Biology subjects

Hu, J.

Publications and source records attributed to Hu, J..

At least 451 records · Page 25Linked to original sources

bioRxiv2020

Autographa californica Multiple Nucleopolyhedrovirus orf13 is Required for Efficient Nuclear Egress of Nucleocapsids

Autographa californica multiple nucleopolyhedrovirus (AcMNPV) orf13 (ac13) is a conserved gene in all sequenced alphabaculoviruses. However, its function in the viral life cycle remains unknown. In this study we found that ac13 was a late gene and that the encoded protein, bearing a putative nuclear localization signal motif in the DUF3627 domain, colocalized with the nuclear membrane. Deletion of ac13 did not affect viral DNA replication, gene transcription, nucleocapsid assembly or occlusion body (OB) formation, but reduced virion budding from infected cells by approximately 400-fold compared with the wild-type virus. Deletion of ac13 substantially impaired the egress of nucleocapsids from the nucleus to the cytoplasm, while the number of occlusion-derived viruses embedded within OBs was unaffected. Taken together, our results indicated that ac13 was required for efficient nuclear egress of nucleocapsids during virion budding, but was dispensable for OB formation. IMPORTANCEEgress of baculovirus nucleocapsids from the nucleus is an essential process for morphogenesis of mature budded viruses, which is required to spread infection within susceptible cells and tissues. Although many viral and host proteins are required for nucleocapsid egress, the specific mechanisms underlying this process in baculoviruses remain somewhat enigmatic. In the present study, we found that the ac13 gene, in addition to ac11, ac51, ac66, ac75, ac78, gp41, ac93, p48, exon0 and ac142, was required for efficient nuclear egress of nucleocapsids. Our results contribute to a better understanding of nucleocapsid egress in baculoviruses.

microbiology↗

bioRxiv2020

The D614G mutation of SARS-CoV-2 spike protein enhances viral infectivity and decreases neutralization sensitivity to individual convalescent sera

Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The spike (S) protein that mediates SARS-CoV-2 entry into host cells is a major target for vaccines and therapeutics. Thus, insights into its sequence variations are key to understanding the infection and antigenicity of SARS-CoV-2. A dominant mutational variant at position 614 of the S protein (aspartate to glycine, D614G mutation) was observed in the SARS-CoV-2 genome sequence obtained from the Nextstrain database. Using a pseudovirus-based assay, we identified that S-D614 and S-G614 protein pseudotyped viruses share a common receptor, human angiotensin-converting enzyme 2 (ACE2), which could be blocked by recombinant ACE2 with the fused Fc region of human IgG1. However, S-D614 and S-G614 protein demonstrated functional differences. First, S-G614 protein could be cleaved by serine protease elastase-2 more efficiently. Second, S-G614 pseudovirus infected 293T-ACE2 cells significantly more efficiently than did the S-D614 pseudovirus, especially in the presence of elastase-2. Third, an elastase inhibitor approved for clinical use blocked elastase-enhanced S-G614 pseudovirus infection. Moreover, 93% (65/70) convalescent sera from patients with COVID-19 could neutralize both S-D614 and S-G614 pseudoviruses with comparable efficiencies, but about 7% (5/70) convalescent sera showed reduced neutralizing activity against the S-G614 pseudovirus. These findings have important implications for SARS-CoV-2 transmission and immune interventions.Competing Interest StatementThe authors have declared no competing interest.View Full Text

microbiology↗

bioRxiv2020

C3: Connect separate Connected Components to form a succinct disease module

Accurate disease module is helpful in understanding the molecular mechanism of disease causation and identifying drug target. However, for the fragmentization of disease module in incomplete human interactome, how to determine connectivity pattern and detect a full neighbourhood of disease is an open problem. In this paper, a topology-based method is developed to dissect the connectivity of intermediate nodes and edges and form a succinct disease module. By applying this Connect separate Connected Components (CCC, C3) method on a large corpus of curated diseases, we find that most Separate Connected Components (SCCs) formed by Disease-Associated Proteins (DAPs) can be connected into a well-connected component as a succinct observable module. This pattern also holds for altered genes from multi-omics data such as The Cancer Genome Atlas. Overall, C3 tool can not only inspire a deeper understanding of interconnectedness of phenotypically related genes and different omics data, but also be used to detect a well-defined neighbourhood that drives complex pathological processes.

bioinformatics↗

bioRxiv2020

TraceQC: An R package for quality control of CRISPR lineage tracing data

MotivationThe CRISPR-based lineage tracing system is emerging as a powerful new sequencing tool to track cell lineages by marking cells with irreversible genetic mutations. Accurate reconstruction of cell lineages from CRISPR-based data is sensitive to noise. Quality control is critical for filtering out low-quality data points. Yet, existing quality control tools for RNA-seq and DNA-seq do not measure features specific for the CRISPR-linear tracing system. ResultsWe introduce TraceQC, a quality control package to overcome challenges with measuring the quality of CRISPR-based lineage tracing data and help in interpreting and constructing lineage trees. AvailabilityThe R package is available at https://github.com/LiuzLab/TraceQC. Contactzhandong.liu@bcm.edu or hyun-hwan.jeong@bcm.edu

bioinformatics↗

bioRxiv2020

Voltage-gated Ca2+ and K+ channel coupling regulates CA1 hippocampal synaptic filtering and spine excitability

The transient K+ current (IA) carried by pore forming Kv4.2 subunits regulates the propagation of synaptic input, dendritic excitability, and synaptic plasticity in CA1 pyramidal neuron dendrites of the hippocampus. We report that the Ca2+ channel subunit Cav2.3 regulates IA in this cell type. We first identified Cav2.3 as a Kv4.2 interacting protein in a proteomic screen and we confirmed Cav2.3-Kv4.2 complex association using multiple techniques. Functionally, Cav2.3 Ca2+-entry increases Kv4.2-mediated whole-cell current due to an increase in Kv4.2 surface expression. Using pharmacology and Cav2.3 knockout mice, Cav2.3 was found to promote whole-cell IA and the increasing gradient of IA in the apical dendrite distal to the neuronal soma. Furthermore, the loss of Cav2.3 function leads to enhancement of synaptic currents and spine Ca2+ influx. These results present Cav2.3 and Kv4.2 as integral constituents of an ion channel complex that impacts synaptic function in the hippocampus.

neuroscience↗

bioRxiv2020

An extracellular protein regulates patched-related/DAF-6-mediated sensory compartment formation in C. elegans

Coordination of neurite extension with surrounding glia development is critical for neuronal function, but the underlying molecular mechanisms remain poorly understood. Through a genome-wide mutagenesis screen in C. elegans, we identified dyf-4 and daf-6 as two mutants sharing similar defects in dendrite extension. DAF-6 encodes a glia-specific patched-related membrane protein that plays vital roles in glial morphogenesis. We cloned dyf-4 and found that DYF-4 encodes a glia-secreted extracellular protein. Intriguingly, DYF-4 colocalizes with DAF-6 along the glial channel. Further investigations revealed that DYF-4 directly interacts with DAF-6 and regulates its proper membrane localization. Notably, reported glial suppressors of daf-6 could also restore dendrite elongation and ciliogenesis in both dyf-4 and daf-6 mutants. Collectively, our data suggest that secreted DYF-4 likely acts as a novel ligand/regulator for the patched-related receptor DAF-6 which promotes the proper formation of the glial channel and indirectly affects neurite extension and ciliogenesis.

developmental biology↗

bioRxiv2020

Immune landscape of a genetically-engineered murine model of glioma relative to human glioma by single-cell sequencing

BackgroundThis study focuses on the evaluation of intratumoral immune infiltrates of the Qk/trp53/Pten (QPP) triple-knockout mouse model of glioblastoma with the purpose of establishing its relevance compared to the human disease. This included an analysis at the single cell level, of immune cells composition in both spontaneous and implanted mouse tumors and of samples obtained from human tumor resections. MethodsWe analyzed a cohort of fifteen spontaneous QPP mice, nine implanted QPP mice and 10 glioma patients by standard immune profiling methods such as IHC. Of those, we analyzed three spontaneous QPP mice, three implanted QPP mice, and 10 glioma patients by single cell RNA sequencing. ResultsIn the QPP samples we identified a predominantly myeloid cell population of monocytes, macrophages, and microglia, with minor populations of T, B, and NK cells. When comparing spontaneous and implanted mouse samples, we found that there were more neutrophil, T and NKT cells in the implanted model. In human samples, most of the lymphoid and myeloid compartments were immunosuppressive. While the complexity of the myeloid cell populations is preserved between the QPP model and the human disease, we observed significantly more immune-stimulatory populations in the implanted mouse model. ConclusionsWe found that, despite differences at the subpopulation level, the immunosuppressive nature of the immune system in glioma, is recapitulated in our mouse model. Although we observed differences in the proportions of immune infiltrates derived from the spontaneous and implanted mouse models and in LGG compared to GBM, the major constituents are present in all cases and these differences may be caused by various etiological or pathogenic influences on tumor-immune interactions. Key PointsO_LINew GBM mouse model for immunotherapy C_LIO_LISingle cell profiling of the immune systems in human and mouse GBM C_LI

cancer biology↗

bioRxiv2020

Multiethnic catalog of structural variants and their translational impact for disease phenotypes across 19,652 genomes

Genome sequencing at population scale provides unprecedented access to the genetic foundations of human phenotypic diversity, but genotype-phenotype association analyses limited to small variants have failed to comprehensively characterize the genetic architecture of human health and disease because they ignore structural variants (SVs) known to contribute to phenotypic variation and pathogenic conditions1-3. Here we demonstrate the significance of SVs when assessing genotype-phenotype associations and the importance of ethnic diversity in study design by analyzing SVs across 19,652 individuals and the translational impact on 4,156 aptamerbased proteomic measurements across 4,021 multi-ethnic samples. The majority of 304,533 SVs detected are rare, although we identified 2,336 protein-coding genes impacted by common SVs.\ We identified 64 significant SV-protein associations that comprise 36 cis- and 28 trans-acting relationships, and 21 distinct SV regions overlapped with genome-wide association study loci. These findings represent a more comprehensive mapping of regulatory and translational endophenotypes underlying health and disease.

genomics↗

bioRxiv2020

Permutation-based Identification of Important Biomarkers for Complex Diseases via Black-box Models

Study of human disease remains challenging due to convoluted disease etiologies and complex molecular mechanisms at genetic, genomic, and proteomic levels. Many machine learning-based methods, including deep learning and random forest, have been developed and widely used to alleviate some analytic challenges in complex human disease studies. While enjoying the modeling flexibility and robustness, these model frameworks suffer from non-transparency and difficulty in interpreting the role of each individual feature due to their intrinsic black-box natures. However, identifying important biomarkers associated with complex human diseases is a critical pursuit towards assisting researchers to establish novel hypotheses regarding prevention, diagnosis and treatment of complex human diseases. Herein, we propose a Permutation-based Feature Importance Test (PermFIT) for estimating and testing the feature importance, and for assisting interpretation of individual feature in various black-box frameworks, including deep neural networks, random forests, and support vector machines. PermFIT (available at https://github.com/SkadiEye/deepTL) is implemented in a computationally efficient manner, without model refitting for each permuted data. We conduct extensive numerical studies under various scenarios, and show that PermFIT not only yields valid statistical inference, but also helps to improve the prediction accuracy of black-box models with top selected features. With the application to the Cancer Genome Atlas (TCGA) kidney tumor data and the HITChip atlas BMI data, PermFIT clearly demonstrates its practical usage in identifying important biomarkers and boosting performance of black-box predictive models.

bioinformatics↗

bioRxiv2020

Cytotoxic lymphocytes use mechanosurveillance to target biophysical vulnerabilities in cancer

Immune cells identify cancer cells by recognizing characteristic biochemical features indicative of oncogenic transformation. Cancer cells have characteristic mechanical features, as well, but whether these biophysical properties also contribute to destruction by the immune system is not known. In the present study, we found that enhanced expression of myocardin related transcription factors (MRTFs), which promote migration and metastatic invasion, paradoxically compromised lung colonization by melanoma and breast carcinoma cells in an immune-mediated manner. Cancer cells with increased MRTF signaling were also more sensitive to immune checkpoint blockade therapy in mice and humans. The basis for this vulnerability was not biochemical, but biophysical. MRTF expression strengthened the actin cytoskeleton, increasing the rigidity of cancer cells and thereby making them more vulnerable to cytotoxic T lymphocytes and natural killer cells. These results reveal a mechanical dimension of immunosurveillance, which we call mechanosurveillance, that is particularly relevant to the targeting of metastatic disease.

immunology↗

bioRxiv2020

Ratio of the interferon-γ signature to the immunosuppression signature predicts anti-PD-1 therapy response in melanoma

Immune checkpoint inhibitor (ICI) treatments produce clinical benefit in many patients. However, better pretreatment predictive biomarkers for ICI are still needed to help match individual patients to the treatment most likely to be of benefit. Existing gene expression profiling (GEP)-based biomarkers for ICI are primarily focused on measuring a T cell-inflamed tumour microenvironment that contributes positively to the response to ICI. Here, we identified an immunosuppression signature (IMS) through analysing RNA sequencing data from a combined discovery cohort (n = 120) consisting of three publicly available melanoma datasets. Using the ratio of an established IFN-{gamma} signature and IMS led to consistently better prediction of the ICI therapy outcome compared to a collection of nine published GEP signatures from the literature on a newly generated internal validation cohort (n = 55) and three published datasets of metastatic melanoma treated with anti-PD-1 (n = 48) and anti-CTLA-4 (n = 42) as well as in patients with gastric cancer treated with antiPD-1 (n = 45), demonstrating the potential utility of IMS as a predictive/prognostic biomarker that complements existing GEP signatures for immunotherapy.

immunology↗

bioRxiv2020

Inhibition of the av integrin-TGF-b axis improves natural killer cell function against glioblastoma stem cells

Glioblastoma, the most aggressive brain cancer, often recurs because glioblastoma stem cells (GSCs) are resistant to all standard therapies. Here, we show that patient-derived GSCs, but not normal astrocytes, are highly sensitive to lysis by healthy allogeneic natural killer (NK) cells in vitro. In contrast, single cell analysis of autologous, tissue infiltrating NK cells isolated from surgical samples of high-grade glioblastoma patient tumors using mass cytometry and single cell RNA sequencing revealed an abnormal phenotype associated with impaired lytic function compared with peripheral blood NK cells from GBM patients or healthy donors. This immunosuppression was attributed to an integrin-TGF-{beta} mechanism, activated by direct cell-cell contact between GSCs and NK cells. Treatment of GSC-engrafted mice with allogeneic NK cells in combination with inhibitors of integrin or TGF-{beta} signaling, or with TGF-{beta} receptor 2 gene-edited NK cells prevented GSC-induced NK cell dysfunction and tumor growth. Collectively, our findings reveal a novel mechanism of NK cell immune evasion by GSCs and implicate the integrin-TGF-{beta} axis as a useful therapeutic target to eliminate GSCs in this devastating tumor.

immunology↗

bioRxiv2020

Rottlerin ameliorates DSS-induced colitis by improving intestinal barrier function via activation of the Epac-2/Rap-1 signaling pathway.

OBJECTIVESRottlerin, a pan PDE inhibitor, has a variety of pharmacological activities, including enhancing barrier function and mediating anti-inflammatory activity by changing the distribution of occludin and ZO-1. Nevertheless, the function of rottlerin on Crohn disease (CD) keep unknown. Our aim of the study is to investigate the role of rottlerin on CD-like colitis and its mechanism. METHODSWild-type mice which were 8-10 weeks old were randomly divided into three treatment groups: (i) the normal feeding, no administration (control) group, (ii) the group administered 3% dextran sodium sulfate (DSS) alone, and (iii) the group administered rottlerin (100 mg/kg) and 3% DSS. In this study, the effect of rottlerin on the function and structure of the intestinal barrier was investigated, and the possible mechanism was discussed. We performed signaling pathway analysis and flow cytometry to identify the detailed mechanisms by which rottlerin (10 g/mL) treatment inhibits cell growth arrest and the attenuation of TJ proteins in LPS-treated FHs 74 int cells. RESULTSRottlerin treatment significantly ameliorated colitis induced by DSS in WT mice, which was manifested by a decrease in inflammation score, the attenuation of inflammatory factors and the inhibition of destruction on intestinal barrier structure. Rottlerin enhanced the levels of occludin and ZO-1, and improved the function of intestinal barrier, which may have been why rottlerin ameliorated colitis in WT mice. The anti-inflammatory effect of rottlerin may be partly due to the activation of Epac-2/Rap-1 signaling. CONCLUSIONSRottlerin may treat CD in humans via enhancing TJ proteins expression and improving the function of intestinal barrier.

cell biology↗

bioRxiv2020

Elucidating the pathogenic mechanisms of AD brain-derived tau-containing extracellular vesicles: Highly transmissible and preferential propagation to GABAergic neurons

Extracellular vesicles (EVs) propagate tau pathology for Alzheimers disease (AD). How EV transmission influences AD are, nonetheless, poorly understood. To these ends, the physicochemical and molecular structure-function relationships of human brain-derived EVs, from AD and prodromal AD (pAD), were compared to non-demented controls (CTRL). AD EVs were shown to be significantly enriched in epitope-specific tau oligomers versus pAD or CTRL EVs assayed by dot-blot and atomic force microscopy tests. AD EVs were efficiently internalized by murine cortical neurons and transferred tau with higher aggregation potency than pAD and CTRL EVs. Strikingly, inoculation of tau-containing AD EVs into the outer molecular layer of the dentate gyrus induced tau propagation throughout the hippocampus. This was seen in 22 months-old C57BL/6 mice at 4.5 months post-injection by semiquantitative brain-wide immunohistochemistry tests with multiple anti-phospho-tau (p-tau) antibodies. Inoculation of the equal amount of tau from CTRL EVs or as oligomer or fibril-enriched fraction from the same AD donor showed little propagation. AD EVs induced tau accumulation in the hippocampus as oligomers or sarkosyl-insoluble proteins. Unexpectedly, p-tau cells were mostly GAD67+ GABAergic neurons and to a lesser extent, GluR2/3+ excitatory mossy cells, showing preferential EV-mediated GABAergic neuronal tau propagation. Whole-cell patch clamp recording of Cornu Ammonis (CA1) pyramidal cells showed significant reduction in the amplitude of spontaneous inhibitory post-synaptic currents. This was accompanied by reductions in c-fos+ GAD67+GABAergic neurons and GAD67+ GABAergic neuronal puncta surrounding pyramidal neurons in the CA1 region confirming reduced interneuronal projections. Our study posits a novel tau-associated pathological mechanism for brain-derived EVs.

neuroscience↗

bioRxiv2020

O-GlcNAcylation of SAMHD1 Indicating a Link between Metabolic Reprogramming and Anti-HBV Immunity

Viruses hijack the host cell machinery to promote viral replication; however, the mechanism by which metabolic reprogramming regulates innate antiviral immunity in the host remains elusive. Herein, we found that Hepatitis B virus (HBV) infection upregulates glucose transporter 1expression, promotes hexosamine biosynthesis pathway (HBP) activity, and enhances O-linked {beta}-N-acetylglucosamine (O-GlcNAc) modification of downstream proteins. HBP-mediated O-GlcNAcylation positively regulates host antiviral response against HBV in vitro and in vivo. Mechanistically, O-GlcNAc transferase (OGT)-mediated O-GlcNAcylation of sterile alpha motif and histidine/aspartic acid domain-containing protein 1 (SAMHD1) on Ser93 stabilizes SAMHD1 and enhances its antiviral activity. In addition, O-GlcNAcylation of SAMHD1 promoted its antiviral activity against human immunodeficiency virus-1 in vitro. In conclusion, the results of our study reveal a link between HBP, O-GlcNAc modification, and innate antiviral immunity by targeting SAMHD1. Therefore, the results of this study demonstrate a strategy for the potential treatment of HBV infection by modulating HBP activity.

microbiology↗

bioRxiv2020

Curvature domains in V4 of Macaque Monkey

An important aspect of visual object recognition is the ability to perceive object shape. How the brain encodes fundamental aspects of shape information remains poorly understood. Models of object shape representation describe a multi-stage process that includes encoding of contour orientation and curvature. While modules encoding contour orientation are well established (orientation domains in V1 and V2 visual cortical areas), whether there are modules for curvature is unknown. In this study, we identify a module for curvature representation in area V4 of monkey visual cortex and illustrate a systematic representation of low to high curvature and of curvature orientation, indicative of curvature hypercolumns in V4. We suggest that identifying systematic modular organizations at each stage of the visual cortical hierarchy signifies the key computations performed. SignificanceWe use intrinsic signal optical imaging in area V4 of anesthetized macaque monkey to study the functional organization of curvature representation. We find a modular basis for cue-invariant curvature representation in area V4 of monkey visual cortex and illustrate a systematic representation from low to high curvature and of curvature orientation, replete with curvature pinwheels. This is the first report of systematic functional organization for curvature representation in the visual system. The use of optical imaging has revealed at a population level spatial details of cortical responses, something which has not been evident from previous studies of single neurons. These data support a representational architecture underlying a curvature hypercolumn in V4.

neuroscience↗

bioRxiv2020

Analyzing Presymptomatic Tissue to Gain Insights into the Molecular and Mechanistic Origins of Late-Onset Degenerative Trinucleotide Repeat Disease

How genetic defects trigger the molecular changes that cause late-onset disease is important for understanding disease progression and therapeutic development. Fuchs endothelial corneal dystrophy (FECD) is an RNA-mediated disease caused by a trinucleotide CUG expansion in an intron within the TCF4 gene. The mutant intronic CUG RNA is present at 1-2 copies per cell, posing a challenge to understand how a rare RNA can cause disease. Late-onset FECD is a uniquely advantageous model for studying how RNA triggers disease because; 1) Affected tissue is routinely removed during surgery; 2) The expanded CUG mutation is one of the most prevalent disease-causing mutations, making it possible to obtain pre-symptomatic tissue from eye bank donors to probe how gene expression changes precede disease; and 3) The affected tissue is a homogeneous single cell monolayer, facilitating accurate transcriptome analysis. Here we use RNA sequencing (RNAseq) to compare tissue from individuals who are pre-symptomatic (Pre_S) to tissue from patients with late stage FECD (FECD_REP). The abundance of mutant repeat intronic RNA in Pre_S and FECD_REP tissue is elevated due to increased half-life in a corneal cell-specific manner. In Pre_S tissue, changes in splicing and extracellular matrix gene expression foreshadow the changes observed in advanced disease and predict the activation of the fibrosis pathway and immune system seen in late-stage patients. The absolute magnitude of splicing changes is similar in presymptomatic and late stage tissue. Our data identify gene candidates for early drivers of disease and biomarkers that may represent diagnostic and therapeutic targets for FECD. We conclude that changes in alternative splicing and gene expression are observable decades prior to the diagnosis of late-onset trinucleotide repeat disease.

molecular biology↗

bioRxiv2020

Lanthanide-dependent alcohol dehydrogenases require an essential aspartate residue for metal coordination and function

The presence of lanthanide elements (Ln3+) and pyrroloquinoline quinone (PQQ) containing cofactors in XoxF methanol dehydrogenases (MDHs) and ExaF ethanol dehydrogenases (EDHs) has expanded the list of biological elements and opened novel areas of metabolism and ecology. Other MDHs known as MxaFIs are related in sequence and structure to these proteins, yet they instead possess a Ca2+-PQQ cofactor. An important missing piece of the Ln3+ puzzle is defining what protein features distinguish enzymes using Ln3+-PQQ cofactors from those that do not. In this study, we investigated the functional importance of a proposed lanthanide-coordinating aspartate using XoxF1 MDH from the model methylotrophic bacterium Methylorubrum extorquens AM1. We report two crystal structures of XoxF1, one containing PQQ and the other free of this organic molecule, both with La3+ bound in the active site region and coordinated by Asp320. Using constructs to produce either recombinant XoxF1 or its D320A variant, we show Asp320 is needed for in vivo catalytic function, in vitro activity of purified enzyme, and coordination of La3+. XoxF1 and XoxF1 D320A, when produced in the absence of La3+, coordinate Ca2+, but exhibit little or no catalytic activity. In addition, we generated the parallel substitution to produce ExaF D319S, and showed the enzyme loses the capacity for efficient ethanol oxidation with La3+. These results provide empirical evidence of an essential Ln3+-coordinating aspartate for the function of XoxF MDHs and ExaF EDHs; thus, supporting the suggestion that sequences of these enzymes, and the genes that encode them, are markers for Ln3+ metabolism.

biochemistry↗