title: Unfolded Protein Response governs an Alternative Splicing program conserved from healthy to malignant cells

city: London

first: Eleni

last: Maniati

address: John Vane Science Centre, Charterhouse Square

department: Barts Cancer Institute

institute: Queen Mary University of London

summary: The unfolded protein response (UPR) is a critical adaptive program triggered upon cellular stresses that profoundly reshapes the transcriptome and translatome. In the very first minutes of cellular stress, translation blockage, RNA decay and RNA granules formation prompt the synthesis of proteins essential to the stress response. Due to the dynamic nature of these processes and ribosome stalling, investigating translation upon stress has proven to be challenging; therefore, our understanding of these mechanisms and translatome rewiring upon stress remains limited. Here, we exploited O-Propargyl-puromycin (OPP) labelling of de novo peptides followed by LC-MS/MS to identify de novo proteins translated upon endoplasmic reticulum stress. Our approach combined to transcriptomic analyses revealed the synthesis of core splicing factor proteins and a profound reshaping of the splicing landscape upon ER stress. We identified a signature of 8 splicing events systematically occurring in eukaryotic cells exposed to ER stress. Using pharmacological, genetic, phosphoproteomic and sequencing approaches, we demonstrated that this specific ERsplice signature is driven by PERK activation and is dependent on splicing factors’ phosphorylation. Our study unveils a new role for ER stress and PERK in splicing regulation, and provide a molecular signature of ER stress through an ERsplice signature conserved from healthy to malignant tissues.

overall_design: RNASeq analysis was performed on the Oci AML3 and the HEK293T cell lines to examine the transcriptional changes in response to ER stress induction by treatment with Tunicamicin. Oci AML3 cells were examined at 4h and 24h after Tunicamicin treatment and DMSO treated cells as well as untreated cells were used as controls. HEK293T cells were examined at 24h and DMSO treated cells were used as controls. N = 3 replicates per experimental group.

first: Celine

last: Philippe

first: Kevin

last: Rouaoult-Pierre


title: Longitudinal scRNA-seq of murine Il10KO colitis

status: Public on Apr 28 2026

city: Stockholm

first: Jennifer

last: Fransson

address: Karolinska Institutet

country: Sweden

institute: Karolinska Institutet

type: Expression profiling by high throughput sequencing

summary: In order to describe in detail the processes involved in chronic colitis, we performed scRNA-seq at multiple time points in murine Il10KO colitis. These data were compared to bulk RNAseq and spatial transcriptomics results within the same study. scRNAseq data were analyzed by annotating cell clusters and comparing proportions and gene expression programs over time.

overall_design: Distal colon tissue was collected from wild-type at 6 weeks of age and from Il10KO mice at intervals of 14 days from 6 weeks to 14 weeks of age. Colons from 3 animals at each time point were pooled and cells were sorted by FACS to separate epithelial (EpCAM+/CD45-), immune (EpCAM-/CD45+) and stroma (EpCAM-/CD45-) populations. Cells were processed using the 10X Chromium system for gene expression and Feature Barcodes.

first: Chiara

last: Sorini

first: Eduardo

middle: J

last: Villablanca


title: Dual-targeting pharmacological UFMylation inhibition reprograms tumor and immune microenvironments to achieve long-term glioblastoma regression

city: Haidian

first: Pengcheng

last: Tan

address: 30 Shuangqing Rd

country: China

institute: Tsinghua University

summary: UFMylation, a recently identified ubiquitin-like modification, is essential for cellular stress homeostasis, particularly endoplasmic reticulum (ER) stress regulation. However, its biological and therapeutic exploration has been hindered by the absence of potent small-molecule inhibitors. Here, we report the first discovery of two compounds targeting the UFMylation E3 ligase complex core protein DDRGK1: Osimertinib, originally designed as an EGFR T790M selective inhibitor, acting through a previously unrecognized covalent mechanism, and CP-24, a novel non-covalent inhibitor. Both compounds disrupt the DDRGK1-UFL1 interaction, globally suppress UFMylation, inhibit ER-phagy, and induce ER stress. In glioblastoma (GBM), pharmacological UFMylation inhibition markedly reduces tumor cell viability and sensitizes cells to Temozolomide and radiotherapy. Both compounds also exert strong immunomodulatory activity, promoting macrophage polarization toward an anti-tumor M1 state. In vivo, Osimertinib, benefiting from superior pharmacokinetics, significantly suppresses tumor growth in immunodeficient models and achieves tumor-free outcomes in 65% of immunocompetent mice. These tumor-free mice develop durable anti-GBM immune memory, rapidly clearing tumors upon rechallenge, an outcome unattainable by previous GBM treatments. Mechanistically, Osimertinib enhances anti-tumor immunity by promoting macrophage M1 polarization, T cell expansion, and reducing PD-1 protein levels. Collectively, our study introduces Osimertinib and CP-24 as valuable chemical probes for dissecting UFMylation biology and highlights Osimertinib’s potential for off-label use in EGFR-wildtype GBM. More broadly, we establish UFMylation inhibition as a dual-targeting therapeutic strategy that disrupts tumor survival pathways and reprograms the immune microenvironment, offering a promising avenue for durable GBM control.

overall_design: U87 cells were treated with DMSO, 5 μM Osimertinib, or 40 μM CP-24 24 h before RNA extraction. Total RNA of each sample was extracted using Trizol as described previous. The RNA-seq libraries were constructed using the using VAHTS Universal V6 RNA-seq Library Prep Kit for Illumina® according to manufacturer’s instructions. Sequencing was performed on Illumina Novaseq Xplus-25B platform.

first: Zhimin

last: Liu

first: Xiaodan

last: Hu

first: Yuxia

last: Zhang

first: Hedi

last: Chen

first: Baoliang

last: Lan

first: Tianhua

last: Ma

first: Sheng

last: Ding


title: Longitudinal bulk RNA-seq of murine Il10KO colitis

summary: In order to describe in detail the processes involved in chronic colitis, we performed bulk RNA-seq at multiple time points in murine Il10KO colitis. These data were compared to scRNAseq and spatial transcriptomics results from the same mice within the same study. Bulk data were analyzed by identifying differentially expressed genes between time points and treatment groups.

overall_design: Distal colon tissue was collected from BALB/c wild-type at 6 weeks of age and Il10KO mice at 2 week intervals from 6 weeks to 14 weeks of age. Bulk RNA-seq was performed on tissue samples. n = 3 mice per time point and genotype


title: Unbiased niche labeling maps immune-excluded niche in bone metastasis

city: Houston

first: Xiang

address: One Baylor Plaza

country: USA

institute: Baylor College of Medicine

summary: Microenvironment niches determine cellular fates of metastatic cancer cells. However, robust and unbiased approaches to identify niche components and their molecular profiles are lacking. We established Sortase A-Based Microenvironment Niche Tagging (SAMENT), which selectively labels cells encountered by cancer cells during metastatic colonization. SAMENT was applied to multiple cancer models colonizing the same organ and the same cancer to different organs. Common niche features include macrophage enrichment and T cell depletion. Macrophage niches are phenotypically diverse in different organs. In bone, niche macrophages express estrogen receptor (ER) and exhibit active ER signaling in male and female hosts. Conditional knockout of ER in macrophages significantly retarded bone colonization by allowing T cell infiltration. ER expression was also discovered in human bone metastases of both genders. Collectively, we identified a unique population of ER+ macrophages in metastatic niche and functionally tie ER signaling in macrophages to T cell exclusion during metastatic colonization.

overall_design: For the preparation of mouse tissue for the SAMENT single-cell experiment, we collected various tissues, including lungs, livers, brains, femurs, and tibias, following an overnight (~10 hr) SAMENT substrate labeling period. Then tissues were processed and went through the cell sorting, to seperate the biotin positive and biotin negative populations. All the sorted cells from each sample were multiplexed following the CellPlex or TotalSeqC instructions, followed by library construction and sequencing.

first: Zhan

last: Xu

first: Fengshuo


title: Integrated Phenotypic and Transcriptomic Profiling Positions ONC212 as a Lead Imipridone in Androgen-Independent Prostate Cancer Models

status: Public on Apr 27 2026

city: Beirut

first: AUB

last: Submission

address: Bliss street

country: Lebanon

institute: American University of Beirut

summary: Prostate cancer (PCa) remains lethal at advanced stages, partly due to stem-like subpopulations known as prostate cancer stem cells (PCSCs) that sustain tumor growth and therapeutic resistance. Imipridones are small-molecule anticancer agents, with next-generation derivatives ONC206 and ONC212 designed for enhanced potency and broader activity. This study compared their antitumor efficacy and mechanisms in advanced androgen-independent PCa (AIPC) models, namely DU145 and PC3 cells, using two- and three-dimensional systems encompassing bulk cancer cells and PCSCs. DU145 and PC3 AIPC cells were treated with ONC201 (parent compound), ONC206, or ONC212. Functional assays assessed proliferation, viability, migration, invasion, PCa spheroids formation, cell cycle distribution, and mitochondrial membrane potential and mass, while RNA sequencing defined transcriptional responses. ONC212 was the most potent derivative, inhibiting proliferation and migration and abolishing PCa spheroids at nanomolar doses, whereas ONC201 and ONC206 required higher concentrations. Transcriptomic analyses revealed shared repression of DNA replication and cell-cycle transition programs, with activation of integrated stress and unfolded protein responses (ISR/UPR) and FOXO signaling. ONC206 favored PERK–ATF4-mediated apoptosis with reduced DNA repair, while ONC212 more strongly impacted oxidative phosphorylation-related pathways and mitochondrial RNA processing. Imipridones induced a time-dependent cell-cycle redistribution with increased sub-G1 accumulation and modulated mitochondrial membrane potential and mass in a context-dependent manner. Collectively, these findings position ONC212 as a leading imipridone candidate in AIPC models, combining potent inhibition of tumor and stem-like cell functions with a coherent stress-response signature that supports further translational evaluation.

overall_design: RNA-seq profiling of PC3 and DU145 cell lines after 48 hours treatment with DMSO (Vehicle), ONC206 (0.5 µM), or ONC212 (0.1 µM).

first: Fatima

last: Ghamlouche

first: Amani

last: Yehya

first: Abdallah

last: Kurdi

first: Sana

last: Hachem

first: Varun

middle: V

last: Prabhu

first: Georges

last: Daoud

first: Wassim

last: Abou-Kheir


title: Longitudinal scRNA-seq of murine T cell transfer colitis

first: Jennifer

last: Fransson

summary: In order to describe in detail the processes involved in chronic colitis, we performed scRNA-seq at multiple time points in CD45RBhi T cell transfer colitis mouse models. These data were compared to bulk RNAseq and spatial transcriptomics results from the same mice within the same study. scRNAseq data were analyzed by annotating cell clusters and comparing proportions and gene expression programs over time.

overall_design: T cell transfer colitis was induced in Rag2 KO and SCID mice. Distal colon tissue was collected at intervals of 10 (Rag2 KO) or 7 (SCID) days. A group of mice were treated with anti-IL-12p40 starting from day 14 (Rag2 KO) or day 10 (SCID). Colons from 3 animals were pooled and cells were sorted by FACS to separate epithelial (EpCAM+/CD45-), immune (EpCAM-/CD45+) and stroma (EpCAM-/CD45-) populations. Cells were processed using the 10X Chromium system for gene expression and Feature Barcodes.

first: Chiara

last: Sorini

first: Eduardo

last: Villablanca


title: Longitudinal bulk RNA-seq of murine T cell transfer colitis

summary: In order to describe in detail the processes involved in chronic colitis, we performed bulk RNA-seq at multiple time points in CD45RBhi T cell transfer colitis mouse models. These data were compared to scRNAseq and spatial transcriptomics results from the same mice within the same study. Bulk data were analyzed by identifying differentially expressed genes between time points and treatment groups.

overall_design: T cell transfer colitis was induced in Rag2 KO and SCID mice. Distal colon tissue was collected at intervals of 10 (Rag2 KO) or 7 (SCID) days. A group of mice were treated with anti-IL-12p40 starting from day 14 (Rag2 KO) or day 10 (SCID). Bulk RNA-seq was performed on tissue samples. n = 2-3 mice per time point and treatment group


title: Longitudinal spatial transcriptomics of murine IL10KO colitis

summary: In order to describe in detail the processes involved in chronic colitis, we performed spatial transcriptomics at multiple time points in a IL10KO colitis mouse model. These data were compared to bulk RNAseq and scRNAseq (GSE255423) results from the same mice within the same study.

overall_design: Colon tissue was collected from WT mice at age 6 weeks and from IL10 KO at ages 10 and 14 weeks. One mouse collected at week 14 showed more severe colitis with a rectal prolapse.


title: EZH2 inhibition reshapes 3D chromatin architecture to induce immunogenic phenotype in small cell lung cancer [ATAC-Seq]

city: Milwaukee

first: Huizi

state: Wisconsin

address: 8701 Watertown Plank Rd

department: Department of Medicine

institute: Medical College of Wisconsin

type: Genome binding/occupancy profiling by high throughput sequencing

summary: EZH2, the enzymatic core of the Polycomb Repressive Complex 2 (PRC2), is overexpressed in small cell lung cancer (SCLC). Previous studies have shown that EZH2 enforces epigenetic silencing of immune and DNA repair genes, including Class I MHC molecules (HLA‑A/B/C) and SLFN11. To investigate how EZH1/2 inhibition reshapes the 3D chromatin landscape and its relationship to transcriptional regulation, we performed multi‑omic profiling of a neuroendocrine SCLC cell line (NCI‑H146) treated with the dual EZH1/2 inhibitor valemetostat for 9 days. We employed Micro‑C sequencing for high‑resolution 3D genome mapping, ATAC‑sequencing for chromatin accessibility profiling, and RNA‑sequencing for whole‑transcriptome analysis. This is the ATAC sequencing dataset.

overall_design: Cells were treated with the dual EZH1/2 inhibitor Valemetostat (2.5 µM) or DMSO vehicle control for 9 days, with drug-containing media replenished every third day. All sequencing experiments were performed on day 9 post-treatment samples collected in biological replicates.

last: Parveen

first: Victor

middle: X

last: Jin

first: Hui-Zi


title: Spatial Transcriptomic Analysis of APOe3 and APOE4 Mice after Chemotherapy

city: Washington

first: Christopher

last: Albanese

address: 3800 Reservoir Road NW

institute: Georgetown university

summary: Spatial transcriptomic profiling of APOE4 mice post-chemotherapy compared with APOE3 knock-in mice to assess genotype-specific regional gene expression changes.

overall_design: Mouse brain sections were fixed, sectioned and mounted on positively charged slides with 8μm thickness in the Histology and Tissue Shared Resource

first: Olga

last: Rodriguez

first: Naazneen

last: Khan

first: Adrian

last: Oblak

first: Jeff

last: Dage

first: Chris


title: RNA-seq analysis of hypothalamus in female mice reveals effects of membrane-initiated estrogen receptor alpha signaling

city: Göteborg

first: Yiwen

last: Jiang

address: Vita Stråket 11, Klinfarmlab

institute: University of Gothenburg

summary: Membrane-initiated estrogen receptor alpha (mERα) signaling plays an important role in bone metabolism, but its contribution in the brain remains unclear. In this study, we performed RNA sequencing of hypothalamic tissue from 16-week-old female mice with selective inactivation of mERα signaling in POMC neurons (POMC-C451Af/f) and wild-type littermate controls. Total RNA was isolated from hypothalamus, and samples were pooled (two animals per sample) resulting in four biological replicates per genotype. Libraries were prepared using poly(A) selection and sequenced on an Illumina NovaSeq 6000 platform with paired-end 150 bp reads. Transcript abundance was quantified using Salmon and summarized to gene-level counts. Differential expression analysis was performed using DESeq2. This dataset provides insight into transcriptional changes associated with disrupted mERα signaling in hypothalamic neurons.

overall_design: Hypothalamic RNA was collected from 16-week-old female mice of two genotypes: POMC-C451Af/f (mutant) and wild-type controls. For each genotype, RNA from eight animals was pooled in pairs to generate four biological replicates per group. RNA-seq libraries were prepared using poly(A) enrichment and sequenced on an Illumina NovaSeq 6000 platform (paired-end, 150 bp). Differential gene expression analysis was performed comparing POMC-C451Af/f and wild-type samples.

first: Karin

last: Horkeby

first: Petra

last: Henning

middle: H

last: Nilsson

first: Jianyao

last: Wu

first: Lei

last: Li

first: Sofia

last: Movérare-Skrtic

first: Claes

last: Ohlsson

first: Marie

middle: K

last: Lagerquist


title: Progressive cell fate specification in morphallactic regeneration

city: Basel

first: Panagiotis

last: Papasaikas

address: Fabrikstrasse 24

country: Switzerland

institute: Friedrich Miescher Institute

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

overall_design: Refer to individual Series


title: EZH2 inhibition reshapes 3D chromatin architecture to induce immunogenic phenotype in small cell lung cancer [RNA-Seq]

first: Huizi

last: Chen

summary: EZH2, the enzymatic core of the Polycomb Repressive Complex 2 (PRC2), is overexpressed in small cell lung cancer (SCLC). Previous studies have shown that EZH2 enforces epigenetic silencing of immune and DNA repair genes, including Class I MHC molecules (HLA‑A/B/C) and SLFN11. To investigate how EZH1/2 inhibition reshapes the 3D chromatin landscape and its relationship to transcriptional regulation, we performed multi‑omic profiling of a neuroendocrine SCLC cell line (NCI‑H146) treated with the dual EZH1/2 inhibitor valemetostat for 9 days. We employed Micro‑C sequencing for high‑resolution 3D genome mapping, ATAC‑sequencing for chromatin accessibility profiling, and RNA‑sequencing for whole‑transcriptome analysis. This is the RNA‑sequencing dataset.

first: Sana

last: Parveen

first: Victor


title: EZH2 inhibition reshapes 3D chromatin architecture to induce immunogenic phenotype in small cell lung cancer [microC]

summary: EZH2, the enzymatic core of the Polycomb Repressive Complex 2 (PRC2), is overexpressed in small cell lung cancer (SCLC). Previous studies have shown that EZH2 enforces epigenetic silencing of immune and DNA repair genes, including Class I MHC molecules (HLA‑A/B/C) and SLFN11. To investigate how EZH1/2 inhibition reshapes the 3D chromatin landscape and its relationship to transcriptional regulation, we performed multi‑omic profiling of a neuroendocrine SCLC cell line (NCI‑H146) treated with the dual EZH1/2 inhibitor valemetostat for 9 days. We employed Micro‑C sequencing for high‑resolution 3D genome mapping, ATAC‑sequencing for chromatin accessibility profiling, and RNA‑sequencing for whole‑transcriptome analysis. This is the Micro‑C sequencing dataset.

last: Jin

first: Hui-Zi


title: Single cell analysis of clinically resolved epidermal skin from nickel-challenged individuals treated with either anakinra or placebo

city: Copenhagen

first: Kelvin

last: Yeung

address: Blegdamsvej 3B

country: Denmark

institute: LEO SIC

summary: Background: Topical corticosteroids are used as a standard treatment option in allergic contact dermatitis (ACD). However, the treatment doesn’t inhibit the generation of skin-resident memory T (TRM) cells. These cells play an important role in the pathogenesis of skin flare ups in ACD. Currently, findings in murine models suggest interleukin (IL)-1 inhibition can suppress the immune response to contact allergens. Objectives: To determine if IL-1 inhibition can attenuate acute skin flare-ups and restrict the development of TRM cells. Methods:  A murine model was used to investigate the role of IL-1 on epidermal TRM cells by measuring changes in ear thickness and T cell composition. Next, we conducted a randomized, double-blinded, placebo-controlled trial (RCT) [H-22001652; EudraCT number: 2021-004750-39]. Nickel allergic participants were patch-tested at 1) baseline, 2) while being treated with either anakinra or placebo, 3) and 21 days after treatment. The primary outcome was the International Contact Dermatitis Research Group (ICDRG) score 48 hours after patch test application. While secondary outcomes were a reduction of immune cells in clinically cleared and actively inflamed skin. Results: In mice, IL-1 inhibition decreased the number of epidermal CD8+ TRM cells and attenuated skin flare-ups upon re-exposure. Twenty participants were enrolled and completed the RCT, but anakinra failed to improve ICDRG scores. Conclusions: Although IL-1 inhibition has been proven successful in murine models for ACD, there is currently no clinical evidence that suggest it being efficacious as a short-term treatment option for acute skin flare-ups in humans.

overall_design:
Analysis of epidermal skin cells obtained by punch biopsies from preiovusly nickel-challenged clinically cleared skin taken 21 days after challenge. Individuals were either treated with anakinra or placebo for six consecutive days during the challenge.

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>>>Raw data are unavailable due to patient privacy concerns<<<
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title: On the way to elucidate the effects of cannabidiol (CBD) on living organisms - assessment of changes in the global transcription profile of the spleen in a mouse model

city: Kraków

first: Artur

last: Gurgul

address: Rędzina 1C

department: Center for Experimental and Innovative Medicine

country: Poland

institute: University of Agriculture in Krakow

summary:
Cannabidiol (CBD) is a phytocannabinoid that has anti-inflammatory, immunomodulatory, antioxidant, anxiolytic, antipsychotic, and anticonvulsant effects. This study aims to examine the effects of CBD on behavior, immune system activity, and stress levels, and to determine its impact on transcriptomic alterations in selected mouse organs.

(Study funded by the National Science Centre, Poland; project no. 2020/39/O/NZ9/00821)

overall_design: Male mice of the standard C57BL/6J strain were used. The animals were classified into two groups - (I) short-term CBD administration (2 days) and (II) long-term CBD administration (28 days). In each of the main groups, there were three subgroups receiving different doses of CBD - from low, commonly used in supplementation (0.2 mg/kg bw), through medium (10 mg/kg bw), to high, used in some clinical trials (20 mg/kg bw). Additionally, a control group (placebo/vehicle) was maintained in each of the main groups. Chemically pure Cannabidiol (CBD) was administered intraperitoneally (in the research group) to ensure that the substances reached the blood vessels of the digestive system. CBD was administered in a single daily dose. The mice were housed under standard conditions and with standard feeding and watering ad libitum. They were also tested daily (group I) or every third day (group II) to determine well-being and health. After the period of CBD administration, the mice were immediately euthanized and organs such as the brain (various brain structures including hippocampus), spleen, and liver were collected. In this part of the experiment, spleen samples were used to isolate total RNA. The recovered RNA was quality assessed and used to prepare directional libraries for high throughput transcriptome sequencing using QuantSeq 3’mRNA-Seq (FWD) V2 kit (Lexogen).  The libraries were sequenced commercially in a 1x150bp run. The obtained sequence data were used to analyze the differential expression between the different CBD dosages and the control group. Differentially expressed genes were classified into particular processes or functions and were subjected to statistical testing for overrepresentation. The synthesis of the obtained data will allow the proposal of potential mechanisms of CBD action and create an atlas of changes taking place in the transcriptome of the main target organs.

first: Jakub

last: Żurowski

first: Sebastian

last: Sawicki

first: Ewa

last: Ocłoń

first: Tomasz

last: Szmatoła

first: Igor

last: Jasielczuk

first: Karolina

last: Mizera- Szpilka


title: Calling a Truce: Enterococcus faecalis as a mediator between E. coli and C. albicans during polymicrobial CAUTIs

city: South Bend

first: Ellsa

last: Wongso

address: 809 Lindenwood Dr S

institute: University of Notre Dame

summary: Polymicrobial catheter-associated urinary tract infections (CAUTIs) are a severe clinical burden. CAUTIs promotes colonization by different microbes, including E. faecalis, E. coli, and C. albicans. These mixed biofilms can compromise antimicrobial efficacy and promote resistance. Despite their prevalence, the cross-kingdom dynamics driving these infections within the catheterized bladder remain poorly understood. Using established in vitro and in vivo CAUTI models, we elucidate the complex, environment-dependent interactions governing these multispecies biofilms. We demonstrate that E. coli strongly antagonizes C. albicans, by limiting iron uptake, inhibiting filamentation, and inducing cell death. Crucially, we identify E. faecalis as a protective keystone species that mitigates this antagonism. E. faecalis facilitates multispecies coexistence by acting as a spatial bridge on the urothelium, enabling E. coli and C. albicans to co-localize rather than occupy segregated niches. These mechanistic insights into polymicrobial synergy and competition provide a crucial framework for developing targeted therapeutics against recalcitrant CAUTIs.

sample_organism:
Escherichia coli
Enterococcus faecalis
Candida albicans

overall_design: The objective of the RNAseq is to compare transcriptomes of single species biofilm with double species and triple species biofilms. In this project, I used three different species of microorganisms: Enterococcus faecalis, Escherichia coli, and Candida albicans. The cells were grown in urine (in 12 well plates) for 48 hours and treated with TRI-zol. EF stands for E. faecalis single biofilm, EC stands for E. coli single biofilm, CA stands for C. albicans biofilm. EF-EC means it is a combination between E. faecalis and E. coli biofilm (two species), EF-CA is E. faecalis with C. albicans, EC-CA is E. coli and C. albicans, and EF-EC-CA is E. faecalis, E. coli, C. albicans biofilm (triple species biofilm). I want to see how the transcriptomes of each species is different in single biofilm compare to two species and three species biofilm. The numbering is of different replicates.

first: Ana

last: Flores-Mireles


title: Translocator protein deficiency promotes splenomegaly with extramedullary hematopoiesis after sepsis

city: Minami-ku

first: Kazuya

last: Kikutani

state: Hiroshima

address: 1-2-3 Kasumi

department: Department of Emergency and Critical Care Medicine, Graduate School of Biomedical and Health Sciences

country: Japan

institute: Hiroshima Univeristy

summary: This study investigates the role of translocator protein (TSPO) in splenic immune responses following sepsis using a mouse cecal ligation and puncture (CLP) model. Bulk RNA sequencing of spleen tissue collected 17 days after sham or CLP surgery revealed that TSPO deficiency promotes splenomegaly characterized by red pulp expansion, enhanced erythropoiesis, and reduced immune responses. Gene expression and deconvolution analyses indicated decreased lymphocyte signatures and increased erythroblast and megakaryocyte signatures in TSPO knockout mice. These findings suggest that TSPO plays a critical role in maintaining splenic immune homeostasis after sepsis.

overall_design: Male C57BL/6J wild-type (WT) and TSPO knockout (TSPOKO) mice were subjected to cecal ligation and puncture (CLP) to induce sepsis or sham surgery. Spleen tissues were collected 17 days after surgery. Bulk RNA sequencing was performed on spleen samples from three groups: WT-Sham, WT-CLP, and TSPOKO-CLP (n = 8 per group). Gene expression profiles were analyzed to investigate the effects of TSPO deficiency on post-septic splenic immune responses and hematopoietic remodeling.

first: Hidenori

last: Aizawa

first: Xiao

last: Lyu


title: Dynamic acetylation/deacetylation dictates TRIM27 nuclear-cytoplasmic localization for cancer progression [CUT&Tag II]

city: Zhengzhou City

first: Leixi

last: Cao

address: Zhengzhou University People's Hospital, Fengchan Road Subdistrict, Jinshui District, Zhengzhou City, Henan Province

institute: Zhengzhou University People's Hospital, Translational Medicine Research Center

summary: Cancer invasion and metastasis are orchestrated by intricate molecular mechanisms that remain not fully understood. We have identified that the tripartite motif-containing protein 27 (TRIM27), typically known as an E3 ubiquitin ligase, plays dichotomous roles in cancer progression. These roles are determined by its subcellular localization, which is regulated by acetylation and deacetylation at lysine 297 (K297) independently of its ubiquitin ligase activity. Acetylation by general control non-repressed protein 5 (GCN5) directs TRIM27 to the nucleus, where it inhibits cancer cell invasion and metastasis by modulating the transcription of paired box 6 (PAX6). Conversely, deacetylation by histone deacetylase 4 (HDAC4) results in cytoplasmic localization, promoting cell proliferation and tumor growth. In the nucleus, TRIM27 binds to poly (ADP-ribose) polymerase 1 (PARP1) and negative elongation factors (NELFs), repressing PARP1-mediated mono-ADP-ribosylation of NELFA, thereby preventing the release of RNA polymerase II from transcriptional pausing, and leading to transcriptional repression of PAX6. Furthermore, we demonstrate that treatment with the PARP1 inhibitor Olaparib limits the metastasis of cancer cells lacking breast cancer gene (BRCA) mutations, uncovering a previously overlooked therapeutic potential. Importantly, the combination of Olaparib and the HDAC4 inhibitor Tasquinimod synergistically inhibits cancer metastasis. Collectively, these findings reveal a complex interplay of post-translational modifications that dictate the subcellular localization and biological functions of TRIM27, highlighting novel strategies for effective cancer treatment.

overall_design: Genome-wide CUT&Tag sequencing was performed to profile the chromatin occupancy of histone marks (H3K4me1, H3K4me2, H3K4me3, H3K27ac) and key transcription regulatory factors (CDK9, RNA polymerase II p-CTD-S5). All samples were prepared with independent biological replicates. This study aims to  elucidate the regulatory role of the target protein TRIM27 in the process of eukaryotic gene transcription and its underlying association with transcriptional initiation and elongation.

first: Xuedan

last: Sun

first: Ruijie

last: Wang


title: Longitudinal spatial transcriptomics of murine T cell transfer colitis

first: Jennifer

last: Fransson

summary: In order to describe in detail the processes involved in chronic colitis, we performed scRNA-seq at multiple time points in a CD45RBhi T cell transfer colitis mouse model. These data were compared to bulk RNAseq and scRNAseq results from the same mice within the same study. Spatial trancriptomics were analyzed by performing NMF to identify spatial patterns of gene expression.

overall_design: T cell transfer colitis was induced in Rag2 KO mice. Distal colon tissue was collected at days 0, 30 and 50. One mouse collected at day 50 was treated with anti-IL-12p40 starting from day 14.

first: Chiara

last: Sorini

first: Eduardo

last: Villablanca


title: Targeting Cyclin K-CDK12 Synergizes with ATR Inhibition by Limiting RPA Chromatin Loading in Triple-Negative Breast Cancer [ChIP-seq]

city: Boston

first: Eunbee

last: Choi

state: MA

address: 440 Brookline Ave, Mayer-434

institute: Dana farber cancer institute

summary: Cyclin K-CDK12 inhibition has emerged as a promising therapeutic strategy to induce homologous recombination deficiency (HRD). However, clinical and genomic studies have not clearly linked CDK12 inactivation to HRD. Here, we demonstrate that cyclin K-CDK12 depletion drives a replication stress (RS)-associated vulnerability through RPA regulation. A DNA damage response-focused CRISPR screen identified DNA replication factors and ATR activators as top sensitizers to cyclin K degradation. Consistently, cyclin K-CDK12 depletion synergized with ATR inhibition in triple-negative breast cancer (TNBC) models. Cyclin K degradation reduces RPA chromatin loading and impairs ATR activation, compromising cellular tolerance to RS and increasing susceptibility to ATR inhibition. We show that CDK12-mediated phosphorylation of CDC5L is required for RPA chromatin loading and mediates resistance to combined cyclin K and ATR depletion. Our findings identify cyclin K-CDK12 as a critical regulator of RPA dynamics and support its inhibition as rational therapy for RS-high tumors, including TNBC.

overall_design: ChIP-seq of RPA2 in asynchronous MDA-MB-231 cells treated with vehicle (DMSO) or the cyclin K protein degrader CT7311 (10 nM, 24 h) to evaluate the global enrichment of RPA2.

first: Eun-Bee

first: Geoffrey

middle: I

last: Shapiro


title: PDHA1 Crotonylation Is Essential for Endometrial Metabolic Reprogramming and Decidualization in Normal Pregnancy

city: Shanghai

first: Yifei

last: Sang

address: 910 Hengshan Rd

institute: Shanghai Jiao Tong University

summary: Endometrial decidualization is essential for pregnancy. Impaired decidualization is a hallmark of preeclampsia (PE), yet the underlying mechanisms remain unclear. Herewe show that uterine pyruvate dehydrogenase E1 subunit α (PDHA1) K385 crotonylation (K385cr) is crucial in metabolic reprogramming and decidualization. In PE women, decidual PDHA1 K385cr is markedly reduced. In mice, Pdha1 K385cr deficiency causes endometrial metabolic dysregulation, poor decidualization, and a PE-like phenotype. Mechanistically, crotonylation promotes PDHA1 phosphorylation, inhibiting pyruvate dehydrogenase activity and enhancing glycolysis and NAD+ generation. In turn, NAD+ increases SIRT2-mediated FOXO1 deacetylation and nuclear translocation, upregulating decidual gene expression. In Pdha1 K385cr deficient mice, treatment of the NAD⁺ precursor β-NMN enhances the NAD⁺-SIRT2 FOXO1 pathway and decidualization, alleviating the PE-like phenotype. Our results indicate that PDHA1 K385cr is a key metabolic regulator in the pregnant uterus and that impaired PDHA1 K385cr may contribute to PE, suggesting that enhancing the NAD⁺-SIRT2-FOXO1 pathway may be exploited for PE intervention.

overall_design: RNA-seq profiling of human endometrial stromal cells before and after in vitro decidualization.

first: Jiajia

last: Chen

first: Tingxuan

last: Yin

first: Yang

last: Yan

first: Chunfang

last: Xu

first: Ling

first: Weijie

last: Zhao


title: Dynamic acetylation/deacetylation dictates TRIM27 nuclear-cytoplasmic localization for cancer progression [CUT&Tag]

overall_design: Genome-wide CUT&Tag profiling of TRIM27, RNA polymerase II CTD, and p-CTD-S2 chromatin occupancy in MDA-MB-231 breast cancer cells, following TRIM27 genetic knockout, PARP1 siRNA-mediated knockdown, and Olaparib-induced PARP1 inhibition. All experimental groups included biological replicates, with the goal of dissecting the TRIM27-PARP1 regulatory axis driving breast cancer invasion and metastasis.

first: Mian

last: Wu


title: Transcriptomic profiling reveals complement activation in chronic pancreatitis adjacent to pancreatic ductal adenocarcinoma

first: Mehran

last: Ghaderi

state: Huddinge

address: Forskningsgatan 46

department: Clintech

summary: Short abstract: Chronic pancreatitis (CP) and pancreatic ductal adenocarcinoma (PDAC) frequently coexist, yet distinguishing inflammatory changes secondary to malignancy from primary pancreatitis remains challenging. Here, we present an integrated multi-omics analysis of spatially distinct pancreatic tissue compartments obtained from a single patient undergoing pancreaticoduodenectomy for PDAC, combining histopathology, transcriptomics, immunofluorescence, and meta-transcriptomic microbial profiling. Morphological assessment identified tumor tissue, adjacent chronic pancreatitis, and normal pancreas. Transcriptomic profiling of macro-dissected samples revealed differential gene expression and enrichment of the classical complement pathway in the CP compartment, which was qualitatively supported by immunofluorescent detection of C1q and C3 along the ductal epithelium. Meta-transcriptomic analysis detected a limited number of bacterial taxa and no viral RNA; these findings were interpreted conservatively given the constraints of low-biomass tissue profiling and the single-patient design. Although causal inference and generalizability are limited by the single-patient design, this study demonstrates the feasibility of integrating pathology with transcriptomic and microbial analyses to generate hypothesis-generating, multi-omics framework for exploring inflammatory-malignant interactions in pancreatic disease.

overall_design: This exploratory single-case study used spatially resolved multi-omics analysis of pancreatic tissue from a PDAC patient. Tumor, adjacent chronic pancreatitis (CP), and normal tissue were identified histologically and macro-dissected from FFPE sections. RNA sequencing was performed to assess differential gene expression and pathway enrichment analysis. Complement activation findings were qualitatively validated by immunofluorescence (C1q, C3) in CP tissue, and RNA-seq data were additionally analyzed for microbial signatures. Transcriptome data were processed and analyzed using the Chipster virtual platform provided by the IT Center for Science (CSC), Finland. Briefly, adapter sequences were trimmed, and sequencing read quality was assessed with FastQC. Paired-end reads (40-45 million per sample) were aligned to the human reference genome (GRCh38.95) using HISAT2 aligner and assembled using Cufflinks.Transcript assembly and quantification were performed and followed by differential expression analysis using Cuffdiff across three datasets. Differentially expressed genes (DEGs) were identified based on standard statistical thresholds. Overrepresentation analysis of differentially expressed genes was performed using ConsensusPathDB, integrating pathways from over 30 publicly available databases.The aim was to integrate molecular and pathological data to characterize differences between tumor, inflamed, and normal pancreatic compartments.

first: Arun Kumar

last: Selvam

first: Ville N

last: Pimenoff

first: Mikael

last: Björnstedt

first: Joakim

last: Dillner

first: Carlos Fernández

last: Moro

first: Soledad Pouso

last: Elduayen


title: ATAD2 is a Novel Regulator of Myogenesis and Autophagy in Skeletal Muscles

city: Montreal

first: Sabah

last: Hussain

address: 1001 Bd Décarie

country: Canada

institute: McGill University Health Centre

summary: MYTHO protein is a recently described novel regulator of autophagy and mass in skeletal muscles.  Using immunoprecipitation assay, we found that the nuclear protein ATAD2 (ATPase family AAA domain containing 2) interacts with MYTHO protein in C2C12 myoblasts. The functional role of ATAD2 in skeletal muscle cells was assessed by using transient and stable Atad2 knockdown (KD) in C2C12 myoblasts. Atad2 KD significantly increased myoblast proliferation rate, S-phase entry, overall cell viability, and early differentiation into myotubes.  However, Atad2 KD also elicited myotube atrophy and accumulation of autophagy-related proteins due to inhibition of autophagosome fusion with lysosomes and/or lysosome dysfunction. LAMP1 staining confirmed the presence of enlarged lysosomes in Atad2 KD cells. Additionally, genes involved in lysosome function and integrity including Rab7, Rab29, Nrbf2, and Cathepsin L were downregulated in Atad2 KD cells. Collectively, these results indicate that ATAD2 is a critical regulator of muscle cell proliferation, differentiation, and autophagy.

overall_design: RNA profiling of wildtype C2C12 myoblasts cells and their ATAD2 knockdown

first: Sami

last: Sedraoui

first: Alaa

last: Moamer

first: Tomer

last: Chaffer

first: Jean-Philippe

last: Leduc-Gaudet

first: Felipe

last: Broering

first: Dominique

last: Mayaki

first: Ryann

last: Lang

first: Minna

last: Woo

first: Yumin

last: Zheng

first: Jun

last: Ding

first: Marco

last: Sandri

first: Gilles

last: Gouspillou


title: Generation of a prop1 knockin zebrafish enables single-cell transcriptomics of early pituitary development

city: Ciudad Autonoma de Buenos Aires

first: Maria

middle: Ines

last: Perez Millan

state: Buenos Aires

address: Ciudad Universitaria

department: IFIBYNE

country: Argentina

institute: UBA-CONICET

summary: The adenohypophyseal placode is a developmental structure that is conserved among vertebrates and gives rise to the anterior pituitary. The earliest steps of its formation remain poorly understood. Here, we present a powerful new genetic tool: a zebrafish GAL4 knockin reporter line for prop1. The targeted integration disrupts prop1 function, generating the first prop1-/- zebrafish, which exhibits dwarfism, impaired sexual maturation, and pigmentation defects. The generation of this new reporter line enabled the study of early pituitary development at single cell resolution, using both scRNA-seq and live imaging. We identified epcam/Epcam as a conserved epithelial marker of the adenohypophysis in both zebrafish and mouse. We further mapped transcriptional programs underlying endocrine lineage specification, discovering several previously unrecognized markers for hormone producing lineages, including tfcp2l1, as a likely regulator of lactotrope differentiation. Cell–cell interaction analyses revealed Notch, Fgf, and Slit/Robo signaling as key regulators of progenitor maintenance and differentiation. Overall, this work provides the first single cell atlas of early zebrafish pituitary development, a robust new genetic tool to study fundamental mechanistic insights into pituitary lineage specification and differentiation

overall_design: Cells sorted from pituitary (prop1+) zebrafish at 28 and 52 hpf

first: Julian

last: Martinez Mayer

first: Maria Ines


title: Melatonin suppresses ILC2-driven airway hyperreactivity via glutathione-dependent metabolic reprogramming

city: Los Angeles

first: Jafar

last: Cain

state: CA

address: 1450 Biggy St

department: MMI

institute: University of Southern California

summary: Allergic asthma is characterized by type 2 inflammation and overnight worsening of symptoms, yet dynamic fluxes in cellular metabolic profiles driving time-of-day variation remain poorly defined. Group 2 innate lymphoid cells (ILC2s) are central mediators of airway hyperreactivity. We identify melatonin as a previously unrecognized regulator of ILC2 metabolism and function. In murine models of allergic airway inflammation, melatonin reduced eosinophilia, type 2 cytokine production, and airway hyperreactivity without altering ILC2 abundance. Mechanistically, melatonin acted independently of canonical melatonin receptors and instead reprogrammed ILC2 metabolism toward pentose phosphate pathway activity, enhancing NADPH generation and NRF2-dependent glutathione accumulation. Metabolic profiling, loss-of-function approaches, and pharmacologic activation studies demonstrated that NRF2 is both necessary and sufficient to restrain ILC2 effector function. Importantly, primary human ILC2s exhibited conserved NRF2 activation, glutathione accumulation, and reduced type 2 cytokine production in response to melatonin, underscoring clinical relevance. Together, these findings identify the melatonin-NRF2-glutathione axis as a metabolic checkpoint regulating innate type 2 immunity and suggest that therapeutic targeting of redox metabolism may represent a strategy for modulating airway inflammation in allergic asthma.

overall_design: mRNA profiles of sorted mouse lung ILC2s treated ex-vivo +/- Melatonin were generated by deep sequencing, in triplicate, using a NextSeq 500 (Illumina) system.

first: Benjamin

middle: P

last: Hurrell

first: Stephen

last: Shen

first: Paul

last: Speliakos

first: Omid

last: Akbari


title: Adulthood depletion of Integrator extends lifespan and healthspan via defective pre-mRNA processing [smallRNA-seq]

city: Oxford

first: Luke

last: Slade

address: Sherrington Rd, Oxford

department: Biochemistry

country: United Kingdom

institute: University of Oxford

type: Non-coding RNA profiling by high throughput sequencing

summary: Identifying strategies to mitigate age-related physiological decline remains a central challenge. Transcriptional regulation is essential to organismal healthspan and a frequent therapeutic target in ageing research. The Integrator complex is a highly conserved regulator of transcriptional quality control and RNA processing, yet its role in ageing within an intact organism remains unknown. Using adult-specific RNAi in Caenorhabditis elegans, we find that depletion of most Integrator subunits extends lifespan and healthspan. Mechanistically, loss of the catalytic subunit INTS11 disrupts 3′ end formation of small nuclear and spliced-leader RNAs, impairing trans-splicing and promoting outron retention on a subset of transcripts enriched for spliceosomal, nucleocytoplasmic import and mitochondrial functions. These RNA-processing defects engage endogenous siRNA pathways, which are required for the longevity and healthspan benefits of INTS11 depletion, consistent with a role in buffering the cytoplasmic burden of misprocessed splicing and import factors. In parallel, outron retention compromises nuclear-encoded mitochondrial gene expression and induces a hormetic decline in mitochondrial function. Together, our findings identify Integrator as an upstream regulator of a coupled RNA-processing–small RNA–mitochondrial axis that shapes the pace of ageing.

overall_design: C. elegans were grown at 20 degrees on regular OP50-seeded NGM plates until the L4 (day 0) larval stage. At day 0, animals were transferred onto either empty vector (L4440) or ints-11 dsRNA bacteria-seeded RNAi plates. Animals were transferred every 2-days onto fresh RNAi plates seeded with empty vector or ints-11 dsRNA bacteria until sample collection. For collections, samples were washed from plates and cleaned five times in 1mL of M9 buffer by gravity flotation. Supernatants were removed to a final volume of 30 uL and 500 uL of TriZol reagent was added before snap freezing in liquid nitrogen. Samples were stored at -70 degrees until further processing for RNA extraction.

first: Peter

last: Sarkies

first: Lidia

last: Vasiljeva


title: Adulthood depletion of Integrator extends lifespan and healthspan via defective pre-mRNA processing [ChIP-seq]

overall_design: ChIP-seq in JCP643 ints-11(jcp31)[ints-11::3xFLAG])) animals to map INTS11 binding loci in day-1 adult C. elegans.

first: Krzysztof

last: Kuś


title: MERVL-driven Pcgf5 plays a critical role in the suppression of zygotic genome activation in preimplantation embryos

city: Kyoto

first: Shinnosuke

last: Honda

address: Kitashirakawa Oiwake-cho, Sakyo-ku, Kyoto-shi

institute: Kyoto University

summary: We obtained transcriptomic profiles in Pcgf5 KD late two-cell embryos and morulae, and Pcgf5MT2C_Mm KO four-cell embryos. H2AK119ub1 ChIP-seq profiles in Pcgf5 KD molurae were also performed.

overall_design: In vitro produced mouse embryos were processed to RNA-Seq and ChIP-seq. The experiments were duplicated using ten embryos per group for RNA-seq and ~40 embryos per group for ChIP-seq.

first: Satoshi

last: Mashiko

first: Yuzuki

last: Goto

first: Takuto

last: Yamamoto

first: Tomoyuki

last: Tsukiyama

first: Naojiro

last: Minami

first: Shuntaro

last: Ikeda


title: Elongin B orchestrates chromatin and transcriptional programs in H3K27M‑mutant diffuse midline glioma [XIII_ELOBKO_RNAseq]

city: Houston

first: Jamie

middle: N.

last: Anastas

state: TX

address: 1 Baylor Plaza

department: Neurosurgery

institute: Baylor College of Medicine

summary: Recurrent driver mutations in genes encoding histone H3 (H3.3K27M and H3.1K27M) are observed in ~80% of diffuse midline gliomas (DMG), which lead to aberrant gene regulation, yet the specific RNA polymerase 2 (Pol2) regulators inducing aberrant DMG transcription are not fully defined. We identified multiple regulators of Pol2 elongation as DMG genetic dependencies in a chromatin-focused CRISPR screen. Additional studies confirm that knockout (KO) of the Pol2 SIII complex gene Elongin B (ELOB) inhibits DMG cell proliferation in tissue culture and tumor growth in xenograft models. Further genomic analyses reveal that ELOB binding sites are enriched in H3K27M oncohistones and that ELOB knockout alters H3K27me3 and H3K27M incorporation at thousands of genomic regions, suggesting a role for ELOB in maintaining dysfunctional chromatin states in DMG. Correspondingly, ELOB loss disrupts Pol2 transcriptional activity and alters the expression of transcripts involved in metabolism, proliferation, and brain development. These findings suggest that Pol2 elongation factors cooperate with H3K27M oncohistones to maintain the epigenetic and transcriptional landscape driving DMG malignancy.

overall_design: Cas9+ BT245 or SU-DIPGXIII cells were transduced with control or ELOB-targeting sgRNAs and selected with 250 ug/mL hygromycin 48 hours later. Equal live cell numbers were collected for RNAseq analysis at day 14 following the start of hygromycin selection and processed for bulk RNAseq of polyA transcripts.

first: Adam

last: Kebede

middle: N


title: LuxR regulates Flavobacterium covae pathogenesis in channel catfish (Ictalurus punctatus)

city: Auburn

first: Mark

last: Liles

state: AL

address: Room 101 Rouse Life Sciences Building

department: Biological Sciences

institute: Auburn University

summary: Columnaris disease, caused by Flavobacterium covae, is one of three primary bacterial diseases that cause significant mortality and economic losses in farmed catfish (Ictalurus spp.). The quorum sensing gene luxR was reported to be upregulated in F. covae biofilm-grown cells stimulated with channel catfish (I. punctatus) mucus. To understand LuxR’s contributions to F. covae virulence, a F. covae C#2 luxR deletion mutant (ΔluxR) was created along with a complemented luxR (ΔluxR + pCE1). Channel catfish fingerlings challenged with the wild type or ΔluxR + pCE1 were observed to have 100% or 90% mortality, respectively; in contrast, only 2% mortality was observed for catfish fingerlings challenged with ΔluxR, indicating the important role for LuxR-regulated genes in F. covae virulence. A transcriptomic analysis indicated that LuxR-regulated genes in F. covae encode many functions including those associated with virulence, bacterial growth and metabolism, and survival during a fish infection. A decrease in biofilm formation was observed for ΔluxR relative to the wild-type and the complemented mutant, but only when mannose was included in the growth medium. The results of this study indicated that LuxR is a critical regulator of F. covae quorum sensing and virulence, warranting further research to identify and describe the relative contributions of LuxR-regulated virulence factors.

overall_design: RNA-seq profiling of wildtype Flavobacterium covae C#2 compared to a luxR gene deletion compared to a luxR gene deletion that was complemented with a plasmid containing the wild-type luxR gene


title: Elongin B orchestrates chromatin and transcriptional programs in H3K27M‑mutant diffuse midline glioma [XIII_K27MKO_CUTandRUN]

overall_design: For CUT&RUN, a single cell suspension of 5x105 control or H3K27MKO SU-DIPGXIII+Cas9 cells were immobilized on ConA beads, permeabilized with 0.01% Digitonin, incubated with primary antibodies, and then cut with pAG-MNase to release DNA fragments as previously described. The following antibodies were used in CUT&RUN analyses: H3K27M (cat# ABE419, Millipore Sigma),  H3K27me3 (cat#07-449, ActiveMotif). Yeast spike-in DNA (S. cerevisiae; Cell Signaling Technology) was added at 0.5% to each reaction following chromatin digestion.

first: Alan

middle: L

last: Jiao


title: Mitochondrial transfer from astrocytes to endothelial cells reinforces redox homeostasis and barrier integrity in brain arteriovenous malformation

city: Luzhou

first: zhenghong

last: peng

address: No. 25, Taiping Street, Jiangyang District

institute: The Affiliated Hospital of Southwest Medical University

summary: Brain arteriovenous malformation (BAVM) is a high-flow vascular shunt that predisposes to intracranial hemorrhage and represents a leading cause of stroke in young adults. Somatic mutations in KRAS or BRAF drive endothelial dysfunction and lesion formation, yet the contribution of non-endothelial cells within the neuro-glial-vascular unit remains incompletely understood.

overall_design: In this study, we investigated the role of astrocyte-to-endothelial mitochondrial transfer in maintaining cerebrovascular integrity during BAVM pathogenesis using a genetically engineered mouse model that recapitulates sporadic human BAVM. We show that endothelial KRAS activation disrupts astrocyte-endothelial crosstalk and impairs mitochondrial donation from astrocytes to endothelial cells. Loss of this metabolic support leads to endothelial redox imbalance, barrier instability, and aggravated hemorrhage. Restoration of mitochondrial transfer via targeted delivery of astrocytic mitochondria rescues endothelial dysfunction and reduces lesion severity. These findings reveal that intercellular mitochondrial transfer is a critical homeostatic mechanism in the neurovascular unit and identifies a novel metabolic vulnerability in BAVM.

first: Tianqi

last: Tu

first: Zhenghong

last: Peng


title: PDGF signaling regulates the adipocyte-to-fibroblast transition in wound healing

city: Oklahoma City

first: Lorin

middle: E.

last: Olson

state: OK

address: 825 NE 13th St,

department: Cardiovascular Biology

institute: Oklahoma Medical Research Foundation

summary: Adipocytes are a plastic cell type with the ability transition into fibroblast-like cells during tissue repair or physiological remodeling. How such transitions are regulated, and whether they exemplify cell plasticity or merely a transient suppression of adipocyte features, is unclear. Fibroblasts express platelet-derived growth factor receptors (Pdgfra and Pdgfrb) while adipocytes express adiponectin (Adipoq). We developed two-step lineage tracing to label Pdgfra+ and Pdgfrb+ cells originating from Adipoq+ adipocytes. Adipocyte-derived Pdgfr+ cells (ADPCs) are induced adjacent to skin wounds and contribute to the wound fibroblast population. ADPCs lose adipocyte characteristics and acquire fibroblast characteristics including collagen expression, smooth muscle actin expression, and cell proliferation. Active PDGFRalpha signaling in ADPCs increases cell proliferation and improves wound angiogenesis and re-epithelialization, while deletion of Pdgfra has the opposite effect. Wound ADPCs persist as fibroblast-like cells after completion of wound healing, indicating an enduring change rather than a transient downregulation of adipocyte characteristics.

overall_design: Translating RNA profiling by bulk RNA sequencing in NuTRAP mice

first: Sunhye

last: Jeong

first: Hae Ryong

last: Kwon

middle: E


title: Elongin B orchestrates chromatin and transcriptional programs in H3K27M‑mutant diffuse midline glioma [XIII_ELOB_CUTandRUN]

overall_design: For CUT&RUN, a single cell suspension of control or H3K27MKO SU-DIPGXIII+Cas9 cells were immobilized on ConA beads, permeabilized with 0.01% Digitonin, incubated with primary antibodies, and then cut with pAG-MNase to release DNA fragments as previously described. The following antibodies were used in CUT&RUN analyses: ELOB (cat#ab45297, Abcam), ELOA (cat#, Abcam), H3K27M (cat# ABE419, Millipore Sigma),  H3K27me3 (cat#07-449, ActiveMotif). Yeast spike-in DNA (S. cerevisiae; Cell Signaling Technology) was added at 0.5% to each reaction following chromatin digestion.


title: Dynamic acetylation/deacetylation dictates TRIM27 nuclear-cytoplasmic localization for cancer progression

first: Leixi

last: Cao

summary: Cancer invasion and metastasis are governed by intricate molecular mechanisms. We demonstrate that TRIM27, an E3 ubiquitin ligase, plays dichotomous roles in cancer depending on its subcellular localization, which is controlled by acetylation/deacetylation at K297 independent of its ligase activity. Acetylation by GCN5 directs TRIM27 to the nucleus, while deacetylation by HDAC4 promotes cytoplasmic localization and cell proliferation. Nuclear TRIM27 binds PARP1 and NELFs, represses PARP1-mediated NELFA ADP-ribosylation, maintains RNA polymerase II pausing, and suppresses PAX6 transcription, thereby inhibiting cancer invasion and metastasis.

overall_design: RNA-seq was performed on MDA-MB-231 cells with TRIM27 knockout, PARP1 knockdown (siPARP1), or PARP1 enzymatic inhibition (Olaparib) to dissect transcriptomic changes underlying the TRIM27-PARP1-PAX6 axis in cancer invasion and metastasis.

first: Ruijie

last: Wang

first: Xuedan

last: Sun

first: Mian

last: Wu


title: Adulthood depletion of Integrator extends lifespan and healthspan via defective pre-mRNA processing [RNA-seq]

first: Luke

last: Slade

overall_design: C. elegans were grown at 20 degrees on regular OP50-seeded NGM plates until the L4 (day 0) larval stage. At day 0, animals were transferred onto either empty vector (L4440) or ints-11 dsRNA bacteria-seeded RNAi plates. Animals were transferred every 2-days onto fresh RNAi plates seeded with empty vector or ints-11 dsRNA bacteria until sample collection at defined ages.

first: Krzysztof

last: Kuś

first: Peter

last: Sarkies

first: Lidia

last: Vasiljeva


title: Elongin B orchestrates chromatin and transcriptional programs in H3K27M‑mutant diffuse midline glioma [XIII_ELOBKO_chIP_ELOB_RNAPII]

first: Jamie

last: Anastas

overall_design: ChIPseq for ELOB or total RNA Polymerase II (RNAPII) was performed on SU-DIPGXIII cells expressing Cas9 and either a cotol or ELOB-targeting sgRNA.


title: A progeria syndrome links DNA hypermethylation to age-related pathology [EM-Seq]

city: Edinburgh

first: Dan

last: Sarni

address: Crewe Road

institute: University of Edinburgh

type: Methylation profiling by high throughput sequencing

summary: Declining tissue function and regenerative capacity underlie many chronic diseases. Experimentally establishing the mechanistic basis for such tissue ageing presents substantial challenges, given decades-long timescales and multifactorial origins. Epigenetic alterations have been proposed to have a key aetiological role, but whether they are correlative or causal remains a key unanswered question, as does their contribution to specific age-related pathologies. Here, we establish a novel epigenetically-driven accelerated ageing syndrome. We demonstrate that DNMT3A gain-of-function mutations in Heyn-Sproul-Jackson syndrome recapitulate age-related gains in DNA methylation, cause multilineage stem cell dysfunction, and phenocopy aspects of ageing in humans and mice. We also show that region-specific DNA hypermethylation at lineage-specific genes can explain reduced stem cell output and lineage skewing. Hence, starting from a Mendelian disorder, we causally implicate DNA methylation-mediated stem cell exhaustion in the aetiology of medically important age-related haematological, bone and metabolic pathologies, that might be targetable by future therapies.

overall_design: Enzymatic Methyl-seq (EM-seq) of Hematopoietic stem cells (HSCs) from wild-type and Dnmt3a W326R/+ mice. Nine individual mice were randomly pooled into three samples per genotype for library preparation; s1,s4,s6 = 9 month old; s2,s5 = 3 month old; s3 = 10 month old.

first: Duncan

last: Sproul

first: Andrew

last: Jackson


title: A progeria syndrome links DNA hypermethylation to age-related pathology [vdj_seq]

overall_design: Pro B cells from bone marrow of 3 Dnmt3a +/+ and 3 Dnmt3a W326R/+ mice were isolated by FACS as: CD43+ve, B220+ve, IgM-ve. DNA was extracted by phenol-chloroform extraction. Libraries were constructed using the VDJ seq protocol (PMID: 29725123)


title: CTCF overexpression in K562

city: Santander

first: Manuel

last: Rosa-Garrido

address: Einstein 22

department: Cellular and Molecular Signaling

country: Spain

institute: The Institute of Biomedicine and Biotechnology of Cantabria

summary:
Erythropoiesis is tightly regulated by lineage-specific transcription factors that govern erythroid commitment, proliferation, and differentiation. A core erythroid transcriptional network, together with additional transcription factors and non–DNA-binding cofactors, occupies regulatory regions of genes essential for erythroid development. This process is further shaped by epigenetic mechanisms, including histone post-translational modifications and long-range chromatin interactions.
We used microarrays to detail the global programme of gene expression underlying erythroid differentiation after CTCF overexpresison.

overall_design: Differential expression analysis was performed by comparing Control KpCDNA cells (baseline group) with CTCF-overexpressing KCTCF-D11 cells (experimental group).

first: Maria

last: Dolores Delgado Villar


title: DDX3X-Mediated Translation of Structured Cardiac mRNAs is Essential for Female Heart Development [eCLIP-seq]

city: Chapel Hill

first: Kayla

last: Mason

state: NC

address: 133 N Medical Drive

institute: University of North Carolina Chapel Hill

summary: Sex differences influence congenital heart disease (CHD) development, yet underlying molecular mechanisms remain largely unclear. We demonstrate that the X-linked RNA helicase DDX3X associates in the heart with ribosomal subunit proteins, and eCLIP mapping reveals its preferential binding to cardiac mRNAs with long, structured 5′ untranslated regions (5′UTRs) that can hinder translation. Using a cardiomyocyte-specific mouse Ddx3x knockout model, we show that female embryos lacking Ddx3x die at mid-gestation from heart failure due to impaired translation of key cardiac regulators, whereas male littermates survive. Ribosome profiling and proteomics demonstrate that DDX3X is required for efficient translation of female-differential cardiac mRNAs. Reporter assays confirm that translation of essential cardiac genes such as Srf and Rcor2 depends on their 5′ UTRs and requires DDX3X. These findings uncover a sex-specific post-transcriptional mechanism by which DDX3X safeguards female heart development through selective mRNA translation, providing insight into how X-linked dosage-sensitive regulators contribute to CHD.

overall_design: Eclipse Bionnovations Inc (San Diego, www.eclipsebio.com) performed eCLIP studies according to the published single-end seCLIP protocol with the following modifications. Approximately 80 mg of tissue (n=3 replicates, 8 Ddx3x-3XHA/+ P0-P2 hearts/replicate) was cryoground and UV crosslinked at 400 mJoules/cm2 with 254 nm radiation. Tissue was separated into 30 mg aliquots for each protein and then stored at -80 C until use. Thirty mg of tissue was lysed using 1 mL of eCLIP lysis mix with a modified composition of 6 uL of Proteinase Inhibitor Cocktail and 20 uL of Murine RNase Inhibitor. Samples were then subjected to two rounds of sonication for 4 minutes with 30 second ON/OFF at 75% amplitude. Validated antibodies were then pre-coupled to Anti Rabbit IgG Dynabeads (ThermoFisher), added to lysate, and incubated overnight at 4 C. Prior to immunoprecipitation, 2% of the sample was taken as the paired input sample, with the remainder magnetically separated and washed with eCLIP high stringency wash buffers. Immunoprecipitation and input samples were cut from the membrane at the relative band size to 75 kDa above. RNA adapter ligation, IP-western, reverse transcription, DNA adapter ligation, and PCR amplification were performed as described. The eCLIP cDNA adapter contained a sequence of 10 random nucleotides at the 5' end. This random sequence served as a unique molecular identifier (UMI) after sequencing primers were ligated to the 3' end of cDNA molecules. Samples were subsequently sequenced on a NextSeq 2000. Data was analyzed using a modified single-end seCLIP protocol with Eclipsebio's proprietary analysis pipeline (v1). Unique molecular identifiers (UMIs) were pruned from read sequences using UMI-tools (v1.1.1). Next, 3' adapters were trimmed from reads using Cutadapt (v3.2), and reads <18 bp were discarded. Reads were then mapped to a custom, curated database of repetitive elements and rRNA sequences. All non-repeat mapped reads were mapped to the genome mm38 using STAR (v2.7.7a); during the alignment multimapping, reads were not retained. PCR duplicates were removed using UMI-tools (v1.1.1). Peak calling was conducted with CLIPper 87. For each peak, IP versus input fold enrichment values were calculated as a ratio of counts of reads overlapping the peak region in the IP and the input samples, removing peaks from IgG control (IDR). Statistical significance was assessed using Yates’ Chi-Square test or Fisher’s Exact Test (if counts <5). For comparisons across sample types, enrichment and p-values were calculated using the same method. Significance was set at IDR log2(fold change) ≥ 3.0 p-adj < 0.001. Peaks were annotated using GENCODE transcript models, with priority given to coding features (CDS, UTRs, introns), followed by noncoding transcripts (exons and introns).

first: Seohyun

last: Park

first: James

last: Emerson

first: Yao Wei

last: Lu

first: Da-Zhi

first: Frank

last: Conlon

first: William

middle: F

last: Marzluff


title: A progeria syndrome links DNA hypermethylation to age-related pathology [mouse]

overall_design: Bisulphite converted DNA from bone marrow, intestine and liver tissue from +/+ and W326R/+ mice at four time ages: 4-day, 23-day, 3-month and 1-year, were hybridised to the Illumina Infiniun Mouse Methylation BeadChip following manufacture's protocols.


title: Thrombospondin-1:CD47 signaling contributes to the development of T cell exhaustion in cancer [scRNA_CD172KO]

city: NEW YORK

first: LEVI

last: MANGARIN

state: NY

address: 413 E 69TH STREET

department: MEYER CANCER CENTER

institute: WEILL CORNELL MEDICINE

summary: T cell exhaustion, a hyporesponsive state of antigen-specific CD8+ T cells exhibiting elevated expression of multiple inhibitory surface receptors, is a major obstacle for the treatment of cancer. Current immune checkpoint blockade (ICB) therapies aimed at reinvigorating exhausted CD8+ T cells have demonstrated clinical effectiveness. However, not all cancer patients achieve long-term disease control due, at least in part, to the refractory nature of terminally exhausted CD8+ T cells to be functionally reinvigorated. Besides persistent antigen stimulation, the environmental sources and mechanisms that lead to CD8+ T cell exhaustion in cancer remain to be thoroughly characterized. Here, we show that the expression of CD47 [a.k.a. integrin-associated protein (IAP) and ‘‘don’t eat me’’ signal is upregulated in tumour-associated, exhausted CD8+ T cell compartments in both human and murine tumours. Our findings reveal a novel role of the extracellular matrix protein thrombospondin-1 (TSP-1) and CD47 in promoting T cell exhaustion. Engagement of TSP-1 with CD47 drives the upregulation of TOX and inhibitory immune checkpoint molecules and compromises the effector function of CD8+ T cells during tumour progression. Mechanistically, the interaction of TSP-1 with CD47 on CD+ T cells activates the calcineurin-NFAT signaling, thereby promoting TOX expression and the T cell exhaustion program in cancer.

overall_design: For the analysis of scRNA-seq of tumor-associated T cells of interests, we performed adoptive transfer of naïve-sorted CD8+ T cells from Pmel-1 CD47+/+ and Pmel-1 CD47+/- mice and combined them in 50% to 50% for adoptive transfer into B16 melanoma implanted mice. After 14 days of transfer, tumors from each mouse were dissected to get all the tumor-associated T cells for sorting. We then sorted live, CD8+ T cells that were transferred to perform scRNA-seq analysis separately.

first: Chien-Huan

last: Weng

first: Levi

last: Mangarin


title: Progeria syndrome links DNA hypermethylation to age-related pathology

type: Methylation profiling by genome tiling array

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

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Refer to individual Series


title: Dynamic expression of CD9 protein in T-cell acute lymphoblastic leukemia [bulk RNA-seq]

city: Marseille

first: Dominique

last: Payet Bornet

address: 27bd Lei Roure

country: France

institute: CRCM

summary: T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematologic malignancy affecting both children and adults. Given its persistently poor prognosis, there is a critical need to identify additional factors involved in T-ALL oncogenesis and progression. CD9, a membrane protein of the tetraspanin family implicated in diverse cellular processes, has been associated with prognosis in several cancers, yet its role in T-ALL remains poorly understood. In this study, using first a mouse model, we found that CD9 overexpression is associated with leukemic T cells that have migrated outside the thymus into peripheral tissues. Then, analysis of a human T-ALL cohort shows that CD9 expression is heterogeneous, tends to increase at relapse and is enriched in the TAL1⁺ molecular subtype. We further demonstrate that CD9⁺ cells display enhanced migratory capacity compared with CD9⁻ counterparts, and that CD9 levels affect extracellular vesicle biogenesis. Altogether our findings support a role for CD9 in T-ALL leukemogenesis and highlight its potential involvement in relapse.

overall_design:
RNA-seq profiling of Patient-Derived Xenograft of primary human T-ALL samples in immunodeficient NSG mice and Jurkat cells.
Submitters state "Raw sequencing data correspond to Patient-Derived Xenografts (PDX) and are therefore subject to privacy restrictions (PDX* samples)."

***************************************************************
Raw files for human/patient samples are being made available in EGA (https://www.ebi.ac.uk/ega/) for controlled access to the personally identifiable sequence data.
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first: Julie

last: Quessada

first: Mathis

last: Nozais

first: Charlotte

last: Savey

first: Julien

last: Rey

first: Delphine

last: Potier

first: Marie

last: Loosveld


title: A progeria syndrome links DNA hypermethylation to age-related pathology [human]

first: Dan

last: Sarni

overall_design: Bisulphite converted DNA from whole blood samples from 7 HESJAS patients and 9 healthy controls were hybridised to the Illumina Infiniun MethylationEPIC v.1.0 BeadChip following manufacture's protocols. SNPs removed from idat and processed files to anonymise patients and healthy controls.

first: Duncan

last: Sproul

first: Andrew

last: Jackson


title: DDX3X-Mediated Translation of Structured Cardiac mRNAs is Essential for Female Heart Development [Ribo-Seq]

first: Kayla

last: Mason

overall_design: E10.5 ribosome profiling and RNA-sequencing were performed by EclipseBio through their proprietary method named eRibo Pro. E10.5 hearts were dissected on ice and individually snap frozen, and genotyping was performed on carcasses against Sry, Ddx3x, and Tnnt2-Cre. Experiments were performed at n=3 replicates/genotype (Ddx3xfl/fl and Ddx3xCM-CM-) with 20 hearts/replicate. Individually snap frozen hearts were pooled in Eclipsebio proprietary eRibo Pro lysis buffer, and subsequently thawed on ice, lysed, and 0.75µg of total RNA was isolated, and heat fragmented for RNA-Seq. 7.1-9µg of total RNA per sample was digested with RNase I to isolate ribosome protected RNA footprints (RPFs). Both sample types were taken through rRNA depletion and adapter ligation. Ligated RNA was reverse transcribed, libraries were generated via PCR and then sequenced on a NextSeq 2000.  After sequencing, samples were processed with Eclipsebio's proprietary analysis pipeline (v1), as was conducted in eCLIP-seq for all RNA sequencing, by using mm39. Additionally, gene coverage was calculated using Eclipsebio's proprietary feature counting algorithm and ribosome occupancy was calculated by dividing coverage in the ribosome protected footprint (RPF) library by the RNA-Seq library. All genes with ribosome occupancy with log2(fold change) ≥ ±1.0 and adjusted p (p-adj) < 0.05 were determined to be significantly different. The RNA-seq library was analyzed separately for differentially expressed genes using DESeq2. The Wald test was used to performed, and genes with adjusted p-value < 0.05 and log2(fold change) ≥ ±0.5  were identified as differentially expressed genes.

first: Seohyun

last: Park

first: James

last: Emerson

first: Yao Wei

last: Wang


title: Transcriptomic Profiling of the Sex-linked Biological Pathways of Severe Pulmonary Arterial Hypertension associated with Endothelial Cell Caveolin-1 Depletion and Chronic Hypoxia

city: Chicago

first: Richard

middle: D.

last: Minshall

state: IL

address: 835 S Wolcott Ave - E709C MSB (M/C 868)

department: Anesthesiology & Pharmacology and Regenerative Medicine

institute: University of Illinois Chicago

summary: Pulmonary arterial hypertension (PAH) is distinguished by elevated blood pressure and vascular resistance in the arteries of the lungs.  Patients with PAH demonstrate pulmonary vascular remodeling, wall thickening, and a high rate of morbidity due to right heart failure.  Notably, while female patients are more likely to develop PAH, male patients suffer from higher morbidity rates after diagnosis.  The molecular mechanism(s) underlying PAH development is poorly understood, though heritable PAH linked to mutations in bone morphogenic protein receptor 2 (BMPR2) and caveolin-1 (CAV1) may provide novel insights into the disease’s pathophysiology.  To interrogate this dynamic, we utilized a global CAV1 knockout (CAV1-KO) mouse model (CAV1-/-) in conjunction with chronic hypoxia to induce symptoms of PAH as demonstrated by hemodynamic and ECHO cardiography recordings.  Both female and male CAV1-/- mice in chronic hypoxia demonstrated elevated right ventricular systolic pressure (RVSP) of 48.49mmHg and 47.78mmHg respectively.  Female knockout mice started to die earlier in hypoxic conditions (4 wks), though male mice showed greater total mortality by the end of the 8 wks of hypoxia.  In addition to wildtype controls, we compared this knockout mouse to endothelial-specific CAV1 reconstituted (CAV1-RC) knockouts and found that restoration of CAV1 expression only in endothelial cells (ECs) is sufficient to ameliorate PAH symptoms, highlighting the importance of vascular CAV1 in maintaining pulmonary artery function.  RNA-sequencing of the lungs revealed that CAV1-/- is associated with downregulation of biological process gene pathways involved in cilium assembly in normoxic conditions for both sexes.  In hypoxic conditions, CAV1 knockout in females leads to downregulation of BMP signaling, while male hypoxic CAV1-/- led to a significant increase in muscle cell development genes.  Reconstitution of CAV1 in ECs lead to upregulation of immune signaling pathways, muscle cell development, and various cell differentiation pathways in both sexes.  Females showed a unique upregulation of cilia-related pathways, while males demonstrated increased BMP signaling.  These data indicate that muscle cell development, angiogenesis, cilia assembly, immune response, and BMP signaling pathways undergo sex-specific transcriptional regulation during PH development that may underlie sex differences in PH patient outcome.

overall_design: This experiment contains bulk RNA-sequencing data from mouse lungs exposed to 6 weeks of normal oxygen levels (Normoxia, Nx) or chronic 10% oxygen (Hypoxia, Hx).  Males (M) and Females (F) were treated as separate groups to investigate sex differences. Genotypes studied were wildtype control (B6/129SJ2), global Caveolin-1 knockout (Cav1(tm1Mls/)J), and a Cav1-knockout model with Cav1 specifically reconstituted and expressed in endothelial cells (Cav1-RC) obtained from William Sessa at Yale University.

first: Joseph

middle: W

last: Leasure

first: Samuel

middle: M

last: Lee

last: Yerlioglu

first: Maricela

last: Castellon

first: Haibin

last: Li

first: Andres

last: Fantuzzi

first: Sunny

last: Chen

first: Sami

middle: B

last: Muslmani

first: Arnav

last: Sharma

first: Jiwang

middle: D


title: Rpd3L regulates Transcription-Replication Conflict via H3K4 Methylation-Dependent and Independent Chromatin Mechanisms

city: Yilan City

first: Shin Yen

last: Chong

state: Yilan County

address: No.1, Shennong Rd.,

department: Department of Food Science

country: Taiwan

institute: National Ilan University

summary: Faithful genome duplication requires coordination between transcription and replication. Disruption of this coordination causes transcription-replication conflicts (TRCs), leading to replication stress and genome instability. How chromatin regulators modulate these processes remains unclear. Here, we show that the Rpd3L histone deacetylase complex dynamically modulates chromatin state to control replication fork progression and buffer TRCs in Saccharomyces cerevisiae. Rpd3L is targeted through both histone H3 lysine 4 methylation-dependent recruitment and methylation-independent mechanisms engaged under replication stress. Loss of H3K4 methylation or Rpd3L function promotes histone acetylation, accelerates fork progression through transcribed regions, and increases transcription-associated genome instability. Balanced acetylation at multiple histone lysines is required to stabilize replication forks under stress. While histone deacetylase complexes have been implicated in repairing damaged forks, our findings reveal that Rpd3L acts preemptively to modulate chromatin state and replication dynamics during TRCs, defining a chromatin-based mechanism that safeguards genome stability.

overall_design: ChIP H3K9/14, K18Ac and Pho23-Myc in indicated strains under HU induced replication stress

first: Ya-Ling

first: Yueh-Tzu

last: Hsu

first: Chia-Ling

first: Tsai-Ming

last: Lu

first: Yi-Chen

last: Lo

first: Cheng-Fu

last: Kao


title: Tumor-intrinsic expression of signal regulatory protein α (SIRPα) contributes to the suppression of anti-tumor immune responses [RNA_CD172KO]

first: LEVI

last: MANGARIN

summary: Signal regulatory protein α (SIRPα) is a well-characterized inhibitory receptor expressed on myeloid immune cells. We and others have recently discovered that human and mouse melanoma cells express high levels of SIRPα. However, whether and how melanoma-intrinsic SIRPα contributes to tumor progression and anti-tumor immunity remains underexplored. Here, we identify a novel role of tumor-intrinsic SIRPα in suppressing anti-tumor immune recruitment and activation. Genetic deletion of SIRPα in melanoma cells enhanced tumor control and increased infiltration of immune cells into the tumor. Transcriptomic analysis revealed that loss of melanoma cell-intrinsic SIRPα leads to upregulation of C-X-C motif chemokine 10 (CXCL10) expression in both human and mouse melanoma cells. Notably, knockdown of Cxcl10 in SIRPα-deficient melanoma cells partially rescued tumor growth and reduced CD8+ T cell infiltration, recapitulating the phenotype observed in SIRPα-expressing tumors. These findings indicate that tumor cell-intrinsic SIRPα suppresses anti-tumor immunity by inhibiting Cxcl10 expression, thereby compromising T cell recruitment to the tumor sites. Thus, our work uncovers a previously unrecognized mechanism by which melanoma cell-intrinsic SIRPα suppresses anti-tumor immunity and highlights the therapeutic potential of silencing SIRPα to enhance anti-tumor immune cell infiltration and promote T cell-mediated tumor control. We envision that SIRPα silencing as a therapeutic strategy that could be applied to other cancer types that express SIRPα.

overall_design: We performed bulk RNA-seq analysis using in vitro cultured B16-F10 melanoma cells that were either Sirpa-wild type (WT) or Sirpa-knockout (KO) to assess steady state gene express patterns among Sirpa-WT and KO murine B16 melanoma tumor cell lines. We also performed scRNA-seq analysis of live-sorted CD45+ cells from tumor-draining lymph nodes and tumors from wild-type C57BL6/j mice that were implanted with Sirpa-WT or Sirpa-KO B16 tumors. The experimental design of these in vivo experiments was to understand the impact of tumor-intrinsic Sirpa expression on immune modulation.

first: Chien-Huan

last: Weng

first: Levi

last: Mangarin


title: DDX3X-Mediated Translation of Structured Cardiac mRNAs is Essential for Female Heart Development [RNA-Seq]

first: Kayla

last: Mason

overall_design: Deep RNA seqeuncing. 4 male 4 female adult (4wk) heart samples were taken an deep RNA seqeuncing was performed with greater than 80M reads per sample. Samples were analyzed with cutoff of Log2FC >0.6 and p vaule <0.05.

first: Seohyun

last: Park

first: James

last: Emerson

first: Yao Wei

first: Da-Zhi

last: Wang

first: Frank

last: Conlon

first: William

last: Marzluff


title: EBNA1 inhibitors reveal CDC7 and POU2F1 as direct functional targets in EBV+ epithelial cancers [RNA-Seq]

city: Philadelphia

first: Priyankara

middle: J

last: Wickramasinghe

state: PA

address: 3601 Spruce Street

department: Bioinformatics

institute: The Wistar Institute

summary: Epstein-Barr virus (EBV) latent infection is ubiquitously associated with nasopharyngeal carcinoma and gastric cancer, expressing very few viral protein components (EBNA1 and LMP1/2A). We previously showed that EBNA1 inhibitor VK1727 blocked cell cycle progression and growth of EBV+ tumors in vivo. However, the underlying molecular mechanism requires more evidence. In this study, we employed VK1727 to remove EBNA1 binding to viral and cellular genomes in three EBV+ epithelial tumors (PDX C15, C666-1 and SNU719). Then we integrated EBNA1 ChIP-seq and transcriptomic analysis to identify direct gene targets of VK1727, which involve in cell cycle progression and cell proliferation. Our analysis characterized a cell cycle dependent kinase CDC7 and a stem cell transcription factor POU2F1. Further analysis illustrated that EBNA1 binding to CDC7 promoter and POU2F1 intron promotes RNA Pol II-pS5 to initiate transcription of these two genes. Moreover, our results revealed CDC7 inhibitor Simurosertib is epistatic with VK1727 in reduction of EBV copy number, induction of cell cycle arrest, and inhibition of cell proliferation in EBV+ epithelial cancer. While POU2F1 knockdown reduced copy number of EBV, decreased expression of viral genes and inhibited proliferation of SNU719 cells. Our study revealed two functional gene targets of VK1727 in EBV+ epithelial cancers, which provides two potential biomarkers for following therapeutical research.

overall_design: Three EBV+ tumor models were treated with EBNA1 inhibitor VK1727 on 3-5 days. Then we integrated RNA-seq and EBNA1 ChIP-seq to identify direct targets of EBNA1 inhibitors in three EBV+ tumors.

first: Songtao

last: HE

first: Niseno

last: Terhuja

first: Leena

last: Yoon

first: Bhanu Chandra

last: Karisetty

first: Paul

last: Lieberman


title: Negative control of Candida albicans biofilm formation by combined action of white-opaque regulator Wor2 and biofilm regulator Bcr1

city: Athens

first: Katharina

last: Goerlich

state: GA

address: 120E Green St

department: Microbiology

institute: University of Georgia

summary: Biofilm formation is vital for survival and pathogenicity of the fungus Candida albicans.  Expression of biofilm-promoting genes is coordinated by a transcription factor network that governs the yeast-filament transition and other processes.  A second cell type transition, the white-opaque transition, that is coordinated by its own transcription factor network.  Initial studies suggested that the two transcriptional networks have a mutually exclusive relationship, driven in part by reciprocal repression of biofilm regulator Efg1 and white/opaque regulator Wor1.  However, recent studies have shown that biofilm regulators and white/opaque regulators can promote one another’s function in many situations.  Here we test the function of white/opaque regulator Wor2 in biofilm formation.  We find that Wor2 has a functional relationship with biofilm regulator Bcr1.  We characterized the phenotype of bcr1Δ/Δ wor2Δ/Δ mutants in five strain backgrounds, and conducted RNA-sequencing (RNA-seq) analysis in the SC5314 reference strain background.  The combined Bcr1-Wor2 impact is unexpected: although Bcr1 is known as a positive regulator of biofilm formation and biofilm-related genes, the bcr1Δ/Δ wor2Δ/Δ mutants have increased biofilm or filamentation capacity, depending on the strain, and increased expression of biofilm-related genes.  Those properties suggest that Wor2 and Bcr1 function together as negative regulators of biofilm formation.  Our findings argue that Bcr1 can act as both a positive and negative regulator of downstream effector genes in the biofilm network and establish a new connection between the biofilm and white-opaque regulatory networks.

overall_design: RNA-seq profiling of WT (SC5314 A22) C. albicans cells and their mutant derivatives (wor2△/△, bcr1△/△, and wor2△/△bcr1△/△). Cells were grown for 4h at 37C in YPD.

first: Aaron

last: Mitchell


title: EBNA1 inhibitors reveal CDC7 and POU2F1 as direct functional targets in EBV+ epithelial cancers


title: Perfluorooctane sulfonate (PFOS) in follicular fluid and human granulosa cell dysfunction: a physiologically based toxicokinetic model translation of low-level, long-term in vitro exposure data

city: Novi Sad

first: Tamara

middle: Miroslav

last: Tomanic

address: Trg Dositeja Obradovica 2

country: Serbia

institute: Faculty of Sciences University of Novi Sad

summary:
STUDY QUESTION
Are bioactive human-equivalent doses (HEDs) of perfluorooctane sulfonate (PFOS), derived from long-term low-level in vitro exposure of human granulosa cells comparable to HEDs inferred from follicular fluid PFOS concentrations in women undergoing ART and in occupationally exposed women?

SUMMARY ANSWER
The bioactive HEDs overlapped with and, in some cases, were lower than the median HEDs inferred from follicular fluid PFOS concentrations.

WHAT IS KNOWN ALREADY
PFOS exposure is a growing public health concern, with evidence suggesting adverse female reproductive effects. However, the relevance of current human exposure levels to granulosa cell function remains unclear.

STUDY DESIGN, SIZE, DURATION
Four independent vials of human granulosa cells (HGrC1 cells) were thawed and expanded into separate flasks (biological replicates). Cells were allocated to four experimental groups and exposed to PFOS (0.01, 0.1, or 1 µM) or vehicle control (0.05% DMSO) for up to 12 weeks, with re-dosing at each passage. Different apical endpoints, along with transcriptomic changes, were evaluated at designated time points. Clinical relevance of PFOS risk to human granulosa cells was assessed by integrating experimental data with physiologically based toxicokinetic (PBTK) modeling.

PARTICIPANTS/MATERIALS, SETTING, METHODS
Viability of HGrC1 cells was assessed using the Alamar Blue assay. Estradiol and progesterone secretion were quantified by enzyme-linked immunosorbent assay. Flow cytometry was used to determine the proportions of live, apoptotic and necrotic cells, as well as cell cycle distribution. Global mRNA expression was assessed by DNA nanoball sequencing technology (DNBSEQ), whereas pathway-level molecular functions were derived using bioinformatic tools. Benchmark concentrations (BMCs) were calculated from key endpoints with concentration-dependent responses and used to estimate HEDs via PBTK modeling. These HEDs were compared with HEDs inferred from follicular fluid PFOS levels reported in the literature to derive bioactivity exposure ratios (BERs) and assess relevance to human exposure.

MAIN RESULTS AND THE ROLE OF CHANCE
In HGrC1 cells, long-term PFOS exposure altered steroidogenesis, apoptosis/necrosis, cell cycle distribution (P < 0.05), and gene expression (at least 2-fold change, Q-value ≤ 0.05). Median transcriptomic HEDs were 18.1 (95% CI: 1.1–35.1) and 17.5 ng/kg bw/day (95% CI: 8–27.1) for 6- and 12-week exposures, respectively, with corresponding 5th percentile HEDs of 3.7 ng/kg bw/day (95% CI: 0.4–9.3) and 1.4 ng/kg bw/day (95% CI: 0.5–3.5). Pathway-level HEDs ranged from 2.8 to 24.1 ng/kg bw/day, with eicosanoid synthesis showing the greatest sensitivity. HEDs for apical endpoints ranged from 0.4 to 203 ng/kg bw/day, with the sub-G1 cell cycle phase being most sensitive. HEDs derived from the 5th percentile transcriptomic data, eicosanoid metabolism, and the sub-G1 phase yielded BERs below 1, indicating that PFOS levels measured in follicular fluid of ART patients may be sufficient to induce these biological effects. For occupational exposure, BERs derived from all endpoints were below 1. A subset of nine granulosa-cell genes, including CYP1B1 and TIPARP (aryl hydrocarbon receptor signaling), showed HEDs that were below the follicular-fluid-inferred HED, highlighting potential high-priority targets and candidate biomarkers.

LARGE SCALE DATA
Raw and processed RNA-sequencing data are deposited in NCBI Gene Expression Omnibus (GEO) under accession number GSE315651.

LIMITATIONS, REASONS FOR CAUTION
The estimated exposure values were based on predictions from a PBTK model rather than empirical human exposure data. Also, differences in protein concentrations in vitro and in vivo may affect free PFOS levels and bioactivity estimates. We addressed this with additional adjustments for PFOS-albumin binding. Finally, follicular fluid PFOS concentrations in occupational settings were approximated from serum concentrations using blood-to-follicular fluid transfer efficiency (BFTE) values.

WIDER IMPLICATIONS OF THE FINDINGS
Our findings suggest that PFOS concentrations in follicular fluid from women undergoing ART and those who have been occupationally exposed may be sufficient to perturb granulosa cell mRNA expression and key pathways, including eicosanoid, interleukin, and GPCR signaling. The identified genes may serve as candidate biomarkers linking PFOS exposure to clinical outcomes in ART settings. Overall, this study provides a framework for interpreting PFOS reproductive toxicity and refining health-protective exposure thresholds.

STUDY FUNDING/COMPETING INTEREST(S)
This research was supported by the Ministry of Science, Technological Development and Innovation of the Republic of Serbia (Faculty of Sciences, Novi Sad: Grants No. 451-03-137/2025-03/200125 & 451-03-136/2025-03/200125), the Institute of Physics, Belgrade, National Institute of the Republic of Serbia, and the Science Fund of the Republic of Serbia, Grant No. 7010, ‘Integration of Biological Responses and PBTK Modeling in Chemical Toxicity Assessment: А Case Study of Perfluorooctanoic Acid (PFOA)—ToxIN’. The authors declare no conflicts of interest.

overall_design: RNA-seq profiling of HGrC1 human granulosa cell line following long-term, repeated exposure (6 or 12 weeks) to PFOS at 0.01, 0.1, or 1 µM, or to vehicle control (0.05% DMSO). Four independent biological replicates were generated per exposure group from separate cryopreserved vials.

first: Dragana

last: Samardzija Nenadov

first: Sava

last: Radovic Pletikosic

first: Bojana

last: Stanic

first: Darija

last: Obradovic

first: Sasa

last: Lazovic

first: Nebojsa

last: Andric


title: Rothia mucilaginosa Alleviates Constipation through Gut Microbiota Remodeling and Neuroactive Pathway Activation

city: Tianjin

first: Peishi

last: Jiang

address: No. 190 Jieyuan Road, Hongqiao District

department: Department of Colorectal Surgery

institute: Tianjin Union Medical Center, Nankai University

summary: Constipation affects approximately 15% of the global population, and gut microbiota dysbiosis is implicated in its pathogenesis. Rothia mucilaginosa, a commensal bacterium with established anti-inflammatory properties, has not been previously investigated for its effects on intestinal function. In this study, we evaluated the therapeutic potential of R. mucilaginosa in a loperamide-induced constipation mouse model using multiomics approaches. Twenty-six SPF male C57BL/6 mice were divided into normal control (NC, n=8), constipation model control (MC, n=8), and R. mucilaginosa-treated (RG, n=10) groups. R. mucilaginosa intervention significantly improved fecal output and induced gut microbiota remodeling, including enrichment of Akkermansia muciniphila and Alistipes finegoldii. To characterize host molecular responses, RNA-seq was performed on colon tissues to identify differentially expressed genes and pathways associated with constipation alleviation, with particular focus on neuroactive pathway activation.

overall_design: Twenty-six SPF male C57BL/6 mice (6 weeks old, 20–22 g) were divided into three groups: normal control (NC, n=8, samples NC1–NC8), loperamide-induced constipation model control (MC, n=8, samples MC9–MC16), and Rothia mucilaginosa gavage-treated constipation model (RG, n=10, samples RG17–RG26). The experiment had three phases: adaptation (days 0–7), modeling (days 8–21, loperamide hydrochloride 10 mg/kg then 15 mg/kg), and intervention (days 22–40, continued loperamide 15 mg/kg; RG received daily oral gavage of R. mucilaginosa BNCC363031; NC and MC received PBS). Colon tissues were collected for RNA-seq (Illumina NovaSeq 6000, paired-end 150 bp).

first: Xiaoqing

last: Yang

first: Chenghao

last: Zhang

first: Zhiyue

last: Guo

first: Yuwei

last: Li

first: Chen

last: Xu


title: Age- and Tissue-dependent Diversity of Human Plasmacytoid Dendritic Cells Uncovers a Cycling Subset Dominant in Early Life and Cancer

city: Saint Louis

first: Shitong

last: Wu

state: MO

address: 660 S Euclid Avenue

institute: Washington University in St. Louis

summary: Plasmacytoid dendritic cells (pDCs) are innate sentinels that produce type I interferons (IFN-I) during infection. Here, we asked how developmental stage and tissue context shape human pDC transcriptional states. Single-cell RNA sequencing of pDCs from blood, thymus, lymph nodes, and tonsils across fetal, infant, and pediatric stages revealed tissue-enriched programs, including IFN-I-imprinted pDCs in the thymus, NFκB-imprinted pDCs in tonsils, and resting pDCs in blood and lymph nodes. Across tissues, we identified a pDC subset characterized by prostaglandin D2 production and dopamine responsiveness, indicating a potential neuromodulatory axis. Cycling pDCs were abundant in fetal and infant lymphoid tissues and declined with age, while remaining enriched in bone marrow (BM) throughout life. Blastic pDC neoplasm associated with enrichment of cycling, mutation-bearing pDCs in the BM, suggesting that this niche serves as a reservoir for malignant pDCs. Thus, early in life, various lymphoid tissues can generate pDCs. This tissue-specific hematopoiesis may result in distinct cellular and functional outputs.

overall_design: Dendritic cells were isolated from the skin, skin-draining lymph nodes (sLN), mesenteric lymph nodes (mLN), spleen, thymus, and lung of three C57BL/6 mice. Tissues were processed into single-cell suspensions and enriched for CD11c⁺ cells. cDC1 (CD19⁻ CD3⁻ Bst2⁻ SiglecH⁻ CD11c⁺ MHC-II⁺ XCR1⁺ CD11b⁻), cDC2 (CD19⁻ CD3⁻ Bst2⁻ SiglecH⁻ CD11c⁺ MHC-II⁺ XCR1⁻ CD11b⁺), and pDC (CD19⁻ CD3⁻ Bst2⁺ SiglecH⁺) subsets were sort-purified from all tissues except the skin (pDCs were not sorted from skin due to their absence at steady state). Prior to sorting, each tissue suspension was labeled with a unique TotalSeq-A Hashtag antibody (HTO). For Sample 1, mouse 1 was labeled with HTO-1 (Skin), HTO-4 (sLN), and HTO-7 (mLN); mouse 2 with HTO-2 (Skin), HTO-5 (sLN), and HTO-8 (mLN); and mouse 3 with HTO-3 (Skin), HTO-6 (sLN), and HTO-9 (mLN). For Sample 2, the same mouse-specific HTO assignments were used, such that spleen samples were labeled with HTO-1, HTO-2, and HTO-3, thymus with HTO-4, HTO-8, and HTO-9, and lung with HTO-5, HTO-6, and HTO-7 across the three mice. Following sort-purification, cells from each tissue were pooled per sample and resuspended in PBS containing 0.04% BSA at approximately 1,000 cells/μL, and subsequently loaded into a 10x Genomics Chromium Single Cell Controller at the McDonnell Genome Institute (Washington University in St. Louis).  Single-cell libraries were prepared using the Chromium Next GEM Single Cell 3′ Kit v3.1, Chromium Next GEM Chip G Single Cell Kit, and Dual Index Kit TT Set A. Approximately 20,000–23,000 cells were partitioned into Gel Bead-in-Emulsions (GEMs), enabling barcoded cDNA synthesis with unique molecular identifiers (UMIs), followed by cDNA amplification (11 cycles) and size-selection using SPRIselect beads. Gene expression (GEX) and Hashtag-oligo (HTO) libraries were generated according to the manufacturer’s protocol (10x Genomics Single Cell 3′ Reagent Kits v3.1, Dual Index). Libraries were quantified using the KAPA Library Quantification Kit (Roche). Sequencing was performed on an Illumina NovaSeq 6000 S4 flow cell using a 28×10×10×150 read configuration. Target sequencing depth was ~50,000 reads per cell for GEX libraries and ~5,000 reads per cell for HTO libraries.

first: Alina

last: Ulezko Antonova

first: Patrick

last: Rodrigues

first: Marina

last: Cella

first: Marco

last: Colonna