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  • Dynamic post-translational modification profiling of M. tuberculosis-infected primary macrophages
    eLife (IF 7.551) Pub Date : 2020-01-17
    Jonathan M Budzik; Danielle L Swaney; David Jimenez-Morales; Jeffrey R Johnson; Nicholas E Garelis; Teresa Repasy; Allison W Roberts; Lauren M Popov; Trevor J Parry; Dexter Pratt; Trey Ideker; Nevan J Krogan; Jeffery S Cox

    Macrophages are highly plastic cells with critical roles in immunity, cancer, and tissue homeostasis, but how these distinct cellular fates are triggered by environmental cues is poorly understood. To uncover how primary murine macrophages respond to bacterial pathogens, we globally assessed changes in post-translational modifications of proteins during infection with Mycobacterium tuberculosis, a notorious intracellular pathogen. We identified hundreds of dynamically regulated phosphorylation and ubiquitylation sites, indicating that dramatic remodeling of multiple host pathways, both expected and unexpected, occurred during infection. Most of these cellular changes were not captured by mRNA profiling, and included activation of ubiquitin-mediated autophagy, an evolutionarily ancient cellular antimicrobial system. This analysis also revealed that a particular autophagy receptor, TAX1BP1, mediates clearance of ubiquitylated Mtb and targets bacteria to LC3-positive phagophores. These studies provide a new resource for understanding how macrophages shape their proteome to meet the challenge of infection.

    更新日期:2020-01-17
  • Mechanisms underlying the response of mouse cortical networks to optogenetic manipulation
    eLife (IF 7.551) Pub Date : 2020-01-17
    Alexandre Mahrach; Guang Chen; Nuo Li; Carl van Vreeswijk; David Hansel

    GABAergic Interneurons can be subdivided into three subclasses: parvalbumin positive (PV), somatostatin positive (SOM) and serotonin positive neurons. With principal cells (PCs) they form complex networks. We examine PCs and PV responses in mouse anterior lateral motor cortex (ALM) and barrel cortex (S1) upon PV photostimulation in vivo. In ALM layer 5 and S1, the PV response is paradoxical: photoexcitation reduces their activity. This is not the case in ALM layer 2/3. We combine analytical calculations and numerical simulations to investigate how these results constrain the architecture. Two-population models cannot explain the results. Four-population networks with V1-like architecture account for the data in ALM layer 2/3 and layer 5. Our data in S1 can be explained if SOM neurons receive inputs only from PCs and PV neurons. In both four-population models, the paradoxical effect implies not too strong recurrent excitation. It is not evidence for stabilization by inhibition.

    更新日期:2020-01-17
  • Kinesin Kif2C in regulation of DNA double strand break dynamics and repair
    eLife (IF 7.551) Pub Date : 2020-01-17
    Songli Zhu; Mohammadjavad Paydar; Feifei Wang; Yanqiu Li; Ling Wang; Benoit Barrette; Tadayoshi Bessho; Benjamin H Kwok; Aimin Peng

    DNA double strand breaks (DSBs) have detrimental effects on cell survival and genomic stability, and are related to cancer and other human diseases. In this study, we identified microtubule-depolymerizing kinesin Kif2C as a protein associated with DSB-mimicking DNA templates and known DSB repair proteins in Xenopus egg extracts and mammalian cells. The recruitment of Kif2C to DNA damage sites was dependent on both PARP and ATM activities. Kif2C knockdown or knockout led to accumulation of endogenous DNA damage, DNA damage hypersensitivity, and reduced DSB repair via both NHEJ and HR. Interestingly, Kif2C depletion, or inhibition of its microtubule depolymerase activity, reduced the mobility of DSBs, impaired the formation of DNA damage foci, and decreased the occurrence of foci fusion and resolution. Taken together, our study established Kif2C as a new player of the DNA damage response, and presented a new mechanism that governs DSB dynamics and repair.

    更新日期:2020-01-17
  • Principles of self-organization and load adaptation by the actin cytoskeleton during clathrin-mediated endocytosis
    eLife (IF 7.551) Pub Date : 2020-01-17
    Matthew Akamatsu; Ritvik Vasan; Daniel Serwas; Michael Alexander Ferrin; Padmini Rangamani; David G Drubin

    Force generation by actin assembly shapes cellular membranes. An experimentally constrained multiscale model shows that a minimal branched actin network is sufficient to internalize endocytic pits against membrane. Around 200 activated Arp2/3 complexes are required for robust internalization. A newly developed molecule-counting method determined that ~200 Arp2/3 complexes assemble at sites of clathrin-mediated endocytosis in human cells. Simulations predict that actin self-organizes into a radial branched array with growing ends oriented toward the base of the pit. Long actin filaments bend between attachment sites in the coat and the base of the pit. Elastic energy stored in bent filaments, whose presence was confirmed by cryo-electron tomography, contributes to endocytic internalization. Elevated membrane tension directs more growing filaments toward the base of the pit, increasing actin nucleation and bending for increased force production. Thus, spatially constrained actin filament assembly utilizes an adaptive mechanism enabling endocytosis under varying physical constraints.

    更新日期:2020-01-17
  • Structure of the human BBSome core complex
    eLife (IF 7.551) Pub Date : 2020-01-17
    Björn Udo Klink; Christos Gatsogiannis; Oliver Hofnagel; Alfred Wittinghofer; Stefan Raunser

    The BBSome is a heterooctameric protein complex that plays a central role in primary cilia homeostasis. Its malfunction causes the severe ciliopathy Bardet-Biedl syndrome (BBS). The complex acts as a cargo adapter that recognizes signaling proteins such as GPCRs and links them to the intraflagellar transport machinery. The underlying mechanism is poorly understood. Here we present a high-resolution cryo-EM structure of a human heterohexameric core subcomplex of the BBSome. The structure reveals the architecture of the complex in atomic detail. It explains how the subunits interact with each other and how disease-causing mutations hamper this interaction. The complex adopts a conformation that is open for binding to membrane-associated GTPase Arl6 and a large positively charged patch likely strengthens the interaction with the membrane. A prominent negatively charged cleft at the center of the complex is likely involved in binding of positively charged signaling sequences of cargo proteins.

    更新日期:2020-01-17
  • The inner junction complex of the cilia is an interaction hub that involves tubulin post-translational modifications
    eLife (IF 7.551) Pub Date : 2020-01-17
    Ahmad Abdelzaher Zaki Khalifa; Muneyoshi Ichikawa; Daniel Dai; Shintaroh Kubo; Corbin Black; Katya Peri; Thomas S McAlear; Simon Veyron; Shun Kai Yang; Javier Vargas; Susanne Bechstedt; Jean-François Trempe; Khanh Huy Bui

    Microtubules are cytoskeletal structures involved in stability, transport and organization in the cell. The building blocks, the α- and β-tubulin heterodimers, form protofilaments that associate laterally into the hollow microtubule. Microtubule also exists as highly stable doublet microtubules in the cilia where stability is needed for ciliary beating and function. The doublet microtubule maintains its stability through interactions at its inner and outer junctions where its A- and B-tubules meet. Here, using cryo-electron microscopy, bioinformatics and mass spectrometry of the doublets of Chlamydomonas reinhardtii and Tetrahymena thermophila, we identified two new inner junction proteins, FAP276 and FAP106, and an inner junction-associated protein, FAP126, thus presenting the complete answer to the inner junction identity and localization. Our structural study of the doublets shows that the inner junction serves as an interaction hub involved tubulin post-translational modification. These interactions contribute to the stability of the doublet and hence, normal ciliary motility.

    更新日期:2020-01-17
  • Transcriptomics: Revisiting the genomes of herpesviruses
    eLife (IF 7.551) Pub Date : 2020-01-16
    Bhupesh K Prusty; Adam W Whisnant

    Combining integrative genomics and systems biology approaches has revealed new and conserved features in the genome of human herpesvirus 6.

    更新日期:2020-01-17
  • Comprehensive annotations of human herpesvirus 6A and 6B genomes reveal novel and conserved genomic features
    eLife (IF 7.551) Pub Date : 2020-01-16
    Yaara Finkel; Dominik Schmiedel; Julie Tai-Schmiedel; Aharon Nachshon; Roni Winkler; Martina Dobesova; Michal Schwartz; Ofer Mandelboim; Noam Stern-Ginossar

    Human herpesvirus-6 (HHV-6) A and B are ubiquitous betaherpesviruses, infecting the majority of the human population. They encompass large genomes and our understanding of their protein coding potential is far from complete. Here, we employ ribosome-profiling and systematic transcript-analysis to experimentally define HHV-6 translation products. We identify hundreds of new open reading frames (ORFs), including upstream ORFs (uORFs) and internal ORFs (iORFs), generating a complete unbiased atlas of HHV-6 proteome. By integrating systematic data from the prototypic betaherpesvirus, human cytomegalovirus, we uncover numerous uORFs and iORFs conserved across betaherpesviruses and we show uORFs are enriched in late viral genes. We identified three highly abundant HHV-6 encoded long non-coding RNAs, one of which generates a non-polyadenylated stable intron appearing to be a conserved feature of betaherpesviruses. Overall, our work reveals the complexity of HHV-6 genomes and highlights novel features conserved between betaherpesviruses, providing a rich resource for future functional studies.

    更新日期:2020-01-17
  • The Natural History of Model Organisms: The Norway rat, from an obnoxious pest to a laboratory pet
    eLife (IF 7.551) Pub Date : 2020-01-17
    Klaudia Modlinska; Wojciech Pisula

    The laboratory rat was the first mammal domesticated for research purposes. It is descended from wild Norway rats, Rattus norvegicus, which despite their name likely originated in Asia. Exceptionally adaptable, these rodents now inhabit almost all environments on Earth, especially near human settlements where they are often seen as pests. The laboratory rat thrives in captivity, and its domestication has produced many inbred and outbred lines that are used for different purposes, including medical trials and behavioral studies. Differences between wild Norway rats and their laboratory counterparts were first noted in the early 20th century and led some researchers to later question its value as a model organism. While these views are probably unjustified, the advanced domestication of the laboratory rat does suggest that resuming studies of wild rats could benefit the wider research community.

    更新日期:2020-01-17
  • Cortical Development: Do progenitors play dice?
    eLife (IF 7.551) Pub Date : 2020-01-17
    Esther Klingler; Denis Jabaudon

    The wide range of cell types produced by single progenitors in the neocortex of mice may result from stochastic rather than deterministic processes.

    更新日期:2020-01-17
  • Transcriptional adaptation in Caenorhabditis elegans
    eLife (IF 7.551) Pub Date : 2020-01-17
    Vahan Serobyan; Zacharias Kontarakis; Mohamed A El-Brolosy; Jordan M Welker; Oleg Tolstenkov; Amr M Saadeldein; Nicholas Retzer; Alexander Gottschalk; Ann M Wehman; Didier YR Stainier

    Transcriptional adaptation is a recently described phenomenon by which a mutation in one gene leads to the transcriptional modulation of related genes, termed adapting genes. At the molecular level, it has been proposed that the mutant mRNA, rather than the loss of protein function, activates this response. While several examples of transcriptional adaptation have been reported in zebrafish embryos and in mouse cell lines, it is not known whether this phenomenon is observed across metazoans. Here we report transcriptional adaptation in C. elegans, and find that this process requires factors involved in mutant mRNA decay, as in zebrafish and mouse. We further uncover a requirement for Argonaute proteins and Dicer, factors involved in small RNA maturation and transport into the nucleus. Altogether, these results provide evidence for transcriptional adaptation in C. elegans, a powerful model to further investigate underlying molecular mechanisms.

    更新日期:2020-01-17
  • Endothelial heterogeneity across distinct vascular beds during homeostasis and inflammation
    eLife (IF 7.551) Pub Date : 2020-01-16
    Ankit Jambusaria; Zhigang Hong; Lianghui Zhang; Shubhi Srivastava; Arundhati Jana; Peter T Toth; Yang Dai; Asrar B Malik; Jalees Rehman

    Blood vessels are lined by endothelial cells engaged in distinct organ-specific functions but little is known about their characteristic gene expression profiles. RNA-Sequencing of the brain, lung, and heart endothelial translatome identified specific pathways, transporters and cell-surface markers expressed in the endothelium of each organ, which can be visualized at http://www.rehmanlab.org/ribo. We found that endothelial cells express genes typically found in the surrounding tissues such as synaptic vesicle genes in the brain endothelium and cardiac contractile genes in the heart endothelium. Complementary analysis of endothelial single cell RNA-Seq data identified the molecular signature shared across the endothelial translatome and single cell transcriptomes. The tissue-specific heterogeneity of the endothelium is maintained during systemic in vivo inflammatory injury as evidenced by the distinct responses to inflammatory stimulation. Our study defines endothelial heterogeneity and plasticity and provides a molecular framework to understand organ-specific vascular disease mechanisms and therapeutic targeting of individual vascular beds.

    更新日期:2020-01-16
  • Muscle function and homeostasis require cytokine inhibition of AKT activity in Drosophila
    eLife (IF 7.551) Pub Date : 2020-01-16
    Katrin Kierdorf; Fabian Hersperger; Jessica Sharrock; Crystal M Vincent; Pinar Ustaoglu; Jiawen Dou; Attila Gyoergy; Olaf Gross; Daria E Siekhaus; Marc S Dionne

    Unpaired ligands are secreted signals that act via a GP130-like receptor, domeless, to activate JAK/STAT signaling in Drosophila. Like many mammalian cytokines, unpaireds can be activated by infection and other stresses and can promote insulin resistance in target tissues. However, the importance of this effect in non-inflammatory physiology is unknown. Here, we identify a requirement for unpaired-JAK signaling as a metabolic regulator in healthy adult Drosophila muscle. Adult muscles show basal JAK-STAT signaling activity in the absence of any immune challenge. Plasmatocytes (Drosophila macrophages) are an important source of this tonic signal. Loss of the dome receptor on adult muscles significantly reduces lifespan and causes local and systemic metabolic pathology. These pathologies result from hyperactivation of AKT and consequent deregulation of metabolism. Thus, we identify a cytokine signal that must be received in muscle to control AKT activity and metabolic homeostasis.

    更新日期:2020-01-16
  • Phrenic-specific transcriptional programs shape respiratory motor output
    eLife (IF 7.551) Pub Date : 2020-01-16
    Alicia N Vagnozzi; Kiran Garg; Carola Dewitz; Matthew T Moore; Jared M Cregg; Lucie Jeannotte; Niccolò Zampieri; Lynn T Landmesser; Polyxeni Philippidou

    The precise pattern of motor neuron (MN) activation is essential for the execution of motor actions; however, the molecular mechanisms that give rise to specific patterns of MN activity are largely unknown. Phrenic MNs integrate multiple inputs to mediate inspiratory activity during breathing and are constrained to fire in a pattern that drives efficient diaphragm contraction. We show that Hox5 transcription factors shape phrenic MN output by connecting phrenic MNs to inhibitory pre-motor neurons. Hox5 genes establish phrenic MN organization and dendritic topography through the regulation of phrenic-specific cell adhesion programs. In the absence of Hox5 genes, phrenic MN firing becomes asynchronous and erratic due to loss of phrenic MN inhibition. Strikingly, mice lacking Hox5 genes in MNs exhibit abnormal respiratory behavior throughout their lifetime. Our findings support a model where MN-intrinsic transcriptional programs shape the pattern of motor output by orchestrating distinct aspects of MN connectivity.

    更新日期:2020-01-16
  • Gab1 mediates PDGF signaling and is essential to oligodendrocyte differentiation and CNS myelination
    eLife (IF 7.551) Pub Date : 2020-01-16
    Liang Zhou; Chong-Yu Shao; Ya-Jun Xie; Na Wang; Si-Min Xu; Ben-Yan Luo; Zhi-Ying Wu; Yue Hai Ke; Mengsheng Qiu; Ying Shen

    Oligodendrocytes (OLs) myelinate axons and provide electrical insulation and trophic support for neurons in the central nervous system (CNS). Platelet-derived growth factor (PDGF) is critical for steady-state number and differentiation of oligodendrocyte precursor cells (OPCs), but its downstream targets are unclear. Here, we show for the first time that Gab1, an adaptor protein of receptor tyrosine kinase, is specifically expressed in OL lineage cells and is an essential effector of PDGF signaling in OPCs in mice. Gab1 is down-regulated by PDGF stimulation and up-regulated during OPC differentiation. Conditional deletions of Gab1 in OLs cause CNS hypomyelination by affecting OPC differentiation. Moreover, Gab1 binds to downstream GSK3β and regulated its activity, and thereby affects the nuclear accumulation of β-catenin and the expression of a number of transcription factors critical to myelination. Our work uncovers a novel downstream target of PDGF signaling, which is essential to OPC differentiation and CNS myelination.

    更新日期:2020-01-16
  • Structure and activation mechanism of the BBSome membrane protein trafficking complex
    eLife (IF 7.551) Pub Date : 2020-01-15
    Sandeep K Singh; Miao Gui; Fujiet Koh; Matthew CJ Yip; Alan Brown

    Bardet-Biedl syndrome (BBS) is a currently incurable ciliopathy caused by the failure to correctly establish or maintain cilia-dependent signaling pathways. Eight proteins associated with BBS assemble into the BBSome, a key regulator of the ciliary membrane proteome. We report the electron cryomicroscopy (cryo-EM) structures of the native bovine BBSome in inactive and active states at 3.1 ­and 3.5 Å resolution, respectively. In the active state, the BBSome is bound to an Arf-family GTPase (ARL6/BBS3) that recruits the BBSome to ciliary membranes. ARL6 recognizes a composite binding site formed by BBS1 and BBS7 that is occluded in the inactive state. Activation requires an unexpected swiveling of the b-propeller domain of BBS1, the subunit most frequently implicated in substrate recognition, which widens a central cavity of the BBSome. Structural mapping of disease-causing mutations suggests that pathogenesis results from folding defects and the disruption of autoinhibition and activation.

    更新日期:2020-01-15
  • Symptom evolution following the emergence of maize streak virus
    eLife (IF 7.551) Pub Date : 2020-01-15
    Adérito L Monjane; Simon Dellicour; Penelope Hartnady; Kehinde A Oyeniran; Betty E Owor; Marion Bezeidenhout; Daphne Linderme; Rizwan A Syed; Lara Donaldson; Shane Murray; Edward P Rybicki; Anders Kvarnheden; Elhman Yazdkhasti; Pierre Lefeuvre; Rémy Froissart; Philippe Roumagnac; Dionne N Shepherd; Gordon W Harkins; Marc A Suchard; Philippe Lemey; Arvind Varsani; Darren P Martin

    For pathogens infecting single host species evolutionary trade-offs have previously been demonstrated between pathogen-induced mortality rates and transmission rates. It remains unclear, however, how such trade-offs impact sub-lethal pathogen-inflicted damage, and whether these trade-offs even occur in broad host-range pathogens. Here, we examine changes over the past 110 years in symptoms induced in maize by the broad host-range pathogen, maize streak virus (MSV). Specifically, we use the quantified symptom intensities of cloned MSV isolates in differentially resistant maize genotypes to phylogenetically infer ancestral symptom intensities and check for phylogenetic signal associated with these symptom intensities. We show that whereas symptoms reflecting harm to the host have remained constant or decreased, there has been an increase in how extensively MSV colonizes the cells upon which transmission vectors feed. This demonstrates an evolutionary trade-off between amounts of pathogen-inflicted harm and how effectively viruses position themselves within plants to enable onward transmission.

    更新日期:2020-01-15
  • Myosin V executes steps of variable length via structurally constrained diffusion
    eLife (IF 7.551) Pub Date : 2020-01-15
    David Hathcock; Riina Tehver; Michael Hinczewski; Dave Thirumalai

    The molecular motor myosin V transports cargo by stepping on actin filaments, executing a random diffusive search for actin binding sites at each step. A recent experiment suggests that the joint between the myosin lever arms may not rotate freely, as assumed in earlier studies, but instead has a preferred angle giving rise to structurally constrained diffusion. We address this controversy through comprehensive analytical and numerical modeling of myosin V diffusion and stepping. When the joint is constrained, our model reproduces the experimentally observed diffusion, allowing us to estimate bounds on the constraint energy. We also test the consistency between the constrained diffusion model and previous measurements of step size distributions and the load dependence of various observable quantities. The theory lets us address the biological significance of the constrained joint and provides testable predictions of new myosin behaviors, including the stomp distribution and the run length under off-axis force.

    更新日期:2020-01-15
  • A sex difference in the response of the rodent postsynaptic density to synGAP haploinsufficiency
    eLife (IF 7.551) Pub Date : 2020-01-15
    Tara L Mastro; Anthony Preza; Shinjini Basu; Sumantra Chattarji; Sally M Till; Peter C Kind; Mary B Kennedy

    SynGAP is a postsynaptic density (PSD) protein that binds to PDZ domains of the scaffold protein PSD-95. We previously reported that heterozygous deletion of Syngap1 in mice is correlated with increased steady-state levels of other key PSD proteins that bind PSD-95, although the level of PSD-95 remains constant (Walkup et al., 2016). For example, the ratio to PSD-95 of Transmembrane AMPA-Receptor-associated Proteins (TARPs), which mediate binding of AMPA-type glutamate receptors to PSD-95, was increased in young Syngap1+/- mice. Here we show that only females and not males show a highly significant correlation between an increase in TARP and a decrease in synGAP in the PSDs of Syngap1+/- rodents. The data reveal a sex difference in the adaptation of the PSD scaffold to synGAP haploinsufficiency.

    更新日期:2020-01-15
  • Cardiac mitochondrial function depends on BUD23 mediated ribosome programming
    eLife (IF 7.551) Pub Date : 2020-01-15
    Matthew Baxter; Maria Voronkov; Toryn Poolman; Gina Galli; Christian Pinali; Laurence Goosey; Abigail Knight; Karolina Krakowiak; Robert Maidstone; Mudassar Iqbal; Min Zi; Sukhpal Prehar; Elizabeth J Cartwright; Julie Gibbs; Laura C Matthews; Antony D Adamson; Neil E Humphreys; Pedro Rebelo-Guiomar; Michal Minczuk; David A Bechtold; Andrew Loudon; David Ray

    Efficient mitochondrial function is required in tissues with high energy demand such as the heart, and mitochondrial dysfunction is associated with cardiovascular disease. Expression of mitochondrial proteins is tightly regulated in response to internal and external stimuli. Here we identify a novel mechanism regulating mitochondrial content and function, through BUD23-dependent ribosome generation. BUD23 was required for ribosome maturation, normal 18S/28S stoichiometry and modulated the translation of mitochondrial transcripts in human A549 cells. Deletion of Bud23 in murine cardiomyocytes reduced mitochondrial content and function, leading to severe cardiomyopathy and death. We discovered that BUD23 selectively promotes ribosomal interaction with low GC-content 5'UTRs. Taken together we identify a critical role for BUD23 in bioenergetics gene expression, by promoting efficient translation of mRNA transcripts with low 5'UTR GC content. BUD23 emerges as essential to mouse development, and to postnatal cardiac function.

    更新日期:2020-01-15
  • A genetic, genomic, and computational resource for exploring neural circuit function
    eLife (IF 7.551) Pub Date : 2020-01-15
    Fred P Davis; Aljoscha Nern; Serge Picard; Michael B Reiser; Gerald M Rubin; Sean R Eddy; Gilbert L Henry

    The anatomy of many neural circuits is being characterized with increasing resolution, but their molecular properties remain mostly unknown. Here, we characterize gene expression patterns in distinct neural cell types of the Drosophila visual system using genetic lines to access individual cell types, the TAPIN-seq method to measure their transcriptomes, and a probabilistic method to interpret these measurements. We used these tools to build a resource of high-resolution transcriptomes for 100 driver lines covering 67 cell types, available at http://www.opticlobe.com. Combining these transcriptomes with recently reported connectomes helps characterize how information is transmitted and processed across a range of scales, from individual synapses to circuit pathways. We describe examples that include identifying neurotransmitters, including cases of apparent co-release, generating functional hypotheses based on receptor expression, as well as identifying strong commonalities between different cell types.

    更新日期:2020-01-15
  • CNApp, a tool for the quantification of copy number alterations and integrative analysis revealing clinical implications
    eLife (IF 7.551) Pub Date : 2020-01-15
    Sebastià Franch-Expósito; Laia Bassaganyas; Maria Vila-Casadesús; Eva Hernández-Illán; Roger Esteban-Fabró; Marcos Díaz-Gay; Juan José Lozano; Antoni Castells; Josep Maria Llovet; Sergi Castellvi-Bel; Jordi Camps

    Somatic copy number alterations (CNAs) are a hallmark of cancer, but their role in tumorigenesis and clinical relevance remain largely unclear. Here we developed CNApp, a web-based tool that allows a comprehensive exploration of CNAs by using purity-corrected segmented data from multiple genomic platforms. CNApp generates genome-wide profiles, computes CNA scores for broad, focal and global CNA burdens, and uses machine learning-based predictions to classify samples. We applied CNApp to the TCGA pan-cancer dataset of 10,635 genomes showing that CNAs classify cancer types according to their tissue-of-origin, and that each cancer type shows specific ranges of broad and focal CNA scores. Moreover, CNApp reproduces recurrent CNAs in hepatocellular carcinoma, and predicts colon cancer molecular subtypes and microsatellite instability based on broad CNA scores and discrete genomic imbalances. In summary, CNApp facilitates CNA-driven research by providing a unique framework to identify relevant clinical implications. CNApp is hosted at https://tools.idibaps.org/CNApp/.

    更新日期:2020-01-15
  • The tumor suppressor PTPRK promotes ZNRF3 internalization and is required for Wnt inhibition in the Spemann organizer
    eLife (IF 7.551) Pub Date : 2020-01-14
    Ling-Shih Chang; Minseong Kim; Andrey Glinka; Carmen Reinhard; Christof Niehrs

    A hallmark of Spemann organizer function is its expression of Wnt antagonists that regulate axial embryonic patterning. Here we identify the tumor suppressor Protein tyrosine phosphatase receptor-type kappa (Ptprk), as a Wnt inhibitor of the Spemann organizer. We show that PTPRK acts via the transmembrane E3 ubiquitin ligase ZNRF3, a negative regulator of Wnt signaling promoting Wnt receptor degradation, which is also expressed in the organizer. Deficiency of ptprk increases Wnt signaling, leading to reduced expression of Spemann organizer effector genes and inducing head and axial defects. We identify a '4Y' endocytic signal in ZNRF3, which Ptprk maintains unphosphorylated to promote Wnt receptor depletion. Our discovery of PTPRK as a negative regulator of Wnt receptor turnover provides a rationale for its tumor suppressive function and reveals that in PTPRK-RSPO3 recurrent cancer fusions both fusion partners, in fact, encode ZNRF3 regulators.

    更新日期:2020-01-14
  • Rank orders and signed interactions in evolutionary biology
    eLife (IF 7.551) Pub Date : 2020-01-14
    Kristina Crona

    Rank orders have been studied in evolutionary biology for almost a hundred years. Constraints on the order in which mutations accumulate are known from cancer drug treatment, and order constraints for species invasions are important in ecology. However, current theory on rank orders in biology is somewhat fragmented. Here we show how our previous work on inferring genetic interactions from comparative fitness data (Crona et al., 2017) is related to an influential approach to rank orders based on sign epistasis. Our approach depends on order perturbations that indicate interactions. We apply our results to malaria parasites and find that order perturbations beyond sign epistasis are prevalent for the antimalarial drug resistance landscape. This finding agrees with the observation that reversed evolution back to the ancestral type is difficult. Another application concerns bacteria adapting to a methanol environment.

    更新日期:2020-01-14
  • TTBK2 and primary cilia are essential for the connectivity and survival of cerebellar Purkinje neurons
    eLife (IF 7.551) Pub Date : 2020-01-14
    Emily Bowie; Sarah C Goetz

    Primary cilia are vital signaling organelles that extend from most types of cells, including neurons and glia. These structures are essential for the development of many tissues and organs, however, their function in adult tissues, particularly neurons in the brain, remains largely unknown. Tau tubulin kinase 2 (TTBK2) is a critical regulator of ciliogenesis, and is also mutated in a hereditary neurodegenerative disorder, spinocerebellar ataxia type 11 (SCA11). Here, we show that conditional knockout of Ttbk2 in adult mice results in degenerative cerebellar phenotypes that recapitulate aspects of SCA11 including motor coordination deficits and defects to Purkinje cell (PC) integrity. We also find that the Ttbk2 conditional mutant mice quickly lose cilia throughout the brain. We show that conditional knockout of the key ciliary trafficking gene Ift88 in adult mice results in nearly identical cerebellar phenotypes to those of the Ttbk2 knockout, indicating that disruption of ciliary signaling is a key driver of these phenotypes. Our data suggest that primary cilia play an integral role in maintaining the function of PCs in the adult cerebellum and reveal novel insights into mechanisms involved in neurodegeneration.

    更新日期:2020-01-14
  • TRF1 averts chromatin remodelling, recombination and replication dependent-Break Induced Replication at mouse telomeres
    eLife (IF 7.551) Pub Date : 2020-01-14
    Rosa Maria Porreca; Emilia Herrera-Moyano; Eleni Skourti; Pui Pik Law; Roser Gonzalez Franco; Alex Montoya; Peter Faull; Holger Kramer; Jean-Baptiste Vannier

    Telomeres are a significant challenge to DNA replication and are prone to replication stress and telomere fragility. The shelterin component TRF1 facilitates telomere replication but the molecular mechanism remains uncertain. By interrogating the proteomic composition of telomeres, we show that mouse telomeres lacking TRF1 undergo protein composition reorganisation associated with the recruitment of DNA damage response and chromatin remodellers. Surprisingly, mTRF1 suppresses the accumulation of promyelocytic leukemia (PML) protein, BRCA1 and the SMC5/6 complex at telomeres, which is associated with increased Homologous Recombination (HR) and TERRA transcription. We uncovered a previously unappreciated role for mTRF1 in the suppression of telomere recombination, dependent on SMC5 and also POLD3 dependent Break Induced Replication at telomeres. We propose that TRF1 facilitates S-phase telomeric DNA synthesis to prevent illegitimate mitotic DNA recombination and chromatin rearrangement.

    更新日期:2020-01-14
  • Delta-like 1 and Delta-like 4 differently require their extracellular domains for triggering Notch signaling in mice
    eLife (IF 7.551) Pub Date : 2020-01-14
    Ken-ichi Hirano; Akiko Suganami; Yutaka Tamura; Hideo Yagita; Sonoko Habu; Motoo Kitagawa; Takehito Sato; Katsuto Hozumi

    Delta-like (Dll) 1 and Dll4 differently function as Notch ligands in a context-dependent manner. As these ligands share structural properties, the molecular basis for their functional difference is poorly understood. Here, we investigated the superiority of Dll4 over Dll1 with respect to induction of T cell development using a domain-swapping approach in mice. The DOS motif, shared by Notch ligands—except Dll4—contributes to enhancing the activity of Dll for signal transduction. The module at the N-terminus of Notch ligand (MNNL) of Dll4 is inherently advantageous over Dll1. Molecular dynamic simulation revealed that the loop structure in MNNL domain of Dll1 contains unique proline residues with limited range of motion. The Dll4 mutant with Dll1-derived proline residues showed reduced activity. These results suggest that the loop structure—present within the MNNL domain—with a wide range of motion ensures the superiority of Dll4 and uniquely contributes to the triggering of Notch signaling.

    更新日期:2020-01-14
  • Intermittent hypoxia mediated by TSP1 dependent on STAT3 induces cardiac fibroblast activation and cardiac fibrosis
    eLife (IF 7.551) Pub Date : 2020-01-14
    Qiankun Bao; Bangying Zhang; Ya Suo; Chen Liu; Qian Yang; Kai Zhang; Ming Yuan; Meng Yuan; Yue Zhang; Guangping Li

    Intermittent hypoxia (IH) is the predominant pathophysiological disturbance in obstructive sleep apnea (OSA), known to be independently associated with cardiovascular diseases. However, the effect of IH on cardiac fibrosis and molecular events involved in this process are unclear. Here, we tested IH in angiotensin II (Ang II)-induced cardiac fibrosis and signaling linked to fibroblast activation. IH triggered cardiac fibrosis and aggravated Ang II-induced cardiac dysfunction in mice. Plasma thrombospondin-1 (TSP1) content was upregulated in both IH-exposed mice and OSA patients. Moreover, both in vivo and in vitro results showed IH-induced cardiac fibroblast activation and increased TSP1 expression in cardiac fibroblasts. Mechanistically, phosphorylation of STAT3 at Tyr705 mediated the IH-induced TSP1 expression and fibroblast activation. Finally, STAT3 inhibitor S3I-201 or AAV9 carrying a periostin promoter driving the expression of shRNA targeting Stat3 significantly attenuated the synergistic effects of IH and Ang II on cardiac fibrosis in mice. This work suggests a potential therapeutic strategy for OSA-related fibrotic heart disease.

    更新日期:2020-01-14
  • Lola regulates Drosophila adult midgut homeostasis via non-canonical Hippo signaling
    eLife (IF 7.551) Pub Date : 2020-01-14
    Xue Hao; Shimin Wang; Yi Lu; Wentao Yu; Pengyue Li; Dan Jiang; Tong Guo; Mengjie Li; Jinhui Li; Jinjin Xu; Wenqing Wu; Margaret S Ho; Lei Zhang

    Tissue homeostasis and regeneration in the Drosophila midgut is regulated by a diverse array of signaling pathways including the Hippo pathway. Hippo signaling restricts intestinal stem cell (ISC) proliferation by sequestering the transcription co-factor Yorkie (Yki) in the cytoplasm, a factor required for rapid ISC proliferation under injury-induced regeneration. Nonetheless, the mechanism of Hippo-mediated midgut homeostasis and whether canonical Hippo signaling is involved in ISC basal proliferation are less characterized. Here we identify Lola as a transcription factor acting downstream of Hippo signaling to restrict ISC proliferation in a Yki-independent manner. Not only that Lola interacts with and is stabilized by the Hippo signaling core kinase Warts (Wts), Lola rescues the enhanced ISC proliferation upon Wts depletion via suppressing Dref and SkpA expressions. Our findings reveal that Lola is a non-canonical Hippo signaling component in regulating midgut homeostasis, providing insights on the mechanism of tissue maintenance and intestinal function.

    更新日期:2020-01-14
  • Imp/IGF2BP levels modulate individual neural stem cell growth and division through myc mRNA stability
    eLife (IF 7.551) Pub Date : 2020-01-14
    Tamsin J Samuels; Aino I Järvelin; David Ish-Horowicz; Ilan Davis

    The numerous neurons and glia that form the brain originate from tightly controlled growth and division of neural stem cells, regulated systemically by important known stem cell-extrinsic signals. However, the cell-intrinsic mechanisms that control the distinctive proliferation rates of individual neural stem cells are unknown. Here, we show that the size and division rates of Drosophila neural stem cells (neuroblasts) are controlled by the highly conserved RNA binding protein Imp (IGF2BP), via one of its top binding targets in the brain, myc mRNA. We show that Imp stabilises myc mRNA leading to increased Myc protein levels, larger neuroblasts, and faster division rates. Declining Imp levels throughout development limit myc mRNA stability to restrain neuroblast growth and division, and heterogeneous Imp expression correlates with myc mRNA stability between individual neuroblasts in the brain. We propose that Imp-dependent regulation of myc mRNA stability fine-tunes individual neural stem cell proliferation rates.

    更新日期:2020-01-14
  • The bottom-up and top-down processing of faces in the human occipitotemporal cortex
    eLife (IF 7.551) Pub Date : 2020-01-14
    Xiaoxu Fan; Fan Wang; Hanyu Shao; Peng Zhang; Sheng He

    Although face processing has been studied extensively, the dynamics of how face-selective cortical areas are engaged remains unclear. Here we uncovered the timing of activation in core face-selective regions using functional Magnetic Resonance Imaging and Magnetoencephalography in humans. Processing of normal faces started in the posterior occipital areas and then proceeded to anterior regions. This bottom-up processing sequence was also observed even when internal facial features were misarranged. However, processing of two-tone Mooney faces lacking explicit prototypical facial features engaged top-down projection from the right posterior fusiform face area to right occipital face area. Further, face-specific responses elicited by contextual cues alone emerged simultaneously in the right ventral face-selective regions, suggesting parallel contextual facilitation. Together, our findings chronicle the precise timing of bottom-up, top-down, as well as context-facilitated processing sequences in the occipital-temporal face network, highlighting the importance of the top-down operations especially when faced with incomplete or ambiguous input.

    更新日期:2020-01-14
  • State-dependent brainstem ensemble dynamics and their interactions with hippocampus across sleep states
    eLife (IF 7.551) Pub Date : 2020-01-14
    Tomomi Tsunematsu; Amisha A Patel; Arno Onken; Shuzo Sakata

    The brainstem plays a crucial role in sleep-wake regulation. However, their ensemble dynamics underlying sleep regulation remain poorly understood. Here we show slow, state-predictive brainstem ensemble dynamics and state-dependent interactions between the brainstem and the hippocampus in mice. On a timescale of seconds to minutes, brainstem populations can predict pupil dilation and vigilance states where they exhibit longer predictable power compared with hippocampal CA1 neurons. On a timescale of sub-seconds, pontine waves (P-waves) are accompanied by synchronous firing of brainstem neurons during both rapid eye movement (REM) and non-REM (NREM) sleep. Crucially, P-waves functionally interact with CA1 activity in a state-dependent manner: during NREM sleep, hippocampal sharp wave-ripples (SWRs) precede P-waves. On the other hand, P-waves during REM sleep are phase-locked with ongoing theta oscillations and are followed by burst firing of CA1 neurons. This state-dependent global coordination between the brainstem and hippocampus implicates distinct functional roles of sleep.

    更新日期:2020-01-14
  • Disease-modifying effects of natural Δ9-tetrahydrocannabinol in endometriosis-associated pain
    eLife (IF 7.551) Pub Date : 2020-01-14
    Alejandra Escudero-Lara; Josep Argerich; David Cabañero; Rafael Maldonado

    Endometriosis is a chronic painful disease highly prevalent in women that is defined by growth of endometrial tissue outside the uterine cavity and lacks adequate treatment. Medical use of cannabis derivatives is a current hot topic and it is unknown whether phytocannabinoids may modify endometriosis symptoms and development. Here we evaluate the effects of repeated exposure to Δ9-tetrahydrocannabinol (THC) in a mouse model of surgically-induced endometriosis. In this model, female mice develop mechanical hypersensitivity in the caudal abdomen, mild anxiety-like behavior and substantial memory deficits associated with the presence of extrauterine endometrial cysts. Interestingly, daily treatments with THC (2 mg/kg) alleviate mechanical hypersensitivity and pain unpleasantness, modify uterine innervation and restore cognitive function without altering the anxiogenic phenotype. Strikingly, THC also inhibits the development of endometrial cysts. These data highlight the interest of scheduled clinical trials designed to investigate possible benefits of THC for women with endometriosis.

    更新日期:2020-01-14
  • Ankyrin-G mediates targeting of both Na+ and KATP channels to the rat cardiac intercalated disc
    eLife (IF 7.551) Pub Date : 2020-01-14
    Hua-Qian Yang; Marta Pérez-Hernández; Jose Sanchez-Alonso; Andriy Shevchuk; Julia Gorelik; Eli Rothenberg; Mario Delmar; William A Coetzee

    We investigated targeting mechanisms of Na+ and KATP channels to the intercalated disk (ICD) of cardiomyocytes. Patch clamp and surface biotinylation data show reciprocal downregulation of each other's surface density. Mutagenesis of the Kir6.2 ankyrin binding site disrupts this functional coupling. Duplex patch clamping and Angle SICM recordings show that INa and IKATP functionally co-localize at the rat ICD, but not at the lateral membrane. Quantitative STORM imaging show that Na+ and KATP channels are localized close to each other and to AnkG, but not to AnkB, at the ICD. Peptides corresponding to Nav1.5 and Kir6.2 ankyrin binding sites dysregulate targeting of both Na+ and KATP channels to the ICD, but not to lateral membranes. Finally, a clinically relevant gene variant that disrupts KATP channel trafficking also regulates Na+ channel surface expression. The functional coupling between these two channels need to be considered when assessing clinical variants and therapeutics.

    更新日期:2020-01-14
  • Prickle isoforms determine handedness of helical morphogenesis
    eLife (IF 7.551) Pub Date : 2020-01-14
    Bomsoo Cho; Song Song; Jeffrey D Axelrod

    Subcellular asymmetry directed by the planar cell polarity (PCP) signaling pathway orients numerous morphogenetic events in both invertebrates and vertebrates. Here, we describe a morphogenetic movement in which the intertwined socket and shaft cells of the Drosophila anterior wing margin mechanosensory bristles undergo PCP-directed apical rotation, inducing twisting that results in a helical structure of defined chirality. We show that the Frizzled/Vang PCP signaling module coordinates polarity among and between bristles and surrounding cells to direct this rotation. Furthermore, we show that dynamic interplay between two isoforms of the Prickle protein determines right- or left-handed bristle morphogenesis. We provide evidence that, Frizzled/Vang signaling couples to the Fat/Dachsous PCP directional signal in opposite directions depending on whether Pkpk or Pksple predominates. Dynamic interplay between Pk isoforms is likely to be an important determinant of PCP outcomes in diverse contexts. Similar mechanisms may orient other lateralizing morphogenetic processes.

    更新日期:2020-01-14
  • NINscope, a versatile miniscope for multi-region circuit investigations
    eLife (IF 7.551) Pub Date : 2020-01-14
    Andres de Groot; Bastijn JG van den Boom; Romano M van Genderen; Joris Coppens; John van Veldhuijzen; Joop Bos; Hugo Hoedemaker; Mario Negrello; Ingo Willuhn; Chris I De Zeeuw; Tycho M Hoogland

    Miniaturized fluorescence microscopes (miniscopes) have been instrumental to monitor neural signals during unrestrained behavior and their open-source versions have made them affordable. Often, the footprint and weight of open-source miniscopes is sacrificed for added functionality. Here, we present NINscope: a light-weight miniscope with a small footprint that integrates a high-sensitivity image sensor, an inertial measurement unit and an LED driver for an external optogenetic probe. We use it to perform the first concurrent cellular resolution recordings from cerebellum and cerebral cortex in unrestrained mice, demonstrate its optogenetic stimulation capabilities to examine cerebello-cerebral or cortico-striatal connectivity, and replicate findings of action encoding in dorsal striatum. In combination with cross-platform control software, our miniscope is a versatile addition to the expanding toolbox of open-source miniscopes that will increase access to multi-region circuit investigations during unrestrained behavior.

    更新日期:2020-01-14
  • Inflammation-induced IgE promotes epithelial hyperplasia and tumour growth
    eLife (IF 7.551) Pub Date : 2020-01-14
    Mark David Hayes; Sophie Ward; Greg Crawford; Rocio Castro Seoane; William David Jackson; David Kipling; David Voehringer; Deborah Dunn-Walters; Jessica Strid

    IgE is the least abundant circulating antibody class but is constitutively present in healthy tissues bound to resident cells via its high-affinity receptor, FcεRI. The physiological role of endogenous IgE antibodies is unclear but it has been suggested that they provide host protection against a variety of noxious environmental substances and parasitic infections at epithelial barrier surfaces. Here we show, in mice, that skin inflammation enhances levels of IgE antibodies that have natural specificities and a repertoire, VDJ rearrangements and CDRH3 characteristics similar to those of IgE antibodies in healthy tissue. IgE-bearing basophils are recruited to inflamed skin via CXCL12 and thymic stromal lymphopoietin (TSLP)/IL-3-dependent upregulation of CXCR4. In the inflamed skin, IgE/FcεRI-signalling in basophils promotes epithelial cell growth and differentiation, partly through histamine engagement of H1R and H4R. Furthermore, this IgE response strongly drives tumour outgrowth of epithelial cells harbouring oncogenic mutation. These findings indicate that natural IgE antibodies support skin barrier defences, but that during chronic tissue inflammation this role may be subverted to promote tumour growth.

    更新日期:2020-01-14
  • Structural basis of substrate recognition by a polypeptide processing and secretion transporter
    eLife (IF 7.551) Pub Date : 2020-01-14
    Virapat Kieuvongngam; Paul Dominic B Olinares; Anthony Palillo; Michael L Oldham; Brian T Chait; Jue Chen

    The peptidase-containing ATP-binding cassette transporters (PCATs) are unique members of the ABC transporter family that proteolytically process and export peptides and proteins. Each PCAT contains two peptidase domains that cleave off the secretion signal, two transmembrane domains forming a translocation pathway, and two nucleotide-binding domains that hydrolyze ATP. Previously the crystal structures of a PCAT from Clostridium thermocellum (PCAT1) were determined in the absence and presence of ATP, revealing how ATP binding regulates the protease activity and access to the translocation pathway. However, how the substrate CtA, a 90-residue polypeptide, is recognized by PCAT1 remained elusive. To address this question, we determined the structure of the PCAT1-CtA complex by electron cryo-microscopy (cryo-EM) to 3.4 Å resolution. The structure shows that two CtAs are bound via their N-terminal leader peptides, but only one is positioned for cleavage and translocation. Based on these results, we propose a model of how substrate cleavage, ATP hydrolysis, and substrate translocation are coordinated in a transport cycle.

    更新日期:2020-01-14
  • Midkine is a dual regulator of wound epidermis development and inflammation during the initiation of limb regeneration
    eLife (IF 7.551) Pub Date : 2020-01-14
    Stephanie L Tsai; Clara Baselga-Garriga; Douglas A Melton

    Formation of a specialized wound epidermis is required to initiate salamander limb regeneration. Yet little is known about the roles of the early wound epidermis during the initiation of regeneration and the mechanisms governing its development into the apical epithelial cap (AEC), a signaling structure necessary for outgrowth and patterning of the regenerate. Here, we elucidate the functions of the early wound epidermis, and further reveal midkine (mk) as a dual regulator of both AEC development and inflammation during the initiation of axolotl limb regeneration. Through loss- and gain-of-function experiments, we demonstrate that mk acts as both a critical survival signal to control the expansion and function of the early wound epidermis and an anti-inflammatory cytokine to resolve early injury-induced inflammation. Altogether, these findings unveil one of the first identified regulators of AEC development and provide fundamental insights into early wound epidermis function, development, and the initiation of limb regeneration.

    更新日期:2020-01-14
  • Degradation of engulfed mitochondria is rate-limiting in Optineurin-mediated mitophagy in neurons
    eLife (IF 7.551) Pub Date : 2020-01-14
    Chantell S Evans; Erika LF Holzbaur

    Mitophagy, the selective removal of damaged mitochondria, is thought to be critical to maintain neuronal homeostasis. Mutations of proteins in the pathway cause neurodegenerative diseases, suggesting defective mitochondrial turnover contributes to neurodegeneration. In primary rat hippocampal neurons, we developed a mitophagy induction paradigm where mild oxidative stress induced low levels of mitochondrial damage. Mitophagy-associated proteins were sequentially recruited to depolarized mitochondria followed by sequestration into autophagosomes. The localization of these mitophagy events had a robust somal bias. In basal and induced conditions, engulfed mitochondria remained in non-acidified organelles for hours to days, illustrating efficient autophagosome sequestration but delayed lysosomal fusion or acidification. Furthermore, expression of an ALS-linked mutation in the pathway disrupted mitochondrial network integrity and this effect was exacerbated by oxidative stress. Thus, age-related decline in neuronal health or expression of disease-associated mutations in the pathway may exacerbate the slow kinetics of neuronal mitophagy, leading to neurodegeneration.

    更新日期:2020-01-14
  • In vitro reconstitution of branching microtubule nucleation
    eLife (IF 7.551) Pub Date : 2020-01-14
    Ammarah Tariq; Lucy Green; J Charles G Jeynes; Christian Soeller; James G Wakefield

    Eukaryotic cell division requires the mitotic spindle, a microtubule (MT)-based structure which accurately aligns and segregates duplicated chromosomes. The dynamics of spindle formation are determined primarily by correctly localising the MT nucleator, γ-Tubulin Ring Complex (γ-TuRC), within the cell. A conserved MT-associated protein complex, Augmin, recruits γ-TuRC to pre-existing spindle MTs, amplifying their number, in an essential cellular phenomenon termed ‘branching’ MT nucleation. Here, we purify endogenous, GFP-tagged Augmin and γ-TuRC from Drosophila embryos to near homogeneity using a novel one-step affinity technique. We demonstrate that, in vitro, while Augmin alone does not affect Tubulin polymerisation dynamics, it stimulates γ-TuRC-dependent MT nucleation in a cell cycle-dependent manner. We also assemble and visualise the MT-Augmin-γ-TuRC-MT junction using light microscopy. Our work therefore conclusively reconstitutes branching MT nucleation. It also provides a powerful synthetic approach with which to investigate the emergence of cellular phenomena, such as mitotic spindle formation, from component parts.

    更新日期:2020-01-14
  • Biochemical reconstitution of branching microtubule nucleation
    eLife (IF 7.551) Pub Date : 2020-01-14
    Raymundo Alfaro-Aco; Akanksha Thawani; Sabine Petry

    Microtubules are nucleated from specific locations at precise times in the cell cycle. However, the factors that constitute these microtubule nucleation pathways and their mode of action still need to be identified. Using purified Xenopus laevis proteins we biochemically reconstitute branching microtubule nucleation, which is critical for chromosome segregation. We found that besides the microtubule nucleator gamma-tubulin ring complex (γ-TuRC), the branching effectors augmin and TPX2 are required to efficiently nucleate microtubules from pre-existing microtubules. TPX2 has the unexpected capacity to directly recruit γ-TuRC as well as augmin, which in turn targets more γ-TuRC along the microtubule lattice. TPX2 and augmin enable γ-TuRC-dependent microtubule nucleation at preferred branching angles of less than 90 degrees from regularly-spaced patches along microtubules. This work provides a blueprint for other microtubule nucleation pathways and helps explain how microtubules are generated in the spindle.

    更新日期:2020-01-14
  • Living Science: Stepping down
    eLife (IF 7.551) Pub Date : 2020-01-14
    Eve Marder

    As Eve Marder stands down as a Deputy Editor of eLife, she reflects on the need for journals to change and respond to their environment

    更新日期:2020-01-14
  • Nanopore direct RNA sequencing maps the complexity of Arabidopsis mRNA processing and m6A modification
    eLife (IF 7.551) Pub Date : 2020-01-14
    Matthew T Parker; Katarzyna Knop; Anna V Sherwood; Nicholas J Schurch; Katarzyna Mackinnon; Peter D Gould; Anthony JW Hall; Geoffrey J Barton; Gordon G Simpson

    Understanding genome organization and gene regulation requires insight into RNA transcription, processing and modification. We adapted nanopore direct RNA sequencing to examine RNA from a wild-type accession of the model plant Arabidopsis thaliana and a mutant defective in mRNA methylation (m6A). Here we show that m6A can be mapped in full-length mRNAs transcriptome-wide and reveal the combinatorial diversity of cap-associated transcription start sites, splicing events, poly(A) site choice and poly(A) tail length. Loss of m6A from 3’ untranslated regions is associated with decreased relative transcript abundance and defective RNA 3′ end formation. A functional consequence of disrupted m6A is a lengthening of the circadian period. We conclude that nanopore direct RNA sequencing can reveal the complexity of mRNA processing and modification in full-length single molecule reads. These findings can refine Arabidopsis genome annotation. Further, applying this approach to less well-studied species could transform our understanding of what their genomes encode.

    更新日期:2020-01-14
  • Flower-like patterns in multi-species bacterial colonies
    eLife (IF 7.551) Pub Date : 2020-01-14
    Liyang Xiong; Yuansheng Cao; Robert Cooper; Wouter-Jan Rappel; Jeff Hasty; Lev Tsimring

    Diverse interactions among species within bacterial colonies lead to intricate spatiotemporal dynamics, which can affect their growth and survival. Here, we describe the emergence of complex structures in a colony grown from mixtures of motile and non-motile bacterial species on a soft agar surface. Time-lapse imaging shows that non-motile bacteria 'hitchhike' on the motile bacteria as the latter migrate outward. The non-motile bacteria accumulate at the boundary of the colony and trigger an instability that leaves behind striking flower-like patterns. The mechanism of the front instability governing this pattern formation is elucidated by a mathematical model for the frictional motion of the colony interface, with friction depending on the local concentration of the non-motile species. A more elaborate two-dimensional phase-field model that explicitly accounts for the interplay between growth, mechanical stress from the motile species, and friction provided by the non-motile species, fully reproduces the observed flower-like patterns.

    更新日期:2020-01-14
  • Duodenum Intestine-Chip for preclinical drug assessment in a human relevant model
    eLife (IF 7.551) Pub Date : 2020-01-14
    Magdalena Kasendra; Raymond Luc; Jianyi Yin; Dimitris V Manatakis; Gauri Kulkarni; Carolina Lucchesi; Josiah Sliz; Athanasia Apostolou; Laxmi Sunuwar; Jenifer Obrigewitch; Kyung-Jin Jang; Geraldine A Hamilton; Mark Donowitz; Katia Karalis

    Induction of intestinal drug metabolizing enzymes can complicate the development of new drugs, owing to the potential to cause drug-drug interactions (DDIs) leading to changes in pharmacokinetics, safety and efficacy. The development of a human-relevant model of the adult intestine that accurately predicts CYP450 induction could help address this challenge as species differences preclude extrapolation from animals. Here, we combined organoids and Organs-on-Chips technology to create a human Duodenum Intestine-Chip that emulates intestinal tissue architecture and functions, that are relevant for the study of drug transport, metabolism, and DDI. Duodenum Intestine-Chip demonstrates the polarized cell architecture, intestinal barrier function, presence of specialized cell subpopulations, and in vivo relevant expression, localization, and function of major intestinal drug transporters. Notably, in comparison to Caco-2, it displays improved CYP3A4 expression and induction capability. This model could enable improved in vitro to in vivo extrapolation for better predictions of human pharmacokinetics and risk of DDIs.

    更新日期:2020-01-14
  • MYC and Twist1 cooperate to drive metastasis by eliciting crosstalk between cancer and innate immunity
    eLife (IF 7.551) Pub Date : 2020-01-14
    Renumathy Dhanasekaran; Virginie Baylot; Minsoon Kim; Sibu Kuruvilla; David I Bellovin; Nia Adeniji; Anand Rajan KD; Ian Lai; Meital Gabay; Ling Tong; Maya Krishnan; Jangho Park; Theodore Hu; Mustafa A Barbhuiya; Andrew J Gentles; Kasthuri Kannan; Phuoc T Tran; Dean W Felsher

    Metastasis is a major cause of cancer mortality. We generated an autochthonous transgenic mouse model whereby conditional expression of MYC and Twist1 enables hepatocellular carcinoma (HCC) to metastasize in >90% of mice. MYC and Twist1 cooperate and their sustained expression is required to elicit a transcriptional program associated with the activation of innate immunity, through secretion of a cytokinome that elicits recruitment and polarization of tumor associated macrophages (TAMs). Systemic treatment with Ccl2 and Il13 induced MYC-HCCs to metastasize; whereas, blockade of Ccl2 and Il13 abrogated MYC/Twist1-HCC metastasis. Further, in 33 human cancers (n = 9502) MYC and TWIST1 predict poor survival (p=4.3×10−10), CCL2/IL13 expression (p<10−109) and TAM infiltration (p<10−96). Finally, in the plasma of patients with HCC (n = 25) but not cirrhosis (n = 10), CCL2 and IL13 were increased and IL13 predicted invasive tumors. Therefore, MYC and TWIST1 generally appear to cooperate in human cancer to elicit a cytokinome that enables metastasis through crosstalk between cancer and immune microenvironment.

    更新日期:2020-01-14
  • Cytomegalovirus protein m154 perturbs the adaptor protein-1 compartment mediating broad-spectrum immune evasion
    eLife (IF 7.551) Pub Date : 2020-01-13
    Ivana Strazic Geljic; Paola Kucan Brlic; Guillem Angulo; Ilija Brizic; Berislav Lisnic; Tina Jenus; Vanda Juranic Lisnic; Gian Pietro Pietri; Pablo Engel; Noa Kaynan; Jelena Zeleznjak; Peter Schu; Ofer Mandelboim; Astrid Krmpotic; Ana Angulo; Stipan Jonjic; Tihana Lenac Rovis

    Cytomegaloviruses (CMVs) are ubiquitous pathogens known to employ numerous immunoevasive strategies that significantly impair the ability of the immune system to eliminate the infected cells. Here, we report that the single mouse CMV (MCMV) protein, m154, downregulates multiple surface molecules involved in the activation and costimulation of the immune cells. We demonstrate that m154 uses its cytoplasmic tail motif, DD, to interfere with the adaptor protein-1 (AP-1) complex, implicated in intracellular protein sorting and packaging. As a consequence of the perturbed AP-1 sorting, m154 promotes lysosomal degradation of several proteins involved in T cell costimulation, thus impairing virus-specific CD8+ T cell response and virus control in vivo. Additionally, we show that HCMV infection similarly interferes with the AP-1 complex. Altogether, we identify the robust mechanism employed by single viral immunomodulatory protein targeting a broad spectrum of cell surface molecules involved in the antiviral immune response.

    更新日期:2020-01-14
  • The structure of species discrimination signals across a primate radiation
    eLife (IF 7.551) Pub Date : 2020-01-13
    Sandra Winters; William L Allen; James P Higham

    Discriminating conspecifics from heterospecifics can help avoid costly interactions between closely related sympatric species. The guenons, a recent primate radiation, exhibit high degrees of sympatry and form multi-species groups. Guenons have species-specific colorful face patterns hypothesized to function in species discrimination. Here, we use a machine learning approach to identify face regions most essential for species classification across fifteen guenon species. We validate these computational results using experiments with live guenons, showing that facial traits critical for accurate classification influence selective attention toward con- and heterospecific faces. Our results suggest variability among guenon species in reliance on single-trait-based versus holistic facial characteristics for species discrimination, with behavioral responses and computational results indicating variation from single-trait to whole-face patterns. Our study supports a role for guenon face patterns in species discrimination, and shows how complex signals can be informative about differences between species across a speciose and highly sympatric radiation.

    更新日期:2020-01-14
  • Structural basis for COMPASS recognition of an H2B-ubiquitinated nucleosome
    eLife (IF 7.551) Pub Date : 2020-01-10
    Evan J Worden; Xiangbin Zhang; Cynthia Wolberger

    Methylation of histone H3K4 is a hallmark of actively transcribed genes that depends on mono-ubiquitination of histone H2B (H2B-Ub). H3K4 methylation in yeast is catalyzed by Set1, the methyltransferase subunit of COMPASS. We report here the cryo-EM structure of a six-protein core COMPASS subcomplex, which can methylate H3K4 and be stimulated by H2B-Ub, bound to a ubiquitinated nucleosome. Our structure shows that COMPASS spans the face of the nucleosome, recognizing ubiquitin on one face of the nucleosome and methylating H3 on the opposing face. As compared to the structure of the isolated core complex, Set1 undergoes multiple structural rearrangements to cement interactions with the nucleosome and with ubiquitin. The critical Set1 RxxxRR motif adopts a helix that mediates bridging contacts between the nucleosome, ubiquitin and COMPASS. The structure provides a framework for understanding mechanisms of trans-histone cross-talk and the dynamic role of H2B ubiquitination in stimulating histone methylation.

    更新日期:2020-01-10
  • Metabolic signature in nucleus accumbens for anti-depressant-like effects of acetyl-L-carnitine
    eLife (IF 7.551) Pub Date : 2020-01-10
    Antoine Cherix; Thomas Larrieu; Jocelyn Grosse; João Rodrigues; Bruce McEwen; Carla Nasca; Rolf Gruetter; Carmen Sandi

    Emerging evidence suggests that hierarchical status provide vulnerability to develop stress-induced depression. Energy metabolic changes in the nucleus accumbens (NAc) were recently related to hierarchical status and vulnerability to develop depression-like behavior. Acetyl-L-carnitine (LAC), a mitochondria-boosting supplement, has shown promising antidepressant-like effects opening therapeutic opportunities for restoring energy balance in depressed patients. We investigated the metabolic impact in the NAc of antidepressant LAC treatment in chronically-stressed mice using 1H-magnetic resonance spectroscopy (1H-MRS). High rank, but not low rank, mice, as assessed with the tube test, showed behavioral vulnerability to stress, supporting a higher susceptibility of high social rank mice to develop depressive-like behaviors. High rank mice also showed reduced levels of several energy-related metabolites in the NAc that were counteracted by LAC treatment. Therefore, we reveal a metabolic signature in the NAc for antidepressant-like effects of LAC in vulnerable mice characterized by restoration of stress-induced neuroenergetics alterations and lipid function.

    更新日期:2020-01-10
  • Two forms of Opa1 cooperate to complete fusion of the mitochondrial inner-membrane
    eLife (IF 7.551) Pub Date : 2020-01-10
    Yifan Ge; Xiaojun Shi; Sivakumar Boopathy; Julie McDonald; Adam W Smith; Luke H Chao

    Mitochondrial membrane dynamics is a cellular rheostat that relates metabolic function and organelle morphology. Using an in vitro reconstitution system, we describe a mechanism for how mitochondrial inner-membrane fusion is regulated by the ratio of two forms of Opa1. We found that the long-form of Opa1 (l-Opa1) is sufficient for membrane docking, hemifusion and low levels of content release. However, stoichiometric levels of the processed, short form of Opa1 (s-Opa1) work together with l-Opa1 to mediate efficient and fast membrane pore opening. Additionally, we found that excess levels of s-Opa1 inhibit fusion activity, as seen under conditions of altered proteostasis. These observations describe a mechanism for gating membrane fusion.

    更新日期:2020-01-10
  • Distinct cytoskeletal proteins define zones of enhanced cell wall synthesis in Helicobacter pylori
    eLife (IF 7.551) Pub Date : 2020-01-09
    Jennifer A Taylor; Benjamin P Bratton; Sophie R Sichel; Kris M Blair; Holly M Jacobs; Kristen E DeMeester; Erkin Kuru; Joe Gray; Jacob Biboy; Michael S VanNieuwenhze; Waldemar Vollmer; Catherine L Grimes; Josh W Shaevitz; Nina Reda Salama

    Helical cell shape is necessary for efficient stomach colonization by Helicobacter pylori, but the molecular mechanisms for generating helical shape remain unclear. The helical centerline pitch and radius of wild-type H. pylori cells dictate surface curvatures of considerably higher positive and negative Gaussian curvatures than those present in straight- or curved-rod H. pylori. Quantitative 3D microscopy analysis of short pulses with either N-acetylmuramic acid or D-alanine metabolic probes showed that cell wall growth is enhanced at both sidewall curvature extremes. Immunofluorescence revealed MreB is most abundant at negative Gaussian curvature, while the bactofilin CcmA is most abundant at positive Gaussian curvature. Strains expressing CcmA variants with altered polymerization properties lose helical shape and associated positive Gaussian curvatures. We thus propose a model where CcmA and MreB promote PG synthesis at positive and negative Gaussian curvatures, respectively, and that this patterning is one mechanism necessary for maintaining helical shape.

    更新日期:2020-01-09
  • Cardiac ryanodine receptor distribution is dynamic and changed by auxiliary proteins and post-translational modification
    eLife (IF 7.551) Pub Date : 2020-01-09
    Parisa Asghari; David RL Scriven; Myles Ng; Pankaj Panwar; Keng C Chou; Filip van Petegem; Edwin DW Moore

    The effects of the immunophilins, FKBP12 and FKBP12.6, and phosphorylation on type II ryanodine receptor (RyR2) arrangement and function were examined using correlation microscopy (line scan confocal imaging of Ca2+ sparks and dual-tilt electron tomography) and dSTORM imaging of permeabilized Wistar rat ventricular myocytes. Saturating concentrations (10 µmol/L) of either FKBP12 or 12.6 significantly reduced the frequency, spread, amplitude and Ca2+ spark mass relative to control, while the tomograms revealed both proteins shifted the tetramers into a largely side-by-side configuration. Phosphorylation of immunophilin-saturated RyR2 resulted in structural and functional changes largely comparable to phosphorylation alone. dSTORM images of myocyte surfaces demonstrated that both FKBP12 and 12.6 significantly reduced RyR2 cluster sizes, while phosphorylation, even of immunophilin-saturated RyR2, increased them. We conclude that both RyR2 cluster size and the arrangement of tetramers within clusters is dynamic and respond to changes in the cellular environment. Further, these changes affect Ca2+ spark formation.

    更新日期:2020-01-09
  • A processive rotary mechanism couples substrate unfolding and proteolysis in the ClpXP degradation machinery
    eLife (IF 7.551) Pub Date : 2020-01-09
    Zev A Ripstein; Siavash Vahidi; Walid A Houry; John L Rubinstein; Lewis E Kay

    The ClpXP degradation machine consists of a hexameric AAA+ unfoldase (ClpX) and a pair of heptameric serine protease rings (ClpP) that unfold, translocate, and subsequently degrade client proteins. ClpXP is an important target for drug development against infectious diseases. Although structures are available for isolated ClpX and ClpP rings, it remains unknown how symmetry mismatched ClpX and ClpP work in tandem for processive substrate translocation into the ClpP proteolytic chamber. Here we present cryo-EM structures of the substrate-bound ClpXP complex from Neisseria meningitidis at 2.3 to 3.3 Å resolution. The structures allow development of a model in which the sequential hydrolysis of ATP is coupled to motions of ClpX loops that lead to directional substrate translocation and ClpX rotation relative to ClpP. Our data add to the growing body of evidence that AAA+ molecular machines generate translocating forces by a common mechanism.

    更新日期:2020-01-09
  • Point of View: Open Exploration
    eLife (IF 7.551) Pub Date : 2020-01-09
    William Hedley Thompson; Jessey Wright; Patrick G Bissett

    Arguments in support of open science tend to focus on confirmatory research practices. Here we argue that exploratory research should also be encouraged within the framework of open science. We lay out the benefits of 'open exploration' and propose two complementary ways to implement this with little infrastructural change.

    更新日期:2020-01-09
  • Coordination of humoral immune factors dictates compatibility between Schistosoma mansoni and Biomphalaria glabrata
    eLife (IF 7.551) Pub Date : 2020-01-09
    Hongyu Li; Jacob R Hambrook; Emmanuel A Pila; Abdullah A Gharamah; Jing Fang; Xinzhong Wu; Patrick Hanington

    Immune factors in snails of the genus Biomphalaria are critical for combating Schistosoma mansoni, the predominant cause of human intestinal schistosomiasis. Independently, many of these factors play an important role in, but do not fully define, the compatibility between the model snail B. glabrata, and S. mansoni. Here, we demonstrate association between four previously characterized humoral immune molecules; BgFREP3, BgTEP1, BgFREP2 and Biomphalysin. We also identify unique immune determinants in the plasma of S. mansoni-resistant B. glabrata that associate with the incompatible phenotype. These factors coordinate to initiate haemocyte-mediated destruction of S. mansoni sporocysts via production of reactive oxygen species. The inclusion of BgFREP2 in a BgFREP3-initiated complex that also includes BgTEP1 almost completely explains resistance to S. mansoni in this model. Our study unifies many independent lines of investigation to provide a more comprehensive understanding of the snail immune system in the context of infection by this important human parasite.

    更新日期:2020-01-09
  • Cell type specific control of basolateral amygdala neuronal circuits via entorhinal cortex-driven feedforward inhibition
    eLife (IF 7.551) Pub Date : 2020-01-09
    E Mae Guthman; Joshua D Garcia; Ming Ma; Philip Chu; Serapio M Baca; Katharine R Smith; Diego Restrepo; Molly M Huntsman

    The basolateral amygdala (BLA) plays a vital role in associating sensory stimuli with salient valence information. Excitatory principal neurons (PNs) undergo plastic changes to encode this association; however, local BLA inhibitory interneurons (INs) gate PN plasticity via feedforward inhibition (FFI). Despite literature implicating parvalbumin expressing (PV+) INs in FFI in cortex and hippocampus, prior anatomical experiments in BLA implicate somatostatin expressing (Sst+) INs. The lateral entorhinal cortex (LEC) projects to BLA where it drives FFI. In the present study, we explored the role of interneurons in this circuit. Using mice, we combined patch clamp electrophysiology, chemogenetics, unsupervised cluster analysis, and predictive modeling and found that a previously unreported subpopulation of fast-spiking Sst+ INs mediate LEC→BLA FFI.

    更新日期:2020-01-09
  • Origin and role of the cerebrospinal fluid bidirectional flow in the central canal
    eLife (IF 7.551) Pub Date : 2020-01-09
    Olivier Thouvenin; Ludovic Keiser; Yasmine Cantaut-Belarif1; Martin Carbo-Tano; Frederik Verweij; Nathalie Jurisch-Yaksi; Pierre-Luc Bardet; Guillaume van Niel; Francois Gallaire; Claire Wyart

    Circulation of the cerebrospinal fluid (CSF) contributes to body axis formation and brain development. Here, we investigated the unexplained origins of the CSF flow bidirectionality in the central canal of the spinal cord of 30 hpf zebrafish embryos and its impact on development. Experiments combined with modeling and simulations demonstrate that the CSF flow is generated locally by caudally-polarized motile cilia along the ventral wall of the central canal. The closed geometry of the canal imposes the average flow rate to be null, explaining the reported bidirectionality. We also demonstrate that at this early stage, motile cilia ensure the proper formation of the central canal. Furthermore, we demonstrate that the bidirectional flow accelerates the transport of particles in the CSF via a coupled convective-diffusive transport process. Our study demonstrates that cilia activity combined with muscle contractions sustain the long-range transport of extracellular lipidic particles, enabling embryonic growth.

    更新日期:2020-01-09
  • Robust perisomatic GABAergic self-innervation inhibits basket cells in the human and mouse supragranular neocortex
    eLife (IF 7.551) Pub Date : 2020-01-09
    Viktor Szegedi; Melinda Paizs; Judith Baka; Pál Barzó; Gábor Molnár; Gabor Tamas; Karri Lamsa

    Inhibitory autapses are self-innervating synaptic connections in GABAergic interneurons in the brain. Autapses in neocortical layers have not been systematically investigated, and their function in different mammalian species and specific interneuron types is poorly known. We investigated GABAergic parvalbumin-expressing basket cells (pvBCs) in layer 2/3 (L2/3) in human neocortical tissue resected in deep-brain surgery, and in mice as control. Most pvBCs showed robust GABAAR-mediated self-innervation in both species, but autapses were rare in nonfast-spiking GABAergic interneurons. Light- and electron microscopy analyses revealed pvBC axons innervating their own soma and proximal dendrites. GABAergic self-inhibition conductance was similar in human and mouse pvBCs and comparable to that of synapses from pvBCs to other L2/3 neurons. Autaptic conductance prolonged somatic inhibition in pvBCs after a spike and inhibited repetitive firing. Perisomatic autaptic inhibition is common in both human and mouse pvBCs of supragranular neocortex, where they efficiently control discharge of the pvBCs.

    更新日期:2020-01-09
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