title: Phenotypic impact of individual conserved neuronal microexons and their regulators in zebrafish

city: Barcelona

first: Manuel

last: Irimia

state: Barcelona

address: 88 Dr. Aiguader

department: MELIS

institute: Universitat Pompeu Fabra - Centre for Genomic Regulation

summary: Microexons show remarkable neuronal switch-like regulation and evolutionary conservation and they can impact protein-protein interactions and other molecular functions. Neuronal microexons are regulated in a tightly coordinated manner by the splicing factors Srrm3 and Srrm4. Depletion of Srrm3/4 in mice causes dramatic neurological phenotypes and their misregulation is associated with human disease. However, which microexon targets are associated with these phenotypes and how individual microexon deletions could impact neurodevelopment, physiology and behavior is unknown. To address this question, we generated zebrafish KO lines for 18 individual microexons and characterized their phenotypes at multiple levels, together with srrm3/4 mutant lines.  We found that, while mutation of the regulators, specially srrm3, shows multiple alterations in neuritogenesis, locomotion and social behavior, the impact of deleting of individual microexons was usually mild or non-existing. Nonetheless, for various microexons, we identified neuritogenesis defects (evi5b, vav2, itsn1, src) and social defects (vti1a, kif1b). In addition, microexon deletions led to transcriptomic alterations that are likely compensatory. Overall, our data suggest that the defects caused by srrm3/4 depletion are due to a cumulative effect of multiple small impacts that can individually be well tolerated by zebrafish.

overall_design: We deleted individual microexons in zebrafish using CRISPR-Cas9 with a double guide-RNA strategy. Stable mutant lines were generated for each microexon deletion, and rise to the F2 generation. To generate RNA-seq data, we cross two adult heterozygous individuals, and genotyped 5 dpf larva through fin clipping.  Larve of WT or DEL (homozygous deletion) genotype were grouped together, RNA extracted and submitted for  polyA-selected stranded RNA-seq.  A similar approach was taken for the mutant of the regulators. All fish had a HuC:GFP background.

first: Laura

last: Lopez-Blanch

first: Cristina

last: Rodriguez-Marin

first: Jon

last: Permanyer


title: Exercise Training-induced Extracellular miR-136-3p Modulates Glucose Uptake and Myogenesis through Targeting of NRDC in Human Skeletal Muscle

status: Public on Oct 31 2025

city: Stockholm

first: Mutsumi

last: Katayama

address: Solnavägen 9

department: Physiology and Pharmacology

country: Sweden

institute: Karolinska Institutet

summary:
We identified several exercise training-responsive miRNAs within exosome-rich EVs in serum, including miR-136-3p. In human myotubes, miR-136-3p enhanced glucose uptake and targeted the nardilysin convertase (NRDC) gene.
To search miR-136-3p and NRDC relating function in human myotubes, we performed transcriptome analysis.

overall_design: We used microarrays to detail the global regulation of gene expression by miR-136-3p and NRDC.

first: Anna

last: Krook


title: NF-κB-dependent redistribution of the GR cistrome recruits GR to inflammatory gene loci and correlates with reduced repression by glucocorticoid [GR_RELA_ChIPseq]

city: Calgary

first: Alex

last: Gao

state: Alberta

address: 3330 Hospital Dr NW

country: Canada

institute: University of Calgary

type: Genome binding/occupancy profiling by high throughput sequencing

summary: While ligand-activated glucocorticoid receptor (GR) binds DNA to activate transcription, glucocorticoids, including budesonide, reduce inflammatory gene expression, yet recruit GR to many such gene loci. In epithelial cells, the inflammatory cytokine, interleukin-1β (IL1B), activates NF-κB to induce gene expression and co-treatment with budesonide produces nanoscale GR-RELA nuclear co-localization. Such co-stimulation orchestrated reciprocal genome-wide redistribution of GR and RELA binding regions (GBRs and RBRs, respectively) relative to each mono-treatment to produce widespread GBR RBR overlap. This correlated with increased RNA polymerase-2 presence and required NF-κB for GR cistrome remodeling. Mapping transcription start sites to the nearest GBR or RBR both revealed associations with upregulated, but not repressed, genes. Importantly, RBR proximity to budesonide upregulated genes and GBR proximity to IL1B-upregulated genes correlated with attenuated repression on co-treatment. As this occurred on a background of glucocorticoid-induced repression, GR presence at specific IL1B-induced gene loci may reduce, or protect, from an otherwise prevalent glucocorticoid induced repression.

overall_design: To understand interactions between the GR and NF-κB cistromes, A549 cells were not treated, or treated with NS, IL1B, budesonide, or both before ChIP was performed using GR and RELA antibodies, and sequenced using Illumina NovaSeq 6000.

first: Mahmoud

middle: M

last: Mostafa

first: Robert

last: Newton

first: Andrew

middle: J

last: Thorne

first: Akanksha

last: Bansal


title: Comparison of gene expression in HeLa cells after treatment with IFN-b, IFN-g, or VPgEMCV.

city: Kyoto

first: Takashi

last: Fujita

state: Kyoto

address: 53 Shogoin Kawahara-cho, Sakyo-ku

department: Biosystems Science

country: Japan

institute: Kyoto University

type: Expression profiling by high throughput sequencing

summary: We uncovered that cells infected with the encephalomyocarditis virus (EMCV) release soluble factors that induce apoptosis in uninfected cells. These factors were identified as the cytokine TNF-α and the viral protein VPgEMCV, which is necessary for viral RNA synthesis. Intriguingly, we discovered that VPgEMCV also acts as a functional mimic of IFN-γ, activating its canonical signaling pathway. This includes phosphorylation of STAT1 and the induction of IRF-1 and iNOS, leading to increased nitric oxide (NO) production. The synergy of NO accumulation and TNF-α signaling initiates powerful apoptosis. To compare gene sets regulated by IFN-I signaling (IFN-b), IFN-II signaling (IFN-g), and VPgEMCV signaling, we utilized RNA-seq analysis. From the deep sequencing data, we further discovered that VPgEMCV induces a gene expression profile similar to IFN-II signaling (IFN-γ), while also uniquely downregulating a distinct subset of genes. Our findings reveal that this cytokine-mimicry function of VPgEMCV regulates genes to control both viral replication and disease pathogenesis.

overall_design: RNA-seq analysis comparing wildtype HeLa cells treated with IFN-b, IFN-g, or VPgEMCV 24h post-treatment.

first: Yutaro

last: Shirasaka


title: Training-induced exosomal miR-136-3p modulates mitochondrial functions by targeting NRDC in human skeletal muscle.

summary:
The purpose of this study was to validate the role of abundant miRNA, miR-136-3p, in extra vesicles collected from serum after training. We transfected miR-136-3p into human skeletal muscle cells.
To search of targets genes of miR-136-3p, we performed transcriptome analysis to compare expression profiles between control and miR-136-3p transfected human skeletal muscle cells.

overall_design: We used microarrays to detail the global regulation of gene expression by miR-136-3p and identified up- and down-regulated genes by miR-136-3p in human skeletal muscle cells.


title: SGSS05-NS3 as a potent SETD8 inhibitor that activates p53 pathway in Neuroblastoma

city: Rome

first: Veronica

last: Veschi

address: Viale Regina Elena, 291

country: Italy

institute: “La Sapienza” University of Rome

summary: Purpose: The intergration of genetic and chemical screens identified SETD8 as a new druggable target in neuroblastoma tumor. The goal of this study is to evaluate the transcriptome profiling (RNA-seq) of Neuroblastoma cell lines after genetic and pharmacological inhibition of SETD8. Methods: mRNA profiles of NB cells after genetic and pharmacological inhibition of SETD8 were generated by deep sequencing in duplicate with Ilumina HiSeq2500 using Illumina TruSeq V4. The sequence reads were analyzed with software Trimmomatic, STAR and edgeR to determine the differetially expressed genes. qRT–PCR validation was performed using SYBR Green assays. Results:  About 60 million sequence reads per sample were mapped to the human genome (hg19).  Approximately 10% of the transcripts showed differential expression between the control and the treated samples, with a fold change ≥1.5 and p value <0.05. Altered expression of 12 genes was confirmed with qRT–PCR, demonstrating the high degree of sensitivity of the RNA-seq method. Hierarchical clustering of differentially expressed genes uncovered several as yet uncharacterized genes that may contribute to SETD8 function.  Conclusions: Our study identifies SETD8 as a new therapeutic target in Neuroblastoma tumor. RNA-seq transcriptome analyses and functional studies revealed that SETD8 ablation rescued the proapoptotic and cell-cycle arrest functions of p53 through reactivation of the p53 canonical pathway by decreasing p53k382me1.

overall_design: mRNA profiles of Neuroblastoma cells after genetic and pharmacological inhibition of SETD8 were generated by deep sequencing in duplicate with Ilumina HiSeq2500 using Illumina TruSeq V4.

first: Carol

last: Thiele

first: Chunhua

last: Yan

first: Ying

last: Hu


title: NF-κB-dependent redistribution of the GR cistrome recruits GR to inflammatory gene loci and correlates with reduced repression by glucocorticoid [RNAP2_ChIPseq]

overall_design: To understand the impact of NF-κB on GR cistrome redistribution, A549 cells were infected with Ad5-GFP (control), or Ad-IκBαΔN (NF-κB inhibitor) before no treatment, or treatment with IL1B, budesonide, or both for 1h. ChIP was performed using RNAP2 antibody, and sequenced using Illumina NovaSeq 6000.

first: Amandah

last: Necker-Brown


title: Ex vivo–expanded human regulatory T cells enhance cholesterol efflux and PON1 expression in oxLDL-exposed macrophages

city: London

first: Cristiano

last: Scotta

address: Kingston Lane

department: Life Science

country: United Kingdom

institute: Brunel University

summary:
Lipid-driven chronic inflammation is a primary driver of atherosclerosis progression. Enhancing the anti-inflammatory response by bolstering regulatory T cell (Treg) numbers or improving their function, holds promise for therapeutic control of the disease. We and others have previously demonstrated in clinical trials the safety and potential efficacy of adoptive Treg transfer in restoring immunological balance. This study investigates the capability of ex vivo expanded regulatory T cells to counterbalance pro-inflammatory signals and lipid accumulation in primary macrophages, thereby contrasting plaque formation and atherosclerosis progression.
To test this hypothesis, freshly isolated Tregs were ex vivo expanded into a clinical grade cell product before co-culturing with monocyte-derived M2-like macrophages. Macrophages were exposed to oxidised low-density lipoprotein (oxLDL) and co-cultured alone or in the presence of Tregs. Changes in macrophage phenotype, gene expression profile and functional properties such as the capacity to regulate the accumulation of cholesterol were analysed by flow cytometry, RNA sequencing and western blotting.
In the co-culture system, the study demonstrated a significant reduction in both pro-inflammatory effects of oxLDL and accumulation of modified cholesterol in macrophages when Tregs were present. We identified that Tregs through the transferring of cAMP into macrophages, could potentiate the ABCA1-mediated cholesterol efflux pathway, and crucially, elevate the anti-atherogenic activity by inducing Paraoxonase-1 expression.
The findings suggest a novel atheroprotective role for Tregs which supports the potential development of new experimental therapies for cardiovascular disease. By influencing macrophage function and cholesterol accumulation, Tregs present a promising avenue for mitigating atherosclerosis.

overall_design: To investigate the effect of regulatory T cell co-culture on the genetic profile of monocyte-derived M2-like macrophages.

first: Caraugh

last: Albany

first: Daniela

last: Mastronicola

first: Wladislaw

last: Stroukov

first: Rocio

last: Martinez-Nunez

last: Scottà


title: Stability and Progressive Differentiation of Tfr cells are intrinsically and extrinsically controlled by Tfh programs

status: Public on Oct 30 2025

city: BOSTON

first: Jeong-Mi

last: Lee

state: MASSACHUSETTS

address: 221 Longwood Avenue, EBRC 3rd Floor

country: USA

institute: Brigham and Women's Hospital

summary: Follicular regulatory T (Tfr) cells restrain follicular helper T (Tfh) cell-mediated B cell responses in the germinal center (GC) reaction to optimize humoral immunity while limiting autoimmunity.  The immune system partially regulates GC responses by controlling the stepwise differentiation of Tfh cells.  Whether Tfr cell development requires sequential developmental stages and how the immune system regulates these processes to control humoral immunity is poorly understood.   Here, we used longitudinal sampling of lymphoid organs along with fate mapping and matched single-cell RNASeq/TCRseq to assess developmental dynamics of Tfr cells. We found that Tfr cells undergo dynamic clonal expansion. During this process, Tfr cells also undergo progressive differentiation through progenitor-like, early effector, and late effector stages.  Late effector Tfr cells possess inherent instability, with a propensity to lose expression of the transcription factor FoxP3 to become ExTfr cells. ExTfr have unique features and can be redeemed to become suppressive Tfr cells.  Acquisition of a Tfh-like transcriptional program in Tfr cells was an intrinsic predictor of progeny instability.  Extrinsically, Tfh cells enhanced late effector Tfr cell differentiation by diverting early effector Tfr cells away from a default Blimp1-expressing pathway to a Bcl6-expressing one.  Moreover, Tfh induce Tcf7 in Tfr cells which promotes late effector Tfr differentiation. Together, these data indicate that Tfr cells are a dynamic and plastic cell subset, the progressive differentiation of which is controlled at later effector stages by intrinsic and extrinsic programs that work together to provide a negative feedback loop to control humoral immunity.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: T follicular regulatory cells from seconday lymphoid tissue

first: Peter

middle: T

last: Sage

first: Paulo

last: Raeder


title: Gene expression changes in postnatal mouse brain Neural Stem Cells differentiated in the presence of platelets.

city: Patras

first: Adamantia

last: Kouvela

address: Asklipiou 1

country: Greece

institute: Department of Biochemistry, School of Medicine, University of Patras

summary: Whole transcriptome sequencing of postnatal mouse brain Neural Stem Cells (NSCs) cultured for three days in serum-free media, in the presence (proliferation conditions) or absence (differentiation conditions) of growth factors EFG and FGF2, as well as when co-cultured with mouse platelets (in a ratio of 75 platelets per NSC) in the absence of growth factors.

overall_design: Expression profiling by the Ion Torrent Genestudio S5 platform with Ion Chef (Ion torrent)

first: Maria

last: Anesti

first: Aggeliki

last: Dimopoulou

first: Dimitrios

last: Dimitrakopoulos

first: Constantinos

last: Stathopoulos

first: Ilias

last: Kazanis


title: NF-κB-dependent redistribution of the GR cistrome recruits GR to inflammatory gene loci and correlates with reduced repression by glucocorticoid

first: Alex

last: Gao

summary: While ligand-activated glucocorticoid receptor (GR) binds DNA to activate transcription, glucocorticoids, including budesonide, reduce inflammatory gene expression, yet recruit GR to many such gene loci. In epithelial cells, the inflammatory cytokine, interleukin-1β (IL1B), activates NF-κB to induce gene expression and co-treatment with budesonide produces nanoscale GR-RELA nuclear co-localization. Such co-stimulation orchestrated reciprocal genome-wide redistribution of GR and RELA binding regions (GBRs and RBRs, respectively) relative to each mono-treatment to produce widespread GBR_x0002_RBR overlap. This correlated with increased RNA polymerase-2 presence and required NF-κB for GR cistrome remodeling. Mapping transcription start sites to the nearest GBR or RBR both revealed associations with upregulated, but not repressed, genes. Importantly, RBR proximity to budesonide_x0002_upregulated genes and GBR proximity to IL1B-upregulated genes correlated with attenuated repression on co-treatment. As this occurred on a background of glucocorticoid-induced repression, GR presence at specific IL1B-induced gene loci may reduce, or protect, from an otherwise prevalent glucocorticoid_x0002_induced repression.

overall_design: To understand transcriptional interactions between GR and NF-κB, mammalian airway epithelial (A549) cells were not stimulated or treated with IL1B, budesonide, or both for 1, 2, 6, 12, and 24h before harvesting for RNA sequencing.

first: Mahmoud

last: Mostafa

first: Robert

last: Newton


title: Cooperativity between head-to-head dimerization and head-to-tail multimerization of FoxP3 transcription factor

city: Boston

first: Sun

last: Hur

state: MA

address: 3 Blackfan St

department: Howard Hughes Medical Institute and Program in Cellular and Molecular Medicine

institute: Boston Children's Hospital

summary: FoxP3, an essential transcription factor for regulatory T cells (Tregs), forms higher-order multimers on TnG microsatellites in the genome. Although a previous study has shown that FoxP3 can form head-to-head (H-H) dimers on inverted repeats of forkhead motifs (IR-FKHM), in vivo evidence for this configuration was elusive. An unbiased FoxP3 pull-down sequencing approach identified a wide range of sequences supporting H-H dimerization, enabling systematic genomic analysis. This work revealed that FoxP3 binds DNA in Tregs as both H-H dimers and higher-order multimers, often incorporating one half-site of the H-H dimer into TnG repeats. A robust H-H interaction can seed multimerization on adjacent TnG repeats, enhancing assembly even on suboptimal sequences. Unlike other FoxP transcription factors, H-H dimerization is unique to FoxP3. Chimeric construct analyses identified a loop adjacent to the forkhead domain as the determinant for this distinctive function, underscoring a unique synergism between two binding modes.

overall_design:
In vitro pulldown assay using a synthetic dsDNA oligonucleotide library to identify FoxP3-binding motifs. MBP-tagged FoxP3 and FoxP1 were used to pull down DNA sequences with potential binding affinity. The input library was included as a control. Pulldown-enriched DNA was PCR-amplified and sequenced using paired-end 150 bp reads on an Illumina NovaSeq 6000 platform.
Follow-up in vitro pulldown experiment using the same synthetic dsDNA library, focused on comparative binding between FoxP3 and FoxP1. Samples were sequenced using single-end 130-cycle reads on an Illumina NextSeq 2000 platform.

first: Fangwei

last: Leng

first: Ryan

last: Clark

first: Wenxiang

last: Zhang

first: Thibault

last: Viennet

first: Haribabu

last: Arthanari

first: Xi

last: Wang


title: Automated chromatin profiling with spa-ChIP-seq uncovers the impacts of condition variations

city: La Jolla

first: Alon

last: Goren

state: CA

address: 9375 Gilman Dr

department: Department of Medicine

institute: University of California San Diego

summary: Chromatin immunoprecipitation followed by sequencing (ChIP-seq) is widely used to study the genomic localization of DNA-associated proteins. However, conventional protocols include multiple manual steps that can introduce inconsistency and limit scalability, thereby restricting the inclusion of appropriate replicates and controls. Although the introduction of liquid handling platforms has improved reproducibility, most existing efforts have automated only a subset of the workflow, and extending automation to efficiently map non-histone proteins, such as chromatin regulators, remains challenging. Here, we present a fully automated implementation of our previously developed single-pot ChIP-seq protocol, named spa-ChIP-seq, which enables scalable processing of 8 to 96 ChIP-seq samples from crosslinked cells to sequencing-ready library in approximately three days with an estimated cost of $70 per sample. Benchmarking spa-ChIP-seq against manual ChIP-seq performed in parallel demonstrates comparable signal-to-noise ratio between the two workflows. Using spa-ChIP-seq, we systematically evaluate multiple parameters including shearing and crosslinking conditions, buffer compositions, and the ratio of antibody to cell-number. We find, for the first time to our knowledge, that weaker genomic localization signals are sensitive to changing the antibody to cell-number ratio, whereas the stronger signals remain unaffected. This finding underscores the importance of maintaining consistent antibody-to-cell-number ratio for comparative studies, such as treatment responses or chromatin-QTL mapping. The spa-ChIP-seq protocol is publicly available, including deck setups, operational parameters, and scripts. We envision that this robust, cost-efficient protocol will facilitate high-throughput, reproducible ChIP-seq analyses, supporting large-scale studies of antibody validation, compound screening, population genomics, and diagnostic frameworks.

overall_design: ChIP-seq of H3K27ac and H3K27me3 comparing crosslinking conditions (DSG+FA and FA only), lysis buffers for sonications (LB3 and EBB) while evaluating the automated workflow versus the manual workflow. We also evaluated the ratios of antibody to chromatin by performing titration ChIP-seq experiments with H3K9ac, BRD4, HA-tag, and P300 antibodies.

first: Yuwei

last: Cao

first: Lauren

last: Patel

last: Alcoser

first: Eric

last: Mendenhall

first: Christopher

last: Benner

first: Sven

last: Heinz


title: Transcriptomic Profiling of Bronchial Epithelium Reveals Dysregulated Interferon and Inflammatory Responses to Rhinovirus in Exacerbation-Prone Pediatric Asthma [scRNA-Seq]

city: Seattle,

first: Stephanie

last: Osmond

state: WA

address: 1201 9th Ave

institute: Benaroya Research Institute

summary: Host factors influencing susceptibility to rhinovirus-induced asthma exacerbations remain poorly characterized. Using organotypic bronchial epithelial cultures from well-characterized children with asthma and healthy children, this study investigated viral load kinetics and resultant host responses by bulk and single-cell transcriptomics and targeted protein analyses. Bronchial epithelium from exacerbation-prone children exhibited greater rhinovirus replication and a cascade of exaggerated downstream interferon (IFN), inflammatory, epithelial stress, and remodeling responses. These transcriptional patterns were confirmed and further refined using single-cell transcriptomics, revealing cell type-specific contributions—particularly from non-ciliated cell populations including secretory immune response, tuft, and basal cells. We observed that these post-infection differences were associated with lower pre-infection IFN-stimulated gene (ISG) expression and protein levels of the ISG CXCL10. Prophylactic IFN-β treatment reduced viral replication and normalized downstream responses, supporting low baseline (pre-infection) IFN tone as a modifiable causal determinant of host susceptibility to adverse rhinovirus-induced responses in exacerbation-prone children with asthma.

overall_design: We compared transcriptional responses across 3 donor groups, severe asthma (SE), non-Severe asthma (NSE), and healthy controls at baseline (uninfected) and post RV infection sampes.

first: Naresh

last: Doni Jayavelu

first: Jason

last: S. Debley

first: Matthew

middle: C

last: Altman


title: Transcriptomic Profiling of Bronchial Epithelium Reveals Dysregulated Interferon and Inflammatory Responses to Rhinovirus in Exacerbation-Prone Pediatric Asthma [RNA-Seq]

overall_design: Bronchial epithelial cells (BECs) from children with asthma (n=37) and healthy children (n=3) ages 6-18 years of age were obtained from subjects while under general anesthesia for an elective surgery using 4-mm Harrell unsheathed bronchoscope cytology brushes (CONMED®) inserted through an endotracheal tube as we have previously described (8, 42). RNA was collected from BEC cultures pre-infection and at 2d, 4d, 7d, and 10d post RV infection.

first: Basilin

last: Benson

last: C. Altman


title: The transcription factor NFIL3 drives innate lymphoid cell specification from lymphoid progenitors [RNA-seq - Tox]

city: Nantes

first: Christelle

last: Harly

address: 8 quai Moncousu

country: France

institute: CNRS

summary: Helper innate lymphoid cells (ILC) play important functions in immunity and tissue homeostasis, but their development remains poorly understood. In this study, we identified NFIL3 as the earliest known requirement during ILC development. Forced expression of NFIL3 in mouse lymphoid progenitors in vitro triggered the staged expression of downstream transcription factors including Tcf7, Gata3, Zbtb16 and Id2 that are implicated in ILC development. Thus, NFIL3 resides at the apex of a hierarchy of transcription factors important for innate lymphocyte development, and is necessary and sufficient to impose the innate lymphoid cell fate.

overall_design: Bulk RNA-seq comparing ILC precursors generated by forcibly expressing TOX or control GFP in lymphoid precursors

first: Joseph

last: Léger

first: Eva

last: Artano

first: Delphine

last: Coulais

first: Naomie

last: Belletoise

first: Fadhloun

last: Raoudha

first: Devin

last: Kenney

first: Avinash

last: Bhandoola


title: The transcription factor NFIL3 drives innate lymphoid cell specification from lymphoid progenitors [CUT&RUN]

overall_design: Cleavage Under Targets and Release Using Nuclease (CUT&RUN) in lymphoid precursors transduced to express NFIL3-HA for specific binding, or NFIL3 for control background with the anti-HA antibody


title: Transcriptomic Profiling of Bronchial Epithelium Reveals Dysregulated Interferon and Inflammatory Responses to Rhinovirus in Exacerbation-Prone Pediatric Asthma

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

overall_design: Refer to individual Series


title: The transcription factor NFIL3 drives innate lymphoid cell specification from lymphoid progenitors [RNA-seq - ex vivo, Nfil3]

overall_design: Bulk RNA-seq comparing ILC precursors generated by forcibly expressing NFIL3 in lymphoid precursors, and ex vivo isolated ILC precursors


title: The transcription factor NFIL3 drives innate lymphoid cell specification from lymphoid progenitors [scRNA-seq]

overall_design: scRNA-seq of ILC precursors at different developmental stages


title: Comparative transcriptomics as a key to understanding the adaptation mechanisms of Baikal sculpins to the deep-water habitat

city: Irkutsk

first: Tuyana

last: Sidorova

address: 3, Ulan-Batorskaya

department: Siberian Branch of the Russian Academy of Sciences

country: Russia

institute: Limnological Institute

summary: In this study, the transcriptional profiles of four deep-sea sculpin fish species (Cottoidei) are described for the first time as part of the study of the adaptation of endemic fishes to different habitats in Lake Baikal. These studies will lead to a deeper understanding of the molecular mechanisms that ensure the adaptation of endemic species to specific habitats in Lake Baikal, even at depths of more than 500 m (up to 1600 m).

sample_organism:
Batrachocottus multiradiatus
Comephorus dybowskii
Batrachocottus nikolskii
Comephorus baikalensis

overall_design: To study the effect of habitat depth on the transcriptional profile, a transcriptome analysis was performed for 4 species: m3 - Comephorus dybowski Korotneff, 1905; m4 - Comephorus baicalensis (Pallas, 1776); m6 - Batrachocottus nikolskii (Berg, 1900); m7 - Batrachocottus multiradiatus Berg, 1907. Fish species were collected during trawl surveys on the R/V Vereshchagin in August 2023 and 2024. The work was supported by State project No. 0279-2021-0005 (121032300224-8).

first: Yulia

middle: P

last: Sapozhnikova

middle: V

first: Anastasiya

middle: G

last: Koroleva

first: Evgenia

middle: A

last: Vakhteeva

first: Alexander

last: Epifantsev

first: Vera

last: Yakhnenko

first: Sergey

last: Potapov

first: Tatyana

last: Butina

last: Kirilchik

first: Lyubov

last: Sukhanova

first: Ivan

last: Nebesnykh

first: Igor

last: Khanaev


title: The transcription factor NFIL3 drives innate lymphoid cell specification from lymphoid progenitors [DNase-seq]

first: Christelle

last: Harly

overall_design: DNase-seq analysis of ILC precursors at different developmental stages

first: Joseph

last: Léger

first: Eva

last: Artano

last: Coulais


title: The transcription factor NFIL3 drives innate lymphoid cell specification from lymphoid progenitors

first: Delphine

first: Naomie

last: Belletoise

first: Raoudha

last: Fadhloun

last: Kenney

last: Bhandoola


title: Podocyte Twin in Culture: Patient-specific ex vivo Model of Genetic FSGS caused by an INF2 Mutation

city: Aachen

first: René

last: Krüger

address: Pauwelsstraße 20

department: Institute for Stem Cell Biology

country: Germany

institute: RWTH Aachen University Medical School

summary: We used an ex vivo cell culture model to generate patient-specific podocytes from a patient with genetic FSGS caused by an INF2 mutation. The patient's entire genetic background was maintained in all cell types of the protocol. Bulk RNA sequencing analysis was performed to first validate the outcome of the protocol by comparing the hiPSC-derived podocytes generated from a healthy donor with a conditionally immortalized podocyte cell line. Subsequently, the patient-specific hiPSC-derived podocytes holding the INF2 mutation were compared to the healthy hiPSC-derived podocytes and mutation specific phenotypic and functional alterations were investigated.

overall_design:
This study consists of bulk RNA sequencing samples from a conditionally immortalized podocyte cell line (M. Saleem) and from different steps of the differentiation protocol for the generation of podocytes derived from human induced pluripotent stem cells (hiPSC) in biological duplicates (n=2). The different steps include hiPSCs before differentiation, nephron progenitor cells and terminal differentiated hiPSC-derived podocytes generated from a healthy donor and a patient with genetic FSGS caused by an INF2 mutation.
***Submitter states that raw data are not available due to patient privacy concerns***

first: Janina

last: Müller-Deile

first: Victoria

last: Rose


title: Bile acids engage the SIPR-STAT3 signaling axis to modulate regulatory T cell responses in fibrosing cholangiopathies

city: Cincinnati

last: Miethke

state: OH

address: 3333 Burnet Ave, MLC 2010

institute: Cincinnati Childrens Hospital Medical Ce

summary:
Background & aims: Regulatory T cells (Tregs), a subset of CD4 lymphocytes, protect against inflammatory tissue injury. However, it is currently unknown how retention of bile acids (BA) in fibrosing cholangiopathies like biliary atresia or PSC shape hepatic Treg responses.

Methods: To induce sclerosing cholangitis (SC), mice were fed a diet containing 0.1% 3,5-diethoxycarbonyl-1,4-dihydrocollidine for 14 days, followed by 28 days on regular chow to assess tissue repair. Serial hepatic cell and nuclear preparations were subjected to single-cell RNA sequencing and ATAC-seq to define gene regulatory networks controlling Tregs under cholestatic conditions. Candidate molecules mediating the effects of tauro- and chenodeoxycholic acid (T/CDCA) on Tregs were validated in vitro, across three murine models of SC, and in liver tissue samples from 130 infants with biliary atresia.

Results: Single-cell analyses revealed that Tregs acquired a Th17-like transcriptional program during cholestasis and upregulated amphiregulin (Areg) during the repair phase. S1P receptors were identified as mediators of T/CDCA effects on Tregs, and this was validated both in vitro and in the Abcb4-/- model of SC. Deletion of Stat3 in CD4+ cells enhanced hepatic Treg responses following bile duct ligation. Pharmacologic reduction of hepatic BA concentrations using an IBAT (ileal bile acid transporter) inhibitor increased hepatic Treg numbers and attenuated liver injury and fibrosis in Abcb4-/- mice. These protective effects were lost upon Treg depletion or AREG neutralization. Finally, in infants with biliary atresia, a liver transcriptional profile consistent with Treg activation and AREG upregulation at diagnosis was associated with improved 2-year native liver survival.

Conclusion: Bile acids suppress Treg regulatory function by promoting a Th17-like phenotype, thereby limiting their capacity to mitigate immune-mediated cholangiocyte injury. Restoring Treg function and amphiregulin expression may represent a novel therapeutic strategy in fibrosing cholangiopathies.

overall_design:
CD25+CD127-/lowCD4+CD3+CD56-CD45+eflour506-cells FACS sorted from peripheral blood of healthy donors. FACs sorted Tregs were cultured with  α-CD3, α-CD28 (Beads 1:1ratio), hIL-2(100U), with either H2O(Vehicle) or with TCDCA (100 μM), 48hrs.
contributor: Zhang, Xiang and his Laboratory for Statistical Genomics and Systems Biology at the University of Cincinnati.

first: Ramesh

last: Kudira

first: Zhang

last: Xiang


title: IsomiR Utility in ALS Prognostication (ALS)

city: ראשון לציון

first: Iddo

last: Magen

state: Not in US

address: בן אליעזר 32, 17

country: Israel

institute: Weizmann Institute

type: Non-coding RNA profiling by high throughput sequencing

summary: Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by progressive motor neuron loss. IsomiRs are microRNA (miRNA) isoforms that arise from alternative processing or editing events during miRNA biogenesis. While isomiRs may carry distinct biological and clinical relevance, their potential as cell-free biomarkers in neurodegeneration remains largely unexplored. Here we investigated the prognostic utility of plasma isomiRs in ALS, using next-generation sequencing. We profiled cell-free isomiRs in 154 ALS patients from a British cohort and identified higher levels of one isomiR, let-7g-5p.t, to be associated with longer survival. This finding was independently validated in an international ALS cohort of 200 patients and was in two orthogonal approaches. let-7g-5p.t prognostic utility was comparable to that of neurofilament light chain (NfL) or miR-181. These results establish isomiRs as a novel class of blood-based biomarkers in ALS with potential to refine prognostication in clinical trials for neurodegenerative diseases.

overall_design: The molecular experimenters were blinded to the identity of the samples. Total RNA was extracted from 500μl plasma using miRNeasy Micro Kit following the manufacturer's protocol except adding 2.5ml of Qiazol to each plasma sample (Qiagen, #217084). RNA was quantified with a Qubit fluorometer using the RNA Broad Range Assay Kit (Thermo Fisher Scientific, #Q10211). Small RNA next-generation sequencing libraries were prepared using the QIAseq miRNA Library Kit (Qiagen, #331502) from 7.5 ng of total RNA. If RNA concentration was below the limit of detection, 5μl of undiluted RNA were used. Precise linear quantification of miRNAs was achieved by 12-nucleotide long unique molecular identifiers (UMIs). Samples were randomly assigned to the preparation of libraries indexed with QIAseq miRNA NGS 48 Index IL (Qiagen, #331595) and sequenced in batches of 48 or fewer samples. Library concentration was determined with a Qubit fluorometer (dsDNA High Sensitivity Assay Kit, Thermo Fisher Scientific, #Q32854) and library size with TapeStation D1000 (Agilent). Libraries with different indices were multiplexed and sequenced on a NextSeq 500/550 v2 (Illumina, #20024906) or a NovaSeq 6000 flow cell (Illumina, #20028401), with a 75-bp single read or paired-end reads with 6-bp index. FASTQ files were de-multiplexed using the user-friendly transcriptome analysis pipeline. Human miRNAs and isomiRs, as defined by miRBase - V22, were mapped using CLC genomic workbench 22.0 (Qiagen) only to the positive strand. Reads were trimmed in two consecutive fashions; Phred score, Q based error was calculated for every nucleotide, high values reflect poor read quality. A trimming score defined by the subtraction of the error from a threshold of 0.05 was set for each nucleotide. Then, a running sum of the nucleotide errors was calculated per read. The region between the first positive value of the running sum and the highest value of the running sum was kept with non-positive score reads being removed completely. Consecutively, reads’ ends were trimmed to keep the maximum length region containing 2 or fewer ambiguous nucleotides. This was followed by the library’s 3’-adaptors (TGGAATTCTCGGGTGCCAAGG) trimming, with reads that did not contain any adaptors being excluded from downstream analyses. Reads were filtered based on lengths of 15-55 bps. IsomiRs were considered as 2bps changes (insertion, deletion, shift, and mismatch) up/downstream from miRBase reference sequence with a maximum of 2 mismatches allowed in alignment. The algorithm prioritizes perfect matches to reference miRNAs, followed by sequences of increasing mismatch nucleotides. When a sequence aligns equally well to multiple isomiRs, counts are assigned uniformly across all matching forms.

first: Yahel

last: Cohen

first: Ilan

last: Sinai

first: Yehuda

last: Danino

first: Joanne

last: Wuu

first: Andrea

last: Malaspina

first: Michael

last: Benatar

first: Eran

last: Hornstein


title: Comparative Single Cell RNA Sequencing of Notch2 and RBPJ-deficient mononuclear phagocyte cells and myeloid progenitors from the spleen and bone marrow

city: Hannover

first: Jaba

last: Gamrekelashvili

address: Carl-Neuberg-Str. 1

department: Nephrology

institute: MHH

summary: Mononuclear phagocytes and their myeloid progenitors are critical in various homeostatic processes as well as immune responses. We employed single-cell RNA sequencing to investigate myeloid progenitors and mononuclear phagocytic cell subsets, including monocytes, splenic red pulp macrophages and dendritic cells in the spleen and in the bone marrow, with a focus on how Notch2 and its nuclear mediator Rbpj regulate these subsets. Our study offers a comparative analysis of mononuclear phagocytes and their progenitors examining cells isolated from the spleens or bone marrow of myeloid Notch2 (N2ΔCx3cr1)-, and Rbpj (RbpjΔCx3cr1)-deficient mice or control animals. Our findings may provide further insights into the heterogeneity of the mononuclear phagocytic cells in the spleen and bone marrow and show the critical role of Notch signaling in their development.

overall_design:
Mononuclear phagocytic cells and their myeloid progenitors from Ctrl, Notch2- and Rbpj deficient mouse spleens or bone marrow (two mice of each genotype) were labeled with different HTOs, pooled as spleen or bone marrow, and isolated using fluorescence-activated cell (FACS)-sorting.

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

first: Florian P

last: Limbourg


title: Transcriptional dysregulation in the spinal cord of SCA3 provides insights into disease mechanisms

status: Public on Oct 29 2025

city: Ann Arbor

first: Hayley

middle: S

last: McLoughlin

state: MI

address: 109 Zina Pitcher Pl

department: Neurology, Human Genetics

institute: University of Michigan

summary: Spinocerebellar ataxia type 3 (SCA3) is a neurodegenerative disease caused by polyglutamine repeat expansion in the ATXN3 gene. Despite the ubiquitous expression of ATXN3 throughout the body, SCA3 pathology is pronounced in select, vulnerable regions within the central nervous system (CNS). Notably, imaging studies of SCA3 patients have revealed spinal cord atrophy prior to ataxia symptom onset and progressing with disease severity. However, the molecular mechanisms underlying SCA3 pathology in the spinal cord remain largely unexplored. Here, we present, for the first time, a comprehensive analysis of the spinal cord transcriptome using SCA3 tissue from humans and mouse models. Our data demonstrates both early and progressive transcriptional dysregulation in the spinal cord, impacting key biological processes such as lipid metabolism, inflammation, cellular structure, and nucleic acid processing. Transcriptomic profiling of Atxn3 knockout mice revealed only weak transcriptional changes in the spinal cord, with minimal overlap compared to the robust alterations observed in SCA3 knock-in mice, indicating that the molecular signature of the disease in the spinal cord is likely not driven by ATXN3 loss of function. In addition, we identified aberrant RNA splicing, particularly affecting genes involved in cytoskeletal organization. Collectively, our findings indicate that SCA3-associated transcriptional dysregulation in the spinal cord contributes to canonical pathological mechanisms of SCA3 both early and progressively. These results underscore the central role of the spinal cord in SCA3 pathogenesis and advocate for its inclusion as a key therapeutic target.

overall_design: RNAseq of RNA collected from the spinal cord of SCA3 KIQ300 and WT mice at 24 and 56 weeks of age.

first: Jacen

last: Emerson

first: Brianna

last: Nelthrope

first: Emma

last: Walker

first: Grace

last: Mao

first: Hannah

last: Shorrock


title: The transcription factor NFIL3 drives innate lymphoid cell specification from lymphoid progenitors [RNA-seq - Il7rLT]

first: Christelle

last: Harly

overall_design: Bulk RNA-seq comparing ex vivo early ILC intermediate progenitors using different sorting strategies

first: Joseph

last: Léger

first: Eva

last: Artano

first: Delphine

last: Coulais

first: Naomie

last: Belletoise

first: Fadhloun

last: Raoudha

first: Devin

last: Kenney

first: Avinash

last: Bhandoola


title: Leukocyte-intrinsic ER stress responses contribute to chemotherapy-induced peripheral neuropathy.

city: Winston Salem

first: Ming

last: Leung

state: NC

address: 1 Medical Center Blvd

department: Comprehensive Cancer Center

institute: Atrium Health Wake Forest

summary: Chemotherapy-induced peripheral neuropathy (CIPN) is the most prevalent and limiting side effect of paclitaxel treatment in cancer patients. CIPN affects sensory neurons through neuroinflammatory mechanisms, but how immune cells sense and interpret systemic paclitaxel exposure during treatment is unclear. Here, we report that paclitaxel administration triggers the endoplasmic reticulum (ER) stress sensor inositol-requiring enzyme 1α (IRE1α) in circulating and dorsal root ganglia-resident myeloid cells, engendering an inflammatory milieu that promotes CIPN. Mechanistically, paclitaxel induced overproduction of mitochondrial-derived reactive oxygen species (ROS) that provoked ER stress and IRE1α hyperactivation in macrophages. This process reprogrammed macrophages towards a highly inflammatory state characterized by IRE1α-dependent production of TNF-α, IL-1β, PGE2, IL-6, IL-5, GM-CSF, MCP-1, and MIP-2. Ablation of IRE1α in leukocytes, or treatment with a selective IRE1α pharmacological inhibitor, prevented dorsal root ganglia neuroinflammation and CIPN-related pain behaviors in mice. Furthermore, the development and severity of CIPN in patients with gynecological cancer was associated with the status of IRE1α activation in their circulating leukocytes. Our study uncovers leukocyte-intrinsic IRE1α as a key mediator of CIPN and suggests that targeting its dysregulated activation could help mitigate CIPN in cancer patients receiving paclitaxel.

overall_design: To investigate whether paclitaxel might impact the global transcriptomic profile of myeloid cells via IRE1a activation, we performed gene profiling analysis using data obtained from RNA-seq of wild-type (WT) versus IRE1a-deficient (cKO) bone marrow differentiated macrophages exposed to paclitaxel (10 µM, n=4) or vehicle control (Dimethyl sulfoxide; DMSO, n=4) treated for 18 hours.

first: Miriam

last: Fonseca

first: Oriana

last: Gelblung

first: Sarah

middle: D

last: Pennypacker

first: Taylor

last: Brooks

first: Michael

last: Limia

first: James

middle: W

last: Morgan

first: Xuewei

last: Zhu

first: Luis

last: Tovias-Sanchez

first: Alejandro

last: Pluma-Pluma

first: Ruth

last: Elena Martinez

first: Mathew

last: Eber

first: Sun

last: Park

first: Cristina

last: Furdui

first: Deepika

last: Awasthi

first: Alexander

last: Emmanuelli

first: Byuri

last: Cho

first: Chen

last: Tan

first: David

middle: I

last: Shalowitz

first: Samuel

last: Lentz

last: Kelly

first: Anderson

middle: O

last: Cox

first: Lindsay

last: Macnamara

first: Fang-Chi

last: Hsu

first: Yusuke

last: Shiozawa

first: Wesley

first: Takao

last: Iwawaki

first: Lance

last: Miller

first: Glenn

last: Lesser

first: Roy

last: Strowd

first: Juan

middle: R

last: Cubillos-Ruiz

first: Edgar

last: Romero-Sandoval


title: Transcriptional dysregulation in the spinal cord of SCA3 provides insights into disease mechanisms [KO_RNAseq]

first: Hayley

last: McLoughlin

overall_design: RNAseq of RNA collected from the spinal cord of Atxn3 KO and WT mice at 24 weeks of age.

first: Jacen

last: Emerson

first: Brianna

last: Nelthrope

first: Emma

last: Walker

first: Grace

last: Mao

first: Hannah

last: Shorrock


title: Mitochondrial RNA cytosolic leakage drives the SASP [CRISPR/Cas9]

city: Rochester

first: Dominik

last: Saul

state: Minnesota

address: 200 First Street SW Rochester

institute: Mayo Clinic

summary: Senescent cells secrete proinflammatory factors known as the senescence-associated secretory phenotype (SASP), contributing to tissue dysfunction and aging. Mitochondrial dysfunction is a key feature of senescence, influencing SASP via mitochondrial DNA (mtDNA) release and cGAS/STING pathway activation. Here, we demonstrate that mitochondrial RNA (mtRNA) also accumulates in the cytosol of senescent cells, activating RNA sensors RIG-I and MDA5, leading to MAVS aggregation and SASP induction. Inhibition of these RNA sensors significantly reduces SASP factors. Furthermore, BAX and BAK play a key role in mtRNA leakage during senescence, and their deletion diminishes SASP expression in vitro and in a mouse model of Metabolic Dysfunction Associated Steatohepatitis (MASH). These findings highlight mtRNA's role in SASP regulation and its potential as a therapeutic target for mitigating age-related inflammation.

overall_design: RNA-seq profiling of proliferating and senescent fibroblasts lacking proteins associated with cytosolic RNA signaling, including DDX58 (RIG-I), IFIH1 (MDA5) and MAVS. Moreover, RNA-seq was also performed in fibroblasts lacking mitochondria, which were subsequently transfected with isolated mitochondrial RNA (mtRNA). Finally, RNA-seq analysis was performed in livers from liver-specific Bax Bak KO animals with diet-induced metabolic dysfunction-associated steatohepatitis (MASH).

first: Victorelli

last: Stella

first: Han

last: Yeaeun

first: Saul

last: Dominik

first: Passos

last: Joao


title: ATACSeq of TET2-mutant human HSCs

first: Hector

last: Huerga Encabo

address: The Francis Crick Institute, 1 Midland Rd

institute: The Francis Crick Institute

summary: The expansion of hematopoietic stem cell clones and their immune progeny carrying somatic mutations is defined as clonal hematopoiesis (CH) and its prevalence increases with age. Individuals with CH are at higher risk of developing hematological malignancies as well as other pathologies, such as cardiovascular or inflammatory pulmonary diseases. Understanding the environmental conditions that favor mutant clones and the CH-immune system response to such environments is key to designing therapeutic strategies to stall the expansion of mutant clones and the development of CH-associated pathologies. In this work, we characterized how mutations in TET2, increase the competitive advantage of human HSCs upon environmental stress conditions and simultaneously shape the immune response of their myeloid progeny. Employing a humanized mouse model, we unravel a cell-specific impact of TET2 mutations on HSCs and their myeloid progeny to inflammatory stress. In such environments, TET2Mut HSCs have a mitigated transcriptional upregulation of inflammatory pathways, and by contrast, TET2Mut myeloid cells have an exacerbated inflammatory response. A distinct monocyte-macrophage trajectory derived from TET2Mut HSCs contributes to pathological inflammation. These findings provide evidence at the human cell level and reconcile the notion that TET2-derived CH is associated with both increased stemness in the HSC compartment and inflammation elicited by the myeloid progeny.

overall_design: Using CRISPR-edited human HSCs and transplantation in immunodeficient mice, we have sorted CD34+CD38-CD45RA- cells and analyzed the chromatin conformation of TET2 mutant (TET2Mut) HSCs.

first: Huerga Encabo

last: Hector

first: Garcia-Albornoz

last: Manuel

first: Llorian

last: Miriam

first: Bonnet

last: Dominique


title: Transcriptional dysregulation in the spinal cord of SCA3 provides insights into disease mechanisms [Human_RNAseq]

overall_design: RNAseq of RNA collected from the spinal cord of Human SCA3 and age-matched control samples.


title: Dynamic neutrophil-keratinocyte communication network centered on IL-36/TNFSF15 responses characterizes inflammatory responses in generalized pustular psoriasis.

first: Poulami

last: Dey

address: 1301 Catherine St

institute: Michigan Medicine

summary: Generalized pustular psoriasis (GPP) is a severe subtype of psoriasis characterized by epidermal neutrophil infiltration, often presenting as acute, potentially life-threatening flares. However, the characterization of the immune micro-environment in GPP lesions remains largely unknown. Here, we used single-cell RNA profiling to interrogate the transcriptomes of 60,000 single cells from GPP lesional skin (n=13) and healthy adult skin (n=4), combined with spatial transcriptomics. We identified a unique neutrophil subset lacking CASP8 expression but exhibiting elevated levels of inflammatory pathway genes, including RIPK1, NFKB1, IL1B, CXCL1, and CXCL8 in GPP flares, illustrating neutrophil transition from pre-inflammatory to a pro-inflammatory state, and activation of a communication network between IL36G+ keratinocytes and neutrophils in GPP lesions, with TNFSF15 (TL1A) released from neutrophils exaggerating the inflammatory crosstalk. We further demonstrated that fibroblasts and capillary endothelial cells function as central communication hubs in GPP, through dynamic receptor-ligand interactions with several spatially proximate immune cells, including T cells, neutrophils, and macrophages. In this work, we provide an in-depth view of immune cell participation and highlight the role of neutrophil-keratinocyte crosstalk in GPP pathogenesis.

overall_design: To investigate changes in cell type composition and transcriptional profiles in lesional skin from GPP patients, compared with skin from healthy controls.

first: Johann

middle: E

last: Gudjonsson

first: Lam

last: Tsoi


title: RNASeq of K562 cell clones for DNMT3A-WT, DNMT3A-W305*, DNMT3A-R882H, DNMT3A-R882C

summary: Donor blood saves lives, yet the potential impact of recurrent large-volume phlebotomy on donor health and hematopoietic stem cells (HSCs) remains largely unexplored. In our study, we conducted a comprehensive screening of 217 older male volunteer donors with a history of extensive blood donation (>100 life-time donations) to investigate the phenomenon of clonal hematopoiesis (CH). No significant difference in the overall incidence of CH was found in frequent donors (FD) compared to sporadic donors (<10 life-time donations, 212 donors). However, upon deeper analysis of mutations in DNMT3A, the most commonly affected gene in CH, we observed distinct mutational patterns between the FD and age/sex matched control donor (CD) cohorts. Functional analysis of FD enriched DNMT3A variants examined in CRISPR-edited human HSCs demonstrated their competitive outgrowth potential upon stimulation with erythropoietin (EPO), a hormone which increases in response to blood loss. In contrast, clones harboring leukemogenic DNMT3A R882 mutations increase upon stimulation with IFNy. Through concurrent mutational and immunophenotypic profiling of primary samples at single cell resolution, a myeloid bias of premalignant R882 mutant HSCs was found, while no significant lineage bias was observed in HSCs harboring EPO responsive DNMT3A variants. The latter exhibited  preferential erythroid differentiation when persistent erythropoietic stress was applied to CRISPR-edited human HSC xenografts. Our data demonstrate a nuanced ongoing Darwinian evolution at the somatic stem cell level, with EPO identified as a novel environmental factor that favors HSCs carrying certain DNMT3A mutations.

overall_design: We performed bulk RNA sequencing in K562 cells after isolation and expansion of single mutant clones for each genotype

first: Miriam

last: Llorian

first: Dominique

last: Bonnet


title: A Y-linked duplication of anti-Mullerian hormone is the sex determination gene in threespine stickleback

city: Athens

first: Matthew

last: Treaster

state: GA

address: 120 E Green St

department: Genetics

institute: University of Georgia

summary: Many taxa have independently evolved genetic sex determination where a single gene located on a sex chromosome controls gonadal differentiation. The gene anti-Mullerian hormone (amh) has convergently evolved as a sex determination gene in numerous vertebrate species, but how this gene has repeatedly evolved this novel function is not well understood. In the threespine stickleback (Gasterosteus aculeatus), amh was duplicated onto the Y chromosome (amhy) ~22 million years ago. To determine whether amhy is the primary sex determination gene, we used CRISPR/Cas9 and transgenesis to show that amhy is necessary and sufficient for male sex determination, consistent with the function of a primary sex determination gene. We find that amhy contributes to a higher total dosage of amh early in development and likely contributes to differential germ cell proliferation key to sex determination. The creation of sex reversed lines also allowed us to investigate the genetic basis of secondary sex characteristics. Threespine stickleback have striking differences in behavior and morphology between sexes. Here we show one of the classic traits important for reproductive success, blue male nuptial coloration, is controlled by both sex-linked genetic factors as well as hormonal factors independent of sex chromosome genotype. This research establishes stickleback as a model to investigate how amh regulates gonadal development and how this gene repeatedly evolves novel function in sex determination. Analogous to the “four core genotypes” model in house mice, sex-reversed threespine stickleback offer a new vertebrate model for investigating the separate contributions of gonadal sex and sex chromosomes to sexual dimorphism.

overall_design: Threespine stickleback fish embryos were sequenced for bulk gene expression on the Illumina platform. This gene expression data was used to investigate differential gene expression between males and females across early development.


title: Circadian regulator REV-ERBα is a master regulator of tumor lineage plasticity and an effective therapeutic target [ChIP-Seq]

city: sacramento

first: Xiong

last: Zhang

address: 4645 2nd Ave

institute: University of California, Davis

summary: Epigenetic and transcriptional dysregulation plays a fundamental role in tumor lineage plasticity (LP). However, the underlying mechanisms, especially for the initial events of LP development, are still poorly understood. Here, we report that in progression of prostate cancer from adenocarcinoma to treatment-induced neuroendocrine prostate cancer (t-NEPC), anti-AR signaling inhibitors (ARSIs) reprogram the function of a circadian regulator/nuclear receptor REV-ERBα by switching its target gene programs from kinase signaling and metabolic programs to programs of LP, which includes neurogenesis, stem cell and epithelia to mesenchymal transition (EMT) as well as over fifteen LP drivers including POU3F2/BRN2, ASCL1, FOXA2, ONECUT2 and MYCN. Unexpectedly, REV-ERBα facilitates the chromatin occupancy of BRN2, ASCL1 and FOXA1 in their activation of LP programs, thus functioning as a master regulator of ARSI-induced LP driver network. Mechanistically, REV-ERBα induces chromatin accessibility and H3K27ac modification at promoters of LP genes through its recruitment of BRD4 and p300. Overexpression of REV-ERBα alone is sufficient to induce LP and neuroendocrine phenotype and confers cell resistance to ARSI in adenocarcinoma cells. Loss of REV-ERBα potently inhibits NEPC cell growth and abolishes the expression of LP drivers and gene programs. Pharmacological inhibition of REV-ERBα exhibits high potency in blocking the growth of NEPC tumors including patient-derived xenografts.  Our findings reveal that therapy-induced LP development entails a coordinated induction of a network of LP drivers and that REV-ERBα is a novel master regulator of the network and a promising therapeutic target for treatment of advanced prostate cancer such as NEPC.

overall_design: ChIP-seq profiling of circadian regulator REV-ERBα, BRN2, ASCL1, FOXA1, BRD4,p300, H3K27ac  in ARSI-resistant prostate cancer cells and PDX tumors.

first: Yatian

last: Yang

first: Ronald M

last: Evans

first: Hongwu

last: Chen


title: The opposing effect of TET2-derived clonal hematopoiesis in stemness and immunity

city: LONDON

first: Giuseppe

middle: Alessandro

last: D'Agostino

state: ENGLAND

address: 1 Midland Road

summary: The expansion of HSC clones and their immune progeny carrying somatic mutations is defined as clonal hematopoiesis (CH) with its prevalence increasing with age. Individuals with CH are at higher risk of developing hematological malignancies but also other pathologies, such as cardiovascular or inflammatory pulmonary diseases. Understanding which environmental conditions favor mutant clones and the CH-immune system response to such environments is key to designing therapeutic strategies to stall the expansion of mutant clones and the development of CH-associated pathologies. In this work, we characterized how mutations in TET2, one of the most frequent in CH, increase the competitive advantage of human HSCs upon environmental stress conditions and at the same time shape the immune response of their myeloid progeny. Employing a humanized mouse model, we show that environmental conditions that drive emergency myelopoiesis promote human TET2-derived CH. We unravel an opposite and cell-specific impact of TET2 mutations on HSCs and their myeloid progeny to inflammatory stress. In such environments, TET2Mut HSCs have a mitigated transcriptional upregulation of inflammatory pathways, and by contrast, TET2Mut myeloid cells have an exacerbated inflammatory response. A distinct monocyte-macrophage trajectory derived from TET2Mut HSCs contributes to pathological inflammation. These findings provide evidence at the human cell level and reconcile the notion that TET2-derived CH is associated with both increased stemness at the HSC compartment and promoted inflammatory environments from the myeloid progeny.

overall_design: We performed single cell RNA sequencing in human TET2-WT and TET2-Mut hematopoietic stem and progenitor cells and immune cells isolated from bone marrow of humanised mice after a regimen of LPS injections to induce systemic inflammation.


title: Circadian regulator REV-ERBα is a master regulator of tumor lineage plasticity and an effective therapeutic target. [ATAC-Seq]

overall_design: ATAC-seq profiling of circadian regulator REV-ERBα inhibition in ARSI-resistant prostate cancer cells.


title: Mitochondrial RNA cytosolic leakage drives the SASP

first: Dominik

last: Saul

overall_design: RNA-seq profiling of proliferating and senescent fibroblasts with and without mitochondria, which were subsequently transfected with isolated mitochondrial RNA (mtRNA).

first: Victorelli

last: Stella

first: Han

last: Yeaeun

first: Saul

last: Dominik

first: Passos

last: Joao


title: RNASeq of TET2-mutant human HSPCs and monocytes

city: London

first: Hector

last: Huerga Encabo

overall_design: We performed bulk RNA sequencing following our competitive assays, where TET2WT and TET2Mut cells are present within the same environment (Figure 2A). For the analysis of HSPCs, we isolated Lineage-CD34+ cells on day 4 of the LTC assay, 6 hours after the LPS treatment (Figure 2B-C). For the analysis of the myeloid compartment, we isolated monocytes (CD14+ cells) on day 14 of the LTC assay, also 6 hours after the last LPS treatment (Figure 2D-E) to compare between the two cell types the impact of TET2 mutations in the transcriptional adaptation to stress.

first: Huerga Encabo

last: Hector

first: Llorian Sopena

last: Miriam

first: Bonnet

last: Dominique


title: Modeling SMAD2 mutations in iPSCs provides insights into cardiovascular disease pathogenesis [RNA-seq]

first: Jon

last: Seidman

state: Massachusetts

address: 77 Avenue Louis Pasteur

institute: Harvard Medical School

summary: Rationale: SMAD2 is a co-regulator that binds a variety of transcription factors and modulates activities during human development. Heterozygous SMAD2 loss-of-function (LoF) variants and missense variants are identified in patients with complex congenital heart disease (CHD), and in patients with arterial aneurysms and dissections. Mechanisms that account for distinct cardiovascular phenotypes caused by SMAD2 variants remain unknown. We aimed to define the transcriptional and epigenetic effects of heterozygous SMAD2 loss-of-function (LoF) and missense variants and identify the involvement of transcription factor networks and genes that contribute to CHD. Compared to LoF variants, we assessed the function of rare SMAD2 missense variants of uncertain significance. Methods: We constructed isogenic induced pluripotent stem cells (iPSCs) with heterozygous or homozygous LoF and rare (MAF ≤ 10-5) missense SMAD2 variants identified in CHD probands. Wildtype and mutant iPSCs were studied by bulk RNA sequencing (RNAseq) and chromatin accessibility (ATACseq). Datasets were integrated with published SMAD2/3 chromatin immunoprecipitation (ChIPseq) studies. Cardiomyocyte differentiation and contractility of mutant iPSCs was assessed. Results: SMAD2 haploinsufficiency reduced open chromatin across promoter regions and altered the expression of 385 direct SMAD regulated genes, including ten previously identified as dominant CHD genes. Motif analyses in regions with differential ATAC peaks predicted that SMAD2 haploinsufficiency disrupts interactions with at least five transcription factors, NANOG, ETS, TEAD3/4, CREB1 and AP1. Compared to SMAD2-haploinsufficient cells, iPSCs with heterozygous R114C or W274C variants exhibited shared and distinct patterns of altered chromatin accessibility, transcription factor binding motifs, and dysregulated genes. Conclusions: SMAD2 haploinsufficiency disrupts chromatin interactions and perturbs transcription factor binding, disrupting normal cardiovascular development. Differences in the molecular consequences of LoF and missense variants likely contribute to clinical phenotypic heterogeneity. Additionally, these data indicate opportunities for molecular functional analyses to improve re-classification of SMAD2 variants of uncertain clinical significance.

overall_design: Examination of genetically engineered iPS cell lines that carry SMAD2 mutations. Two biological replicates for SMAD2 HET LoF, W274C, R114C cell lines and two technical replicates for each

first: Tarsha

last: Ward

first: Warren

last: Tai

first: Sarah

last: Morton

first: Joshua

last: Gorham

first: Christine


title: A 3D in vitro mouse ovarian follicle culture system preserves molecular signatures of ovulation for studying ovulation biology and novel non-hormonal discovery

city: Piscataway

first: Shuo

last: Xiao

state: NJ

address: 170 Frelinghuysen Rd

department: Department of Pharmacology and Toxicology

institute: Rutgers University

summary: We previously established an alginate hydrogel-based 3D encapsulated in vitro follicle growth (eIVFG) system to culture mouse and human immature follicles in vitro. The alginate encapsulation maintained the 3D architecture of follicles and support their growth from the secondary to the antral stage to acquire maturation. Upon the stimulation of human chorionic gonadotropin (hCG), a LH analog, the grown antral follicle from eIVFG was able to rupture, undergo cumulus cell expansion, and ovulate a fertilizable metaphase II (MII) oocytes. We have also demonstrated that mouse follicles cultured using eIVFG preserved dynamic transcriptomic profile of many key genes that are essential for gonadotropin-dependent folliculogenesis, such as genes related to gonadotropin hormone receptors and ovarian steroidogenesis. However, it is unknown whether these follicles preserve molecular signatures of ovulation, which would enable this system to serve as a truly scalable and highly controllable system for nominating therapeutic and contraceptive candidates. In the present study, we treated mouse mature follicles from eIVFG with hCG to induce in vitro ovulation. Follicles were collected at 0, 1, 4, and 8 hours post-hCG for single-follicle RNA sequencing (RNA-seq) analysis. Our results showed that follicles grown from eIVFG preserve key ovulatory genes and signaling pathways.

overall_design: After 8 days of eIVFG culture, follicles were stimulated for ovulation by hCG, and we collected time-course samples for RNA-seq analysis.Among all 46 samples collected in two batches, 9, 13, 13 and 11 follicles were collected at  0, 1, 4, and 8 hours post-hCG treatment, resepectively.

first: Jiyang

last: Zhang

first: Brittany

last: Goods

first: Pawat

last: Pattarawat

first: Yingzheng

last: Wang

first: Tessa

last: Haining

first: Qiang

first: Alex

middle: K

last: Shalek

first: Francesca

last: Ducan

first: Teresa

last: Woodruff


title: Transcriptomic Analysis of Zebrafish Larvae Exposed to Tire Wear Particle Leachates

status: Public on Oct 28 2025

city: Busan

first: Tae Hwan

last: Shin

address: Nakdong-daero 550 beon-gil

department: Health Sciences

country: South Korea

institute: Dong-A University

summary: For transcriptomic analysis, total RNA was extracted from pooled zebrafish larvae (80 per sample, three biological replicates per group) preserved in RNAlater™. High-quality RNA (RIN > 7.0), isolated using the RNeasy Mini Kit, was quantified and assessed for integrity. Ribosomal RNA was depleted prior to cDNA library construction using the TruSeq Stranded Total RNA Library Prep Gold Kit. The resulting libraries were quantified, size-validated, and sequenced using the Illumina NovaSeqX platform with 2×100 bp paired-end reads.

overall_design: Adult wild-type zebrafish were acclimated under controlled laboratory conditions for three months prior to breeding. Fertilized embryos were collected and exposed to vehicle control or one of three concentrations of tire wear particle (TWP) leachates (TWP 250, 350, and 700) from 4 to 168 hours post-fertilization (hpf). Each group included 200 embryos and was tested in triplicate. At 72 hpf, larvae were transferred to clean medium with adjusted water quality parameters while maintaining consistent temperature and photoperiod. For transcriptomic analysis, 80 larvae per replicate were pooled, and total RNA was extracted from three biological replicates per group. RNA samples with RIN > 7.0 were used for cDNA library construction, followed by high-throughput sequencing using the Illumina NovaSeqX platform.

first: Beomsu

last: Kim

first: Songyeon

first: Yunwi

last: Heo

first: Jung-Man

last: Jo

first: Soyoung

last: Ahn

first: Soyeon

last: Jeon

first: Sang-Hee

last: Woo

first: Seokhwan

last: Lee

first: Jae-Jun

first: June-Woo

last: Park

first: Wan-Seob

last: Cho


title: Single cell RNA sequencing identifies a novel tenogenic heterologous differentiation in endometrial carcinosarcomas

city: Madrid

first: Silvia

last: Gonzalez-Martinez

address: M-607, Km. 9, 100, Fuencarral-El Pardo

department: Pathological Anatomy

country: Spain

institute: Ramon y Cajal University Hospital

summary: This study presents a single-cell RNA sequencing (scRNAseq) analysis of six carcinosarcomas and two normal endometrial samples, profiling over 96,000 cells. By integrating transcriptomic data with inferred copy number variations (CNVs), immunohistochemistry (IHC), and fluorescence in situ hybridization (FISH), we resolve the complex cellular architecture and differentiation dynamics of these tumors. We identify distinct lineage-specific programs and uncover heterologous differentiation trajectories—including rhabdomyogenic, osteogenic, and a previously undescribed tenogenic lineage defined by SCX, MKX, and TNMD expression. Our results reveal that carcinosarcomas exhibit multilineage plasticity within their mesenchymal component and highlight the power of single-cell analysis to refine tumor classification and uncover hidden developmental programs.

overall_design: Six carcinosarcoma samples and two normal endometrial controls were analyzed using scRNAseq. Tumors were dissociated and processed for droplet-based scRNAseq, followed by integration with inferred CNVs, IHC, and FISH validation. The study aimed to characterize epithelial and mesenchymal lineages, identify novel differentiation programs, and explore the genomic architecture underlying carcinosarcoma plasticity.

last: González Martínez

first: José

last: Palacios


title: EZH2 alters aggressive variant prostate cancer subtype evolution, but is not required for neuroendocrine prostate cancer. [DNA methylation]

city: Buffalo

first: David

last: Goodrich

state: NY

address: Elm and Carlton Streets

department: Department of Pharmacology & Therapeutics

institute: Roswell Park Comprehensive Cancer Center

type: Methylation profiling by genome tiling array

summary: Advanced prostate cancer (PrCa) remains a leading cause of cancer-related death among men because nearly all patients progress on standard therapy that targets androgen receptor (AR) signaling. An important mechanism driving therapeutic resistance is lineage plasticity, the ability of PrCa cells to reprogram into AR independent lineage variants like neuroendocrine (NE) PrCa. The histone methyltransferase EZH2 has been implicated in PrCa lineage plasticity and EZH2 inhibitors are being evaluated clinically for the treatment of PrCa. Here we test how Ezh2 impacts the evolution of PrCa lineage plasticity using genetically engineered mice and human patient data. Ezh2 deficiency in mice diversifies the evolution of PrCa lineage variants, including a newly discovered variant with high expression of neurofilament genes. Analogous PrCa lineage variants are detectable in human specimens where they similarly correlate with EZH2 expression. Lineage variant diversification is associated with activation of transcription factor programs and AR cistrome reprogramming that is normally repressed by Ezh2. These findings advance the understanding of PrCa lineage plasticity and have implications for targeting EZH2 as a PrCa therapy.

overall_design: Intact (non-castrated) mouse endstage prostate tumor tissue from GEMMs (SKO, DKO, TKO) or with Ezh2 deletions (DKO_EP and DKO_EE) were profiled using the Infinium Mouse Methylation MM285 BeadChip array.

first: Justine

last: Jacobi


title: RNASeq of DNMT3A mutant K562

first: Hector

last: Huerga Encabo

summary: Donor blood saves lives, yet the potential impact of recurrent large-volume phlebotomy on donor health and hematopoietic stem cells (HSCs) remains largely unexplored. In our study, we conducted a comprehensive screening of 217 older male volunteer donors with a history of extensive blood donation (> 100 life-time donations) to investigate the phenomenon of clonal hematopoiesis (CH). No significant difference in the overall incidence of CH was found in frequent donors (FD) compared to sporadic donors (< 10 life-time donations, 212 donors). However, upon deeper analysis of mutations in DNMT3A, the most commonly affected gene in CH, we observed distinct mutational patterns between the FD and control donor (CD) cohorts. Functional analysis of FD enriched DNMT3A variants examined in CRISPR-edited human HSCs demonstrated their competitive  outgrowth potential upon stimulation with erythropoietin (EPO). In contrast, clones harboring leukemogenic DNMT3A R882 mutations exhibited a stronger response to inflammatory cues rather than EPO. Through concurrent mutational and immunophenotypic profiling of primary samples at single cell resolution as well as experiments in humanized mice, a myeloid bias of R882 mutant HSCs was detected. Remarkably, no significant lineage bias was observed in HSCs harboring EPO responsive DNMT3A variants. Our data demonstrate a nuanced ongoing Darwinian evolution at the somatic stem cell level, with EPO identified as a novel environmental factor that favors HSCs carrying certain non-malignant DNMT3A mutations. Crucially, our findings robustly demonstrate that frequent blood donation over many years does not promote the selection of malignant clones, thus strongly reaffirming its safety.

overall_design: Using CRISPR-mediated genetic engineering of K562 we generated K562 clones harbouring DNMT3A W305*, R882H or R882C mutations and compare their trnascriptome with wild type K562 cells.

middle: H

last: Encabo

first: Miriam

middle: L

last: Sopena

first: Dominique

last: Bonnet


title: EZH2 alters aggressive variant prostate cancer subtype evolution, but is not required for neuroendocrine prostate cancer [RNAseq]

overall_design: Bulk RNA-seq profiling of prostate tumor tissue from genetically engineered mouse models (SKO, DKO, TKO) or DKO mouse models with Ezh2 deletions (DKO_EP and DKO_EE). Prostate tissue was collected at 12 weeks of age (intact, pre-castration), endstage (castrated at 12 wks.), or intact endstage. Select TKO mice were treated with either Vehicle or Ezh2-inhibitor (CPI1688) following castration.


title: Distinct Molecular Responses of Human Intestinal Organoids to Proton and Photon Radiation

city: Houston

first: Sarah

middle: Elizabeth

last: Blutt

state: Texas

address: One Baylor Plaza

department: Molecular Virology and Microbiology

institute: Baylor College of Medicine

summary: Radiation exposure causes damage to tissues with high numbers of rapidly dividing cells such as the intestine. Mouse models have been widely used to understand the effects of radiation injury, repair following exposure, and to develop and test potential therapies for radiation-induced damage. However, although the mouse intestine shares physiological and molecular similarities with human intestine, translation and validation of findings in mouse radiation models to the human intestine have been limited by the lack of models. The development of ex vivo human intestinal organoids provides a new model in which the effects of radiation on the human intestinal epithelium can be examined and compared to in vivo animal models. Using human small intestinal jejunal organoids, multi-omic profiles were analyzed using bioinformatic pipelines to explore the response to a dose of proton and photon radiation exposure which may be encountered during a radiotherapy treatment or over time in high background radiation environments. Transcriptional signatures following radiation exposure were indicative of reprogramming of cells to a more primitive state that has been associated with regeneration and repair. Integrative analyses of transcriptomic and metabolomic datasets revealed enrichment of amino acid synthesis and metabolism pathways. Human intestinal organoids can complement and validate findings from traditional animal models and are emerging as a powerful tool to study responses to radiation exposure.

overall_design: RNAseq analysis of crypt-like or villus-like  human intestinal organoids (HIO) exposed to 1Gy gamma irradiation, 1Gy LET proton irradiation or left unirradiated.  Three technical replicates for each donor crypt and villus HIO +/- radiation exposure were pooled prior to RNA extraction.

first: Victoria

last: Poplaski

first: Tajhal D.

last: Patel

first: Mary K.

last: Estes

first: Cristian

last: Coarfa

first: Sarah E.


title: EZH2 alters aggressive variant prostate cancer subtype evolution, but is not required for neuroendocrine prostate cancer. [scRNA-seq]

overall_design: scRNA-seq profiling of prostate tumor tissue from mouse models (SKO,DKO, TKO) and DKO mouse models with Ezh2 deletions (DKO_EP and DKO_EE).


title: The Piwi-piRNA complex initiates transposon silencing via transcription termination factors PNUTS and Senataxin

city: Tokyo

first: Kensaku

last: Murano

address: 35 Shinanomachi, Shinjuku

department: Department of Molecular Biology

institute: Keio University School of Medicine

summary: Transcriptional silencing of transposable elements (TEs) is guided by Piwi and PIWI-interacting RNAs (piRNAs) through the SFiNX complex to induce heterochromatin formation. We previously reported that members of the SFiNX complex can induce TE silencing prior to heterochromatin formation, suggesting that Piwi-mediated silencing can be separated into two steps: initiation and maintenance. While many studies have investigated the heterochromatin-dependent maintenance step of TE silencing, the mechanisms underlying silencing initiation remain elusive. In this study, we found that SFiNX can induce silencing independent of H1 and HP1a through an association between the SFiNX-interaction partner Sov and two proteins involved in transcription termination, phosphatase 1 nuclear targeting subunit (PNUTS) and Senataxin. Mechanistically, PNUTS and Senataxin affect the elongation speed or stalling of RNA polymerase II (RNA Pol II) to induce repression. Our findings suggest that piRNAs act as transcription termination signals in the initiation step of piRNA-guided transcriptional repression preceding heterochromatinization of TEs.

overall_design:
To investigate the function of SUMO, SUMO-related genes, Piwi, and Setx in the silencing of transposable elements in ovarian somatic cells (OSCs), we performed gene knockdown experiments by siRNA transfection, followed by RNA-seq.
To investigate the function of Panx, PNUTS, and Setx in the silencing of transposable elements in OSCs, we performed ChIP-seq using anti-Pol II Ser5-P antibody.

first: Yaning

last: Wu

first: Toshiya

last: Nakahara


title: EZH2 alters aggressive variant prostate cancer subtype evolution, but is not required for neuroendocrine prostate cancer [ChIP-seq]

overall_design: ChIP-seq profiling of prostate tumor tissue from mouse models (SKO,DKO, TKO) and DKO mouse models with Ezh2 deletions (DKO_EP and DKO_EE) for androgen receptor (Ar) or histone marks (H3K27ac, H3K27me3, or H3K4me3). Paired-end sequencing reads were mapped to mm10 using bwa and deduplicated with Picard. deeptools was used to generate bigwig files.


title: Stress controls heterochromatin inheritance through histone H3 ubiquitylation [ChIP-Seq]

city: Bethesda

first: Shiv

last: Grewal

state: MD

address: 9000 Rockville Pike

department: National Cancer Institute

institute: National Institutes of Health

summary: Heterochromatin, marked by histone H3 lysine 9 methylation, can be inherited through cell divisions, maintaining gene repression that preserves cell identity and enables adaptation to environmental challenges. Studies in S. pombe have shown that heterochromatin propagation depends on the read-write mechanism wherein sufficient density of H3K9me3-modified nucleosomes, stabilized by histone deacetylases (HDACs), concentrate Clr4SUV39H on chromatin to promote further deposition of H3K9 methylation. Whether additional mechanisms control heterochromatin propagation via Clr4SUV39H, a part of the E3 ubiquitin ligase complex ClrC, was unknown. Here we uncover a ubiquitin-dependent heterochromatin-heritability regulatory hub (HRH) that broadly governs heterochromatin propagation, even without HDAC activity. The HRH is tuned by the limiting factor Raf1DDB2, a substrate receptor for ClrC ubiquitin ligase. In addition to linking Clr4SUV39H to other ClrC components on chromatin, Raf1DDB2 acts in a dosage-dependent manner to promote ubiquitination of histone H3 at lysine 14 (H3K14ub), which is critical for heterochromatin self-propagation. Importantly, HRH is intricately linked to environmentally responsive pathways, including nonsense-mediated decay (NMD) and TOR signaling, allowing cells to adapt to changing conditions. By modulating heterochromatin propagation, cells leverage the HRH to gain resistance to antifungal agents and to adapt to high temperature. Thus, heterochromatin self-propagation is actively regulated by the HRH in response to external stimuli, with broad implications for understanding mechanisms governing rapid changes in the epigenetic landscape in physiology and disease.

overall_design:
Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) in fission yeast. Input control and immunoprecipitation samples are provided for each experiment.
ChIP-Seq of H3K9me3 and H3K14ub histone modification in Schizosaccharomyces pombe cells

first: Bharat

last: Bhatt

first: Yi

last: Wei

first: Ashis

last: Pradhan

first: Jothy

last: Dhakshnamoorthy

first: Martin

last: Zofall

first: Hua

last: Xiao

first: Drisya

last: Vijayakumari

first: Shweta

last: Jain

first: Hernan

last: Folco

first: Hongyun

last: Qi

last: Ball

first: Tatiana

last: Karpova

last: Wheeler

first: Jiemin

last: Wong


title: DNGR-1 signalling limits dendritic cell activation for optimal antigen cross-presentation

first: Probir

last: Chakravarty

institute: Francis Crick Institute

summary: Innate immune receptors often induce activation of conventional dendritic cells (cDCs) and enhance antigen (cross-)presentation, favouring immune responses. DNGR-1 (CLEC9A), a receptor expressed by type 1 cDCs (cDC1s) and implicated in immune responses to viruses and cancer, recognises F-actin exposed on dead cell remnants and promotes cross-presentation of associated antigens. Here, we show that recruitment of phosphatase SHIP1, a process governed by a single amino acid residue adjacent to the signalling motif of the receptor, partly explains how DNGR-1 fails to trigger cDC1 activation in vitro. Substituting this residue converts DNGR-1 into an activating receptor but decreases induction of cross-presentation of dead cell-associated antigens. Introducing the reverse mutation into the related receptor Dectin-1 impairs its activation capacity while enhancing its ability to promote cross-presentation. These findings reveal a functional trade-off in receptor signalling and suggest that DNGR-1 has evolved to prioritise antigen cross-presentation over cellular activation, possibly to minimise inflammatory responses to dead cells.

overall_design: DNGR-1 or mutant DNGR-1 expressing MuTuDCs were treated with 12.5 nM DNGR-1 ligand for 6 hours

first: Michael

last: Buck

first: Caetano

last: ReiseSousa


title: RNA-sequencing of SVF-derived preadipocytes and adipocytes from inguinal white adipose tissue of male mice treated with vehicle, CGRPa, NPY, NE, or SP

city: ANN ARBOR

first: Jorge

last: Ruas

state: Michigan

address: 2800 Plymouth RD, B025-2681 NCRC - Ruas Lab NCRC B025

summary: To investigate the effects of sensory neuropeptides on preadipocyte and adipocyte function, we treated cultures with vehicle, calcitonin gene-related peptide-alpha (CGRPa), neuropeptide y (NPY), norepinephrine (NE), or substance P (SP) for 10 hours and collected RNA for RNA-sequencing.

overall_design: RNA sequencing analysis of stromal vascular fraction (SVF)-derived preadipocytes from inguinal white adipose tissue (iWAT) of male mice (n = 3 per group, C57BL/6N). Preadipocyte or adipocytes were treated with vehicle, CGRPa, NPY, or SP for 10 hours. Total RNA was extracted using TRI reagent (Sigma-Aldrich, T9424) according to the manufacturer's instructions. RNA was further purified, and DNase treated (Zymo, R1013), and integrity confirmed prior to sequencing (Agilent Technologies, 2100 Bioanalyzer), with an RNA integrity number (RIN) greater than 7 as a threshold. Samples were purified by poly-A enrichment before library preparation using the Illumina NovaSeq X Plus Series platform with a paired-end 150 base pair strategy.

first: Kyle

last: Dumont


title: FeaSion Decodes the Regulatory Landscape and Functional Diversity of RNA Polymerase II CTD Phosphorylation

city: Beijing

first: Junyi

last: Zhu

address: Peking University, No.5 YiHeYuan Road, Haidian District

department: School of Life sciences

country: China

institute: Peking University

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

overall_design: Refer to individual Series


title: Targeting IGF2BP1 Enhances Immune Checkpoint Therapy in High-grade Serous Ovarian Cancer

city: Halle

first: Markus

last: Glaß

address: Kurt-Mothes-Str. 3a

department: Institute of Molecular Medicine

institute: Martin Luther University Halle-Wittenberg


title: Targeting IGF2BP1 Enhances Immune Checkpoint Therapy in High-grade Serous Ovarian Cancer [HGSC]

summary: High-grade serous ovarian cancer (HGSC) accounts for the vast majority (>70%) of ovarian cancer-associated deaths with minor therapeutic improvements. Among four proposed molecular subtypes of HGSC, the C5 subtype is distinguished by high proliferative potential and elevated immune evasion which is indicated by an unfavorable MHCI/PD-L1 ratio. However, the underlying key drivers of the C5 subtype remained elusive. Here we identify oncofetal RNA-binding proteins (RBPs) promoting the immune evasion of C5-HGSC. The IGF2 mRNA binding protein 1 (IGF2BP1) enhances expression of the E3 ligase MDM2, which induces degradation of IRF1 resulting in reduced MHCI presentation. Concomitantly, IGF2BP1 elevates PD-L1 synthesis by impairing its microRNA-directed silencing. This shifts the intra-tumoral MHCI/PD-L1 ratio, limits immune cell infiltration, and promotes evasion of tumor cells from cytotoxic T cells (CTLs) in human and syngeneic mouse models of ovarian cancer. The small molecule inhibitor BTYNB impairs IGF2BP1-directed regulation of MHCI/PD-L1 ratios, promotes CTL-directed killing as well as activation, and strongly synergizes with immune checkpoint inhibition (ICI) by Nivolumab in vitro and in vivo.

overall_design: HGSC patient samples were analyzed by scRNA-seq.

first: Nadine

last: Bley

first: Alexander

last: Rausch


title: Stress controls heterochromatin inheritance through histone H3 ubiquitylation [RNA-Seq]

first: Shiv

last: Grewal

overall_design: RNA-Seq of transcripts in Schizosaccharomyces pombe cells

first: Bharat

last: Bhatt

first: Yi

last: Wei

first: Ashis

last: Pradhan

first: Jothy

last: Dhakshnamoorthy

first: Martin

last: Zofall

last: Xiao

last: Vijayakumari

first: David


title: Targeting IGF2BP1 Enhances Immune Checkpoint Therapy in High-grade Serous Ovarian Cancer [polyA RNA-seq, unsorted co-cultures]

city: Halle (Saale)

first: Danny

last: Misiak

department: Molecular Cell Biology

overall_design: PolyA-RNA sequencing of unsorted co-cultures from ES-2 cells and PBMCs, treated with different conditions (3x DMSO, 3x BTYNB 10µM, 3x CUB (cucurbitacine B, 10 nM)).


title: Developing exocrine glands harbour a distinct macrophage population that is maintained by ILC2 derived GM-CSF [CD45 scRNA-seq]

city: Zurich

first: Burkhard

last: Becher

address: Winterthurerstrasse 190

country: Switzerland

institute: University of Zurich

summary: Granulocyte-macrophage colony-stimulating factor (GM-CSF) has a non-redundant role in the emergence and maintenance of alveolar macrophages (AMs). However, its role in steady-state myelopoiesis outside the lung is largely unexplored. We discovered a strong signal of GM-CSF production by type 2 innate lymphoid cells (ILC2s) in the developing salivary glands (SG). ILC2 derived GM-CSF fosters the development of a hitherto undescribed phagocyte subset, which we named adenophages. Detailed analysis focusing on phenotypic and transcriptional profiling revealed that adenophages represent a non-canonical macrophage subset displaying shared aspects of both, macrophages and dendritic cells. We found them to be homogenously distributed across the SG, but always forming a niche-triad with GM-CSF producing ILC2s and myoepithelial cells. Importantly, adenophages were present throughout all analyzed exocrine glands such as lacrimal glands and mammary glands, and were also identified in human SG, indicating a conserved role in exocrine glands across species.

overall_design: Cells from salivary glands of 20 day old mice were released by digestion. Total CD45+ cells were pre-selected using CD45 MicroBeads (Milteny Biotec, #130-052-301) on LS columns and further facs sort-purified for CD45+ cells. Each sample was pooled from two individual P20 mice.

first: Frederike

last: Westermann

first: Selma

last: Tuzlak

first: Victor

last: Kreiner


title: Cellular and molecular mediators of thymic dendritic cell homeostasis

city: Nashville

first: Bryan

middle: Robert

last: Helm

state: Tennessee

address: 2525 West End Avenue, Suite 1100

department: Center for Quantitative Science

institute: Vanderbilt University Medical Center

summary: To determine changes in cellular composition and gene signatures of hematopoietic antigen-presenting cells (APCs) in thymi harvested from mice where T-cell development is blocked at different maturation stages.

overall_design: 1) CD45+ MHC-II lo to + hematopoietic APCs were FACS-isolated from WT (C57BL6/J), Rag2-/-, B2m-/- and MHC-II-/- mice to determine the role of thymocytes in regulating DC homeostasis and activation (N=2 experimental repeats per genotype). We used scRNA-seq to determine the transcriptional heterogeneity of DCs across the four genotypes. 2) To determine if our flow-validated DC subsets correlate with transcriptional DC clusters, we FACS-isolated 7 DC subsets and labeled each DC subset with a unique TotalSeqBTM hashtag antibody (BioLegend), prior to pooling all subsets for scRNA-seq and hashtag-derived oligonucleotide (HTO) amplification.

first: Ehrlich

last: Lauren

first: Srinivisan

last: Jayashree

first: Liu

last: Qi

first: Lau

last: Ken

first: Helm

last: Bryan


title: Developing exocrine glands harbour a distinct macrophage population that is maintained by ILC2 derived GM-CSF [bulk RNA-seq]

overall_design: Myoepithelial cells were facs sorted from the salivary glands of 1 month old Csf2-KO (N=4) or Control (N=4) mice (male and female) and RNA-seq was performed


title: Targeting IGF2BP1 Enhances Immune Checkpoint Therapy in High-grade Serous Ovarian Cancer [mouse]

first: Markus

last: Glaß

overall_design: Intraperitoneal syngeneic mouse model using the IGF2BP1-directed small molecule inhibitor BTYNB (closed conformation) with or without PD-1 blocking antibodies. Ascites was analyzed by scRNA-seq.

first: Nadine

last: Bley

first: Alexander

last: Rausch

first: Robin

last: Rolnik


title: Arabidopsis DXO1 is an evolutionarily diverged homolog that impacts ribosome loading and mRNA surveillance

city: Canberra

first: Diep

last: Ganguly

state: ACT

address: 46 Sullivans Creek Rd

department: Research School of Biology

country: Australia

institute: The Australian National University

summary: The Rai1/Dxo1/DXO protein family are involved in eukaryotic 5′-end RNA quality control by hydrolyzing non-canonical mRNA cap structures, such as the NAD+ 5′-cap. The plant DXO1 ortholog shows distinct biochemical properties, compared to its fungal and mammalian counterparts, and only minor phenotypic traits can be attributed to its NAD+-decapping (deNADding) activity. On the other hand, the chloroplast and growth defects observed in dxo1 null mutants appear linked to the presence of a serine-rich domain at its N-terminus. Here, we report that plant DXO1 is a distinct ortholog that likely diverged earlier than its fungal and mammalian counterparts. We attribute this to the presence of a serine-rich N-terminal domain, which is found almost exclusively in plants and likely arose during the emergence of angiosperms. Degradome profiling revealed that Arabidopsis DXO1 contributes to mRNA surveillance, via examination of exon junction complex and ribosome footprints, and this activity is reliant on the presence of the serine-rich N-terminal domain in Arabidopsis. Furthermore, polysome profiling revealed that plants lacking DXO1 have severe ribosome loading defects, which may explain the loss of mRNA surveillance and alterations in pre-rRNA processing. Taken together, our data leads us to propose a model whereby Arabidopsis DXO1 is involved in 5′-end RNA quality control, which is critical for ribosome loading that impacts translational fidelity and translation-dependent mRNA surveillance.

overall_design: GMUCT was performed on two biological replicates of 12-day-old transgenic Arabidopsis seedlings expressing DXO1 variants fused to GFP under the control of the 35S promoter from the cauliflower mosaic virus (Kwasnik et al 2019 Nucleic Acids Research).

first: Xiang

last: Yu

first: Brian

last: Gregory


title: Early female germline development in Xenopus laevis: Stem cells, nurse cells and germline cysts

status: Public on Oct 27 2025

city: Baltimore

first: Asya

last: Davidian

state: MARYLAND

address: 3520 San Martin Dr

department: Embryology

institute: Carnegie Institution for Science

summary: In Drosophila, germline cysts arise through synchronous mitotic divisions and acquire a polarized architecture organized by the fusome, which guides oocyte specification and supports meiotic progression. Similar cyst structures exist in non-mammalian vertebrate ovaries, but their polarity and function have remained uncertain. Using single-cell RNA sequencing and high-resolution imaging, we reconstructed the germ cell differentiation trajectory in Xenopus laevis and uncovered striking parallels with invertebrate and mouse cyst development. We identified a distinct germline stem cell (GSC) population marked by piwil4, low translational activity, and expression of neuronal-specific and transposon-silencing genes. Downstream from GSCs, during cyst development, an asymmetric fusome-like structure (FLS) composed of stable microtubules forms a rosette-like connection between cystocytes and co-localizes with Golgi vesicles and ER, suggesting polarized trafficking. In contrast to previous claims, ∼80% of EdU-prelabeled cyst cells turned over rather than forming oocytes, consistent with a nurse cell fate. The striking parallels described here between cyst and fusome formation, polarization, cyst breakdown, and nurse cell turn over to produce relatively few oocytes, argue that amphibian cysts have important functions in female gametogenesis.

overall_design: We performed single-cell RNA sequencing on juvenile Xenopus laevis ovaries collected at developmental stages NF62–66 to characterize germline populations during early ovarian development. Dissected ovaries were enzymatically dissociated, and approximately 10,000 viable cells per sample were loaded onto the 10X Genomics Chromium platform. Libraries were generated from three biological replicates and sequenced on an Illumina NextSeq 500. Raw reads were processed using Cell Ranger v8.0.1, and the resulting count matrices were used for downstream analysis.

first: Allan

middle: C

last: Spradling


title: Targeting IGF2BP1 Enhances Immune Checkpoint Therapy in High-grade Serous Ovarian Cancer [small RNA-seq]

first: Danny

last: Misiak

overall_design: MiTRAP experiments using 3´UTR of PD-L1 or MS2 control RNA were essentially performed as described recently (Busch et al. 2016). MBP-MS2BP (maltose-binding protein – MS2 binding protein) was coupled to amylose resin. Bait RNA was in vitro transcribed form linearized templates containing MS2 loops and coupled to the pre-coupled MBP-MS2BP. Cell-free extracts of 5x106 ES-2 cells per condition generated in binding buffer (20mM Tris, pH 7.5, 150mM NaCl, 1.5mM MgCl2, 8.6% glycerol and 0.05% NP40) supplemented with protease inhibitor cocktail (1:200; Sigma Aldrich) were cleared by centrifugation and incubated with bait-coupled resin and supplements (11 mg/ml heparin, 1mM DTT and 400 U/ml RNAsin (Promega)) for 30 min at room temperature. After three washing steps with heparin-supplemented binding buffer, protein–RNA complexes were eluted twice in 150 μl binding buffer supplemented with 15mM maltose. Protein enrichment was tested by Western blotting. To identify associated microRNAs, RNA was isolated by phenol-chloroform extraction and quality checked by a Bioanalyzer.  Library preparation and single-end short-read RNA sequencing was performed on Illumina HiSeq 1500 platform at Novogene (Hong Kong).


title: Developing exocrine glands harbour a distinct macrophage population that is maintained by ILC2 derived GM-CSF [scRNA-seq]

first: Burkhard

last: Becher

overall_design: Cells from salivary glands of 20 day old mice were released by digestion. Nuclei were isolate ….

first: Frederike

last: Westermann

first: Selma

last: Tuzlak

first: Victor

last: Kreiner


title: Small molecule inhibition rescues the skeletal dysplasia phenotype of Trpv4 mutant mice

city: Los Angeles

first: Jennifer

last: Zieba

state: CA

address: 615 Charles E. Young Dr.

institute: University of California, Los Angeles

summary: The TRPV4 skeletal disorders are characterized by short stature and scoliosis but the cellular mechanisms behind this phenotype are unclear. We isolated tibial and femoral cartilage growth plate chondrocytes from postnatal day 3 wild type and Col2a1-Cre driven Trpv4 p.R594H conditional knock-in mice, which models the phenotype, and analyzed differential expression patterns using single cell RNA-seq.

overall_design: Growth plate chondrocytes from wild type and Col2a1-Cre induced Trpv4 p.R594H conditional knock-in mice were isolated from femoral and tibial cartilage tissue at postnatal day 3 and analyzed using scRNAseq.

first: Lisette

last: Nevarez

first: Taylor

last: Ismaili

first: Jorge

last: Martin

first: Derick

last: Diaz

first: Jocelyn

last: Ramirez

first: David

last: Goldstein

first: Sunil

last: Sahdeo

first: Deborah

last: Krakow

first: Daniel

last: Cohn