title: An orally active, gut-enriched AhR agonist demonstrates therapeutic efficacy in colitis through regulation of inflammation, barrier homeostasis, and fibrosis

city: Suzhou

first: zhong

last: huiting

address: Building B, Phase 3, Suzhou BioBay, Suzhou Industrial Park

institute: Allianthera bio-pharmaceutical Co., Ltd CN

summary: Inflammatory bowel disease (IBD) is characterised by chronic intestinal inflammation, epithelial barrier dysfunction, and maladaptive stromal remodelling that can progress to fibrosis. Current therapies primarily target inflammatory pathways and rarely achieve durable remission or structural disease modification, highlighting a major unmet clinical need. Here, we evaluated the therapeutic potential of YUQ-A1007, an orally active, gut-enriched small-molecule aryl hydrocarbon receptor (AhR) agonist, across multiple translational models of colitis. The anti-inflammatory, barrier-restorative, and anti-fibrotic effects of YUQ-A1007 were assessed using in vitro cell systems, patient-derived colonic biopsies and organoids, permeability assays, and three murine colitis models. YUQ-A1007 partially activated AhR signalling, suppressed pro-inflammatory cytokine production, enhanced regulatory T cell differentiation, increased tight junction protein expression, and inhibited fibrotic gene programs. In vivo, oral YUQ-A1007 administration reduced disease severity, restored epithelial barrier integrity, and broadly downregulated inflammatory cytokines, chemokines, fibrosis-associated genes, pro-inflammatory mesenchymal stromal cell biomarkers, and matrix metalloproteinases. In ulcerative colitis patient-derived colonic explants, YUQ-A1007 dose-dependently reduced inflammatory mediator secretion while preserving barrier function. Collectively, these findings support further development of YUQ-A1007 as a gut-targeted AhR therapeutic for ulcerative colitis and broader inflammatory bowel disease.

overall_design: To define the molecular basis for the therapeutic effects observed in TNBS colitis, we performed RNA-seq on distal colon tissue from vehicle- and YUQ-A1007–treated mice. TNBS treatment triggered robust immune activation, as evidenced by elevated expression of a broad array of inflammatory cytokines and chemokines.

first: Pan

last: Ren-You

first: Zhang

last: Yongli


title: FOXO/DAF-16 modulates the transcription factor ROR/NHR-23 and inhibits the let-7 microRNA to maintain multipotency during dauer

status: Public on Jul 17 2026

city: Baltimore

first: John

middle: K

last: Kim

state: MD

address: 3400 North Charles St

department: Biology

country: USA

institute: Johns Hopkins University

type: Expression profiling by high throughput sequencing

summary: Animals rapidly reprogram gene expression to adapt development to environmental stress. How gene regulatory programs that drive continuous development are repressed during stress-induced developmental arrest remains poorly understood. In Caenorhabditis elegans, starvation and overcrowding trigger entry into the stress-resistant, quiescent dauer stage. Here, we identify interactions among the conserved transcription factors DAF-16/FOXO and NHR-23/ROR, and the let-7 family of microRNAs as key regulators of the switch from continuous development to dauer. We show that loss of daf-16 during dauer causes elevated let-7 family microRNAs and premature expression of the adult collagen reporter col-19p::GFP. Reducing let-7 family activity suppresses this phenotype, whereas dauer-specific let-7 expression is sufficient to induce col-19p::GFP expression. Mechanistically, DAF-16 inhibits let-7 transcription in part by repressing nhr-23, which encodes a transcriptional activator of the let-7 family and molting-cycle genes. ChIP-seq analysis reveals DAF-16 binding upstream of nhr-23, and daf-16; daf-7 mutant dauers exhibit increased nhr-23 mRNA and NHR-23 protein, supporting a model in which DAF-16 directly represses nhr-23. Integrated ChIP-seq and transcriptomic analyses identify 1,183 genes activated and 681 genes repressed by DAF-16 during dauer. Repressed targets are enriched for pro-growth genes involved in mitotic DNA replication and translational elongation. DAF-16 targets include 59 transcription factors that may mediate broader transcriptional reprogramming during dauer to maintain multipotency and establish quiescence. Together, these findings reveal that DAF-16/FOXO establishes stress-induced developmental arrest by coupling activation of protective pathways with repression of conserved developmental timing, growth, and differentiation programs.

overall_design: RNA-seq in developing L2d larvae of 1. QK216 nhr-23(xk40[nhr-23::aid::3xflag]) I; ieSi57[eft-3p::tir1::mRuby::unc-54 3'UTR + Cbr-unc-119(+)] II; daf-7(e1372) III; maIs105[col-19p::gfp] V these samples are annotated daf-7 in the file names, 2. QK215 daf-16(mgDf50) nhr-23(xk40[nhr-23::aid::3xflag]) I; ieSi57[eft-3p::tir1::mRuby::unc-54 3'UTR + Cbr-unc-119(+)] II; daf-7(e1372) III; maIs105[col-19p::gfp] V without auxin, and 3. QK215 daf-16(mgDf50) nhr-23(xk40[nhr-23::aid::3xflag]) I; ieSi57[eft-3p::tir1::mRuby::unc-54 3'UTR + Cbr-unc-119(+)] II; daf-7(e1372) III; maIs105[col-19p::gfp] V with 1mM auxin these samples are annotated daf-7daf-16 in the file names

first: Himani

last: Galagali

first: Matthew

middle: J

last: Wirick

first: Amelia

middle: F

last: Alessi

first: Margaret

middle: R

last: Starostik

first: Priya

last: Balamurugan

first: Isabella

middle: G

last: Sims

first: Laurianne

last: Pene

first: Ruhi

last: Patel

first: Alison

last: Frand

first: Suhua

last: Feng

first: Steven

middle: E

last: Jacobsen

first: Xantha

last: Karp


title: Single cell RNA-seq of EpCAM+ epithelial cells from mouse esophagus

city: Fuzhou

first: Qingxiang

last: Gao

state: FuJian

address: Qiuyang Road No1.

country: China

institute: Fujian University of Traditional Chinese Medicine

summary: Esophageal squamous epithelial homeostasis depends on a tightly controlled balance between basal cell self-renewal and stepwise differentiation along the basal-suprabasal-superficial axis. Disruption of this balance leads to barrier dysfunction and contributes to diseases such as eosinophilic esophagitis (EoE), which is characterized by basal cell hyperplasia. N6-methyladenosine (m6A), the most abundant internal modification of eukaryotic mRNA, is a key regulator of tissue homeostasis through control of mRNA stability and other post-transcriptional processes. However, the role of m6A, catalyzed by the METTL3/METTL14 complex, in the esophageal epithelium and contribution to disease pathogenesis remain largely unknown.

overall_design: To resolve cell-state changes underlying the early remodeling phenotype, we performed scRNA-seq on esophageal epithelial cells from Mettl3L/L (WT) and Sox2CreER;Mettl3L/L (KO) mice

first: Shuzhen

last: Xu

first: Tala

last: Fnu

first: Chao

last: Jing

first: Jianwen

last: Que


title: RNA profiles of hematopoietic stem cells before and after long-term culture [RNA-Seq]

city: Kumamoto

first: Terumasa

last: Umemoto

address: 2-2-1, Honjo, Chuo-ku

country: Japan

institute: Kumamoto University

summary: Self-renewing hematopoietic stem cells (HSCs) during hematopoietic regeneration showed lower intracellular glutamate (Glu) levels compared with dividing progenitor cells in the steady state and HSCs attenuate stem cell function during ex vivo culture. R162, an inhibitor of glutamine dehydrogenase (Glud1) that catabolize Glu into α-ketoglutarate, contributed to stem cell maintenance during ex vivo culture. These data suggest that lower intracellular Glu levels during HSC division contributed to self-renewing divisions by suppressing Glud1-mediated reactions.

overall_design: EPCRHigh cells within the HSC fraction (CD150+ CD48- EPCR+ Lineage-) were cultured for 5 or 30 days in the absence or presence of R162. Before and after the culture, EPCRHigh or EPCRLow cells within the HSC fraction (CD150+ CD48- EPCR+ Lineage-) were sorted and subsequently subjected to RNA-seq.

first: Michie

last: Uchikawa


title: Inflammatory Neuropathy in Mouse and Primate Models of Colorectal Cancer

city: Houston

first: Rajasekaran

last: Mahalingam

state: TX

address: 6565 MD Anderson Blvd

department: Symptom Research

institute: University of Texas MD Anderson Cancer Center

summary: Colorectal cancer survivors are at increased risk of developing neurological issues, particularly peripheral neuropathy and chronic pain. Although pre-existing neuropathy is a risk factor for chronic pain, tumor-induced neuropathy has not been firmly established in pre-clinical models. Consistent with clinical observations, we show that mice with colorectal cancer develop peripheral neuropathy, which was associated with subtle locomotor deficits, without overt hypersensitivity. We detected widespread differences in pro-inflammatory cytokines and lipid metabolites in peripheral nerves from tumor-bearing mice. Macrophage accumulation, myelin decompaction and ryanodine receptor oxidation were associated with dysfunctional calcium homeostasis and reduced spike amplitude in sensory neurons. Similar alterations in plasma inflammatory mediators and lipid metabolites were associated with neuropathy and macrophage accumulation in peripheral nerves of rhesus macaques with colorectal cancer. These findings suggest colorectal cancer is causally linked to a subacute form of chronic inflammatory demyelinating polyneuropathy across species, which may represent an under-reported, yet important risk factor for neurological dysfunction in colorectal cancer survivors.

overall_design: RNA-seq profiling of Bone Marrow Derived Macrophages and Scwann cells from control and MC38 tumor mice and also with lipid depletion in Bone Marrow Derived Macrophages

first: Caitlyn

last: M. Gaffney

first: Angela

last: M. Casaril

first: Iqbal

last: Mahmud

first: Bo

last: Wei

first: Yanyan

last: Jiang

first: Nathan

last: T. Fiore

first: Jialu

last: Liu

first: Karen

last: M. Valadez

first: Elizabeth

last: A. Kolb

first: Fisher

last: R. Cherry

first: Lei

last: Shi

first: Kenneth

last: Dunner Jr.

first: Philip

last: L. Lorenzi

last: R. Reiken

first: Peter

last: M. Grace

first: Theresa

last: A. Guise

first: Carolyn

last: L. Hodo

first: Andrew

last: J. Shepherd


title: Vascular function underlies enhanced intestinal regeneration after FLASH radiation [scRNAseq]

city: Lausanne

first: Tania

last: Wyss

address: Rue du Bugnon 25A

country: Switzerland

institute: SIB Swiss Institute of Bioinformatics

summary: Ultra-high dose-rate FLASH radiotherapy spares normal tissue while promoting tumor control, yet the basis of this protection remains unknown. Using whole-abdominal irradiation, intestinal organoids, single-cell transcriptomics, spatial vascular phenotyping, and genetic, pharmacological, and physiological perturbations, we show that the intestinal FLASH effect is the result of active heterotypic vascular-epithelial crosstalk rather than passive radiochemical protection. FLASH preserves endothelial tone and perfusion along the crypt–villus axis, averting phospho-MLC–dependent vascular collapse caused by conventional irradiation. This maintains the physiological oxygen gradient and licenses early, YAP-dependent regeneration that shields transit-amplifying secretory progenitors from persistent DNA damage and drives rapid crypt reconstitution. Disrupting the oxygen gradient directly, or indirectly through endothelial VEGFR2 blockade, abolishes FLASH sparing—reinstating progenitor DNA damage, loss of DCLK1⁺ and CHGA⁺ cells, and impaired regeneration. We identify an oxygen–vasculature–YAP–progenitor axis as a central regulator of radiation-induced repair and a target to widen the therapeutic window.

overall_design: C57BL/6 mice were maintained under specific pathogen-free conditions with ad libitum access to food and water on a 12 h light/dark cycle. Mice were used at 8–12 weeks of age. Animals received either FLASH or conventional (CONV) whole-abdominal irradiation at a dose of 15 Gy, or were left untreated as controls. Mice were subjected to a single fraction of 15 Gy whole-abdominal irradiation delivered either at ultra-high dose rate (FLASH) or conventional dose rate (CONV). Unirradiated mice served as controls. Small intestinal tissue was harvested 24 h after irradiation. Blood endothelial cells were isolated by fluorescence-activated cell sorting (FACS) for single-cell RNA sequencing analysis.

first: Anet

last: Valdés-Zayas

first: Preethi

last: Devanand

first: Jeremiah

last: Bernier-Latmani

first: Mehdi

last: Hicham

first: David

last: Tarussio

first: Laure

last: Garnier

first: Gaspard

last: Pardon

first: Stephanie

last: Tissot

first: Nadine

last: Fournier

first: Tatiana

last: Petrova

first: Fernanda

last: G Herrera


title: Vascular function underlies enhanced intestinal regeneration after FLASH radiation

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

overall_design: Refer to individual Series


title: MYCN sustains a progenitor-stage differentiation block in human neuroepithelial stem cells to drive ETMR-like tumourigenesis [scRNA-seq]

city: London

first: Diego

last: Chillon Pino

address: Great Ormond Street Hospital for Children NHS

department: Cancer development and genomics

country: United Kingdom

institute: UCL

summary: Embryonal tumours with multilayered rosettes (ETMRs) are aggressive paediatric brain tumours driven in most cases by amplification of the chromosome 19 microRNA cluster (C19MC), which upregulates the oncogene MYCN. Whether MYCN dysregulation alone is sufficient to produce ETMR-like tumours, independent of the broader C19MC machinery, has not been tested. Here, we show that sustained MYCN expression in human neuroepithelial stem (NES) cells, i.e. primitive neural progenitors resembling the proposed cell of origin of ETMR, is sufficient to generate ETMR-like brain tumours in vivo. These tumours share methylation and transcriptional signatures with ETMR and reproduce its primitive, stem-like cellular hierarchy. MYCN-expressing tumour cells are arrested at an early neural progenitor stage, and MYCN withdrawal arrests growth and engages neuronal differentiation. Our findings identify sustained MYCN as sufficient to drive an ETMR-like developmental arrest in primitive human neural cells.

overall_design: Single-cell RNA-sequencing (10x Genomics Chromium GEM-X Flex, v2 chemistry) of three NESt xenograft tumours, generated by intracranial implantation of Sai2 NES cells carrying doxycycline-induced MYCN(T58A) into doxycycline-fed NSG mice. The three tumours were sample-multiplexed by probe barcode on a single Chromium chip (~5,000 cells/sample target) and sequenced on Illumina NovaSeq (~20,000 reads/cell). 10,438 cells passed quality control.

first: Sumana

last: Shrestha

last: Chillón Pino

first: Colin

last: Kwok

first: Yura

last: Grabovska

first: Barbara

last: Martins da Costa

first: Kevin

last: Greenslade

first: Karen

last: Barker

first: Yann

last: Jamin

first: Angela

last: Hayes

first: Maximilian

last: Crane

first: Laurie

last: Liu

first: Fupan

last: Yao

first: Otis

last: Agbaimoni

first: Claire

last: Lynn

first: Thomas

middle: S

last: Jacques

first: Ulrich

last: Schüller

first: Annie

last: Huang

first: Laura

last: Donovan

first: Louis

last: Chesler


title: KAT7-Driven Lactylation of CCAR2 Promotes Macrophage Extracellular Traps Formation and Accelerates Atherosclerosis Progression

city: Hefei

first: Xianshe

last: Meng

state: Anhui

address: No. 96 Jinzhai Road

institute: University of Science and Technology of China

summary: KAT7 regulates macrophage foam cell formation and atherosclerosis progression. To identify transcriptional changes associated with KAT7 deficiency in macrophages, bulk RNA-seq was performed using bone marrow-derived macrophages (BMDMs) from Ctrl and KAT7KO mice. Three biological replicates were included for each group. Differential expression analysis was performed for KAT7KO versus Ctrl to identify KAT7-regulated genes and pathways in macrophages.

overall_design: This bulk RNA-seq study compared BMDMs from two genotypes: Ctrl and KAT7KO. The experiment included three biological replicates per group, generating six paired-end RNA-seq libraries in total.


title: Breast cancer-associated fibroblast-derived exosomal miR-1290 promotes triple-negative breast cancer cell proliferation

city: Daejeon

first: Sugi

last: Lee

address: 125, Gwahak-ro, Yuseong-gu, Daejeon 34141, Republic of Korea

country: South Korea

institute: KRIBB

summary: Breast cancer is a major cause of cancer-associated mortality in females. Although therapeutic strategies targeting subtype-specific receptors have been developed for various breast cancer subtypes, effective targeted therapies for triple-negative breast cancer (TNBC) are lacking. Recently, the role of the tumor microenvironment (TME), particularly cancer-associated fibroblasts (CAFs), in the progression and metastasis of breast cancer, especially TNBC, has attracted increasing attention. However, the role of exosomes secreted by CAFs in TNBC cell proliferation is poorly understood. Thus, in this study, we analyzed exosomes derived from both CAFs and MDA-MB-231 cells and identified exosomal miR-1290 as a key component. When delivered to TNBC cells (BT-549 and Hs578T), exosomal miR-1290 directly suppressed the expression of the putative tumor suppressor TBC1D4, which was accompanied by reduced TSC1 expression and activation of the mTOR pathway. The promotion of TNBC cell growth by miR-1290 through direct targeting of TBC1D4 was validated using 2D culture models, 3D spheroid models, and in vivo xenograft models. Furthermore, we demonstrated that treatment with the mTOR inhibitor rapamycin and the FDA-approved rapamycin analog everolimus effectively suppressed miR-1290-induced TNBC cell proliferation, suggesting a potential therapeutic strategy for TNBC. Moreover, in patient cohorts, elevated levels of circulating miR-1290 significantly distinguished patients with breast cancer from healthy individuals and correlated with poor survival outcomes. Therefore, miR-1290 and its direct target TBC1D4 have potential value in both targeted treatment and diagnosis of TNBC.

overall_design: Total RNA was extracted from the samples using the RNeasy Mini Kit (Qiagen, 74106). RNA-seq libraries were prepared from total RNA and sequenced on the Illumina NextSeq 1000 platform using a paired-end 2 × 100 bp configuration. Transcriptomic profiles were generated to compare gene expression between experimental groups.


title: Bulk RNA-seq of U251N orthotopic glioblastoma xenografts in athymic RNU/RNU rats before and after MRI-guided laser interstitial thermal therapy (LITT)

city: Detroit

first: Tavarekere

last: Nagaraja

state: MI

address: 2799 W. Grand Blvd.

department: Neurosurgery

institute: Henry Ford Health

summary: Laser interstitial thermal therapy (LITT) is increasingly used for deep-seated and recurrent glioblastoma, but the transcriptional state of cells that survive the sublethal thermal penumbra and seed recurrence is poorly defined. We performed bulk RNA-seq on matched primary (pre-LITT) and recurrent (post-LITT) tumors from a U251N orthotopic xenograft in athymic RNU/RNU rats treated with MRI-guided LITT. Reads were sorted into human (graft) and rat (host) streams with xengsort to isolate the human tumor transcriptome from host signal. Recurrent tumors showed a pronounced translational shutdown and integrated-stress-response activation, with a shift toward the Garofano Mitochondrial molecular subtype, nominating the translation/ISR axis as a candidate target for LITT-adjuvant therapy.

sample_organism:
Homo sapiens
Rattus norvegicus

platform_organism:
Homo sapiens
Rattus norvegicus

overall_design: Six orthotopic U251N xenograft tumors were profiled: primary/pre-LITT (n=3: IL67B, IL68B, IL69B) versus recurrent/post-LITT (n=3: IL66B, NL70B, NL71B). Three tumor-free in vivo controls (IL64B failed graft; N168B, N269B procedural) and one in vitro U251 culture reference (C2B) were sequenced through the identical pipeline; controls and the in vitro reference were used for QC / contamination assessment and were excluded from the differential-expression contrast. Differential expression was computed between recurrent and primary tumors.

first: Gregory

last: Schwing

first: Indrani

last: Datta

first: Ian


first: John

last: Kim

address: 3400 Charles Street

department: Department of Biology

type: Genome binding/occupancy profiling by high throughput sequencing

overall_design: ChIP-seq of DAF16 in starved dauer from wild-type and daf-16(ar620[daf-16::zf1-wrmScarlet-3xFLAG]) C. elegans grown at 20°C.

first: Himani

last: Galagali

first: Matthew

last: Wirick

first: Amelia

last: Alessi

first: Margaret

last: Starostik

first: Priya

last: Balamurugan

first: Isabella

last: Sims

first: Laurianne

last: Pene

first: Ruhi

last: Patel

first: Alison

last: Frand

first: Suhua

last: Feng

first: Steven

last: Jacobsen

first: Xantha

last: Karp


title: Metabolites from Bifidobacterium adolescentis provide protection against inflammation-induced colonic barrier disruption in vitro and in vivo

city: Ann Arbor

first: Thomas

last: Schmidt

state: Michigan

address: 1150 W Medical Ctr 1520C MSRB I

department: Medical School Internal Medicine Infectious Diseases

institute: University of Michigan

summary: Bifidobacteria are common and beneficial members of the gastrointestinal (GI) microbiome. Cultivars of bifidobacterial species common in infants have been better studied and used more frequently as probiotics than adult-type bifidobacteria, in part because they are more tolerant of oxygen. We sought to identify interactions between B. adolescentis – an oxygen-sensitive species that is the most prevalent and abundant Bifidobacterium species in the adult gut microbiome – and the colonic epithelium. Using an asymmetric oxygen system (AOS) that maintains constant oxygen concentrations on the basolateral side of colonic epithelial monolayers and anoxic conditions on the luminal side, we found that monolayers in the AOS behaved more like in vivo epithelia than monolayers cultured under 20% oxygen atmospheres. Co-culturing colonic monolayers with Bifidobacterium adolescentis U269-1 in the anoxic apical chamber mitigated loss of the barrier function in monolayers exposed to inflammatory cytokines. We identified indole lactic acid (ILA) as one metabolite from B. adolescentis U269-1 that provides dose-dependent protection against inflammatory cytokines. Oral administration of ILA was well tolerated in mice and provided protection against immunotherapy-induced (anti-CTLA4) colitis in female (but not male) mice. The present study yielded strong evidence for the beneficial effects of Bifidobacterium adolescentis on the maintenance of colonic barrier function in an in vitro organoid model under physiological conditions. Bacterial metabolite ILA was implicated in this protection, and beneficial effects were observed in a mouse model of colitis. The AOS scaled reproducibly to 96-well plates, providing the capacity for future mid-throughput screening of microbes and microbial metabolites under physiologically relevant conditions.

overall_design: RNA extracted from primary human colonoid monolayers incubated at ambient oxygen (~21%) or asymmetrically delivered 10% oxygen gas mix (5% CO2, 37°C for both); marked S21 and A10 respectively

first: Nicole

middle: M

last: Cady

first: Sophia

last: Meyer

first: Kwi

first: Bernard

middle: C

last: Lo

first: Ngai

last: So

first: Grace

middle: Y

last: Chen

first: Stephen

last: Rosshart

first: Yatrik

last: Shah

first: Gabriel

last: Nunez

first: Jonathan

middle: Z

last: Sexton


title: Linker histone H1.0 and JUNB Interaction Maintains Lineage-Specific and Age-Dependent Chromatin Architecture to Safeguard Cell Identity [RNA-Seq]

first: Chulan

last: Kwon

address: 720 Rutland Ave, Ross 954

department: Medicine, Cardiology, Heart & Vascular Institute, Institute for Cell Engineering, and Cellular and Molecular Medicine

summary: Maintenance of differentiated cell identity is essential for tissue homeostasis and depends on active molecular mechanisms that reinforce cell fate stability. This thesis focuses on identifying the molecular factors involved in this maintenance system and defining how they function as barriers that oppose cell fate reprogramming and protect cellular identity. More broadly, this work addresses a fundamental question in biology: how differentiated cells actively preserve their identity while resisting inappropriate lineage programs, and how aging erodes these mechanisms, contributing to loss of cell identity in aged cells.  In particular, this work examines how transcription factor barriers cooperate with chromatin regulators to maintain cell fate stability. In a previous study, JUNB was identified as a transcription factor barrier that restricts cell fate reprogramming through regulation of chromatin accessibility at both open and closed chromatin regions. Notably, transcription factors are not typically characterized for their ability to regulate closed chromatin states. Given that transcription factors frequently function in cooperation with other regulatory proteins, it was hypothesized that JUNB interacts with additional chromatin regulators to control chromatin accessibility more broadly.  In this study, H1F0, a linker histone with chromatin-regulatory properties, was identified as a novel functional interactant of JUNB that acts as a barrier to cell fate reprogramming. Mechanistic investigation further demonstrated that JUNB and H1F0 cooperate at shared regulatory regions, including sites associated with the chromatin architectural regulator CTCF, to modulate chromatin accessibility. Through this coordinated action, engagement of lineage-reprogramming transcription factors with gene programs required for alternative fate activation was restricted, thereby preserving cell fate stability.

overall_design: Bulk RNA-seq of iMGT (Mef2c, Gata4, Tbx5)-Myh6-eGFP-MEF cell line reprogrammed into induced-cardiomyocytes at day 3 post-MGT overexpression comparing siControl, siJunb or siH1f0 conditions. GFP+ Cells were FACS sorted and used for RNA extraction.

first: Yu-Ling

last: Chang

first: Sean

last: Murphy

first: Alexandre

last: Colas


title: Linker histone H1.0 and JUNB Interaction Maintains Lineage-Specific and Age-Dependent Chromatin Architecture to Safeguard Cell Identity [ATAC-seq]

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design:
Bulk ATAC of iMGT (Mef2c, Gata4, Tbx5)-Myh6-eGFP-MEF cell line reprogrammed into induced-cardiomyocytes at day 1 post-MGT overexpression comparing siControl, siJunb or siH1f0 conditions or Ground state (no reprogramming).
Bulk ATAC of IMR90-doxy-inducible (MYOD) skeletal muscle cell reprogramming cell line at day 1 post-MYOD overexpression comparing siControl, siJunb or siH1f0 conditions or Ground state (no reprogramming).


title: Linker histone H1.0 and JUNB Interaction Maintains Lineage-Specific and Age-Dependent Chromatin Architecture to Safeguard Cell Identity [CUT&Run]

overall_design: CUT&RUN of iMGT (Mef2c, Gata4, Tbx5)-Myh6-eGFP-MEF cell line reprogrammed into induced-cardiomyocytes at day 1 post-MGT overexpression comparing siControl, siJunb or siH1f0 conditions or Ground state (no reprogramming). Antibody target to MEF2C,


title: Single-Cell RNA sequencing reveals clonally-expanded CD4+ tissue-resident memory T cells in Histidyl-tRNA Synthetase-induced myositis [RNA-seq]

city: Pittsburgh

first: Alyxzandria

last: Gaydosik

state: PA

address: 200 Lothrop St

institute: University of Pittsburgh

summary: The precise mechanisms underlying the pathogenesis of idiopathic inflammatory myopathy (IIM) remain undefined.  However, there has been increasing recognition that tissue-resident memory cells (TRMs) play an important role in the pathogenesis of systemic autoimmune disease.  In IIM, TRM-associated transcriptional signatures have been reported, but on a very limited basis.  By using multimodal single-cell RNA sequencing analysis in our established murine model of histidyl-tRNA synthetase (HRS)-induced myositis, we identified a prominent population of CD4+ TRMs in inflamed skeletal muscle. Muscle CD4+ TRMs exhibited high expression of genes encoding Cd69, Cxcr6, Runx3, and Prdm1, alongside low expression of Klf2, Ccr7, Sell, S1pr1, and Tcf7—a profile that is generally consistent with previous reports of TRM gene signature and that we validate through comparison to transcriptomic profiles of human muscle tissue.  Detailed pathway analysis in our model indicates that muscle CD4+ TRMs contribute to innate immune regulatory pathways enriched for TNF and IFN-γ signaling. Furthermore, analysis of TCR clonotype distribution and CDR3 sequence similarity revealed pronounced clonal expansion of CD4⁺ TRMs relative to other T-cell subsets—a pattern that remained stable from 2 to 6 weeks post-immunization. Collectively, these results suggest a potential role for CD4+ TRMs in the pathogenesis of autoimmune myositis.

overall_design: Mice Treatment: 100 uL of affinity-purified, filter-sterilized recombinant HRS (3-5 mg/mL) was administered via bilateral intramuscular hamstring injection (50 ml/side) of 8-10 week old female C57BL/6 mice to induce myositis; control mice received PBS only.  Mice were sacrificed 14 days post-immunization for collection of muscle tissue.

first: Decheng

last: Li

first: Daniel

last: Reay

first: Iago

last: Pinal-Fernadez

first: Maria

last: Casal-Dominguez

first: Andrew

last: Mammen

first: Sarah

last: Gaffen

first: Timothy

last: Oriss

first: Dana

last: Ascherman


title: PD-1 regulates CD4+ T cell-mediated CD8+ T cell responses in the brain to balance viral control and neuroinflammation

city: Hershey

first: Vonn

last: Walter

address: 500 University Drive

department: Public Health Sciences

institute: Penn State College of Medicine

summary: Programmed cell death protein 1 (PD-1) is expressed by T cells during progressive multifocal leukoencephalopathy (PML), a life-threatening brain disease caused by the human-only JC polyomavirus. Why PD-1 blockade finds variable success in PML patients is unclear. Brain CD4+ and CD8+ T cells are PD-1high during mouse polyomavirus (MuPyV) encephalitis. Here, we show that PD-1 loss during MuPyV infection acts in a brain-autonomous manner to increase the magnitude of brain-infiltrating CD4+ and CD8+ T cells and the function of virus-specific CD8+ T cells; in concert, brain virus levels decline and neuroinflammation increases. Deletion of PD-1 in CD4+ T cells, but not CD8+ T cells, recapitulates effects of global PD-1 loss. Single-cell RNA sequencing shows that PD-1-deficient CD8+ T cells cluster as effectors while transcripts associated with proliferation and function are upregulated with loss of PD-1. Thus, CD4+ T cell-intrinsic PD-1 signaling balances antiviral defense against neural injury during polyomavirus CNS infection.

overall_design: Following A2 MuPyV infection and tamoxifen-induced knockout of PD-1 in wild-type Rosa-Cre and PD-1fl/fl Rosa-Cre mice, brains were collected, and virus-specific CD8 and antigen-experienced CD4 T cells were sorted through  fluorescence-activated cell sorting (FACS). Cells were then processed for scRNA-sequencing.

first: Arrienne

middle: B

last: Butic

first: Elia

last: Afanasiev

first: Samantha

middle: A

last: Spencer

first: Mofida

last: Abdelmageed

first: Anirban

last: Paul

first: Kalynn

last: Alexander

first: Katelyn

middle: N

last: Ayers

first: Todd

middle: D

last: Schell

first: Matthew

last: Lauver

first: Ge

last: Jin

last: Borys

first: Rachel

last: Kang

first: ChaeMin

last: Kim

first: Elizabeth

last: Proctor

first: Laurent

last: Brossay

first: Jo Anne

last: Stratton

first: Aron

last: Lukacher


title: Vascular function underlies enhanced intestinal regeneration after FLASH radiation [bulk RNAseq]

first: Tania

last: Wyss

summary: Ultra-high dose-rate FLASH radiotherapy spares normal tissue while promoting tumor control, yet the basis of this protection remains unknown. Using whole-abdominal irradiation, intestinal organoids, single-cell transcriptomics, spatial vascular phenotyping, and genetic, pharmacological, and physiological perturbations, we show that the intestinal FLASH effect is the result of active heterotypic vascular-epithelial crosstalk rather than passive radiochemical protection. FLASH preserves endothelial tone and perfusion along the crypt–villus axis, averting phospho-MLC–dependent vascular collapse caused by conventional irradiation. This maintains the physiological oxygen gradient and licenses early, YAP-dependent regeneration that shields transit-amplifying secretory progenitors from persistent DNA damage and drives rapid crypt reconstitution. Disrupting the oxygen gradient directly, or indirectly through endothelial VEGFR2 blockade, abolishes FLASH sparing—reinstating progenitor DNA damage, loss of DCLK1⁺ and CHGA⁺ cells, and impaired regeneration. We identify an oxygen–vasculature–YAP–progenitor axis as a central regulator of radiation-induced repair and a target to widen the therapeutic window

overall_design: Bulk RNA sequencing was performed to characterize transcriptional responses to ultra-high dose-rate FLASH versus conventional (CONV) radiotherapy in the intestine. Mice received 15 Gy whole-abdominal irradiation, and whole jejunum tissue was harvested 16 hours post-irradiation. Unirradiated mice served as controls.

first: Anet

last: Valdés-Zayas

first: Preethi

last: Devanand

first: Jeremiah

last: Bernier-Latmani

first: Mehdi

last: Hicham

first: David

last: Tarussio

first: Laure

last: Garnier

first: Jessica

last: Sordet Dessimoz

first: Stephanie

last: Tissot

first: Gaspard

last: Pardon

first: Nadine

last: Fournier

first: Tatiana

last: V Petrova

first: Fernanda

last: G Herrera


title: Single-Cell RNA sequencing reveals clonally-expanded CD4+ tissue-resident memory T cells in Histidyl-tRNA Synthetase-induced myositis [2]

first: Alyxzandria

last: Gaydosik

overall_design: Mice Treatment (similar to above but some details different): 100 uL of affinity-purified, filter-sterilized recombinant HRS (3-5 mg/mL) was administered via bilateral, intramuscular hamstring injection (50 ul/side) of 8-10 week old female C57BL/6 mice to induce myositis; control mice received PBS only. Mice were sacrificed 2 weeks or 6 weeks post-immunization for collection of muscle tissue. Physical and enzymatic digestion of groups of 2-3 pooled muscle samples yielded single cell preparations that were then subjected to scRNAseq using the 10X Genomics platform.

first: Decheng

last: Li

first: Daniel

last: Reay

first: Iago

last: Pinal-Fernadez

first: Maria

last: Casal-Dominguez

first: Andrew

last: Mammen

first: Sarah

last: Gaffen

first: Timothy

last: Oriss

first: Dana

last: Ascherman


title: Impaired mechanotransduction in second heart field-derived smooth muscle cells causes NOTCH1 haploinsufficiency-associated aortopathy

city: Columbus

first: Vidu

last: Garg

state: OH

address: 700 Children's Drive WB4239

department: Cardiovascular Research

institute: Research Institute at Nationwide Children's Hospital

summary: Ascending aortic aneurysms (AscAA) are associated with aortic dissection/rupture and arise from genetic, developmental or biomechanical etiologies. NOTCH1 pathogenic variation is associated with AscAA found with congenital heart defects, such as bicuspid aortic valve and tetralogy of Fallot. This study sought to characterize a Notch1 haploinsufficient murine model of AscAA and investigate mechanisms by which Notch1 deficiency contributes to AscAA.

overall_design: Human NOTCH1+/- induced pluripotent stem cell (iPSC) lines were differentiated to iPSC-derived vascular smooth muscle-like cells (iVSMCs) via a second heart field (SHF) lineage; followed by RNA-sequencing at baseline and after exposure to 24h of cyclic stress.

first: Ruth

last: Ackah


title: Single-Cell RNA sequencing reveals clonally-expanded CD4+ tissue-resident memory T cells in Histidyl-tRNA Synthetase-induced myositis


title: Single-Cell RNA sequencing reveals clonally-expanded CD4+ tissue-resident memory T cells in Histidyl-tRNA Synthetase-induced myositis [1]

overall_design: Single-cell transcriptome libraries were generated from formaldehyde-fixed, paraffin embedded (FFPE) human muscle biopsy specimens (n=16) using the 10x Genomics Chromium Single Cell Flex assay, a probe-based approach designed to profile fixed cells. Libraries prepared with this approach were sequenced at a depth of approximately 10,000 mean reads per cell, in line with 10X Genomics’ recommendations for Flex gene expression datasets.  After sequencing FLEX samples, FASTQ files were processed with the 10x Cell Ranger multiplex Flex (1 probe barcode/sample) workflow, performing alignment, filtering, and unique molecular identifier counting through the “multi” function and outputting per-sample gene-barcode count matrices. The reference human genome GRCh38 was used for mapping along with the associated 10X Genomics probe set, Chromium Human Transcriptome v1. Further analyses were carried out using the Seurat v5 R package.


title: Microglia integrated neural spheroids enable neuroinflammatory responses and correct network dysfunction induced by alpha-synuclein mutation.

city: Rockville

first: Marc

last: Ferrer

address: 9800 Medical Center Dr.

department: NCATS (National Center for Advancing Translational Sciences)

institute: NIH (National Institutes of Health)

summary: Microglia are the primary resident immune cells of the brain, playing both protective and deleterious roles in neurological diseases, which make them an enticing therapeutic target. Here we developed a Neural Microglia Integrated Multicellular iPSC-derived Cultured Spheroids (NeuroMIMICS) system that enables measurements of both neural network activity and immune responses using human iPSC-derived microglia, astrocytes, and neurons. We validated microglia functionalities in these neural tri-cultures including phagocytic activity, directed motility, and inflammatory responses. RNAseq analysis revealed a phenotypical acquisition of immune functionality via microglia incorporation, supported by increased cytokine production in spheroids challenged with pathogen-like insults.

overall_design: RNA-seq profiling of spheroids of either microglia, neural co-culture spheroids of neurons and astrocytes, or neural triculture spheroids of neurons, astrocytes, and microglia

first: Jiajing

last: Zhang

first: Yi Wei

last: Lim

first: Angelica

last: Medina

first: Yu-Chi

last: Chen

first: Yining

last: Wang

first: Martin

last: Carrasco Carvajal

first: Emily

last: Lee


title: Single-cell transcriptomics provides insights into the stress-immune interplay and informs disease risk

city: Cary

first: Sofia

last: Benavides

state: NC

address: 1134 Dominion Oak Cir

institute: UNC Chapel Hill

summary: Environmental stress contributes to many disease conditions – including cardiovascular disease, mental illness, and autoimmune disorders – through immune dysregulation. Stress operates through the hypothalamic-pituitary-adrenal and sympathoadrenal axes, leading to peripheral secretion of cortisol and norepinephrine, respectively. While stress hormones can profoundly influence immune function, their mechanistic effects at the cell-type level remain poorly understood. To address this knowledge gap, we employed an ex vivo model investigating the transcriptomic responses of primary peripheral blood mononuclear cells and neutrophils to physiological stress levels (i.e., levels reached during in vivo stress) of cortisol and norepinephrine at single-cell resolution.  We identified novel stress-hormone-dependent and cell-type-specific transcriptomic changes, including uniquely regulated genes and inflammation-related signatures within distinct innate (e.g., CD163+ monocytes/macrophages, neutrophils) and other immune cell types. The single-cell-derived signatures effectively distinguished individuals with stress-related diseases in two independent human cohorts. Our findings provide cell-type-level insights into how stress hormones modulate immune function relevant to stress-related conditions.

overall_design: Human peripheral blood mononuclear cells (PBMCs) and neutrophils were treated with cortisol, norepinephrine, or DMSO (control) for 24 hours (PBMCs) or 4 hours (neutrophils), followed by single-cell RNA sequencing.

first: Oksana

last: Kosyk

first: Haiyang

first: Alexander B.

last: Bastian

first: Christina

last: Georgiou

first: Brian C.

last: Miller

first: Natalie

last: Stanley

first: Anthony S.

last: Zannas


title: Lactylation-Dependent DDX18 Nucleolar Escape Regulates CD44 mRNA Stability to Drive Fibrotic Progression in Kidney Disease [RIP-Seq]

city: Guangzhou

first: Lijun

last: Dong

address: 1023-1063 Shatai South Road, Baiyun District

institute: Southern Medical University

summary: To investigate the function of K116 lactylation of DDX18, we performed RIP-seq analysis in DDX18-deficient HK-2 cells transfected with either DDX18-WT or the DDX18-K116R mutant plasmid to compare the RNA-binding profiles of DDX18-WT and DDX18-K116R.

overall_design: Comparative analysis of the RNA-binding profiles of DDX18-WT and the DDX18-K116R mutant in H/R-treated DDX18-deficient HK-2 cells based on RIP-seq data.

first: Jingwen

last: Xie

first: Mengyuan

last: Tao


title: Lactylation-Dependent DDX18 Nucleolar Escape Regulates CD44 mRNA Stability to Drive Fibrotic Progression in Kidney Disease

summary: To investigate the function of k116 lactylation of DDX18, we then performed gene expression profiling analysis using data obtained from RNA-seq of DDX18-deficient HK-2 cells transfected with DDX18-WT or the DDX18-K116R mutant plasmid.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for H/R-treated DDX18-deficient HK-2 cells transfected with DDX18WT or the DDX18K116R mutant.


title: Bulk RNA-sequencing after estrogen-related receptorr knockdown (ESRRA) knockdown in human GIST-T1 cell line

city: Philadelphia

first: Ronald

middle: P

last: DeMatteo

address: 3400 Spruce Street

department: Surgery

institute: University of Pennsylvania Perelman School of Medicine

summary: Gastrointestinal stromal tumor (GIST) is the most common sarcoma and primarily driven by a gain-of-function mutation in either the KIT or PDGFRA receptor tyrosine kinase. Estrogen-related receptor alpha (ERRα, encoded by ESRRA) is an orphan nuclear receptor whose role in GIST is unknown. We found that ERRα was highly expressed in human GIST cell lines and human GIST surgical specimens. Pharmacologic inhibition of ERRα using XCT90 or siRNA-mediated knockdown reduced viability, colony formation, and migration of human GIST cell lines.To further examine ERRα's role in regulating gene transcription in  human GIST, we performed RNA-sequencing after siRNA-mediated knockdown of ERRα to identify changes in mRNA expression. We also sequenced T1 cells after imatinib treatment to compare the two treatments with each other in addition to the non-target controls.

overall_design: GIST-T1 cellswere plated in replicates of three. The three treatment groups included  1) Non-target control:  transfected with  non-target siRNA  2) ESRRA knockdown: transfected with ESRRA siRNA  3) Non-target+imatinib: transfected with  non-target siRNA and 50 nM imatinib  Cells were harvestd 48h later and the nine samples were processed for RNA sequencing. Bulk RNA sequencing was performed using Illumina HiSeq 2500 platform.

first: Taylor

last: Hartlein

first: Jonathan

middle: H

last: Sussman

first: Juan

last: Perez


title: Detection of N3-methylcytidine in RNA at single base resolution using Protoscript II to read through and generate high mutation rates [m3C_tRNA]

city: Chicago

first: Caraline

last: Sepich

state: Illinois

address: 929 E 57th Street

department: Chemistry

institute: University of Chicago

summary: N3-methylcytidine (m3C), an RNA modification involving methylation of cytidine at the Watson-Crick face, has been identified in human tRNA, mitochondrial tRNA (mt-tRNA), and mRNA. In tRNA and mt-tRNA, it has been shown to play important roles in tRNA structure and translation. m3C remains understudied in mRNA and other low-abundance RNAs due to its markedly lower levels, the lack of high-resolution techniques to reproducibly map its location, and under-characterization of its effector proteins on these RNA types. Here we describe a new approach which utilizes the high readthrough and mutation rates of Protoscript II (PSII) at m3C sites in combination with AlkB demethylation to detect m3C at single-base resolution. We first validate the method on synthetic probes. Then, using this method, we successfully identify the known m3C sites on tRNA and mt-tRNA and show that none of the m3C sites identified in cytosolic tRNA are likely METTL8 substrates. In contrast, the two m3C sites detected in mt-tRNA are installed by METTL8 since they are sensitive to METTL8 knockdown (KD) and over expression (OE). These results demonstrate that the method can be used to investigate changes in the m3C level in many sites using an unbiased approach. Overall, this study introduces a convenient high-resolution method for m3C mapping with high readthrough rates, paving the way for future mechanistic studies on the biological function of m3C and its effector proteins.

overall_design: 16 samples: Three input samples and one AlkB-treated sample from each of the following HeLa cells: shCtrl, shMETTL8, Control-overexpression, METTL8-overexpression. 3’ RNA adaptor: 5-’rApp-NNNNNATCACGAGATCGGAAGAGCACACGTCT/3BioTEG/-3’; SR RT primer: 5'-AGACGTGTGCTCTTCCGATCT; 3’ cDNA adaptor: 5'-/Phos/NNNNNGATCGTCGGACTGTAGAACTCTGAAC/3SpC3.

first: Hui-Lung

last: Sun

first: Kinga

last: Pajdzik

first: Chang

last: Ye

first: Xiaoyang

last: Dou

first: Boyang

last: Gao

last: Sepich-Poore

first: Qing

last: Dai

first: Chuan

last: He


title: Transcriptional profiling of retinoic acid responses in stem cell-enriched organoids

city: Cincinnati

first: Theresa

last: Alenghat

state: Ohio

address: 3333 Burnet Avenue

department: Division of Immunobiology

institute: Cincinnati Children's Hospital Medical Center

summary: To define retinoic acid-responsive transcriptional programs in the intestinal stem cells, we performed RNA sequencing on stem cell-enriched intestinal organoids following 24-hour retinoic acid treatment. This dataset provides a resource for examining gene expression changes associated with retinoic acid signaling in intestinal stem cell-enriched epithelial cultures.

overall_design: Ileal crypts were isolated from wild-type mice and cultured as stem cell-enriched intestinal organoids in growth media supplemented with 10 μM CHIR99021 for 96 hours, followed by treatment with 10 μM retinoic acid or vehicle control for 24 hours. RNA was extracted and subjected to bulk RNA sequencing on an Illumina platform. Reads were processed, aligned to the mouse reference genome (mm10), and gene-level counts were generated for downstream differential expression analysis comparing retinoic acid-treated and control organoids.

first: Eshleman

last: Emily M

first: Alenghat

last: Theresa


title: Phagosome-mediated activation of STING by purine and pyrimidine-based bacterial cyclic dinucleotides

city: Boston

first: Stephanie

last: Ragland

state: MA

address: 1 Blackfan Circle

department: Gastroenterology

institute: Boston Children's Hospital

summary: The cytoplasm harbors a spectrum of microbial sensory proteins. Beyond pathogens and their membrane disruptive activities, the contexts and mechanisms whereby bacterial molecules access cytoplasmic sensors are unknown. In this study, we describe the intracellular cyclic dinucleotide (CDN) sensor STING from humans, mice, and other animals as a driver of immune responses to a spectrum of CDNs produced by evolutionarily diverse bacteria, including those containing pyrimidine bases. Diverse bacterial CDNs were present in murine tissues and increased in human inflammatory bowel diseases patients. After bacteriolysis in phagolysosomes, bacterial CDNs could activate STING via two CDN transporters. Systemic delivery of dead, CDN-laden bacteria promoted resistance to subcutaneous and liver-localized tumors in mice. X-ray crystallography of STING-CDN complexes revealed distinct binding modes for purine- and pyrimidine-based CDNs. We propose a connection between phagocytosis, bacterial CDNs, and STING that ensures cytoplasmic innate immune responses are a common outcome of host-bacteria interactions.

overall_design: Immortalized macrophages (iBMDMs) of indicated genotypes were exposed to bacteria (different species indicated or ClearColi expressing (or not) the indicated CDN synthases) for the indicated time points

first: Katherine

last: Lindblad

first: Yao

last: Li

first: Joon

last: Choi

first: Daniel

last: Okin

first: Joshua

last: Mao

first: Desmond

last: Richmond-Buccola

first: Daniela

last: Coronado

first: Madison

last: Brown

first: Scott

last: Snapper

first: Philip

last: Kranzusch

last: Kagan


title: KPNA3 drives temozolomide resistance in glioblastoma by upregulating MGMT and activating STAT3 to sustain glioma stem cells

first: Yanru

last: Gao

address: 81 Meishan Road

institute: Anhui Medical University

summary: Glioblastoma (GBM) invariably develops resistance to temozolomide (TMZ), the frontline chemotherapeutic agent, leading to treatment failure. The molecular mechanisms underlying this resistance remain incompletely understood. Here, we identify karyopherin subunit alpha 3 (KPNA3) as a novel and critical driver of TMZ resistance. Through integrated bioinformatics analysis of temozolomide-resistant glioma cells (SF126R) and patient databases, we found KPNA3 expression is elevated in TMZ-resistant contexts and correlates with poor prognosis in TMZ-treated patients. Functional studies demonstrated that KPNA3 knockdown in resistant cells impaired malignant behaviors, re-sensitized cells to TMZ-induced apoptosis, and suppressed glioma stem cell (GSC) properties, including sphere formation and expression of stemness factors (OCT4, ALDH1A1). Mechanistically, KPNA3 promotes resistance via a dual pathway: it upregulates the expression of the DNA repair enzyme MGMT and activates the STAT3 signaling pathway. KPNA3 knockdown reduced both MGMT levels and STAT3 phosphorylation (Tyr705). Furthermore, pharmacological inhibition of STAT3 mirrored the effect of KPNA3 knockdown by potently inhibiting GSC generation. Clinically, KPNA3 and p-STAT3 protein levels were positively correlated in high-grade glioma tissues. Collectively, our findings unveil KPNA3 as a central regulator that concurrently enhances DNA repair and sustains the GSC population to foster TMZ resistance, nominating it as a promising therapeutic target for overcoming chemoresistance in GBM.

overall_design: RNA-seq analysis of sensitive SF126 and U118MG cells and their derived temozolomide-resistant SF126R and U118MGR cells.

first: He

last: Yuan

first: Houling

last: Zhou

first: Chao

first: Suling

last: Sun

first: Chuanjie

first: Zhiyou

last: Fang


title: Detection of N3-methylcytidine in RNA at single base resolution using Protoscript II to read through and generate high mutation rates [m3C_screen]

first: Caraline

last: Sepich

overall_design: Libraries were prepared beginning with a mixture of total RNA (0.5 μg), small RNA (1.5 μg) and synthetic RNA oligonucleotides to test different reverse transcriptases (HIV WT RT, HIV-1306 RT, PSII). Additionally, control library was prepared by treating RNA first with AlkB demethylase followed by RT with PSII.

first: Hui-Lung

first: Kinga

last: Pajdzik

first: Chang

last: Ye

first: Xiaoyang

last: Dou

first: Boyang

last: Sepich-Poore

first: Qing

last: Dai

first: Chuan

last: He


title: METTL1-Mediated m7G Modification of Valine tRNAs Drives Metabolic Adaptation in Pancreatic Ductal Adenocarcinoma

city: Monrovia

first: Ajay

last: Goel

state: CA

address: 1218 south 5th avenue

department: Molecular Diagnostics and Experimental Therapeutics

institute: City of Hope National Medical Center

summary: Transfer RNA (tRNA) modifications play a critical role in regulating codon-specific mRNA translation and enabling tumor cell adaptation. The RNA methyltransferase METTL1 installs N7-methylguanosine (m⁷G) modifications on tRNAs, thereby shaping codon usage and translational output. However, the function and mechanistic contribution of the METTL1–tRNA axis in pancreatic ductal adenocarcinoma (PDAC) remain poorly defined. Here, we show that METTL1 is overexpressed in PDAC tissues and that elevated METTL1 expression is associated with poor patient survival. Genetic ablation of METTL1 markedly suppresses PDAC cell proliferation, migration, and tumor growth in vitro and in vivo. Mechanistically, METTL1 loss selectively reduces m⁷G-modified valine tRNAs – particularly, Val-AAC, Val-CAC, and Val-TAC – leading to impaired translation of valine-enriched oxidative phosphorylation transcripts. As a consequence, METTL1 deficiency disrupts mitochondrial respiration and energy production in PDAC cells. Consistent with this model, valine tRNA levels are elevated in PDAC tissues, and their selective depletion phenocopies METTL1 loss by impairing mitochondrial bioenergetics and tumor cell fitness. Thus, the METTL1–valine tRNA axis promotes PDAC progression through codon-dependent translational control of mitochondrial complex I and oxidative metabolism. Together, our findings identify a METTL1–tRNA–mitochondrial signaling axis as a previously unrecognized metabolic vulnerability and a promising therapeutic target in pancreatic cancer.

overall_design: Polysome profiling followed by RNA sequencing (polysome-seq) in BxPC-3 and MiaPaCa-2 cells after deleting METTL1.

first: Jiabei

last: Zhu


title: Single-cell transcriptomic profiling unveils tumor-mediated reprogramming of neutrophils and their unique vulnerability that inhibits metastasis

first: Nissim

last: Hay

state: IL

address: 900 South Ashland Ave

department: BMG

institute: University of Illinois Chicago

summary: We performed single cell RNA sequencing (scRNAseq) with the 10x Genomics platform to study how neutrophils are reprogrammed during breast tumorigenesis. Neutrophils acquire a pro-metastatic phenotype induced by signals eminating from the tumor. This comprehensive scRNAseq study assessed neutrophils from various tissues in naive and tumor-bearing MMTV-PyMT mice to uncover how neutrophils are transcriptionally modified. We found that neutrophils are reprogrammed similarly amongst tissues in tumor-bearing mice, and that G-CSF is the main driver of this.

overall_design: Unsorted single cell suspensions were prepared from the BM, blood, lungs, spleen, and tumor from naïve and MMTV-Pymt (Fvb background) tumor-bearing mice. Additionally, we collected BM cells from naive mice treated with vehicle or recombinant G-CSF and MMTV-PyMT mice treated with an IgG control or neutralizing anti-G-CSF antibody (Fvb background). We also isolated BM cells from naive naive and E0771 orthotopically transplanted mice (C57BL/6 background). These cell were processed for scRNAseq. Neutrophils were isolated in silico using well-established neutrophil markers.

first: Will

last: Putzbach

first: Abul

last: Islam

first: Maria

last: Zappia

first: Maxim

middle: V

last: Frolov


title: The U6 snRNA m6A Methyltransferase METTL16 is required for accurate and efficient splicing of Drosophila pre-mRNAs

city: Shang Hai

first: song

last: penghui

address: No. 2005, Songhu Road, Yangpu District

institute: FuDan University

summary: In this study, we generated viable mettl16 knockout Drosophila and GFP knock-in Drosophila via the CRISPR/Cas9 technique, which provides an excellent in vivo model for investigating the precise functions of the Mettl16 protein. Subsequent staining assays revealed that Drosophila Mettl16 is a ubiquitously expressed nuclear protein, and its depletion does not affect the translational level in flies. Loss of Mettl16 results in multiple developmental phenotypes, including developmental delay, reduced locomotor activity, flightlessness, and sterility. Through MeRIP-seq and splicing analyses, we found that Mettl16 mediates m6A methylation of only a small subset of mRNAs. The reduced m6A modification level on its conserved methylation substrate U6 snRNA induces significant global changes in splicing events

overall_design: w1118 adult flies as control(WT) and Mettl161-9 adult flies as experimental group, each genotype included three biological replicates

first: Yan

last: Dong


title: RSPO2 synergizes with BMP2 to enhance wnt-dependent osteoblast differentiation

first: Buchan

last: Jiang

address: 109 Zina Pitcher Pl, Room 2013

department: Department of Orthopaedic Surgery

summary: Canonical Wnt signaling plays a critical role in osteoblast differentiation and bone formation and is enhanced by R-spondin (RSPO) proteins. Bone morphogenetic protein-2 (BMP2) is a clinically used osteogenic factor whose regenerative efficacy is limited by dose-dependent adverse effects. We recently identified RSPO2 as a contextual enhancer of BMP2-induced bone formation, but the transcriptional mechanisms underlying this synergy remain incompletely understood. To define the molecular basis of BMP2–RSPO2 cooperation, we performed RNA sequencing (RNA-seq) analysis of MC3T3-E1 osteoblast precursor cells treated with vehicle control, RSPO2, BMP2, or BMP2+RSPO2 for 5 days under osteogenic conditions. Transcriptomic analysis revealed distinct and overlapping gene expression programs regulated by BMP2 and RSPO2, with co-treatment inducing a unique transcriptional signature enriched for genes associated with osteoblast differentiation, Wnt signaling, and cytoskeletal regulation. These data provide mechanistic insight into how RSPO2 potentiates BMP2-induced osteogenesis and support a role for RSPO2 as a therapeutic co-factor to enhance BMP2-mediated bone regeneration.

overall_design: RNA-seq was performed on MC3T3-E1 osteoblast precursor cells cultured under osteogenic differentiation conditions and treated for 5 days with one of four conditions: Vehicle control (Control), recombinant RSPO2 (RSPO2), recombinant BMP2 (BMP2), combined BMP2 and RSPO2 (BMP2+RSPO2). Each condition included three independent biological replicates (n=3 per group), for a total of 12 samples. The objective of this experiment was to identify transcriptional programs regulated by BMP2 and RSPO2 individually and in combination, and to define molecular mechanisms underlying their synergistic effects on osteoblast differentiation.

first: Michael

last: Daratzis

first: Alexander

middle: J

last: Knights

first: Tristan

last: Maerz

first: Yadav

last: Wagley

first: Kurt

last: Hankenson


title: Non-canonical mTORC1-TFEB activation due to loss of TSC1 determines hepatocyte plasticity and drives cholangiocarcinogenesis

city: Kansas City

first: chen

last: zhang

state: KS

address: 3901 Rainbow Blvd, MS-1018

institute: University of Kansas Medical Center

summary: The aim of the study is to investigate the role and mechanisms of tuberous sclerosis complex 1 (TSC1) and TFEB in liver cancer study。Hepatic mRNA profiles of WT , hepatic TSC1 KO mice and hepatic TSC1 and TFEB DKO mouse liver

overall_design: Comparative gene expression profiling analysis of RNA-seq data for  TSC1 WT and hepatic TSC1 KO,hepatic TSC1 and TFEB DKO mouse livers.

first: WenXing

last: Ding

first: Chen

last: Zhang

first: HongMin

last: Ni


title: Immunization by mobile virus-derived siRNAs is crucial for plant antiviral silencing

city: Zurich

first: Genome

last: Informatics

address: Winterthurerstrasse 190

department: Functional Genomics Center Zurich (FGCZ)

institute: ETH ZURICH / University of Zurich

type: Non-coding RNA profiling by high throughput sequencing

summary: The movement of virus-derived (v)siRNAs between plant cells has prompted the widely held assumption that they help immunize as-yet-uninfected tissues. Ascertaining this role is challenging because it requires uncoupling vsiRNA mobility from viral movement while preserving viral virulence. Consequently, the contribution of non-cell-autonomous RNAi to plant antiviral immunity remains essentially unknown. Here, we investigated the naturally phloem-restricted infection of turnip yellows virus (TuYV) in the stereotypical Arabidopsis root system at cellular resolution. We uncovered intertwined, complex mechanisms operating in both vsiRNA-emitting and -recipient cells, that drive reiterated short-range (~3-4 cell) immunization events. We found that, while intracellular antiviral RNAi is nullified by TuYV, non-cell-autonomous immunization accounts mainly —if not exclusively— for the containment of viral tissue invasion, titers and symptoms severity at the whole-plant scale. Mobile vsiRNA-mediated immunization likewise spatially confines a TuYV unrelated virus over ~3-4 cells, uncovering a generic, essential, yet previously unexplored component of plant antiviral defence.

overall_design: Plants were grown hydroponically following standard procedures developed in our laboratory (Devers. et al, 2020). 3-week-old plants were infected with TuYV/TuYV-GFP through agroinfiltration of source leaves. Infected roots were analyzed at 15 days-post-infiltration (TuYV-WT) or at 21 days-post-infiltration (TuYV-GFP).

first: Daniele

last: Albertini

first: Emanuel

last: Devers

first: Gregory

last: Schott

first: Rajendran

last: Rajeswaran

first: Maëlle

last: Jacquemettaz

first: Olivier

last: Voinnet


title: Fibroblast maturation in burned tissue is modified by neutrophils

city: Madison

first: Anna

last: Huttenlocher

state: WI

address: 1550 Linden Drive

department: Pediatrics

institute: University of Wisconsin School of Medicine and Public Health

summary: Behavior of fibroblasts contributes to fibrosis, but less is known about the burn wound environment and cell-cell interactions that influence fibroblast behavior. Here we identify the temporal overlap between inflammatory and remodeling phases during burn healing. We generate mesenchymal cell reporter fish to investigate the developmental origin and burn wound response of mesenchymal, fibroblast-like cells in larval zebrafish. We show that maturation of these cells is perturbed following burn injury and that the presence of neutrophils in burned tissue is inhibitory to their behavior. This work adds to our understanding of fibroblast development in zebrafish and shows that collagen expressing mesenchymal cells regulate ECM remodeling in coordination with immune cells during burn wound healing.

overall_design: Tail fins from 50 unwounded or burn wounded larvae were pooled and collected in ice cold PBS for each time point. Three biological replicates, representing unique breeding clutches, were collected for each time point. RNA was extracted from pooled fins using TRIzol reagent and RNAqueous Micro Kit (Invitrogen). Extracted RNA was submitted to GENEWIZ for library preparation and sequencing. Pooled RNA libraries were sequenced on Illumina HiSeq 4000 to obtain 150 bp paired end reads.

first: Adam

last: Horn

first: Yiran

last: Hou

first: Jayne

last: Squirrell

first: Alexandra

last: Fister

first: Julie

last: Rindy

first: Veronika

last: Miskolci

first: Jonathan

last: Schrope

first: Russell

last: Burke

first: Colin

last: Dewey

first: Kevin

last: Eliceiri


title: RNA Terminal Uridylyl-Transferases Are Druggable Vulnerabilities in AML, but are Dispensable for Normal Hematopoiesis

city: Cambridge

first: Bertie

last: Gottgens

email: bg200@cam.ac.uk, nkw22@cam.ac.uk

address: Jeffrey Cheah Biomedical Centre

institute: University of Cambridge

summary: Acute myeloid leukemia (AML) is an aggressive hematological malignancy arising from blood-forming hematopoietic stem and progenitor cells (HSPCs). Current therapies often fail to eradicate AML, therefore, identification of novel therapeutic targets is essential. Here, we reveal Terminal Uridylyl Transferase Enzymes 4 and 7 (TUT4/7) as druggable novel therapeutic targets whose genetic deletion in AML suppresses growth, induces apoptosis and improves the survival of in vivo mouse models. Use of a pre-clinical TUT4/7 inhibitor is cytotoxic to AML patient samples, dysregulates mevalonate pathway genes and synergizes with Venetoclax. AML therapies are often limited by hematopoietic toxicity. Remarkably, we find that although Tut4/7 deletion results in mild inflammatory activation throughout the hematopoietic system, this activation is largely permissive, and mice exhibit no overt pathology for at least one year. Thus, we reveal TUT4/7 as novel druggable therapeutic targets, whose inactivation compromises AML while sparing normal hematopoiesis.

overall_design: scRNA-Seq analysis of mouse hematopoietic progenitors from mice: WT or conditional knockout of Tut4 and Tut7 genes (Vav-Cre).

first: Iwo

last: Kucinski

first: Elise

last: Georges

first: Donal

last: O'Carroll

first: Kamil

last: Kranc


title: Cohesin mutations suppress HLA class II gene expression to promote immune evasion during the progression to acute leukemia in children with Down syndrome

city: Memphis

first: Wojciech

last: Rosikiewicz

state: TN

address: 262 Danny Thomas Place

department: Center for Applied Bioinformatics

institute: St. Jude Childrens Research Hospital

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

overall_design: Refer to individual Series


title: Substantial Adaptability of Mononuclear Polyploid Cardiomyocytes Revealed by Perinatal Loss of Chaf1a

city: Shanghai

first: Yuan

last: Min

address: 1239 Siping Road

institute: Tongji university

summary: Here, we show that chromatin assembly factor 1 subunit A (Chaf1a), a key regulator of nucleosome deposition during DNA replication, is essential for postnatal cardiomyocyte proliferation and binucleation. Using cardiomyocyte-specific Chaf1a knockout mice (Myh6-Cre; Chaf1af/f), we demonstrate that Chaf1a deficiency disrupts karyokinesis, leading to a marked increase in the proportion of mononuclear polyploid cardiomyocytes and a reduction in binucleated cells. This results in nearly 50% fewer cardiomyocytes, yet strikingly, the hearts preserve baseline function through substantial compensatory hypertrophy without significant fibrosis. Notably, following cardiac apex resection, neonatal Myh6-Cre; Chaf1af/f mice fail to regenerate myocardium, highlighting the essential role of Chaf1a in cardiac regeneration.

overall_design: RNA-seq profiling of cardiomyocyte-specific Chaf1a knockout mice (Myh6-Cre; Chaf1af/f) and control mice (Chaf1af/f).

first: Ke

last: Wei

first: Zhaohui

last: Ouyang

first: Min

last: Yuan

first: Rongrong

last: Huang


title: LRRK2-G2019S Impairs Astrocyte Differentiation and Triggers a senescence-like phenotype in Parkinson's Disease Models - Brain samples

city: Belval

first: Matthieu

last: Gobin

address: 6, avenue du Swing

country: Luxembourg

institute: LCSB

summary: Astrocytes are increasingly recognized as essential contributors to both physiological brain function and neurodegenerative diseases. Here, we describe how the Parkinson’s disease (PD)-associated mutation LRRK2-G2019S affects astrocytes using autoptic brain samples and PD patient-specific 3D midbrain organoids and 2D astrocytes derived from induced pluripotent stem cells. In autoptic midbrain samples from LRRK2-G2019S patients, we observed a reduction in GFAP⁺ astrocytes but increased branching, together with transcriptional signatures consistent with altered astrocyte function. We also observed delayed astrocyte differentiation in PD patient-specific midbrain organoids, accompanied by altered astrocyte transcriptomic profiles revealed by single-cell RNA sequencing. This defective differentiation contributes to the acquisition of a senescent-like phenotype. In 2D cultures, astrocyte differentiation from LRRK2-G2019S precursor cells was associated with early apoptosis and altered Wnt/β-catenin and TGFβ signalling compared to LRRK2-WT cultures. Notably, pharmacological activation of the developmental transcription factor NR2F1, downregulated in LRRK2-G2019S models, reduced astrocyte cell death and senescence-like phenotypes. Together, these data show that LRRK2-G2019S impairs astrocyte specification and predisposes to a senescent phenotype.

overall_design: Data on the impact of LRRK2-G2019S on early astrocyte development and fate specification are limited. To investigate further this aspect, we used autoptic brain samples and human neuroepithelial stem cells (NESCs) derived from patient-specific iPSCs carrying the LRRK2-G2019S mutation and from healthy individuals, both with their isogenic controls. NESCs were either differentiated into astrocytes in 2D or used for the generation of midbrain organoids. We showed that astrocyte differentiation was impaired both in 3D midbrain organoids and 2D cultures from PD patients carrying the LRRK2-G2019S mutations compared to isogenic lines and healthy individual controls. Using single-cell RNA-sequencing (scRNA-seq) data, we observed a significantly increased expression of astrocyte-specific genes at 35 days in LRRK2-G2019S organoids compared to LRRK2-WT, which resulted in an abnormal developmental trajectory. This defective astrocyte specification is associated with the acquisition of a senescent phenotype in LRRK2-G2019S organoids. In summary, we propose a novel role of the LRRK2-G2019S mutation in impairing astrocytic differentiation and predisposing to senescence.

first: Lisa

last: Smits

first: Stefano

last: Magni

first: Jens

last: Schwamborn

first: Silvia

last: Bolognin


title: Alpha retinal ganglion cell-derived SPP1 instructs microglia to limit neurodegeneration

first: Tatjana

last: Jakobs

address: 20 Staniford Street

department: Howe Laboratory of Ophthalmology

institute: Massachusetts Eye and Ear Infirmary

summary: Communication between neurons and microglia is essential for microglial functions and neuronal survival.  Here we report that alpha retinal ganglion cells (αRGCs) respond to glaucomatous or traumatic axonal injury by increasing the expression and secretion of secreted phosphoprotein 1 (SPP1), which in turn instructs retinal microglia to assume a neuroprotective phenotype. Alpha RGCs increase the secretion of SPP1, which acts on its specific receptor, ItgαV, on microglia and elicits transcriptional and morphological changes. SPP1 functionally increases the capacity of microglia to clear protein aggregates and inhibits neuroinflammation. SPP1 positively regulates microglial debris clearance through an upregulation of autophagy-selective receptors. This shifts microglia towards the expression of neuroprotective mediators, such as interleukin-10, protecting ganglion cells and limiting vision decline. Our findings identify SPP1 as a key mediator of αRGC-microglia crosstalk in autophagy and debris clearance, and a promising candidate for therapy in neurodegeneration.

overall_design: Isolated retinal microglia from wild-type and Spp1 knock-out mice. Baseline condition (naïve), 4 weeks after induction of experimental glaucoma (high IOP), and 7 days after optic nerve crush (ONC).

first: Song

last: Li


title: Fragmentation Patterns of Human Telomeric Chromatin in Plasma cfDNA

city: Aurora

first: Srinivas

last: Ramachandran

state: CO

department: Biochemistry and Molecular Genetics

institute: University of Colorado School of Medicine

summary: The unique chromatin structure at telomeres protects the ends of our chromosomes from being recognized as DNA damage and preserves genome integrity by preventing recombination. Here, we demonstrate that circulating cell-free DNA (cfDNA) from plasma can be used to map chromatin structure at telomeres. Using an alignment-free approach to overcome challenges in characterizing repetitive regions of DNA using sequencing, we find that the telomeric 6-mer repeats (TTAGGG/CCCTAA) are the most abundant circulating 6-mers in cfDNA. Telomeric sequences in cfDNA contain subnucleosomal footprints distinct from the rest of the genome, arising from specific cleavages in the C-rich strand. To map nucleosomal footprints from telomeric sequences in cfDNA, we performed long-read sequencing. We identified a nucleosome repeat length of 145 bp for telomeric sequences, which is significantly shorter than the repeat length of ~170 bp for the rest of the genome. The abundance of cfDNA telomeric footprints decreases with age, and this decline is exacerbated by Dyskeratosis Congenita (DC), a telomere biology disorder. DC also alters subtelomeric chromatin accessibility, as determined from cfDNA, allowing us to capture genome-wide effects of shortened telomeres non-invasively. Promoter subnucleosome enrichment (PSE) from cfDNA identifies DC-specific gene signatures that reflect disease states observed in transcriptomic studies. Notably, DC-specific gene signatures correlate with decreased cfDNA telomere footprint abundance and show enrichment of genes towards the ends of chromosomes. In summary, cfDNA provides an unexpected means to map telomere chromatin state and the impact of telomere shortening on the rest of the genome, offering a non-invasive way to track gene expression changes in DC.

overall_design: Plasma cfDNA was isolated and sequenced from healthy donors. Illumina 150 bp paired-end and Oxford Nanopore Long Read Sequencing conducted on the human blood plasma cfDNA.

first: Brandon

last: Buck

first: Abigail

last: Weirich

first: Stephanie

last: Eramo


title: Dysregulated ribonucleoprotein granules impair mitochondrial activity in RBM20-dilated cardiomyopathy

city: Heidelberg

first: Lars

last: Steinmetz

state: Baden Wurttemberg

address: Meyerhofstraße 1

department: Genome Biology Unit

country: Germany

institute: EMBL

summary: Pathogenic variants in the RNA-binding motif protein 20 (RBM20) cause severe dilated cardiomyopathy (DCM). While loss-of-function (LoF) variants lead to splicing dysfunction, gain-of-function (GoF) variants also mislocalize the protein to cytoplasmic ribonucleoprotein (RNP) granules resulting in a more aggressive disease phenotype with a yet unknown pathogenic mechanism. Here, we show that the induction of severe mitochondrial dysfunction is a key consequence of RBM20 mislocalization. Proteomic profiling of RBM20-GoF mouse hearts reveals a predominant reduction in mitochondrial membrane protein solubility and widespread post-transcriptional downregulation of mitochondrial proteins, including Transmembrane Protein 65 (TMEM65), a regulator of mitochondrial calcium efflux. Ultrastructural analysis shows enlarged mitochondria with aberrant morphology. Functional assays confirm impaired mitochondrial respiration, transmembrane potential, and calcium handling in RBM20-GoF, but not LoF models. Using human iPSC-derived cardioids, we validate that RBM20 mislocalization, not splicing deficiency, drives mitochondrial dysfunction in DCM. These findings establish a mechanistic link between dysregulated RNP granules and mitochondrial defects in RBM20-DCM, resembling pathogenic features observed in Fused in Sarcoma protein (FUS)-associated neurodegeneration causing amyotrophic lateral sclerosis (ALS), where cytoplasmic mislocalization of an RNA-binding protein disrupts granule dynamics and organelle function.

overall_design: Libraries for total RNA-sequencing were prepared from D30 cardioids. Total RNA was isolated by douncing cardioids to generate cell suspensions, followed by RNA extraction using Trizol LS according to the manufacturer’s instructions (Thermo Fisher). RNA samples were subjected to RNase H-based rRNA depletion using the NEBNext rRNA Depletion Kit v2 (Human/Mouse/Rat). Ribo-depleted RNA was then used for library preparation with the NEBNext Ultra™ II Directional RNA Library Prep Kit for Illumina. Paired-end sequencing (101 bp reads) was performed on the Illumina NextSeq2000 platform. Sequencing data were processed using the RNA-sequencing module from snakePipes72 , with the flags --trim and -m "alignment-free,alignment". Reads were mapped to the Homo sapiens reference genome (Ensembl assembly GRCh38) and transcriptome annotation release 114 using STAR. For alternative exon usage analysis, we used DEXSeq73, and significant exons were defined by an adjusted p-value < 0.05. RNA stability analysis was performed using the default eisar workflow74. For all analyses, comparisons were made using the model formula ~batch + condition to account for batch-specific effects of cardioid differentiation.

first: Julia

last: Kornienko

first: Linda

last: Müller

first: Alexander

last: Nickel

first: Michael

last: Kohlhaas

first: Mandy

last: Börmel

first: Carlos

last: Alfonso-Gonzalez

first: Viola

last: Oorschot

first: Michel

last: Fungate

first: Susanne

last: Lewey

first: Jacob

last: Kees

first: Dominik

last: Lindenhofer

first: Kai

last: Fenzl

first: Laura

last: Schraft

first: Sandra

last: Clauder-Münster

first: Per

last: Haberkant

first: Frank

last: Montoya

first: Vita

last: Dauksaite

last: Gotthardt

first: Maarten

last: van den Hoogenhof

first: Christoph

last: Maack


title: Co-targeting EZH2 and TEAD elicits apoptosis through tumor-intrinsic innate immune signaling in Hippo pathway-mutated cancers - ChIP-seq data from genetic disruption of EZH2

first: Zhaorong

last: Li

address: 60 Fenwood Road

institute: Dana Farber Cancer Institute

summary: TEA/TEF-domain [TEAD] inhibitors are being evaluated in clinical trials for cancers with alterations in the Hippo pathway including mesothelioma. We recently developed and showcased the potency of TEAD palmitoylation inhibitors MYF-03-69 and MYF-03-176 in mesothelioma cell lines. However, TEAD inhibition results in cell cycle arrest in cell line models with Hippo pathway alterations without inducing cell death, potentially limiting their long-term clinical efficacy. Using a genome-wide CRISPR/Cas9 screen, we identified EZH2 as a critical modulator of the cellular response to TEAD inhibition. Compared to single agent treatments, EZH2i/TEADi robustly triggered apoptosis and suppressed the growth of Hippo-mutated cells in vitro and in vivo. Mechanistically, EZH2i/TEADi-treated cells exhibited heightened activation of tumor-intrinsic innate immune signaling which resulted in DNA damage and subsequent apoptosis. Taken together, we propose this novel combinatorial strategy as a potential approach to enhancing the anti-tumor efficacy of single agent TEAD targeting therapies in Hippo pathway altered tumors. This GEO accession includes the data series associated with ChIP-seq derived from genetic disruption of EZH2.

overall_design: MSTO-211H malignant pleural mesothelioma cells were transiently transfected with Cas9 and either an sgRNA control guide (sgCTRL) or a guide targeting EZH2 (EZH2-KO). ChIP-seq profiling of sgCTRL and EZH2-KO cells  treated with DMSO or a TEAD inhibitor, MYF-03-69 for 24h, was then performed.

first: Antja-Voy

last: Hartley

first: Shweta

last: Kukreja

first: Sonsoles

middle: L

last: Veiga

first: Rong

first: Xintao

last: Qiu

first: Henry

middle: W

last: Long

first: Pasi

last: Jänne


title: Identification of a novel isoform of Slc26a4 by single-cell RNA-sequencing of pendrin-expressing cells in the cochlea

city: Baltimore

first: Jin-Young

last: Koh

state: MD

address: 670 W Baltimore Street

department: Otorhinolaryngology-Head & Neck Surgery

institute: University of Maryland, Baltimore

overall_design: Refer to individual Series.


title: Identification of a novel isoform of Slc26a4 by single-cell RNA-sequencing of pendrin-expressing cells in the cochlea - Nanopore

summary: Pathogenic variation in the coding sequence of SLC26A4 causes both Pendred syndrome (PDS) and non-syndromic enlarged vestibular aqueduct (NSEVA/DFNB4), two autosomal recessive disorders. The former accounts for approximately 6% of human genetic hearing loss, making it the second most common form of syndromic deafness after Usher syndrome, while the latter is the most common radiological malformation associated with childhood sensorineural hearing loss (SNHL). Here, we used short- and long-read single-cell RNA sequencing (scRNA-seq) of pendrin-expressing cells in the murine cochlea to identify a novel short isoform of Slc26a4. We demonstrate that the short Slc26a4 isoform is expressed in both the inner ear and kidney and investigate its interactions and functions. We also characterize the genotype-phenotype association for SLC26A4-related hearing loss in the context of these two isoforms. These results provide a new reference for molecular profiling of pendrin and offer novel insights into cell-type-specific splicing events and SLC26A4-related hearing loss.

overall_design: Spindle/root cells of 129S6 WT mice were isolated and analyzed using Illumina short-read sequencing and Oxford Nanopore long-read sequencing.

first: Richard

last: Smith


title: Identification of a novel isoform of Slc26a4 by single-cell RNA-sequencing of pendrin-expressing cells in the cochlea - Illumina


title: Receptor-tethered orthogonal IL-2 enhances regulatory T cell therapy

city: San Francisco

first: Huiyun

last: Lyu

state: CALIFORNIA

address: 513 Parnassus Ave

department: Diabetes Center

institute: University of California, San Francisco

summary: Regulatory T cell (Treg) therapy is an emerging platform for controlling immune overactivation. The efficacy of Treg therapy is limited by the poor persistence of infused Tregs due to insufficient IL-2, which is essential for Treg survival and function. IL-2 activates many immune cells besides Tregs, imposing a challenge for the selective provision of IL-2 to infused Tregs. In this study, we found infusions of orthogonal (ortho) IL-2 failed to enhance Tregs engineered with a corresponding orthoIL-2 receptor (IL-2R) in a mouse model of autoimmune diabetes. We then developed a receptor-tethered orthoIL-2 by optimizing the combination of IL-2, IL-2R, and the linker connecting them to achieve autocrine signaling selectively in engineered Tregs. Tregs expressing the tethered orthoIL-2 showed autocrine IL-2 signaling in vitro, enhanced CD25, CTLA-4, and Foxp3 expression, persisted without exogenous IL-2 in vivo, and prevented autoimmune diabetes using as few as 2,000 Tregs. Knocking the tethered orthoIL-2 construct into the Foxp3 locus enabled Treg-specific expression, with the additional benefit of positively reinforcing tethered orthoIL-2 expression through the activation of the Foxp3 gene by enhanced IL-2 signaling. Together, these results illustrate a safe and effective cell-engineering solution for overcoming Tregs’ dependency on exogenous IL-2, thereby achieving superior therapeutic efficacy.

overall_design: we compared transcriptomic profiles of 3A10t- and 1G12t-transduced Tregs. NOD Tregs were transduced with EV as a control. On day 2 after transduction (day 4 after activation), the cells were washed to remove exogenous IL-2 and cultured for 5 additional days. Half of the EV-transduced cell was left in IL-2-containing medium as a positive control. The transduced cells were FACS purified based on the expression of Thy1.1 prior to RNA extraction and bulk RNAseq.

first: Alyssa

last: Indart

first: Qizhi

last: Tang


title: Charting the transition from in vitro gliogenesis to the in vivo maturation of human glial progenitor cells transplanted into the hypomyelinated mouse brain

city: Rochester

first: Steven

last: Goldman

state: NY

address: 601 Elmwood Ave

department: Center for Translational Neuromedicine

institute: University of Rochester Medical Center

summary: Neither rodent models nor in vitro studies of human cells adequately reflect the molecular regulation of human glial progenitor cell (hGPC) expansion, differentiation, and myelination in vivo. To better understand this process, we used scRNA-seq of >60,000 human cells to track the differentiation of hGPCs from pluripotent stem cells (PSCs), both in vitro and in vivo. In vitro, the hGPC pool comprised 4 transcriptionally-distinct subpopulations, the heterogeneity of which was associated with histone modifications of stage-dependent genes. After the neonatal transplant of these progenitor cells into immunodeficient and myelin-deficient shiverer mice, they differentiated further; when extracted back from their host brains, scRNA-Seq revealed that the donor cells were comprised solely of hGPCs, astrocytes and oligodendrocytes. A combination of gene co-expression, motif enrichment, cell-trajectory, and cell-cell interaction analyses revealed that the tissue environment potentiated the further, context-dependent differentiation of hGPCs, via ligand-mediated activation of gene regulatory networks. Together, these data chart the process by which manufactured human GPCs differentiate into mature astrocytes and oligodendrocytes in vivo, while establishing the transcriptional and phenotypic stability of these cells following transplantation.

overall_design: We ran 10X Chromium 3' scRNA seq on samples consisting of either WA09 (ESC) or C27 (iPSC) lines that were differentiated and  sequenced at the pluripotent, GPC, or post murine engraftment stages.

first: John

last: Mariani

last: Schanz

first: Benjamin

last: Mansky

first: Xiaolu

last: Wei

first: Carter

first: Devin

last: Chandler-Militello

first: Nguyen

last: Huynh


title: Immune dysregulation and stem-like CD8+ T cell enrichment in type 1 diabetes pancreatic lymph nodes

city: Gainesville

first: Todd

last: Brusko

state: FL

address: 1275 Center Dr BMSB J515

department: Pathology

institute: University of Florida

summary: Effector CD8+ T cells are key drivers of type 1 diabetes (T1D) pathogenesis, yet questions remain regarding the molecular defects leading to altered cytotoxicity, peripheral tissue phenotype, and receptor specificity. We analyzed human pancreatic lymph nodes (pLN) using mass cytometry and single-cell RNA sequencing (scRNAseq) with combined T cell receptor (TCR) profiling. Cytometric analysis revealed enrichment of T stem-cell memory (TSCM)-like cells (CD8+CD45RA+CD27+CD28+CCR7+CXCR3+) in T1D pLNs. scRNAseq indicated an elevated inflammatory cytokine gene signature (IFITM3, LTB) along with regulators of terminal differentiation (BCL6, BCL3), coupled with downregulation of exhaustion-associated genes (DUSP2, NR4A2, TSC22D3) in CD8+ T cells in T1D pLN. Immune Response Enrichment Analysis (IREA) indicated IL-15 signaling as a significant driver of these phenotypes. Integrated TCR and transcriptomic analysis revealed a cluster of diverse naïve-like CD8+ T cell clones in T1D pLN. Comparison of pLN and pancreatic slice isolates supported sharing of effector CD8+ T cells, with enhanced terminal effector signatures within the pancreas relative to paired pLN. Multiplex imaging revealed differential localization of TCF1 and TOX-expressing T cells in the pancreas, with islet-proximal TCF1+TOX+ cells displaying a mixture of activation and exhaustion-associated phenotypes. Thus, we provide multimodal cellular profiles enriched in T1D tissues for consideration in therapeutic targeting.

overall_design: Single cell RNA sequencing with T cell receptor (TCR) sequencing was conducted on pancreatic lymph node from donors with Type 1 Diabetes (n=9) as well as donors without diabetes (n=7) using the 10x genomics platform.

first: Leeana

last: Peters

first: Howard

last: Seay

first: Justin

first: Amanda

last: Posgai

first: Reed

last: Berkowitz

first: Clive

last: Wasserfall

first: Mark

last: Atkinson

first: Rhonda

last: Bacher

first: Maigan


title: Complete phenotype rescue through the restoration of full-length dystrophin using CRISPR/Cas9 genome editing in Duchenne muscular dystrophy patient-derived iPSCs carrying the deletion of two exons

city: Milan

first: Mattia

last: Chiesa

address: Via Parea 4

department: Bioinformatics and Artificial Intelligence facility

country: Italy

institute: Centro Cardiologico Monzino IRCCS

summary: Duchenne (DMD) muscular dystrophy is an X-linked disease affecting skeletal and cardiac muscle due to deficient dystrophin protein expression. Here we describe the full-length dystrophin restoration by CRISPR/Cas9 in an induced pluripotent stem cell (iPSC) line from a patient carrying a deletion of exons 49-50.  Using sinle nuclei-RNA seq we compared the transcriptomic profile of engineered heart tissue (EHTs) derived from original Duchenne iPSC cells and edited iPSC. This analyses allowed us to discriminate the effects of dystrophin restoration in specific cell types present in EHTs.

overall_design: EHTs were generated from  cardiomyocytes derived from original DMD (DMD1) and edite (ISO-50) iPSCs. After 50 days of colture EHTs were dissocieted and nuclei were isolated.

first: Davide

last: Rovina

first: Josè

last: Pioner

first: Angela

last: Maione


title: Spatial transcriptomics stereo-seq was used to reveal the functional compartmentalization of Unicentric Castleman Disease.

city: Singapore

first: Zexi

last: Guo

address: 90 Tanglin Halt Road

country: Singapore

institute: National Cancer Centre Singapore

summary: The cell-of-origin of UCD and the cell-cell communication in lymphnode tissue in UCD context is still not well-studied. We performed stereo-seq to profile heteigeneous cell population within UCD lymphnode tissue as well as to determine their importance for UCD development. Distinct subpopulations of Follicular dendritic cells were identified in the microenvironment that contribute the growth of the disease.

overall_design: Stereo-seq was performed on a frozen section of the diseased lymph node.


title: Gene expression profile at single cell level of lymphnode cells (LN) and peripheral blood mononuclear cells (PBMC)

summary: LN was considered to be the cells of origin of Unicentric Castleman Disease. We used scRNAseq to analuze the microenvironment of LN and PBMC.

overall_design: The diseased lymph node was dissociated into single cells with the human Tumor Dissociation kit (Miltenyi Biotec) and the the PBMCs were isolated from the patient's whole blood. The cells were were then subjected to single cell isolation with the 10x Genomics Chromium iX instrument and single-cell RNA-seq libraries were prepared as per protocol.


title: REV-ERBs agonist SR9009 inhibits liver fibrosis progression in a MASH fibrosis model invivo.

city: Strasbourg

first: Frank

last: Jühling

address: 3 Rue Koeberlé

country: France

institute: Université de Strasbourg, Inserm, Institut de Recherche sur Les Maladies Virales et Hépatiques UMR_S1110

summary: To explore the CC as a therapeutic target for MASH-induced fibrosis, we studied whether SR9009, a pharmacological activator of REV-ERBα, modulates fibrosis in CDA-HFD fed C57Bl/6J mice, a state-of-the-art model for MASH fibrosis

overall_design: Forty male C57BL/6J mice (8-week-old; RRID: IMSR_JAX:000664, Charles River Laboratories) were fed with standard chow diet (n=10) or with a choline-deficient amino-acid adapted high-fat diet (CDA-HFD, Research Diet #A 06071302) (n=30) for 12 weeks. This CDA-HFD diet contains 0.023% Cholesterol (~233 mg/Kg). After 12 weeks of diet, the mice fed with CDA-HFD were randomized in 2 groups, receiving daily IP injections of either SR9009 (100 mg/kg/day) or vehicle control (Kolliphor ELP 15% in injectable NaCl) for 6 weeks (n=15 per group). SR9009 was bulk synthesized (Cayman# 11929) and the entire experiment was performed with the SR9009 prepared from a single batch. All the mice were anesthetized and sacrificed after a total of 18 weeks. Livers were fixed in formalin, mounted in optimal cutting temperature (OCT) compound or snap frozen and kept at -80 °C until further analyses. Experiment was approved by the local ethic committee and authorized by the French ministry of higher education and research (APAFIS #44275-2023072512518363 v3).

first: Romain

last: Martin

first: Laurent

last: Mailly

first: Nicolas

last: Brignon

first: Emilie

last: Crouchet

first: Marine

last: Oudot

first: Cloé

last: Gadenne

first: Thomas

middle: F

last: Baumert


title: MASH fibrosis mouse model for circadian analyses of HSC

summary: Aim was to identify the modulation of circadian clock (CC) genes and their target genes in hepatic cells (HSC) in chronic liver disease and fibrosis

overall_design: HSC were isolated from normal mouse liver of MASH/fibrosis mouse liver at different time points (Zeitgeber ZT0, ZT6, ZT12 and ZT18) for gene expression analysis by RNA-Seq

first: Clara

last: Ponsolles


title: Fat cadherin cleavage releases a transcriptionally active nuclear fragment to regulate target gene expression

status: Public on Jul 16 2026

city: St. Louis

first: Helen

last: McNeill

state: Missouri

address: 660 S. Euclid Avenue

department: Developmental Biology

institute: Washington University School of Medicine in St. Louis

summary: The atypical cadherin fat (ft) controls growth, planar cell polarity (PCP) tissue organization, and mitochondrial function, in organisms ranging from fruit flies to mammals.  Working at the apical-junctional plasma membrane, the intracellular domain of the Ft protein, FtICD, binds to and regulates components of the Hippo and PCP pathways.  Unexpectedly, we find that a fragment of the FtICD is present in the nucleus in cultured cells as well as in embryonic and larval tissues, and we identify nuclear localization and nuclear export signals in FtICD required for this localization.  We show that membrane-bound FtICD is cleaved and enters nuclei in vivo.  Using endogenously tagged Ft as well as overexpressed FtICD, we conducted ChIP-seq experiments and identified putative Ft targets, including genes involved in signaling pathways, chromatin organization, pattern formation, and neural development.  RNAseq demonstrates that some of these genes are dysregulated in ft mutants.  We observe strong co-localization of Ft binding regions with peaks for other factors such as DREF and BEAF-32, as well as the Hippo pathway components Yorkie (Yki) and Scalloped (Sd), suggesting that Ft may act in conjunction with these factors to regulate gene expression.  Supporting this hypothesis, we found that Ft physiclly interacts with both Yki and Sd in co-immunoprecipitation experients in S2 cells.  We propose that the modulation of Hippo pathway activity constitutes one of the nuclear functions of Ft, complementing its estalished function as an upstream regulator of Hippo signaling.

overall_design: RNA-seq profiling of Drosophila melanogaster 16–20 h embryos, third-instar larval imaginal wing discs, and third-instar larval eye-brain complexes. Samples were collected from w^1118 (wild-type), Ubi>FtICD (Ft overexpression), and  ft^fd5-5/ft^X13 (Ft null) mutant genotypes to characterize transcriptomic changes associated with altered fat gene activity across developmental stages and tissues.

first: Jannette

last: Rusch

first: Nattapon

last: Thanintorn

last: Mcneill


title: Transcriptional effects of lysosome stress on mouse bone marrow-derived macrophages and primary microglia

city: South San Francisco

last: Sandmann

email: genomics@dnli.com, sandmann@dnli.com

state: California

address: 161 Oyster Point Blvd

institute: Denali Therapeutics

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

overall_design: Refer to individual Series


title: Integrating machine learning with transcriptomic data for comprehensive assessment of resistance genes against powdery scab in potato

city: Baoding

first: dai

last: zhang

address: No. 2596, Lekai South Street, Lianchi District

department: College of Plant Protection

institute: Hebei Agricultural University

summary: Powdery scab of potato (Solanum tuberosum L.), caused by Spongospora subterranea f. sp. subterranea, is a devastating disease that causes substantial economic losses. However, the molecular mechanisms underlying tuber specific resistance remain poorly understood, largely due to phenotyping bottlenecks and the dynamic nature of infection.

overall_design: We developed an integrated AI omics pipeline combining deep learning based phenotyping with multi algorithm machine learning to systematically dissect resistance against powdery scab. AAF DeepLabv3+, a semantic segmentation network with an adaptive activation function, enabled high throughput, pixel level quantification of disease severity from ~20,000 RGB images across 12 cultivars over a 118 day cycle, achieving 98.56% accuracy. Dynamic monitoring of disease progression throughout the entire potato growth cycle pinpointed tuber initiation and tuber expansion as the critical windows for S. subterranea infection. Integrating phenotypic data with transcriptomic profiling and a consensus ML framework (Random Forest, SVM RFE, and LASSO) prioritized 292 high confidence candidate genes. Functional categorization revealed lignin biosynthesis as a core defense hub, with seven lignin related genes, including CCR and three MYB transcription factors, consistently selected. Cross dataset validation confirmed directional robustness (68.8% concordance, Spearman's ρ = 0.317, p = 0.028). Virus induced gene silencing of St4CL1 and StCCoAOMT1 compromised resistance, confirming lignin's contribution to tuber defense. Beyond lignin, our pipeline ranked several poorly characterized genes, including EDNR2GH7 and DIV3B, as high priority candidates, suggesting additional novel resistance components await functional exploration. This integrative AI omics framework provides a powerful, interpretable strategy for resistance gene discovery and offers a blueprint for accelerating molecular breeding in other pathogen-systems constrained by phenotyping limitations.

first: Dai

last: Zhang