Biology subjects

Woo, J.

Publications and source records attributed to Woo, J..

At least 37 records · Page 2Linked to original sources

bioRxiv2024

Dual-mode action of scalable, high-quality engineered stem cell-derived SIRP alpha extracellular vesicles for treating acute liver failure

Acute liver failure (ALF) is a critical inflammatory condition characterized by rapid hepatocyte death, impaired liver regeneration due to the delayed removal of necroptotic cells, and high mortality rates. This study introduces a novel dual-mode action therapeutic approach using extracellular vesicles expressing Signal Regulatory Protein Alpha (SIRP-EVs) derived from genetically engineered mesenchymal stem cells (MSCs). These SIRP-EVs are designed to concurrently resolve necroptosis and promote liver regeneration. Our studies identified CD47 and SIRP as promising therapeutic targets for ALF. We developed a scalable 3D bioreactor-based process that produces high-purity SIRP-EVs, which preserve MSC properties and achieve significant production levels. SIRP-EVs target both macrophages and necroptotic hepatocytes in ALF models, enhancing macrophage phagocytic activity against necroptotic cells via CD47 blockade and promoting liver regeneration by reprogramming macrophages with MSC-derived cargo. Comprehensive in vitro and in vivo studies demonstrate that SIRP-EVs decrease CD47+ necroptotic cells and promote liver regeneration in ALF models, leading to reduced liver damage markers and enhanced survival rates. These findings highlight the potential of SIRP-EVs as a dual-mode action therapeutic for ALF, offering promising prospects for their application in other inflammatory diseases. Moreover, these results pave the way for advancing engineered EV-based therapies toward clinical implementation.

bioengineering↗

bioRxiv2024

Deep Learning Detection of Subtle Torsional Eye Movements: Preliminary Results

The control of torsional eye position is a key component of ocular motor function. Ocular torsion can be affected by pathologies that involve ocular motor pathways, spanning from the vestibular labyrinth of the inner ears to various regions of the brainstem and cerebellum. Timely and accurate diagnosis enables efficient interventions and management of each case which are crucial for patients with dizziness, vertical double vision, or imbalance. Such detailed evaluation of eye movements may not be possible in all frontline clinical settings, particularly for detecting torsional abnormalities. These abnormalities are often more challenging to identify at the bedside compared to horizontal or vertical eye movements. To address these challenges, we used a dataset of torsional eye movements recorded with video-oculography (VOG) to develop deep learning models for detecting ocular torsion. Our models achieve 0.9308 AUROC and 86.79 % accuracy, leveraging ocular features particularly pertinent to tracking torsional eye position.

neuroscience↗

bioRxiv2024

Proteome-scale tissue mapping using mass spectrometry based on label-free and multiplexed workflows

Multiplexed bimolecular profiling of tissue microenvironment, or spatial omics, can provide deep insight into cellular compositions and interactions in healthy and diseased tissues. Proteome-scale tissue mapping, which aims to unbiasedly visualize all the proteins in a whole tissue section or region of interest, has attracted significant interest because it holds great potential to directly reveal diagnostic biomarkers and therapeutic targets. While many approaches are available, however, proteome mapping still exhibits significant technical challenges in both protein coverage and analytical throughput. Since many of these existing challenges are associated with mass spectrometry-based protein identification and quantification, we performed a detailed benchmarking study of three protein quantification methods for spatial proteome mapping, including label-free, TMT-MS2, and TMT-MS3. Our study indicates label-free method provided the deepest coverages of [~]3500 proteins at a spatial resolution of 50 {micro}m and the highest quantification dynamic range, while TMT-MS2 method holds great benefit in mapping throughput at >125 pixels per day. The evaluation also indicates both label-free and TMT-MS2 provide robust protein quantifications in identifying differentially abundant proteins and spatially co-variable clusters. In the study of pancreatic islet microenvironment, we demonstrated deep proteome mapping not only enables the identification of protein markers specific to different cell types, but more importantly, it also reveals unknown or hidden protein patterns by spatial co-expression analysis.

systems biology↗

bioRxiv2024

The Updated Mouse Universal Genotyping Array Bioinformatic Pipeline Improves Genetic QC in Laboratory Mice

The MiniMUGA genotyping array is a popular tool for genetic QC of laboratory mice and genotyping of samples from most types of experimental crosses involving laboratory strains, particularly for reduced complexity crosses. The content of the production version of the MiniMUGA array is fixed; however, there is the opportunity to improve arrays performance and the associated reports usefulness by leveraging thousands of samples genotyped since the initial description of MiniMUGA in 2020. Here we report our efforts to update and improve marker annotation, increase the number and the reliability of the consensus genotypes for inbred strains and increase the number of constructs that can reliably be detected with MiniMUGA. In addition, we have implemented key changes in the informatics pipeline to identify and quantify the contribution of specific genetic backgrounds to the makeup of a given sample, remove arbitrary thresholds, include the Y Chromosome and mitochondrial genome in the ideogram, and improve robust detection of the presence of commercially available substrains based on diagnostic alleles. Finally, we have made changes to the layout of the report, to simplify the interpretation and completeness of the analysis and added a table summarizing the ideogram. We believe that these changes will be of general interest to the mouse research community and will be instrumental in our goal of improving the rigor and reproducibility of mouse-based biomedical research.

genetics↗

bioRxiv2023

An RXLR effector targets ER-Golgi interface to induce ER stress and necrotic cell death

To achieve successful colonization, the pathogen secretes hundreds of effectors into host cells to manipulate the hosts immune response. Despite numerous studies, the molecular mechanisms underlying effector-induced necrotic cell death remain elusive. In this study, we identified a novel virulent RXLR effector named Pc12 from P. capsici. Pc12 induces necrosis by triggering a distinct ER stress response through its interaction with Rab13-2. Unlike conventional hypersensitive response cell death associated with effector-triggered immunity, Pc12-induced cell death does not coincide with defense gene expression. Instead, it induces the aggregation of ER-resident proteins and confines secretory proteins within the ER. Pc12 interacts with Rab13-2, exhibiting a specific affinity for the active form of Rab13-2. Thus, the complex of Pc12 and Rab13-2 mimics the conformation of the inactive state of Rab13-2, subsequently recruiting the Rab-escort protein (REP). This process results in disruptions in vesicle formation within the ER-Golgi trafficking pathway. Furthermore, the substitution of a single amino acid of Rab13-2 structurally predicted to be crucial for the Pc12 interaction decreased the interaction with Pc12 while maintaining the interaction with REP1. These findings offer valuable insights into the ER stress-mediated cell death as well as a potential strategy for enhancing resistance against pathogens.

plant biology↗

bioRxiv2023

Tumor-wide RNA splicing aberrations generate immunogenic public neoantigens

T-cell-mediated immunotherapies are limited by the extent to which cancer-specific antigens are homogenously expressed throughout a tumor. We reasoned that recurrent splicing aberrations in cancer represent a potential source of tumor-wide and public neoantigens, and to test this possibility, we developed a novel pipeline for identifying neojunctions expressed uniformly within a tumor across diverse cancer types. Our analyses revealed multiple neojunctions that recur across patients and either exhibited intratumor heterogeneity or, in some cases, were tumor-wide. We identified CD8+ T-cell clones specific for neoantigens derived from tumor-wide and conserved neojunctions in GNAS and RPL22, respectively. TCR-engineered CD8+ T-cells targeting these mutations conferred neoantigen-specific tumor cell eradication. Furthermore, we revealed that cancer-specific dysregulation in splicing factor expression leads to recurrent neojunction expression. Together, these data reveal that a subset of neojunctions are both intratumorally conserved and public, providing the molecular basis for novel T-cell-based immunotherapies that address intratumoral heterogeneity.

cancer biology↗

bioRxiv2023

Non-coding sequence variation reveals fragility within interleukin 2 feedback circuitry and shapes autoimmune disease risk

Genetic variants associated with human autoimmune diseases commonly map to non-coding control regions, particularly enhancers that function selectively in immune cells and fine-tune gene expression within a relatively narrow range of values. How such modest, cell-type-selective changes can meaningfully shape organismal disease risk remains unclear. To explore this issue, we experimentally manipulated species-conserved enhancers within the disease-associated IL2RA locus and studied accompanying changes in the progression of autoimmunity. Perturbing distinct enhancers with restricted activity in conventional T cells (Tconvs) or regulatory T cells (Tregs)--two functionally antagonistic T cell subsets--caused only modest, cell-type-selective decreases in IL2ra expression parameters. However, these same perturbations had striking and opposing effects in vivo, completely preventing or severely accelerating disease in a murine model of type 1 diabetes. Quantitative tissue imaging and computational modelling revealed that each enhancer manipulation impinged on distinct IL-2-dependent feedback circuits. These imbalances altered the intracellular signaling and intercellular communication dynamics of activated Tregs and Tconvs, producing opposing spatial domains that amplified or constrained ongoing autoimmune responses. These findings demonstrate how subtle changes in gene regulation stemming from non-coding variation can propagate across biological scales due to non-linearities in intra- and intercellular feedback circuitry, dramatically shaping disease risk at the organismal level.

immunology↗

bioRxiv2023

Remote neuronal activity drives glioma infiltration via Sema4f

The tumor microenvironment (TME) plays an essential role in malignancy and neurons have emerged as a key component of the TME that promotes tumorigenesis across a host of cancers. Recent studies on glioblastoma (GBM) highlight bi-directional signaling between tumors and neurons that propagates a vicious cycle of proliferation, synaptic integration, and brain hyperactivity; however, the identity of neuronal subtypes and tumor subpopulations driving this phenomenon are incompletely understood. Here we show that callosal projection neurons located in the hemisphere contralateral to primary GBM tumors promote progression and widespread infiltration. Using this platform to examine GBM infiltration, we identified an activity dependent infiltrating population present at the leading edge of mouse and human tumors that is enriched for axon guidance genes. High-throughput, in vivo screening of these genes identified Sema4F as a key regulator of tumorigenesis and activity-dependent infiltration. Furthermore, Sema4F promotes the activity-dependent infiltrating population and propagates bi-directional signaling with neurons by remodeling tumor adjacent synapses towards brain network hyperactivity. Collectively, our studies demonstrate that subsets of neurons in locations remote to primary GBM promote malignant progression, while revealing new mechanisms of tumor infiltration that are regulated by neuronal activity.

cancer biology↗

bioRxiv2023

Inhibitory input directs astrocyte morphogenesis through glial GABABR

Communication between neurons and glia plays an important role in establishing and maintaining higher order brain function. Astrocytes are endowed with complex morphologies which places their peripheral processes in close proximity to neuronal synapses and directly contributes to their regulation of brain circuits. Recent studies have shown that excitatory neuronal activity promotes oligodendrocyte differentiation; whether inhibitory neurotransmission regulates astrocyte morphogenesis during development is unknown. Here we show that inhibitory neuron activity is necessary and sufficient for astrocyte morphogenesis. We found that input from inhibitory neurons functions through astrocytic GABABR and that its deletion in astrocytes results in a loss of morphological complexity across a host of brain regions and disruption of circuit function. Expression of GABABR in developing astrocytes is regulated in a region-specific manner by SOX9 or NFIA and deletion of these transcription factors results in region-specific defects in astrocyte morphogenesis, which is conferred by interactions with transcription factors exhibiting region-restricted patterns of expression. Together our studies identify input from inhibitory neurons and astrocytic GABABR as universal regulators of morphogenesis, while further revealing a combinatorial code of region-specific transcriptional dependencies for astrocyte development that is intertwined with activity-dependent processes.

neuroscience↗

bioRxiv2023

Activity-dependent induction of astrocytic Slc22a3 regulates sensory processing through histone serotonylation

Neuronal activity drives global alterations in gene expression within neurons, yet how it directs transcriptional and epigenomic changes in neighboring astrocytes in functioning circuits is unknown. Here we show that neuronal activity induces widespread transcriptional upregulation and downregulation in astrocytes, highlighted by the identification of a neuromodulator transporter Slc22a3 as an activity-inducible astrocyte gene regulating sensory processing in the olfactory bulb. Loss of astrocytic Slc22a3 reduces serotonin levels in astrocytes, leading to alterations in histone serotonylation. Inhibition of histone serotonylation in astrocytes reduces expression of GABA biosynthetic genes and GABA release, culminating in olfactory deficits. Our study reveals that neuronal activity orchestrates transcriptional and epigenomic responses in astrocytes, while illustrating new mechanisms for how astrocytes process neuromodulatory input to gate neurotransmitter release for sensory processing.

neuroscience↗

bioRxiv2023

Isolation may select for earlier and higher peak viral load but shorter duration in SARS-CoV-2 evolution

During the COVID-19 pandemic, human behavior change as a result of nonpharmaceutical interventions such as isolation may have induced directional selection for viral evolution. By combining previously published empirical clinical data analysis and multi-level mathematical modeling, we found that the SARS-CoV-2 variants selected for as the virus evolved from the pre-Alpha to the Delta variant had earlier and higher infectious periods but a shorter duration of infection. Selection for increased transmissibility shapes the viral load dynamics, and the isolation measure is likely to be a driver of these evolutionary transitions. In addition, we showed that a decreased incubation period and an increased proportion of asymptomatic infection were also positively selected for as SARS-CoV-2 mutated to the extent that people did not isolate. We demonstrated that the Omicron variants evolved in these ways to adapt to human behavior. The quantitative information and predictions we present here can guide future responses in the potential arms race between pandemic interventions and viral evolution.

evolutionary biology↗

bioRxiv2022

Disease-causing variant recommendation system for clinical genome interpretation with adjusted scores for artefactual variants

BackgroundIn the process of finding the causative variant of rare diseases (RD), accurate assessment and prioritization of genetic variants is essential. Although quality control (QC) of genetic variants is strictly performed, the presence of artefactual variants in the remaining set of variants can deteriorate the process. Variant QC and prioritization have been treated as separate processes, leading to limited efficiency and risk of misdiagnosis. ResultsWe developed a disease-causing variant recommendation system that integrates quality control into variant prioritization by adjusting scores for artefactual variants. We confirmed that the QC-related features of the variants contribute to a significant performance improvement. For genomic data from 2,878 patients with rare disorders, the recall rate of finding causative variants was 0.961 for the top 5 ranked variants. We also found that our system recognized the anomaly of QC-related features, so that the scores of artifactual variants to be disease-causing were assessed relatively low. ConclusionsIntegration of variant QC and prioritization help reduce the risk of misdiagnosis based on artefactual variants and increase the effectiveness of clinical genome interpretation.

genetics↗

bioRxiv2022

Diversity, composition, altitude, and seasonality of high-altitude windborne migrating mosquitoes in the Sahel: Implications for disease transmission

Recent studies have reported Anopheles mosquitoes captured at high-altitude (40-290 m above ground) in the Sahel. Here, we describe this migration modality across genera and species of African Culicidae and examine its implications for disease transmission and control. As well as Anopheles, six other genera--Culex, Aedes, Mansonia, Mimomyia, Lutzia, and Eretmapodites comprised 90% of the 2,340 mosquitoes captured at altitude. Of the 50 molecularly confirmed species (N=2,107), 33 species represented by multiple specimens were conservatively considered high-altitude windborne migrants, suggesting it is a common migration modality in mosquitoes (31-47% of the known species in Mali), and especially in Culex (45-59%). Overall species abundance varied between 2 and 710 specimens/species (in Ae. vittatus and Cx. perexiguus, respectively). At altitude, females outnumbered males 6:1, and 93% of the females have taken at least one blood meal on a vertebrate host prior to their departure. Most taxa were more common at higher sampling altitudes, indicating that total abundance and diversity are underestimated. High-altitude flight activity was concentrated between June and November coinciding with availability of surface waters and peak disease transmission by mosquitoes. These hallmarks of windborne mosquito migration bolster their role as carriers of mosquito-borne pathogens (MBPs). Screening 921 mosquitoes using pan-Plasmodium assays revealed that thoracic infection rate in these high-altitude migrants was 2.4%, providing a proof of concept that vertebrate pathogens are transported by windborne mosquitoes at altitude. Fourteen of the 33 windborne mosquito species had been reported as vectors to 25 MBPs in West Africa, which represent 32% of the MBPs known in that region and include those that inflict the heaviest burden on human and animal health, such as malaria, yellow fever, dengue, and Rift Valley fever. We highlight five arboviruses that are most likely affected by windborne mosquitoes in West Africa: Rift Valley fever, Onyongnyong, Ngari, Pangola, and Ndumu. We conclude that the study of windborne spread of diseases by migrating insects and the development of surveillance to map the sources, routes, and destinations of vectors and pathogens is key to understand, predict, and mitigate existing and new threats of public health.

ecology↗

bioRxiv2022

Helminth egg derivatives as pro-regenerative immunotherapies

The immune system is increasingly recognized as an important regulator of tissue repair. We developed a regenerative immunotherapy from the helminth Schistosoma mansoni soluble egg antigen (SEA) to stimulate production of interleukin (IL)-4 and other type 2-associated cytokines without negative infection-related sequelae. The regenerative SEA (rSEA) applied to a murine muscle injury induced accumulation of IL-4 expressing T helper cells, eosinophils, and regulatory T cells, and decreased expression of IL-17A in gamma delta ({gamma}{delta}) T cells, resulting in improved repair and decreased fibrosis. Encapsulation and controlled release of rSEA in a hydrogel further enhanced type 2 immunity and larger volumes of tissue repair. The broad regenerative capacity of rSEA was validated in articular joint and corneal injury models. These results introduce a new regenerative immunotherapy approach using natural helminth-derivatives. One-Sentence SummaryHelminth-derived soluble egg antigen regenerative immunotherapies promote tissue repair in multiple injury models.

bioengineering↗

bioRxiv2022

Network analysis reveals a major role for 14q32 cluster miRNAs in determining transcriptional differences between IGHV-mutated and unmutated CLL

Tumour cells from patients with chronic lymphocytic leukaemia (CLL) can express unmutated (U-CLL) or mutated (M-CLL) immunoglobulin heavy chain (IGHV) genes with differing clinical behaviours, variable B cell receptor (BCR) signalling capacity and distinct transcriptional profiles. As it remains unclear to what extent these differences reflect the tumour cells innate pre/post germinal centre origin or their BCR signalling competence, we applied RNA sequencing, small RNA sequencing and DNA methylation array analysis to 38 CLL cases categorised into three groups by IGHV mutational status and BCR signalling capacity. We identified 492 mRNAs and 38 miRNAs differentially expressed between U-CLL and M-CLL, but only 9 mRNAs and 0 miRNAs associated with BCR competence within M-CLL. A significant proportion of the IGHV-associated miRNAs derived from chr14q32 clusters (14/38 (37%)), where all miRNAs were co-expressed with the MEG3 lncRNA, as part of the DLK1-DIO3 genomic imprinted region, a locus of known importance in the pathogenesis of other human tumours. Integrative in silico analysis of miRNA/mRNA data revealed pronounced regulatory potential for the 14q32 miRNAs, potentially accounting for up to 25% of the IGHV-related transcriptome signature. GAB1, a positive regulator of BCR signalling, was predicted to be regulated by five 14q32 miRNAs and we confirmed that two of these (miR-409-3p and miR-411-3p) significantly repressed activity of the GAB1 3UTR. Our analysis demonstrates a potential key role of the 14q32 miRNA locus in the regulation of CLL-related gene regulation.

cancer biology↗

bioRxiv2022

Precise temporal control of GEVI conformations enables the visualization of charge migration in a fluorescent protein resulting in an improved optical response

Rapid and reproducible optical transitions of a fluorescent protein (FP) can be achieved with a Genetically Encoded Voltage Indicator (GEVI) via manipulation of the membrane potential. These transitions revealed novel effects of internal mutations near the chromophore that would not be detected under steady state conditions. Mutating an internal threonine (T203) affected the speed of the voltage-dependent fluorescence transition suggesting a conformational change inside the protein. These optical transitions also demonstrated interplay between internal and externally oriented sidechains of the {beta}-can structure. Replacing the steric hindrance of a phenylalanine near the chromophore with threonine (F165T) did not alter the resting fluorescence but resulted in a more complex fluorescent transition providing evidence for a flexible chromophore undergoing conformational changes. F165T orientation was influenced by the flanking external amino acids at positions 164 and 166 with 164F/165T/166T exacerbating the complexity of the voltage-dependent transition while 164T/165T/166F reduced the flexibility of the chromophore resembling the transition pattern of the original F165 version. Alphafold predictions reveal a threonine switch with different orientations of the F165T internal side chain depending on the direction of the offset in polarity at external positions 164 and 166. The crystal structures of the pH-sensitive FP, Super Ecliptic pHluorin and two derivatives solved in varying pH conditions also indicate interactions between the external protein surface and the internal environment providing another example of a threonine switch near the chromophore at T203. This ability to orient internal sidechains has led to the development of a novel GEVI that gets brighter upon depolarization of the plasma membrane, works at low light levels, is less susceptible to physiological pH, and provides in vivo signals. These observations affecting fluorescent transitions should also prove valuable to the development of any FP-based biosensor.

neuroscience↗

bioRxiv2022

Potent universal-coronavirus therapeutic activity mediated by direct respiratory administration of a Spike S2 domain-specific human neutralizing monoclonal antibody

Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) marks the third novel {beta}-coronavirus to cause significant human mortality in the last two decades. Although vaccines are available, too few have been administered worldwide to keep the virus in check and to prevent mutations leading to immune escape. To determine if antibodies could be identified with universal coronavirus activity, plasma from convalescent subjects was screened for IgG against a stabilized pre-fusion SARS-CoV-2 spike S2 domain, which is highly conserved between human {beta}-coronavirus. From these subjects, several S2-specific human monoclonal antibodies (hmAbs) were developed that neutralized SARS-CoV-2 with recognition of all variants of concern (VoC) tested (Beta, Gamma, Delta, Epsilon, and Omicron). The hmAb 1249A8 emerged as the most potent and broad hmAb, able to recognize all human {beta}-coronavirus and neutralize SARS-CoV and MERS-CoV. 1249A8 demonstrated significant prophylactic activity in K18 hACE2 mice infected with SARS-CoV-2 lineage A and lineage B Beta, and Omicron VoC. 1249A8 delivered as a single 4 mg/kg intranasal (i.n.) dose to hamsters 12 hours following infection with SARS-CoV-2 Delta protected them from weight loss, with therapeutic activity further enhanced when combined with 1213H7, an S1-specific neutralizing hmAb. As little as 2 mg/kg of 1249A8 i.n. dose 12 hours following infection with SARS-CoV Urbani strain, protected hamsters from weight loss and significantly reduced upper and lower respiratory viral burden. These results indicate in vivo cooperativity between S1 and S2 specific neutralizing hmAbs and that potent universal coronavirus neutralizing mAbs with therapeutic potential can be induced in humans and can guide universal coronavirus vaccine development.

immunology↗

bioRxiv2022

Different efficacies of neutralizing antibodies and antiviral drugs on SARS-CoV-2 Omicron subvariants, BA.1 and BA.2

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron subvariant BA.2 has spread in many countries, replacing the earlier Omicron subvariant BA.1 and other variants. Here, using a cell culture infection assay, we quantified the intrinsic sensitivity of BA.2 and BA.1 compared with other variants of concern, Alpha, Gamma, and Delta, to five approved-neutralizing antibodies and antiviral drugs. Our assay revealed the diverse sensitivities of these variants to antibodies, including the loss of response of both BA.1 and BA.2 to casirivimab and of BA.1 to imdevimab. In contrast, EIDD-1931 and nirmatrelvir showed a more conserved activities to these variants. The viral response profile combined with mathematical analysis estimated differences in antiviral effects among variants in the clinical concentrations. These analyses provide essential evidence that gives insight into variant emergences impact on choosing optimal drug treatment.

microbiology↗