Biology subjects

Lin, S.

Publications and source records attributed to Lin, S..

At least 127 records · Page 7Linked to original sources

bioRxiv2023

Towards establishing a rapid constant temperature detection method for canine parvovirus based on endonuclease activities

Canine parvovirus can cause a high morbidity and mortality rate in puppies, posing a significant threat to both pet dogs and the breeding industry. Rapid, accurate and convenient detection methods are important for the early intervention and treatment of canine parvovirus. In this study, we propose a visual CPV detection system called nucleic acid mismatch enzyme digestion (NMED). This system combines reverse transcription-loop mediated isothermal amplification (RT-LAMP), endonuclease for gene mismatch detection and colloidal gold lateral chromatography. We demonstrated that NMED can induce the binding of the amplicon from the sample to the specific labeling probe, which in turn triggers digestion by the endonuclease. The sensitivity and visual visibility of RT-LAMP were increased by combining of endonuclease and colloidal gold lateral chromatography assisted by a simple temperature-controlled device. The sensitivity of the NMED assay was 1 copy/L, which was consistent with qPCR. The method was validated with 10 clinical samples that potentially had CPV infection and achieved 100% accuracy. As a rapid, accurate and convenient molecular diagnostic method, NMED has great potential for application in the field of pathogenic microorganism detection.

molecular biology↗

bioRxiv2023

Small molecule inhibitor binds to NLRP3 and prevents inflammasome activation

Despite recent advances in the mechanism of oxidized DNA activating NLRP3, the molecular mechanism and consequence of oxidized DNA associating with NLRP3 remains unknown. Cytosolic NLRP3 binds oxidized DNA which has been released from the mitochondria, which subsequently triggers inflammasome activation. Human glycosylase (hOGG1) repairs oxidized DNA damage which inhibits inflammasome activation. The fold of NLRP3 pyrin domain contains amino acids and a protein fold similar to hOGG1. Amino acids that enable hOGG1 to bind and cleave oxidized DNA are conserved in NLRP3. We found NLRP3 could bind and cleave oxidized guanine within mitochondrial DNA. The binding of oxidized DNA to NLRP3 was prevented by small molecule drugs which also inhibit hOGG1. These same drugs also inhibited inflammasome activation. Elucidating this mechanism will enable design of drug memetics that treat inflammasome pathologies, illustrated herein by NLRP3 pyrin domain inhibitors which suppressed interleukin-1{beta} (IL-1{beta}) production in macrophages. One-Sentence SummaryNLRP3 cleaves oxidized DNA and small molecule drug binding inhibits inflammasome activation.

immunology↗

bioRxiv2023

Human naive stem cell models reveal the role of FGF signalling in hypoblast specification in the human embryo

The hypoblast is an essential extra-embryonic tissue set aside within the inner cell mass early in mammalian embryo development, in the blastocyst. Research with human embryos is challenging. Thus, stem cell models that reproduce hypoblast differentiation provide valuable alternatives. We show here that human naive PSC to hypoblast differentiation proceeds via reversion to a transitional ICM-like state, from which hypoblast emerges in concordance with the trajectory in human blastocysts. We identified a window when fibroblast growth factor (FGF) signalling is critical for hypoblast specification. Revisiting FGF signalling in human embryos revealed that inhibition in the early blastocyst suppresses hypoblast formation. In vitro, the induction of hypoblast is synergistically enhanced by limiting trophectoderm and epiblast fates. This finding revises previous reports and establishes a conservation in lineage specification between mouse and human. Overall, this study demonstrates the utility of human naive PSC-based models in elucidating mechanistic features of early human embryogenesis.

developmental biology↗

bioRxiv2023

A high-performance brain-sentence communication designed for logosyllabic language

Many severe neurological diseases, such as stroke and amyotrophic lateral sclerosis, can impair or destroy the ability of verbal communication. Recent advances in brain-computer interfaces (BCIs) have shown promise in restoring communication by decoding neural signals related to speech or motor activities into text. Existing research on speech neuroprosthesis has predominantly focused on alphabetic languages, leaving a significant gap of logosyllabic languages such as Mandarin Chinese which are spoken by more than 15% of the world population. Logosyllabic languages pose unique challenges to brain-to-text decoding due to extended character sets (e.g., 50,000+ for Mandarin Chinese) and complex mapping between characters and pronunciation. To address these challenges, we established a speech BCI designed for Mandarin, decoding speech-related stereoelectroencephalography (sEEG) signals into coherent sentences. We leverage the unique acoustic features of Mandarin Chinese syllables, constructing prediction models for syllable components (initials, tones, and finals), and employ a language model to resolve pronunciation to character ambiguities according to the semantic context. This method leads to a high-performance decoder with a median character accuracy of 71.00% over the full character set, demonstrating huge potentials for clinical application. To our knowledge, we are the first to report brain-to-sentence decoding for logosyllabic languages over full character set with a large intracranial electroencephalography dataset.

neuroscience↗

bioRxiv2023

Using behavioral biomarkers to redefine epochs of spontaneous recovery following spinal cord injury

The brain-spinal cord axis generates movement by assembling motor primitives into coordinated sequences. Spinal cord injury (SCI) disrupts this neuroaxis, impairing not only locomotion, but the full repertoire of behavior. Traditional scales for quantifying recovery collapse this complexity into predefined locomotor-focused criteria that obscure heterogeneity in recovery. To quantify the full behavioral repertoire following SCI, we adapted motion sequencing (MoSeq) to identify sub-second behavioral "syllables" and capture their usage and sequential organization without predefined features. We identified biomechanically distinct variants within syllable classes that are shared across injury severities and found that recovery is jointly structured by injury severity and individual mouse identity. Changes in sequences, however, unfold along a conserved temporal trajectory. By compressing behavior into a single metric, we uncovered clusters of coevolving locomotor and non-locomotor behaviors. These results frame SCI recovery with repertoire-level changes, where adaptive strategies emerge from constrained access to motor primitives and their sequences.

neuroscience↗

bioRxiv2023

Reliability of high-quantity human brain organoids for modeling microcephaly, glioma invasion, and drug screening

Brain organoids offer unprecedented insights into brain development and disease modeling and hold promise for drug screening. Significant hindrances, however, are morphological and cellular heterogeneity, inter-organoid size differences, cellular stress, and poor reproducibility. Here, we describe a method that reproducibly generates thousands of organoids across multiple iPSC lines. These High Quantity brain organoids (Hi-Q brain organoids) exhibit reproducible cytoarchitecture, cell diversity, and functionality, are free from ectopically active cellular stress pathways, and allow cryopreservation and re-culturing. Using patient-derived Hi-Q brain organoids, we recapitulated distinct forms of microcephaly pathogenesis: primary microcephaly due to a mutation in centrosomal CDK5RAP2 and progeria-associated microcephaly in Cockayne syndrome. When modeling glioma invasion, hi-Q brain organoids displayed a similar invasion pattern for a given patient-derived glioma cell line. This enabled a medium-throughput screen to identify Selumetinib and Fulvestrant, which also perturbed glioma invasion in vivo. Thus, the Hi-Q approach can easily be adapted to reliably harness brain organoids utility for personalized disease modeling and drug discovery.

cell biology↗

bioRxiv2023

Spatially Resolved Gene Expression is Not Necessary for Identifying Spatial Domains

The development of Spatially Resolved Transcriptomics (SRT) technologies has revolutionized the study of tissue organization. We introduce a graph convolutional network with an attention and positive emphasis mechanism, named "BINARY," relying exclusively on binarized SRT data to delineate spatial domains. BINARY outperforms existing methods across various SRT data types while using significantly less input information. Our study suggests that precise gene expression quantification may not always be essential, inspiring further exploration of the broader applications of spatially resolved binarized gene expression data.

bioinformatics↗

bioRxiv2023

Co-transcriptional production of programmable RNA condensates and synthetic organelles

Condensation of RNA and proteins is central to cellular functions, and the ability to program it would be valuable in synthetic biology and synthetic cell science. Here we introduce a modular platform for engineering synthetic RNA condensates from tailor-made, branched RNA nanostructures that fold and assemble co-transcriptionally. Up to three orthogonal condensates can form simultaneously and selectively accumulate guest molecules. The RNA condensates can be expressed within synthetic cells to produce membrane-less organelles with controlled number, size, morphology and composition, and that display the ability to selectively capture proteins. The in situ expression of programmable RNA condensates could underpin spatial organisation of functionalities in both biological and synthetic cells.

synthetic biology↗

bioRxiv2023

Metabotropic signaling within somatostatin interneurons regulates thalamocortical inputs during development

During brain development, neural circuits undergo major activity-dependent restructuring. Circuit wiring mainly occurs through synaptic strengthening following the Hebbian "fire together, wire together" precept. However, select connections, essential for circuit development, are transient. They are effectively connected early in development, but strongly diminish during maturation. The mechanisms by which transient connectivity recedes are unknown. To investigate this process, we characterize transient thalamocortical inputs, which depress onto somatostatin inhibitory interneurons during development, by employing optogenetics, chemogenetics, transcriptomics and CRISPR-based strategies. We demonstrate that in contrast to typical activity-dependent mechanisms, transient thalamocortical connectivity onto somatostatin interneurons is non-canonical and involves metabotropic signaling. Specifically, metabotropic-mediated transcription, of guidance molecules in particular, supports the elimination of this connectivity. Remarkably, we found that this developmental process impacts the development of normal exploratory behaviors of adult mice.

neuroscience↗

bioRxiv2023

Rapid, biochemical tagging of cellular activity history in vivo

Intracellular calcium (Ca2+) is ubiquitous to cell signaling across all biology. While existing fluorescent sensors and reporters can detect activated cells with elevated Ca2+ levels, these approaches require implants to deliver light to deep tissue, precluding their noninvasive use in freely-behaving animals. Here we engineered an enzyme-catalyzed approach that rapidly and biochemically tags cells with elevated Ca2+ in vivo. Ca2+-activated Split-TurboID (CaST) labels activated cells within 10 minutes with an exogenously-delivered biotin molecule. The enzymatic signal increases with Ca2+ concentration and biotin labeling time, demonstrating that CaST is a time-gated integrator of total Ca2+ activity. Furthermore, the CaST read-out can be performed immediately after activity labeling, in contrast to transcriptional reporters that require hours to produce signal. These capabilities allowed us to apply CaST to tag prefrontal cortex neurons activated by psilocybin, and to correlate the CaST signal with psilocybin-induced head-twitch responses in untethered mice.

neuroscience↗

bioRxiv2023

Rational design of a SOCS1-edited tumor infiltrating lymphocyte therapy for solid tumors using CRISPR/Cas9 screens

Cell therapies such as Tumor Infiltrating Lymphocyte (TIL) therapy have shown promise in the treatment of patients with refractory solid tumors, with improvement in response rates and durability of responses nevertheless sought. To identify targets capable of enhancing the anti-tumor activity of T cell therapies, large-scale in vitro and in vivo CRISPR/Cas9 screens were performed, with the suppressor of cytokine signaling 1 (SOCS1) gene identified as a top T cell-enhancing target. In murine CD8 T cell therapy models, SOCS1 served as a critical checkpoint in restraining the accumulation of T central memory cells in lymphoid organs as well as intermediate (Texint) and effector (Texeff) exhausted T cell subsets derived from progenitor exhausted T cell (Texprog) cells in tumors. A comprehensive CRISPR tiling screen of the SOCS1 coding region identified sgRNAs targeting the SH2 domain of SOCS1 as the most potent, with a sgRNA with minimal off-target cut sites used to manufacture KSQ-001, an engineered TIL therapy with SOCS1 inactivated by CRISPR/Cas9. KSQ-001 possessed increased responsiveness to cytokine signals and enhanced in vivo anti-tumor function in mouse models. These data demonstrate the use of CRISPR/Cas9 screens in the rational design of T cell therapies.

immunology↗

bioRxiv2023

Structural Basis of Streptomyces Antibiotic Regulatory Proteins Activating Transcription

Streptomycetes are renowned antibiotic producers, with Streptomyces antibiotic regulatory proteins (SARPs) acting as activators for antibiotic biosynthesis. However, the precise mechanism underlying SARPs transcriptional activation remains elusive. Here, we used cryo-electron microscopy (cryo-EM) to unravel the interplay between SARP, DNA, and RNA polymerase (RNAP) during transcriptional activation. The SARP domain of Streptomyces coelicolor AfsR (SAS) forms a side-by-side dimer contacting the afs box centered at -29.5 relative to the transcription start site. The upstream protomer binds to the direct repeat encompassing the -35 element while the {sigma}HrdB region 4 (R4) is positioned on top of both protomers, causing the removal of R4 from the major groove of the -35 element. Both SAS protomers establish interactions with C-terminal domain of one RNAP subunits, while specific regions of the RNAP {beta} flap tip helix and {beta} zinc-binding domain also engage with SAS. Key interfacial residues accounting for transcriptional activation were confirmed by mutational studies and in vitro transcriptional assays. Overall, our results present a detailed molecular view of how SARPs serve to activate transcription.

molecular biology↗

bioRxiv2023

Distinct mechanisms by which antioxidant transcription factors Nrf1 and Nrf2 as drug targets contribute to the anticancer efficacy of Cisplatin on hepatoma cells

Cisplatin (cis-Dichlorodiamineplatinum[II], CDDP) is generally accepted as a platinum-based alkylating agent type of the DNA-damaging anticancer drug, which is widely administrated in clinical treatment of many solid tumors. The pharmacological effect of CDDP is mainly achieved by replacing the chloride ion (Cl-) in its structure with H2O to form active substances with the strong electrophilic properties and then react with any nucleophilic molecules, primarily leading to genomic DNA damage and subsequent cell death. In this process, those target genes driven by the consensus electrophilic and/or antioxidant response elements (EpREs/AREs) in their promoter regions are also activated or repressed by CDDP. Thereby, we here examined the expression profiling of such genes regulated by two principal antioxidant transcription factors Nrf1 and Nrf2 (both encoded by Nfe2l1 and Nfe2l2, respectively) in diverse cellular signaling responses to this intervention. The results demonstrated distinct cellular metabolisms, molecular pathways and signaling response mechanisms by which Nrf1 and Nrf2 as the drug targets differentially contribute to the anticancer efficacy of CDDP on hepatoma cells and xenograft tumor mice. Interestingly, the role of Nrf1, rather than Nrf2, is required for the anticancer effect of CDDP, to suppress malignant behavior of HepG2 cells by differentially monitoring multi-hierarchical signaling to gene regulatory networks. To our surprise, it was found there exists a closer relationship of Nrf1 than Nrf2 with DNA repair, but the hyperactive Nrf2 in Nrf1-/-cells manifests a strong correlation with its resistance to CDDP, albeit their mechanistic details remain elusive.

cell biology↗

bioRxiv2023

NaV1.6 inhibition drives the efficacy of voltage-gated sodium channel inhibitors to prevent electrically induced seizures in both wild type and Scn8aN1768D/+ gain-of-function mice

Inhibitors of voltage-gated sodium channels (NaVs) are important anti-epileptic drugs, but the contribution of specific channel isoforms is unknown since available inhibitors are nonselective. We created a series of compounds with diverse selectivity profiles enabling block of NaV1.6 alone or together with NaV1.2. Mice with a heterozygous gain-of-function mutation (N1768D/+) in Scn8a (encoding NaV1.6) responded with a tonic-clonic seizure to a mild 6 Hz stimulus that was innocuous to wild-type mice. Pharmacologic inhibition of NaV1.6 in Scn8aN1768D/+ mice prevented seizures. Inhibitors were also effective in a direct current maximal electroshock seizure assay in wild-type mice. NaV1.6 inhibition correlated with efficacy in both models, even without inhibition of other CNS NaV isoforms. Our data suggest NaV1.6 inhibition is a driver of efficacy for NaV inhibitor anti-seizure medicines. Selective NaV1.6 inhibitors may provide targeted therapies for human Scn8a developmental and epileptic encephalopathies and better tolerated treatments for idiopathic epilepsies. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=199 SRC="FIGDIR/small/551823v2_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@39c09borg.highwire.dtl.DTLVardef@19413b8org.highwire.dtl.DTLVardef@9aa226org.highwire.dtl.DTLVardef@b9207_HPS_FORMAT_FIGEXP M_FIG C_FIG

pharmacology and toxicology↗

bioRxiv2023

MultiPPIMI: a multimodal deep learning framework for predicting interactions between protein-protein interaction targets and modulators

Protein-protein interactions (PPIs) are essential for various biological processes and diseases. However, most existing computational methods for identifying PPI modulators require either target structure or reference modulators, which restricts their applicability to novel PPI targets. To address this challenge, we propose MultiPPIMI, a sequence-based deep learning framework that predicts the interaction between any given PPI target and modulator. MultiPPIMI integrates multimodal representations of PPI targets and modulators, and uses a bilinear attention network to capture inter-molecular interactions. Experimental results on our curated benchmark dataset show that MultiPPIMI achieves an average AUROC of 0.837 in three cold-start scenarios, and an AUROC of 0.994 in the random-split scenario. Furthermore, the case study show that MultiPPIMI can assist molecular simulations in screening inhibitors of Keap1/Nrf2 PPI interactions. We believe that the proposed method provides a promising way to screen PPI-targeted modulators.

bioinformatics↗

bioRxiv2023

Analysis of the Neuron-specific IIS/FOXO Transcriptome in Aged Animals Reveals Regulators of Neuronal and Cognitive Aging

Cognitive decline is a significant health concern in our aging society. Here, we used the model organism C. elegans to investigate the impact of the IIS/FOXO pathway on age-related cognitive decline. The daf-2 Insulin/IGF-1 receptor mutant exhibits a significant extension of learning and memory span with age compared to wild-type worms, an effect that is dependent on the DAF-16 transcription factor. To identify possible mechanisms by which aging daf-2 mutants maintain learning and memory with age while wild-type worms lose neuronal function, we carried out neuron-specific transcriptomic analysis in aged animals. We observed downregulation of neuronal genes and upregulation of transcriptional regulation genes in aging wild-type neurons. By contrast, IIS/FOXO pathway mutants exhibit distinct neuronal transcriptomic alterations in response to cognitive aging, including upregulation of stress response genes and downregulation of specific insulin signaling genes. We tested the roles of significantly transcriptionally-changed genes in regulating cognitive functions, identifying novel regulators of learning and memory. In addition to other mechanistic insights, comparison of the aged vs young daf-2 neuronal transcriptome revealed that a new set of potentially neuroprotective genes is upregulated; instead of simply mimicking a young state, daf-2 may enhance neuronal resilience to accumulation of harm and take a more active approach to combat aging. These findings suggest a potential mechanism for regulating cognitive function with age and offer insights into novel therapeutic targets for age-related cognitive decline.

neuroscience↗

bioRxiv2023

Multifaceted effects on even-skipped transcriptional dynamics upon Kruppel dosage changes

Although fluctuations in transcription factor (TF) dosage are often well tolerated, little is known about how TF dosage modulation changes the target gene expression dynamics and results in subsequent developmental phenotypes. Using MS2/MCP-mediated quantitative live imaging in early Drosophila embryos, we characterized transcriptional dynamics of the pair-rule gene even-skipped (eve) upon changing the gap gene Kruppel (Kr) level. Decreasing Kr dosage by half leads to a transient posterior expansion of the eve stripe 2 and an anterior shift of stripe 5. Surprisingly, the most significant changes are observed in eve stripes 3 and 4, whose enhancers do not contain Kr binding sites. For both stripes, the widths of stripes are narrowed, boundaries are located more anteriorly, and the mRNA production level is reduced in Kr heterozygous embryos. We show that Kr dosage indirectly affects stripes 3 and 4 dynamics by modulating other gap gene dynamics. We quantitatively correlate moderate body segment phenotypes of Kr heterozygotes with spatiotemporal changes in eve expression. Our results indicate that nonlinear relationships between TF dosage and phenotypes underlie direct TF-DNA and indirect TF-TF interactions. Summary statementThis study analyzed the changes in transcriptional dynamics of even-skipped upon halving Kruppel dosage, and revealed how the combination of direct TF-DNA and indirect TF-TF interactions shape proper patterning.

developmental biology↗

bioRxiv2023

Lysine vitcylation is a novel vitamin C-derived protein modification that enhances STAT1-mediated immune response

Vitamin C (vitC) is a vital nutrient for health and also used as a therapeutic agent in diseases such as cancer. However, the mechanisms underlying vitCs effects remain elusive. Here we report that vitC directly modifies lysine without enzymes to form vitcyl-lysine, termed "vitcylation", in a dose-, pH-, and sequence-dependent manner across diverse proteins in cells. We further discover that vitC vitcylates K298 site of STAT1, which impairs its interaction with the phosphatase PTPN2, preventing STAT1 Y701 dephosphorylation and leading to increased STAT1-mediated IFN pathway activation in tumor cells. As a result, these cells have increased MHC/HLA class-I expression and activate immune cells in co-cultures. Tumors collected from vitC-treated tumor-bearing mice have enhanced vitcylation, STAT1 phosphorylation and antigen presentation. The identification of vitcylation as a novel PTM and the characterization of its effect in tumor cells opens a new avenue for understanding vitC in cellular processes, disease mechanisms, and therapeutics.

biochemistry↗