TY - JOUR AB - Glutamine synthetases (GS) are central enzymes essential for the nitrogen metabolism across all domains of life. Consequently, they have been extensively studied for more than half a century. Based on the ATP-dependent ammonium assimilation generating glutamine, GS expression and activity are strictly regulated in all organisms. In the methanogenic archaeon Methanosarcina mazei, it has been shown that the metabolite 2-oxoglutarate (2-OG) directly induces the GS activity. Besides, modulation of the activity by interaction with small proteins (GlnK1 and sP26) has been reported. Here, we show that the strong activation of M. mazei GS (GlnA1) by 2-OG is based on the 2-OG dependent dodecamer assembly of GlnA1 by using mass photometry (MP) and single particle cryo-electron microscopy (cryo-EM) analysis of purified strep-tagged GlnA1. The dodecamer assembly from dimers occurred without any detectable intermediate oligomeric state and was not affected in the presence of GlnK1. The 2.39 Å cryo-EM structure of the dodecameric complex in the presence of 12.5 mM 2-OG demonstrated that 2-OG is binding between two monomers. Thereby, 2-OG appears to induce the dodecameric assembly in a cooperative way. Furthermore, the active site is primed by an allosteric interaction cascade caused by 2-OG-binding towards an adaption of an open active state conformation. In the presence of additional glutamine, strong feedback inhibition of GS activity was observed. Since glutamine dependent disassembly of the dodecamer was excluded by MP, feedback inhibition most likely relies on the binding of glutamine to the catalytic site. Based on our findings, we propose that under nitrogen limitation the induction of M. mazei GS into a catalytically active dodecamer is not affected by GlnK1 and crucially depends on the presence of 2-OG. AU - Herdering, E.* AU - Reif-Trauttmansdorff, T.* AU - Kumar, A.* AU - Habenicht, T.* AU - Hochberg, G.K.A.* AU - Bohn, S. AU - Schüller, J.* AU - Schmitz, R.A.* C1 - 73924 C2 - 57243 CY - 95 Regent Street, Cambridge, England TI - 2-oxoglutarate triggers assembly of active dodecameric Methanosarcina mazei glutamine synthetase. JO - eLife VL - 13 PB - Elife Sciences Publ Ltd PY - 2025 SN - 2050-084X ER - TY - JOUR AB - The identification of cell types remains a major challenge. Even after a decade of single-cellRNA sequencing (scRNA-seq), reasonable cell type annotations almost always include manualnon-automated steps. The identification of orthologous cell types across species complicatesmatters even more, but at the same time strengthens the confidence in the assignment. Here,we generate and analyze a dataset consisting of embryoid bodies (EBs) derived from inducedpluripotent stem cells (iPSCs) of four primate species: humans, orangutans, cynomolgus, andrhesus macaques. This kind of data includes a continuum of developmental cell types,multiple batch effects (i.e. species and individuals) and uneven cell type compositions andhence poses many challenges. We developed a semi-automated computational pipelinecombining classification and marker based cluster annotation to identify orthologous celltypes across primates. This approach enabled the investigation of cross-species conservationof gene expression. Consistent with previous studies, our data confirm that broadly expressedgenes are more conserved than cell type-specific genes, raising the question how conserved -inherently cell type-specific - marker genes are. Our analyses reveal that human markergenes are less effective in macaques and vice versa, highlighting the limited transferability ofmarkers across species. Overall, our study advances the identification of orthologous celltypes across species, provides a well-curated cell type reference for future in vitro studies andinforms the transferability of marker genes across species. AU - Janssen, P.* AU - Jocher, J.* AU - Vieth, B.* AU - Edenhofer, F.C.* AU - Dietl, T. AU - Térmeg, A.* AU - Spurk, P.* AU - Geuder, J.* AU - Enard, W.* AU - Hellmann, I.* C1 - 74734 C2 - 57586 TI - Identification and comparison of orthologous cell types from primate embryoid bodies shows limits of marker gene transferability. JO - eLife PY - 2025 SN - 2050-084X ER - TY - JOUR AB - Accumulation of extracellular matrix (ECM) in liver fibrosis is associated with changes in protein abundance and composition depending upon etiology of the underlying liver disease. Current efforts to unravel etiology-specific mechanisms and pharmacological targets rely on several models of experimental fibrosis. Here, we characterize and compare dynamics of hepatic proteome remodeling during fibrosis development and spontaneous healing in experimental mouse models of hepatotoxic (carbon tetrachloride [CCl4] intoxication) and cholestatic (3,5-diethoxycarbonyl-1,4-dihydrocollidine [DDC] feeding) injury. Using detergent-based tissue extraction and mass spectrometry, we identified compartment-specific changes in the liver proteome with detailed attention to ECM composition and changes in protein solubility. Our analysis revealed distinct time-resolved CCl4 and DDC signatures, with identified signaling pathways suggesting limited healing and a potential for carcinogenesis associated with cholestasis. Correlation of protein abundance profiles with fibrous deposits revealed extracellular chaperone clusterin with implicated role in fibrosis resolution. Dynamics of clusterin expression was validated in the context of human liver fibrosis. Atomic force microscopy of fibrotic livers complemented proteomics with profiles of disease-associated changes in local liver tissue mechanics. This study determined compartment-specific proteomic landscapes of liver fibrosis and delineated etiology-specific ECM components, providing thus a foundation for future antifibrotic therapies. AU - Jirouskova, M.* AU - Harant, K.* AU - Cejnar, P.* AU - Ojha, S.* AU - Korelova, K.* AU - Sarnova, L.* AU - Sticova, E.* AU - Mayr, C.H. AU - Schiller, H.B. AU - Gregor, M.* C1 - 74004 C2 - 57297 CY - 95 Regent Street, Cambridge, England TI - Dynamics of compartment-specific proteomic landscapes of hepatotoxic and cholestatic models of liver fibrosis. JO - eLife VL - 13 PB - Elife Sciences Publ Ltd PY - 2025 SN - 2050-084X ER - TY - JOUR AB - Bacterial cell division relies on the Z ring, a cytoskeletal structure that acts as a scaffold for the assembly of the divisome. To date, the detailed mechanisms underlying the assembly and stabilization of the Z ring remain elusive. This study highlights the role of the FtsZ-associated protein (Zap) ZapD in the assembly and stabilization of Z-ring-like structures via filament crosslinking. Using cryo-electron tomography and biochemical analysis, we show that, at equimolar concentrations of ZapD and FtsZ, ZapD induces the formation of toroidal structures composed of short, curved FtsZ filaments that are crosslinked vertically, but also laterally and diagonally. At higher concentrations of ZapD, regularly spaced ZapD dimers crosslink FtsZ filaments from above, resulting in the formation of straight bundles. Despite the simplicity of this reconstituted system, these findings provide valuable insights into the structural organization and stabilization of the Z ring by Zap proteins in bacterial cells, revealing the key role of optimal crosslinking density and geometry in enabling filament curvature and ring formation. AU - Merino‐Salomón, A.* AU - Schneider, J.* AU - Babl, L.* AU - Krohn, J.F.* AU - Sobrinos‐Sanguino, M.* AU - Schäfer, T.* AU - Luque-Ortega, J.R.* AU - Alfonso, C.* AU - Jimenez, M.* AU - Jasnin, M. AU - Schwille, P.* AU - Rivas, G.* C1 - 74736 C2 - 57587 TI - Crosslinking by ZapD drives the assembly of short FtsZ filaments into toroidal structures in solution. JO - eLife PY - 2025 SN - 2050-084X ER - TY - JOUR AB - Thylakoid membranes coordinate the light reactions of photosynthesis across multiple scales, coupling the architecture of an elaborate membrane network to the spatial organization of individual protein complexes embedded within this network. Previously, we used in situ cryo- electron tomography (cryo-ET) to reveal the native thylakoid architecture of the green alga Chlamydomonas reinhardtii [1] and then map the molecular organization of these thylakoids with single-molecule precision [2]. However, it remains to be shown how generalizable this green algal blueprint is to the thylakoids of vascular plants, which possess distinct membrane architecture subdivided into grana stacks interconnected by non-stacked stromal lamellae. Here, we continue our cryo-ET investigation to reveal the molecular architecture of thylakoids within intact chloroplasts isolated from spinach (Spinacia oleracea). We visualize the fine ultrastructural details of grana membranes, as well as interactions between thylakoids and plastoglobules. We apply and further develop AI-based computational approaches for automated membrane segmentation and membrane protein picking [3], enabling us to quantify the organization of photosynthetic complexes within the plane of the thylakoid membrane and across adjacent stacked membranes. Our analysis reveals that, despite different 3D architecture, the molecular organization of thylakoid membranes in vascular plants and green algae is strikingly similar. In contrast to isolated plant thylakoids, where semi- crystalline arrays of photosystem II (PSII) appear to hold some membranes together, we find in intact chloroplasts that PSII is non-crystalline and has uniform concentration both within the membrane plane and across stacked grana membranes. Similar to C. reinhardtii, we observe strict lateral heterogeneity of PSII and PSI at the boundary between appressed and non-appressed thylakoid domains, with no evidence for a distinct grana margin region where these complexes have been proposed to intermix. Based on these measurements, we support a simple two-domain model for the molecular organization of thylakoid membranes in both green algae and plants. AU - Wietrzynski, W.* AU - Lamm, L. AU - Wood, W.H.* AU - Loukeri, M.* AU - Malone, L.A.* AU - Peng, T. AU - Johnson, M.P.* AU - Engel, B.D.* C1 - 74731 C2 - 57585 TI - Molecular architecture of thylakoid membranes within intact spinach chloroplasts. JO - eLife VL - 14 PY - 2025 SN - 2050-084X ER - TY - JOUR AB - Alterations in the function of K+ channels such as the voltage- and Ca2+-activated K+ channel of large conductance (BKCa) reportedly promote breast cancer (BC) development and progression. Underlying molecular mechanisms remain, however, elusive. Here, we provide electrophysiological evidence for a BKCa splice variant localized to the inner mitochondrial membrane of murine and human BC cells (mitoBKCa). Through a combination of genetic knockdown and knockout along with a cell permeable BKCa channel blocker, we show that mitoBKCa modulates overall cellular and mitochondrial energy production, and mediates the metabolic rewiring referred to as the 'Warburg effect', thereby promoting BC cell proliferation in the presence and absence of oxygen. Additionally, we detect mitoBKCa and BKCa transcripts in low or high abundance, respectively, in clinical BC specimens. Together, our results emphasize, that targeting mitoBKCa could represent a treatment strategy for selected BC patients in future. AU - Bischof, H.* AU - Maier, S.* AU - Koprowski, P.* AU - Kulawiak, B.* AU - Burgstaller, S.* AU - Jasińska, J.* AU - Serafimov, K.* AU - Zochowska, M.* AU - Gross, D.* AU - Schroth, W.* AU - Matt, L.* AU - Juarez Lopez, D.A.* AU - Zhang, Y.* AU - Bonzheim, I.* AU - Büttner, F.A.* AU - Fend, F.* AU - Schwab, M.* AU - Birkenfeld, A.L. AU - Malli, R.* AU - Lämmerhofer, M.* AU - Bednarczyk, P.* AU - Szewczyk, A.* AU - Lukowski, R.* C1 - 70772 C2 - 55887 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - mitoBKCa is functionally expressed in murine and human breast cancer cells and potentially contributes to metabolic reprogramming. JO - eLife VL - 12 PB - Elife Sciences Publ Ltd PY - 2024 SN - 2050-084X ER - TY - JOUR AB - Parkinson's disease (PD) is characterized by the progressive loss of dopaminergic neurons in the substantia nigra of the midbrain. Familial cases of PD are often caused by mutations of PTEN-induced kinase 1 (PINK1) and the ubiquitin ligase Parkin, both pivotal in maintaining mitochondrial quality control. CISD1, a homodimeric mitochondrial iron-sulfur-binding protein, is a major target of Parkin-mediated ubiquitination. We here discovered a heightened propensity of CISD1 to form dimers in Pink1 mutant flies and in dopaminergic neurons from PINK1 mutation patients. The dimer consists of two monomers that are covalently linked by a disulfide bridge. In this conformation CISD1 cannot coordinate the iron-sulfur cofactor. Overexpressing Cisd, the Drosophila orthologue of CISD1, and a mutant Cisd incapable of binding the iron-sulfur cluster in Drosophila reduced climbing ability and lifespan. This was more pronounced with mutant Cisd and aggravated in Pink1 mutant flies. Complete loss of Cisd, in contrast, rescued all detrimental effects of Pink1 mutation on climbing ability, wing posture, dopamine levels, lifespan, and mitochondrial ultrastructure. Our results suggest that Cisd, probably iron-depleted Cisd, operates downstream of Pink1 shedding light on PD pathophysiology and implicating CISD1 as a potential therapeutic target. AU - Bitar, S.* AU - Baumann, T.* AU - Weber, C.* AU - Abusaada, M.* AU - Rojas-Charry, L.* AU - Ziegler, P.* AU - Schettgen, T.* AU - Randerath, I.E.* AU - Venkataramani, V.* AU - Michalke, B. AU - Hanschmann, E.M.* AU - Arena, G.* AU - Krueger, R.* AU - Zhang, L.* AU - Methner, A.* C1 - 71494 C2 - 56215 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - Iron-sulfur cluster loss in mitochondrial CISD1 mediates PINK1 loss-of-function phenotypes. JO - eLife VL - 13 PB - Elife Sciences Publ Ltd PY - 2024 SN - 2050-084X ER - TY - JOUR AB - The unlimited expansion of human progenitor cells in vitro could unlock many prospects for regenerative medicine. However, it remains an important challenge as it requires the decoupling of the mechanisms supporting progenitor self-renewal and expansion from those mechanisms promoting their differentiation. This study focuses on the expansion of human pluripotent stem (hPS) cell-derived pancreatic progenitors (PP) to advance novel therapies for diabetes. We obtained mechanistic insights into PP expansion requirements and identified conditions for the robust and unlimited expansion of hPS cell-derived PP cells under GMP-compliant conditions through a hypothesis-driven iterative approach. We show that the combined stimulation of specific mitogenic pathways, suppression of retinoic acid signaling, and inhibition of selected branches of the TGFβ and Wnt signaling pathways are necessary for the effective decoupling of PP proliferation from differentiation. This enabled the reproducible, 2000-fold, over 10 passages and 40-45 d, expansion of PDX1+/SOX9+/NKX6-1+ PP cells. Transcriptome analyses confirmed the stabilization of PP identity and the effective suppression of differentiation. Using these conditions, PDX1+/SOX9+/NKX6-1+ PP cells, derived from different, both XY and XX, hPS cell lines, were enriched to nearly 90% homogeneity and expanded with very similar kinetics and efficiency. Furthermore, non-expanded and expanded PP cells, from different hPS cell lines, were differentiated in microwells into homogeneous islet-like clusters (SC-islets) with very similar efficiency. These clusters contained abundant β-cells of comparable functionality as assessed by glucose-stimulated insulin secretion assays. These findings established the signaling requirements to decouple PP proliferation from differentiation and allowed the consistent expansion of hPS cell-derived PP cells. They will enable the establishment of large banks of GMP-produced PP cells derived from diverse hPS cell lines. This approach will streamline SC-islet production for further development of the differentiation process, diabetes research, personalized medicine, and cell therapies. AU - Jarc, L. AU - Bandral, M. AU - Zanfrini, E. AU - Lesche, M.* AU - Kufrin, V. AU - Sendra, R. AU - Pezzolla, D.* AU - Giannios, I. AU - Khattak, S.* AU - Neumann, K.* AU - Ludwig, B. AU - Gavalas, A. C1 - 68993 C2 - 55176 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - Regulation of multiple signaling pathways promotes the consistent expansion of human pancreatic progenitors in defined conditions. JO - eLife VL - 12 PB - Elife Sciences Publ Ltd PY - 2024 SN - 2050-084X ER - TY - JOUR AB - BACKGROUND: The development of obesity-associated comorbidities such as type 2 diabetes (T2D) and hepatic steatosis has been linked to selected microRNAs in individual studies; however, an unbiased genome-wide approach to map T2D induced changes in the miRNAs landscape in human liver samples, and a subsequent robust identification and validation of target genes are still missing. METHODS: Liver biopsies from age- and gender-matched obese individuals with (n=20) or without (n=20) T2D were used for microRNA microarray analysis. The candidate microRNA and target genes were validated in 85 human liver samples, and subsequently mechanistically characterized in hepatic cells as well as by dietary interventions and hepatic overexpression in mice. RESULTS: Here, we present the human hepatic microRNA transcriptome of type 2 diabetes in liver biopsies and use a novel seed prediction tool to robustly identify microRNA target genes, which were then validated in a unique cohort of 85 human livers. Subsequent mouse studies identified a distinct signature of T2D-associated miRNAs, partly conserved in both species. Of those, human-murine miR-182-5 p was the most associated with whole-body glucose homeostasis and hepatic lipid metabolism. Its target gene LRP6 was consistently lower expressed in livers of obese T2D humans and mice as well as under conditions of miR-182-5 p overexpression. Weight loss in obese mice decreased hepatic miR-182-5 p and restored Lrp6 expression and other miR-182-5 p target genes. Hepatic overexpression of miR-182-5 p in mice rapidly decreased LRP6 protein levels and increased liver triglycerides and fasting insulin under obesogenic conditions after only seven days. CONCLUSIONS: By mapping the hepatic miRNA-transcriptome of type 2 diabetic obese subjects, validating conserved miRNAs in diet-induced mice, and establishing a novel miRNA prediction tool, we provide a robust and unique resource that will pave the way for future studies in the field. As proof of concept, we revealed that the repression of LRP6 by miR-182-5 p, which promotes lipogenesis and impairs glucose homeostasis, provides a novel mechanistic link between T2D and non-alcoholic fatty liver disease, and demonstrate in vivo that miR-182-5 p can serve as a future drug target for the treatment of obesity-driven hepatic steatosis. FUNDING: This work was supported by research funding from the Deutsche Forschungsgemeinschaft (KI 1887/2-1, KI 1887/2-2, KI 1887/3-1 and CRC-TR296), the European Research Council (ERC, CoG Yoyo LepReSens no. 101002247; PTP), the Helmholtz Association (Initiative and Networking Fund International Helmholtz Research School for Diabetes; MB) and the German Center for Diabetes Research (DZD Next Grant 82DZD09D1G). AU - Krause, C.* AU - Britsemmer, J.H.* AU - Bernecker, M. AU - Molenaar, A. AU - Taege, N.* AU - Lopez-Alcantara, N.* AU - Geißler, C.* AU - Kaehler, M.* AU - Iben, K.* AU - Judycka, A.* AU - Wagner, J.* AU - Wolter, S.* AU - Mann, O.* AU - Pfluger, P.T. AU - Cascorbi, I.* AU - Lehnert, H.* AU - Stemmer, K.* AU - Schriever, S.C. AU - Kirchner, H.* C1 - 71251 C2 - 56024 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - Liver microRNA transcriptome reveals miR-182 as link between type 2 diabetes and fatty liver disease in obesity. JO - eLife VL - 12 PB - Elife Sciences Publ Ltd PY - 2024 SN - 2050-084X ER - TY - JOUR AB - S100A8/A9 is an endogenous alarmin secreted by myeloid cells during many acute and chronic inflammatory disorders. Despite increasing evidence of the proinflammatory effects of extracellular S100A8/A9, little is known about its intracellular function. Here, we show that cytosolic S100A8/A9 is indispensable for neutrophil post-arrest modifications during outside-in signaling under flow conditions in vitro and neutrophil recruitment in vivo, independent of its extracellular functions. Mechanistically, genetic deletion of S100A9 in mice caused dysregulated Ca2+ signatures in activated neutrophils resulting in reduced Ca2+ availability at the formed LFA-1/F-actin clusters with defective β2 integrin outside-in signaling during post-arrest modifications. Consequently, we observed impaired cytoskeletal rearrangement, cell polarization, and spreading, as well as cell protrusion formation in S100a9-/- compared to wildtype (WT) neutrophils, making S100a9-/- cells more susceptible to detach under flow, thereby preventing efficient neutrophil recruitment and extravasation into inflamed tissue. AU - Napoli, M.* AU - Immler, R.* AU - Rohwedder, I.* AU - Lupperger, V. AU - Pfabe, J.* AU - Gonzalez Pisfil, M.* AU - Yevtushenko, A.* AU - Vogl, T.* AU - Roth, J.* AU - Salvermoser, M.* AU - Dietzel, S.* AU - Slak Rupnik, M.* AU - Marr, C. AU - Walzog, B.* AU - Sperandio, M.* AU - Pruenster, M.* C1 - 72856 C2 - 56754 CY - 95 Regent Street, Cambridge, England TI - Cytosolic S100A8/A9 promotes Ca2+ supply at LFA-1 adhesion clusters during neutrophil recruitment. JO - eLife VL - 13 PB - Elife Sciences Publ Ltd PY - 2024 SN - 2050-084X ER - TY - JOUR AB - Mutations in the human PURA gene cause the neurodevelopmental PURA syndrome. In contrast to several other monogenetic disorders, almost all reported mutations in this nucleic acid-binding protein result in the full disease penetrance. In this study, we observed that patient mutations across PURA impair its previously reported co-localization with processing bodies. These mutations either destroyed the folding integrity, RNA binding, or dimerization of PURA. We also solved the crystal structures of the N- and C-terminal PUR domains of human PURA and combined them with molecular dynamics simulations and nuclear magnetic resonance measurements. The observed unusually high dynamics and structural promiscuity of PURA indicated that this protein is particularly susceptible to mutations impairing its structural integrity. It offers an explanation why even conservative mutations across PURA result in the full penetrance of symptoms in patients with PURA syndrome. AU - Proske, M. AU - Janowski, R. AU - Bacher, S. AU - Kang, H.-S. AU - Monecke, T.* AU - Koehler, T.* AU - Hutten, S.* AU - Tretter, J. AU - Crois, A.* AU - Molitor, L. AU - Varela-Rial, A.* AU - Fino, R.* AU - Donati, E.* AU - De Fabritiis, G.* AU - Dormann, D.* AU - Sattler, M. AU - Niessing, D. C1 - 70568 C2 - 55766 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - PURA syndrome-causing mutations impair PUR-domain integrity and affect P-body association. JO - eLife VL - 13 PB - Elife Sciences Publ Ltd PY - 2024 SN - 2050-084X ER - TY - JOUR AB - Mapping neurotransmitter identities to neurons is key to understanding information flow in a nervous system. It also provides valuable entry points for studying the development and plasticity of neuronal identity features. In the Caenorhabditis elegans nervous system, neurotransmitter identities have been largely assigned by expression pattern analysis of neurotransmitter pathway genes that encode neurotransmitter biosynthetic enzymes or transporters. However, many of these assignments have relied on multicopy reporter transgenes that may lack relevant cis-regulatory information and therefore may not provide an accurate picture of neurotransmitter usage. We analyzed the expression patterns of 16 CRISPR/Cas9-engineered knock-in reporter strains for all main types of neurotransmitters in C. elegans (glutamate, acetylcholine, GABA, serotonin, dopamine, tyramine, and octopamine) in both the hermaphrodite and the male. Our analysis reveals novel sites of expression of these neurotransmitter systems within both neurons and glia, as well as non-neural cells, most notably in gonadal cells. The resulting expression atlas defines neurons that may be exclusively neuropeptidergic, substantially expands the repertoire of neurons capable of co-transmitting multiple neurotransmitters, and identifies novel sites of monoaminergic neurotransmitter uptake. Furthermore, we also observed unusual co-expression patterns of monoaminergic synthesis pathway genes, suggesting the existence of novel monoaminergic transmitters. Our analysis results in what constitutes the most extensive whole-animal-wide map of neurotransmitter usage to date, paving the way for a better understanding of neuronal communication and neuronal identity specification in C. elegans. AU - Wang, C.* AU - Vidal, B.* AU - Sural, S.* AU - Loer, C.* AU - Aguilar, G.R.* AU - Merritt, D.M.* AU - Toker, I.A.* AU - Vogt, M.C. AU - Cros, C.C.* AU - Hobert, O.* C1 - 75208 C2 - 57847 CY - 95 Regent Street, Cambridge, England TI - A neurotransmitter atlas of C. elegans males and hermaphrodites. JO - eLife VL - 13 PB - Elife Sciences Publ Ltd PY - 2024 SN - 2050-084X ER - TY - JOUR AB - Brain size and cortical folding have increased and decreased recurrently during mammalian evolution. Identifying genetic elements whose sequence or functional properties co-evolve with these traits can provide unique information on evolutionary and developmental mechanisms. A good candidate for such a comparative approach is TRNP1, as it controls proliferation of neural progenitors in mice and ferrets. Here, we investigate the contribution of both regulatory and coding sequences of TRNP1 to brain size and cortical folding in over 30 mammals. We find that the rate of TRNP1 protein evolution (ω) significantly correlates with brain size, slightly less with cortical folding and much less with body size. This brain correlation is stronger than for >95% of random control proteins. This co-evolution is likely affecting TRNP1 activity, as we find that TRNP1 from species with larger brains and more cortical folding induce higher proliferation rates in neural stem cells. Furthermore, we compare the activity of putative cis-regulatory elements (CREs) of TRNP1 in a massively parallel reporter assay and identify one CRE that likely co-evolves with cortical folding in Old World monkeys and apes. Our analyses indicate that coding and regulatory changes that increased TRNP1 activity were positively selected either as a cause or a consequence of increases in brain size and cortical folding. They also provide an example how phylogenetic approaches can inform biological mechanisms, especially when combined with molecular phenotypes across several species. AU - Kliesmete, Z.* AU - Wange, L.E.* AU - Vieth, B.* AU - Esgleas Izquierdo, M. AU - Radmer, J.* AU - Hülsmann, M.* AU - Geuder, J.* AU - Richter, D.* AU - Ohnuki, M.* AU - Götz, M. AU - Hellmann, I.* AU - Enard, W.* C1 - 67699 C2 - 54006 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - Regulatory and coding sequences of TRNP1 co-evolve with brain size and cortical folding in mammals. JO - eLife VL - 12 PB - Elife Sciences Publ Ltd PY - 2023 SN - 2050-084X ER - TY - JOUR AB - The hypothalamus-pituitary-adrenal (HPA) axis is activated in response to inflammation leading to increased production of anti-inflammatory glucocorticoids by the adrenal cortex, thereby representing an endogenous feedback loop. However, severe inflammation reduces the responsiveness of the adrenal gland to adrenocorticotropic hormone (ACTH), although the underlying mechanisms are poorly understood. Here, we show by transcriptomic, proteomic, and metabolomic analyses that LPS-induced systemic inflammation triggers profound metabolic changes in steroidogenic adrenocortical cells, including downregulation of the TCA cycle and oxidative phosphorylation, in mice. Inflammation disrupts the TCA cycle at the level of succinate dehydrogenase (SDH), leading to succinate accumulation and disturbed steroidogenesis. Mechanistically, IL-1β reduces SDHB expression through upregulation of DNA methyltransferase 1 (DNMT1) and methylation of the SDHB promoter. Consequently, increased succinate levels impair oxidative phosphorylation and ATP synthesis and enhance ROS production, leading to reduced steroidogenesis. Together, we demonstrate that the IL-1β-DNMT1-SDHB-succinate axis disrupts steroidogenesis. Our findings not only provide a mechanistic explanation for adrenal dysfunction in severe inflammation, but also offer a potential target for therapeutic intervention. AU - Mateska, I.* AU - Witt, A.* AU - Hagag, E.* AU - Sinha, A.* AU - Yilmaz, C.* AU - Thanou, E.* AU - Sun, N. AU - Kolliniati, O.* AU - Patschin, M.* AU - Abdelmegeed, H.* AU - Henneicke, H.* AU - Kanczkowski, W.* AU - Wielockx, B.* AU - Tsatsanis, C.* AU - Dahl, A.* AU - Walch, A.K. AU - Li, K.W.* AU - Peitzsch, M.* AU - Chavakis, T.* AU - Alexaki, V.I.* C1 - 68102 C2 - 54580 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - Succinate mediates inflammation-induced adrenocortical dysfunction. JO - eLife VL - 12 PB - Elife Sciences Publ Ltd PY - 2023 SN - 2050-084X ER - TY - JOUR AB - The freshwater polyp Hydra is a popular biological model system; however, we still do not understand one of its most salient behaviors, the generation of spontaneous body wall contractions. Here, by applying experimental fluid dynamics analysis and mathematical modeling, we provide functional evidence that spontaneous contractions of body walls enhance the transport of chemical compounds from and to the tissue surface where symbiotic bacteria reside. Experimentally, a reduction in the frequency of spontaneous body wall contractions is associated with a changed composition of the colonizing microbiota. Together, our findings suggest that spontaneous body wall contractions create an important fluid transport mechanism that (1) may shape and stabilize specific host-microbe associations and (2) create fluid microhabitats that may modulate the spatial distribution of the colonizing microbes. This mechanism may be more broadly applicable to animal-microbe interactions since research has shown that rhythmic spontaneous contractions in the gastrointestinal tracts are essential for maintaining normal microbiota. AU - Nawroth, J. AU - Giez, C.* AU - Klimovich, A.* AU - Kanso, E.* AU - Bosch, T.C.G.* C1 - 68606 C2 - 54635 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - Spontaneous body wall contractions stabilize the fluid microenvironment that shapes host-microbe associations. JO - eLife VL - 12 PB - Elife Sciences Publ Ltd PY - 2023 SN - 2050-084X ER - TY - JOUR AB - We used non-invasive real-time genomic approaches to monitor one of the last surviving populations of the critically endangered kākāpō (Strigops habroptilus). We first established an environmental DNA metabarcoding protocol to identify the distribution of kākāpō and other vertebrate species in a highly localized manner using soil samples. Harnessing real-time nanopore sequencing and the high-quality kākāpō reference genome, we then extracted species-specific DNA from soil. We combined long read-based haplotype phasing with known individual genomic variation in the kākāpō population to identify the presence of individuals, and confirmed these genomically informed predictions through detailed metadata on kākāpō distributions. This study shows that individual identification is feasible through nanopore sequencing of environmental DNA, with important implications for future efforts in the application of genomics to the conservation of rare species, potentially expanding the application of real-time environmental DNA research from monitoring species distribution to inferring fitness parameters such as genomic diversity and inbreeding. AU - Urban, L. AU - Miller, A.K.* AU - Eason, D.* AU - Vercoe, D.* AU - Shaffer, M.* AU - Wilkinson, S.P.* AU - Jeunen, G.J.* AU - Gemmell, N.J.* AU - Digby, A.* C1 - 69027 C2 - 53812 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - Non-invasive real-time genomic monitoring of the critically endangered kākāpō. JO - eLife VL - 12 PB - Elife Sciences Publ Ltd PY - 2023 SN - 2050-084X ER - TY - JOUR AB - Delta-like homolog 1 (Dlk1), an inhibitor of adipogenesis, controls the cell fate of adipocyte progenitors. Experimental data presented here identify two independent regulatory mechanisms, transcriptional and translational, by which Ifrd1 (TIS7) and its orthologue Ifrd2 (SKMc15) regulate Dlk1 levels. Mice deficient in both Ifrd1 and Ifrd2 (dKO) had severely reduced adipose tissue and were resistant to high-fat diet-induced obesity. Wnt signaling, a negative regulator of adipocyte differentiation, was significantly upregulated in dKO mice. Elevated levels of the Wnt/β-catenin target protein Dlk1 inhibited the expression of adipogenesis regulators Pparg and Cebpa, and fatty acid transporter Cd36. Although both Ifrd1 and Ifrd2 contributed to this phenotype, they utilized two different mechanisms. Ifrd1 acted by controlling Wnt signaling and thereby transcriptional regulation of Dlk1. On the other hand, distinctive experimental evidence showed that Ifrd2 acts as a general translational inhibitor significantly affecting Dlk1 protein levels. Novel mechanisms of Dlk1 regulation in adipocyte differentiation involving Ifrd1 and Ifrd2 are based on experimental data presented here. AU - Vietor, I.* AU - Cikes, D.* AU - Piironen, K.* AU - Vasakou, T.* AU - Heimdörfer, D.* AU - Gstir, R.* AU - Erlacher, M.D.* AU - Tancevski, I.* AU - Eller, P.* AU - Demetz, E.* AU - Hess, M.W.* AU - Kuhn, V.* AU - Degenhart, G.* AU - Rozman, J. AU - Klingenspor, M.* AU - Hrabě de Angelis, M. AU - Valovka, T.* AU - Huber, L.A.* C1 - 67977 C2 - 54455 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - The negative adipogenesis regulator Dlk1 is transcriptionally regulated by Ifrd1 (TIS7) and translationally by its orthologue Ifrd2 (SKMc15). JO - eLife VL - 12 PB - Elife Sciences Publ Ltd PY - 2023 SN - 2050-084X ER - TY - JOUR AB - Neuroinflammation after stroke is characterized by the activation of resident microglia and the invasion of circulating leukocytes into the brain. Although lymphocytes infiltrate the brain in small number, they have been consistently demonstrated to be the most potent leukocyte subpopulation contributing to secondary inflammatory brain injury. However, the exact mechanism of how this minimal number of lymphocytes can profoundly affect stroke outcome is still largely elusive. Here, using a mouse model for ischemic stroke, we demonstrated that early activation of microglia in response to stroke is differentially regulated by distinct T cell subpopulations - with TH1 cells inducing a type I INF signaling in microglia and regulatory T cells (TREG) cells promoting microglial genes associated with chemotaxis. Acute treatment with engineered T cells overexpressing IL-10 administered into the cisterna magna after stroke induces a switch of microglial gene expression to a profile associated with pro-regenerative functions. Whereas microglia polarization by T cell subsets did not affect the acute development of the infarct volume, these findings substantiate the role of T cells in stroke by polarizing the microglial phenotype. Targeting T cell-microglia interactions can have direct translational relevance for further development of immune-targeted therapies for stroke and other neuroinflammatory conditions. AU - Benakis, C.* AU - Simats, A.* AU - Tritschler, S. AU - Heindl, S.* AU - Besson-Girard, S.* AU - Llovera, G.* AU - Pinkham, K.* AU - Kolz, A.* AU - Ricci, A.* AU - Theis, F.J. AU - Bittner, S.* AU - Gökçe, A.* AU - Peters, A.* AU - Liesz, A.* C1 - 66991 C2 - 53399 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - T cells modulate the microglial response to brain ischemia. JO - eLife VL - 11 PB - Elife Sciences Publ Ltd PY - 2022 SN - 2050-084X ER - TY - JOUR AB - The iron hormone hepcidin is transcriptionally activated by iron or inflammation via distinct, partially overlapping pathways. We addressed how iron affects inflammatory hepcidin levels and the ensuing hypoferremic response. Dietary iron overload did not mitigate hepcidin induction in LPS-treated wt mice but prevented effective inflammatory hypoferremia. Likewise, LPS modestly decreased serum iron in hepcidin-deficient Hjv-/- mice, model of hemochromatosis. Synthetic hepcidin triggered hypoferremia in control but not iron-loaded wt animals. Furthermore, it dramatically decreased hepatic and splenic ferroportin in Hjv-/- mice on standard or iron-deficient diet, but only triggered hypoferremia in the latter. Mechanistically, iron antagonized hepcidin responsiveness by inactivating IRPs in the liver and spleen, to stimulate ferroportin mRNA translation. Prolonged LPS treatment eliminating ferroportin mRNA permitted hepcidin-mediated hypoferremia in iron-loaded mice. Thus, de novo ferroportin synthesis is critical determinant of serum iron and finetunes hepcidin-dependent functional outcomes. Our data uncover a crosstalk between hepcidin and IRE/IRP systems that controls tissue ferroportin expression and determines serum iron levels. Moreover, they suggest that hepcidin supplementation therapy is more efficient combined with iron depletion. AU - Charlebois, E.* AU - Fillebeen, C.* AU - Katsarou, A.* AU - Rabinovich, A.* AU - Wisniewski, K.* AU - Venkataramani, V.* AU - Michalke, B. AU - Velentza, A.* AU - Pantopoulos, K.* C1 - 66174 C2 - 53112 TI - A crosstalk between hepcidin and IRE/IRP pathways controls ferroportin expression and determines serum iron levels in mice. JO - eLife VL - 11 PY - 2022 SN - 2050-084X ER - TY - JOUR AB - Anthracyclines are among the most used and effective anticancer drugs. Their activity has been attributed to DNA double-strand breaks resulting from topoisomerase II poisoning and to eviction of histones from select sites in the genome. Here we show that the extensively used anthracyclines Doxorubicin, Daunorubicin and Epirubicin decrease the transcription of nuclear factor kappa B (NF-κB)-dependent gene targets, but not interferon responsive genes in primary mouse (Mus musculus) macrophages. Using an NMR-based structural approach, we demonstrate that anthracyclines disturb the complexes formed between the NF-kB subunit RelA and its DNA binding sites. The anthracycline variants Aclarubicin, Doxorubicinone and the newly developed Dimethyl-doxorubicin, which share anticancer properties with the other anthracyclines but do not induce DNA damage, also suppressed inflammation, thus uncoupling DNA damage from the effects on inflammation. These findings have implications for anticancer therapy and for the development of novel anti-inflammatory drugs with limited side effects for life-threatening conditions such as sepsis. AU - Chora, A.F.* AU - Pedroso, D.* AU - Kyriakou, E. AU - Pejanovic, N.* AU - Colaço, H.* AU - Gozzelino, R.* AU - Barros, A.* AU - Willmann, K.* AU - Velho, T.R.* AU - Moita, C.* AU - Santos, I.* AU - Pereira, P.* AU - Carvalho, S.* AU - Martins, F.B.* AU - Ferreira, J.A.* AU - de Almeida, S.F.* AU - Benes, V.* AU - Anrather, J.* AU - Weis, S.* AU - Soares, M.P.* AU - Geerlof, A. AU - Neefjes, J.J.* AU - Sattler, M.* AU - Messias, A.C. AU - Neves Costa, A.* AU - Moita, L.F.* C1 - 67006 C2 - 53406 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - DNA damage independent inhibition of NF-kB transcription by anthracyclines. JO - eLife VL - 11 PB - Elife Sciences Publ Ltd PY - 2022 SN - 2050-084X ER - TY - JOUR AB - Cleavage of membrane proteins in the lipid bilayer by intramembrane proteases is crucial for health and disease. Although different lipid environments can potently modulate their activity, how this is linked to their structural dynamics is unclear. Here we show that the carboxy-peptidase-like activity of the archaeal intramembrane protease PSH, a homolog of the Alzheimer's disease-associated presenilin/γ-secretase is impaired in micelles and promoted in a lipid bilayer. Comparative molecular dynamics simulations revealed that important elements for substrate binding such as transmembrane domain 6a of PSH are more labile in micelles and stabilized in the lipid bilayer. Moreover, consistent with an enhanced interaction of PSH with a transition-state analog inhibitor, the bilayer promoted the formation of the enzyme´s catalytic active site geometry. Our data indicate that the lipid environment of an intramembrane protease plays a critical role in structural stabilization and active site arrangement of the enzyme-substrate complex thereby promoting intramembrane proteolysis. AU - Feilen, L.P.* AU - Chen, S.Y.* AU - Fukumori, A.* AU - Feederle, R. AU - Zacharias, M.* AU - Steiner, H.* C1 - 64985 C2 - 52601 TI - Active site geometry stabilization of a presenilin homolog by the lipid bilayer promotes intramembrane proteolysis. JO - eLife VL - 11 PY - 2022 SN - 2050-084X ER - TY - JOUR AB - Protein biomarkers have been identified across many age-related morbidities. However, characterising epigenetic influences could further inform disease predictions. Here, we leverage epigenome-wide data to study links between the DNAm signatures of the circulating proteome and incident diseases. Using data from four cohorts, we trained and tested epigenetic scores (EpiScores) for 953 plasma proteins, identifying 109 scores that explained between 1% and 58% of the variance in protein levels after adjusting for known protein quantitative trait loci (pQTL) genetic effects. By projecting these EpiScores into an independent sample, (Generation Scotland; n=9,537) and relating them to incident morbidities over a follow-up of 14 years, we uncovered 137 EpiScore - disease associations. These associations were largely independent of immune cell proportions, common lifestyle and health factors and biological aging. Notably, we found that our diabetes-associated EpiScores highlighted previous top biomarker associations from proteome-wide assessments of diabetes. These EpiScores for protein levels can therefore be a valuable resource for disease prediction and risk stratification. AU - Gadd, D.A.* AU - Hillary, R.F.* AU - McCartney, D.L.* AU - Zaghlool, S.B.* AU - Stevenson, A.J.* AU - Cheng, Y.* AU - Fawns-Ritchie, C.* AU - Nangle, C.* AU - Campbell, A.* AU - Flaig, R.* AU - Harris, S.E.* AU - Walker, R.M.* AU - Shi, L.* AU - Tucker-Drob, E.M.* AU - Gieger, C. AU - Peters, A. AU - Waldenberger, M. AU - Graumann, J.* AU - McRae, A.F.* AU - Deary, I.J.* AU - Porteous, D.J.* AU - Hayward, C.* AU - Visscher, P.M.* AU - Cox, S.R.* AU - Evans, K.L.* AU - McIntosh, A.M.* AU - Suhre, K.* AU - Marioni, R.E.* C1 - 64070 C2 - 52029 TI - Epigenetic scores for the circulating proteome as tools for disease prediction. JO - eLife VL - 11 PY - 2022 SN - 2050-084X ER - TY - JOUR AB - Protozoa and fungi are known to have extraordinarily diverse mechanisms of genetic exchange. However, the presence and epidemiological relevance of genetic exchange in Trypanosoma cruzi, the agent of Chagas disease, has been controversial and debated for many years. Field studies have identified both predominantly clonal and sexually recombining natural populations. Two of six natural T. cruzi lineages (TcV and TcVI) show hybrid mosaicism, using analysis of single-gene locus markers. The formation of hybrid strains in vitro has been achieved and this provides a framework to study the mechanisms and adaptive significance of genetic exchange. Using whole genome sequencing of a set of experimental hybrids strains, we have confirmed that hybrid formation initially results in tetraploid parasites. The hybrid progeny showed novel mutations that were not attributable to either (diploid) parent showing an increase in amino acid changes. In long-term culture, up to 800 generations, there was a variable but gradual erosion of progeny genomes towards triploidy, yet retention of elevated copy number was observed at several core housekeeping loci. Our findings indicate hybrid formation by fusion of diploid T. cruzi, followed by sporadic genome erosion, but with substantial potential for adaptive evolution, as has been described as a genetic feature of other organisms, such as some fungi. AU - Matos, G.M.* AU - Lewis, M.D.* AU - Talavera Lopez, C.N. AU - Yeo, M.* AU - Grisard, E.C.* AU - Messenger, L.A.* AU - Miles, M.A.* AU - Andersson, B.* C1 - 65013 C2 - 52614 TI - Microevolution of Trypanosoma cruzi reveals hybridization and clonal mechanisms driving rapid genome diversification. JO - eLife VL - 11 PY - 2022 SN - 2050-084X ER - TY - JOUR AB - Early events during axolotl limb regeneration include an immune response and the formation of a wound epithelium. These events are linked to a clearance of damaged tissue prior to blastema formation and regeneration of the missing structures. Here, we report the resorption of calcified skeletal tissue as an active, cell-driven, and highly regulated event. This process, carried out by osteoclasts, is essential for a successful integration of the newly formed skeleton. Indeed, the extent of resorption is directly correlated with the integration efficiency, and treatment with zoledronic acid resulted in osteoclast function inhibition and failed tissue integration. Moreover, we identified the wound epithelium as a regulator of skeletal resorption, likely releasing signals involved in recruitment/differentiation of osteoclasts. Finally, we reported a correlation between resorption and blastema formation, particularly, a coordination of resorption with cartilage condensation. In sum, our results identify resorption as a major event upon amputation, playing a critical role in the overall process of skeletal regeneration. AU - Riquelme-Guzmán, C.* AU - Tsai, S.L.* AU - Carreon Paz, K.* AU - Nguyen, C.* AU - Oriola, D.* AU - Schuez, M.* AU - Brugués, J.* AU - Currie, J.D.* AU - Sandoval-Guzmán, T. C1 - 66498 C2 - 53195 TI - Osteoclast-mediated resorption primes the skeleton for successful integration during axolotl limb regeneration. JO - eLife VL - 11 PY - 2022 SN - 2050-084X ER - TY - JOUR AB - The eLife Early-Career Advisory Group discusses eLife's new peer review and publishing model, and how the whole process of scientific communication could be improved for the benefit of early-career researchers and the entire scientific community. AU - Urban, L. AU - De Niz, M.* AU - Fernández-Chiappe, F.* AU - Ebrahimi, H.* AU - Han, L.K.M.* AU - Mehta, D.* AU - Mencia, R.* AU - Mittal, D.* AU - Ochola, E.* AU - Paz Quezada, C.* AU - Romani, F.* AU - Sinapayen, L.* AU - Tay, A.* AU - Varma, A.* AU - Yahia Mohamed Elkheir, L.* C1 - 67005 C2 - 53405 TI - eLife's new model and its impact on science communication. JO - eLife VL - 11 PY - 2022 SN - 2050-084X ER - TY - JOUR AB - Accurate brain tissue extraction on magnetic resonance imaging (MRI) data is crucial for analyzing brain structure and function. While several conventional tools have been optimized to handle human brain data, there have been no generalizable methods to extract brain tissues for multimodal MRI data from rodents, nonhuman primates, and humans. Therefore, developing a flexible and generalizable method for extracting whole brain tissue across species would allow researchers to analyze and compare experiment results more efficiently. Here, we propose a domain-adaptive and semi-supervised deep neural network, named the Brain Extraction Net (BEN), to extract brain tissues across species, MRI modalities, and MR scanners. We have evaluated BEN on 18 independent datasets, including 783 rodent MRI scans, 246 nonhuman primate MRI scans, and 4,601 human MRI scans, covering five species, four modalities, and six MR scanners with various magnetic field strengths. Compared to conventional toolboxes, the superiority of BEN is illustrated by its robustness, accuracy, and generalizability. Our proposed method not only provides a generalized solution for extracting brain tissue across species but also significantly improves the accuracy of atlas registration, thereby benefiting the downstream processing tasks. As a novel fully automated deep-learning method, BEN is designed as an open-source software to enable high-throughput processing of neuroimaging data across species in preclinical and clinical applications. AU - Yu, Z.* AU - Han, X.* AU - Xu, W.* AU - Zhang, J.* AU - Marr, C. AU - Shen, D.* AU - Peng, T. AU - Zhang, X.Y.* AU - Feng, J.* C1 - 67136 C2 - 53456 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - A generalizable brain extraction net (BEN) for multimodal MRI data from rodents, nonhuman primates, and humans. JO - eLife VL - 11 PB - Elife Sciences Publ Ltd PY - 2022 SN - 2050-084X ER - TY - JOUR AB - 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. AU - Bespalov, A.* AU - Bernard, R.* AU - Gilis, A.* AU - Gerlach, B.* AU - Guillen, J.* AU - Castagne, V.* AU - Lefevre, I.* AU - Ducrey, F.* AU - Monk, L.* AU - Bongiovanni, S.* AU - Altevogt, B.* AU - Arroyo Araujo, M.* AU - Bikovski, L.* AU - de Bruin, N.* AU - Castanos-Velez, E.* AU - Dityatev, A.* AU - Emmerich, C.H.* AU - Fares, R.* AU - Ferland-Beckham, C.* AU - Froger-Colléaux, C.* AU - Gailus-Durner, V. AU - Hölter, S.M. AU - Hofmann, M.C.* AU - Kabitzke, P.* AU - Kas, M.J.H.* AU - Kurreck, C.* AU - Moser, P.* AU - Pietraszek, M.* AU - Popik, P.* AU - Potschka, H.* AU - Prado Montes de Oca, E.* AU - Restivo, L.* AU - Riedel, G.* AU - Ritskes-Hoitinga, M.* AU - Samardzic, J.* AU - Schunn, M.* AU - Stoeger, C. AU - Voikar, V.* AU - Vollert, J.* AU - Wever, K.E.* AU - Wuyts, K.* AU - MacLeod, M.R.* AU - Dirnagl, U.* AU - Steckler, T.* C1 - 62077 C2 - 50634 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - Introduction to the EQIPD quality system. JO - eLife VL - 10 PB - Elife Sciences Publications Ltd PY - 2021 SN - 2050-084X ER - TY - JOUR AB - From 1985 to 2016, the prevalence of underweight decreased, and that of obesity and severe obesity increased, in most regions, with significant variation in the magnitude of these changes across regions. We investigated how much change in mean body mass index (BMI) explains changes in the prevalence of underweight, obesity, and severe obesity in different regions using data from 2896 population-based studies with 187 million participants. Changes in the prevalence of underweight and total obesity, and to a lesser extent severe obesity, are largely driven by shifts in the distribution of BMI, with smaller contributions from changes in the shape of the distribution. In East and Southeast Asia and sub-Saharan Africa, the underweight tail of the BMI distribution was left behind as the distribution shifted. There is a need for policies that address all forms of malnutrition by making healthy foods accessible and affordable, while restricting unhealthy foods through fiscal and regulatory restrictions. AU - NCD Risk Factors Collaboration (Döring, A. AU - Meisinger, C. AU - Müller-Nurasyid, M. AU - Peters, A. AU - Stieber, J. AU - Stöckl, D.) C1 - 61715 C2 - 50168 TI - Heterogeneous contributions of change in population distribution of body mass index to change in obesity and underweight NCD Risk Factor Collaboration (NCD-RisC). JO - eLife VL - 10 PY - 2021 SN - 2050-084X ER - TY - JOUR AB - Eukaryotic DNA replication initiates during S phase from origins that have been licensed in the preceding G1 phase. Here, we compare ChIP-seq profiles of the licensing factors Orc2, Orc3, Mcm3, and Mcm7 with gene expression, replication timing and fork directionality profiles obtained by RNA-seq, Repli-seq and OK-seq. ORC and MCM are significantly and homogeneously depleted from transcribed genes, enriched at gene promoters, and more abundant in early- than in late-replicating domains. Surprisingly, after controlling these variables, no difference in ORC/MCM density is detected between initiation zones, termination zones, unidirectionally replicating and randomly replicating regions. Therefore, ORC/MCM density correlates with replication timing but does not solely regulate the probability of replication initiation. Interestingly, H4K20me3, a histone modification proposed to facilitate late origin licensing, was enriched in late replicating initiation zones and gene deserts of stochastic replication fork direction. We discuss potential mechanisms specifying when and where replication initiates in human cells. AU - Kirstein, N. AU - Buschle, A. AU - Wu, X.* AU - Krebs, S.* AU - Blum, H.* AU - Kremmer, E. AU - Vorberg, I.M.* AU - Hammerschmidt, W. AU - Lacroix, L.* AU - Hyrien, O.* AU - Audit, B.* AU - Schepers, A. C1 - 61435 C2 - 50243 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - Human ORC/MCM density is low in active genes and correlates with replication time but does not delimit initiation zones. JO - eLife VL - 10 PB - Elife Sciences Publications Ltd PY - 2021 SN - 2050-084X ER - TY - JOUR AB - Analysing the characteristics of the SARS-CoV-2 virus makes it possible to estimate the length of quarantine that reduces the impact on society and the economy, while minimising infections. AU - Kretzschmar, M.* AU - Müller, J. C1 - 61661 C2 - 50372 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - Should I stay or should I go? JO - eLife VL - 10 PB - Elife Sciences Publications Ltd PY - 2021 SN - 2050-084X ER - TY - JOUR AB - SNARE proteins have been described as the effectors of fusion events in the secretory pathway more than two decades ago. The strong interactions between SNARE domains are clearly important in membrane fusion, but it is unclear whether they are involved in any other cellular processes. Here, we analyzed two classical SNARE proteins, syntaxin 1A and SNAP25. Although they are supposed to be engaged in tight complexes, we surprisingly find them largely segregated in the plasma membrane. Syntaxin 1A only occupies a small fraction of the plasma membrane area. Yet, we find it is able to redistribute the far more abundant SNAP25 on the mesoscale by gathering crowds of SNAP25 molecules onto syntaxin clusters in a SNARE-domain-dependent manner. Our data suggest that SNARE domain interactions are not only involved in driving membrane fusion on the nanoscale, but also play an important role in controlling the general organization of proteins on the mesoscale. Further, we propose these mechanisms preserve active syntaxin 1A-SNAP25 complexes at the plasma membrane. AU - Mertins, J.* AU - Finke, J.* AU - Sies, R.* AU - Rink, K.M.* AU - Hasenauer, J. AU - Lang, T.* C1 - 63753 C2 - 51552 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England SP - 2624-2624 TI - The mesoscale organization of syntaxin 1A and SNAP25 is determined by SNARE-SNARE interactions. JO - eLife VL - 10 IS - 11 PB - Elife Sciences Publ Ltd PY - 2021 SN - 2050-084X ER - TY - JOUR AB - Triple-negative breast cancer (TNBC) is an aggressive tumor with limited treatment options and poor prognosis. We applied the in vivo phage display technology to isolate peptides homing to the immunosuppressive cellular microenvironment of TNBC as a strategy for non-malignant target discovery. We identified a cyclic peptide (CSSTRESAC) that specifically binds to a vitamin D receptor, protein disulfide-isomerase A3 (PDIA3) expressed on the cell surface of tumor-associated macrophages (TAM), and targets breast cancer in syngeneic TNBC, non-TNBC xenograft, and transgenic mouse models. Systemic administration of CSSTRESAC to TNBC-bearing mice shifted the cytokine profile toward an antitumor immune response and delayed tumor growth. Moreover, CSSTRESAC enabled ligand-directed theranostic delivery to tumors and a mathematical model confirmed our experimental findings. Finally, in silico analysis showed PDIA3-expressing TAM in TNBC patients. This work uncovers a functional interplay between a cell surface vitamin D receptor in TAM and antitumor immune response that could be therapeutically exploited. AU - Staquicini, F.I.* AU - Hajitou, A.* AU - Driessen, W.H.* AU - Proneth, B. AU - Cardó-Vila, M.* AU - Staquicini, D.I.* AU - Markosian, C.* AU - Hoh, M.* AU - Cortez, M.* AU - Hooda-Nehra, A.* AU - Jaloudi, M.* AU - Silva, I.T.* AU - Buttura, J.* AU - Nunes, D.* AU - Dias-Neto, E.* AU - Eckhardt, B.* AU - Ruiz-Ramírez, J.* AU - Dogra, P.* AU - Wang, Z.* AU - Cristini, V.* AU - Trepel, M.* AU - Anderson, R.* AU - Sidman, R.L.* AU - Gelovani, J.G.* AU - Cristofanilli, M.* AU - Hortobagy, G.* AU - Bhujwalla, Z.M.* AU - Burley, S.* AU - Arap, W.* AU - Pasqualini, R.* C1 - 62213 C2 - 50734 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - Targeting a cell surface vitamin D receptor on tumor-associated macrophages in triple-negative breast cancer. JO - eLife VL - 10 PB - Elife Sciences Publications Ltd PY - 2021 SN - 2050-084X ER - TY - JOUR AB - Interleukin-4-induced-1 (IL4i1) is an amino acid oxidase secreted from immune cells. Recent observations have suggested that IL4i1 is pro-tumorigenic via unknown mechanisms. As IL4i1 has homologues in snake venoms (LAAO, L-amino acid oxidases), we used comparative approaches to gain insight into the mechanistic basis of how conserved amino acid oxidases regulate cell fate and function. Using mammalian expressed recombinant proteins, we found venom LAAO kills cells via hydrogen peroxide generation. By contrast, mammalian IL4i1 is non-cytotoxic and instead elicits a cell productive gene expression program inhibiting ferroptotic redox death by generating indole-3-pyruvate (I3P) from tryptophan. I3P suppresses ferroptosis by direct free radical scavenging and through the activation of an anti-oxidative gene expression program. Thus, the pro-tumor effects of IL4i1 are likely mediated by local anti-ferroptotic pathways via aromatic amino acid metabolism, arguing that an IL4i1 inhibitor may modulate tumor cell death pathways. AU - Zeitler, L.* AU - Fiore, A.* AU - Meyer, C.* AU - Russier, M.* AU - Zanella, G.* AU - Suppmann, S.* AU - Gagaro, M.* AU - Sidhu, S.S.* AU - Seshagiri, S.* AU - Ohnmacht, C. AU - Köcher, T.* AU - Fallarino, F.* AU - Linkermann, A.* AU - Murray, P.J.* C1 - 61466 C2 - 50274 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - Anti-ferroptotic mechanism of IL4i1-mediated amino acid metabolism. JO - eLife VL - 10 PB - Elife Sciences Publications Ltd PY - 2021 SN - 2050-084X ER - TY - JOUR AB - All eukaryotes require iron. Replication, detoxification, and a cancer-protective form of regulated cell death termed ferroptosis, all depend on iron metabolism. Ferrous iron accumulates over adult lifetime in Caenorhabditis elegans. Here, we show that glutathione depletion is coupled to ferrous iron elevation in these animals, and that both occur in late life to prime cells for ferroptosis. We demonstrate that blocking ferroptosis, either by inhibition of lipid peroxidation or by limiting iron retention, mitigates age-related cell death and markedly increases lifespan and healthspan. Temporal scaling of lifespan is not evident when ferroptosis is inhibited, consistent with this cell death process acting at specific life phases to induce organismal frailty, rather than contributing to a constant aging rate. Because excess age-related iron elevation in somatic tissue, particularly in brain, is thought to contribute to degenerative disease, post-developmental interventions to limit ferroptosis may promote healthy aging. AU - Jenkins, N.L.* AU - James, S.A.* AU - Salim, A.* AU - Sumardy, F.* AU - Speed, T.P.* AU - Conrad, M. AU - Richardson, D.R.* AU - Bush, A.I.* AU - McColl, G.* C1 - 59782 C2 - 49040 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - Changes in ferrous iron and glutathione promote ferroptosis and frailty in aging Caenorhabditis elegans. JO - eLife VL - 9 PB - Elife Sciences Publications Ltd PY - 2020 SN - 2050-084X ER - TY - JOUR AB - Perturbation of addition of second heart field (SHF) cardiac progenitor cells to the poles of the heart tube results in congenital heart defects (CHD). The transcriptional programs and upstream regulatory events operating in different subpopulations of the SHF remain unclear. Here, we profile the transcriptome and chromatin accessibility of anterior and posterior SHF subpopulations at genome-wide levels and demonstrate that Hoxbl negatively regulates differentiation in the posterior SHF. Spatial mis-expression of Hoxbl in the anterior SHF results in hypoplastic right ventricle. Activation of Hoxbl in embryonic stem cells arrests cardiac differentiation, whereas Hoxbl-deficient mouse embryos display premature cardiac differentiation. Moreover, ectopic differentiation in the posterior SHF of embryos lacking both Hoxbl and its paralog Hoxal results in atrioventricular septal defects. Our results show that Hoxbl plays a key role in patterning cardiac progenitor cells that contribute to both cardiac poles and provide new insights into the pathogenesis of CHD. AU - Stefanovic, S.* AU - Laforest, B.* AU - Desvignes, J.P.* AU - Lescroart, F.* AU - Argiro, L.* AU - Maurel-Zaffran, C.* AU - Salgado, D.* AU - Plaindoux, E.* AU - De Bono, C.* AU - Pazur, K. AU - Théveniau-Ruissy, M.* AU - Béroud, C.* AU - Puceat, M.* AU - Gavalas, A. AU - Kelly, R.G.* AU - Zaffran, S.* C1 - 60022 C2 - 49170 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - Hox-dependent coordination of mouse cardiac progenitor cell patterning and differentiation. JO - eLife VL - 9 PB - Elife Sciences Publications Ltd PY - 2020 SN - 2050-084X ER - TY - JOUR AB - In recent years, functional genomics approaches combining genetic information with bulk RNA-sequencing data have identified the downstream expression effects of disease-associated genetic risk factors through so-called expression quantitative trait locus (eQTL) analysis. Single-cell RNA-sequencing creates enormous opportunities for mapping eQTLs across different cell types and in dynamic processes, many of which are obscured when using bulk methods. Rapid increase in throughput and reduction in cost per cell now allow this technology to be applied to large-scale population genetics studies. To fully leverage these emerging data resources, we have founded the single-cell eQTLGen consortium (sc-eQTLGen), aimed at pinpointing the cellular contexts in which disease-causing genetic variants affect gene expression. Here, we outline the goals, approach and potential utility of the sc-eQTLGen consortium. We also provide a set of study design considerations for future single-cell eQTL studies. AU - van der Wijst, M.* AU - de Vries, D.H.* AU - Groot, H.E.* AU - Trynka, G.* AU - Hon, C.C.* AU - Bonder, M.J.* AU - Stegle, O.* AU - Nawijn, M.C.* AU - Idaghdour, Y.* AU - van der Harst, P.* AU - Ye, C.J.* AU - Powell, J.* AU - Theis, F.J. AU - Mahfouz, A.* AU - Heinig, M. AU - Franke, L.* C1 - 58615 C2 - 48368 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - The single-cell eQTLGen consortium. JO - eLife VL - 9 PB - Elife Sciences Publications Ltd PY - 2020 SN - 2050-084X ER - TY - JOUR AB - High-throughput testing of drugs across molecular-characterised cell lines can identify candidate treatments and discover biomarkers. However, the cells' response to a drug is typically quantified by a summary statistic from a best-fit dose-response curve, whilst neglecting the uncertainty of the curve fit and the potential variability in the raw readouts. Here, we model the experimental variance using Gaussian Processes, and subsequently, leverage uncertainty estimates to identify associated biomarkers with a new Bayesian framework. Applied to in vitro screening data on 265 compounds across 1074 cancer cell lines, our models identified 24 clinically established drug-response biomarkers, and provided evidence for six novel biomarkers by accounting for association with low uncertainty. We validated our uncertainty estimates with an additional drug screen of 26 drugs, 10 cell lines with 8 to 9 replicates. Our method is applicable to any dose-response data without replicates, and improves biomarker discovery for precision medicine. AU - Wang, D.* AU - Hensman, J.* AU - Kutkaite, G. AU - Toh, T.S.* AU - Galhoz, A. AU - Dry, J.R.* AU - Saez-Rodriguez, J.* AU - Garnett, M.J.* AU - Menden, M.P. AU - Dondelinger, F.* C1 - 60913 C2 - 49743 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - A statistical framework for assessing pharmacological responses and biomarkers using uncertainty estimates. JO - eLife VL - 9 PB - Elife Sciences Publications Ltd PY - 2020 SN - 2050-084X ER - TY - JOUR AB - Thylakoid membranes scaffold an assortment of large protein complexes that work together to harness the energy of light. It has been a longstanding challenge to visualize how the intricate thylakoid network organizes these protein complexes to finely tune the photosynthetic reactions. Previously, we used in situ cryo-electron tomography to reveal the native architecture of thylakoid membranes (Engel et al., 2015). Here, we leverage technical advances to resolve the individual protein complexes within these membranes. Combined with a new method to visualize membrane surface topology, we map the molecular landscapes of thylakoid membranes inside green algae cells. Our tomograms provide insights into the molecular forces that drive thylakoid stacking and reveal that photosystems I and II are strictly segregated at the borders between appressed and non-appressed membrane domains. This new approach to charting thylakoid topology lays the foundation for dissecting photosynthetic regulation at the level of single protein complexes within the cell. AU - Wietrzynski, W. AU - Schaffer, M.* AU - Tegunov, D.* AU - Albert, S.* AU - Kanazawa, A.* AU - Plitzko, J.M.* AU - Baumeister, W.* AU - Engel, B.D. C1 - 58867 C2 - 48407 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - Charting the native architecture of Chlamydomonas thylakoid membranes with single-molecule precision. JO - eLife VL - 9 PB - Elife Sciences Publications Ltd PY - 2020 SN - 2050-084X ER - TY - JOUR AB - Islet vascularization is essential for intact islet function and glucose homeostasis. We have previously shown that primary cilia directly regulate insulin secretion. However, it remains unclear whether they are also implicated in islet vascularization. At eight weeks, murine Bbs4-/-islets show significantly lower intra-islet capillary density with enlarged diameters. Transplanted Bbs4-/- islets exhibit delayed re-vascularization and reduced vascular fenestration after engraftment, partially impairing vascular permeability and glucose delivery to β-cells. We identified primary cilia on endothelial cells as the underlying cause of this regulation, via the vascular endothelial growth factor-A (VEGF-A)/VEGF receptor 2 (VEGFR2) pathway. In vitro silencing of ciliary genes in endothelial cells disrupts VEGF-A/VEGFR2 internalization and downstream signaling. Consequently, key features of angiogenesis including proliferation and migration are attenuated in human BBS4 silenced endothelial cells. We conclude that endothelial cell primary cilia regulate islet vascularization and vascular barrier function via the VEGF-A/VEGFR2 signaling pathway. AU - Xiong, Y.* AU - Scerbo, M.J. AU - Seelig, A. AU - Volta, F. AU - O'Brien, N. AU - Dicker, A.* AU - Padula, D. AU - Lickert, H. AU - Gerdes, J.M. AU - Berggren, P.O.* C1 - 60646 C2 - 49558 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - Islet vascularization is regulated by primary endothelial cilia via VEGF-A-dependent signaling. JO - eLife VL - 9 PB - Elife Sciences Publications Ltd PY - 2020 SN - 2050-084X ER - TY - JOUR AB - Lung cancer and chronic lung diseases impose major disease burdens worldwide and are caused by inhaled noxious agents including tobacco smoke. The cellular origins of environmental-induced lung tumors and of the dysfunctional airway and alveolar epithelial turnover observed with chronic lung diseases are unknown. To address this, we combined mouse models of genetic labeling and ablation of airway (club) and alveolar cells with exposure to environmental noxious and carcinogenic agents. Club cells are shown to survive KRAS mutations and to form lung tumors after tobacco carcinogen exposure. Increasing numbers of club cells are found in the alveoli with aging and after lung injury, but go undetected since they express alveolar proteins. Ablation of club cells prevents chemical lung tumors and causes alveolar destruction in adult mice. Hence club cells are important in alveolar maintenance and carcinogenesis and may be a therapeutic target against premalignancy and chronic lung disease. AU - Spella, M.* AU - Lilis, I.* AU - Pepe, M. AU - Chen, Y.* AU - Armaka, M.* AU - Lamort, A.-S. AU - Zazara, D.E.* AU - Roumelioti, F.* AU - Vreka, M. AU - Kanellakis, N.I.* AU - Wagner, D.E. AU - Giannou, A.D.* AU - Armenis, V.* AU - Arendt, K.A.M. AU - Klotz, L.V. AU - Toumpanakis, D.* AU - Karavana, V.* AU - Zakynthinos, S.G.* AU - Giopanou, I.* AU - Marazioti, A.* AU - Aidinis, V.* AU - Sotillo, R.* AU - Stathopoulos, G.T. C1 - 56181 C2 - 46873 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - Club cells form lung adenocarcinomas and maintain the alveoli of adult mice. JO - eLife VL - 8 PB - Elife Sciences Publications Ltd PY - 2019 SN - 2050-084X ER - TY - JOUR AB - We use a genome-wide association of 1 million parental lifespans of genotyped subjects and data on mortality risk factors to validate previously unreplicated findings near CDKN2B-AS1, ATXN2/BRAP, FURIN/FES, ZW10, PSORS1C3, and 13q21.31, and identify and replicate novel findings near ABO, ZC3HC1, and IGF2R. We also validate previous findings near 5q33.3/EBF1 and FOXO3, whilst finding contradictory evidence at other loci. Gene set and cell-specific analyses show that expression in foetal brain cells and adult dorsolateral prefrontal cortex is enriched for lifespan variation, as are gene pathways involving lipid proteins and homeostasis, vesicle-mediated transport, and synaptic function. Individual genetic variants that increase dementia, cardiovascular disease, and lung cancer - but not other cancers - explain the most variance. Resulting polygenic scores show a mean lifespan difference of around five years of life across the deciles. Editorial note: This article has been through an editorial process in which the authors decide how to respond to the issues raised during peer review. The Reviewing Editor's assessment is that all the issues have been addressed (see decision letter). AU - Timmers, P.R.* AU - Mounier, N.* AU - Lall, K.* AU - Fischer, K.* AU - Ning, Z.* AU - Feng, X.* AU - Bretherick, A.D.* AU - Clark, D.W.* AU - eQTLGen Consortium (Agbessi, M.* AU - Ahsan, H.* AU - Alves, I.* AU - Andiappan, A.* AU - Awadalla, P.* AU - Battle, A.* AU - Bonder, M.J.* AU - Boomsma, D.* AU - Christiansen, M.* AU - Claringbould, A.* AU - Deelen, P.* AU - van Dongen, J.* AU - Esko, T.* AU - Favé, M.* AU - Franke, L.* AU - Frayling, T.* AU - Gharib, S.A.* AU - Gibson, G.* AU - Hemani, G.* AU - Jansen, R.* AU - Kalnapenkis, A.* AU - Kasela, S.* AU - Kettunen, J.* AU - Kim, Y.* AU - Kirsten, H.* AU - Kovacs, P.* AU - Krohn, K.* AU - Kronberg-Guzman, J.* AU - Kukushkina, V.* AU - Kutalik, Z.* AU - Kähönen, M.* AU - Lee, B.* AU - Lehtimäki, T.* AU - Loeffler, M.* AU - Marigorta, U.* AU - Metspalu, A.* AU - van Meurs, J.* AU - Milani, L.* AU - Müller-Nurasyid, M. AU - Nauck, M.* AU - Nivard, M.* AU - Penninx, B.* AU - Perola, M.* AU - Pervjakova, N.* AU - Pierce, B.* AU - Powell, J.* AU - Prokisch, H. AU - Psaty, B.M.* AU - Raitakari, O.* AU - Ring, S.* AU - Ripatti, S.* AU - Rotzschke, O.* AU - Ruëger, S.* AU - Saha, A.* AU - Scholz, M.* AU - Schramm, K. AU - Seppälä, I.* AU - Stumvoll, M.* AU - Sullivan, P.* AU - Teumer, A.* AU - Thiery, J.* AU - Tong, L.* AU - Tönjes, A.* AU - Verlouw, J.* AU - Visscher, P.M.* AU - Võsa, U.* AU - Völker, U.* AU - Yaghootkar, H.* AU - Yang, J.* AU - Zeng, B.* AU - Zhang, F.*) AU - Shen, X.* AU - Esko, T.* AU - Kutalik, Z.* AU - Wilson, J.F.* AU - Joshi, P.K.* C1 - 55165 C2 - 46333 TI - Genomics of 1 million parent lifespans implicates novel pathways and common diseases and distinguishes survival chances. JO - eLife VL - 8 PY - 2019 SN - 2050-084X ER - TY - JOUR AB - The transcription factor c-Myc amplifies the transcription of many growth-related genes in cancer cells, but its role as an oncogene is not fully understood. AU - Eick, D. C1 - 52739 C2 - 44297 CY - Cambridge TI - Getting to grips with c-Myc. JO - eLife VL - 7 PB - Elife Sciences Publications Ltd PY - 2018 SN - 2050-084X ER - TY - JOUR AB - The pancreatic islet, a cellular community harboring the insulin-producing beta-cells, is known to undergo age-related alterations. However, only a handful of signals associated with aging have been identified. By comparing beta-cells from younger and older zebrafish, here we show that the aging islets exhibit signs of chronic inflammation. These include recruitment of -expressing macrophages and the activation of NF-kB signaling in beta-cells. Using a transgenic reporter, we show that NF-kB activity is undetectable in juvenile beta-cells, whereas cells from older fish exhibit heterogeneous NF-kB activity. We link this heterogeneity to differences in gene expression and proliferation. Beta-cells with high NF-kB signaling proliferate significantly less compared to their neighbors with low activity. The NF-kB signaling cells also exhibit premature upregulation of , an age-related gene that inhibits beta-cell proliferation. Together, our results show that NF-kB activity marks the asynchronous decline in beta-cell proliferation with advancing age. AU - Janjuha, S. AU - Singh, S.P.* AU - Tsakmaki, A.* AU - Mousavy Gharavy, S.N.* AU - Murawala, P.* AU - Konantz, J.* AU - Birke, S.* AU - Hodson, D.J.* AU - Rutter, G.A.* AU - Bewick, G.A.* AU - Ninov, N. C1 - 53490 C2 - 44592 TI - Age-related islet inflammation marks the proliferative decline of pancreatic beta-cells in zebrafish. JO - eLife VL - 7 PY - 2018 SN - 2050-084X ER - TY - JOUR AB - The P2X7 channel is involved in the pathogenesis of various CNS diseases. An increasing number of studies suggest its presence in neurons where its putative functions remain controversial for more than a decade. To resolve this issue and to provide a model for analysis of P2X7 functions, we generated P2X7 BAC transgenic mice that allow visualization of functional EGFP-tagged P2X7 receptors in vivo. Extensive characterization of these mice revealed dominant P2X7-EGFP protein expression in microglia, Bergmann glia, and oligodendrocytes, but not in neurons. These findings were further validated by microglia- and oligodendrocyte-specific P2X7 deletion and a novel P2X7-specific nanobody. In addition to the first quantitative analysis of P2X7 protein expression in the CNS, we show potential consequences of its overexpression in ischemic retina and post-traumatic cerebral cortex grey matter. This novel mouse model overcomes previous limitations in P2X7 research and will help to determine its physiological roles and contribution to diseases. AU - Kaczmarek-Hajek, K.* AU - Zhang, J.* AU - Kopp, R.* AU - Grosche, A.* AU - Rissiek, B.* AU - Saul, A.* AU - Bruzzone, S.* AU - Engel, T.* AU - Jooss, T.* AU - Krautloher, A.* AU - Schuster, S.* AU - Magnus, T.* AU - Stadelmann, C.* AU - Sirko, S. AU - Koch-Nolte, F.* AU - Eulenburg, V.* AU - Nicke, A.* C1 - 54415 C2 - 45529 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - Re-evaluation of neuronal P2X7 expression using novel mouse models and a P2X7-specific nanobody. JO - eLife VL - 7 PB - Elife Sciences Publications Ltd PY - 2018 SN - 2050-084X ER - TY - JOUR AB - Oligomeric assemblies of intraflagellar transport (IFT) particles build cilia through sequential recruitment and transport of ciliary cargo proteins within cilia. Here we present the 1.8 Å resolution crystal structure of the Chlamydomonas IFT-B protein IFT80, which reveals the architecture of two N-terminal β-propellers followed by an α-helical extension. The N-terminal β-propeller tethers IFT80 to the IFT-B complex via IFT38 whereas the second β-propeller and the C-terminal αa-helical extension result in IFT80 homo-dimerization. Using CRISPR/Cas to create biallelic Ift80 frameshift mutations in IMCD3 mouse cells, we demonstrate that IFT80 is absolutely required for ciliogenesis. Structural mapping and rescue experiments reveal that human disease-causing missense mutations do not cluster within IFT80 and form functional IFT particles. Unlike missense mutant forms of IFT80, deletion of the C-terminal dimerization domain prevented rescue of ciliogenesis. Taken together our results may provide a first insight into higher order IFT complex formation likely required for IFT train formation. AU - Täschner, M.* AU - Lorentzen, A.* AU - Mourao, A. AU - Collins, T.* AU - Freke, G.M.* AU - Moulding, D.* AU - Basquin, J.* AU - Jenkins, D.* AU - Lorentzen, E.* C1 - 53409 C2 - 44681 TI - Crystal structure of intraflagellar transport protein 80 reveals a homodimer required for ciliogenesis. JO - eLife VL - 7 PY - 2018 SN - 2050-084X ER - TY - JOUR AB - Blood is arguably the most important bodily fluid and its analysis provides crucial health status information. A first routine measure to narrow down diagnosis in clinical practice is the differential blood count, determining the frequency of all major blood cells. What is lacking to advance initial blood diagnostics is an unbiased and quick functional assessment of blood that can narrow down the diagnosis and generate specific hypotheses. To address this need, we introduce the continuous, cell-by-cell morpho-rheological (MORE) analysis of diluted whole blood, without labeling, enrichment or separation, at rates of 1000 cells/sec. In a drop of blood we can identify all major blood cells and characterize their pathological changes in several disease conditions in vitro and in patient samples. This approach takes previous results of mechanical studies on specifically isolated blood cells to the level of application directly in blood and adds a functional dimension to conventional blood analysis. AU - Toepfner, N.* AU - Herold, C.* AU - Otto, O.* AU - Rosendahl, P.* AU - Jacobi, A.* AU - Kraeter, M.* AU - Staechele, J.* AU - Menschner, L.* AU - Herbig, M.* AU - Ciuffreda, L.* AU - Ford-Cartwright, L.* AU - Grzybek, M. AU - Coskun, Ü. AU - Reithuber, E.* AU - Garriss, G.* AU - Mellroth, P.* AU - Henriques-Normark, B.* AU - Tregay, N.* AU - Suttorp, M.* AU - Bornhaeuser, M.* AU - Chilvers, E.R.* AU - Berner, R.* AU - Guck, J.* C1 - 52900 C2 - 44311 CY - Cambridge TI - Detection of human disease conditions by single-cell morpho-rheological phenotyping of blood. JO - eLife VL - 7 PB - Elife Sciences Publications Ltd PY - 2018 SN - 2050-084X ER - TY - JOUR AB - Despite the intrinsically stochastic nature of damage, sensory organs recapitulate normal architecture during repair to maintain function. Here we present a quantitative approach that combines live cell-lineage tracing and multifactorial classification by machine learning to reveal how cell identity and localization are coordinated during organ regeneration. We use the superficial neuromasts in larval zebrafish, which contain three cell classes organized in radial symmetry and a single planar-polarity axis. Visualization of cell-fate transitions at high temporal resolution shows that neuromasts regenerate isotropically to recover geometric order, proportions and polarity with exceptional accuracy. We identify mediolateral position within the growing tissue as the best predictor of cell-fate acquisition. We propose a self-regulatory mechanism that guides the regenerative process to identical outcome with minimal extrinsic information. The integrated approach that we have developed is simple and broadly applicable, and should help define predictive signatures of cellular behavior during the construction of complex tissues. AU - Viader Llargues, O. AU - Lupperger, V. AU - Pola-Morell, L. AU - Marr, C. AU - López-Schier, H. C1 - 53336 C2 - 44830 CY - Sheraton House, Castle Park, Cambridge, Cb3 0ax, England TI - Live cell-lineage tracing and machine learning reveal patterns of organ regeneration. JO - eLife VL - 7 PB - Elife Sciences Publications Ltd PY - 2018 SN - 2050-084X ER - TY - JOUR AB - Gene transcription can be activated by decreasing the duration of RNA polymerase II pausing in the promoter-proximal region, but how this is achieved remains unclear. Here we use a 'multi-omics' approach to demonstrate that the duration of polymerase pausing generally limits the productive frequency of transcription initiation in human cells ('pause-initiation limit'). We further engineer a human cell line to allow for specific and rapid inhibition of the P-TEFb kinase CDK9, which is implicated in polymerase pause release. CDK9 activity decreases the pause duration but also increases the productive initiation frequency. This shows that CDK9 stimulates release of paused polymerase and activates transcription by increasing the number of transcribing polymerases and thus the amount of mRNA synthesized per time. CDK9 activity is also associated with long-range chromatin interactions, suggesting that enhancers can influence the pause-initiation limit to regulate transcription. AU - Gressel, S.* AU - Schwalb, B.* AU - Decker, T.-M. AU - Qin, W.* AU - Leonhardt, H.* AU - Eick, D. AU - Cramer, P.* C1 - 52067 C2 - 43711 CY - Cambridge TI - CDK9-dependent RNA polymerase II pausing controls transcription initiation. JO - eLife VL - 6 PB - Elife Sciences Publications Ltd PY - 2017 SN - 2050-084X ER - TY - JOUR AB - The evolutionary origins of the hypoxia-sensitive cells that trigger amniote respiratory reflexes - carotid body glomus cells, and 'pulmonary neuroendocrine cells' (PNECs) - are obscure. Homology has been proposed between glomus cells, which are neural crest-derived, and the hypoxia-sensitive 'neuroepithelial cells' (NECs) of fish gills, whose embryonic origin is unknown. NECs have also been likened to PNECs, which differentiate in situ within lung airway epithelia. Using genetic lineage-tracing and neural crest-deficient mutants in zebrafish, and physical fate-mapping in frog and lamprey, we find that NECs are not neural crest-derived, but endoderm-derived, like PNECs, whose endodermal origin we confirm. We discover neural crest-derived catecholaminergic cells associated with zebrafish pharyngeal arch blood vessels, and propose a new model for amniote hypoxia-sensitive cell evolution: endoderm-derived NECs were retained as PNECs, while the carotid body evolved via the aggregation of neural crest-derived catecholaminergic (chromaffin) cells already associated with blood vessels in anamniote pharyngeal arches. AU - Hockman, D.* AU - Burns, A.* AU - Schlosser, G.* AU - Gates, K.P.* AU - Jevans, B.* AU - Mongera, A.* AU - Fisher, S.* AU - Unlu, G.* AU - Knapik, E.W.* AU - Kaufman, C.K.* AU - Mosimann, C.* AU - Zon, L.I.* AU - Lancman, J.J.* AU - Dong, P.D.* AU - Lickert, H. AU - Tucker, A.S.* AU - Baker, C.V.* C1 - 50905 C2 - 42995 TI - Evolution of the hypoxia-sensitive cells involved in amniote respiratory reflexes. JO - eLife VL - 6 PY - 2017 SN - 2050-084X ER - TY - JOUR AB - Cool ambient temperatures are major cues determining flowering time in spring. The mechanisms promoting or delaying flowering in response to ambient temperature changes are only beginning to be understood. In Arabidopsis thaliana, FLOWERING LOCUS M (FLM) regulates flowering in the ambient temperature range and FLM is transcribed and alternatively spliced in a temperature-dependent manner. We identify polymorphic promoter and intronic sequences required for FLM expression and splicing. In transgenic experiments covering 69% of the available sequence variation in two distinct sites, we show that variation in the abundance of the FLM-ß splice form strictly correlate (R2 = 0.94) with flowering time over an extended vegetative period. The FLM polymorphisms lead to changes in FLM expression (PRO2+) but may also affect FLM intron 1 splicing (INT6+). This information could serve to buffer the anticipated negative effects on agricultural systems and flowering that may occur during climate change. AU - Lutz, U.* AU - Nussbaumer, T. AU - Spannagl, M. AU - Diener, J.* AU - Mayer, K.F.X. AU - Schwechheimer, C.* C1 - 50715 C2 - 42869 CY - Cambridge TI - Natural haplotypes of FLM non-coding sequences fine-tune flowering time in ambient spring temperatures in Arabidopsis. JO - eLife VL - 6 PB - Elife Sciences Publications Ltd PY - 2017 SN - 2050-084X ER - TY - JOUR AB - The recent advent of methods for high-throughput single-cell molecular profiling has catalyzed a growing sense in the scientific community that the time is ripe to complete the 150-year-old effort to identify all cell types in the human body. The Human Cell Atlas Project is an international collaborative effort that aims to define all human cell types in terms of distinctive molecular profiles (such as gene expression profiles) and to connect this information with classical cellular descriptions (such as location and morphology). An open comprehensive reference map of the molecular state of cells in healthy human tissues would propel the systematic study of physiological states, developmental trajectories, regulatory circuitry and interactions of cells, and also provide a framework for understanding cellular dysregulation in human disease. Here we describe the idea, its potential utility, early proofs-of-concept, and some design considerations for the Human Cell Atlas, including a commitment to open data, code, and community. AU - Regev, A.* AU - Teichmann, S.A.* AU - Lander, E.S.* AU - Amit, I.* AU - Benoist, C.* AU - Birney, E.* AU - Bodenmiller, B.* AU - Campbell, P.J.* AU - Carninci, P.* AU - Clatworthy, M.* AU - Clevers, H.* AU - Deplancke, B.* AU - Dunham, I.* AU - Eberwine, J.* AU - Eils, R.* AU - Enard, W.* AU - Farmer, A.* AU - Fugger, L.* AU - Göttgens, B.* AU - Hacohen, N.* AU - Haniffa, M.* AU - Hemberg, M.* AU - Kim, S.K.* AU - Klenerman, P.* AU - Kriegstein, A.* AU - Lein, E.* AU - Linnarsson, S.* AU - Lundberg, E.* AU - Lundeberg, J.* AU - Majumder, P.* AU - Marioni, J.C.* AU - Merad, M.* AU - Mhlanga, M.* AU - Nawijn, M.C.* AU - Netea, M.* AU - Nolan, G.* AU - Pe'er, D.* AU - Phillipakis, A.* AU - Ponting, C.P.* AU - Quake, S.R.* AU - Reik, W.* AU - Rozenblatt-Rosen, O.* AU - Sanes, J.R.* AU - Satija, R.* AU - Schumacher, T.N.* AU - Shalek, A.K.* AU - Shapiro, E.* AU - Sharma, P.* AU - Shin, J.W.* AU - Stegle, O.* AU - Stratton, M.R.* AU - Stubbington, M.J.T.* AU - Theis, F.J. AU - Uhlén, M.* AU - van Oudenaarden, A.* AU - Wagner, A.* AU - Watt, F.M.* AU - Weissman, J.S.* AU - Wold, B.J.* AU - Xavier, R.J.* AU - Yosef, N.* C1 - 52478 C2 - 44000 CY - Cambridge TI - The Human Cell Atlas. JO - eLife VL - 6 PB - Elife Sciences Publications Ltd PY - 2017 SN - 2050-084X ER - TY - JOUR AB - Membrane-assisted amyloid formation is implicated in human diseases, and many of the aggregating species accelerate amyloid formation and induce cell death. While structures of membrane-associated intermediates would provide tremendous insights into the pathology and aid in the design of compounds to potentially treat the diseases, it has not been feasible to overcome the challenges posed by the cell membrane. Here we use NMR experimental constraints to solve the structure of a type-2 diabetes related human islet amyloid polypeptide intermediate stabilized in nanodiscs. ROSETTA and MD simulations resulted in a unique b-strand structure distinct from the conventional amyloid b-hairpin and revealed that the nucleating NFGAIL region remains flexible and accessible within this isolated intermediate, suggesting a mechanism by which membrane-associated aggregation may be propagated. The ability of nanodiscs to trap amyloid intermediates as demonstrated could become one of the most powerful approaches to dissect the complicated misfolding pathways of protein aggregation. AU - Rodriguez Camargo, D.C.* AU - Korshavn, K.J.* AU - Jussupow, A.* AU - Raltchev, K.* AU - Goricanec, D.* AU - Fleisch, M. AU - Sarkar, R.* AU - Xue, K. AU - Aichler, M. AU - Mettenleiter, G. AU - Walch, A.K. AU - Camilloni, C.* AU - Hagn, F. AU - Reif, B. AU - Ramamoorthy, A.* C1 - 52364 C2 - 43917 CY - Cambridge TI - Stabilization and structural analysis of a membrane-associated hIAPP aggregation intermediate. JO - eLife VL - 6 PB - Elife Sciences Publications Ltd PY - 2017 SN - 2050-084X ER - TY - JOUR AB - The ubiquitin ligase TRAF6 is a key regulator of canonical IκB kinase (IKK)/NF-κB signaling in response to interleukin-1 (IL-1) stimulation. Here, we identified the deubiquitinating enzyme YOD1 (OTUD2) as a novel interactor of TRAF6 in human cells. YOD1 binds to the C-terminal TRAF homology domain of TRAF6 that also serves as the interaction surface for the adaptor p62/Sequestosome-1, which is required for IL-1 signaling to NF-κB. We show that YOD1 competes with p62 for TRAF6 association and abolishes the sequestration of TRAF6 to cytosolic p62 aggregates by a non-catalytic mechanism. YOD1 associates with TRAF6 in unstimulated cells but is released upon IL-1β stimulation, thereby facilitating TRAF6 auto-ubiquitination as well as NEMO/IKKγ substrate ubiquitination. Further, IL-1 triggered IKK/NF-κB signaling and induction of target genes is decreased by YOD1 overexpression and augmented after YOD1 depletion. Hence, our data define that YOD1 antagonizes TRAF6/p62-dependent IL-1 signaling to NF-κB. AU - Schimmack, G. AU - Schorpp, K.K. AU - Kutzner, K. AU - Gehring, T. AU - Brenke, J.K. AU - Hadian, K. AU - Krappmann, D. C1 - 50631 C2 - 42536 CY - Cambridge TI - YOD1/TRAF6 association balances p62-dependent IL-1 signaling to NF-κB. JO - eLife VL - 6 PB - Elife Sciences Publications Ltd PY - 2017 SN - 2050-084X ER - TY - JOUR AB - FGF2 is secreted from cells by an unconventional secretory pathway. This process is mediated by direct translocation across the plasma membrane. Here, we define the minimal molecular machinery required for FGF2 membrane translocation in a fully reconstituted inside-out vesicle system. FGF2 membrane translocation is thermodynamically driven by PI(4,5)P2-induced membrane insertion of FGF2 oligomers. The latter serve as dynamic translocation intermediates of FGF2 with a subunit number in the range of 8-12 FGF2 molecules. Vectorial translocation of FGF2 across the membrane is governed by sequential and mutually exclusive interactions with PI(4,5)P2 and heparan sulfates on opposing sides of the membrane. Based on atomistic molecular dynamics simulations, we propose a mechanism that drives PI(4,5)P2 dependent oligomerization of FGF2. Our combined findings establish a novel type of self-sustained protein translocation across membranes revealing the molecular basis of the unconventional secretory pathway of FGF2. AU - Steringer, J.P.* AU - Lange, S.* AU - Čujová, S.* AU - Šachl, R.* AU - Poojari, C.* AU - Lolicato, F.* AU - Beutel, O.* AU - Müller, H.M.* AU - Unger, S.* AU - Coskun, Ü. AU - Honigmann, A.* AU - Vattulainen, I.* AU - Hof, M.* AU - Freund, C.* AU - Nickel, W.* C1 - 52116 C2 - 43705 TI - Key steps in unconventional secretion of fibroblast growth factor 2 reconstituted with purified components. JO - eLife VL - 6 PY - 2017 SN - 2050-084X ER - TY - JOUR AB - Glioblastoma multiforme (GBM) is the most aggressive human primary brain cancer. Using a Trp53-deficient mouse model of GBM, we show that genetic inactivation of the Atm cofactor Atmin, which is dispensable for embryonic and adult neural development, strongly suppresses GBM formation. Mechanistically, expression of several GBM-associated genes, including Pdgfra, was normalized by Atmin deletion in the Trp53-null background. Pharmacological ATM inhibition also reduced Pdgfra expression, and reduced the proliferation of Trp53-deficient primary glioma cells from murine and human tumors, while normal neural stem cells were unaffected. Analysis of GBM datasets showed that PDGFRA expression is also significantly increased in human TP53-mutant compared with TP53-wild-type tumors. Moreover, combined treatment with ATM and PDGFRA inhibitors efficiently killed TP53-mutant primary human GBM cells, but not untransformed neural stem cells. These results reveal a new requirement for ATMIN-dependent ATM signaling in TP53-deficient GBM, indicating a pro-tumorigenic role for ATM in the context of these tumors. AU - Blake, S.M.* AU - Stricker, S.H. AU - Halavach, H.* AU - Poetsch, A.R.* AU - Cresswell, G.* AU - Kelly, G.* AU - Kanu, N.* AU - Marino, S.* AU - Luscombe, N.M.* AU - Pollard, S.M.* AU - Behrens, A.* C1 - 48378 C2 - 41620 CY - Cambridge TI - Inactivation of the ATMIN/ATM pathway protects against glioblastoma formation. JO - eLife VL - 5 PB - Elife Sciences Publications Ltd PY - 2016 SN - 2050-084X ER - TY - JOUR AB - Mg2+ regulates many physiological processes and signalling pathways. However, little is known about the mechanisms underlying the organismal balance of Mg2+. Capitalizing on a set of newly generated mouse models, we provide an integrated mechanistic model of the regulation of organismal Mg2+ balance during prenatal development and in adult mice by the ion channel TRPM6. We show that TRPM6 activity in the placenta and yolk sac is essential for embryonic development. In adult mice, TRPM6 is required in the intestine to maintain organismal Mg2+ balance, but is dispensable in the kidney. Trpm6 inactivation in adult mice leads to a shortened lifespan, growth deficit and metabolic alterations indicative of impaired energy balance. Dietary Mg2+ supplementation not only rescues all phenotypes displayed by Trpm6-deficient adult mice, but also may extend the lifespan of wildtype mice. Hence, maintenance of organismal Mg2+ balance by TRPM6 is crucial for prenatal development and survival to adulthood. AU - Chubanov, V.* AU - Ferioli, S.* AU - Wisnowsky, A.* AU - Simmons, D.G.* AU - Leitzinger, C. AU - Einer, C. AU - Jonas, W.* AU - Shymkiv, Y.* AU - Bartsch, H.* AU - Braun, A.* AU - Akdogan, B.* AU - Mittermeier, L.* AU - Sytik, L.* AU - Torben, F. AU - Jurinovic, V.* AU - van der Vorst, E.P.C.* AU - Weber, C.* AU - Yildirim, A.Ö. AU - Sotlar, K.* AU - Schürmann, A.* AU - Zierler, S.* AU - Zischka, H. AU - Ryazanov, A.G.* AU - Gudermann, T.* C1 - 50347 C2 - 42268 CY - Cambridge TI - Epithelial magnesium transport by TRPM6 is essential for prenatal development and adult survival. JO - eLife VL - 5 PB - Elife Sciences Publications Ltd PY - 2016 SN - 2050-084X ER - TY - JOUR AB - Being taller is associated with enhanced longevity, and higher education and earnings. We reanalysed 1472 population-based studies, with measurement of height on more than 18.6 million participants to estimate mean height for people born between 1896 and 1996 in 200 countries. The largest gain in adult height over the past century has occurred in South Korean women and Iranian men, who became 20.2 cm (95% credible interval 17.5-22.7) and 16.5 cm (13.3- 19.7) taller, respectively. In contrast, there was little change in adult height in some sub-Saharan African countries and in South Asia over the century of analysis. The tallest people over these 100 years are men born in the Netherlands in the last quarter of 20th century, whose average heights surpassed 182.5 cm, and the shortest were women born in Guatemala in 1896 (140.3 cm; 135.8- 144.8). The height differential between the tallest and shortest populations was 19-20 cm a century ago, and has remained the same for women and increased for men a century later despite substantial changes in the ranking of countries. AU - NCD Risk Factors Collaboration (Döring, A. AU - Meisinger, C. AU - Müller-Nurasyid, M. AU - Stieber, J. AU - Stöckl, D.) C1 - 49219 C2 - 33300 TI - A century of trends in adult human height. JO - eLife VL - 5 PY - 2016 SN - 2050-084X ER - TY - JOUR AB - Metzincin metalloproteases have major roles in intercellular communication by modulating the function of membrane proteins. One of the proteases is the a-disintegrin-and-metalloprotease 10 (ADAM10) which acts as alpha-secretase of the Alzheimer's disease amyloid precursor protein. ADAM10 is also required for neuronal network functions in murine brain, but neuronal ADAM10 substrates are only partly known. With a proteomic analysis of Adam10-deficient neurons we identified 91, mostly novel ADAM10 substrate candidates, making ADAM10 a major protease for membrane proteins in the nervous system. Several novel substrates, including the neuronal cell adhesion protein NrCAM, are involved in brain development. Indeed, we detected mistargeted axons in the olfactory bulb of conditional ADAM10-/- mice, which correlate with reduced cleavage of NrCAM, NCAM and other ADAM10 substrates. In summary, the novel ADAM10 substrates provide a molecular basis for neuronal network dysfunctions in conditional ADAM10-/- mice and demonstrate a fundamental function of ADAM10 in the brain. AU - Kuhn, P.H.* AU - Colombo, A.V.* AU - Schusser, B.* AU - Dreymueller, D.* AU - Wetzel, S.* AU - Schepers, U.* AU - Herber, J.* AU - Ludwig, A.* AU - Kremmer, E. AU - Montag, D.* AU - Müller, U.* AU - Schweizer, M.* AU - Saftig, P.* AU - Bräse, S.* AU - Lichtenthaler, S.F.* C1 - 47737 C2 - 39492 TI - Systematic substrate identification indicates a central role for the metalloprotease ADAM10 in axon targeting and synapse function. JO - eLife VL - 5 PY - 2016 SN - 2050-084X ER - TY - JOUR AB - The multi-domain splicing factor RBM5 regulates the balance between antagonistic isoforms of the apoptosis-control genes FAS/CD95, Caspase-2 and AID. An OCRE (OCtamer REpeat of aromatic residues) domain found in RBM5 is important for alternative splicing regulation and mediates interactions with components of the U4/U6.U5 tri-snRNP. We show that the RBM5 OCRE domain adopts a unique b–sheet fold. NMR and biochemical experiments demonstrate that the OCRE domain directly binds to the proline-rich C-terminal tail of the essential snRNP core proteins SmN/B/B’. The NMR structure of an OCRE-SmN peptide complex reveals a specific recognition of poly-proline helical motifs in SmN/B/B’. Mutation of conserved aromatic residues impairs binding to the Sm proteins in vitro and compromises RBM5-mediated alternative splicing regulation of FAS/CD95. Thus, RBM5 OCRE represents a poly-proline recognition domain that mediates critical interactions with the C-terminal tail of the spliceosomal SmN/B/B’ proteins in FAS/ CD95 alternative splicing regulation. AU - Mourao, A. AU - Bonnal, S.* AU - Soni, K. AU - Warner, L. AU - Bordonné, R.* AU - Valcárcel, J.* AU - Sattler, M. C1 - 50137 C2 - 42063 CY - Cambridge TI - Structural basis for the recognition of spliceosomal SmN/B/B’ proteins by the RBM5 OCRE domain in splicing regulation. JO - eLife VL - 5 PB - Elife Sciences Publications Ltd PY - 2016 SN - 2050-084X ER - TY - JOUR AB - The timely transition from neural progenitor to post-mitotic neuron requires down-regulation and loss of the neuronal transcriptional repressor, REST. Here, we have used mice containing a gene trap in the Rest gene, eliminating transcription from all coding exons, to remove REST prematurely from neural progenitors. We find that catastrophic DNA damage occurs during S-phase of the cell cycle, with long-term consequences including abnormal chromosome separation, apoptosis, and smaller brains. Persistent effects are evident by latent appearance of proneural glioblastoma in adult mice deleted additionally for the tumor suppressor p53 protein (p53). A previous line of mice deleted for REST in progenitors by conventional gene targeting does not exhibit these phenotypes, likely due to a remaining C-terminal peptide that still binds chromatin and recruits co-repressors. Our results suggest that REST-mediated chromatin remodeling is required in neural progenitors for proper S-phase dynamics, as part of its well-established role in repressing neuronal genes until terminal differentiation. AU - Nechiporuk, T.* AU - McGann, J.* AU - Mullendorff, K.* AU - Hsieh, J.* AU - Wurst, W. AU - Floß, T. AU - Mandel, G.* C1 - 47659 C2 - 39278 TI - The REST remodeling complex protects genomic integrity during embryonic neurogenesis. JO - eLife VL - 5 PY - 2016 SN - 2050-084X ER - TY - JOUR AB - Due to their economic relevance, the study of plant pathogen interactions is of importance. However, elucidating these interactions and their underlying molecular mechanisms remains challenging since both host and pathogen need to be fully genetically accessible organisms. Here we present milestones in the establishment of a new biotrophic model pathosystem: Ustilago bromivora and Brachypodium sp. We provide a complete toolset, including an annotated fungal genome and methods for genetic manipulation of the fungus and its host plant. This toolset will enable researchers to easily study biotrophic interactions at the molecular level on both the pathogen and the host side. Moreover, our research on the fungal life cycle revealed a mating type bias phenomenon. U. bromivora harbors a haplo-lethal allele that is linked to one mating type region. As a result, the identified mating type bias strongly promotes inbreeding, which we consider to be a potential speciation driver. AU - Rabe, F.* AU - Bosch, J.A.* AU - Stirnberg, A.* AU - Guse, T.* AU - Bauer, L.* AU - Seitner, D.* AU - Rabanal, F.A.* AU - Czedik-Eysenberg, A.* AU - Uhse, S.* AU - Bindics, J.* AU - Genenncher, B.* AU - Navarrete, F.* AU - Kellner, R.* AU - Ekker, H.* AU - Kumlehn, J.* AU - Vogel, J.P.* AU - Gordon, S.P.* AU - Marcel, T.C.* AU - Münsterkötter, M. AU - Walter, M.* AU - Sieber, C.M.K. AU - Mannhaupt, G. AU - Güldener, U. AU - Kahmann, R.* AU - Djamei, A.* C1 - 50037 C2 - 41984 CY - Cambridge TI - A complete toolset for the study of Ustilago bromivora and Brachypodium sp. as a fungal-temperate grass pathosystem. JO - eLife VL - 5 PB - Elife Sciences Publications Ltd PY - 2016 SN - 2050-084X ER - TY - JOUR AB - The neuronal DNA-/RNA-binding protein Pur-alpha is a transcription regulator and core factor for mRNA-localization. Pur-alpha deficient mice die after birth with pleiotropic neuronal defects. Here we report the crystal structure of the DNA-/RNA-binding domain of Pur-alpha in complex with ssDNA. It reveals base-specific recognition and offers a molecular explanation for the effect of point mutations in the 5q31.3 microdeletion syndrome. Consistent with the crystal structure, biochemical and NMR data indicate that Pur-alpha binds DNA and RNA in the same way, suggesting binding modes for tri- and hexanucleotide repeat RNAs in two neurodegenerative RNAopathies. Additionally, structure-based in vitro experiments resolved the molecular mechanism of Pur-alpha's unwindase activity. Complementing in vivo analyses in Drosophila demonstrated the importance of a highly conserved phenylalanine for Pur-alpha's unwinding and neuroprotective function. By uncovering the molecular mechanisms of nucleic-acid binding, this study contributes to understanding the cellular role of Pur-alpha and its implications in neurodegenerative diseases. AU - Weber, J. AU - Bao, H.* AU - Hartlmüller, C.* AU - Wang, Z.* AU - Windhager, A. AU - Janowski, R. AU - Madl, T. AU - Jin, P.* AU - Niessing, D. C1 - 47660 C2 - 39719 TI - Structural basis of nucleic-acid recognition and double-strand unwinding by the essential neuronal protein Pur-alpha. JO - eLife VL - 5 PY - 2016 SN - 2050-084X ER - TY - JOUR AB - Actin filament dynamics govern many key physiological processes from cell motility to tissue morphogenesis. A central feature of actin dynamics is the capacity of the filament to polymerize and depolymerize at its ends in response to cellular conditions. It is currently thought that filament kinetics can be described by a single rate constant for each end. Here, using direct visualization of single actin filament elongation, we show that actin polymerization kinetics at both filament ends are strongly influenced by proteins that bind to the lateral filament surface. We also show that the less dynamic end, called the pointed-end, has a non-elongating state that dominates the observed filament kinetic asymmetry. Estimates of filament flexibility and Brownian dynamics simulations suggest that the observed kinetic diversity arises from structural alteration. Tuning filament kinetics by exploiting the natural malleability of the actin filament structure may be a ubiquitous mechanism to generate the rich variety of observed cellular actin dynamics. AU - Crevenna, A.H.* AU - Arciniega, M.* AU - Dupont, A.* AU - Mizuno, N.* AU - Kowalska, K.* AU - Lange, O.F. AU - Wedlich-Soeldner, R.* AU - Lamb, D.C.* C1 - 43911 C2 - 36647 CY - Cambridge TI - Side-binding proteins modulate actin filament dynamics. JO - eLife VL - 4 PB - Elife Sciences Publications Ltd PY - 2015 SN - 2050-084X ER - TY - JOUR AB - Calcium in the flagellum controls sperm navigation. In sperm of marine invertebrates and mammals, Ca2(+) signalling has been intensely studied, whereas for fish little is known. In sea urchin sperm, a cyclic nucleotide-gated K(+) channel (CNGK) mediates a cGMP-induced hyperpolarization that evokes Ca2(+) influx. Here, we identify in sperm of the freshwater fish Danio rerio a novel CNGK family member featuring non-canonical properties. It is located in the sperm head rather than the flagellum and is controlled by intracellular pH, but not cyclic nucleotides. Alkalization hyperpolarizes sperm and produces Ca2(+) entry. Ca2(+) induces spinning-like swimming, different from swimming of sperm from other species. The 'spinning' mode probably guides sperm into the micropyle, a narrow entrance on the surface of fish eggs. A picture is emerging of sperm channel orthologues that employ different activation mechanisms and serve different functions. The channel inventories probably reflect adaptations to species-specific challenges during fertilization. AU - Fechner, S.* AU - Alvarez, L.* AU - Bönigk, W.* AU - Müller, A.* AU - Berger, T.* AU - Pascal, R.* AU - Trötschel, C.* AU - Poetsch, A.* AU - Stölting, G.* AU - Siegfried, K.R.* AU - Kremmer, E. AU - Seifert, R.* AU - Kaupp, U.B.* C1 - 47521 C2 - 40642 CY - Cambridge TI - A K+ selective CNG channel orchestrates Ca2+ signalling in zebrafish sperm. JO - eLife VL - 4 PB - Elife Sciences Publications Ltd PY - 2015 SN - 2050-084X ER - TY - JOUR AB - In mammals, the carboxy-terminal domain (CTD) of RNA polymerase (Pol) II consists of 52 conserved heptapeptide repeats containing the consensus sequence Tyr1-Ser2-Pro3-Thr4-Ser5-Pro6-Ser7. Post-translational modifications of the CTD coordinate the transcription cycle and various steps of mRNA maturation. Here we describe Tyr1 phosphorylation (Tyr1P) as a hallmark of promoter (5' associated) Pol II in mammalian cells, in contrast to what was described in yeast. Tyr1P is predominantly found in antisense orientation at promoters but is also specifically enriched at active enhancers. Mutation of Tyr1 to phenylalanine (Y1F) prevents the formation of the hyper-phosphorylated Pol IIO form, induces degradation of Pol II to the truncated Pol IIB form and results in a lethal phenotype. Our results suggest that Tyr1P has evolved specialized and essential functions in higher eukaryotes associated with antisense promoter and enhancer transcription, and Pol II stability. AU - Descostes, N.* AU - Heidemann, M. AU - Spinelli, L.* AU - Schüller, R. AU - Maqbool, M.A.* AU - Fenouil, R.* AU - Koch, F.* AU - Innocenti, C.* AU - Gut, M.* AU - Gut, I.* AU - Eick, D. AU - Andrau, J.C.* C1 - 31323 C2 - 34407 CY - Cambridge TI - Tyrosine phosphorylation of RNA polymerase II CTD is associated with antisense promoter transcription and active enhancers in mammalian cells. JO - eLife VL - 3 PB - Elife Sciences Publications Ltd PY - 2014 SN - 2050-084X ER - TY - JOUR AB - The response of the brain to sugar is determined by specific cell populations in the brain, including neurons that secrete melanin-concentrating hormone, and culminates in the release of dopamine. AU - García-Cáceres, C. AU - Tschöp, M.H. C1 - 29170 C2 - 32729 TI - The emerging neurobiology of calorie addiction. JO - eLife VL - 3 IS - 0 PB - Elife Sciences Publications Ltd PY - 2014 SN - 2050-084X ER - TY - JOUR AB - Planar cell polarity (PCP) regulates basal body (BB) docking and positioning during cilia formation, but the underlying mechanisms remain elusive. Here, we investigate the uncharacterized gene Flattop (Fltp) that is transcriptionally activated during PCP acquisition in ciliated tissues. Fltp knock-out mice show BB docking and ciliogenesis defects in multiciliated lung cells. Furthermore, Fltp is necessary for kinocilium positioning in monociliated inner ear hair cells. In these cells, the core PCP molecule Dishevelled 2, the BB/spindle positioning protein Dlg3 and Fltp localize directly adjacent at the apical plasma membrane, physically interact and surround the BB at the interface of the microtubule and actin cytoskeleton. Dlg3 and Fltp knock-outs suggest that both cooperatively translate PCP cues for BB positioning in the inner ear. Taken together, the identification of novel BB/spindle positioning components as potential mediators of PCP signaling might have broader implications for other cell types, ciliary disease and asymmetric cell division. AU - Gegg, M. AU - Böttcher, A. AU - Burtscher, I. AU - Hasenöder, S. AU - van Campenhout, C.A.* AU - Aichler, M. AU - Walch, A.K. AU - Grant, S.G.* AU - Lickert, H. C1 - 32506 C2 - 35110 TI - Flattop regulates basal body docking and positioning in mono- and multiciliated cells. JO - eLife VL - 3 PY - 2014 SN - 2050-084X ER - TY - JOUR AB - MicroRNAs (miRNAs) are conserved noncoding RNAs that function as posttranscriptional regulators of gene expression. miR-9 is one of the most abundant miRNAs in the brain. Although the function of miR-9 has been well characterized in neural progenitors, its role in dendritic and synaptic development remains largely unknown. In order to target miR-9 in vivo, we developed a transgenic miRNA sponge mouse line allowing conditional inactivation of the miR-9 family in a spatio-temporal-controlled manner. Using this novel approach, we found that miR-9 controls dendritic growth and synaptic transmission in vivo. Furthermore, we demonstrate that miR-9-mediated downregulation of the transcriptional repressor REST is essential for proper dendritic growth. AU - Giusti, S.A.* AU - Vogl, A.M.* AU - Brockmann, M.M.* AU - Vercelli, C.A.* AU - Rein, M.L.* AU - Trümbach, D. AU - Wurst, W. AU - Cazalla, D.* AU - Stein, V.* AU - Deussing, J.M.* AU - Refojo, D.* C1 - 42792 C2 - 35364 TI - MicroRNA-9 controls dendritic development by targeting REST. JO - eLife VL - 3 PY - 2014 SN - 2050-084X ER -