title: Physiologically relevant media are associated with overlapping metabolic responses in primary human hepatocytes and Huh7 cells

city: Oxford

first: Leanne

last: Hodson

address: OCDEM, Churchill Hospital

department: OCDEM

country: United Kingdom

institute: University of Oxford

summary: Background and aims: The initiation and progression of metabolic dysfunction-associated steatotic liver disease (MASLD) is challenging to study in vivo in humans and robust in vitro high-fidelity disease models are limited.  Although primary human hepatocytes (PHH) are often considered to be the gold-standard, immortalized hepatic cell lines are often utilised because of their scalability and experimental tractability. Therefore, the aim of this study was to compare the metabolic responses of PHHs with our characterized Huh7-based model when exposed to the physiologically-relevant fatty acid (FA) mixtures. Methods: PHH and Huh7 cells were treated with 2% human serum and a combination of sugar and FAs enriched in either unsaturated (OPLA) or saturated (POLA) FAs for 4 or 7 days, respectively.  Stable isotope tracers were utilized to investigate basal changes in hepatocyte metabolism in response to different treatment regimes. Changes in cell viability, media biochemistry, intracellular metabolism, lipid droplet morphology and gene expression were quantified. Results:  Huh7 cells had greater viability than PHH, while NEFA uptake and triglyceride secretion were similar. Both OPLA and POLA increased the proportion of large lipid droplets in Huh7 cells, whereas only OPLA produced comparable effects in PHH. Despite higher baseline TG in PHH, both models showed similar lipid composition, de novo lipogenic responses, and glycogen levels. Compared to Huh7 cells PHH exhibited higher media 3-hydroxybutyrate, lower media lactate, reduced glucose uptake, and donor-dependent transcriptomic variability. Conclusions:  Our data demonstrate that Huh7 cells are metabolically adaptable and when cultured in physiologically relevant media metabolic readouts are more similar to those observed in PHH cells, thus making Huh7 a potentially useful workhorse model to investigate relevant pathways that may underpin the development of MASLD.

overall_design: Cell Culture: Huh7 and PHH cells were used. Huh7 cells were kindly provided by Dr Camilla Pramfalk (Karolinska Institute) and were cultured as previously described. PHH cells were obtained from Thermofisher (ThermoFisher, HMCPSQ) originating from both male and female donors (see supplementary data, Table 1 for donor characteristics. Frozen PHH cells were thawed in a 37°C water bath rapidly. 500µL of warm plating media (500mL Williams E Medium (ThermoFisher, A1217601) supplemented with a Primary Hepatocyte Thawing and Plating Supplements (ThermoFisher, CM3000) which contained FBS, dexamethasone, and a cocktail of penicillin-streptomycin, insulin, GlutaMAX™ and HEPES solution, was then added to the cryovial. Cells were then suspended and transferred to a falcon tube containing 4.5mL of warm plating media. Cell number and viability was then quantified using Trypan blue stain (ThermoFisher, 15250061) and a Cellometer Auto T4 Bright Field Cell Counter. Cells were then diluted with plating media and seeded into plates which had been pre-coated with collagen type I (ThermoFisher, A1048301) at a density of 35,000 cells/well for 96-well plates, 150,000 cells/well for 24-well plates or 50,000 cells/well for 12mm diameter glass cover slips for microscopy experiments. Both Huh7 and PHH cells were maintained at 37°C in 5% CO2. Experimental Treatments: Huh7 cells were seeded into relevant plates and cultured in experimental media comprised of 2% HS, 5.5mM glucose and 800µM of 4 FAs (oleic acid (O), palmitic acid (P), linoleic acid (L) and alpha-linoleic acid (A)) in one of two physiological ratios: OPLA (45:30:24:1) or POLA (44:45:10:1), or a no fat control as previously described. Huh7 cells were cultured in experimental media for seven days with media changes every other day as previously described. Following seeding, PHHs were cultured in plating media overnight then cultured in experimental media which consisted of Williams E Medium (ThermoFisher, A1217601) and Primary Hepatocyte Maintenance Supplements (ThermoFisher, CM4000) containing dexamethasone and a cocktail of penicillin-streptomycin, insulin, transferrin, selenium complex, BSA, GlutaMAX™, and HEPES solution, supplemented with 2% HS. PHH cells were cultured in 800µM OPLA and POLA or no fat control experimental media for four days with media changes every other day.

first: Eloise

last: Cross

first: Felix

last: Westcott

first: Kieran

last: Smith

first: Shilpa

last: Nagarajan

first: Fabio

last: Sanna

first: Kaitlyn

last: Dennis


title: RNA transcriptomics of MUP+HFD Tumors with oncogenic β-Catenin

status: Public on Jul 08 2026

city: Philadelphia

first: Abhinav

last: Illendula

address: 333 Cottman Avenue

country: USA

institute: Fox Chase Cancer Center

type: Expression profiling by high throughput sequencing

summary: This study analyzes RNA transcriptomics of tumors isolated from MUP+HFD mice with oncogenic β-Catenin 6.5 months after injection.

overall_design: Two CMR mice were started on high-fat diet (HFD) starting at 6 weeks. They were injected with AAV8-TBG-Cre to recombine expression of oncogenic β-Catenin at 8 weeks. 6.5 months later, macroscopic tumors were isolated from the livers for RNA isolation. A total of three tumors from each mouse were processed for total RNA sequencing.

first: Charles

last: Hewett

first: Yuki

last: Hayata

first: Joan

last: Font-Burgada


title: Folic acid alleviates X-ray irradiation-induced jaw malformation in zebrafish

city: Tsu

first: Yuhei

last: Nishimura

state: Mie

address: 2-174 Edobashi

department: Integrative Pharmacology

country: Japan

institute: Mie University Graduate School of Medicine

summary: X-rays are a form of ionizing radiation that has sufficient energy to remove electrons from atoms, thereby creating potentially harmful ions. X-ray irradiation during organogenesis can have profound detrimental effects, depending on the developmental stage and irradiating energy. In this study, we examined the impact of X-ray irradiation on zebrafish development at 36 h post-fertilization and observed marked jaw malformation, which was alleviated by folic acid pretreatment. Mechanistic studies revealed that folic acid pretreatment suppressed irradiation-induced production of reactive oxygen species. Transcriptome analysis performed at 24 h and 48 h post-irradiation revealed dysregulated expression of apolipoprotein A-IV a (apoa4a), methionine adenosyltransferase 1A (mat1a), heat shock protein 90 alpha class A member 1 tandem duplicate 1 (hsp90aa1.1), FKBP prolyl isomerase 5 (fkbp5), plac8 onzin related protein 3 (ponzr3), and prostaglandin E synthase 3a (ptges3a), which was not observed in zebrafish treated with folic acid-treated before the irradiation. These findings suggest that folic acid alleviates X-ray irradiation-induced jaw malformation in zebrafish, at least in part, by reducing oxidative stress and ameliorating dysregulated gene expression.

overall_design: Zebrafish embryos were randomly assigned into four groups; Control, X-rays (XR), Folic acid (FA), and FA+XR. Each group was incubated in a separate well of a 6-well plate. At 35 h post-fertilization (hpf), a freshly prepared sterile solution of FA was added to the FA and FA+XR embryo groups to give a final concentration of 200 µM, and the Control and XR groups received an equivalent volume of 0.3× Danieau’s solution. After 1 h preincubation, the 6-well plates for the XR and FA+XR groups were placed in the X-ray irradiation apparatus and exposed to XR (8Gy). At 1, 24, or 48 h post-irradiation, total RNA was extracted from each larva using Direct-TRI, and RNA concentrations were determined using a QuantiFluor RNA System Kit with a Quantus Fluorometer. Six larvae were used for each group at each time point. RNA was diluted to a final concentration of 1 ng/µl and a library was prepared for RNA-Seq. Sequencing of 150-bp paired-end reads was conducted using a HiSeq X Ten system. Quantification of gene expression was conducted using Danio_rerio.GRCz11.cdna.all.fa.

first: Aina

last: Higuchi

first: Miho

last: Hosaka

first: Makoto

last: Kashima


title: Palmitate drives mitochondrial and ER stress through disruption of the CD73-Adenosine axis

city: Augusta

first: Ana

middle: Carolina

last: Morandini

state: GA

address: 1120 15th Street

institute: Augusta University

summary: Bulk RNA sequencing was used to characterize transcriptional changes associated with the dysfunctional metabolic state in murine gingival fibroblasts (mGF). The mGF were treated with BSA control, palmitate, IL-1β, or palmitate plus IL-1β. Gene ontology analysis demonstrated enrichment of pathways related to innate immune activation, oxidative stress, mitochondrial dysfunction, ER stress, and purine metabolism. Palmitate disrupts the Cd73-adenosine axis while promoting mitochondrial dysfunction, oxidative stress, and Perk-mediated ER stress in gingival fibroblasts. Adenosine signaling protects against lipotoxic-induced ER stress, highlighting the relevance of the Cd73-adenosine pathway for  metabolic and inflammatory diseases.

overall_design: mGF were treated with BSA control, palmitate, IL-1β, or palmitate plus IL-1β, followed by bulk RNA sequencing.

first: S

last: Dawson

last: Batan

first: N

last: Adams

last: Bombin

first: ES

last: Ramos-Junior

first: AC


title: Expression landscape and circadian regulation of lncRNAs in the kidney [RNA-seq_circadian]

city: Lausanne

first: Leonore

last: Wigger

address: Amphipôle Building

country: Switzerland

institute: Swiss Institute of Bioinformatics

summary: Recent multi-omics work shows that the diurnal rhythmicity of kidney function is associated with circadian oscillations in renal mRNA, protein and metabolite abundances. Yet circadian regulation of renal lncRNAs, which are emerging as important modulators of diverse (patho)physiological processes, remains largely unexplored. Here, we provide a comprehensive characterization of renal lncRNA expression across cell types and circadian time points using bulk RNA-Seq and single-nucleus RNA-Seq (snRNA-seq) using kidney samples from male mice. Although the majority of lncRNAs are lowly expressed and only a fraction is annotated, their profiles sufficed to distinguish nearly all renal cell types. Compared with mRNAs, lncRNAs were more cell-type specific and an inverse correlation between cell-type specificity and expression level was evident. To assess the role of the circadian clock, we used mice lacking the core-clock regulator BMAL1 in renal tubules. Approximately one-sixth of lncRNAs and one-third of mRNAs were rhythmic. In contrast with mRNAs, lncRNAs exhibited an asymmetric temporal distribution between night and day, with peak expression occurring preferentially during the inactive phase of the mice. Bmal1 deletion partially disrupted rhythmicity in both biotypes and altered overall expression levels; lncRNAs were predominantly upregulated. These findings uncover the diversity of renal lncRNAs, their cellular distribution and their circadian regulation in male mice.

overall_design:
RNA was extracted from the renal tubules of 60 mice. Of these, 30 were conditional knockouts of Arntl (Bmal1) and 30 were of control genotype. They were housed under 12-hours light/12-hours dark cycles and were sacrificed at six different time points: zeitgeber time ZT 0, ZT 4, ZT 8, ZT 12, ZT 16, ZT 20 ( ZT 0 being the time of light on and ZT 12 the time of light off).  Five replicates per genotype and time point were included.     Technical note: The FASTQ file for sample 56 ("Renal tubule, cKO genotype, ZT12, rep 5") is  incomplete: The last 8201575 of 27714258 reads are missing. The original file was corrupted (truncated) at the sequencing facility due to a technical problem on a server cluster.    The raw data (FASTQ) files are the same as in GSE216252. This is a follow-up study. Processed data have been generated using a more recent annotation release and a different software pipeline.

***************************************************************
The table below lists GEO accessions reused/reanalyzed for this study.
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first: Dmitri

last: Firsov

first: Fanny

last: Durussel

first: Muriel

last: Auberson

first: Yohan

last: Bignon


title: Single Nuclei RNA-Sequencing of Mettl14 KO mouse islets

city: Boston

first: Hui

last: Pan

address: 1 Joslin Place

department: Bioinformatics and Biostatistics Core

institute: Joslin Diabetes Center

summary: Islet samples from a control and a Mettl14 KO mouse

overall_design: Islets were isolated from 4 months old males, cultured ON in 7mM RPMI and collected as a dry pellet snap frozen.

first: Dario

middle: F

last: De Jesus


title: Transcriptome of aTC6 cells transfected with Yy1-overexpressing construct

summary: We overexpressed Yy1 in mouse aTC6 cells for 72h

overall_design: aTC6 cells were seeded and after 24h were forward transfected with Yy1 overexpressing plasmid or a stuffer (empty) in 11mM DMEM at a confluency of 800K in a 1x6 well


title: Expression landscape and circadian regulation of lncRNAs in the kidney [RNA-Seq]

overall_design: RNAseq profiling was performed on kidney RNA samples from 16 mice. Of these, 8 were conditional knockouts of Arntl (Bmal1) in the renal tubule (cKOt), and 8 were of a control genotype (Ctrl). They were housed under 12-hours light/12-hours dark cycles and were sacrificed at two different circadian time points: zeitgeber time ZT 4 and ZT 16 (ZT 0 being the time of light on and ZT 12 the time of light off). Four replicates per genotype and time point were included.  The RNA libraries used for this experiment were a subset of those used in record GSE216252 by the same research group. They were re-sequenced with greater sequencing depth on a newer Illumina sequencer and analyzed separately.  The genotpyes and time points were the same in the single-nucleus RNASeq data set (GEO accession <>) , which was produced for the same study from a new set of mice.


title: Persistent transcriptomic changes following repeated exposure to wood smoke in non-human primate airway epithelial cells [RNA-Seq]

city: Davis

first: Hong

last: Ji

state: California

address: One Shields Avenue

department: California National Primate Research Center

institute: University of California, Davis

summary: Exposure to wildfire smoke is a significant public health concern with the respiratory system heavily impacted. Wildfire events are increasing in number and severity, and inhalation of wildfire smoke can lead to airway inflammation, oxidative stress-induced lung injury, impaired mucociliary clearance, and exacerbation of respiratory conditions such as asthma and chronic obstructive pulmonary disease (COPD). Despite these well-documented health risks, the molecular mechanisms underlying the respiratory effects of wildfire smoke exposure and recovery remain incompletely understood. In this study, we utilized transcriptomic and epigenomic analyses to investigate how exposure to wood smoke (WS) influences the expression patterns of gene networks in differentiated tracheobronchial epithelial cells from adult female rhesus monkeys. Our analysis identified exposure-induced differentially methylated regions, differentially expressed genes and gene networks that are implicated in lung disease, including dysregulation of immune responses, oxidative stress and cell death, compromise of epithelial barrier integrity and function, and epigenetic remodeling. Strikingly, significant transcriptomic changes were still detected one week after exposure cessation, enriched in pathways involved in inflammation, wound healing and tissue repair. Despite these transcriptomic and epigenomic perturbations, histology staining revealed no significant changes in epithelial tissue morphology following WS exposure. However, we found a significant number of pulmonary disease-associated genes and pathways whose transcription were affected by WS exposure. Our study enhances understanding of the molecular basis of wildfire smoke-induced respiratory effects and highlights the potential for WS to leave a lasting imprint on airway epithelium, with important implications for respiratory health in exposed populations.

overall_design: RNA-seq profiling of differentiated primary tracheobronchial airway epithelial cells from adult female rhesus macaques cultured at air–liquid interface and exposed to filtered air or diluted whole wood smoke for 6 h/day for 1 day, 5 consecutive days, or 5 days followed by 7 days of recovery.

first: Holmes

last: Evan

first: Brown

middle: P

last: Anthony

first: Elston

last: Sweeney

first: Royer

middle: M

last: Christopher

first: Ji

last: Hong


title: Persistent transcriptomic changes following repeated exposure to wood smoke in non-human primate airway epithelial cells [bisulfite-Seq]

type: Methylation profiling by high throughput sequencing


title: Multi-omics profiling reveals systemic rejuvenation of the aged kidney through senolytic therapy

city: Greifswald

first: Yao

last: Xu

state: State

address: Felix-Hausdorff-Straße 3

country: Germany

institute: University Medicine Greifswald

summary: Cellular senescence is a key driver of kidney aging, leading to functional decline and increased susceptibility to chronic kidney disease. While the senolytic combination of dasatinib and quercetin (D+Q) has shown promise in mitigating age-related pathologies, its long-term effects and underlying multi-level systemic mechanisms in the aging kidney remain poorly defined. Here, we systematically evaluated the long-term effects of D+Q in naturally aged mice using multi-omics approaches. We show that D+Q treatment reduces senescence markers (p16, p21, SA-β-gal), restores the anti-aging protein Klotho, and attenuates renal fibrosis and inflammation. Proteomic profiling reveals that D+Q enhances apoptotic clearance of senescent cells and promotes proliferative and regenerative pathways. Moreover, D+Q reactivates PPARα signaling, improves fatty acid oxidation, and reduces lipid accumulation in aged kidneys. Single-cell transcriptomics further demonstrates that D+Q reverses transcriptional aging signatures across multiple renal cell types and remodels cell-type-specific pathways associated with metabolism, inflammation, and fibrosis. Cell-cell communication analysis reveals that D+Q normalizes the hyperconnected intercellular network in aged kidneys, particularly by modulating inflammation-related signaling. Our findings offer a comprehensive, systems-level understanding of how senolytic therapy restores renal homeostasis, emphasizing its potential as a multifaceted intervention to combat kidney aging.

overall_design: This study included three groups of male mice: young (3/4-month), aged vehicle-treated (20-month), and aged mice receiving D+Q treatment (20-month). All animals were housed in individually ventilated cages in the Animal Center of Affiliated Nanshan Hospital of Shenzhen University under controlled environmental conditions, with a 12-h light–dark cycle and unrestricted access to standard chow and drinking water. Allocation to experimental groups was performed by random number tables, and investigators were kept blinded to group assignments during baseline characterization, administration of interventions and outcome assessments. At the experimental endpoint, mice were deeply anaesthetized by inhalation of isoflurane until breathing ceased and subsequently euthanized by cervical dislocation. Both kidneys were rapidly excised, decapsulated and either fixed or snap-frozen for histological, molecular, proteomic and single-cell RNA-sequencing analyses, as detailed in the following sections.


title: Single-cell RNA-seq profiles of intestinal mucosa in IBS-D rats to explore the mechanism of immune barrier regulation in pathogenesis

city: Guangzhou

first: Liyan

address: No. 232 Waihuan East Road, Higher Education Mega Center, Panyu District

country: China

institute: Guangzhou university of Traditional Chinese Medicine

summary: We have employed a single cell sequencing approach using 10x Genomics scRNAseq to study isolated endothelial cells, immune cells and stromal cells from the intestinal mucosa of the control group and diarrhea-predominant irritable bowel syndrome (IBS-D) rats. This study systematically deciphered the transcriptional landscape of the colonic mucosa in a rat model of IBS-D at single-cell resolution, revealing compositional shifts, transcriptional signatures, and potential regulatory networks among various cellular subpopulations, including epithelial, immune, and stromal cells.

overall_design: Following the establishement of neonatal maternal separation induced IBS-D model, we collected the colon sigmoid tissues from IBS-D model and normal rats. Then the intestinal mucosal cells were isolated and seqeunced by 10x Genomics.

first: Yongquan

last: Huang

first: Jiahe

last: Zhang

first: Jiaman

last: Lin

first: Zefang

last: Yang

first: Xiling

first: Fengbin

last: Liu

first: Qiuke

last: Hou


title: Hepatic transcriptomic changes with subchronic exposure to a dioxin mixture

city: Louisville

first: Banrida

last: Wahlang

address: 505 S Hancock Street

department: Medicine

institute: University of Louisville

summary: In this study, the primary focus was to determine changes in hepatic gene expression with exposure to a mixture of dioxins and dioxin-like compounds. This study addresses a critical knowledge gap by investigating sex-dependent effects from low-dose exposure to a mixture of dioxin and dioxin-like compounds in the context of sex differences and dietary patterns.

overall_design: Male and female C57BL/6J mice were fed either a low- (LFD) or high-fat diet (HFD) and administered a weekly dose of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD-10ng/kg), 3,4,7,8-pentachlorodibenzofuran (PeCDF-80ng/kg) and polychlorinated biphenyl 126 (PCB126-140ng/kg) or vehicle control. After 12 weeks, the study was terminated and samples collected for downstream analyses

first: Oluwanifemi Esther

last: Bolatimi


title: Protocol for Robust Genomic Mapping of Chromatin Targets Using High-Throughput CUT&RUN

city: Houston

first: Yue

last: Lu

state: TX

address: 1881 East Road

department: Epigenetics & Molecular Carcinogenesis

institute: The University of Texas MD Anderson Cancer Center

type: Genome binding/occupancy profiling by high throughput sequencing

summary: The genomic distribution of proteins that define chromatin state including transcription factors, chromatin modifiers and histone modifications, is critical for understanding gene regulation, cellular identity, and disease development. CUT&RUN assay is a powerful tool for high-resolution genomic mapping of factors that define the epigenome. Here, we present a high-throughput CUT&RUN protocol performed in 96-well plate format, from targeted DNA enrichment to high-quality library preparation, enabling reproducible, efficient, and robust genomic mapping of protein of interests and epigenomic profiling.

overall_design: CUT&RUN for histone marks and CTCF in MCF7 cells cultured in normal groth media.

first: Abhinav

middle: K

last: Jain


title: Identification of long non-coding RNAs involved in leukemogenesis and venetoclax response in acute myeloid leukemia through functional CRISPR-dCas9 interference screens

city: Solna

first: Bernhard

last: Schmierer

state: (optional)

address: Biomedicum 9B, Solnavagen 9

department: Medical Biochemistry and Biophysics

country: Sweden

institute: Karolinska Institutet

summary: Acute myeloid leukemia (AML) is a malignant hematologic disease with poor prognosis, and improved understanding of its biology is critical for patient outcomes. While protein-coding genes in AML are well characterized, the role of long non-coding RNAs (lncRNAs) remains largely unexplored. Here, we investigated how lncRNAs contribute to AML biology and treatment resistance. Three high-throughput lncRNA-CRISPR-interference (CRISPRi) screens were performed in the AML cell line MOLM-13, targeting 7,996 lncRNAs expressed in hematopoietic cells, with knockdowns directed to transcription start sites defined through cap analysis of gene expression (CAGE) sequencing. Effects on proliferation, differentiation, and response to the Bcl-2 inhibitor venetoclax were assessed. In total, 58, 4, and 23 lncRNAs were found to affect proliferation, differentiation, and venetoclax sensitivity, respectively. Three proliferation-associated lncRNAs (MIR17HG, CATG00000106133.1, and CATG00000056792.1) and one venetoclax-associated lncRNA (AC009299.3) were further investigated. CATG00000106133.1 was enriched in de novo and cytogenetically normal AML and correlated with NPM1 and IDH2 mutations. Gene Ontology and Reactome analyses of RNA-seq from CATG00000106133.1 knockout cells revealed roles in cytokine signaling and immune pathways. AC009299.3, linked to venetoclax response, was associated with poor outcome, adverse risk, and increased age in AML patients. Collectively, this study identifies four lncRNAs implicated in key leukemogenic processes, highlighting their potential for understanding AML biology, prognosis, and therapeutic response.

overall_design: Pooled CRISPRi screen. A custom sgRNA library was transduced into MOLM13 cells expressing dCas9-KRAB. Phenotypic enrichment was done by either cell proliferation, or cell differentiation followed by FACS sorting. Guide frequencies in the isolated populations were determined by NGS.

first: Elisabetta

last: Cozzi

first: Anne

last: Neddermeyer

first: Sophia

last: Milaria

first: Xiangfu

last: Zhong

first: Tyler

last: Weirick

first: Chung-Chau

last: Hon

first: Andreas

last: Lennartsson

first: Sören

last: Lehmann


city: Stockholm

address: Blickagången 16

department: the Department of Medicine

institute: Karolinska Institute

overall_design: HL60 cells were genetically modified using CRISPR-Cas9 ribonucleoprotein (RNP) complexes knockout of CATG00000106133.1, employing two guide RNAs (gRNAs) per deletion.


title: Dynamic interaction between tentacle-tethered microdomains in Integrator and NELF regulates the promoter-proximal pause checkpoint [TTchem]

city: Copenhagen N

first: Jesper

last: Svejstrup

state: Denmark

address: Blegdamsvej 3, 18.6

department: Department of Cellular and Molecular Medicine

country: Denmark

institute: University of Copenhagen

summary: NELF and Integrator are key regulators of promoter-proximal pausing and premature termination. Integrator subunit 12 (INTS12) contains a PHD domain within a flexible linker. While PHDs generally bind histones, we identify an interaction between the INTS12 PHD and a domain in the flexible C-terminal tentacle of NELF-A. Mutation of either domain disrupts the interaction, and although RNAPII-associated pausing complexes form, they are dysfunctional. Importantly, upon NELF-A mutation, interactions of the Integrator-containing pausing complex with transcriptional activators become apparent that correlate with increased pause-escape. Indeed, while INTS12 PHD loss functionally mimics Integrator loss, NELF-A mutation mimics loss of NELF, although the RNAPII pause position does not change. This supports a model in which Integrator is an integral component of the promoter-proximal pausing complex, and where INTS12’s PHD either interacts with NELF-A to pause/terminate transcription or enables interaction with activators and positive transcription regulators to allow pause-escape.

overall_design: TTchem-seq. Nascent transcript analysis following endogenous INTS12 knockdown and replacement of WT PHD mutated INTS12 forms.


title: Dynamic interaction between tentacle-tethered microdomains in Integrator and NELF regulates the promoter-proximal pause checkpoint

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

overall_design: Refer to individual Series


title: Whole transcriptome sequencing of HCT116 cells with altered DNA repair capacities treated with RTX or 5FdUR

city: Budapest

first: Angela

last: Bekesi

address: Műegyetem rakpart 3.

department: Department of Applied Biotechnology and Food Science

country: Hungary

institute: Budapest University of Technology and Economics, Faculty of Chemical Technology and Biotechnology

type: Non-coding RNA profiling by high throughput sequencing

summary: Thymidylate synthase (TS) is a key enzyme in thymidylate biosynthesis and an established target of chemotherapeutics such as 5-fluoro-2’-deoxyuridine (5FdUR) and raltitrexed (RTX). Inhibition of TS disrupts the dUTP:dTTP balance, leading to uracil misincorporation, futile base excision repair cycles, DNA strand breaks, and ultimately cell death. However, active U-DNA repair is apparently not required for the cytotoxic effect of these drugs, indicating that alternative pathways contribute to the observed S-phase arrest. To dissect these mechanisms, we investigated the transcriptomic effects of TS inhibition in HCT116 colon cancer cell lines with altered DNA repair capacities treated with 5-fluoro-2′-deoxyuridine (5FdUR) or raltitrexed (RTX). Both drugs induced similar DNA damage responses and S-phase arrest, yet displayed distinct transcriptional signatures. Our findings underscore the multifaceted impact of TS inhibition and reveal drug-specific differences in cellular responses.

overall_design: Three cell lines were used: HCT116 colon cancer cells, which are mismatch repair deficient (HCT116); the HCT116_UGI cells, which stably express the UGI transgene (an inhibitor of the major uracil-DNA glycosylase, UNG); and the HCT116_MMR_UGI cells, in which mismatch repair was restored by the introduction of a healthy third chromosome. Cells were treated for 48 hours with drugs (100 nM RTX or 20 µM 5FdUR) or left untreated (NT). Total RNA was isolated using TRIzol reagent (Invitrogen) according to the manufacturer's protocol. RNA quality was assessed by agarose gel electrophoresis, and concentration was measured using a NanoDrop 2000C UV-Vis spectrophotometer (Thermo Fisher Scientific). Following size selection, stranded RNA sequencing libraries for short RNAs (<200 nt) and long RNAs (>200 nt) were prepared separately. Small and long RNA libraries were sequenced on a Novaseq 6000 platform using Illumina 50 pb single-end (20M reads) and 150 paired-end (150M reads) protocols, respectively. Library preparations, including rRNA depletion and quality control, as well as the sequencing, were performed by Novogene Co., Ltd. (China) as a whole-transcriptome sequencing service.

first: Eszter

last: Holub

first: Anna

last: Felföldi

first: Milda B

last: Szajkó

first: Gábor

last: Papp

first: Beáta

last: G. Vértessy

first: Angéla

last: Békési


title: Dynamic interaction between tentacle-tethered microdomains in Integrator and NELF regulates the promoter-proximal pause checkpoint [eNET-seq]

overall_design: eNET-seq in INTS12 WT and INTS12 KO. 2x replicates


title: Dynamic interaction between tentacle-tethered microdomains in Integrator and NELF regulates the promoter-proximal pause checkpoint [PRO-seq]

overall_design: PRO-seq analysis following siRNA-mediated knockdown of either INTS12 or NELFA, followed by re-expression of INTS12 PHD finger and NELFA R453A mutants.


title: Generation and single-cell characterization of functional megakaryocytes derived from umbilical cord blood

city: Hangzhou

first: Yanmin

last: He

state: Zhejiang

address: 789 Jianye Road

department: Institute of Transfusion medicine

institute: Blood center of Zhejiang Province

summary: The stable culture system derived from umbilical cord blood (UCB) was established, which differentiates into megakaryocytes (MKs)in vitro and the transcriptome data of UCB-derived MKs produced by our system was analyzed.

overall_design: Here, the cultured  cells were collected and subjected to single-cell RNA sequencing(sc-RNAseq) using the 10×Genomics Chromium platform.  The subpopulations of UCB-derived MKs were further analyzed based on the captured of 8,064 cells.

first: Yue

last: Liang

first: Ji

first: Yu

last: Shen

first: Zhipan

last: Wu

first: Shanshan

last: Pei

first: Faming

last: Zhu


title: BCKDK Protects against Obesity-Induced Cardiac Remodelling and Dysfunction by Modulating Mitochondrial ROS-Driven JNK/p38 MAPK Signaling

city: Nanjing

last: Liu

state: Jiangsu

address: No.87 Dingjiaqiao Street

institute: Southeast University

summary: Obesity cardiomyopathy (OCM), characterized by diastolic dysfunction, often progresses to obese heart failure with preserved ejection fraction and remains a major clinical challenge. Accumulating evidence indicates that impaired branched-chain amino acid (BCAA) catabolism is present in OCM, and systemic enhancement of BCAA catabolism through global inhibition of branched-chain ketoacid dehydrogenase kinase (BCKDK), a key negative regulator of BCAA oxidation, alleviates cardiac dysfunction. However, whether cardiac-restricted manipulation of BCAA catabolism is sufficient to confer therapeutic benefits remains unknown. Here, we generated an inducible cardiomyocyte-specific BCKDK knockout mouse model to enhance cardiac BCAA catabolism. Unexpectedly, this model fails to improve and instead exacerbates cardiac dysfunction and ventricular remodelling in OCM. Notably, cardiac BCKDK expression is reduced in both OCM mice and obese patients. Cardiomyocyte-specific BCKDK overexpression robustly improves cardiac function and pathological remodelling in OCM, accompanied by further lowering myocardial BCAA levels. Bulk-RNA sequencing and untargeted metabolomics reveal that the cardiomyocyte-specific BCKDK overexpression restore mitochondrial respiratory function and suppress MAPK-driven inflammatory response. Overall, these findings uncover an unanticipated BCAA-independent role of BCKDK in maintaining cardiomyocyte mitochondrial function and restraining inflammatory signalling, positioning BCKDK as a promising therapeutic target for OCM.

overall_design: We performed RNA sequencing on left ventricular tissues from ND+AAV-CON, HFD+AAV-CON, and HFD+AAV-BCKDK mice (3 biological replicates per group).


title: Non-ablative fractional laser 1940nm treatment series modulates epigenetic pathways and signs of skin aging: a split face investigation

city: London

first: Harry

last: Pink

address: Innovation and Translation Hub, 84 Wood Lane,

institute: Mitra Bio

summary: Skin aging is driven by cumulative environmental damage and epigenetic dysregulation, yet it is unclear whether energy-based rejuvenation therapies can durably remodel the cutaneous methylome. To address this, we conducted a split-face, paired longitudinal study in 22 adults treated with a 1940-nm non-ablative fractional laser (NAFL). Epidermal samples were collected by tape stripping from treated and contralateral control sites at baseline, immediately after the first treatment, and at 1, 3, and 6 months. Enzymatic methyl-sequencing profiled ~3.8 million CpGs per sample; clinical endpoints included pigmentation, texture, and global aesthetic scores. No significant differentially methylated regions (DMRs) were detected immediately post-treatment, but 635 DMRs emerged from 1 month after the final session, expanding through 3 months and stabilizing by 6 months. These loci were enriched for pathways involved in epidermal differentiation, collagen organization, wound response, and stem-cell maintenance, and showed selective resetting at Polycomb-regulated and WNT-signaling genes. NAFL reversed age-associated methylation change at >83% of age-linked CpGs and engaged gene sets previously implicated in molecular rejuvenation. Treatment also induced transient, treatment-specific DMRs related to immune and stress responses. Epigenetic remodeling paralleled significant clinical improvements, indicating that 1940-nm NAFL drives durable molecular rejuvenation rather than transient repair.

overall_design: In a randomised split-face study, one side of the participants forehead was treated with 1940nm non-ablative fraction laser (NAFL). Samples were taken at baseline (before treatment), and from both treated and untreated sides of the forehead at 4 timepoints; post-treatment 1 (after 1 out of 3 NAFL treatments), 1-month follow up (1MFU), 3MFU & 6MFU.

first: Konika Patel

last: Schallen

first: Kevin

last: Schomacker

first: Cristiana

last: Banila

first: Nicolle

last: Dest

first: Katherine

middle: L

last: Coleman


title: SARS-CoV-2 spike protein exerts an anti-cancer effect by regulating MEG3 and BCYRN1 in A549 cells

city: Seoul

first: Hyo-Jeong

last: Lee

state: --- Select One ---

address: 26, Kyungheedae-ro,Dondaemun-gu, Seoul, 02447, Korea

country: South Korea

institute: Kyung Hee University

summary: Background Few studies have reported that severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein suppresses cancer cell growth. Here, we investigated the effect of the SARS-CoV-2 spike protein in A549 lung cancer cells using recombinant spike protein (SP), spike protein transfection (SPT), and pseudo-SARS-CoV-2 virus (PSV). Methods To evaluate its anticancer activity, we performed RNA sequencing, colony formation, immunofluorescence, cell viability, migration, FACS, western blotting, 3D spheroid, molecular docking, siRNA,qRT-PCR, and apoptosis assays. Results Spike protein significantly inhibited the long-term growth of A549 cells. Among the delivery methods, PSV showed the most potent anticancer effect, followed by SPT and SP, as evidenced by suppressed migration, spheroid growth, and sub-G1 arrest. RNA-Seq revealed distinct expression profiles between control and SP-treated cells, identifying two differentially expressed lncRNAs: MEG3 was upregulated, while BCYRN1 was downregulated. Knockdown of BCYRN1 or overexpression of MEG3 reproduced the spike protein–induced antiproliferative and pro-apoptotic effects, including enhanced cleaved caspase-3 expression. Docking analysis suggested potential binding of spike protein to these lncRNAs. To determine whether spike protein affects their stability, transcription was inhibited using actinomycin D (ActD). Spike protein attenuated the ActD-induced decrease of MEG3, indicating enhanced stability, whereas it accelerated BCYRN1 reduction. Conclusion Collectively, these findings demonstrate that SARS-CoV-2 spike protein exerts antitumor activity in A549 lung cancer cells by differentially regulating the stability of lncRNAs MEG3 and BCYRN1.

overall_design: RNA-seq profiling of A549 human lung cancer cells treated with SARS-CoV-2 spike protein for 24 h compared with untreated control cells.

first: Jung-Eun

last: Kim

first: Hengmin

last: Han

first: Leeseon

last: An

first: Eunseo

last: Park


title: Repeated TLR7 Activation Increases Alcohol Consumption and Induces Cell Type-Specific Transcriptome Changes in Mice

city: Austin

first: Roy

middle: Dayne

last: Mayfield

state: Texas

address: 2500 Speedway, MBB 1.124

department: Neuroscience

institute: University of Texas at Austin

summary: Alcohol consumption triggers neuroinflammation, potentially creating a feed-forward loop that increases drinking. Previous studies have shown that activation of the toll-like receptor 7 (TLR7) leads to escalated drinking. In this study, we aim to identify cell type-specific transcriptomic patterns linking TLR7-induced neuroinflammation to drinking. Therefore, male C57BL/6J mice were treated with the selective TLR7 agonist R848 every-other day for 20 days in total. After 10 treatment-free days, brains were collected for single-nucleus RNA-Sequencing (snRNA-Seq) in the medial prefrontal cortex (mPFC) and central amygdala (AMG). The AMG showed a greater number of differentially expressed genes (DEGs), primarily in inhibitory and excitatory neurons. Among glial cells, AMG astrocytes exhibited the greatest number of DEGs, which were involved in blood-brain barrier (BBB) regulation (e.g., Cldn5, Mecom, Nrg1), a finding supported by secondary validation using Xenium in situ spatial transcriptomics on AMG-containing sections. BBB-regulatory genes, including those in the Wnt signaling pathway (e.g., Notch3, Top2a, Aldoc), were altered across multiple cell types in both regions. These findings suggest that repeated TLR7 activation induces persistent BBB dysfunction, which may contribute to escalated alcohol consumption.

overall_design: snRNA-seq from dmPFC and AMG of mice repeatedly treated with the TLR-7 agonist R848 or saline

first: Marion

last: Friske

first: Riccardo

last: Barchiesi

first: Nihal

middle: A

last: Salem

first: Ruth

last: Allard

first: Anna

middle: C

last: Dobre

first: Wen

last: Chen

middle: D


title: Colon-restricted Pten haploinsufficiency models PI3K pathway-driven invasion in human colorectal cancer

city: Hiroshima-shi

first: Hikaru

last: Nakahara

address: kasumi 1-2-3, minami-ku

institute: Hiroshima University

summary: Background and Aims: The multistep accumulation of driver gene mutations is associated with early-stage colorectal tumorigenesis. To observe the phenotype-genotype correlation, we developed a series of genetically engineered mouse models based on Apc inactivation led by CDX2P-Cre and found invasive adenocarcinomas developed by inducing the loss of one Pten copy with Apc inactivation in the colonic epithelium. Here, we aimed to study the mechanisms underlying Pten haploinsufficiency and its clinical relevance. Methods: Tumor number, volume, and histology were compared between CDX2P-Cre;Apcflox/+ (CPC;Apc) and CDX2P-Cre;Apcflox/+;Ptenflox/+ (CPC;ApcPten) mice, and a multi-omics analysis was used to investigate the mechanism of the invasive phenotype in CPC;ApcPten mice. Rapamycin was administered to tumor organoids and in vivo to evaluate its dependency on the AKT-mTORC1 pathway. We also performed phylogenetic analysis of tumor evolution during human colorectal carcinogenesis using multiple targeted biopsies. Results: A greater number of tumors developed in CPC;ApcPten mice, half of which invaded the submucosal layer. Although both Apc alleles were inactivated by Cre-LoxP recombination and loss of heterozygosity in tumors, a wild-type Pten allele remained in the tumors of CPC;ApcPten mice, suggesting the haploinsufficient effect of Pten on tumorigenesis. Loss of one Pten copy decreased Pten expression levels and induced downstream activation of the AKT-mTORC1 signaling pathway in CPC;ApcPten mice. Rapamycin administration markedly inhibited tumorigenesis in CPC;ApcPten and CPC;Apc mice. In clinical biopsies, PI3K genetic alteration was predominantly observed during the transition from tubular adenoma to adenocarcinoma. Conclusion:The effect of Pten haploinsufficiency on colonic tumor formation in mice reaffirms the significance of PI3K alterations in tumor evolution in human colorectal cancers.

overall_design: RNA sequencing were performed using colonic tumors from CPC;ApcPten (n =3) mice compared with those in CPC;Apc mice as control (n = 3).

first: Hiroaki

last: Niitsu

first: Haruki

last: Sada


title: The Rho effector ARHGAP18 coordinates a Hippo pathway feedback loop through YAP and Merlin to regulate the cytoskeleton and epithelial cell polarity.

city: Buffalo

first: Jonathan

middle: E

last: Bard

state: New York

address: 701 Ellicott Street

department: Genomics and Bioinformatics Core

institute: University at Buffalo

summary: The organization of the cell’s cytoskeletal filaments is coordinated through a complex symphony of signaling cascades originating from internal and external cues. Two major actin regulatory pathways are signal transduction through Rho family GTPases and growth and proliferation signaling through the Hippo pathway. .  These two pathways define the actin cytoskeleton, controlling foundational cellular attributes such as morphology and polarity and are hijacked to promote proliferation and motility in aggressive cancers. In this study, we use human epithelial cells to investigate the interplay between the Hippo and Rho Family signaling pathways, which have predominantly been characterized as independent actin regulatory mechanisms.  We identify that the RhoA effector, ARHGAP18, forms a complex with the Hippo pathway transcription factor YAP to address a long-standing enigma in the field. Using super resolution STORM microscopy, we characterize the changes in the actin cytoskeleton, on the single filament level, that arise from CRISPR/CAS9 knockout of ARHGAP18. We report that the loss of ARHGAP18 results in alterations of the cell that derive from both aberrant RhoA signaling and inappropriate nuclear localization of YAP. These findings indicate that the Hippo and Rho family GTPase signaling cascades are coordinated in their temporal and spatial control of the actin cytoskeleton.

overall_design: RNAseq profiling of WT Jeg-3 cells and their knockout derivitives of ARHGAP18

first: Emma

last: Murray

first: Gillian

last: Hodge

first: Khanh

last: Pham

first: Leighton

middle: S

first: Cameron

last: Mitchell

first: Yongho

last: Bae

first: Andrew

middle: T

last: Lombardo


title: MDI1228, a Topical Pan-JAK Inhibitor, Disrupts Dermal Fibroblast-T cell Chemokine Crosstalk to Resolve Autoimmune Skin Diseases

city: Xiamen

first: Lingjuan

last: Zhang

address: Xiangan South Street

institute: Xiamen University

summary: Delayed typehypersensitivities are driven by distinct T cell programs and are limited safe long-term topical therapies. Dermal fibroblasts (dFBs) have emerged as active immunomodulators, but whether they can be therapeutically targeted remains unexplored.  Here we developed MDI1228, a novel topical pan JAK inhibitor with nanomolar potency against JAK1/2/3/TYK2 (IC50 0.11-0.85 nM) and high selectivity. Topical MDI1228 ameliorated both DNFB induced ACD and MC903 induced AD in mice, reducing T cell infiltration and cytokine production. Mechanistically, MDI1228 not only directly inhibits T cell activation and cytokine production but also disrupts fibroblast-T cell crosstalk by reducing dFB-derived chemokine expression. Single cell transcriptomics identified dFBs as the primary source of CXCL9/10 in ACD and CCL2 in AD. Conditioned medium and neutralization experiments demonstrate that CXCL9/10-CXCR3 and CCL2-CCR2 signaling axes contribute to T cell polarization in a context-dependent manner. Compared with glucocorticoids, prolonged topical application of MDI1228 shows minimal systemic toxicity and preserves tissue homeostasis. These findings identify dFBs as a central therapeutic node and demonstrate that MDI1228, by directly targeting T cells and disrupting dFB derived chemokine axes via JAK inhibition, offers a potent and safe topical treatment for both ACD and AD.

overall_design: Single-cell suspensions were prepared from mouse dorsal skin samples as previously described (6). Briefly, skin was minced and digested with 0.25% collagenase IV (Gibco) and 0.1% DNase I (Sigma) for 1 hr at 37°C. The resulting cell suspension was filtered through a 40 μm strainer, and red blood cells were removed using ACK lysis buffer. Viability (>85%) was confirmed by trypan blue exclusion. Libraries were constructed using the Chromium Next GEM Single Cell 3ʹ Reagent Kit v3.1 (10x Genomics) according to the manufacturer’s protocol. Sequencing was performed on an Illumina NovaSeq 6000 platform with a target depth of 50,000 reads per cell. Raw sequencing data were processed with Cell Ranger (v6.0.0) using the mouse reference genome (mm10-2020-A). Downstream analysis including quality control, dimensionality reduction, clustering, and marker gene identification was performed using the Seurat package (v4.3.0) in R. Cells with <200 or >6000 detected genes, or >10% mitochondrial reads, were excluded. Clusters were annotated based on canonical marker genes (e.g., Pdgfra for fibroblasts, Cd3e for T cells, Cd68 for macrophages). Cell-cell communication was inferred using CellChat (v1.6.0) with default parameters.

first: Ling-juan


title: Dek Loss Induces Sex-Dependent, Task-Specific Cognitive Deficits and Reprograms the Hippocampal Transcriptome in Mice

city: Cincinnati

first: Taylor

last: Lange

state: OH

address: 240 Albert Sabin Way

department: CBDI

institute: Cincinnati Children's Hospital Medical Center

summary: Cognitive decline with aging, and some neurodegenerative conditions like Alzheimer’s disease, disproportionately affects females yet few mechanisms beyond steroid hormone signaling fully explain this sex-specific vulnerability. The chromatin-remodeling DEK protein, upregulated by estrogen and progesterone and broadly expressed in the brain, including the hippocampus, may be one such mechanism. We have previously linked DEK loss with indices of neuronal dysfunction, including increased DNA damage, impaired neurite development, and apoptosis, suggesting a potential neuroprotective role. Here, we investigated the molecular and behavioral consequences of Dek loss in vivo. Female Dek constitutive knockout (cKO) mice exhibited a sex-specific behavioral phenotype, with impairments in sensorimotor gating, as measured by pre-pulse inhibition, and in reversal learning in the Morris Water Maze. These findings are suggestive of deficits in pre-attentive sensory processing and cognitive flexibility, respectively. Notably, these cognitive deficits were not observed in male Dek cKO mice and were not attributable to differences in general learning ability, locomotor activity, or anxiety-like behavior. The absence of impairment in object recognition and conditioned fear learning and memory in females suggests that the effects of DEK loss are task-specific and likely brain region-specific. Transcriptomic analysis of hippocampal tissue revealed differentially expressed genes related to inflammation, metabolism, and neuropeptide signaling in all Dek-cKO mice, along with a distinct female-specific transcriptomic profile indicative of impaired neuronal function. Combined, we report for the first time that DEK supports certain aspects of cognitive function, particularly in females. These data may be relevant for understanding sex differences in some cognitive disorders.

overall_design: We performed gene expression profiling of mouse hippocampus with and without DEK knockout using bulk RNA-seq

first: Kaitlyn

last: Gardner

first: Marla

last: Perna

first: Susanne

middle: I

last: Wells

first: Michael

last: Williams

first: Charles

middle: V

last: Vorhees

first: Matia

middle: B

last: Solomon

first: Lisa

last: Privette Vinnedge


title: Deciphering resistance mechanisms to auxin-inducible protein degradation in mammalian cells

city: Memphis

first: Beisi

last: Xu

state: Tennessee

address: 262 Danny Thomas Pl

department: Center for Applied Bioinformatics

institute: St Jude Children's Research Hosipital


title: Deciphering resistance mechanisms to auxin-inducible protein degradation in mammalian cells [ChIP-Seq]

summary: Targeted protein degradation is a favorable strategy for studying the immediate downstream effects of protein loss-of-function, such as the auxin-inducible degron (AID) system. Although this system has been applied extensively to cell and animal models, degradation resistance to long-term auxin treatment has not been studied. With the advent of the new AID2 system, cellular toxicity caused by the high auxin concentrations required in the original AID1 system is no longer a concern, enabling the study of protein degradation over extended periods. In this study, we derived multiple miniAID-tagged knock-in human cell lines and a Ctcf-miniAID knock-in mouse strain to investigate mechanisms of degradation resistance. We revealed four independent resistance mechanisms, including a nonsense mutation in the CTCF coding sequence that removed the miniAID peptide, a missense point mutation in the miniAID coding region that disrupted ubiquitin complex targeting, and reduced expression of OsTIR1 adaptor protein. Resistance to auxin degradation was also observed in mouse primary CtcfminiAID/miniAID knock-in B-cell acute lymphoblastic leukemia cells through missense mutations of the OsTIR1(F74G) protein ex vivo and in vivo. By systematically characterizing degron resistance mechanisms in mammalian cells, we identified potential limitations of the AID system for long-term protein degradation studies.

overall_design: To assess the chromatin binding affinity of a mis-sense mutation form of CTCF identified from auxin treatment resistance, we ectopically expressed CTCF-WT and CTCF-Q666* mutant forms in the doxycycline inducible vector TRE3G in the CTCF-miniAID knockin cell lines. Doxycycline and 5-Ph-IAA co-treatment would induce ectopic expression of CTCF in the absense of endogenous CTCF-miniAID fusion protein. ChIP-seq was conducted in this model system to interrogate genome-wide bniding occupancy.

first: Judith

last: Hyle

first: Zhenling

last: Liu

first: Qiong

first: Chunliang

last: Li


title: Thymidylate synthase RIP-seq from DNA-repair-deficient HCT116 cells treated with RTX or 5FdUR [RIP-Seq]

first: Angela

last: Bekesi

summary: Thymidylate synthase (TS) is a key enzyme in thymidylate biosynthesis and an established target of chemotherapeutics such as 5-fluoro-2’-deoxyuridine (5FdUR) and raltitrexed (RTX). Inhibition of TS disrupts the dUTP:dTTP balance, leading to uracil misincorporation, futile base excision repair cycles, DNA strand breaks, and ultimately cell death. Beyond its catalytic role, TS also binds RNA, autoregulating its own translation and interacting with transcripts such as p53 and c-myc, thereby linking TS activity to broader post-transcriptional regulatory networks. To better understand how TS inhibitory drugs influence this network, we performed TS-coupled RNA-coimmunoprecipitation (TS-RIP) and sequencing from mismatch repair-deficient, UNG-inhibited HCT116 colon cancer cells treated with 5FdUR or RTX. TS-RIP revealed RNA-binding targets of TS, in addition to the known p53 and TS mRNAs. Our findings underscore the multifaceted impact of TS inhibition, linking enzymatic disruption to RNA-level regulation and revealing drug-specific differences in cellular responses.

overall_design: HCT116 cells were treated for 48 hours with drugs (100 nM RTX or 20 µM 5FdUR) or left untreated (NT), then lysis and subcellular fractionation were performed, and cytoplasmic and nuclear fractions were combined, then pre-cleared with magnetic Protein A/G  beads. TS-specific RNA immunoprecipitation (TS-RIP) was performed using anti-TS-antibody (Proteintech 15047-1-AP) captured on Protein A/G beads, followed by washing and elution. RNA was isolated from the immunoprecipitated (TS-RIP) and control (RIPctr) samples, quality-checked by agarose gelelectrophoresis, and prepared for sequencing.

first: Eszter

last: Holub

first: Anna

last: Felföldi

first: Beáta

last: G. Vértessy

first: Angéla

last: Békési


title: Dose-dependent sensitivity of human 3D chromatin to a heart disease-linked transcription factor [RNA-Seq 2]

city: San Francisco

first: Benoit

middle: G.

last: Bruneau

state: CA

address: 1650 Owens St

institute: Gladstone Institutes

summary: Dosage-sensitive transcription factors (TFs) underlie altered gene regulation in human developmental disorders, and cell type-specific gene regulation is linked to the reorganization of three-dimensional (3D) chromatin during cellular differentiation. Here, we show dose-dependent regulation of chromatin organization by the congenital heart disease (CHD)-linked, lineage-restricted TF TBX5 in human cardiomyocyte differentiation. Genome organization, including compartments, topologically associated domains, and chromatin loops, are sensitive to reduced TBX5 dosage in a human model of CHD, with variations in response across individual cells. Cohesin binding was dose-dependently reduced at TBX5-bound enhancer elements in the absence of TBX5, providing a potential mechanism for disrupted loop formation. These results highlight the importance of lineage-restricted TF dosage in cell type-specific 3D chromatin dynamics, suggesting a new mechanism for TF-dependent disease.

overall_design: RNA-seq of a TBX5-degron cell line treated with DMSO or 500 nM dTAGv-1, each sample has three biological replicates.

first: Zoe

last: Grant

first: Shuzhen

last: Kuang

first: Shu

last: Zhang

first: Abraham

middle: J

last: Horrillo

first: Zhe

last: Chen

first: Kavitha

last: Rao

first: Cemre

last: Celen

first: Vasumathi

last: Kameswaran

first: Carine

last: Joubran

first: Pik Ki

last: Lau

first: Keyi

last: Dong

first: Bing

last: Yang

first: Weronika

last: Bartosik

first: Nathan

middle: R

last: Zemke

last: Ren

first: Deepak

last: Srivastava

first: Irfan

last: Kathiriya

first: Katherine

last: Pollard

first: Benoit. G


title: Dose-dependent sensitivity of human 3D chromatin to a heart disease-linked transcription factor [ChIP-Seq 2]

first: Benoit

last: Bruneau

overall_design: RAD21 ChIP-seq of cells from atrial cardiomyocyte differentation at D20 from WT (TBX5+/+), heterozygous (TBX5in/+) and null (TBX5in/del) WTC11 iPSCs, H3K27ac ChIP-seq of cells from atrial cardiomyocyte differentation at D20 from WT (TBX5+/+), heterozygous (TBX5in/+) and null (TBX5in/del) WTC11 iPSCs used for differential analyses, and H3K27ac ChIP-seq of cells from atrial cardiomyocyte differentation at D20 from WT

first: Zoe

last: Grant

first: Shuzhen

last: Kuang


title: Single-cell transcriptomics of oncogenic β-catenin in hepatocytes

city: Philadelphia

first: Abhinav

last: Illendula

address: 333 Cottman Avenue

institute: Fox Chase Cancer Center

summary: This study analyzes hepatocytes recombined to express oncogenic β-catenin in the normal liver and chronically damaged liver background.

overall_design: 1 or 3 months post-AAV8-TBG-Cre injection in 8-week-old CR mice, liver parenchymal cells were isolated and sorted by Fluorescence Activated Cell Sorting (FACS). tdT- (WT) hepatocytes and tdT+ (Ctnnb1-ΔExon3) were sorted and subsequently mixed in a 1:1 ratio. This process was repeated in each mouse. CMR mice modelled chronic liver damage and were started on high-fat diet at 6-weeks old. At 8-weeks old, mice were injected with AAV8-TBG-Cre and cells were sorted by FACS 3 or 5 months post-injection similar to CR mice. All samples were then analyzed for their trancriptional profile by scRNAseq.

first: Charles

last: Hewett

first: Yuki

last: Hayata

first: Joan

last: Font-Burgada


title: Dose-dependent sensitivity of human 3D chromatin to a heart disease-linked transcription factor [Hi-C 2]

overall_design: Hi-C 3.0 of cells from atrial cardiomyocyte differentation at D20 with DMSO or 500 nM dTAGv-1 ligand treated for 24h

first: Shu

last: Zhang

first: Abraham

last: Horrillo

first: Zhe

last: Chen

first: Kavitha

last: Rao

first: Cemre

last: Celen

first: Vasumathi

last: Kameswaran

first: Carine

last: Joubran


title: Mov10 directly targets Camp to promote its protein synthesis

city: Chongqing

first: Chang

middle: Lin

last: Zhen

address: Baianba

institute: Chongqing Three Gorges Medical College

summary: Further research has revealed that the mRNA levels of Camp did not change but its protein expression was decreased after Mov10 depletion evidently, suggesting that Mov10 may regulates the protein translation efficiency of Camp without affecting its mRNA abundance . To validate the above hypothesis, we isolated Lin−c-Kit+ cell from Mov10fl/fl and Mov10-CKO mice after 12 days 4.5 Gy IR for translation sequencing (Ribo-seq) to analyze the effect of Mov10 on the translation efficiency of differentially expressed proteins and the abundance of ribosome-protected fragments (RPFs), funding protein translation efficiency of Camp is significantly downregulated after 12 days 4.5 Gy IR . Ribo-seq genome browser views re-vealed strong ribosome stalling both at the Camp’s TISs and during the protein translation pro-cess of Camp after 12 days 4.5 Gy IR

overall_design: RPF abundance (Ribo-seq) in Lin-c-Kit+ cells from Mov10 knockout and control mice 12 days post-4.5 Gy IR based on Transcriptomics data.


title: BUI1, an actin cytoskeleton protein that confers durable and broad-spectrum resistance in rice

city: Shanghai

first: yangxuan

last: liu

address: 300 Fenglin Road

institute: CAS Center for Excellence in Molecular Plant Science (CEMPS), Chinese Academy of Sciences

summary: The cytoskeleton functions as a dynamic intracellular structural scaffold, enabling cells to rapidly perceive biotic and abiotic stresses. During plant innate immune responses, the cytoskeleton undergoes actin remodeling. Using the rice-Magnaporthe oryzae pathosystem, we investigated changes in actin organization within host plant cells. Our results demonstrate that pattern-triggered immunity (PTI), induced by pathogen-associated molecular patterns (PAMPs), triggers a rapid but transient increase in actin filament abundance. This process depends on BUI1, a type-II formin protein. Disruption of actin filaments with latrunculin B not only suppresses this increase but also enhances host susceptibility. Furthermore, we show that BUI1 positively regulates broad-spectrum resistance against multiple pathogens by suppressing reactive oxygen species (ROS) scavenging. Together, these findings uncover a key mechanism by which receptor-mediated signaling reorganizes the host cytoskeleton during plant-pathogen interactions.

overall_design: The leaves of ZJ22 and bui1 plants following Xoo inoculation were sampled at 0h, 6h, 12h, 24h and 48h for transcriptome analysis. Around  5-6 leaves were pooled as one replicate. The libraries were prepared and sequenced using Illumina.

first: Jingyi

last: Wang

first: Yanlong

last: Jin

first: Mingjun

last: Gao


title: Biomarker-guided responses in patient-derived organoids predict effective therapies in breast cancer

first: Jennifer

middle: M

last: Rosenbluth

address: 10 Koret Way

institute: University of California San Francisco

summary: Poor therapeutic response in subsets of breast cancer (BC) patients poses an ongoing challenge. Here, we present a series of 14 patient-derived triple-negative BC organoids, established with the aim of modeling resistant disease with greater fidelity using biomarkers and developing an in-vitro system grounded in clinical data for testing alternative treatment strategies.

overall_design: RNA-seq profiling of triple-negative breast cancer organoids

first: Tam Binh

last: Bui


title: GLI2 mediates response to paclitaxel in triple-negative breast cancer

city: Nashville

first: Shristi

last: Shrestha

address: 2213 Garland Ave

institute: Vanderbilt University

summary: Bone is the most frequent site of both metastasis and recurrence for patients with breast cancer, and there are currently no FDA-approved therapeutics which target cancer in bone. Skeletal metastases upregulate the transcription factor Glioma-associated oncogene 2 (Gli2), resulting in secretion of parathyroid hormone-related protein (PTHrP), which accelerates bone turnover and enables tumor proliferation. Gli2 has been implicated as a mediator of chemoresistance in a wide range of cancers, but its unique role in drug resistance has yet to be defined in treatment-resistant skeletal metastases. Therefore, this dataset aims to define the impact of GLI2 CRISPR knockout on transcriptional response to paclitaxel (PTX), the current standard of care for breast cancer patients with distant metastases.

overall_design: MDA-MB-231 cells were passaged intratibially in athymic nude mice, isolated from bone marrow post mortem, injected intracardiac, and isolated again from bone marrow to make bone-tropic cells of the MDA-MB-231b lineage. CRISPR knockout  was performed via lentiviral transduction using Dharmacon Edit-R CRISPR-Cas9 products (Horizon) using sgRNA targeted to GLI2 (sgGLI2) or non-targeted sgRNA.

first: Jade

last: Miller

first: Emily

last: Jaremba

first: Julie

last: Rhoades


title: MDM2 suppresses c-Myc synthesis by binding to the 5’ mRNA translation regulatory sequence.

city: Umeå

first: Lixiao

state: Sverige

address: Medicinsk biovetenskap, M21, Försörjningsvägen 1

department: Department of Medical Biosciences

institute: Umeå University

summary: The p53 tumor suppressor and the c-Myc oncogene are among the most frequently deregulated genes in human cancers, yet the molecular crosstalk between these pathways remains poorly understood. MDM2 is a key negative regulator of p53 and a target for emerging cancer therapies designed to activate p53. Likewise, targeting c-Myc is a longstanding but challenging goal in cancer therapy. Here we report that the small MDM2 binding drug Milademetan promotes an interaction between MDM2 and the 5’ untranslated region (UTR) of the c-Myc mRNA, causing a suppression of c-Myc mRNA translation without affecting c-Myc RNA levels. The interaction also occurs under non-proliferative conditions in the absence of drug. Milademetan-mediated c-Myc depletion is accompanied by the induction of apoptosis and suppression of cell proliferation and prevents tumor growth, independently of p53 status. These findings reveal an unexpected mechanism by which MDM2 coordinates two of the most frequently altered pathways in cancer and provide a rationale for targeting c-Myc-driven tumours, including those lacking functional p53, through MDM2 modulators.

overall_design: Spatial transcriptomic profiling was performed using the NanoString GeoMx Digital Spatial Profiler (DSP) platform with the Human NGS Whole Transcriptome Atlas (WTA v1.0) panel. Tissue sections from milademetan-treated (n = 5) and vehicle control (n = 5) tumors were distributed across four slides (S1–S4), with each slide containing both treatment groups. Within each region of interest (ROI), four cell populations were defined by fluorescent marker co-staining for Ki67 (proliferation) and MYC (oncogene expression): Ki67+MYC+, Ki67−MYC+, Ki67−MYC−, and Ki67+MYC−. Each segment was collected as an individual area of illumination (AOI), yielding a total of 92 AOIs across all slides.

first: Justine

last: Habault

first: Norman

last: Salomao

first: Laurence

last: Malbert-Colas

first: Chrysoula

last: Daskalogianni

first: Sivakumar Vadivel

last: Gnanasundram

first: Mitesh

last: Dongre

first: Marzia

last: Spagnardi

first: Lucia

last: Martinkova

first: Sa

first: Lenka

last: Hernychova

first: Lucas

last: Robidas

first: Josef

last: Kucera

first: Ondrej

last: Bonczek

first: Daniel

last: Ohlund

first: Michael J.

last: Garabedian

first: Susan K.

last: Logan

first: Borek

last: Vojtesek

first: Robin

last: Fahraeus


title: Depletion of H1.3 in AML cells prompts redistribution of H1.2, causing chromatin remodeling and cell cycle deregulation [RNA-Seq]

city: Marseille

first: Estelle

middle: Anna

last: Duprez

address: 23 bvd lei roure

department: OHIO

country: France

institute: CRCM-INSERM

summary: Linker histone H1 variants play critical, yet distinct, roles in chromatin organization and gene regulation. However, very little is known about their specificity in cancer cells. In this study, we investigated the specificity of the H1.3 variant in acute myeloid leukaemia (AML) cells. Through chromatin mapping and transcriptomic analyses, we revealed that H1.3 was specifically enriched in regions with a high GC content and co-localized with the repressive mark H3K27me3, supporting its role in chromatin compaction and transcriptional repression. Knockout of H1.3 induced specific changes in gene expression profiles and chromatin dynamics, characterized by the relocalization of H1.2, which was redistributed from its usual chromatin regions to the freed H1.3 regions. Consequently, locus-specific chromatin alterations associated with interferon-related signalling pathways and cell cycle deregulation were observed. Our findings emphasized an important and locus-specific function of H1.3 in AML cells. Overall, our study revealed a mechanistic connection between H1 variant imbalance, activation of the immune response, and regulation of the cell cycle, with implications for our understanding of epigenetic regulation in cancer cells.

overall_design: RNA-seq profiling of wildtype OCI-AML-3 cells and their knockout derivates (H1.3 KO clone 1 and H1.3 KO clone 2)

first: Clara

last: Tellez Quijorna

first: Lia

last: N'Guyen

first: Núria

last: Serna Pujol

first: Julien

last: Vernerey

first: Albert

last: Jordan


title: Depletion of H1.3 in AML cells prompts redistribution of H1.2, causing chromatin remodeling and cell cycle deregulation [ATAC-Seq]

overall_design: Genome accessibility profiling by high throughput sequencing of OCI-AML-3 parental cells and OCI-AML-3 H1-3 KOs (KO clone #1 and clone #2)


title: Research on the Application of Lubricated Catheters in Perioperative Mucosal Analgesia and Comfortable Medical Care

city: guangzhou

first: Jinyan

last: Guo

state: guangdong

address: No. 600, Tianhe Road, Tianhe District

department: The Third Affiliated Hospital

institute: Sun Yat-sen University

summary: This study aims to investigate the improving effects of lubricated catheters on mucosal injuries and complications associated with perioperative endotracheal intubation.

overall_design: By establishing a rhesus monkey endotracheal intubation model to simulate the clinical process of general anesthesia intubation, we compared the degree of pharyngeal and laryngeal mucosal irritation caused by modified lubricated catheters versus traditional catheters during intubation. The incidence of postoperative complications, such as mucosal bleeding, throat pain, and hoarseness, was evaluated. Additionally, mucosal tissue samples were collected for transcriptome sequencing to identify potential molecular targets related to intubation-induced injuries.

first: Jiyan


title: KMT2D depletion promote KRAS-induced pancreatic carcinogenesis independent of TP53

city: Rochester

first: Martin

last: Fernandez-Zapico

state: MN

address: 200 First Street SW

institute: Mayo Clinic

summary: We examined the role Kmt2d in PDAC development using two independent genetically engineered mouse models driven by mutant Kras in the presence and absence of the tumor suppressor Tp53. Loss Kmt2d reduces survival and promotes disease progression in both models even in absence of Tp53. Similarly, in human PDAC the loss of TP53 does not affect the prognosis value of KMT2D low cases. Of note, the sole Kmt2d depletion did not affect normal pancreas development or mice survival. RNA-seq analysis demonstrates the enrichment of pathways controlling cell growth in Kmt2df/+ PDAC cells. Interestingly, inactivation of Kmt2d resulted in increased levels of Ki67 positive cells in vivo. Together, these findings define the role of this methyltransferase in Kras-driven PDAC biology as tumor suppressor candidate for this dismal condition.

overall_design: We performed RNA-seq analysis in murine cell lines derived from GEMM pancreas tumor tissue with KPC (3 mice) or KPC Kmt2df/+ (5 mice) genotype under basal conditions. Cells were grown in tissue culture before collecting for RNA-seq. Each mouse has three biological replicates.

first: Fernandez-Zapico

last: Martin

first: Hogenson

last: Tara

first: Pena Ruiz

last: Nicole


title: XRCC1 Deficiency Drives Telomeric Chromatin Leakage, Inflammatory signaling and Senescence in Cancer Cells

city: Naples

first: Stefano

last: Amente

address: via pansini, 5

department: Department of  Molecular Medicine and Medical Biotechnologies

country: Italy

institute: University of Naples "FEDERICO II"

summary: Genome integrity is constantly challenged by oxidative DNA damage, which, if left unrepaired, can result in replication stress, chromosomal instability, and tumorigenesis. The base excision repair (BER) pathway is essential for resolving oxidative base lesions such as 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxodG). Among BER factors, APE1 and XRCC1 play critical roles not only in lesion repair but also in maintaining replication fork stability. Here, we investigate the cellular consequences of APE1 and XRCC1 deficiency in isogenic breast epithelial models (MCF10A and MCF7), under basal and oxidative stress conditions. We show that XRCC1 deficiency, more than APE1 depletion, leads to accumulation of unrepaired BER intermediates, increased DNA damage signaling, and formation of cytoplasmic chromatin fragments (CCFs) containing telomeric DNA. XRCC1-deficient cells also exhibit activation of the cGAS-STING pathway, inflammatory signaling, and features of senescence. Our findings identify XRCC1 as a central suppressor of oxidative stress-induced chromatin instability, cytosolic DNA accumulation, and immune signaling. These results suggest that targeting BER in carcer, particularly XRCC1, may offer a therapeutic avenue to trigger tumor-selective senescence and promote anti-tumor immunity.

overall_design: RNA-seq profiling MCF10A Control-KO (expressing Cas9) and their derived KO derivaties (APE1 and XRCC1)

first: Anna

last: Piscone

first: Francesca

last: Gorini

first: Susanna

last: Ambrosio

first: Carmen

last: Caiazza

last: Noviello

first: Giovanni

last: Scala

first: Barnara

last: Majello


title: Adaptive immunity shapes baseline pImmunity Shapes Baseline Physiology of M. Tuberculosistuberculosis in High-Dose Versus Low-Dose I in high-dose versus low-dose infection BALB/c mouse drug treatment mMouse Drug Treatment Models

city: Aurora

first: Jo

last: Hendrix

state: CO

department: Pulmonology

institute: University of Colorado

summary: Around day 16 of infection with Mycobacterium tuberculosis (Mtb), the BALB/c mouse adaptive immunity activates to combat the infection. Usual dual RNA-seq, we investigated the transcriptional changes that occur in both mouse host and Mtb pathogen during an untreated low dose aerosol (LDA) infection—focusing on the transition from innate to adaptive immunity. We found that the onset of adaptive immunity coincided with a rapid reprogramming of Mtb physiology, characterized by suppressed respiration, metabolism, and biosynthesis, together with induction of stress and nutrient acquisition pathways. Changes in key Mtb processes were concordant with changes in host expression of canonical features of adaptive immunity.

sample_organism: Mycobacterium tuberculosis str. Erdman = ATCC 35801

overall_design: BALB/c female mice were infected with a low dose aerosol (LDA) of Mtb Erdman. Treatment was not administered at any time. Five mice were sacrificed on days 1, 7, 11, 19, 28, and 56 post-infection for lung specimen collection, respectively. Murine host and Mtb pathogen RNA were extracted from whole lung homogenates. Murine RNA was sequenced with RNA-seq at every timepoint. Mtb RNA was sequenced with SEARCH-TB at every timepoint starting on day 11.

first: Reem

last: Al Mubarak

first: Gregory

last: Robertson

first: Chuangqi

last: Wang

first: Nicholas

last: Walter


title: Therapy-resistance in AT/RT is driven by epithelial-mesenchymal transition, extracellular matrix changes, and enrichment of CD1A+ CD207+ dendritic cells [RNA-seq_FFPE]

city: Hamburg

first: Lea

last: Altendorf

address: Martinistraße 52

institute: Universitätsklinikum Hamburg-Eppendorf

summary: Atypical teratoid/rhabdoid tumors (AT/RT) are the most common malignant brain tumors during infancy and associated with a dismal prognosis. 50% of the patients suffer from progressions or recurrences, but the underlying mechanisms remain largely unknown. To identify mechanisms of progression and recurrence, we performed single-nucleus RNA sequencing of eight primary-recurrence pairs. Tumor cells and cells of the tumor microenvironment (TME) were analysed separately. Therapy-resistant tumor cells were identified through the comparison of global gene expression profiles between primary and recurrent tumor cell populations using CIBERSORT. Histology, in vitro experiments, bulk RNA sequencing, and survival analyses were performed to validate our findings. Paired primary and recurrent AT/RT tumor samples showed significant differences in their gene expression profiles. The TME of AT/RT recurrences was characterized by enrichment of CD1A+ CD207+ dendritic cells (DCs), which were per se associated with inferior survival in relapsed patients. Differentially expressed genes of AT/RT-MYC primary tumor cells that potentially persist after therapy, were involved in the organization of the extracellular matrix (ECM), developmental processes, and diverse immune signalling pathways. In addition, recurrences demonstrated a significantly higher proportion of cells undergoing partial epithelial-mesenchymal transition (pEMT) compared to primary tumors, a feature that we confirmed following treating AT/RT-MYC tumor cells in vitro. Finally, patients with primary tumors strongly expressing respective genes displayed a significantly worse survival. We identified pEMT, ECM remodeling, and enrichment of CD1A+ CD207+ DCs as potential mechanisms of therapy resistance in AT/RT which could be employed to improve therapy in the future.

overall_design: RNA sequencing of twelve AT/RT primary tumors using SMART-Seq Stranded kit (Takara Bio) for library construction and NovaSeq X Plus (Illumina) for sequencing.

first: Anton

last: Altehammer

first: Rajanya

last: Roy

first: Karoline

last: Hack

first: Flavia

middle: W

last: de Faria

first: Arend

last: Koch

first: Vanessa

last: Thaden

first: Melanie

last: Schoof

middle: U

last: Schuhmann

first: Peter

last: Hauser

first: Pascal

last: Johann

last: Hasselblatt

first: Michael

last: Frühwald

first: Kornelius

last: Kerl

first: Ulrich

last: Schüller


title: Therapy-resistance in AT/RT is driven by epithelial-mesenchymal transition, extracellular matrix changes, and enrichment of CD1A+ CD207+ dendritic cells [RNA-seq_cell_lines]

overall_design: RNA sequencing of two AT/RT-MYC cell lines before and after treatment using SMART-Seq Stranded kit (Takara Bio) for library construction and NovaSeq X Plus (Illumina) for sequencing.

last: Althammer

first: Martin


title: Therapy-resistance in AT/RT is driven by epithelial-mesenchymal transition, extracellular matrix changes, and enrichment of CD1A+ CD207+ dendritic cells [snRNA-seq]

overall_design: snRNA sequencing of 16 samples using FFPE tissue was performed using the Chromium Single Cell Flex Gene Expression Assay from 10x Genomics.


title: Hepatic delivery of miroRNA-30c analog reduces plasma cholestrol in mice and monkeys and atherosclerosis in mice without causing liver injury

city: Mineola

first: M. Mahmood

last: Hussain

address: 101 Mineola Blvd.

department: Foundations of Medicine

institute: NYU Grossman Long Island School of Medicine

summary: Despite the many useful drugs that are currently available for this indication, cardiovascular disease remains a major cause of premature mortality worldwide. High plasma cholesterol levels contribute substantially to this problem. Here, we show that hepatic delivery of a microRNA-30c analog C2 reduces plasma cholesterol levels in apoB-containing lipoproteins in hypercholesterolemic C57Bl6 mice, and in African green monkeys that spontaneously developed diabetes and hyperlipidemia. In addition, it prevented diet-induced hypercholesterolemia in FRG-mice with humanized livers. Furthermore, C2 significantly reduced plasma cholesterol and atherosclerosis in LDL receptor knockout mice indicating that the cholesterol-lowering effects of C2 do not rely on the presence of LDL receptors. In all these studies, C2 had no effect on hepatic triglyceride and cholesterol, plasma ALT, AST, CK-MB, ALP, IL-6, TNF-α, and INF-ϒ indicating absence of tissue lipid accumulation, inflammatory and immune responses. Mechanistic studies revealed that C2 reduced hepatic microsomal triglyceride transfer protein activity, hepatic secretion of apolipoprotein B-containing lipoproteins and enhanced fecal cholesterol excretion indicating that this could contribute to lowering of plasma cholesterol. Furthermore, C2 increased hepatic FA oxidation, and reduced hepatic FA synthesis and these mechanisms may contribute to avoidance of hepatic steatosis associated with reduced lipoprotein production. C2 could be a first-in-class microRNA therapeutic to reduce plasma cholesterol and atherosclerosis without causing hepatic injury and inflammatory response.

overall_design: Two-month-old female C57BL/6 mice were fed a WD for 2 months and injected with C2 or miR-30c (100-800 mg/kg b.wt). After final dosage of 800 mg/kg, we sacrificed the mice and collected the livers. Total RNA was isoalted from the liver tissue for RNA-sequencing.

first: Binu

last: Prakash

first: Zhihua

last: Chang

first: Shannon

last: Scarberry

first: Bhargavi

last: Gangula

first: Sujith

last: Rajan

first: Chandana

last: Prakashmurthy

first: Swati

last: Valmiki

first: Otabek

last: Pulatov

first: Ryan

last: Temel

first: Steven

last: Carsons

first: Kylie

last: Kavanagh

first: Jia

last: Sheng

first: Mahmood


title: Single-nucleus transcriptomics of CLRN1 knockout rabbit retina reveals Müller glia-driven mechanisms of retinal degeneration

city: Ann Arbor

first: Dongshan

last: Yang

address: 2800 Plymouth Rd

institute: University of Michigan

summary: Single-nucleus RNA sequencing of wild-type and CLRN1 knockout rabbit retina at 10 months of age (pre-symptomatic timepoint). Three biological replicates per genotype were pooled and processed using 10x Genomics Chromium Single Cell 3' v3.1 platform. SNP-based demultiplexing with Vireo was used to assign nuclei to individual animals. Analysis of 6,542 high-quality nuclei across 7 retinal cell types revealed that CLRN1 is exclusively expressed in Müller glia and identified a primary adherens junction defect (CTNNA2 downregulation) leading to secondary photoreceptor degeneration while inner retinal neurons mount robust compensatory responses.

overall_design: Two pools were sequenced: Pool 1 contains 3 wild-type rabbits, Pool 2 contains 3 CLRN1 knockout rabbits. All animals were 10 months old. Retinal tissue from the visual streak region (2-3mm ventral to medullary rays) was collected, snap-frozen, and nuclei were isolated using the 10x Genomics Chromium Nuclei Isolation Kit. Nuclei from 3 biological replicates per genotype were pooled at equal concentration prior to library preparation. Libraries were prepared using 10x Chromium Single Cell 3' v3.1 chemistry and sequenced on Illumina NovaSeq X Plus. Computational demultiplexing using Vireo assigned nuclei back to individual animals based on natural genetic variation.

last: Chen


title: Molecular heterogeneity and differential metabolic signatures in thymic adipocytes

city: Salt Lake City

first: Dean

last: Tantin

state: Utah

address: N Campus Dr

institute: University of Utah

summary: Adipocyte depots throughout the body are physiologically and molecularly distinct. With age, adipocytes increase in and around aged thymi. However, thymic adipocytes completely lack molecular characterization. We developed and optimized methods to isolate adipocyte nuclei from mouse thymi of different ages and sexes. Single-nucleus multiomic analysis of male and female mice aged 4-9 months reveals that thymic adipocytes are heterogeneous, with at least two distinct populations. One subpopulation harbors a transcription and chromatin signature consistent with beige/brown fat. A larger subpopulation more strongly resembles classic white adipose tissue and expresses genes associated with epithelial-to-mesenchymal transition (EMT) and antigen presentation. Analysis of differentially open chromatin in the white compared to beige adipose population identifies binding sites for Foxn1 and HIF-1a/Arnt, consistent with a situation in which thymic white adipose cells emerge from thymic epithelial cells, possibly under hypoxic conditions. Immunofluorescence microscopy confirmed the expression of UCP1 protein in cells within the thymic parenchyma, most prominently in subcapsular cortical regions. These results reveal a complex milieu of thymic adipocytes and identify multiple avenues for directly probing their ontogeny, dynamics and functional significance.

overall_design: Cleaned, intact thymi were pooled from 16 naïve 6mo Female C57/BL6 (Multiome-seq) or 6 naïve 4mo Male C57/BL6 (snRNAseq) on standard chow diets. Adipocytes were enriched by enzymatic digestion followed by fractionation containing buoyant cells. Nuclei were isolated for library prep and subsequent multiome (snRNAseq+snATACseq) or snRNAseq alone.

first: Joon

last: Schwakopf

first: Amber

last: Syage

first: Anca

last: Franzini

first: Katherine

last: Varley


title: Alyref Deficiency Exacerbates Chronic Inflammatory Pain by Impairing m5C-Dependent Nuclear Export of Ccn3 mRNA

city: Shenzhen

first: Jian

last: Zhang

address: 1088 Xueyuan Avenue

institute: Southern University of Science and Technology

summary: The 5-methylcytosine (m⁵C) RNA modification regulates multiple aspects of RNA metabolism; however, its contribution to pathological pain remains poorly understood. Here, we investigated the role of the m⁵C reader Alyref in a mouse model of complete Freund’s adjuvant (CFA)-induced chronic inflammatory pain. We observed a marked and sustained upregulation of Alyref in nociceptive neurons of the dorsal root ganglion (DRG) following CFA administration. Conditional deletion of Alyref in NaV1.8⁺ nociceptive neurons significantly exacerbated thermal and cold hypersensitivity, mechanical allodynia, and hyperalgesia. These behavioral abnormalities were accompanied by pronounced activation of microglia and astrocytes in the spinal dorsal horn, along with elevated expression of proinflammatory mediators, indicating enhanced neuroinflammation and central sensitization. Mechanistically, Alyref directly binds to m⁵C-modified Ccn3 mRNA and promotes its nuclear export, thereby maintaining cytoplasmic Ccn3 expression in DRG neurons. Loss of Alyref impaired Ccn3 mRNA export, reduced Ccn3 expression, and was associated with increased level of MMP-9, a key mediator of neuroinflammatory pain signaling. Collectively, these findings identify Alyref as a protective regulator in chronic inflammatory pain by restraining neuroinflammation and limiting the emergence of neuropathic pain-like features. Our study highlights a previously unrecognized role of m⁵C-dependent RNA regulation in nociceptive sensitization and suggests Alyref as a potential therapeutic target for pathological pain.

overall_design: We performed high-throughput sequencing of mouse DRGs (dorsal root ganglia) with or without deletion of Alyref to profile RNA expression. For the RNA-seq experiments, we conducted 4 batches of sequencing experiments using different biological replicates to account for technical variation.

first: Yuxuan

last: Shui

first: Shuxin

last: Li

first: Hanlin

last: Wang

first: Sheng-Jian

last: Ji


title: Secreted Frizzled-Related Protein 1 Controls Distal Lung Formation via Wnt and PDGF Signaling

city: New York

first: Jeanine

last: D'Armiento

address: 622 W. 168th street

institute: Columbia University Medical Center

summary: Normal lung development is an intricate process that depends on temporally regulated mesenchymal signaling to coordinate structural maturation, cell differentiation, and extracellular matrix organization. How endogenous Wnt modulators such as secreted frizzled-related protein 1 (SFRP1), influence mesenchymal behavior in the developing lung remains incompletely understood, but a deeper understanding could reveal potential therapeutic targets relevant for neonatal lung diseases. Using a genetic Sfrp1 knockout model, we studied the role of Sfrp1 in distal lung development and mesenchymal signaling. Histological and morphometric studies were performed to compare lung developmental stages in the Sfrp1-/- and Sfrp1+/+ mice. Single cell RNA sequencing and proximity ligation in situ hybridization studies determined cell specific Sfrp1 expression in early development. Bulk RNA sequencing and pathway enrichment analysis detected differentially expressed genes in mesenchymal cells in the saccular stage of development. Ablation of Sfrp1 led to abnormal morphometry in the pseudoglandular, canalicular, saccular, and alveolar stages. Single cell RNA sequencing and proximity ligation in situ hybridization studies confirmed expression of Sfrp1 in mesenchymal cells, predominantly fibroblasts, in early development. Pathway enrichment analysis revealed changes in signaling pathways within the whole lung and PDGFR𝛼+ cells in the saccular stage.These findings provide new evidence that Wnts exert their control over sacculation and alveologenesis through fibroblast signaling and differentiation. These results provide an important link between studies demonstrating Wnt signaling patterns in alveologenesis, aberrant Wnt signaling in BPD, and Sfrp1 mediated control of fibroblast differentiation after injury.

overall_design: Bulk RNA sequencing of whole lung and sorted PDGFRa+ cells on postnatal day 0

first: Nooralam

last: Rai


title: Dynamic interaction between tentacle-tethered microdomains in Integrator and NELF regulates the promoter-proximal pause checkpoint [RNA-Seq]

city: Copenhagen N

first: Jesper

last: Svejstrup

address: Blegdamsvej 3, 18.6

institute: University of Copenhagen

summary: NELF and Integrator are key regulators of promoter-proximal pausing and premature termination. Integrator subunit 12 (INTS12) contains a PHD domain within a flexible linker. While PHDs generally bind histones, we identify an interaction between the INTS12 PHD and a domain in the flexible C-terminal tentacle of NELF-A. Mutation of either domain disrupts the interaction, and although RNAPII-associated pausing complexes form, they are dysfunctional. Importantly, upon NELF-A mutation, interactions of the Integrator-containing pausing complex with transcriptional activators become apparent that correlate with increased pause-escape. Indeed, while INTS12 PHD loss functionally mimics Integrator loss, NELF-A mutation mimics loss of NELF, although the RNAPII pause position does not change. This supports a model in which Integrator is an integral component of the promoter-proximal pausing complex, and where INTS12’s PHD either interacts with NELF-A to pause/terminate transcription or enables interaction with activators and positive transcription regulators to allow pause-escape.

overall_design: TTchem-seq. Nascent transcript analysis following endogenous NELFA knockdown and replacement with WT and R453A mutated NELFA forms. Samples were also analysed following DRB addition and subsequent washout.


title: The E-cadherin-Wnt-mir-994 Axis Repurposes a Cadherin Switch for Niche Robustness and Germline Stem Cell Maintenance

city: Kansas City

first: Shiyuan

middle: (Cynthia)

state: MO

department: Computational Biology

institute: Stowers Institute for Medical Research

summary: E-cadherin is encoded by the shg gene and is also expressed in Drosophila ovarian IGS cells. Its knockdown leads to a reduction in GSCs. To investigate the downstream molecular mechanisms and changes in gene expression, we labeled IGS cells with GFP, sorted GFP-positive cells one week after shg knockdown, and performed RNA sequencing. Finally, differential gene expression analysis along with GO and KEGG analyses were conducted.

overall_design: By labeling Drosophila ovarian IGS cells with GFP, at the adult stage, RNAi knockdown of shg was performed through culturing at 29°C for one week. Subsequently, GFP-positive cells were sorted by flow cytometry, and RNA was extracted from the collected cells for sequencing.

first: Renjun

last: Tu

first: Ting

last: Xie


title: The role of the renal tubules in diabetic kidney disease (DKD)

city: Beijing

first: Qi

last: Jin

address: No.5, Beixiange, Xicheng District, Beijing

institute: Guang'anmen Hospital, China Academy of Chinese Medical Sciences

summary: To assess the renal tubular condition in DKD patients，

overall_design: To explore the renal tubules in DKD patients, we performed digital spatial profiling (DSP) sequencing of the tubular regions of paraffin sections of renal puncture samples from the normal (paraneoplastic tissue) and DKD groups.

first: Fang

last: Ma

first: Lanfang

first: Peng

last: Qu

first: Yongli

last: Zhan

first: Liang

last: Peng


title: Spatial Transcriptomic Mapping of Postnatal Mouse Uterine Development

city: Callaghan

first: M. Fairuz

last: B. Jamaluddin

state: NSW

address: University drive

country: Australia

institute: University of Newcastle

summary: The uterus undergoes extensive remodeling after birth to acquire the epithelial structure required for adult reproductive function, yet how this process is spatially organized has remained unclear. This study provides a high-resolution spatial map of postnatal uterine development, revealing how epithelial cells diversify and mature within their native tissue context. By linking gene expression patterns to precise anatomical locations, the work shows how local signaling environments guide epithelial differentiation over time. The findings have broad relevance for understanding how epithelial tissues are patterned, maintained, and remodeled, providing a reference for studies of normal development and disease.

overall_design: We performed Xenium in situ spatial transcriptomics (10x Genomics) to uterine sections collected at day 3, d8, d12, and d21


title: Depletion of H1.3 in AML cells prompts redistribution of H1.2, causing chromatin remodeling and cell cycle deregulation [ChIP-Seq]

first: Estelle

last: Duprez

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for histone variants H1.2 and H1.3, as well as the histone modifications H3K27me3 and H3K9me3 in OCI-AML-3 WT (parental) and OCI-AML-3 H1.3 KO cell lines. OCI-AML-3 cells overexpressing V5 tagged H1.3 were subjected to ChIP-seq with anti-V5 antibody. OCI-AML-3 V5 tagged H1.3 cells expressing doxycline.

first: Clara

last: Tellez Quijorna

first: Lia

last: N'Guyen

first: Núria

last: Serna Pujol

first: Maeva

last: Rodies

first: Julien

last: Vernerey

first: Albert

last: Jordan


title: IL4Ra blockade inhibits proliferation of malignant lymphocytes and the immunosuppressive tumor microenvironment of mycosis fungoides

city: Pittsburgh

first: Patrizia

last: Fuschiotti

state: PA

address: 200 Lothrop St

institute: University of Pittsburgh

summary: The IL4 and IL13 receptors are highly expressed in the mycosis fungoides (MF) cutaneous tumor microenvironment (TME). By combining single-cell RNA sequencing with inhibition of IL4Ra with Dupilumab (REGN668), a monoclonal antibody that targets the common subunit of the IL4 and IL13 receptors, we have identified the genes and pathways regulated by IL4/IL13 signaling in the MF TME. We found that Dupilumab inhibits patient-specific processes in malignant lymphocytes, and we also identified downregulation of common pathways among patient samples including those associated with cell cycle progression, PI3K/AKT signaling, JAK/STAT signaling, DNA damage/repair, and TP53 regulation. Using ex vivo skin explants assays from advanced MF skin tumors we demonstrated that Dupilumab inhibits proliferation of MF malignant lymphocytes without affecting their survival. Furthermore, we showed that IL4Ra blockade in the TME of advanced MF downregulates multiple immunosuppressive pathways associated with myeloid-derived suppressive cell and LAMP3+ dendritic cell recruitment and function, cytokine production and antigen presentation in B cells as well as differentiation, and degranulation of mast cells. These results provide new insights into advanced MF tumorigenesis including the nature of Dupilumab interactions within the MF TME. However, the extensive genetic, transcriptional, and functional heterogeneity across MF patients indicates that inhibition of multiple driver pathways and a personalized therapeutic approach, which may include Dupilumab, will be essential for developing new therapeutic strategies against this incurable disease.

overall_design:
MF samples and healthy skin samples were cultured for 2 days and analyzed by scRNAseq.  Raw data are unvailable due to patient privacy concerns

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

first: Alyxzandria

last: Gaydosik

first: Brigit

last: Lapolla

first: Akilov

last: Oleg

first: Larisa

last: Geskin


title: Effect of SIRT6 on renal tubular cells in diabetic kidney disease [CUT&Tag]

first: Qi

last: Jin

summary: To explore the role of SIRT6 in renal tubular epithelial cells in DKD mice, we extracted primary renal tubular epithelial cells from wild-type and tubular epithelial SIRT6-specific knockout DKD mice, respectively.

overall_design: We then analysed promoter region binding peaks in CUT&Tag data from primary renal tubular cells from different groups using the H3K9ac antibody.

first: Fang

last: Ma

first: Lanfang

last: Li


title: Epigenetic landscape of synthetic human centromeres: Euchromatic modifications coexist interdependently at CENP-A dense regions and prevent intrusion of other modifications [ChIP-seq]

city: Yonago

first: Junichiro

last: Ohzeki

address: 86 Nishi-cho

department: Chromosome Engineering Research Center

institute: Tottori University

summary: Centromere is specialized chromatin structure essential for equal chromosome segregation. Human centromere is organized on a portion of homogeneously repeated DNA sequence, and contains nucleosomes including centromere specific histone H3, CENP-A. Histone modifications around the CENP-A dense region are also thought to have important roles in centromere function. Here, we used human artificial chromosome (HAC) technology to create functional centromeric chromatin on a synthetic centromeric repeat DNA sequence that allowed for structural analysis at 2-kb resolution. Analysis using this HAC centromere showed that the CENP-A dense regions are approximately 18–50 kb in size and coexisted with euchromatic histone modifications in a mutually dependent manner. We also find that when DNA replication reduces CENP-A density, a heterochromatin modification H3K9me3 transiently intrudes CENP-A-dense regions, and that both the CENP-A deposition factor HJURP and the histone acetyltransferase KAT7 are important for pushing back this intrusion. The synthetic centromere DNA generated in this study is more effective at elucidating centromere chromatin structure than relying on naturally occurring DNA mutations, allowing for a more precise understanding and manipulation of centromere function.

overall_design: ChIP analysis using antibodies against CENP-A and histone modifications in human HT1080 cells carrying artificial chromosomes. Cells were treated with drugs or siRNA depending on the purpose. The position code sequence of the ChIPed DNA was amplified by competitive PCR, and the amount ratio of the position code sequence was counted using a sequencer.

first: Kazuto

last: Kugou