title: Reorganization of PBAF/cBAF remodelers on H3.3 chromatin regulates BMAL1 activity in the absence of circadian negative feedback

city: Lyon

first: Kiran

last: Padmanabhan

address: 32-34 Av. Tony Garnier

institute: IGFL - ENS Lyon

summary: Circadian rhythms in gene expression are coincident with 24hr dynamics in the recruitment of the core-clock transcription factor heterodimer CLOCK-BMAL1 to chromatin. In the liver, circadian chromatin is characterized by rhythmic histone modifications and the deposition of histone variant H2A.Z. However, other histone variants and the remodelers that work in conjunction with CLOCK-BMAL1 on variant chromatin remain poorly understood. Here, we reveal that H3.3 variant histone deposition peaks during the daytime in liver chromatin and that CLOCK-BMAL1 is recruited to H3.3 nucleosomes. Moreover, H3.3:CLOCK-BMAL1 associates with PBAF and BRG1/cBAF complexes - members of the SWI/SNF remodeler family - only during the active phase of the circadian cycle. In clock-disrupted Per1-/-; Per2-/- livers, we observe a depletion in ARID2, the central cog in the molecular assembly of the PBAF complex, accompanied by an increase in H3.3 incorporation. Remarkably, a disassembly of PBAF complex and the concurrent reduction in BRG1 triggers a remodeler reorganization in Per knockout livers, where BRM/cBAF now targets BMAL1 at readily-accessible genomic sites. An abundance of fragile acetylated H3.3 nucleosomes and a remodeler reorganization provide a mechanistic basis for BMAL1 activity in the absence of PER-mediated negative feedback.

overall_design: Circadian H3.3A, H3.1/2 and BMAL1 ChIP-seq experiment performed on MNase-digested soluble chromatin from wild-type and PerKO murine livers. Experiments were done in three independent biological replicates.

first: Dominika

last: Letková

first: Aurélie

last: Peticca

first: Damien

last: Sery

first: Yutaka

last: Suzuki


title: RNA-seq analysis of spinal lumbar (L3-5) samples after SCI

status: Public on Aug 12 2025

city: Mudanjiang

first: Ying

last: Wang

state: Heilongjiang

address: No 3,Tongxiang Road, Aiming district

country: China

institute: Mudanjiang Medical University

type: Expression profiling by high throughput sequencing

summary:
To explore the influence of gene expression in different time sequences and pathway changes in spinal lumbar segment after SCI
To investigate the key transcriptional regulatory factors regulating spinal lumbar axon regeneration after SCI
We then performed gene expression profiling analysis using data obtained from RNA-seq of spinal lumbar (L3-5) samples of SCI model at four time points.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for spinal lumbar (L3-5) samples  after SCI in acute stage (3dpi ) and subacute stage (7dpi  and 14dpi).

first: Wenyuan

last: Li


title: Bulk RNA-seq of mouse bone marrow derived macrophages co-cultured with platelets

status: Public on Aug 11 2025

city: Würzburg

first: Clement

last: Cochain

address: Josef Schneider Str.2 Haus D16

country: Germany

institute: University Hospital Würzburg

summary: We performed bulk RNA-seq gene expression analysis of macrophages co-cultured with resting or activated platelets

overall_design: Macrophages were generated from mouse bone marrow cells and treated with vehicle (PBS), thrombin alone, resting mouse platelets, or thrombin activated mouse platelets for 4 hours. RNA was extracted from the macrophages and gene expression analyzed by bulk RNA-seq.

first: Anna

last: Rizakou

first: Panagiota

last: Arampatzi


title: Gene expression profiling of the murine central nervous system at different developmental time points

city: Freiburg

first: Geoffroy

last: Andrieux

address: Breisacherstr 153

institute: University clinics Freiburg

summary: In this study, we performed bulk RNA-sequencing analysis to compare the gene expression profile of the developing CNS at different time points. We aimed to identify genes which are specifically enriched in the CNS around embryonic day E 9.5 compared to E14.5 and postnatal day P60.

first: Melanie

last: Boerries

first: Philippe

last: Petry

first: Alexander

last: Oschwald

first: Katrin

last: Kierdorf


title: Co-regulation of HexC fine-tunes HexA-dependent transcription and acid stress response in Pectobacterium carotovorum.

status: Public on Aug 10 2025

city: Seoul

first: EUNHYE

last: GOO

address: 1, Gwanak-ro, Gwanak-gu

department: Agricultural Biotechnology

country: South Korea

institute: Seoul National University

summary: HexA is a global transcriptional regulator involved in the modulation of virulence and environmental adaptation in Pectobacterium carotovorum. To identify the genes regulated by HexA, we performed RNA-seq analysis comparing the wild-type strain PCC27 and a hexA deletion mutant at mid-log and early stationary growth phases. This study provides comprehensive transcriptomic profiles that reveal the influence of HexA on gene expression dynamics across growth stages.

overall_design: Total RNA was extracted from P. carotovorum PCC27 wild-type and ΔhexA mutant strains grown in LB medium. Samples were collected at two time points: mid-log phase and early stationary phase. Three independent biological replicates were prepared for each condition. rRNA-depleted libraries were sequenced using the Illumina NovaSeq 6000 platform with a paired-end 2 × 101 bp configuration.

first: Eunhye

last: Goo

first: Serim

last: Lee

first: Jae Hoon


title: Expansion of the Cra regulatory network and conservation of transcriptional regulation in carbon metabolism of Escherichia coli [ChIP-exo]

city: Ulsan

first: Donghyuk

last: Kim

address: 50 UNIST-gil, Eonyang-eup, Ulju-gun

department: Department of Chemical Engineering

institute: UNIST

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Carbon metabolism in diverse bacteria is primarily governed by two key transcription factors, CRP and Cra. Although numerous studies have investigated their regulatory mechanisms under glucose conditions, the understanding of their role in controlling central metabolism across different carbon sources, such as galactose, remains limited, with scarce in vivo binding information available. In this study, we focused on the Cra binding profile of Escherichia coli grown on galactose, an unfavorable carbon source. While genome-wide Cra binding sites on galactose closely resemble that on glucose, however, further examination of transcriptomic and flux simulation data suggests a complex regulatory mechanism by which Cra modulates central carbon metabolism on galactose. Additionally, conservation analysis across a wide range of bacterial taxa revealed that Cra and its regulons are more broadly conserved compared to CRP and its regulons, indicating a potential role for Cra in the early evolution of bacterial central carbon metabolism.

sample_organism: Escherichia coli str. K-12 substr. MG1655

overall_design: [ChIP-exo] cra-8myc tagged cells were cultured in M9 minimal media with 0.2% galactose up to mid-log phase with a biological replicate.


title: Co-regulation of HexC fine-tunes HexA-dependent transcription and acid stress response in Pectobacterium carotovorum

summary: HexA is a global transcriptional regulator involved in quorum sensing and virulence regulation in Pectobacterium carotovorum PCC27. To identify genome-wide DNA binding sites of HexA, we performed chromatin immunoprecipitation followed by sequencing (ChIP-seq) using a FLAG-tagged HexA strain. Samples were collected at two different growth phases—mid-log and early stationary—to investigate condition-dependent differences in HexA binding profiles. A total of six samples (three biological replicates per condition) were sequenced on the Illumina NovaSeq 6000 platform using paired-end 101 bp reads. Peaks were called using MACS2, and downstream analysis was performed with ChIPseeker. This study provides insight into the dynamic regulation of HexA across bacterial growth stages.

overall_design: ChIP-seq was performed on P. carotovorum PCC27 expressing FLAG-tagged HexA. Samples were collected at mid-log and early stationary growth phases, with three biological replicates per condition (n=6 total). Paired-end sequencing was performed using the Illumina NovaSeq 6000 platform (2 × 101 bp). ChIP DNA was enriched using anti-FLAG antibody and libraries were prepared with the TruSeq ChIP Sample Preparation Kit. Peaks were called using MACS2 and annotated with ChIPseeker.


title: Medium-chain triglycerides improve cognition and systemic metabolism in mouse models of Alzheimer's disease

city: Sherbrooke

first: Mariano

last: Avino

address: 3201 Jean Mignault

department: Biochemistry and Functional Genomics

country: Canada

institute: Sherbrooke University

summary: Lifestyle-based interventions, including dietary modifications, can reduce dementia risk. In this regard, dietary supplementation with medium-chain triglycerides (MCT) has shown potential therapeutic benefits in individuals with Alzheimer's disease (AD). These effects are widely presumed to be mediated by hepatic conversion of MCT into circulating ketones. However, the physiological and cellular mechanisms underlying the benefits of MCT remain understudied, particularly in the context of AD. Here, we investigated the cellular and molecular changes occurring in the brain and systemically in response to dietary supplementation with MCT versus a ketogenic diet (KD). The experimental design consisted of comparing a 70% carbohydrate control diet to either a control diet supplemented with 10% MCT or a carbohydrate-free high fat KD. Diets were tested in two AD mouse models, slow-progressing 3xTg-AD mice that model pre-symptomatic/early stages and rapidly-progressing 5xFAD mice that model late stages of the disease. We found that MCT supplementation and KD both improved hippocampal-dependent spatial learning and memory, increased dendritic spine density of hippocampal neurons, and modulated hippocampal expression of genes associated with mitochondrial functions, synaptic structure, and insulin signaling in AD mouse models. However, unlike KD, MCT supplementation did not elevate circulating ketones, suggesting different mechanisms. Indeed, MCT enhanced the peripheral insulin response of AD mice, while KD conversely unveiled their latent metabolic vulnerability, increasing their hyperglycaemia, body weight gain, and adiposity. The systemic metabolic disturbances of AD mice correlated with transcriptomic alterations in hepatic lipid metabolism and ketogenesis genes and increased lipid droplet accumulation. These liver metabolic abnormalities were partially reversed by both MCT supplementation and KD, but in distinct ways. Notably, KD selectively triggered hepatic neutral lipid depletion and prominent proinflammatory gene expression while MCT down-regulated expression of cholesterol-related genes. Collectively, these findings reveal that MCT supplementation in the context of AD improves cognition and systemic metabolism without elevating circulating ketone levels.

overall_design: Hippocampus 3 months-old mice received for 6 months, either a Control diet (C, 70% carbohydrate, 20% long-chain triglycerides, 10% protein, 4.1 kcal/g, Research Diets), a Control diet enriched with medium-chain triglycerides (MCT or M, 70% carbohydrate, 10% long-chain triglycerides, 10% medium-chain triglycerides (C8:0-C10:0 ratio of 3:2), 10% protein, 4.1 kcal/g, Research Diets) or a carbohydrate-free high fat ketogenic diet (KD or K, 90% long-chain triglycerides, 10% protein, 6.7 kcal/g, Research Diets). WT and 3xTg-AD mice hippocampus were used with a sample size of 3-5 mice per genotype/diet giving a total of 24 hippocampi (WT-C=4, WT-K=4, WT-M=4, 3xTg-C=3, 3xTg-K=5, 3xTg-M=4).

first: Paule

middle: E

last: M'Bra

first: Laura

middle: K

last: Hamilton

first: Gaël

last: Moquin-Beaudry

first: Chenicka

middle: L

last: Mangahas

first: Federico

last: Pratesi

first: Anne

last: Castonguay

first: Sophia

last: Mailloux

first: Manon

last: Galoppin

first: Jessica

last: Avila Lopez

first: Megan

last: Bernier

first: Marta

last: Turri

first: Marian

last: Mayhue

last: Aumont

first: Martine

last: Tétreault

first: Stephen

middle: C

last: Cunnane

first: Karl

middle: J

last: Fernandes


title: Half of annotated human microRNAs are expressed at levels of questionable biological significance

city: Adelaide

first: Katherine

middle: Anne

last: Pillman

state: SA

address: North Terrace

department: Centre for Cancer Biology

country: Australia

institute: Adelaide University

type: Non-coding RNA profiling by high throughput sequencing

summary: MicroRNAs (miRNAs) are widely studied for their role in post-transcriptional gene regulation, often using exogenous overexpression to infer function. However, such approaches may not reflect physiological activity, as they rely on unnaturally high expression levels. To address this, we determined the minimal threshold required for detectable endogenous miRNA function using sensitive reporters. Comparison with small RNA sequencing data from thousands of cell lines and tissues revealed that over half of all miRNAs never reach functionally relevant levels, with more recently evolved miRNAs generally exhibiting lower expression. This challenges the validity of numerous studies reporting roles for lowly expressed miRNAs, suggesting many findings reflect artefacts of overexpression rather than true biological activity. Our work underscores the importance of evaluating miRNA function in a native context and supports the view that the functional human “microRNAome” is substantially smaller than estimates based solely on sequencing data.

overall_design: RNA-seq profiling of small RNAs expressed in wildtype HEK-293T, HeLa, MDA-MB-231 and C32 cells

first: Saba

middle: A

last: Kachooei

first: Julie

middle: M

last: Bracken

first: Philip

last: Gregory

first: Cameron

middle: P


title: Expansion of the Cra regulatory network and conservation of transcriptional regulation in carbon metabolism of Escherichia coli [RNA-seq]

first: Donghyuk

last: Kim

platform_organism: Escherichia coli str. K-12 substr. MG1655

overall_design: [RNA-seq] Wild type E. coli MG1655 cells were cultured in M9 minimal media with 0.2% galactose and glucose upto mid-log phase with a biological replicate


title: Expansion of the Cra regulatory network and conservation of transcriptional regulation in carbon metabolism of Escherichia coli

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

overall_design: Refer to individual Series


title: Chronic ISG15 Exposure Accelerates CD8 T Cell Dysfunction while Increasing PD-1 Blockade Sensitivity in Oral Squamous Cell Carcinoma

city: Columbus

first: Yu-Lin

last: Chen

state: OH

address: 1645 Neil Avenue

department: Department of Microbial Infection and Immunity

country: USA

institute: The Ohio State University

summary: Immunotherapy has emerged as a promising treatment for head and neck squamous cell carcinoma (HNSCC), but clinical responses remain limited. Elevated expression of interferon-stimulated gene 15 (ISG15), associated with oral squamous cell carcinoma (OSCC), may contribute to this limited efficacy. While chronic interferon signaling impairs CD8 T cell function, the specific role of secreted ISG15 in T cell exhaustion remains unclear. Analyzing human OSCC datasets, we observed significant enrichment of the ISG core score, including ISG15, in tumors compared to adjacent non-tumor tissues. Using a novel in vitro model of human T cell dysfunction, we found that acute ISG15 exposure enhances CD8 T cell effector functions, whereas prolonged ISG15 exposure induces severe dysfunction via an endocytic mechanism independent of LFA-1 signaling. In an immunocompetent orthotopic OSCC model, ISG15-expressing tumors exhibited accelerated growth and recruited more tumor-reactive CD8 T cells though these cells were dysfunctional. Importantly, PD-1 blockade treatment significantly slowed tumor progression and restored T cell functionality in ISG15-expressing tumors. Our findings indicate that chronic ISG15 exposure promotes CD8 T cell dysfunction; however, these cells remain responsive to PD-1 blockade. This study reveals ISG15 as a potential biomarker for identifying patients likely to benefit from PD-1 blockade therapy.

overall_design: In vitro chronic TCR stimulation of cord blood–derived human CD8⁺ T cells in the presence or absence of TGF-β1 and ISG15

first: Hazem E

last: Ghoneim

first: Ava

last: Lowin


title: Comparative Analysis of Teratogen-Induced Malformations and Gene Expression Across Zebrafish Strains in Early Development

city: Sagamihara

first: Hiromi

last: Hirata

state: Kanagawa

address: 5-10-1, Fuchinobe, Chuo-ku

country: Japan

institute: Aoyama Gakuin University

summary: Zebrafish embryos are widely used in developmental toxicity testing. However, the extent to which genetic background influences susceptibility to teratogenic compounds remains incompletely understood. We here evaluated inter-strain variability in both phenotypic and transcriptomic responses to six model teratogens using five commonly utilized zebrafish strains, AB, TU, RW, WIK, and PET. All test compounds, valproic acid, hydroxyurea, methotrexate, acitretin, topiramate, and ibuprofen, elicited concentration-dependent developmental hazards characterized by malformations at moderate doses and lethality at higher concentrations. Despite distinct toxicodynamic profiles, the incidence and severity of phenotypic outcomes were highly consistent across strains. Transcriptomic analysis was performed following exposure to valproic acid, hydroxyurea, and warfarin, revealing strong, dose-dependent gene expression changes that were largely conserved among strains. Principal component analysis demonstrated that chemical concentration, rather than strain, was the dominant driver of transcriptional variation. Minor strain-specific differences were observed at baseline or low-dose levels but did not alter the overall direction or magnitude of response. These findings demonstrate that zebrafish embryos from diverse genetic backgrounds exhibit broadly conserved developmental and molecular responses to teratogens. The minimal inter-strain variability supports the use of any wild-type strain, transgenic line, or even outbred population in developmental toxicity testing without compromising sensitivity or reproducibility. Our study reinforces the suitability of zebrafish as a robust vertebrate model in regulatory toxicology.

overall_design: AB, TU, RW, WIK, and PET were treated with seven teratogens; valproic acid, hydroxyurea, methotrexate, acitretin, topiramate, ibprofen and warfarin.

first: Chitose

last: Taya

first: Kota

last: Ujibe

first: Shinnosuke

last: Shimodaira

first: Aoto

last: Sakamoto

first: Seiji

last: Wada

first: Makoto

last: Kashima


title: Ribosome biogenesis in embryo-like grand states as potential therapeutic target for KRAS mutant CRC

city: Ariake

first: Ryoji

last: Yao

state: Tokyo

institute: Cancer Institute, JFCR

summary: KRAS signaling is a promising target for cancer treatment, and the development of molecular targeted therapies has significantly improved the treatment outcome. However, cancer cells often adopt drug tolerant state to impede efficacy of chemotherapeutic agents. In colorectal cancer (CRC), therapeutic agents targeting KRAS have often failed to demonstrate a significant benefit, and the elucidation of its mechanism has been major clinical challenge. Here we show that MEK inhibitor, trametinib, and allele specific KRAS inhibitor, AMG510, induce the cellular state resembling the pluripotent naïve grand state of embryonic stem cells (ESCs). These cells are characterized by elevated ribosome biogenesis with Myc expression profile, yet global protein synthesis rate is low. We found they were vulnerable to the inhibition of ribosome biogenesis, and RNA polymerase I (RNA PolI) inhibition showed synergistic anti-tumor effects with trametinib and AMG510 in the autochthonous mouse model of intestinal tumors and in human patient-derived organoids with KRAS G12C mutation, respectively. These observations in PDOs demonstrated that CRC cells are equipotent to enter the pluripotent stem-like state by exploiting the cellular program of early embryogenesis and become tolerant to KRAS inhibition. High ribosome biogenesis is indispensable to maintain this cell state and is a potential therapeutic target of these cells. Our findings uncover previously unrecognized mechanisms of drug tolerance of CRC to KRAS inhibition, and provide the clue for the development of therapeutic strategies.

overall_design: PDO established from colorectal cancer patients were subjected to scRNA-seq analysis.


title: Assessing the impact of TET2 and TET3 deletion in TCRa and TCRb expression and repertoire  in murine CD4 T cells in physiological and pathological conditions [TCR-SEQ]

city: New York

first: Tarmo

last: Äijö

address: 162 5th Avenue

department: Center for Computational Biology

institute: Flatiron Institute

summary: TET proteins mediate DNA demethylation by oxidizing 5-methylcytosine to 5-hydroxymethylcytosine (5hmC) and other oxidative derivatives. While we have previously demonstrated a dynamic enrichment of 5hmC during T cell lineage specification, the impact of TET proteins in CD4 cell lineage differentiation remains largely unknown. Here, we discover that TET proteins exert a fundamental role in regulating the differentiation and maturation of the CD4 lineage by controlling the expression of the lineage specifying factor Th-POK, which is encoded by Zbtb7b.  We demonstrate that TET proteins mediate DNA demethylation of a proximal enhancer, critical for the stability of Th-POK expression. In addition, TET proteins drive the DNA demethylation of site A at the Zbtb7b locus to facilitate GATA3 binding, an event critical for Th-POK expression and subsequent CD4 maturation.  The impaired maturation of Tet2/3 DKO CD4 cells leads to the acquisition of a stemness signature and results in acquisition of malignant traits.

overall_design: We evaluated TCRa and TCRb chain repertoire to assess clonality in control, Tet2/3 DKO CD4 SP, Tet2/3 DKO CD4 peripheral T cells as well as transferred and expanded cells

first: Ageliki

last: Tsagaratou


title: Assessing the impact of TET2 and TET3 deletion in TCRa and TCRb expression and repertoire in murine CD4 T cells in physiological and pathological conditions [TCR-seq]

summary: TET proteins are tumor suppressors that through their catalytic activity oxidize 5-methylcytosine to 5-hydroxymethylcytosine, to promote DNA demethylation and to regulate gene expression. Notably, TET2 is one of the most frequently mutated genes in hematological malignancies, including T cell lymphomas. However, murine models with deletion of TET2 do not exhibit T cell expansion, presumably due to redundancy with other members of the TET family of proteins. In order to gain insight on the TET mediated molecular events that safeguard T cells from aberrant proliferation we performed serial adoptive transfers of murine CD4 T cells that lack concomitantly TET2 and TET3 to fully immunocompetent congenic mice. Our data reveal a progressive acquisition of malignant traits upon loss of TET2 and TET3 that is characterized by loss of genomic integrity, acquisition of aneuploidy and upregulation of the protooncogene Myc.


title: Assessing the impact of TET2 and TET3 deletion in TCRa and TCRb expression and repertoire in murine CD4 T cells in physiological and pathological conditions


title: CUT&Tag Identifies Repetitive Genomic Loci that are Excluded from ChIP Assays [chromatin fractionation and sequencing]

city: Rochester

first: Patrick

last: Murphy

state: New York

address: 601 Elmwood Avenue

department: Biomedical Genetics and Genomics

institute: University of Rochester

summary: Determining the genomic localization of chromatin features is an essential aspect of investigating gene expression control, and ChIP-Seq has long been the gold standard technique for interrogating chromatin landscapes. Recently, the development of alternative methods, such as CUT&Tag, have provided researchers with alternative strategies that eliminate the need for chromatin purification, and allow for in situ investigation of histone modifications and chromatin bound factors. Mindful of technical differences, we set out to investigate whether distinct chromatin modifications were equally compatible with these different chromatin interrogation techniques. We found that ChIP-Seq and CUT&Tag performed similarly for modifications known to reside at gene regulatory regions, such as promoters and enhancers, but major differences were observed when we assessed enrichment over heterochromatin-associated loci. Unlike ChIP-Seq, CUT&Tag detects robust levels of H3K9me3 at a substantial number of repetitive elements, with especially high sensitivity over evolutionarily young retrotransposons.  IAPEz-int elements for example, exhibited underrepresentation in mouse ChIP-Seq datasets but strong enrichment using CUT&Tag.  Additionally, we identified several euchromatin-associated proteins that co-purify with repetitive loci and are similarly depleted when applying ChIP-based methods.  This study reveals that our current knowledge of chromatin states across the heterochromatin portions of the mammalian genome is extensively incomplete, largely due to limitations of ChIP-Seq. We also demonstrate that newer in situ chromatin fragmentation-based techniques, such as CUT&Tag and CUT&RUN, are more suitable for studying chromatin modifications over repetitive elements and retrotransposons.

overall_design: Chromatin fractionation sequencing and analysis for pellet and supernatant samples. MEFs were crosslinked and sonicated for 1 or 3 cycles, then centrifuged and separated into pellet and supernatant fractions.  Supernatant samples sonicated for 1 or 3 cycles, and pellet samples sonicated for 1 cycle were sequenced. 3 replicates per sample.

first: Park

last: Brandon

first: Hua

last: Shan

first: Casler

last: Karli

first: Cefaloni

last: Eric

first: Ayers

last: Michael

first: Lake

last: Rahiim

first: Murphy

last: Kristin

first: Vertino

last: Paula

first: O'Connell

last: Mitchell

last: Patrick


title: Pancreatic cancer-intrinsic HuR regulates the pro-tumorigenic properties of extracellular vesicles

status: Public on Aug 09 2025

city: Portland

first: Yifei

last: Guo

address: 2730 S Moody Ave

institute: Oregon Health & Science University

summary: Pancreatic ductal adenocarcinoma (PDAC) tumors contain chaotic vasculature that limits immune surveillance and promotes early events in the metastatic cascade. However, current antiangiogenic therapies have failed in PDAC, and thus, it remains important to uncover mechanisms by which cancer cells signal to endothelial cells to increase angiogenesis. Our lab has shown that the tumor-intrinsic RNA-binding protein HuR (ELAVL1) plays an important role re-shaping the tumor microenvironment (TME) by regulating the stability and translation of cytokine encoding transcripts. Herein, we demonstrate that PDAC-intrinsic HuR influences endothelial cell function in the TME via extracellular vesicle (EV) signaling, an underexplored signaling axis in tumor progression. We found that HuR knockout (KO) tumors have impaired growth in an immunocompetent mouse model, and that administering purified wildtype (WT) EVs can increase tumor growth. Further, we observed that PDAC EVs contain HuR-dependent cargoes relating to endothelial cell function and angiogenesis. Treatment of endothelial cells with HuR WT EVs strongly increased the expression of genes involved in barrier function and endothelial cell development, and directly increased their migratory and tube forming functions. In an immunocompetent orthotopic mouse model of PDAC, we showed that HuR increases endothelial cell presence and sprouting, while decreasing ICAM-1 expression. Importantly, we found utilizing a genetic EV reporter, that decreased ICAM-1 within WT tumors occurs in endothelial cells that have imported PDAC EVs, suggesting that this signaling axis is directly modulating endothelial cell behavior in vivo. Collectively, our data reveal a new role of HuR in EV signaling to endothelial cells, promoting angiogenesis while restricting endothelial cell leukocyte trafficking behavior.

overall_design: EV cargo profiling analysis of RNA-seq from PANC-1 WT and HuR KO EVs. Samples were collected from 48-hour conditioned media via size exclusion chromatography. HPaMEC profiling analysis of RNA-seq from HPaMECs treated with PANC-1 WT vs. HuR KO EVs for 24 hours.

first: Jennifer

last: Finan

first: Matthew

last: Ryer

first: Kevin

last: Hawthorne

first: Rosalie

last: Sears

first: Jonathan

middle: R

last: Brody


title: Faithful Modeling of Terminal CD8 T Cell Dysfunction and Epigenetic Stabilization In Vitro [RNA-seq]

status: Public on Aug 08 2025

first: Hazem

last: Ghoneim

state: Ohio

address: 460 W 12th Ave

department: Microbial Infection and Immunity

summary: Epigenetic scarring of terminally dysfunctional CD8 T cells hinders long-term protection and response to immune checkpoint blockade during chronic infections and cancer. We developed a faithful in vitro model for CD8 T cell terminal dysfunction as a platform to advance T cell immunotherapy. Using TCR-transgenic CD8 T cells, we found that 1-week peptide stimulation, mimicking conditions in previous models, failed to induce a stable exhaustion program. In contrast, prolonged stimulation for 2-3 weeks induced T cell dysfunction but triggered activation-induced cell death, precluding long-term investigation of exhaustion programs. To better mimic in vivo exhaustion, we provided post-effector, chronic TGFβ1 signals, enabling survival of chronically stimulated CD8 T cells for over 3 weeks. These conditions induced a stable state of terminal dysfunction (TDysf), marked by a stable loss of effector, cytotoxicity, and memory programs, along with mitochondrial stress and impaired protein translation. Importantly, transcriptomic and epigenetic analyses confirmed the development of terminal exhaustion-specific signatures in TDysf cells. Adoptive transfer of TDysf cells revealed their inability to recall effector functions or proliferate after acute LCMV rechallenge. This novel tractable model system enables investigation of molecular pathways driving T cell terminal dysfunction and discovery of new therapeutic targets for cancer or chronic infections.

overall_design: RNA-seq profiling of D7 Effector, D19 Acute-2d, D19 Acute-7d, D19 Dysfunctional P14 CD8 T cells, with 2 biological replicates per group.

first: Amir

last: Yousif

first: Abbey

last: Saadey

first: Ava

last: Lowin

first: Ankita

last: Saini


title: RNA and PTBP1-RIP sequencing in BEAS-2B cells with circFCHO2 overexpression and knockdown

city: Hefei

first: Yucong

last: Wang

state: Anhui

address: 443 Huangshan Road

institute: USTC

summary: Circular RNAs (circRNAs) have emerged as key regulators in human diseases, yet their mechanisms of action in chronic obstructive pulmonary disease (COPD) remain largely unknown. We report a mammalian conserved circRNA circFCHO2 is identified to play critical roles in COPD. The expression level of circFCHO2 was significantly increased in COPD cell models, mouse models, and human lung tissue samples. Meanwhile, we demonstrated that circFCHO2 promotes epithelial-to-mesenchymal transition (EMT) of bronchial epithelial cells and extracellular matrix (ECM) remodeling. Mechanistically, circFCHO2 binds to and facilitates the nuclear translocation of PTBP1, thereby inhibiting the splicing of GRN pre-mRNA, which reduces PGRN protein expression levels. This activation of the NF-κB signaling pathway regulates the expression of EMT and ECM remodeling-related proteins, leading to the occurrence of airway remodeling. Overall, our study advanced the understanding of the molecular mechanisms by which circRNAs contribute to the airway remodeling of COPD.

overall_design: Measurement of RNA expression level and PTBP1 binding level in BEAS-2B cells with circFCHO2 overexpression and knockdown

first: Yanfei

last: Liu

first: Dahai

last: Zhao


first: Mariano

last: Avino

overall_design: 3 months-old mice received for 6 months, either a Control diet (C, 70% carbohydrate, 20% long-chain triglycerides, 10% protein, 4.1 kcal/g, Research Diets), a Control diet enriched with medium-chain triglycerides (MCT or M, 70% carbohydrate, 10% long-chain triglycerides, 10% medium-chain triglycerides (C8:0-C10:0 ratio of 3:2), 10% protein, 4.1 kcal/g, Research Diets) or a carbohydrate-free high fat ketogenic diet (KD or K, 90% long-chain triglycerides, 10% protein, 6.7 kcal/g, Research Diets). WT and 3xTg-AD mice liver and hippocampus were used with a sample size of 3-5 mice per genotype/diet/tissue giving a total of 48 samples (24 livers, 24 hippocampi).

first: Paule

last: M'Bra

first: Laura

last: Hamilton

first: Gaël

last: Moquin-Beaudry

first: Chenicka

last: Mangahas

first: Federico

last: Pratesi

first: Anne

last: Castonguay

first: Sophia

last: Mailloux

first: Manon

last: Galoppin

first: Jessica

last: Avila Lopez

first: Megan

last: Bernier

first: Marta

last: Turri

first: Marian

last: Mayhue

last: Aumont

first: Martine

last: Tétreault

first: Stephen

last: Cunnane

first: Karl

last: Fernandes


title: mRNA Sequencing Analysis of MCM7 Knockdown in Primary Hepatocytes Under IL-6 Stimulation

city: Wuhan

first: RUI

last: Zhou

address: Donghu Road 185

institute: Wuhan University

summary: Liver fibrosis, marked by excessive extracellular matrix deposition, remains a significant challenge due to its complex and not fully understood mechanisms. This study explores the role of Minichromosome Maintenance Complex Component 7 (MCM7) in the development of liver fibrosis, with a focus on its regulatory network and interactions. We knocked down MCM7 in primary hepatocytes and exposed them to IL-6 to simulate inflammatory conditions. RNA sequencing was then utilized to identify the differentially expressed genes and pathways influenced by MCM7 depletion. The results revealed notable alterations in gene expression, particularly in fibrosis- and inflammation-related pathways. Our study identified MCM7 as a key regulator of IL11 expression through the SHCBP1-RACGAP1-STAT3 signaling pathway, which contributes to the activation of hepatic stellate cells and the advancement of liver fibrosis. These findings underscore the critical role of MCM7 in liver fibrosis by promoting IL11 expression, thereby driving the activation of hepatic stellate cells and the progression of fibrosis. Overall, this research sheds light on a mechanistic pathway where MCM7 plays a pivotal role in liver fibrosis progression through the regulation of IL11 expression under inflammatory conditions.

overall_design: To understand the role of MCM7 in fibrotic hepatocytes, we employed shRNA to knock down MCM7 in primary hepatocytes. Following the knockdown, the cells were stimulated with IL-6, and RNA sequencing was conducted to profile gene expression. A comparative analysis was performed on the RNA-seq data from primary hepatocytes, both with and without IL-6 stimulation, to assess the effects of MCM7 knockdown.

first: R

first: C

last: Wang


title: High-throughput mRNA-sequence of MCM7 and SHCBP1 knock down hepatocytes with IL-1β stimulation

summary: Liver fibrosis, characterized by the excessive deposition of extracellular matrix, remains a formidable challenge due to its complex and poorly understood mechanisms. In this study, we aimed to elucidate the role of Minichromosome Maintenance Complex Component 7 (MCM7) and SHC SH2-Domain Binding Protein 1 (SHCBP1) in liver fibrosis, focusing on their regulatory network and interaction. Stable hepatocyte cell lines with knockdown of MCM7 and SHCBP1 were constructed and subsequently stimulated with IL-1β to mimic inflammatory conditions. RNA sequencing was conducted to identify differentially expressed genes and pathways affected by the knockdown. Our analysis revealed significant changes in gene expression profiles, particularly in pathways related to fibrosis and inflammation. We discovered that MCM7 interacts with SHCBP1, which in turn binds to RACGAP1. This interaction activates the SHCBP1-RACGAP1-STAT3 signaling axis, leading to increased IL11 expression. Elevated IL11 levels contribute to the activation of hepatic stellate cells and the progression of liver fibrosis. The findings demonstrate that MCM7 is an important regulatory factor in liver fibrosis and promotes IL11 expression through the SHCBP1-RACGAP1-STAT3 signaling pathway, driving hepatic stellate cell (HSC) activation and fibrosis progression. Overall, our findings highlight a mechanistic pathway where MCM7 and SHCBP1 collaborate to regulate IL11 expression, thereby influencing liver fibrosis progression.

overall_design: To investigate the cooperative function of MCM7 and SHCBP1 in hepatocytes with IL-1β stimulation, we established cell lines in which each target gene has been knocked down by shRNA. We then performed gene expression profiling analysis using data obtained from RNA-seq of 3 different cell lines with or without IL-1β stimulation. Comparative gene expression profiling analysis of RNA-seq data for hepatocytes and their KD derivatives (shMCM7 and shSHCBP1) under IL-1β stimulation was conducted to determine the impact of these knockdowns


title: PKM splice-switching ASOs induce upregulation of dual-specificity phosphatases and dephosphorylation of ERK1/2 in hepatocellular carcinoma

city: Cold Spring Harbor

first: Adrian

last: Krainer

state: NY

address: 1 Bungtown Rd

institute: Cold Spring Harbor Laboratory

summary: Pyruvate kinase M isoform 2 (PKM2) is preferentially expressed in nearly all cancers. It primarily functions as the last enzyme in glycolysis, but has other reported non-canonical functions, including recruiting transcription factors to oncogenes, and phosphorylating proteins. We previously described antisense oligonucleotides that disrupt alternative splicing of PKM pre-mRNA (PKM-ASOs), resulting in PKM2-to-PKM1 isoform switching in hepatocellular carcinoma (HCC), which reduces HCC growth. PKM1 has higher enzymatic activity than PKM2, which potentially drives metabolism away from macromolecule synthesis, and may explain decreased HCC growth upon PKM-ASO treatment. As PKM-ASOs also reduce PKM2 levels, how PKM splice-switching inhibits HCC cell proliferation was unclear. Here, we characterized the individual consequences of altering PKM1 or PKM2 protein levels in HCC, and observed that reducing PKM2 alone was sufficient to decrease HCC cell proliferation, whereas overexpressing PKM1 had no effect. Moreover, increasing PK activity via a small-molecule PKM2 activator had no effect on HCC cell proliferation, suggesting that PKM-ASOs affect PKM2’s non-metabolic functions. Transcriptomic and RT-qPCR analyses of HCC cells treated with PKM-ASO or PKM2-siRNA revealed upregulation of dual-specificity phosphatase 2 (DUSP2) and other related DUSPs, which act directly on ERK1/2 in the MAPK signaling pathway. Luciferase reporter assays demonstrated that PKM-ASO treatment activated the DUSP2 promoter, which correlated with decreased ERK1/2 phosphorylation. Lenvatinib is a second-line HCC therapy that indirectly reduces ERK1/2 phosphorylation, and combined treatment with PKM-ASOs inhibited proliferation of HCC cells more than either treatment alone. In summary, our results reveal a mechanism by which PKM-ASOs affect PKM2 dependency in HCC.

overall_design:
To investigate changes in transcription after altering PKM1 or PKM2 protein levels in HCC cells, we generated a dox-inducible SNU449-PKM1OE cell line and treated with either 0.25 ug/ml dox, 5 nM of siPKM2, or 60 nM of a PKM2-to-PKM1 splice switching antisense oligonucleotide (PKM-ASO) for 48 hr. We included a lipofectamine-only control, and an siCtrl. We then performed bulk RNA-Seq analysis. There were 3 biological replicates for each group.
We then used the RNA-seq data to perform comparative gene expression analysis between the treatment groups.

first: Voss

last: Dillon

first: Krainer

last: Adrian


title: Mouse placentae generated by in vitro fertilization exhibit altered gene expression, activated hypoxia responses and reduced fitness.

city: San Francisco

first: Reza

last: Oqani

state: CA

address: 513 Parnassus Ave.

institute: UCSF

summary: In this study, we utilized single-nucleus RNA sequencing to quantify alterations in the gene expression programs of mouse placentas conceived through in vitro fertilization (IVF). We identified genetic programs exhibiting both global and cell type specific differences between IVF and natural in vivo fertilization groups. Gene set enrichment analysis revealed pathways associated with parietal trophoblast giant cell differentiation and implicated in the regulation of lactation (placental lactogens), along with Hypoxia-inducible factor dependent gene expression. Importantly, IVF-derived conceptuses showed increased abortion rates when their surrogate mothers were exposed to hypoxia (10.5% O2) during pregnancy (from E7.5 to 12.5). Collectively, our findings shed light on the cellular and molecular underpinnings driving differences in pregnancy outcomes associated with these conception methods and indicate that IVF can sensitize embryos to additional stressful events, as proposed by the developmental origin of health and disease hypothesis.

overall_design: Mouse placentas were generated by transferring E3.5 blastocysts conceived either in vivo (FB) or by in vitro fertilization (IVF) into pseudopregnant recipients. On embryonic day 12.5, placentas were collected, flash-frozen, and processed for single-nucleus RNA sequencing. Nuclei were isolated with the Singulator 100, counted, and barcoded using the 10X Genomics Chromium Next GEM Single Cell 3 v3.1 kit. Libraries were sequenced on an Illumina NovaSeq 6000. Data processing included quality control, clustering, and differential gene expression analysis to compare IVF and FB placentas.

first: Emin

last: Maltepe

first: Paolo

last: Rinaudo

first: Daniel

last: Wagner


title: Aurora A Regulates the Material Property of Spindle Poles to Orchestrate Nuclear Organization at Mitotic Exit

city: Gif-sur-Yvette

first: Daan

last: Noordermeer

address: Avenue de la Terrasse

country: France

institute: Université Paris-Saclay

summary: Animal cells dismantle their nuclear envelope (NE) at the beginning and reconstruct it at the end of mitosis. This process is closely coordinated with spindle pole organization: poles enlarge at mitotic onset and reduce in size as mitosis concludes. The significance of this coordination remains unknown. Here, we demonstrate that Aurora A maintains a pole-localized protein NuMA in a dynamic state during anaphase. Without Aurora A activity, NuMA shifts from a dynamic to a solid state and abnormally accumulates at the poles, causing the segregated chromosome sets to bend around the NuMA-enriched poles. NuMA localization at the poles relies on interactions with dynein/dynactin, its coiled-coil domain, and an intrinsically disordered region (IDR). Mutagenesis experiments revealed that cation-𝜋 interactions within IDR are key for NuMA pole localization, while glutamine residues trigger the solid-state transition of NuMA upon Aurora A inhibition. We propose that maintaining the proper material properties of the spindle poles is a key step in choreographing accurate organization of the nucleus and genome post-mitosis.

overall_design: 4C-seq on the rDNA array and FLT3 gene in synchronized human HeLa cells with or without 1.5 hrs MLN8237 incubation

first: François

last: Charon


title: Human multiethnic radiogenomics reveals low-abundancy microRNA signature in plasma-derived extracellular vesicles for early diagnosis and molecular subtyping of pancreatic cancer

city: Magdeburg

first: Ulf

last: Kahlert

address: Leipziger Str. 44

department: Molekulare und Experimentelle Chirurgie

institute: Universitaetsklinikum Magdeburg

summary: Pancreatic cancer (PC) is a highly lethal malignancy, and its early diagnosis remains a clinical challenge. Extracellular vesicles (EVs) derived from patient plasma contain diverse RNA species that may serve as minimally invasive biomarkers. In this study, we performed RNA sequencing (RNA-seq) on EVs isolated from plasma samples of 85 individuals, including 65 patients with pancreatic cancer and 20 patients with benign pancreatic diseases. The benign group consisted of 10 intraductal papillary mucinous neoplasms (IPMN) and 10 chronic pancreatitis (CP) cases. Our aim was to identify differentially expressed RNA signatures in plasma-derived EVs that can distinguish pancreatic cancer from benign conditions. This dataset provides a valuable resource for biomarker discovery in liquid biopsy-based diagnosis of pancreatic cancer.

overall_design: A total of 85 plasma EV samples were analyzed using RNA-sequencing (RNA-seq), including 65 samples from patients with pancreatic cancer (PC) and 20 samples from patients with benign pancreatic conditions. Among the 20 benign cases, 10 were intraductal papillary mucinous neoplasms (IPMN) and 10 were chronic pancreatitis (CP).

first: Christoph

middle: D


title: Nociceptor to macrophage communication through CGRP/RAMP1 signaling drives endometriosis-associated pain and lesion growth

city: Boston

first: Victor

last: Fattori

state: MA

address: 1 Blackfan Circle, Karp Family Research Building, RB10006C

institute: Boston Children's Hospital

summary: Endometriosis is a debilitating and painful gynecological inflammatory disease affecting approximately 15% of women. Current treatments are ineffective for a significant fraction of patients, underscoring the need for new medical therapies with long-term benefits. Given the relevance of neuroimmune communication in different disease outcomes, we investigated the role of CGRP-mediated neuroimmune communication in endometriosis. We found that mouse and human endometriosis lesions contained CGRP and RAMP1. In mice, nociceptor ablation reduced pain, monocyte recruitment, and lesion size, suggesting that nociceptor activation and neuropeptide release contribute to endometriosis lesion growth and pain. In vitro, CGRP-induced pro-endometriosis macrophages (PEMs) showed impaired efferocytosis and supported endometrial cell growth in a RAMP1-dependent manner. Treatment with FDA-approved drugs that block CGRP-RAMP1 signaling reduced mechanical hyperalgesia, spontaneous pain, and lesion size in mice. Altogether, our data demonstrates the effectiveness, cellular mechanisms and pre-clinical safety of non-hormonal and non-opioid CGRP/RAMP1 blocking therapies, which may lead to clinical benefit for endometriosis patients.

overall_design: Peritoneal cavity washes from sham littermate controls (no lesions) and lesion-bearing TRPV1-creDTA and littermate control mice were collected at 28 days post endometriosis (dpi) induction

first: Michael

middle: S

last: Rogers


title: Loss of tumor cell MHC Class II drives MAPK-inhibitor insensitivity of BRAF-mutant anaplastic thyroid cancers

first: Richard

last: Koche

institute: Memorial Sloan Kettering Cancer Center

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

overall_design: Refer to individual Series


title: Loss of tumor cell MHC Class II drives MAPK-inhibitor insensitivity of BRAF-mutant anaplastic thyroid cancers [ATAC-seq]

summary: Cancer cells present neoantigens dominantly through MHC class I (MHCI) to drive tumor rejection through cytotoxic CD8+ T-cells. There is growing recognition that a subset of tumors express MHC class II (MHCII), causing recognition of antigens by TCRs of CD4+ T-cells that contribute to the anti-tumor response. We find that mouse BrafV600E-driven anaplastic thyroid cancers (ATC) respond markedly to the RAF + MEK inhibitors dabrafenib and trametinib (dab/tram) and that this is associated with upregulation of MhcII in cancer cells and increased CD4+ T-cell infiltration. A subset of recurrent tumors lose MhcII expression due to silencing of Ciita, the master transcriptional regulator of MhcII, despite preserved interferon gamma signal transduction, which can be rescued by EZH2 inhibition. Orthotopically-implanted Ciita-/- and H2-Ab1-/- ATC cells into immune competent mice become unresponsive to the MAPK inhibitors. Moreover, depletion of CD4+, but not CD8+ T-cells, also abrogates response to dab/tram. These findings implicate MHCII-driven CD4+ T cell activation as a key determinant of the response of Braf-mutant ATCs to MAPK inhibition.

overall_design: To probe into the underlying mechanisms of MhcII loss and attenuated Ciita expression we performed bulk RNA-seq and ATAC-seq. We tested mouse cell lines generated from thyroid tumors derived from a genetic engineered anaplastic thyroid cancer mouse model (Tpo-Cre/eYFP/BRaf-CA/Trp53fl/fl; termed Braf-CAV600E/p53). We included one primary (B92) and one recurrent (B36934) cell line after a 96h treatment with DMSO, IFNγ (20ng/ml), trametinib (10nM) or the combination of IFNγ and trametinib in vitro.

first: Vera

last: Tiedje

first: James

last: Fagin


title: Faithful Modeling of Terminal CD8 T Cell Dysfunction and Epigenetic Stabilization In Vitro [ATAC-seq]

first: Hazem

last: Ghoneim

overall_design: ATAC-seq profiling of D7 Effector, D19 Acute-2d, D19 Acute-7d, D19 Dysfunctional P14 CD8 T cells with 2 biological replicates per group.

first: Amir

last: Yousif

first: Abbey

last: Saadey

first: Ava

last: Lowin

first: Ankita

last: Saini


title: Metabolite enhanced perfusion solution improves kidney transplantation viability

city: Hamburg

first: Fabian

last: Hausmann

address: Falkenried 94

department: Institute of Medical Systems Biology

institute: University Medical Center Hamburg-Eppendorf

summary: Normothermic machine perfusion (NMP) has become a valuable tool to expand the pool of transplantable organs. However, the application of NMP to kidneys presents substantial challenges, mostly due to high variability in the configuration of perfusion buffers. Here we introduce a multimodal atlas of kidney cell injury associated with NMP using a literature-based consensus buffer. This resource provided the foundation for a systematic cross-species framework with integrated metabolic, genomic, spatial proteomic, and histopathologic profiling that was used to re-engineer the consensus perfusion buffer, mitigating cellular damage ex vivo and in vivo, thus extending organ viability and improving adequacy for transplantation.

sample_organism:
Sus scrofa
Mus musculus
Rattus norvegicus

overall_design: Perfused and unperfused snRNAseq samples from Mouse (3 perfused/3 unperfused), Pig (2 perfused/2 unperfused) and Rat (3 perfused / 3 unperfused).

first: Jan

last: Czogalla

first: Sonja

last: Hänzelmann


title: Loss of tumor cell MHC Class II drives MAPK-inhibitor insensitivity of BRAF-mutant anaplastic thyroid cancers [RNA-seq]


title: Integrator complex bridges TFIID and RNA polymerase II to maintain pluripotency during development (RNA-Seq)

city: MIAMI

first: Raghu

middle: Ram

last: Edupuganti

state: FL

address: 1501 NW 10th Ave

department: Human Genetics

institute: University of Miami

summary: The mechanisms by which the expression of pluripotency and Polycomb networks are harmonized to allow the transition from the pluripotency to a differentiated state has not been fully elucidated. Integrator complex regulates transcription pause release and RNA processing in metazoans. Here, we show that acute depletion of INTS11 in mouse embryonic stem cells or its conditional deletion in mouse results in loss of stemness and block of early embryonic development, respectively. We show that Integrator plays a role in stemness and cellular differentiation by bridging the association of RNA polymerase II (RNAPII) and basal transcription factor (TFIID). While the catalytic endonuclease activity is required to enhance cellular reprogramming, depletion of INTS11 diminish RNAPII recruitment to promoters and super-enhancers of pluripotency and Polycomb genes leading to their repression. We place Integrator at pre-initiation complex formation linking the pluripotency and Polycomb pathways to ensure orderly cellular differentiation during development.

overall_design:
To determine how acute INTS11 loss affects RNAPII activity in mouse ESCs , we generated INTS11 degron ESCs. Upon removal of Trimethoprim drug to induce INTS11 degradation for 0h, 3h, 6h and 24h, samples were isolated at indicated timepoints.
RNA-seq was performed to identify gene expression changes upon INTS11 depletion.

first: Pradeep

last: Cingaram

first: Saeid

last: Parast

first: Felipe

last: Beckedorff

first: Jingyin

last: Yue

first: Helena

middle: G

last: Dos Santos

first: Monica

last: Valencia

first: Mingjiang

last: Xu

first: Brooke

last: DeRosa

first: Derek

last: Dykxhoorn

first: Lluis

last: Morey

first: Ali

last: Shilatifard

first: Ramin

last: Shiekhattar


title: Integrator complex bridges TFIID and RNA polymerase II to maintain pluripotency during development (ChIP-Seq)

overall_design:
Chromatin immunoprecipitation and DNA-sequencing of INTS11, HA-INTS11, INTS12, RING1B, RNA Polymerase II, OCT4, SOX2, IgG, and Input.
Chromatin immunoprecipitation and DNA-sequencing of TAF1 and TBP.

last: Ram  Edupuganti

last: Reddy Cingaram

last: Mohammad  Parast

first: Gary

last: LeRoy

first: Sadat

last: Dokaneheifard

last: Gomes  Dos Santos

first: Omar

last: Lopez Ocejo

last: M Dykxhoorn

first: Guilherme

last: Lavezzo

first: Daniel

last: Bilbao

first: Danny

last: Reinberg


title: Integrator complex bridges TFIID and RNA polymerase II to maintain pluripotency during development (PRO-Seq)

first: Raghu

last: Edupuganti

overall_design: To determine how acute INTS11 loss affects RNAPII activity in mouse ESCs, we generated INTS11 degron ESCs. Upon removal of drug to induce INTS11 degradation for 0h, 1h, 3h and 6h, samples were isolated at indicated timepoints.  Nuclear run-on assays (PRO-seq) were performed with ten million purified nuclei per sample.

first: Pradeep

last: Cingaram

first: Saeid

last: Parast

last: Beckedorff

last: Yue

last: Dos Santos

last: Valencia

last: Xu


title: TFAP2A orchestrates gene regulatory networks and tubular architecture in kidney outer medullary collecting ducts.

city: Hannover

first: Karen

middle: I.

last: Lopez-Cayuqueo

address: Carl-Neuberg-Str.1,30625 Hannover

department: Department of Nephrology and Hypertension

institute: Hannover Medical School (MHH)

summary: Mutations of the transcription factor TFAP2A are associated with congenital anomalies of the kidney and urinary tract in humans. In mice, deficiency of its ortholog Tfap2a in collecting duct cells causes widened epithelial tubules in the outer medulla of the adult kidney, but the molecular mechanisms of this phenotype are unknown. In this study, we identified gene regulatory networks controlled by Tfap2a in mouse kidney collecting ducts by combining gene knockout and transcriptomics. Integrated analyses of single-nucleus and bulk RNA-sequencing data from kidneys of adult Tfap2a knockout and control mice indicated deregulated expression of genes associated with cell adhesion and Wnt signaling pathways. Validation studies revealed that Tfap2a controls Wnt9b and Alcam, known regulators of kidney tubule morphogenesis. Our data provide novel insights into kidney epithelial gene regulatory networks controlled by Tfap2a and its potential functional role in congenital renal disease.

overall_design: Mouse kidneys (single nucleus preparation) of 12-week-old male Hoxb7Cre+;Tfap2afl/fl (n=2) and control mice (n=2)

first: Janna

last: Leiz

middle: I

last: López-Cayuqueo

first: Shuang

last: Cao

first: Louisa

last: Gerhardt

first: Christian

last: Hinze

first: Kai

last: Schmidt-Ott


title: The effects of Cryopreservation on PBMCs transcriptome profile

status: Public on Aug 07 2025

city: Frederick

first: Tomozumi

last: Imamichi

state: MD

address: Frederick National Laboratory for Cancer Research

department: Applied and Developmental Research Directorate

institute: National Institutes of Health

summary: Cryopreservation is a key method for long-term storage of biological specimens. The development of optimal cryopreservation condition will reduce the damage from the storage as least as possible. The effect of the cryopreservation condition we used on peripheral blood mononuclear cells (PBMCs) has been previously evaluated with microarray analysis. The emerging single-cell RNA sequencing (scRNA-seq) technology enable deeper exploring cellular heterogeneity and function. In the current study, we further optimized the cryopreservation procedure using FACS analysis, and the method were further evaluated by ScRNA-Seq for two different length of storage time: six months and 12 months in comparison to fresh cells. We identified major immune cell types from both fresh and recovered cryopreserved PBMCs, including monocytes, dendritic cells (DCs), natural Killer (NK) cells, CD4+ T cells, CD8+ T cells, and B cells. The cell viability of all identified immune cell types was relatively stable during the initial 6-month of cryopreservation, while showed10 ~ 40 % decline after cryopreserved for 12 months for different cell types. Furthermore, transcriptome profile of cryopreserved samples did not show obvious perturbation during whole 12 months testing time, although a few key genes involved in stress response or apoptosis, exhibited significant change, especially in monocytes, DC and NK cells. Our results validate that the optimized cryopreservation is a reliable procedure for maintaining these major immune cell types in PBMCs, also highlight the importance of careful assessment of individual sample as variation could be introduced by cryopreservation.

overall_design: Fresh and thawed PBMCs samples from three donors were analyzed by FACS, and scRNA-seq to study the impact of cryopreservation on cell viability and transcriptome profile.

first: Xuan

last: Li

first: Qian

last: Chen

first: Jeanette

last: Higgins

first: Kendal

last: Cook

first: Michael

middle: W

last: Baseler

first: Weizhong

last: Chang


title: TUMOR-INFILTRATING NOCICEPTOR NEURONS PROMOTE IMMUNOSUPPRESSION [MDSC]

city: Kingston

first: Sebastien

last: Talbot

state: ON

address: 18 Stuart st

department: DBMS

institute: Queen's University

summary: Nociceptor neurons impact tumor immunity. Removing nociceptor neurons reduced myeloid-derived suppressor cell (MDSCs) tumor infiltration in mouse models of head and neck carcinoma and melanoma. Carcinoma-released small extracellular vesicles (sEVs) attract nociceptive nerves to tumors. sEV-deficient tumors fail to develop in mice lacking nociceptor neurons. Exposure of dorsal root ganglia (DRG) neurons to cancer sEVs elevated expression of Substance P, IL-6 and injury-related neuronal markers while treatment with cancer sEVs and cytotoxic CD8 T-cells induced an immunosuppressive state (increased exhaustion ligands and cytokines). Cancer patient sEVs enhanced DRG responses to capsaicin, indicating increased nociceptor sensitivity. Conditioned media from DRG and cancer cell co-cultures promoted expression of MDSC markers in primary bone marrow cells while DRG conditioned media together with cancer sEVs induced checkpoint expression on T-cells. Our findings indicate that nociceptor neurons facilitate CD8+ T cell exhaustion and enhance MDSC infiltration. Targeting nociceptor-released IL-6 emerges as a novel strategy to disrupt harmful neuro-immune interactions in cancer and enhance anti-tumor immunity.

overall_design: Eight-week-old female TRPV1Cre::DTAfl/wt mice, in which nociceptor neurons are genetically ablated (n = 4), and their TRPV1Cre::DTAwt/wt littermate controls (n = 3) each received 2 × 10^5 B16F10-OVA-mCherry melanoma cells suspended in 50 µL PBS, injected subcutaneously into the right flank. Fourteen days later the mice were euthanized and the tumors were harvested. The tumors were finely minced and digested for 40 min at 37 °C in DMEM supplemented with 5 % FBS, collagenase D (2 mg mL⁻¹), collagenase IV (1 mg mL⁻¹) and DNase I (40 µg mL⁻¹). The resulting suspension was purified on a 70 %/40 % Percoll gradient, passed through a 70-µm strainer and resuspended in PBS containing 2 % FBS and 2 mM EDTA, while lymph nodes were processed in parallel by mechanical dissociation and filtration.  Single-cell suspensions were stained on ice and sorted on a BD FACSAria II. Non-viable cells were excluded with Zombie Aqua (BioLegend 423102), and Fc receptors were blocked with anti-CD16/32 (BD 553141, 0.5 mg mL⁻¹, 15 min, 4 °C). For lymphoid analyses, cells were labeled with CD45-BV421 (clone 30-F11, 1:100) and CD3ε-FITC (clone 145-2C11, 1:100); for myeloid analyses, CD45-BV421 was combined with CD3ε-PerCP-Cy5.5 (clone 145-2C11, 1:100). Intrinsic mCherry fluorescence identified melanoma cells. Tumor-infiltrating lymphocytes were further resolved with CD4-PerCP-Cy5.5 (clone GK1.5, 1:100), CD8α-Alexa Fluor 700 (clone 53-6.7, 1:100) and CD25-PE (clone PC61.5, 1:100), defining CD8⁺ T cells as Zombie⁻ CD45⁺ CD3⁺ CD8α⁺ CD4⁻ CD25⁻ and regulatory T cells as Zombie⁻ CD45⁺ CD3⁺ CD4⁺ CD25^hi. Myeloid subsets were distinguished with CD11b-APC-Cy7 (clone M1/70, 1:100), Gr-1-APC (clone RB6-8C5, 1:100), CD64-PE-Cy7 (clone X54-5/7.1, 1:100), MHC II-FITC (clone M5/114.15.2, 1:100) and CD11c-PE (clone N418, 1:100), gating myeloid-derived suppressor cells as Zombie⁻ CD45⁺ CD3⁻ CD11b⁺ Gr-1⁺ MHC II⁻ and antigen-presenting cells as Zombie⁻ CD45⁺ CD3⁻ CD11c⁺ MHC II⁺ CD11b⁻ Gr-1⁻. Melanoma cells were sorted as Zombie⁻ mCherry⁺ CD45⁻ and stromal cells as Zombie⁻ mCherry⁻ CD45⁻. Fluorescence-minus-one controls set all gates, and doublets were excluded by FSC-A/FSC-H and SSC-A/SSC-H discrimination.  Six purified populations—CD8⁺ T cells, regulatory T cells, myeloid-derived suppressor cells, antigen-presenting cells, mCherry⁺ melanoma cells and mCherry⁻/CD45⁻ stromal cells—yielded 1.5 × 10^3 to 1.8 × 10^5 cells per library. Sorted cells were collected directly into 500 µL TRIzol reagent (Invitrogen 15596026) and stored at −80 °C until RNA extraction according to established protocols. RNA quality assessed on an Agilent Bioanalyzer yielded RNA Integrity Numbers of 8.2–10. Poly-A-selected libraries were prepared with NEBNext Ultra II; a total of 48 libraries (eight per population) were constructed at Fulgent Genetics and sequenced on an Illumina HiSeq 4000 in 75-cycle single-end mode (project IEM-048-02, 10 May 2020), generating approximately 20 million reads per sample. Base calling used Illumina RTA 2.4.11, and demultiplexing was performed with bcl2fastq 2.20 (one index mismatch allowed). Adapter trimming and quality filtering were carried out with Trimmomatic, and high-quality reads were aligned to the GRCm38 mouse genome with STAR v2.5.1, which also generated gene-level counts. Differential expression analysis in DESeq2 used normalized counts; genes with an adjusted P value (false-discovery rate) < 0.05 were considered differentially expressed. Log2 fold changes and −log10 P values were calculated from the normalized data, and all downstream analyses and visualizations were performed in RStudio.

first: Mohammad

last: Balood

first: Amin

last: Nikpoor


title: Single-Cell Transcriptomics Shows Cellular Heterogeneity, Intercellular Communication, and Extracellular Matrix Remodeling in Corneal Fibrosis In Vivo

city: Columbia

first: Rajiv

last: Mohan

state: Missouri

address: 1520 East Rollins University of Missouri

department: Department of Veterinary Medicine and Surgery

institute: University of Missouri

summary: Alkali burns cause rapid corneal remodeling, which is characterized by fibrosis, neovascularization, and immune infiltration; however, the cellular dynamics driving these responses remain poorly understood. Here, we present the first single-cell transcriptomic atlas of the rabbit cornea on day 14 postalkali injury, capturing the cellular and molecular architecture of stromal fibrosis. Single-cell RNA sequencing of naïve and injured corneas identified 14 transcriptionally distinct populations, including keratocytes, EMT-like epithelial cells, immune infiltrates, and heterogeneous fibroblast subtypes. Injury induced the expansion of ECM-remodeling fibroblasts and immune cells, accompanied by widespread transcriptional reprogramming. Stromal subclustering revealed four fibroblast states—quiescent, activated, proliferative, and progenitor-like—each with distinct gene signatures and functions. Pseudotime analysis revealed a trajectory from proliferative fibroblasts to ECM-secreting myofibroblasts, marked by stage-specific activation of mitotic, matrix, and contractile programs. Ligand–receptor inference via CellChat identified coordinated signaling through the TGFβ, PDGF, VEGF, and FGF pathways, alongside lesser-known axes such as NAMPT and NECTIN, implicating stromal–vascular crosstalk and metabolic stress in fibrotic progression. This study provides a comprehensive cellular framework for alkali-induced corneal fibrosis and identifies actionable targets for antifibrotic and antiangiogenic intervention in ocular surface disease.

overall_design: This study employed a rabbit model of alkali-induced corneal fibrosis, where six New Zealand White rabbits underwent controlled corneal wounding, followed by longitudinal clinical imaging and intraocular pressure monitoring. Corneal tissues were collected 14 days post-injury and processed for histology, immunofluorescence, and scRNA-seq. scRNA-seq libraries were prepared using the 10x Genomics Chromium platform and sequenced on an Illumina NovaSeq. Raw data were processed using Cell Ranger, and downstream analyses were performed with Seurat, Harmony, Slingshot, tradeSeq, and CellChat for clustering, batch correction, trajectory inference, cell–cell communication, and gene expression dynamics. Functional enrichment was conducted via clusterProfiler using human orthologs.

first: Rajnish

last: Kumar

first: Nishant

last: Sinha


title: The role of CD61 in pancreatic cancer

city: London

first: Huafu

address: 237 Fulham Road, Fulham Road

country: United Kingdom

institute: Institute of Cancer Research- Fulham Road (ICR)

summary: We performed RNA-seq analysis on Cd61 wild-type and CRISPR/Cas9-mediated Cd61 knock out pancreatic tumour organiod from Pdx1-Flp;KrasFSF-G12D;Trp53frt/frt; Vim-EGFP (KPFV) mice. We compared the transcriptome between these two groups of them with the aim to understand their molecular signatures and the relevance to human pancreatic ductal adenocarcinoma subtypes.

overall_design: Cd61 wild-type compare to Cd61 knock out pancreatic tumour organiod

first: Linxiang

last: Lan

first: Axel

last: Behrens


title: Effect of overexpression of aging-associated transcription factors on epithelial cells.

city: Calgary

first: Myriam

last: Hemberger

address: 3330 Hospital Dr NW

institute: University of Calgary

summary: RNA-seq profiling of whole, intact uteri at embryonic day 3.5 (E3.5) from young and aged virgin wild-type C57BL/6 mice—each mated with vasectomized wild-type males—identified the transcription factors Foxc1 and Six1 as aging-associated markers. Immunostaining of mouse uteri across an age range of 8 to 41 weeks revealed a progressive upregulation of FOXC1, initially localized to individual stromal cells in the endometrium, followed by prominent expression in both luminal and glandular epithelial cells. To investigate the functional role of Foxc1 and Six1, their full-length open reading frames were cloned into a PiggyBac expression vector, sequence-verified, and overexpressed in Ishikawa cells. Phenotypic characterization and RNA-seq analysis of these cells provided a framework for understanding FOXC1-mediated aging-related effects.

overall_design: RNA-seq profiling of Ishikawa cells (human endometrial adenocarcinoma cell line) overexpressing Foxc1, Six1, alone and in combination. Ishikawa cells treated with / without progesterone-estrodiol-cyclic AMP were processed for RNA-sequencing analysis.

first: Anshul

last: Jadli

first: Aleksandra

middle: O

last: Tsolova

first: Alexa

last: Krala

first: Wendy

last: Dean


title: TUMOR-INFILTRATING NOCICEPTOR NEURONS PROMOTE IMMUNOSUPPRESSION

overall_design: Isolation of CD8+ T cells Six-to-eight-week-old male and female naïve mice were euthanized, and their spleens were collected in ice-cold PBS (5% FBS) and mechanically dissociated. The cells were strained (70 μm), RBC lysed (Life Technologies, A1049201; 2 min), and counted using a haemocytometer. Total CD8+ T cells were magnet sorted (Stem Cell, 19853A) and cultured (DMEM + FBS 10%, penicillin–streptomycin 1% + nonessential amino acids (Corning, 25-025-Cl) + vitamin + β-mercaptoethanol (Gibco, 21985-023) + l-glutamine (VWR, 02-0131) + sodium pyruvate (Corning, 25-000-Cl)). Cell purity was systematically confirmed after magnet sorting and the numbers of CD8+CD62Lhi were immunophenotyped by flow cytometry. To generate cytotoxic T lymphocytes, 2 × 105 CD8+ T cells were seeded and stimulated for 48 h under Tc1 inflammatory conditions (2 μg ml−1 plate bounded anti-CD3 and anti-CD28 (Bio X Cell, BE00011, BE00151) + 10 ng ml−1 rIL-12 (BioLegend, 577008) + 10 μg ml−1 of anti-IL-4 (Bio X Cell, BE0045). Neuro culture. Naïve TRPV1cre::tdTomatofl/WT mice were euthanized, and their dorsal root ganglia (DRG) were immediately transferred into DMEM (Corning, 10 013 CV) supplemented with 50 U ml−1 penicillin, 50 μg ml−1 streptomycin (Corning, MT 3001 Cl) and 10 % fetal bovine serum (Seradigm, 3100). The tissue was dissociated for 80 min at 37 °C in HEPES buffered saline (Sigma, 51558) containing 1 mg ml−1 collagenase IV (Sigma, C0130) and 2.4 U ml−1 dispase II (Sigma, 04942078001). After digestion, ganglia were gently triturated through glass Pasteur pipettes of decreasing diameter, centrifuged through a 10 % BSA cushion and plated onto laminin coated (Sigma, L2020) culture dishes. Approximately 4 × 104 neurons per dish were maintained for 48 h in Neurobasal A medium (Gibco, 21103 049) supplemented with 0.05 ng μl−1  nerve growth factor (Life Technologies, 13257 019), 0.002 ng μl−1 glial cell line derived neurotrophic factor (PeproTech, 450 51 10), 0.01 mM cytosine β D arabinoside (Sigma, C6645), 200 mM L glutamine (VWR, 02 0131) and B 27 (Gibco, 17504044).' Neurons were then co-cultured at a 1:10 ratio with mEERL derived small extracellular vesicles, cytotoxic CD8+ T-cells or both, each condition prepared in T cell medium supplemented with the same concentrations of NGF and GDNF to support neuronal viability. After 48 h, cells were detached with a scraper, washed in flow cytometry buffer (PBS containing 2 % FBS and 1 mM EDTA), passed through a 70 μm filter, and TRPV1+ neurons (FSChigh SSChigh tdTomatohigh) were isolated by fluorescence activated cell sorting directly into TRIzol. RNA extraction, library construction and sequencing were performed at the IRIC Genomics Platform (Université de Montréal). RNA integrity was confirmed with an Agilent Bioanalyzer (RIN > 7.5). Poly adenylated transcripts were captured and libraries prepared with the KAPA mRNA HyperPrep Kit (Kapa Biosystems, KR1352). Bar coded libraries were sequenced on an Illumina NextSeq 500 using 75 cycle single end reads. Adapter trimming and low quality base removal were carried out with Trimmomatic v0.35, and reads were aligned to the mouse reference genome GRCm38 (Gencode M25, Ensembl 100) with STAR v2.7.1. Gene level counts were generated with STAR and RSEM, normalized to fragments per kilobase per million mapped reads, and differential expression was assessed with DESeq2. Genes with a false discovery rate–adjusted P value below 0.20 were considered differentially expressed. Downstream analyses and visualization were performed in RStudio or Microsoft Excel.

first: Maryam

last: Ahmadi


title: Plaque-induced transcriptional reprogramming of microglia towards lymphoid receptor expression protects against Alzheimer’s disease. [RNA-seq]

city: New York

first: Pinar

last: Ayata

address: 1425 Madison Ave

department: Neuroscience

institute: Icahn School of Medicine at Mt Sinai

summary: Microglia, the innate immune cells of the brain, play a defining role in the progression of Alzheimer’s disease (AD). The microglial response to amyloid plaques can be either neuroprotective or neurotoxic. Here we show that the protective function of microglia is associated with and functionally linked to a transcriptional shift toward lymphoid gene expression. Microglial contact with the plaques induces downregulation of PU.1, a lineage-specifying transcription factor. PU.1 downregulation plays a key role in mitigating the severity of AD, as evidenced by a reduction in neurotoxic microglia activation, preservation of neuronal connections, maintenance of cognitive function, and increased survival in mice with genetically reduced PU.1 expression in microglia. The neuroprotective effect of PU.1 downregulation is linked to the de-repression of lymphoid-specific proteins, particularly CD28—a surface protein critical for T cell activation. Microglia-specific deficiency in CD28, which is expressed exclusively in a small subset of plaque-associated PU.1low microglia, promotes an inflammatory microglial state and associated increase in amyloid plaque load. Our findings suggest that PU.1low lymphoid/CD28-expressing microglia that emerge in response to AD pathology exert a suppressive, anti-inflammatory effect during AD. This novel role of CD28 -and potentially other lymphoid co-stimulatory or co-inhibitory receptor proteins- in governing microglia responses in AD points to possible new immunotherapy approaches for AD treatment.

overall_design: Microglial RNAs were isolated from human HPSC-induced microglia (iMgls)

first: Jessica

last: Crowley

first: Matthew

last: Challman

first: Vinaya

last: Sahasrabuddhe

first: Maud

last: Gratuze

first: Sebastian

last: Werneburg

first: Diogo

last: Ribeiro

first: Emma

last: Hays

first: Violeta

last: Durán-Laforet

first: Travis

last: Faust

first: Philip

last: Hwang

first: Francisco

last: Lopes

first: Chrysa

last: Nikopoulou

first: Sarah

last: Buchholz

first: Robert

last: Murphy

first: Taoyu

last: Mei

first: Anna

last: Pimenova

first: Carmen

last: Romero-Molina

first: Francesca

last: Garretti

first: Tulsi

last: Patel

first: Claudia

last: DeSanctis

first: Angie

last: Ramirez

first: Megan

last: Crow

first: Felix

last: Weiss

first: Jason

last: Ulrich

first: Edoardo

last: Marcora

first: John

last: Murray

last: Meissner

first: Andreas

last: Beyer

first: Dan

last: Hasson

last: Crary

first: Dorothy

last: Schafer

first: David

last: Holtzman

first: Alison

last: Goate

first: Alexander

last: Tarakhovsky

first: Anne

last: Schaefer


title: MYOD represses gene expression from non-Ebox motifs

city: La jolla

first: Chiara

last: Nicoletti

address: 10901 North Torrey Pines Road

department: Development, Aging and Regeneration Program

institute: Sanford Burnham Prebys Medical Discovery Institute

summary: We report on the identification of a previously unrecognized property of MYOD as repressor of gene expression, via E-box-independent chromatin binding, during the process of somatic cell trans-differentiation into skeletal muscle. When ectopically expressed in proliferating human fibroblasts or endogenously induced in activated muscle stem cells (MuSCs) MYOD was detected at accessible regulatory elements of expressed genes, invariably leading to reduced chromatin accessibility and gene repression. At variance with conventional E-box-driven increased chromatin accessibility and H3K27 acetylation at previously silent loci of MYOD-activated genes, MYOD-mediated chromatin compaction and repression of transcription was associated with high occurrence of non-E-box motifs, did not lead to reduced levels of H3K27ac and coincided with reduced levels of H4 acetyl-methyl lysine modification (Kacme). Using MYOD mutants we dissected the molecular mechanism of MYOD-mediated repression, whereby repression of mitogen- and growth factor-responsive genes occurred via promoter binding, which requires a conserved domain within the first helix; conversely, repression of cell-of-origin/alternative lineage genes occurred via binding and decommissioning of distal regulatory elements, such as super-enhancers (SE), it required either the N-terminal activation domain or the two chromatin-remodeling domains, and it coincided with reduced strength of CTCF-mediated chromatin interactions. These data extend MYOD biological properties beyond the current dogma that restricts MYOD function to a monotone transcriptional activator. They also reveal an unprecedented functional versatility arising from alternative chromatin recruitment through E-box or non-E-box sequences, whereby genetic determinants dictate differential usage of MYOD functional domains.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Integration of RNA-seq, ATAC-seq, CUT&RUN, ChIP-seq, Hi-C, H3K27ac and CTCF HiChIP experiments to characterize the 3D regulatory mechanisms for the myogenic conversion of fibroblasts. For this project we used human Hi-C IMR90 rep1 (GSM2597682), Hi-C IMR90 rep2 (GSM2597683), Hi-C IMR90/MyoD rep1 (GSM2597684), Hi-C IMR90/MyoD rep2 (GSM2597685), ChIP-seq H3K27ac IMR90 (GSM3938576), ChIP-seq Input IMR90 (GSM3938577). Integration of RNA-seq, ATAC-seq and CUT&RUN experiments to characterize MyoD regulatory mechanisms during muscle stem cell activation and progression towards terminal differentiation. Mouse RNA-seq and ATAC-seq data from GSE189074, MyoD ChIP-seq from GSE56077.

first: Luca

last: Caputo

first: Jimmy

last: Massenet

first: Leah

last: Connor

first: Matthew

last: Simon

first: Pier Lorenzo

last: Puri


title: Plaque-induced transcriptional reprogramming of microglia towards lymphoid receptor expression protects against Alzheimer's disease

first: Pinar

last: Ayata

summary: We have utilized a single nucleus sequencing approach using 10x Genomics snRNA-seq to understand the role of PU.1 expression level in the determination of microglia activation states relavent to Alzheimer's disease. Microglia have been shown to have both neuroprotective and neurotoxic functions during disease, however it remains to be understood how these states are regulated. This study provides new insights into the protective function of microglia mediated by a transcriptional shift toward lymphoid gene expression.

overall_design: Lineage traced microglia nuclei (Cx3cr1CreErt2/+(Litt);Eef1a1LSL.eGFPL10a/+) from 5xFAD, 5xFAD; PU.1-low, 5xFAD; PU.1-high, and 5xFAD with CSF1R inhbition were sorted using fluorescence-activated nucleus sorting (FANS) and analyzed using snRNA-seq.

first: Jessica

last: Crowley

last: Challman


title: Plaque-induced transcriptional reprogramming of microglia towards lymphoid receptor expression protects against Alzheimer's disease [scRNA-seq]

summary: Amyloid plaque associated microglia induce a transcriptional state characterized by downregulation of the transcription factor Spi1 and upregulation of a subset of canonically lymphoid-specific signaling genes. Knockout of one such gene, Cd28, in microglia exacerbates interferon-responses in response to amyloid pathology.

overall_design: Wild type, 5xFAD, and 5xFAD; Cd28-cKO (microglia specific KO generated by breeding 5xFAD, Cd28fl/fl mice to Cx3cr1creERT2/+ mice and gavaged 6x with tamoxifen) were aged to 3, 6, or 8 months of age. Microglia were isolated by removing forebrain, homogenizing tissue, preparing single cell suspensions using Percoll, and sorting microglia using fluorescent antibodies against CD45 and CD11b (collecting CD45-intermediate, CD11B+ cells as microglia). 10,000 cells per sample were submitted for 10X Single Cell RNA-sequencing.

first: Vinaya

last: Sahasrabuddhe


title: Plaque-induced transcriptional reprogramming of microglia towards lymphoid receptor expression protects against Alzheimer’s disease

first: BiNGS

last: Core

address: 1470 Madison Ave

institute: Icahn School of Medicine at Mount Sinai

overall_design: ATAC-sequencing of FACS isolated microglia from control, PU.1 knockdown, and PU.1 over-expressing mice, FACS isolated T cells from control mice

middle: F

first: Maud

last: Gratuze

first: Sebastian

last: Werneburg

first: Diogo

last: Ribeiro

first: Emma

last: Hays

first: Violeta

last: Durán-Laforet

first: Travis

middle: E

last: Faust

first: Philip

last: Hwang

first: Francisco

last: Mendes Lopes

first: Chrysa

last: Nikopoulou

first: Sarah

last: Buchholz

first: Robert

last: Murphy

first: Taoyu

last: Mei

first: Anna

last: Pimenova

first: Carmen

last: Romero-Molina

first: Francesca

last: Garretti

first: Tulsi

last: Patel

first: Claudia

last: De Sanctis

first: Angie

middle: V

last: Ramirez Jimenez

first: Megan

last: Crow

first: Felix

last: Weiss

first: Jason

last: Ulrich

first: Edoardo

last: Marcora

first: John

last: Murray

last: Meissner

last: Beyer

last: Hasson


title: TFAP2A orchestrates gene regulatory networks and tubular architecture in kidney outer medullary collecting ducts [bulk RNA-seq]

first: Karen

last: Lopez-Cayuqueo

overall_design: Bulk RNAseq from whole kidney samples from Hoxb7Cre+;Tfap2afl/fl or control mice (n = 4 per group).

first: Janna

last: Leiz

last: López-Cayuqueo

first: Shuang

last: Cao

first: Louisa

last: Gerhardt

first: Christian

last: Hinze

first: Kai

last: Schmidt-Ott


title: Loss of Free Fatty Acid Receptor 2 leads to impaired islet mass and beta cell survival

city: Chicago

first: Brian

middle: T

last: Layden

state: Illinois

address: 835 S Wolcott

department: Division of Endocrinology, Diabetes & Metabolism, Department of Medicine,

institute: University of Illinois at Chicago

summary: The regulation of pancreatic β cell mass is a critical factor to help maintain normoglycemia during insulin resistance. Nutrient-sensing G protein-coupled receptors (GPCR) contribute to aspects of β cell function, including regulation of β cell mass. Nutrients such as free fatty acids (FFAs) contribute to precise regulation of β cell mass by signaling through cognate GPCRs, and considerable evidence suggests that circulating FFAs promote β cell expansion by direct and indirect mechanisms. Free Fatty Acid Receptor 2 (FFA2) is a β cell-expressed GPCR that is activated by short chain fatty acids, particularly acetate. Recent studies of FFA2 suggest that it may act as a regulator of β cell function. Here, we set out to explore what role FFA2 may play in regulation of β cell mass. Interestingly, Ffar2(-/-) mice exhibit diminished β cell mass at birth and throughout adulthood, and increased β cell death at adolescent time points, suggesting a role for FFA2 in establishment and maintenance of β cell mass. Additionally, activation of FFA2 with Gαq/11-biased agonists substantially increased β cell proliferation in in vitro and ex vivo proliferation assays. Collectively, these data suggest that FFA2 may be a novel therapeutic target to stimulate β cell growth and proliferation.

overall_design: Islet from normal chow fed 10-12 weeks global FFA2 KO and WT mice were harvested. Islets from two mice were pool as one sample. Total RNA is extracted and sequenced.

first: Stephanie

last: Villa

first: Medha

last: Priyadarshini

first: Miles

middle: H

last: Fuller

first: Tanya

last: Bhardwaj

first: Michael

last: Brodsky

first: Anthony

last: Angueira

first: Rockann

last: Mosser

first: Bethany

last: Carboneau

first: Sarah

last: Tersey

first: Helena

last: Mancebo

first: Annette

last: Gilchrist

first: Raghavendra

last: Mirmira

first: Maureen

last: Gannon


title: Intestinal mast cell-derived leukotrienes mediate the anaphylactic response to ingested antigens

city: Seoul

first: Jaechul

last: Lim

state: CT

address: 1 Gwanak-ro, 1 Gwanak-gu

institute: Seoul National University

summary: Anaphylaxis is a life-threatening complication of allergen exposure. While mechanisms governing anaphylaxis after intravenous injection have been defined in mice, these models neglect mucosal exposure that accompanies food ingestion. We investigated the role of mast cell populations within the intestine of mice in response to antigens delivered orally. Oral anaphylaxis required IgE-FcεR1 signaling, and profiling of intestinal mast cells revealed a rapidly developing population shaped by epithelial cues. Intestinal mast cells were largely epithelium-resident and displayed divergent transcriptomes and effector functions from connective tissue mast cells found throughout the body. Histamine synthesis was diminished and leukotriene generation enhanced. Mice genetically deficient in cysteinyl leukotriene synthesis, or those treated with aLOX5 antagonist, Zileuton, were protected from oral anaphylaxis whereas that elicited by intravenous injection was unaltered.

overall_design: BALB/cJ female mice were sensitized subcutaneously with  5ug endograde OVA + 1mg aluminum hydroxide in 200ul PBS on d0 and d7 under ketamine/xylazine treatment. Mice were then challenged intragastrically with 50mg of OVA (Sigma grade III) in 200ul PBS starting on day 14. These mice were challenged every other day, 5 times in total, by oral gavage.  5 days after the 5th OVA challenge, MCs were sorted via FACS Aria for scRNAseq analysis.

first: Nathaniel

last: Bachtel

first: Min

last: Liu

first: Ruslan

last: Medzhitov


overall_design: BALB/cJ female mice were sensitized subcutaneously with OVAxAlum on d0 and d7 . Mice were challenged intragastrically with 20mg of OVA (Sigma grade III) in 200ul water on d14.  MCs were sorted 24 hr later

first: Esther

middle: B

last: Florsheim


title: RBPMS and RBPMS2 Cooperate to Safeguard Cardiac Splicing E11.5

city: San Diego

first: Zeyu

last: Chen

address: 9500 Gilman Drive, BFRII 1217

department: School of Medicine

institute: UCSD

summary: RBPMS and RBPMS2 share a high amino acid sequence similarity and are interchangeable in in vitro splicing assays. However, it remains unclear whether RBPMS and RBPMS2 play similar or distinct function during heart development in vivo. To address this question, we constructed Rbpms and Rbpms2 floxed mice with an established CRISPR-Cas9 method, and validated the efficacy of Rbpms or Rbpms2 floxed alleles in ablating RBPMS or RBPMS2 proteins by crossing them with germline-expressing Sox2-Cre. Transcriptome were abtained by RNA seq to investigate the cardiac function of RBPMS and RBPMS2. We performed RNA sequencing on RbpmscmKO, Rbpms2cmKO, and Rbpms/Rbpms2dcmKO hearts along with their littermate controls.

overall_design: In this study, we generated cardiomyocyte-specific knockout (KO) mice for Rbpms, Rbpms2, and both genes combined (double KO). The experimental groups included Rbpms single KO, Rbpms2 single KO, and Rbpms/Rbpms2 double KO mice, which were analyzed to assess cardiac phenotypes. RNA sequencing was conducted on E11.5 heart tissue from each KO group to evaluate changes in gene expression and splicing patterns. In silico analyses were used to dissect the mechanisms behind the distinct and overlapping roles of RBPMS and RBPMS2 in splicing regulation during heart development. The study focused on identifying the effects of RBPMS and RBPMS2 on sarcomere assembly and the regulation of cardiac-specific splicing programs, as well as determining the impact of their binding locations on pre-mRNA.

first: Tongbi

last: Wu

first: Ju


title: Prelimbic cortex to ventral tegmental area projection regulates early social isolation stress-potentiated heroin seeking in mice

city: Lawrence

first: Zijun

last: Wang

state: Kansas

address: 1251 Wescoe Hall Drive, Malott 5064

department: Pharmacology and Toxicology

institute: University of Kansas

summary: Early-life adversities increase vulnerability to substance use disorders, which are characterized by persistent, uncontrollable drive to seek drugs, often leading to relapse. Previously, we reported that early social isolation (ESI) during adolescence potentiates heroin-seeking in mice. However, the underlying neurobiology remains unknown. Here, we found that ESI aggravated heroin-induced neuronal dysfunction in prelimbic cortex (PrL) to ventral tegmental area (VTA) projecting neurons. Activating PrL->VTA projection attenuated ESI-potentiated heroin seeking, alongside normalized neuronal function. RNA-seq revealed that ESI and heroin convergently altered genes regulating morphogenesis and metabolism, with Tmsb4x (thymosin β4) as a key gene. ESI and heroin interaction affected genes regulating cell cycle and DNA damage response, with Mcm3 and Mcm7 (minichromosome maintenance proteins 3/7) as hubs. PrL thymosin β4 infusion or CRISPR-Cas9-mediated PrL->VTA projection-specific Mcm3/7 knockdown attenuated ESI-potentiated heroin-seeking and neuronal hypofunction. Our study suggests that ESI-potentiated heroin relapse is associated with neuronal and transcriptional alterations in PrL->VTA projection.

overall_design: RNA-seq profiling of TRAP pull-down PrL -> VTA neurons from GFP-L10a mice after 14 days of saline/heroin abstinece.

first: Yunwanbin

first: Shuwen

last: Yue

first: Fengwei

last: Yang

first: Lu

first: Archana

last: Singh

first: Magnus

last: Marciniak

first: Wei

last: Wei

first: Zi-Jun


title: RBPMS and RBPMS2 Cooperate to Safeguard Cardiac Splicing P0

overall_design: In this study, we generated cardiomyocyte-specific knockout (KO) mice for Rbpms, Rbpms2, and both genes combined (double KO). The experimental groups included Rbpms single KO, Rbpms2 single KO, and Rbpms/Rbpms2 double KO mice, which were analyzed to assess cardiac phenotypes. RNA sequencing was conducted on P0 heart tissue from each KO group to evaluate changes in gene expression and splicing patterns. In silico analyses were used to dissect the mechanisms behind the distinct and overlapping roles of RBPMS and RBPMS2 in splicing regulation during heart development. The study focused on identifying the effects of RBPMS and RBPMS2 on sarcomere assembly and the regulation of cardiac-specific splicing programs, as well as determining the impact of their binding locations on pre-mRNA.


first: Jaechul

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

overall_design: Refer to individual Series


title: Plaque-induced transcriptional reprogramming of microglia towards lymphoid receptor expression protects against Alzheimer’s disease.

city: Worcester

first: Violeta

last: Duran Laforet

address: 365 Plantation St

department: Neurobiology

institute: University of Massachusetts Chan Medical School

overall_design: Mouse brains from two control Cx3cr1CreErt2/+(Litt) mouse, three Cx3cr1CreErt2/+(Litt); 5xFAD mice, two 5xFAD; Cx3cr1CreErt2/+(Litt);Spi1fl/+  5xFAD;PU.1-low mouse, and one  5xFAD; Cx3cr1CreErt2/+(Litt);R26FLAG-Spi1/+ 5xFAD;PU.1-high mouse  were used to perform spatial transcriptomics (MERFISH).

first: Pinar

last: Ayata

first: Jessica

middle: M

last: Crowley

first: Matthew

last: Challman

first: Vinaya

last: Sahasrabuddhe

first: Maud

last: Gratuze

first: Sebastian

last: Werneburg

first: Diogo

last: Ribeiro

first: Emma

middle: C

last: Hays

last: Durán-Laforet

first: Travis

last: Faust

first: Philip

last: Hwang

first: Francisco

last: Mendes Lopes

first: Chrysa

last: Nikopoulou

first: Sarah

last: Buchholz

first: Robert

last: Murphy

first: Taoyu

last: Mei

first: Anna

middle: A

last: Pimenova

first: Carmen

last: Romero-Molina

first: Francesca

last: Garretti

first: Tulsi

last: Patel

first: Claudia

last: De Sanctis

first: Angie

last: Ramirez

first: Megan

last: Crow

first: Felix

last: Weiss

first: Jason

last: Ulrich

first: Edoardo

last: Marcora

first: John

last: Murray

last: Meissner

first: Andreas

last: Beyer

first: Dan

last: Hasson

last: Crary

first: Dorothy

last: Schafer

first: David

last: Holtzman

first: Alison

last: Goate

first: Alexander

last: Tarakhovsky

first: Anne

last: Schaefer


title: Microglia-astrocyte crosstalk regulates synapse remodeling via Wnt signaling

first: Pinar

last: Ayata

summary: Unilateral whisker lesioning at age P4 leads to removal of thalamocortical synapses in layer IV of the barrel cortex over the course of the next 6 days. Synapse removal is mediated by microglia and induced by CX3CL1-CX3CR1 signaling between neurons and microglia. Downstream of CX3CL1-CX3CR1 activation, microglia induce astrocyte morphological changes. To identify molecular crosstalk between astrocytes and microglia, we performed translating ribosome affinty purification followed by RNA sequencing (TRAP-Seq).

overall_design: Astrocyte ribosomes were labeled using the transgenic Aldh1l1EGFP-L10a/+ mice and microglial ribosomes were labeled using Cx3cr1CreER/+; Eef1a1LSL-EGFP-L10a/+ mice. 24 hours after unilateral whisker lesioning in P4 mice, mice were euthanized with CO2, and brain regions of interest were dissected and flash-frozen for TRAP. The deprived and non-deprived barrel cortices were micro-dissected and GFP-tagged ribosomes from the cell type of interest were immunoprecipitated with a GFP antibody. RNA sequencing was performed on both the GFP antibody-bound and unbound fractions.

first: Travis

last: Faust

first: Georgia

last: Gunner

first: Ana

last: Badimon

middle: P


title: Alpha-ketoglutarate mitigates insulin resistance and metabolic inflexibility in a mouse model of Ataxia-Telangiectasia

city: Hong Kong

first: Hei-Man Kim

last: Chow

address: Room 604, Mong Man Wai Building, CUHK

department: School of Life Sciences

country: Hong Kong

institute: The Chinese University of Hong Kong

summary: Bulk RNA sequencing of cerebellum brain samples from ATM-KO and their control littermates.

overall_design:
Frozen cerebellar tissues from the transgenic mice were harvested and sent to the Novogene for total RNA extraction and RNA sequencing with the Illumina HiSeq X Ten platform. FASTQ files obtained were subsequently analysed by the FastQC (V. 0.11.9) of Babraham Bioinformatics, serving as a quality control assessment tool for these data. After assuring the quality of the data files, genome indexes were generated using the STAR software. These indexes were generated with the mouse annotation and
mouse genome assembly provided by the Gencode Release 28 (GRCm39, released in May 2021).

The cleaned dataset was then aligned with the genome indexes using the STAR alignment software. FeatureCounts was then used to quantify the sequencing data. Data extraction, matrix construction and differential gene expression analysis was then performed using the DESeq2 package on R.

The sequenced FASTQ files and the raw counts matrix generated by FeatureCounts has been uploaded.

first: Hei-Man

middle: K

first: Ka-Li

middle: J

last: Sun


title: Epigenetic regulation of the mesenchymal niche during pancreatic epithelial development

city: TORONTO

first: Adrian

middle: Kwan Ho

last: Loe

address: 686 Bay Street

institute: The Hospital for Sick Children

summary: Exploring the role of epigenetic regulation in the E13.5 pancreatic mesenchyme by Eed to understand its role in chromatin regulation using ATAC-seq, transcriptional regulation using RNA-seq, and in the regulation of transcriptional heterogeneity using scRNA-seq.

overall_design:
N=2 independent replicates performed for RNA-seq and ATAC-seq of control E13.5 pancreatic mesenchyme versus mutant E13.5 Eed knockout mesenchyme. scRNA-seq was performed on whole control E13.5 pancreata versus whole mutant E13.5 Eed knockout pancreata.

This series includes re-analyzed samples from GSE147418.

first: Taehee

last: Kim

first: Theodora

last: Yung

first: Minggao

last: Liang


title: Retinoic acid receptor-related orphan receptor α regulates bystander activation of memory CD8+ T cells [RNA-Seq]

city: Sapporo

first: Kensuke

last: Takada

address: N21W11 Kita-ku

institute: Hokkaido University

summary: Background: Memory CD8+ T cells sense inflammation and rapidly produce interferon-γ (IFN-γ) independent of cognate antigens. This innate-like property, called bystander activation, is involved in early host defense before the antigen-specific memory response. However, the molecular mechanisms underlying this activation remain unknown. Retinoic acid receptor-related orphan receptor α (RORα) belongs to the nuclear receptor family and regulates gene transcription in ligand-dependent manner. Although RORα is highly expressed in memory CD8+ T cells, its functional relevance has not been investigated. Methods: Primary and secondary memory T cells that are sufficient or deficient of RORα were induced by adoptive transfer of naïve OT-I T cells to recipient mice and subsequent infection with Listeria monocytogenes expressing ovalbumin (LM-OVA). RORα expression in memory T cells were examined by quantitative PCR. The target genes of RORα in memory T cells were explored by RNA-sequencing and verified by RORα overexpression in postactivated T cells. The impact of RORα-deficiency on bystander activation was assessed by stimulating memory T cells with inflammatory cytokines in vitro or injecting lipopolysaccharide (LPS) into mice bearing memory T cells. Results: RORα expression was remarkably elevated in secondary memory CD8+ T cells along with the enrichment of effector-like memory T cells. RORα primarily acted as a transcription factor in regulating the gene expression of the TL1A receptor. RORα deficiency abrogated the IFN-γ production by memory CD8+ T cells in response to IL-12 + TL1A in vitro and diminished the bystander response to LPS-induced inflammation in vivo. Conclusion: This study revealed a regulatory mechanism of bystander activation. The findings also improve our understanding of how memory T cells increase their immediate protective capacity through repeated infections and vaccinations.

overall_design: RNA-seq profiling of wild-type and Rora-deficient secondary memory CD8+ T cells.

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