Biology subjects

Song, S.

Publications and source records attributed to Song, S..

At least 55 records · Page 3Linked to original sources

bioRxiv2025

Atypical activation and molecular glue-like dimerization mechanism of an intrinsically-biased chemokine receptor

CXCR7, also known as atypical chemokine receptor 3 (ACKR3), is a naturally-biased, {beta}-arrestin-coupled seven transmembrane receptor (7TMR) that lacks productive coupling with heterotrimeric G-proteins. Despite a critical involvement in cancer metastasis, cardiovascular pathophysiology, and inflammatory disorders, the molecular basis of non-canonical activation and functional divergence of CXCR7 remains elusive. Here, we present a complete landscape of CXCR7 activation using a series of cryo-EM structures, and discover an atypical activation mechanism that is distinct from prototypical GPCRs. CXCR7 is maintained in a basal conformation by a unique tripartite ionic-lock involving TM5-TM6, in contrast to a broadly conserved TM3-TM6 ionic-lock in GPCRs, which is disrupted upon receptor activation. Importantly, activation of CXCR7 results in a constricted pocket and distinct surface topology on the intracellular side compared to prototypical GPCRs. Serendipitously, we capture novel dimeric arrangements of CXCR7 with an inter-protomer stitching by a native phospholipid serving as a molecular glue, and identify previously unanticipated extrahelical allosteric sites on the receptor. Surprisingly, in an intermediate state structure of CXCR7, the second extracellular loop (ECL2) displays a self-blocking conformation, in stark contrast to ECL2-mediated self-activating mechanism reported recently for some orphan GPCRs. Finally, we unequivocally establish CXCR7 as an atypical opioid receptor via a large peptide library screening and structure elucidation in complex with distinct opioid peptides imparting full receptor activation. In summary, our study elucidates an atypical mechanism of CXCR7 activation, and establishes it as an alternative, non-canonical opioid receptor target with potential for novel pain therapeutics.

biochemistry↗

bioRxiv2025

Transcriptomic and protein analysis of human cortex reveals genes and pathways linked to NPTX2 disruption in Alzheimer's disease

BackgroundCognitive resilience to Alzheimers disease (AD) pathology is associated with preserved expression of NPTX2, an activity-regulated synaptic protein involved in circuit plasticity, excitation-inhibition balance, and complement-linked synapse regulation. However, the broader molecular programs coordinated with NPTX2 in resilient individuals remain unclear. MethodsWe analyzed postmortem middle temporal gyrus tissue using targeted PRM-MS proteomics in 135 individuals and bulk RNA-seq in an expanded 575-sample cohort. NPTX2-associated molecular coordination was assessed within cognitively normal low-pathology controls (CN-Lo), cognitively normal high-pathology controls (CN-Hi), mild cognitive impairment (MCI), and AD. Correlation-based approaches were applied using NPTX2 protein and NPTX2 mRNA expression as anchors to define resilience mechanisms in CN-Hi subjects. ResultsNPTX2 protein abundance was preserved across all controls regardless of age and pathology but reduced in MCI and AD. NPTX2 mRNA expression was also invariant across pathology within controls and reduced in MCI and AD but decreased markedly with age. Targeted proteomics identified NPTX2 relationships with synaptic and inhibitory-circuit proteins that were preserved across control groups, alongside CN-Hi-specific recruitment of trafficking, lysosomal, metabolic, and proteostasis-associated proteins. Transcriptome-wide correlations with NPTX2 revealed differences in gene co-expression between groups, identifying a prominent activity-dependent program including BDNF, VGF, SCG2, SST, SERTM1, DUSP4, and EGR4, that was preserved in both CN-Lo and CN-Hi subjects, while genes recruited to the NPTX2 network specifically in CN-Hi implicated immune, neuroprotective, translation, and proteostasis-related pathways. Coupling differential gene expression analysis with co-expression, we further identified five candidate resilience genes whose expression and NPTX correlation was preserved across controls, but lost in MCI and AD: SST, MAL2, TAC1, SERTM1, and RFK. Expression of genes in distinct NPTX2 co-expression classes can be freely explored in our bulk RNA-seq data and other public AD transcriptomic datasets at NeMO Analytics. ConclusionFindings suggest that cognitive resilience in the context of AD neuropathology engages a coordinated molecular state distinct from both persevered cognition without pathology and MCI/AD, which is organized around preserved and selectively remodeled NPTX2-associations. Rather than reflecting broad transcript abundance changes, resilience was characterized by maintained synaptic and inhibitory programs, and adaptive proteostasis and trafficking pathways that distinguish resilient high-pathology individuals from low-pathology controls or symptomatic AD.

bioinformatics↗

bioRxiv2025

Transcriptomic profiling of the middle temporal gyrus reveals differential glial/neuronal dysregulation across Alzheimer disease and aging.

Alzheimers disease (AD), the most common cause of dementia, is characterized by amyloid-{beta} plaques, neurofibrillary tangles, and widespread neuronal dysfunction. Aging, the strongest risk factor for AD, is also associated with some overlapping processes, such as neuronal cell transcriptional downregulation and glial cell activation. The middle temporal gyrus (MTG) is a brain region that supports semantic processing and default-mode connectivity and shows early vulnerability in both aging and AD. Here we profile bulk RNA-seq from 606 postmortem MTG samples with the goal of understanding the transcriptional changes associated with AD and aging. In 217 clinical and neuropathologically confirmed AD versus 290 no-dementia controls donors, we identify 613 differentially expressed genes (390 up, 223 down; |log2 fold change| [≥] 0.5; BH P < 0.05), with NPNT and ADAMTS2 among the top upregulated signals. Cell set enrichment indicates reduced excitatory neuronal signatures together with increased microglial, astrocytic, endothelial, and pericyte programs. Gene-set analyses reveal strong activation of angiogenesis, extracellular-matrix organization, wound response, adaptive immunity, and coordinated suppression of neuronal and mitochondrial processes, including synaptic signaling and respiratory-chain complexes. Multiscale coexpression mapping resolves three disease clusters: a neuron-mitochondrial module suppressed in AD (M5; hub PJA2; key driver GABRB3), a microglial immune module upregulated in AD (M6; hub C1QC; key driver FCER1G), and an increased astrocyte-vascular extracellular-matrix module in AD (M8; hub ESAM; key driver TAGLN). Across 324 non-AD controls aged 24-108 years, aging is associated with declines in gene expression associated with translation, proteostasis, and mitochondrial function and increases in those linked to oligodendrocyte and myelination programs (for example M4; hub CNTN2; key driver MOBP); in a 65+ subset, neuronal and protein-folding modules show the strongest decrements with reduced glial gene expression upregulatio. Our results indicate that late-life aging involves increased glial responses and neuronal/proteostasis suppression, whereas AD is also associated with immune- vascular-ECM activation and suppression of neuronal programs.

genomics↗

bioRxiv2025

Continuous Production of Recombinant Adeno-Associated Virus in the Insect Cell/Baculovirus Expression Vector System

Continuous production processes may offer significant advantages for biotherapeutic manufacturing, including increased productivity, consistent product quality, reduced facility footprint, and decreased process turnaround time. Despite these benefits, the in-situ formation of defective recombinant baculovirus expression vectors (BEVs) hinders the continuous manufacturing of recombinant adeno-associated viruses (rAAV) in the baculovirus expression vector system. This study investigates an approach of reducing defective viruses through the infusion of standard recombinant baculovirus (rBV) and the compartmentalization of early- and late-stage infected cells, resulting in stable rAAV production. In this study, rAAVs were continuously produced in a series of cascading reactors, comprising a feeder reactor, an infection reactor, and a production reactor. Residence times and transfer rates across the three reactors were optimized based on the production kinetics of rBV and rAAV derived from our mechanistic model. The majority of rBV was produced within the production reactor, thereby reducing the impact of defective viruses in the infection reactor, enabling continuous rAAV production. This study demonstrates the successful implementation of a continuous rAAV production process, yielding over 5x1010 vg/mL per day for 4 weeks. This work represents the first reported continuous rAAV production process utilizing the Sf9/BEVS platform and establishes engineering know-how for overcoming manufacturing challenges associated with rAAV-based gene therapies.

bioengineering↗

bioRxiv2025

Metadomain and metaloop genome interactions in mammalian T cells

Recent studies have advanced our understanding of chromosomal organization and its principal role in gene regulation. However, most analyses have focused on short-range interactions (<2 Mb), limiting insight into broader regulatory architecture. In particular, the relationships between topologically associating domains (TADs), sub-TAD loops, long-range cross-TAD interactions, and higher-order chromosomal compartmentalization remain poorly understood. Here, we identify extensive multi-megabase and interchromosomal interactions (metaloops) in T lymphocytes, which organize into larger meta-TAD associations (metadomains). Metaloops bridge distal promoters and regulatory elements of key T cell-specific genes such as Ctla4, Ikzf2, Il2ra, Ets1, Lef1, Runx1, Bach2, Foxo1 and others, and are both shared and cell type-specific across functionally distinct T cell lineages. Reanalysis of published data confirms the reproducibility of these interactions in both mouse and human T cells and their dependence on superenhancers. Genome-wide clustering of metadomains reveals three interchromosomal hubs with distinct epigenomic profiles, including a superenhancer-enriched hub associated with T cell-specific gene activation. By integrating a compendium of new and public T cell epigenomic data, we infer distinct architectural factors associated with short-range loops and long-range metaloops. Altogether, our study reveals new features of T cell-specific 3D genome organization across scales, and our computational framework is broadly applicable to analyses of chromatin architecture across different cell types and experimental systems. Highlights- Ultra-long-range (>2 Mb) chromatin interactions linked to gene regulation in T cells - A new algorithm identifies distal and interchromosomal meta-TADs and metaloops - An interchromosomal T cell-specific active hub emerges from metadomain clustering - Epigenomic compendium implicates TFs associated with long-range interactions

genomics↗

bioRxiv2025

Query Augmented Generation (QAG) from the Genomic DataCommons for Accurate Variant Statistics

In precision oncology, researchers often use public knowledgebases to check somatic variant frequencies against their cohort data. Large language models (LLMs) can quickly answer questions on somatic variant frequencies, but often hallucinate and give inaccurate results for factual data. Using synthetic queries, we show that somatic variant frequencies in baseline LLM responses are underestimated compared to the Genomic Data Commons (GDC), the worlds largest data commons for cancer research. We present a modular architecture called Query Augmented Generation (QAG) for integrating LLMs with high-quality data from a third party data source such as a data commons, knowledgebase or database. We apply QAG to the GDC to help researchers obtain accurate frequencies for somatic variants, copy number variants, and MSI status--even for complex queries requiring multiple steps in the GDC portal and API. Our software is deployed as a model context protocol (MCP) server on Hugging Face and available on GitHub.

bioinformatics↗

bioRxiv2025

Influ-BERT: An Interpretable Model for Enhancing Low-Frequency Influenza A virus Subtype Recognition

Influenza A Virus (IAV) poses a persistent threat to global public health due to its broad host adaptability, frequent anti-genic variation, and potential for cross-species transmission. Accurate identification of IAV subtypes is essential for effective epidemic surveillance and precise disease control. Here, we present Influ-BERT, a domain-adaptive pretrained model based on the Transformer architecture. Optimized from DNABERT-2, Influ-BERT was developed using a dedicated corpus of approximately 900,000 influenza genome sequences. We constructed a custom Byte Pair Encoding (BPE) tokenizer, and employed a two-stage training strategy involving domain-adaptive pretraining followed by task-specific fine-tuning. This approach significantly enhanced identification performance for IAV subtypes. Experimental results demonstrate that Influ-BERT outperforms both traditional machine learning approaches and general genomic language models, such as DNABERT-2 and MegaDNA, in the task of IAV subtype identification. The model achieved F1-scores consistently above 97% and exhibited stable performance gains for subtypes that are underrepresented in sequencing data, including H5N8, H5N1, H7N9, and H9N2. Beyond subtype identification, Influ-BERT was successfully applied to additional tasks including respiratory virus identification, IAV pathogenicity prediction, and identification of IAV genomic fragments and functional genes, demonstrating robust performance throughout. Further interpretability analysis using sliding window perturbation confirmed that the model focuses on biologically significant genomic regions, providing insight into its improved predictive capability. Contactsongshh@big.ac.cn (Song S), atrv@lncc.br (Ana Tereza Ribeiro Vasconcelos)

bioinformatics↗

bioRxiv2025

MTARC1 Regulates Lipid Droplet Degradation via Phospholipid Remodeling in Metabolic Fatty Liver Disease

Metabolic dysfunction-associated steatotic liver disease (MASLD) is a spectrum of liver disease, including simple fatty liver, hepatic steatosis, fibrosis, cirrhosis, and hepatocellular carcinoma, with limited treatment options. MTARC1 p.A165T variant is associated with reduced risk of MASLD. We previously showed that this variant promotes MTARC1 protein degradation, suggesting MTARC1 inactivation may represent a potential therapeutic strategy. Here, we report that global or liver specific Mtarc1 knockout markedly suppresses diet-induced hepatic TG accumulation, liver injury, inflammation, and fibrosis in a manner dependently on lipolysis and lipophagy. MTARC1 inactivation post-transcriptionally upregulates PEMT and CEPT1 expression, consequently elevating GPL levels, including PC, PE, and lyso-PE, three major GPLs of hepatic LDs. Furthermore, we observed that MTARC1-deficient hepatocytes exhibited smaller but more numerous LDs compared to controls, under comparable cellular TG levels during lipolysis/lipophagy inhibition. Importantly, inhibition of CEPT1 or PEMT could attenuated the hepatoprotective effect of MTARC1 deficiency. Mechanistically, subcellular GPL remodeling induced by MTARC1 deficiency reduce LD size while increase their surface-to-volume ratio, which in turn promote TG degradation through lipolysis and lipophagy. Collectively, our findings identify MTARC1-GPL biosynthesis-LD degradation axis as a key regulator of fatty liver disease and highlight MTARC1 inhibition as a promising therapeutic strategy for MASLD.

cell biology↗

bioRxiv2025

Epitranscriptome-wide profiling identifies RNA editing events regulated by ADAR1 that are associated with DNA repair mechanisms in human TK6 cells

Adenosine-to-inosine (A-to-I) editing is an endogenous RNA modification in eukaryotes, catalyzed by adenosine deaminases acting on RNA (ADARs). This modification modulates the gene expression by influencing splicing, RNA stability, and coding potential, depending on the site of editing. Although recent studies suggest a crosstalk between A-to-I editing and transcripts involved in DNA repair, the extent and functional significance of this interaction remain unclear. To investigate this, we applied the EpiPlex RNA assay--a method enabling epitranscriptome-wide detection of RNA modifications--in human lymphoblastoid TK6 cells. Across two biological replicates, we identified 869 transcripts bearing A-to-I-modifications. Gene Ontology analysis revealed significant enrichment in genome maintenance pathways, including "chromatin remodeling" and "DNA repair." Notably, 27 transcripts encoding DNA repair proteins--such as ATM, FANCA, BRCA1, POLH, and XPA--contained A-to-I sites within introns or 3' untranslated regions. To assess the isoform-specific contributions of ADAR enzymes--specifically ADAR1 p150 and p110--to RNA editing, we generated p150-deficient (p150 KO) and p150/p110-deficient (p150/p110 KO) TK6 cells. A-to-I editing peaks were reduced by [~]73.4% in p150 KO cells and nearly abolished (99.9%) in p150/p110 KO cells, indicating that most editing sites are p150-dependent, while a notable subset relies on p110. Importantly, a novel splice variant of XPA emerged in ADAR1-deficient cells, suggesting a role for RNA editing in alternative splicing regulation. Our epitranscriptomic analysis of A-to-I RNA editing underscores a multifaceted role for ADAR1-dependent editing in preserving genome integrity through posttranscriptional regulation of DNA repair genes, laying the groundwork for future studies into RNA-based mechanisms of genome maintenance.

genetics↗

bioRxiv2025

HaploThread: A Scalable Integrated Desktop Platform for Constructing and Visualizing Haplotype Networks for Large-sample Sequences

This note announces HaploThread, a user-friendly GUI desktop software designed for haplotype network construction and visualization. HaploThread is written in C++ using the Qt library, integrating network visualization and multiple multi-threaded haplotype construction algorithms such as McAN and fastHaN (includes MSN, MJN and TCS) based on plugin mechanisms. It offers a straightforward approach to constructing and visualizing haplotype networks from large sample sets, and to extending functionality with plugins, facilitating the analysis of genetic variations and their evolutionary relationships. HaploThread is an open-source software released under the GNU General Public License (GPL). Its precompiled executables for Windows is freely available for download at https://ngdc.cncb.ac.cn/biocode/tool/BT007948.

bioinformatics↗

bioRxiv2025

Bridging Scales: Spectral Theory Reveals How Local Connectivity Rules Sculpt Global Neural Dynamics in Spatially Extended Networks

The brains diverse spatiotemporal activity patterns are fundamental to cognition and consciousness, yet how these macroscopic dynamics emerge from microscopic neural circuitry remains a critical challenge. We address this by developing a spatially extended neural network model integrated with a spectral theory of its connectivity matrix. Our theory quantitatively demonstrates how local structural parameters, such as E/I neuron projection ranges, connection strengths, and density determine distinct features of the eigenvalue spectrum, specifically outlier eigenvalues and a bulk disk. These spectral signatures, in turn, precisely predict the networks emergent global dynamical regime, encompassing asynchronous states, synchronous states, oscillations, localized activity bumps, traveling waves, and chaos. Motivated by observations of shifting cortical dynamics in mice across arousal states, our framework not only explains this repertoire of behaviors but also offers a principled approach to inferring underlying effective connectivity changes from macroscopic brain activity. By mechanistically linking neural structure to dynamics, this work provides a powerful tool for understanding brain function and paves the way for identifying potential biomarkers for neurological disorders.

biophysics↗

bioRxiv2025

Receptor agonist of NFκB signaling ligand directs lung epithelial cell expansion through RANK signaling

Chronic obstructive pulmonary disease (COPD) is the third leading cause of death globally, with progressive emphysema driven by repeated epithelial damage and impaired repair. Recently, we found that secretion of the osteokine "receptor agonist of nuclear factor {kappa}B signaling ligand" (RANKL) is higher in lung fibroblasts from patients with COPD compared to control and that RANKL represses lung epithelial cell death. However, the underlying mechanism, its cross-species conservation and the involved epithelial cell types remain unclear. To investigate how RANKL affects lung epithelial cells, we used primary lung organoids from human and mouse epithelial cells and showed that RANKL supplementation increased alveolar organoid forming capacity in these cultures compared to vehicle-treated controls. In elastase-treated mice, RANKL was able to rescue the elastase-induced loss of epithelial cells compared to vehicle-treated controls and it augmented the proportion of epithelial cells in transitional states expressing Krt8 and MHCII. Using A549 epithelial cells to investigate whether RANK or the alternative receptor for RANKL, "leucine-rich repeat-containing G-protein coupled receptor 4" (LGR4) were responding to RANKL treatment, we found that RANKL most likely acts through RANK These results suggest that RANKL may enhance stem cell survival of primarily alveolar epithelial cells in both mice and humans. We therefore conclude that RANKL is another osteokine, in addition to periostin, osteopontin, osteoglycin, and osteoprotegerin, that has a role in lung tissue repair and that its signaling pathway could be explored for therapeutic applications. New & NoteworthyRANKL enhances human and murine alveolar epithelial cell expansion. Using organoids and an elastase injury model, we show RANKL promotes alveolar epithelial type II cell expansion and rescues transitional cell type loss. RANKL/RANK signaling therefore emerges as a conserved lung regenerative pathway and may be a potential therapeutic target for diseases like COPD with impaired epithelial repair.

cell biology↗

bioRxiv2025

The mitochondrial NAD transporter SLC25A51 is a modulator of beta cell senescence and type 2 diabetes

AbstractNicotinamide adenine dinucleotide (NAD+) is an essential redox cofactor and signaling molecule linked to age-dependent metabolic decline, with its compartmentalization regulated by the mitochondrial carrier SLC25A51. The mechanisms contributing to declining NAD+ levels during aging and the consequences of altered NAD+ homeostasis across tissues are poorly understood. Here, we show that SLC25A51 is upregulated in aging and aging-associated conditions, particularly in senescent cells. In a mouse model of beta-cell senescence, upregulated SLC25A51 was associated with beta-cell identity loss, senescence progression, and a reduced NAD+/NADH ratio. SLC25A51 was elevated following p16INK4a-, replicative-, irradiation-, and H2O2-induced senescence, with NRF2 implicated as a potential transcriptional regulator. Overexpression of SLC25A51, but not a transport-dead mutant, induced senescence factors, while its deletion prevented this effect. Beta-cell-specific deletion of SLC25A51 lowered p16INK4a levels in pancreatic islets, circulating insulin, and glucose levels, improving insulin sensitivity and indicating its role in cellular senescence and the metabolic control of beta-cell function.

cell biology↗

bioRxiv2025

The GSA Family in 2025: A Broadened Sharing Platform for Multi-Omics and Multimodal Data

The Genome Sequence Archive Family (GSA family) provides a suite of database resources for archiving, retrieving, and sharing multi-omics data for the global academic and industrial communities. It currently comprises four distinct database members: the Genome Sequence Archive (GSA, https://ngdc.cncb.ac.cn/gsa/), the Genome Sequence Archive for Human (GSA-Human, https://ngdc.cncb.ac.cn/gsa-human/), the open Archive for Miscellaneous Data (OMIX, https://ngdc.cncb.ac.cn/omix/), and the Open Biomedical Imaging Archive (OBIA, https://ngdc.cncb.ac.cn/obia). Compared to the 2021 version, the GSA family has significantly expanded by introducing a new database member, the OBIA, and through comprehensive upgrades to the existing ones. Notable enhancements to the existing members include broadening the range of accepted data types, enhancing the quality control systems, fine-tuning the data retrieval system, and refining the data-sharing management mechanisms.

genomics↗

bioRxiv2025

SORDINO for Silent, Sensitive, Specific, and Artifact-Resisting fMRI in awake behaving mice

Blood-oxygenation-level-dependent (BOLD) functional magnetic resonance imaging (fMRI) has revolutionized our understanding of the brain activity landscape, bridging circuit neuroscience in animal models with noninvasive brain mapping in humans. This immensely utilized technique, however, faces challenges such as acoustic noise, electromagnetic interference, motion artifacts, magnetic-field inhomogeneity, and limitations in sensitivity and specificity. Here, we introduce Steady-state On-the-Ramp Detection of INduction-decay with Oversampling (SORDINO), a transformative fMRI technique that addresses these challenges by maintaining a constant total gradient amplitude while acquiring data during continuously changing gradient direction. When benchmarked against conventional fMRI on a 9.4T system, SORDINO is silent, sensitive, specific, and resistant to motion and susceptibility artifacts. SORDINO offers superior compatibility with multimodal experiments and carries novel contrast mechanisms distinct from BOLD. It also enables brain-wide activity and connectivity mapping in awake, behaving mice, overcoming stress- and motion-related confounds that are among the most challenging barriers in current animal fMRI studies.

neuroscience↗

bioRxiv2025

Dickkopf1 is a Novel Endogenous Ligand for priming NLRP3 Inflammasome in Macrophages via TLR4

Dickkopf1(DKK1) is a quintessential Wnt antagonist and immunomodulator in various inflammatory diseases. The underlying molecular mechanisms of DKK1-mediated immunomodulation remain elusive. Here, we identified TLR4 as a new receptor for DKK1 to activate NF{kappa}B pathway-mediated gene expressions and pyroptosis via NLRP3 inflammasome in human and mouse macrophages. DKK1 employed TLR4 to initiate NF{kappa}B signaling cascade via MyD88. MyD88-TAK1-NF{kappa}B pathway activation by DKK1 increased HIF1, NF{kappa}B, and NLRP3 protein expression levels, leading to pyroptosis. Unlike LPS, DKK1 did not induce IRAK4 phosphorylation, while the interaction between MyD88 and IRAK4 was maintained for downstream signaling activation. DKK1 did not induce IRF3 phosphorylation in the nucleus and failed to induce IFN{beta} gene expression, indicating differential signaling from LPS. DKK1 primed macrophages via TLR4-MyD88, resulting in NLRP3 inflammasome-mediated pyroptosis via Caspase-1 and Gasdermin D maturation with various NLRP3 inflammasome activators, including Nigericin. Our results demonstrated that DKK1 is a novel endogenous priming ligand that augments differential NF{kappa}B pathway from LPS and NLRP3 inflammasome activation via TLR4 in mouse and human macrophages.

immunology↗

bioRxiv2025

CDK9 degradation Inhibits Gastroesophageal Cancer growth and Overcomes Radiation Resistance by Increasing Chromatin Accessibility and Downregulating YAP1/TEAD signaling

BackgroundGastroesophageal cancer (GEAC) remains a major health burden and urgently needs novel therapeutic targets. The inhibition of CDK9s activity holds the potential to be a highly effective anti-cancer therapeutic. However, the functional role of CDK9, and its potential targeting in GEAC, remain largely unknown. ObjectiveWe aim to evaluate the potential of degradation CDK9 in GEAC treatment and explore its mechanisms. DesignWe evaluated the expression and distribution of CDK9 in GEAC tissue. We designed and synthesized novel CDK9 degraders using proteolysis targeting chimeras (PROTACs) strategy and selected the promising one for further anti-tumor activity evaluation both in vitro and in vivo. We evaluated the effects of CDK9 degradation on epigenetic reactivation, gene expression and chromatin accessibility. We evaluated the co-targeting of CDK9 and YAP/Tead signaling for GEAC treatment, especially for radiation-resistant tumor treatment. ResultsWe demonstrated significantly elevated CDK9 expression in primary GEAC tumor tissues compared to normal tissues, in association with poor survival. We developed a novel CDK9 degrader, YX0597, reducing RNA Pol II Serine 2 phosphorylation, and inhibition MCL-1; this was accompanied by potent inhibition of GEAC cell growth, especially in radiation-resistant tumor cells. Mechanistically, YX0597 strongly enhanced chromatin accessibility, activating epigenetically silenced genes; and dramatically inhibited YAP/TEAD signaling. CDK9 closely interacts with YAP/TEAD signaling, and co-targeting these two mediators could be a novel treatment strategy for the treatment of GEAC. ConclusionOur studies reveal a new avenue for targeting CDK9-hyperactivated GEAC tumors, especially in combination with YAP1/TEAD inhibition in radiation-resistant GEAC tumors. What is already known on this subject?Gastroesophageal cancer (GEAC), the third-leading cause of global cancer death and urgently needs novel targeted therapies. Targeted protein degradation has emerged as an attractive strategy to fight cancer, complementing the activity of traditional small-molecule inhibitors. Cyclin-dependent kinase 9 (CDK9) has been implicated in various cancers. Recently, CDK9 degraders have now been developed for future clinical benefit in targeting tumors with highly activated CDK9. What are the new findings?In this study, we show that CDK9 expression was significantly elevated in primary GEAC tumor tissues including PDXs compared to normal tissues, in association with poor survival. We developed a novel CDK9 degrader XY0597 and proved that it inhibits the progression of GEAC by increasing chromatin accessibility and down-regulating YAP/Tead signaling. We also proved a strong connection between CDK9 and YAP/TEAD signaling and Co-targeting them synergistically inhibits GEAC tumor cell growth, especially in tumors with high YAP signaling activity, such as radiation-resistant cancer. How might it impact on clinical practice in the foreseeable future?Our data suggests that CDK9 is a viable anticancer target in GEAC. We demonstrate that a novel CDK9 degrader, YX0597, has high potential clinical application for GEAC treatment, especially in radiation-resistant cancer. We reveal the crosstalk of CDK9 and YAP/TEAD signaling, and that co-targeting these two mediators could be a new avenue for targeting CDK9-hyperactivated GEAC primary and radiation-resistant tumors.

cancer biology↗

bioRxiv2025

Fluorescence-barcoded cell lines stably expressing membrane-anchored influenza neuraminidases

The discovery of broadly protective antibodies to the influenza virus neuraminidase (NA) has raised interest in NA as a vaccine target. However, recombinant, solubilized tetrameric NA ectodomains are often challenging to express and isolate, hindering the study of anti-NA humoral responses. To address this obstacle, we established a panel of 22 non-adherent cell lines stably expressing native, historical N1, N2, N3, N9, and NB NAs anchored on the cell surface. The cell lines are barcoded with fluorescent proteins, enabling high-throughput, 16-plex analyses of antibody binding with commonly available flow cytometers. The cell lines were at least as efficient as a Luminex multiplex binding assay at identifying NA antibodies from a library of unselected clonal IgGs derived from human memory B cells. The cell lines were also useful for measuring the magnitude and breadth of the serum antibody response elicited by experimental infection of rhesus macaques with influenza virus. The membrane-anchored NAs are catalytically active and are compatible with established sialidase activity assays. NA-expressing K530 cell lines therefore represent a useful tool for studying NA immunity and evaluating influenza vaccine efficacy.

immunology↗