Biology subjects

Ge, X.

Publications and source records attributed to Ge, X..

At least 37 records · Page 2Linked to original sources

bioRxiv2025

RNA triggers chronic stress during neuronal aging

Neurodegenerative diseases are linked with dysregulation of the integrated stress response (ISR), which coordinates cellular homeostasis during and after stress events. Cellular stress can arise from several sources, but there is significant disagreement about which stress might contribute to aging and neurodegeneration. Here, we leverage directed transdifferentiation of human fibroblasts into aged neurons to determine the source of ISR activation. We demonstrate that increased accumulation of cytoplasmic double-stranded RNA (dsRNA) activates the eIF2 kinase PKR, which in turn triggers the ISR in aged neurons and leads to sequestration of dsRNA in stress granules. Aged neurons accumulate endogenous mitochondria-derived dsRNA that directly binds to PKR. This mitochondrial dsRNA leaks through damaged mitochondrial membranes and forms cytoplasmic foci in aged neurons. Finally, we demonstrate that PKR inhibition leads to the cessation of stress, resumption of cellular translation, and restoration of RNA-binding protein expression. Together, our results identify a source of RNA stress that destabilizes aged neurons and may contribute to neurodegeneration.

cell biology↗

bioRxiv2025

Phosphoinositide 3-kinase regulates wild-type RAS signaling to confer resistance to KRAS inhibition

Despite the availability of RAS inhibitors and the dependence of >90% of pancreatic ductal adenocarcinomas (PDAC) on oncogenic KRAS mutations, resistance to KRAS inhibition remains a serious obstacle. We show here that phosphoinositide 3-kinase (PI3K) plays a major role in this resistance through upstream activation of wild-type RAS signaling - beyond its known KRAS effector function. Combining proximity labeling, CRISPR screens, live-cell imaging, and functional assays we found that PI3K orchestrates phosphoinositide-mediated GAB1 recruitment to the plasma membrane, nucleating assembly of RAS signaling complexes that activate mitogen-activated protein kinase (MAPK) in an EGFR/SHP2/SOS1-dependent manner. We further demonstrate that inhibiting PI3K enhances sensitivity to mutant-specific KRAS inhibitors in PDAC cells, including cells with clinically identified PIK3CA mutations. Our findings refine RAS-PI3K signaling paradigms, reveal that PI3K-driven wild-type RAS activation drives resistance to KRAS inhibition, and illuminate new avenues for augmenting KRAS-targeted therapies in PDAC.

cancer biology↗

bioRxiv2025

Proteomic profiling of primary cilia in the developing brain uncovers newregulators of cortical development

In the developing brain, the primary cilia of radial glial cells extend from the surface of the lateral ventricle, serving as the signaling hub to integrate environmental cues critical for brain formation. Dysfunctions in ciliary proteins contribute to a wide range of brain structural abnormalities in ciliopathies, yet identifying the bona fide ciliary protein components in the brain remains a significant challenge. Here, using proximity labeling and quantitative proteomics, we systematically charted proteins localized to the cilia of radial glial cells in the dorsal and ventral regions of the embryonic brain. From this dataset, we identified cohorts of new molecules intrinsic to the cilia at distinct regions of the developing brain. We validated the cilium localization of several components of the translation machinery and studied the mechanistic roles of ciliary candidates previously linked to brain malformation, including Marcks, a key regulator of radial glial polarity, and Ckap2l, a protein associated with Filippi syndrome. These results revealed previously unrecognized ciliary mechanisms that regulate brain development. Thus, our brain proteomic dataset provides a unique resource for understanding ciliary functions in brain development and the molecular etiology of developmental disorders.

developmental biology↗

bioRxiv2025

2H MRI-based quantification of leucine uptake in glioblastoma multiforme

Glioblastoma (GBM) brain tumors are among the most lethal of all human cancers, with a median survival time of [~]15 months. Treatment planning requires radiologic demarcation of tumor boundaries with contrast-enhanced magnetic resonance imaging (CE MRI); however, significant tumor burden extends beyond the contrast-enhancing margins of the tumor. GBM tumors have an increased expression of amino acid (AA) transporters, including the Alanine, Serine, Cysteine Transporter 2 (ASCT2) and the L-Type Amino Acid Transporter 1 (LAT1). This upregulation has been leveraged in positron emission tomography (PET) studies to detect tumor burden beyond the contrast-enhancing margins identified by standard-of-care CE MRI. Here we leverage recent approaches in deuterium metabolic magnetic resonance with this known upregulation of AA transporters in GBM to demonstrate that 2H MR can detect glioma based on enhanced branched- chain amino acid (BCAA) uptake. To the best of our knowledge, these data represent the first non- invasive quantification of AA concentrations in brain tumor and raises the potential to (i) detect tumor burden beyond contrast-enhancing margins and (ii) quantify AA metabolism using 2H MR spectroscopy.

cancer biology↗

bioRxiv2025

Single-Cell eQTL Mapping Reveals Cell Type-Specific Genetic Regulation in Lung Cancer

Genome-wide association studies (GWAS) have identified over 50 lung cancer risk loci; however, the precise cellular context of these genetic mechanisms remains unclear due to limitations in bulk tissue eQTL analyses. Here, we present the largest single-cell eQTL atlas of human lung tissue to date, profiling 222 donors using multiplexed scRNA-seq. We identified 16,785 eQTLs across 17 cell types, with over 90% of sc-eQTLs and 59% eGene being cell-type-specific, and fewer than 23% were detectable in paired bulk datasets. Integration with GWAS for non-small cell lung cancer highlighted epithelial and immune cells as key contributors to genetic susceptibility, identifying 28 candidate genes within known risk loci and 24 in novel regions. Notably, 47% of established NSCLC susceptibility loci exhibited cell-type-specific pleiotropic genetic regulation. This study provides a valuable resource of lung sc-eQTLs and illuminates how genetic variation modulates gene expression in a cell-type-specific fashion, contributing to lung cancer susceptibility.

genetics↗

bioRxiv2025

Diet-induced phospholipid remodeling dictates ferroptosis sensitivity and tumorigenesis in the pancreas

High-fat diet (HFD) intake has been linked to an increased risk of pancreatic ductal adenocarcinoma (PDAC), a lethal and therapy-resistant cancer. However, whether and how specific dietary fats drive cancer development remains unresolved. Leveraging an oncogenic Kras-driven mouse model that closely mimics human PDAC progression, we screened a dozen isocaloric HFDs differing solely in fat source and representing the diversity of human fat consumption. Unexpectedly, diets rich in oleic acid - a monounsaturated fatty acid (MUFA) typically associated with good health - markedly enhanced tumorigenesis. Conversely, diets high in polyunsaturated fatty acids (PUFAs) suppressed tumor progression. Relative dietary fatty acid saturation levels (PUFA/MUFA) governed pancreatic membrane phospholipid composition, lipid peroxidation, and ferroptosis sensitivity in mice, concordant with circulating PUFA/MUFA levels being linked to altered PDAC risk in humans. These findings directly implicate dietary unsaturated fatty acids in controlling ferroptosis susceptibility and tumorigenesis, supporting potential "precision nutrition" strategies for PDAC prevention.

cancer biology↗

bioRxiv2025

Proximity based proteomics reveals Git1 as a regulator of Smoothened signaling

The GPCR-like protein Smoothened (Smo) plays a pivotal role in the Hedgehog (Hh) pathway. To initiate Hh signaling, active Smo binds to and inhibits the catalytic subunit of PKA in the primary cilium, a process facilitated by G protein-coupled receptor kinase 2 (Grk2). However, the precise regulatory mechanisms underlying this process, as well as the events preceding and following Smo activation, remain poorly understood. To address this question, we leveraged the proximity labeling tool TurboID and conducted a time-resolved proteomic study of Smo-associated proteins over the course of Hh signaling activation. Our results not only confirmed previously reported Smo interactors but also uncovered new Smo-associated proteins. We characterized one of these new Smo interactors, Grk-interacting protein 1 (Git1), previously known to modulate GPCR signaling. We found that Git1 localizes to the base of the primary cilium, where it controls the cilium transport of Grk2, an early event in Hh signaling. Loss of Git1 impairs Smo phosphorylation by Grk2, a critical step for Smo-PKA interaction, leading to attenuated Hh signaling and reduced cell proliferation in granule neuron precursors. These results revealed a critical regulatory mechanism of Grk2 phosphorylation on Smo in the primary cilium. Our Smo-TurboID proteomic dataset provides a unique resource for investigating Smo regulations across different stages of Hh pathway activation.

cell biology↗

bioRxiv2024

De novo functional discovery of peptide-MHC restricted CARs from recombinase-constructed large-diversity monoclonal T cell libraries

Chimeric antigen receptors (CAR) that mimic T cell receptors (TCR) on eliciting peptide-major histocompatibility complex (pMHC) specific T cell responses hold great promise in the development of immunotherapies against solid tumors, infections, and autoimmune diseases. However, broad applications of TCR-mimic (TCRm) CARs are hindered to date largely due to lack of a facile approach for the effective isolation of TCRm CARs. Here, we establish a highly efficient process for de novo discovery of TCRm CARs from human naive antibody repertories by combining recombinase-mediated large-diversity monoclonal library construction with T cell activation-based positive and negative screenings. Panels of highly functional TCRm CARs with peptide-specific recognition, minimal cross-reactivity, and low tonic signaling were rapidly identified towards MHC-restricted intracellular tumor-associated antigens MAGE-A3, NY-ESO-1, and MART-1. Transduced TCRm CAR-T cells exhibited pMHC-specific functional avidity, potent cytokine release, and efficacious and persistent cytotoxicity. The developed approach could be used to generate safe and potent immunotherapies targeting MHC-restricted antigens.

bioengineering↗

bioRxiv2024

Rubisco packaging and stoichiometric composition of a native β-carboxysome

Carboxysomes are anabolic bacterial microcompartments that play an essential role in carbon fixation in cyanobacteria. This self-assembling proteinaceous organelle encapsulates the key CO2-fixing enzymes, Rubisco and carbonic anhydrase, using a polyhedral shell constructed by hundreds of shell protein paralogs. Deciphering the precise arrangement and structural organization of Rubisco enzymes within carboxysomes is crucial for understanding the formation process and overall functionality of carboxysomes. Here, we employed cryo-electron tomography and subtomogram averaging to delineate the three-dimensional packaging of Rubiscos within {beta}-carboxysomes in the freshwater cyanobacterium Synechococcus elongatus PCC7942 that were grown under low light. Our results revealed that Rubiscos are arranged in multiple concentric layers parallel to the shell within the {beta}-carboxysome lumen. We also identified the binding of Rubisco with the scaffolding protein CcmM in {beta}-carboxysomes, which is instrumental for Rubisco encapsulation and {beta}-carboxysome assembly. Using QconCAT-based quantitative mass spectrometry, we further determined the absolute stoichiometric composition of the entire {beta}-carboxysome. This study and recent findings on the {beta}-carboxysome structure provide insights into the assembly principles and structural variation of {beta}-carboxysomes, which will aid in the rational design and repurposing of carboxysome nanostructures for diverse bioengineering applications.

biochemistry↗

bioRxiv2024

Wilting Wildflowers and Bummed-Out Bees: Climate Change Threatens U.S. State Symbols

Species designated as state symbols in the United States carry cultural importance and embody historical heritage. However, they are threatened by climate change and even face the risk of local or global extinction. The responses of these species to climate change have received little attention. In this study, we examine the effects of climate change on 64 state flowers and 68 state insects in the United States by employing correlative species distribution models (SDMs). We select a variety of commonly used SDM algorithms to construct an ensemble forecasting framework aimed at predicting the potential climatic habitats for each species under both historical (1981-2010) and future (2071-2100) climate scenarios (SSP1-2.6 and SSP5-8.5), and how these changes might influence the habitat suitability of flower and insect species within their symbolic states and across the United States. Our results indicate that 30 - 66% of state flowers and 18 - 51% of state insects are projected to experience substantial losses of climatically suitable habitat within their symbolic states. Under the high-emissions scenario (SSP5-8.5), ten state flowers and three state insects are likely to face local extinction by the 2080s. Although most of these species may find suitable habitats in other states, only two are projected to have such areas located adjacent to their current symbolic states, potentially limiting natural dispersal. Nationally, 85% of flower species and 71 - 79% of insect species are expected to shift their suitable habitat both poleward and uphill, with the magnitude of latitudinal and elevational shifts significantly greater under SSP5-8.5 than under SSP1-2.6. These findings highlight the vulnerability of culturally significant species to climate change and underscore the urgency of integrating climate adaptation into conservation planning. Proactive, forward-looking conservation and management strategies may be critical for preserving cultural heritage and maintaining ecosystem resilience.

ecology↗

bioRxiv2024

Structural mechanism of FusB-mediated rescue from fusidic acid inhibition of protein synthesis

Antibiotic resistance protein FusB rescues protein synthesis from inhibition by fusidic acid (FA), which locks elongation factor G (EF-G) to the ribosome after GTP hydrolysis. Here, we present time-resolved single-particle cryo-EM structures explaining the mechanism of FusB-mediated rescue. FusB binds to the FA-trapped EF-G on the ribosome, causing large-scale conformational changes of EF-G that break ribosome interactions. This leads to dissociation of EF-G from the ribosome, followed by FA release. We also observe two independent binding sites of FusB on the classical-state ribosome, overlapping with the binding site of EF-G to each of the ribosomal subunits, yet not inhibiting tRNA delivery. Our results reveal an intricate resistance mechanism involving specific interactions of FusB with both EF-G and the ribosome, and a non-canonical release pathway of EF-G.

molecular biology↗

bioRxiv2024

Comparative and population genomics analyses of eared pheasants inhabiting highly varying altitudes

Oxygen pressure varies dramatically with altitudes on Earth; however, humans and animals thrive at almost all altitudes. To better understand genetic basis underlying adaptation of closely related species to varying altitudes, we annotated and compared the genome of a white eared pheasant (WT) (Crossoptilon crossoptilon) inhabiting high altitudes and the genome of a brown eared pheasant (BR) (C. mantchuricum) inhabiting low altitudes. Moreover, we compared genetic variations in populations of WT and BR as well as of blue eared pheasants (BL) (C. auritum) inhabiting intermediate altitudes, and identified thousands of selective sweeps in each species. Intriguingly, the unique genes and pseudogenes in the two genomes converge on the same set of altitude adaptation-related pathways of four functional categories as genes in selective sweeps in each species. Thus, these species appear to adapt to highly varying altitudes by diverging selection on the same traits via loss-of-function mutations and fine-tuning genes in common pathways.

bioinformatics↗

bioRxiv2024

Large scale loss-of-function mutations during chicken evolution and domestication

Evolution and domestication are often driven by genetic innovations, yet the role of gene loss remains debated. Here, we present comparative genomic analyses of four indigenous chicken breeds from Yunnan Province, China and a reference red jungle fowl genome (GRCg6a). We identify extensive gene presence-absence variation and large numbers of pseudogenes, revealing highly dynamic gene repertoires among closely related chickens. By reconstructing ancestral gene content, we estimate that the common ancestor harbored at least 21,972 genes, of which 7,993 are dispensable. Each lineage has independently lost thousands of genes through both complete gene loss and pseudogenization. These loss-of-function events are non-random: pseudogenization mutations are biased toward gene termini, frequently fixed in populations, and enriched in specific biological pathways. Notably, patterns of gene loss recapitulate phylogenetic relationships, suggesting that loss-of-function mutations are shaped by selection rather than neutral drift. Analysis of four other chicken genomes assembled using PacBio HiFi reads draws the same conclusion. Thus, our results support a model in which large-scale loss-of-function mutations are a major driver of chicken evolution and domestication, consistent with the "less-is-more" hypothesis of adaptive evolution.

bioinformatics↗

bioRxiv2024

SS18::SSX redistributes BAF chromatin remodelers selectively to activate and repress transcription

Synovial sarcoma (SyS) is driven by the expression of a chromosomal translocation-generated SS18::SSX fusion oncoprotein1,2. This oncoprotein fuses the SSX carboxy terminal tail that binds ubiquitylated histone H2A (H2AK119ub)3-6 to the SS18 component of both canonical (CBAF) and non-canonical (GBAF, GLTSCR1-containing) BAF-family chromatin remodeling complexes7-11, wherein SS18::SSX reprograms the epigenetic landscape. Mice that express SS18::SSX develop tumors that faithfully recapitulate human SyS histopathologically and molecularly12-14, allowing dissection of transcriptional reprogramming in vivo15,16. Here, we show that SS18::SSX redistributes GBAF complexes broadly to promoters and distal enhancers decorated by H2AK119ub, which uncharacteristically lack the transcription-silencing trimethylation of H3K27 that otherwise accompanies H2AK119ub. Fusion-GBAF instead associates with H2AK119ub and transcription-enabling H3K4 trimethylation (promoters), monomethylation (distal enhancers), and H3K27 acetylation (both). CBAF with the fusion redistributes away from typical loci, avoids H2AK119ub-bearing regions, and instead narrowly flanks transcription start sites (TSSs), co-distributed with polybromo BAF (PBAF). Accelerated tumorigenesis in our SyS mouse model followed deletion of Smarcb1 (PBAF and CBAF), Pbrm1 (PBAF-specific), and Arid1a or Arid1b (CBAF-specific). While tumors lacking Arid1a or Arid1b retained SyS character, loss of Smarcb1 or Pbrm1 resulted in tumors lacking SyS features. These findings suggest that SyS transcriptional reprogramming includes both improper GBAF localization and gene activation at H2AK119ub-bearing regulatory regions and improper gene silencing through loss of CBAF.

cancer biology↗

bioRxiv2024

Uncovering the roles of the scaffolding protein CsoS2 in mediating the assembly and shape of the α-carboxysome shell

Carboxysomes are proteinaceous organelles featuring icosahedral protein shells that enclose the carbon-fixing enzymes, Rubisco, alone with carbonic anhydrase. The intrinsically disordered scaffolding protein CsoS2 plays a vital role in the construction of -carboxysomes through bridging the shell and cargo enzymes. The N-terminal domain of CsoS2 binds Rubisco and facilitates Rubisco packaging within the -carboxysome, whereas the C-terminal domain of CsoS2 (CsoS2-C) anchors to the shell and promotes shell assembly. However, the role of the middle region of CsoS2 (CsoS2-M) has remained elusive. Here, we conducted indepth examinations on the function of CsoS2-M in the assembly of the -carboxysome shell by generating a series of recombinant shell variants in the absence of cargos. Our results reveal that CsoS2-M assists CsoS2-C in the assembly of the -carboxysome shell and plays an important role in shaping the -carboxysome shell through enhancing the association of shell proteins on both the facet-facet interfaces and flat shell facets. Moreover, CsoS2-M is responsible for recruiting the C-terminal truncated isoform of CsoS2, CsoS2A, into -carboxysomes, which is crucial for Rubisco encapsulation and packaging. This study not only deepens our knowledge of how the carboxysome shell is constructed and regulated but also lays the groundwork for engineering and repurposing carboxysome-based nanostructures for diverse biotechnological purposes.

synthetic biology↗

bioRxiv2024

Systematic Evaluation of Cell Type Deconvolution Methods for Plasma Cell-free DNA

Plasma cell-free DNA (cfDNA) is derived from cellular death in various tissues. Investigating the origin of cfDNA through tissue/cell type deconvolution allows us to detect changes in tissue homeostasis that occur during disease progression or in response to treatment. Consequently, cfDNA has emerged as a valuable noninvasive biomarker for disease detection and treatment monitoring. Although there are numerous methylation-based methods of cfDNA cell type deconvolution available, a comprehensive and systematic evaluation of these methods has yet to be conducted. In this study, we thoroughly benchmarked five previously published methods: MethAtlas, cfNOMe, CelFiE, CelFEER, and UXM. Utilizing deep whole-genome bisulfite sequencing data from 35 human cell types, we generated cfDNA mixtures with known ground truth to assess the deconvolution performance under various scenarios. Our findings indicate that different factors, including sequencing depth, reference marker selection, and reference completeness, influence cell type deconvolution performance. Notably, omitting cell types present in a mixture from the reference leads to suboptimal results. Despite each method exhibited distinct performances under various scenarios, CelFEER and UXM exhibit overall superior performance compared to the others. In summary, we comprehensively evaluated factors influencing methylation-based cfDNA cell type deconvolution and proposed general guidelines to maximize the performance.

bioinformatics↗

bioRxiv2024

Click-free imaging of carbohydrate trafficking in live cells using an azido photothermal probe

Real-time tracking of intracellular carbohydrates remains challenging. While click chemistry allows bio-orthogonal tagging with fluorescent probes, the reaction permanently alters the target molecule and only allows a single snapshot. Here, we demonstrate click-free mid-infrared photothermal (MIP) imaging of azide-tagged carbohydrates in live cells. Leveraging the micromolar detection sensitivity for 6-azido-trehalose (TreAz) and the 300-nm spatial resolution of MIP imaging, the trehalose recycling pathway in single mycobacteria, from cytoplasmic uptake to membrane localization, is directly visualized. A peak shift of azide in MIP spectrum further uncovers interactions between TreAz and intracellular protein. MIP mapping of unreacted azide after click reaction reveals click chemistry heterogeneity within a bacterium. Broader applications of azido photothermal probes to visualize the initial steps of the Leloir pathway in yeasts and the newly synthesized glycans in mammalian cells are demonstrated.

cell biology↗

bioRxiv2024

Phylogenomics resolves a 100-year-old debate regarding the evolutionary history of caddisflies (Insecta: Trichoptera)

Trichoptera (caddisfly) phylogeny provides an interesting example of aquatic insect evolution, with rich ecological diversification, especially for underwater architecture. Trichoptera provide numerous critical ecosystem services and are also one of the most important groups of aquatic insects for assessing water quality. The phylogenetic relationships of Trichoptera have been debated for nearly a century. In particular, the phylogenetic position of the "cocoon-makers" within Trichoptera has long been contested. Here, we designed a universal single-copy orthologue and sets of ultraconserved element markers specific for Trichoptera for the first time. Simultaneously, we reconstructed the phylogenetic relationship of Trichoptera based on genome data from 111 species, representing 29 families and 71 genera. Our phylogenetic inference clarifies the probable phylogenetic relationships of "cocoon-makers" within Integripalpia. Hydroptilidae is considered as the basal lineage within Integripalpia, and the families Glossosomatidae, Hydrobiosidae, and Rhyacophilidae formed a monophyletic clade, sister to the integripalpian subterorder Phryganides. The resulting divergence time and ancestral state reconstruction suggest that the most recent common ancestor of Trichoptera appeared in the early Permian and that diversification was strongly correlated with habitat change.

zoology↗