Biology subjects

Wu, G.

Publications and source records attributed to Wu, G..

At least 145 records · Page 8Linked to original sources

bioRxiv2021

Machine learning approaches to identify sleep genes

Genetics impacts sleep, yet, the molecular mechanisms underlying sleep regulation remain elusive. We built machine learning (ML) models to predict genes based on their similarity to known sleep genes. Using a manually curated list of 109 labeled sleep genes, we trained a prediction model on thousands of published datasets, representing circadian, immune, sleep deprivation, and many other processes. Our predictions fit with prior knowledge of sleep regulation and also identify several key genes/pathways to pursue in follow-up studies. We tested one of our findings, the NF-{kappa}B pathway, and showed that its genetic alteration affects sleep duration in mice. Our study highlights the power of ML to integrate prior knowledge and genome-wide data to study genetic regulation of sleep and other complex behaviors.

genomics↗

bioRxiv2021

Lack of Tgfbr1 and Acvr1b synergistically stimulates myofibre hypertrophy and accelerates muscle regeneration

In skeletal muscle, transforming growth factor-{beta} (TGF-{beta}) family growth factors, TGF-{beta}1 and myostatin, are involved in atrophy and muscle wasting disorders. Simultaneous interference with their signalling pathways may improve muscle function, however little is known about their individual and combined receptor signalling. Here we show that inhibition of TGF-{beta} signalling by simultaneous muscle-specific knockout of TGF-{beta} type I receptors Tgfbr1 and Acvr1b simultaneously, induces substantial hypertrophy, while such effect does not occur by single receptor knockout. Hypertrophy is induced by increased phosphorylation of Akt and p70S6K and reduced E3 ligases expression, while myonuclear number remains unaltered. Combined knockout of both TGF-{beta} type I receptors increases the number of satellite cells, macrophages and improves regeneration post cardiotoxin-induced injury by stimulating myogenic differentiation. Extra cellular matrix gene expression is exclusively elevated in muscle with combined receptor knockout. Tgfbr1 and Acvr1b are synergistically involved in regulation of myofibre size, regeneration and collagen deposition.

physiology↗

bioRxiv2021

Segmentor: A tool for manual refinement of 3D microscopy annotations

BackgroundRecent advances in tissue clearing techniques, combined with high-speed image acquisition through light sheet microscopy, enable rapid three-dimensional (3D) imaging of biological specimens, such as whole mouse brains, in a matter of hours. Quantitative analysis of such 3D images can help us understand how changes in brain structure lead to differences in behavior or cognition, but distinguishing features of interest, such as nuclei, from background can be challenging. Recent deep learning-based nuclear segmentation algorithms show great promise for automated segmentation, but require large numbers of manually and accurately labeled nuclei as training data. ResultsWe present Segmentor, an open-source tool for reliable, efficient, and user-friendly manual annotation and refinement of objects (e.g., nuclei) within 3D light sheet microscopy images. Segmentor employs a hybrid 2D-3D approach for visualizing and segmenting objects and contains features for automatic region splitting, designed specifically for streamlining the process of 3D segmentation of nuclei. We show that editing simultaneously in 2D and 3D using Segmentor significantly decreases time spent on manual annotations without affecting accuracy. ConclusionsSegmentor is a tool for increased efficiency of manual annotation and refinement of 3D objects that can be used to train deep learning segmentation algorithms, and is available at https://www.nucleininja.org/ and https://github.com/RENCI/Segmentor.

neuroscience↗

bioRxiv2021

Covid19Risk.ai: An open source repository and online calculator of prediction models for early diagnosis and prognosis of Covid-19

ObjectiveThe current pandemic has led to a proliferation of predictive models being developed to address various aspects of COVID-19 patient care. We aimed to develop an online platform that would serve as an open source repository for a curated subset of such models, and provide a simple interface for included models to allow for online calculation. This platform would support doctors during decision-making regarding diagnoses, prognoses, and follow-up of COVID-19 patients, expediting the models transition from research to clinical practice. MethodsIn this proof-of-principle study, we performed a literature search in PubMed and WHO database to find suitable models for implementation on our platform. All selected models were publicly available (peer reviewed publications or open source repository) and had been validated (TRIPOD type 3 or 2b). We created a method for obtaining the regression coefficients if only the nomogram was available in the original publication. All predictive models were transcribed on a practical graphical user interface using PHP 8.0.0, and published online together with supporting documentation and links to the associated articles. ResultsThe open source website https://covid19risk.ai/ currently incorporates nine models from six different research groups, evaluated on datasets from different countries. The website will continue to be populated with other models related to COVID-19 prediction as these become available. This dynamic platform allows COVID-19 researchers to contact us to have their model curated and included on our website, thereby increasing the reach and real-world impact of their work. ConclusionWe have successfully demonstrated in this proof-of-principle study that our website provides an inclusive platform for predictive models related to COVID-19. It enables doctors to supplement their judgment with patient-specific predictions from externally-validated models in a user-friendly format. Additionally, this platform supports researchers in showcasing their work, which will increase the visibility and use of their models.

bioinformatics↗

bioRxiv2020

Landscapes and dynamic diversifications of B-cell receptor repertoires in COVID-19 patients

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused the pandemic of coronavirus disease 2019 (COVID-19). Great international efforts have been put into the development of prophylactic vaccines and neutralizing antibodies. However, the knowledge about the B cell immune response induced by the SARS-CoV-2 virus is still limited. Here, we report a comprehensive characterization of the dynamics of immunoglobin heavy chain (IGH) repertoire in COVID-19 patients. By using next-generation sequencing technology, we examined the temporal changes in the landscape of the patients immunological status, and found dramatic changes in the IGH within the patients immune system after the onset of COVID-19 symptoms. Although different patients have distinct immune responses to SARS-CoV-2 infection, by employing clonotype overlap, lineage expansion and clonotype network analyses, we observed a higher clonotype overlap and substantial lineage expansion of B cell clones during 2-3 weeks of illness, which is of great importance to B-cell immune responses. Meanwhile, for preferences of V gene usage during SARS-CoV-2 infection, IGHV3-74 and IGHV4-34 and IGHV4-39 in COVID-19 patients were more abundant than that of healthy controls. Overall, we present an immunological resource for SARS-CoV-2 that could promote both therapeutic development as well as mechanistic research.

immunology↗

bioRxiv2020

Patient-Derived Orthotopic Xenografts and Cell Lines from Pediatric High-Grade Glioma Recapitulate the Heterogeneity of Histopathology, Molecular Signatures, and Drug Response

Pediatric high-grade glioma (pHGG) is a major contributor to cancer-related death in children. In vitro and in vivo disease models reflecting the intimate connection between developmental context and pathogenesis of pHGG are essential to advance understanding and identify therapeutic vulnerabilities. We established 21 patient-derived pHGG orthotopic xenograft (PDOX) models and eight matched cell lines from diverse groups of pHGG. These models recapitulated histopathology, DNA methylation signatures, mutations and gene expression patterns of the patient tumors from which they were derived, and included rare subgroups not well-represented by existing models. We deployed 16 new and existing cell lines for high-throughput screening (HTS). In vitro HTS results predicted variable in vivo response to inhibitors of PI3K/mTOR and MEK signaling pathways. These unique new models and an online interactive data portal to enable exploration of associated detailed molecular characterization and HTS chemical sensitivity data provide a rich resource for pediatric brain tumor research.

cancer biology↗

bioRxiv2020

A Novel SARS-CoV-2 Multitope Protein/Peptide Vaccine Candidate is Highly Immunogenic and Prevents Lung Infection in an Adeno Associated Virus Human Angiotensin-Converting Enzyme 2 (AAV hACE2) Mouse Model

A novel multitope protein-peptide vaccine against Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) infection and disease is described in this report. The initial development and characterization experiments are presented along with proof-of-concept studies for the vaccine candidate UB-612. UB-612 consists of eight components rationally designed for induction of potently neutralizing antibodies and broad T cell responses against SARS-CoV-2: the S1-RBD-sFc fusion protein, six synthetic peptides (one universal peptide and five SARS-CoV-2-derived peptides), a proprietary CpG TLR-9 agonist at low concentration as an excipient, and aluminum phosphate adjuvant. Through immunogenicity studies in Guinea pigs and rats, we optimized the design of protein/peptide immunogens and selected an adjuvant system, yielding a vaccine that provides excellent S1-RBD binding and high neutralizing antibody responses, robust cellular responses, and a Th1-oriented response at low doses. In challenge studies, UB- 612 vaccination reduced viral load and prevented development of disease in mouse and non-human primate challenge models. With a Phase 1 trial completed, a Phase 2 trial ongoing in Taiwan, and additional trials planned to support global authorizations, UB-612 is a highly promising and differentiated vaccine candidate for prevention of SARS-CoV-2 infection and COVID-19 disease. Author SummarySARS-CoV-2 virus, the causative agent of Coronavirus Disease 2019 (COVID-19), has spread globally since its origin in 2019, causing an unprecedented public health crisis that has resulted in greater than 4.7 million deaths worldwide. Many vaccines are under development to limit disease spread and reduce the number of cases, but additional candidates that promote a robust immune response are needed. Here, we describe a multitope protein-peptide vaccine platform that is unique among COVID-19 vaccines. The advantages of our approach are induction of both high levels of neutralizing antibodies as well as a Th/CTL response in the vaccinated host, which mimics the immune response that occurs after natural infection with SARS-CoV-2. We demonstrate that our vaccine is immunogenic and effective in preventing disease in several animal models, including AAV- hACE-2 transduced mice, and both rhesus and cynomolgus macaques. Importantly, no immunopathology was observed in the lungs of immunized animals, therefore showing that antibody-dependent enhancement (ADE) does not occur. Our study provides an additional, novel vaccine candidate for advancement in clinical trials to treat and prevent SARS-CoV-2 infection and COVID-19 disease.

immunology↗

bioRxiv2020

Pruriception and neuronal coding in nociceptor subtypes in human and nonhuman primates

In human, intradermal administration of {beta}-alanine (ALA) and bovine adrenal medulla peptide 8-22 (BAM8-22) evokes the sensation of itch. Currently, it is unknown which human dorsal root ganglion (DRG) neurons express the receptors of these pruritogens, MRGPRD and MRGPRX1 respectively, and which cutaneous afferents these pruritogens activate in primate. In situ hybridization studies revealed that MRGPRD and MRGPRX1 are co-expressed in a subpopulation of TRPV1+ human DRG neurons. In electrophysiological recordings in nonhuman primates (Macaca nemestrina), subtypes of polymodal C-fiber nociceptors are preferentially activated by ALA and BAM8-22, with significant overlap. When pruritogens ALA, BAM8-22 and histamine, that activate different subclasses of C-fiber afferents, are administered in combination, human volunteers report itch and nociceptive sensations similar to those induced by a single pruritogen. Our results provide evidence for differences in pruriceptive processing between primates and rodents, and do not support the spatial contrast theory of coding of itch and pain.

neuroscience↗

bioRxiv2020

idCOV: a pipeline for quick clade identification of SARS-CoV-2 isolates

idCOV is a phylogenetic pipeline for quickly identifying the clades of SARS-CoV-2 virus isolates from raw sequencing data based on a selected clade-defining marker list. Using a public dataset, we show that idCOV can make equivalent calls as annotated by Nextstrain.org on all three common clade systems using user uploaded FastQ files directly. Web and equivalent command-line interfaces are available. It can be deployed on any Linux environment, including personal computer, HPC and the cloud. The source code is available at https://github.com/xz-stjude/idcov. A documentation for installation can be found at https://github.com/xz-stjude/idcov/blob/master/README.md.

bioinformatics↗

bioRxiv2020

Architecture of the Tuberous Sclerosis Protein Complex

The Tuberous Sclerosis Complex (TSC) protein complex (TSCC), comprising TSC1, TSC2, and TBC1D7, is widely recognised as a key integration hub for cell growth and intracellular stress signals upstream of the mammalian target of rapamycin complex 1 (mTORC1). The TSCC negatively regulates mTORC1 by acting as a GTPase-activating protein (GAP) towards the small GTPase Rheb. Both human TSC1 and TSC2 are important tumour suppressors, and mutations in them underlie the disease tuberous sclerosis. We used single-particle cryo-EM to reveal the organisation and architecture of the complete human TSCC. We show that TSCC forms an elongated scorpion-like structure, consisting of a central "body", with a "pincer" and a "tail" at the respective ends. The "body" is composed of a flexible TSC2 HEAT repeat dimer, along the inner surface of which runs the TSC1 coiled-coil backbone, breaking the symmetry of the dimer. Each end of the body is structurally distinct, representing the N- and C-termini of TSC1; a "pincer" is formed by the highly flexible N-terminal TSC1 core domains and a barbed "tail" makes up the TSC1 coiled-coil-TBC1D7 junction. The TSC2 GAP domain is found abutting the centre of the body on each side of the dimerisation interface, poised to bind a pair of Rheb molecules at a similar separation to the pair in activated mTORC1. Our architectural dissection reveals the mode of association and topology of the complex, casts light on the recruitment of Rheb to the TSCC, and also hints at functional higher order oligomerisation, which has previously been predicted to be important for Rheb-signalling suppression.

cell biology↗

bioRxiv2020

RNA helicase DDX5 enables STAT1 mRNA translation and interferon signaling in hepatitis B virus replicating hepatocytes

ObjectiveRNA helicase DDX5 is downregulated during hepatitis B virus (HBV) replication, and poor prognosis HBV-related hepatocellular carcinoma (HCC). The objective of this study is to understand the significance of DDX5 downregulation during HBV infection and poor prognosis HCC, as it pertains to therapy of chronically HBV-infected patients with HCC. IFN- therapy is effective only for a subgroup of HBV-infected patients, for reasons not understood. Herein, we investigated a novel mechanism of STAT1 translational control, involving DDX5-mediated resolution of a G-quadruplex structure, located at the 5UTR of STAT1 mRNA, enabling STAT1 translation. Design and ResultsMolecular, pharmacologic, and biophysical assays were used together with cellular model of HBV replication, HCC cell lines, and HBV-related liver tumors. We observed, the protein level of DDX5 correlated with that of transcription factor STAT1. We show, DDX5 regulates STAT1 mRNA translation by resolving a G-quadruplex (rG4) RNA structure, proximal to the 5 end of STAT1 5UTR. We employed luciferase reporter assays comparing wild type (WT) vs. mutant (MT) rG4 sequence, rG4-stabilizing compounds, CRISPR/Cas9 editing of the rG4 sequence, and circular dichroism determination of rG4 structure. Ribonucleoprotein assays identified direct DDX5 binding to WT but not MT rG4. Importantly, liver cancer cell lines expressing low DDX5 exhibited reduced interferon response, while immunohistochemistry of HBV-related HCCs exhibiting absence of DDX5, also lacked STAT1. ConclusionDDX5 resolves a G-quadruplex structure in 5UTR of STAT1 mRNA, enabling STAT1 translation. DDX5 is a key regulator of the dynamic range of interferon response during innate immunity and adjuvant IFN- therapy.

cancer biology↗

bioRxiv2020

Changes in biochemistry and histochemical characteristics during somatic embryogenesis in Ormosia henryi Prain

Mature embryos were used as an explant for embryogenic callus (EC) induction, and then EC was further developed to form somatic embryos during somatic embryogenesis (SE) of Ormosia henryi Prain; however, some mature embryos could induced non-embryogenic callus (NEC), browning callus (BC) or snowflake callus (SC). These phenomena might be related to the biochemical and histochemical differences during somatic embryo induction. The present study was conducted to analyze the biochemical events and histochemical changes at different SE stages in 0. henryi. The contents of soluble sugar, starch, soluble protein, H2O2, and endogenous hormones and the activities of polyphenoloxidase (PPO), superoxide dismutase (SOD), ascorbate peroxidase (APX), peroxidase (POD) and catalase (CAT) were measured at different SE stages, such as EC, globular embryo (GE), and cotyledon embryo (CE), and in abnormal tissue, such as NEC, BC, and SC. The results showed that the contents of soluble sugar and starch; the activities of PPO, SOD, APX and POD; and the ratios of indole-3-acetic acid/abscisic acid (IAA/ABA), IAA/gibberellins (IAA/GAs), auxin /GAs (AUX/GAs), and AUX/ABA decreased gradually at different SE stages. In contrast, the contents of soluble protein, H2O2, all endogenous hormones gradually increased. However, CAT activity and the ratios of IAA/cytokinins (IAA/CKs), AUX/CKs, ABA/CKs, and GAs/CKs first increased and then decreased. The high contents of GAs and ABA, high ratios of ABA/CKs and GAs/CKs and low ratios of IAA/ABA, IAA/GAs, AUX/GAs and AUX/ABA were responsible for the inability of the callus to form EC. The low enzyme activities, low contents of energy substances and H2O2 were related to NEC formation. The high contents of soluble sugar, H2O2, AUX, CKs and PPO activity and the low content of soluble protein were the basic causes of BC formation. The high-energy substances contents and low activities of SOD and POD facilitated SC formation. Histochemical observation showed that starch granule staining gradually lightened with SE development, but protein granules were darkly stained. Compared with EC, starch and protein granules were stained darker in SC, and lighter in NEC and BC. These results showed that energy substances were the material basis of SE, which affected enzyme activities, regulated reactive oxygen species (ROS) metabolism, and thus regulated the morphogenesis and development of somatic embryos. In addition, the contents and ratios of endogenous hormones affected the dedifferentiation, dedifferentiation and embryogenesis of somatic cells. To induce EC from mature embryos and further develop their formation into somatic embryos, it is necessary to adjust the energy supply and hormone ratio in the medium. One-sentence summarySomatic embryogenesis and abnormal callus tissues formation of Ormosia henryi Prain were associated with energy substances, reactive oxygen species, enzyme activities and endogenous hormones, as well as histochemical characteristics.

biochemistry↗

bioRxiv2020

Molecular basis for cysteine oxidation by Plant Cysteine Oxidases from Arabidopsis thaliana

Plant Cysteine Oxidases (PCOs) play important roles in controlling the stability of Group VII ethylene response factors (ERF-VIIs) via N-Arg/degron pathway through catalyzing the oxidation of their N-Cys for subsequent Arginyl-tRNA--protein transferase 1 (ATE1) mediated arginine installation. Here we presented structures of PCO2, PCO4, and PCO5 from Arabidopsis thaliana (AtPCOs) and examined their in vitro activity by MS. On the basis of Tris-bound AtPCO2, we modelled the Cys-bound AtPCO2 structure and identified key residues involved in N-Cys oxidation. Alanine substitution of potential N-Cys interaction residues impaired the activity of AtPCO5 remarkably. The structural research, complemented by mutagenesis and mass spectrometry experiments, not only uncovers the substrate recognition and catalytic mode by AtPCOs, but also sheds light on the future design of potent inhibitors for plant cysteine oxidases.

molecular biology↗

bioRxiv2020

NuMorph: tools for cellular phenotyping in tissue cleared whole brain images

Tissue clearing methods allow every cell in the mouse brain to be imaged without physical sectioning. However, the computational tools currently available for cell quantification in cleared tissue images have been limited to counting sparse cell populations in stereotypical mice. Here we introduce NuMorph, a group of image analysis tools to quantify all nuclei and nuclear markers within the mouse cortex after tissue clearing and imaging by a conventional light-sheet microscope. We applied NuMorph to investigate two distinct mouse models: a Topoisomerase 1 (Top1) conditional knockout model with severe neurodegenerative deficits and a Neurofibromin 1 (Nf1) conditional knockout model with a more subtle brain overgrowth phenotype. In each case, we identified differential effects of gene deletion on individual cell-type counts and distribution across cortical regions that manifest as alterations of gross brain morphology. These results underline the value of 3D whole brain imaging approaches and the tools are widely applicable for studying 3D structural deficits of the brain at cellular resolution in animal models of neuropsychiatric disorders.

neuroscience↗

bioRxiv2020

St. Jude Cloud a Pediatric Cancer Genomic Data Sharing Ecosystem

Effective data sharing is key to accelerating research that will improve the precision of diagnoses, efficacy of treatments and long-term survival of pediatric cancer and other childhood catastrophic diseases. We present St. Jude Cloud (https://www.stjude.cloud), a cloud-based data sharing ecosystem developed via collaboration between St. Jude Childrens Research Hospital, DNAnexus, and Microsoft, for accessing, analyzing and visualizing genomic data from >10,000 pediatric cancer patients, long-term survivors of pediatric cancer and >800 pediatric sickle cell patients. Harmonized genomic data totaling 1.25 petabyes on St. Jude Cloud include 12,104 whole genomes, 7,697 whole exomes and 2,202 transcriptomes, which are freely available to researchers worldwide. The resource is expanding rapidly with regular data uploads from St. Judes prospective clinical genomics programs, providing public access as soon as possible rather than holding data back until publication. Three interconnected apps within the St. Jude Cloud ecosystem--Genomics Platform, Pediatric Cancer Knowledgebase (PeCan) and Visualization Community--provide a unique experience for simultaneously performing advanced data analysis in the cloud and enhancing the pediatric cancer knowledgebase. We demonstrate the value of the St. Jude Cloud ecosystem through use cases that classify 48 pediatric cancer subtypes by gene expression profiling and map mutational signatures across 35 subtypes of pediatric cancer.

cancer biology↗

bioRxiv2020

Recombinant SARS-CoV-2 RBD molecule with a T helper epitope as a built in adjuvant induces strong neutralization antibody response

Without approved vaccines and specific treatment, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is spreading around the world with above 20 million COVID-19 cases and approximately 700 thousand deaths until now. An efficacious and affordable vaccine is urgently needed. The Val308 - Gly548 of Spike protein of SARS-CoV-2 linked with Gln830 - Glu843 of Tetanus toxoid (TT peptide) (designated as S1-4) and without TT peptide (designated as S1-5), and prokaryotic expression, chromatography purification and the rational renaturation of the protein were performed. The antigenicity and immunogenicity of S1-4 protein was evaluated by Western Blotting (WB) in vitro and immune responses in mice, respectively. The protective efficiency of it was measured by virus neutralization test in Vero E6 cells with SARS-CoV-2. S1-4 protein was prepared to high homogeneity and purity by prokaryotic expression and chromatography purification. Adjuvanted with Alum, S1-4 protein stimulated a strong antibody response in immunized mice and caused a major Th2-type cellular immunity compared with S1-5 protein. Furthermore, the immunized sera could protect the Vero E6 cells from SARS-CoV-2 infection with neutralization antibody GMT 256. The candidate subunit vaccine molecule could stimulate strong humoral and Th1 and Th2-type cellular immune response in mice, giving us solid evidence that S1-4 protein could be a promising subunit vaccine candidate.

bioengineering↗

bioRxiv2020

Regulatory T cell activation triggers specific changes in glycosylation associated with Siglec-1-dependent inflammatory responses

Siglec-1 is a macrophage lectin-like receptor that mediates sialic acid-dependent cellular interactions. It was shown previously to promote inflammation in autoimmune disease through suppressing the expansion of regulatory T cells (Tregs). We have investigated the molecular basis for Siglec-1 binding to these cells using in vitro-induced Tregs. Siglec-1 binding was strongly upregulated on activated cells, but lost under resting conditions. Glycosylation changes that affect Siglec-1 binding were studied by comparing activated and resting Tregs using RNA-Seq, glycomics, proteomics and binding of selected antibodies and lectins. A proximity labelling and proteomics strategy identified 49 glycoproteins expressed by activated Tregs that may function as Siglec-1 counter-receptors. These represent [~]5% of the total membrane protein pool and were mainly related to T cell activation and proliferation. We demonstrate that several of these counter-receptors are upregulated following activation of Tregs and provide initial evidence that their altered glycosylation may also be important for Siglec-1 binding.

immunology↗

bioRxiv2020

Integrator is a genome-wide attenuator of non-productive transcription

Termination of RNA polymerase II (RNAPII) transcription in metazoans relies largely on the Cleavage and Polyadenylation (CPA) and Integrator (INT) complexes originally found to act at the ends of protein-coding and snRNA genes, respectively. Here we monitor CPA- and INT-dependent termination activities genome-wide, including over 8000 previously unannotated transcription units (TUs), that produce unstable RNA. We verify the global activity of CPA, that occurs at pA sites indiscriminately of their positioning relative to the TU promoter. We also identify a global activity of INT, which is, however, largely sequence-independent and restricted to a ~3 kb promoter-proximal region. Our analyses suggest two functions of genome-wide INT activity; it dampens transcriptional output from weak promoters and it provides quality-control of RNAPII complexes, that are unfavorably configured for transcriptional elongation. We suggest that the function of INT in stable snRNA production is an exception from its general cellular role, attenuation of non-productive transcription.

cell biology↗