title: Intranasal ΔNS1-86 Live Attenuated Vaccines Confer Sterilizing Immunity to Mammalian-Adapted H5N1

city: Daejeon

first: Woohyun

last: Kwon

address: expo-ro

institute: Institute for Basic Science

summary: Influenza A viruses continue to evolve across avian and mammalian hosts, highlighting the need for vaccines that elicit broad and durable immunity. Here, we developed a modular live-attenuated influenza vaccine (LAIV) platform based on a short NS1 truncation (ΔNS1-86) engineered to express heterologous HA1 antigens through a multibasic-cleavage and 2A-peptide system. ΔNS1-86 LAIVs replicated in an attenuated manner while maintaining stable antigen expression. In mice, a single intranasal dose induced strong neutralizing antibodies and robust cross-reactive IFN-γ–producing T cell responses against divergent H5N1 clades and H9N2 lineages. In ferrets, the ΔNS1-86 LAIV provided sterilizing protection against a highly mammalian-adapted North American clade 2.3.4.4b H5N1 virus, completely blocking viral shedding and systemic dissemination—efficacy that surpassed a two-dose inactivated vaccine despite similar neutralization titers. The platform was adaptable to multiple HA/NA backbones and compatible with DIVA diagnostics. These findings establish ΔNS1-86 LAIVs as versatile candidates for broad influenza protection and pandemic preparedness.

overall_design: Single cell RNA seq profiling was performed on cervical lymph nodes collected at 5dpi from vaccinated and boosted with either an inactivated vaccine or LAIV and subsequently challenged with the H5N1 strain GA/W22-145E/22.

first: Boyeong

last: Jeong

first: Eun-Ha

last: Kim

first: Seung-Gyu

last: Jang

first: Se-Mi

first: Dongbin

last: Park

first: Young-Ki

last: Choi


title: Dual degradation of IKK1 and NR4A1 for AML therapy

status: Public on Jul 27 2026

city: Gainesville

first: Chandra Kumar

last: Maharjan

state: Florida

address: 1333 Center Drive Rm BG003

department: Pathology, Immunology and Laboratory Medicine

country: USA

institute: University of Florida

type: Expression profiling by high throughput sequencing

summary: Acute myeloid leukemia (AML) is an aggressive hematologic malignancy with poor clinical outcomes and limited therapeutic options. Aberrant activation of the IKKβ–NF-κB pathway occurs in approximately 40% of AML cases and contributes to leukemogenesis. However, pharmacological inhibition of IKKβ has been limited by serious toxicities, including neutrophilia. Here we identify IKKβ and NR4A1 as cooperative drivers of AML progression and develop a proteolysis-targeting chimera (PROTAC) to simultaneously degrade both proteins. Although NR4A1 has previously been described as a tumor suppressor in AML, our findings demonstrate that NR4A1 exhibits context-dependent oncogenic functions in AML. Notably, IKKβ and NR4A1 are highly expressed in AML associated with poor clinical outcomes and mutually enhance each other’s expression, playing synergistic roles to promote AML pathogenesis. To therapeutically target this pathway, we designed and synthesized a series of celastrol-based PROTACs that exploit celastrol’s ability to bind both IKKβ and NR4A1. Among these compounds, the lead A9 induced potent cytotoxicity in multiple AML cell lines and primary AML samples through CRBN E3 ligase–dependent dual degradation of IKKβ and NR4A1. In vivo, A9 suppressed leukemia progression in a KMT2A::MLLT3 AML mouse model without inducing neutrophilia, supporting dual PROTAC-mediated degradation as a promising therapeutic strategy.

overall_design: MOLM-13 cells and MONO-MAC-6 cells were treated with DMSO or A9 PROTAC degrader of IKK2 and NR4A1 (at 0.125 µM for MOLM-13 and 0.25 µM MONO-MAC-6) for 72 h, and bulk RNA-sequencing and analysis were performed. N =  3 per treatment condition per cell line

first: Yi

last: Liu

first: Zeng

last: Jin

first: Gruangrong

last: Zheng

first: Weizhou

last: Zhang


title: Postnatal Pax7-expressing limb cells are multipotent and generate non-myogenic lineages that persist into adulthood.

city: Paris

first: Takuto

last: HAYASHI

address: 160 Rue de Vaugirard

country: France

institute: Institut Necker Enfants Malades

summary: Organs are composed of diverse cell types that arise from distinct or shared progenitor cells. Skeletal muscle originates from mesoderm-derived Pax7⁺ stem/progenitor cells, which typically differentiate into myoblasts and form muscle fibers. During embryonic development, however, Pax7⁺ somitic cells can also generate non-muscle  lineages such as dermis and adipocytes.In this study, we examined whether Pax7⁺ cells in postnatal growing limb muscle retain similar multipotency. Lineage-tracing analyses revealed unexpected plasticity during the early postnatal period, with Pax7⁺ cells giving rise not only to myogenic cells but also to multiple non-myogenic lineages. Among these, we identified a previously unrecognized Pax7-derived fibro-adipogenic progenitor population (Pax7FAPs). Using mouse models, we further found that Notch signaling promotes a fibrogenic molecular program in neonatal Pax7⁺ cells, at the expense of myogenic differentiation, potentially biasing their fate toward the Pax7FAP lineage. Long-term tracing showed that Pax7-derived FAPs generated during the neonatal stage persist into adulthood and display enhanced proliferative capacity following muscle injury. Moreover, adult muscle injury can also induce the generation of Pax7FAPs, which proliferate more robustly than resident stromal cells. Together, these findings reveal previously unappreciated multipotency in postnatal Pax7⁺ cells and provide new insights into their contribution to muscle development and regeneration.

overall_design:
We analyzed the diversity of Pax7 progeny during the early postnatal period using Pax7CreERT2; Rosa26HTB::GFP mice.
In addition, to investigate the effect of Notch signaling activation on Pax7-positive cells, we used Pax7CreERT2; NICD mice.
In both experimental settings, cells were isolated by fluorescence-activated cell sorting (FACS). Sorted cells were subsequently subjected to single-nucleus RNA sequencing (snRNA-seq) for the lineage-tracing samples and single-cell RNA sequencing (scRNA-seq) for the NICD samples and muscle injury experiment.

first: Stamatia

last: Gioftsidi

last: Hayashi

first: Sidy

last: Fall

first: Sabrine

last: Jagot

first: Matthieu

middle: D

last: Santos

first: Fabien

middle: L

last: Grand

first: Frederic

last: Relaix

first: Philippos

last: Mourikis


title: Distinct transcriptional and functional modules mediate resistance to targeted therapy in glioblastoma

city: San Francsico

first: Sixuan

last: Pan

state: California

institute: UCSF

summary: Glioblastomas (GBM) harbor recurrent mutations in receptor tyrosine kinase (RTK) signaling, yet targeted therapy has shown limited efficacy. To dissect mechanisms underlying drug resistance, we combined single cell RNA-sequencing (scRNA-seq) and functional genomic screens following MEK or SHP2 inhibition in preclinical GBM models. ScRNA-seq of mouse intracranial xenografts revealed that although mesenchymal-like GBM subpopulation exhibits heightened sensitivity to both MEK or SHP2 inhibition, each targeted therapy induces distinct transcriptional signatures. Genome-wide CRISPR interference (CRISPRi) screens identified shared mediators of MEK or SHP2 inhibitor sensitivity enriched for the conserved downstream effector Ras/RAF/MEK pathway. However, genes uniquely mediating SHP2 inhibitor response were enriched for Ras/RAF/MEK-independent signaling modules such as SRC family kinases, GSK3B, and SMAD proteins regulating cell adhesion and differentiation. Perturb-seq of 39 target genes following MEK or SHP2 inhibition identified 10 discrete clusters with distinct effects on Ras signaling and cellular phenotypes. Proliferation and differentiation gene expression signatures clustered genetic perturbations into 4 broad classes: cell cycle regulators, positive effectors of Ras/RAF/MEK signaling, negative regulators of Ras/RAF/MEK signaling, or parallel regulators of Ras/RAF/MEK independent pathways. Combinatorial linear modeling from transcriptional signatures identified multiple gene-drug relationships including expected synergistic and potentiation combinations between MEK, SHP2, and CDKN2A/B and unexpected antagonistic and neomorphic interactions with indirect effectors such as LZTR1 and RAC1. Finally, preclinical testing of predicted synergistic inhibitor combinations with MEK, SHP2, and CDK4/6 inhibition demonstrated improved efficacy compared to molecular monotherapy in multiple GBM preclinical models. Taken together, our approach provides a systems-level framework for understanding RTK-Ras signaling network principles and targeted therapy resistance in glioblastoma.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Perturb dataset: The Perturb-seq experiment was designed to systematically profile how genetic perturbations interact with targeted therapy in glioblastoma. SB28 mouse glioma cells engineered to express dCas9-KRAB were transduced at low multiplicity with a dual-sgRNA CRISPRi library targeting 45 genes selected from genome-wide screens, along with non-targeting controls, ensuring that each cell received a single dual-guide construct. After FACS enrichment of sgRNA-positive cells, cells were exposed for five days to selumetinib (MEK inhibitor), RMC-4550 (SHP2 inhibitor), or vehicle. Cells from each sgRNA–drug condition were uniquely labeled via MULTI-seq lipid barcoding, pooled, and loaded into the 10x Chromium system for single-cell transcriptome and direct sgRNA capture. This design enabled simultaneous measurement of gene knockdown, drug response, and transcriptional state across all perturbation–drug combinations, supporting integrated analysis of gene–drug interactions and resistance mechanisms.  Xeno dataset: The scRNA-seq study analyzed human NF1-mutant GBM43 in mice intracranial xenografts treated in vivo with selumetinib (MEK inhibitor), RMC-4550 (SHP2 inhibitor), or vehicle (DMSO). Tumors were harvested after treatment to profile treatment-specific transcriptional changes and cell-state shifts.

first: John

first: Joanna

last: Pak

first: Kanish

last: Mirchia

first: Emily

last: Payne


title: High-throughput mapping of 6,888 RAD51D variants identifies distinct biochemical functions needed for homologous recombination and olaparib response

city: Ann Arbor

first: Jacob

last: Kitzman

state: MI

address: 4811 Med Sci II, 1241 E Catherine St

department: Department of Human Genetics

institute: University of Michigan

summary: The tumor suppressor RAD51D is essential for homologous recombination (HR). Pathogenic variants in RAD51D are associated with breast and ovarian cancers. However, most clinical missense variants are of unknown significance. We performed a multiplex assay of variant effect to test 6,888 RAD51D coding variants for loss-of-function. The resulting variant-to-function map perfectly separates known pathogenic and benign variants and is validated by orthogonal HR and biochemical assays across 70 clinical variants. Our screen shows that variants in the DNA- binding or ATPase core most severely compromise HR, and we identify the RAD51D-RAD51C interface within the BCDX2 complex as essential for regulating its ATPase activity. We hypothesize that, paradoxically, the primary function of RAD51D is to slow the ATPase activity of BCDX2, thereby allowing sufficient time and space for RAD51 filament assembly. Together, we identify hotspots of deleterious RAD51D variants and uncover the mechanisms by which variants compromise its biochemical functions.

overall_design: We performed saturation mutagenesis of the RAD51D cDNA, split across 6 tiles. Mutational libraries were stably integrated into U2OS RAD51D KO cells such that each resulting cell expressed a single RAD51D variant. Mixed cellular populations were treated with 250 nM olaparib for two passages to enrich for cells expressing functionally normal RAD51D mutants. Variant frequencies were quantified by deep amplicon sequencing before and after selection.

first: Hemker

last: Shelby

first: Darrah

last: Kristie

first: Rawal

last: Yashpal

first: Goff

last: Noah

first: Sung

last: Patrick

first: Bernstein

last: Kara

first: Kitzman

last: Jacob


title: The clinical and molecular spectrum of AGO2-associated Lessel-Kreienkamp neurodevelopmental syndrome

city: Regensburg

first: Davor

last: Lessel

address: Franz-Josef-Strauss-Allee 11

department: Institut für Humangenetik

country: Germany

institute: Universität Regensburg

type: Non-coding RNA profiling by high throughput sequencing

summary: Pathogenic variants in AGO2, encoding a central component of the RNA-induced silencing complex (RISC), cause the neurodevelopmental disorder Lessel-Kreienkamp syndrome (LESKRES). The variant spectrum and associated molecular mechanisms underlying phenotypic variability and disease severity remain incompletely understood. We investigated 45 newly identified individuals carrying 33 distinct AGO2 variants, 30 of which were previously unreported. Phenotypic data from these and previously reported cases (n = 70) were integrated to delineate the LESKRES-associated clinical spectrum and genotype-phenotype correlations. Functional studies included shRNA-based silencing, co-immunoprecipitation, subcellular localization, and sequencing of AGO2-bound miRNAs. All individuals presented with a neurodevelopmental disorder of variable severity. Delayed speech and language development (97%), intellectual disability (97%), and motor delay (93%) were the most consistent features, frequently accompanied by muscular hypotonia, autistic traits, attention deficit hyperactivity disorder, visual impairment and structural brain anomalies. Systemic manifestations, including skeletal, craniofacial, cardiac, and male urogenital anomalies were common, underscoring AGO2’s multisystemic role. Moreover, we report occurrence of gonadal mosaicism and reveal the presence of interfamilial and variant-specific clinical heterogeneity. Variants clustered in defined regions of AGO2, including the L1 loop, helix-7, and multiple loops of the PIWI domain, highlight structural hotspots critical for RISC activity. Not all pathogenic variants impaired shRNA-mediated silencing; this was restricted to p.(Arg714Trp) and p.(Asn729His). Biochemical analyses revealed that p.(Asp619Asn) impaired GW182 binding and P-body assembly. Variants p.(Arg506Gln), p.(Glu531Gln) p.(Gly604Arg) and p.(Asp619Asn), reduced C-terminal phosphorylation, implicating defective AGO2 recycling. AGO2–miRNA co-immunoprecipitation and sequencing demonstrated variant-specific perturbations in miRNA association, strand selectivity, and isomiR generation. Variants near the hinge of the helix-7 region, especially p.(Phe182del), induced extensive changes in miRNA association and 3′-end modification, suggesting impaired anchoring within the miRNA-binding pocket. Our findings substantially broaden the clinical and molecular landscape of LESKRES, establishing AGO2 as a pivotal regulator of neurodevelopment whose structural integrity is essential for precise miRNA-mediated gene regulation. Pathogenic variants disrupt distinct interconnected processes: P-body association, phosphorylation-dependent turnover, and miRNA interactions, culminating in dysregulated post-transcriptional gene silencing. These mechanistic insights link specific structural perturbations in AGO2 to graded clinical outcomes and underscore the critical role of AGO2 conformational dynamics in human neurodevelopment.

overall_design: HEK293T cells were transfected with Flag/HA-tagged AGO2 variants (WT, F182del, N359H, G604R, D619N), AGO2-miRNA complexes were immunoprecipitated using the anti-HA antibody and miRNA sequencing was performed.

first: Debora

last: Tibbe

first: Christina

last: Kiel

first: Hans-Jürgen

last: Kreienkamp


title: Deconvolution of Low-count RNA Sequencing Data for Tumor Cells Using Embedded Negative Binomial Distributions

city: Houston

first: George

last: Calin

state: TX

address: 1515 Bertner Ave

institute: The University of Texas MD Anderson Cancer Center

summary: A major challenge in studying tumor heterogeneity is the presence of mixed signals in gene expression data, especially when working with sparse count datasets such as microRNA and spatial transcriptomics. Estimating tumor-specific transcript proportions can provide critical insights into tumor heterogeneity and cell plasticity. However, current methods often require comprehensive single-cell reference data, which is not always obtainable, and may suffer from technical biases. To overcome these issues, we introduce DeMixNB, a semi-reference-based deconvolution method designed specifically for estimating transcript proportions from sparse expression matrices using a negative binomial model, without the reliance on high-quality external reference data. Through simulations and analyses of real microRNA sequencing and spatial transcriptomics data from breast and lung cancers, we demonstrate that DeMixNB accurately estimates tumor-specific transcript proportions and provides meaningful biological insights from real-world datasets.

overall_design: Benchmarking dataset with precisely known miRNA proportions, comprised of thirty-nine samples created by systematically mixing HS-5 bone marrow stromal fibroblast cells with either wild-type HCT116 colorectal cancer cells or CRISPR-Cas9-engineered Dicer1 knockout HCT116 colorectal cancer cells

first: Matthew

last: Montierth

first: Kinga

last: Nemeth

first: Wenyi

last: Wang


title: Cannabinoid-driven epigenetic remodeling of a chromatin-lipid homeostatic axis via ARID1A uncovers a metabolic vulnerability in cancer

city: L'Hospitalet de Llobregat

first: David

last: Llobet

state: Barcelona

address: Gran Via de L'Hospitalet 199-203

department: Gynecologic cancer

country: Spain

institute: Bellvitge Biomedical Research Institute (IDIBELL)

type: Genome binding/occupancy profiling by high throughput sequencing

summary: A substantial body of evidence supports a tumor suppressor role for the chromatin remodeler ARID1A, whose inactivating mutations and loss of function are common across human cancers. These alterations have spurred extensive efforts to uncover vulnerabilities associated with ARID1A deficiency. However, emerging studies suggest that ARID1A may also exert context-dependent tumor-promoting functions, opening new therapeutic avenues in ARID1A-proficient tumors. In endometrial cancer (EC), ARID1A mutations typically arise late during tumor progression and are dispensable for tumor initiation, providing a relevant context to interrogate its noncanonical homeostatic roles. Here, through integrated multiomics analyses, cellular bioenergetics and in vivo studies using EC patient-derived xenografts and genetically engineered mouse models, we demonstrate that functional ARID1A is required for acquired resistance to nutrient deprivation induced by antiangiogenic (starvation) therapy. Mechanistically, ARID1A orchestrates an epigenetically regulated chromatin-lipid homeostatic program involving lipid metabolism and mitochondrial fatty acid oxidation. Unexpectedly, we find that Δ9- tetrahydrocannabinol (THC), a phytocannabinoid and orthosteric CB1/CB2 agonist, exerts its antitumor activity by repressing ARID1A, thereby dismantling this chromatin- lipid metabolic axis and restoring sensitivity to starvation therapy. These findings position ARID1A as a central node linking chromatin remodeling to metabolic plasticity, and identify THC as a pharmacologic disruptor of this epigenetic metabolic program with therapeutic potential in ARID1A-proficient tumors.

overall_design: RNA-seq of HEC-1A cells: untreated controls and THC-treated cells at 24 h and 72 h; ATAC-seq analysis in HEC-1A cells: untreated controls and THC-treated cells at 24h and 72h.; CUT&RUN of the histone modification H3K4me3 in HEC-1A cells: untreated controls and THC-treated cells.

last: LLobet- Navas


title: RPL41 stabilizes ribosome dynamics and supports long-protein homeostasis in mammals

city: Chikusa-ku, Nagoya, Aichi

first: Kazuya

last: Ichihara

address: Furo-cho

country: Japan

institute: Nagoya University

summary: Ribosomal protein L41 (RPL41 or eL41) is the smallest ribosomal protein and forms the eukaryote-specific bridge, eB14, near the decoding center; however, its role in mammalian translation remains unclear. In this study, we established RPL41-deficient models of human HEK293T cells and mice to define its function. Cryo-electron microscopy revealed that RPL41 constrains intersubunit conformational dynamics without inducing major local static rearrangements. Loss of RPL41 altered A-site dynamics, slowed elongation, modestly increased amino acid misincorporation, and modestly enhanced readthrough of collision-inducing reporter sequences. Quantitative proteomic analysis suggested that these translational defects compromise long-protein homeostasis, as evidenced by increased insolubility and reduced abundance of long proteins. In vivo, Rpl41−/− mice were viable but exhibited growth retardation and decreased abundance of long proteins in tissues. Our findings reveal a conserved role for RPL41 in maintaining ribosome dynamics and translational fidelity, indicating that RPL41 supports ribosome function and long-protein homeostasis in mammals.

overall_design: Translatome and transcriptome analysis of RPL41-deficient HEK293T cells and mice

first: Mina

last: Hirata

first: Akinobu

last: Matsumoto


title: Macrophage Reprogramming Induced by Hypertensive Stress Underlies Pathogenesis of Non-Obese Heart Failure with Preserved Ejection Fraction

city: Suita

first: Manami

last: Katoh

state: Osaka

address: 6-1 Kishibe-simmachi

department: Department of Heart Failure Pathophysiology  and Therapeutics

institute: National Cerebral and Cardiovascular Center Research Institute

summary: Heart failure with preserved ejection fraction (HFpEF) is a leading cause of morbidity and mortality worldwide; nonetheless, effective therapies remain elusive. In Western populations, HFpEF is typically obesity-related, whereas in Asia it frequently develops in non-obese patients with hypertension and underlying mechanisms are poorly defined. Here, analysis of two Japanese hospital cohorts revealed that obesity and metabolic comorbidities are uncommon in HFpEF, while sympathetic overactivation and hypertension predominated. To model this condition, we combined isoproterenol (ISO)–induced sympathetic activation with L-arginine methyl ester (L-NAME)-induced endothelial dysfunction, establishing the first non-obese HFpEF mouse model. These mice exhibited hallmark HFpEF features, whereas ISO alone caused hypertensive heart disease (HHD) without HFpEF, indicating that a "second hit" is essential for HFpEF progression. HFpEF mice showed expansion and reprogramming of cardiac macrophages to Pla2g7⁺ phenotypes and fibroblast activation via PDGF signaling, as loss of Timd4⁺ resident macrophages impaired cardiomyocyte energetics due to diminished BMP signaling. These outcomes drove maladaptive remodeling and inflammation, which precedes HFpEF onset. Notably, inhibition of the inflammatory mediator ANGPTL2 attenuated cardiac inflammation and prevented HFpEF progression. Our findings identify hypertensive stress-induced macrophage reprogramming as a central mechanism of non-obese HFpEF and highlight cardiac inflammation as a promising therapeutic target.

overall_design: Nuclei were extracted from hearts of model mice and control mice, and single-nucleus RNA-seq was performed on each sample. Mice heart tissue samples were obtained from the left ventricular apex myocardium of hearts.

first: Michio

last: Sato

first: Yuichi

last: Oike


title: RHOAY42C-driven cell lineage plasticity and immune remodeling of diffuse gastric cancer [scRNA-Seq]

first: Kyung Pil

last: Ko

state: Tx

address: 1515 Holcombe Blvd

institute: The University of Texas M.D. Anderson Cancer Center

summary: Diffuse gastric adenocarcinoma (DGA) is an aggressive gastric cancer subtype with a poor prognosis. RHOAY42C is a recurrent mutation in DGA, yet its independent oncogenic role remains unclear. We used genetically engineered gastric organoid models to demonstrate that RHOAY42C, along with KrasG12D activation and Trp53 loss (RKP), is sufficient to drive DGA without loss of E-cadherin. Integrated single-cell RNA sequencing (scRNA-seq) and spatial transcriptomics (Xenium In Situ) revealed that RKP tumors exhibit mucinous histopathology and a unique lineage trajectory originating from Aqp5+ cells and differentiating into Muc1/Muc4+ cells via YAP1 signaling. RKP tumors establish a highly immunosuppressive tumor microenvironment in which AREG-mediated activation of EGFR promotes T cell exhaustion. Unlike E-cadherin loss-induced DGA, RKP tumors display strong susceptibility to PD-1 blockade. These findings redefine the oncogenic role of RHOAY42C, highlighting its sufficiency for DGA development and identifying PD-1 inhibition as a promising therapeutic strategy for patients with DGA harboring this mutation.

overall_design: EKP and RKP cells were subcutaneously transplanted into immunocompetent mice and the tumors were comparatively analyzed.

first: Kyung-Pil

first: Sangmin

last: Kim

first: Jinho

last: Jang

first: Yoojeong

last: Seo

first: Gengyi

last: Zou

first: Jie

last: Zhang

first: Sohee

last: Jun

first: Brittany

last: Morrow

first: Ryan

last: Park

first: Yuan-Hung

last: Lo

first: Jaffer

last: A. Ajani

first: Michael

last: A. Curran

first: Hyunki

first: Jae-Il


title: RHOAY42C-driven cell lineage plasticity and immune remodeling of diffuse gastric cancer [CUT&Run]

overall_design: 2D-cultured EKP and RKP cells were comparatively analyzed for genes regulated by TEAD1 transcription factor.


title: Transcriptomic profiling of BYSL knockdown in MDA-MB-231 human breast cancer cells

city: DaLian

first: Luo

last: Kai

state: Liaoning Province

address: No. 467, Zhongshan Road, Shahekou District, Dalian 116011, China

country: China

institute: The Second Hospital of Dalian Medical University

summary: To characterize the transcriptional effects of BYSL depletion in human breast cancer cells, MDA-MB-231 cells were transfected with negative-control siRNA or SI-BYSL-Homo-838 targeting BYSL. Strand-specific poly(A)+ RNA sequencing was performed with three biological replicates per condition.

overall_design: Bulk RNA-seq analysis of MDA-MB-231 cells under two conditions: negative-control siRNA and siRNA-mediated BYSL knockdown. Each condition contains three independent biological replicates.


title: Mechanisms of β-cell adaptation to increased insulin demand

city: Bucharest

first: Ana

last: Vacaru

address: BP Hasdeu 8

country: Romania

institute: ICBP-NS

summary: β-cells have various ways to adapt to stress, but in diabetes, these pathways become dysregulated, driving β-cell dysfunction, resulting in poor glucose control. Unfolded protein response (UPR) plays central roles within these pathways, cooperating to restore endoplasmic reticulum function under stress. In this study, we followed the dynamic interaction between UPR players during stress onset in response to targeted ~50% β-cell ablation in mice, using diphtheria toxin (DT) in the NSG RIP-DTR system. Importantly, these ablated mice remained normoglycemic for up to 10 weeks. By transcriptomic profiling of the islets we assessed two polarized UPR responses, with different dynamics. The first response was characterized by activation of apoptosis and clearance pathways, while the second, at a later time point, was characterized by adaptive pathways, including increased insulin processing. These findings offer potential targets for strategies aimed at promoting enhanced resistance to stress of the β-cells.

overall_design: NSG RIP-DTR mice were followed at different time points after partial β-cell ablation  (0, 1, 3, 5 days post DT-injection). Islets from these mice were isolated for bulk RNAseq to identify molecular mechanisms that beta cells use to withstand stress.

first: Laura

middle: M

last: Daian

first: Andrei

first: Andreas

middle: F

last: Mathisen

first: Luiza

last: Ghila

first: Simona

last: Chera


title: RHOAY42C-driven cell lineage plasticity and immune remodeling of diffuse gastric cancer

first: Kyung Pil

last: Ko

overall_design: For Xenium In Situ experiment, a single FFPE block was prepared from EKP and RKP tumor samples and placed onto a Xenium slide. Alongside the 379 Mouse Tissue Atlassing gene panel, additional 100 genes were incorporated for further analysis.


title: Loss of tolerance in cow milk allergic endotypes is linked to allergen-specific CD4+ T cell subset outgrowth and microbial dysbioses

city: Doha

first: Asmma

last: Doudin

address: Al Gharrafa Street, Ar Rayyan

department: Research

country: Qatar

institute: Sidra Medicine

summary: Cow milk allergy (CMA) is the most prevalent pediatric food allergy, presenting in IgE and non-IgE mediated forms. Currently, the CD4+ T cell inflammatory responses underlying  non-IgE CMA remain poorly characterized. By applying an allergen-reactive CD4+ T cell sorting and differential gene expression analysis approach, we characterized the CD4+ T cell subsets associated with both IgE and non-IgE mediated CMA endotypes and assessed their associations with gut bacterial compositions. Analysis of conventional (Tcon) and regulatory (Treg) CD4+ T cells revealed an expansion of inflammatory Tcon in CMA subjects, coincident with TH-like subset polarization and alterations in Treg signaling pathways. TH2-skewing was dominant in IgE mediated allergies, whereas subjects with non-IgE mediated allergies exhibited a CD4+ T cell compartment dominated by a novel TH response. Microbial dysbiosis was observed across all CMA endotypes, and network mapping revealed significant associations between a decrease in butyrate producing bacteria and the presence of inflammatory TH-like subsets. These insights link the composition of the allergen-reactive Tcon/Treg compartment to CMA pathology and identifies differences in bacterial dysbioses associated with specific disease endotypes.

overall_design:
RNAseq analysis of sorted TconCM+ (CD154+) and TregCM+ (CD137+) subsets from healthy controls and allergic subjects after 6hrs of cow milk protein stimulation.

Sample groups:
IgE mediated CMA subjects without atopic dermatitis (ALL)
non-IgE mediated CMA subjects (ENP)
healthy controls (CON)
CMA subjects with atopic dermatitis (AD)

first: Tracy

last: Augustine

first: Nicholas

last: Van Panhuys


title: LMNA p.H222P mutation causes contractile dysfunction via impaired mitochondrial calcium uptake in human cardiac laminopathy

first: Jean-Sébastien

last: Hulot

address: Rue Leblanc

institute: Paris Centre de Recherche Cardiovasculaire

summary: Mutations in the LMNA gene, encoding lamin A/C, are implicated in laminopathies and can lead to dilated cardiomyopathy and heart failure. However, the underlying mechanisms remain unclear. Here, we generated human induced pluripotent stem cells-derived cardiomyocytes (hiPSC-CMs) from a patient carrying the LMNA p.His222Pro (c.665A>C) mutation and an isogenic CRISPR/Cas9-corrected control. Although differentiation efficiency and sarcomere organization were comparable, 3D cardiac organoids composed of LMNA mutant hiPSC-CMs displayed significantly reduced contractility compared to control organoids. Transcriptomic, biochemical and functional analyses revealed impaired mitochondrial calcium uniporter (MCU) complex activity in LMNA mutant hiPSC-CMs, marked by decreased MCU expression, an elevated MICU1:MCU ratio, and increased phosphorylation of pyruvate dehydrogenase (PDH) at Ser232 and PDH Ser300 – indicative of reduced mitochondrial calcium uptake. These defects were accompanied by decreased mitochondrial respiration and ATP production. Notably, treatment with the MCU activator amorolfine restored mitochondrial calcium uptake and contractility in LMNA mutant hiPSC-CMs. These findings identify mitochondrial calcium dysregulation as a key downstream consequence of the LMNA p.H222P mutation. Our results establish a mechanistic link between LMNA-associated cardiomyopathy and impaired mitochondrial function, highlighting the MCU complex as a potential therapeutic target in human cardiac laminopathy.

overall_design: RNA-seq profiling of human induced pluripotent stem cell-derived cardiomyocytes from a patient heterozygous for the LMNA p.His222Pro (c.665A>C) mutation and the corrected isogenic control generated with CRISPR-Cas9.

first: Magali

last: Seguret

first: Charlène

last: Jouve

first: Andrea

last: Ruiz-Velasco


title: The Core Subunit NDUFS2 in Mitochondrial Complex I is Critical to Oxygen Sensing in Human Ductus Arteriosus Smooth Muscle Cells

city: Kingston

first: Rachel

middle: Emily Teresa

last: Bentley

state: Ontario

address: 18 Stuart St, 342 Botterell Hall

department: Medicine

country: Canada

institute: Queen's University

summary: Rationale: Mitochondria in ductus arteriosus smooth muscle cells (DASMC) are oxygen sensors triggering vasoconstriction at birth; however, the mitochondrial oxygen sensing mechanisms are not fully understood. Given the conserved role of mitochondrial Complex I subunit NDUFS2 in other oxygen-sensing tissues, we examined its role in DASMC oxygen sensing, comparing it to other Complex I subunits (NDUFS1 and NDUFS7) and putative O2-sensor subunits (UQCRFS1 and COX4I2). Methods: Human DASMC were grown in hypoxia (pO2=41mmHg). Oxygen responsiveness was assessed, measuring oxygen-induced changes in intracellular calcium, [Ca2+]i, cell length, and mitochondrial reactive oxygen species (mROS) production. DASMC were treated for 48-hours with control or targeting. Knockdown was confirmed using qPCR and immunoblot. 3’RNA sequencing assessed molecular changes following siRNA manipulation. Results: Oxygen increased mitochondrial fission, [Ca2+]I, and constricted DASMC. 48-hours post-treatment, siNDUFS2 selectively depressed oxygen-induced increase in [Ca2+]i (siControl +18.6±2.3%, siNDUFS2 +5.5±1.5%, p<0.0001), DASMC shortening (from 18.4±1.1% to 8.9±0.8%, p<0.0001), and mROS (+24±4.9% untreated, -6.6±5.4% siNDUFS2, p<0.0001), without altering the KCl response or depressing respiration. The mitochondrial antioxidant MitoTEMPO reduced mROS (2.9±4.5%, p=0.001) and attenuated oxygen-induced cell shortening (8.4±0.9%, p=0.0003). Transcriptomics revealed distinct changes in mitochondrial pathways 48-hours post siNDUFS2. Conclusions: NDUFS2 regulates mROS and is a mitochondrial oxygen sensor in human DASMC.

overall_design: 3' RNA-seq profiling of primary human ductus arteriosus smooth muscle cells grown in hypoxia (pO2=~40mmHg) treated for 48-hours with control siRNA or siRNA targeting mitochondrial electron transport chain subunits (NDUFS1, NDUFS2, NDUFS7, UQCRFS1, or COX4I2).

first: Rachel ET

first: Kimberly

middle: J

last: Dunham-Snary

first: Ashley

middle: Y

last: Martin

first: Jeffrey

last: Mewburn

first: Asish

last: Dasgupta

first: Kuang-Hueih

last: Chen

first: Benjamin

middle: P

last: Ott

first: Tanvi

last: Nandani

first: Bernard

last: Thébaud

first: Mark

middle: K

last: Friedberg

first: Charles CT

last: Hindmarch

first: Stephen

last: Archer


title: Single-cell profiling of human liver and peripheral-blood γδ T cells according to HCMV serostatus

first: Bae-Hyeon

last: Moon

address: 193, Munji-ro, Yuseong-gu

country: South Korea

institute: Korea Advanced Institute of Science and Technology

summary: This study generated single-cell transcriptomic and surface-protein datasets to examine the heterogeneity of human liver sinusoidal γδ T cells and their association with latent human cytomegalovirus infection. γδ T cells isolated from liver perfusates of healthy living donors were profiled using single-cell RNA sequencing with antibody-derived tags. The dataset includes Vγ9Vδ2 and non-Vγ9Vδ2 γδ T-cell populations from HCMV-seropositive and HCMV-seronegative donors and enables comparison of their transcriptional states, surface phenotypes, differentiation patterns, and TCR repertoires. Paired peripheral-blood γδ T-cell samples were additionally profiled for comparison with liver-derived populations.eir diverse functions.

overall_design:
Liver perfusate and paired peripheral-blood samples were obtained from healthy living donors during liver transplantation. Liver sinusoidal mononuclear cells were isolated from perfusates, followed by depletion of CD14⁺ and CD19⁺ cells and fluorescence-activated sorting of γδ T cells. Liver-derived γδ T cells were analyzed using the Chromium Next GEM Single Cell 5′ Gene Expression platform with feature barcoding.

For paired peripheral-blood samples, γδ T cells were directly isolated by fluorescence-activated cell sorting without prior CD14⁺ or CD19⁺ cell depletion. Sorted blood γδ T cells were analyzed using the Chromium Next GEM Single Cell 5′ Gene Expression platform with feature barcoding and V(D)J enrichment. Custom primers for γδ T-cell receptor amplification were based on the method reported by Song et al. (2023, J Leukoc Biol., 114(6):630-638).

Samples were grouped according to tissue source and donor HCMV serostatus to compare liver and blood γδ T-cell populations and to evaluate transcriptional, phenotypic, and TCR-repertoire differences associated with latent HCMV infection.


title: MYD88 maintains skin homeostasis in filaggrin-dificient mice, wild-type mice

first: Poulami

last: Dey

address: 1301 Catherine St

institute: Michigan Medicine

summary: This dataset contains RNA-seq profiles from wild-type balb/c mouse eye-skin and was generated as a control cohort for comparison with a previously published MyD88-deficient dataset. These data provide a baseline transcriptomic reference for normal skin immune homeostasis.

overall_design:
The eye-skin biopsies of ft/ft and ft/ftMyD88 /- mice were excised from 6-week-old female BALB/c mice. Smaples were perfomed RNA extraction, purification, library preparation, and cDNA sequencing.
The eye-skin biopsies were excised from 6-week-old wild-type female BALB/c mice. Samples were perfomed RNA extraction, purification, library preparation, and cDNA sequencing.

first: Wu

last: Meng-Jen

first: Tsoi

last: Alex

first: Gudjohnsson

last: Johann

first: Archer

last: Nathan


title: CTCF regulates wild-type and recombinant AAV gene expression by shaping viral chromatin [V3C-seq]

city: Madison

first: Kinjal

last: Majumder

state: WI

address: 1525 Linden Drive

department: Oncology

institute: University of Wisconsin

summary: Recombinant Adeno-Associated Viruses (rAAV) gene therapy vectors have been engineered from wild-type AAV (wtAAV2) by inserting the viral telomeres (that serve as replication and packaging signals) on either side of therapeutic transgenes. However, efficient expression of rAAV transgenes require large doses, leading to sporadic toxic side effects.  We have identified a novel cis-acting element in wtAAV2 bound by the cellular architectural protein CTCF (CCCTC-binding Factor). This CTCF binding element is necessary for efficient wtAAV2 gene expression and is sufficient to enhance the efficiency of rAAV vector’s ability to express reporter transgenes. Chromatin profiling studies reveal that this CTCF binding element regulates the chromatin landscape of the virus and viral vectors. Our discovery of adding merely 19bp of AAV’s CTCF binding element has generated a promising new rAAV gene therapy platform with enhanced transgene expression without impacting vector packaging capacity.

overall_design: Identification of the cellular interaction site of Adeno-Associated Virus, and the localization of the resultant DNA damage response. Independent replicates of chromosome conformation capture (designated as V3C in the file name) have been performed. For each time-point or treatment, at least two independent replicates of the indicated assays have been performed.

first: Claireine

last: Larsen

first: Rhiannon

last: Abrahams


title: AP2-HS and a GDV1 regulatory feedback loop mediate environmental induction of sexual conversion in Plasmodium falciparum [RNA-Seq]

city: Barcelona

first: Elisabet

last: Tinto-Font

address: c/Rossello 153

department: Malaria

institute: ISGlobal

summary: Transmission of Plasmodium spp. from a human host to a mosquito vector requires that some parasites abandon the asexual replicative cycle in the blood, associated with all clinical symptoms of malaria, and convert into non-replicative sexual precursors called gametocytes. The sexual conversion rate (i.e., the proportion of parasites that convert at each cycle) is variable, which enables parasites to adjust the balance between sexual and asexual development to the conditions of the human blood environment. Under some specific types of stress, parasites enhance sexual conversion rates to increase their relative investment in transmission. Sexual conversion is orchestrated by the master regulator AP2-G, a transcription factor of the ApiAP2 family. In Plasmodium falciparum, the species that produces the vast majority of human malaria cases and deaths, activation of ap2-g expression requires the GDV1 protein, which displaces heterochromatin from the ap2-g promoter. An antisense long noncoding RNA (lncRNA) encoded in the same locus as GDV1, named gdv1-as, is a negative regulator of GDV1 expression, but how the expression of this lncRNA is regulated or how specific types of stress enhance sexual conversion is not known. Here we show that environmental induction of sexual conversion is initiated by activation of gdv1 expression in a process dependent on the AP2-HS transcription factor, previously described as the activator of the malarial protective HS response. Next, GDV1 activates the expression of its own repressor, gdv1-as, by removing heterochromatin from its putative promoter. This positive-negative regulatory feedback loop limits GDV1 activation to a short burst of expression. This same mechanism operates for stimulation of sexual conversion by different environmental conditions, and AP2-HS is also needed for the activation of metabolic pathways that compensate depletion of phospholipid precursors. These findings provide new insight on the mechanisms underlying the fundamental developmental decision of malaria parasites and explain their plasticity to increase the investment in transmission when the conditions of the environment are unfavourable.

overall_design: RNA-seq profiling of P. falciparum parasites exposed to choline depletion, dihydroartemisinin or heat shock, or control conditions.

last: Tintó-Font

first: Núria

last: Casas-Vila

first: César

last: Martínez-Guardiola

first: Alba

last: Pérez-Cantero

first: Neus

last: Ràfols

first: Prince B.

last: Nyarko

first: Alfred

last: Cortés


title: AP2-HS and a GDV1 regulatory feedback loop mediate environmental induction of sexual conversion in Plasmodium falciparum [H3K9me3 ChIP-seq]

overall_design: H3K9me3 ChIP-seq profiling of P. falciparum parasites exposed to choline depletion, dihydroartemisinin or heat shock, or control conditions.


title: Maternal immune activation delays DNA methylation and physiological maturation of cortical excitatory neurons

city: La Jolla

first: Joseph

middle: R

last: Ecker

state: CA

address: 10010 North Torrey Pines Road

department: Genomic Analysis Laboratory

institute: HHMI-Salk-Institute

type: Methylation profiling by high throughput sequencing

summary: Elevated levels of maternal pro-inflammatory cytokines during gestation can disrupt offspring neural development, increasing the risk of neurodevelopmental disorders. We studied the effects of Poly(I:C)-induced maternal immune activation (PIC-MIA) during mid-gestation on the DNA methylation and transcriptome of developing cortical excitatory neurons. PIC-MIA disrupted the developmental regulation of synapse-related genes and of genes implicated in autism spectrum disorders. Genomic regions that gain or lose DNA methylation during normal development were altered following PIC-MIA, including neurodevelopmental transcription factor binding sites. The DNA methylation and transcriptional changes were consistent with a delay in excitatory neuron maturation. Whole-cell recordings showed that PIC-MIA preferentially altered the physiological development of layer 5 excitatory neurons. Taken together, present results suggest that alterations in the epigenome, through the disruption of circuit formation, may drive the long-term consequences of maternal infection during gestation.

overall_design: RNA-seq and MethylC-seq profiling of whole cortex and INTACT purified nuclei of excitatory pyramidal neurons from perinatal mouse frontal cortex following Poly(I:C)-induced maternal immune activation or saline treatment.

first: Chi-Yu

last: Lai

first: Jessica

last: Arzavala

first: Antonio

last: Pinto-Duarte

first: Shiyuan

last: Wang

first: Junhao

last: Li

first: Hanqing

last: Liu

first: Julia

last: Osteen

first: Rosa

last: Gomez-Castanon

last: Nery

first: Susan

middle: B

last: Powell

first: Eran

middle: A

last: Mukamel

first: M

last: Behrens


title: CAR design equips diverse immune cells with unique antigen-specific activation identities

city: Tokyo

first: Satoshi

last: Yamazaki

address: 4 Chome-6-1 Shirokanedai, Minato City

institute: The Institute of Medical Science, The University of Tokyo

summary: Haematopoietic stem and progenitor cells (HSPCs) are multipotent cells, capable of generating all haematopoietic lineages, representing a powerful platform for regenerative immunotherapy. In parallel, CAR-engineering has extended beyond T cells, which have achieved success in treating B cell malignancies, to broader immune cell types alongside cell type-specific CAR design. However, how CAR architecture modulates activation phenotypes across immune lineages remains poorly understood. Here, we systematically compared previously validated, immune cell-optimised CARs across immune cell types using a human HSPC-derived multilineage CAR-immune cell platform. HSPC-derived T, NK and myeloid cells demonstrated immune activation across different CAR designs whereas cell-specific activation showed CAR design dependency in B cells. Hallmark pathway analysis revealed that immune lineage was the dominant determinant of activation-induced transcriptional states while CAR design modulated signalling programmes within lineage constrained programmes, notably proliferation, inflammation and cytokine responses. Single cell InTraSeq highlights differential activation of signalling pathways depending on CAR design and lineage identity with links to downstream transcriptional programmes across T, B and myeloid cells. Although exploratory and performed with limited sample sizes, this demonstrates the activated phenotypes and pathways mediated by CAR design and lineage identity that may inform the development of next-generation CAR-immune cells with tailored activation-induced characteristics.

overall_design:
InTraSeq: NOG-w41 mice irradiated at 1Gy and xenotransplanted with human cord-blood HSPCs that underwent ex vivo lentiviral transduction with a CD8⍺-CD8⍺-4-1BB-CD3z (4-1BB/z) CAR construct (Addgene #199457, Silva et al Biomolecules. 2023 Mar 2;13(3):459. doi: 10.3390/biom13030459), 4-1BB/z derived anti-CD19 scFv ligated to IgG4-NKG2D-2B4-CD3z (2B4/z) CAR construct (Addgene #157744, Chang et al Bioact Mater. 2023 Apr 5;27:168-180. doi: 10.1016/j.bioactmat.2023.03.018). At 16 weeks post-xenotransplantation, cells were isolated from the spleen and were cryopreserved.  At a later date, cells were thawed and fluorescence activated cell sorted into live single human CD45+ (hCD45+) murine CD45- (mCD45-)GFP+CD3+CD19-CD33-, hCD45+mCD45-GFP+CD56+CD3-, hCD45+mCD45-GFP+CD19+CD3-CD33-, hCD45+mCD45-GFP+CD33+CD3-CD19-, hCD45+mCD45-GFP-CD3+CD19-CD33-, hCD45+mCD45-GFP-CD56+CD3-, hCD45+mCD45-GFP-CD19+CD3-CD33- and hCD45+mCD45-GFP-CD33+CD3-CD19- cell populations for 4-1BB/z and 2B4/z-CAR immune cells and respective non-transduced control immune cells. The samples were multiplexed using 10x genomics Protocol 3: Cell Multiplexing Oligo Labelling for samples with >80% viability, according to the manufacturer's instructions. Cells were combined and CD19 antigen stimulated for 10 minutes at 37°C. Cells were counted and immediately fixed in ice-cold methanol according to the InTraSeq protocol (Cell Signalling Technologies).  InTraSeq datasets were generated from n=1 biological replicates and are therefore presented descriptively.
RNA-sequencing: NOG-w41 mice irradiated at 1Gy and xenotransplanted with human cord-blood HSPCs that underwent ex vivo lentiviral transduction with a CD8⍺-CD8⍺-4-1BB-CD3z (4-1BB/z) CAR construct (Addgene #199457, Silva et al Biomolecules. 2023 Mar 2;13(3):459. doi: 10.3390/biom13030459), 4-1BB/z derived anti-CD19 scFv ligated to IgG4-NKG2D-2B4-CD3z (2B4/z) CAR construct (Addgene #157744, Chang et al Bioact Mater. 2023 Apr 5;27:168-180. doi: 10.1016/j.bioactmat.2023.03.018) or 4-1BB/z derived anti-CD19 scFv ligated to IgG4-CD4-CD3z (CD3z) CAR construct (Addgene #157743, Chang et al Cell Rep. 2022 July 19;40(3):111128. doi: 10.1016/j.celrep.2022.111128). At 16 weeks post-xenotransplantation, cells were isolated from the spleen and were cryopreserved. At a later date, cells were thawed, activated for 24 hours at 37°C with recombinant human CD19 antigen and were fluorescence activated cell sorted into live single human CD45+ (hCD45+) murine CD45- (mCD45-)GFP+CD3+CD19-CD33-, hCD45+mCD45-GFP+CD56+CD3-, hCD45+mCD45-GFP+CD19+CD3-CD33-, hCD45+mCD45-GFP+CD33+CD3-CD19-, hCD45+mCD45-GFP-CD3+CD19-CD33-, hCD45+mCD45-GFP-CD56+CD3-, hCD45+mCD45-GFP-CD19+CD3-CD33- and hCD45+mCD45-GFP-CD33+CD3-CD19- cell populations for each CAR design. Cell populations were directly sorted into TRIZOL LS. GFP+ cells are CAR-expressing immune cells and GFP- cells are non-transduced control immune cells arising from the same xenograft.   RNA-sequencing datasets were generated from n=1 biological replicates and are therefore presented descriptively.   A technical negative control of TRIZOL LS and a technical positive control of 50,000 Jurkat cells were used to validate RNA extraction.

first: Takaharu

last: Kimura

first: Takao

last: Yogo

first: Ayano

last: Sugiyama-Finnis


title: SnRK2.4 and SnRK2.10 redundantly control developmental leaf senescence by sustaining ABA production and signaling

city: Warsaw

first: Michal

last: Krzyszton

address: Pawinskiego 5a

country: Poland

institute: Institute of Biochemistry and Biophysics, Polish Academy of Sciences

summary: Plants constantly and precisely control their growth by inducing distinct developmental programs to survive and produce high-quality offspring in the changing environment. The fine-tuning of the development according to endogenous and environmental signals requires exact intercellular signaling and a balanced response. Kinases of the Sucrose non-fermenting-1-Related protein Kinases type 2 (SnRK2s) family primarily take part in the response and adaptation to environmental stress factors. Notably, here we show that two ABA-non-activated SnRK2s, SnRK2.4 and SnRK2.10, are also activated in non-stress conditions in developmentally senescing leaves of Arabidopsis thaliana. Phenotypic, biochemical, and molecular analyses performed on single snrk2.4 or snrk2.10, and double snrk2.4/2.10 kinase mutants showed that SnRK2.4 and SnRK2.10, acting redundantly, promote developmental leaf senescence. Further, SnRK2.4 and SnRK2.10 enhance ABA accumulation in senescing leaves by inducing NCED2, one of key ABA biosynthesis-related genes. The two kinases induce developmental leaf senescence by modulating the expression of multiple ABA-responsive, osmotic stress, and senescence-related genes, such as the senescence master regulators ORE1, ORS1, WRKY33, WRKY75, and ANAC087. Furthermore, we show that SnRK2.4 and SnRK2.10 act upstream of MAPK signaling by enhancing the expression and activity of MAPKKK18, a senescence-inducing kinase. These results document a new regulatory function of SnRK2.4 and SnRK2.10: they are activated in Arabidopsis leaves in response to endogenous signals and redundantly induce developmental leaf senescence by stimulating ABA production and sustaining major ABA-dependent and –independent signaling pathways.

overall_design: Using 3’RNA-seq, we analyzed leaves transcriptomes (5 replicas) of Col-0 (WT) and snrk2.4/snrk2.10 double mutant of A. thaliana. Leaves were collected at 25 and 45 days after sowing (DAS).

first: Anna

last: Anielska-Mazur

last: Rachowka

last: Kulik


title: Functional dissection of enhancers in 2-Cell embryo with an in vitro model [STARR-seq]

city: Guangzhou

first: Man

last: Zhang

state: Guangdong

address: No.9, XingDaoHuanBei Road, Guangzhou International Bio-island, Haizhu District

department: Department of Basic Research

institute: Guangzhou Laboratory

summary: Zygotic genome activation (ZGA) marks the initial transcription event in embryogenesis, yet the mechanisms cis-regulating gene activation remain unclear due to material constraints. Here, we functionally dissect the enhancers across the mouse genome in DUX-induced 2C-like cells. Integrated analysis with the epigenetic modifications and RNA transcriptiome in both 2C-like cells and 2C embryos, regions which show enhancer activities and their potential downstream target gens were depicted. Among them, a notable proportion of promoters exhibits enhancer activities in totipotent cells. Those promoter regions are characterized by elevated levels of open chromatin-associated epigenetic modifications, which correlate with enhanced gene expression during ZGA. Our data further reveal that class Ⅲ mouse endogenous retrovirus (MERV)-related enhancers are enriched in 2C-like cells and nearly half of MERVL-LTR (Mt2_mm) elements exhibit enhancer activities, which have been mapped and characterized within the genome. Intriguingly,  enhancers identified in 2C-like cells augment the transcription in vivo when injected into the embryos. Furthermore, the deletion of enhancer regions in both 2C-like cells and embryos demonstrated their role in promoting the transcription of their adjacent downstream target genes, with one of the deletions impaired the early embryo development. This study enhances our understanding of the regulatory elements guiding ZGA.

overall_design: The ES-E14Tg2a cells were cultured on 0.1% gelatin-coated (07903, Stem cell) plates with standard LIF/serum medium. Doxycycline (Dox)-inducible DUX overexpression to induce 2C-like cells (2CLCs). UMI-STARR-Seq, ChIP-seq, RNA-seq and ATAC-seq were performed in mESCs (GFP negative) and 2C-like cells (GFP positive).

first: Lingyue

last: Yang


title: Functional dissection of enhancers in 2-Cell embryo with an in vitro model [RNA-seq]


title: Functional dissection of enhancers in 2-Cell embryo with an in vitro model [scRNA-seq]


title: Functional dissection of enhancers in 2-Cell embryo with an in vitro model [ChIP-seq]


title: Functional dissection of enhancers in 2-Cell embryo with an in vitro model [ATAC-seq]


title: Unique epigenomic signatures identify biologically significant subtypes of MDS and predict response to azacytidine [eRRBS]

city: Miami

first: Qin

state: FL

address: 1501 NW 10th Avenue

department: Sylvester Comprehensive Cancer Center

institute: University of Miami Miller School of Medicine

summary: Myelodysplastic syndromes (MDS) are characterized by genetic mutations affecting epigenetic modifiers and aberrant DNA methylation. Although DNA methyltransferase inhibitors (DMTis) are used to treat MDS, response variability remains a challenge, with limited predictive markers. Through integration of comprehensive genomic, epigenomic, and transcriptomic data, we have gained valuable insights into the intricate interplay between genetic and epigenetic alterations in MDS. We identified both hypermethylated and hypomethylated regions in MDS compared to normal, with a significant presence of long-range gene regulatory interactions. Moreover, distinct molecular subtypes enriched for specific mutations were characterized, providing critical insights into MDS biology. Notably, we identified approximately 2,500 differentially methylated regions (DMRs) at baseline that distinguish responders from non-responders to AZA treatment. Leveraging these DMRs and their associated genes within the same topologically associating domains (TADs), we developed robust classifiers integrating DNA methylation (DNAme), gene expression (GE), mutations, Hemoglobin, age, Platelet, Blast BM Blast, and absolute neutrophil count (ANC) data, highlighting the potential of personalized treatment approaches for MDS patients.

overall_design: Enhanced reduced representation bisulfite sequencing (ERRBS) of 90 MDS patients from CD34+ cells from bone marrow.


title: The Effects of Refined Carbohydrates on Systemic Immunity: Results from a Randomized Controlled Feeding Trial

city: New York

first: Ezequiel

middle: Carlos

last: Dantas

state: Y

address: 550E First Street

institute: NYU Langone

summary:
Objective
To explore the effects of three diets differing in amount and type of refined carbohydrates on peripheral immunity.
Methods
After a weight loss of 15% with a very low carbohydrate diet (VLCD), participants were randomized to isocaloric diets: VLCD diet; high carbohydrate diet with high starch content (HC-Starch); or high carbohydrate diet with high sugar content (HC-Sugar). Changes in peripheral immunity were assessed using a multimodal analyses before randomization and after 10 weeks of diet.
Results
Multiplexed cytokine analysis shows increased adipsin in HC-Starch and decreased adiponectin in HC-Sugar. Transcriptomics analysis shows upregulation of immune and metabolic pathways with HC-Starch, but reductions with HC-Sugar. CyTOF revealed significant shifts in NK cell populations and increased inflammatory markers in neutrophils and monocytes following consumption of HC-Starch.
Conclusions
Consumption of refined starch following weight loss on a VLCD increases circulating markers of immune activity, whereas sugar consumption reduces these markers.
Keywords: dietary nutrients, immunity, refined starch, added sugar, VLCD, cytokines, gene expression, CyTOF, inflammation, immune cells.
ClinicalTrials.gov registration: NCT03394664

overall_design:
Dietary nutrients impact immune function, with specific metabolic demands for different immune cells.This study explores the effects of three diets differing in amount and type of refined carbohydrates on the immune system during a period of sustained weight loss maintenance following a very low carbohydrate diet (VLCD).
After reaching 15.25% (±2.20%) weight loss using a hypocaloric VLCD, 54 participants aged 18–50 years with a BMI of 27 kg/m² or higher were randomly assigned to one of three isocaloric diets: a VLCD diet; a high carbohydrate diet with high starch content (HC-Starch); or a high carbohydrate diet with high sugar content (HC-Sugar) for 10 weeks. Changes in immunity were assessed before randomization and after 10 weeks of diet

first: Marcus

last: Goncalves


title: Unique epigenomic signatures identify biologically significant subtypes of MDS and predict response to azacytidine [RNA-Seq]

overall_design: To characterize gene expression changes occurring in MDS or in response to AZA treatment, RNA-seq data were obtained from 77 MDS patients and 6 healthy donors.


title: JAK inhibition results in aggressive cutaneous squamous cell carcinoma through MAPK pathway activation and shapes the tumor immune microenvironment to create novel therapeutic opportunities [RNA-seq]

city: Paris

first: Camille

last: Lobry

address: 16 rue de la Grange aux Belles

department: IRSL Inserm U944 / CNRS UMR7212

institute: Inserm

summary: Secondary malignancies are the most feared adverse effects of cancer therapies. In this study, we explore the oncogenic effects of the JAK inhibitor ruxolitinib on the skin. First, we demonstrate an epidemiological association between ruxolitinib exposure and development of aggressive cutaneous squamous cell carcinoma (cSCC), by conducting a disproportionality analysis on international pharmacovigilance data and a large local cohort study. We next demonstrate the causal link between ruxolitinib treatment and cSCC development, using in vitro and in vivo models. Mechanistically, we reveal that ruxolitinib induces transformation of pre-malignant keratinocytes into aggressive cSCC by activating the MAPK pathway through the COT kinase. Intriguingly, ruxolitinib also reshapes the immune microenvironment of cSCC resulting in T regulatory cells reduction and T CD8+ cells increase. Ruxolitinib-mediated immune microenvironment effects create new therapeutic opportunities by enhancing anti-PD1 treatment efficacy in cSCC and other immunotherapy resistant cancer types such as pancreatic cancer and acute myeloid leukemia.

overall_design: cSCC mice tumor treated with RUXOLITINIB or VEHICLE were subsequently lysed, total RNA extracted and RNA-sequencing libraries were prepared using TruSeq Stranded mRNA LT Sample Prep Kit. Sequencing librarie were sequenced on Illumina NovaSeq 600 sequencer.

first: Hélène

last: Pasquer

first: Lina

last: Benajiba

first: Emmanuelle

last: Latour


title: Chromatin Accessibility Analysis Reveals Regulatory Dynamics in hESC-Derived Kidney Organoids

city: guangzhou

first: chen

last: huan

address: guangzhou

institute: guangzhou laboratory

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Chromatin Accessibility Analysis Reveals Regulatory Dynamics in hESC-Derived Kidney Organoids [RNA-seq]

summary: Kidney organoid model can better simulate the development process, thereby providing an optimal platform for elucidating the mechanisms underlying human kidney development. In this study, we systematically characterized the chromatin availability dynamics and transcriptional regulation during the induction of human embryonic stem cells (hESCs) into kidney organoids (E-iKOs) for the first time.  We demonstrated that HNF1B is essential for nephrogenesis, as its deficiency results in absence of tubule structures. Furthermore, DOT1L, a histone H3K79 methyltransferase, is activated during E-iKOs induction and is predominantly responsible for opening pivotal loci and promotes the mesenchymal to epithelial transition (MET) required for kidney epithelial differentiation. Collectively, our study provides valuable resources and insights into the mechanistic study of human kidney development in vitro at the chromatin remodeling level.

overall_design: To determine the chromatin accessibility dynamics (CADs) during the E-iKOs induction, samples from seven time points (day 0, day 3, day 6, day 8, day 9, day 14, and day 16) were collected to perform both ATAC-seq and RNA-seq. And we collected the samples during E-iKOs induction using DOT1L KO cells  to explore the role of DOT1L.

first: Huan

last: Chen

first: Yapei

last: Yuan

first: Shangtao

last: Cao


title: Chromatin Accessibility Analysis Reveals Regulatory Dynamics in hESC-Derived Kidney Organoids [ATAC-seq]


title: Chromatin accessibility classification of TAD boundaries discloses new chromatin insulators (Hi-C 1)

city: Mexico City

first: Mayra

last: Furlan-Magaril

state: Mexico City

address: Circuito exterior s/n

department: Molecular Genetics

country: Mexico

institute: IFC-UNAM

summary: Perturbation of domain boundaries causes restructuration of domains and gene expression misregulation. Structural proteins such as CTCF and the cohesin complex have been identified enriched at domain boundaries. However, we still lack a complete understanding of the function of boundaries and what makes one boundary more effective at insulating genomic interactions than others. To understand how domains are structured, we experimentally classified boundaries according to their accessibility degree as a proxy of protein occupancy in K562 human cells. To determine if accessibility indeed reflects more robust boundaries and domains, we experimentally explored whether domain boundaries exhibit different degrees of accessibility in K562 cell line. To do so, we performed Hi-C experiments varying the incubation time of digestion with the restriction enzyme. The incorporation of this digestion time course into the Hi-C enables us to directly assess boundary accessibility and TAD formation from the same experiments.

overall_design: Hi-C experiments were performed varying the enzymatic restriction incubation time. We digested chromatin with HindIII enzyme for 0.25, 1, 3 and 12 hours to capture the most accessible boundaries at the shortest incubation times. After digestion, we continued the Hi-C experiment and sequenced the generated libraries.

first: Karina

last: Jácome-López


title: Tumor Cell JAGGED-1 Promotes Lymph Node Metastasis and Predicts Recurrence in Lymph Node Positive Breast Cancer Patients [MDA231-P_MDA231-LN]

city: Chicago

first: Naiche

last: Adler

address: 909 S Wolcott Ave, room 2160

institute: University of Illinois Chicago

summary: Purpose: Lymph node invasion is a hallmark of breast cancer disease progression, but current treatment strategies lack guidance from lymph node biomarkers. We investigated JAGGED1 (JAG1) as a promoter of lymph node metastasis and a prognostic biomarker in metastatic lymph node specimens.  Experimental Design: We used mouse models to assess the role of JAG1 expression in human and mouse breast cancer cells on lymphovascular invasion, lymph node metastatic potential, transcriptional profiles, and tumor interactions with lymphatic endothelium. We examined breast and lymph node samples from 284 breast cancer patients to determine the correlative and prognostic value of tumoral JAG1 expression in the lymph node. Results: In matched human breast tumor and lymph node samples, tumor cells that invaded lymph nodes showed higher JAG1 expression than their associated primary tumors (P value < 0.0001). In multiple models, breast cancer cells with high JAG1 expression showed increased lymphovascular invasion, lymph node metastasis, lymph node metastatic outgrowth, and migration through lymphatic endothelium. Transcriptomic analysis indicated that tumoral JAG1 regulates both juxtacrine and paracrine signaling pathways that induce inflammatory and pro-metastatic genes in lymphatic endothelium. When examining patients with identical surgical treatments, patients with lymph node JAG1 H-scorelow showed increased 5-year recurrence free survival rates than patients with lymph node JAG1 H-scorehi (87% vs 70%, P=0.016). Conclusions: JAG1 expression promotes lymphovascular invasion and lymph node metastasis in murine models. In patients, high expression of JAG1 in tumor cells in lymph node metastases predicts reduced recurrence free survival.

overall_design: RNA from MDA-MB-231 (samples labelled S1A,B,C) and MDA-MB-231-D3H2LN (samples labelled S2A,B,C) were transcriptionally profiled and analyzed for gene expression variances using bulk RNAseq strategies.

first: Gordon

last: Benjamin

first: Swaminathan

last: Bhairavi

first: Vadakath

last: Rahul

first: Teneqexhi

last: Pamela

first: Youn

last: Seock Won

first: Alvarez-Lopez

last: Isabel

first: Rezola

last: Marta

first: Xu

last: Ziqiao

first: Chen

last: Zhengjia

first: Er

last: Ekrem Emrah

last: LA

first: Caffarel

last: María Muñoz

first: Kitajewski

last: Jan


title: Chromatin accessibility classification of TAD boundaries discloses new chromatin insulators (Hi-C 2)

summary: Perturbation of domain boundaries causes restructuration of domains and gene expression misregulation. Structural proteins such as CTCF and the cohesin complex have been identified enriched at domain boundaries. However, we still lack a complete understanding of the function of boundaries and what makes one boundary more effective at insulating genomic interactions than others. Here we have classified domain boundaries by chromatin accessibility and found there is a collection of transcription factors (ZNF316, REST and EMSY among others) that occupy boundaries with or without CTCF. We functionally demonstrate that DNA-binding motif edition for ZNF316 leads to domain boundary disruption and increased inter-domain interactions, suggesting ZNF316 as a new factor structuring chromatin and insulating interactions between regulatory elements.

overall_design: Hi-C experiments were performed in K562 cell line edited by CRISPR. We generated a homozygous clone with a deletion including 612pb within a highly accessible boundary (Type I) without CTCF, disrupting the ZNF316 DNA binding site. We digested chromatin with DpnII enzyme. After digestion, we continued the Hi-C experiment and sequenced the generated libraries.


title: Tumor Cell JAGGED-1 Promotes Lymph Node Metastasis and Predicts Recurrence in Lymph Node Positive Breast Cancer Patients [hdLEC_JAG1_TLA]

overall_design: To assess JAG1/Notch LEC signaling, we seeded HdLECs on JAG1-coated (Samples labelled hdLEC_JAG1) or control FC-coated plates (Samples labelled hdLEC_Fc) and collected RNA after 6 hours which was subjected to mRNA sequencing.


title: Tumor Cell JAGGED-1 Promotes Lymph Node Metastasis and Predicts Recurrence in Lymph Node Positive Breast Cancer Patients [MDA231LN_JAG1_KO1_KO2]

overall_design: Wildtype MDA231-LN cells (samples labelled R) and two JAG1-KO clones (samples labelled E and F) were transcriptionally profiled and alayzed for gene expression variances using bulk RNAseq strategies.


title: Tumor Cell JAGGED-1 Promotes Lymph Node Metastasis and Predicts Recurrence in Lymph Node Positive Breast Cancer Patients [MDA231LN_JAG1_KO1cnt_KO1rescue]

overall_design: Transcription profiles of JAG1-KO MDA231-LN cells (Samples S3 A,B,C) were compared to JAG1-KO MDA231-LN cells transfected with JAG1-OE lentiviral plasmids (Samples labelled S4 A,B,C) and analyzed for gene expression variances using bulk RNAseq strategies.


title: The evolution of the type I IFN transcriptional response across human populations [PRO-seq]

city: Boulder

first: Robin

middle: D.

last: Dowell

state: CO

address: 596 UCB

department: Molecular, Cellular and Developmental Biology

institute: University of Colorado Boulder

summary: Host-pathogen dynamics have been observed to entail rapid evolution of genes involved in the host innate immune system. Type I interferon cytokines trigger an antiviral cellular state by binding to membrane receptors, encoded in chromosome 21, that transduce a signaling response to induce the transcription of hundreds of interferon-stimulated genes. However, it remains poorly understood how gene transcription regulation has been rewired through evolutionary time, even at relatively short time scales, to achieve inter- and intra-species variation in interferon-controlled changes in gene expression. We generated nascent transcription (PRO-seq) and steady-state mRNA expression (RNA-seq) datasets from human individuals from diverse ethnic backgrounds, and from individuals with trisomy 21, to investigate differences in the transcriptional response triggered by type I interferon within human populations, and to study the interferonopathy related to trisomy 21.

overall_design: We used lymphoblastoid cell lines derived from 10 human individuals: 7 unrelated individuals from diverse ethnic backgrounds; and 2 siblings, one with trisomy 21 and the other with disomy 21, as well as another unrelated individual with trisomy 21. PRO-seq datasets were treated for 1 hour with either BSA (negative control), or 100 ng/mL of human interferon beta. RNA-seq datasets were treated for 3 hours with either BSA (negative control), or 100 ng/mL of human interferon beta.

first: Daniel

last: Ramirez


title: Hyaluronan underlies the emergence of form, fate, and function in human cardioids

city: Vienna

first: Stefan

middle: M.

last: Jahnel

address: Dr. Bohrgasse 3

country: Austria

institute: IMBA

summary: The extracellular matrix (ECM) is crucial for organ development and disease. Yet, the interplay among cells, function, and the ECM during human cardiogenesis remains obscure. Using human cardioids, we discovered that cardiac mesoderm-synthesized hyaluronan (HA) underlies early cardiac functional development. HA drives cardioid cavity formation through hydrogel swelling and bioscaffolding, critical functions of the cardiac jelly in the early vertebrate heart. During an early developmental window, HA is essential for establishing cardiac cell identity, while at later stages, HA-generated forces promote beating function through mechanosensitive channels. Chamber-specific differences in mechanical sensitivity ensure robust contractions in multi-chambered tissues. Our findings reveal how a single endogenous ECM component orchestrates the co-emergence of form, fate, and function during human organogenesis, opening new avenues for bioengineering physiologically relevant organ models.

overall_design: RNA-seq profiling of WT, untreated, HAS2-KO and hyaluronidase-treated cardioids at d1.5, d2.5, d4.5 and d7.5

first: Anna

last: Dimitriadi

first: Julia

last: Kodnar

first: Vasileios

last: Gerakopoulos

first: Yajushi

last: Khurana

first: Maximilian

last: Mayrhauser

first: Tobias

last: Ilmer

first: Keisuke

last: Ishihara

first: Sasha

last: Mendjan


title: Caloric restriction suppresses colorectal tumors and melanoma through CD8 T cells in aged mice

status: Public on Jul 26 2026

city: Kumamoto City

first: Yuichi

last: Oike

state: Kumamoto

address: 1-1-1 Honko, Chuo-Ku

department: Department of Molecular Genetics

institute: Kumamoto University

summary: Caloric restriction (CR) has shown the potential to extend lifespan and reduce cancer risk; however, mechanisms underlying CR-mediated tumor suppression are not fully understood. Here, we investigate age-dependent CR effects on tumor progression and anti-tumor immune responses in a murine CR model. In aged mice, CR, defined as a 30% reduction in caloric intake, significantly suppressed tumor growth in murine syngeneic models of colorectal cancer or melanoma. CR also enhanced tumor infiltration by CD8+ T cells, which when depleted limited the tumor-suppressive effects of CR in aged mice. RNA-seq analysis of intratumoral CD8+ T cells revealed that CR upregulated expression of genes associated with T cell function. Furthermore, mechanistic studies of effects of CR on age-related changes in CD8+ T cells and immunohistochemical analysis suggested that normalization of the vasculature in the tumor microenvironment of aged CR mice is accompanied by decreased expression of angiogenic growth factors secreted by intratumoral CD8+ T cells. Our findings overall provide insight into age-dependent tumor-suppressive effects of CR and illustrate the essential role of CD8+ T cells in CR-mediated tumor suppression.

overall_design: Young (5-month-old) ad libitum-fed mice, aged (20-month-old) ad libitum-fed mice, and aged (20-month-old) caloric restriction-fed mice were subcutaneously inoculated with MC38 colon carcinoma cells. Twenty-one days after tumor implantation, tumor-infiltrating CD8⁺ T cells were isolated using magnetic-activated cell sorting (MACS). Bulk RNA sequencing was performed using three independent biological replicates from each group (young ad libitum, aged ad libitum, and aged caloric restriction; n = 3 per group, 9 samples total) to characterize transcriptional changes associated with aging and caloric restriction.

first: Taichi

last: Horino

first: Haruki

last: Horiguchi


title: HEK293 polysome profiling of ribosome-load remodeling by minocycline

city: San Diego

first: Khalyd

last: Clay

address: 10550 N. Torry Pines. Rd.

institute: Scripps Research

summary: Polysome-fraction RNA sequencing of HEK293 cells treated with vehicle control or minocycline. Cytoplasmic lysates were separated into monosome, light-polysome, and heavy-polysome fractions, ERCC spike-ins were added after fractionation and before RNA extraction, and fraction RNA was sequenced to quantify transcript ribosome-load state. The processed matrix reports control fraction distributions, fraction-enrichment classifications, polyEN, and mean ribosome load (MRL).

overall_design: Eighteen single-end libraries: vehicle/control and minocycline-treated HEK293 cells, each with three biological replicates split into monosome, light-polysome, and heavy-polysome fractions. The uploaded processed matrix supports reviewer recomputation of control MRL values.

first: Michael

last: Petrascheck


title: Multi-omics profiling links epigenetic and lncRNA changes to early human endochondral ossification priming

city: Lund

first: Paul

last: Bourgine

state: Skane

address: Sölvegatan 17

country: Sweden

institute: Lund University

summary: Differentiation programs remain incompletely understood across stem cell types, including for human bone marrow mesenchymal stromal/stem (BM-MSCs) cells, a heterogenous population orchestrating bone formation and establishing a functional hematopoietic niche in the bone marrow. BM-MSCs form and repair bone through the evolutionarily conserved process of endochondral ossification (EO), initiated by deposition of a transient cartilage template subsequently remodeled into bone and bone marrow tissues. Despite their considerable potential for skeletal regeneration, the early molecular and cellular events underlying BM-MSCs commitment to EO remain elusive. To overcome donor dependent variability in chondrogenic potential that limits mechanistic studies, we here exploit OssiGel as a potent chondro-inductive extracellular matrix offering robust recapitulation of EO by BM-MSCs. Through multi-omics profiling of OssiGel-primed BM-MSCs, we identify rapid chromatin remodeling at chondrogenic loci as concomitant for lineage commitment. The emergence of a chondro-progenitor population is detected as early as 3 days in vivo and correlates with successful EO recapitulation. Mechanistically, we identify LINC02511 as novel enhancer-associated element involved in the onset of EO. We confirm presence of LINC02511 in human skeletal atlases, and its CRISPR-mediated silencing significantly impaired EO. By integrating human tissue engineering strategies with single cell multi-omics profiling, our study provides a framework for deciphering BM-MSCs fate decisions, highlighting the role of enhancer and non-coding elements as key determinants of early lineage specification. These findings advance our understanding of BM-MSCs biology and will prompt their translational exploitation in regenerative medicine.

overall_design:
primary hBM-MSCs were expanded and primed towards endochondral ossificaiton in vivo, and single nulcei multiome sequencing was performed in vitro and 3 days after priming

***************************************************************
Raw files for human/patient samples were not submitted to GEO due to concerns about submitting personally identifiable sequence data for open access.
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first: David

last: Hidalgo Gil


title: Bulk RNA sequencing of human forebrain organoids following Aβ42 treatment

city: Shenzhen

first: Yilin

last: Feng

state: Please Choose

address: XILI Street

institute: Tsinghua University

summary: Human iPSC-derived forebrain organoids were exposed to amyloid-β42 (Aβ42) at day90 to model Alzheimer's disease-associated pathology. Bulk RNA sequencing was performed to characterize transcriptional changes induced by Aβ42 treatment. The resulting dataset provides genome-wide expression profiles of treated and untreated organoids.

overall_design: Human forebrain organoids were cultured under standard conditions and treated with either vehicle or 5 μM Aβ42 for 6 days. RNA was isolated and subjected to bulk RNA sequencing. Three independent biological replicates were analyzed for each condition.


title: CHI3L1 regulates gene methylation modifications in ovarian cancer cells

city: Shanghai

first: Yaodi

last: Shao

address: 2699 West Gaoke Road

institute: Shanghai First Maternity and Infant Hospital

summary: Adipocyte-tumor interaction plays a critical role in tumor microenvironment. Typically, high-grade serous ovarian carcinoma (HGSOC) preferentially metastasizes to the omentum, an adipocyte-rich tissue, and obesity correlates with worse patient prognosis. However, the underlying mechanisms remain poorly understood. Here, we identify a bidirectional adipocyte–cancer cell axis in which HGSOC cells stimulate lactate metabolism in adjacent adipocytes, leading to hypoxia-inducible factor 1α (HIF-1α) activation and subsequent secretion of chitinase 3-like 1 (CHI3L1). On the one hand, CHI3L1 acts as an inflammatory factor that promotes the secretion of additional cytokines, perpetuating omental inflammation, driving CD8⁺ T cell exhaustion, and shaping an immunosuppressive microenvironment that favors tumor metastasis. On the other hand, CHI3L1 binds to and stabilizes protein arginine methyltransferase 5 (PRMT5) in tumor cells, thereby enhancing its methyltransferase activity. This upregulation suppresses dual-specificity phosphatase 1 (DUSP1), a phosphatase of mitogen-activated protein kinase (MAPK), and sustains oncogenic MAPK signaling across multiple tumor models. Obesity amplifies this pathway by heightening HIF-1α activity and elevating CHI3L1 levels in adipose tissue, which drives both tumor progression and resistance to anti-angiogenic therapy. Pharmacological inhibition of CHI3L1 disrupts this signaling loop and restores therapeutic sensitivity, while also alleviating immune exhaustion in the tumor microenvironment. Collectively, our findings establish CHI3L1 as a key mediator of a unified mechanism that drives omental tropism and obesity-associated aggressive phenotypes, highlighting its potential as a prognostic biomarker and therapeutic target to improve anti-angiogenic therapy.

overall_design: Cut&Run assay targeting H4R3me2s in ovarian cancer cells


title: BPAF-induced inflammatory response via estrogen signaling: insights from in vivo and in vitro evidence

first: jiaying

last: li

address: 99 Shangda Road, Baoshan District, Shanghai

institute: Shanghai University

summary: This study investigated the effects of BPAF on the indicators related to inflammatory response and explored the roles of G protein estrogen membrane receptor 1 (GPER1) in this process. The results found that BPAF promoted the secretion of pro-inflammatory cytokines such as IL-1β, IL-8 and TNF-α, increased the protein expression of IL-1β and IL-6, and upregulated the relative mRNA levels of inflammatory related genes in Jurkat cells. In vivo SD rat experiments also found that BPAF exposure significantly upregulated the levels of serum inflammatory cytokines, elevated the relative mRNA expression of inflammatory related genes in immune organs, and induced damage of immune organs. In addition, BPAF upregulated the protein expression of GPER1 and elevated the relative mRNA expression of PI3K, AKT and MAPKs genes in Jurkat cells. GPER1 inhibitor G15 suppressed the upregulation of MAPK3, PI3K and AKT genes and concurrently inhibited the relative mRNA expression of inflammatory response genes, indicating that GPER1 activates the PI3K/AKT and MAPK signaling pathways and that this activation contributes to BPAF-induced inflammatory response at the transcriptional level. Molecular docking further confirmed that BPAF directly binds to GPER1. These results suggest that BPAF induces immunotoxicity by activating GPER1-mediated PI3K/AKT and MAPK signaling pathways.

overall_design: RNA-seq profiling was conducted on Jurkat cells treated with 0.1% DMSO or 1 μM BPAF for 12 h, with three biological replicates per group.

first: Jiaying

last: Li

first: Mengjie

last: Yu

first: Xiaoyu

last: Lu

first: Junmei

last: Xia

first: Fen

first: Shufang

last: Lou

first: Bingli

last: Lei


title: Myofiber-specific pathological programs in mTORC1-driven myopathy

city: Ann Arbor

first: Jun Hee

last: Lee

state: MI

address: 109 Zina Pitcher Pl

institute: University of Michigan

summary: Skeletal muscle contains diverse myofiber types and non-myocyte populations, but the molecular basis of many myopathies remains poorly defined because conventional methods cannot directly connect histopathology with gene expression at single-fiber resolution. We applied high-resolution Seq-Scope spatial transcriptomics to extensor digitorum longus and soleus muscles from a mouse model of mTORC1 hyperactivation. The analysis revealed fiber-type-specific pathological responses. Type I and IIa fibers showed limited morphological changes, whereas type IIx fibers followed distinct trajectories. In soleus, type IIx fibers exhibited abnormal enlargement associated with growth signaling, cytoskeletal remodeling, and impaired proteostasis. In extensor digitorum longus, type IIx fibers developed basophilia associated with increased RNA content and transcriptional signatures related to lipid, oxidative, and nucleotide metabolism. Type IIb fibers also displayed multiple transcriptional states. Macrophages and fibroblasts preferentially accumulated in soleus, where they contributed to an inflammatory and fibrotic microenvironment. These data link muscle histopathology with spatially resolved molecular states at single-fiber resolution.

overall_design: Extensor digitorum longus (EDL) and soleus (SOL) muscles were collected from four 10-week-old male Ckm-Cre Tsc1F/F Depdc5F/F mice and four Tsc1F/F Depdc5F/F littermate controls. One EDL and one SOL muscle from each mouse were analyzed, producing 16 muscle cross-sections in total. The same tissue sections were subjected to DAPI/WGA imaging, H&E histology, and Seq-Scope spatial transcriptomic profiling.

first: Jer-En

last: Hsu

first: Weiqiu

last: Cheng

first: Hyun-Min

last: Kang

first: Myungjin

last: Kim


title: CD36-mediated lipid-accumulation in reactive microglia drives retinal degeneration through Nlrp3-IL-1β pathway

first: Yang

last: Ziqi

address: Jinsui Road

institute: Zhongshan opythalmic center

summary: Lipid accumulation in microglia through CD36 enhances Nlrp3 inflammasome activation, contributing to retinal neurodegeneration.

overall_design: RNA-seq transcriptomic profiles were from HMC3 cells treated with oxLDL at concentrations of 0, 1, 2.5, 10, 50, or 100 μg/mL for 24 h, using Illumina NovaSeq platform.

first: Tian

last: Zhou

first: Ziqi

last: Yang

first: Hong

first: Biyan

last: Ni

first: Jingpeng

first: Minglu

last: Ma

first: Huaicheng

last: Wang

first: Peng

last: An

first: Huiyi

last: Xu

first: Xiaojing

last: Lin

first: Shiya

first: Shida

last: Chen

first: Lixia

first: Wei

last: Yi

first: Xialin

last: Liu

first: Chang

last: He


title: Multi-omics analysis of ion channels expressed by chicken embryo vestibular type I and type II hair cells

city: Pavia

first: Sergio

last: Masetto

address: Viale Golgi 19

department: Department of Brain and Behavioral Sciences

country: Italy

institute: University of Pavia

summary: Balance and gaze rely on the rapid and accurate detection and signalling of head movements by vestibular type I and type II hair cells. Signal transduction and transmission involve several types of ion channels, which are progressively acquired during hair cells differentiation, and whose identity is only partially known. In the present study, we adopted a multi-omics approach to identify the molecular nature of ion channels expressed by chicken vestibular hair cells between embryonic day (E)15 and E21, the day of hatching, when both type I and II hair cells appear functionally mature. Type I hair cells are characterized by the expression of a low-voltage activated outward rectifying K+ current, termed IKL. In adult mice, IKL is carried by the KV1.8 α-subunit, while it is still controversial if the KV7.4 α-subunit contributes to IKL in immature mouse type I hair cells. In this study, no effect of Linopirdine, a specific blocker of KV7 channels, was found in either immature or mature type I or II hair cells. Furthermore, whole-organ RT-PCR and transcriptome analysis at E16 and E21 showed no expression of the KV7.4 α-subunit transcript, while detecting the transcript for the KV1.8 α-subunit. Its expression in type I and II hair cells was finally confirmed by Patch-seq experiments. Further analysis of the transcriptomes of individual hair cells revealed the presence of transcripts for several other α and auxiliary subunits of Na+, K+ and Ca2+ channels, which are presumably responsible for the ionic currents recorded in both birds and mammals.

overall_design: Single cell RNAseq profiling of chicken embryo vestibula type I and II hair cells

first: Giulia

last: Cheli

first: Roberta

last: Giunta

first: Francesca

last: Stella

first: Eleonora

last: Guidi

first: Umberto

last: Laforenza

first: Mariarosa

last: Polimeni

first: Giancarlo

last: Russo

last: Borgo

first: Dejan

last: Lazarević


title: PAX3::FOXO1-Targeting PROTAC Induces Myogenic Differentiation of Fusion-Positive Rhabdomyosarcoma Cells

city: Washington

first: Nikola

last: Knoll

state: DC

address: 3700 O St. NW

institute: Georgetown University

summary: Fusion-positive rhabdomyosarcoma (FP-RMS) is characterized by the presence of tumor-specific chromosomal translocation products, most commonly PAX3::FOXO1, and typically results in lower survival rates compared to fusion-negative RMS cases. PAX3::FOXO1 plays a critical role in FP-RMS oncogenesis in both tumor initiation and maintenance, making it an excellent target for therapeutic intervention in FP-RMS. We created Proteolysis Targeting Chimeras (PROTACs) by combining PAX3::FOXO1 binding small molecules with E3 ligase recruiters for cereblon (CRBN) or S-Phase Kinase Associated Protein 1 (SKP1). PROTACs achieved degradation of endogenous PAX3::FOXO1 protein in FP-RMS cell lines in a concentration-, time-, and proteasome-dependent manner. Moreover, bulk RNA-seq revealed that PAX3::FOXO1-targeting PROTAC treatment impaired endogenous PAX3::FOXO1 target gene expression and induced myogenic differentiation. Importantly, treatment of FP-RMS cells with PAX3::FOXO1-PROTACs synergized with vincristine treatment and demonstrated an impairment in anchorage-independent growth in soft agar. Taken together, we demonstrate the proof of principle of PROTACs targeting the oncogenic fusion protein PAX3::FOXO1 in FP-RMS cells. PROTACs created in this study will not only be useful tools in studying PAX3::FOXO1 biology in laboratory models but could also serve as scaffolds for designing clinical-grade molecules to assess the therapeutic potential of PAX3::FOXO1-targeting PROTACs  in FP-RMS patients.

overall_design: RNA-seq profiling of RH41 fusion-positive rhabdomyosarcoma cells treated for 7 days with DMSO control or 5 uM G3_C (PFP-16) PAX3::FOXO1-targeting PROTAC.


title: CD36-mediated lipid-accumulation in reactive microglia drives retinal degeneration through NLRP3-IL-1β pathway

city: Guangzhou

first: Yang

last: Ziqi

summary: Microglia are central regulators of retinal immune homeostasis, yet their dysregulated responses can trigger neuroinflammation and drive retinal degeneration, a leading cause of irreversible blindness with limited therapeutic options. The specific pathogenic subsets and underlying mechanisms remain poorly defined. In this study, using single-cell RNA sequencing and spatial transcriptomics, we identified a distinct subset of lipid-accumulated reactive microglia (aLARM), predominantly localized to the ventral inferior retina in NaIO3-induced retinal degeneration mice, a region partially analogous to the human macula in terms of its susceptibility to neurodegenerative injury. In both patients and mouse models, this aLARM uniquely expresses high levels of CD36, which is critical for their formation and pathogenic function. Importantly, microglia-specific CD36 deletion abolished the aLARM-mediated retinal inflammation and degeneration, while subretinal transplantation of isolated CD36+ aLARM exacerbated the destruction of retinal structures and functional impairment. Furthermore, these aLARM subsets exhibited NLRP3 inflammasome activation and IL-1β production, which would activate IL-1R1 on microglia/macrophages and pericytes/ smooth muscle cells, amplifying a feed-forward inflammatory loop that worsened neurodegeneration. Finally, therapeutic blockade of CD36 using by the neutralizing antibody FA6-152 effectively reduced aLARM formationand protected against retinal degeneration. Together, our findings highlight CD36+ aLARM as a novel and targetable pathogenic microglial population that linksneuroinflammation to retinal degeneration, providing a potential foundation for microglia-based precision therapies in retinal degenerative disorders.

overall_design: To establish the retinal degeneration model, mice were injected intravenously with 20 mg/kg of NaIO3 and PBS as control. Mouse eyes enucleated at day 7 following NaIO3 or PBS treatment  were embedded together. Two eyeballs were placed into the same container with their optic nerves facing each other and then stored by freezing. Sagittal cryosections crossing the optic nerve head were obtained and then analyzed using spatial transcriptomics.

first: Tian

last: Zhou

first: Ziqi

last: Yang

first: Hong

first: Biyan

last: Ni

first: Jingpeng

last: Li

first: Minglu

last: Ma

first: Huaicheng

last: Wang

first: Peng

last: An

first: Huiyi

last: Xu

first: Xiaojing


city: GuangZhou

state: GuangDong

department: Zhongshan Ophthalmic Center

institute: Sun Yat-sen University

overall_design: Single cell profilings of retinal CD11b+ cells were from  NaIO3 induced retinal degeneration mice and PBS treated control mice, using 10X genomics platform. Spatial transcriptomics samples of eyeballs were from  NaIO3 induced retinal degeneration mice and PBS treated control mice, using 10X genomics platform.

last: Lin

first: Shiya

first: Shida

last: Chen

first: Lixia

first: Wei

last: Yi

first: Xialin

last: Liu

first: Chang

last: He


title: RNA-seq analysis of rat liver under drug treatment

city: Nanjing

first: Zhang

last: Sasa

address: Tongjiaxiang 24

institute: China Pharmaceutical University

summary: Epimedii Folium (EF) is widely used in traditional Chinese medicine, yet its hepatotoxic risk is increasingly recognized to be highly context dependent. This study demonstrates that EF induces significant liver injury only under conditions of immune stress, characterized by a pro-inflammatory and metabolically vulnerable hepatic state. Importantly, herb compatibility critically modulates this risk. Co-administration of Psoraleae Fructus (PF) markedly exacerbated EF-induced hepatotoxicity through amplification of inflammatory and cell death–related pathways, whereas Rhizoma Drynariae (RD) partially attenuated liver injury by stabilizing cholesterol and bile acid metabolism and restoring hepatic homeostasis

overall_design: 4-5 control vs. 4-5 treated samples, biological replicates

first: Hua

first: Kun

last: Hao

first: Sasa

last: Zhang

first: Hayaki H

last: Hamidu

first: Jiaqi

last: Sun

first: Shuyu

first: Jianheng

last: Zheng


title: Single-allele nanoscale mapping of regulatory variants [ATAC]

city: Oxford

first: Joseph

last: Hamley

address: John Radcliffe Hospital/Headley Way

department: Weatherall Institute of Molecular Medicine

country: United Kingdom

institute: University of Oxford

summary: Millions of genetic variants are linked to human disease but identifying underlying mechanisms is challenging because most variants lie within the non-coding genome. We developed a Micro Capture-C variant-to-function platform (MCCv) based on analysis of single-allele chromatin structure. This can identify changes in nanoscale chromatin architecture and link variants in cis-regulatory elements to associated genes. Furthermore, MCCv can phase other heterozygous variants within a locus to link regulatory variants to allelically imbalanced gene expression and directly read out variant effects on chromatin interactions following genome editing. With this approach, we investigated 405 cis-regulatory elements linked to immune-mediated inflammatory disease in CD4+ T cells. We uncover a previously undescribed gain-of-function mechanism, which increases risk of autoimmunity through creation of a neo-CTCF motif that blocks super-enhancer contacts with the SESN3 promoter. We show SESN3 regulates mammalian target of rapamycin (mTOR) by sensing tryptophan and demonstrate its role in autoimmunity using mouse models.

overall_design: Two technical replicates of ATAC with capture enrichment from healthy donor activated CD4+ T cells.

middle: C

first: James

last: Davies