Biology subjects

Tan, L.

Publications and source records attributed to Tan, L..

At least 19 recordsLinked to original sources

bioRxiv2026

Oncolytic Newcastle disease virus promotes tumor cell apoptosis through SUMO-1-dependent PRKN-mediated degradation of c-Myc

The oncogenic transcription factor c-Myc is aberrantly upregulated in many cancers and plays a central role in tumor cell proliferation and survival. However, direct pharmacological targeting of c-Myc remains challenging because of its intrinsically disordered structure. Newcastle disease virus (NDV) is an oncolytic virus with selective antitumor activity, but the molecular mechanisms by which NDV disrupts oncogenic signaling remain incompletely understood. Here, we show that NDV induces apoptosis in multiple tumor cell lines and suppresses tumor growth in an A549 xenograft model. NDV infection markedly reduced c-Myc protein levels in tumor cells in a time- and dose-dependent manner, and c-Myc knockdown further enhanced NDV-induced intrinsic apoptosis. Mechanistically, NDV promoted ubiquitin-proteasome-dependent degradation of c-Myc through ubiquitination at Lys142. Screening of candidate E3 ubiquitin ligases identified PRKN as a key mediator of NDV-induced c-Myc degradation. NDV infection induced SUMO-1-dependent nuclear translocation of PRKN, allowing PRKN to interact with nuclear c-Myc and promote its degradation. Disruption of this pathway by SUMO-1 or PRKN knockdown impaired PRKN nuclear accumulation, restored c-Myc stability, and attenuated NDV-induced apoptosis. Functionally, expression of the degradation-resistant c-Myc K142R mutant or restoration of c-Myc in c-Myc-knockdown cells protected tumor cells from NDV-induced apoptosis. Together, these findings identify a SUMO-1-PRKN-c-Myc signaling axis that mediates NDV-induced tumor cell apoptosis and reveal c-Myc destabilization as an important mechanism underlying the oncolytic activity of NDV.

cell biology↗

bioRxiv2026

Role of Uridine Phosphorylase 1 (UPP1) in Pancreatic β-Cell Physiology and Its Dysregulation in Obesity and Type 2 Diabetes

Aims/hypothesis Fasting plasma uridine concentrations are elevated in obese mice and in individuals with type 2 diabetes (T2D), suggesting that uridine may function as a metabolic signal that adjusts insulin secretion to changes in metabolic demand. Here, we tested this hypothesis by determining whether uridine potentiates glucose-stimulated insulin secretion (GSIS) in mouse and human islets and by identifying uridine phosphorylase 1 (UPP1), the principal enzyme responsible for systemic uridine catabolism, as a key mediator of this effect. Methods Whole-body Upp1 knockout (KO) and {beta}-cell-specific Upp1 knockout (KO{beta}) mice were used to determine the requirement for UPP1 in uridine-potentiated glucose-stimulated insulin secretion (GSIS) and glucose-stimulated MAPK activation. Leptin-deficient and diet-induced obese mouse models were used to assess the impact of obesity on uridine responsiveness. UPP1 protein abundance and subcellular localization were examined in islets from obese mice and human donors spanning a range of glycemic control. Integrated single-nucleus RNA sequencing and ATAC sequencing were performed on wild-type and Upp1 KO mouse islets to characterize {beta}-cell molecular remodeling associated with UPP1 deficiency and altered responses to uridine. Results Uridine potentiated GSIS in islets from chow-fed and high-fat diet-fed mice, but not in leptin-deficient obese mice. Uridine also potentiated GSIS in human islets, although this effect progressively declined with worsening glycemic control. In mouse islets, uridine potentiation was accompanied by transient activation of the MAPK pathway. Both uridine-potentiated GSIS and uridine-enhanced glucose-stimulated MAPK activation were abolished in islets from Upp1 KO and KO{beta} mice. The requirement for a transient MAPK signaling was further supported by pharmacological inhibition of MEK and constitutive activation of MEK1. Leptin-deficient islets exhibited impaired glucose-induced MAPK activation together with increased UPP1 protein expression. In human islets, UPP1 protein abundance positively correlated with donor HbA1c and displayed heterogeneous subcellular localization. Integrated single-nucleus multiomic analysis identified UPP1 as an important regulator of {beta}-cell identity, function, and transcriptional responses to uridine. Conclusions These findings establish UPP1 as a key regulator linking pyrimidine metabolism to {beta}-cell function and identify impaired uridine responsiveness as a potential contributor to and/or consequence of {beta}-cell dysfunction during the progression of T2D.

molecular biology↗

bioRxiv2026

A brain locus for viable gestation

The expectant mother's body is systemically remodeled by her hormones to weather the physiological vicissitudes of pregnancy. In contrast to other reproductive organs, how the expectant mother's brain promotes these system-wide changes is largely unexplored. Here we show that pregnancy induces profound changes in gene expression and identity of hormone-sensitive neurons and report a previously unknown class of such cells that is required for viable pregnancy. We performed RNA sequencing from late mid-gestation mice of four estrogen receptor alpha-expressing populations from hypothalamus and amygdala that regulate reproductive behaviors altered during gestation. Each of these populations undergoes such large, specific transcriptional shifts during pregnancy that these changes even exceed their transcriptional differences between the sexes. The gene expression changes during pregnancy also imbue particular transcriptomically-defined neuronal types within these four populations with new molecular identities. Targeted ablation of one such neuronal type, POANpy2r cells, precludes implantation and abrogates viable pregnancy. Taken together, we have uncovered an essential role of hormone-sensitive neurons in the brain in sustaining pregnancy. The etiology of spontaneous gestational loss, which afflicts ~15% of human pregnancies, remains idiopathic in many cases, and our findings suggest a brain-based mechanism that contributes to such events. More broadly, we provide a molecular and cellular foundation to study gestational processes in health and disease from the perspective of the pregnant brain-body axis.

neuroscience↗

bioRxiv2026

PlantRegMoD: An integrative and AI-driven multi-omics database for plant regeneration research

Plant regeneration underpins plant developmental plasticity, tissue culture, genetic transformation and crop improvement. Although numerous related omics datasets have been generated, specialized multi-omics databases for this field are still scarce. Here, we constructed PlantRegMoD, an AI-powered integrated database dedicated to plant regeneration. This platform hosts 20.54 TB standardized multi-omics data from 147 projects across 32 plant species and 2,593 samples. We established a unified hierarchical classification system covering five major categories and nine regeneration models, and curated 236 regeneration genes as well as their 28,190 homologs across 58 representative plant species. It contains extensive transcriptomic resources across all regeneration models, together with 196,423 single cells and over 8.81 million epigenetic peaks to dissect cellular heterogeneity and multi-layered epigenetic regulation. Equipped with nine online omics-related tools and a RAG-based intelligent Q&A system, PlantRegMoD greatly reduces bioinformatic barriers and serves as a robust resource for mechanistic, functional and evolutionary studies of plant regeneration.

plant biology↗

bioRxiv2026

Benchmarking CUT&RUN analysis using motif enrichment

Background. Cleavage under targets and release using nuclease (CUT&RUN) maps the genome-wide locations of chromatin-associated proteins and provides an improved alternative to chromatin immunoprecipitation sequencing (ChIP-seq) for profiling sequence-specific transcription factor binding sites. Identifying these binding sites plays a critical role in understanding gene regulation, and transcription factors provide a useful setting for benchmarking because their well-defined sequence motifs serve as built-in controls for evaluating performance. Compared with ChIP-seq, CUT&RUN achieves higher resolution and lower background by avoiding cross-linking and bulk precipitation. Its distinct fragment length and cleavage characteristics, however, limit the direct transfer of existing computational tools, which primarily target ChIP-seq data. The performance of these tools on CUT&RUN can depend strongly on preprocessing choices. In this work, we investigate preprocessing strategies for transcription factor CUT&RUN, focusing on fragment length filtering and spike-in calibration. We aim to improve peak detection and provide practical guidance for analysis. Results. We designed a benchmarking method to evaluate peak-calling procedures for CUT&RUN data and the effects of preprocessing approaches, including fragment length filtering and spike-in calibration. We benchmarked the two most widely used peak callers, MACS2 and SEACR, by assessing motif enrichment---the degree to which identified peaks contain the expected transcription factor binding motifs. Filtering for fragments with a length [≤]120 bp generally improved target motif enrichment. Spike-in calibration using heterologous Saccharomyces cerevisiae DNA improved motif elucidation substantially for MACS2, with little benefit for SEACR. By contrast, using Escherichia coli DNA as a spike-in control often failed to produce valid results unless we could meticulously control E. coli contamination. MACS2 performed robustly across samples. SEACR performed especially well on clean, sparse-background datasets, but performed poorly on some datasets with denser background signal and often produced numerous apparent false positives. While MACS2 provided robust results under minor perturbations in fragment length filtering, SEACR exhibited greater sensitivity to such changes. Discussion. Our benchmarking highlights how both peak caller choice and preprocessing strategy shape the analysis of transcription factor CUT&RUN data. By comparing the robustness and limitations of two widely used peak callers, we provide practical guidance on fragment length filtering, spike-in calibration, and tool selection. These findings help improve the processing and interpretation of CUT&RUN data, allowing researchers to more rapidly and reliably utilize this new technology. We expect that our work will guide more informed choices in CUT&RUN analysis and support the development of improved computational methodologies.

bioinformatics↗

bioRxiv2026

Integrating Narrow-Window DIA with AI-Powered Search Enables Deep and Reliable Functional Proteomics

Narrow-window data-independent acquisition (nDIA) is emerging as a powerful technique for bottom-up proteomics. Here, we systematically benchmarked nDIA, wide-window DIA (wDIA), narrow-window data-dependent acquisition (nDDA), and wide-window DDA (wDDA) for rapid, single-shot proteomic analysis. For data processing, we introduced Tesorai Search, a new search engine leveraging a large pre-trained model and compared it with DIA-NN and FragPipe across both DIA and DDA datasets. Among 12 acquisition-analysis pipelines evaluated, nDIA combined with DIA-NN and Tesorai Search delivered the highest proteome coverage, identifying 10,255 and 10,766 protein groups from benchmark samples, respectively. Both search engines maintained rigorous false-discovery rate (FDR) control. While nDIA generally outperformed nDDA in sensitivity, FragPipe-DDA+ approach proved to be the most sensitive within the nDDA pipelines. However, entrapment analyses indicate that this sensitivity comes at the cost of less robust FDR control compared to Tesorai Search. As a proof of concept, we applied nDIA-MS to 17 cancer cell lines harboring DNA damage response (DDR) gene knockouts, successfully detecting significant downregulation of all targeted proteins and uncovering 81 DDR-related proteins modulated in at least one cell line. These results underscore nDIA-MS, together with DIA-NN and Tesorai Search, as a robust and scalable platform for high-throughput functional proteomic screening.

systems biology↗

bioRxiv2026

Deep phenotyping and multi-omics analyses reveal systems-wide metabolic dysregulation in a refined trisomy mouse model of Down syndrome

Trisomy 21 or Down syndrome (DS) affects multi-organ systems across the lifespan. The presence of an extra chromosome, along with genome dosage imbalance due to triplicated genes, contributes to the DS phenotypes. Of the DS mouse models, few are aneuploid with a freely segregating extra chromosome. We previously showed that the aneuploid Ts65Dn mice exhibit metabolic deficits consistent with the metabolic profile of DS. However, the genotype-phenotype relationships in Ts65Dn mice are complicated by the presence of triplicated genes unrelated to human chromosome 21 (Hsa21). To address this issue, we leveraged a refined model, Ts66Yah, where the extra triplicated genes in Ts65Dn have been removed. Deep phenotyping and multi-omics analyses showed that Ts66Yah mice develop pronounced and widespread metabolic disturbances. Despite sexual dimorphism in weight gain, body temperature, lipid and lipoprotein profiles, hepatic injury and adipose fibrosis, both male and female Ts66Yah mice share a common phenotype of pronounced glucose intolerance and insulin resistance, reduced mitochondrial respiratory capacity in visceral fat, altered serum inflammatory cytokine profile, and dysregulated serum and liver metabolomes. Pan-tissue transcriptomes also reveal signatures of immune activation, disrupted metabolic processes and cellular respiration, altered cytokine signaling, enhanced oxidative stress, and extracellular matrix remodeling. These combined changes across tissues disrupt metabolic homeostasis more severely in Ts66Yah than in Ts65Dn mice. Several phenotypes, including glucose intolerance, insulin resistance, tissue fibrosis, and oxidative stress were further exacerbated by an obesogenic diet. This foundational data establishes Ts66Yah as a valuable reference model for the mechanistic and comparative study of metabolic dysfunction in DS.

physiology↗

bioRxiv2026

Coronavirus papain-like protease antagonizes innate immunity by cleaving Importin α1 to disrupt nuclear transport

The Importin family, as key mediators of nucleocytoplasmic transport, represents a common target for viral immune evasion. However, whether coronaviruses directly manipulate Importin to disrupt nuclear trafficking and suppress antiviral immunity has remained unclear. In this study, we identify a previously unrecognized mechanism by which coronaviruses from all four genera subvert host innate immunity through the proteolytic inactivation of Importin 1, a key mediator of nuclear import of cargo proteins. We demonstrate that the membrane-associated papain-like protease (PLpro-TM) directly cleaves Importin 1 at specific glycine residues, G129 for PEDV and IBV PLpro, and G119 for MHV and PDCoV PLpro, thereby disrupting its nuclear import function. This cleavage impairs the nuclear translocation of multiple transcription factors (IRF3, STAT1, STAT2, and p65) and suppresses the expression of downstream antiviral genes, including IFN-{beta} and IFN-stimulated genes (ISGs). Importantly, cleavage-resistant mutants of Importin 1 (G129A or G119A) restore nuclear import capability and rescue IFN-{beta} signaling. Consequently, they exert a more potent inhibitory effect on viral replication than the wild-type Importin 1, fulfilling an antiviral role. Our work establishes PLpro-TM-mediated cleavage of Importin 1 as a conserved immune evasion strategy across coronaviruses and highlights this interaction as a potential target for broad-spectrum antiviral intervention. Author summaryThe nuclear transport of transcription factors is a critical checkpoint for the initiation of innate antiviral immunity. Here, we identify the PLpro-TM protein as a pan-coronavirus antagonist of nucleocytoplasmic trafficking and innate immune response. We demonstrate that PLpro-TM from four distinct coronavirus genera directly cleaves Importin 1 at specific glycine residues, thereby disabling its ability to mediate the nuclear import of key transcription factors and subsequent transcription of anti-viral genes. This work reveals a previously unrecognized, evolutionarily conserved immune evasion strategy shared across , {beta}, {gamma}, and {delta} coronaviruses. By uncovering the proteolytic inactivation of Importin 1 targeting by PLpro-TM, our findings not only resolve a long-standing question about how coronaviruses disrupt nuclear trafficking, but also establish PLpro-TM and Importin 1 as promising targets for the development of broad-spectrum antiviral therapeutics against current and emerging coronaviruses.

microbiology↗

bioRxiv2026

Chromatin topology control by a muscle-specific ribosomal protein

Three-dimensional genome organization stabilizes cell-type-specific gene expression, yet the tissue-restricted factors that maintain chromatin insulation remain poorly understood. Here, we identify the muscle-specific ribosomal protein Rpl3l as an unexpected nuclear regulator of genome architecture in atrial cardiomyocytes. Rpl3l is enriched in the nucleus and nucleolus, where it binds its own genomic locus and stabilizes a CTCF-anchored chromatin boundary that represses the T-type calcium channel gene Cacna1h. Loss of Rpl3l weakens local chromatin insulation, increases long-range contacts across the Rpl3l-Cacna1h locus, derepresses Cacna1h, and increases susceptibility to atrial fibrillation (AF), which is suppressed by pharmacological inhibition of T-type calcium channels. Furthermore, AF-associated RPL3L variants exhibit impaired nucleolar localization, reduced rRNA binding, and defective repression of CACNA1H in human iPSC-derived atrial cardiomyocytes. Together, these findings reveal a ribosomal protein-chromatin axis linking genome insulation to ion-channel dosage control and cardiac rhythm stability, expanding the repertoire of cell-type-specific genome architecture regulators. HighlightsThe muscle-specific ribosomal protein Rpl3l exhibits unexpected nuclear and nucleolar enrichment. Rpl3l stabilizes a CTCF-anchored chromatin boundary to maintain atrial-specific local genome insulation. Loss of Rpl3l derepresses Cacna1h which encodes a T-type Ca2+ channel and increases atrial fibrillation susceptibility. Atrial fibrillation-associated RPL3L variants impair nucleolar targeting and CACNA1H repression

physiology↗

bioRxiv2026

Longitudinal analysis reveals myeloid cell contributions to neuroPASC pathogenesis

Neurological and neuropsychiatric symptoms, collectively termed neuroPASC, are among the most prevalent Post-Acute Sequelae of COVID-19 (PASC). Neuroinflammation - particularly microglia reactivity - has been implicated in neuroPASC. Current insights are largely derived from post-mortem tissues of acutely infected patients and may not reflect PASC-related neuropathology. We previously established a PASC model in which SARS-CoV-2-infected mice developed persistent behavioral alterations and prolonged neuroinflammation for up to 120 days post-infection (dpi) in the absence of viral neuroinvasion. Here, we extended these results to a longitudinal single-cell RNA sequencing analysis of brain immune cells collected at 0, 6, 30, and 100 dpi. We identified a coordinated contribution of infiltrating and resident myeloid cells to the initiation and persistence of neuroinflammation. In specific, microglia displayed sustained expansion of subclusters characterized by inflammatory, stress response, and metabolic signatures. Border-associated macrophages upregulated monocyte attractants during acute infection. Concurrently, monocytes and neutrophils showed marked brain recruitment and mounted transient inflammatory responses at 6 dpi, potentially triggering long-term microglial reactivity. Together, these findings provide a high-resolution atlas of brain myeloid immune dynamics during neuroPASC and highlight a central role for microglia in sustaining chronic neuroinflammation.

immunology↗

bioRxiv2026

Whole-genome 3D architectural screen reveals modulators of brain DNA structure

Three-dimensional (3D) genome architecture is the foundation of gene regulation, and plays a critical role in normal physiology and disease. However, our understanding of its biochemical determinants has long been limited by technology: imaging-based screens only profile a small number of loci, while sequencing-based studies rarely exceed 100 samples or conditions. Here we present "in-plate chromosome conformation capture" (Plate-C), a high-throughput, cost-effective platform that profiles thousands of whole-genome architectures in a day. Plate-C enabled the first chemical screen for whole-genome structural changes--profiling 2,956 samples from 834 conditions across 5 neuronal and glial types, accompanied by 6,081 single cells using "easy diploid chromosome conformation capture" (Easy Dip-C) and 200,893 single-cell transcriptomes. We discovered that diverse, dose/time-dependent, and cell type/species-specific modes of DNA structural changes can be rapidly induced by manipulating epigenetic (HDAC, BET), metabolic (mTOR), proteostatic (UPR), developmental (GSK3/Wnt, Hedgehog), immune (cGAS/STING), and neurotransmission pathways. To validate our finding in vivo, we demonstrated in newborn mice that HDAC inhibition drives brain-wide genome rewiring within hours, highly correlated with changes in vitro and inducing a latent structural and transcriptional state orthogonal to normal differentiation. By enabling massively parallel profiling of whole-genome structures, Plate-C paves the way for systematic discovery of DNA folding principles to better understand and engineer the human genome in 3D.

genomics↗

bioRxiv2026

Genome expansions and regulatory contact entanglement help preserve ancestral metazoan synteny

Chromosomes constitute deeply conserved evolutionary units in many metazoan genomes, with chromosomal fusions and fissions, accompanied by sub-chromosomal rearrangements, rewiring three dimensional genome architecture. How chromatin loops and compartments that define distal regulatory interactions within chromosomes impose functional constraints that affect this long-term evolutionary process (and vice-versa) is an emerging research topic. Genome expansions, especially through transposable element (TE) activity, test these constraints by increasing the genomic distances over which regulatory interactions must function and were thus suggested to be the drivers of chromosomal rearrangements. To study dynamics and stability of such distal interactions in the light of genome expansions, we focus on the cnidarian Hydra vulgaris, which, based on its simple and well-understood biology as well as one of the largest genomes among cnidarians, is particularly suited to test how chromatin loop and genome architecture respond to genome expansion. We investigate genome architecture using Micro-C, single-cell Hi-C (Dip-C), and DNA FISH, and perform comparative analysis using available genomic and epigenomic data. Contrary to prior expectations, our analysis of whole-genome data and particular loci (e.g., Wnt) suggests a scenario where genome expansion did not only result in chromatin loops often reaching several megabases in hydra, but also led to regulatory contact mixing and entanglement, introducing additional constraints to maintain ancestral genomic architecture. Generalizing these findings across hundreds of metazoan genomes, we show a new mechanistic role for genome expansion in yielding entangled long-range regulatory configurations that, in turn, decelerate chromosomal rearrangements, thus maintaining (and not breaking) ancestral regulatory states and synteny. Significance statementSome of the largest and most repetitive animal genomes retain a surprisingly high level of deeply conserved metazoan synteny. As repetitive regions are often associated with chromosomal rearrangements, it has been enigmatic why syntenic retention is so frequently observed. In this study, using hydra as a model system for both stem cell biology and an evolutionary history rich in transposable-element driven genome expansion, a multi-level conformational landscape dissection reveals a multitude of long-range regulatory states. We show that mixing of multiple such regulatory links accompanied by genomic expansion is associated with maintained ancestral synteny, thus pointing to the counter-intuitive role of genome expansions as "fossilization" agents across metazoan genomes.

evolutionary biology↗

bioRxiv2026

nELAVL phosphorylation by CDKL5 regulates inter-condensates composition and communication to promote experience-dependent maturation of the visual cortex

Mutations in Cyclin-Dependent Kinase-Like 5 (CDKL5) cause CDKL5 deficiency disorder (CDD), an X-linked neurodevelopmental condition. Through a phosphoproteomic screen, we identified the neuron-specific nELAVL family of RNA-binding proteins as direct activity-dependent substrates of CDKL5. In support of this regulatory axis, single-nuclei transcriptomics of Cdkl5 knockout (KO) cortices revealed an enriched reduction in activity-dependent mRNAs. Mechanistically, we show that nELAVL proteins undergo phase separation to form biomolecular condensates, the size of which is gated by CDKL5 phosphorylation. Loss of CDKL5 leads to enlarged nELAVL condensates, which exhibit reduced binding affinity for the target mRNA Fos, resulting in its accelerated degradation. This disruption extends to inter-condensate communication: phosphodeficient nELAVL show diminished interaction with P-bodies, which themselves become enlarged in CDD mutant iNeurons. Functionally, the absence of nELAVL phosphorylation recapitulates the deficits in experience-dependent visual function observed in Cdkl5 KO mice. Our findings establish a critical molecular mechanism by which CDKL5-mediated phosphorylation governs mRNA metabolism by tuning the properties of nELAVL condensates and their communication with other biomolecular condensates, ultimately promoting experience-dependent maturation of the visual cortex.

neuroscience↗

bioRxiv2026

Fiber optical parametric amplification of low-photon-flux microscopy signals

The performance of a laser scanning microscope inevitably depends on the performance of the point detector. As laser scanning approaches aim to penetrate deeper in tissue, there is a commensurate need for detectors that can operate with high sensitivity, bandwidth, and dynamic range at near-infrared wavelengths where scattering is reduced. Here, we demonstrate that fiber optical parametric amplification can be used to boost low-power microscopy signals to levels that can be detected by near-infrared photodiodes without introducing prohibitive noise. We construct amplifiers that achieve >50 dB of parametric gain at wavelengths within the third near-infrared transparency window and have similar sensitivity to near-infrared photomultiplier tubes. Furthermore, these amplifiers outperform detection with a photodiode and subsequent electrical amplification, providing a factor of 10-100-fold improvement in sensitivity. We demonstrate amplifier bandwidths up to ~1.6 GHz, a factor of 10 faster than conventional detectors, including near-infrared photo-multiplier tubes, with sensitivity of ~8 nW (corresponding to ~20 photons/pixel). Finally, the increased performance of the optical amplifier is confirmed in diagnostic imaging experiments where >10x less power is required to achieve the same signal-to-noise ratio and contrast as images using electrical amplification. Accordingly, fiber optical parametric amplification is a new path forward for extending the performance of laser scanning microscopes in the near infrared.

biophysics↗

bioRxiv2026

A Genetic Tool for Specific Tracking of Mature Neutrophils

Tracking mature neutrophils remains challenging due to the lack of reliable cell surface markers. Although CD101 is a promising candidate for mature neutrophils, its stability under pathological conditions is unclear. Using a CD101-tdTomato reporter mouse model, we confirmed that the reporting system does not alter CD101 expression, and tdTomato fluorescence is predominantly expressed in mature neutrophils across peripheral tissues. Further analysis revealed that CD101+ and tdTomato+ neutrophils display identical characteristics of mature neutrophil, including poly-segmented nuclei, cell size, and key functions under homeostasis. By comparing tdTomato fluorescence with CD101 protein levels, we demonstrate that reduced CD101 expression under pathological states was not attributed to shedding or degradation. Our finding enhances CD101 as a robust and reliable marker of neutrophil maturity, providing a foundation for future applications in spatial transcriptomics and lineage tracing studies to dissect neutrophil heterogeneity and function. Highlights of the studyO_LIIn CD101-tdTomato homozygous mice, tdTomato is predominantly expressed in neutrophils and labels nearly 100% of mature neutrophils, aligning with the phenotype of CD101+ mature neutrophils; C_LIO_LIThe CD101-tdTomato reporting system does not interrupt CD101 expression or neutrophil functions; C_LIO_LICD101 remains a stable and reliable cell surface marker for labeling mature neutrophils, even under pathological conditions. C_LI

immunology↗

bioRxiv2026

EasyPseudogene: an easy-to-use and multithreaded pipeline for pseudogene detection

Pseudogenes are recognized as essential components for reconstructing adaptive evolutionary trajectories and understanding genomic remodeling. However, identifying these sequences in large eukaryotic genomes remains technically challenging due to fragmented workflows, complex manual configurations, and the lack of high-performance, parallelized tools capable of processing rapidly growing data volumes. We present EasyPseudogene, an automated and multithreaded pipeline designed for the end-to-end identification of pseudogenes across diverse eukaryotic lineages. Unlike traditional self-mapping tools that often fail to detect unitary pseudogenes when functional counterparts are absent, EasyPseudogene introduces an inter-species reference-driven paradigm that utilizes high-quality proteomes as probes to scan target genomes for evolutionary relics. The pipeline employs a modular "hierarchical screening and precision detection" architecture, integrating high-speed homology searching via MMseqs2 and spliced alignments via miniprot with high-fidelity, three-frame alignments using GeneWise. Performance benchmarking on cetacean genomes demonstrates that EasyPseudogene can replicate known gene loss events, such as the functional decay of the ADRB3 gene, with 100% consistency relative to established manual workflows. By encapsulating complex comparative genomics logic into a standardized framework with interactive HTML visualization for mutation auditing at single-base resolution, EasyPseudogene provides a versatile and reproducible solution for marine ecology and evolutionary research.

bioinformatics↗

bioRxiv2026

Aberrant oxidative metabolism selects for TET2-deficient hematopoietic stem and progenitor cells

The mechanism(s) driving selective expansion of mutant hematopoietic stem and progenitor cells (HSPC) in clonal hematopoiesis (CH) are incompletely understood. Here, we address the role of metabolism in selection for HSPC with loss of function mutations in TET2. Loss of Tet2 in murine HSPC triggers overexpression of glycolysis and oxidative phosphorylation genes and increased oxidative metabolism via an enlarged mitochondrial network. However, Tet2-deficient HSPC maintain a normal redox state. Strikingly, compound loss of the rate-limiting pentose phosphate pathway (PPP) enzyme glucose-6-phosphate dehydrogenase (G6PD) triggers increased reactive oxygen species and impairs the fitness of Tet2-deficient HSPC. We find that aberrant oxidative metabolism is also a feature of HSPC in human CH and clonal cytopenia of unknown significance (CCUS). Overall, our data point to aberrant metabolism as a critical and conserved driver of selection in TET2-deficient CH and identify the PPP as a crucial compensatory pathway needed to maintain their selective advantage. Statement of SignificanceThis study identifies oxidative metabolism as a critical driver of selection for TET2-deficient HSPC in clonal hematopoiesis (CH). It also demonstrates that cellular redox state is a vulnerability that impairs their fitness. These insights establish targetable metabolic pathway(s) that could be exploited in the setting of TET2 mutant CH.

cancer biology↗

bioRxiv2026

Transcriptomic profiling of human γδ T cells reveals non-linear immune aging characterized by childhood transitions and relative stability in adulthood

{gamma}{delta} T cells are one of the first T cell subsets developing in early ontogeny and show various effector functions in immune homeostasis and response in the young and the old. However, their maturation trajectories from infancy to children, adults and elderly have not been systematically defined. Here, we generated a single-cell transcriptome atlas of 106,711 {gamma}{delta} T cells from 223 individuals spanning infancy to old age. Our analysis reveals that {gamma}{delta} T cell aging is non-linear, characterized by pronounced childhood transitions followed by relative stability throughout adulthood despite marked inter-individual variability. In childhood, changes from developmental and mitochondrial programs toward cytotoxicity and inflammaging were evident. This includes maturation trajectories from GZMK intermediates to GZMB+Perforin+ effectors at both RNA and protein levels. Taken together, our study delineates the aging trajectories of human {gamma}{delta} T cells, establishes {gamma}{delta} T cells as a cellular paradigm of non-linear immune aging, and provides a comprehensive resource for investigating {gamma}{delta} T cell biology across the human lifespan.

immunology↗