title: Revealing the transcriptional profile of fish muscle cells in vitro and in vivo: insights to improve fish muscle culture

city: Jatai

first: Bruna

middle: Tereza Thomazini

last: Zanella

state: Goias

institute: Federal University of Jatai (UFJ)

summary: In vitro experiments have contributed to numerous fields of knowledge, including fish skeletal muscle. Despite improved strategies, in vitro assays still show discrepancies with in vivo systems, especially for non-model organisms. In this sense, we characterized the transcriptional profile of pacus (Piaractus mesopotamicus) muscle cells in vitro and in vivo. Processes related to proliferation, glycolytic metabolism, and extracellular matrix were enriched in vitro, while energy production, muscle contraction, and amino acid pro-cessing were enriched in vivo. Through qPCR, the genes fn1a (fibronectin 1a), hk1 (hexokinase 1) and ctnnb1 (catenin beta 1), respectively related to extracellular matrix, glycolytic metabo-lism, and cell proliferation and differentiation, were highly expressed in vitro. The genes ckma (creatine kinase, muscle a), acat1 (acetyl-CoA acetyltransferase 1), mdh2 (malate dehydrogenase 2) and pkmb (pyruvate kinase M1/2b), respectively associated with ATP production, fatty acid oxidation, oxidative and glycolytic metabolism, and the genes musk (muscle, skeletal, receptor tyrosine kinase), chrna1 (cholinergic receptor, nicotinic, alpha 1) and clu (clusterin), in-volved in cell signaling, were highly expressed in vivo. Our findings may support the de-velopment of molecular strategies to enhance muscle cell culture technology and expand knowledge for in vitro meat production, improving sustainable fish farming.

overall_design: Juvenile pacu (Piaractus mesopotamicus) (12-20g) were farmed at 28°C in storage tanks equipped with a water circulation system under 12h light:12h dark photoperiod. The animals were fed ad libitum once a day with commercial food. The fish were euthanized with an overdose of benzocaine, and the fast-twitch muscles were collected from the epaxial region for in vitro (muscle cells: MC) and in vivo (muscle tissue: MT) analysis. For the in vitro experiments, we collected the skeletal muscle of twelve animals for each cell isolation process, and for in vivo experiments, the skeletal muscle of six animals.

first: Érika

last: Perez

first: Mirely

last: Barbosa

first: Jéssica

last: Valente

first: Bruno

last: da Silva Duran

first: Maeli

last: Dal-Pai-Silva


title: Gene differences between chemoresistant and chemosensitive variants of the human bladder cancer cell line T24 [RNA-Seq UC3]

status: Public on Mar 11 2026

city: Chongqing

first: Weiyang

last: He

state: chongqing

address: No.1 Youyi Road, Yuzhong District

country: China

institute: The First Affiliated Hospital of Chongqing Medical University

type: Expression profiling by high throughput sequencing

summary: The chemoresistant cell line was established using a concentration-gradient increment method. Transcriptome sequencing was performed after stable resistance was achieved to identify differentially expressed genes.

overall_design: The chemoresistant cell line was developed by repeatedly exposing the chemosensitive human bladder cancer cell line T24 to increasing concentrations of the chemotherapeutic drug. This cyclic process of treatment, followed by outgrowth of surviving resistant cells, was continued until a stably resistant cell population was established.

first: Yan

last: Sun

first: Tinghao

last: LI

first: Junlong

last: Zhu

first: Xiaoyu

last: Zhang

first: Linfeng

last: Wu

first: Kunyao

first: Aimin

last: Wang


title: Impact of SliP4 deletion on the high-light acclimation in Synechocystis sp. PCC 6803

city: Freiburg

first: Wolfgang

middle: R.

last: Hess

state: Baden-Würtemberg

address: Schänzlestrasse 1

department: Institute of Biology III

country: Germany

institute: University of Freiburg

summary:
SliP4 is a small, 37 amino acids protein that is strongly induced when the cyanobacterium Synechocystis sp. PCC 6803 is exposed to high-light conditions. Deletion mutants of the sliP4 gene manifest a light-sensitive phenotype due to impaired cyclic electron flow and state transitions. In this study, we aimed to investigate the consequences of SliP4 deficiency on the process of high-light acclimation.
The sliP4 deletion mutant demonstrated a complete gene regulatory response 30 minutes after the light intensity was increased from 50 to 250 μmol photons m-2 s-1, a process that was mediated by the RpaB-PsrR1 system. However, after the shift to high light, the mutant increased the production of extracellular polysaccharides and the cells began to aggregate, thereby effectively reducing the potential impact of light stress effects caused by the impaired capacity for cyclic electron flow and state transitions. This effect included the upregulation of xssA-E and xssN-P genes for the production of the sulfated exopolysaccharide synechan. Our results demonstrate that the unicellular cyanobacterium Synechocystis attempts to compensate for the crucial role of SliP4 by activating a population-level response to cope with light stress conditions, thereby revealing several genes involved in this response.

overall_design: Total RNA (5 µg) was extracted from the WT and ∆sliP4 mutant grown at 50 μmol photons m-2 s-1 and after a shift to 250 μmol photons m-2 s-1 for 30 min and 48 h, respectively. Total RNA was DNase-treated using TURBO DNase (Thermo Fisher Scientific). 2 µg DNase-treated RNA were labeled with Cy3 using the ULS Fluorescent Labeling Kit for Agilent arrays (Kreatech). A high-resolution microarray manufactured by Agilent (Design ID 075764, format 8 × 60 K, slide Layout = IS-62976-8-V2) was used. 600 ng labeled RNA were used for hybridization for 17 h at 65 °C, per array.

first: Luna

last: Alvarenga-Lucius

first: Sandra

last: Maaß

first: Viktoria

last: Reimann

first: Dörte

last: Becher

middle: R

first: Martin

last: Hagemann


title: AURKA-Mediated 7-Dehydrocholesterol Biosynthesis Drives Ferroptosis Evasion and Chemoresistance

city: Guangzhou

first: Jialing

last: Gao

state: Guangdong

address: Zhongshan No.2 Road 1th, Nonglin Street

institute: Sun Yat-Sen University

summary: Chemoresistance remains a major obstacle to effective colorectal cancer (CRC) treatment. While ferroptosis induction offers promise for chemosensitization, its clinical utility is limited by intrinsic resistance mechanisms. Here, we identify Aurora kinase A (AURKA) as a central ferroptosis suppressor that sustains chemoresistance by rewiring cholesterol metabolism through the SAPS3–AMPK–SREBP2 axis. Mechanistically, AURKA phosphorylates SAPS3 at Ser523/524, promoting its degradation and relieving AMPK suppression. Activated AMPK subsequently inhibits SREBP2 nuclear translocation and DHCR7 transcription, resulting in the accumulation of 7-dehydrocholesterol (7-DHC), a lipid antioxidant that confers ferroptosis resistance. Both genetic and pharmacologic inhibition of AURKA restore ferroptosis sensitivity and enhance chemotherapy efficacy in vitro and in patient-derived xenograft models. Clinically, elevated AURKA expression correlates with poor prognosis and reduced chemotherapy response in CRC patients. These findings define a novel AURKA-driven chemoresistance mechanism, revealing a regulatory nexus between cholesterol homeostasis and ferroptosis evasion, and position AURKA as a promising therapeutic target for chemosensitization in CRC.

overall_design: Colorectal cancer cell SW480 stably expressing shAURKA or control shRNA were harvested for RNA preparation. Total RNA was extracted using TRIzol reagent and sequenced on the MGISEQ2000 Platform (BGI, China). There were two experimental groups (shNC, shAURKA) with 3 replicates for each.


title: Systematic Perturbation of Cultures of K. Phaffii Show Gene Signatures Associated with 1 Specific Productivity of Recombinant Proteins

city: Cambridge

first: Charles

middle: Arthur

last: Whittaker

state: MA

country: USA

institute: Koch Institute

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

overall_design: Refer to individual Series


title: Cdkn2a/p16INK4A Loss affects hippocampal transcription in Young Adult Mice Independently of Alzheimer ’s-Associated Genetic Pathways

city: ILLKIRCH

first: Yann

last: HERAULT

address: 1 rue Laurent Fries

country: France

institute: IGBMC/PHENOMIN-ICS

summary: Background – The cyclin-dependent kinase inhibitor CDKN2A/p16INK4A is a central regulator of cellular senescence, widely studied in aging and cancer. Although its expression increases with age and disease, its role in the adult brain remains poorly defined. Recent evidence suggests a possible contribution to neuroinflammation and cognitive decline in Alzheimer’s disease (AD), but underlying mechanisms are unclear. We investigated the functional impact of p16INK4A inactivation on hippocampal gene expression in adult mice under normal and amyloidogenic conditions using the AppNL-G-F AD model.  Methods –. Hippocampal transcriptomes from control, p16INK4Aknockout (p16INK4AKO), AppNL-G-F, and double mutant mice were analysed by RNA sequencing. We performed differential gene expression analysis, gene set enrichment analysis (GSEA), and interaction modelling to assess the molecular consequences of p16INK4A loss alone and in combination with amyloid pathology.  Results –Transcriptomic analysis revealed that p16INK4AKO upregulated metabolic, mitochondrial, and translational pathways, while downregulating synaptic and cytoskeletal genes. In contrast, AppNL-G-F mice displayed strong immune activation. The double mutants showed additive transcriptional changes, yet GSEA indicated non-linear interactions in synaptic and immune-related pathways. Unexpectedly, p16INK4A deletion alone enriched senescence-associated signatures, despite the loss of this canonical senescence gene.  Conclusion – Constitutive p16INK4A loss causes broad transcriptomic remodelling in the adult hippocampus, independently of amyloid pathology. These findings uncover non-canonical roles for p16INK4A in neuronal homeostasis and emphasise the need for temporally controlled models to better dissect the brain-specific functions of senescence regulators in ageing and neurodegeneration.

overall_design: Mice were euthanized via cervical dislocation at ~5 months of age. For transcriptomic analysis, both hippocampi were collected from a subset of randomly selected animals and stored at −80°C. Tissue homogenization was performed using the Precellys 24 Touch (Bertin Instruments, Montigny-le-Bretonneux, France) at 4500 rpm for 20 seconds. RNA extraction was carried out using the RNeasy Lipid Tissue Mini Kit (74804, Qiagen), following the manufacturer’s instructions. RNA concentration was measured using the Qubit RNA Assay Kit (Thermo Fisher Scientific), and RNA integrity was assessed using the Agilent Bioanalyzer 2100 (Agilent Technologies, Les Ulis, France). Library preparation was performed using the Illumina Stranded mRNA Prep Ligation kit (Reference Guide – PN 1000000124518). RNA-Seq libraries were generated from 350 ng of total RNA following the manufacturer’s instructions (Illumina Stranded mRNA Prep, Ligation kit and IDT for Illumina RNA UD Indexes, Illumina, San Diego, USA). After polyA selection, mRNA was fragmented at 94 °C for 8 minutes. DNA libraries were then amplified using 12 PCR cycles, and excess primers were removed by two rounds of purification using SPRIselect beads (Beckman-Coulter, Villepinte, France). Library quality and concentration were assessed using the Bioanalyzer 2100 system (Agilent Technologies, Les Ulis, France). Sequencing was performed on an Illumina NextSeq 2000 platform using single-end 50 base reads. Image analysis and base calling were conducted using RTA (v2.7.7) and BCL Convert (v3.8.4). Raw sequencing reads were preprocessed to remove adapter sequences, polyA tails, and low-quality reads (defined as those with a Phred score < 20). Reads shorter than 40 bases after trimming were excluded from downstream analysis. Preprocessing was performed using the R package cutadapt (v4.2) (26). Reads were first aligned to ribosomal RNA (rRNA) reference sequences from RefSeq (27) and GenBank (28) using bowtie (v2.3.5) (29), and rRNA-matching reads were discarded. Remaining reads were then aligned to the Mus musculus GRCm38 genome assembly using STAR (v2.7.10b) (30). Gene body coverage was assessed across all samples using the geneBodyCoverage function from RSeQC (v5.0.1) (31), and transcript integrity number (TIN) values were calculated for transcripts with at least 10 uniquely aligned reads. In cases of overlapping genomic annotations, priority was assigned as follows: exonic > intronic > antisense genic > intergenic regions. Strand specificity was also evaluated using RSeQC (v5.0.1) (31). Gene expression levels were quantified using uniquely aligned reads with STAR (v2.7.10b) (30), based on Ensembl annotation version 102. Only reads unambiguously assigned to a gene were retained for downstream analysis. To reduce noise from lowly expressed genes, those with fewer than 15 reads in more than 10% of the samples were excluded, yielding a final set of 18,768 genes. Outlier samples were identified using Principal Component Analysis (PCA) on variance-stabilized expression data (VST)-normalized using DESeq2 (v1.45.3) (32). The number of PCs retained was determined using the Elbow Method, automated with the FindPC package (v1.0) (33), which identifies the point of maximal deviation from a straight line between the first and last PC. Mahalanobis distances were then calculated based on the selected PCs to detect statistical outliers. Squared Mahalanobis distances were compared to the 95th percentile of the Chi-squared distribution; samples exceeding this threshold were classified as outliers.

last: Herault

first: Pauline

last: Stephan

first: Damien

last: Plassard


title: Systematic Perturbation of Cultures of K. Phaffii Show Gene Signatures Associated with 1 Specific Productivity of Recombinant Proteins [K. phaffii S-953 RNA-seq]

summary: Demand for recombinant proteins is rapidly growing, driven by their use as 16 biotherapeutics, vaccine components, industrial enzymes, and food ingredients. The growing market requires novel strategies for increasing specific productivity (qp) of cellular hosts. Systems-level frameworks have been used to improve productivity, but have had difficulty relating complex cellular pathways with protein expression. Here, we demonstrate a method for mapping relationships between gene expression signatures and complex phenotypes related to recombinant protein productivity. Our approach induces systematic perturbations in cultures of K. Phaffii using varied co-feeds of carbon sources. The different carbon sources significantly impacted cell growth, specific productivity, and transcriptional states. With these data, we identified scale-independent metagenes that explained significant transcriptomic variance and strongly associated with complex cellular phenotypes, including qp and response to methanol induction for both IgG1 and VHH. We used these results to identify and knockout 31 novel gene targets whose expression inversely correlated with productivity. Ten of these genes improved productivity of IgG1 by up to 3x, and 17 genes increased productivity of VHH by up to 1.7x without impacting cell growth. This study identified ten groups of carbon co-feeds which induce similar transcriptomic states. We systematically perturbated cultures of K. phaffii using these co-feeds and identified phenotype-associated metagenes. Using the productivity-correlated metagene, we identified 10-17 genes which improved recombinant protein productivity when knocked out. This framework for relating gene signatures to complex cellular phenotypes could provide useful metrics for assessing 37 production processes and identifying new targets for cellular engineering.

overall_design: Gene expression data for K. phaffii strain (S-953) expressing 3B2 Variable domain of the Heavy Chain (VHH) nanobody grown in glycerol-containing complex media, then exchanged into complex media containing eight different carbon coifeeds, three different co-feed concentrations, and three different methanol concentrations (8x3x3=72 conditions) with 2-3 replicates per condition.

first: Raghav

last: Acharya

first: Joshua

last: Hinckley

first: Rachel

last: Barry

first: Eugenie

last: Cha

first: Harini

last: Narayanan

first: Brittney

middle: C

last: Sunday

first: Hayley

last: Ford

middle: A

first: John

last: Love


title: Dysmetabolic-dependent α-ketoglutarate/succinate ratio imbalance impairs TDG function and base excision repair process increasing susceptibility to pancreatic cancer

city: Rome

first: Valerio

last: Licursi

address: via degli Apuli, 4

department: c/o Dept. Biology and Biotechnologies "C. Darwin", Sapienza University of Rome

country: Italy

institute: Institute of Molecular Biology and Pathology (IBPM), National Research Council (CNR)

summary: Pancreatic ductal adenocarcinoma (PDAC) is a high lethal cancer with poor prognosis and lack of effective therapeutic options. Chronic metabolic disorders, including obesity and type 2 diabetes, are established risk factors for PDAC and contribute to its unfavorable prognosis. Prolonged exposure to altered metabolic states induces aberrant fluctuations in specific metabolites, which can impact the epigenetic landscape and facilitate the acquisition of neoplastic lesions. Indeed, certain metabolites function as cofactors for epigenetic enzymes, modulating their activity. In this study, we explored the interplay between metabolism and epigenetics in PDAC tumorigenesis under dysmetabolic conditions, aiming to uncover novel vulnerabilities in pancreatic cancer. The link between dysmetabolism and PDAC was examined in LSL-KrasG12D; PDX-1-Cre mice (KC mice) exposed to high-fat diet (HFD). Untargeted metabolomic analysis of pancreatic tissue revealed altered free fatty acid levels and elevated S-adenosyl methionine (SAM) during tumor development in HFD-fed mice. Targeted metabolomics further showed increased succinate (SA) levels alongside reduced α-ketoglutarate (αKG) levels, indicating dysregulation of the DNA methylation pathway. Given these findings, the DNA demethylation complex of ten-to-eleven-translocation methylcytosine 1 (TET1) and thymine DNA glycosylase (TDG) was further investigated in human pancreatic ductal epithelial cells bearing KRAS mutation exposed to dysmetabolic mimicking culture conditions. We demonstrated that dysmetabolic stress disrupts quickly TET1/TDG function, leading to accumulation of iterative cytosine modifications, including 5-formylcytosine (5fC), and increased apurinic/apyrimidinic (AP) sites. Specifically, AP site accumulation was a consequence of TDG hyperactivation induced by direct binding of SA on its arginine residue 275. This TDG alteration, together with the methylation-prone metabolic environment, impairs the base excision repair (BER) pathway by hypermethylation of the promoters of DNA ligases 1 (LIG1) and 3 (LIG3) and thus inducing their downregulation. The resulting inhibition of single-strand break repair predisposes to genomic instability and promotes pancreatic preneoplastic lesion development. Our results reveal an unprecedented role of metabolic-epigenetic crosstalk in dysmetabolism-driven PDAC tumorigenesis. These findings highlight TDG as a novel therapeutic target and propose 5fC and AP sites as potential biomarkers for pancreatic cancer.

overall_design: RNASeq from PDX-1-Cre LSL-KrasG12D (KC) mouse model fed either with low fat diet (LFD) or high fat diet (HFD)

first: Chiara

last: Cencioni

first: Silvia

last: Malatesta

first: Francesco

last: Spallotta


title: KIF5A downregulation in spinal muscular atrophy links axonal regeneration defects with ALS

city: Stanford

first: Yi

last: Zeng

address: 300 Pasteur Drive

institute: Stanford University

summary: Spinal muscular atrophy (SMA) is a devastating neuromuscular disorder caused by mutations in the Survival Motor Neuron 1 (SMN1) gene, leading to decreased SMN levels and motor neuron dysfunction. SMN-restoring therapies offer clinical benefit, but the downstream molecular consequences of SMN reduction remain incompletely understood. Here, we demonstrate that SMN deficiency results in downregulation of KIF5A in human neurons and in a mouse model of SMA. We provide evidence that reduced SMN levels impair axon regeneration, which is rescued by KIF5A overexpression and that the RNA-binding protein SMN functions to stabilize KIF5A mRNA. These findings provide evidence of a molecular link between SMA and ALS pathophysiology, highlighting KIF5A as a new SMN target. Our findings suggest SMN-independent interventions targeting KIF5A could represent a complementary therapeutic approach for SMA and other motor neuron diseases.

overall_design: RNA sequencing profiing of human stem cell derived corticial neurons and motor neurons treated with SMN shRNA

first: Tetsuya

last: Akiyama

first: Aaron

middle: D

last: Gitler


title: Microglial NUS1 deficiency drives dopaminergic neurodegeneration by activation of the LCN2/SLC22A17 axis-mediated iron transport

city: Suzhou

first: Lele

last: Liu

state: Jiangsu

address: No. 199, Ren'ai Road

department: college of pharmacy

institute: Soochow University

summary: The mechanisms underlying selective loss of dopaminergic (DAergic) neurons in substantia nigra pars compacta in Parkinson’s disease (PD) remain unclear. The NUS1 gene, encoding NgBR, has recently been identified as a novel PD risk gene. Here, we show that NgBR is highly expressed in microglia, and microglia-specific NgBR deficiency drives progressive PD-like motor deficits and DAergic neurodegeneration via lipocalin-2 (LCN2)/SLC22A17 axis-mediated iron transport. NgBR deficiency in microglia triggers endoplasmic reticulum stress and PERK-ATF4-NUPR1 pathway to selectively upregulate LCN2-mediated iron efflux rather than cytokines. Strikingly, microglia-derived iron-loaded Holo-LCN2 but not iron-free Apo-LCN2 is internalized by SLC22A17, a receptor we identified as specifically highly expressed in DAergic neurons that induces iron overload/ferroptosis. Pharmacological inhibition of the LCN2/SLC22A17 axis or ferroptosis alleviate DAergic neuron loss and PD-like symptoms caused by microglial NgBR deficiency in vitro and in vivo. Our findings establish intercellular iron homeostasis imbalance as a selective pathological driver in PD and highlight the LCN2/SLC22A17 axis for specific therapeutic targeting.

overall_design: RNA-seq profiling of control BV2 cells (NC) and Nus1 knockdown BV2 cells (644 and 878) 48 hours following the introduction of the knockdown system.

first: Yan

last: Hong

first: Yanni

last: Ma

first: Rui

last: Wang

first: Qian

last: Sun

first: Chenchen

last: Mu

first: Jifeng

last: Guo

first: Beisha

last: Tang

first: Guanghui

first: Haigang

last: Ren


title: The roles of GTPases in biogenesis of the large ribosomal subunit

city: Göttingen

first: Katherine

last: Bohnsack

address: Humboldtallee 23

institute: University Medical Centre Göttingen

summary: Ribosomes are ribonucleoprotein complexes responsible for protein synthesis, and their biogenesis is a complex, highly orchestrated process. Formation of the ribosomal subunits is coordinated by trans-acting ribosome biogenesis factors (RBFs) that mediate ribosomal RNA (rRNA) modification, folding and processing, as well as the incorporation and positioning of ribosomal proteins (RPs). GTPases are among the RBFs that consume energy to drive structural rearrangements of pre-ribosomal particles, and several of these enzymes play important roles in assembly of the large subunit. Protein-RNA crosslinking approaches were used to identify GNL3L binding sites on pre-ribosomes and map RNA contact sites on the GTPase. The findings of this study support the involvement of GNL3L in large subunit biogenesis and provide new insights into its functions in this process.

overall_design: Flp-In T-Rex HEK293 cell lines expressing GNL3L-His-FLAG or the His-FLAG tag alone were crosslinked using UV and analysed by CRAC.

first: Chairini

last: Thomé

first: Nicolas

last: Lemus-Diaz

first: Philipp

last: Hackert

middle: E


title: Systematic Perturbation of Cultures of K. Phaffii Show Gene Signatures Associated with 1 Specific Productivity of Recombinant Proteins [K. phaffii S-1076 RNA-seq]

first: Charles

last: Whittaker

overall_design: Gene expression data for K. phaffii strain (S-1076) expressing the monoclonal antibody (mAb) trastuzumab grown in glycerol-containing complex media, then exchanged into complex media with 10 mM Glutathione containing eight different carbon coifeeds, three different co-feed concentrations, and three different methanol concentrations (8x3x3=72 conditions) with 2-3 replicates per condition.

first: Raghav

last: Acharya

first: Joshua

last: Hinckley

first: Rachel

last: Barry

first: Eugenie

last: Cha

first: Harini

last: Narayanan

first: Brittney

last: Sunday

first: Hayley

last: Ford

first: John

last: Love


title: Systemic administration of CD28-deprived in vitro-induced regulatory T cell attenuates pathological remodeling caused by pressure overload-induced cardiac fibrosis

city: Suita

first: Takura

last: Taguchi

state: Osaka

address: 2-15, Yamadaoka

country: Japan

institute: Department of Cardiovascular Surgery, The University of Osaka Graduate School of Medicine

summary: In heart failure, myocardial fibrosis driven by immune-mediated inflammation contributes to progressive cardiac dysfunction. We investigated the therapeutic efficacy of CD28-deprived in vitro–induced regulatory T cells (iTreg), a stable Treg subset, in a murine model of pressure overload–induced heart failure. CD28-deprived iTreg were intravenously administered one week after transverse aortic constriction (TAC). Treated mice showed improved cardiac function, reduced ventricular fibrosis, and Treg localization in the heart. Transcriptomic analysis revealed downregulation of profibrotic and proinflammatory pathways, including TNF-α/NF-κB and TGF-β signaling. These findings suggest that systemic delivery of CD28-deprived iTreg effectively modulates the inflammatory microenvironment and suppresses cardiac remodeling, offering a promising immunomodulatory strategy for treating heart failure.

overall_design: RNA sequencing of left ventricular myocardium from mice undergoing TAC surgery, including control and Treg groups.

first: Takuji

last: Kawamura

first: Norihisa

last: Mikami

first: Kosuke

last: Torigata

first: Akima

last: Harada

first: Lisa

last: Fujimura

first: Emiko

last: Ito

first: Maki

last: Takeda

first: Kenji

last: Miki

first: Shimon

last: Sakaguchi

first: Shigeru

last: Miyagawa


title: Percentage of CD56⁺ monocytes at neutrophil engraftment is associated with the incidence of acute graft-versus-host disease

city: Nagoya

first: Takahiko

last: Sato

state: Aichi

address: 65 Tsurumai, Showa-ku

department: Department of Hematology

institute: Nagoya University Graduate School of Medicine

summary: Acute graft-versus-host disease (GVHD) is one of the serious complications following allogeneic hematopoietic stem cell transplantation (HSCT) that leads to non-relapse mortality. Although several biomarkers for acute GVHD have been proposed, no definitive predictive markers are clinically available. Recent studies have shown that a specific subset of peripheral blood monocytes expresses CD56 (neural cell adhesion molecule [NCAM]), particularly under inflammatory conditions; however, their role in allogeneic HSCT remains unclear. Single-cell RNA sequencing of peripheral blood mononuclear cells from patients with acute GVHD identified a unique monocyte subset characterized by an NCAM signature. Prospective multicolor flow cytometry analysis of peripheral blood samples revealed a transient increase in CD56⁺ monocytes at the time of neutrophil engraftment. Gene set enrichment analysis demonstrated pro-inflammatory transcriptome signatures of the CD56⁺ monocyte fraction. Notably, a reduced frequency of CD56⁺ monocytes was significantly associated with a lower incidence of acute GVHD, suggesting their potential as novel predictive cellular biomarkers of acute GVHD.

overall_design:
CD56+ monocytes and CD56- monocytes were sorted at the neutrophil engraftment of allogeneic HSCT. Total RNA was extracted using TRIsol Reagent (Thermo Fisher Scientific) and RNeasy® Micro Kit (QIAGEN, Netherlands), and sequenced by the Illumina NovaSeq 6000 (Illumina, CA, USA). Raw FASTQ files were processed using fastp version 0.19.5 20, and reads were aligned to the GRCh38 human reference using STAR software version 2.7.11b 21. RSEM version 1.3.1 was used for transcript quantification.

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

first: Ken

last: Hashimoto

first: Yuichi

last: Ishikawa

first: Yuki

last: Okuhiro

first: Daisuke

last: Sugiyama

first: Zhang

last: He

first: Sachiko

first: Yuichiro

last: Inagaki

first: Kotaro

last: Miyao

first: Masashi

last: Sawa

first: Takanobu

last: Morishita

first: Tatsunori

last: Goto

first: Tetsuya

last: Nishida

first: Nobuaki

last: Fukushima

first: Kazutaka

last: Ozeki

first: Ryo

last: Hanajiri

first: Seitaro

last: Terakura

first: Hiroyoshi

last: Nishikawa

first: Hitoshi

last: Kiyoi


title: The Dual Immunoregulatory Role of Creb3l1 Underlying Latent and Severe Tuberculosis Clinical Manifestation

city: Ribeirao Preto

first: Luiz Gustavo

middle: Araujo

last: Gardinassi

state: Sao Paulo

address: Av. dos Bandeirantes, 3900

country: Brazil

institute: Ribeirao Preto Nursing School/ University of São Paulo

summary: During tuberculosis (TB), organ-specific immune responses and intracellular pathways play critical roles in disease progression and prognosis. Identifying genes that regulate these immune mechanisms remains a key challenge in improving TB management strategies. To investigate genes potentially associated with enhanced resistance to TB and the modulation of immune responses, we analyzed RNA-seq data from whole cells isolated from the lungs and livers of mice infected with Mycobacterium tuberculosis (Mtb). We hypothesized that these two organs mount distinct responses to infection, supported by differences in the immune response and bacterial burden kinetics observed in each tissue. Our analysis revealed differential gene expression profiles between the lungs and livers, primarily involving metabolic and immune-related pathways. Through meta-analysis, we identified orthologous genes shared between Mtb infected mice and human patients with latent pulmonary TB. In this omics analysis, the four genes, Creb3l1, Myo7b, Cyyr1, and Cbs, were differentially expressed and associated with either resistance or susceptibility. In vitro assays further demonstrated that knockdown of Creb3l1 in Mtb infected monocytes impaired key effector functions, including phagocytosis, bacterial killing, and apoptosis. These findings suggest that Creb3l1 contributes to bacterial containment in the lungs and may play a protective role during infection. Notably, Creb3l1 expression was also elevated in the peripheral blood of patients with severe TB, and correlated with increased levels of pro-inflammatory cytokines such as IL-17, IL-2, and IFN-γ. This pattern points to a potential dual role for Creb3l1 in TB. Under controlled conditions, such as latent infection, it may help regulate pulmonary inflammation, whereas during active disease, the expression is elevated in circulating leukocytes and associated with heightened cytokine production, which may reflect an immune attempt to counteract progressive infection.

overall_design: RNA-seq profiling of lung and liver tissue from Balb/c mice infected with Mycobacterium tuberculosis for 14 and 19 days.

first: Felipe

last: Lima

first: Luiz

first: Fabiani

last: Frantz


title: CUT-TAG for AML cell line (CMK)

first: Fang

last: Fang

address: 92 Zhongnan street

institute: Children’s Hospital of Soochow University

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to explore CUT-TAG results in AML cell line.

overall_design: CUT-TAG for CTNNB1 in CMK cell line

first: Jian

last: Pan

first: Jia

last: Cheng


title: Effect of combined deletions of ZFP36L1 and ZFP36L2 on gene expression in the mouse liver

city: New Hill

first: Yogesh

last: Saini

state: NC

address: 3644 Rail Overlook Drive

institute: NCSU

summary: RNA-binding proteins, Zinc Finger Protein 36 Like 1 (ZFP36L1) and Zinc Finger Protein 36 Like 2 (ZFP36L2), post-transcriptionally regulate the expression of genes involved in various cellular processes. Here, we show that the combined liver-specific loss of ZFP36L1 and ZFP36L2 in mice (L1/L2dKO) results in cholestatic liver injury as evidenced by the elevated hepatic and serum levels of total bile acids (TBA) and liver injury biomarkers, including ALP, ALT, and AST and characterized by the presence of bile infarcts followed by marked inflammation, cellular proliferation, and fibrosis. L1/L2dKO mice also showed reduced bile flow rate and decreased biliary TBA excretion. Subsequent analyses revealed impaired bile canalicular morphogenesis by postnatal day 7, as evidenced by F-actin and Zona Occludens-1 staining patterns. Additionally, RNA sequencing of whole liver samples of L1/L2dKO mice showed significant perturbation of ZFP36L1/ ZFP36L2 target genes associated with cholestasis, inflammation, fibrosis, and cell cycle. Overall, the study reveals critical beneficial roles of ZFP36L1 and ZFP36L2 in maintaining liver homeostasis.

overall_design: RNA-seq profiling of livers from combined liver-specific ZFP36L1 and ZFP36L2 knockout and flox-only control adult male mice


title: Decoding PRC2 stage-specific roles during germline-embryo-germline transition

city: Boston

first: Yi

last: Zhang

address: 200 Longwood Avenue WAB 149

department: PCMM

institute: Boston Children's Hospital / Harvard Medical School

summary: Early mammalian development undergoes a series of precisely regulated processes, including the maternal-to-zygotic transition (MZT), the first lineage specifications, gastrulation, and the primordial germ cell (PGC) specification. While key transcription factors (TFs) and epigenetic modifiers that regulate these events have been investigated, the intricate transcriptional and epigenetic networks, such as interplays between TFs and chromatin modifications remain to be fully established. The Polycomb Repressive Complex 2 (PRC2) mediates transcriptional silencing by depositing H3K27me3 and is essential for proper gastrulation. However, the PRC2 function in early development may not be fully captured due to limitations of conventional knockout approaches. Here we applied the degradation tag (dTAG) system to deplete the PRC2 acutely and stage-specifically in mouse pre- and early post-implantation development. This work revealed previously underappreciated functions of PRC2 in regulating key events in early development including the MZT, the first and second cell lineage specifications. In addition, we demonstrated that PRC2 restricts premature activation of developmental genes in embryonic day (E)4.5 inner cell mass (ICM) cells, facilitating proper epiblast specification. Moreover, systematic analysis of H3K27me3 and H3K4me3 dynamics reveals that epigenetics interplay drives the chromatin bivalency establishment at E4.5 ICM, which is earlier than previously thought that the bivalency appears at post-implantation stages. Lastly, we elucidated the lineage crosstalk dependent PRC2-ESRRB-BMP4 signal axis in regulating PGCs specification during gastrulation. Together, our results demonstrate that PRC2 plays stage-specific roles through dual interplay with epigenetic modification and transcription factors during germline-embryo-germline transition.

overall_design: Eed-dTAG mice were generated. RNA-seq, ATAC-seq and CUT&RUN were used to study the Eed's function during mouse early embryo development.

first: Chengjie

last: Zhou

first: Meng

last: Wang

first: Zhiyuan

last: Chen


title: Reduction of di-methyl histone H3K9 (H3K9me2) in CUX1-deficient cells.

city: Chicago

first: Angela

last: Stoddart

state: IL

address: 900 E. 57th Street

department: Pathology

institute: University of Chicago

summary: CUX1 represses endogenous retroelements leading to an increased inflammatory signature after CUX1 loss.  CUX1 interacts with the histone H3K9 methyltransferase, EHMT2 (G9a). EHMT2, in turn, represses endogenous retroelements (EREs) via the deposition of H3K9me2. CUX1-deficient K562 cells exhibited a broad reduction in H3K9me2 deposition, with particularly marked losses at LINEs, SINEs, and LTRs, supporting the model that CUX1 represses ERE expression through EHMT2-mediated H3K9me2 deposition.

overall_design: Cut & Run of di-methyl Histone H3K9 (H3K9me2) in gControl and gCUX1 KO K562 cell lines.

first: Tanner

last: Martinez

first: Megan

last: McNerney


title: Single-cell RNA sequencing of primary cultures of neonatal digit amputation wound cells 4 days post-amputation and treated with FGF2 for 24H

city: College Station

first: ling

last: yu

state: TEXAS

address: VMR Bldg 1197  Room 327  Mail Stop 4466

department: VTPP

institute: Texas A&M university

summary: Primary wound cells were isolated   and  treated with FGF2 (100 ng/ml) for 24 hours, and analyzed by single cell RNA sequencing (scRNAseq). The dataset from FGF2 treated wound fibroblasts was compared to our previously published dataset for un-treated primary wound cells(GSE185197)  to determine  the   transcriptomic changes in treated fibroblasts. We  found  Hmga1 and Hmga2 as novel FGF2-inducible genes potentially linked to fibroblast dedifferentiation during FGF2 induced blastema formation.

overall_design: First, FGF2 treated 24H wound cells was prepared using a 10X Genomics Chromium Controller and deeply sequenced.  Then , perform a 2-dimensional t-SNE projection of un-treated and FGF2-treated dataset and differentially expressed gene (DEG) analysis.

first: Ling

last: Yu

first: Mingquan

last: Yan

first: Katherine

last: Zimmel

first: Osama

last: Qureshi

first: Yu-Lieh

last: Lin

first: Lindsay

last: Dawson

first: Regina

last: Brunauer

first: Larry

last: J. Suva

first: Connor

last: P. Dolan

first: Ken

last: Muneoka


title: Targeting EBF2 condensates modulates adipose thermogenesis

first: Li

state: Guangdong province

address: 190 Kai Yuan Avenue, Science Park

institute: Guangzhou Institutes of Biomedicine and Health,Chinese Academy of Sciences

summary: Brown adipose tissue (BAT) actively dissipates energy and represents a promising therapeutic target for combating obesity. The establishment of the thermogenic gene program is orchestrated by the pioneering transcription factor Early B Cell Factor 2 (EBF2). Here, we identified a low-complexity C-terminal domain (CTD) within EBF2 that drives biomolecular condensation and is essential for thermogenic activation. Deletion of the CTD (EBF2 ΔCTD) or mutation of key proline residues within this region did not affect the genomic binding of EBF2, but profoundly impaired its ability to undergo phase separation and to drive brown adipocyte differentiation and thermogenic function both in vitro and in vivo. Fusion of the intrinsic disorder region (IDR) of FUS to EBF2 ΔCTD fully restored its function, whereas fusion with MED1-IDR resulted in distinct phenotypic outcomes. Mechanistically, EBF2 formed condensates with ZFP423, a key repressor of brown adipocyte differentiation. This co-condensation excluded HDAC1 from the complex, establishing a permissive chromatin environment that promotes active transcription. Furthermore, a phase separation-based phenotypic drug screen identified compounds that modulated EBF2 condensate formation and thermogenic genes expression. Our findings reveal phase separation as an emerging regulatory mechanism underlying brown fat determination and suggest that targeting biomolecular condensation offers a novel therapeutic strategy for obesity.

overall_design: We collected multi-omics data from the initial and terminal stages of the LGT differentiation process, including RNA-seq, ATAC-seq, and CUT&Tag assays targeting Ebf2, H3K27ac, Zfp423, Pprag, and Hdac1,Chd4,Brg1.

first: Xianju

first: Chen

last: Li

first: Zongzheng

last: Gui


title: Hormonal and transcriptional responses to 1,4-dimethylnaphthalene reveal tissue-specific mechanisms of sprout suppression in potato tubers during storage

city: Fargo

first: Evandro

last: Fortini

address: 1605 Albrecht Blvd N

institute: United States Department of Agriculture

summary: This study employed an transcriptomic and targeted metabolomic approach to investigate the hormonal and molecular changes induced by 1,4-dimethylnaphthalene (DMN) in potato tubers (cv. 'Russet Burbank') during a long-term storage period.

overall_design: Comparative gene expression analyses were conducted on potato tubers subjected to single (1x), double (2x), or triple (3x) DMN applications. Samples were collected at various storage times (0, 3, 6, 9, and 12 weeks) following DMN application. Tissue-specific responses were assessed in the primary and secondary meristems, and the tuber flesh.

first: Tadeu

last: Oliveira

first: Dipayan

last: Sarkar

first: Munevver

last: Dogramaci


title: de novo H3.2K9me2 deposition pathway establishes heterochromatin for suppressing transposon mobilizations during somatic development [Nanopore]

city: Shanghai

first: Yu

last: Liang

address: Yueyang Road 320

department: SIBCB

institute: CEMCS, CAS

summary: Histone variants along with their associated chaperons have been considered as one of the major complexes to provide versatility in organizing chromatin structure, contributing to define dynamic landscapes and maintaining genome integrity. Post translational modifications (PTMs) of H3 variants serves as very important factors in promoting heterochromatin assembly, protecting telomere stability and suppressing transposon activity. However, the precise mechanism regarding the specific PTMs of H3 variants in performing the function of heterochromatin formation and transposon silencing during somatic development remains elusive. Here, by monitoring retrotransposon activity and mobilizations during somatic development, we found that DNA synthesis-coupled (DSC) H3.2K9me2 deposition pathway plays a pivotal role in heterochromatin establishment and transposon suppression. Depleting the factors involving in the complex of DSC H3.2 chaperons, but not DNA synthesis-independent (DSI) H3.3 chaperons, led to the failure of heterochromatin formation, unleashing massive activation of retrotransposons. DSC chaperones specifically establish di-methylation at H3.2K9 site through directly interacting and recruiting histone methyltransferase, G9a, to heterochromatic regions. Intriguingly, we also discovered that the crosstalk of DSC H3.2K9me2 and DSI H3.3K9me3 in heterochromatin during development is dynamically regulated and properly balanced. Although the DSI H3.3K9me3 could efficiently incorporate into transposon DNA when DSC H3.2K9me2 deposition pathway was compromised, H3.3K9me3 was still unable to establish functional heterochromatin to achieve transposon silencing during development. Collectively, our discoveries provide a framework to understand how cells employ deposition of specific histone modifications to establish and underpin heterochromatin states for silencing transposon mobilizations and safeguarding genome stability.

overall_design: We employed Nanopore sequencing to capture the bona fide mobilization events within Drosophila hindguts and RNA-seq to quantify transposon expression. Combined with ChIP-seq, we were able to gain a comprehensive understanding of the breakpoints generated at transposon insertion sites and the changes in histone modifications on the transposons.

first: Yini

last: Luo

first: Yazi

last: Deng

first: Wei

last: Wu

first: Lu

last: Wang


title: de novo H3.2K9me2 deposition pathway establishes heterochromatin for suppressing transposon mobilizations during somatic development

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

overall_design: Refer to individual Series


title: Cold-inducible RNA-binding protein promotes lung fibroblast activation via TLR2/4-dependent IL-6 signaling: implications in pulmonary fibrosis

city: hamamatsu

first: Yasutaka

last: Mochizuka

address: handayama

institute: Hamamatsu University School of Medicine

summary: This study investigates the role of Cold-inducible RNA-binding protein (CIRBP) in the pathogenesis of pulmonary fibrosis. Using RNA-seq, we analyzed gene expression changes in primary mouse lung fibroblasts stimulated with recombinant CIRBP or PBS control. CIRBP significantly upregulated cytokine-related gene expression, including IL-6, and promoted collagen production and fibroblast proliferation via TLR2/4 pathways.

overall_design: Primary lung fibroblasts were isolated from wild-type C57BL/6 mice and treated with recombinant mouse CIRBP (1.0 µg/mL) or PBS control for 24 hours after serum starvation. Total RNA was extracted and subjected to RNA sequencing (n = 3 per group). The sequencing was performed in a single batch to avoid technical variation.

first: Hironao

last: Hozumi


title: Transcriptomes of human whole blood cells in sepsis patients

city: Beijing

first: Haiyan

last: Xue

address: 11th Xizhimen South Street

institute: Peking University People's Hospital

summary: This study focuses on the transcriptomic characteristics of whole blood cells in sepsis patients. Sepsis is a severe systemic infection that often leads to multiple organ dysfunction. Its pathogenesis is complex and involves the abnormal activation of various cellular and molecular pathways. By analyzing the transcriptomes of whole blood cells in sepsis patients through sequencing, we aim to reveal the key changes in gene expression and their potential regulatory mechanisms during the onset and progression of sepsis. The study included sepsis patients with different underlying diseases and also set up a healthy control group for comparative analysis.

overall_design: RNA-seq profiling from the whole blood cells from 10 healthy controls and 30 sepsis patients.

first: Fengxue

last: Zhu


title: UVI-5008: A A First-in-Class Reversible, Non-Covalent Dual Epigenetic and Kinase Inhibitor Targeting Bruton's Tyrosine Kinase in Resistant B-Cell Malignancies

city: Napels

first: Wout

last: Megchelenbrink

address: Vico de Crecchio 7

department: Precision Medicine

institute: University of Campania "Luigi Vanvitelli”

summary: Bruton Tyrosine Kinase (BTK) is a crucial target for anticancer, anti-inflammatory, and antiviral therapies. BTK inhibitors (BTKis) have revolutionized the therapeutic options for B-cell malignancies, including mantle cell lymphoma (MCL), chronic lymphocytic leukemia (CLL), lymphoplasmacytic lymphoma/Waldenström macroglobulinemia and small lymphocytic lymphoma. Despite the therapeutic efficacy achieved with BTKis, some patients develop relapse and drug resistance. Different new approaches offer an alternative way to prevent or overcome the resistance: by targeting bypass mechanisms by combination therapies;temporal sequencing of therapies;chimeric antigen receptor-modified T (CAR-T) cell therapy;real-time monitoring of patient response; next-generation BTKis (ngBTKis). Above all, ngBTKis and combined treatment are proposed as strategies more effective and less toxic, to improve disease management and patient survival.   Epigenomics of B-cell malignancies may explain the intraclonal diversification and help elucidating the reason of emerging relapse and treatment refractoriness/resistance. We identified the first next-generation, selective, reversible, non-covalent BTK-epigenetic inhibitor (BTK-epi-inhibitor) capable of downregulating BTK at both the transcriptional and enzymatic levels.To characterize this molecule, we employed a comprehensive and integrative experimental approach, encompassing genomic, epigenomic, transcriptional, and on-target analyses. We investigated the molecular and functional mechanisms underlying the activity of UVI5008, while also evaluating its therapeutic efficacy, toxicity, pharmacokinetics, and safety profile. These studies were conducted through a combination of in vitro (cell culture), ex vivo, and in vivo (animal model) experiments, aiming to minimize side effects and off-target toxicity. Importantly, we demonstrated that UVI5008 inhibits both wild-type BTK and the C481S mutant isoform—a key mutation associated with resistance to first-generation BTK inhibitors—across in silico, ex vivo, and in vivo models of chronic lymphocytic leukemia (CLL) and B-cell lymphoma. Treatment with UVI5008 led to a significant increase in cell death, accompanied by a reduction in BTK protein levels, its phosphorylation, and downstream signaling activity. Taken together, these data highlight the novelty, potency, and clinical relevance of UVI5008 as a new  therapeutic strategy for BTK-driven malignancies, especially in relapsed/refractory (R/R) CLL and B-cell lymphoma, where current therapies often fail.

overall_design: Rna-seq profiling of Mec-1, Mino and Granta-519 cell lines ctr and after 24h of treatment with uvi5008 5 µM

first: Lucia

last: Altucci

first: Carmela

last: Dell'aversana


title: The deubiquitinase OTULIN regulates tau expression in neurons

city: Albuquerque

first: Kiran

last: Bhaskar

state: New Mexico

address: 915 Camino De Salud NE

department: Molecular Genetics and Microbiology

institute: University of New Mexico

summary: Degradation of aggregation-prone tau is regulated by the ubiquitin-proteasome system (UPS) and autophagy, which are impaired in Alzheimer’s disease (AD) and related tauopathies causing tau aggregation. Protein ubiquitination with linkage specificity determines the fate of proteins that can be either degradative or stabilization signals. While the linear M1-linked ubiquitination on protein aggregates is a signaling hub that recruits various ubiquitin-binding proteins for coordinated actions of protein aggregates turnover and inflammatory NF-kB activation, the deubiquitinase OTULIN counteracts with the M1-linked ubiquitin signaling. However, the exact role of OTULIN on tau aggregate clearance in AD is unknown. We did bulk RNA sequencing in wild-type and OTULIN knockout (by CRISPR-Cas9) SH-SY5Y human neuroblastoma cells. Our results showed a 14.39-fold down-regulation of tau mRNA levels and differential expression of many other genes associated with autophagy, the ubiquitin-proteasome system (UPS), NF-kB pathway, and RNA metabolism. Together, our results suggest for the first time a non-canonical function for OTULIN in regulating gene expression and RNA metabolism, which may have a significant pathogenic role in AD and related tauopathies.

overall_design: Poly (A) RNA sequencing experiment was designed to compare and quantify the levels of differential gene expression and RNA transcripts between human neuroblastoma SH-SY5Y wild type (WT) and CRISPR-Cas9-mediated OTULIN knockout (KO) cells. Three replicates were included from each genotype for quantitative analysis.

first: Karthikeyan

last: Tangavelou

first: Virginie

last: Bondu

first: Mingqi

last: Li

first: Francesca-Fang

last: Liao


title: Selective Janus kinase inhibitors exhibit different effects in murine bone marrow failure

city: Bethesda

first: Shouguo

last: Gao

state: MD

address: 9000 Rockville Pike

department: NHLBI

institute: National Institutes of Health

summary: We compared transcriptomes of bone marrow mononuclear cells (BMMNCs) and flow-sorted Lin-Sca-1+c-kit+ cells (LSK/HSPCs) from untreated bone marrow failure (BMF) mice and the mice treated with JAK inhibitors (JAKi): baricitinib (BAR), solcitinib (SOL), fedratinib (FED), and tofacitinib (TOF) at day 10 post BMF induction.

overall_design: RNA profiles of single bone marrow mononuclear cells (BMMNCs) and HSPCs were generated by deep sequencing using Illumina NovaSeq. Two pooled BMMNCs from each group of the untreated bone marrow failure (BMF) mice and the mice treated with baricitinib, solcitinib, fedratinib, and tofacitinib. One pooled flow-sorted HSPCs from individual groups.

first: Xingmin

last: Feng

first: Zhijie


title: The deubiquitinase OTULIN regulates tau expression in neurons [sAD2.1_WTC11_iPSNs]

summary: Degradation of aggregation-prone tau is regulated by the ubiquitin-proteasome system (UPS) and autophagy, which are impaired in Alzheimer’s disease (AD) and related tauopathies causing tau aggregation. Protein ubiquitination with linkage specificity determines the fate of proteins that can be either degradative or stabilization signals. While the linear M1-linked ubiquitination on protein aggregates is a signaling hub that recruits various ubiquitin-binding proteins for coordinated actions of protein aggregates turnover and inflammatory NF-kB activation, the deubiquitinase OTULIN counteracts with the M1-linked ubiquitin signaling. However, the exact role of OTULIN on tau aggregate clearance in AD is unknown. We did bulk RNA sequencing in human inducible pluripotent stem cell (iPSC)-derived neurons (iPSNs) from a healthy control (WTC11) and an individual with late-onset sporadic AD (sAD2.1), which shows downregulation of ubiquitin ligase activating factors (MAGEA2B and MAGEA) and OTULIN long non-coding RNA (lncRNA-OTULIN) compared to healthy control WTC11 iPSNs. In sAD2.1 iPSNs, downregulated lncRNA-OTULIN is inversely correlated with increased levels of OTULIN protein and phosphorylated tau at p-S202/p-T205 (AT8), p-T231 (AT180), and p-S396/p-S404 (PHF-1). Loss of OTULIN deubiquitinase function using pharmacological inhibitor UC495 or CRISPR-Cas9-mediated OTULIN gene knockout causes a significant reduction of total tau and phosphory-lated tau at AT8 epitope in sAD2.1 iPSNs.  Together, our results suggest for the first time a non-canonical function for OTULIN in regulating gene expression and RNA metabolism, which may have a significant pathogenic role in AD and related tauopathies.

overall_design: Poly (A) RNA sequencing experiment was designed to compare and quantify the levels of differential gene expression and RNA transcripts between human inducible pluripotent stem cell (iPSC)-derived neurons (iPSNs) from a healthy control (WTC11) and an individual with late-onset sporadic AD (sAD2.1). Three replicates were included from each genotype for quantitative analysis.

first: Wei


title: CUT-TAG for AML cell line CMK

first: Fang

last: Fang

overall_design: CUT-TAG for USP20 in CMK cell line

first: Jian

last: Pan

first: Jia

last: Cheng


title: CUT&Tag for AML cell line CMK

overall_design: CUT-TAG in CMK cell line


title: Bulk RNA sequencing of bone marrow CD8+ T cells in BMF mice treated with or without baricitinib at different time points

summary: We performed bulk RNA sequencing of bone marrow  CD8+ T cells in bone marrow failure (BMF) mice treated with or without baricitinib (BAR) at day 10 and day 14 post BMF induction to investigate the time-dependent gene expression changes of T cells induced by JAK1/2 inhibition (BAR).

overall_design: RNA profiles of flow-sorted bone marrow CD8+ T cells from BMF mice treated with or without BAR were generated by deep sequencing using Illumina NovaSeq. Five pooled cells for BMF and BMF+BAR at day10; 3 pooled cells for BMF and BMF+BAR at day 14.


title: Kupffer Cell-Derived IL-6 Aggravates Radiation-Induced Liver Disease by Activating Hepatocyte STAT3

city: cahgnzhou

first: Wendong

last: Gu

address: NO.185, juqianjie road

institute: The Third Affiliated Hospital of Soochow University

summary: Radiation-induced liver disease (RILD) is a serious complication of radiotherapy for upper abdominal tumors, the cellular and molecular basis of which remain mostly unclear. Sprague‒Dawley (SD) rats were exposed to a single dose of 30 gray (Gy) at a dose rate of 400 cGy/min on the right upper quadrant of the abdomen to induce liver injury in vivo. At 2 weeks after IR, Kupffer cells (KCs) characteristically expressed Il6. IL-6 activated the JAK‒STAT signaling pathway in hepatocytes. However, the critical downstream target genes were unclear.

overall_design: ChIP-seq of P-STAT3 in primary hepatocytes isolated from SD rats 2 weeks after a single 30 Gy IR.

first: Xing

last: Song


title: de novo H3.2K9me2 deposition pathway establishes heterochromatin for suppressing transposon mobilizations during somatic development [ChIP-Seq]

first: Yu

last: Liang

first: Yini

last: Luo

first: Yazi

last: Deng

last: Wu

first: Lu

last: Wang


title: Mechanism of radiation-induced liver disease based on single-cell transcriptomics

summary: We report single-cell transcriptomic analyses of whole livers from irradiated and controlled Sprague‒Dawley (SD) rats. The aim of the study was to elucidate the molecular and cellular drivers of radiation-induced liver disease (RILD), a serious complication of radiation therapy for upper abdominal tumors in humans.

overall_design: Six male Sprague‒Dawley (SD) rats were split into control and irradiation groups, with three rats in each group. For the irradiation group, rats were exposed to a single dose of 30 gray (Gy) at a dose rate of 400 cGy/min on the right upper quadrant of the abdomen to induce liver injury in vivo. The rats were sacrificed 2 weeks after irradiation, and the liver were collected for scRNA-seq. The transcriptomic information of single cells was captured using the BD Rhapsody system.


title: RNA-seq for AML cell lines in either the absence or presence of USP20

city: Suzhou

summary: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare RNA-seq results for AML cell lines in either the absence or presence of USP20.

overall_design: 2 MV4-11 samples with no treatment and 2 MV4-11 samples with USP20 knockdown；2 CMK samples with no treatment and 2 CMK samples with USP20 knockdown


title: Antibody responses against bacterial glycans affinity mature and diversify in germinal centers.

city: Clayton

first: Nitika

last: Kandhari

state: VIC

address: Wellington Road

country: Australia

institute: Monash University

summary: Anti-carbohydrate antibodies (Abs) play crucial roles in pathogen control, but their generation remains poorly understood. By studying responses to Streptococcus pyogenes in humans, we reveal that the glycan-targeted response shifts from IgM towards IgG and IgA memory with age and antigen exposure across blood, spleen, and tonsils. Both natural colonization and controlled human infection with S. pyogenes increased class-switched B cells, with evidence of within-clone switching. Glycan-specific B cells readily participated in germinal center (GC) responses and showed robust somatic hypermutation despite a molecular signature consistent with receiving reduced T cell help. We conclude that mucosal pathogen encounters elicit glycan responses that class-switch, evolve and diversify through the GC. These findings reveal how age and infection history can influence the quality, quantity, and isotype use of glycan-specific B cells, with implications for the design and schedule of glycan-containing vaccines.

overall_design: Cryopreserved human tonsil cells from 4 Strep A-colonised paediatric donors were thawed, stained with GAC and SpyCEP antigen tetramers separately, and magnetically enriched for tetramer-binding cells. These cells were stained with surface marker antibodies and either TotalSeq™-C0252 anti-human Hashtag 2 Antibody (for SpyCEP tetramer-binding cells) or TotalSeq™-C0254 anti-human Hashtag 4 Antibody (for GAC tetramer-binding cells). Antigen tetramer-binding isotype switched (IgD–) B cells were isolated by Fluorescence-activated cell sorting (FACS) and sequenced by single cell sequencing.

first: Danika L.

last: Hill

first: Holly

last: Fryer

first: Catherine

last: Pitt


title: A longitudinal single-cell and spatial multiomic atlas of pediatric high-grade glioma

city: Philadelphia

first: Kai

last: Tan

state: PA

address: 3501 Civic Center Boulevard

department: Pediatrics

institute: Children's Hospital of Philadelphia

summary: Pediatric high-grade glioma (pHGG) is an incurable central nervous system malignancy that is a leading cause of pediatric cancer death. While pHGG shares many similarities to adult glioma, it comprises distinct disease entities. In this study, we longitudinally profile a molecularly diverse cohort of 16 pHGG patients through single-nucleus RNA and ATAC sequencing, whole-genome sequencing, and CODEX spatial proteomics to capture the evolution of neoplastic and microenvironmental features during disease progression and treatment. We define a set of core pHGG neoplastic cell states and observe differential tumor-myeloid interactions between malignant cell phenotypes. We find that essential neuromodulators and the interferon response are upregulated post-therapy and validated malignant cell-intrinsic targets. We observe an increase in oligodendrocytes upon progression and that they coordinate spatial motifs with proneural tumor cells. This multiomic atlas of longitudinal pHGG captures features of therapy response and provides a scalable reference for the study of pediatric brain tumors.

overall_design: Pediatric HGG cell lines treated with 0 or 4 Gy of ionizing radiation

first: Jonathan

last: Sussman

first: Thomas

last: De Raedt


title: de novo H3.2K9me2 deposition pathway establishes heterochromatin for suppressing transposon mobilizations during somatic development [RNA-Seq]

first: Wei


title: DMEM and EMEM as alternate growth media for pathogenic Leptospira

city: Storrs

first: Elsio

middle: Augusto

last: Wunder

state: CT

address: 61 N Eagleville

department: Pathobiology and Veterinary Sciences

institute: University of Connecticut

summary: Pathogenic Leptospira spp. must adapt to diverse environments during their life cycle, yet most laboratory studies rely on in vitro culture conditions that poorly reflect the host environment. This limitation restricts the relevance of transcriptomic studies for understanding leptospiral pathogenesis. In this study, we evaluated genome-wide gene expression of Leptospira interrogans grown in four different in vitro media—EMJH, DMEM, EMEM, and HAN—and compared their transcriptomic profiles to those obtained from bacteria recovered from the whole blood of infected hamsters.

sample_organism: Leptospira interrogans serovar Copenhageni str. Fiocruz L1-130

overall_design: RNA-seq of Leptospira interrogans serovar Copenhageni L1-130 in in vitro conditions like DMEM, EMEM, HAN, EMJH and Whole blood of hamster to mimic host environment.

first: Leandro

last: Garcia

first: Emillie

last: Karovic

first: Candace

last: Banasiewicz

first: Zheng

first: Francesc

last: Lopez-Giraldez

first: Jeffrey

last: P Townsend


title: Study on the Mechanism of Action of Hippuric Acid Targeting the UGDH/FOXK1/CD36 Signaling Axis in Regulating Metabolic Dysfunction-AssociatedSteatotic Liver Diseases

city: Changzhou

first: Shuai

last: Chen

address: 68 Gehu Rd, Wujin District

institute: The Affiliated Changzhou NO.2 People’s Hospital of Nanjing Medical University

summary: Metabolic dysfunction-associated steatotic liver diseases (MASLD)  is the most prevalent chronic liver disease globally, and the systemic metabolic disorders it triggers have become a serious public health concern. The endogenous metabolite hippuric acid (HA) is closely associated with a variety of metabolic disorders, but its molecular mechanisms in the progression of MASLD are not yet clear. Preliminary studies of this project have found that serum HA levels are significantly reduced in obese individuals and animals, and exogenous HA supplementation can alleviate body weight and liver fat accumulation in obese mice. It was also discovered that HA can reduce intracellular lipid deposition in hepatocytes by inhibiting CD36 expression. Mechanistic studies have revealed that HA can target UGDH, enhancing its binding with the transcription factor FOXK1, and inhibiting FOXK1 expression significantly reduces the transcription level of CD36. Therefore, the scientific hypothesis is proposed: HA inhibits liver fat accumulation by regulating the UGDH/FOXK1/CD36 signaling axis. This project will combine clinical sample analysis with in vitro and in vivo experiments to systematically elucidate the molecular mechanisms by which HA targets UGDH to inhibit nuclear expression of FOXK1, thereby regulating CD36 transcription. This will enhance our understanding of HA's role in lipid metabolism regulation and the development of MASLD and lay a theoretical foundation for uncovering effective therapeutic targets for MASLD.

overall_design: The obese mouse model was intraperitoneally injected with HA at a dose of 60 mg/kg twice a week for 5 weeks.


title: BULK RNAseq - SV40 large T antigen immortalization of primary mouse intestinal epithelial stem and progenitor cells

city: Cleveland

first: Julie

last: Zhou

state: OH

address: 9500 Euclid Ave

institute: Cleveland Clinic Research Institute

summary: Developing new tools for studying the intestinal epithelium is essential for addressing unan-swered questions in development, physiology, and disease pathology. Primary intestinal epitheli-al stem cell (ISC) cultures are an invaluable in vitro model system. However, their cultivation re-mains technically demanding and costly, limiting their accessibility and use. Though commonly utilized, most extant intestinal epithelial cell lines were derived from colon adenomas from vari-ous animals, are largely uncharacterized, and possess a plethora of genetic abnormalities, thus restricting rigor, reproducibility, and physiological relevance. Here we derived immortalized intestinal epithelial cell lines (iIECs) from six jejunal and six colonic stem cell-enriched spheroid cultures from wild type C57BL/6 mice. We transduced ISCs with a lentiviral vector encoding the SV40 large T antigen, and subsequently selected lines through adaptation to growth on plastic. Additionally, we weaned these lines off conditioned medium including the growth factors Wnt3a, Rspo3, and Noggin. We found that iIECs have growth characteristics similar to conventional plastic adapted cell lines. While we observed transcriptional signatures of epithelial-mesenchymal transition (EMT), these cells still exhibited epithelial characteristics including ex-pression of junctional proteins, mounted an innate immune response to viral infection, and se-creted cytokines in response to LPS stimulation. We further demonstrated the utility of iIECs through transient and stable genetic manipulation. iIECs occupy an important experimental niche through offering a scalable, practical, and physiologically relevant model system that can func-tion as a discovery platform before transitioning to primary spheroid cultures and/or animal models. We propose that they will be a valuable tool for advancing understanding of intestinal epithe-lial biology.

overall_design: Bulk RNA sequencing (RNAseq) comparing SV40 large T antigen immortalized mouse intestinal epithelial cells (iIEC) and their parental counterparts consisting of cultured primary mouse intestinal epithelial spheroids comprised of intestinal stem and progenitor cells. Jejunal and colonic spheroid lines derived from 3 different mice were used for this analysis. iIECs from of 3 clonal populations were used per each tissue site. Isolated mRNA was submitted for high-throughput sequencing.

middle: Y

first: Ryan

middle: J

last: Musich

first: Amalu

last: Job

first: Thaddeus

middle: S

last: Stappenbeck


title: TORC1-driven translation of Nucleoporin44A promotes chromatin remodeling and germ cell-to_x0002_4 maternal transition in Drosophila

city: New York

first: BiNGS

last: Core

address: 3 E 101st St

institute: Icahn School of Medicine at Mount Sinai

overall_design: Refer to individual Series


title: TORC1-driven translation of Nucleoporin44A promotes chromatin remodeling and germ cell-to_x0002_4 maternal transition in Drosophila [cr_nup44a_zfrp8]

summary: Oocyte specification is a critical developmental transition that requires the coordinated repression of germ cell specific genes and activation of the maternal program to support embryogenesis. In Drosophila, the timely repression of germ cell and early oogenesis genes is essential for this transition, yet the mechanisms that coordinate this process remain unclear. Here, we uncover an unexpected translation-chromatin axis, where transient Target of Rapamycin Complex 1 (TORC1)-driven translation triggers chromatin remodeling, ensuring irreversible oocyte fate commitment. Through a screen, we identified ribosome biogenesis regulators, including Zinc finger protein RP-8 (Zfrp8) and TORC1 components, as key mediators of gene silencing. We show that TORC1 activity increases during oocyte specification, and disrupting ribosome biogenesis, translation, or TORC1 function prevents proper heterochromatin formation, leading to epigenetic instability. Polysome-seq analysis of zfrp8-depleted ovaries revealed that Zfrp8 is required for the translation of Nucleoporin 44A (Nup44A), a key nuclear pore complex (NPC) component. Given the role of the NPC in chromatin organization, independent disruption of Nup44A results in defective silencing of the germ cell and early oogenesis genes. Our findings reveal a mechanism in which translation-driven NPC remodeling coordinates heterochromatin establishment, facilitating the germ cell-to-maternal transition and ensuring proper oocyte fate commitment.

overall_design: Sequencing of samples after performing CUT&RUN on Zfrp8 GKD, Nup44A GKD ovaries and WT in differentiated stages for the histone marks H3k9me3 and H3k27me3

first: Noor

middle: M

last: Kotb

first: Prashanth

last: Rangan

first: Gulay

last: Ulukaya

first: Dan

last: Hasson

first: Anupriya

last: Ramamoorthy

first: Lina

last: Park

first: Julia

last: Tang


title: TORC1-driven translation of Nucleoporin44A promotes chromatin remodeling and germ cell-to_x0002_4 maternal transition in Drosophila [polysome_zfrp8]

overall_design: Total mRNA control of nosGAL4 and >zfrp8 GKD ovaries and polysome isolation of control nosGAL4 and >zfrp8 GKD ovaries using sucrose gradients are performed in duplicates.


title: TORC1-driven translation of Nucleoporin44A promotes chromatin remodeling and germ cell-to_x0002_4 maternal transition in Drosophila [rna_zfrp8_mio]

overall_design: RNA-seq was performed on duplicates of each genotype: Raptor GKD, Mio GKD, Zfrp GKD ovaries as well as five WT ovaries.


title: c-Rel drives pancreatic cancer metastasis through Fibronectin-Integrin signaling-induced isolation stress resistance and EMT activation [Cut&Run]

city: Chicago

first: Harun

last: Ozturk

state: Illinois

address: 303. E. Superior, Lurie cancer center

institute: Northwestern University

summary: Pancreatic ductal adenocarcinoma remains one of the deadliest malignancies, with limited treatment options and a high recurrence rate. Recurrence happens often with metastasis, for which cancer cells must adapt to isolation stress to successfully colonize distant organs. While the fibronectin-integrin axis has been implicated in this adaptation, its regulatory mechanisms require further elaboration. Here, we identify c-Rel as an oncogenic driver in PDAC, promoting epithelial-to-mesenchymal transition (EMT) plasticity, extracellular matrix (ECM) remodeling, and resistance to isolation stress. Mechanistically, c-Rel directly regulates fibronectin (Fn1) and CD61 (itgb3) transcription, enhancing cellular plasticity and survival under anchorage-independent conditions. Fibronectin is not essential for EMT, but its absence significantly impairs metastatic colonization, highlighting a tumor-autonomous role for FN1 in isolation stress adaptation. These findings establish c-Rel as a key regulator of PDAC metastasis by controlling circulating tumor cell (CTC) niche and survival, suggesting that targeting the c-Rel–fibronectin–integrin axis could provide new therapeutic strategies to mitigate disease progression and recurrence.

overall_design: Cut&Run to determine c-Rel binding sites in A2929 cells treated with leptomycin B

first: Derya

last: Bakirdogen

first: Hana

last: Algul


title: c-Rel drives pancreatic cancer metastasis through Fibronectin-Integrin signaling-induced isolation stress resistance and EMT activation

overall_design: 2 Rel knock out mouse PDAC cell lines are stably transfected with either empty pAY15 or Rel-pAY15 constructs. For rescue experiment, Rel-pAY15 stable transfectants are treated with an auxin analog-5phIAA to induce c-Rel degradation. The cells are cultured to form spheroids under nonadherent conditions and used for RNAseq.


title: RBBP6 orchestrates diabetic endothelial dysfunction viadisrupting JUNB-centric chromatin topology

city: Guangzhou

first: Lei

last: Jiang

address: No.106 Zhongshan 2nd Road

institute: Guangdong Provincial People's Hospital

summary: Diabetes mellitus, a chronic metabolic disease affecting over 536.6 million people globally, is closely associated with vascular endothelial dysfunction, an early hallmark of diabetic cardiovascular complications. This dysfunction is characterized by impaired endothelial nitric oxide synthase (eNOS) activity, reduced nitric oxide (NO) production, and diminished angiogenic capacity, ultimately contributing to tissue ischemia and complications such as diabetic nephropathy, coronary artery disease, and peripheral arterial disease. These complications significantly increase morbidity and mortality in diabetic patients. Endothelial cells, which regulate vascular development and function, rely on the coordinated activity of cis-regulatory elements, including enhancers and promoters, which are influenced by the three-dimensional (3D) genome architecture and transcription factors (TFs) such as AP-1, ETS, and GATA families. However, the mechanisms by which these TFs mediate enhancer-promoter interactions and how these interactions are altered under diabetic conditions remain poorly understood.  To address this, we employed multi-omics profiling to map the 3D genome architecture in endothelial cells under hyperglycemic conditions in both diabetic mice and human vein endothelial cells. Our findings identify the disruption of JUNB-centric genome topology as a key feature of hyperglycemic endothelial cells. Moreover, we demonstrate that RBBP6 mediates ubiquitin-dependent degradation of JUNB, leading to changes in enhancer-promoter interactions. This study provides novel mechanistic insights into the chromatin 3D structural basis of endothelial dysfunction in diabetes and highlights the RBBP6-JUNB axis as a potential therapeutic target for preventing diabetic cardiovascular complications.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design:
ChIP-seq: ChIP-seq with JUNB or H3K27ac antibodies in three groups of mouse aortic endothelial cells and human umbilical vein endothelial cells. Mouse aortic endothelial cells were derived from C57BL6 db/db mice, BKS db/db mice, STZ diabetic model mice and their respective healthy control mice, and HUVECs were subjected to LG control, HG control, JUNB overexpression and RBBP6 knockdown experiments.
RNA-seq: RNA-seq analysis of BKS db/db mouse aortic endothelial cells, BKS control aortic endothelial cells, HUVECs cultured in LG or HG for 72h and their JUNB overexpression or RBBP6 knockdown derivatives.
ATAC-seq: ATAC-seq analysis of primary aortic endothelial cells from C57BL6 db/db mice, BKS db/db mice, STZ diabetic model mice and their respective healthy control mice.
HiChIP: HUVECs cultured under HG conditions for 72h were infected with adenovirus for 2h, and HUVECs cultured under LG and HG conditions for 72h were used as controls. Chromatin pull-down HiChIP analysis was performed with H3K27ac.

first: Xiaofei

last: Yang

first: Ruixin

first: Furong

last: Li

first: Wei

last: Huang

first: Liuyang

last: Cai

first: Yuliang

last: Feng


title: Transcriptomic Analysis of Candida boidinii Wild-Type Strain CICC#33029 and Its Engineered L-Lactic Acid-Producing Chassis Strain LA001-14 Cultured with Glucose or Methanol as the Sole Carbon Source

city: Tianjin

first: Feiyu

last: Fan

state: Tianjin

address: 32 West 7th Avenue, Tianjin Airport Economic Area

institute: Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences

summary: Single colonies of the wild-type Candida boidinii strain CICC#33029 and its derived L-lactic acid-producing chassis strain LA001-14 were inoculated into test tubes containing 3 mL of YPD medium and cultured overnight. Subsequently, 3 mL of the culture was transferred to a 250 mL Erlenmeyer flask containing 100 mL of liquid YPD medium and incubated at 30°C with shaking at 250 rpm for 12 hours. Cells were then harvested by centrifugation at 6,000 × g for 5 minutes and washed twice with phosphate-buffered saline (PBS). The cell pellets were resuspended in 100 mL of liquid YPD medium (supplemented with 2% w/v glucose) or YPM medium (supplemented with 2% w/v methanol) in 250 mL Erlenmeyer flasks. Following 6 hours of incubation at 30°C and 250 rpm, yeast cells were collected by centrifugation at 6,000 × g for 5 minutes and flash-frozen in liquid nitrogen. Total RNA was extracted from the samples using TRIzol reagent (Invitrogen, Grand Island, USA) and subjected to high-throughput RNA sequencing. The 150-nt paired-end RNA-seq libraries were commercially prepared by Novogene Biotechnology Co. Ltd (Tianjin, China) and sequenced on the Illumina NovaSeq 6000 platform (Illumina, San Diego, USA).

overall_design: RNA-seq profiles of the Candida boidinii wild-type strain CICC#33029 and its engineered L-lactic acid-producing chassis strain LA001-14, cultured under conditions of 2% (w/v) glucose or 2% (w/v) methanol as the sole carbon source, were obtained via deep sequencing using the Illumina NovaSeq 6000 platform.

first: Kaiyu

last: Ma

first: Qilong

last: Wen

first: Xueli

last: Zhang


title: TORC1-driven translation of Nucleoporin44A promotes chromatin remodeling and germ cell-to_x0002_4 maternal transition in Drosophila [rna_nup44a]

first: BiNGS

last: Core

overall_design: RNA-seq was performed on triplicates of each genotype: Control nosGAL4 and nosGAL4>Nup44a GKD ovaries

first: Noor

last: Kotb

first: Prashanth

last: Rangan

first: Gulay

last: Ulukaya

first: Dan

last: Hasson

first: Anupriya


title: Hepatic Transcriptomic Responses in Pregnant and Non-pregnant Rats Exposed to HFPO-DA: Analyses to Inform the Role of Maternal Effects on Neonatal Toxicity

city: Asheville

first: Melissa

middle: Marie

last: Heintz

address: 31 College Place, Suite B118

institute: ToxStrategies

summary: Like other per- and polyfluoroalkyl substances (PFAS), toxicity studies for short-chain HFPO-DA (ammonium, 2,3,3,3-tetrafluoro-2-(heptafluoropropoxy)-propanoate) indicate the liver is the primary target of toxicity in rodents. However, neonatal mortality and decreased birth weight were also reported in rats following oral exposure to HFPO-DA in utero. Exposure-related effects in neonatal rats including hypoglycemia, decreased liver glycogen, and perturbed hepatic gene expression of glucose metabolism and peroxisome proliferator activated receptor (PPAR) pathways also accompanied these findings. A putative rodent-specific adverse outcome pathway (AOP) network was recently developed using these endpoints and assessed for its applicability to PFAS. AOP 1 in this putative AOP network consists of PPARα activation as one of multiple initiating events, and placental insufficiency, neonatal hepatic glycogen deficit, and hypoglycemia as key events leading to neonatal mortality and lower birth weight. To further inform AOP 1 of this putative AOP network and investigate whether this altered carbohydrate metabolism liver phenotype observed in rat neonates also occurs in HFPO-DA-exposed pregnant and non-pregnant adult rats, transcriptomic analysis and glycogen staining were performed on female rat livers from a 15-d developmental or 90-d subchronic toxicity study. HFPO-DA-mediated changes in hepatic gene expression in female rats were consistent with PPAR signaling. Changes in hepatic glycogen content and glucose metabolism-related gene expression were independent of HFPO-DA exposure, indicating that the altered carbohydrate metabolism phenotype observed in neonatal livers does not occur in adult female rats, regardless of pregnancy status. Therefore, key events in this AOP for neonatal mortality and lower birthweight are likely to directly affect the perinatal rat and are not expected to affect the maternal rat liver. Findings from this study were consistent with previous mechanistic studies supporting the rodent-specific PPARα mode of action for HFPO-DA-mediated liver effects in rodents.

overall_design: Comparative gene expression profiling analysis of TempO-Seq data from FFPE liver tissue of pregnant (female) Sprague Dawley rats administered deionized water (vehicle control) or HFPO-DA in deionized water at 10, 100 or 1000 mg/kg by oral gavage once daily during gestation days (GD) 6 to 20, for a total of 15 days (developmental study), or non-pregnant female Sprague Dawley rats orally administered the same HFPO-DA doses daily for 90 days (subchronic study).

first: Chad

last: Thompson

first: Laurie

last: Haws

first: Grace

last: Chappell


title: Comprehensive Transcriptomic Analysis of DLD1 Colon Cancer Cells Overexpressing TRMT6 and TRMT61A Genes [RNA methylation]

city: GuangZhou

first: Mingzhe

address: Guangzhou, Guangdong

institute: Sun Yat-sen University

summary: This study investigates the transcriptomic changes in DLD1 colon cancer cells upon overexpression of TRMT6 and TRMT61A genes, with a focus on identifying downstream genes associated with tumor cell senescence. We performed RNA-seq, tRNA m1A-MeRIP-seq, and ribo-seq on DLD1 cells overexpressing TRMT6 and TRMT61A, as well as control cells, each with two biological replicates. The integrated analysis of these sequencing data aims to elucidate the molecular mechanisms underlying the regulation of tumor cell senescence by TRMT6 and TRMT61A.

overall_design: The experimental design includes three types of sequencing: RNA-seq, tRNA m1A-MeRIP-seq, and ribo-seq. DLD1 colon cancer cells were transduced with lentiviruses to overexpress TRMT6 and TRMT61A genes, respectively, and control cells were transduced with an empty vector. Each group (control, TRMT6 overexpression, and TRMT61A overexpression) was cultured in duplicate. RNA-seq was performed to profile the overall gene expression changes. tRNA m1A-MeRIP-seq was conducted to identify m1A modification sites in tRNA. Ribo-seq was used to assess ribosome occupancy on mRNA. The combined data from these three sequencing approaches were analyzed to identify key genes and pathways related to tumor cell senescence.

first: Tuoyang

last: Li

first: Huichuan

last: Yu

first: Jinxin

last: Lin

first: Meijin

first: Yanxin

last: Luo


title: Influenza A Virus Infection Induces Immune Dysregulation in the Placenta and Fetus Without Vertical Transmission in Nonhuman Primates

city: Seattle

first: Kristina

last: Adams Waldorf

state: Washington

address: 750 Republican Street, E Building, Room E360

department: OBGYN

institute: University of Washington

summary: Influenza A virus (IAV) infection during pregnancy is associated with stillbirth and preterm birth. We hypothesized that maternal IAV infection disrupts placental and fetal immune networks in ways that correlate with disease severity. Pregnant pigtail macaques were inoculated with IAV [A/California/07/2009 (H1N1)] and underwent delivery and fetal necropsy 5 days post-infection (N=11), with comparison to uninfected controls (N=16). Stillbirth occurred in 18% of infected pregnancies and in no controls. Although vertical transmission was not observed, low levels of viral RNA were detected in two placentas. Maternal infection was associated with induction of a type I interferon response in the placenta and altered innate and adaptive immune cell populations in the fetus. Maternal disease metrics (e.g., lung viral load, IFN-) were rarely associated with placental immune perturbations but consistently linked to fetal immune cell populations (CD4+ and CD8+ T cells, monocyte subsets). Collectively, these data indicate that maternal IAV disease severity is associated with dysregulated fetal immunity.

overall_design: Pregnant female pigtail macaques (Macaca nemestrina) were inoculated with 10e7.4 PFU IAV [A/California/07/2009 (H1N1)] using a combination of intranasal, intratracheal, intraocular, and intraoral routes and underwent delivery and fetal necropsy 5 days post-infection (N=11), with comparison to uninfected controls inoculated with sterile saline or media (N=16). Placenta samples were collected after Cesarean section.

first: Orlando

last: Cervantes

first: Sidney

last: Sun

first: Hazel

last: Huang

first: Briana

last: Del Rosario

first: Hong

last: Zhao

first: Amanda

first: Gygeria

last: Manuel

first: Roslyn

last: Van Abel

first: Jeff

last: Munson

first: John

last: Cornelius

first: Raj

middle: P

last: Kapur

first: Miranda

first: Edmunda

first: Hanning

first: Alexandra

last: Christodoulou

last: Berg

first: Britni

last: Curtis

first: Elizabeth

last: Miller

first: Shannon

last: Ruff

first: Brenna

last: Menz

first: Solomon

last: Wangari

first: Audrey

last: Germond

last: Baldessari

first: Chris

last: English

first: W McIntyre

last: Durning

first: Thomas

middle: B

last: Lewis

first: Megan

middle: N

last: Fredericks

first: Mara

last: Maughan

first: Austyn

last: Orvis

first: Michelle

last: Coleman

first: Lakshmi

last: Rajagopal

first: Deborah

middle: H

last: Fuller


title: Melatonin analog-agomelatine alleviates hypoxia-induced pulmonary arterial hypertension by activating mitophagy via the SIRT1/FoxO1/ULK1 signaling pathway

city: Nanjing

first: yuxian

last: Qian

address: baiziting no.42

institute: Nanjing medical university

summary: To explore the protective effect and molecular mechanism of agomelatine in hypoxic pulmonary arterial hypertension, we used Su5416 combined with hypoxia-induced rat PAH model and hypoxia-induced PASMC model. We verified agomelatine's therapeutic effect both in vivo and in vitro. Then, we performed RNA sequencing on control, SuHx, and SuHx+AGM groups. Pathway enrichment analysis revealed increased mitophagy, confirmed by WB, laser confocal microscopy, and electron microscopy. Finally, we found that agomelatine alleviates hypoxic pulmonary arterial hypertension by regulating mitophagy through the SIRT1/FOXO1/ULK1 pathway.

overall_design: To investigate the mechanisms by which agomelatine attenuates PAH, we conducted a comprehensive transcriptome analysis on PASMCs with normoxia, hypoxia and hypoxia plus agomelatine treatment.

first: Jiayi

last: Zhang

first: Ling

last: Chen

first: Lan

last: Wang


title: Comprehensive Transcriptomic Analysis of DLD1 Colon Cancer Cells Overexpressing TRMT6 and TRMT61A Genes [RNA-seq]

first: Mingzhe

last: Huang

first: Tuoyang

last: Li

first: Huichuan

last: Yu

first: Jinxin

last: Lin

first: Meijin

first: Yanxin

last: Luo


title: Tumor-secreted SPP1 activates CD44-Hedgehog signaling in hepatic stellate cells to promote fibrosis

city: Tokyo

first: Atsushi

last: Nara

state: Tokyo

address: 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8510, Japan

institute: Institute of Science Tokyo

summary: Background: Advanced hepatic fibrosis is a pivotal event leading to cirrhosis and hepatocellular carcinoma (HCC). We identified osteopontin (SPP1) as a fibrosis-associated gene through transcriptome and immunohistochemical analyses and investigated its role in intratumoral fibrosis and HCC development. Methods: Gene expression analysis was conducted using publicly available datasets to screen for fibrosis-related genes. Immunohistochemical staining was performed on HCC samples to assess SPP1 expression and its correlation with fibrosis and prognosis. The biological effects of SPP1 overexpression (SPP1-OE) in HCC cells were examined in xenograft mouse models. Co-culture assay with hepatic stellate cells (HSCs) and single-cell RNA sequencing (scRNA-seq) analysis were used to explore HCC-HSC interactions. Results: Transcriptomic and prognostic analyses revealed that SPP1 was significantly upregulated in fibrotic HCC tissues and associated with unfavorable outcomes. Immunohistochemical analysis confirmed a strong correlation between SPP1 expression and intratumoral fibrosis. In xenograft models, SPP1-OE HCC cells exhibited enhanced tumor growth and extensive fibrosis. Co-culture assay demonstrated that SPP1-OE cells stimulated HSCs, and gene set enrichment analysis and differential gene expression analysis elucidated activation of the Hedgehog signaling pathway and upregulation of GLI1 in HSCs. scRNA-seq analysis discovered SPP1-CD44 signaling transduction as a key mediator of HSC activation. Pharmacological inhibition of GLI1 with the SMO inhibitor vismodegib suppressed HSC activation in vitro and reduced fibrosis and tumor growth in vivo. Conclusions: SPP1 promotes intratumoral fibrosis and HCC progression through the SPP1-CD44-GLI1 axis, highlighting its potential as a prognostic biomarker and therapeutic target. Inhibition of Hedgehog signaling may provide a promising strategy to mitigate fibrosis and improve HCC patient outcomes.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for the HuH7 cells and LX2 cells subclones.

first: Shu

last: Shimada


title: Diverse and Location-Specific Roles of PlexinA2, PlexinA4, and NCAM in Developing Hippocampal Mossy Fibers

city: Ann Arbor

first: Michigan

middle: CDB

last: Bioinformatics

state: MI

address: 109 Zina Pitcher Pl

department: Cell & Developmental Biology

institute: University of Michigan

summary: Mossy fibers (MFs) originate from dentate granule cells and innervate area CA3 of the hippocampus. Upon entry of CA3, MFs partition into two prominent axon bundles, the suprapyramidal tract (SPT) and infrapyramidal tract (IPT) and form lamina specific synaptic contacts in the stratum lucidum (SL) and stratum oriens (SO), respectively. Here we employed new mouse lines to dissect the function of Sema6A and its receptors, PlexinA2 (PlxnA2) and PlxnA4, in developing MFs. In Sema6a-/- mice, MF partitioning into SPT and IPT bundles is incomplete and IPT axons in the SO are overextended, while the SPT correctly innervates the SL. Loss of neuronal Sema6a results in defective MF patterning and we show that this involves Sema6A reverse signaling. Plxna4 controls MF partitioning, SPT axon bundling and laminar targeting to the SL, as well as IPT length. Many of these defects are replicated in mice deficient for PlxnA4 GAP catalytic activity, underscoring the importance of this GAP domain. MFs are tightly fasciculated in Plxna2-/- mice and fail to separate into SPT and IPT bundles, and defects are significantly reduced in PlxnA2 GAP mutants, highlighting the involvement of GAP-independent signaling events. To further explore the molecular basis of aberrant axon fasciculation, we employed anti-PlxnA2 dependent proximity biotinylation and identified several PlxnA2-associated Ig-CAM family members. We observed a genetic interaction between Plxna2 and Ncam1, but not Plxna4 and Ncam1, for SPT and IPT formation and positioning in CA3.  Together, our studies provide insights into the multifaceted and overlapping, yet distinct, functions of PlxnA family members in orchestrating specific guidance decisions in developing MFs.

overall_design: P7 WT mouse hippocampi were micro-dissected and 10X Next GEM Chip G used for single-nucleus RNA sequencing.

first: Xiao-Feng

last: Zhao

first: Roman

last: Giger

first: Craig

last: Johnson


title: Enhanced Antitumoral Activity of the Academic CAR-T ARI0002h Against Normal and Low BCMA Expressing Myeloma Cells After Incorporating a Transmembrane CD28 Domain

city: Barcelona

first: David

middle: F.

last: Moreno

state: Barcelona

address: Villarroel 170

department: Hematology

country: Spain

institute: IDIBAPS - Hospital Clínic de Barcelona

summary: BCMA is the main target for CAR-T cells in multiple myeloma (MM), demonstrating promising outcomes. However, unlike what happens with CART19 in lymphoblastic leukemia and non-hodgkin lymphoma, a high proportion of patients will relapse after CART BCMA therapy due to insufficient antigen expression, low CAR-T cell persistence and/or T-cell exhaustion. CD28 transmembrane domains (CD28-TMD) in CAR receptors promote dimerization with CD28, forming CAR-CD28 heterodimers that enhance sensitivity to low-abundance antigens.  In other B cell malignancies, second-generation anti-CD19 4-1BB CARs with CD28-TMD have shown high efficacy and a favorable toxicity profile. We have developed a second-generation CD8α-TMD BCMA-4-1BBζ CAR-T product, ARI0002h (ARI2h; cesni-cel) for patients with relapsed/refractory MM. We hypothesized that replacing the TMD of ARI0002h with a CD28-TMD could increase efficacy and reduce tumor escape while maintaining a tolerable toxicity profile. We generated CAR-T cells using T-cells isolated from buffy coats and evaluated the efficacy and fitness of CAR-Ts at day 8-10 of expansion against several MM cell lines. In vitro analyses included cytotoxicity, proliferation, cytokine secretion, T-cell subset markers, activation and exhaustion profiling, metabolomic assays, and RNA-seq after multiple tumor challenges. In in vivo xenograft studies using NSG mice, with tumor cells expressing GFP-ffLuc, disease progression was monitored weekly via bioluminescence imaging.

overall_design: CAR-T cells were co-cultured with ARP-1 GFP+ target cells at a 1:2 E:T for 48 hours, after which the remaining CAR-T cells were rechallenged with fresh target cells under identical conditions. This process was repeated for up to four rounds. Post-rechallenge, CD8+ CAR-T cells were isolated using a magnetic bead-based CD8 Positive Selection Kit (EasySep).

first: Oriol

last: Cardús

first: Gerardo

last: Rodríguez-Lobato

middle: F


title: Upregulation of E-cadherin Promotes Spheroid Formation in Crizotinib-Resistant ALK-rearranged Lung Cancer

city: Yongin-si

first: Dawon

last: Hong

state: Gyeonggi-do

address: 152 Jukjeon-ro

country: South Korea

institute: Dankook University

summary: The anaplastic lymphoma kinase (ALK) gene fusion functions as a potent oncogene in lung cancer. The development of ALK tyrosine kinase inhibitors has improved patient outcomes. However, cancer progression commonly occurs even after treatment. Chemoresistance arises mostly by the acquisition of gatekeeper mutations in ALK as well as by off-target bypass mechanisms arising during chemotherapy. To investigate these bypass mechanisms, we studied crizotinib resistance in ALK-rearranged lung cancer by using 2D and 3D cell line models and longitudinal human biopsy samples. We found that crizotinib-resistant H2228 cells exhibited increased tumor cell clustering and spheroid formation, with elevated E-cadherin levels. Notably, depletion of E-cadherin profoundly disrupted spheroid formation and restored drug sensitivity. Transcriptomic profiling revealed the upregulation of tissue morphology-related genes; in particular, EpCAM was highly expressed in the protrusive regions of spheroids. Remarkably, longitudinal biopsy analysis also demonstrated enhanced expression of E-cadherin and EpCAM in patients with ALK-rearranged lung cancer during chemotherapy. We propose that off-target resistance is mediated by the upregulation of morphogenic genes, which drive cell clustering and spheroid formation during tumor evolution. To overcome chemoresistance, it is crucial to monitor tumor evolution during treatment. Tracking the collective migration of cells may present a novel therapeutic target.

overall_design: RNA-seq profiling of parental H2228 cells and duplicates of crizotinib-resistant cell lines (H2228-CR1 and H2228-CR2) in ALK-rearranged lung cancer.

first: Hyun Jung

last: Kwon

first: Joo Eon

last: Kim

first: Seokhyun

last: Yoon

first: Sunjoo

last: Jeong

first: Jin-Haeng

last: Chung


title: Herpes simplex virus 1 (HSV-1) R-loops are targets for APOBEC-mediated innate immunity [RNaseH_DRIP-seq]

city: Debrecen

first: Márton

last: Miskei

address: Egyetem tér1.

country: Hungary

institute: University of Debrecen

summary: APOBEC cytidine deaminase enzymes play a crucial role in antiviral innate immunity as they introduce mutations in viral genomes, restrict viral replication, and promote host defense against infections. Special three-stranded nucleic acid structures, called R-loops, have been considered as efficient targets for APOBEC-mediated mutagenesis because they present an ideal substrate, a persistent single-stranded DNA, for the enzyme. However, the relationship between R-loops and APOBEC enzyme activity has not been verified and remains to be elucidated. Here, we reveal a mechanistic link between R-loop formation, APOBEC enzyme binding, and APOBEC mutagenesis at viral R-loop targets during virus infection. In a cellular model involving human T lymphoblastoid leukemia cells infected with Herpes simplex virus 1 (HSV-1), we show that APOBEC3A and APOBEC3G binding sites and mutagenesis signatures occur mainly in viral R-loop structures. C-to-T mutagenesis primarily targets R-loops, which are specifically concentrated in HSV-1 genes that are essential for virus assembly and function. Among these genes, we identified mutation hotspots resulting in amino acid changes, likely leading to inactivation of HSV-1 protein functions. Collectively, our findings show that viral R-loops serve as potent targets for APOBEC-induced mutagenesis, representing an effective strategy to inactivate essential viral proteins and compromise viral activity.

overall_design: The specificity of S9.6 immunoselection was monitored by RNaseH treatment, which degrades the RNA strand of RNA-DNA hybrids

first: Marton

first: Lorant

last: Szekvolgyi