title: Antagonistic roles of histone 3 lysine methyltransferases dictate tumor immune surveillance

city: Philadelphia

first: Amy

last: Gladstein

state: Pennsylvania

address: 421 Curie Blvd Rm 729

department: Cancer Biology

institute: University of Pennsylvania

summary: Histone H3 lysine 36 (H3K36) methyltransferases are recurrently altered in cancer, but how distinct H3K36 methylation states influence tumorigenesis remains incompletely understood. Using KRAS-driven lung cancer models, we show that NSD2, SETD2, and EZH2 form an epigenetic circuit that regulates endogenous retroviral elements (ERVs) and tumor immune surveillance. Although SETD2 is a potent tumor suppressor in KRAS-driven lung adenocarcinoma, expression of the H3K36M oncohistone, which broadly inhibits H3K36 methylation, does not phenocopy Setd2 loss. Instead, H3K36M expression suppresses tumor growth by reducing H3K36me2, derepressing ERV transcripts, inducing cytoplasmic dsRNA, and activating RIG-I/MDA5-dependent antiviral-like immune clearance. Genetic inactivation of histone dimethyltransferase Nsd2 phenocopies H3K36M expression, nominating H3K36me2 loss as a trigger of this viral-mimicry response. In Setd2-deficient contexts, EZH2-dependent H3K27me3 restores ERV repression, prevents dsRNA accumulation, and permits immune escape. Pharmacologic inhibition of NSD2 induces dsRNA in human lung adenocarcinoma cells, whereas concurrent SETD2 inhibition blunts this response while additional EZH2 inhibition restores it. These findings define opposing roles for NSD2- and SETD2-dependent H3K36 methylation in lung tumor immunity and reveal an NSD2–SETD2–EZH2 axis that can be targeted to modulate viral mimicry and immune surveillance in cancer.

overall_design: This is an in vivo pooled CRISPR knockout enrichment screen designed to identify genes whose loss alters fitness during tumor lung colonization in the context of H3K36M mutation.  The experiment consists of two baseline/control parental cell line groups (KP-H3K36M-Ctrl and KP-H3WT-Ctrl) sequenced immediately post-transduction to establish initial sgRNA library representation.  Following tail-vein injection, the in vivo screen tracks selection across 18 individual experimental mice divided into two main biological cohorts:  An H3K36M-mutant cohort consisting of 8 biological replicates (mice IDs 34108, 34109, 34110, 34111, 34112, 34113, 34115, and 34117)  An H3WT wild-type control cohort consisting of 10 biological replicates (mice IDs 34098 through 34107).  The primary design objective is to compare relative sgRNA enrichment and depletion in the lung tumor tissue of the H3K36M mutant model against the H3WT control model, using MAGeCK to normalize all tumor counts back to their respective pre-injection baseline library representation.

first: Akino Mercy

last: Charles Solomon

first: Carson

last: Poltorack

first: Sharan

last: Venkatesh

first: Keren

middle: M

last: Adler

first: Maggie

middle: R

last: Robertson

first: Stephanie

last: Stransky

first: Valerie

last: Irizarry-Negron

first: Dain

last: Ruiz

first: Dingwen

last: Tao

first: Nelson

last: Freeburg

first: Simone

last: Sidoli

first: Irfan

last: Asangani

first: Sydney

last: Shaffer

first: David

last: Feldser


title: Localized PD-1 CAR T therapy reprograms neuroinflammation.

status: Public on Jul 21 2026

city: Rehovot

first: Ido

last: Amit

address: 234 Herzl st.

department: Immunology

country: Israel

institute: Weizmann Institute of Science

type: Expression profiling by high throughput sequencing

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

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Refer to individual Series


title: Temporal phase-resolved transcriptomics reveals host determinants of Marek’s disease virus reactivation in transformed chicken T cells

status: Public on Jul 20 2026

city: Langston

first: haji

last: akbar

address: 100 Success Drive Ag building

department: Agriculture Sciences

country: USA

institute: Langston University

summary: Marek’s disease virus (MDV)  establishes latency in CD4⁺ T cells and can reactivate to initiate productive replication and viral dissemination.To define host and viral mechanisms governing the transition from latency to lytic replication, we developed recombinant MDV strains expressing RLORF4-mRFP and UL47-eGFP reporters. These reporters were used to isolate latent (Lo; mRFP⁻), early lytic (Hi; mRFP⁺), and late lytic (DP; mRFP⁺eGFP⁺) populations from MDV-induced lymphoblastoid cell lines (LCLs) following temperature-induced reactivation. Fluorescence-activated cell sorting-purified populations from two independent LCLs (Lines 82 and 62) were subjected to Illumina RNA sequencing. Viral transcription increased progressively from Lo to Hi to DP populations; however, initiation of viral gene expression differed markedly between cell lines. During the early transition (Lo to Hi), Line 82 exhibited robust induction of 17 viral genes, including US3, UL16, UL51, and UL34, whereas Line 62 showed only two differentially expressed viral genes. In contrast, both lines converged on a highly conserved late-lytic transcriptional program during Lo-to-DP transitions, sharing 118 viral transcripts, dominated by structural and assembly-associated genes. Among the host transcriptional responses, the Line 82 exhibited activation of TP53-associated stress signaling, chromatin remodeling factors, RNA biogenesis pathways, consistent with a permissive cellular state. In contrast, Line 62 displayed enhanced inflammatory, metabolic and proteostasis-associated signatures, suggesting a restrictive environment that limits early viral induction.  Integrated pathway analysis revealed coordinated remodeling of cytoskeletal trafficking, RNA processing, proteostasis, and lipid metabolism during reactivation, while TP53, MTOR, XBP1, NFKB1/RELA, EIF2AK2, DDX58, and STUB1 emerged as candidate host determinants of lytic permissiveness. Collectively, these findings demonstrate that MDV reactivation is a host-gated process in which cellular state governs initiation of the lytic switch, whereas the downstream reproductive replication proceeds through a conserved viral program.

overall_design: RNA-seq was performed on fluorescence-activated cell sorting (FACS)-purified latent (Lo), early lytic (Hi), and late lytic (DP) populations isolated from two independent MDV-transformed chicken lymphoblastoid cell lines (Lines 82 and 62) following low-temperature (32°C)-induced reactivation. Three biological replicates were analyzed for each condition (18 samples total). Libraries were sequenced as single-end 100-nt reads on an Illumina NovaSeq 6000 platform to define stage-specific host and viral transcriptional responses during MDV reactivation.

first: Haji

last: Akbar

first: Yung-Tien

last: Tien

first: Nagendraprabhu

last: Ponnuraj

first: Kathrine

middle: V

last: Etten

first: Keith

middle: W

last: Jarosinski


title: A Blood-Derived Factor Rescues ALS: Platelet Factor 4 Activates OPTN-Dependent Autophagy to Clear SOD1 Aggregates Independently of PINK1

city: Wenzhou

first: Binbin

last: Deng

state: -- Select --

address: 0 Chashan Street, Ouhai District, Wenzhou City,

country: China

institute: wenzhou medical university

summary: Peripheral factors that systemically regulate amyotrophic lateral sclerosis (ALS) have remained elusive-until now. Here, by integrating population-scale epidemiology with mechanistic dissection, we identify platelet factor 4 (PF4) as the central driver of a circulating neuroprotective axis that restores proteostasis and rescues ALS. In a prospective cohort of >500,000 UK Biobank participants, platelet indices were strongly associated with ALS risk, and serum PF4 levels were significantly reduced in ALS patients. Systemic administration of recombinant PF4 in hSOD1G93A mice produced dramatic therapeutic effects: extended survival, preserved motor function, attenuated neuroinflammation, and reduced neuromuscular junction denervation. Remarkably, this efficacy appears pathology-selective--robust in SOD1-driven models but shows no observable effect in TDP-43 or C9orf72 ALS models. Mechanistically, PF4 achieves what few molecules can: it engages the cell surface receptor LRP1 to activate the TBK1-OPTN signaling axis, restoring impaired autophagic flux through a PINK1/Parkin-independent pathway requiring ATG7, establishing a previously unrecognized peripheral platelet-autophagy-neuron axis that facilitates the co-clearance of pathological SOD1 aggregates and damaged mitochondria. This study unveils PF4 as a first-in-class circulating autophagy regulator with therapeutic potential in ALS. Beyond identifying a candidate biomarker and drug lead, it reveals that systemic factors can directly engage central proteostatic machinery--opening a new frontier for ALS therapy.

overall_design: NSC-34 cells were transfected with SOD1G93A plasmid and cultured to the logarithmic growth phase. Four biological replicates were treated with PF4 (100ng/ml) for 24 hours, and four replicates were treated with Vehicle (control). Total RNA was extracted using TRIzol for bulk RNA-sequencing to investigate the transcriptomic changes induced by PF4.

first: Qingjian

last: Xie

first: Yanbo

last: Zhu

first: Wenhua

last: Jiang

first: Haobo

first: Yaojia

last: Li

first: Mengjie

last: Hu

first: Kezheng

first: Huan

last: Yu

first: Yixuan

last: Pan

first: Chaoyi

first: Xueqin

last: Song

first: Dongsheng

last: Fan


title: Plasmodium blood-stage induces trained immunity in hepatocytes - ATAC-seq

city: Seattle

first: Alexis

last: Kaushansky

state: Washington

address: 1900 9th Ave

department: Center for Global Infectious Disease Research

institute: Seattle Children’s Research Institute

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Plasmodium parasites, the causative agents of malaria, are transmitted at high levels in endemic areas and sequential infections are common. Using a mouse model of infection we discovered liver burden was suppressed in blood-stage experienced, compared to naïve, animals, independently of adaptive responses and inflammation. We observed greater chromatin accessibility of a subset of interferon stimulated genes in blood-stage experienced animals, and a rapid increase in transcription of these genes upon sporozoite challenge. Ex vivo stimulation of hepatocytes from blood-stage experienced mice also led to a rapid and elevated response upon treatment with an unrelated antigen, ctDNA. Taken together, these data are consistent with a model in which hepatocytes are reprogramed by blood stage Plasmodium infection to exhibit trained memory akin to what has been described in innate immune cells. The consequences of training of hepatocytes could be wide-reaching and might alter responses to diverse pathogens and other stimuli.

overall_design: ATAC-seq profiling of liver tissue from P. chabaudi (Pc) blood-stage infected mice 20 days post-infection or age-matched uninfected (UI) controls

first: Elizabeth

middle: K

last: Glennon


title: Dual lineages of Langerhans cells cooperate to restore the immune barrier after skin injury (scRNA-seq)

city: East Lansing

first: Axel

middle: Dizan

last: Schmitter-Sanchez

state: MI

address: 775 Woodlot Dr

department: Medicine

institute: Michigan State University

summary: Langerhans cells (LCs) are key immune sentinels of the epidermis. How this network reorganizes to safeguard epidermal immunity after injury has remained unclear. Here, we uncover a previously unrecognized two-lineage program of LC repopulation during wound repair. Classically, tissue-resident embryonically derived LCs (eLCs) migrate to lymph nodes in response to antigens. In contrast, we find that injury triggers nearby eLCs to migrate into wounds, providing immediate coverage. In parallel, circulating monocytes infiltrate the skin and differentiate into long-lived monocyte-derived LCs (mLCs) that integrate stably into the network. We identify the chemokine receptor CXCR2 as a novel regulator of eLC migration into wounds, distinct from the CXCR4/CCR7 pathways mediating LC egress to lymph nodes. Pharmacological inhibition of CXCR2 impairs directional eLC migration and is accompanied by increased mLC infiltration, preserving immune barrier density. These findings reveal a coordinated and flexible two-lineage repair program that ensures robust restoration of epidermal immunity.

overall_design: To isolate both progenitor-derived and embryonic Langerhans cells, we collected and pooled the dorsal ear epidermis of six mice into a single sample. These mice were all male littermates. Ears were collected on Day 11 after wound induction. Samples were then enriched for LCs by magnetic sorting following the standard protocol from the Epidermal Langerhans Cell MicroBead Kit (Miltenyi Biotec).

middle: D

last: Schmitter-Sánchez

first: Sangbum

last: Park


title: Dual lineages of Langerhans cells cooperate to restore the immune barrier after skin injury (bulkRNA-seq)

overall_design: LCs and epithelial cells were collected from homeostasis mouse skin and sorted using fluorescence-activated cell sorting (FACS) based on their red (LCs) and blue (epithelial cells) fluorescence. Four samples representing a single mouse each.


title: Characterization of cancer-associated fibroblast populations that promote tertiary lymphoid structure formation in murine melanoma tumors

city: Charlottesville

first: Robert

middle: Weldon

last: Barnes

state: VA

address: 409 Lane Rd

department: Biomedical Engineering

institute: University of Virginia

summary: In recent years, the clinical relevance of tertiary lymphoid structures (TLS) in cancer has become increasingly clear. However, the mechanisms that promote TLS development have remained obscure, largely because of a lack of animal models in which cause and effect studies can be performed. In this study, we used a mouse model in which TLS develop spontaneously in intraperitoneal tumors to address this issue. Extending earlier studies in which we showed that CXCR5-expressing B cells are essential for TLS development, we found that CXCL13, the ligand for CXCR5, is expressed exclusively by a population of cancer-associated fibroblasts (CAF), and that these CAF are essential to promote B cell accumulation in these tumors. In keeping with earlier work showing that the presence of TLS was associated with smaller tumor size, we also found that the lack of B cell accumulation was associated with larger tumor size. Using scRNA-seq, we identified 7 subpopulations of CAF in TLS containing tumors. While many of these subpopulations appear similar to those de ned in other human and murine tumor models, we also discriminate new immunologically relevant subpopulations of CAF. One of these CAF subpopulations was substantially enriched for CXCL13-expressing cells. This population was also enriched for additional genes that could promote B cell recruitment and enable regulation by a variety of cytokines. However, other genes that we have shown or hypothesize to be important in TLS development were enriched for expression in other CAF subpopulations. Overall, this work indicates that TLS development in this model depends on multiple CAF populations operating at distinct points in the developmental process.

overall_design: Cxcl13-cre-tdTomato x  R26R-eGFPflox mice were implanted intraperitoneally with B16-OVA melanoma tumors. Cancer associated fibroblasts expressing tdTomato, eGFP, both, or neither were separately collected from these tumors by cell sorting.  Each collected population was  analyzed for transcript expression using scRNA-seq, clustered into groups using the Seurat package, and profiled using pathway analysis and differential gene analysis.

first: Kara

last: Cummings

first: Mirna

last: Perusina-Lafranca

first: Anthony

middle: B

last: Rodriguez

first: Katarzyna

last: Stasiak

first: Burkhard

last: Ludewig

first: Sepideh

last: Dolatshahi

first: Victor

last: Engelhard


title: Insulin Signaling Functions as a Topological Switch That Couples Aging and Heat Stress Responsiveness

city: Chicago

first: Manuel

middle: Jospeh

last: Widuch

state: IL

address: 1040 West Granville Avenue, 1021

department: Molecular Biology Sciences

institute: Northwestern University

summary: While aging and thermal stress are phenotypically intertwined, their transcriptomic signatures remain paradoxically distinct in Drosophila. Here, we demonstrate that this parallel modularity is not an inherent genomic constraint but is actively maintained by Insulin/IGF-1 signaling (IIS). Using transcriptomic analysis, we show that high IIS activity partitions the genome into orthogonal regulatory layers where aging and heat stress function as independent additive inputs. Systemic attenuation of IIS triggers a transition to an integrated architecture. In this state, the genome’s perception of heat stress becomes age-dependent, enabling a hyper-adaptive induction of heat shock proteins in old adults—effectively reversing the typical age-related decline in proteostasis. We propose that the IIS pathway functions as a topological switch, shifting the genome from a modular growth-mode to an integrated maintenance-mode. Thus, dampening the response to heat stress with age is an adaptable regulatory state rather than an inevitable consequence of cellular senescence.

overall_design: RNA-seq profiling of young and aged wild-type and insulin-producing-cell-ablated Drosophila following heat-shock stress. After heat shock, flies were classified as climbers or non-climbers using a negative geotaxis assay, and gene-expression responses were assessed across multiple recovery time points.

first: Manny

middle: J

first: Sanda

last: Siepka

first: Richard

last: Carthew


title: Plasmodium blood-stage induces trained immunity in hepatocytes

overall_design: RNA-seq profiling of whole livers from uninfected (UI) mice, day-20 P. chabaudi (Pc) blood-stage infected mice, P. yoelii (Py) liver-stage infected mice at 6, 24 and 44 hours post-infection (hpi), and sequentially infected mice (PcPy) at 6, 24, and 44 hours post-Py infection.


title: Cell culture effects on isolated fibrotic lung CD103+ DCs

city: Ann Arbor

first: Stephen

middle: J.

last: Gurczynski

address: 109 Zina Pitcher Pl, Room 4062

department: Internal Medicine

institute: University Of Michigan

summary: The trypophan metabolite kynurenine is a well known AHR ligand and drives inflammatory AHR respones, especially in dendritic cels.  Ex vivo studies of dendrtic cells are complicated by the fact that culture conditions do not reproduce complex tissue microenvironments. To study these effects we compared transcriptomes of CD103+ lung DCs isolated from mice with blm-induced fibrosis and compared with DCs that had been cultured in standard culture media for 18h.

overall_design: Groups of mice were treated with bleomycin for 7-days, after which lungs were harvested.  Single cell suspensions were produced via collagenase digestion and CD103+ DCs were isolated via magnetic bead. One group of DCs (pre-culture) was harvested directly, the second group (post-culture) was cultured ex-vivo in RPMI media supplemented with 10% FBS and 1% pen/strep. RNA was processed for RNAseq by Azenta following their standard RNAseq pipline.

first: Bethany

last: Moore


title: EKLF/KLF1 coordinates specialized transcriptional networks required to maintain the integrity of terminal erythropoiesis

city: New York

first: James

last: Bieker

state: NY

address: One Gustave Levy Place

department: Cell, Developmental, and Regenerative Biology

institute: Mount Sinai School of Medicine

summary: Erythroid Krüppel Like Factor (EKLF/ KLF1) is a C2H2 zinc finger transcription factor that plays a critical role in all aspects of erythropoiesis. Mutations in KLF1 lead to diverse phenotypes ranging from mild to severe anemias. Patients with a heterozygous E325K mutation (CDA type IV) exhibit impaired erythroid terminal differentiation and increased presence of binucleate erythroblasts. We previously showed that KLF1 is necessary for cell cycle exit and enucleation in mouse primary cells. In the current study we discovered that genes involved in cell motility, cell division, and mitotic pathways are all directly regulated by KLF1. Klf1-/- cells exhibit increased numbers of binucleated erythroblasts and DNA bridges, and differentiating Klf1-/- erythroblasts display an increased percentage of cytokinesis failure events and defective microtubule bundling. Klf1-/- erythroblasts produce frequent aberrant F-actin-rich membrane protrusions and anucleate cell fragments. Human CDA type IV cells exhibit similar patterns of dysregulation of cytokinesis and cell motility genes. Collectively, we show that KLF1 is necessary for maintaining the integrity of erythroid cell divisions by direct regulation of genes involved in cytokinesis and motility pathways during terminal erythroid differentiation.

overall_design: RNA seq on triplicate samples of erythroid cells obtained from Klf1+/+ and -/- E12.5 fetal liver ex vivo cultures, harvested during expansion and differentiation (day 2) stages.

first: Merlin

middle: N

last: Gnanapragasam


title: Genome instability induced by Cyclin E overexpression and ionizing radiation in WT and RECQL4-deficient U2OS cells

city: Kitakyushu

first: Masaoki

last: Kohzaki

state: Fukuoka

address: 1-1 Iseigaoka Yahatanishi-ku

department: Institute of Industrial Ecological Sciences

country: Japan

institute: University of Occupational and Environmental Health, Japan

summary:
RECQL4 is a RecQ helicase implicated in DNA replication and genome maintenance. To investigate the effects of RECQL4 deficiency on genome instability induced by replication stress, genome-wide copy number alterations were analyzed using the Affymetrix CytoScan HD Array in WT and RECQL4-deficient U2OS cells with or without Cyclin E overexpression and/or ionizing radiation.
Genome-wide copy number alterations were analyzed using the Affymetrix CytoScan HD Array. Raw CEL files are provided together with processed CYCHP files generated using Chromosome Analysis Suite (ChAS) version 4.5.

overall_design: Ten genomic DNA samples from WT and RECQL4-deficient U2OS cells were analyzed using the Affymetrix CytoScan HD Array. Two parental clones were included as baseline controls. Eight experimental samples consisted of cells cultured under non-induced (NE) or Cyclin E overexpression (OE) conditions, in the presence or absence of 3 Gy ionizing radiation. Genome-wide copy number alterations were analyzed using Chromosome Analysis Suite (ChAS).


title: Dissecting the molecular pathways through which long-chain PFAS disrupts dendritic cell maturation [RNAseq_mDCs_96h]

city: Wageningen

first: Benthe

last: van der Lugt

address: Akkermaalsbos 2

country: Netherlands

institute: Wageningen University & Research

summary: Per- and polyfluoroalkyl substances (PFAS) have been associated with impaired vaccine responsiveness and altered immune function, but the mechanisms underlying their immunotoxic effects remain incompletely understood. Because dendritic cells (DCs) play a central role in antigen presentation and initiation of adaptive immunity, this study investigated the effects of selected PFAS on DC maturation and the underlying molecular pathways. Human THP-1-derived DCs were exposed to four long chain PFAS followed by transcriptomic analysis, quantitative gene expression analysis, and pharmacological modulation of peroxisome proliferator-activated receptor alpha (PPARα)- and glucocorticoid receptor (GR)-associated pathways. Perfluorooctanoic acid (PFOA) induced the strongest transcriptional response, characterized by enrichment of pathways involved in fatty acid oxidation, peroxisomal metabolism, and PPAR signaling. Among the affected genes, pyruvate dehydrogenase kinase 4 (PDK4) emerged as a robust marker of PFAS-induced metabolic reprogramming. Functionally, PFAS exposure reduced expression of HLA-DR isotype, a key marker of DC maturation, with PFOA showing greater potency than the other compounds tested. Pharmacological inhibition experiments demonstrated that both PPARα-associated and GR-associated mechanisms contribute to the observed phenotype, although neither pathway alone fully accounted for the impairment. Overall, these findings indicate that PFAS disrupt DC maturation through coordinated immunometabolic reprogramming involving interconnected nuclear receptor signaling pathways, providing mechanistic support for the immunotoxic effects associated with PFAS exposure.

overall_design: THP-1 human monocytic cells were seeded at a density of 1×10^5 cells/mL and cultured for five days in cell culture medium (RPMI 1640 medium, 2 mM L-glutamine, 100 U/mL penicillin, 0.1 mg/mL streptomycin, 50 uM 2-mercaptoethanol, 10% heat-inactivated FBS) supplemented with rhIL-4 (1500 U/mL) and GM-CSF (1500 U/mL) to induce differentiation into immature dendritic cells. The effects of PFAS on DC maturation were investigated. On day five of differentiation, immature DCs were seeded at a density of 1×10⁶ cells/mL in 12-well plates containing complete treatment medium and exposed for 24 h to PFOA (10 µg/mL), PFOS (10 µg/mL) or vehicle control (0.1% DMSO, v/v). Following the initial 24-h exposure, a maturation cocktail consisting of rhIL-4 (3000 IU/mL), rhGM-CSF (1500 IU/mL), rhTNF-α (2000 IU/mL), and ionomycin (200 ng/mL) was added to induce maturation into mature dendritic cells (mDCs). PFAS and control compounds remained present throughout the maturation period. Cells were harvested after 24 or 72 h of maturation, corresponding to total exposure times of 48 and 96 h, respectively.

first: Martina

last: Iulini

first: Karsten

last: Beekmann

first: Ron

last: Hoogenboom

first: Valentina

last: Galbiati

first: Giulia

last: Russo

first: Francesco

last: Pappalardo

first: Styliani

last: Fragki

first: Alicia

last: Paini

first: Emanuela

last: Corsini

first: Aafke

last: Janssen


title: Dissecting the molecular pathways through which long-chain PFAS disrupts dendritic cell maturation [RNAseq_mDCs_48h]


title: Transcriptomic changes in human AC16 cardiomyocytes induced by electronic cigarette aerosol exposure

city: Beijing

first: 唐

last: 伟评

address: 2 Anzhen Rd, Chaoyang District,

institute: Beijing Institute of Heart Lung and Blood Vessel Diseases

type: Non-coding RNA profiling by high throughput sequencing

summary: Electronic cigarette (e-cig) use has raised increasing concerns regarding its cardiovascular effects, yet the underlying molecular mechanisms in cardiomyocytes remain unclear. In this study, human AC16 cardiomyocytes were exposed to e-cigarette aerosol using an air–liquid interface system, and transcriptomic profiling was performed to identify differentially expressed miRNAs and mRNAs. The analysis revealed marked alterations in calcium signaling–related genes, most notably suppression of miR-6504-3p and upregulation of its target gene PLN, providing mechanistic insight into e-cig–induced cardiomyocyte injury.

overall_design: Human AC16 cardiomyocytes were assigned to two groups: a filtered air control group and an e-cigarette aerosol exposure group. Cells were exposed using an air–liquid interface (ALI) system. Total RNA was isolated with TRIzol reagent (Invitrogen) according to the manufacturer's instructions, and RNA sequencing was performed for both mRNA and miRNA profiling, with five biological replicates per group (n = 5). Differentially expressed genes and miRNAs were identified between the e-cigarette and control groups.

first: Linyuan

last: Huang

first: Ruiyang

last: Ding

first: Xiaoke

last: Ren

first: Qinglin

last: Sun

first: Shiqian

last: Liu

first: Qing

last: Xu

first: Jinlin

last: Zheng

first: Yang

last: Li

first: Zhiwei


title: Small RNA sequencing of human sural nerves

city: Richardson

first: Diana

last: Tavares-Ferreira

address: 800 W Campbell Rd

department: Department of Neuroscience

institute: University of Texas at Dallas

summary: Small RNA (miRNA) sequencing of human sural nerves with (n=6) and without (n=6) diabetic peripheral neuropathy

overall_design:
We performed small RNA-sequencing to profile the miRNA landscape of human sural nerves from individuals with (n=6) and without DPN (n=6).

***************************************************************
Raw files for human/patient samples were not submitted to GEO due to concerns about submitting personally identifiable sequence data for open access.
***************************************************************

first: Sneha

middle: A

last: Gummadi

first: Ryan

last: Ju

first: Victoria

last: Pastor

first: Eric

middle: C

last: Meyers


title: Genome-wide DNA methylation motifs of Alicyclobacillus acidocaldarius subsp. acidocaldarius strain KCTC 1825 determined by PacBio HiFi sequencing

city: Seodaemun-gu

first: Jae-Yoon

last: Sung

state: Seoul

address: YERP B118DA, Yonsei University

department: Biotechnology

country: South Korea

institute: Yonsei University

type: Methylation profiling by high throughput sequencing

summary: PacBio HiFi sequencing was used to characterize genome-wide DNA base modifications and methylation motifs in Alicyclobacillus acidocaldarius subsp. acidocaldarius strain KCTC 1825 (DSM 446). Base-modification kinetics were mapped to the complete genome assembly and analyzed for m6A and m4C signals. The processed dataset provides per-base modification calls, motif-associated calls, and motif summaries linked to the public genome assembly and SRA run.

overall_design: Genomic DNA from a single culture of Alicyclobacillus acidocaldarius subsp. acidocaldarius strain KCTC 1825 (DSM 446) was sequenced on a PacBio Revio instrument. One HiFi sequencing library and one biological sample were analyzed. HiFi reads were assembled into a complete circular chromosome and plasmid, aligned to the assembly, and evaluated for m6A and m4C base modifications and enriched sequence motifs. No treatment or comparative condition was applied.


title: Characterization of transcriptomic profiles underlying gross morphological changes observed in semelparous pink salmon (Oncorhynchus gorbuscha)

city: PULLMAN

first: Michael

last: Phelps

state: WA

address: 1815 Ferdinand's Lane, VBRB 160

department: Animal Sciences

institute: Washington State University

summary: Pacific salmon undergo dramatic physiological remodeling during their terminal spawning migration, culminating in reproductive maturation, senescence, and death. To investigate the molecular mechanisms underlying these processes, we characterized transcriptomic changes in wild pink salmon (Oncorhynchus gorbuscha) sampled at three migratory stages (marine, riverine, and spawning ground) and across three tissues central to endocrine regulation and energy metabolism: gonads, head kidney, and skeletal muscle. RNA sequencing and dDifferential gene expression analyses waswere performed to identify biological pathways associated with maturation and semelparity. Transcriptomic changesremodeling wereas most pronounced during the final transition from river migrating fish entry to spawning, with the gonads exhibiting the greatest number of differentially expressed genes, followed by skeletal muscle and head kidney. Gene ontology analyses revealed coordinated upregulation of pathways associated with starvation responses, lipid mobilization, tissue catabolism, proteolysis, autophagy, and cell death across tissues. Immune and inflammatory processes were elevated in skeletal muscle and head kidney, while gonads showed strong enrichment of insulin signaling, steroid metabolism, and hormone-response pathways. Analysis of endocrine receptor expression identified tissue-specific and ohnolog-specific regulation of receptors involved in thyroid, growth hormone, insulin, gonadotropin, sex steroid, and glucocorticoid signaling, suggesting functional divergence among duplicated salmonid genes. These findings identified in pink salmon provide evidence that the demonstrate that the most extreme physiological changes occur extensive molecular reprogramming in pink salmon occurs duringat the time of  final maturation rather than freshwater entry. The coordinated activation of endocrine, metabolic, and degenerative pathways supports a mechanistic link between reproductive development and semelparous senescence, providing new insight into the endocrine  and transcriptomic networks that govern spawning-associated physiological decline in Pacific salmon.

overall_design: Transcriptomic study examining progressive physiologies of muscle/headkidney/gonads across both sexes in maturing pink salmon. Samples were taken from three distinct migratory stages (ocean, river, spawn) depicting sequenctially emerging phenotypes presenting during return migration.

first: Max

last: Butensky


title: Dissecting the molecular pathways through which long-chain PFAS disrupts dendritic cell maturation

first: Benthe

last: van der Lugt

overall_design: THP-1 human monocytic cells were seeded at a density of 1×10^5 cells/mL and cultured for five days in cell culture medium (RPMI 1640 medium, 2 mM L-glutamine, 100 U/mL penicillin, 0.1 mg/mL streptomycin, 50 uM 2-mercaptoethanol, 10% heat-inactivated FBS) supplemented with rhIL-4 (1500 U/mL) and GM-CSF (1500 U/mL) to induce differentiation into immature dendritic cells. Two independent RNA sequencing experiments were performed. THP-1-derived immature DCs were used to evaluate the early transcriptional response to PFAS exposure. On day five of differentiation, iDCs were seeded at a density of 1×10⁶ cells/mL in 12-well plates containing complete treatment medium, consisting of phenol red-free RPMI 1640 supplemented with 2 mM L-glutamine, 100 U/mL penicillin, 0.1 mg/mL streptomycin, 50 µM 2-mercaptoethanol, and 5% heat-inactivated delipidated FBS (dFBS). Cells were exposed for 24 h to 10 µg/mL PFOA or PFOS (corresponding to approximately 24.4 µM and 20.0 µM, respectively), DEX (150 µg/mL), the PPARα agonist GW7647 (1 µM), or vehicle control (0.1% DMSO v/v). Following 24 h exposure, cells were collected for transcriptomic analysis.

first: Martina

last: Iulini

first: Karsten

last: Beekmann

first: Ron

last: Hoogenboom

first: Valentina

last: Galbiati

first: Giulia

last: Russo

first: Francesco

last: Pappalardo

first: Styliani

last: Fragki

first: Alicia

last: Paini

first: Emanuela

last: Corsini

first: Aafke

last: Janssen


title: CALB2 is a mechanoresistance gene in metastatic prostate cancer

city: Houston

first: Abigail

last: Fabiano

state: TX

address: 6100 Main St

department: Bioengineering

institute: Rice University

summary: Metastatic progression hinges on the ability of circulating tumor cells (CTCs) to withstand the extreme mechanical forces of the bloodstream, yet the mechanisms that enable this survival remain poorly defined. Understanding how prostate cancer (PCa) cells adapt to fluid shear stress (FSS) is essential for identifying vulnerabilities that could be leveraged to prevent metastatic spread. Here, we generate “mechanoresistant” (MR) LNCaP and PC3 PCa cell lines by repeatedly exposing cells to high‑intensity (HI) FSS (3950 dyn/cm²), modeling the forces encountered by CTCs in circulation. LNCaP MR cells acquire marked resistance to HI FSS, exhibiting significantly reduced apoptosis compared to controls, while PC3 cells demonstrate a more innate resistance. Morphologically, PC3 MR cells adopt an amoeboid phenotype, whereas LNCaP MR cells retain parental characteristics. Both MR populations show significantly increased Piezo1 expression. Bulk RNA‑sequencing reveals that LNCaP and PC3 MR cells developed distinct molecular programs. Notably, CALB2 (calretinin) emerges as a mechanoresistance gene selectively upregulated in PC3 MR cells, and CALB2 knockout significantly reduces viability upon re‑exposure to HI FSS. Analysis of CALB2‑high PCa cases capture patient‑level molecular heterogeneity, with reproducible co‑expression correlated with EPB41L3 and DGKI, suggesting a mechanoadaptive state rather than a uniform aggressiveness-driven tumor phenotype. This is consistent with the absence of correlation between CALB2 levels and Gleason Scores. In an orthotopic PCa mouse model, the PC3 MR condition displays the most aggressive early tumor growth and the largest endpoint tumor volumes, consistent with enhanced proliferation observed in vitro. Together, these findings demonstrate that PCa cells acquire discrete mechanoadaptive phenotypes under extreme shear stress, revealing targetable pathways that may ultimately be exploited to limit metastatic competence.

overall_design: RNA was isolated from immortalized human prostate cancer cell lines (LNCaP and PC3) to identify transcriptional differences specific and unique to the mechanoresistant (MR) phenotype within each cell lines. Differential gene expression analysis was performed to identify genes and pathways unique to the MR condition.

first: Allen

last: Luo

first: Paul

last: Taufalele

first: Cynthia

last: Reinhart-King

last: King


title: Easy-Multiome enables joint profiling of gene expression and chromatin accessibility in single cells

city: ATHENS

first: Robert

last: Schmitz

state: GA

address: B416 Davison Life Sciences Department of Genetics University of Georgia 120 East Green Street

department: Genetics

institute: University of Georgia

summary: Gene expression and chromatin accessibility provide complementary insights into the regulatory mechanisms that define cell states. Although methods for jointly profiling these modalities exist, plant applications remain limted because of complex workflows, inconsistent performance, and prohibitive costs. Here, we present easy-Multiome, a streamlined single-cell multiomic workflow that integrates a single in situ reverse transcription step into the standard droplet-based scATAC-seq protocol. Using easy-Multiome, we profiled more than 20,000 nuclei from maize seedlings generating paired gene expression and chromatin accessibility data, with approximately 90% of nuclei containing both high-quality RNA chromatin accessibility profiles. The resulting transcriptome data resolved 16 clusters corresponding to nine major maize seedling cell types and enabled direct characterization of cell-type-specific chromatin accessibility from the same nuclei. Furthermore, easy-Multiome simultaneously captured cell-type-specific transcription factor expression and the accessibility of their cognate DNA-binding motifs, providing direct links between transcriptional programs and regulatory landscapes. Together, these results demonstrate that easy-Multiome enables robust and efficient joint profiling of plant gene expression and chromatin accessibility while requiring only minimal modifications to existing droplet-based scATAC-seq workflows.

overall_design: To evaluate easy-Multiome, maize seedling tissues were collected as follows. Seeds of the maize genotype **B104** were sown in Sun Gro Horticulture Professional Growing Mix (Sun Gro Horticulture Canada Ltd.). The soil was saturated with tap water, and seedlings were grown under a 50:50 mixture of 4100 K fluorescent lamps (Sylvania Supersaver Cool White Deluxe F34CWX/SS, 34 W) and 3000 K fluorescent lamps (GE Ecolux with Starcoat F40CX30ECO, 40 W). Plants were maintained under a 16-h light/8-h dark photoperiod at approximately 25°C during the light period with a relative humidity of approximately 54%. Six days after sowing, a 2-cm stem segment centered on the first visible node (1 cm on either side of the node) was harvested between 8:00 and 9:00 AM. Approximately three to five seedlings were pooled for each biological replicate.

first: Xuan

last: Zhang


title: ZFP36L2 orchestrates stress-adaptive plasticity in intestinal regeneration and colorectal cancer metastasis (PDO scRNAseq)

first: Qingwen

last: Jiang

address: 417 East 68th Street, 1725

institute: memorial sloan kettering cancer center

summary: Single-cell RNA sequencing (scRNA-seq) was performed on patient-derived primary colorectal cancer (CRC) organoids (OKG146P) to investigate the role of ZFP36L2 in regulating cell state plasticity during differentiation and dedifferentiation. Organoids expressing a doxycycline-inducible shRNA targeting ZFP36L2 (shZFP36L2) or a non-targeting control (shCtrl) were cultured with 2 μg/mL doxycycline and subjected to three conditions: (1) HISC medium for 7 days (baseline ISC state); (2) HISC for 7 days followed by intestinal growth factor-free (IGFF) medium for 7 days (differentiation); and (3) HISC for 7 days, IGFF for 7 days, then HISC for 7 days (dedifferentiation). Organoids were dissociated into single cells and multiplexed using TotalSeq hashtag antibodies (BioLegend). Viable (DAPI-negative) cells were FACS-sorted and pooled in equal numbers across conditions for library preparation using the Chromium Single Cell 3′ v3.1 platform (10x Genomics), targeting up to 10,000 cells per sample. Libraries were sequenced on an Illumina NovaSeq S4 (Read 1: 28 cycles; i7 index: 8 cycles; Read 2: 90 cycles). ZFP36L2 knockdown decreased the proportion of ISC-like cells across HISC and dedifferentiation conditions and impaired re-acquisition of the ISC state following dedifferentiation, with the greatest phenotypic divergence observed upon dedifferentiation.

overall_design: scRNA-seq experiment in OKG146P CRC organoids across 3 culture conditions (HISC, IGFF, Dediff) × 2 knockdown conditions (shCtrl, shZFP36L2), multiplexed using TotalSeq hashtag antibodies and sequenced on the 10x Genomics Chromium 3′ v3.1 platform.

first: Karuna

last: Ganesh

first: Jura

last: Pintar

first: Britney

last: Forsyth


title: Survival wisdom of Tradescantia spathacea Sw: Trade-off be-tween anthocyanin mediated photoprotection and photosyn-thetic capacity under high light stress

city: Chaozhou

first: Xiaoting

last: Zheng

state: Guangdong

address: Qiaodong

institute: Hanshan Normal University

summary: Light heterogeneity shapes plant survival, yet how shade-tolerant species balance photoprotection and photosynthesis under changing light remains unclear. Tradescantia spathacea Sw, with green upper and purple-red lower leaf surfaces, exhibits both shade tolerance and high-light acclimation, but the functional specialization of its leaf sides and the underlying trade-off mechanisms are poorly understood. This study exposed T. spathacea to 100% (FL) and 30% (LL) sunlight for 2 years and systematically assessed light adaptation mechanisms through morphological observation, physiological and biochemical detection, and transcriptomic analyses. Results showed that under FL conditions, leaf morphology underwent adaptive remodeling. Anthocyanins accumu-lated specifically in the lower side, thermal dissipation and antioxidant systems were fully activated, while photosynthetic capacity was markedly inhibited, reflecting a survival-priority strategy. Under LL conditions, leaves became broader and flatter, chlorophyll content increased, and the expression of photosynthesis-related genes was upregulated. This enhanced light capture and carbon assimilation efficiency, leading to significantly greater biomass accumulation, reflecting a growth-oriented resource al-location strategy. Transcriptomic and physiological data consistently showed that FL upregulated anthocyanin biosynthetic genes, while LL activated photosystem and photosynthetic electron transport genes. T. spathacea achieves a flexible photoadapta-tion strategy across different light environments through leaf surface functional spe-cialization and an anthocyanin mediated trade-off between photoprotection and pho-tosynthesis.

overall_design: RNA-seq profiling of Tradescantia spathacea Sw plants under two light intensity conditions (100% full sunlight, FL and 30% low sunlight, LL) after 2 years of light acclimation treatment.

first: Qingyi

last: Cai

first: Hui

last: Zhu

first: Yuzhong

first: Ziyuan

last: Li

first: Kaicheng

first: Zhengchao

last: Yu


title: ZFP36L2 orchestrates stress-adaptive plasticity in intestinal regeneration and colorectal cancer metastasis

summary:
Phenotypic plasticity is a hallmark of cancer, yet the molecular switches required for cell fate reprogramming are poorly understood. During intestinal wound healing and colorectal cancer (CRC) metastasis, differentiated cells can dynamically dedifferentiate into an intestinal stem cell (ISC) state to drive epithelial regeneration and metastatic outgrowth. Here we show that the RNA-binding protein ZFP36L2, which is mutated in 5-10% of CRC, is a critical stress-responsive orchestrator of dynamic dedifferentiation into an LGR5+ ISC state. In mouse colon regeneration models, ZFP36L2 ablation inhibits dedifferentiation, ISC gene expression and function, and impairs intestinal regeneration. In human CRC, loss of ZFP36L2 function abrogates metastatic seeding and the outgrowth of LGR5+ canonical metastases, while promoting lineage plasticity and non-canonical differentiation into heterogeneous cell states. Mechanistically, we find that ZFP36L2 binds to stress-associated mRNAs containing AU-rich 3' UTRs, inducing the formation of dynamic biomolecular condensates associated with mRNA degradation and termination of the stress response. Together, this work defines ZFP36L2 acts as a critical molecular switch coupling stress sensing with phenotypic plasticity, driving cellular dedifferentiation essential for re-establishing the ISC state during wound healing and metastasis. In ZFP36L2 deficient CRC, inability to re-enter the LGR5+ state during metastatic outgrowth drives non-canonical lineage plasticity, associated with poor clinical outcomes.

This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series

first: Cyrus

middle: L

last: Tam


title: Spatial transcriptomics supports a role for SOX-4 driven signaling throughout the disease course of biliary atresia

city: London

first: Tallulah

middle: Suzanne

last: Andrews

state: Ontario

address: 1151 Richmond St.

country: Canada

institute: University of Western Ontario

summary: Biliary atresia (BA) is a fibroinflammatory cholangiopathy of infancy that is the most common indication for pediatric liver transplantation. We aimed to define the molecular mechanisms responsible for differences in the rate of disease progression among children with BA. We performed spatial transcriptomics (ST) analysis on frozen liver tissue at transplant from 16 children: BA with survival with native liver (SNL) <2 years (BA1, n=3, BA1_3 was excluded from analysis), BA with SNL >2 years (BA2, n=4), non-BA cholestasis (n=4), and non-diseased donors (n=4). Transcriptional signatures were compared between patient groups by tissue region (scar, hepatocyte, cholangiocyte). Findings were validated in larger patient cohorts that included BA samples at diagnosis. ST analysis of BA1 patients showed the most aggressive disease phenotype, characterized by reduced hepatocyte zonation, low expression of homeostatic metabolic signatures, and increased scar heterogeneity enriched for pathways including extracellular matrix remodeling, interferon response, and leukocyte activation. Notably, genes involved in SOX4 hepatocyte-to-cholangiocyte reprogramming were most enriched in BA1 patients. Liver immunohistochemistry with in situ mRNA hybridization showed that BA patients at diagnosis had increased SOX4 quantification as compared to BA patients at transplant. Lastly, previously published liver bulk RNA-sequencing data demonstrated higher SOX4 gene-set expression in BA patients at diagnosis with SNL < 2 years. Children with BA and worse outcome exhibit increased SOX4 gene-set expression at diagnosis with greater loss of hepatocyte zonation and immune-driven scar heterogeneity at transplant. Further mechanistic studies are needed to determine whether SOX4-driven biliary reprogramming plays a maladaptive role in the reparative response to obstructive cholestasis in BA.

overall_design: Liver tissue was obtained from 16 patients, fresh-frozen and biobanked at -80. Tissue was obtained during liver transplant either from explanted diseased liver, or from health donor tissue (NL). Four sample (BA1) were obtained from pediatric transplant for patients with BA and recieved their transplant prior to age of 2 (mean age 0.7 years). Four samples (BA2) were obtained from pediatric transplant for patients with BA over the age of 2 (mean age 11.8 years). Four samples were obtained from non-BA cholestatic liver disease (mean age 4.9 years). Four samples from donor healthy livers (mean age 2.8 years). Tissue was sliced to 10um thickness and mounted in Visium slides. Fluorescent imagining was performed to visualize the CD68 distribution across the tissue slices using Abcam Alexa Fluor® 647 Anti-CD8 (Abcam Alexa Fluor® 647, rabbit, 1:10 dilution) and Anti-CD68 antibodies (Abcam Alexa Fluor® 488, mouse, 1:50 dilution) following the 10x immunofluorescence protocol. Tissue was permeablized and RNA was captured and sequenced following standard kits, and sequenced on an Illumina NovaSeq 6000.

middle: S

first: Sarah

last: Taylor


title: ZFP36L2 orchestrates stress-adaptive plasticity in intestinal regeneration and colorectal cancer metastasis (SLAMseq)

summary: Thiol-linked alkylation for the metabolic sequencing of RNA (SLAM-seq) was used to measure genome-wide mRNA half-lives in patient-derived colorectal cancer (CRC) liver metastasis organoids (MSK107Li). Organoids expressing a doxycycline-inducible short hairpin RNA targeting ZFP36L2 (shZFP36L2) or a non-targeting control (shCtrl) were metabolically labeled with 500 μM 4-thiouridine (4sU) for 6 hours, followed by a uridine chase. Total RNA was harvested in triplicate at t=0, 2, 5, and 12 hours post-chase, along with matched unlabeled control samples for background T>C correction. RNA was subjected to iodoacetamide alkylation and libraries were prepared using the QuantSeq 3′ mRNA-seq V2 Library Prep Kit FWD (Lexogen). T>C conversion rates were used to quantify labeled RNA fractions and fit single-exponential decay models to estimate per-gene mRNA half-lives. ZFP36L2 knockdown significantly stabilized transcripts associated with stress-response gene programs, consistent with ZFP36L2 promoting post-transcriptional decay of AU-rich stress-associated mRNAs.

overall_design: We performed 4sU pulse-chase labeling to measure mRNA decay kinetics in PDO MSK107Li cells expressing pTRIPZ control shRNA or ZFP36L2-targeting shRNAs. After doxycycline-induced shRNA expression, organoids were labeled with 500 μM 4sU for 6 hours, followed by replacement with uridine-containing medium to initiate RNA chase. Samples were collected at 0, 2, 5, and 12 hours after chase initiation to quantify RNA decay dynamics upon ZFP36L2 knockdown. Matched unlabeled control samples are used for background T>C correction.


title: Comparison of transcriptional programs between undifferentiated (0 hr) and differentiated N2A-A5 (96 hr) cells

city: Burlington

first: Princess

last: Rodriguez

state: Vermont

address: 89 Beaumont Avenue

department: Microbiology and Molecular Genetics

institute: University of Vermont

summary: Kinesins are molecular motor proteins essential for organizing and remodeling the cytoskeleton during neuronal development and maintenance. One key regulator is kinesin family binding protein (KIFBP), which inhibits a subset of kinesins by blocking motor-microtubule interactions. Homozygous nonsense mutations in KIFBP cause Goldberg-Shprintzen Syndrome (GOSHS), a neurodevelopmental disorder characterized by intellectual disability, microcephaly, and axonal neuropathy. Although loss of KIFBP has been linked to reduced neurite length and microtubule disorganization, the specific kinesins underlying these phenotypes remain unclear. This work uses a variety of approaches, including transcriptomics, to establish Neuro-2a cells as a model for studying KIFBP function and provide new insight into the regulation of kinesin activity and cytoskeletal dynamics in neurons.

overall_design: RNA-Seq of undifferentiated versus differentiated N2A-A5 cells. These cells were differentiated for 96 hours by seeding in differentiation culture media without blasticidin, supplemented with 1% FBS and 10 µM retinoic acid. We then performed gene expression profiling analysis using pair-end sequencing for three biological replicates per condition.

first: Sarah-Catherine

last: Paschall

first: Jason

last: Stumpff


title: ZFP36L2 orchestrates stress-adaptive plasticity in intestinal regeneration and colorectal cancer metastasis (HyperTRIBE)

first: Qingwen

summary: HyperTRIBE (Targets of RNA-binding proteins Identified By Editing) was used to identify genome-wide mRNA targets of ZFP36L2 in patient-derived colorectal cancer (CRC) and normal colon organoids. Full-length wild-type ZFP36L2 or the RNA-binding-incompetent frameshift mutant fsZFP36L2 (G144Afs*43) were fused to the hyperactive E488Q mutant catalytic domain of adenosine deaminase acting on RNA (ADAR) and expressed under a doxycycline-inducible Tet-on promoter, with a P2A-EGFP reporter to mark expressing cells. Constructs were stably integrated by lentiviral transduction into four organoid lines: two CRC liver metastasis lines (MSK107Li, OKG146Li), one matched primary tumour line (OKG146P), and one matched normal colon line (OKG146N). On day 5 post-seeding, organoids were treated with 2 μg/mL doxycycline for 2 days to induce transgene expression. GFP-positive live cells were isolated by flow cytometry, and total RNA was extracted using the RNeasy Mini Kit (Qiagen). mRNA libraries were prepared using the TruSeq Stranded mRNA Kit and sequenced on the Illumina NovaSeq 6000 (PE100). ZFP36L2-bound mRNAs were identified by comparing A-to-I editing events (read as A-to-G substitutions) enriched in ZFP36L2-ADAR versus fsZFP36L2-ADAR expressing organoids. Over 80% of ZFP36L2-specific edits occurred in mRNA 3′UTRs, predominantly at the conserved UAUUUA motif, consistent with known ZFP36 family RNA-binding properties.

overall_design: Patient-derived normal colon or CRC organoids expressing lentivirally integrated ZFP36L2-ADAR or fsZFP36L2-ADAR constructs were cultured in WRENAFI (OKG146N) or HISC (CRC PDOs) medium with puromycin selection for 5 days. Doxycycline was added at 2 μg/mL on day 5 to induce construct expression, and GFP-positive cells were sorted on day 7 for bulk RNA-seq to profile ZFP36L2 target mRNA.


title: ZFP36L2 orchestrates stress-adaptive plasticity in intestinal regeneration and colorectal cancer metastasis (bulk RNAseq)

summary: Bulk RNA sequencing was performed on MSK107Li patient-derived colorectal cancer (CRC) liver metastasis organoids to characterize transcriptional changes resulting from CRISPR-Cas9-mediated ZFP36L2 knockout during dedifferentiation. ZFP36L2 was disrupted using two independent sgRNAs; two validated knockout clones per sgRNA were pooled for sequencing, with control organoids serving as comparators. To assess dedifferentiation capacity, organoids were cultured in intestinal growth factor-free (IGFF) medium for 13 days to induce differentiation, then dissociated into single cells and passaged into HISC medium; samples were harvested on day 7 of dedifferentiation. Total RNA was extracted using the RNeasy Mini Kit (Qiagen) and submitted to Plasmidsaurus for 3′ end RNA-seq library preparation and sequencing. Libraries were generated using a 3′ end counting approach with unique molecular identifiers (UMIs) incorporated during cDNA synthesis. ZFP36L2 knockout organoids exhibited decreased ISC-associated gene signatures and increased expression of canonical and non-canonical differentiation programs, including osteoblast, squamous, and neuroendocrine lineages, relative to controls.

overall_design: Bulk 3′ end RNA-seq of MSK107Li ZFP36L2 CRISPR knockout (two sgRNAs, two clones each, pooled) versus control organoids harvested at day 7 of dedifferentiation. Libraries sequenced on the Illumina platform (~10M deduplicated reads per sample); reads aligned to the hg38 reference genome.

first: Karuna


title: Systematic transcriptomic comparison of the time-serial interferon responses in two widely used human cell lines

city: 江苏省 / 扬州市 / 邗江区

first: JIANG

last: JIAYAO

address: 江苏省扬州市邗江区扬州大学文汇路校区

department: 扬州大学

institute: 扬州大学

summary: Here, we systematically compared the transcriptomic dynamics of IFN‑γ stimulation in HEK293T and HeLa cells across a time series (0–24 h) in duplicate. Our analysis uncovered both shared and cell‑type‑specific transcriptional responses, providing insights into the temporal regulation of interferon signaling in distinct human epithelial backgrounds.

overall_design: HEK293T and HeLa cells were cultured in high‑glucose DMEM supplemented with 10% FBS and 1% penicillin‑streptomycin at 37 °C in 5% CO₂. Cells were passaged using 0.25% trypsin‑EDTA, neutralized with serum‑containing medium upon detachment. For time‑course stimulation, cells were treated with IFN‑γ and harvested at indicated time points: HEK293T at 0 h, 30 min, 1 h, 3 h, 6 h, 12 h, and 24 h; HeLa at 0 h, 15 min, 30 min, 1 h, 3 h, 6 h, 12 h, and 24 h. All stimulations were performed in duplicate.

first: Jiayao

first: Liangliang

first: Shuai

last: Chen

first: Mingan

last: Sun


title: Resmetirom ameliorates fibrogenesis in hepatic stellate cells via thyroid hormone receptor alpha-fatty-acid amide hydrolase 1 signaling pathway

city: Santiago de Compostela

first: Rubén

last: Nogueiras Pozo

state: A Coruña

address: Campus Vida Avenida Barcelona, s/n, CIMUS

department: Fisiology

country: Spain

institute: CIMUS - USC

summary: Resmetirom is a liver-directed, thyroid hormone receptor β (THRβ)-selective agonist approved for treating metabolic-associated steatohepatitis (MASH). While Resmetirom hepatocyte-specific effects are well-established, its impact on other hepatic cells, particularly hepatic stellate cells (HSCs), the main fibrogenic cells, remain unknown.  Using animal models, immortalized cell lines, and primary murine and human HSCs, we combined pharmacological treatments with genetic manipulation of THRα and fatty acid amide hydrolase (FAAH) to examine the effects of Resmetirom. Resmetirom ameliorates MASH through actions in both hepatocytes and HSCs. The overexpression of solute carrier organic anion transporter family member 1B1(SLCO1β1) in both cell types increases the efficiency of Resmetirom. THRα expression is higher in HSCs than in hepatocytes in both human and murine cells. Resmetirom attenuates TGF-β1-induced HSC activation via THRα and increased FAAH expression and activity, while their inhibition prevents Resmetirom from reducing fibrotic marker expression and the elevated glycolytic activity typical of activated HSCs.  These results uncover an unrecognized mechanism of action for Resmetirom, demonstrating that its antifibrotic efficacy extends beyond hepatocytes to include direct effects on HSCs via THRα and FAAH. These results may support the development of future therapeutic strategies aimed at THRα in HSC, in combination with existing approaches targeting hepatocyte THRβ.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for human primary hepatic stellate cells (PhHSCs). Transcriptomic analysis of 8 samples in total, comprising 4 biological replicates of TGF-β1 36h 8ng/mL -treated PhHSCs and 4 biological replicates of TGF-β1 36h 8ng/mL + Resmetirom 300uM 24h co-treated PhHSCs.

first: Eva

last: Novoa

first: Alba

last: Cabaleiro

first: Borja

last: López-Picallo

first: Natalia

last: da Silva Lima

first: Cristina

last: Riobello

first: Carmen

last: Pena

first: Jose

last: Iglesias-Moure

first: Ana

last: Senra

first: Maria Luz

last: Martínez-Chantar

first: Anabel

last: Fernandez-Iglesias

first: Jordi

last: Gracia-Sancho

first: Susana

last: Bravo

first: Carlos

last: Dieguez

first: Marta

last: Varela-Rey

first: Vincent

last: Prevot

first: Markus

last: Shcwaninger

first: Miguel

last: López

last: Nogueiras


title: Fibroblast-derived Spondin-2 promotes dysplastic transition in metaplastic gastric epithelial cells

city: NASHVILLE

first: James

middle: R

last: Goldenring

state: TN

address: 2213 Garland Avenue

department: Epithelial Biology Center

institute: Vanderbilt University

summary: Background and Aims: Fibroblasts are increasingly recognized as active drivers of early gastric carcinogenesis, yet the specific fibroblast-derived factors that promote the transition from metaplasia to dysplasia remain unknown. We have recently found that coculture of metaplasia or cancer-derived fibroblasts with gastroids dominated by spasmolytic polypeptide-expressing metaplasia (SPEM) lineages leads to a decrease of metaplastic markers and a concomitant upregulation of dysplasia markers. We have now identified the matricellular protein Spondin-2 (SPON2) as a key fibroblast-secreted mediator of this transition. Methods: Single-cells RNAsequencing of patient-derived fibroblast populations and proteomic profiling of conditioned media were used to identify fibroblast-secreted factors. Spatial transcriptomics was used to evaluate localization of SPON2+ fibroblasts in human gastric tissue. Knockdown of SPON2 in cancer-derived fibroblasts was performed in coculture with SPEM lineage gastroids, and recombinant SPON2 treatment was applied to metaplastic gastroids. Results: Single-cell RNAsequencing and proteomic profiling revealed SPON2 as the most upregulated and selectively secreted protein in metaplasia- and cancer-derived fibroblasts compared to normal fibroblasts. Spatial transcriptomics demonstrated PDGFRA/SPON2 co-expressing fibroblasts localized adjacent to metaplastic glands in human gastric tissue. Knockdown of SPON2 in cancer-derived fibroblasts abrogated induction of dysplastic promotion in cocultured SPEM lineage gastroids, while preserving metaplastic markers. Conversely, recombinant SPON2 treatment promoted down-regulation of metaplastic markers and upregulation of dysplasia-associated proteins in metaplastic gastroids. Conclusions: These findings identify fibroblast-derived SPON2 as a potent secreted factor driving the metaplasia-to-dysplasia transition in gastric epithelium. Therefore, SPON2 represents the first fibroblast-derived secreted factor that can promote preneoplastic progression.

overall_design: Human IntestiCult Medium (StemCell Technologies) containing dissociated gastroids was plated onto the upper surface of collagen-coated Transwell inserts. Concurrently, IntestiCult Medium was added to the lower chamber beneath the insert. Media in both compartments were replaced after 48 hours. Gastroid cells were allowed to proliferate and adhere for a total of 7 days atop the collagen-coated membrane. On day 7, the apical medium was aspirated to expose the epithelial surface to air, initiating the air-liquid interface (ALI). At the same time, the basal compartment was replenished with either fresh IntestiCult Medium (negative control) or with IntestiCult medium consisting of or 20 ng/mL of recombinant human SPON2. ALI culture was maintained for an additional 14 days. During this period, the medium in the lower wells was changed every other day to ensure nutrient availability, while any secretions accumulating on the apical surface of the Transwell were removed daily.At the conclusion of the 14-day ALI period, samples were collected in DPBS, centrifuged, and stored in RNALater. Bulk RNA sequencing was performed to identify transcriptomic changes in gastroid-derived epithelial cells in response to SPON2 treatment compared to negative control conditions.

first: Jared

last: Rhodes

first: Sumin

last: Hur

first: Changqing

last: Zhang

first: Isaac

last: Tamayo-Sarver

first: Eric

last: Sha

first: Elizabeth

last: Manning

first: Jean René

last: Clemenceau

first: Bogun

last: Jang

first: Eunyoung

last: Choi

first: Tae Hyung

last: Hwang

first: Su-Hyung

last: Lee


title: Latexin deletion in the bone marrow niche suppresses leukemia via LIFR signaling downregulation and immune activation

city: Rye

first: Liming

last: H ou

address: 601 Midland Ave

institute: New York Blood Center

summary: Acute myeloid leukemia (AML) is the most common adult leukemia diagnosis. Bone marrow (BM) niche significantly influences the initiation and progression of AML. However, our knowledge about the impact of leukemic niche on leukemia stem cells (LSC) and leukemogenesis is still limited. In this study, we identified a novel function of latexin (Lxn) in the cell-extrinsic regulation of leukemogenesis. Using a MLL-AF9-induced AML mouse model in wild-type (WT) and Lxn knockout (Lxn-/-) recipient mice, we found that Lxn deletion in the BM niche enhances the survival of AML mice by suppressing LSCs and reducing blood blasts. Single-cell RNA sequencing of stromal cells and cell communication analysis uncovered downregulation of the leukemia inhibitory factor receptor (LIFR) signaling pathway in the Lxn-/- niche, particularly within mesenchymal stromal cells (MSCs). Mechanistically, reduced LIFR level in Lxn-/- MSCs upregulates Cxcl9 expression, leading to increased recruitment of CD8 T cells and enhanced cytotoxicity against leukemic cells. Combination of Lxn niche deletion and immune checkpoint inhibitor PD-L1 further prolonged survival. The findings have important clinical implications, suggesting that Lxn inhibition could improve the efficacy of AML therapies by targeting the leukemia niche and enhancing immune surveillance.

overall_design: The single-cell RNA (sc-RNA) sequencing were performed on both stromal and hematopoietic cells from WT and Lxn-/- normal and GFP+ MLL-AF9-induced AML mice. Stromal cells were sorted as CD45-, Lin-, Ter119-, CD71- and 7-AAD- population, while hematopoietic cells were Lin- in the normal condition, and GFP+ Lin- in AML condition.

first: Cuiping

first: Xia

last: Yao

first: Pinpin

last: Sui

last: Hou

first: Bowen

last: Yan

first: Gang

last: Huan

first: Sheng

last: Tong

first: Youwen

first: Hui

last: Zhong

first: Bojing

last: Shao

first: Hong

last: Qian

last: Zheng

first: Fengchun

last: Yang

first: Ying

last: Liang


title: Translational remodeling across serial xenotransplantation in acute myeloid leukemia patient-derived xenografts

city: paris

first: Jerome

last: Tamburini

address: 22 rue mechain

country: France

institute: Cochin Institute

summary: We performed ribosome profiling (Ribo-seq) on acute myeloid leukemia patient-derived xenograft models across serial transplantation passages to characterize translational remodeling associated with leukemia propagation.

overall_design: Ribo-seq was performed on FACS-purified human AML blasts (hCD45+ hCD33+) isolated from patient-derived xenograft models across serial transplantation passages (P1–P3). Each passage represents an independent biological sample reflecting leukemia propagation in vivo. A total of 8 samples were analyzed.

first: Clement

last: Larrue

first: Paolo

last: Angelino

first: Petros

last: Tsantoulis


title: DNA methylation remodeling across serial xenotransplantation in acute myeloid leukemia patient-derived xenografts

summary: We performed Illumina Infinium MethylationEPIC array profiling of acute myeloid leukemia patient-derived xenograft models across serial transplantation passages P0, P1, and P2 to characterize DNA methylation changes associated with leukemia propagation.

overall_design: Genome-wide DNA methylation profiling was performed using the Illumina Infinium MethylationEPIC BeadChip on 27 AML patient-derived xenograft samples collected across serial transplantation passages P0, P1, and P2. Raw IDAT files were processed in R using the minfi package. Data were normalized with preprocessNoob, and beta values were extracted for downstream analyses.


title: Single-cell RNA-seq profiling of AML patient-derived xenografts across serial transplantation

summary: Single-cell RNA sequencing was performed on AML PDX samples across serial transplantation (P0, P1, P2) to investigate transcriptional evolution and leukemic stem cell dynamics.

overall_design: scRNA-seq (10x Genomics) was performed on multiple AML PDX samples at different transplantation stages (P0, P1, P2). Each sample corresponds to a distinct biological condition.


title: Single-cell RNA-seq analysis of benign prostatic hyperplasia prostate specimens stratified by prostate weight

city: Guangzhou

first: Ning

last: Jiang

address: 253 Middle Industrial Road

institute: Southern Medical University

summary: Six de-identified benign prostatic hyperplasia prostate specimens were profiled by 10x Genomics single-cell RNA-seq. Samples were stratified into Large BPH and Small BPH groups by prostate weight. Raw FASTQ files and sequencing-provider CellRanger filtered feature-barcode matrices are provided; downstream quality control, integration, annotation, and endothelial-focused analyses were performed by the authors.

overall_design: Discovery cohort of six BPH prostate specimens: three Large BPH samples and three Small BPH samples. Each sample has paired-end FASTQ files and per-sample CellRanger filtered feature-barcode matrix files.

first: Jiang

last: Ning

first: Gao

last: Wei


title: Evolution of leukemic stem cell states and transcriptional programs across serial xenotransplantation in acute myeloid leukemia

summary: We performed bulk RNA-seq profiling of acute myeloid leukemia (AML) patient-derived xenograft models across serial transplantation passages (P0, P1, P2) to characterize transcriptional changes associated with leukemia propagation and stem cell activity.

overall_design: Bulk RNA-seq was performed on AML patient-derived xenograft models across serial transplantation passages P0, P1, and P2. A total of 27 samples were analyzed, representing 9 independent models. Gene-level counts and TPM matrices were generated using Salmon and used for downstream analyses.


title: Single-cell gene programs define subtype identity and metastatic trajectories in renal cell carcinoma

city: Montréal

first: Hamed

last: Najafabadi

state: Québec

address: 740 Dr. Penfield Avenue

department: Human Genetics

institute: McGill University

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


title: The regulation of COX-2, ABCA1 and ABCG1 by the lncRNA PACERR links the inflammatory response and cholesterol homeostasis

city: Philadelphia

first: Nicholas

last: Lahens

state: PA

address: Smilow Center for Translational Research 10-110 3400 Civic Center Blvd, Bldg 421

department: ITMAT

institute: University of Pennsylvania

summary: The COX-2 antisense lncRNA PACERR interacts with discrete RNA binding proteins to mediate a cell specific COX-2 response to HDL. PACERR exerts its action both in cis and in trans, controlling the expression of COX-2 in macrophages, while also regulating the levels of macrophage ABCA1 and ABCG1, cholesterol transport genes involved in maintaining cholesterol homeostasis. When cells are exposed to HDL, PACERR is transcribed, leading to the release of bound histone acetyltransferase p300 and its relocation to the promoter of COX-2 which is then transcribed, leading to PG formation. In concert, PACERR also releases heterogeneous nuclear ribonucleoprotein L (hnRNPL) which relocates to the ABCA1 and ABCG1 gene loci, controlling mRNA alternative splicing and subsequent cholesterol efflux to HDL. A strong hnRNPL binding site motif is found in PACERR RNA and when this was deleted, the interaction between PACERR RNA and hnRNPL is abrogated both at baseline and following HDL treatment of macrophages. THP-1 macrophages stably overexpressing PACERR (P1-3) or deleted of the hnRNPL binding site (Delta1-3) from the PACERR sequence. In delta cells, PACERR no longer binds to hnRNPL but is free to bind to ABCA1 and ABCG1 impacting their transcription. In summary, we found that PACERR controls the levels of the cholesterol transporters through its interaction with the splicing factor, hnRNPL. It is released from PACERR RNA in response to HDL where it relocates to introns of cholesterol transporters determining the alternative splicing fate of transcripts required for cholesterol efflux. Through use of the delta cells, we have Illustrated the specificity for the interaction between hnRNPL and PACERR.

overall_design: RNA-seq profiling of THP-1 differentiated macrophages stably expressing the lncRNA PACERR and PACERR with the hnRNPL bidning motif deleted and treated with HDL

first: Elizabeth

last: Hennessy

first: Takae

last: Tanosaki

first: Ronan

last: Lordan

first: Soon Yew

last: Tang

first: Vladimir

middle: V

last: Shuvaev

first: Soumita

last: Ghosh

first: Ujjalkumar

last: Das

first: Robin

last: Joshi

first: Hu

last: Meng

first: John

last: Snouwaert

first: Hanna

last: Winter

first: Lars

last: Maegdefessel

first: Beverly

middle: H

last: Koller

first: Garret

last: FitzGerald


title: Single-cell resolution spatial transcriptomics of a mucinous tubular and spindle cell carcinoma using CosMX

summary: This dataset investigates the spatial organization of a human mucinous tubular and spindle cell carcinoma using single-cell resolution spatial transcriptomics. The dataset comprises 22 fields of view from a primary tumor specimen profiled using the CosMX Human 1K panel. Gene program activity scores were estimated using gene programs previously identified from single-cell RNA sequencing data. Local malignant cell density, defined as the fraction of neighboring cells classified as malignant, was quantified and analyzed in relation to gene program activity to assess spatial associations within the tumor microenvironment.

overall_design: Single-cell–resolution spatial transcriptomic profiling of a human mucinous tubular and spindle cell carcinoma was performed using the CosMX Human 1K panel.

first: Ariel

last: Madrigal

first: Minjun

last: Kim

first: Zohreh

last: Mehrjoo

first: Tamiko

last: Nishimura

first: Yasser

last: Riazalhosseini


title: Effects of CRISPR-mediated knockout of Mitf, Tfe3 and Tfeb on the transcriptome of mouse BMDMs from WT and Grn KO animals [hex001130]

city: South San Francisco

first: Thomas

last: Sandmann

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

state: California

address: 161 Oyster Point Blvd

institute: Denali Therapeutics

summary: Previous single-cell RNA sequencing of microglia from aged WT and Grn KO mice revealed a microglial subset with transcriptomic signatures that were highly specific to Grn KO. Further epigenetic analysis demonstrated that these signatures were likely regulated by Mitf/Tfe family transcription factors. To investigate the necessity of these transcription factors in mediating the Grn KO transcriptomic response, we turned to an experimental in vitro validation. A multiplexed CRISPR approach was used to knockout Mitf, Tfe3 and Tfeb in BMDMs from WT or Grn KO mice. 10 days post-CRISPR, bulk RNA sequencing was performed to analyze the transcriptional profiles. These profiles identified the expected enrichment of Grn KO-specific genes in BMDMs from Grn KO mouse background. Strikingly, the levels of many of these genes were reversed with knockout of Mitf, Tfe3 and Tfeb.

overall_design: This experiment profiles mouse bone-marrow-derived macrophages (BMDMs) harvested from 4 experimental groups, each containing 4 animals (16 animals in total): 1) WT, Rosa26 (control), 2) WT, CRISPR KO of Mitf/Tfe3/Tfeb, 3) Grn KO, Rosa26 (control) 4) Grn KO, CRISPR KO of Mitf/Tfe3/Tfeb (TKO). Samples were collected in four batches, each containing one animal from each experimental group.

first: Catherine

last: Heffner

first: Connie

last: Ha

first: Gilbert

last: Di Paolo


title: Transcription profiles of mouse microglia isolated by FACS from WT and Grn KO animals, separated by lipofuscin levels [hex001110]

summary: Previous single-cell RNA sequencing of microglia from aged WT and Grn KO mice revealed a microglial subset with transcriptomic signatures that were highly specific to Grn KO. Immunohistochemistry analysis suggested that this population was also characterized by high lipofuscin burden. In this study, we used bulk RNAseq to investigate whether these transcriptomic markers were affected by lipofuscinosis. Microglia were sorted by FACS from dissociated brain tissue of 19-month-old mice of WT or Grn KO background. Cells expressing microglial markers (CD45+/CD11b+) were selected and stratified into lipofuscin ‘low’ or ‘high’ subpopulations, based on autofluorescence. 1,000 cells were collected per sample and were processed and analyzed for bulk RNA sequencing. Results indicated a consistent upregulation of Grn KO-specific genes in microglia from the Grn KO background, which were further enriched ‘high’ vs. ‘low’ lipofuscin cells.

overall_design: In this study, microglia were isolated from the brains of 12 mice, either WT (3 male, 3 female animals) or Grn knockout (KO) (3 male, 3 female) animals using FACS. Microglia from each animal were split into two bins (high / low) according to their lipofuscin content, leading the the following four experimental groups of samples: 1) WT, lipofuscin low, 2) WT, lipofuscin high, 3) Grn KO, lipofuscin low, 1) Grn KO, lipofuscin high,


title: Active histone modifications restrict off-target 6mA deposition and maintain transcriptional stability in Rhizopus microsporus [RNA-seq]

city: Murcia

first: Carlos

last: Lax

address: Facultad de Biología

department: Genetics and Microbiology

institute: University of Murcia

summary: Epigenetic mechanisms provide sophisticated regulatory layers that modulate gene expression across diverse organisms, yet their organization and crosstalk remain poorly understood in non-dikarya fungi. Here, we characterize the genome-wide landscape of chromatin organization in Rhizopus microsporus, revealing a compartmentalized architecture where active histone modifications (H3K4me1, H3K4me3, H3K27ac) define transcriptionally active euchromatin distinct from H3K9me3-marked constitutive heterochromatin. Through comprehensive ChIP-seq analysis, we demonstrate that these modifications exhibit distinct distribution patterns over gene bodies and co-localize with 6-methyladenine (6mA) clusters, the predominant DNA modification in this fungus. We identified functional specialization among Set1 and Gcn5 paralogs, where Set1a primarily deposits H3K4me3, Set1b regulates H3K4me1, and both Gcn5 variants function redundantly in H3K27ac deposition. Knockout analysis reveals that these enzymes are critical for sporulation, stress resistance, and pathogenicity. Importantly, we uncover a hierarchical crosstalk where histone modifications restrict off-target 6mA deposition, regulate methylation cluster stability, and buffer transcriptional variation. Our findings establish R. microsporus as a model for understanding epigenetic compartmentalization in non-dikarya fungi and reveal conserved principles of epigenetic crosstalk that may be fundamental to eukaryotic chromatin regulation.

overall_design: RNA-seq of Rhizopus microsporus WT, set1 mutant, and gcn5 mutant. ChIP-seq: 18 libraries (WT/set1 mutant/gcn5 mutant × H3K4me1, H3K4me3, H3K27ac, RNApol II, beads, input). RNA-seq: 12 libraries (WT × 3 replicates; set1 mutant allele a × 3; set1 mutant allele db × 3; gcn5 mutant × 3). Differential expression by DESeq2. ChIP-seq peaks by MACS2.

first: Macario

last: Osorio

first: Natalia

last: Nicolás

first: Ghizlane

last: Tahiri

first: Stephen

last: Mondo

first: Vivian

last: Ng

first: Eusebio

last: Navarro

first: Igor

last: Grigoriev

first: Francisco

first: Victoriano

last: Garre


title: Active histone modifications restrict off-target 6mA deposition and maintain transcriptional stability in Rhizopus microsporus [ChIP-seq]

overall_design: ChIP-seq of Rhizopus microsporus WT, set1 mutant, and gcn5 mutant. ChIP-seq: 18 libraries (WT/set1 mutant/gcn5 mutant × H3K4me1, H3K4me3, H3K27ac, RNApol II, beads, input). RNA-seq: 12 libraries (WT × 3 replicates; set1 mutant allele a × 3; set1 mutant allele db × 3; gcn5 mutant × 3). Differential expression by DESeq2. ChIP-seq peaks by MACS2.


title: Transcriptomic and functional analysis reveals the role of photoperiod and Cryptochrome1 in regulating immunity of sea urchins (Strongylocentrotus intermedius)

city: Yan Tai

first: Qiao

middle: Zi

last: Zhao

address: China Agr Univ, Yantai Ins

institute: CAU

summary: Photoperiod is a key environmental factor regulating circadian rhythm and physiology in aquatic organisms, but its effects on the immune function of the sea urchin Strongylocentrotus intermedius (S. intermedius) remain unclear. This study investigated photoperiodic regulation of immune resistance, transcriptomes, and circadian rhythm in S. intermedius, and the role of circadian clock gene Cryptochrome1 (Cry1) in immunity. The sea urchins were acclimated to four photoperiods (LL, DD, LD, DL) for one month, followed by Pseudoalteromonas sp. challenge. Bacterial load and gene expression were detected by qRT-PCR; transcriptomic analysis identified DEGs between LD-ZT0 and LD-ZT12, and Cry1 was knocked down via dsRNA for verification. LD photoperiod significantly enhanced antibacterial resistance, with the lowest bacterial load and highest expression of immune genes (SOD, CAT, TNFSF14, C3, ACP, NLRP3) at 12/24 hpi. Transcriptome analysis identified 1450 DEGs enriched in immune (innate immunity, NLRP3 inflammasome assembly) and circadian pathways. LD maintained robust 24-hour rhythms of circadian (Cry1, Cycle, Clock, Rev-erbα) and immune genes, while DL induced phase inversion and LL/DD abolished rhythms. Cry1 knockdown reduced bacterial load and upregulated NLRP3, C3, IL-17D, and TNFSF14. Collectively, LD is optimal for S. intermedius circadian rhythm and immune homeostasis, and Cry1 negatively regulates its immune response. This study provides insights into photoperiodic regulation of echinoderm immunity and support for S. intermedius aquaculture. Collectively, these findings indicate that LD photoperiod is optimal for maintaining circadian rhythm and immune homeostasis of S. intermedius, and Cry1 plays a negative regulatory role in its immune response. This study provides novel insights into the molecular mechanisms of photoperiod regulating immune function in echinoderms and theoretical support for aquaculture management of S. intermedius.

overall_design: RNA‑seq analysis of coelomocytes from Strongylocentrotus intermedius at ZT0 and ZT12 under LD conditions


title: In vivo metatranscriptomics reveals sRNA-driven zinc-sparing responses in chronic Staphylococcus aureus infection

city: Atlanta

first: Paul

last: Briaud

address: 311 Fesrt Dr

institute: Georgia Institute of Technology

summary: Understanding bacterial gene regulation during human infection remains challenging, as most regulatory networks are inferred from in vitro models that poorly recapitulate host environments. Here, we present INTENSE (In-human Transcriptional Expression Networks in Site-specific Environments), a pangenome-based metatranscriptomic pipeline designed to profile Staphylococcus aureus transcription directly from human infection samples. Using a curated S. aureus pangenome and stringent decoy-based filtering, we analyzed RNA-seq data from chronic wound and cystic fibrosis sputum samples and uncovered infection-specific transcriptional signatures that clearly diverge from in vitro growth conditions. Small RNAs (sRNAs) emerged as major contributors to transcriptomic variability during chronic infection, with nine sRNAs consistently induced in human samples. Among these, we identify rsaX20 as a previously uncharacterized, zinc-responsive sRNA that also encodes a small peptide. Meta-analysis of over 2,000 publicly available S. aureus RNA-seq datasets revealed that rsaX20 is recurrently induced under low-zinc and host-associated stress conditions and displays co-regulation patterns with known metal-responsive sRNAs. Genetic, transcriptomic, and proteomic analyses demonstrate that rsaX20 is repressed by the zinc uptake regulator Zur, induced upon zinc limitation, and functions as a dual-purpose sRNA/sORF. Functional dissection revealed that the sRNA and peptide exert distinct and often opposing regulatory effects on protein expression, collectively targeting a network of metal-binding proteins. Notably, rsaX20-mediated repression of zinc-dependent enzymes supports a zinc-sparing adaptive response during infection. Together, these findings establish pangenome-based metatranscriptomics as a powerful approach to interrogate bacterial regulation in vivo and uncover rsaX20 as a key dual-function regulator shaping zinc homeostasis during chronic S. aureus infection

overall_design: RNA-seq profiling of Human Chronic Wound.This includes both human and microbial transcriptomes

first: Julia

last: Schumacher

first: Marvin

last: Whiteley


title: RNA-seq analysis of pulmonary fibrosis induced by alveolar epithelial mitochondrial ND1 deficiency in rats

city: beijing

first: Zhang

last: Ryan

address: panjiayuan chaoyang china

institute: Chinese Academy of Medical Sciences  AND Peking Union Medical College

summary: Mitochondrial dysfunction in alveolar epithelial cells has been implicated in the pathogenesis of idiopathic pulmonary fibrosis (IPF), yet direct in vivo evidence remains limited. In this study, we generated a novel rat model with lung epithelium-specific knockout of the mitochondrial ND1 gene by crossing Sftpc-Cre rats with LSL-DdCBE-ND1 rats. The resulting ND1Sftpc-cre rats spontaneously developed pulmonary fibrosis with restrictive ventilatory defects, metabolic abnormalities, and characteristic histopathological changes including alveolar septal thickening, inflammatory infiltration, and collagen deposition. To investigate the molecular mechanisms underlying this phenotype, we performed bulk RNA sequencing on lung tissues from ND1Sftpc-crerats and ND1LSL littermate controls. Differential expression analysis identified genes significantly altered in ND1-deficient lungs. Gene Ontology and KEGG pathway enrichment analyses revealed significant upregulation of extracellular matrix (ECM) organization, p53 signaling pathway, and cell cycle-related processes. This dataset provides transcriptomic insights into the molecular changes associated with epithelial mitochondrial dysfunction-induced pulmonary fibrosis and identifies potential therapeutic targets for interstitial lung diseases.

overall_design: This study aimed to characterize the transcriptomic landscape of pulmonary fibrosis induced by alveolar epithelial mitochondrial dysfunction. We generated ND1Sftpc-cre rats by crossing Sftpc-Cre driver rats with LSL-DdCBE-ND1 rats, resulting in lung epithelium-specific knockout of the mitochondrial ND1 gene. Age-matched ND1LSL littermates served as controls. Lung tissues were harvested from 12 week old female rats (ND1Sftpc-cre: n = 3; ND1LSL control: n = 3 ). Total RNA was extracted using TRIzol reagent. RNA quality was assessed using Agilent 2100 Bioanalyzer. Libraries were prepared using VAHTS Universal V10 RNA-seq Library Prep Kit and sequenced on Illumina Novaseq 6000 platform with 150 bp paired-end reads. Differential expression analysis was performed using DESeq2. Genes with adjusted p-value < 0.05 and |log2 fold change| > 1 were considered significantly differentially expressed. Functional enrichment analysis was conducted using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases.

first: Lei

last: Zhang

first: Gefan

last: Wan

first: Xiaolong

last: Qi

first: Rui

last: Fan

first: Chenyang

last: Liu

first: Qi

last: Kong

first: Yuanwu

last: Ma


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

city: Memphis

first: Wojciech

last: Rosikiewicz

address: 262 Danny Thomas Place

department: Center for Applied Bioinformatics

institute: St. Jude Childrens Research Hospital

summary: Children with Down syndrome have a 150-fold increased risk of developing the myeloid leukemia of Down syndrome (ML-DS). ML-DS is preceded by transient abnormal myelopoiesis (TAM), which spontaneously resolves in most cases but progresses to AML with additional mutations most commonly in the cohesin complex or signaling genes. However, the mechanisms by which these alterations promote leukemia are unknown. Here, we leveraged isogenic cell lines and patient data to investigate the role of cohesin mutations in leukemia progression. Multi-omic analyses revealed that haploinsufficiency of the cohesin complex suppresses the occupancy of CIITA at HLA class II loci, leading to a reduction in HLA Class II gene expression. Surface levels of HLA-DR are lower in ML-DS relative to TAM and these decreased levels are associated with an increased risk of TAM progression. Of note, a decrease in HLA-Class II gene expression is also a feature of non-DS AML with reduced expression of cohesin family members. These data suggest that unlike TAM, ML-DS blasts may evade the immune system by reduced HLA class II expression, providing a new model for leukemia progression in DS.

overall_design: Genome-edited AML cell lines were generated and compared to control parental lines to investigate the impact of genetic perturbations on chromatin organization and gene regulation.

first: Austin

last: Boucher

first: Yichao

last: Li

first: Yong

last: Cheng

first: Beisi

last: Xu

first: John

last: Crispino


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

first: Lahiri

last: Konada


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

first: Te

last: Ling


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


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

first: Shilpa

last: Narina

first: Shondra

middle: M

last: Pruett-Miller


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

middle: D


title: Effects of CDC5L knockdown on gene transcription levels in HeLa cell lines

city: Shanghai

first: Shunyi

state: Shanghai

address: 100 Haike Rd, Building 13

department: Interdisciplinary Research Center on Biology and Chemistry, Shanghai Institute of Organic Chemistry

institute: Chinese Academy of Sciences

summary: Stress granules (SGs) are membraneless condensates that assemble in response to cellular stress. We identified cell division cycle 5-like protein (CDC5L) as a regulator of SG dynamics, where CDC5L knockdown triggered SG formation even in the absence of exogenous stress. Reduced CDC5L levels led to upregulation of PERK at both mRNA and protein levels, suggesting transcriptional regulation. To examine the global impact of CDC5L on gene expression, particularly stress-related pathways, we performed RNA-seq in HeLa cells following siRNA-mediated CDC5L knockdown. In parallel, ChIP-PCR demonstrated that CDC5L directly binds to the PERK promoter, indicating a role as a transcriptional repressor. To further characterize CDC5L-bound genomic regions, we established a stable HeLa cell line expressing CDC5L-HA via lentiviral transduction and performed ChIP-seq to identify promoter regions directly occupied by CDC5L. Together, these transcriptomic and epigenomic datasets define CDC5L as a transcriptional regulator of stress-responsive genes.

overall_design: To elucidate the role of CDC5L in transcriptional repression and cellular stress response, CDC5L was knocked down in HeLa cells using siRNA, followed by RNA-seq to assess transcriptomic changes. In parallel, ChIP-seq was performed to profile CDC5L genomic binding, allowing integration of occupancy and expression data to identify direct transcriptional targets.

first: Beituo

last: Qian

first: Feng

last: Qiu

first: Wanjin

first: Yanshan

last: Fang


title: Posphorylation-Dependent Activation of Transcription Factors Drives Cell Fate Switching in Cortical Development

city: Odense M

first: Vijay

last: Tiwari

state: Denmark

address: Campusvej 55

department: Institut for Molekylær Medicin (IMM)

country: Denmark

institute: Queen’s University Belfast

summary: Cortical development involves precise regulation of transcription factors (TFs) and signalling pathways. Here, we show that phosphorylation of TFs is a key mechanism driving cell fate transitions during neurogenesis and gliogenesis. Using multi-omics profiling of embryonic mouse cortex, we identify stage-specific phosphorylation of Hmgn3 and Nfib as critical for neuronal differentiation and maturation. Phospho-mutant analyses reveal impaired migration and astrocyte differentiation. CUT&RUN and scATAC-seq integration show phosphorylation-dependent enhancer binding by Nfib. Additionally, Hmgn3 and Nfib regulate distinct signaling pathways mediating cell-cell communication. These findings highlight phosphorylation as a central regulator of TF activity and cortical lineage specification.

overall_design: We disected cortex from E14.5 and E18.5 developing mouse brains and performed RNA isolation using Trizol. Purified RNA samples were quantified using Qubit high-sensitivity RNA Quantification assay (Thermo Fisher Scientific, Q32852). The quality of the RNA was analyzed using Agilent TapeStation systems with high-sensitivity RNA tapes with RINs. Following extraction, RNA samples were normalized to 1 ng/μL, and 1 μL of RNA was input to prepare RNA libraries using a bulk input optimized FLASH-Seq protocol.

first: Anuj

last: Dwivedi


title: Interplay of ELF5, GATA-3 and FLI-1 Regulate Malignant Transformation of Keratinocytes

city: Nottingham

first: Mohammed

last: Ahmed

address: Clifton Lane

country: United Kingdom

institute: Nottingham Trent University

summary: Cutaneous squamous cell carcinoma (cSCC) arises from epidermal keratinocytes and is the second most common epithelial malignancy. E74-like factor 5 (ELF5) is a member of the epithelium specific ETS subfamily of transcription factors and is essential for epithelial development, homeostasis and prevention of epithelial tumourigenesis. Malignant transformation of keratinocytes involves the disrupts the progenitor differentiation program causing keratinocytes to fail to complete the differentiation process, leading to dysplastic epithelium and precancerous cells in skin. However, the precise mechanisms that causes the failure of keratinocytes to differentiate completely is not well understood. Our study brings new insights into novel role of ELF5 as tumour suppressor in keratinocytes. We found that ELF5 is abundantly expressed in healthy human skin (both basal and suprabasal layers), while its expression is lost in cSCC tissue biopsies. Using CRISPR-Cas9 knockout technology, functional studies along with transcript and protein analysis, loss of ELF5 in immortalised healthy keratinocytes (HaCaT) leads to decreased ability of cells to differentiation and increased migration, invasion and tumourigenicity, in vitro. While overexpression of ELF5 leads to inhibition of cell migration and proliferation in keratinocytes and in skin cancer cell lines (A431 and SCC-9), in vitro. Further analysis using RNA transcriptomics has uncovered that GATA-3 and FLI-1 are key molecular targets of ELF5 in keratinocytes. ELF5 regulation of both GATA-3 and FLI-1 is required to maintain the balance between healthy keratinocytes (proliferation/differentiation) and to prevent precancerous cell formation (tumourigenicity). Our data indicates that ELF5 plays pivotal role in preventing malignant transformation of keratinocytes and potentially suppresses skin tumourigenesis, which could be targeted for future drug treatments for cSCC. Better understanding of the underlying mechanisms by which ELF5 regulates gene expression and consequently cell activity will provide new knowledge that can be translated into long-term benefits for patients with precancerous legions to cSCC.

overall_design: RNA seq analysis of CRISPR-Cas9 mediated Knockout of ELF5 in Immortalized Human Keratinocytes (HaCaT) compared to  CRISPR-Cas9 Control RNA samples

first: Hetal

last: Patel

first: Maximilian

middle: E

last: Pickup

first: Maryam

last: Lawal

first: Chandrakala

last: Khadka

middle: I


title: The effects of the hepatocyte targeted RSPO mimetic, SZN-043, on liver cells

first: Russell

last: Fletcher

state: CA

address: 171 Oyster Point Blvd, Suite 400

department: Discovery Biology

institute: Surrozen, Inc.

summary: We assessed the effects on liver cells of repeated dosing the hepatocyte targeted RSPO mimetic, SZN-043, on liver cells by applying scRNA-seq to dissociated liver cells at 24-hours after one dose, 24-hours after 14 consecutive daily doses, and upon a 2-month washout.

overall_design: Mice were dosed IP with a control antibody (Anti-GFP) or a hepatocyte targeted RSPO mimetic. Two mice from each treatment for each timepoint were assessed at 24-hours after one dose, 24-hours after 14 daily doses, or after a 2-month washout period, and the livers were perfused with a collagenase solution to dissociate the liver cells, and the dissociated cells were purified by FACS, and 10x Genomics 3' v3 scRNA-seq was applied to them.

middle: B

first: Jacqueline

last: Phipps

first: Darshini

last: Shah

first: Trevor

last: Fisher

first: Mehaben

first: Hélène

last: Baribault

first: Yang

last: Li

first: Wen-Chen

last: Yeh

first: Chenggang

last: Lu