Biology subjects

Xu, K.

Publications and source records attributed to Xu, K..

At least 253 records · Page 14Linked to original sources

bioRxiv2019

Direct comparison of clathrin-mediated endocytosis in budding and fission yeast reveals conserved and evolvable features

AbstractConserved proteins drive clathrin-mediated endocytosis (CME), which universally involves a burst of actin assembly. To gain fundamental mechanistic insights into this process, a side-by-side quantitative comparison of CME was performed on two distantly related yeast species. Though endocytic protein abundance in S. pombe and S. cerevisiae are more similar than previously thought, membrane invagination speed and depth are two-fold greater in fission yeast than in budding yeast. In both yeasts, Arp2/3 complex activation drives membrane invagination when triggered by the accumulation of [~]70 WASP molecules. In contrast to budding yeast, WASP-mediated actin nucleation activity plays an essential role in fission yeast endocytosis. Genetics and live-cell imaging revealed core CME spatiodynamic similarities between the two yeasts, though two-zone actin assembly is a fission yeast-specific mechanism, which is not essential for CME. These studies identified conserved CME mechanisms and species-specific adaptations and have broad implications that extend from yeast to humans.

cell biology↗

bioRxiv2019

Super-resolution microscopy unveils FIP200-scaffolded, cup-shaped organization of mammalian autophagic initiation machinery

Autophagy is an essential physiological process by which eukaryotic cells degrade and recycle cellular materials. Although the biochemical hierarchies of the mammalian autophagy pathway have been identified, questions remain regarding the sequence, subcellular location, and structural requirements of autophagosome formation. Here, we characterize the structural organization of key components of the mammalian autophagic initiation machinery at [~]20 nm spatial resolution via three-color, three-dimensional super-resolution fluorescence microscopy. We thus show that upon cell starvation, FIP200, a large structural protein of the ULK1 complex with no direct yeast homolog, scaffolds the formation of cup-like structures located at SEC12-enriched remodeled ER-exit sites prior to LC3 lipidation. This cup scaffold, then, provides a structural asymmetry to enforce the directional recruitment of downstream components, including the Atg12-Atg5-Atg16 complex, WIPI2, and LC3, to the cup inside. Moreover, we provide evidence that the early autophagic machinery is recruited in its entirety to these cup structures prior to LC3 lipidation, and gradually disperses and dissociates on the outer face of the phagophore membrane during elongation. We thus shed new light on the physical process of mammalian autophagic initiation and development at the nanometer-scale.

cell biology↗

bioRxiv2019

Mitochondrial dysfunction is signaled to the integrated stress response by OMA1, DELE1 and HRI

In mammalian cells, mitochondrial dysfunction triggers the integrated stress response (ISR), in which eIF2 phosphorylation upregulates the transcription factor ATF4. However, how mitochondrial stress is relayed to the ISR is unknown. We found that HRI is the eIF2 kinase necessary and sufficient for this relay. Using an unbiased CRISPRi screen, we identified factors upstream of HRI: OMA1, a mitochondrial stress-activated protease, and DELE1, a little-characterized protein we found to be associated with the inner mitochondrial membrane. Mitochondrial stress stimulates the OMA1-dependent cleavage of DELE1, leading to its accumulation in the cytosol, where it interacts with HRI and activates its eIF2 kinase activity. Blockade of the OMA1-DELE1-HRI pathway is beneficial during some, but not all types of mitochondrial stress, and leads to an alternative response that induces specific molecular chaperones. Therefore, this pathway is a potential therapeutic target enabling fine-tuning of the ISR for beneficial outcomes in diseases involving mitochondrial dysfunction.

cell biology↗

bioRxiv2019

Shared and Distinct Dysfunction of Dynamic Connectivity Networks across Schizophrenia, Bipolar Disorder and Major Depression Disorder

Dynamic functional connectivity (DFC) analysis can capture time-varying properties of connectivity and may provide further information about transdiagnostic psychopathology across major psychiatric disorders. In this study, we used resting state functional MRI and a sliding-window method to study DFC in 150 schizophrenia (SZ), 100 bipolar disorder(BD), 150 major depressive disorder (MDD), and 210 healthy controls (HC). DFC were clustered into two functional connectivity states. Significant 4-group differences in DFC were found only in state 2. Post hoc analyses showed that transdiagnostic dysconnectivity among there disorders featured decreased connectivity within visual, somatomotor, salience and frontoparietal networks. Our results suggest that decreased connectivity within both lower-order (visual and somatomotor) and higher-order (salience and frontoparietal) networks may serve as transdiagnostic marker of these disorders, and that these dysconnectivity is state-dependent. Targeting these dysconnectivity may improve assessment and treatment for patients that having more than one of these disorders at the same time.

neuroscience↗

bioRxiv2019

Scalable Spike Source Localization in Extracellular Recordings using Amortized Variational Inference

Determining the positions of neurons in an extracellular recording is useful for investigating functional properties of the underlying neural circuitry. In this work, we present a Bayesian modelling approach for localizing the source of individual spikes on high-density, microelectrode arrays. To allow for scalable inference, we implement our model as a variational autoencoder and perform amortized variational inference. We evaluate our method on both biophysically realistic simulated and real extracellular datasets, demonstrating that it is more accurate than and can improve spike sorting performance over heuristic localization methods such as center of mass.

neuroscience↗

bioRxiv2019

Graphene-enabled, spatially controlled electroporation of adherent cells for live-cell super-resolution microscopy

The incorporation of exogenous molecules into live cells is essential for both biological research and therapeutic applications. In particular, for the emerging field of super-resolution microscopy of live mammalian cells, reliable fluorescent labeling of intracellular targets remains a challenge. Here, utilizing the unique mechanical, electrical, and optical properties of graphene, a single layer of bonded carbon atoms, we report a facile approach that enables both high-throughput delivery of fluorescent probes into adherent live cells and in situ super-resolution microscopy on the same device. [~]90% delivery efficiencies are achieved for free dyes and dye-tagged affinity probes, short peptides, and whole antibodies, thus enabling high-quality super-resolution microscopy. Moreover, we demonstrate excellent spatiotemporal controls, which, in combination with the ready patternablity of graphene, allow for the spatially selective delivery of two different probes for cells at different locations on the same substrate. We thus open up a new pathway to the microscopic manipulation and visualization of live cells.

bioengineering↗

bioRxiv2019

Retrospective Association Analysis of Longitudinal Binary Traits Identifies Important Loci and Pathways in Cocaine Use

Longitudinal phenotypes have been increasingly available in genome-wide association studies (GWAS) and electronic health record-based studies for identification of genetic variants that influence complex traits over time. For longitudinal binary data, there remain significant challenges in gene mapping, including misspecification of the model for the phenotype distribution due to ascertainment. Here, we propose L-BRAT, a retrospective, generalized estimating equations-based method for genetic association analysis of longitudinal binary outcomes. We also develop RGMMAT, a retrospective, generalized linear mixed model-based association test. Both tests are retrospective score approaches in which genotypes are treated as random conditional on phenotype and covariates. They allow both static and time-varying covariates to be included in the analysis. Through simulations, we illustrated that retrospective association tests are robust to ascertainment and other types of phenotype model misspecification, and gain power over previous association methods. We applied L-BRAT and RGMMAT to a genome-wide association analysis of repeated measures of cocaine use in a longitudinal cohort. Pathway analysis implicated association with opioid signaling and axonal guidance signaling pathways. Lastly, we replicated important pathways in an independent cocaine dependence case-control GWAS. Our results illustrate that L-BRAT is able to detect important loci and pathways in a genome scan and to provide insights into genetic architecture of cocaine use.

genetics↗

bioRxiv2019

Single-cell Transcriptome Mapping Identifies Common and Cell-type Specific Genes Affected by Acute Delta9-tetrahydrocannabinol in Humans

Delta 9-tetrahydrocannabinol (THC), the principal psychoactive constituent of cannabis, is also known to modulate immune response in peripheral cells. The mechanisms of THCs effects on gene expression in human immune cells remains poorly understood. Combining a within-subject design with single cell transcriptome mapping, we report that administration of THC acutely alters gene expression in 15,973 human blood immune cells. Controlled for high inter-individual transcriptomic variability, we identified 294 transcriptome-wide significant genes among eight cell types including 69 common genes and 225 cell-type specific genes affected by acute THC administration, including those genes involving not only in immune response, cytokine production, but signal transduction, and cell proliferation and apoptosis. We revealed distinct transcriptomic sub-clusters affected by THC in major immune cell types where THC perturbed cell type-specific intracellular gene expression correlations. Gene set enrichment analysis further supports the findings of THCs common and cell-type specific effects on immune response and cell toxicity. We found that THC alters the correlation of cannabinoid receptor gene, CNR2, with other genes in B cells, in which CNR2 showed the highest level of expression. This comprehensive cell-specific transcriptomic profiling identified novel genes regulated by THC and provides important insights into THCs acute effects on immune function that may have important medical implications.

genetics↗

bioRxiv2019

Hypotonic stress induces fast, reversible degradation of the vimentin cytoskeleton via intracellular calcium release

The dynamic response of the cell to osmotic changes is critical to its physiology and has been widely exploited for cell manipulation. Using 3D-STORM super-resolution microscopy, here we examine the hypotonic stress-induced ultrastructural changes of the cytoskeleton of a common fibroblast cell type. Unexpectedly, we observe a fast, yet reversible dissolution of the vimentin intermediate filament system that precedes ultrastructural changes of the supposedly more dynamic actin and tubulin cytoskeletal systems, as well as changes in cell morphology. In combination with calcium imaging and biochemical analysis, we next show that the vimentin-specific fast cytoskeletal degradation under hypotonic stress is due to proteolysis by the calcium-dependent protease calpain. We find the process to be activated by the hypotonic stress-induced calcium release from intracellular stores, and so is efficiently suppressed by inhibiting any part of the IP3-Ca2+-calpain pathway we establish. Together, our findings highlight an unexpected, fast degradation mechanism for the vimentin cytoskeleton in response to the external stimuli, and point to the significant, yet previously overlooked physiological impacts of hypotonic stress-induced intracellular calcium release on cell ultrastructure and function.

cell biology↗

bioRxiv2019

VRmol: an Integrative Cloud-Based Virtual Reality System to Explore Macromolecular Structure

Past decades have seen much effort devoted to the tool development for biomacromolecule structural visualization and analysis. In recent years, we have witnessed tremendous advancement in computational technologies, especially in graphics and internet. Here we present an integrative cloud-based, virtual reality (VR) system -- VRmol that exploits these technology advances to visualize and study macromolecule structures in various forms. The immersive environment can improve user experiences in visualization, calculation, and editing of complex structures with more intuitive perception. VRmol is also a system with cloud-based capability to directly access disease-related databases, and couples the structure with genomic variation as well corresponding drugs information in a visual interface, thus serving as an integrative platform to aid structure-based translational research and drug design. VRmol is freely available at https://VRmol.net.

bioinformatics↗

bioRxiv2019

A failed top-down control from prefrontal cortex to amygdala in generalized anxiety disorder: evidence from resting-state fMRI with Granger causality analysis

ObjectiveIn generalized anxiety disorder (GAD), abnormal top-down control from prefrontal cortex (PFC) to amygdala is a widely accepted hypothesis through which \"emotional dysregulation model\" may be explained. However, whether and how the PFC directly exerted abnormal top-down control on amygdala remained largely unknown. We aim to investigate the amygdala-based effective connectivity by using Granger causality analysis (GCA).\n\nMethodsThirty-five drug-naive patients with GAD and thirty-six healthy controls (HC) underwent resting-state functional MR imaging. We used seed-based Granger causality analysis to examine the effective connectivity between the bilateral amygdala and the whole brain. The amygdala-based effective connectivity was compared between the two groups.\n\nResultsIn HC, the left middle frontal gyrus exerted inhibitory influence on the right amygdala, while in GAD group, this influence was disrupted (single voxel P < 0.001, Gaussian random field corrected with P < 0.01).\n\nConclusionOur finding might provide new insight into the \"insufficient top-down control\" hypothesis in GAD.

neuroscience↗

bioRxiv2019

Super-resolution displacement mapping of unbound single molecules reveals nanoscale heterogeneities in intracellular diffusivity

Intracellular diffusion underlies vital processes of the cell. However, it remains difficult to elucidate how an average-sized protein diffuses in the cell with good spatial resolution and sensitivity. Here we report single-molecule displacement/diffusivity mapping (SMdM), a super-resolution strategy that enables the nanoscale mapping of intracellular diffusivity through the local statistics of instantaneous displacements of freely diffusing single molecules. We thus show that diffusion in the cytoplasm and in the nucleus to both be spatially heterogeneous at the nanoscale, and such variations in local diffusivity correlate strongly with the ultrastructure of the actin cytoskeleton and the chromosome, respectively. Moreover, we identify the net charge of the diffuser as a key determinant of diffusion rate: intriguingly, the possession of positive, but not negative, net charges significantly impedes diffusion, and the exact degree of slowdown is determined by the specific subcellular environments.

biophysics↗

bioRxiv2019

Comprehensive network analysis reveals alternative splicing-related lncRNAs in hepatocellular carcinoma

A number of recent studies have highlighted the findings that certain lncRNAs are associated with alternative splicing (AS) in tumorigenesis and progression. Although existing work showed the importance of linking certain misregulations of RNA splicing with lncRNAs, a primary concern is the lack of genome-wide comprehensive analysis for their associations. We analyzed an extensive collection of RNA-seq data, quantified 198,619 isoform expressions, and found systematic isoform usage changes between hepatocellular carcinoma (HCC) and normal liver tissue. We identified a total of 1375 splicing switched isoforms and further analyzed their biological functions. To predict which lncRNAs are associated with these AS genes, we integrated the co-expression networks and epigenetic interaction networks collected from text mining and database searching, linking lncRNA modulators such as splicing factors, transcript factors, and miRNAs with their targeted AS genes in HCC. To model the heterogeneous networks in a single framework, we developed a multi-graphic random walk (RWMG) network method to prioritize the lncRNAs associated with AS in HCC. RWMG showed a good performace evaluated by ROC curve based on cross-validation and bootstrapping strategy. As a summary, we identified 31 AS-related lncRNAs including MALAT1 and HOXA11-AS, which have been reported before, as well as some novel lncRNAs such as DNM1P35, HAND2-AS1, and DLX6-AS1. Survival analysis further confirmed the clinical significance of identified lncRNAs.

bioinformatics↗

bioRxiv2019

Genome-wide Association Study of Alcohol Consumption and Use Disorder in Multiple Populations (N = 274,424)

Although alcohol consumption level and alcohol use disorder (AUD) diagnosis are both moderately heritable, their genetic risks and overlap are not well understood. We conducted genome-wide association studies of these traits using longitudinal Alcohol Use Disorder Identification Test-Consumption (AUDIT-C) scores (reflecting alcohol consumption) and AUD diagnoses from electronic health records (EHRs) in a single, large multi-ancestry Million Veteran Program sample. Meta-analysis across population groups (N = 274,424) identified 18 genome-wide significant loci, 5 of which were associated with both traits and 13 with either AUDIT-C (N = 8) or AUD (N = 5). A significant genetic correlation between the traits reflects this overlap. However, downstream analyses revealed biologically meaningful points of divergence. Cell-type group partitioning heritability enrichment analyses indicated that central nervous system was the most significant cell type for AUDIT-C and the only significant cell type for AUD. Polygenic risk scores (PRS) for both traits were associated with alcohol-related disorders in two independent samples. Genetic correlations for 188 non-alcohol-related traits were significantly different for the two traits, as were the phenotypes associated with the traits polygenic risk scores. We conclude that EHR-derived, longitudinal, repeated measures of alcohol consumption level and AUD diagnosis can facilitate genetic discovery and help to elucidate the relationship between drinking level and AUD risk. Finally, although heavy drinking is a key risk factor for AUD, it is not a sufficient cause of the disorder.

genetics↗

bioRxiv2019

Reproducibility of functional brain alterations in major depressive disorder: evidence from a multisite resting-state functional MRI study with 1,434 individuals

Resting-state functional MRI (R-fMRI) studies have demonstrated widespread alterations in brain function in patients with major depressive disorder (MDD). However, a clear and consistent conclusion regarding a repeatable pattern of MDD-relevant alterations is still limited due to the scarcity of large-sample, multisite datasets. Here, we address this issue by including a large R-fMRI dataset with 1,434 participants (709 patients with MDD and 725 healthy controls) from five centers in China. Individual functional activity maps that represent very local to long-range connections are computed using the amplitude of low-frequency fluctuations, regional homogeneity and distance-related functional connectivity strength. The reproducibility analyses involve different statistical strategies, global signal regression, across-center consistency, clinical variables, and sample size. We observed significant hypoactivity in the orbitofrontal, sensorimotor, and visual cortices and hyperactivity in the frontoparietal cortices in MDD patients compared to the controls. These alterations are not affected by different statistical analysis strategies, global signal regression and medication status and are generally reproducible across centers. However, these between-group differences are partially influenced by the episode status and the age of disease onset in patients, and the brain-clinical variable relationship exhibits poor cross-center reproducibility. Bootstrap analyses reveal that at least 400 subjects in each group are required to replicate significant alterations (an extent threshold of P<.05 and a height threshold of P<.001) at 50% reproducibility. Together, these results highlight reproducible patterns of functional alterations in MDD and relevant influencing factors, which provides crucial guidance for future neuroimaging studies of this disorder.

neuroscience↗

bioRxiv2018

FT/FD-GRF5 repression loop directs growth to increase soybean yield

Major advances in crop yield are eternally needed to cope with population growth. To balance vegetative and reproductive growth plays an important role in agricultural yield. To extend vegetative phase can increase crop yield, however, this strategy risks loss of yield in the field as crops may not mature in time before winter come. Here, we identified a repression feedback loop between GmFTL/GmFDL and GmGRF5-1 (Glycine-max-Flowering-Locus-T/Glycine-max-FDL and Glycine-max-GROWTH-REGULATING-FACTOR5-1), which functions as a pivotal regulator in balancing vegetative and reproductive phases in soybean. GmFTL/GmFDL and GmGRF5-1 directly repress gene expression each other. Additionally, GmGRF5-1 enhances vegetative growth by directly enhancing expression of photosynthesis- and auxin synthesis-related genes. To modulate the loop, such as fine-tuning GmFTL expression to trade-off vegetative and reproductive growth, increases substantially soybean yield in the field. Our findings not only uncover the mechanism balancing vegetative and reproductive growth, but open a new window to improve crop yield.

plant biology↗

bioRxiv2018

TANGO1 and SEC12 are co-packaged with procollagen I to facilitate the generation of large COPII carriers

Large COPII-coated vesicles serve to convey the large cargo procollagen I (PC1) from the endoplasmic reticulum (ER). The link between large cargo in the lumen of the ER and modulation of the COPII machinery remains unresolved. TANGO1 is required for procollagen (PC) secretion and interacts with PC and COPII on opposite sides of the ER membrane, but evidence suggests that TANGO1 is retained in the ER, and not included in normal size (<100nm) COPII vesicles. Here we show that TANGO1 is exported out of the ER in large COPII-coated PC1 carriers, and retrieved back to the ER by the retrograde coat, COPI, mediated by the C-terminal RDEL retrieval sequence of HSP47. TANGO1 is known to target the COPII initiation factor SEC12 to ER exit sites through an interacting protein, cTAGE5. SEC12 is important for the growth of COPII vesicles, but it is not sorted into small budded vesicles. We found both cTAGE5 and SEC12 were exported with TANGO1 in large COPII carriers. In contrast to its exclusion from small transport vesicles, SEC12 was particularly enriched around ER membranes and large COPII carriers that contained PC1. We constructed a split GFP system to recapitulate the targeting of SEC12 to PC1 via the luminal domain of TANGO1. The minimal targeting system enriched SEC12 around PC1 and generated large PC1 carriers. We conclude that TANGO1, cTAGE5, and SEC12 are co-packed with PC1 into COPII carriers to increase the size of COPII thus ensuring the capture of large cargo.

cell biology↗

bioRxiv2018

Preliminary Study of the Potential Mechanism of CTSD in the Aging Process of Sepiella japonica: Fundamental Function Analysis

Cathepsin D, a kind of endopeptidase, can degrade peptides and proteins in lysosomes, which are involved in cell apoptosis. Previous transcriptome analysis of optic glands of Sepiella japonica across four growth stages, expression of a cathepsin D-like segment was found to be significantly different. Based on the complete cDNA sequence of S. japonica, the CTSD gene (also called sjCTSD, GenBank accession no. KY745896.1) was cloned using RACE amplification; this gene is 1389 bp in length and encodes proteins composed of 393 amino acids. Spatio-temporal expression profiles of the sjCTSD gene were determined using qPCR assays, which showed that the expression levels of sjCTSD constantly increased across four growth stages in 9 of 11 tissues that were investigated. In the optic glands, as well as pancreas and liver cells, sjCTSD expression levels sharply increased during the post-spawning phase. To investigate the potential role of the sjCTSD gene in aging progress, we constructed the prokaryotic expression vector of pET28a/sjCTSD. After induced by IPTG, the recombinant protein sjCTSD was obtained in the form of inclusion bodies, with a molecular size of approximately 40.3 kDa, the inclusion body of sjCTSD can be converted to soluble protein through the denaturation and renaturation. The results of functional experiments showed that sjCTSD could degrade bovine hemoglobin under acidic conditions, and inhibit the growth of Escherichia coli and Vibrio alginolyticus, which was speculated that the increased expression of sjCTSD may help inhibit the invasion of pathogenic bacteria with the immune function of cuttlefish declines during the aging process. To a certain extent, these results indicated the potential functional role of the sjCTSD gene in the aging process of S. japonica. This study provides insights to further understand the roles of lysosomal proteins on anti-aging effects in S. japonica and other cephalopoda species.

genetics↗