title: Temporal transcriptomic profiling of bone autograft healing reveals dynamic immune, vascular, and osteogenic programs [scRNA-seq]

city: Rochester

first: Jared

last: Mereness

state: New York

address: 204 Robert B. Goergen Hall, Department of Biomedical Engineering, Box 270168

department: Biomedical Engineering

institute: University of Rochester

summary: Bone autograft healing is a highly orchestrated process that integrates immune activation, vascular ingrowth, and osteogenic remodeling. To define the molecular and cellular programs driving early autograft integration, bulk and single-cell RNA sequencing was used to analyze graft-associated tissues over 14 days in a murine periosteal-mediated autograft model. Global transcriptomic analysis revealed rapid and dynamic remodeling, with maximal gene expression changes occurring within the first week. The first 48 h were dominated by pro-inflammatory signaling, including TNF, IL-1, TLR, and MAPK pathways, accompanied by transcriptional signatures of phagocytosis and cellular clearance. These early inflammatory programs gave way to pro-regenerative signals, including activation of HIF-1, PI3K-AKT, Wnt, and BMP pathways, coincident with angiogenesis, osteogenesis, and matrix deposition. By day 14, extracellular matrix production and remodeling predominated, marked by metalloproteinase activity and structural matrix gene enrichment. Single-cell RNA sequencing revealed that donor-derived (eGFP+) graft cells were rare and transient, whereas host-derived immune cells were progressively replaced by myofibroblasts, endothelial cells, and neurogenic cell types, including GABAergic neurons and IGSF21+ dendritic cells, suggesting active neurovascular crosstalk during healing. Together, these data define a temporal immune-to-regenerative cascade in bone autograft repair and highlight candidate cellular and molecular targets to enhance graft performance.

overall_design: Single cell RNAseq of mouse periosteum harvested from sham and autografted femurs at days 1, 7, and 14 post-surgery.  Grafts were derived from GFP+ C57BL/6 mice (C57BL/6-Tg(UBC-GFP)30Scha/J, Jackson Lab strain 004353) to differentiate graft cells from host).

first: Sayantani

last: Basu

first: Lauren

last: Benoodt

first: Danielle

last: Benoit


title: A cell-intrinsic glucocorticoid biosynthesis and sensing circuit maintains homeostatic Th17 cell state [ATAC-seq]

status: Public on Jun 12 2026

city: Boston

first: The Gene Lay Institute of

last: Immunology and Inflammation

state: MA

address: 60 Fenwood Road,

country: USA

institute: Brigham and Women's Hospital

type: Genome binding/occupancy profiling by high throughput sequencing

summary: T helper 17 (Th17) cells are heterogeneous; homeostatic Th17 (Th17Hom) cells maintain tissue homeostasis and pro-inflammatory Th17 (Th17Inf) cells drive autoimmune inflammation. IL-23 drives Th17Inf, but the signals that maintain Th17Hom remain unclear. Here, we found that differential glucocorticoid (GC) production downstream of CYP11A1, a critical enzyme for steroidogenesis, distinguished Th17Hom cells from Th17Inf cells. Although TCR signaling opened the Cyp11a1 locus, TGFβ1 and IL-6, key cytokines for Th17Hom cell differentiation, maintained and amplified Cyp11a1. Th17Hom cell-derived GC signaled through the glucocorticoid receptor (GR), which was higher in Th17Hom cells compared to Th17Inf cells, thereby forming a circuit that maintained the homeostatic state. Integration of multi-omics data from CYP11A1- and GR-deficient Th17Hom cells revealed a gene network underlying this circuit. TGFβ1, a key node in the network, restored GC sensing to Th17Inf. We thus identify a GC signaling circuit that maintains Th17 homeostasis with implications for treating Th17-mediated autoimmunity.

overall_design: Naïve CD4+ T cells isolated from GR WT and GR cKO mice were differentiated into Th17Hom and then treated with or without GC (Dex) for two days. Cells were then collected and analyzed by bulk ATAC-seq (n=3 per group).

first: Dandan

last: Yang

first: Linglin

last: Huang

first: Ana

middle: C

last: Anderson


title: Blood-based Molecular Endotyping of Fibrotic Hypersensitivity Pneumonitis Identifies Two Immune Subtypes with Divergent Clinical Outcomes

city: Charlottesville

first: Yong

state: VA

address: 1340 Jefferson Park Ave

department: Medicine

institute: University of Viginia

type: Expression profiling by high throughput sequencing

summary: Fibrotic hypersensitivity pneumonitis (fHP) is an antigen driven deadly interstitial lung disease with limited treatment options. Using multiple analysis algorithms, we identified two molecular endotypes with divergent immune signatures in transcriptome of whole blood from a fHP cohort and validated in transcriptomes from whole blood, peripheral blood mononuclear cells, bronchoalveolar lavage, surgical lung biopsy tissue, and in the proteome from blood plasma. The lymphocytic endotype (L-fHP) comprising two-thirds of fHP cases showed enrichment of adaptive immune pathways and lymphocyte predominance, while the non-lymphocytic endotype (N-fHP) exhibited enrichment of innate pathways with neutrophil and macrophage predominance. The N-fHP cases had lower baseline pulmonary function, and reduced transplant-free survival compared to L-fHP cases, yet corticosteroid therapy was associated with poorer transplant-free survival in L-fHP and without impact in N-fHP. These results show that blood-based molecular endotyping can uncover prognostic and therapeutic insights and advance precision medicine otherwise inaccessible to traditional clinical phenotyping.

overall_design: Venous whole blood was collected in PAXgene blood RNA tube (BD Biosciences, cat# 762165). RNA was extracted using PAXgene Blood RNA kit (Qiagen, cat#762164) following manufacturer’s protocol. RNA quality and integrity were confirmed by Quant-iT RiboGreen RNA Assay Kit (ThermoFisher, cat#R11490) and TapeStation RNA ScreenTape (Agilent, Santa Clara, CA) at Psomagen Inc., (https://www.psomagen.com). All RNA samples displayed an RNA Integrity Number (RIN)>7 were proceeded to cDNA library preparation following the manufacturer’s guidelines. Total RNA in the amount of 0.1μg per sample was depleted of ribosomal RNA using the Illumina Stranded Total RNA Prep, Ligation with Ribo-Zero Plus (Illumina, cat# 470-2020-003-A). RNA was fragmented at 94°C for 2 min, followed by the directional (first strand) cDNA generation. Deoxy-UTP was incorporated in second strand synthesis to effectively quench the second strand during PCR amplification. After adenylation of the 3' end and ligation of adapters, fragments were selected and enriched with 13 cycles of PCR amplification. Libraries sizes were checked by Agilent TapeStation 2200 or 4200 with an average size range of 330 to 445 bp. Libraries concentrations were quantified by Picogreen method according to each protocol guidelines to create optimum cluster densities across every lane of every flow cell to achieve the higher quality of data on Illumina sequencing platforms. Clusters were generated using ExAmp chemistry on patterned flow cells and sequenced with the NovaSeq 6000 S4 Reagent Kit v1.5 (300 cycles) on the NovaSeq 6000 system (Illumina, San Diego, CA), following the manufacturer's instructions. Approximately 60 million reads by were generated for each cDNA library by 150bp paired end (PE) sequencing cycle.

first: Shwu-Fan

last: Ma

first: Evan R.

last: Fernandez Perez

first: Justin M.

last: Oldham

first: Fernando J.

last: Martinez

first: Imre

last: Noth


title: Mapping the spatial architecture of glioblastoma from core to edge delineates niche-specific tumor cell states and intercellular interactions [scRNA-seq]

first: Allegra

middle: A

last: Petti

state: Massachusetts

address: 185 Cambridge St.

department: Department of Neurosurgery

institute: Massachusetts General Hospital

summary: Treatment resistance in glioblastoma (GBM) is largely driven by the extensive multi-level heterogeneity that typifies this disease. Despite significant progress toward elucidating GBM’s genomic and transcriptional heterogeneity, a critical knowledge gap remains in defining this heterogeneity at the spatial level. To address this, we employed spatial transcriptomics to map the architecture of the GBM ecosystem. This revealed tumor cell states that are jointly defined by gene expression and spatial localization, and multicellular niches whose composition varies along the tumor core-edge axis. Ligand-receptor interaction analysis uncovered a complex network of intercellular communication, including niche- and region-specific interactions. Finally, we found that CD8 +ve GZMK +ve T cells colocalize with LYVE1 positive CD163 positive myeloid cells in vascular regions, suggesting a potential mechanism for immune evasion. These findings provide novel insights into the GBM tumor microenvironment, highlighting previously unrecognized patterns of spatial organization and intercellular interactions, and novel therapeutic avenues to disrupt tumor-promoting interactions and overcome immune resistance.

overall_design: We used the 10x Genomics 3’ v2 and 5’ v2 Chromium Gene Expression Solution to generate high-quality scRNA-seq data for 11 GBM samples from nine patients, three of whom also contributed Visium data.e could not obtain scRNA-seq samples from every patient analyzed using Visium. Sequencing reads were demultiplexed, aligned, and counted using the Cellranger 5.0 pipeline. Downstream processing steps, including filtering, doublet removal, normalization, scaling, identification of variable genes, dimensionality reduction, clustering, and cell typing were performed using a WDL pipeline we developed for tailored analysis of GBM scRNA-seq data (https://dockstore.org/organizations/PettilabMGH).

first: Saad

middle: M

last: Khan

first: Anthony

middle: Z

last: Wang

first: Rupen

middle: R

last: Desai

first: Colin

last: McCornack

first: Rui

last: Sun

first: Sonika

last: Dahiya

first: Jennifer

last: Foltz

first: Ngima

middle: D

last: Sherpa

first: Lydia

last: Leavitt

first: Timothy

last: West

first: Alexander

middle: F

first: Aleksandar

last: Krbanjevic

first: Bryan

last: Choi

first: Eric

last: Leuthardt

first: Bhuvic

last: Patel

first: Albert

middle: H

last: Kim

first: Gavin

middle: P

last: Dunn


title: Mapping the spatial architecture of glioblastoma from core to edge delineates niche-specific tumor cell states and intercellular interactions [Spatial Transcriptomics]

first: Allegra

last: Petti

overall_design: We used the Visium V1 platform to survey the spatial landscape of 22 samples along the edge-core axis of nine patients with primary isocitrate dehydrogenase (IDH) wildtype (WT) GBM, and the Xenium platform to analyze nine samples from six patients with primary IDHWT GBM. Each sample was preserved in OCT or FFPE, stained with H&E, and analyzed using the Visium V1 spatial transcriptomic platform. We then used two independent approaches to complement the core and edge designations established intraoperatively. First, we incorporated histopathologic tumor core and edge designations made by a board-certified neuropathologist (S.D.). These were consistent with those based on 5-ALA fluorescence, MRI, and surgical evaluation, but indicated that most samples - even those from the tumor edge - contained some regions of high tumor cell density. Within each sample, manual histologic annotation indicated regions of high tumor cell density (consistent with tumor “core”), low tumor cell density (infiltrative tumor “edge”), microvascular proliferation (MVP), palisading necrosis, predominantly non-malignant white matter, and predominantly non-malignant gray matter.

first: Saad


title: Interplay between the RBP SYNCRIP and RNA methylation in determining EV miRNA-cargo and function in cell-to-cell communication

city: Rome

first: Luca

last: Quattrocchi

state: Lazio

address: Viale Regina Elena 324

country: Italy

institute: Sapienza University of Rome

type: Non-coding RNA profiling by high throughput sequencing

summary: Small extracellular vesicles (sEVs) are critical mediators of tumour–microenvironment communication, largely through the selective transfer of microRNAs (miRNAs) that reprogram  recipient cells. Active miRNA sorting into sEVs depends on RNA‑binding proteins, sequence determinants and RNA modifications. Here, a functional interplay between the RBP SYNCRIP and N6‑methyladenosine (m6A) RNA methylation in controlling miRNA loading into hepatocellular carcinoma (HCC)‑derived sEVs has been disclosed. It is reported that i) METTL3‑dependent m6A modification is required for efficient binding of SYNCRIP to specific miRNAs, thereby enabling their selective incorporation into sEVs; ii) silencing of SYNCRIP markedly reshapes the sEV miRNA-cargo and impairs the ability of HCC‑derived sEVs to induce epithelial–mesenchymal transition (EMT) in non‑tumorigenic hepatocytes. Notably, iii) depletion of METTL3 produces an even stronger effect, indicating that m6A methylation represents an upstream and essential determinant of SYNCRIP‑mediated miRNA export. Mechanistically, the data identify SYNCRIP as an m6A‑dependent miRNA reader, adding epitranscriptomic regulation to sequence‑based miRNA sorting into sEVs. Functionally, disruption of this interaction attenuates sEV‑driven EMT and pro‑tumorigenic signalling. Collectively, these findings uncover a novel regulatory axis governing sEV miRNA cargo selection and highlight the m6A–SYNCRIP interplay as a potential therapeutic target to interfere with EV‑mediated tumour progression and metastasis.

overall_design: miRNA-seq profiling of small extracellular vesicles derived from control HepG2 cells (shCTR) and their knockdown derivatives (shSYNCRIP)


title: Mapping the spatial architecture of glioblastoma from core to edge delineates niche-specific tumor cell states and intercellular interactions

overall_design:
We designed a panel of oligonucleotides targeting transcripts from 362 genes, comprising 266 genes from the predesigned 10X Genomics human brain-and-glioblastoma panel and 96 custom genes indicated by our scRNA-seq analyses (Table S4). Together, this customized panel captures diverse neurons, glia, endothelial cells, microglia, immune cells, GBM cells (including both spatial GBM cell states derived from our Visium data and published scRNA-seq-derived GBM cell states), and ligand-receptor pairs that putatively mediate tumor/neuron, tumor/T cell, and tumor/macrophage interactions.
Sample Preparation - Primary Cohort
Formalin-fixed, paraffin-embedded (FFPE) tumor samples were sectioned into 5-um sections using a microtome onto Xenium slides according to Protocol CG000578 published by 10X Genomics. The Xenium slides were then deparaffinized and decrosslinked according to the Protocol CG000580. Both a pre-designed gene expression panel (Human Brain Probe set) and a custom probe set were used during probe hybridization, ligation, and amplification according to Protocol CG000749, which also contained cell segmentation staining. The Xenium slide was then loaded into the Xenium Analyzer using Protocol CG000584 and regions of interest (ROIs) were initially selected using the on-instrument software.

last: Khan

first: Anthony

last: Wang

first: Rupen

first: Colin


title: Piezo1 Modulates Hematopoietic Homeostasis and Chronic Myelomonocytic Leukemia Progression [low-put RNA-seq]

city: Xi'an

first: xiao

last: yu

state: Shaanxi

address: Innovation Harbor Campus, Xi'an Jiaotong University, Chang'an District

country: China

institute: Xi'an Jiaotong University

summary: We demonstrated that PIEZO1 is essential for sustaining hematopoietic homeostasis. To elucidate the underlying mechanism of Piezo1 in normal hematopoiesis, we analyzed and compared the gene expression profiles of hematopoietic stem cells (HSCs) — which were isolated via flow cytometry  from Piezo1 floxed/floxed; Vav-iCre mice and control mice. Our findings revealed that, among the differentially expressed genes (DEGs) in Piezo1 conditional knockout (cKO) HSCs, the cell adhesion pathway was significantly enriched and exhibited downregulation. This downregulation of the cell adhesion pathway was associated with reduced expression of integrin beta1.

overall_design: We performed transcriptional profiling using low-input RNA-seq(Smart-seq2) in sorted HSC from the bone marrow of control and Piezo1 cKO mice.

first: Guangyao

last: Kong

first: Yi

last: Yang

first: Xiao

last: Yu


title: Differential adenosine to inosine RNA editing of SINE B2 non-coding RNAs in mouse unveils a novel type of epi-transcriptome response to amyloid beta neuro-toxicity

city: Winnipeg

first: Athanasios

last: Zovoilis

state: MB

address: 745 Bannatyne Ave

department: Biochemistry and Medical Genetics

country: Canada

institute: University of Manitoba

summary: Alzheimer's disease (AD) is characterized by early molecular responses to amyloid beta neuro-toxicity that remain poorly defined. RNA editing by adenosine-to-inosine (A-to-I) conversion is a major epitranscriptomic mechanism, yet its contribution to non-coding RNA regulation during neurodegeneration is largely unknown. Short Interspersed Nuclear Element (SINE)-derived RNAs, including mouse B2 RNAs, represent the dominant substrates of A-to-I editing and have recently been shown to regulate gene expression through ribozyme-mediated processing. Here, we introduce and validate a repeat-aware bioinformatics framework that enables position-specific quantification of A-to-I editing within highly repetitive SINE RNAs - an analysis that has been challenging using standard genome-based pipelines. Applying this approach, we uncover discrete editing hotspots in B2 RNAs whose modification is selectively increased during the earliest stages of amyloid beta pathology. Elevated B2 RNA editing is consistently observed  in hippocampal neurons exposed to acute amyloid beta toxicity. Functional perturbation of ADAR activity alters both B2 RNA editing levels and expression of B2 RNA-regulated stress response genes, directly linking RNA editing to SINE-mediated transcriptional control. Independent validation using Nanopore direct RNA sequencing supports increased RNA modification signals at the same B2 RNA regions identified by short-read Illumina sequencing. Together, our findings establish a previously unrecognized epitranscriptomic response to amyloid beta neurotoxicity mediated by site-specific A-to-I editing of SINE RNAs. This work defines a new analytical paradigm for studying RNA editing in RNAs from repetitive elements and reveals a regulatory axis connecting amyloid beta toxicity, ADAR activity, SINE RNA editing, and stress-responsive gene expression, with implications for conserved mechanisms underlying Alzheimer's disease and related neurodegenerative disorders.

overall_design: HT22 mouse hippocampal cells were subjected to treatment with i) siRNAs against Adar1 and Adar 2, and ii) a scramble (control siRNA). Subsequently RNA was extracted and subjected to Illumina directional RNA sequencing and  Nanopore direct RNA-sequencing (RNA-seq).

first: Liam

last: Mitchell

first: Luke

last: Saville


title: Piezo1 Modulates Hematopoietic Homeostasis and Chronic Myelomonocytic Leukemia Progression

summary: We identified that PIEZO1 is crucial for maintaining hematopoietic homeostasis and is highly expressed in patients with chronic myelomonocytic leukemia (CMML), correlating with disease progression. To elucidate the underlying mechanism, we analyzed the gene expression profiles of c-Kit⁺ cells isolated from Nrasᴳ¹²ᴰ/ᴳ¹²ᴰ; Mx1-Cre CMML mice, CMML mice with conditional Piezo1 knockout, and control mice for comparative analysis. Our findings revealed that the cell adhesion pathway was one of the significantly enriched terms among the differentially expressed genes in Piezo1 conditional knockout (cKO) c-Kit⁺ cells, which was associated with altered expression of integrins.

overall_design: We performed transcriptional profiling using RNA-seq in sorted c-Kit+ from the bone marrow of control,CMML mice and CMML Piezo1 cKO mice.


title: A longitudinal single-nucleus transcriptomic atlas of bovine placentation reveals dynamic cellular hierarchies and regulatory programs

city: Xianyang City

first: Guanghui

last: Tan

address: No.22, Xinong Road, Yangling Demonstration Zone, Yangling District, Xianyang City

institute: Northwest A&F University

summary: Supporting healthy pregnancy outcomes requires a comprehensive understanding of the cellular hierarchy and underlying molecular mechanisms in the ruminant placenta during gestation. Here, we constructed a single-nucleus transcriptomic atlas of 304,726 placental nuclei spanning 13 developmental stages in cows, identifying 13 major cell types and 14 trophoblast subtypes. Bioinformatics analyses and multiple validation experiments suggested that placental trophoblast cells exhibited stage-specific differences across gestation, and identified HAND1 and DLX5 as novel key regulators of maternal recognition of pregnancy. We found that binucleate (BNC) cells arise from specific uninucleate (UNC) subpopulations at E24, with their differentiation trajectories co-regulated by imprinted genes and metabolic reprogramming. Integrated genome-wide association study (GWAS) analysis identified early trophoblast subtypes significantly associated with gestation length, along with key pathways and risk genes (e.g., CYCS, HMGA1, VDAC1), with these signals under strong evolutionary constraint. Furthermore, we found that placental macrophages originate at E30 in cow and are significantly associated with pregnancy loss in both cattle and humans, sharing common risk pathways. Collectively, this study provides a cross-stage cellular and molecular map of the ruminant placenta, delineates the key regulatory networks at the maternal–fetal interface, and offers a theoretical framework and potential molecular targets for understanding pregnancy maintenance and improving reproductive traits in ruminants.

overall_design: Seventeen bovine placental samples were collected at six gestation stages (E60, E85, E110, E180, E240, E280) with 2–3 replicates per stage. Nuclei were isolated and processed with 10x Genomics Chromium snRNA-seq, followed by sequencing on Illumina NovaSeq 6000.


title: Assessment of the role of genetic variation in microRNAs on the susceptibility and physiopathology of Gastric Cancer

city: Punta Arenas

first: Yolanda

last: Espinosa-Parrilla

state: Magallanes

address: Manuel Bulnes 01855

country: Chile

institute: Universidad de Magallanes

summary: Gastric Cancer (GC) continues to be a disease with a poor prognosis being a leading cause of cancer death worldwide. Varying incidence among regions depends on several factors such as Helicobacter pylori infection, environmental exposure and genetic background. GC is the result of a multifactorial complex process for which a multistep model of carcinogenesis is currently accepted. The clinical outcome of GC is highly dependent on the local progression and metastasis development, which is partially controlled by oncogenes and tumor suppressor genes, whose expression is further regulated by microRNAs (miRNAs). MiRNAs are non-protein-coding small RNAs that act as post-transcriptional gene repressors participating in the regulation of most biological processes. Several miRNAs have been found differentially expressed between tumor and normal tissues in GC, also circulating miRNAs in serum/plasma have been proposed as a novel class of non-invasive biomarkers with a great potential for early detection and prognostic of cancer. We propose that the analysis of the differential expression of miRNAs in the serum of controls and patients at different disease stages of GC might lead to the discovery of new GC causative factors as well as new non-invasive diagnostic biomarkers for GC.

overall_design: Total RNA was extracted from serum using the Plasma/Serum RNA Purification Mini Kit following the manufacturer’s protocol. Small RNA NGS was performed on an Illumina NextSeq 500 instrument at Norgen Biotek (Canada) to evaluate circulating miRNAs in the serum of controls and GC patients at different disease stages of GC

first: Daniela

last: Zapata-Contreras

first: Claudio

last: Urrea

first: Fernando

last: Orellana

first: Maria José

last: Iriarte

first: Carlos

last: Delgado

first: Marcela

last: Puente

first: Lissette

last: Leiva

first: Luis

last: Godoy

first: Alejandro

last: Altamirano

first: Stanko

last: Karelovic


title: CDK8 Inhibition Releases the Muscle Differentiation Blockade in Fusion-driven Alveolar Rhabdomyosarcoma [ATAC-seq]

first: Susu

last: Zhang

address: 360 Longwood Ave, LC6102

department: Pediatric Oncology

institute: Dana-Farber Cancer Institute

summary: Alveolar rhabdomyosarcoma (aRMS) is a fusion-driven pediatric cancer with poor survival and limited therapeutic options. To uncover novel vulnerabilities, we employed complex-based analysis of the functional genomic dataset (Depmap), identifying CDK8 as a novel dependency in aRMS. Both CDK8 knockout or pharmacologic inhibition impaired tumor cell growth and induced myogenic differentiation in vitro and in vivo. Compared to genetic loss, CDK8 inhibition induced more dynamic transcriptional changes, suggesting a distinct gain-of-function mechanism of the CDK8 inhibitor. With a genome-scale CRISPR-Cas9 drug modifier screen, we determined that the maximal anti-tumor activity of the CDK8 inhibitor requires the presence of the Mediator kinase module and transcriptional cooperation with the SAGA complex. We further identified SIX4 as a key transcription factor mediating CDK8 inhibitor-induced transcriptional activation of myogenic differentiation genes and tumor cell proliferation. These findings establish a strong rationale for CDK8 inhibition as a differentiation-inducing therapeutic strategy in aRMS.

overall_design: Rh30 cells treated with DMSO, BI-1347 for 4 hrs, 24 hrs, and 72 hrs were collected for PRO-seq experiment to measure the nascent transcribed RNA.

first: Clare

last: Malone

first: Kenneth

last: Ross

first: Diyana

last: Granum

first: Kristianne

last: Oristian

first: Assil

last: Fahs

first: Corinne M

last: Linardic

first: Kimberly

last: Stegmaier


title: MInD: Multi-organ Invasion Device incorporating a 3D printed granular elastomer reveals cardiac resistance to cancer metastasis

city: Toronto

first: Milica

last: Radisic

state: Ontario

address: 160 College St

institute: University of Toronto

summary: Despite the systemic spread of cancer, the ventricular myocardium remains one of the least common sites of metastasis—a phenomenon that remains poorly understood. To investigate this, we developed the Multi-organ Invasion Device (MInD), a dynamic organ-on-a-chip platform that enables multi-organ culture under flow. Organ compartments are connected in MInD using PermeoTubes—3D-printed conduits with multi-scale fractal porosity fabricated from a granular poly(octamethylene maleate (anhydride) citrate) (POMaC) ink to impart viscoelasticity and ECM-mimetic permeability that supported cancer cell intravasation, migration, and extravasation.  Highly aggressive breast cancer cells preferentially invaded hepatic tissue, while invasion into cardiac tissue was markedly suppressed in both co- and tri-organ culture. Cytokine profiling and RNA sequencing revealed that cardiac co-culture downregulated pro-metastatic and immunosuppressive cytokines and upregulated immune-activating and anti-invasive genes. In contrast, hepatic co-culture promoted matrix remodeling, angiogenesis, and immune suppression. Overall, this platform provides a new approach for uncovering organ-specific drivers of metastasis and cardiac resistance to metastasis, paving the way for future discovery of metastasis-inhibiting therapies.

overall_design: This RNA-seq dataset includes gene expression profiles of breast cancer cells (GFP+ MDA-MB-231) cultured under three different conditions using the AngioPlate system under flow: cancer monoculture, cancer-hepatic co-culture, and cancer-cardiac co-culture. Each condition includes biological replicates (n = 4). Hepatic tissue was created from HepG2 mixed with lung fibroblasts (2%), and cardiac tissue was created from iPSC-derived cardiomyocytes mixed with cardiac fibroblasts (2%) to create cardiac tissues. The cancer cells and all the tissues were embedded in fibrin gels. RNA was extracted from cancer tissues five days after cancer cell seeding.

first: Amid

last: Shakeri

first: Dhana

last: Abdo

first: Matthew Ho Cheong

last: Lei

first: Sargol

last: Okhovatian

first: Richard

last: Jiang

first: Chuan

last: Liu

first: Karl T

last: Wagner

first: Daniel

last: Vosoughi

first: Jennifer

last: Kieda


title: Deletion of Mettl3 in adult cardiomyocytes in vivo.

city: bad nauheim

first: Stefan

last: Günther

address: ludwigtr. 43

country: Germany

institute: MPI for heart and lung research

summary: Deletion of Mettl3 in adult cardiomyocytes reveals the impact of Mettl3 on the regulation of genome expression in postmitotic cardiomyocytes.

overall_design: RNAseq profiling to compare the transcriptome of hearts deficient for Mettl3 in cardiomyocytes to respective control hearts.

first: Theresa

last: Hofmann

first: Thomas

last: Boettger

first: Carsten

last: Künne


title: CDK8 Inhibition Releases the Muscle Differentiation Blockade in Fusion-driven Alveolar Rhabdomyosarcoma [CUT&RUN]

first: Susu

last: Zhang

overall_design: Rh30 cells were treated with DMSO or BI1347 from 24 hr to 72hr.

first: Clare

last: Malone

first: Kenneth

last: Ross

first: Diyana

last: Granum

first: Kristianne

last: Oristian

first: Assil

last: Fahs


title: Deletion of Trdn-as transcription in the adult heart.

summary: Deletion of Trdn-as transcription in the mouse heart reveals a function of trdn-as in the control of TRDN processing as well as secondary changes induced be the alteration of the cardiac calcium release complex.

overall_design: RNAseq profiling to compare the transcriptome of hearts deficient for trdn-as in cardiomyocytes to respective control hearts.


title: Deletion of Trdn-as or Mettl3 transcription in the adult heart

summary: Deletion of trdn-as and Mettl3 transcription in the mouse heart reveals a function of trdn-as in the control of TRDN processing..

overall_design: RNA PolII and CTCF ChIP seq for mouse heart lysates of WT mice, trdn-as deficient mice and mice with cardiomyocyte-specific deletion of Mettl3


title: CDK8 Inhibition Releases the Muscle Differentiation Blockade in Fusion-driven Alveolar Rhabdomyosarcoma

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

overall_design: Refer to individual Series


title: CDK8 Inhibition Releases the Muscle Differentiation Blockade in Fusion-driven Alveolar Rhabdomyosarcoma [RNA-seq]

overall_design: Rh30 cells were collected and RNA was extracted with Trizol.

first: Corinne M

last: Linardic

first: Kimberly

last: Stegmaier


title: CDK8 Inhibition Releases the Muscle Differentiation Blockade in Fusion-driven Alveolar Rhabdomyosarcoma [PRO-seq]


title: Tracing Xanthomonas translucens evolution reveals gene loss for broad host range emergence

city: Columbus

first: Jonathan

last: Jacobs

state: OH

address: 2021 Coffey Rd

department: Plant Pathology

institute: Ohio State University

summary: Epidemic preparedness depends on tracking microbial evolution that drives disease emergence. For outbreaks caused by host jump, we often identify the causal agent, but functional validation of pathogen emergence lags behind. Here, we defined the role of effector gene loss in the emergence of a generalist lineage of Xanthomonas translucens, an important bacterial plant pathogen of cereals and grasses. This lineage, X. translucens pv. undulosa, causes disease in multiple genera of cereals and grasses and encodes significantly fewer virulence effector genes than the specialist sister lineage X. translucens pv. translucens, which infects barley only. Genomic analyses suggested events contributing to host expansion, and we experimentally reproduced effector gene loss evolution by deleting the effectors unique to the specialist lineage. Notably, deletion of the previously uncharacterized effector gene xopAL1 promoted host expansion to wheat but reduced fitness in barley, supporting a fitness tradeoff for niche expansion. Furthermore, transcriptomic analysis revealed specific XopAL1-dependent defense response components of wheat. Three candidate defense response genes were identified and validated for their functional role in defense against X. translucens via individual targeted gene induction using artificial transcription activator-like effectors (arTALEs). Conversely, we identified that the barley specialist lineage Xtt gained the effector gene xopAJ, which limits disease development in oat but may contribute to barley adaptation. This research provides an experimentally validated evolutionary framework for predicting pathogen emergence based on gene loss, and host adaptation via gene gain, and identifies key host defense components for durable disease control.

overall_design: Transcriptional profiles of wheat infected with water, Xanthomonas translucens pv. undulosa, Xanthomonas translucens pv. translucens, Xanthomonas translucens pv. undulosa with genomic insertion of effector xopAL1, and Xanthomonas translucens pv. translucens with xopAL1 deletion with three replicates per treatment.

last: Jacobs (Administrator)

first: Tracy

last: Hawk

first: Marcus

last: Merfa


title: Nuclear Myosin 1 links genomic architecture to adipose tissue remodeling, metabolic inflammation and obesity in mice

city: Abu Dhabi

first: Piergiorgio

last: Percipalle

state: abu dhabi

address: NYUAD Saadiyat Island

country: United Arab Emirates

institute: Biology

summary: Nuclear myosin 1c (NM1) is a chromatin-associated motor protein with known roles in transcriptional regulation and metabolic control. However, its involvement in adipogenesis and adipose tissue remodeling remains unexplored. Here, we demonstrate that NM1 is a critical regulator of adipocyte differentiation and visceral fat homeostasis. Using an NM1 knockout (KO) mouse model, we observed increased adipose tissue mass, adipocyte hypertrophy, and impaired adipogenic differentiation  in mesenchymal stem cell–derived adipocytes ex vivo. Integrative analysis of RNA-seq and ATAC-seq data from wild-type (WT) and NM1 KO mouse embryonic fibroblasts (MEFs) revealed a subset of genes with concordant changes in transcription and chromatin accessibility, many of which are involved in lipid metabolism and adipogenic regulation, including Insig1, Mylip, Lipg, and Fat1. Gene regulatory network modeling using single-cell RNA-seq and TF motif data identified 19 transcription factors (TFs) dysregulated in NM1 KO cells, eight of which (E2f8, Foxo3, Klf6, Npas4, Smad5, Zeb2, Gata4, and Tbx2) also showed significant alterations in chromatin accessibility. Hi-C–based chromatin compartment analysis revealed switching in several adipogenesis-linked genes (Ncoa2, Prdm14, Cops5) in the absence of NM1, suggesting broader effects on 3D genome organization. Together, these findings establish NM1 as a central chromatin-regulatory node that integrates transcription factor activity, enhancer accessibility, and higher-order genome architecture to control adipogenic differentiation. Our results position NM1 and its human paralog MYO1C as potential regulators of obesity and targets for metabolic disease intervention.

overall_design: RNA seq of viseral adipose tissues isolated from abdomen of NM1 WT and KO mice

first: Samira

last: Khalaji

first: Tomas

last: Venit

first: Valentina

last: Fambri

first: Zuzana

last: Lukacova

first: Rahul

last: Shrestha

first: Maylis

last: Boitet

first: Maud

last: Fagny

first: Guiseppe

last: Saldi


title: Gene expression profiling of hindpaw tissues in a spontaneous arthritis mouse model to explore pathological ossification and the effects of non-pharmacological interventions

city: Chihuahua

first: Susana

middle: Aideé

last: González-Chávez

state: Chihuahua

address: Circuito Universitario Campus II

department: Facultad de Medicina

country: Mexico

institute: Universidad Autónoma de Chihuahua

summary:
Spondyloarthritis is a chronic inflammatory disease with limited therapeutic options, particularly for pathological bone formation. The spontaneous arthritis (SpAD) mouse model reproduces key features of human axial spondyloarthritis, including abnormal joint ossification. This dataset captures transcriptional profiles from joint-associated tissues under various non-pharmacological interventions, such as exercise and intermittent fasting, providing insights into mechanisms of inflammation and ectopic ossification.
Gene expression profiling was conducted on distal hindpaw tissues from DBA/1 mice with spontaneous arthritis using GeneChip Clariomª D microarrays. Mice were assigned to groups with or without interventions including treadmill exercise and intermittent fasting. Healthy BALB/c mice were included as controls. Several samples correspond to animals with distinct ossification scores, enabling exploration of gene expression signatures related to pathological ossification.

overall_design: Distal hindpaw tissues were collected from DBA/1 mice with spontaneous arthritis and from healthy BALB/c controls. Diseased mice were assigned to different treatment groups, including treadmill exercise, every-other-day fasting, or no intervention. Histological scoring of ectopic ossification was performed in selected samples. Total RNA was extracted and hybridized on Clariomª D Mouse Arrays. The study design enables transcriptomic comparison across disease states, treatment conditions, and degrees of pathological ossification.

first: Gonz‡lez-Ch‡vez

last: Susana AideŽ

first: Pacheco-Tena

last: CŽsar Francisco

first: Chaparro-Barrera

last: Eduardo


title: Effects of cannabigerol on gene expression of human decidualized endometrial stromal cells

city: Coventry

first: Jan

last: Brosens

address: University Hospital Coventry and Warwickshire

department: Warwick Medical School

country: United Kingdom

institute: University of Warwick

summary: Decidualization denotes the inflammatory reprogramming of endometrial stromal cells (EnSC) into progesterone-dependent decidual cells (DC), a process essential for embryo implantation and placenta formation. Decidualization also leads to the emergence of progesterone-resistant cells, including decidual-like senescent cells (dSC), involved in extracellular matrix (ECM) remodelling, tissue breakdown and menstruation. The use of cannabinoids is increasing due to their therapeutic potential, though the impacts on reproductive health are poorly understood. Cannabigerol (CBG), a non-psychotropic minor phytocannabinoid, affects multiple cellular processes, including proliferation, differentiation and inflammation. In this study, we performed RNA-sequencing and identified differentially expressed genes in CBG-treated versus untreated primary EnSC after eight days of decidualization, mostly associated with mitochondrial activity, lipid biosynthesis, inflammatory response and ECM remodelling. Our findings suggest a modulation by CBG of DC metabolism, ECM restructuring, and inflammatory responses, which may constitute a significant risk factor for adverse pregnancy outcome.

overall_design: RNA-seq was performed on three biological replicates of primary human endometrial stromal cells at day 8 of decidualization, in the absence or presence of cannabigerol (CBG)

first: Mireia

middle: T

last: Nebot

first: Chow Seng

last: Kong

first: Patrícia

last: Alves

middle: J


title: Reduced Representation Bisulfite Sequencing of Mouse Hearts Following Cardiomyocyte-Specific DNMT3A Overexpression

first: Dogacan

last: Yucel

address: 300 Longwood Ave

department: Cardiology

institute: Boston Children's Hospital

type: Methylation profiling by high throughput sequencing

summary: Aging is associated with extensive epigenetic remodeling in cardiomyocytes, including increased DNA methylation. To investigate the causal role of DNA hypermethylation in age-related cardiac dysfunction, we overexpressed DNA methyltransferase 3A (DNMT3A) specifically in cardiomyocytes of young adult mice using a cardiac-specific AAV vector. Reduced representation bisulfite sequencing (RRBS) was performed on isolated cardiomyocyte nuclei from DNMT3A-overexpressing (Dnmt3a OE) and control (GFP) hearts. This dataset provides genome-wide DNA methylation profiles for both groups, enabling analysis of DNMT3A-driven methylome remodeling in the heart.

overall_design: Young adult C57BL/6J mice (aged 3 months) were injected retro-orbitally with AAV9-Tnnt2-Dnmt3a or AAV9-Tnnt2-GFP vectors to overexpress DNMT3A or GFP, respectively, specifically in cardiomyocytes. Three months post-injection, hearts were harvested, and nuclei were isolated and sorted. Cardiomyocyte nuclei were purified by flow cytometry and subjected to reduced representation bisulfite sequencing (RRBS) using the Zymo-Seq RRBS kit. Differential methylation analysis was performed between DNMT3A-overexpressing (Dnmt3a OE) and control (GFP) groups to identify methylation changes induced by DNMT3A overexpression. Each group contains biological replicates.

first: William

last: Pu


title: Single-cell atlas of mandarin fish ovary reveals the trajectories of oogenesis and its immune microenvironment development in teleosts

city: Jingzhou

first: Feng

last: Chen

address: No. 88 Jingmi Road, Jingzhou District

department: College of Animal Science and Technology

institute: Yangtze University

summary: The establishment of the immune microenvironment during oogenesis in teleosts plays an important role in individual survival and population continuation. However, the maternal influence on offspring immunity is poorly understood, particularly regarding the dynamic development and underlying mechanisms of the oocyte immune microenvironment. Here, we characterized the single-cell atlas of the mandarin fish ovary at the late stage IV, and identified three germ cells and five somatic cells. The distinctive features of the three germ cell types were further explored through the enrichment analysis, suggesting that the immune microenvironment of fish oocytes is formed during the mature oocyte stage. Subsequently, based on the pseudotemporal analysis of three germ cells, as well as the clustering analysis and the cell-cell communication analysis, we depicted the trajectories of oogenesis and its immune microenvironment formation in the mandarin fish ovary. Notably, we identified a high-low-high expression pattern in immune-related pathways across different development stages of oocytes. Furthermore, the spatiotemporal expression dynamics of six key markers were validated using RNA-FISH. These findings provide novel insights into oogenesis and the establishment of maternal/innate immunity in teleosts.

overall_design: Three mandarin fish ovaries at the late stage IV were collected for snRNA-seq

first: Yuting

last: Wang

first: Tilin

last: Yi

first: Yingbing

last: Su

first: Xuejun

last: Gao

first: Weiguang


title: Longitudinal single cell RNA-sequencing reveals evolution of micro- and macro-states in chronic myeloid leukemia

city: Duarte

first: Russell

last: Rockne

state: CA

address: 1500 E Duarte Rd

department: Beckman Research Institute

institute: City of Hope

summary: Single cell RNA sequencing (scRNA-seq) has revolutionized our understanding of cancer, yet identifying meaningful disease states from single cell data remains challenging. Here, we systematically explore the cancer specific information content encoded in single cell versus bulk transcriptomics to resolve this paradox and clarify how discrete disease-defining states emerge from inherently noisy single cell data. Using chronic myeloid leukemia (CML) progression as a model, we demonstrate that, while single cell transcriptomes exist along continuous transcriptional microstates, clinically relevant leukemia phenotypes clearly manifest only at the pseudobulk (macrostate) level. By leveraging state-transition theory, we reveal how robust disease phenotype state-transitions are governed by cell type specific contributions. Our results establish a theoretical framework explaining why discrete disease phenotypes remain hidden at the single cell scale but emerge clearly at the aggregated macrostate level, enabling previously inaccessible biological insights into leukemia evolution. Broadly applicable across cancers and other complex diseases, our approach fundamentally advances single cell analysis by clarifying how microscopic transcriptional variation collectively shapes macroscopic disease dynamics.

overall_design: Londitudinal single cell RNA-seq analysis on peripheral blood mononuclear cells (PBMC) samples collected over time from a BCR::ABL inducible mouse model of CP CML

first: David

middle: E

last: Frankhouser

first: Dandan

last: Zhao

first: Yu-Hsuan

last: Fu

first: Anupam

last: Dey

first: Ziang

first: Jihyun

last: Irizarry

first: Jennifer

last: Rangel Ambriz

first: Sergio

last: Branciamore

first: Denis

last: O'Meally

first: Lucy

last: Ghoda

first: Jeffery

last: Trent

first: Stephen

last: Forman

first: Adam

middle: L

last: MacLean

first: Ya-Huei

last: Kuo

first: Bin

last: Zhang

first: Guido

last: Marcucci


title: Taxodione Inhibits Glioblastoma Proliferation and Potentiates the Cytotoxicity of Paclitaxel

city: Tallahassee

first: Cynthia

last: Vied

state: Florida

address: 1115 W. Call St

department: Translational Science Laboratory

institute: Florida State University

summary: Glioblastoma rapidly acquires resistance to conventional genotoxic therapy. This behavior is closely associated with the enhancement of stem-cell-like character during disease progression. Farnesyl diphosphate synthase (FDPS) plays an important role in maintaining such stem-cell-like features. This finding has stimulated interest in FDPS as neuro-oncology drug target; however, the lack of CNS-permeable inhibitors has hampered further development. In this study, we explored the utility of taxodione, a diterpenoid described as an FDPS inhibitor and predicted to penetrate the blood-brain-barrier. The effects of taxodione were compared to its congener 7-(2-oxohexyl)-taxodione and a known FDPS inhibitor in glioblastoma cells. Taxodione was the only treatment that significantly reduced the size of tumor spheroids in a temporal and dose-dependent manner. This activity was linked to FDPS inhibition and transcriptional downregulation of other Mevalonate Pathway genes. Consistent with this putative mechanism-of-action, taxodione sensitized glioblastoma cells to nanomolar concentrations of paclitaxel.

overall_design: RNA-Seq profiling of human glioblastoma U87 cells treated with compounds 1, 4, 5, or vehicle.

first: Kyle

last: Parella

first: Megan

last: Solans

first: Svetlana

middle: N

last: Kononenko

first: Hanna

last: Colegrove

first: Diego

last: Zorio

first: James

last: Frederich


title: A Transient Increase in Cardiomyocyte Protein O-GlcNAcylation Enhances Susceptibility to Pressure Overload-Induced Cardiac Remodeling

city: Birmingham

first: Samuel

last: Chang

state: Alabama

institute: University of Alabama Heersink School of Medicine

summary: The concept of metabolic memory comes from findings that diabetic patients always under tight-glycemic control consistently show better cardiovascular disease outcomes compared to patients who transition to tight-glycemic control after prior conventional glycemic control; mechanisms like epigenetics possibly mediate the lasting metabolic memory effects, however more research into underlying mechanisms is needed. Within the diabetic milieu, increased cardiac protein posttranslational O-linked β-N-acetylglucosamine (O-GlcNAc) modification is implicated in cardiac remodeling, and our previous work shows chronically elevated cardiac O-GlcNAc causes adverse cardiac changes. Therefore, the current study hypothesized that transiently increased cardiomyocyte O-GlcNAcylation leads to exacerbated adverse cardiac remodeling after subsequent pressure-overload. Using our previously described inducible cardiomyocyte specific, dominant-negative O-GlcNAcase (dnOGAh) mouse or single transgenic littermate controls (Con), we induced O-GlcNAc levels for 2wk (ON), followed by a 2wk washout (OFF); mice then underwent transverse-aortic constriction (TAC) or Sham surgery. We observed the expected cardiac remodeling in TAC groups, like decreased cardiac function, and increased hypertrophy and fibrosis. However, we see exacerbation of these pathologic measures in the ON/OFF-TAC vs. Con-TAC mice; additionally, transcriptomic analysis of LV-tissue from each experimental group showed pathways which not only supported phenotypic results of furthered cardiac remodeling, but also, revealed potential novel molecular pathways underlying this pathologic remodeling. We observed exacerbated cardiac pathology between ON/OFF-TAC vs. Con-TAC groups supporting the concept of “O-GlcNAc memory” as a component of metabolic memory. Moreover, transcriptomic analysis provides insight into potential molecular pathways underpinning this metabolic/O-GlcNAc memory such as Ccn2/CTGF driven fibrosis, and/or Nox4 driven oxidative stress.

overall_design: Comparative gene expression analysis of Bulk RNA-seq data from left ventricle tissue of control, or temporary cardiac specific overexpressed dominant negative OGA (dnOGA)  for 2wk induction and then halting of induction for rest of study, with either subseqent transverse aortic constriction or sham surgery 2 weeks after stop of induction period in either control or dnOGA mice.

first: Chae-Myeong

last: Ha

first: Manoja

middle: K

last: Brahma

first: Luke

last: Potter

first: Mahima

middle: S

last: Reddy

first: Sayan

last: Bakshi

first: Kerstin

last: Preuß

first: Md Saimoon

last: Rahman

first: Johannes

last: Fischer

first: Caitlin

last: Harrell

first: Zhihuan

last: Sun

first: John

last: Chatham

last: Wende


title: A cell-intrinsic glucocorticoid biosynthesis and sensing circuit maintains homeostatic Th17 cell state

city: Boston

first: The Gene Lay Institute of

last: Immunology and Inflammation

address: 60 Fenwood Road,

institute: Brigham and Women's Hospital


title: A cell-intrinsic glucocorticoid biosynthesis and sensing circuit maintains homeostatic Th17 cell state [RNA-seq]

summary: T helper 17 (Th17) cells are heterogeneous; homeostatic Th17 (Th17Hom) cells maintain tissue homeostasis and pro-inflammatory Th17 (Th17Inf) cells drive autoimmune inflammation. IL-23 drives Th17Inf, but the signals that maintain Th17Hom remain unclear. Here, we found that differential glucocorticoid (GC) production downstream of CYP11A1, a critical enzyme for steroidogenesis, distinguished Th17Hom cells from Th17Inf cells. Although TCR signaling opened the Cyp11a1 locus, TGFβ1 and IL-6, key cytokines for Th17Hom cell differentiation, maintained and amplified Cyp11a1. Th17Hom cell-derived GC signaled through the glucocorticoid receptor (GR), which was higher in Th17Hom cells compared to Th17Inf cells, thereby forming a circuit that maintained the homeostatic state. Integration of multi-omics data from CYP11A1- and GR-deficient Th17Hom cells revealed a gene network underlying this circuit. TGFβ1, a key node in the network, restored GC sensing to Th17Inf. We thus identify a GC signaling circuit that maintains Th17 homeostasis with implications for treating Th17-mediated autoimmunity.

overall_design: Naive 2D2⁺ T cells isolated from CYP11A1 WT or cKO, and GR WT or cKO mice, were differentiated into Th17Hom cells in vitro and then adoptively transferred into Rag1 KO recipient mice to induce EAE.  At the peak of disease, CNS-infiltrating 2D2⁺ Th17 cells were sorted for RNA-seq analysis in the following groups: (1) CYP11A1 WT(n=4) vs. cKO(n=6) ; (2) GR WT(n=5) vs. cKO(n=4). Separately, in vitro–differentiated Th17Inf cells were treated with dexamethasone (Dex), TGF-β1, or a combination of both for 2 days, followed by RNA-seq analysis: (3) Control (n = 3), Dex (n = 3), TGF-β1 (n = 3), and TGF-β1 + Dex (n = 3).

first: Dandan

last: Yang

first: Linglin

last: Huang

first: Ana

middle: C

last: Anderson


title: Interleukin-23 promotes a pro-inflammatory Th17 cell state by stabilizing RORγt and suppressing glucocorticoid receptor activity

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus


title: Interleukin-23 promotes a pro-inflammatory Th17 cell state by stabilizing RORγt and suppressing glucocorticoid receptor activity I

summary: Interleukin-23 receptor (IL-23R) signaling is critical for the generation of pro-inflammatory CD4+ IL-17-producing T cells (Th17) that can drive autoimmune tissue inflammation, but the underlying mechanisms are not clear. We integrated phosphoproteomic and transcriptomic data downstream of IL-23R and IL-12R, which share a common subunit, to identify mechanisms engaged specifically by IL-23. We identified chromodomain helicase DNA-binding protein 1 (CHD1), an epigenetic regulator, and the glucocorticoid receptor (GR), a transcription factor (TF), as mediators of IL-23R signaling. IL-23R activation promoted CHD1 interaction with TF STAT3 and co-binding at the TF RORγt locus to enforce a pro-inflammatory Th17 state. Conversely, IL-23R signaling altered phosphorylation of the GR, thereby preventing its activation and nuclear translocation, ultimately impairing GR-driven inhibition of pro-inflammatory Th17 gene programs. Our findings uncover two mechanisms by which IL-23 promotes a pro-inflammatory Th17 cell state, offering potential therapeutic targets for treating Th17-driven autoimmune tissue inflammation and restoring homeostasis.

overall_design: Human memory T cells were transduced with lentiviral vectors encoding either IL-23R or IL-12Rβ2. Following transduction, IL-23R⁺ or IL-12Rβ2⁺ cells were sorted and stimulated with IL-23 or IL-12, respectively, for 4 or 12 hours, in the presence or absence of anti-CD3/CD28 antibodies. Cells from all 14 resulting conditions (n = 2 per condition) were subjected to RNA-seq analysis.

first: Jianfei

first: Oier

last: Etxezarreta-Arrastoa

first: Alex

last: V Khodaverdian

first: Nir

last: Yoself

last: C. Anderson

first: Vijay

last: K. Kuchroo


title: Population ATAC-seq of in vitro polarized Th0 cells

summary: To understand the molecular programs driving T helper cell heterogeneity and fate, we profiled in vitro polarized Th0 using ATAC-seq. This data were generated as part of the timecourse experiment in GSE206607.

overall_design: We in vitro polarized Th0 cells from mouse naive CD4+ T cells and harvested the cells at 48 hours after polarization for ATAC-seq (n=3).

first: Alexandra

last: Schnell

last: Kuchroo


title: Interleukin-23 promotes a pro-inflammatory Th17 cell state by stabilizing RORγt and suppressing glucocorticoid receptor activity II

overall_design: Naive 2D2⁺ T cells isolated from CHD1 WT or cKO, and GR WT or cKO mice, were differentiated into Th17Inf cells in vitro and subsequently adoptively transferred into WT recipient mice to induce EAE. At peak disease, 2D2⁺ Th17 cells were isolated from the CNS or draining lymph nodes (dLN) for RNA-seq analysis across the following groups: (1) dLN: CHD1 WT (n = 3) vs. cKO (n = 4); (2) CNS: CHD1 WT (n = 3) vs. cKO (n = 4); (3) CNS: GR WT (n = 4) vs. cKO (n = 4). Separately, naive T cells from WT or IL-23R KO mice were differentiated into Th17Inf cells in vitro and treated with or without dexamethasone (Dex) for two days. Cells were then subjected to RNA-seq analysis under the following conditions: (4) WT Control (n = 4), WT + Dex (n = 4), IL-23R KO Control (n =4), and IL-23R KO + Dex (n = 4).


title: Sustained PRC1.1 activity preserves gene repression independently of PRC2.2 and restrains leukemic cell differentiation

city: Nijmegen

first: Joost

last: Martens

state: Nederland

address: Geert Grooteplein 28

department: RIMLS

country: Netherlands

institute: Radboud University

summary: Loss-of-function mutations in BCOR, a subunit of the non-canonical Polycomb Repressive Complex 1.1 (PRC1.1), are frequently observed in acute myeloid leukemia (AML) and associate with adverse risk. Paradoxically, leukemic stem cell viability in BCOR wild type AMLs strongly depends on PRC1.1 activity. Here, we use BCOR and KDM2B degron models to study PRC1.1 dependency in leukemic cells and find that BCOR is a bridging factor tethering the catalytic and chromatin-binding moieties of the PRC1.1 complex. BCOR degradation induces a quick localized loss of H2AK119ub at PRC1.1 target genes, whereas PRC2-induced H3K27me3 remains unaffected. Degron-mediated depletion of BCOR or KDM2B induces a rapid but time-dependent transcriptional induction, whereby late-upregulated genes are more heavily decorated with H3K27me3 compared to early-upregulated genes. Combined PRC1.1 inactivation and PRC2 inhibition further amplifies gene induction suggesting collaborative yet distinct control over target genes. Strikingly, both JARID2/AEBP2 and SUZ12 knockout cells, devoid of PRC2.2 or PRC2.1/PRC2.2 respectively, retain PRC1.1 loss-induced transcriptional activation, underscoring that PRC1.1 can repress target genes independently of a downstream PRC2.2-canonical PRC1 repressive axis. Finally, combined targeting of PRC1.1 and PRC2 induces differentiation of leukemic cells emphasizing that co-targeting PRC1.1 and PRC2 represents a promising strategy to improve treatment of AML patients.

overall_design: Investigate the binding of PRC1.1 complex members in AML

first: Fatemeh

last: Mojallali

first: Tjerk-Jan

last: Atsma

first: Shanna

last: Hogeling

first: Rianne

last: Kloosterman

first: Eve

last: Ioannou

first: Albertus

last: Wierenga

first: Gerwin

last: Huls

first: Jan-Jacob

last: Schuringa

first: Vincent

last: van den Boom


title: Proper nuclear compartment reorganization in G1 phase is essential for DNA replication control [scRepli-seq]

city: Kobe

first: Ichiro

last: Hiratani

state: Hyogo

address: 2-2-3 Minatojima-minamimachi, Chuo-ku

department: Center for Developmental Biology

country: Japan

institute: RIKEN

summary: Upon G1 entry after mitosis, mammalian chromosomal DNA is spatially segregated into A and B compartments, which correlate well with classic euchromatin and heterochromatin, respectively. The significance of this spatial segregation, however, has never been explored for lack of means to manipulate this level of chromosomal organization. Through a genome-wide CRISPR screen, we identify GINS4, a component of the replicative DNA helicase complex, as an essential factor for A/B compartment segregation in G1. By manipulating the G1 compartment reorganization process through GINS4, we show that it is required for efficient DNA synthesis and proper DNA replication timing (RT) regulation in the subsequent S-phase. Our findings reveal an unexpected role of GINS4 and highlight the significance of nuclear compartmentalization in DNA replication control.

overall_design: scRepli-seq experiment was performed using the male C57BL/6N mouse embrypnic stem cells (mESCs). Genotype and treatment for each cell line is following: Gins4-mAID control, Gins4-mAID thymidine-released early-S samples, Gins4-mAID 5-Ph-IAA released early-S samples. Metadata for each sample can be found in the Supplementary Table 4.

first: Asami

last: Oji


first: Alessandra

last: De Meis

first: Sinne

last: Alkema


title: LDB1 regulates gene expression and chromatin structure in pluripotency and lineage differentiation [ATAC-seq]

city: Bethesda

first: HeungSun

last: Kwon

state: MD

address: 50 South Drive Building 50

department: NIDDK

institute: NIH

summary: Chromatin organization is a pivotal factor in stem cell pluripotency and differentiation. However, despite considerable study, the role of enhancer looping protein LDB1 in these processes has not been explored. Here, we generated Ldb1(-/-) embryonic stem cells (ESC) using CRISPR/Cas9 editing to understand the significance of LDB1 during development. Ldb1(-/-) ESC exhibited a reduction in key stem cell factors SOX2 and KLF4. Embryoid bodies (EB) derived from Ldb1(-/-) ESC displayed reduced expression of lineage-specific markers and impaired ability to undergo terminal differentiation to erythroblasts. Transcriptome analysis revealed differential gene expression between WT and Ldb1(-/-) ESC and EB but altered gene expression was most pronounced after differentiation to erythroblasts. The Lin28-mediated stem cell self-renewal pathway was dysregulated in Ldb1(-/-) cells. Our data reveal that LDB1 occupies super enhancers of pluripotency genes in ESC together with pluripotency factors. LDB1 loss resulted in a global decrease in chromatin accessibility in ESC and EB. Conditional LDB1-deficient mice showed reduced hematopoietic stem cell markers on bone marrow cells, and dysregulation of the Lin28/Let-7/Hmga2 pathway. We conclude LDB1 function is critical for ESC and EB development and becomes progressively more important during differentiation to erythroblasts.

overall_design: To investigate the role of LDB1 in development, we established Ldb1 (-/-) stem cell lines using CRISPR-Cas9 technology. To assess the differential accessibility of chromatin upon loss of LDB1, we conducted a genome wide analysis using Assay for Transposase-Accessible Chromatin sequencing (ATAC-seq).

first: Heungsun

first: Juhyun

last: Kim

first: Lecong

last: Zhou

first: Ann

last: Dean


title: LDB1 regulates gene expression and chromatin structure in pluripotency and lineage differentiation [ChIPmentation]

overall_design: To investigate the role of LDB1 in development, we established Ldb1 (-/-) stem cell lines using CRISPR-Cas9 technology. To investigate protein-DNA interactions, histone markers, and transcription factors on chromatin, we performed ChIPmentation experiment.


title: LDB1 regulates gene expression and chromatin structure in pluripotency and lineage differentiation [RNA-seq]

overall_design: To investigate the role of LDB1 in development, we established Ldb1 (-/-) stem cell lines using CRISPR-Cas9 technology. We performed a comparative analysis of gene expression using RNA-seq data from ESCs, EBs, and differentiated cells


title: LDB1 regulates gene expression and chromatin structure in pluripotency and lineage differentiation [Cut&Tag]

overall_design: To investigate the role of LDB1 in development, we established Ldb1 (-/-) stem cell lines using CRISPR-Cas9 technology. To investigate protein-DNA interactions, histone markers, and transcription factors on chromatin, we performed Cut-and-Tag experiment.


title: Single-cell TCR  sequencing from skin, blood and cerebral spinal fluid (CSF) samples from Degos disease patients.

first: Stephen

last: Brooks

address: 50 SOUTH DR, RM 1140

department: Biodata Mining and Discovery Section

institute: NIAMS/NIH

summary: Single-cell TCR-sequencing was pefromed to identify clonal expansion of T cells in Skin, blood and CSF samples from Degos disease patients.

overall_design: Single cell suspension from skin was prepared by enzymatic digestion and live cells were enriched by fluorescence-activated cell sorting (FACS) using Sytox green. Peripheral blood mononuclear cells (PBMCs) were isolated from blood according to density centrifugation method. PBMCs and CSF were subjected to ACK lysis treatment to remove RBCs. Single cell suspension prepared from skin, PBMCs and CSF were subjected to 10X capturing using 5'v2 or 5'-HT kit followed by TCR cDNA and libraray preparation. Finally, TCR libraries were sequenced and data was analysed using Sc-repertoire package (ScRepertoire 2.0.7).

first: Goel

last: Shubham

first: Jin

last: Seon-Pil

first: Sakamoto

last: Keiko

first: Keisuke

last: Nagao


title: Single-cell RNA transcriptomic profile from skin, blood and cerebral spinal fluid (CSF) from Degos disease patients.

summary: Skin, blood and CSF samples from Degos disease patients were subjected to single-cell RNA-sequencing analysis to characterize the disease pathophysiology and identify for a therapeutic target.

overall_design: Single cell suspension from skin was prepared by enzymatic digestion and live cells were enriched by fluorescence-activated cell sorting (FACS) using Sytox green. Peripheral blood mononuclear cells (PBMCs) were isolated from blood according to desity centrifugation method. PBMCs and CSF were subjected to ACK lysis treatment to remove red bllod cells (RBCs). Finally, their transcriptome were assessed by single-cell RNA-sequencing using the 10X genomics platform.


title: DREAMS Illuminates DNA & RNA Modification Landscapes  with a Spotlight on TET1's New Role in m1A

city: Beijing

first: Liang

last: Xue

state: Beijing

address: Bldg.3, NO.9, YIKE Rd, Zhongguancun Life Science Park, Changping District, Beiji

institute: Chinese Institute for Brain Research, Beijing(CIBR)

summary: We developed DNA RNA Elements Areal Mass Spectrometry (DREAMS), a mass spectrometry imaging platform that spatially maps diverse nucleic acid modifications. Applying DREAMS to TET-deficient mouse brains, we uncover TET1’s unexpected role in regulating N1-methyladenosine (m1A), a pivotal RNA modification. While DREAMS reveals broad modification landscapes altered across TET knockouts, we establish TET1-m1A as a central regulatory node, revealing its regulation of neuronal activity-related transcripts and interplay with other modification-altering systems. Our work establishes DREAMS as a transformative tool for spatial epigenomics/epitranscriptomics and redefines TET enzymes as integrators of interconnected nucleic acid modification systems.

overall_design:
We applied RNA immunoprecipitation (RIP) technique to locate the specific loci within the transcriptome where TET1 modulates m1A. Total RNA extracted from CPu of Tet1 KO mice was compared with Tet1 floxed mice. m1A-specific antibodies were utilized to enrich and conduct high-throughput sequencing of m1A-modified RNA molecules (m1A RIP-seq).
Please note that each processed data file was generated from both input and m1A pull-down samples, and is linked to the corresonding m1A pull-down sample records.

first: Yan

last: Peng

first: Yujie

last: Zhang

first: Shuangshuang

last: Feng

first: Houyu

first: Mingjie

last: Dong

first: Yiting

last: Lu

first: Chenyang

last: Wang

first: Meng

last: Bao

first: Waleed

last: Alam

first: Lei

first: Jie

last: Yin

first: Jiuming

last: He

first: Magdalena

last: Koziol


title: Adipogenesis of human mesenteric tissue

city: Guangzhou

first: Ren

last: Mao

address: 58 Zhongshan Road 2nd, Guangzhou

institute: The First Affiliated Hospital, Sun Yat-Sen University

summary: Creeping fat (CrF), defined as the hyperplasia of mesenteric adipose tissue surrounding inflamed intestinal segments, is a characteristic pathological feature of Crohn’s disease and is associated with disease progression and postoperative recurrence. To investigate the cellular composition and transcriptional alterations underlying CrF formation, we generated a single-nucleus RNA sequencing atlas of human mesenteric tissues from patients with Crohn’s disease and healthy controls. This dataset includes intestinal creeping fat (ICF) and corresponding intestinal non-inflamed fat (INF) from patients with Crohn’s disease, as well as normal mesenteric tissue (NL) from healthy controls. The data were used to characterize disease-associated changes in mesenteric tissue cell populations and to explore potential cellular and molecular mechanisms involved in CrF formation.

overall_design:
Single-nucleus RNA sequencing was performed on human mesenteric tissue samples, including five intestinal creeping fat (ICF) samples and five corresponding intestinal non-inflamed fat (INF) samples from patients with Crohn’s disease, together with three normal mesenteric tissue (NL) samples from healthy controls. Nuclei were isolated from frozen mesenteric tissues and subjected to single-nucleus transcriptomic profiling. Processed gene-barcode matrices were generated for each sample and used to construct a cellular atlas of human mesenteric tissues across disease-associated and control conditions.

For GSM9812225-GSM9812238:
Submitters state "Raw read files are not provided due to patient privacy concerns and institutional data-sharing restrictions"

***For GSM9812225-GSM9812238, raw files for human/patient samples were not submitted to GEO due to concerns about submitting personally identifiable sequence data for open access***


title: Spatial mapping of RNA turnover kinetics in the mouse brain

city: Philadelphia

first: Hao

last: Wu

state: PA

address: 415 Curie Boulevard

department: Genetics

institute: University of Pennsylvania

summary: Gene regulation requires coordinated control of RNA synthesis and degradation, yet measuring RNA turnover across intact tissues remains challenging. Here we present spatial NT-seq, a method that combines transgenesis-free metabolic RNA labeling with in situ chemical recoding on spatial transcriptomics platforms to co-map newly synthesized and pre-existing RNAs. Applying spatial NT-seq to the mouse brain reveals pronounced regional heterogeneity in RNA turnover and identifies the dentate gyrus as a spatial hotspot marked by coordinated up-regulation of basal RNA synthesis and decay. Moreover, spatial NT-seq uncovers rapid, brain region-specific transcriptional and post-transcriptional responses to electroconvulsive stimulation, a clinically relevant treatment for refractory depression. Finally, we leverage computational modeling to identify sequence features and post-transcriptional regulators that shape transcriptome-wide mRNA stability across spatial and cellular contexts in the mouse brain. Together, this integrated “in vivo Timescope” framework provides a spatially resolved view of RNA turnover kinetics and reveals the regulatory architecture of RNA stability in vivo.

overall_design: Refer to individual Series


title: Spatial mapping of RNA turnover kinetics in the mouse brain [Spatial NT-seq]

overall_design: To spatially co-map RNA abundance and turnover in intact tissues, we developed spatially resolved New RNA Tagging Sequencing technology (spatial NT-Seq). The newly synthesized RNAs in postnatal day 7 (P7) or 56 (P56) mouse brains were labeled by intraperitoneally injecting 4-thiouridine (4sU), and whole brains were dissected and embedded in OCT for spatial NT-seq analysis. This method combines transgenesis-free in vivo 4sU labeling, fresh frozen tissue sectioning, methanol fixation, and histological staining with an optimized in situ SLAM reaction  to chemically recode 4sU-labeled new mRNAs directly within tissue sections. This strategy is broadly compatible with sequencing-based spatial transcriptomics (ST) methods, including the Visium patform used in this study, which contains spatially barcoded mRNA capture oligonucleotides. After tissue permeabilization, modified reverse transcription (RT) and second-strand synthesis reactions were used to efficiently capture newly synthesized (“new”) and pre-existing (“old”) transcriptomes from the same tissue section.

first: Qi

last: Qiu

first: Hongjie

first: Zijie

last: Xia

first: Zhaolan

last: Zhou


title: Spatial mapping of RNA turnover kinetics in the mouse brain [scNT-seq2]

overall_design:
The lack of systematic validation has limited the application of in vivo metabolic RNA labeling for studying RNA turnover and stability in tissues. Although 4sU enables direct RNA labeling in non-neural tissues,  its labeling efficiency and potential toxicity in the brain remain uncertain. To address these gaps and enable analysis of challenging brain cell types, we optimized droplet-based single-cell metabolically labeled new RNA tagging sequencing (scNT-seq) to improve chemical conversion efficiency and library complexity (scNT-seq2). Using scNT-seq2, we systematically validated transgenesis-free 4sU labeling for profiling RNA turnover in embryonic and postnatal mouse brains.

First, using the ‘TU tagging’ as a benchmark, we evaluated the in vivo RNA labeling efficiency with 4sU in the brain. We generated “Ex-UPRT” mice by crossing a Nex/Neurod6-Cre driver with a conditional UPRT line (CA>GFPstop>UPRT), achieving excitatory (Ex) neuron-specific Uprt expression. Postnatal day 12 (P12) “Ex-UPRT mice” were injected intraperitoneally with 4tU or 4sU at equimolar doses, and scNT-seq2 identified major cortical cell-types and confirmed Uprt transgene expression restricted to Neurod6-expressing Ex neurons. Both 4sU and 4tU labeling minimally altered cell-type specific transcriptomic states, as shown by closely matched distributions of cortical cell-types in treated and control mice. Quantification of labeled new RNA fractions indicated robust labeling in Neurond6-positive Ex neurons with both 4sU and 4tU, with highly concordant transcriptome-wide gene level measurements from 4sU and 4tU labeling in cortical Ex neurons. Notably, 4sU also efficiently labeled new RNAs in Neurod6-negative cell types independent of UPRT expression, whereas 4tU labeling in these cell-types was substantially lower, likely due to low endogenous UPRT-like enzymatic activity. Thus, 4sU enables efficient and broader RNA labeling in the brain, matching 4tU labeling efficiency in UPRT-expressing cells.

Second, we evaluated transgenesis-free 4sU RNA labeling in wild-type animals. Time-pregnant females were injected 4sU, and embryonic day 16.5 (E16.5) embryonic cortices were compared with primary E16.5 cortical cells labeled in vitro.  This comparison showed similar increases in 4sU-labeled new RNA fractions (in vitro: 16.5%, in vivo: 15.8%) compared to saline controls (<1.0%), with comparable intrinsic cell-to-cell variability. Transcriptome-wide comparisons of gene-level new-to-total RNA ratios (NTRs) showed high concordance between in vitro and in vivo labeling, demonstrating that 4sU efficiently crosses both fetal-maternal and blood-brain barriers to label new RNAs in developing embryonic brains.

Third, we evaluated the effects of 4sU labeling duration and dosage on RNA labeling across 26,640 cortical cells from nine wild-type P7 mice. Consistent with “Ex-UPRT” mice, 4sU minimally affected cell-type composition. Labeled RNA fractions increased almost linearly with labeling duration, were reproducible across biological replicates and cortical cell-types, and plateaued at ~1 mg/g with increased 4sU dose. Importantly, 4sU labeling did not affect scNT-seq2 performance.


title: Proper nuclear compartment reorganization in G1 phase is essential for DNA replication control [Hi-C]

first: Ichiro

last: Hiratani

overall_design: Hi-C experiment was performed using the male C57BL/6N mouse embrypnic stem cells (mESCs). Genotype of each cell line is following: mAID-Mcm6 , Gins4-mAID Metadata for each sample can be found in the Supplementary Table 4.

first: Asami

last: Oji

first: Takako

last: Ichinose

first: Izumi

last: Noda


title: Nuclear compartmentalization at the G1/S transition plays a key role in DNA replication control

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


title: HIF1 Counteracts FOXO1/Bim-associated Follicle Atresia in Mouse Ovaries by Promoting Bcl-XL Expression.

status: Public on Jun 11 2026

first: Ming

last: Hao

address: Shuangqing Rd.

institute: Tsinghua Univ

summary:
Background
In mammalian ovaries, the majorities of developing follicles (up to 95% in human) undergo atresia. A considerable part of follicle atresia is initiated by granulosa cell (GCs) apoptosis. Our previous work identified hypoxia-inducible factors 1 (HIF1) pathway induced by follicle-stimulating hormone as a protective factor against atresia, yet the molecular mechanism of follicles’ “norm” death, and how HIF1 pathway protect follicles has not been fully delineated.
Methods
We employed a single-follicle, multi-dimensional profiling strategy based on serial sectioning, to dissect the signaling logic underlying physiological follicle fate decisions.
Results
HIF1 and forkhead box protein O1 (FOXO1) pathway show complementary activation patterns in GCs of healthy versus atretic mouse follicles. In growing mouse follicles, HIF1 pathway activity is associated with increased reactive oxygen species (ROS)-related signals, likely through enhanced granulosa-cell metabolic activity. Under the physiological conditions, ROS-related signals appear to help maintain HIF1 pathway activity in mouse granulosa cells during follicle growth, and are not by themselves associated with follicle atresia. HIF1 pathway appears to promote Bcl-XL expression, which may counterbalance FOXO1-associated atretic pressure.
Conclusion
In mouse ovaries, follicular fate appears to be associated with the dynamic balance between the pro-survival HIF1/Bcl-XL axis and the pro-apoptotic FOXO1/Bim axis.

overall_design: Hif1aflox/flox and Foxl2-CreERT2 mice were crossed, and the Hif1aflox/flox; Foxl2-CreERT2 and Hif1aflox/flox female mice were kept, and bred up to 3-week-old, and injected with 100 mg/kg tamoxifen each day for 5 days. Ovaries were collected 7 days after the last tamoxifen injection.Ovaries was cryosectioned and Granulosa cells were collected by laser microdissection, followed by RNA extraction, reverse transcription, amplification and sequencing.

first: Longping

last: Liu

first: Jianbing


title: Gene Expression Profiling of Synechococcus elongatus PCC 7942 and Rhodotorula toruloides NBRC 0880 (Rhodosporidium toruloides IFO0880) in Co-culture and Membrane-Separated Bioreactor Configurations Under Various Stress and Nutrient Conditions

city: Richland

first: Predictive Phenomics

last: Initiative

state: WA

address: 902 Battelle Boulevard

institute: Pacific Northwest National Laboratory

summary: This RNA-seq study examines transcriptional responses of a cyanobacterium–yeast co-culture system comprising Synechococcus elongatus PCC 7942 and Rhodotorula toruloides NBRC 0880 (Rhodosporidium toruloides IFO0880) under multiple conditions. Experiments included planktonic co-cultures in 50 mL tubes subjected to chemical stressors (oxidative stress, photosynthesis inhibition), temperature stress, nutrient limitation (nitrogen, phosphorus), pH extremes, and heterotrophic conditions, as well as physically separated co-cultures in a 600 mL membrane bioreactor. Axenic controls for Synechococcus elongatus PCC 7942 were included for comparison. Differential gene expression analysis provides insight into interspecies interactions and individual organism responses across co-culture configurations.

sample_organism: Synechococcus elongatus PCC 7942 = FACHB-805

overall_design: Transcriptional analysis of S. elongatus PCC 7942 and R. toruloides NBRC 0880 was performed across three experimental setups: (1) co-cultures in 50 mL tubes under ~20 perturbation conditions with 2–3 biological replicates each, compared against axenic controls of S. elongatus PCC 7942; (2) membrane-separated bioreactor (~600 mL) where the two organisms were physically separated by a membrane but shared a common medium, sampled at multiple time points with 3 biological replicates, with axenic controls for both organisms maintained in parallel; and (3) mixed co-culture bioreactor (~600 mL) where S. elongatus PCC 7942 and R. toruloides NBRC 0880 were physically co-located in the same reactor space, sampled at the same time points and serving as a direct comparison to the membrane-separated configuration.

first: Pavlo

last: Bohutskyi

first: Zachary

last: Johnson


title: Glucokinase activity suppresses hepatic cholesterol synthesis and triglyceride accumulation: A new model for the effects of the GKRP P466L common human variant

first: Patrick

last: Seale

address: 3400 Civic Center Blvd.

department: Institute of Diabetes, Obesity, and Metabolism

summary: Objective: Glucokinase Regulatory Protein (GKRP) controls the activity of Glucokinase (GCK) to regulate liver glucose uptake and storage.  Coding variants in GCKR, the gene encoding GKRP, strongly associate with fatty liver disease, hypertriglyceridemia, and hypercholesterolemia. Here, we sought to investigate the mechanisms by which a common GKRP variant affects hepatic lipid and cholesterol metabolism. Methods: We developed mouse models to examine how the human GKRP P446L variant influences liver and systemic metabolism. Endogenous Gckr expression was ablated in adult mouse hepatocytes, together with re-expression of either human GKRP-P446L or the reference GKRP protein. We assessed body weight, adiposity, systemic glucose homeostasis, and hepatic metabolites in mice expressing reference GKRP or GKRP P446L under multiple metabolic conditions. To determine whether the effects of GKRP P446L may result from reduced GCK activity, we analyzed mice with liver-specific deletion of Gck. Results: Hepatic expression of GKRP P446L resulted in reduced GKRP and GCK protein levels and elevated serum cholesterol. Hepatic deletion of Gck in mice recapitulated several effects of GKRP-P446L, including increased hepatic cholesterol and triglyceride content. The elevated cholesterol was associated with increased cholesterogenic gene expression and cholesterol synthesis. Hepatic expression of an alternative hexokinase (HKII) normalized the effects of GCK-deficiency, suggesting that impaired glucose phosphorylation underlies the phenotype. Conclusions: The GKRP P446L variant reduced GKRP protein abundance, and diminished GCK activity while increasing cholesterol levels. Loss of GCK elevated cholesterol and hepatic triglyceride levels. Collectively, these findings demonstrate that GCK suppresses hepatic cholesterol synthesis and lipid accumulation, suggesting that reduced GCK activity underlies the metabolic abnormalities associated with the GKRP P446L variant.

overall_design: RNA-seq profiling of liver samples from 10-12 week old control and L-GCK-KO mice after 1 week on high carbohydrate diet that were overnight fasted followed by a 6 hour refeed

first: Santoleri

last: Dominic

first: Traynor

last: Sarah

first: Calhoun

last: Ryan

first: Gavin

last: Matthew J.

first: Merrick

last: David

first: Seale

last: Patrick

first: Titchenell

last: Paul M.


title: Protocol for scalable generation of uniform 3D porcine oviduct epithelial cell spheroids

city: Urbana

first: Hao-Chun

last: Fan

address: 1207 West Gregory Drive

department: College of Agricultural, Consumer and Environmental Sciences

institute: University of Illinois Urbana-Champaign

summary: While previous studies have reported in vitro formation of oviduct epithelial spheroid models in porcine species (sus scrofa), these approaches often rely on manual selection of the formed aggregates. Here, we establish a scalable and reproducible system for generating oviduct spheroids. We describe steps for the isolation of primary secretory and ciliated oviduct epithelial cells and their assembly into spheroids using the AggreWell platform. This approach enables the mass production of uniformly sized spheroids ready for use in experiments such as sperm-oviduct co-incubation experiments.

overall_design: Bulk 3′ mRNA-seq profiling of freshly isolated porcine oviductal epithelial cells and spheroids formed in DMEM/F12 and Incubation Medium

first: Leonardo

last: Molina

first: Adrienne

last: Antonson

first: David

last: Miller


title: Genome-wide characterization of extrachromosomal circular DNA and its functional role in colon cancer carcinogenesis and progression [miRNA]

city: Suzhou

first: Wenbo

last: Jia

state: Jiangsu Province

address: No.199 Ren'ai Road

institute: Soochow University

summary: Extrachromosomal circular DNAs (eccDNAs) are defined as distinct genomic entities of circular and mobile DNA molecules, but their molecular functions in and impact on breast cancer (BC) are rarely known. This study used Circle-seq to analyze eccDNAs from 19 BC tissues and 17 adjacent normal tissues. We found that eccDNAs are present on all chromosomes and enriched in seven eccDNA hotspot genes (HSGs) associated with the BC pathway. Several eccDNAs harboring entire genes (eccGenes) and eccDNAs harboring miRNAs (eccMIRs) were identified and linked to cancer-relevant pathways. Synthetic eccMIR6748, eccMIR6508, and eccMIR3142 elevated miRNA expression in MCF-7 cells, with eccMIR6748 promoting BC cell migration and invasion by upregulating miR-6748, which suppresses tumor suppressor candidate factor 5 (TUSC5) at the post-transcriptional level. eccMIR6748 also influences BC progression via the p38 mitogenactivated protein kinase (MAPK) signaling pathway. These findings suggest that eccDNAs, which contain functional genomic segments, play a role in BC initiation and progression, offering a dynamic source of genomic plasticity and potential as biomarkers and therapeutic targets.

overall_design: This study includes three complementary sequencing assays on human samples to investigate circular DNA, microRNA, and mRNA expression profiles.A total of 6 samples were analyzed, comprising two experimental groups with 3 biological replicates per group. A: Circle seq (circular DNA). Paired end 150 bp sequencing on Illumina NovaSeq 6000. Linear DNA was removed by exonuclease treatment, followed by rolling circle amplification. Clean reads were aligned to the reference genome (HG38) using BWA, and circular DNAs were identified with circle map. Differential expression was analyzed with edgeR. B: miRNA seq. Single end sequencing. Small RNA libraries were constructed and sequenced. Reads were trimmed with cutadapt, aligned to miRBase pre miRNAs (v22) plus newly predicted pre miRNAs using Novoalign. miRNA expression was normalized by TPM. Novel miRNAs were predicted with miRDeep2. Differential expression was calculated using edgeR. C: mRNA seq. Paired end 150 bp sequencing on Illumina NovaSeq 6000. mRNA was enriched via poly A selection, strand specific libraries were prepared. Raw reads were preprocessed with fastp, aligned to HG38 using HISAT2, and gene counts were obtained with featureCounts. Differential expression was analyzed with edgeR). All differential analyses were followed by GO and KEGG pathway enrichment. Data visualization was performed with IGV, cluster heatmaps, scatter plots, and volcano plots.

first: Ji

last: Wang

first: Kehan


title: Targeting chordoma via an Isocitrate Dehydrogenase-1 dependent susceptibility to redox metabolism

city: Ann Arbor

first: Sriram

last: Venneti

state: Michigan

address: 1500 E Medical Center Dr

department: Pathology

institute: University of Michigan

summary: Chordomas are rare cancers that arise along the axial skeleton. Alterations in metabolism are a hallmark of cancer, and we sought to identify metabolic vulnerabilities in chordoma. We discovered that the tricarboxylic acid (TCA)-related enzyme isocitrate dehydrogenase-1 (IDH1) was expressed highly in bulk and single-cell patient-derived chordomas and was associated with worse survival outcomes. IDH1 catalyzes the conversion of isocitrate and nicotinamide adenine dinucleotide phosphate (NADP+) to alphaketoglutarate (a-KG) and NADPH. This critical reaction influences TCA cycle metabolism, regulates epigenetic pathways, and affects redox balance. Both IDH1 knockdown and treatment with an inhibitor targeting IDH1 were toxic to chordoma cells. An integrated analysis of the transcriptomic, chromatin, and metabolomic responses on IDH1 inhibition converged on deregulated glutathione metabolism. IDH1 inhibition was associated with increased expression and enrichment of activating H3K27ac at NRF2 (Nuclear factor erythroid 2-related factor 2) signature genes including those in the glutathione biosynthetic pathway. This was accompanied by reduction of both NADPH/NADP+ and reduced/oxidized glutathione (GSH/GSSG) ratios. Importantly, IDH1 inhibitor-driven toxicity was rescued via media supplementation with the antioxidant N-acetyl cysteine, suggesting that IDH1 inhibition in chordomas creates a redox-dependent metabolic vulnerability. Finally, IDH1 inhibitor treatment reduced tumor growth in two independent chordoma mouse xenograft models. Our findings suggest a potential therapeutic avenue for further exploration in chordoma.

overall_design: To understand the role of IDH1 protein expression in Chordoma, Chordoma cells (UM-Chor1, CH22 and JHC7) were treated with 3uM IDH1 inhibitor and RNAseq was performed.


title: Stromal-immune interactions promote immune suppression in intraperitoneal metastasis of ovarian carcinoma

city: Rehovot

first: Neta

last: Erez

address: Haim Holtsman St 1

department: Systems Immunology

country: Israel

institute: Weizmann Institute of Science

summary: High-grade serous (tubo-ovarian) carcinoma (HGSC) is the most lethal gynecological malignancy and the most common form of ovarian cancer. Most patients (75%) are diagnosed at advanced disease stages, with widespread peritoneal dissemination leading to poor prognosis. Cancer-associated fibroblasts (CAFs) were shown to modulate the immune microenvironment in multiple cancer types. However, their interactions with immune cells and their role in facilitating peritoneal metastasis in ovarian cancer are largely unresolved. Here, we characterized the interactions between fibroblasts and immune cells that facilitate the formation of a hospitable niche in metastatic peritoneal ovarian cancer. We isolated and characterized CAFs from HGSC patients and mouse models, and investigated the events that promote peritoneal metastases. Our findings reveal the reciprocal communication between fibroblasts and T cells in distinct peritoneal metastatic niches, positioning the TIGIT-CD155 immune checkpoint signaling axis as a target for therapeutic intervention in metastatic HGSC. Our study provides a potential novel avenue for therapeutic intervention to inhibit metastatic spread of ovarian carcinoma.

overall_design: Fibroblasts were isolated by cell sorting from normal, early stage and late-stage metastatic peritoneum of Col1a-YFP+ transgenic mice.

last: Solomon

first: Lea

last: Monteran

first: Dan

last: Grisaru


title: Genome-wide RNA binding landscapes of PABPC1 and CCT3 in GC-2spd(ts) and verification of their direct interactions with lncRNA Tug1

city: Nanjing

first: lingxiu

last: Tong

state: Jiangsu

address: Nanjing Medical University 101 Longmian Avenue, Jiangning District, Nanjing 211166, Jiangsu Province, P.R. China

institute: Nanjing Medical University

summary: Verification of direct interactions between lncRNA Tug1 and RNA-binding proteins PABPC1 and CCT3 and genome-wide RNA binding maps of PABPC1 and CCT3 in GC-2spd(ts)

overall_design: eCLIP-seq profiling of GC-2spd(ts) cells and their immunoprecipitation derivatives (PABPC1, CCT3, and Input control)

first: Lingxiu

first: Yiting

last: Ye

first: Ke

last: Zheng


title: The challenges of single cell transcriptomics on difficult human tissue: the placenta [Xenium]

city: London

first: Theodoros

last: Xenakis

address: 20 Guilford Street

institute: UCL

summary: Demand for the application of single cell transcriptomics on difficult tissues, processed and stored in disparate conditions, has led to the development of various single cell modalities. We focus on the placenta, a challenging tissue to transcriptomically interrogate due to senescence, intermittent hypoxia, high levels of RNase activity and tissue trauma at delivery. We performed single cell and nuclei transcriptomics on two samples with probe-based and native molecule transcript capture, droplet based and in situ plate based cell isolation. We explored sample and storage variations, including freshly dissociated tissues, fixed cells, snap frozen and FFPE processing. We find that variations in sample processing and storage have much larger effect on cell type proportions than differences in chemistry, impacting the biology that can be learned or the identification of disease markers. Further, the transcriptomic output of in situ combinatorial indexing consistently overlaps, with single nucleus transcriptomics, sharing little with other modalities. This may limit combinatorial indexing for sampling cytoplasmstic mRNAs. Our comprehensive analysis of the varying effect of single cell transcriptomic modalities, including their sample management strategies, provides novel and essential considerations for the experimental design and analysis of single cell transcriptomics applicable to challenging tissues, offering actionable guidance for future experiments.

overall_design: High throughput in situ hybridisation using 10X Genomics Xenium platform on a control placenta at 35 gestational weeks.


title: Clinical development of tacrolimus-resistant regulatory T cells to enable simultaneous immunosuppression and support of immune regulation

city: Berlin

first: Julia

last: Polansky

address: Augustenburger Platz1

institute: Berlin Institute of Health Center for Regenerative Therapies

type: Methylation profiling by genome tiling array

summary: Unwanted alloimmune responses are a central driver of solid organ transplant rejection and currently managed with life-long immunosuppression, which imposes substantial risks and burdens on patients. Adoptive transfer of regulatory T cells (Tregs) offers a strategy to restore immunological balance and reduce long-term adverse effects of generalized immunosuppression. However, while Tregs can potently inhibit incipient immune activation, they struggle to suppress established memory effector T cells, necessitating the continued use of immunosuppression. Calcineurin inhibitors such as Tacrolimus effectively control both, newly activated and pre-existing effector-T-cells, but unfortunately, they also impair Treg function. Therefore, we hypothesized that gene-editing of Tregs inducing Tacrolimus resistance (FKBP12KO) would enable combined therapy that curbs effector T-cell responses without compromising Treg efficacy. Here, we developed FKBP12KO-Tregs using a ribonucleoprotein-based CRISPR/Cas9 approach and characterized them extensively in-vitro. FKBP12KO-Tregs retained phenotype, high viability, and suppressive function comparable to unedited TregWT and they remained functionally impervious to Tacrolimus, while preserving sensitivity to alternative CNIs. We additionally established a Good-Manufacturing-Practice process for FKBP12KO-Tregs. Comprehensive in vitro phenotypic, functional, and molecular characterization, together with the established manufacturing, provide the rationale for a proof-of- concept clinical trial assessing the feasibility and safety of co-administration of FKBP12KO-Tregs with Tacrolimus in living-donor kidney transplant recipients.

overall_design: Isolation and culture of Treg CD4+ T-cells were isolated from peripheral whole blood either by a research-grade or a GMP protocol; i.e. EasySep™ Human CD4+ T-cell Enrichment Kit (STEMCELL Technologies) and CliniMACS Plus device (Miltenyi Biotec), respectively. CD4+ cells subsequently underwent fluorescently activated cell sorting (FACS) using MACSQuant-Tyto (Table S1, Miltenyi Biotec). Tregs were thereafter cultured with x- vivo-15 medium (Lonza) with 1% penicillin/streptomycin and 10% fetal calf serum (FCS) supplemented with Rapamycin at a concentration of 100nM/ml and recombinant human IL2 at a concentration of 1,250 IU/ml (Table S2). Tregs were subjected to expansion over the first 7 days with anti-CD3/CD28-MACSiBeads at 4:1 ratio and following 14 days at 1:1 ratio replenished every other day when cultivated in 24 well plates (research grade, pre-GMP). For GMP grade expansion, Tregs were cultured in G-Rex bioreactors (G-Rex 10 and 100, Wilson Wolf Manufacturing). An aliquot of the negative fraction of the FACSort, i.e. non-Tregs (Tcon) served as control, and was expanded using identical conditions. In addition, expanded and cyro-preserved CMV- antigen specific FKBP12KO-Teff were included as control samples for the proteomics and CITEseq analysis.      FKBP12 knock-out in Treg    On day 7, the Tregs were electroporated with ribonucleoprotein (RNP) complexes using the Amaxa-P3-primary-cell-4D-Nucleofector-X-Kit-L and the Amaxa- Nucleofector-4D (Lonza); after pre-complexing 10mg Alt-R™ S.p. (HiFi) Cas9 Nuclease V362 (Integrated DNA Technologies) with 15mg 2O’-methyl- 3'phosphothioate-modified sgRNA (Synthego) 17,18. MOCK controls were treated with Cas9 Nuclease only. The same protocol was applied to the control CMV-antigen specific FKBP12KO-Teff.

first: Ghazaleh

last: Zarrinrad

first: Lisa-Marie

last: Burkhardt

first: Claudia

last: Beltran-Mestres

first: Silvina

last: Romero-Suárez

first: Dimitrios

last: Laurin Wagner

first: Stephan

last: Schlickeiser

first: Maik

last: Stein

first: Désirée

last: Jacqueline Wendering

first: Iván

last: Juky Carrera Diaz de la Cebosa

first: Lukas

last: Ehlen

first: Frederik

last: Hamm

first: Gavin

last: Kurgan

first: Pawel

last: Durek

last: Heinrich

first: Anamika

last: Giri

first: Yaolin

last: Pu

first: Kristy

last: Ou

first: Henrike

last: Hoffmann

first: Sandra

last: Muench

first: Insa

last: Stuewe

first: Sven

last: Dolling

first: Oliver

last: McCallion

first: Jaspal

last: Kaeda

first: Jonas

last: Kath

first: Sarah

last: Schulenberg

first: Niklas

last: Wiese

first: Abdolreza

last: Nazari

first: Lena

last: Peter

first: Samira

last: Picht

first: Andrea

last: Sanchez-Pena

first: Olalekan

last: Usman

first: Christian

last: Brommel

first: Rolf

last: Turk

first: Garrett

last: Rettig

first: Morgan

last: Sturgeon

first: Thomas

last: Osborne

first: Ashley

last: Jacobi

first: Rebecca

last: Friedrich

first: Masako

last: Monika Kaufmann

last: Klermund

first: Simon

last: Fink

first: Markus

last: Templin

first: Andy

last: Roemhild

first: Daniel

last: Kaiser

last: Klein