Biology subjects

Chen, X.

Publications and source records attributed to Chen, X..

At least 127 records · Page 7Linked to original sources

bioRxiv2026

Proton FLASH preserves neurocognition across delivery techniques: implications for clinical translation in pediatric brain tumors

BackgroundRadiation therapy is integral to the curative treatment of childhood brain tumors but contributes to late neurocognitive impairment in survivors. FLASH (ultra-high dose rate, >40Gy/s) reduces normal-tissue toxicity in preclinical models, and proton-FLASH is currently the only modality capable of delivering ultra-high dose rates to the deep targets, such as pediatric brain tumors. However, two questions remain unresolved before clinical translation: (1) whether the FLASH effect can be achieved on synchrotron-based proton systems, which deliver protons in discrete spills that may be insufficient to cover a clinical target within a single delivery, and (2) which dose-rate metric, among the multiple definitions currently used in the field, best predicts the biological FLASH effect. MethodsC57BL/6 mice (7-8 weeks) received 10 Gy whole-brain RT via a clinical Hitachi ProBEAT synchrotron with CBCT-guided delivery, using three transmission-beam techniques: single-spill pencil beam scanning (SS PBS), multi-spill PBS with [~]2-second inter-spot delay (MS PBS), and passive scatter (PS), compared to conventional (CONV) delivery and unirradiated controls (n=24-28/group, equal sex distribution). Dose rate was quantified using three frameworks: field dose rate (FDR), PBS dose rate (PBSDR), and dose-averaged dose rate (DADR). Recognition memory was assessed by novel object recognition (NOR) at 6 weeks post-RT, and cognitive flexibility was assessed via touchscreen visual discrimination and reversal learning at 14 weeks. Hippocampal neuroinflammation was evaluated by immunofluorescence and immunohistochemistry for Iba1, NeuN, and GFAP. ResultsFLASH conditions were met by SS PBS and PS under all three dose-rate definitions, but MS PBS qualified as FLASH only by DADR. Despite this, neuroprotection was preserved across all three FLASH techniques: discrimination index was significantly higher for SS PBS (P=0.021), MS PBS (P=0.008), and PS (P<0.001) versus CONV, with no significant difference between FLASH techniques. On touchscreen testing, FLASH-treated females demonstrated preserved cognitive flexibility (P=0.047 vs. CONV on reversal learning correct trials). Iba1+ microglia were reduced in FLASH compared to CONV mice, with morphology suggestive of preserved homeostatic state. ConclusionsSynchrotron-based proton FLASH preserves neurocognitive function across all delivery techniques, including under multi-spill delivery essential for treating clinical-scale pediatric brain tumors. Critically, this neuroprotection was observed even for deliveries that qualified as FLASH only by DADR, identifying DADR as the dose-rate metric most relevant to the biological FLASH effect with direct implications for clinical trial design and dose-rate reporting standards.

cancer biology↗

bioRxiv2026

A Sandwich-Structured Silk Fibroin Mesh with ROS-Responsive and Immunoregulatory Functions for Pelvic Floor Repair

Current biodegradable meshes for pelvic floor repair are constrained by a limited ability to actively modulate the hostile immune microenvironment following implantation. To address these challenges, we developed a functionalized degradable silk fibroin mesh (SFM) integrated with a reactive oxygen species (ROS)-responsive nanocomposite hydrogel. The resulting composite mesh, SFM@Gel-NP, features a "hydrogel-mesh-hydrogel" sandwich structure, wherein the hydrogel layers are loaded with self-assembled nanoparticles (NP-H-CRFP) for the co-delivery of the STING inhibitor H-151 and the ROS-scavenging agent catechin. This design provides immediate mechanical reinforcement while enabling microenvironment-triggered drug release. In vitro, NP-H-CRFP demonstrated efficient cellular uptake, significant ROS clearance, and effective attenuation of macrophage inflammation and apoptosis. In vivo, SFM@Gel-NP remodeled the local immune milieu by inhibiting the ROS/cGAS-STING/NF-{kappa}B axis, thereby promoting a shift from pro-inflammatory M1 toward pro-regenerative M2 macrophage polarization. This immunomodulatory effect, coupled with enhanced and well-organized collagen deposition--particularly of early type III collagen--resulted in improved tissue integration and repair. This work presents a novel strategy that combines structural reinforcement with active immune regulation, offering a promising next-generation solution for durable and functional pelvic floor reconstruction.

bioengineering↗

bioRxiv2026

The b0+ Amino Acid Transporter Defines a Selenium-utilization Enterocyte Program in the Human Intestine

Selenium is an essential trace element incorporated into selenoproteins as selenocysteine, yet the intestinal cellular programs associated with selenium utilization remain poorly defined. Here, we performed a large-scale single-cell transcriptomic analysis of 169,068 human enterocytes from 105 donors to systematically profile nine candidate selenium transporter systems and their relationship to selenoprotein expression. Among all transporters examined, the b0+ amino acid transporter system (SLC3A1/SLC7A9) showed the strongest and most maturation-independent association with selenoproteins, including SELENOP (Spearman r = 0.489) and GPX4 (r = 0.392), with enrichment across 21 of 24 detected selenoproteins in b0+-complete versus transporter-negative enterocytes. These associations were robust across individual donors and confirmed by pseudobulk validation, and were only partially explained by enterocyte maturation state, intestinal segment, and sequencing depth after covariate adjustment, indicating both differentiation-dependent and differentiation-independent components. Furthermore, enterocytes carrying the b0+ complex expressed a non-canonical LRP repertoire (LRP1, LRP5 and LRP6) rather than the canonical SELENOP receptors LRP2 and LRP8; this co-expression was maturation-dependent, nominating these LRPs as candidate intestinal SELENOP-handling receptors. Together, these single-cell data identify b0+ transporter expression as a marker of a selenoprotein-enriched enterocyte state in the human intestine.

molecular biology↗

bioRxiv2026

Immunomodulatory impact of PTPN11/SHP2-based vertical RAS-MAPK pathway inhibition in pancreatic cancer

Protein tyrosine phosphatase non-receptor 11 (PTPN11/SHP2) is a critical upstream mediator of RAS-MAPK signaling and a central node in adaptive resistance mechanisms evolving with RAS and MEK/ERK inhibition. Accordingly, clinical trials are currently evaluating allosteric SHP2 inhibitors in vertical RAS pathway combination therapies for various KRAS-mutant malignancies, including pancreatic ductal adenocarcinoma (PDAC). Here, we aimed to delineate the immunomodulatory effects of SHP2-based vertical RAS pathway inhibition in notoriously immunotherapy-refractory PDAC, spanning from early treatment response to invariably evolving adaptive resistance. Employing human and murine PTPN11 knockout and wild-type PDAC cell lines, an autochthonous murine PDAC model (KPC), and patient-derived PDAC organoids, we find that short term dual MEK/SHP2 inhibition induces increased T cell infiltration and a reduction in immunosuppressive M2-like macrophages. However, these effects are accompanied by a decrease in mature dendritic cells and a concomitant expansion of monocytic myeloid-derived suppressor cells, indicative of a mixed immunological response with both immune-activating and immune-suppressing features. These changes are associated with tumor cell-intrinsic upregulation of CXCR3 ligands and TGF-{beta}, as well as increased expression of checkpoint ligands for TIGIT and TIM-3 across molecular subtypes and species. With prolonged treatment and transition to an adaptive resistant tumor cell state, the initial immune-sensitizing effects are lost and the immune-suppressive features prevail. M2-like macrophages re-accumulate, dendritic cell maturation remains impaired, TGF-{beta} expression persists, and TIGIT and TIM-3 ligand expression is further enhanced. Notably, dual SHP2/RAS inhibition recapitulates the observed induction of TGF-{beta} and checkpoint ligands. Collectively, these findings identify a dynamic but ultimately immunosuppressive remodeling of the tumor microenvironment in response to SHP2-based vertical RAS pathway inhibition in PDAC and provide a rationale for combinatorial immunotherapy strategies. In particular, concurrent targeting of TGF-{beta}, combined TIGIT/TIM-3 checkpoint blockade, and likely CD40 agonism may help sustain early immune activation while counteracting emerging suppressive features, thereby improving the durability of tumor control.

immunology↗

bioRxiv2026

Genos-m: a foundation model for human-associated microbial genomes

Human-associated microbial genomes encode extensive strain-level diversity and niche-specific gene repertoires that are critical to host health. However, these complex sequence features remain difficult to capture using general-purpose DNA foundation models, highlighting the need for dedicated representation learning tailored to the human microbiome. Here, we introduce Genos-m, an open-source foundation model for human-associated microbial genome representation. Genos-m was pretrained on approximately 1.2 trillion nucleotide tokens from a curated microbial genome corpus, including human-associated prokaryotic isolates, high-quality metagenome-assembled genomes (MAGs) and bacteriophages, supplemented with GTDB species-level representative genomes to broaden prokaryotic taxonomic breadth. The model uses a sparsely activated Mixture-of-Experts (MoE) Transformer architecture, with 4.7 billion total parameters, approximately 330 million activated parameters per forward pass and a maximum context length of one million base pairs. We evaluated frozen Genos-m representations across short-sequence and gene-level tasks, biosynthetic gene cluster (BGC)-based regional sequence tasks, whole-genome strain phenotype prediction, and zero-shot transfer on prokaryote-related RNAfitness assays. Across these benchmarks, Genos-m consistently ranked among the leading comparison models, with the best performance in five of eight gene-fitness regression tasks and in BGC type classification. Using sparse autoencoders, we identified sparse features in Genos-m hidden activations that aligned with annotated ORFs, intergenic regions, and tRNA and rRNA loci. In downstream applications, Genos-m-derived genome-informed species representations in-corporated into a human microbiome self-supervised learning model improved colorectal cancer (CRC)-control classification over conventional species-abundance random forest models. Genos-m also generated stable sample-level embeddings from as few as 10,000 metagenomic reads, retaining gut microbial community structure that distinguished geographic origin and aligned with enterotypes defined from full-depth taxonomic profiles. Together, these results support Genos-m as a reusable representation model for microbial genomes and metagenomes, with conclusions bounded by the reported datasets, task definitions and evaluation protocols. Genos-m model weights, inference code, and usage documentation are publicly available on GitHub (https://github.com/BGI-HangzhouAI/Genos-m) and Hugging-Face (https://huggingface.co/BGI-HangzhouAI/Genos-m).

bioinformatics↗

bioRxiv2026

STRIP2 Stabilizes LCN2 to Suppress Ferroptosis and Drives Colorectal Cancer Malignancy

Colorectal cancer (CRC) presents a significant global health challenge with high incidence rates, mortality, and poor five-year survival rates, particularly in advanced cases, despite the availability of existing treatments. This study investigates the role of Striatin-interacting protein 2 (STRIP2) in CRC progression and the underlying mechanisms. To evaluate the impact of STRIP2 on CRC, we conducted in vitro experiments to assess cell proliferation, migration, invasion, and apoptosis in CRC cell lines. Furthermore, in vivo studies were performed utilizing nude mouse models. Transcriptomic analysis, Western blotting, co-immunoprecipitation assays, and ferroptosis validation were employed to elucidate the role of STRIP2 in CRC. The findings indicate that STRIP2 is overexpressed in CRC and drives malignant behaviors in both in vitro and in vivo settings. Overexpressing STRIP2 (oe-STRIP2) suppresses ferroptosis, leading to increased cell proliferation and reduced oxidative stress, whereas silencing STRIP2 (si-STRIP2) has the opposite effect. Mechanistically, STRIP2 stabilizes LCN2, an effector downstream of IL-17, by preventing its K48-linked ubiquitination and subsequent degradation. This stabilization enhances the anti-ferroptosis capacity of CRC cells, thereby promoting cell survival and facilitating malignant progression in CRC. These findings suggest STRIP2 as a promising diagnostic biomarker and therapeutic target for CRC intervention and warrant further investigation.

cancer biology↗

bioRxiv2026

Direct RNA Sequencing reveals epitranscriptomic regulation of brain cells and Alzheimer's Disease pathology

Alternative mRNA splicing and post-transcriptional RNA modification are key mechanisms that regulate transcript function; however, their role in neuronal activity and neurodegenerative disease remains poorly defined. In this study, we evaluated two nanopore-based long-read sequencing (LR-seq) formats: cDNA-PCR sequencing (CPS) and direct RNA sequencing (DRS). We then applied DRS to profile both full-length isoforms and RNA modifications in major brain cell types derived from induced pluripotent stem cells (iPSCs) and post-mortem Alzheimers disease (AD) brains. Relative to CPS, DRS achieved higher accuracy and sensitivity for transcript quantification, de novo transcript model construction, and open reading frame (ORF) annotation across neuropathological gene sets. Focusing on iPSC-derived neurons, we built a multi-omic atlas to connect transcriptional output with translational engagement and protein abundance, by integrating DRS-based mRNA abundance, N6-methyladenosine (m6A) status and poly(A) tail length with ribosome profiling (Ribo-seq) and mass spectrometry (MS). The combination of DRS and Ribo-seq data demonstrated synergism in predicting protein abundance. This analysis also uncovered a significant inverse relationship between m6A modification and mRNA abundance, which was dependent on the engagement of the ribosomal A-site. Lastly, we applied DRS to the epitranscriptomic analysis of AD brain samples, demonstrating that m6A profiles can be used to distinguish early-versus late-stage disease.

neuroscience↗

bioRxiv2026

Applications of adeno-associated virus for 3D single-cell morphometric analysis in iPSC-derived midbrain organoids.

Human midbrain organoids (hMBOs) are emerging in vitro models to mirror the cellular diversity and the structural complexity of the developing human brain. However, the dense neural network, limits the investigation of individual cells morphology or cell-cell connectivity, which is mostly restricted to fixed organoids following extensive optical clearing techniques. To better resolve individual cells within a brain organoid and for longitudinal tracking of its growth and development, we turned to adeno-associated virus (AAVs) for targeted gene delivery. In particular, we applied AAVs for expressing specific markers that provide the foundation to image individual cells within 3D hMBOs. Thus, we developed a phenotypic platform to specifically inspect the neuronal and astrocytic cytoarchitecture and to examine their connectivity in living hMBOs derived from two genetically unrelated control iPSC lines. We demonstrate that through AAV transduction, we could capture and reconstruct the 3D architecture of both neurons and astrocytes within the hMBO as a whole. Transduced cells exhibited an intrinsic heterogeneity in term of soma volume, arbor complexity and territory covered, regardless of both genetic background, age, and cell-type. Yet, these cellular morphometrics remained equivalent between the two cell lines, indicative of homogeneity in hMBO cellular development. We were able to establish longitudinal profiling of transduced cells, demonstrating how neurons and astrocytes could expand their network over time. Lastly, we describe time-lapse studies to track cellular motility and morphology fluctuations in neurons and astrocytes over time, highlighting the dynamic nature of these cells within the ramified architecture of the neural network in the developing hMBOs. Overall, our platform underscores the versatility of AAVs in studying single cell-morphometrics and cellular connectivity for longitudinal monitoring of cellular dynamics in live 3D hMBOs instead of a static snapshot.

neuroscience↗

bioRxiv2026

Chloroplast genome engineering of potato enables diterpene production without agronomic penalty

Terpenes constitute the largest and most structurally diverse class of plant secondary metabolites, with critical roles in plant-environment interactions and broad industrial applications. Although nuclear genome engineering of terpene pathways has been extensively explored, chloroplast genome engineering remains largely undeveloped, with all reported studies restricted to the model plant Nicotiana. Here we report successful chloroplast genome engineering for diterpene production in the crop plant potato (Solanum tuberosum). First we identified the trnT/trnL plastomic locus as optimal for minimizing integration-associated growth penalties. Insertion of a bifunctional diterpene synthase gene into this plastomic site yielded transplastomic plants with successful diterpene production, but with reduced growth. The co-expression of a geranylgeranyl diphosphate synthase gene to enhance precursor supply restored normal growth while elevating diterpene accumulation. Transplastomic plants were otherwise agronomically comparable to wild-type. This work expands chloroplast engineering as a viable strategy for terpene pathway engineering in crop improvement and high-value terpene production.

bioengineering↗

bioRxiv2026

TDP-43 regulates chromatin looping and gene transcription through binding and stabilizing DNA G-quadruplex structures

TAR DNA-binding protein 43 (TDP-43) is a multifunctional DNA/RNA-binding protein implicated in transcriptional and post-transcriptional regulation. Dysregulation of TDP-43 is closely correlated with human diseases such as cancer and neurodegenerative diseases. Although its roles in RNA metabolism are well characterized, its function in transcriptional regulation remains largely underexplored. DNA G-quadruplexes (dG4s) are non-canonical nucleic acid structures enriched at gene promoters and regulatory elements, where they facilitate chromatin looping and gene transcription. Here, we investigated the transcriptional regulatory role of TDP-43 by integrating multi-omics datasets, including Hi-C, dG4 ChIP-seq, TDP-43 ChIP-seq, RNA-seq and ATAC-seq from K562 and HepG2 cells. Our analyses demonstrate TDP-43 binding and dG4s formation are highly colocalized at chromatin loop anchors, particularly at promoter and enhancer regions. TDP-43 occupancy at these anchors correlates with increased dG4 stability, chromatin loop interaction frequency, elevated chromatin accessibility, and upregulated gene expression. Morover, TDP-43 knockdown in HepG2 cells revealed a significant reduction in dG4 formation and loop interaction strength, accompanied by widespread transcriptional dysregulation. Collectively, our findings highlight a novel regulatory role of TDP-43 in facilitating long-range chromatin interactions and transcriptional activation through binding to and stabilizing dG4 structures, providing a mechanistic basis for gene dysregulation driven by TDP-43 dysfunction in diseases.

bioinformatics↗

bioRxiv2026

Mapping of Stripe Rust and Leaf Rust Resistance Genes in the Hard Red Winter Wheat Population Green Hammer/Lonerider

Stripe rust and leaf rust, caused by Puccinia striiformis f. sp. tritici and P. triticina, respectively, are the most destructive wheat diseases in the southern Great Plains. Green Hammer is a hard red winter wheat (HRWW) cultivar released by Oklahoma State University in 2018 and has demonstrated a stable adult plant resistance to stripe rust and race-specific seedling resistance to leaf rust. To identify and map rust resistance loci, 109 doubled haploid (DH) lines derived from the cross between Green Hammer and another HRWW cultivar, Lonerider, were developed. Lonerider showed adult plant resistance to stripe rust but was susceptible to multiple P. triticina races. The DH lines were evaluated for stripe rust at the adult plant stage in greenhouse and field environments across Oklahoma, Kansas, and Washington, and for leaf rust at the seedling stage against seven U.S. P. triticina races and at the adult plant stage in Oklahoma and Texas. Genotyping-by-sequencing generated 6,078 polymorphic single-nucleotide polymorphisms used for genetic mapping. Quantitative trait loci (QTL) analysis identified 14 stripe rust and 8 leaf rust resistance QTL. For stripe rust, a major QTL in Green Hammer, QYr.osughln-2AS, was identified in the proximity of the 2NvS translocation. Three other major stripe rust resistance QTL were identified in Lonerider on chromosomes 2AL (two QTL) and 2BS (one QTL). For leaf rust, QLr.osughln-1DS and QLr.osughln-2DS.1 were the two major QTL identified in Green Hammer and most likely correspond to the all-stage resistance genes Lr21 and Lr39, respectively. In this study, we identified previously characterized genes as well as unknown genes that can be utilized in wheat breeding programs to enhance resistance to leaf rust and stripe rust.

genetics↗

bioRxiv2026

A Conserved Obg-type GTPase sustains protein synthesis and its physiological roles in eukaryotic cells

Translation is regulated both temporally and spatially to ensure that proteins are synthesized at the correct time and delivered to the proper subcellular location. Temporary slowing or halting of ribosomal movement along an mRNA, a physiological pausing, allows cells to coordinate protein production with internal cues and external signals. However, how cells distinguish such constructive pauses from harmful pathological stalls remains a fundamental unresolved question in molecular biology. Here, we report that a ribosome-binding GTPase (Rbg in yeast, and so-called Developmentally Regulated GTP-binding (Drg) in human) promotes protein biosynthesis in halted ribosomes in an evolutionarily conserved manner. We show that the essential bacterial Obg GTPase rescues cell growth and translation in Saccharomyces cerevisiae and human cells lacking their endogenous Rbg/Drg proteins in a GTPase-dependent manner. Furthermore, Obg enhances puromycin incorporation into stalled ribosomes in a manner requiring GTP hydrolysis. We reason that Rbg/Drg proteins act to prevent premature engagement of ribosome-associated quality-control or stress-response pathways. Under physiological conditions, proteomic analyses of Rbg1-bound ribosomes in yeast reveal that Rbg GTPases associate with ribosomes translating proteins essential for diverse functions, including those of the nucleus, ER, Golgi, mitochondria, and protein transport pathways. Finally, loss of even one of the two eukaryotic Rbg/Drg paralogs impedes cell-cycle progression, disrupts DNA replication, and compromises mitochondrial function. Together, these findings suggest that Rbg/Drg proteins enable cells across different domains of life to protect physiological ribosomal pauses, revealing a previously unrecognized layer of regulation in gene expression. Significance StatementHow cells distinguish productive ribosomal pauses from harmful stalls remains unclear. We identify a ribosome-binding GTPase--Rbg in yeast and Drg in humans--as a conserved regulator that protects physiological pauses and sustains protein synthesis. Remarkably, bacterial Obg functionally substitutes for eukaryotic Rbg/Drg in both yeast and human cells, revealing deep evolutionary conservation of this mechanism. Our data show that Obg promotes continued peptide-bond formation in a GTPase-dependent manner. Positioned near the ribosomal GTPase-associated center, these GTPases are also poised to compete with stringent-response and stress-activation factors that trigger translation shutdown. Together, the findings suggest that translation is controlled not only by GTPases that drive protein synthesis forward but also by Rbg/Drg GTPases that protect the catalytic cycle against premature arrest.

biochemistry↗

bioRxiv2026

AAV tools enable functional modulation and readout of central and peripheral nervous systems in spiny mice

Among mammals, spiny mice (Acomys spp.) exhibit the unique capacity to regenerate parts of their nervous system. Studying this phenomenon has the potential to reveal new targets that can slow or halt human neurodegenerative disorders. Unfortunately, research tools (e.g., transgenic lines, gene delivery vehicles) are lacking compared to those available for other rodent models. Here, we tested systemic adeno-associated viral vectors (AAVs) in Acomys dimidiatus and identified three promising candidates: X1.1, CAP-Mac, and MaCPNS1. Characterizing their tropism following intravenous delivery, we found that in the brain, MaCPNS1 and X1.1 primarily transduced astrocytes. In the peripheral nervous system, MaCPNS1 efficiently transduced dorsal root ganglia, axon bundles of the ear pinnae, and enteric neurons throughout the gastrointestinal tract. As a proof-of-concept, we used MaCPNS1 to chemogenetically modulate the activity of enteric neurons, successfully decreasing gastric motility in vivo and increasing colonic motility ex vivo. We expect these findings to enable functional studies of the uniquely regenerative nervous system of Acomys, which may in turn help advance neuroregenerative therapeutics for humans. Summary StatementIdentification of an AAV tool to efficiently deliver transgenes to the central and peripheral nervous systems of spiny mice enables functional studies of the nervous system in a mammalian model of regeneration.

neuroscience↗

bioRxiv2026

Chemogenetic Inhibition of the Cortical Amygdala Reduces Alcohol Intake and Restores Thalamic Connectivity in Dependent Female Mice

BackgroundAlcohol use disorder is a chronic relapsing condition characterized by excessive drinking and withdrawal symptoms. Alcohol dependence disrupts function across multiple brain regions, and recent evidence implicates the cortical amygdala (CoA) as a critical node in alcohol-related circuits. However, how CoA activity influences alcohol intake and brain-wide network function during withdrawal remains unclear. MethodsAlcohol dependence was induced using chronic intermittent ethanol vapor (CIE). In one cohort, electrophysiological activity of CoA neurons was assessed during withdrawal. In a second cohort, mice underwent CIE paired with two-bottle choice drinking, and inhibitory DREADDs (hM4Di) were used to suppress CoA activity during drinking and withdrawal while behavioral outcomes were measured. Brains were then collected for Fos immunolabeling and iDISCO+ based whole-brain activity mapping to determine how CoA inhibition during withdrawal altered network organization. ResultsRepeated CIE increased alcohol sensitivity in CoA neurons during withdrawal. Chemogenetic inhibition of the CoA reduced alcohol intake in dependent mice without affecting withdrawal-related behaviors. Whole-brain Fos mapping showed that CoA inhibition reduced activity within the CoA while enhancing functional connectivity across multiple brain regions, particularly in the isocortex, thalamus, and anterior hypothalamic nucleus. During withdrawal without CoA inhibition, thalamic regions exhibited negative connectivity, consistent with disrupted network function; CoA inhibition reversed this pattern, producing strongly positive thalamic and medial prefrontal cortex connectivity. ConclusionsThese findings demonstrate that alcohol dependence alters CoA sensitivity, alcohol dependence-induced drinking and brain-wide network organization during withdrawal. The CoA appears to selectively regulate withdrawal-associated alcohol drinking, and its inhibition may reduce intake by restoring thalamic and cortical connectivity. HighlightsO_LIThis study identifies the cortical amygdala as a previously underexplored brain region involved in alcohol-related behaviors. C_LIO_LIBy integrating chemogenetic inhibition with brain-wide network analysis, the study reveals candidate circuit connections through which the CoA may regulate alcohol dependence-related brain activity. C_LIO_LIThis study establishes the CoA as a potential driver of excessive alcohol drinking and alcohol-related network dysfunction. C_LI

neuroscience↗

bioRxiv2026

NFIC-mediated CaSR endocytosis defines a hyperactive TOMM20highCHGAhigh oxyphil cell state as a pathological driver of autonomous secondary hyperparathyroidism

Secondary hyperparathyroidism (SHPT) is a debilitating complication of chronic kidney disease. Its clinical management is frequently compromised by calcimimetic resistance, a refractory state primarily driven by the progressive downregulation of the membrane calcium-sensing receptor (CaSR). Despite the centrality of CaSR as a therapeutic target, the mechanisms governing its aberrant expression and membrane localization remain incompletely elucidated. Here, we generated a single-cell transcriptomic atlas of human parathyroid tissues from SHPT and primary hyperparathyroidism (PHPT) patients, uncovering a unique stromal-immune niche that is specifically induced by uremic stress in SHPT. Our data also observed a striking dissociation between CaSR mRNA and its abundance as a membrane protein in SHPT tissues. Pseudotime trajectory analysis showed a progressive decline in CaSR pathway activity and a concomitant increase in endo-lysosomal activity along the trajectory, terminating in pathological oxyphil cells as the endpoint of chief cell differentiation in SHPT. Mechanistically, Nuclear Factor I C (NFIC) transcriptionally activated clathrin light chain B (CLTB) and Ras-related protein Rab7a (RAB7A) to trigger clathrin-mediated endocytosis and lysosomal degradation of CaSR, thus reducing its membrane abundance. This degradative program was further validated by multiplex immunofluorescence in TOMM20highCHGAhigh pathological oxyphil cells in human SHPT tissues. To translate these mechanistic findings into a clinically actionable strategy, we repurposed the clinically approved lysosomal inhibitor hydroxychloroquine (HCQ) to block this CaSR degradation pathway. In patient-derived xenograft (PDX) mouse models, co-administration of HCQ and cinacalcet acted synergistically to restore membrane CaSR expression, normalize serum PTH and calcium levels and suppress parathyroid tumor growth more effectively than monotherapies. Collectively, our single-cell-guided study identifies an NFIC-driven endo-lysosomal program as a previously unrecognized mechanism underlying CaSR downregulation and calcimimetic resistance in SHPT, and establishes HCQ repurposing as a clinically tractable therapeutic strategy for patients with refractory SHPT.

cell biology↗

bioRxiv2026

Genetic suppression of myeloid receptor Clec7a attenuates microglia neuroinflammation and promotes microglial phagocytosis to delay disease progression in ALS models

Microglial activation has been closely associated with accelerated ALS disease progression. However, specific microglial pathways that regulate microglial activation and ALS disease progression remain limitedly understood. Here, we determined the role of Clec7a (or Dectin-1), a core signature gene of disease-associated microglia (DAM) in ALS, in regulating microglial activation and ALS disease progression. Our spinal cord scRNA-Seq results found that Clec7a deficiency specifically attenuated microglial neuroimmune gene expression in SOD1G93A mice and human ALS. In addition, in vivo two-photon imaging of human (h) TDP43 phagocytosis by microglia in the cortex showed that Clec7a deficiency promotes microglial phagocytosis of pathological hTDP43 by enhancing microglial process dynamics. Subsequent survival analysis further showed that selective deletion of Clec7a in microglia mitigates motor neuron degeneration and delays disease progression in SOD1G93A ALS mice. Together, our results establish that Clec7a is a key regulator in shaping disease microglial functions and promotes disease progression in ALS.

neuroscience↗

bioRxiv2026

Prediction of Stimulation-Defined Eloquent Cortex Using Graph-Theoretical Connectivity from Electrocorticography During Presurgical Mapping

Background and ObjectivesElectrical stimulation mapping (ESM) is the clinical gold standard for identifying eloquent cortex during presurgical evaluation but is time-intensive, constrained by incomplete cortical sampling, and limited by patient tolerance. We investigated whether features derived from electrocorticography (ECoG), including functional connectivity measures, can provide complementary information for identifying functionally critical cortex and examined how predictive performance varies across functional domains, behavioral tasks, and data quantity. MethodsFourteen patients undergoing intracranial monitoring for epilepsy surgery performed speech production tasks while ECoG was recorded independently of stimulation mapping. Graph-theoretic functional connectivity features derived from high-gamma activity (70-150 Hz), combined with anatomical region encoding, were used to train machine learning classifiers, and predictive performance was evaluated using leave-one-subject-out validation with ESM-defined functional deficits used as ground-truth outcomes. ResultsFourteen patients were included. Performance differed across functional domains, with motor-critical electrodes identified with the highest accuracy (ROC-AUC 0.929 {+/-} 0.061; PR-AUC 0.755 {+/-} 0.191), followed by speech arrest (ROC-AUC 0.793 {+/-} 0.103; PR-AUC 0.550 {+/-} 0.196), whereas language-critical electrodes were more difficult to robustly predict (ROC-AUC 0.761 {+/-} 0.160; PR-AUC 0.385 {+/-} 0.167). Additional signal-derived features provided limited benefit beyond anatomical and connectivity features. Performance also varied across tasks and with the number of available trials, with motor prediction remaining stable across trial counts and speech arrest prediction improving with increasing trial counts up to approximately 10 trials before plateauing. In addition, evaluation at the level of stimulation pairs increased sensitivity, with only modest changes in overall performance. DiscussionGraph-theoretic connectivity analysis of ECoG provides complementary information for identifying stimulation-defined functional criticality and supports presurgical mapping. Differences in predictive performance across functional domains likely reflect underlying neuro-physiologic organization, with connectivity providing the clearest improvement for speech arrest prediction. Combined-label prediction strategies increased sensitivity at the expense of specificity, reflecting a clinically relevant tradeoff between broad detection of critical cortex and precise localization. These findings suggest that connectivity-informed approaches may help guide task selection and improve mapping efficiency while complementing electrical stimulation mapping.

neuroscience↗

bioRxiv2026

Reactivation of a TAL1 progenitor cell enhancer region by non-coding somatic variants in T-lineage acute lymphoblastic leukemia

Aberrant activation of TAL1, a key oncogenic driver, defines a major subgroup comprising [~]30% of childhood T-lineage acute lymphoblastic leukemias (T-ALLs). We and others have shown that somatic non-coding mutations within upstream and intronic cis-regulatory regions of TAL1 contribute to transformation by creating binding sites for MYB and other transcription factors. Here we investigated cis-regulatory mechanisms mediated by somatic mutations occurring in an intergenic region located 29 kilobase pairs downstream of the canonical TAL1 transcription initiation site, implicated in 6% of TAL1-expressing T-ALLs. These somatic variants include i) complex indels resulting in de novo MYB transcription factor binding sites (TFBSs) and ii) internal tandem duplications (ITDs) encompassing canonical MYB TFBSs. Chromatin immunoprecipitation sequencing (ChIP-seq) revealed binding of the TAL1 core regulatory circuit (CRC) transcription factors MYB, GATA3, and RUNX1, resulting in enhancer activity mediated by sequences with the mutant allele. Strikingly, ChIP-seq peaks for the repressive H3K27me3 mark and the active H3K27ac mark co-existed across TAL1 regulatory sequences but enriched for different haplotypes. TAL1 transcription from the mutant haplotype initiated from a promoter located within exon 4 of the canonical TAL1 transcript, resulting in a short isoform normally expressed by hematopoietic stem cells (HSC). Interestingly, neither the isoform expression nor the enhancer activity could be predicted by the sequence-to-function deep learning artificial intelligence (AI) model AlphaGenome, emphasizing the importance of experimental validation. Our findings indicate that selection for cis-regulatory, non-coding variants leads to reactivation of enhancers normally active in HSC but silenced in differentiated lineages during normal hematopoietic cell development.

cancer biology↗