificial intelligence, including machine learning and deep learning; outline a collaborative approach that integrates reflection, dialogues and societal engagement on ethical and societal opportunities and challenges as part of future neuroscience research

The coming decade of digital brain research - A vision for neuroscience at the intersection of technology and computing

<p>Brain research has in recent years indisputably entered a new epoch, driven by substantial methodological advances and digitally enabled data integration and modeling at multiple scales – from molecules to the whole system. Major advances are emerging at the intersection of neuroscience with technology and computing. This new science of the brain integrates high-quality basic research, systematic data integration across multiple scales, a new culture of large-scale collaboration and translation into applications. A systematic approach, as pioneered in Europe's Human Brain Project (HBP), will be essential in meeting the pressing medical and technological challenges of the coming decade. The aims of this paper are</p><ul><li>To develop a concept for the coming decade of digital brain research</li><li>To discuss it with the research community at large, with the aim of identifying points of convergence and common goals</li><li>To provide a scientific framework for current and future development of EBRAINS</li><li>To inform and engage stakeholders, funding organizations and research institutions regarding future digital brain research</li><li>To identify and address key ethical and societal issues</li></ul><p>While we do not claim that there is a 'one size fits all' approach to addressing these aspects, we are convinced that discussions around the theme of digital brain research will help drive progress in the broader field of neuroscience.</p><p><strong>As the final version 5 has now been published, comments on this manuscript are now closed. We thank everyone who made a valuable contribution to this paper.</strong></p><p>This manuscript has been developed in a participatory process. The work has been initiated by the Science and Infrastructure Board of the Human Brain Project (HBP), and the entire research community was invited to contribute to shaping the vision by submitting comments. </p><p>All submitted comments were considered and discussed. The final decision on whether edits or additions was made to each version of the manuscript based on an individual comment was made by the Science and Infrastructure Board (SIB) of the Human Brain Project (HBP).</p><p><strong>Supporters of the paper</strong>: Pietro Avanzini, Marc Beyer, Maria Del Vecchio, Jitka Annen, Maurizio Mattia, Steven Laureys, Rosanne Edelenbosch, Rafael Yuste, Jean-Pierre Changeux, Linda Richards, Hye Weon Jessica Kim, Chrysoula Samara, Luis Miguel González de la Garza, Nikoleta Petalidou, Vasudha Kulkarni, Cesar David Rincon, Isabella O'Shea, Munira Tamim Electricwala, Bernd Carsten Stahl, Bahar Hazal Yalcinkaya, Meysam Hashemi, Carola Sales Carbonell, Marcel Carrère, Anthony Randal McIntosh, Hiba Sheheitli, Abolfazl Ziaeemehr, Martin Breyton, Giovanna Ramos Queda, Anirudh NIhalani Vattikonda, Gyorgy Buzsaki, George Ogoh, William Knight, Torbjørn V Ness, Michiel van der Vlag, Marcello Massimini, Thomas Nowontny, Alex Upton, Yaseen Jakhura, Ahmet Nihat Simsek, Michael Hopkins, Addolorata Marasco, Shamim Patel, Jakub Fil, Diego Molinari, Susana Bueno, Lia Domide, Cosimo Lupo, Mu-ming Poo, George Paxinos, Huifang Wang.</p&gt