Biology subjects

Lee, J.

Publications and source records attributed to Lee, J..

At least 379 records · Page 21Linked to original sources

bioRxiv2024

Synergistic Cytotoxicity of Permethrin and N,N-Diethyl-Meta-Toluamide on Sinonasal Epithelial Cells

BackgroundN,N-Diethyl-Meta-Toluamide (DEET) and permethrin are pesticides commonly used in combination due to their synergistic insecticidal and repellent properties. This study investigates whether simultaneous exposure to these compounds elicits synergistic cytotoxicity in sinonasal epithelial cells (SNECs). Material and MethodsEthmoid sinus mucosal specimens were procured from eight patients during endoscopic sinus surgery. SNECs were expanded on culture plates and exposed to various concentrations of DEET and permethrin (0-5m), individually and concurrently, for up to 156 hours. Experiments were replicated in triplicates and cell viability was recorded every 2 hours using IncuCyte real-time cell imaging system. Synergy score was calculated on the basis of Loewe additivity synergy finder model. ResultsDEET and permethrin exhibited synergistic cytotoxicity across all eight tissues, albeit with variations in onset and magnitude. Peak synergy was observed at 144h for tissue 1 (SLoewe 4.2, 95% CI 1.8-7.3; [permethrin concentration, DEET concentration] [2.5M, 1.25M]), 48h for tissue 2 (19.9, 16.5-23.6; [1.25, 0.625]), 144h for tissue 3 (15.2, 3.9-30.4; [0.625, 1.25]), 144h for tissue 4 (6.4, -3.6 to 18.0; [0.625, 0.625]), 48h for tissue 5 (10.1, 8.9-12.5; [0.625, 1.25]), 96h for tissue 6 (24.7, 12.2-36.3; [0.625, 0.625]), 48h for tissue 7 (47.7, 29.6-62.2; [0.625, 1.25]), and 96h for 8 (47.4, 26.8-67.0; [0.625, 0.625]). ConclusionThe concurrent exposure of DEET and permethrin can lead to synergistic cytotoxicity in sinonasal epithelia. Further research is warranted in preclinical animal models to explore whether this synergy accelerates the pathogenesis of chronic rhinosinusitis.

pharmacology and toxicology↗

bioRxiv2024

Molecular basis of convergent evolution of ACE2 receptor utilization among HKU5 coronaviruses

DPP4 was considered a canonical receptor for merbecoviruses until the recent discovery of African bat-borne MERS-related coronaviruses using ACE2. The extent and diversity with which merbecoviruses engage ACE2 and their receptor species tropism remain unknown. Here, we reveal that HKU5 enters host cells utilizing Pipistrellus abramus (P.abr) and several non-bat mammalian ACE2s through a binding mode distinct from that of any other known ACE2-using coronaviruses. These results show that several merbecovirus clades independently evolved ACE2 utilization, which appears to be a broadly shared property among these pathogens, through an extraordinary diversity of ACE2 recognition modes. We show that MERS-CoV and HKU5 have markedly distinct antigenicity, due to extensive genetic divergence, and identified several HKU5 inhibitors, including two clinical compounds. Our findings profoundly alter our understanding of coronavirus evolution and pave the way for developing countermeasures against viruses poised for human emergence.

biochemistry↗

bioRxiv2024

Comparative transcriptomics in ferns reveals key innovations and divergent evolution of secondary cell wall

Despite ferns being crucial to understanding plant evolution, their large and complex genomes has kept their genetic landscape largely uncharted, with only a handful of genomes sequenced and sparse transcriptomic data. Addressing this gap, we generated extensive RNA-sequencing data for multiple organs across 22 representative species over the fern phylogeny, assembling high-quality transcriptomes. These data facilitated the construction of a time-calibrated fern phylogeny covering all major clades, revealing numerous whole-genome duplications and highlighting the uniqueness of fern genetics, with half of the uncovered gene families being fern-specific. Our investigation into fern cell walls through biochemical and immunological analyses identified occurrences of the lignin syringyl unit and its independent evolution in ferns. Moreover, the discovery of an unusual sugar in fern cell walls hints at a divergent evolutionary path in cell wall biochemistry, potentially driven by gene duplication and sub-functionalization. We provide an online database preloaded with genomic and transcriptomic data for ferns and other land plants, which we used to identify an independent evolution of lignocellulosic gene modules in ferns. Our data provide a framework for the unique evolutionary path that ferns have navigated since they split from the last common ancestor of euphyllophytes more than 360 million years ago.

plant biology↗

bioRxiv2024

Molecular basis of pathogenicity of the recently emerged FCoV-23 coronavirus

The ability of coronaviruses to recombine and cross species barriers affects human and animal health globally and is a pandemic threat. FCoV-23 is a recently emerged, highly pathogenic recombinant coronavirus responsible for a widespread outbreak of feline infectious peritonitis (FIP) likely linked to in-host viral evolution. Here, we report cryoEM structures of two FCoV-23 spike (S) isoforms explaining that the in-host loss of domain 0 observed in clinical samples enhances entry into cells and fusogenicity by facilitating protease access, leading to biotype switching and lethality. We show that FCoV-23 can use several aminopeptidase N (APN) orthologs as receptors and reveal the molecular determinants of receptor species tropism, including a glycan modulating human receptor utilization. We define antigenic relationships among alphacoronaviruses infecting humans and other mammalian species and identify a cross-reactive alphacoronavirus monoclonal antibody inhibiting FCoV-23 pseudovirus entry, paving the way for vaccine and therapeutic development targeting this highly pathogenic virus.

biochemistry↗

bioRxiv2024

Tracking clonal evolution of drug resistance in ovarian cancer patients by exploiting structural variants in cfDNA

Drug resistance is the major cause of therapeutic failure in high-grade serous ovarian cancer (HGSOC). Yet, the mechanisms by which tumors evolve to drug resistant states remains largely unknown. To address this, we aimed to exploit clone-specific genomic structural variations by combining scaled single-cell whole genome sequencing with longitudinally collected cell-free DNA (cfDNA), enabling clonal tracking before, during and after treatment. We developed a cfDNA hybrid capture, deep sequencing approach based on leveraging clone-specific structural variants as endogenous barcodes, with orders of magnitude lower error rates than single nucleotide variants in ctDNA (circulating tumor DNA) detection, demonstrated on 19 patients at baseline. We then applied this to monitor and model clonal evolution over several years in ten HGSOC patients treated with systemic therapy from diagnosis through recurrence. We found drug resistance to be polyclonal in most cases, but frequently dominated by a single high-fitness and expanding clone, reducing clonal diversity in the relapsed disease state in most patients. Drug-resistant clones frequently displayed notable genomic features, including high-level amplifications of oncogenes such as CCNE1, RAB25, NOTCH3, and ERBB2. Using a population genetics Wright-Fisher model, we found evolutionary trajectories of these features were consistent with drug-induced positive selection. In select cases, these alterations impacted selection of secondary lines of therapy with positive patient outcomes. For cases with matched single-cell RNA sequencing data, pre-existing and genomically encoded phenotypic states such as upregulation of EMT and VEGF were linked to drug resistance. Together, our findings indicate that drug resistant states in HGSOC pre-exist at diagnosis and lead to dramatic clonal expansions that alter clonal composition at the time of relapse. We suggest that combining tumor single cell sequencing with cfDNA enables clonal tracking in patients and harbors potential for evolution-informed adaptive treatment decisions.

cancer biology↗

bioRxiv2024

Accurate antibody loop structure prediction enables zero-shot design of target-specific antibodies

Protein loops, characterized by their versatile structures with varying sizes and shapes, can recognize a wide range of targets with high specificity and affinity. The variable loops of the antibody complementarity-determining region (CDR) are particularly crucial for immune responses and therapeutic applications due to their effective target recognition capabilities. Accurate structure prediction of these antibody loops is essential for the efficient in silico design of target-binding antibodies for therapeutic or industrial use. However, predicting antibody loop structures is challenging due to the lack of evolutionary information from related proteins. Thus, a successful ab initio structure prediction method, which operates without structural templates or related sequences, is crucial for the effective design of antibody loop-mediated interactions. This study demonstrates that highly accurate antibody loop structure prediction enables the effective zero-shot design of target-binding antibody loops. The performance of loop design has been shown to depend on the accuracy of ab initio loop structure prediction, as tested with two versions of our design model. The high affinity, diversity, novelty, and specificity of the antibody loops designed with these new methods were validated experimentally on four target proteins.

bioengineering↗

bioRxiv2024

Vagal TRPV1+ sensory neurons regulate myeloid cell dynamics and protect against influenza virus infection

Influenza viruses are a major global cause of morbidity and mortality. Vagal TRPV1+ nociceptive sensory neurons, which innervate the airways, are known to mediate defenses against harmful agents. However, their function in lung antiviral defenses remains unclear. Our study reveals that both systemic and vagal-specific ablation of TRPV1+ nociceptors reduced survival in mice infected with influenza A virus (IAV), despite no significant changes in viral burden or weight loss. Mice lacking nociceptors showed exacerbated lung pathology and elevated levels of pro-inflammatory cytokines. The increased mortality was not attributable to the loss of the TRPV1 ion channel or neuropeptides CGRP or substance P. Immune profiling through flow cytometry and single-cell RNA sequencing identified significant nociceptor deficiency-mediated changes in the lung immune landscape, including an expansion of neutrophils and monocyte-derived macrophages. Transcriptional analysis revealed impaired interferon signaling in these myeloid cells and an imbalance in distinct neutrophil sub-populations in the absence of nociceptors. Furthermore, anti-GR1-mediated depletion of myeloid cells during IAV infection significantly improved survival, underscoring a role of nociceptors in preventing pathogenic myeloid cell states that contribute to IAV-induced mortality. One Sentence Summary: TRPV1+ neurons facilitate host survival from influenza A virus infection by controlling myeloid cell responses and immunopathology.

immunology↗

bioRxiv2024

Characterization and analysis of neuronal signaling using microelectrode array combined with rapid and localized cooling device for cryo-neuromodulation

Cryoanesthesia--a purely physical anesthesia treatment that freezes tissue and attenuate nerve activity--can provide fast treatment through freezing and thawing of cryo-machine and is inexpensive compared to other anesthetics. However, cryoanesthesia has not been widely adopted because securing safe and effective conditions requires quantitative measurement and analysis of neuronal signaling during freezing and recovery, for which research tools are limited. A lack of rapid and localized cooling technologies for quantitative cellular level analysis, in particular, hinders research on not only the optimal cryo-modulation of neuronal activities but also its influence to neighboring cells via cellular networks. Here, we introduce a novel cryo-neuromodulation platform, a high-speed precision probe-type cooling device ([~]20{degrees}C/s at cooling) that provides localized cooling combined with a microelectrode array (MEA) system. We explored the temperature conditions for efficient silencing and recovery of neuronal activities without cell damage. We found that electrical activities of neurons were fully recovered within 1 minute of cooling duration with the maximum cooling speed, which was also confirmed with calcium imaging. The impact of silenced neurons on the neighboring neural network was explored using the localized cooling and we perceived that its influence can be transmitted if the neuronal network is well organized. Our new cryo-device provides rapid and reversible control of neural activities, which allows not just quantitative analysis of the network dynamics, but also new applications in clinical settings.

neuroscience↗

bioRxiv2024

When monkeys meet an ANYmal robot in the wild

Animal-robot interaction studies have been of increasing interest in research, but most of these studies have involved robots interacting with insects, birds, and frogs in laboratory settings. To date, only two studies used non-human primates and no behavioral study has tested the social integration of a robot in a group of wild primates. To fill this gap, we studied the interactions between the quadruped ANYmal robot and a group of 37 wild vervet monkeys in South Africa. The ANYmal robot is a remote-controlled sheep-sized robot with an open box of food on its back. We gradually introduced the robot to the monkeys following five different steps over 6 days for a total exposition time of about 10h. The monkeys habituated to the robot very quickly with six individuals eating the food in the robots box from the second day. A few individuals, mostly juveniles, emitted alarm calls towards the robot. In total, seven individuals, high rankers, spent time near the robot and 21 monkeys approached the robot from a greater distance. High rankers displayed significantly more vigilant and self-centered behaviors and they, with females and juveniles, ate more food in the robots box compared to low rankers, males, and adults conversely. This study offers exciting perspectives on the phenomena of social acceptance of machines in mammalian societies and the automation of field data collection.

animal behavior and cognition↗

bioRxiv2024

B cells targeting parasites capture spatially linked antigens to secure T cell help

Our understanding of T-cell-dependent humoral responses has been largely shaped by studies involving model antigens such as recombinant proteins and viruses 1,2. In these contexts, B cells internalize the entire antigen or pathogen, and present a range of antigens to helper CD4+ T cells to initiate the humoral response. However, this model does not account for large pathogens (such as parasites) that are too large to be taken up by individual B cells, and the mechanisms by which B cells acquire and present antigens from large complex pathogens to T cells remain poorly understood. Here we used Plasmodium, the causative parasite of malaria, as a model to investigate the requirements for T cell help for B cells targeting the Plasmodium surface circumsporozoite protein (CSP). Upon Plasmodium sporozoite (SPZ) immunization, CSP-specific B cells can form a synapse-like structure with SPZs and take up CSP and non-CSP surface antigens. As a result, CSP-specific B cells can receive help from CD4+ T cells specific to antigens that are located on the surface but not cytosol of the Plasmodium SPZ. Therefore, B cells can obtain help, not only from T cells with the same protein specificity, but also from T cells specific for spatially linked antigens. This flexibility in T cell help may enhance the initiation and maintenance of humoral immune responses to complex pathogens.

immunology↗

bioRxiv2024

A pan-variant miniprotein inhibitor protects against SARS-CoV-2 variants

The continued evolution of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has compromised neutralizing antibody responses elicited by prior infection or vaccination and abolished the utility of most monoclonal antibody therapeutics. We previously described a computationally-designed, homotrimeric miniprotein inhibitor, designated TRI2-2, that protects mice against pre-Omicron SARS-CoV-2 variants. Here, we show that TRI2-2 exhibits pan neutralization of variants that evolved during the 4.5 years since the emergence of SARS-CoV-2 and protects mice against BQ.1.1, XBB.1.5 and BA.2.86 challenge when administered post-exposure by an intranasal route. The resistance of TRI2-2 to viral escape and its direct delivery to the upper airways rationalize a path toward clinical advancement.

biochemistry↗

bioRxiv2024

Centrocortin potentiates co-translational localization of its mRNA to the centrosome via dynein

Centrosomes rely upon proteins within the pericentriolar material to nucleate and organize microtubules. Several mRNAs also reside at centrosomes, although less is known about how and why they accumulate there. We previously showed that local Centrocortin (Cen) mRNA supports centrosome separation, microtubule organization, and viability in Drosophila embryos. Here, using Cen mRNA as a model, we examine mechanisms of centrosomal mRNA localization. We find that while the Cen N-terminus is sufficient for protein enrichment at centrosomes, multiple domains cooperate to concentrate Cen mRNA at this location. We further identify an N-terminal motif within Cen that is conserved among dynein cargo adaptor proteins and test its contribution to RNA localization. Our results support a model whereby Cen protein enables the accumulation of its own mRNA to centrosomes through a mechanism requiring active translation, microtubules, and the dynein motor complex. Taken together, our data uncover the basis of translation-dependent localization of a centrosomal RNA required for mitotic integrity. SummaryEnrichment of Centrocortin (Cen) mRNA at centrosomes is required for mitotic fidelity. This study describes a mechanism underlying co-translational Cen mRNA targeting involving microtubules, the dynein motor, and a highly conserved dynein binding motif within the Cen coding sequence.

cell biology↗

bioRxiv2024

Hybrid Protocells based on Coacervate-Templated Fatty Acid Vesicles combine Improved Membrane Stability with Functional Interior Protocytoplasm

Prebiotically-plausible compartmentalization mechanisms include membrane vesicles formed by amphiphile self-assembly and coacervate droplets formed by liquid-liquid phase separation. Both types of structures form spontaneously and can be related to cellular compartmentalization motifs in todays living cells. As prebiotic compartments, they have complementary capabilities, with coacervates offering excellent solute accumulation and membranes providing superior boundaries. Herein, we describe protocell models constructed by spontaneous encapsulation of coacervate droplets by mixed fatty acid/phospholipid and by purely fatty acid membranes. Coacervate-supported membranes formed over a range of coacervate and lipid compositions, with membrane properties impacted by charge-charge interactions between coacervates and membranes. Vesicles formed by coacervate-templated membrane assembly exhibited profoundly different permeability than traditional fatty acid or blended fatty acid/phospholipid membranes without coacervate interiors, particularly in the presence of Mg2+ ions. While fatty acid and blended membrane vesicles were disrupted by addition of 25 mM MgCl2, the corresponding coacervate-supported membranes remained intact and impermeable to externally-added solutes even in the presence of MgCl2. With the more robust membrane, fluorescein diacetate (FDA) hydrolysis, which is commonly used for cell viability assays, could be performed inside the protocell model due to the simple diffusion of FDA and then following with the coacervate-mediated abiotic hydrolysis to fluorescein.

biophysics↗

bioRxiv2024

Th1 polarization in Bordetella pertussis vaccine responses is maintained through a positive feedback loop

Outbreaks of Bordetella pertussis (BP), the causative agent of whooping cough, continue despite broad vaccination coverage and have been increasing since vaccination switched from whole-BP (wP) to acellular BP (aP) vaccines. wP vaccination has been associated with more durable protective immunity and an induced Th1 polarized memory T cell response. Here, a multi-omics approach was applied to profile the immune response of 30 wP and 31 aP-primed individuals and identify correlates of T cell polarization before and after Tdap booster vaccination. We found that transcriptional changes indicating an interferon response on day 1 post-booster along with elevated plasma concentrations of IFN-{gamma} and interferon-induced chemokines that peaked at day 1-3 post-booster correlated best with the Th1 polarization of the vaccine-induced memory T cell response on day 28. Our studies suggest that wP-primed individuals maintain their Th1 polarization through this early memory interferon response. This suggests that stimulating the interferon pathway during vaccination could be an effective strategy to elicit a predominant Th1 response in aP-primed individuals that protects better against infection.

immunology↗

bioRxiv2024

Activated Interferon Signaling Suppresses Age-Dependent Liver Cancer

Age is a major risk factor for liver cancer, as is the case for most adult human cancers. However, the underlying mechanisms are not well defined. A better understanding of the role of aging in liver and other cancers can facilitate approaches for risk assessment, early detection and prevention. We hypothesize that age-driven changes render aged liver more sensitive to oncogenic stress and hence tumorigenesis. To investigate how the liver changes with age, we documented the immune profile, transcriptome and epigenome of healthy livers from both young and aged mice, revealing pronounced alterations with aging. Notably, in aged hepatocytes, we identified heightened interferon (IFN) signaling, as well as simultaneous tumor suppressor and oncogene signaling at both bulk and single cell level, suggestive of an aged liver that is poised for neoplasia. To challenge this seemingly poised state, we employed adeno-associated virus (AAV)-mediated expression of a c-Myc oncogene in young and aged mouse liver hepatocytes in vivo. Analysis of aged hepatocytes expressing c-Myc revealed further elevated expression of IFN Stimulated Genes (ISGs). This ISG upregulation was evident in multiple models of oncogenic stress and transformation in older mice and also observed in aged humans with Metabolic dysfunction-Associated Steatohepatitis (MASH). We determined that Stat1 is both necessary and sufficient for the age specific elevated ISG expression in old wild type mice. Remarkably, inhibiting Jak/Stat signaling alongside ectopic c-Myc expression led to high-grade hepatocyte dysplasia and tumor formation, selectively in aged mice. Together, these results suggest that an aged liver is in a state of "precarious balance", due to concurrent activation of oncogenic and tumor suppressor pathways, but protected against neoplastic progression by IFN-signaling. Age-dependent activation of IFN signaling has been observed in many tissues and recent studies have demonstrated its detrimental consequences on aging, raising the question as to why IFN-signaling is activated during aging. We propose that aged tissues are intrinsically at higher risk of cancer and age-dependent activation of IFN-signaling is an adaptive process to protect from tumorigenesis, but one that also has maladaptive consequences.

cancer biology↗

bioRxiv2024

Social Isolation During Adolescence Differentially Affects Spatial Learning in Adult Male and Female Mice

Social isolation is a risk factor for cognitive impairment. Adolescents may be particularly vulnerable to these effects, because they are in a critical period of development marked by significant physical, hormonal, and social changes. However, it is unclear if the effects of social isolation on learning and memory are similar in both sexes or if they persist into adulthood after a period of recovery. We socially isolated male and female 129Sv/Ev mice throughout adolescence (post-natal days 29-56), provided a 2-week re-socialization recovery period, and then tested spatial learning and cognitive flexibility in the active place avoidance task. After behavioral testing, mice were injected with 5-bromo-2-deoxyuridine (BrdU) so that lasting effects of social isolation on cell proliferation in the dentate gyrus could be examined. We found that in males, isolation led to a modest impairment in the rate of initial spatial learning, whereas in females, initial learning was unaffected. However, when the location of the shock zone was switched during the conflict variant of the task, cognitive flexibility was impaired in females only. Similarly, social isolation reduced cell proliferation in the ventral dentate gyrus only in females. Together, these findings indicate that social isolation during adolescence differentially impairs spatial processing in males and females, with effects that persist into adulthood.

neuroscience↗

bioRxiv2024

Senescent fibroblasts in the tumor stroma rewire lung cancer metabolism and plasticity

Senescence has been demonstrated to either inhibit or promote tumorigenesis. Resolving this paradox requires spatial mapping and functional characterization of senescent cells in the native tumor niche. Here, we identified senescent p16Ink4a+ cancer-associated fibroblasts with a secretory phenotype that promotes fatty acid uptake and utilization by aggressive lung adenocarcinoma driven by Kras and p53 mutations. Furthermore, rewiring of lung cancer metabolism by p16Ink4a+ cancer- associated fibroblasts also altered tumor cell identity to a highly plastic/dedifferentiated state associated with progression in murine and human LUAD. Our ex vivo senolytic screening platform identified XL888, a HSP90 inhibitor, that cleared p16Ink4a+ cancer- associated fibroblasts in vivo. XL888 administration after establishment of advanced lung adenocarcinoma significantly reduced tumor burden concurrent with the loss of plastic tumor cells. Our study identified a druggable component of the tumor stroma that fulfills the metabolic requirement of tumor cells to acquire a more aggressive phenotype.

cancer biology↗

bioRxiv2024

CardioMEA: Comprehensive Data Analysis Platform for Studying Cardiac Diseases and Drug Responses

In recent years, high-density microelectrode arrays (HD-MEAs) have emerged as a valuable tool in preclinical research for characterizing the electrophysiology of human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs). HD-MEAs enable the capturing of both extracellular and intracellular signals on a large scale, while minimizing potential damage to the cell. However, a gap exists between technological advancements of HD-MEAs and the availability of effective data-analysis platforms. To address this need, we introduce CardioMEA, a comprehensive data-analysis platform designed specifically for HD-MEA data that have been obtained from iPSC-CMs. CardioMEA features scalable data processing pipelines and an interactive web-based dashboard for advanced visualization and analysis. In addition to its core functionalities, CardioMEA incorporates modules designed to discern crucial electrophysiological features between diseased and healthy iPSC-CMs. Notably, CardioMEA has the unique capability to analyze both extracellular and intracellular signals, thereby facilitating customized analyses for specific research tasks. We demonstrate the practical application of CardioMEA by analyzing electrophysiological signals from iPSC-CM cultures exposed to seven antiarrhythmic drugs. CardioMEA holds great potential as an intuitive, user-friendly platform for studying cardiac diseases and assessing drug effects.

bioengineering↗