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

Forward-central two-particle correlations in p-Pb collisions at root s(NN)=5.02 TeV

Two-particle angular correlations between trigger particles in the forward pseudorapidity range (2.5 2GeV/c. (C) 2015 CERN for the benefit of the ALICE Collaboration. Published by Elsevier B. V.Peer reviewe

Event-shape engineering for inclusive spectra and elliptic flow in Pb-Pb collisions at root(NN)-N-S=2.76 TeV

Peer reviewe

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

Elliptic flow of muons from heavy-flavour hadron decays at forward rapidity in Pb-Pb collisions at root s(NN)=2.76TeV

The elliptic flow, v(2), of muons from heavy-flavour hadron decays at forward rapidity (2.5 <y <4) is measured in Pb-Pb collisions at root s(NN)= 2.76TeVwith the ALICE detector at the LHC. The scalar product, two- and four-particle Q cumulants and Lee-Yang zeros methods are used. The dependence of the v(2) of muons from heavy-flavour hadron decays on the collision centrality, in the range 0-40%, and on transverse momentum, p(T), is studied in the interval 3 <p(T)<10 GeV/c. A positive v(2) is observed with the scalar product and two-particle Q cumulants in semi-central collisions (10-20% and 20-40% centrality classes) for the p(T) interval from 3 to about 5GeV/c with a significance larger than 3 sigma, based on the combination of statistical and systematic uncertainties. The v(2) magnitude tends to decrease towards more central collisions and with increasing pT. It becomes compatible with zero in the interval 6 <p(T)<10 GeV/c. The results are compared to models describing the interaction of heavy quarks and open heavy-flavour hadrons with the high-density medium formed in high-energy heavy-ion collisions. (C) 2015 CERN for the benefit of the ALICE Collaboration. Published by Elsevier B.V.Peer reviewe