bluuebunny/crossref_metadata_2025 huggingface dataset:
South Asian Patient Population Genetics Reveal Strong Founder Effects and High Rates of Homozygosity – New Resources for Precision Medicine
Authors: Wall Jeffrey D. , Sathirapongsasuti J. Fah , Gupta Ravi , Barik Anamitra , Rai Rajesh Kumar , Rasheed Asif , Radha Venkatesan , Belsare Saurabh , Menon Ramesh , Phalke Sameer , Mittal Anuradha , Fang John , Tanneeru Deepak , Robinson Jacqueline , Chaudhary Ruchi , Fuchsberger Christian , Forer Lukas , Schoenherr Sebastian , Bei Qixin , Bhangale Tushar , Tom Jennifer , Krishna Gadde Santosh Gopi , Priya B. V. , Naik Naveen Kumar , Wang Minxian , Kwok Pui-Yan , Khera Amit V. , Lakshmi B. R. , Butterworth Adam , Danesh John , Seshagiri Sekar , Kathiresan Sekar , Ghosh Arkasubhra , Mohan V. , Chowdhury Abhijit , Saleheen Danish , Stawiski Eric , Peterson Andrew S.
Published: October 2020

Population-scale genetic studies can identify drug targets and allow disease risk to be predicted with resulting benefit for management of individual health risks and system-wide allocation of health care delivery resources. Although population-scale projects are underway in many parts of the world, genetic variation between population groups means that additional projects are warranted. South Asia has a population whose genetics is the least characterized of any of the world’s major populations. Here we describe GenomeAsia studies that characterize population structure in South Asia and that create tools for economical and accurate genotyping at population-scale. Prior work on population structure characterized isolated population groups, the relevance of which to large-scale studies of disease genetics is unclear. For our studies we used whole genome sequence information from 4,807 individuals recruited in the health care delivery systems of Pakistan, India and Bangladesh to ensure relevance to population-scale studies of disease genetics. We combined this with WGS data from 927 individuals from isolated South Asian population groups, and developed a custom SNP array (called SARGAM) that is optimized for future human genetic studies in South Asia. We find evidence for high rates of reproductive isolation, endogamy and consanguinity that vary across the subcontinent and that lead to levels of homozygosity that approach 100 times that seen in outbred populations. We describe founder effects that increase the power to associate functional variants with disease processes and that make South Asia a uniquely powerful place for population-scale genetic studies.
CoffeeProt: An online tool for correlation and functional enrichment of proteome-wide systems genetics
Authors: Molendijk Jeffrey , Seldin Marcus M. , Parker Benjamin L.
Published: October 2020

The integration of genomics, proteomics and phenotypic traits across genetically diverse populations is a powerful approach to discover novel biological regulators. The increasing volume of complex data require new and easy-to-use tools accessible to a variety of scientists for the discovery and visualization of functionally relevant associations. To meet this requirement, we developedCoffeeProt, an open-source tool that analyzes genetic variants associated to protein networks and phenotypic traits.CoffeeProtuses proteomics data to perform correlation network analysis and annotates protein-protein interactions and subcellular localizations. It then integrates genetic and phenotypic associations along with variant effect predictions. We demonstrate its utility with the analysis of mouse and human population data enabling the rapid identification of genetic variants associated with protein complexes and clinical traits. We expectCoffeeProtwill serve the proteomics and systems genetics communities, leading to the discovery of novel biologically relevant associations.CoffeeProtis available atwww.coffeeprot.com.
Amyloidogenic core of a human λ-III immunoglobulin light chain fibril and their germline variants probed by MAS solid state NMR
Authors: Pradhan Tejaswini , Annamalai Karthikeyan , Sarkar Riddhiman , Huhn Stephanie , Hegenbart Ute , Schönland Stefan , Fändrich Marcus , Reif Bernd
Published: October 2020

Systemic antibody light chains (AL) amyloidosis is characterized by deposition of amyloid fibrils derived from a particular antibody light chain. Cardiac involvement is a major risk factor for mortality. Using MAS solid-state NMR, we study the fibril structure of a recombinant light chain fragment corresponding to the fibril protein from patient FOR005, together with fibrils formed by protein sequence variants that reflect the closest germline (GL) sequence. Both analyzed fibril structures were seeded withex-vivoamyloid fibrils purified from the explanted heart of this patient. We find that residues 11-42 and 69-102 adopt β-sheet conformation in patient protein fibrils. We identify glycine-49 that is mutated with respect to the germline sequence into arginine-49 as a key residue that forms a salt bridge to aspartate-25 in the patient protein fibril structure. Fibrils from the GL protein and from the patient protein harboring the single point mutation R49G can be both heterologously seeded using patientex-vivofibrils. Seeded R49G fibrils show an increased heterogeneity for the C-terminal residues 80-102 which is reflected by the disappearance of all resonances of these residues. By contrast, residues 11-42 and 69-77, which are visible in the MAS solid-state NMR spectra show13Cα chemical shifts that are highly similar to patient fibrils. The mutation R49G thus induces a conformational heterogeneity at the C-terminus in the fibril state, while the overall fibril topology is retained.
U-Infuse: Democratization of Customizable AI for Object Detection
Authors: Shepley Andrew , Falzon Greg , Lawson Christopher , Meek Paul , Kwan Paul
Published: October 2020

Image data is one of the primary sources of ecological data used in biodiversity conservation and management worldwide. However, classifying and interpreting large numbers of images is time and resource expensive, particularly in the context of camera trapping. Deep learning models have been used to achieve this task but are often not suited to specific applications due to their inability to generalise to new environments and inconsistent performance. Models need to be developed for specific species cohorts and environments, but the technical skills required to achieve this are a key barrier to the accessibility of this technology to ecologists. There is a strong need to democratise access to deep learning technologies by providing an easy to use software application allowing non-technical users to custom train custom object detectors.

U-Infuse addresses this issue by putting the power of AI into the hands of ecologists. U-Infuse provides ecologists with the ability to train customised models using publicly available images and/or their own camera trap images, without the constraints of annotating and pre-processing large numbers of images, or specific technical expertise. U-Infuse is a free and open-source software solution that supports both multiclass and single class training and inference, allowing ecologists to access state of the art AI on their own device, customised to their application without sharing IP or sensitive data.

U-Infuse provides ecological practitioners with the ability to (i) easily achieve camera trap object detection within a user-friendly GUI, generating a species distribution report, and other useful statistics, (ii) custom train deep learning models using publicly available and custom training data, (iii) achieve supervised auto-annotation of images for further training, with the benefit of editing annotations to ensure quality datasets.

Broad adoption of U-Infuse by ecological practitioners will improve camera trap image analysis and processing by allowing significantly more image data to be processed with minimal expenditure of time and resources. Ease of training and reliance on transfer learning means domain-specific models can be trained rapidly, and frequently updated without the need for computer science expertise, or data sharing, protecting intellectual property and privacy.
Effects of relocation on Immunological and physiological measures in squirrel monkeys (Saimiri boliviensis boliviensis)
Authors: Nehete Pramod , Nehete Bharti P , Wilkerson Greg K , Schapiro Steve J , Williams Lawrence E
Published: October 2020

In the present study, we have quantified the effects of transport, relocation and acclimate/adapt to their new surroundings on squirrel monkey. These responses are measured in blood samples obtained from squirrel monkeys, at different time points relative to their relocation from their old home to their new home. A variety of immunological assays are performed on the phenotype and function of peripheral blood mononuclear cells (PBMCs) in a group of squirrel monkeys that were transported by road for approximately 10 hours from one facility to another. Using a panel of human antibodies and a set of standardized human immune assays, we evaluated the phenotype of lymphocyte subsets by flow, mitogen-specific immune responses of PBMCs in vitro, and levels of cytokines at various time points including immediately before transport, immediately upon arrival, and after approximately 150 days of acclimation. We observed significant changes in T cells and subsets, NK and B cells (CD4+, CD8+, CD4+/CD8+, CD16+, and CD20+). Mitogen specific (e.g. PHA, PWM and LPS) proliferation responses, IFN-g by ELISPOT assay, and cytokines (IL-2, IL-4 and VEGF) significant changes were observed. Changes seen in the serum chemistry measurements mostly complement those seen in the hematology data. The specific goal was to empirically assess the effects of relocation stress in squirrel monkeys in terms of changes in the numbers and functions of various leukocyte subsets in the blood and the amount of time require for acclimating to their new environment. Such data will help to determine when newly arrived animals become available for use in research studies.
Angulin-1 seals tricellular contacts independently of tricellulin and claudins
Authors: Sugawara Taichi , Furuse Kyoko , Otani Tetsuhisa , Furuse Mikio
Published: October 2020

Tricellular tight junctions (tTJs) are specialized tight junctions (TJs) that seal the intercellular space at tricellular contacts (TCs), where the vertices of three epithelial cells meet. Tricellulin and angulin family membrane proteins are known constituents of tTJs, but the molecular mechanism of tTJ formation remains elusive. Here, we investigated the roles of angulin-1 and tricellulin in tTJ formation in MDCK II cells by genome editing. Angulin-1-deficient cells lost the plasma membrane contact at TCs with impaired epithelial barrier function. The COOH-terminus of angulin-1 bound to the TJ scaffold protein ZO-1 and disruption of their interaction influenced the localization of claudins at TCs, but not the tricellular sealing. Strikingly, the plasma membrane contact at TCs was formed in tricellulin- or claudin-deficient cells. These findings demonstrate that angulin-1 is responsible for the plasma membrane seal at TCs independently of tricellulin and claudins.
Breeding and hibernation of captive meadow jumping mice (Zapus hudsonius)
Authors: Brem Ethan A. , McNulty Alyssa D. , Israelsen William J.
Published: October 2020

Hibernating mammals exhibit unique metabolic and physiological phenotypes that have potential applications in medicine or spaceflight, yet our understanding of the genetic basis and molecular mechanisms of hibernation is limited. The meadow jumping mouse, a small North American hibernator, exhibits traits – including a short generation time – that would facilitate genetic approaches to hibernation research. Here we report the collection, captive breeding, and laboratory hibernation of meadow jumping mice. Captive breeders in our colony produced a statistically significant excess of male offspring and a large number of all-male and all-female litters. We confirmed that short photoperiod induced pre-hibernation fattening, and cold ambient temperature facilitated entry into hibernation. During pre-hibernation fattening, food consumption exhibited non-linear dependence on both body mass and temperature, such that food consumption was greatest in the heaviest animals at the coldest temperatures. Meadow jumping mice exhibited a strong circadian rhythm of nightly activity that was disrupted during the hibernation interval. We quantified the length and timing of torpor bouts and arousals obtained from an uninterrupted recording of a hibernating female. Over a 90.6 day hibernation interval, torpor bouts ranged from 2.1 to 12.8 days (mean 7.7 days), and arousal length was relatively constant with a mean length of 9.6 hours. We conclude that it is possible to study hibernation phenotypes using captive-bred meadow jumping mice in a laboratory setting.
The nuclear receptor RORα preserves cardiomyocyte mitochondrial function by regulating mitophagy through caveolin-3
Authors: Beak JY , Kang HS , Huang W , Aghajanian A , Gerrish K , Jetten AM , Jensen BC
Published: October 2020

Preserving optimal mitochondrial function is critical in the heart, which is the most ATP-avid organ in the body. Recently, we showed that global deficiency of the nuclear receptor RORα in the “staggerer” (RORαsg/sg) mouse exacerbates angiotensin II-induced cardiac hypertrophy and compromises cardiomyocyte mitochondrial function. The mechanisms underlying these observations have not been defined. Here we present evidence that RORα regulates cardiomyocyte mitophagy using pharmacological and genetic gain- and loss-of-function tools, including a novel cardiomyocyte-specific RORα knockout mouse. Cardiomyocyte RORα is upregulated by hypoxia and the loss of RORα blunts hypoxia-induced mitophagy and broadly compromises mitochondrial function. We show that RORα is a direct transcriptional regulator of the mitophagy mediator caveolin-3 in cardiomyocytes and that increased expression of RORα increases caveolin-3 abundance and enhances mitophagy. Knockdown of RORα impairs cardiomyocyte mitophagy, but this defect can be rescued by caveolin-3 overexpression. Collectively, these findings reveal a novel role for RORα in regulating mitophagy through caveolin-3 and expand our currently limited understanding of the mechanisms underlying RORα-mediated cardioprotection.
ROS-dependent innate immune mechanisms controlStaphylococcus aureusMRSA virulence in theDrosophilalarval model
Authors: Ramond Elodie , Jamet Anne , Ding Xiongqi , Bouvier Clémence , Lallemant Louison , He Xiangyan , Arbibe Laurence , Coureuil Mathieu , Charbit Alain
Published: October 2020

Antibiotics multi-resistantStaphylococcus aureusstrains constitute a major public health concern worldwide and are responsible of both healthcare- and community-associated infections. Here we have established a robust and simpleS. aureusoral infection model, usingDrosophila melanogasterlarva, which allowed to followS. aureusfate at the whole organism level as well as the host immune responses. Fluorescence microscopy and Light sheet 3D imaging revealed bacterial clustering at the posterior midgut that displays neutral pH. Our study demonstrates thatS. aureusinfection triggers host H2O2production through Duox enzyme, consequently empowering antimicrobial peptides production through Toll pathway activation. We also show that catalase-mediated quenching of H2O2not only enhancesS. aureussurvival but also minimizes host antimicrobial response, hence reducing bacterial clearancein vivo.Finally, we confirm the versatility of this model by demonstrating the colonization and host stimulation capacities of two other bacterial pathogens:SalmonellaTyphimurium andShigella flexneri.Overall, the drosophila larva may constitute a general model to followin vivohost innate immune responses triggered upon infection with human pathogens.
Neural tube patterning: from a minimal model for rostrocaudal patterning towards an integrated 3D model
Authors: Brambach Max , Ernst Ariane , Nolbrant Sara , Drouin-Ouellet Janelle , Kirkeby Agnete , Parmar Malin , Olariu Victor
Published: October 2020

The rostrocaudal patterning of the neural tube is a key event in early brain development. This process is mainly driven by a gradient of WNT, which defines the fate of the present neural progenitor cells in a dose dependent matter and leads to a subdivision of the tube into forebrain, midbrain and hindbrain. Although this process is extensively studied experimentally bothin vivoandin vitro, an integrated view of the responsible genetic circuitry is currently lacking. In this work, we present a minimal gene regulatory model for rostrocaudal neural tube patterning. The model's nodes and architecture are determined in a data driven way, leading to a tristable configuration of mutually repressing brain regions. Analysis of the parameter sensitivity and simulations of knockdown and overexpression cases show that repression of hindbrain fate is a promising strategy for the improvement of current protocols for the generation of dopaminergic neuronsin vitro. Furthermore, we combine the model with an existing model for dorsoventral neural tube patterning, to test its capabilities in anin vivosetting, by predicting the steady state pattern of a realistic three-dimensional neural tube. This reveals that the rostrocaudal pattern stacks dorsoventrally in the caudal half of the neural tube. Finally, we simulate morphogen secretion overexpression, which highlights the sensitivity of neural tube patterning to the morphogen levels.
Intestinal microbiota modulates adrenomedullary response through Nod1 sensing in chromaffin cells
Authors: Xiang Chen , Chen Peihua , Zhang Qin , Li Yinghui , Pan Ying , Xie Wenchun , Sun Jianyuan , Liu Zhihua
Published: October 2020

The intestinal microbiota closely interacts with the neuroendocrine system and exerts profound effects on host physiology. Many of the molecular mechanisms underlying these interactions await discovery. Here we report that Nod1 ligand derived from intestinal bacteria directly modulates catecholamine storage and secretion in mouse adrenal chromaffin cells. The cytosolic peptidoglycan receptor Nod1 is involved in chromogranin A (CHGA) retention in dense core granules (DCGs) in chromaffin cells in a cell-autonomous manner. Mechanistically, upon recognizing its cognate ligands, Nod1 localizes to DCGs, and recruits Rab2a, which is critical for CHGA retention in DCGs. Loss of Nod1 ligands leads to a profound defect in epinephrine storage in chromaffin cells, and subsequently, less secretion upon stimulation. The intestine-adrenal medulla crosstalk bridged by Nod1 ligands modulates adrenal medullary responses during the immobilization-induced stress response in mice. Thus, our study uncovers a mechanism by which intestinal microbes directly modulate a major pathway in response to stress, which may provide further understanding of the gut-brain axis.
Enhancement of synaptic AMPA receptors depends mutually on Src and PSD-95
Authors: Huang Xiaojie , Krüger Juliane M. , Beroun Anna , Xu Weifeng , Dong Yan , Schlüter Oliver M.
Published: October 2020

Synaptic incorporation and removal of AMPA receptors is highly regulated to modulate the strength of synaptic transmission for long-term synaptic plasticity during brain development and associative learning. PSD-93α2 and PSD-95α, two paralogs of the DLG-MAGUK protein family of signaling scaffolds govern the synaptic incorporation and stabilization of AMPA receptors opposingly, with PSD-95α promoting and PSD-93α2 inhibiting it. The associated signaling mechanisms that control the synaptic incorporation and stabilization remain elusive. Here, we used domain swapping between the antagonizing signaling scaffolds to identify the protein motifs responsible for enhancing synaptic AMPA receptors and the associated signaling protein. We narrowed down multiple motifs in the N-terminal domain that are principally responsible for governing the enhancement by Src. Specific activation and inhibiting peptides revealed continuous activity of Src. Together, the results depict a mutual dependence of Src and PSD-95α in enhancing and maintaining synaptic AMPA receptors.
ER-embedded UBE2J1/RNF26 ubiquitylation complex in spatiotemporal control of the endolysosomal pathway
Authors: Cremer Tom , Jongsma Marlieke L.M. , Trulsson Fredrik , Vertegaal Alfred C.O. , Neefjes Jacques J.C. , Berlin Ilana
Published: October 2020

The endolysosomal system fulfills a wide variety of cellular functions, many of which are modulated through interactions with other organelles. In particular, the ER exerts spatiotemporal constraints on the organization and motility of endosomes and lysosomes. We have recently described the ER transmembrane E3 ubiquitin ligase RNF26 to control perinuclear positioning and transport dynamics of the endolysosomal vesicular network. We now report that the ubiquitin conjugating enzyme UBE2J1, also anchored in the ER membrane, collaborates with RNF26 in this context, and that the cellular activity of this E2/E3 pair, localized in a perinuclear ER subdomain, is underpinned by transmembrane interactions. Through modification of its substrate SQSTM1/p62, the ER-embedded UBE2J1/RNF26 ubiquitylation complex recruits endosomal adaptors to immobilize their cognate vesicles in the perinuclear region. The resulting spatiotemporal compartmentalization of the endolysosomal system between the perinuclear vesicle cloud and the cell periphery facilitates timely downregulation of endocytosed cargoes, such as EGFR.
Trophectoderm differentiation to invasive syncytiotrophoblast is induced by endometrial epithelial cells during human embryo implantation
Authors: Ruane Peter T , Garner Terence , Parsons Lydia , Babbington Phoebe A , Kimber Susan J , Stevens Adam , Westwood Melissa , Brison Daniel R , Aplin John D
Published: October 2020

At implantation, trophoblast derived from the trophectoderm of the blastocyst-stage embryo invades the endometrium to establish pregnancy. To understand how embryos breach the endometrial epithelium, we modelled human implantation using blastocysts or trophoblast stem cell spheroids cultured with endometrial epithelial cells (EEC). Blastocyst invasion of the EEC layer was initiated by multinuclear syncytiotrophoblast. Spheroids also invaded the epithelium with syncytiotrophoblast, and EEC induced upregulation of syncytiotrophoblast markers. Modelling implantation in silico using blastocyst and EEC transcriptomes revealed gene networks that exhibited greater connectivity and organisation in trophectoderm of the polar region of the embryonic axis. However, gene ontologies and machine learning suggested that EEC drives syncytiotrophoblast differentiation in polar and mural trophectoderm. This is the first evidence for endometrial epithelium-induced trophectoderm differentiation to invasive syncytiotrophoblast as the cellular mechanism of embryonic breaching of the endometrium in humans, with implications for reproductive medicine and our understanding of human embryonic development.
Structural basis for recruitment of the CHK1 DNA damage kinase by the CLASPIN scaffold protein
Authors: Day Matthew , Morris Sarah P. , Houghton-Gisby Jack , Oliver Antony W. , Pearl Laurence H.
Published: October 2020

CHK1 is a protein kinase that functions downstream of activated ATR to phosphorylate multiple targets as part of intra-S and G2/M DNA damage checkpoints. Its role in allowing cells to survive replicative stress has made it an important target for anti-cancer drug discovery. Activation of CHK1 by ATR depends on their mutual interaction with CLASPIN – a natively unstructured protein that interacts with CHK1 through a cluster of phosphorylation sites in its C-terminal half. We have now determined the crystal structure of the kinase domain of CHK1 bound to a high-affinity motif from CLASPIN. Our data show that CLASPIN engages a conserved site on CHK1 adjacent to the substrate-binding cleft, involved in phosphate sensing in other kinases. The CLASPIN motif is not phosphorylated by CHK1, nor does it affect phosphorylation of a CDC25 substrate peptide, suggesting that it functions purely as a scaffold for CHK1 activation by ATR.
Human hypertrophic cardiomyopathy mutation R712L suppresses the working stroke of cardiac myosin and can be rescued by omecamtiv mecarbil
Authors: Snoberger Aaron , Barua Bipasha , Atherton Jennifer L. , Shuman Henry , Forgacs Eva , Goldman Yale E. , Winkelmann Donald A. , Ostap E. Michael
Published: October 2020

Hypertrophic cardiomyopathies (HCMs) are the leading cause of acute cardiac failure in young individuals. Over 300 mutations throughout β-cardiac myosin, including in the motor domain, are associated with HCM. A β-cardiac myosin motor mutation (R712L) leads to a severe form of HCM. Actin-gliding motility of R712L-myosin is inhibited, despite near normal ATPase kinetics. By optical trapping, the working stroke of R712L-myosin was decreased 4-fold, but actin-attachment durations were normal. A prevalent hypothesis that HCM mutants are hypercontractile is thus not universal. R712 is adjacent to the binding site of the heart failure drug omecamtiv mecarbil (OM). OM suppresses the working stroke of normal β-cardiac myosin, but remarkably, OM rescues the R712L-myosin working stroke. Using a flow chamber to interrogate a single molecule during buffer exchange, we found OM rescue to be reversible. Thus, the R712L mutation uncouples lever arm rotation from ATPase activity and this inhibition is rescued by OM.
Fine scale sampling unveils diazotroph patchiness in the South Pacific Ocean
Authors: Benavides Mar , Conradt Louis , Bonnet Sophie , Berman-Frank Ilana , Barrillon Stéphanie , Petrenko Anne , Doglioli Andrea M.
Published: October 2020

Diazotrophs are important contributors to reactive nitrogen availability in the ocean. Oceanographic cruise data accumulated along decades has revealed a heterogeneous distribution of diazotroph species at regional to global scales. However, the role of dynamic fine scale structures in distributing diazotrophs is not well understood. This is due to typical insufficient spatiotemporal resolution sampling and the lack of detailed physical studies in parallel. Here we show the distribution of five groups of diazotrophs in the South Pacific at an unprecedented resolution of 7-16 km. We find a patchy distribution of diazotrophs, with each group being differently affected by parameters describing fine scale structures. The observed variability could not have been revealed with a lower resolution sampling, highlighting the need to consider fine scale physics to resolve the distribution of diazotrophs in the ocean.
Abstract neural choice signals during action-linked decisions
Authors: Sandhaeger Florian , Omejc Nina , Pape Anna-Antonia , Siegel Markus
Published: October 2020

Humans can make abstract choices independent of motor actions. However, in laboratory tasks, choices are typically reported with an associated action. Consequentially, knowledge about the neural representation of abstract choices is sparse, and choices are often thought to evolve as motor intentions. Here, we show that in the human brain, perceptual choices are represented in an abstract, motor-independent manner, even when they are directly linked to an action. We measured MEG signals while participants made choices with known or unknown motor response mapping. Using multivariate decoding, we quantified stimulus, perceptual choice and motor response information with distinct cortical distributions. Choice representations were invariant to whether the response mapping was known during stimulus presentation, and they occupied distinct representational spaces from both stimulus and motor signals. Furthermore, their strength predicted decision confidence and accuracy, as expected from an internal decision variable. Our results uncover abstract neural choice signals that generalize to action-linked decisions, suggesting a general role of an abstract choice stage in human decision-making.
LRRK2-dependent Rab GTPase phosphorylation in response to endolysosomal damage depends on macrophage differentiation
Authors: Herbst Susanne , Gutierrez Maximiliano G.
Published: October 2020

The Parkinson’s Disease (PD) kinase LRRK2 is highly expressed in immune cells such as macrophages. In these cells, LRRK2 regulates innate immune pathways and it is activated after membrane damage leading to the phosphorylation of the Rab GTPases Rab8A and Rab10. Due to their wide-range of functions in immunity and tissue remodelling, macrophagesin vivoare phenotypically heterogeneous.In vitrosystems are used to differentiate these cells into diverse macrophage subsets to mimic the populations observedin vivo. M-CSF and GM-CSF differentiated human blood monocytes are often used to generate monocyte-derived macrophages as a model for tissue macrophages. However, how LRRK2 is activated in different macrophage subsets after membrane damage is unknown.

Here, we report that bone marrow derived macrophages and human monocyte-derived macrophages differentiated with either M-CSF or GM-CSF show different levels of LRRK2 activation after membrane damage. Notably, the membrane damaging agent LLOMe triggered LRRK2-dependent Rab8A and Rab10 phosphorylation primarily in GM-CSF differentiated macrophages. Moreover, LRRK2 and Rab8A were recruited to damaged endolysosomes in GM-CSF differentiated macrophages. Strikingly, GM-CSF differentiated macrophages recruited significantly more CHMP4B and Galectin-3 into damaged endolysosomes. These results suggest that LRRK2-regulated pathways of endolysosomal membrane damage and repair differ between macrophage subsets.
A Multiscale Coarse-grained Model of the SARS-CoV-2 Virion
Authors: Yu Alvin , Pak Alexander J. , He Peng , Monje-Galvan Viviana , Casalino Lorenzo , Gaieb Zied , Dommer Abigail C. , Amaro Rommie E. , Voth Gregory A.
Published: October 2020

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the COVID-19 pandemic. Computer simulations of complete viral particles can provide theoretical insights into large-scale viral processes including assembly, budding, egress, entry, and fusion. Detailed atomistic simulations, however, are constrained to shorter timescales and require billion-atom simulations for these processes. Here, we report the current status and on-going development of a largely “bottom-up” coarse-grained (CG) model of the SARS-CoV-2 virion. Structural data from a combination of cryo-electron microscopy (cryo-EM), x-ray crystallography, and computational predictions were used to build molecular models of structural SARS-CoV-2 proteins, which were then assembled into a complete virion model. We describe how CG molecular interactions can be derived from all-atom simulations, how viral behavior difficult to capture in atomistic simulations can be incorporated into the CG models, and how the CG models can be iteratively improved as new data becomes publicly available. Our initial CG model and the detailed methods presented are intended to serve as a resource for researchers working on COVID-19 who are interested in performing multiscale simulations of the SARS-CoV-2 virion.

Significance Statement

This study reports the construction of a molecular model for the SARS-CoV-2 virion and details our multiscale approach towards model refinement. The resulting model and methods can be applied to and enable the simulation of SARS-CoV-2 virions.
The TRIM9/TRIM67 neuronal interactome reveals novel activators of morphogenesis
Authors: Menon Shalini , Goldfarb Dennis , Ho Tsungyo , Cloer Erica W. , Boyer Nicholas P. , Hardie Christopher , Bock Andrew J. , Johnson Emma C. , Anil Joel , Major M. Ben , Gupton Stephanie L.
Published: October 2020

TRIM9 and TRIM67 are neuronally-enriched E3 ubiquitin ligases essential for appropriate morphogenesis of cortical and hippocampal neurons and fidelitous responses to the axon guidance cue netrin-1. Deletion of murineTrim9orTrim67results in neuroanatomical defects and striking behavioral deficits, particularly in spatial learning and memory. TRIM9 and TRIM67 interact with cytoskeletal and exocytic proteins, but the full interactome is not known. Here we performed the unbiased proximity-dependent biotin identification (BioID) approach to define TRIM9 and TRIM67 protein-protein proximity network in developing cortical neurons and identified neuronal putative TRIM interaction partners. Candidates included cytoskeletal regulators, cytosolic protein transporters, exocytosis and endocytosis regulators, and proteins necessary for synaptic regulation. A subset of high priority candidates was validated, including Myo16, Coro1A, SNAP47, ExoC1, GRIP1, PRG-1, and KIF1A. For a subset of validated candidates, we utilized TIRF microscopy to demonstrate dynamic colocalization with TRIM proteins at the axonal periphery, including at the tips of filopodia. Further analysis demonstrated the RNAi-based knockdown of the unconventional myosin Myo16 in cortical neurons altered axonal branching patterns in a TRIM9 and netrin-1 dependent manner. Future analysis of other validated candidates will likely identify novel proteins and mechanisms by which TRIM9 and TRIM67 regulate neuronal form and function.
White matter microstructural changes in short-term learning of a continuous visuomotor sequence
Authors: Tremblay Stefanie A. , Jager Anna-Thekla , Huck Julia , Giacosa Chiara , Beram Stephanie , Schneider Uta , Grahl Sophia , Villringer Arno , Tardif Christine L. , Bazin Pierre-Louis , Steele Christopher J. , Gauthier Claudine J.
Published: October 2020

Efficient neural transmission is crucial for optimal brain function, yet the plastic potential of white matter (WM) has long been overlooked. Growing evidence now shows that modifications to axons and myelin occur not only as a result of long-term learning, but also after short training periods. Motor sequence learning (MSL), a common paradigm used to study neuroplasticity, occurs in overlapping learning stages and different neural circuits are involved in each stage. However, most studies investigating short-term WM plasticity have used a pre-post design, in which the temporal dynamics of changes across learning stages cannot be assessed. In this study, we used multiple magnetic resonance imaging (MRI) scans at 7 Tesla to investigate changes in WM in a group learning a complex visuomotor sequence (LRN) and in a control group (SMP) performing a simple sequence, for 5 consecutive days. Consistent with behavioral results, where most improvements occurred between the two first days, structural changes in WM were observed only in the early phase of learning (d1-d2), and in overall learning (d1-d5). In LRNs, WM microstructure was altered in the tracts underlying the primary motor and sensorimotor cortices. Moreover, our structural findings in WM were related to changes in functional connectivity, assessed with resting-state functional MRI data in the same cohort, through analyses in regions of interest (ROIs). Significant changes in WM microstructure were found in a ROI underlying the right supplementary motor area. Together, our findings provide evidence for highly dynamic WM plasticity in the sensorimotor network during short-term MSL.
Accurate Microbiome Sequencing with Synthetic Long Read Sequencing
Authors: Chung Nico , Van Goethem Marc W. , Preston Melanie A. , Lhota Filip , Cerna Leona , Garcia-Pichel Ferran , Fernandes Vanessa , Giraldo-Silva Ana , Kim Hee Shin , Hurowitz Evan , Balamotis Michael , Wu Indira , Ben-Yehezkel Tuval
Published: October 2020

The microbiome plays a central role in biochemical cycling and nutrient turnover of most ecosystems. Because it can comprise myriad microbial prokaryotes, eukaryotes and viruses, microbiome characterization requires high-throughput sequencing to attain an accurate identification and quantification of such co-existing microbial populations. Short-read next-generation-sequencing (srNGS) revolutionized the study of microbiomes and remains the most widely used approach, yet read lengths spanning only a few of the nine hypervariable regions of the 16S rRNA gene limit phylogenetic resolution leading to misclassification or failure to classify in a high percentage of cases. Here we evaluate a synthetic long-read (SLR) NGS approach for full-length 16S rRNA gene sequencing that is high-throughput, highly accurate and low-cost. The sequencing approach is amenable to highly multiplexed sequencing and provides microbiome sequence data that surpasses existing short and long-read modalities in terms of accuracy and phylogenetic resolution. We validated this commercially-available technology, termed LoopSeq, by characterizing the microbial composition of well-established mock microbiome communities and diverse real-world samples. SLR sequencing revealed differences in aquatic community complexity associated with environmental gradients, resolved species-level community composition of uterine lavage from subjects with histories of misconception and accurately detected strain differences, multiple copies of the 16S rRNA in a single strain’s genome, as well as low-level contamination in soil cyanobacterial cultures. This approach has implications for widespread adoption of high-resolution, accurate long-read microbiome sequencing as it is generated on popular short read sequencing platforms without the need for additional infrastructure.
Safe and effective two-in-one replicon-and-VLP minispike vaccine for COVID-19
Authors: Hennrich Alexandru A. , Banda Dominic H. , Oberhuber Martina , Schopf Anika , Pfaffinger Verena , Wittwer Kevin , Sawatsky Bevan , Riedel Christiane , Pfaller Christian K. , Conzelmann Karl-Klaus
Published: October 2020

The large SARS-CoV-2 spike (S) protein is the main target of current COVID-19 vaccine candidates but can induce non-neutralizing antibodies, which may cause vaccination-induced complications or enhancement of COVID-19 disease. Besides, encoding of a functional S in replication-competent virus vector vaccines may result in the emergence of viruses with altered or expanded tropism. Here, we have developed a safe single round rhabdovirus replicon vaccine platform for enhanced presentation of the S receptor-binding domain (RBD). Structure-guided design was employed to build a chimeric minispike comprising the globular RBD linked to a transmembrane stem-anchor sequence derived from rabies virus (RABV) glycoprotein (G). Vesicular stomatitis virus (VSV) and RABV replicons encoding the minispike not only allowed expression of the antigen at the cell surface but also incorporation into the envelope of secreted non-infectious particles, thus combining classic vector-driven antigen expression and particulate virus-like particle (VLP) presentation. A single dose of a prototype replicon vaccine, VSVΔG-minispike-eGFP (G), stimulated high titers of SARS-CoV-2 neutralizing antibodies in mice, equivalent to those found in COVID-19 patients. Boost immunization with the identical replicon further enhanced neutralizing activity. These results demonstrate that rhabdovirus minispike replicons represent effective and safe alternatives to vaccination approaches using replication-competent viruses and/or the entire S antigen.

Highlights

SARS-CoV-2 S RBD antigen is preferred over entire S to preclude potential disease enhancing antibodies

construction of a chimeric rhabdovirus minispike protein presenting RBD in native conformation

construction of single round VSV and rabies virus replicon vaccines

presentation of minispike antigen on cells and on noninfectious VLPs

strong induction of SARS-CoV-2 neutralizing antibodies by the VSV replicon/VLP system in vaccinated mice
A Generative Approach toward Precision Antimicrobial Peptide Design
Authors: Ferrell Jonathon B. , Remington Jacob M. , Van Oort Colin M. , Sharafi Mona , Aboushousha Reem , Janssen-Heininger Yvonne , Schneebeli Severin T. , Wargo Matthew J. , Wshah Safwan , Li Jianing
Published: October 2020

As the emergence of bacterial resistance is outpacing the development of new antibiotics, we must find cost-effective and innovative approaches to discover new antibacterial therapeutics. Antimicrobial peptides (AMPs) represent one promising solution to fill this void, since they generally undergo faster development, display rapid onsets of killing, and most importantly, show lower risks of induced resistance. Despite prior success in AMP design with physics- and/or knowledge-based approaches, an efficient approach to precisely design peptides with high activity and selectivity is still lacking. Toward this goal, we have invented a novel approach which utilizes a generative model to predict AMP-like sequences, followed by molecular modeling to rank the candidates. Thus, we can identify peptides with desirable sequences, structures, and potential specific interactions with bacterial membranes. For the proof of concept, we curated a dataset that comprises 500,000 non-AMP peptide sequences and nearly 8,000 labeled AMP sequences to train the generative model. For 12 generated peptides that are cationic and likely helical, we assessed the membrane binding propensity via extensive all-atom molecular simulations. The top six peptides were promoted for synthesis, chemical characterizations, and antibacterial assays, showing various inhibition to bacterial growth. Three peptides were validated with broad-spectrum antibacterial activity. In aggregate, the combination of AMP generator and sophisticated molecular modeling affords enhanced speed and accuracy in AMP design. Our approach and results demonstrate the viability of a generative approach to develop novel AMPs and to help contain the rise of antibiotic resistant microbes.
SARS-CoV-2 infected cells present HLA-I peptides from canonical and out-of-frame ORFs
Authors: Weingarten-Gabbay Shira , Klaeger Susan , Sarkizova Siranush , Pearlman Leah R. , Chen Da-Yuan , Bauer Matthew R. , Taylor Hannah B. , Conway Hasahn L. , Tomkins-Tinch Christopher H. , Finkel Yaara , Nachshon Aharon , Gentili Matteo , Rivera Keith D. , Keskin Derin B. , Rice Charles M. , Clauser Karl R. , Hacohen Nir , Carr Steven A. , Abelin Jennifer G. , Saeed Mohsan , Sabeti Pardis C.
Published: October 2020

T cell-mediated immunity may play a critical role in controlling and establishing protective immunity against SARS-CoV-2 infection; yet the repertoire of viral epitopes responsible for T cell response activation remains mostly unknown. Identification of viral peptides presented on class I human leukocyte antigen (HLA-I) can reveal epitopes for recognition by cytotoxic T cells and potential incorporation into vaccines. Here, we report the first HLA-I immunopeptidome of SARS-CoV-2 in two human cell lines at different times post-infection using mass spectrometry. We found HLA-I peptides derived not only from canonical ORFs, but also from internal out-of-frame ORFs in Spike and Nucleoprotein not captured by current vaccines. Proteomics analyses of infected cells revealed that SARS-CoV-2 may interfere with antigen processing and immune signaling pathways. Based on the endogenously processed and presented viral peptides that we identified, we estimate that a pool of 24 peptides would provide one or more peptides for presentation by at least one HLA allele in 99% of the human population. These biological insights and the list of naturally presented SARS-CoV-2 peptides will facilitate data-driven selection of peptides for immune monitoring and vaccine development.
Heat Stress Interferes with Formation of Double-Strand Breaks and Homolog Synapsis inArabidopsis thaliana
Authors: Ning Yingjie , Liu Qingpei , Wang Chong , Qin Erdai , Wu Zhihua , Wang Minghui , Yang Ke , Elesawi Ibrahim Eid , Chen Chunli , Liu Hong , Qin Rui , Liu Bing
Published: October 2020

Meiotic recombination (MR) drives novel combination of alleles and contributes to genomic diversity in eukaryotes. In this study, we showed that heat stress (36-38°C) over fertile threshold fully abolished crossover (CO) formation in Arabidopsis. Cytological and genetic studies in wild-type plants, and thesyn1andrad51mutants suggested that heat stress reduces generation of SPO11-dependent double-strand breaks (DSBs). In support, the abundance of recombinase DMC1, which is required for MR-specific DSB repair, was significantly reduced under heat stress. In addition, we showed that high temperatures induced disassembly and/or instability of ASY4-but not SYN1-mediated chromosome axis. At the same time, ASY1-associated lateral element of synaptonemal complex (SC) was partially affected, while the ZYP1-dependent central element of SC was disrupted, indicating that heat stress impairs SC formation. Moreover, quantitative RT-PCR revealed that genes involved in DSB formation; e.g.SPO11-1,PRD1,2and3, were not impacted; however, recombinaseRAD51and chromosome axis factorsASY3andASY4were significantly downregulated under heat stress. Taken together, these findings revealed that heat stress inhibits MR via compromised DSB formation and homolog synapsis, which are possible downstream effects of the impacted chromosome axis. Our study thus provides evidence shedding light on how increase of environmental temperature influences MR in Arabidopsis.
Genome-Wide CRISPR Screen Identifies Non-Canonical NF-κB Signaling as a Potent Regulator of Density-dependent Proliferation
Authors: Fomicheva Maria , Macara Ian G.
Published: October 2020

Epithelial cells possess intrinsic mechanisms to maintain an appropriate cell density for normal tissue morphogenesis and homeostasis. Defects in such mechanisms likely contribute to hyperplasia and cancer initiation. To identify genes that regulate the density-dependent proliferation of murine mammary epithelial cells, we developed a fluorescence-activated cell sorting assay based on FUCCI, which marks different stages of the cell cycle with distinct fluorophores. Using this powerful assay, we performed a genome-wide CRISPR/Cas9 knockout screen, selecting for cells that proliferate normally at low density but continue to divide at high density. Unexpectedly, one top hit wasTraf3, a negative regulator of NF-κB signaling that has never previously been linked to density-dependent proliferation. We demonstrate that loss ofTraf3specifically activates non-canonical NF-κB signaling. This in turn triggers an innate immune response and drives cell division independently of known density-dependent proliferation mechanisms, including YAP/TAZ signaling and cyclin kinase inhibitors, by blocking entry into quiescence.
A study of adaptive gambling behaviors of visitors from Mainland China to Macau
Authors: Zeng Zhonglu , Wang Xing , Wong IpKin Anthony
Published: October 2020

Purpose

The adaptation hypothesis suggests that gambling participation would gradually decline after an initial exposure to this activity. While this hypothesis was tested in pathological gambling among residents in Western countries, the present inquiry explores the hypothesis in a tourism context.

Design/methodology/approach

This research is focused on the Mainland Chinese gamblers. Convenience sampling was used. Data were collected outside participating casinos and at major attractions. A total of 498 valid responses were collected.

Findings

By assessing changes of the Mainland Chinese gambling perceptions (e.g. excitement and fallacy) and behaviors, results point to visitor gamblers' decrease in gambling excitement and fallacy as well as budget to income ratio.

Originality/value

By assessing changes of the Mainland Chinese gambling perceptions and behaviors, this research aims to contribute to the literature by demonstrating whether the Chinese gamblers have adapted and hence, are more rational about this recreational activity.