Biology subjects

Shi, Y.

Publications and source records attributed to Shi, Y..

At least 217 records · Page 12Linked to original sources

bioRxiv2024

5-HT Neurons Integrate GABA and Dopamine Inputs to Regulate Meal Initiation

Obesity is a growing global health epidemic with limited effective therapeutics. Serotonin (5-HT) is one major neurotransmitter which remains an excellent target for new weight-loss therapies, but there remains a gap in knowledge on the mechanisms involved in 5-HT produced in the dorsal Raphe nucleus (DRN) and its involvement in meal initiation. Using a closed-loop optogenetic feeding paradigm, we showed that the 5-HTDRN{lozenge}arcuate nucleus (ARH) circuit plays an important role in regulating meal initiation. Incorporating electrophysiology and ChannelRhodopsin-2-Assisted Circuit Mapping, we demonstrated that 5-HTDRN neurons receive inhibitory input partially from GABAergic neurons in the DRN, and the 5-HT response to GABAergic inputs can be enhanced by hunger. Additionally, deletion of the GABAA receptor subunit in 5-HT neurons inhibits meal initiation with no effect on the satiation process. Finally, we identified the instrumental role of dopaminergic inputs via dopamine receptor D2 in 5-HTDRN neurons in enhancing the response to GABA-induced feeding. Thus, our results indicate that 5-HTDRN neurons are inhibited by synergistic inhibitory actions of GABA and dopamine, which allows for the initiation of a meal.

neuroscience↗

bioRxiv2024

A posterior approach to correct for focal plane offsets in lattice light sheet structured illumination microscopy

SignificanceLattice light sheet structured illumination microscopy (latticeSIM) has proven highly effective in producing 3D images with super resolution rapidly and with minimal photobleaching. However, due to the use of two separate objectives, sample-induced aberrations can result in an offset between the planes of excitation and detection, causing artifacts in the reconstructed images. AimWe introduce a posterior approach to detect and correct for the axial offset between the excitation and detection focal planes in latticeSIM and provide a method to minimize artifacts in the reconstructed images. ApproachWe utilized the residual phase information within the overlap regions of the laterally shifted structured illumination microscopy (SIM) information components in frequency space to retrieve the axial offset between the excitation and the detection focal planes in latticeSIM. ResultsWe validated our technique through simulations and experiments, encompassing a range of samples from fluorescent beads to subcellular structures of adherent cells. We also show utilizing transfer functions with the same axial offset as that which was present during the data acquisition results in reconstructed images with minimal artifacts and salvages otherwise unusable data. ConclusionWe envision that our method will be a valuable addition to restore image quality in latticeSIM datasets even for those acquired under non-ideal experimental conditions.

biophysics↗

bioRxiv2024

Comparative genomics of macaques and integrated insights into genetic variation and population history

The crab-eating macaques (Macaca fascicularis) and rhesus macaques (M. mulatta) are widely studied nonhuman primates in biomedical and evolutionary research. Despite their significance, the current understanding of the complex genomic structure in macaques and the differences between species requires substantial improvement. Here, we present a complete genome assembly of a crab-eating macaque and 20 haplotype-resolved macaque assemblies to investigate the complex regions and major genomic differences between species. Segmental duplication in macaques is [~]42% lower, while centromeres are [~]3.7 times longer than those in humans. The characterization of [~]2 Mbp fixed genetic variants and [~]240 Mbp complex loci highlights potential associations with metabolic differences between the two macaque species (e.g., CYP2C76 and EHBP1L1). Additionally, hundreds of alternative splicing differences show post-transcriptional regulation divergence between these two species (e.g., PNPO). We also characterize 91 large-scale genomic differences between macaques and humans at a single-base-pair resolution and highlight their impact on gene regulation in primate evolution (e.g., FOLH1 and PIEZO2). Finally, population genetics recapitulates macaque speciation and selective sweeps, highlighting potential genetic basis of reproduction and tail phenotype differences (e.g., STAB1, SEMA3F, and HOXD13). In summary, the integrated analysis of genetic variation and population genetics in macaques greatly enhances our comprehension of lineage-specific phenotypes, adaptation, and primate evolution, thereby improving their biomedical applications in human diseases.

genomics↗

bioRxiv2024

PhenoMultiOmics: an enzymatic reaction inferred multi-omics network visualization web server

Enzymatic reactions play a pivotal role in regulating cellular processes with a high degree of specificity to biological functions. When enzymatic reactions are disrupted by gene, protein, or metabolite dysfunctions in diseases, it becomes crucial to visualize the resulting perturbed enzymatic reaction-induced multi-omics network. Multi-omics network visualization aids in gaining a comprehensive understanding of the functionality and regulatory mechanisms within biological systems. In this study, we designed PhenoMultiOmics, an enzymatic reaction-based multi-omics web server designed to explore the scope of the multi-omics network across various cancer types. We first curated the PhenoMultiOmics Database (PMODB), which enables the retrieval of cancer-gene-protein-metabolite relationships based on the enzymatic reactions. We then developed the MultiOmics network visualization module to depict the interplay between genes, proteins, and metabolites in response to specific cancer-related enzymatic reactions. The biomarker discovery module facilitates functional analysis through differential omic feature expression and pathway enrichment analysis. PhenoMultiOmics has been applied to analyze transcriptomics data of gastric cancer and metabolomics data of lung cancer, providing insights into interrupted enzymatic reactions and the associated multi-omics network. PhenoMultiOmics is freely accessed at https://phenomultiomics.shinyapps.io/cancer/ with a user-friendly and interactive web interface.

bioinformatics↗

bioRxiv2024

Decoding Missense Variants by Incorporating Phase Separation via Machine Learning

Computational models have made significant progress in predicting the effect of protein variants. However, deciphering numerous variants of unknown significance (VUS) located within intrinsically disordered regions (IDRs) remains challenging. To address this issue, we introduced phase separation (PS), which is tightly linked to IDRs, into the investigation of missense variants. Phase separation is vital for multiple physiological processes. By leveraging missense variants that alter phase separation propensity, we developed a machine learning approach named PSMutPred to predict the impact of missense mutations on phase separation. PSMutPred demonstrated robust performance in predicting missense variants that affect natural phase separation. In vitro experimental findings further underscore its validity. By applying PSMutPred on over 522,000 ClinVar missense variants, it significantly contributes to decoding the pathogenesis of disease variants, especially those in IDRs. Our work provides unique insights into the understanding of a vast number of VUSs in IDRs, thereby expediting clinical interpretation and diagnosis of disease variants.

bioinformatics↗

bioRxiv2024

The Molecular Architecture of the Nuclear Basket

The nuclear pore complex (NPC) is the sole mediator of nucle-ocytoplasmic transport. Despite great advances in understanding its conserved core architecture, the peripheral regions can exhibit considerable variation within and between species. One such structure is the cage-like nuclear basket. Despite its crucial roles in mRNA surveillance and chromatin organization, an architectural understanding has remained elusive. Using in-cell cryo-electron tomography and subtomogram analysis, we explored the NPCs structural variations and the nuclear basket across fungi (yeast; S. cerevisiae), mammals (mouse; M. musculus), and protozoa (T. gondii). Using integrative structural modeling, we computed a model of the basket in yeast and mammals that revealed how a hub of Nups in the nuclear ring binds to basket-forming Mlp/Tpr proteins: the coiled-coil domains of Mlp/Tpr form the struts of the basket, while their unstructured termini constitute the basket distal densities, which potentially serve as a docking site for mRNA preprocessing before nucleocytoplasmic transport

biophysics↗

bioRxiv2024

Map based cloning of CT2 and the pilot functional exploration in abiotic stress

Heterotrimeric G-proteins are multifunctional modulators that participate in a wide range of growth and developmental processes in eukaryotic species, from yeast to plants and animals. Component detection and the study of G protein signaling in most plants, including maize, are in the initial stages. In this study, we characterized a maize mutant, ct2, that showed a compact architecture and reproductive organ-related phenotypic variation. The target gene CT2 was cloned using bulked segregant analysis and map-based cloning. Gene structure prediction and phylogenetic analysis indicated that CT2 is a canonical G protein belonging to the monocotyledonous group. Promoter analysis of CT2 and RNA sequencing revealed cis-acting regulatory elements and differentially expressed genes involved in JA signaling and stress tolerance. The transcription of CT2 was repressed by NaCl and PEG treatments, and ct2 mutation in the ct2 line compromised stress tolerance in maize. On the basis of our results, we proposed a schema diagram of CT2-regulated biological process and their feedback on CT2 transcription. This research provides clues for further studies of CT2 function in hormone signaling and stress tolerance, which is beneficial for maize breeding through the screening and application of beneficial alleles.

molecular biology↗

bioRxiv2024

A high resolution model of the grapevine leaf morphospace predicts synthetic leaves

O_LIGrapevine leaves are a model morphometric system. Sampling over ten thousand leaves using dozens of landmarks, the genetic, developmental, and environmental basis of leaf shape has been studied and a morphospace for the genus Vitis predicted. Yet, these representations of leaf shape fail to capture the exquisite features of leaves at high resolution. C_LIO_LIWe measure the shapes of 139 grapevine leaves using 1672 pseudo-landmarks derived from 90 homologous landmarks with Procrustean approaches. From hand traces of the vasculature and blade, we have derived a method to automatically detect landmarks and place pseudo-landmarks that results in a high-resolution representation of grapevine leaf shape. Using polynomial models, we create continuous representations of leaf development in 10 Vitis spp. C_LIO_LIWe visualize a high-resolution morphospace in which genetic and developmental sources of leaf shape variance are orthogonal to each other. Using classifiers, V. vinifera, Vitis spp., rootstock and dissected leaf varieties as well as developmental stages are accurately predicted. Theoretical eigenleaf representations sampled from across the morphospace that we call synthetic leaves can be classified using models. C_LIO_LIBy predicting a high-resolution morphospace and delimiting the boundaries of leaf shapes that can plausibly be produced within the genus Vitis, we can sample synthetic leaves with realistic qualities. From an ampelographic perspective, larger numbers of leaves sampled at lower resolution can be projected onto this high-resolution space; or, synthetic leaves can be used to increase the robustness and accuracy of machine learning classifiers. C_LI Societal Impact StatementGrapevine leaves are emblematic of the strong visual associations people make with plants. At a glance, leaf shape is immediately recognizable, and it is because of this reason it is used to distinguish grape varieties. In an era of computationally-enabled, machine learning-derived representations of reality, we can revisit how we view and use the shapes and forms that plants display to understand our relationship with them. Using computational approaches combined with time-honored methods, we can predict theoretical leaves that are possible to understand the genetics, development, and environmental responses of plants in new ways.

plant biology↗

bioRxiv2024

The ZmHSF20-ZmHSF4-ZmCesA2 module regulates heat stress tolerance in maize

Temperature shapes the geographical distribution and behavior of plants. Understanding the regulatory mechanisms behind plant heat response is important for developing climate-resilient crops, including maize (Zea mays). To identify transcription factors that may contribute to heat response, we generated a dataset of short- and long-term transcriptome changes following a heat treatment time course in the maize inbred line B73. Co-expression network analysis highlighted several transcription factors, including the class B2a heat shock factor ZmHSF20. ZmHsf20 mutant seedlings exhibited enhanced tolerance of heat stress. Furthermore, DNA affinity purification sequencing and CUT&Tag assays demonstrated that ZmHSF20 binds the promoters of Cellulose synthase A2 (ZmCesA2) and three class A HSF genes, including ZmHSF4, repressing their transcription. We showed that ZmCesA2 and ZmHSF4 positively regulate heat response, with ZmHSF4 directly activating ZmCesA2 transcription. In agreement with the transcriptome analysis, ZmHSF20 negatively modulated cellulose accumulation and repressed the expression of cell wall-related genes. Importantly, the ZmHsf20 ZmHsf4 double mutant exhibited decreased thermotolerance, placing ZmHSF4 downstream of ZmHSF20. Based on our results, we propose an expanded model of the heat stress response in maize, whereby ZmHSF20 lowers heat tolerance of seedlings by repressing ZmHSF4 and ZmCesA2, thus balancing growth and defense at the seedling stage. One-sentence summaryZmHSF20, as a negative factor, acts upstream of ZmHSF4 and ZmCesA2, which are involved in positively regulating the cell wall development under heat stress, thereby improving maize heat tolerance.

plant biology↗

bioRxiv2024

RNA-MobiSeq: Deep mutational scanning and mobility-based selection for RNA structure inference

Tertiary structures of RNAs are increasingly found to be essential for their functions and yet are notoriously challenging to be determined by experimental techniques or predicted by energy-based and AlphaFold2-like deep-learning-based computational methods. Here, we coupled deep mutational scanning with mobility-based selection to separate structurally stable from nonstable RNA variants. The subsequent high-throughput sequencing allows the detection of the covariational signals of key secondary and tertiary base pairs with significantly improved, RNA-specific, unsupervised analysis of covariation-induced deviation of activity (CODA2) and amplification of these signals by Monte-Carlo simulated annealing. The resulting base-pairing structures can serve as the restraints for final energy-based structure predictions. This MobiSeq technique was tested on four structurally different RNAs. The results show that the detection of tertiary base pairs by MobiSeq allows reasonably accurate prediction of RNA tertiary structures. The proposed method should be useful for structural inference and improved mechanistic understanding of all RNAs with detectable changes in mobility due to deleterious mutations.

molecular biology↗

bioRxiv2024

UXS1 regulates UDP-GlcA levels to support growth of UGDH-high cancer cells

Identifying genes that are crucial for cancer cell survival but dispensable in normal cells holds immense therapeutic potential. The DepMap Consortiums extensive datasets have paved the way for uncovering such selectively essential genes in cancer. However, it remained challenging to efficiently prioritize understudied, selectively essential genes for validation and characterization. To this end, our lab has previously ranked and prioritized potentially understudied, selectively essential genes based on their PubMed publication numbers. This approach led to successful identification and detailed characterization of two top understudied genes. Building on this methodology, our current research identified UXS1, an enzyme responsible for catalyzing the conversion from UDP-glucuronic acid (UDP-GlcA) to UDP-xylose, as a selectively essential gene in cancer cells expressing elevated levels of UGDH, an enzyme responsible for producing UDP-GlcA. Through an integrated approach combining genetic and biochemical assays, we discovered that UXS1 plays a critical role in these UGDH-overexpressing cancer cells by preventing the harmful buildup of UDP-GlcA, which otherwise would lead to cellular toxicity. Our findings not only validate our strategy for prioritizing underexplored but potentially pivotal selectively essential genes but also highlight UXS1 as a potential vulnerability and therapeutic target in cancers characterized by high UGDH expression.

cell biology↗

bioRxiv2024

AhR-Ovol1-Id1 regulatory axis in keratinocytes promotes skin homeostasis against atopic dermatitis

Skin is our outer permeability and immune defense barrier against myriad external assaults. Aryl hydrocarbon receptor (AhR) senses environmental factors and regulates barrier robustness and immune homeostasis. AhR agonist is in clinical trial for atopic dermatitis (AD) treatment, but the underlying mechanism of action remains ill-defined. Here we report OVOL1/Ovol1 as a conserved and direct transcriptional target of AhR in epidermal keratinocytes. We show that OVOL1/Ovol1 impacts AhR regulation of keratinocyte gene expression, and Ovol1 deletion in keratinocytes hampers AhRs barrier promotion function and worsens AD-like inflammation. Mechanistically, we identify Ovol1s direct downstream targets genome-wide, and provide in vivo evidence for Id1s critical role in barrier maintenance and disease suppression. Furthermore, our findings reveal an IL-1/dermal {gamma}{delta}T cell axis exacerbating both type 2 and type 3 immune responses downstream of barrier perturbation in Ovol1-deficient AD skin. Finally, we present data suggesting the clinical relevance of OVOL1 and ID1 function in human AD. Our study highlights a keratinocyte-intrinsic AhR-Ovol1-Id1 regulatory axis that promotes both epidermal and immune homeostasis against AD-like inflammation, implicating new therapeutic targets for AD.

molecular biology↗

bioRxiv2023

Ultrasound programmable hydrogen-bonded organic frameworks for sono-chemogenetics

The precise control of mechanochemical activation within deep tissues via non-invasive ultrasound holds profound implications for advancing our understanding of fundamental biomedical sciences and revolutionizing disease treatments. However, a theory-guided mechanoresponsive materials system with well-defined ultrasound activation has yet to be explored. Here we present the concept of using porous hydrogen-bonded organic frameworks (HOFs) as toolkits for focused ultrasound programmably triggered drug activation to control specific cellular events in the deep brain, through on-demand scission of the supramolecular interactions. A theoretical model is developed to visualize the mechanochemical scission and ultrasound mechanics, providing valuable guidelines for the rational design of mechanoresponsive materials at the molecular level to achieve programmable and spatiotemporal activation control. To demonstrate the practicality of this approach, we encapsulate designer drug clozapine N-oxide (CNO) into the optimal HOF nanoparticles for FUS gated release to activate engineered G-protein-coupled receptors in the mice and rat ventral tegmental area (VTA), and hence achieved targeted neural circuits modulation even at depth 9 mm with a latency of seconds. This work demonstrates the capability of ultrasound to precisely control molecular interaction and develops ultrasound programmable HOFs to minimally invasive and spatiotemporally control cellular events, thereby facilitating the establishment of precise molecular therapeutic possibilities. We anticipate that this research could serve as a source of inspiration for precise and non-invasive molecular manipulation techniques, potentially applicable in programming molecular robots to achieve sophisticated control over cellular events in deep tissues.

bioengineering↗

bioRxiv2023

Rapid, concerted switching of the neural code in inferotemporal cortex

A fundamental paradigm in neuroscience is the concept of neural coding through tuning functions1. According to this idea, neurons encode stimuli through fixed mappings of stimulus features to firing rates. Here, we report that the tuning of visual neurons can rapidly and coherently change across a population to attend to a whole and its parts. We set out to investigate a longstanding debate concerning whether inferotemporal (IT) cortex uses a specialized code for representing specific types of objects or whether it uses a general code that applies to any object. We found that face cells in macaque IT cortex initially adopted a general code optimized for face detection. But following a rapid, concerted population event lasting < 20 ms, the neural code transformed into a face-specific one with two striking properties: (i) response gradients to principal detection-related dimensions reversed direction, and (ii) new tuning developed to multiple higher feature space dimensions supporting fine face discrimination. These dynamics were face specific and did not occur in response to objects. Overall, these results show that, for faces, face cells shift from detection to discrimination by switching from an object-general code to a face-specific code. More broadly, our results suggest a novel mechanism for neural representation: concerted, stimulus-dependent switching of the neural code used by a cortical area.

neuroscience↗

bioRxiv2023

ERK3 Is Involved in Regulating Cardiac Fibroblast Function

ERK3/MAPK6, an atypical MAPK, activates MAP kinase-activated protein kinase (MK)-5 in selected cell types. MK5 haplodeficient mice show reduced hypertrophy and attenuated increase in Col1a1 mRNA in response to increased cardiac afterload. In addition, MK5 deficiency alters cardiac fibroblast function. This study was to determine the effect of reduced ERK3 on cardiac hypertrophy following transverse aortic constriction (TAC) and fibroblast biology. Three wk post-surgery, ERK3, but not ERK4 or p38, was co-immunoprecipitated with MK5 from both sham and TAC heart lysates. The increase in left ventricular mass and myocyte diameter was lower in TAC-ERK3+/- than TAC-ERK3+/+ hearts, whereas ERK3 haploinsufficiency did not alter systolic or diastolic function. Furthermore, the TAC-induced increase in Col1a1 mRNA abundance was diminished in ERK3+/- hearts. ERK3 immunoreactivity was detected in atrial and ventricular fibroblasts but not myocytes. In both quiescent fibroblasts and activated myofibroblasts isolated from adult mouse heart, siRNA-mediated knockdown of ERK3 reduced the TGF-{beta}-induced increase in Col1a1 mRNA. In addition, intracellular type 1 collagen immunoreactivity was reduced following ERK3 depletion in quiescent fibroblasts but not myofibroblasts. Finally, knocking down ERK3 impaired motility in both atrial and ventricular myofibroblasts. These results suggest that ERK3 plays an important role in multiple aspects of cardiac fibroblast biology.

cell biology↗

bioRxiv2023

SARS-CoV-2 NSP13 suppresses the Hippo pathway downstream effector YAP

The Hippo pathway controls organ development, homeostasis, and regeneration primarily by modulating YAP/TEAD-mediated gene expression. Although emerging studies report Hippo-YAP dysfunction after viral infection, it is largely unknown in the context of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Here, we analyzed RNA sequencing data from induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) and SARS-CoV-2-infected human lung samples, and observed a decrease in YAP target gene expression. In screening SARS-CoV-2 nonstructural proteins, we found that nonstructural protein 13 (NSP13), a conserved coronavirus helicase, inhibits YAP transcriptional activity independent of the upstream Hippo kinases LATS1/2. Consistently, introducing NSP13 into cardiomyocytes suppresses an active form of YAP (YAP5SA) in vivo. Subsequent investigations on NSP13 mutants revealed that NSP13 helicase activity, including DNA binding and unwinding, is crucial for suppressing YAP transactivation. Mechanistically, TEAD4 serves as a platform to recruit NSP13 and YAP. NSP13 likely inactivates the YAP/TEAD4 transcription complex by remodeling chromatin to recruit proteins, such as transcription termination factor 2 (TTF2), to bind the YAP/TEAD/NSP13 complex. These findings reveal a novel YAP/TEAD regulatory mechanism and uncover molecular insights into Hippo-YAP regulation after SARS-CoV-2 infection.

cell biology↗

bioRxiv2023

Fast and accurate disulfide bridge detection

Recombinant expression of proteins, propelled by therapeutic antibodies, has evolved into a multi-billion-dollar industry. Essential here is quality control assessment of critical attributes such as sequence fidelity, proper folding, and post-translational modifications (PTMs). Errors can lead to diminished bioactivity and, in the context of therapeutic proteins, an elevated risk for immunogenicity. Over the years, many techniques were developed and applied to validate proteins in a standardized and high-throughput fashion. One parameter has, however, so far been challenging to assess. Disulfide bridges, covalent bonds linking two Cysteine residues, assist in the correct folding and stability of proteins and thus have a major influence on their efficacy. Mass spectrometry promises to be an optimal technique to uncover them in a fast and accurate fashion. In this work, we present a unique combination of sample preparation, data acquisition and analysis facilitating the rapid and accurate assessment of disulfide bridges in purified proteins. Through microwave-assisted acid hydrolysis (MAAH), the proteins are digested rapidly and artifact-free into peptides, with a substantial degree of overlap over the sequence. The nonspecific nature of this procedure, however, introduces chemical background which is efficiently removed by integrating ion mobility preceding the mass spectrometric measurement. The nonspecific nature of the digestion step additionally necessitates new developments in data analysis, for which we extended the XlinkX node in Proteome Discoverer (XlinkX/PD) to efficiently process the data and ensure correctness through effective false discovery rate correction. The entire workflow can be completed within one hour, allowing for high-throughput, high-accuracy disulfide mapping.

biochemistry↗

bioRxiv2023

Integrative multiomics analysis of neointima proliferation in human saphenous vein: implications for bypass graft disease

IntroductionHuman saphenous veins (SV) are widely used as grafts in coronary artery bypass (CABG) surgery but often fail due to neointima proliferation (NP). NP involves complex interplay between vascular smooth muscle cells (VSMC) and fibroblasts. Little is known, however, regarding the transcriptomic and proteomic dynamics of NP. Here, we performed multi-omics analysis in an ex vivo tissue culture model of NP in human SV procured for CABG surgery. Methods and resultsHistological examination demonstrated significant elastin degradation and NP (indicated by increased neointima area and neointima/media ratio) in SV subjected to tissue culture. Analysis of data from 73 patients suggest that the process of SV adaptation and NP may differ according to sex and body mass index. RNA sequencing confirmed upregulation of pro-inflammatory and proliferation-related genes during NP and identified novel processes, including increased cellular stress and DNA damage responses, which may reflect tissue trauma associated with SV harvesting. Proteomic analysis identified upregulated extracellular matrix-related and coagulation/thrombosis proteins and downregulated metabolic proteins. Spatial transcriptomics detected transdifferentiating VSMC in the intima on the day of harvesting and highlighted dynamic alterations in fibroblast and VSMC phenotype and behavior during NP. Specifically, we identified new cell subpopulations contributing to NP, including SPP1+, LGALS3+ VSMC and MMP2+, MMP14+ fibroblasts. ConclusionDynamic alterations of gene and protein expression occur during NP in human SV. Identification of the human-specific molecular and cellular mechanisms may provide novel insight into SV bypass graft disease.

genomics↗