Biology subjects

Chu, J.

Publications and source records attributed to Chu, J..

At least 37 records · Page 2Linked to original sources

bioRxiv2024

Spyglass: a data analysis framework for reproducible and shareable neuroscience research

Scientific progress depends on reliable and reproducible results. Progress can be accelerated when data are shared and re-analyzed to address new questions. Current approaches to storing and analyzing neural data involve bespoke formats and software that make replication and reuse of data difficult. To address these challenges, we created Spyglass, an open-source data management and analysis framework written in Python. Spyglass provides reproducible pipelines for common neuroscience analyses and sharing of raw data, intermediate analyses, and final results within and across labs. Spyglass uses the Neurodata Without Borders (NWB) standard and includes pipelines for spectral filtering, spike sorting, pose tracking, and neural decoding. Spyglass can be extended to apply existing and newly developed pipelines to datasets from multiple sources. We demonstrate these features in the context of a cross-laboratory replication by applying advanced state space decoding algorithms to publicly available data. New users can try out Spyglass on a Jupyter Hub hosted by HHMI and 2i2c: https://spyglass.hhmi.2i2c.cloud/.

neuroscience↗

bioRxiv2024

Mannose Supplementation Curbs Liver Steatosis and Fibrosis in Murine MASH by Inhibiting Fructose Metabolism

Metabolic dysfunction-associated steatohepatitis (MASH) can progress to cirrhosis and liver cancer. There are no approved medical therapies to prevent or reverse disease progression. Fructose and its metabolism in the liver play integral roles in MASH pathogenesis and progression. Here we focus on mannose, a simple sugar, which dampens hepatic stellate cell activation and mitigates alcoholic liver disease in vitro and in vivo. In the well-validated FAT-MASH murine model, oral mannose supplementation improved both liver steatosis and fibrosis at low and high doses, whether administered either at the onset of the model ("Prevention") or at week 6 of the 12-week MASH regimen ("Reversal"). Thein vivo anti-fibrotic effects of mannose supplementation were validated in a second model of carbon tetrachloride-induced liver fibrosis. In vitro human and mouse primary hepatocytes revealed that the anti-steatotic effects of mannose are dependent on the presence of fructose, which attenuates expression of ketohexokinase (KHK), the main enzyme in fructolysis. KHK is decreased with mannose supplementation in vivo and in vitro, and overexpression of KHK abrogated the anti-steatotic effects of mannose. Our study identifies mannose as a simple, novel therapeutic candidate for MASH that mitigates metabolic dysregulation and exerts anti-fibrotic effects.

cell biology↗

bioRxiv2023

Scm6A: A fast and low-cost method for quantifying m6A modifications at the single-cell level

It is widely accepted that m6A exhibits significant intercellular specificity, which poses challenges for its detection using existing m6A quantitative methods. In this study, we introduce Scm6A, a machine learning-based approach for single-cell m6A quantification. Scm6A leverages input features derived from the expression levels of m6A trans regulators and cis sequence features, and found that Scm6A offers remarkable prediction efficiency and reliability. To further validate the robustness and precision of Scm6A, we applied a winscore-based m6A calculation method to conduct m6A-seq analysis on CD4+ and CD8+ T-cells isolated through magnetic-activated cell sorting (MACS). Subsequently, we employed Scm6A for analysis on the same samples. Notably, the m6A levels calculated by Scm6A exhibited a significant positive correlation with m6A quantified through m6A-seq in different cells isolated by MACS, providing compelling evidence for Scm6As reliability. We also used the scm6A-seq method to validate the reliability of our approach. Additionally, we performed single-cell level m6A analysis on lung cancer tissues as well as blood samples from COVID-19 patients, and demonstrated the landscape and regulatory mechanisms of m6A in different T-cell subtypes from these diseases. In summary, our work has yielded a novel, dependable, and accurate method for single-cell m6A detection. We are confident that Scm6A will have broad applications in the realm of m6A-related research.

bioinformatics↗

bioRxiv2023

Pan-cancer Analysis Reveals m6A Variation and Cell-specific Regulatory Network in Different Cancer Types

As the most abundant mRNA modification in mRNA, N6-methyladenosine (m6A) plays a crucial role in RNA fate, impacting cellular and physiological processes in various tumor types. However, our understanding of the function and role of the m6A methylome in tumor heterogeneity remains limited. Herein, we collected and analyzed m6A methylomes across nine human tissues from 97 m6A-seq and RNA-seq samples. Our findings demonstrate that m6A exhibits different heterogeneity in most tumor tissues compared to normal tissues, which contributes to the diverse clinical outcomes in different cancer types. We also found that the cancer type-specific m6A level regulated the expression of different cancer-related genes in distinct cancer types. Utilizing a novel and reliable method called "m6A-express", we predicted m6A- regulated genes and revealed that cancer type-specific m6A-regulated genes contributed to the prognosis, tumor origin and infiltration level of immune cells in diverse patient populations. Furthermore, we identified cell-specific m6A regulators that regulate cancer-specific m6A and constructed a regulatory network. Experimental validation was performed, confirming that the cell-specific m6A regulator CAPRIN1 controls the m6A level of TP53. Overall, our work reveals the clinical relevance of m6A in various tumor tissues and explains how such heterogeneity is established. These results further suggest the potential of m6A for cancer precision medicine for patients with different cancer types.

bioinformatics↗

bioRxiv2023

ntsm: an alignment-free, ultra low coverage, sequencing technology agnostic, intraspecies sample comparison tool for sample swap detection

BackgroundDue to human error, sample swapping in large cohort studies with heterogeneous data types (e.g. mix of Oxford Nanopore, Pacific Bioscience, Illumina data, etc.) remains a common issue plaguing large-scale studies. At present, all sample swapping detection methods require costly and unnecessary (e.g. if data is only used for genome assembly) alignment, positional sorting, and indexing of the data in order to compare similarly. As studies include more samples and new sequencing data types, robust quality control tools will become increasingly important. FindingsThe similarity between samples can be determined using indexed k-mer sequence variants. To increase statistical power, we use coverage information on variant sites, calculating similarity using a likelihood ratio-based test. Per sample error rate, and coverage bias (i.e. missing sites) can also be estimated with this information, which can be used to determine if a spatially indexed PCA-based pre-screening method can be used, which can greatly speed up analysis by preventing exhaustive all-to-all comparisons. ConclusionsBecause this tool processes raw data, is faster than alignment, and can be used on very low coverage data, it can save an immense degree of computational resources in standard QC pipelines. It is robust enough to be used on different sequencing data types, important in studies that leverage the strengths of different sequencing technologies. In addition to its primary use case of sample-swap detection, this method provides other useful information useful in QC, such as error rate and coverage bias, as well as population-level PCA ancestry analysis visualization.

bioinformatics↗

bioRxiv2023

Forsythoside A blocks the infection of Avian Infectious Bronchitis Virus through binding S1 subunit

Avian infectious bronchitis (IB) is an acute highly contagious respiratory infectious disease caused by avian infectious bronchitis virus (IBV), causing serious economic losses to poultry industry. Forsythiaside A (FTA) is a phenylethanol glycoside compound in the fruit of Forsythia suspensa, which is widely used for inflammatory respiratory tract disease, including viral infected inflammation. In this study, the mechanism of FTA against IBV was researched. To determine the inhibitory effect of FTA on IBV infection and the interaction between FTA and IBV particles, immunofluorescence assay, plaque assay, RT-qPCR and enzyme-linked immunosorbent assay (ELISA) were performed and it was discovered that FTA inhibited IBV infection and viral attachment to host receptor under non-cytotoxic concentrations, and FTA may bind to virus particles. Then ELISA and isothermal titration calorimetry (ITC) were employed to further study the interaction between FTA and S protein. The results indicated that FTA bound to S1 subunit rather S2 subunit. Besides, the molecular docking showed that FTA form hydrogen bonds with eight amino acids in the S1 subunit. When these amino acids were mutated. the affinity disappeared between the S1 mutant and FTA. In conclusion, FTA prevents IBV infection by blocking viral attachment and contacts with the S1, but not the S2 subunit of IBV. At last, the bind sites were identified by comparing the affinity of wild S1 and mutant S1.

pharmacology and toxicology↗

bioRxiv2023

The Neonatal Gyrencephalic Cortex Maintains Regionally Distinct Streams of Neuroblasts

Neurodevelopmental mechanisms have evolved to support the formation of diverse brain structures, such as in humans, during the perinatal period. Here, we demonstrate that neonatal gyrencephalic brains harbor an expanded subventricular zone, termed the Arc, defined by tiered arrangement of doublecortin (DCX)-expressing neuroblasts and vascular enrichment at the ventricular wall. The Arc is the origin of dorsal and ventral populations of migratory neuroblasts that target multiple regions involved in higher cognitive functions. Arc-derived migratory streams, primarily from the caudal ganglionic eminence, are composed of diverse neuronal subtypes with distinct spatial and migratory-receptor profiles. Our findings indicate the Arc is a structure present in phylogenetically divergent species that supports the expansion of postnatal neuronal migration, contributing to a protracted formation of cortical circuits in gyrencephalic brains. One-Sentence SummaryThe ventricular cytoarchitecture of gyrencephalic brains supports an ongoing supply of migratory neurons to the neonatal cortex.

neuroscience↗

bioRxiv2023

All-optical interrogation of brain-wide activity in freely swimming larval zebrafish

We introduce an all-optical technique that enables volumetric imaging of brain-wide calcium activity and targeted optogenetic stimulation of specific brain regions in freely swimming larval zebrafish. The system consists of three main components: a 3D tracking module, a dual color fluorescence imaging module, and a real-time activity manipulation module. Our approach uses a sensitive genetically encoded calcium indicator in combination with a long Stokes shift red fluorescence protein as a reference channel, allowing the extraction of Ca2+ activity from signals contaminated by motion artifacts. The method also incorporates rapid 3D image reconstruction and registration, facilitating real-time selective optogenetic stimulation of different regions of the brain. By demonstrating that selective light activation of the midbrain regions in larval zebrafish could reliably trigger biased turning behavior and changes of brain-wide neural activity, we present a valuable tool for investigating the causal relationship between distributed neural circuit dynamics and naturalistic behavior. HighlightsO_LIWe develop an all-optical technique that enables simultaneous whole brain imaging and optogenetic manipulation of selective brain regions in freely behaving larval zebrafish. C_LIO_LIA combination of a genetically encoded calcium indicator and a long Stokes-shift red fluorescence protein, together with the adaptive filter algorithm, enables us to reliably distinguish calcium activity from motion-induced signal contamination. C_LIO_LIRapid 3D image reconstruction and registration enables real-time targeted optogenetic stimulation of distinct brain regions in a freely swimming larval zebrafish. C_LI

neuroscience↗

bioRxiv2023

Tomato CYP94C1 terminates jasmonate signaling during fruit ripening by inactivating bioactive jasmonoyl-L-isoleucine

Ripe fruits are more susceptible to necrotrophic pathogens than unripe fruits. Although this phenomenon is widespread across different fruit species and results in substantial economic losses, the underlying mechanism is still poorly understood. Previous studies revealed that ethylene (ET) is a key signal controlling climacteric fruit ripening and that jasmonate (JA) regulates plant resistance to necrotrophs. We investigated the function of tomato cytochrome P450 94 (CYP94) family genes in JA signaling and report here that ET-mediated ripening suppresses JA-mediated defense by promoting the deactivation of bioactive JA-Ile. ETHYLENE-INSENSITIVE 3 (EIN3)/EIN3-LIKE (EIL) transcription factors directly activated CYP94C1 to convert JA-Ile to its inactive form 12-COOH-JA-Ile, thereby terminating JA signaling during fruit ripening. Mutation of CYP94C1 led to increased resistance of ripe fruits to the necrotrophic pathogen Botrytis cinerea without affecting the ripening process. Additionally, the master transcription factor MYC2 directly activated two other CYP94 members CYP94B1 and CYP94B2 to convert JA-Ile to its less active form 12-OH-JA-Ile, thereby attenuating JA signaling in wounded leaves. Simultaneous mutation of CYP94B1 and CYP94B2 increased the resistance of leaves to B. cinerea. Thus, differences in the expression and enzymatic activities of CYP94 family gene members precisely control JA-mediated defense responses in tomato.

plant biology↗

bioRxiv2023

Genetic control of cell layer interactions in plants via tissue mechanics

Plant development depends on coordination of growth between different cell layers. Coordination may be mediated by molecular signalling or mechanical connectivity between cells, but evidence for genetic control via direct mechanics has been lacking. We show that a brassinosteroid-deficient dwarf mutant of the aquatic plant Utricularia gibba has twisted internal tissue, likely caused by a mechanical constraint from a slow-growing epidermis creating tissue stresses. This conclusion is supported by showing that inhibition of brassinosteroid action in an Arabidopsis mutant compromised for cell adhesion, enhances epidermal crack formation, an indicator of increased tissue tension. Thus, genes driving brassinosteroid synthesis can promote growth of internal tissue by reducing mechanical epidermal constraint, showing that tissue mechanics plays a key role in coordinating growth between cell layers. One-Sentence SummaryInternal twists in a mutant carnivorous plant reveal how genes control growth via tissue mechanics.

plant biology↗

bioRxiv2023

ATP-releasing SWELL1 channel in spinal microglia contributes to neuropathic pain

Following peripheral nerve injury, extracellular ATP-mediated purinergic signaling is crucial for spinal cord microglia activation and neuropathic pain. However, the mechanisms of ATP release remain poorly understood. Here, we show that volume-regulated anion channel (VRAC) is an ATP-releasing channel and is activated by inflammatory mediator sphingosine-1-phosphate (S1P) in microglia. Mice with microglia-specific deletion of Swell1 (also known as Lrrc8a), a VRAC essential subunit, had reduced peripheral nerve injury-induced increase in extracellular ATP in spinal cord. The mutant mice also exhibited decreased spinal microgliosis, dorsal horn neuronal hyperactivity, and both evoked and spontaneous neuropathic pain-like behaviors. We further performed high-throughput screens and identified an FDA-approved drug dicumarol as a novel and potent VRAC inhibitor. Intrathecal administration of dicumarol alleviated nerve injury-induced mechanical allodynia in mice. Our findings suggest that ATP-releasing VRAC in microglia is a key spinal cord determinant of neuropathic pain and a potential therapeutic target for this debilitating disease.

physiology↗

bioRxiv2022

Bright and sensitive red voltage indicators for imaging action potentials in brain slices and pancreatic islets

As fast developing tools for observing cellular membrane potential, red-emitting genetically encoded voltage indicators (GEVIs) reduce auto-fluorescence background, allow multiplexed recordings, and enable all-optical electrophysiology, but have been limited by either insensitivity or dimness. Here, we report a pair of red GEVIs, Cepheid1b/s, with improved sensitivity, brightness, and photostability. Cepheid1 indicators faithfully report cellular excitability in pancreatic islets and neural activity in acute brain slices.

neuroscience↗

bioRxiv2022

Splicing quality control mediated by DHX15 and itsG-patch activator, SUGP1

Pre-mRNA splicing is surveilled at different stages by quality control (QC) mechanisms. The leukemia-associated DExH-box family helicase hDHX15/scPrp43, is known to disassemble spliceosomes after splicing. Here, using rapid protein depletion and analysis of nascent and mature RNA to enrich for direct effects, we identified a widespread splicing QC function for DHX15 in human cells, consistent with recent in vitro studies. We found that suboptimal introns with weak splice sites, multiple branch points, and cryptic introns are repressed by DHX15, suggesting a general role in promoting splicing fidelity. We identified SUGP1 as a G-patch factor that activates DHX15s splicing QC function. This interaction is dependent on both DHX15s ATPase activity and on SUGP1s ULM domain. Together, our results support a model in which DHX15 plays a major role in splicing QC when recruited and activated by SUGP1.

molecular biology↗

bioRxiv2022

Characterization of Three Variants of SARS-CoV-2 in vivo Shows Host-Dependent Pathogenicity in Hamsters

Animal models are used in preclinical trials to test vaccines, antivirals, monoclonal antibodies, and immunomodulatory drug therapies against SARS-CoV-2. However, these drugs often do not produce equivalent results in human clinical trials. Here, we show how different animal models infected with some of the most clinically relevant SARS-CoV-2 variants, WA1/2020, B.1.617.2/Delta, B.1.1.529/Omicron and BA5.2/Omicron, have independent outcomes. We show that in mice, B.1.617.2 is more pathogenic, followed by WA1, while B.1.1.529 showed an absence of clinical signs. Only B.1.1.529 was able to infect C57BL/6J mice, which lack the human ACE2 receptor. B.1.1.529-infected ACE2 mice had different T cell profiles compared to infected K18-hACE2 mice, while viral shedding profiles and viral titers in lungs were similar between the ACE2 and the C57BL/6J mice. These data suggest B.1.1.529 virus adaptation to a new host and shows that asymptomatic carriers can accumulate and shed virus. Next, we show how B.1.617.2, WA1 and BA5.2/Omicron have similar viral replication kinetics, pathogenicity, and viral shedding profiles in hamsters, demonstrating that the increased pathogenicity of B.1.617.2 observed in mice is host-dependent. Overall, these findings suggest that small animal models are useful to parallel human clinical data, but the experimental design places an important role in interpreting the data. IMPORTANCEThere is a need to investigate SARS-CoV-2 variants phenotypes in different animal models due to the lack of reproducible outcomes when translating experiments to the human population. Our findings highlight the correlation of clinically relevant SARS-CoV-2 variants in animal models with human infections. Experimental design and understanding of correct animal models are essential to interpreting data to develop antivirals, vaccines, and other therapeutic compounds against COVID-19.

microbiology↗

bioRxiv2022

The metabolite alpha-ketobutyrate increases health and life spans by activating AMPK

Aging is a complex process that is directly related to human health and disease. The extraordinary finding that aging is malleable, as shown in model organisms whose life and health spans are extended by specific gene mutations or dietary or pharmacological perturbations 1-3, has offered enormous hope for our understanding and treatment of aging and related diseases. Although many molecules have been identified that can extend the lifespan of model organisms, few have been shown to alleviate age-related symptoms or illness in mammals 4. Here we show that supplementation with the endogenous metabolite -ketobutyrate (-KB) increases the lifespan of adult C. elegans. Using Gelfree DARTS-PROTOMAP, we identified microtubule-actin cross-linking factor (MACF1) that was protected against proteolysis in the presence of -KB. MACF1 belongs to the spectraplakin family of giant, evolutionarily conserved proteins with versatile functions 5, but their link to longevity regulation has not been explored. -KBs longevity effect in C. elegans is abrogated by loss-of-function mutation in vab-10, encoding the worm ortholog of mammalian MACF1 6. Like -KB treatment, vab-10 knockdown activates AMP-activated protein kinase (AMPK), and AMPK is required for -KB effects on longevity. The findings suggest a model in which -KB increases longevity by activating AMPK via VAB-10/MACF1 modulation. -KB also delays aging in mammals, increasing the lifespan of aged male mice and the healthspan of both male and female animals. Targeting of broadly expressed scaffolding proteins in connection to cellular energy homeostasis seems to be a clever way that nature has devised for metabolite signals to impinge upon multiple organ and tissue systems, which may have utility for controlling aging and related diseases.

physiology↗

bioRxiv2022

A Draft Human Pangenome Reference

The Human Pangenome Reference Consortium (HPRC) presents a first draft human pangenome reference. The pangenome contains 47 phased, diploid assemblies from a cohort of genetically diverse individuals. These assemblies cover more than 99% of the expected sequence and are more than 99% accurate at the structural and base-pair levels. Based on alignments of the assemblies, we generated a draft pangenome that captures known variants and haplotypes, reveals novel alleles at structurally complex loci, and adds 119 million base pairs of euchromatic polymorphic sequence and 1,529 gene duplications relative to the existing reference, GRCh38. Roughly 90 million of the additional base pairs derive from structural variation. Using our draft pangenome to analyze short-read data reduces errors when discovering small variants by 34% and boosts the detected structural variants per haplotype by 104% compared to GRCh38-based workflows, and by 34% compared to using previous diversity sets of genome assemblies.

genomics↗

bioRxiv2022

A CRISPR-engineered Isogenic Model Reveals Altered Neuronal Phenotypes of the 22q11.2 A-B Syndromic Deletion

The 22q11.2 deletion syndrome (22q11.2DS), associated with congenital and neuropsychiatric anomalies, is the most common copy number variant (CNV)-associated syndrome. Patient-derived, induced pluripotent stem cell (iPS) models have provided important insight into the mechanisms of phenotypic features of this condition. However, patient-derived iPS models may harbor underlying genetic heterogeneity that can confound analysis of pathogenic CNV effects. Furthermore, the [~]1.5 Mb "A-B" deletion at this locus is inherited at higher frequency than the more common [~]3 Mb "A-D" deletion, but remains under-studied due to lack of relevant models. To address these issues, here we leveraged a CRISPR-based strategy in Cas9-expressing iPS cells to engineer novel isogenic models of the 22q11.2 "A-B" deletion. After in vitro differentiation to excitatory neurons, integrated transcriptomic and cell surface proteomics identified deletion-associated alterations in surface adhesion and cell signaling. Furthermore, implantation of iPS-derived neuronal progenitor cells into the cortex of neonatal mice found accelerated neuronal maturation within a relevant microenvironment. Taken together, our results suggest pathogenic mechanisms of the 22q11.2 "A-B" deletion in driving neuronal and neurodevelopmental phenotypes, both in vitro and in vivo. We further propose that the isogenic models generated here will provide a unique resource to study this less-common variant of the 22q11.2 microdeletion syndrome.

genetics↗

bioRxiv2022

DESIGN OF A CHIMERIC ACE-2/Fc-SILENT FUSION PROTEIN WITH ULTRAHIGH AFFINITY AND NEUTRALIZING CAPACITY FOR SARS-CoV-2 VARIANTS

As the coronavirus SARS-CoV-2 continues to mutate into Variants of Concern (VOC), there is a growing and urgent need to develop effective antivirals to combat the newly emerged infectious disease COVID-19. Recent data indicate that monoclonal antibodies developed early in the pandemic are no longer capable of effectively neutralizing currently active VOCs. This report describes the design of a class of variant-agnostic chimeric molecules consisting of an Angiotensin Converting Enzyme-2 (ACE-2) domain mutated to retain ultrahigh affinity binding to a wide variety of SARS-CoV-2 variants, coupled to an Fc-silent immunoglobulin domain that eliminates antibody-dependent enhancement (ADE) and simultaneously extends biological half-life compared to existing mABs. Molecular modeling revealed that ACE-2 mutations L27, V34 and E90 resulted in ultrahigh affinity binding of the LVE-ACE-2 domain to the widest variety of VOCs, with KDs of 93 pM, 507 pM and 73 pM for binding to the Alpha B1.1.7, Delta B.1.617.2 and Omicron B.1.1.529 variants, and notably, 78fM affinity to the Omicron BA.2 variant, respectively. Surrogate viral neutralization assays (sVNT) revealed titers of[≥] 4.9ng/ml, for neutralization of recombinant viral proteins corresponding to the Alpha, Delta and Omicron variants. The values above were obtained with LVE-ACE-2/mAB chimeras containing the Y-T-E sequence that enhances binding to the FcRn receptor, which in turn is expected to extend biological half-life 3-4-fold. It is proposed that this new class of chimeric ACE-2/mABs will constitute variant-agnostic and cost-effective prophylactics against SARS-CoV-2, particularly when administered by nasal delivery systems.

microbiology↗