@article{9607, abstract = {While high risk of failure is an inherent part of developing innovative therapies, it can be reduced by adherence to evidence-based rigorous research practices. Numerous analyses conducted to date have clearly identified measures that need to be taken to improve research rigor. Supported through the European Union's Innovative Medicines Initiative, the EQIPD consortium has developed a novel preclinical research quality system that can be applied in both public and private sectors and is free for anyone to use. The EQIPD Quality System was designed to be suited to boost innovation by ensuring the generation of robust and reliable preclinical data while being lean, effective and not becoming a burden that could negatively impact the freedom to explore scientific questions. EQIPD defines research quality as the extent to which research data are fit for their intended use. Fitness, in this context, is defined by the stakeholders, who are the scientists directly involved in the research, but also their funders, sponsors, publishers, research tool manufacturers and collaboration partners such as peers in a multi-site research project. The essence of the EQIPD Quality System is the set of 18 core requirements that can be addressed flexibly, according to user-specific needs and following a user-defined trajectory. The EQIPD Quality System proposes guidance on expectations for quality-related measures, defines criteria for adequate processes (i.e., performance standards) and provides examples of how such measures can be developed and implemented. However, it does not prescribe any pre-determined solutions. EQIPD has also developed tools (for optional use) to support users in implementing the system and assessment services for those research units that successfully implement the quality system and seek formal accreditation. Building upon the feedback from users and continuous improvement, a sustainable EQIPD Quality System will ultimately serve the entire community of scientists conducting non-regulated preclinical research, by helping them generate reliable data that are fit for their intended use.}, author = {Bespalov, Anton and Bernard, René and Gilis, Anja and Gerlach, Björn and Guillén, Javier and Castagné, Vincent and Lefevre, Isabel A. and Ducrey, Fiona and Monk, Lee and Bongiovanni, Sandrine and Altevogt, Bruce and Arroyo-Araujo, María and Bikovski, Lior and De Bruin, Natasja and Castaños-Vélez, Esmeralda and Dityatev, Alexander and Emmerich, Christoph H. and Fares, Raafat and Ferland-Beckham, Chantelle and Froger-Colléaux, Christelle and Gailus-Durner, Valerie and Hölter, Sabine M. and Hofmann, Martine Cj and Kabitzke, Patricia and Kas, Martien Jh and Kurreck, Claudia and Moser, Paul and Pietraszek, Malgorzata and Popik, Piotr and Potschka, Heidrun and Prado Montes De Oca, Ernesto and Restivo, Leonardo and Riedel, Gernot and Ritskes-Hoitinga, Merel and Samardzic, Janko and Schunn, Michael and Stöger, Claudia and Voikar, Vootele and Vollert, Jan and Wever, Kimberley E. and Wuyts, Kathleen and Macleod, Malcolm R. and Dirnagl, Ulrich and Steckler, Thomas}, issn = {2050-084X}, journal = {eLife}, publisher = {eLife Sciences Publications}, title = {{Introduction to the EQIPD quality system}}, doi = {10.7554/eLife.63294}, volume = {10}, year = {2021}, } @article{9244, abstract = {Organ function depends on tissues adopting the correct architecture. However, insights into organ architecture are currently hampered by an absence of standardized quantitative 3D analysis. We aimed to develop a robust technology to visualize, digitalize, and segment the architecture of two tubular systems in 3D: double resin casting micro computed tomography (DUCT). As proof of principle, we applied DUCT to a mouse model for Alagille syndrome (Jag1Ndr/Ndr mice), characterized by intrahepatic bile duct paucity, that can spontaneously generate a biliary system in adulthood. DUCT identified increased central biliary branching and peripheral bile duct tortuosity as two compensatory processes occurring in distinct regions of Jag1Ndr/Ndr liver, leading to full reconstitution of wild-type biliary volume and phenotypic recovery. DUCT is thus a powerful new technology for 3D analysis, which can reveal novel phenotypes and provide a standardized method of defining liver architecture in mouse models.}, author = {Hankeova, Simona and Salplachta, Jakub and Zikmund, Tomas and Kavkova, Michaela and Van Hul, Noémi and Brinek, Adam and Smekalova, Veronika and Laznovsky, Jakub and Dawit, Feven and Jaros, Josef and Bryja, Vítězslav and Lendahl, Urban and Ellis, Ewa and Nemeth, Antal and Fischler, Björn and Hannezo, Edouard B and Kaiser, Jozef and Andersson, Emma Rachel}, issn = {2050-084X}, journal = {eLife}, publisher = {eLife Sciences Publications}, title = {{DUCT reveals architectural mechanisms contributing to bile duct recovery in a mouse model for alagille syndrome}}, doi = {10.7554/eLife.60916}, volume = {10}, year = {2021}, } @article{9437, abstract = {The synaptic connection from medial habenula (MHb) to interpeduncular nucleus (IPN) is critical for emotion-related behaviors and uniquely expresses R-type Ca2+ channels (Cav2.3) and auxiliary GABAB receptor (GBR) subunits, the K+-channel tetramerization domain-containing proteins (KCTDs). Activation of GBRs facilitates or inhibits transmitter release from MHb terminals depending on the IPN subnucleus, but the role of KCTDs is unknown. We therefore examined the localization and function of Cav2.3, GBRs, and KCTDs in this pathway in mice. We show in heterologous cells that KCTD8 and KCTD12b directly bind to Cav2.3 and that KCTD8 potentiates Cav2.3 currents in the absence of GBRs. In the rostral IPN, KCTD8, KCTD12b, and Cav2.3 co-localize at the presynaptic active zone. Genetic deletion indicated a bidirectional modulation of Cav2.3-mediated release by these KCTDs with a compensatory increase of KCTD8 in the active zone in KCTD12b-deficient mice. The interaction of Cav2.3 with KCTDs therefore scales synaptic strength independent of GBR activation.}, author = {Bhandari, Pradeep and Vandael, David H and Fernández-Fernández, Diego and Fritzius, Thorsten and Kleindienst, David and Önal, Hüseyin C and Montanaro-Punzengruber, Jacqueline-Claire and Gassmann, Martin and Jonas, Peter M and Kulik, Akos and Bettler, Bernhard and Shigemoto, Ryuichi and Koppensteiner, Peter}, issn = {2050-084X}, journal = {eLife}, publisher = {eLife Sciences Publications}, title = {{GABAB receptor auxiliary subunits modulate Cav2.3-mediated release from medial habenula terminals}}, doi = {10.7554/ELIFE.68274}, volume = {10}, year = {2021}, } @article{8127, abstract = {Mechanistic modeling in neuroscience aims to explain observed phenomena in terms of underlying causes. However, determining which model parameters agree with complex and stochastic neural data presents a significant challenge. We address this challenge with a machine learning tool which uses deep neural density estimators—trained using model simulations—to carry out Bayesian inference and retrieve the full space of parameters compatible with raw data or selected data features. Our method is scalable in parameters and data features and can rapidly analyze new data after initial training. We demonstrate the power and flexibility of our approach on receptive fields, ion channels, and Hodgkin–Huxley models. We also characterize the space of circuit configurations giving rise to rhythmic activity in the crustacean stomatogastric ganglion, and use these results to derive hypotheses for underlying compensation mechanisms. Our approach will help close the gap between data-driven and theory-driven models of neural dynamics.}, author = {Gonçalves, Pedro J. and Lueckmann, Jan-Matthis and Deistler, Michael and Nonnenmacher, Marcel and Öcal, Kaan and Bassetto, Giacomo and Chintaluri, Chaitanya and Podlaski, William F. and Haddad, Sara A. and Vogels, Tim P and Greenberg, David S. and Macke, Jakob H.}, issn = {2050-084X}, journal = {eLife}, publisher = {eLife Sciences Publications}, title = {{Training deep neural density estimators to identify mechanistic models of neural dynamics}}, doi = {10.7554/eLife.56261}, volume = {9}, year = {2020}, } @article{7490, abstract = {In plants, clathrin mediated endocytosis (CME) represents the major route for cargo internalisation from the cell surface. It has been assumed to operate in an evolutionary conserved manner as in yeast and animals. Here we report characterisation of ultrastructure, dynamics and mechanisms of plant CME as allowed by our advancement in electron microscopy and quantitative live imaging techniques. Arabidopsis CME appears to follow the constant curvature model and the bona fide CME population generates vesicles of a predominantly hexagonal-basket type; larger and with faster kinetics than in other models. Contrary to the existing paradigm, actin is dispensable for CME events at the plasma membrane but plays a unique role in collecting endocytic vesicles, sorting of internalised cargos and directional endosome movement that itself actively promote CME events. Internalized vesicles display a strongly delayed and sequential uncoating. These unique features highlight the independent evolution of the plant CME mechanism during the autonomous rise of multicellularity in eukaryotes.}, author = {Narasimhan, Madhumitha and Johnson, Alexander J and Prizak, Roshan and Kaufmann, Walter and Tan, Shutang and Casillas Perez, Barbara E and Friml, Jiří}, issn = {2050-084X}, journal = {eLife}, publisher = {eLife Sciences Publications}, title = {{Evolutionarily unique mechanistic framework of clathrin-mediated endocytosis in plants}}, doi = {10.7554/eLife.52067}, volume = {9}, year = {2020}, } @article{7878, abstract = {Type 1 metabotropic glutamate receptors (mGluR1s) are key elements in neuronal signaling. While their function is well documented in slices, requirements for their activation in vivo are poorly understood. We examine this question in adult mice in vivo using 2-photon imaging of cerebellar molecular layer interneurons (MLIs) expressing GCaMP. In anesthetized mice, parallel fiber activation evokes beam-like Cai rises in postsynaptic MLIs which depend on co-activation of mGluR1s and ionotropic glutamate receptors (iGluRs). In awake mice, blocking mGluR1 decreases Cai rises associated with locomotion. In vitro studies and freeze-fracture electron microscopy show that the iGluR-mGluR1 interaction is synergistic and favored by close association of the two classes of receptors. Altogether our results suggest that mGluR1s, acting in synergy with iGluRs, potently contribute to processing cerebellar neuronal signaling under physiological conditions.}, author = {Bao, Jin and Graupner, Michael and Astorga, Guadalupe and Collin, Thibault and Jalil, Abdelali and Indriati, Dwi Wahyu and Bradley, Jonathan and Shigemoto, Ryuichi and Llano, Isabel}, issn = {2050-084X}, journal = {eLife}, publisher = {eLife Sciences Publications}, title = {{Synergism of type 1 metabotropic and ionotropic glutamate receptors in cerebellar molecular layer interneurons in vivo}}, doi = {10.7554/eLife.56839}, volume = {9}, year = {2020}, } @article{7909, abstract = {Cell migration entails networks and bundles of actin filaments termed lamellipodia and microspikes or filopodia, respectively, as well as focal adhesions, all of which recruit Ena/VASP family members hitherto thought to antagonize efficient cell motility. However, we find these proteins to act as positive regulators of migration in different murine cell lines. CRISPR/Cas9-mediated loss of Ena/VASP proteins reduced lamellipodial actin assembly and perturbed lamellipodial architecture, as evidenced by changed network geometry as well as reduction of filament length and number that was accompanied by abnormal Arp2/3 complex and heterodimeric capping protein accumulation. Loss of Ena/VASP function also abolished the formation of microspikes normally embedded in lamellipodia, but not of filopodia capable of emanating without lamellipodia. Ena/VASP-deficiency also impaired integrin-mediated adhesion accompanied by reduced traction forces exerted through these structures. Our data thus uncover novel Ena/VASP functions of these actin polymerases that are fully consistent with their promotion of cell migration.}, author = {Damiano-Guercio, Julia and Kurzawa, Laëtitia and Müller, Jan and Dimchev, Georgi A and Schaks, Matthias and Nemethova, Maria and Pokrant, Thomas and Brühmann, Stefan and Linkner, Joern and Blanchoin, Laurent and Sixt, Michael K and Rottner, Klemens and Faix, Jan}, issn = {2050-084X}, journal = {eLife}, publisher = {eLife Sciences Publications}, title = {{Loss of Ena/VASP interferes with lamellipodium architecture, motility and integrin-dependent adhesion}}, doi = {10.7554/eLife.55351}, volume = {9}, year = {2020}, } @article{7466, abstract = {Unpaired ligands are secreted signals that act via a GP130-like receptor, domeless, to activate JAK/STAT signalling in Drosophila. Like many mammalian cytokines, unpaireds can be activated by infection and other stresses and can promote insulin resistance in target tissues. However, the importance of this effect in non-inflammatory physiology is unknown. Here, we identify a requirement for unpaired-JAK signalling as a metabolic regulator in healthy adult Drosophila muscle. Adult muscles show basal JAK-STAT signalling activity in the absence of any immune challenge. Plasmatocytes (Drosophila macrophages) are an important source of this tonic signal. Loss of the dome receptor on adult muscles significantly reduces lifespan and causes local and systemic metabolic pathology. These pathologies result from hyperactivation of AKT and consequent deregulation of metabolism. Thus, we identify a cytokine signal that must be received in muscle to control AKT activity and metabolic homeostasis.}, author = {Kierdorf, Katrin and Hersperger, Fabian and Sharrock, Jessica and Vincent, Crystal M. and Ustaoglu, Pinar and Dou, Jiawen and György, Attila and Groß, Olaf and Siekhaus, Daria E and Dionne, Marc S.}, issn = {2050-084X}, journal = {eLife}, publisher = {eLife Sciences Publications}, title = {{Muscle function and homeostasis require cytokine inhibition of AKT activity in Drosophila}}, doi = {10.7554/eLife.51595}, volume = {9}, year = {2020}, } @article{8740, abstract = {In vitro work revealed that excitatory synaptic inputs to hippocampal inhibitory interneurons could undergo Hebbian, associative, or non-associative plasticity. Both behavioral and learning-dependent reorganization of these connections has also been demonstrated by measuring spike transmission probabilities in pyramidal cell-interneuron spike cross-correlations that indicate monosynaptic connections. Here we investigated the activity-dependent modification of these connections during exploratory behavior in rats by optogenetically inhibiting pyramidal cell and interneuron subpopulations. Light application and associated firing alteration of pyramidal and interneuron populations led to lasting changes in pyramidal-interneuron connection weights as indicated by spike transmission changes. Spike transmission alterations were predicted by the light-mediated changes in the number of pre- and postsynaptic spike pairing events and by firing rate changes of interneurons but not pyramidal cells. This work demonstrates the presence of activity-dependent associative and non-associative reorganization of pyramidal-interneuron connections triggered by the optogenetic modification of the firing rate and spike synchrony of cells.}, author = {Gridchyn, Igor and Schönenberger, Philipp and O'Neill, Joseph and Csicsvari, Jozsef L}, issn = {2050-084X}, journal = {eLife}, publisher = {eLife Sciences Publications}, title = {{Optogenetic inhibition-mediated activity-dependent modification of CA1 pyramidal-interneuron connections during behavior}}, doi = {10.7554/eLife.61106}, volume = {9}, year = {2020}, } @article{8284, abstract = {Multiple resistance and pH adaptation (Mrp) antiporters are multi-subunit Na+ (or K+)/H+ exchangers representing an ancestor of many essential redox-driven proton pumps, such as respiratory complex I. The mechanism of coupling between ion or electron transfer and proton translocation in this large protein family is unknown. Here, we present the structure of the Mrp complex from Anoxybacillus flavithermus solved by cryo-EM at 3.0 Å resolution. It is a dimer of seven-subunit protomers with 50 trans-membrane helices each. Surface charge distribution within each monomer is remarkably asymmetric, revealing probable proton and sodium translocation pathways. On the basis of the structure we propose a mechanism where the coupling between sodium and proton translocation is facilitated by a series of electrostatic interactions between a cation and key charged residues. This mechanism is likely to be applicable to the entire family of redox proton pumps, where electron transfer to substrates replaces cation movements.}, author = {Steiner, Julia and Sazanov, Leonid A}, issn = {2050-084X}, journal = {eLife}, publisher = {eLife Sciences Publications}, title = {{Structure and mechanism of the Mrp complex, an ancient cation/proton antiporter}}, doi = {10.7554/eLife.59407}, volume = {9}, year = {2020}, } @article{6230, abstract = {Great care is needed when interpreting claims about the genetic basis of human variation based on data from genome-wide association studies.}, author = {Barton, Nicholas H and Hermisson, Joachim and Nordborg, Magnus}, issn = {2050-084X}, journal = {eLife}, publisher = {eLife Sciences Publications}, title = {{Why structure matters}}, doi = {10.7554/eLife.45380}, volume = {8}, year = {2019}, } @article{6868, abstract = {Hyperpolarization-activated cyclic-nucleotide-gated (HCN) channels control electrical rhythmicity and excitability in the heart and brain, but the function of HCN channels at the subcellular level in axons remains poorly understood. Here, we show that the action potential conduction velocity in both myelinated and unmyelinated central axons can be bidirectionally modulated by a HCN channel blocker, cyclic adenosine monophosphate (cAMP), and neuromodulators. Recordings from mouse cerebellar mossy fiber boutons show that HCN channels ensure reliable high-frequency firing and are strongly modulated by cAMP (EC50 40 mM; estimated endogenous cAMP concentration 13 mM). In addition, immunogold-electron microscopy revealed HCN2 as the dominating subunit in cerebellar mossy fibers. Computational modeling indicated that HCN2 channels control conduction velocity primarily by altering the resting membrane potential and are associated with significant metabolic costs. These results suggest that the cAMP-HCN pathway provides neuromodulators with an opportunity to finely tune energy consumption and temporal delays across axons in the brain.}, author = {Byczkowicz, Niklas and Eshra, Abdelmoneim and Montanaro-Punzengruber, Jacqueline-Claire and Trevisiol, Andrea and Hirrlinger, Johannes and Kole, Maarten Hp and Shigemoto, Ryuichi and Hallermann, Stefan}, issn = {2050-084X}, journal = {eLife}, publisher = {eLife Sciences Publications}, title = {{HCN channel-mediated neuromodulation can control action potential velocity and fidelity in central axons}}, doi = {10.7554/eLife.42766}, volume = {8}, year = {2019}, } @article{7340, abstract = {Coupling of endoplasmic reticulum stress to dimerisation‑dependent activation of the UPR transducer IRE1 is incompletely understood. Whilst the luminal co-chaperone ERdj4 promotes a complex between the Hsp70 BiP and IRE1's stress-sensing luminal domain (IRE1LD) that favours the latter's monomeric inactive state and loss of ERdj4 de-represses IRE1, evidence linking these cellular and in vitro observations is presently lacking. We report that enforced loading of endogenous BiP onto endogenous IRE1α repressed UPR signalling in CHO cells and deletions in the IRE1α locus that de-repressed the UPR in cells, encode flexible regions of IRE1LD that mediated BiP‑induced monomerisation in vitro. Changes in the hydrogen exchange mass spectrometry profile of IRE1LD induced by ERdj4 and BiP confirmed monomerisation and were consistent with active destabilisation of the IRE1LD dimer. Together, these observations support a competition model whereby waning ER stress passively partitions ERdj4 and BiP to IRE1LD to initiate active repression of UPR signalling.}, author = {Amin-Wetzel, Niko Paresh and Neidhardt, Lisa and Yan, Yahui and Mayer, Matthias P. and Ron, David}, issn = {2050-084X}, journal = {eLife}, publisher = {eLife Sciences Publications}, title = {{Unstructured regions in IRE1α specify BiP-mediated destabilisation of the luminal domain dimer and repression of the UPR}}, doi = {10.7554/eLife.50793}, volume = {8}, year = {2019}, } @article{7202, abstract = {The cerebral cortex contains multiple areas with distinctive cytoarchitectonical patterns, but the cellular mechanisms underlying the emergence of this diversity remain unclear. Here, we have investigated the neuronal output of individual progenitor cells in the developing mouse neocortex using a combination of methods that together circumvent the biases and limitations of individual approaches. Our experimental results indicate that progenitor cells generate pyramidal cell lineages with a wide range of sizes and laminar configurations. Mathematical modelling indicates that these outcomes are compatible with a stochastic model of cortical neurogenesis in which progenitor cells undergo a series of probabilistic decisions that lead to the specification of very heterogeneous progenies. Our findings support a mechanism for cortical neurogenesis whose flexibility would make it capable to generate the diverse cytoarchitectures that characterize distinct neocortical areas.}, author = {Llorca, Alfredo and Ciceri, Gabriele and Beattie, Robert J and Wong, Fong Kuan and Diana, Giovanni and Serafeimidou-Pouliou, Eleni and Fernández-Otero, Marian and Streicher, Carmen and Arnold, Sebastian J. and Meyer, Martin and Hippenmeyer, Simon and Maravall, Miguel and Marín, Oscar}, issn = {2050-084X}, journal = {eLife}, publisher = {eLife Sciences Publications}, title = {{A stochastic framework of neurogenesis underlies the assembly of neocortical cytoarchitecture}}, doi = {10.7554/eLife.51381}, volume = {8}, year = {2019}, } @article{9445, abstract = {Cytosine methylation regulates essential genome functions across eukaryotes, but the fundamental question of whether nucleosomal or naked DNA is the preferred substrate of plant and animal methyltransferases remains unresolved. Here, we show that genetic inactivation of a single DDM1/Lsh family nucleosome remodeler biases methylation toward inter-nucleosomal linker DNA in Arabidopsis thaliana and mouse. We find that DDM1 enables methylation of DNA bound to the nucleosome, suggesting that nucleosome-free DNA is the preferred substrate of eukaryotic methyltransferases in vivo. Furthermore, we show that simultaneous mutation of DDM1 and linker histone H1 in Arabidopsis reproduces the strong linker-specific methylation patterns of species that diverged from flowering plants and animals over a billion years ago. Our results indicate that in the absence of remodeling, nucleosomes are strong barriers to DNA methyltransferases. Linker-specific methylation can evolve simply by breaking the connection between nucleosome remodeling and DNA methylation.}, author = {Lyons, David B and Zilberman, Daniel}, issn = {2050-084X}, journal = {eLife}, publisher = {eLife Sciences Publications}, title = {{DDM1 and Lsh remodelers allow methylation of DNA wrapped in nucleosomes}}, doi = {10.7554/elife.30674}, volume = {6}, year = {2017}, }