Biology subjects

Huang, Y.

Publications and source records attributed to Huang, Y..

At least 235 records · Page 13Linked to original sources

bioRxiv2025

Himito: a Graph-based Toolkit for Mitochondrial Genome Analysis using Long Reads

Understanding the genetic and epigenetic regulation of mitochondrial DNA (mtDNA) is essential for elucidating mechanisms of aging and disease. Long-read sequencing can span the entire mitochondrial genome and directly capture base-modification signals, yet analytical tools for such data remain limited. We developed Himito, a graph-based toolkit for analyzing mitochondrial genome using long reads. Himito filters reads originating from nuclear mitochondrial insertions (NUMTs), constructs a sequence graph to represent mtDNA diversity, assembles primary haplotypes, calls variants, and analyzes 5-methylcytosine (5mC) modifications within a unified framework. Benchmarking on high-quality reference datasets shows Himito achieves superior performance in assembly and variant calling compared with existing tools. Applied to the All of Us (AoU) v8 dataset, Himito identified pathogenic mtDNA variants, revealed population-scale haplogroup diversity, and uncovered age-related genetic and epigenetic patterns. These results demonstrate that long-read sequencing, combined with graph-based analysis, enables integrated characterization of mitochondrial genomic and epigenomic variation. Himito is available at https://github.com/broadinstitute/Himito.

bioinformatics↗

bioRxiv2025

Brown Adipose Tissue Dysfunction Links Obesogen Exposure to Reduced Energy Expenditure and Transgenerational Obesity

Obesity has become a global health challenge, and increasing evidence suggests that environmental obesogens contribute to its prevalence. Tributyltin (TBT) is a model obesogen known to activate PPAR{gamma} and RXR and to promote transgenerational obesity in mice, but the mechanisms linking TBT exposure to impaired energy balance remain poorly defined. Brown adipose tissue (BAT) is a central regulator of energy expenditure (EE) and basal metabolic rate (BMR) through both UCP1-dependent and UCP1-independent pathways. Here, we tested whether ancestral TBT exposure disrupts BAT function across generations. To assess whether TBT alters body composition and thermogenic capacity, we measured fat and lean mass, BAT gene expression, mitochondrial abundance, and core body temperature in F1 and F3 offspring. TBT-group males accumulated more fat mass without changes in lean mass or food intake scaled to metabolic size, indicating reduced EE/BMR. Expression of BAT lineage and oxidative genes (Zic1, Ebf2, Pgc1a, Pdk4) was suppressed in TBT-group males across generations. Ucp1 expression was unchanged at baseline but decreased after high-fat diet (HFD) challenge, whereas Ucp2 and Ucp4 were reduced even at baseline. In addition, key UCP1-independent thermogenic genes involved in creatine (Slc6a8, Ckb) and calcium (Atp2a2, Itpr) futile cycles were significantly decreased, while lipid-cycle genes (Pnpla2, Abhd5) were unaffected. Mitochondrial DNA content was largely unchanged, but core body temperature was reduced in TBT-group males prior to diet challenge, confirming impaired basal thermogenesis. These findings demonstrate that ancestral TBT exposure produces male-specific and heritable defects in BAT identity and thermogenic capacity. By suppressing both UCP1-dependent and UCP1-independent pathways without altering mitochondrial abundance, TBT-group males exhibit reduced energy expenditure and basal metabolic rate. These results identify BAT dysfunction as a mechanistic link between environmental obesogen exposure and transgenerational susceptibility to obesity.

pharmacology and toxicology↗

bioRxiv2025

A single-component optogenetic toolkit for programmable control of microtubule

Microtubules (MTs) form dynamic cytoskeletal scaffolds essential for intracellular transport, organelle positioning, and spatial organization of signaling. Their architecture and function are continuously remodeled through the concerted actions of microtubule-associated proteins (MAPs), post-translational modifications (PTMs), and molecular motors. To precisely interrogate these processes in living systems, we developed a genetically encoded optogenetic toolkit for spatiotemporal control of MT organization and dynamics. By replacing native multimerization motifs with a blue light-responsive oligoermization domain, we have engineered single-component probes, OptoMT and OptoTIP, that reversibly label MT polymers or track plus-ends with tunable kinetics from seconds to minutes. When coupled to enzymatic effectors, these modules enable localized tubulin acetylation or detyrosination, directly linking PTMs to MT stability. We further engineered OptoMotor, a light-activatable kinesin platform that reconstitutes tail-dependent cargo transport along MTs, and OptoSAW, a light-triggered severing actuator for controlled MT disassembly. Using these tools, we reveal how local MT integrity governs lysosomal trafficking and ER-associated signaling dynamics. Collectively, this versatile single-component toolkit bridges molecular design with cytoskeletal function, offering new avenues to illuminate how dynamic cytoskeletal architectures coordinate intracellular organization, transport, and signaling.

cell biology↗

bioRxiv2025

Hi-C informed kernel association test: integrating 3-dimensional genome structure into variant-set association for whole-genome sequencing data

Variant-set association analysis is a powerful strategy for genetic studies of whole genome sequence (WGS) data, especially for rare variants. By aggregating variant signals, variant-set analysis can improve statistical power, result interpretability, and study replicability. Motivated by evidence that three-dimensional (3D) genome architecture plays a critical role in regulating gene transcription, several works have incorporated 3D genome architecture into gene-based association tests and demonstrated great promise. In this work, we extend the idea of 3D-genome guided test from gene-centric to gene-agnostic, whole-genome testing by introducing a Hi-C informed kernel association test. We present a principled procedure that converts Hi-C contact confidence into borrowing weights and integrates these weights into genetic similarity kernels so that higher-confidence interacting loci contribute more to the association test of the target variant set. We use a controlling parameter to adaptively determine the appropriate degree of information borrowing from its interacting loci during association testing. We assess the performance of the Hi-C informed test using simulations and illustrate its advantage in detecting rare-variant sets using WGS data from the ARIC study in the Trans-Omics for Precision Medicine (TOPMed) program.

bioinformatics↗

bioRxiv2025

Single cell whole genome and transcriptome sequencing links somatic mutations to cell identity and ancestry

The role of somatic mutations in human development and disease is obscured by difficulties in characterizing mutations at the single cell level and identifying cell types carrying them. Here we analysed somatic genomes of clonal iPSC lines and of single-cells after whole-genome amplification (scWGA) by PTA and ResolveOme from skin fibroblasts, blood and urine of a live donor. Mutation burden and spectra converged across approaches, revealing heterogeneous mutational footprints across cells driven by environmental exposures (UV damage and chemotherapy) and lymphocyte differentiation. Aneuploidies in single cells were detected by all the approaches and were orthogonally validated by Strand-seq. Uniquely, ResolveOme enabled cell-type identification using single-cell transcriptomes. Using a newly developed method accounting for noise and allele drop-out in scWGA, we de novo reconstructed the cell phylogenetic tree for this donor. Together, scWGA establishes a powerful foundation for comprehensive, cell type-aware, lineage-aware profiling of somatic mutations at single cell level.

genomics↗

bioRxiv2025

VRC01 Selects Rare HIV Escape Mutations After Acquisition in Antibody-Mediated Prevention Trials

Broadly neutralizing antibodies (bnAbs) show promise in HIV prevention, yet viral escape remains a challenge. In the Antibody Mediated Prevention (AMP) trials, the CD4 binding site (CD4bs) bNAb VRC01 blocked acquisition by VRC01-sensitive strains. However, its influence on viral evolution post-acquisition is not fully understood. Here we analyzed >12,000 HIV env sequences from 47 participants from the AMP trials, identifying VRC01-mediated de novo escape mutations in 8 of 26 VRC01-treated participants but none in 21 placebo participants. These mutations were found at very low frequency (<1%) in global viruses. Escape mutations, primarily located in the Loop-D and {beta}23/V5 regions of Env, conferred cross-resistance to several CD4bs bnAbs, while more potent CD4bs bnAbs like N6 and 1-18 largely retained their activity. Our findings demonstrate that prophylactic VRC01 can select for viral escape after infection, underscoring the need for next-generation bnAbs with improved breadth and potency to enhance durability and efficacy of antibody-based HIV prevention.

microbiology↗

bioRxiv2025

Stress drives the hippocampus to prioritize statistical prediction over episodic encoding

The hippocampus plays a critical role in encoding individual experiences into episodic memory, while also detecting patterns shared across these experiences that allow us to anticipate future events via statistical learning. Acute stress is known to impact the hippocampus but may have opposite effects on these competing functions. That is, although stress impairs episodic encoding, it may instead enhance statistical learning. Using functional magnetic resonance imaging, we tested how stress influences the hippocampal pathways and subfields that support episodic encoding and statistical learning while participants performed a task that engaged both processes. Across several analyses, stress biased hippocampal processing in favour of statistical learning, suppressing pattern separation between events in the dentate gyrus and enhancing the representation of statistically predictive features. Furthermore, stress drove the hippocampal monosynaptic pathway to support statistical learning rather than episodic encoding during predictive situations. Together, these data suggest that acute stress elicits targeted, adaptive changes in hippocampal pathways which may facilitate predicting and responding to upcoming events.

neuroscience↗

bioRxiv2025

Pigs lacking Natural Killer T cells have altered cellular responses to influenza

It is increasingly recognized that innate T cells such as natural killer T (NKT) cells, mucosal associated invariant T (MAIT) cells, and {gamma}{delta} T cells play an important role in shaping adaptive immune responses following influenza infection or vaccination. This is largely through the multiple cytokines these cells release upon activation, which have downstream effects on the scope and magnitude of virus-specific T and B cells, and antibodies which form. Here, we examined the contribution of NKT cells using pigs, which are considered a highly translational model of human influenza A infection. CD1D-expressing and CD1D-deficient pigs that respectively possess and lack NKT cells were infected with the swine influenza virus H3N2 A/Swine/Colorado/23619/1999 (CO99), with or without prior mucosal immunization with a recombinant H3N2 A/Swine/Texas/4199-2/1998 (TX98) modified live vaccine encoding a truncated NS1 protein (TX98 NS1{Delta}126). Vaccination reduced virus load and pulmonary pathology by similar amounts in both genotypes. However, NKT cells status had a significant impact on the underlying immune response: Unlike following vaccination, virus-specific T cell expansion after infection was greater in CD1D-deficient than -intact pigs, indicating that NKT cells play opposing roles in different phases of the immune response. NKT cell-deficient pigs also had reduced T cells cuffing around airways and higher numbers of expanded T and B cell clones according to single-cell {beta} T cell receptor (TCR) and B cell receptor (BCR) sequencing. Using our newly developed porcine-specific {gamma} and {delta} chain primers, we characterized the pulmonary {gamma}{delta} TCR repertoire. This revealed a high frequency of expanded clones, especially within the CD2- subset, and a tendency for larger expansion in NKT cell-expressing pigs. Overall, our results indicate a homeostatic role for NKT cells on several important features of the influenza immune response, including the dynamics of T cell expansion and contraction.

immunology↗

bioRxiv2025

Fixing Hybrid Rice: >99% Efficient Apomixis with Near-Normal Seed Set

Apomixis, a form of clonal seed reproduction, offers a transformative approach to agriculture by enabling the stable fixation of hybrid vigor and elite heterozygosity across generations. However, the practical implementation of synthetic apomixis has been hindered by highly variable clonal efficiency or significant yield penalties. In this study, through integrated transcriptomic analyses, we identified a sperm-specific transcription factor in rice that likely functions as a key initiator of embryogenesis. Ectopic expression of this factor in egg cells effectively induces parthenogenesis and produces haploid progeny. When combined with clonal gametogenesis, this system achieved nearly complete synthetic apomixis, with clonal seed production rates exceeding 99% across all derived hybrid rice lines. Moreover, we generated apomictic hybrid lines that not only consistently produced over 99% clonal seeds but also exhibited seed yields comparable to conventional F1 hybrids. These findings establish a scalable and agriculturally viable platform for synthetic apomixis, achieving stable fixation of heterosis through clonal seeds and paving the way for the commercial deployment of self-perpetuating hybrid crops.

plant biology↗

bioRxiv2025

Optimized reference region and the effect on test-retest reliability and detection of Parkinson's disease with UCB-J.

[11C]UCB-J is a radioligand targeting synaptic vesicle glycoprotein 2A, used to image synaptic density. For quantification, a small-volume centrum semiovale area was previously optimized as a [11C]UCB-J reference region (CS2mL); however, its high variability resulted in reduced reliability. Herin, we evaluated an alternative reference region method to assess longitudinal test-retest reliability and detection of Parkinsons disease (PD). For estimating distribution volume ratio (DVR), CS2mL and eleven white matter (WM) reference regions (range: 0.5-200 mL) were generated using the Freesurfer WM map. Same-day and longitudinal test-retest variability (TRV) were assessed (24 healthy subjects (HS); n=10 same-day and n=20 longitudinal HRRT scans, range: 7-1028 days). Each reference region was used to evaluate the substantia nigra (SN) and caudate DVRs in HS (n=25) and PD (n=20). The 10mL WM reference region yielded [11C]UCB-J DVR measurements with reduced variability in TRV (same-day: 10mL: 1.2{+/-}5.7%, same-day: CS2mL: -0.9{+/-}9.2% longitudinal: 10mL: 1.5{+/-}7.0%, CS2mL: 1.6{+/-}11.9%,) while maintaining <10% volume of distribution difference, compared to CS2mL. Further, a significant difference between PD and HS groups in SN and caudate DVRs was found using 10mL, with greater effect size (Cohens d 0.61 for SN and 0.66 for caudate) compared to CS2mL (0.38 for SN and 0.43 for caudate).

neuroscience↗

bioRxiv2025

Molecular mechanism of the key dormancy regulator DosR-dependent transcription activation in Mycobacterium tuberculosis

Mycobacterium tuberculosis (Mtb) is the etiological agent responsible for the worldwide disease Tuberculosis. DosR is recognized as the key dormancy regulator of Mtb due to its global transcriptional regulation in intracellular adaptation and long-term persistence prevailing in latent infection. However, the molecular mechanism regarding DosR-dependent transcription activation remains inadequately defined. Here, we successfully determined the cryo-EM structure of an intact DosR-dependent transcriptional activation complex (DosR-TAC), comprising Mtb RNA polymerase (RNAP), DosR, and the hypoxic promoter DNA. In DosR-TAC, two DosR monomers symmetrically dimerize through their N-terminal receiver domains (DosR_RECs) and the distinct 10 helices of the C-terminal DNA-binding domains (DosR_DBDs). Unlike its inhibitory configuration, the DosR dimer undergoes significant conformational changes within the original linker, isomerizing DosR_REC into a canonical ({beta})5 fold. This extended linker affords the DosR_DBDs and DosRI_REC to contact promoter DNA, while concurrently interacting with the conserved domains of RNAP ({sigma}AR4, CTD, and NTD). Additionally, RT-qPCR and growth curve assays of the wild-type and DosR mutant strains, also unravel the physiological importance of this divergent linker. These findings, together with prior hypotheses regarding DosR, support an allosteric activation-recruitment model for DosR. Altogether, these results highlight the molecular mechanism of DosR-dependent transcription activation critical for dormancy survival of Mtb, and offer potential targets for developing anti-dormancy strategies against persistent tuberculosis.

microbiology↗

bioRxiv2025

Chronic kidney disease exacerbates hyperlipidemia and atherosclerosis via miR-148a-3p targeting SIK1/AMPKα1 in liver

BackgroundCardiovascular complications are the leading cause of death in patients with chronic kidney disease (CKD). As a major risk factor for atherosclerotic cardiovascular disease (ASCVD), hyperlipidemia is a common sequela of CKD, which is linked to increased lipid synthesis, impaired lipoprotein clearance, and maladapted reverse cholesterol transport. Because exosomes can transport miRNAs to modulate intercellular and tissue communication, we hypothesized that exosomal miRNAs can act as endocrine-like molecules to modulate the CKD-associated hyperlipidemia and ASCVD. MethodsSmall RNA-seq and qPCR were used to determine the differential plasma level of exosomal miR-148a-3p in ASCVD patients with and without CKD. RNA-seq and in silico analysis were used to establish the regulation of lipid metabolism via miR-148a-3p/salt-inducible kinase 1 (SIK1)/AMP-activated protein kinase alpha 1 subunit (AMPK1) in the liver. Inhibition of miR-148a-3p or overexpression of SIK1/AMPK1 in ApoE-/- mice with 5/6 nephrectomy was used to elucidate the role of the miR-148a-3p/SIK1/AMPK1 axis in CKD-related hyperlipidemia and atherosclerosis. ResultsThe plasma level of exosomal miR-148a-3p was significantly elevated in patients with ASCVD and CKD (ASCVD/CKD) and ApoE-/- mice with 5/6 nephrectomy. Via circulating exosomes, kidney-generated miR-148a-3p was delivered to the liver, where it targeted SIK1/AMPK1 transcripts, thereby upregulating SREBP2/PCSK9 and reciprocally downregulating LDLR. Levels of SIK1 and AMPK1 were decreased in hepatocytes transfected with miR-148a-3p or treated with plasma exosomes from ASCVD/CKD patients, with attendant SREBP2 activation and attenuated LDL-C binding. The decreased LDL-C binding was rectified by SIK1/AMPK1 or LNA-miR-148a-3p overexpression. LNA-miR-148a-3p or AAV8-SIK1/AMPK administration in 5/6 nephrectomy/ApoE-/- mice significantly ameliorated hyperlipidemia and atherosclerosis. ConclusionIn ASCVD/CKD patients and nephrectomy/ApoE-/- mice, kidney-originating miR-148a-3p targeted hepatic SIK1/AMPK1 mRNA. This exosome-mediated endocrine effect upregulated SREBP2/PCSK9 and reciprocally reduced LDLR level, which elevated plasma LDL-C level and exacerbated ASCVD. These findings underscore a kidney-liver-artery axis involved in cardiovascular-kidney-metabolic syndrome.

pathology↗

bioRxiv2025

RIG-I-like receptor-dependent type I Interferon regulates antigen dose and activation in yellow fever vaccine 17D-infected antigen presenting cells

The live-attenuated yellow fever vaccine 17D-204 (YF17D) activates robust innate immune responses followed by rapid induction of adaptive immunity resulting in long-lasting protection. YF17D triggers the production of type I interferons (IFNs) which have a dual role in antigen presenting cells regulating their infection and contributing to their activation. Infection with YF17D was detected in primary human blood monocytes and conventional dendritic cells (DCs) and in monocyte-derived DCs but was highly restricted by type I IFN. Blocking IFNAR signaling in YF17D-infected PBMC from vaccinated donors resulted in increased activation of YF17D-specific CD8+ T cells. Consistently, peak IFN-alpha plasma levels correlated inversely with the CD8+ T cells response in YF17D vaccinees. Loss of function experiments demonstrated a dominant role of retinoic acid inducible gene I (RIG-I)-like receptors (RLRs) and mitochondrial antiviral signaling protein (MAVS) for type I IFN induction and restriction of YF17D. The type I IFN response was mediated by 5 tri- or diphosphate dsRNA intermediates that are formed during YF17D infection. In vivo proximity labelling (IPL) of RIG-I and next-generation sequencing confirmed interaction of RIG-I with YF17D-dsRNA in infected cells. Thus, YF17D-triggered RLR-signaling restricts viral replication through type I IFN and thus limits the production of viral antigens that can be presented to T cells.

immunology↗

bioRxiv2025

Return of the GEDAI: Unsupervised EEG Denoising based on Leadfield Filtering

Current electroencephalogram (EEG) denoising methods struggle to remove the complex physiological and environmental artifacts typical of real-world settings, which both hinders the isolation of true neural activity and limits the technologys translational potential. We present the Generalized Eigenvalue De-Artifacting Instrument (GEDAI), a novel algorithm for denoising highly contaminated EEG. GEDAI employs leadfield filtering to selectively remove noise and artifacts that diverge from a theoretically defined EEG forward model. This approach offers unique advantages over existing solutions, including 1) denoising of highly corrupt recordings without "clean" reference data, 2) single-step correction of artifactual epochs and bad channels, 3) unsupervised detection of brain and noise components based on the signal and noise subspace alignment index (SENSAI). In ground-truth simulations with synthetic and empirical EEG contaminated with realistic artifacts (EOG, EMG, noise), GEDAI globally outperformed leading denoising techniques based on principal component analysis (ASR) and independent component analysis (IClabel, MARA), revealing large effect sizes in challenging scenarios with simultaneous artifact mixtures, low signal-to-noise ratio (-9 dB), and high temporal contamination (up to 100%). Its superior denoising also enhanced neurobehavioral predictions, yielding highest accuracies in ERP classification and brain fingerprinting. GEDAIs autonomy, computational speed and noise-resilience could find future applications in 1) real- world medical, mobile and dry electrode EEG recordings 2) magnetoenecephalography (MEG) denoising (given the shared M/EEG forward model), and 3) real-time brain-computer interfaces (BCIs). The Matlab code for GEDAI is available as an open-source EEGLAB plugin at https://github.com/neurotuning/GEDAI-master

neuroscience↗

bioRxiv2025

Six New Species and Two Reinstatements of Viola (Violaceae) from China

We describe six new species and reinstate two species of Viola (Violaceae) from China, based on material collected in Guangxi, Jiangxi, Hubei, and Chongqing. Field surveys, detailed morphological comparisons, and phylogenetic analyses of ITS and GPI gene sequences placed these taxa within section Plagiostigma subsections Diffusae. The GPI data provided higher resolution, revealing complex relationships and suggesting ancient hybridization or incomplete lineage sorting in subsect. Diffusae. Our findings clarify species boundaries and contribute to understanding the diversity and evolution of Viola in China.

plant biology↗

bioRxiv2025

METTL3 Uncouples Chromatin Accessibility from Transcription during Retinal Development

METTL3 is a key regulator of RNA metabolism, yet its genomic and epitranscriptomic roles in tissue development are largely unexplored. Using embryonic stem cell-derived 3D retinal organoids to model retinal progenitor cell (RPC) differentiation, we integrated transcriptome-wide m6A profiling (GLORI), protein-DNA (ChIP-seq and CUT&RUN) and chromatin accessibility (ATAC-seq) mapping, and targeted m6A engineering (dCas13b-FTO) to dissect METTL3 function. Loss of METTL3 nuclear m6A activity disrupted Rx+ retinal anlage formation in vitro, with dCas13b-FTO epitranscriptome engineering revealing that m6A at the Six3 3UTR governs its stability. Surprisingly, while METTL3 loss altered histone modifications and chromatin accessibility, its direct chromatin targets showed little transcriptional correlation. A degron-based METTL3 degradation strategy, paired with protein-RNA interaction profiling, exposed rapid regulatory shifts in RPCs, revealing a METTL3-Ythdf1 protein-RNA axis. Our multi-omics approach establishes METTL3-dependent m6A as a critical epitranscriptomic layer in retinal development, unveiling a genomic paradigm in which chromatin accessibility diverges from transcriptional output.

developmental biology↗

bioRxiv2025

A Micro-Engineered Heart Tissue Model of Desmin-related Cardiomyopathy Caused by Mutant αB Crystalin

Protein quality control (PQC) is essential for maintaining sarcomere integrity in cardiomyocytes. Crystallin B chain (CRYAB) R120G mutation disrupts CRYABs chaperone activity, leading to aggregation of CRYAB and its client proteins (including Desmin), leading to Desmin-related cardiomyopathy (DRM). Prior experimental systems for modeling DRM linked to CRYAB require massive overexpression of CRYAB mutant isoforms, raising questions about translational relevance. Here, we establish the first model of CRYAB-linked DRM that uses genome-edited hiPSC together with isogenic controls, allowing us to study the impact of mutant CRYAB expressed at near endogenous levels. Within micro-engineered heart tissues (HT), CRYAB-R120G mutant hiPSC-derived cardiomyocytes recapitulated key DRM hallmarks, including Desmin and CRYAB aggregation, contractile dysfunction, and increased vulnerability to PQC pathway inhibition. CRYAB-R120G mutant HT also exhibited dysfunctional calcium-contraction coupling, which exacerbated contractile deficits at higher pacing frequencies. JAK1 inhibition with Itacitinib partially restored contractile function at higher pacing frequencies, suggesting JAK1 inhibition as a viable therapeutic strategy. By preserving human-specific structural and functional features, our {micro}HT platform enables mechanistic characterization of proteotoxic cardiomyopathies and offers a scalable system for targeted drug screening.

bioengineering↗

bioRxiv2025

NQO1 phase condensation promotes stress granule assembly to facilitate pancreatic carcinogenesis

G3BP1 promotes pancreatic ductal adenocarcinoma (PDAC) tumorigenesis driven by oncogenic KRAS mutations through liquid-liquid phase separation (LLPS)-mediated assembly of stress granules (SGs). However, the regulatory mechanisms remain elusive. We identify the antioxidant enzyme NAD(P)H quinone dehydrogenase 1 (NQO1) as a novel SG regulator that enhances SG assembly in pancreatic cancer cells independently of reactive oxygen species (ROS). Mechanistic studies reveal that NQO1 does not regulate the expression of G3BP1 or other SG-associated genes. Instead, NQO1 undergoes LLPS dependent on its RNA-binding K homology (KH) like domain. Further analysis demonstrates that residues 121-131 of NQO1 directly interact with G3BP1s RNA-binding domain (RBD), enhancing the multivalency of G3BP1 complexes to potentiate LLPS-driven SG assembly. This interaction accelerates cell proliferation, KRAS mutation-induced acinar-to-ductal metaplasia (ADM), and pancreatic carcinogenesis. Notably, the interaction between NQO1 residues 121-131 and G3BP1s RBD is essential for NQO1 phase condensation under stress condition. Integrative analysis of human PDAC transcriptomic datasets reveals a weak association between NQO1 and G3BP1 levels. Importantly, both NQO1 and G3BP1 formed biological condensates and co-localized in the lesions of human PDAC tissue sections. Our study uncovers a novel KRAS mutation-driven mechanism of pancreatic carcinogenesis through the lens of phase separation, transcending conventional gene expression regulation and offering new insights into non-canonical KRAS-NQO1-G3BP1-SG regulatory networks in PDAC initiation.

cancer biology↗