title: RNA-seq of phytoene desaturase knockout carotenoid-deficient microalgal mutants generated by cas9-ribonucleoprotein complexes

city: Huelve

first: Ana

last: Molina Márquez

state: Huelve

address: Avd. Fuerzas Armadas, S/N

department: Chemistry

institute: University of Huelva

summary: Phytoene desaturase (PDS; EC 1.3.5.5) is a key enzyme of the carotenoid biosynthetic pathway, catalyzing the desaturation of phytoene, precursor of all carotenoids. In this study, several PDS knockout (PDS-KO) transformants of the chlorophyte microalga Chlamydomonas reinhardtii were generated using a reverse genetics strategy. Two single guide RNAs (sgRNA) were designed to target the first exon of the PDS gene, and pre-assembled Cas9 ribonucleoprotein (RNPs) complexes were delivered into microalgal nuclei by electroporation. Multiple white PDS-KO transformants were successfully obtained by this approach, and three independent transformant lines were subsequently characterized. By integrating ultrastructural, pigment and transcriptomic analyses of dark-grown cells of several PDS-KO carotenoid-deficient mutants in comparison with the parental strain, it was demonstrated that carotenoids are indispensable components of multiple cellular architectures. Chromatographic analysis confirmed that the only carotenoid accumulated in these transformants was phytoene, which lacks the critical structural and photoprotective functions of its colored derivatives. Transmission Electron Microscopy (TEM) observations revealed profound ultrastructure alterations, including poorly developed chloroplasts and effects on other cellular structures that were either absent or severely disorganized. Consistently, clustering differentially expressed genes into functional groups revealed downregulation of pathways associated with photosynthesis, chlorophyll and carotenoid biosynthesis, ribosome biogenesis, and vesicle and membrane trafficking in the PDS-KO lines. Conversely, upregulation of regulatory and retrotransposon-inducing genes was observed. These findings underscore the central metabolic role of colored carotenoids in plant cells, highlighting their essential contribution to cellular homeostasis and photosynthetic competence

overall_design: RNA-seq profiling of wild-type Chlamydomonas reinhardtii 3403 cells and their PDS Knockout derivatives, E10, A2 y D2, after four weeks of growth in darkness at 25 ºC

first: Ana M.

last: Molina-Márquez

first: Simon

last: Kelterborn

first: Peter

last: Hegemann

first: Miguel

last: Pérez-Rodrígez

first: Javier

last: Vigara

first: Rosa

last: León


title: Syntaxin 4 Enrichment in β-Cells Prevents Conversion to Autoimmune Diabetes in Non-Obese Diabetic (NOD) Mice

status: Public on Feb 12 2026

city: Duarte

first: Debbie

middle: C.

last: Thurmond

state: California

address: 1500 E Duarte Rd

country: USA

institute: City of Hope

type: Expression profiling by high throughput sequencing

summary: Syntaxin 4 (STX4), a plasma membrane–localized SNARE protein, regulates human islet β-cell insulin secretion and preservation of β-cell mass. We found that human type 1 diabetes (T1D) and NOD mouse islets show reduced β-cell STX4 expression, consistent with decreased STX4 expression, as a potential driver of T1D phenotypes. To test this hypothesis, we generated inducible β-cell–specific STX4-expressing NOD mice (NOD-iβSTX4). Of NOD-iβSTX4 mice, 73% had sustained normoglycemia vs. <20% of control NOD (NOD-Ctrl) mice by 25 weeks of age. At 12 weeks of age, before diabetes conversion, NOD-iβSTX4 mice demonstrated superior whole-body glucose tolerance and β-cell glucose responsiveness than NOD-Ctrl mice. Higher β-cell mass and reduced β-cell apoptosis were also detected in NOD-iβSTX4 pancreata compared with pancreata of NOD-Ctrl mice. Single-cell RNA sequencing revealed that islets from NOD-iβSTX4 had markedly reduced interferon-γ signaling and tumor necrosis factor-α signaling via nuclear factor-κB in islet β-cells, including reduced expression of the chemokine CCL5; CD4+ regulatory T cells were also enriched in NOD-iβSTX4 islets. These results provide a deeper mechanistic understanding of STX4 function in β-cell protection and warrant further investigation of STX4 enrichment as a strategy to reverse or prevent T1D in humans or protect β-cell grafts.

overall_design: Islets were isolated from female NOD-Ctrl or NOD-iβSTX4 transgenic mice and dispersed into single cells with TrypLE (Invitrogen, Carlsbad, CA). Cell numbers and viability were measured using a TC20 Automated Cell Counter (Bio-Rad); only samples showing at least 70% viability were used. Cells were then loaded onto a chromium controller (10x Genomics, Pleasanton, CA) to generate single-cell Gel Bead-In Emulsions (GEMs) captured in droplets at a targeted cell recovery of 3,000 cells. GEM reverse transcriptions were performed in a Veriti 96-Well Thermal Cycler (Thermo Fisher Scientific). The Chromium Single Cell 3' Reagent Kit V2 (10x Genomics) was used to process samples into single-cell RNA sequencing (scRNA-seq) libraries according to the manufacturer’s protocol. The libraries were sequenced with the paired-end setting of 28 cycles of read 1, 101 cycles of read 2, and 8 cycles of the index read on the Illumina NovaSeq 6000 platform at the Translational Genomics Research Institute (TGen) (Phoenix, AZ).

first: Eunjin

last: Oh

first: Erika

middle: M

last: McCown

first: Miwon

last: Ahn

first: Pablo

middle: A

last: Garcia

first: Sergio

last: Branciamore

first: Shanshan

last: Tang

first: De-Fu

last: Zeng

first: Bart

middle: O

last: Roep

middle: C


title: CUT&Tag profiling of Irx3 binding sites in AC16 cardiomyocytes supporting studies on cardiac regulatory mechanisms

city: Changchun

first: BIng

last: Li

address: 130021, No. 1, Xinmin Street, Chaoyang District, Changchun, Jilin Province

country: China

institute: First Hospital of Jilin University

type: Genome binding/occupancy profiling by high throughput sequencing

summary: This study performed CUT&Tag sequencing to identify genome-wide binding sites of Irx3 in AC16 cardiomyocytes, aiming to reveal the regulatory role of Irx3 in cardiac biology.

overall_design: This dataset presents CUT&Tag sequencing data targeting the transcription factor Irx3 in human AC16 cardiomyocytes (encompassing both IgG control and Irx3-overexpressing groups), which profiles the genome-wide binding landscape of Irx3.

first: Li

last: Bing


title: Physical Activity attenuates Sperm DNA Damage induced by Sedentary Behavior via Testicular Thermoregulation

city: Wuhan

first: Yu

last: Fang

state: Hubei

address: Wuhan University of Science and Technology

summary: The contribution of male factors to infertility has been increasing progressively, with growing attention to non-genetic factors (lifestyle, environment). As a prevalent suboptimal lifestyle, the impact of sedentary behavior on male reproductive dysfunction remains to be fully elucidated. In this study, 6-week-old male C57BL/6 mice were randomly assigned to control (CT), sedentary (ST), and physical activity compensation (CS) groups for 10-week exposure. No observable differences in growth or behavior indicators were identified among groups. The ST group exhibited a pathological cascade: gonadal histopathological changes, sex hormone secretion disordered, and spermatogenic impairment, which were significantly alleviated in CS. Mechanistically, testicular RNA-seq revealed sedentary behavior disrupted thermoregulation, with DNA damage. Further analysis confirmed significant upregulation of Gpx4 (DNA damage marker) and Hsp70 (core temperature-sensing gene) in ST, along with increased testicular temperature and compromised sperm motility. Collectively, sedentary behavior may induce sperm DNA damage by disrupting testicular temperature homeostasis. Our findings provide a novel perspective for elucidating the mechanism underlying male infertility induced by sedentary behavior.

overall_design: In this study, 6-week-old male C57BL/6 mice were randomly assigned to control (CT), sedentary (ST), and physical activity compensation (CS) groups for 10-week exposure.Total RNA was extracted from mouse testicular tissues (n = 3 biological replicates per group) using Trizol reagent (Invitrogen, Carlsbad, CA, USA).

first: Xi

last: Wen

first: Xingcheng

last: Yang

first: Jiahui

last: Wang

first: Menghan

last: Chen

first: Linxiang

last: Han

first: Ying

last: Zhang


title: ALDH18A1 Promotes Gastric Cancer Progression and Regulates the PI3K-Akt Signaling Pathway

city: Urumqi City

first: abulajiang·abudoukelimu

middle: abudoukelimu

last: abulajiang

address: 新疆医科大学第三临床医学院

institute: The Third Clinical Medical College of Xinjiang Medical University

summary: To elucidate the mechanism of ALDH18A1-driven gastric carcinogenesis and identify its downstream transcriptional targets, we conducted genome-wide RNA sequencing in HGC-27 cells knockdowning ALDH18A1, with comparison to negative control (si-NC) group.

overall_design: RNA-seq of ALDH18A1 knockdown (si ALDH18A1) to study the gene expression profile pattern of HGC-27 cells in vitro.


title: The VgrG2 effector of Acinetobacter baumannii mediates immune evasion by repressing Csu pilus assembly and triggering phagocyte methuosis

city: Hangzhou

first: xu

last: wenzeng

address: Department of Respiratory Medicine, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou, 310016, China.

institute: Zhejiang University

summary: Acinetobacter baumannii is a formidable nosocomial pathogen whose multidrug resistance and immune evasion capabilities present a major therapeutic challenge. This study identifies the type VI secretion system (T6SS) effector VgrG2 as a critical virulence regulator in A. baumannii. VgrG2 employs a two-pronged strategy to undermine host innate immunity. First, it transcriptionally represses the major adhesin Csu pilus, reducing bacterial recognition, phagocytic uptake, and neutrophils recruitment. Second, VgrG2 directly targets and hyperactivates the small GTPase Rac1 within phagocytes. This leads to actin cytoskeletal disarray, which drives excessive macropinocytosis, resulting in compromised phagocytosis and methuosis—a non apoptotic, vacuole dependent death specific to phagocytes. Thus, VgrG2 selectively eliminates key immune sentinels and alters pulmonary inflammation, promoting bacterial persistence and dissemination. These findings extend the role of T6SS effectors beyond interbacterial competition to include sophisticated eukaryotic directed attacks, and highlight the VgrG2 Rac1 axis as a promising therapeutic target.

overall_design: AB5075 in vivo VS in vitro; AB 5075 wild type VS vgrG2-ko

first: Wenzeng

last: Xu

first: Hua

last: Zhou


title: SLC33A1 exports oxidized glutathione to maintain endoplasmic reticulum redox homeostasis [ERIP_RNASeq]

city: New York

last: Liu

state: NY

address: 430 East 63rd Street,

institute: The Rockefeller University

summary: The endoplasmic reticulum (ER) is the primary site of synthesis of secretory and membrane proteins, suggesting that specific mRNAs may be enriched at the ER surface. To characterize ER-associated transcripts, ER fractions were isolated using ER immunoprecipitation (ER-IP) and RNA was extracted from ER-bound material and whole-cell lysates. RNA sequencing was performed to compare transcript composition between ER-associated and total cellular RNA. ER-IP RNA sequencing revealed enrichment of transcripts encoding proteins localized to the plasma membrane, ER, Golgi apparatus, lysosome, and extracellular space. This dataset provides transcriptomic profiling of ER-associated RNA and corresponding whole-cell controls.

overall_design: ER-associated RNA was isolated by ER immunoprecipitation in the presence of RNase inhibitor. RNA was extracted from ER-IP material and from whole-cell lysates collected prior to ER purification. Libraries were prepared and sequenced to compare ER-associated transcripts with total cellular RNA. Biological replicates were analyzed for each condition. Sequencing reads were pseudoaligned using Salmon and differential enrichment was analyzed using limma-voom.

first: Alon

last: Millet

first: Kivanc

last: Birsoy


title: SLC33A1 exports oxidized glutathione to maintain endoplasmic reticulum redox homeostasis [KPK_Slc33a1_KO_TUNC]

first: Shanshan

summary: Endoplasmic reticulum stress and redox homeostasis influence protein folding and unfolded protein response signaling. To examine transcriptional responses associated with Slc33a1 loss and complementation, we performed RNA sequencing of KPK cells with Slc33a1 knockout and cells complemented with either empty vector or cDNA constructs. Cells were treated with either DMSO or tunicamycin to induce ER stress, and total RNA was collected for sequencing. This dataset provides transcriptome profiles that enable comparison of gene expression changes across genotypes and treatment conditions.

overall_design: KPK cells with Slc33a1 knockout and complemented lines were cultured and treated with either DMSO or tunicamycin. Total RNA was extracted and libraries were prepared for RNA sequencing. Biological replicates were generated for each condition to compare transcriptional responses to ER stress in different genetic backgrounds.


title: Single-cell multiomic profiling of lung immune cells identifies novel asthma risk genes and cell-type specific functions

city: Chicago

first: Nathan

last: Schoettler

state: IL

address: 5841 S Maryland Ave

institute: the University of Chicago

summary: Genetic variants associated with asthma are mostly in non-coding regions, so it is challenging to identify their target genes and the cellular contexts. Functional profiling of blood immune cell types has helped link asthma risk variants to functions. However, lung immune cells differ from those circulating in blood, and high-resolution functional genomic data of lung immune cells are lacking. Here we profiled single-cell multi-omics (RNA-seq and ATAC-seq) on lymphocytes of lung and spleen tissues. Cross-tissue comparison identified distinct transcriptomes for each immune cell type, but subtle differences in chromatin accessibility. We then assessed open chromatin regions of lung vs. blood, using a public dataset, for their enrichment of asthma risk. Strikingly, lung T cells show unique contributions to heritability of adult-onset (AOA) and childhood-onset asthma (COA), beyond blood T cells. We further identified 43 cis-regulatory elements (CREs) likely contributing to asthma risk. With enhancer-gene maps built from our single-cell data, we identified target genes for these CREs, including CCR4 and LRRC32 that show cell-type specific regulatory activities. These findings demonstrate the utility of single-cell multiomics to identify asthma risk genes and understand their cell-type specific functions.

overall_design: Single-nuclei were extracted from lung and spleen tissues of 9 deceased donors. Joint snRNA-seq and snATAC-seq libraries were prepared using the 10x Genomics platform.

first: Jing

last: Gu

first: Donna

last: Decker

first: Xiaoyuan

last: Zhong

first: Anne

middle: I

last: Sperling

first: Carole

last: Ober

first: Marcelo

last: Nóbrega

first: Xin

last: He


title: Integrating Single-Cell Transcriptomics with Phosphoproteomics Dissects the EGFR-Dependent Network Governing Cortical Gliogenesis

city: Shanghai

first: Guoping

address: 138 Yi Xue Yuan Road

institute: Fudan University

summary: Deciphering gliogenesis is fundamental to understanding glial biology. Epidermal growth factor receptor (EGFR) plays an essential role in this process, with its expression initiating at the onset of gliogenesis and peaking perinatally, coinciding with key stages of glial development. Conditional ablation of EGFR in the glial lineage resulted in a pronounced reduction of cortical glia, particularly astrocytes. To systematically map the EGFR‑dependent molecular landscape during gliogenesis, we generated an integrated multi‑omics atlas through lineage‑restricted deletion of Egfr using hGFAP‑Cre. This high‑resolution resource delineates the transcriptional and phosphoproteomic reprogramming of radial glial lineages upon EGFR loss. We profiled 24,738 cells at embryonic day 18 (E18) and 18,343 cells at postnatal day 2 (P2) from control and Egfr‑CKO cortices by single‑cell RNA sequencing. In parallel, phosphoproteomic analysis quantified 6,409 high‑confidence phosphorylation sites, 6,183 unique phosphopeptides, and 2,510 phosphorylated proteins. Together, these datasets establish a comprehensive framework for dissecting EGFR‑dependent regulatory networks and signaling crosstalk, providing mechanistic insight into glial fate specification and potential developmental origins of gliopathies.

overall_design: Egfr-CKO embryos were subjected to lateral ventricle injection of Cell Trace Yellow at embryonic day 16 (E16) . The cerebral cortices were harvested at E18, corresponding to the injection timepoints. Fluorescent dye-labeled cells were isolated via fluorescence-activated cell sorting using a BD FACSAriaII flow cytometer.

last: Jiang

first: Zhejun


title: Fragmentation through cell-cell adhesion maintains senescent cell viability but promotes debris deposition

city: Illkirch

first: Matej

last: Durik

state: Alsace

address: 1 rue Laurent Fries

department: Development and stem cells

country: France

institute: IGBMC

summary: We identified that senescent cells dispose of large fragments of themselves through cell-to-cell adhesion, which we term senescent-cell adhesion fragments (SCAFs). Dynamic analyses show that SCAFs ultimately rupture, releasing a complex proteome including damage-associated molecular patterns (DAMPs) and proteins linked to neurodegenerative disease. Functionally, SCAFs activate wound-healing and cancer-related programs, promoting migration and invasion.

overall_design: The aim is to identify the genes and pathways activated by Senescent-Cell Adhesion Fragments (SCAFs) in M0 macrophages. THP-1 derived macrophages were treated either by inert beads (CTR) or SCAFs (SCAFs) for 20 hrs and processed for RNA sequencing.

first: William

last: Keyes


title: Hap complex regulates the development of Penicillium oxalicum fungi and the expression of cellulolytic genes

city: Qingdao 266237

first: Kexuan

last: Ma

state: China

address: No. 72 Binhai Road

department: State Key Laboratory of Microbial Technology

institute: Shandong University

summary: The goals of this study are to compare different gene expressions for Penicillium oxalicum wild type strain (WT) and Hap3/5 deletion strain (Dhap3/5) under conditions of  2% glucose and 1% bran + 1% microcrystalline cellulose as carbon source. The results of correlation analysis between each sample showed that the gene expression level of Dhap3/5 was significantly different from that of the WT strain. The deletion of hap3/5 significantly affect the expression levels of sporulation process genes and the expression of cellulase-encoding genes.

overall_design: Examination of differential gene expressions by digital gene expression tag profiling in Penicillium oxalicum wild type strain and hap3/5 deletion mutant strains in 24h in modifeid Mendal’s medium with 2% glucose or 1% bran + 1% microcrystalline cellulose as carbon sources. Three replicates were included for each samples.

first: Yuqi

last: Qin


title: Sympathetic nerve–fibroblast crosstalk drives nerve injury, fibroblast activation, and matrix remodeling in pancreatic cancer

city: Portland

first: Sebnem

middle: Ece

last: Eksi

state: Oregon

address: 2730 S Moody Ave.

institute: Oregon Health & Science University

summary: Pancreatic cancer is a highly innervated gastrointestinal disease in which sympathetic nerves play a critical role in modulating tumor growth and the tumor microenvironment (TME). While recent studies suggest that sympathetic nerves influence various TME components, including lymphoid and myeloid immune cells, their interactions with cancer-associated fibroblasts (CAFs) remain poorly understood. CAFs are a hallmark of pancreatic tumors and are known to upregulate axon guidance and neuroactive cues, suggesting a potential feedback loop with tumor-innervating nerves. Here, we investigated the bidirectional crosstalk between sympathetic nerves and CAFs in human and mouse pancreatic tumors. Using a chemo-genetic ablation model, we selectively eliminated pancreatic sympathetic nerves and found that denervation significantly reduced tumor size in female mice. To further dissect this interaction, we established co-culture systems with immortalized pancreatic fibroblasts and primary sympathetic neuron explants, identifying key transcriptional changes driven by CAF-sympathetic nerve signaling. Our findings demonstrate that sympathetic signaling enhances CAF activation and extracellular matrix remodeling, while activated CAFs, in turn, induce transcriptional programs in sympathetic neurons associated with nerve injury response. These results establish CAFs as central mediators of the tumor-supportive role of sympathetic nerves, offering new insights into the neural regulation of pancreatic cancer progression.

overall_design: In this experiment, we indirectly cocultured p1-p4 murine pup superior cervical ganglia (SCG) with pancreatic fibroblasts, immortalized pancreatic stellate cells (PSCs), and control fibroblasts, mouse embryonic fibroblasts (MEFs, [NIH/3T3 CRL-1658]). Between 8 and 13 explants were plated in a coated 6-well plate and allowed to grow with 10.0 X 103 PSC or MEF cell lines on cell culture inserts (Falcon; 353090) for about 70 hours. RNA was isolated using the RNeasy Kit (Qiagen; 74104) for RNA purification. The samples: Nerve (SCG alone), Nerve_PSC (SCG cocultured with PSCs), Nerve_MEF (SCG cocultured with MEFs), PSC (PSCs alone), and PSC+N (PSCs cocultured with SCGs).

first: Ariana

middle: L

last: Sattler

first: Kevin

last: MacPherson-Hawthorne

middle: E


title: Liver RNA-seq from hepatocyte Tet1-overexpression MASH mice

city: Nanjing

first: junjie

last: yu

address: Address: #639 Longmian Avenue, Jiangning District, Nanjing, China, 211198

institute: China Pharmaceutical University

summary: To determine the mechanism underlying hepatocyte Tet1 regulated fibrogenesis in MASH liver, we performed RNA sequencing of liver samples from hepatocyte-specific Tet1-overexpression MASH mice. Differentially expressed genes were analyzed and potential targets were validated.

overall_design: 6 weeks old C57BL/6J wild-type (WT) mice were hydrodynamically injected with plive-Con or plive-Tet1 plasmid and fed with HFD-CDAA for 4 weeks. Mice were sacrificed after short fast.

first: Fengyi

last: Wang

first: Tianshu

last: Gao

first: Junjie

last: Yu


title: Erastin-Induced Ferroptosis Enhances Natural Killer Cell Anti-Tumor Activity  and Offers Therapeutic Potential in Neuroblastoma

city: Göteborg

first: Ganesh

last: Umapathy

address: Medicinaregatan 9 a-b

department: Sahlgrenska University Hospital

country: Sweden

institute: University of Gothenburg

summary: High-risk neuroblastoma (NB) often relapses and becomes refractory to treatment, making it a significant contributor to childhood cancer mortality despite its relative rarity. Addressing the challenges posed by NB’s resistance to conventional apoptosis-inducing therapies (e.g., chemotherapy and radiation) has become a pressing concern in pediatric oncology research. In recent years, the growing comprehension of  alternative cell death modalities distinct from apoptosis has revealed a promising avenue in the endeavor to combat treatment-resistant cancers. One such mechanism, ferroptosis, has attracted increasing attention for its potential role in combating therapy-resistant cancer cells. High-risk NB typically exhibits an immune-excluded phenotype, characterized by minimal immune cell infiltration. Despite this characteristic, the mechanisms underlying immune exclusion and impaired immune activation in NB remain unclear. Here, we demonstrate that the induction of ferroptosis  using Erastin, a cysteine-glutamate antiporter inhibitor, reduces NB cell proliferation and foci formation. Furthermore, our transcriptomics analysis revealed that treatment  with Erastin in NB cells led to increased expression of ULBP1, a ligand to the activating natural killer (NK) cell receptor, NKG2D. Upon ferroptosis induction, the transcription factor ATF4 was found to drive ULBP1 expression in NB cells. Consistent with this, pre-treatment with Erastin sensitized NB cells to NK cell cytotoxicity in co-culture experiments. These results suggest the NK cell’s cytotoxic function can be enhanced with Erastin-mediated ferroptosis, which may be beneficial for NB patients.

overall_design: Human neuroblastoma cell lines KELLY (Kelly) and SK-N-AS (AS) were treated with Erastin (Era) for 24 hours or vehicle control. Each condition was performed in biological triplicate. Bulk RNA-seq was performed to characterize transcriptional changes associated with ferroptosis induction.

first: Umapathy

last: Ganesh

first: Anthonydhason

last: Vimala


title: The Effector Differentiation of TCRαβ+CD8αα+ Intraepithelial Lymphocytes Is Reciprocally Regulated by BCL6 and BLIMP1

city: Beijing

first: Qi

last: Xing

address: 30 Shuangqing Road

institute: Tsinghua University

summary: CD8αα+TCRαβ+ intraepithelial lymphocytes (IELs) play crucial roles in maintaining intestinal homeostasis and host protection. However, the functional regulation of these cells remains unclear. Here, we have discovered and characterized two distinct developmental stages within intestinal CD8αα+ αβ IELs: a stem-like, defined by BCL6, TCF1 and CD160 expression, and an effector-like, with Granzyme B expression and Prdm1 transcription, by sorting and transcriptional sequencing BCL6+CD160+ CD8αα+ αβ IELs and BCL6-CD160- CD8αα+ αβ IELs. Comparing transcriptional profiles of total CD8αα+ αβ IELs from BCL6-deficient, BLIMP1-deficient and TCF1-deficient mice compared with that from their WT littermate control mice, we found a critical antagonistic function between BCL6/TCF1 and BLIMP1 in governing the fate decisions of CD8αα+ αβ IEL subsets.

overall_design: RFP+CD160+ CD8αα+ αβ IELs and RFP-CD160- CD8αα+ αβ IELs were isolated from small intestine of steady state Bcl6RFP mice for bulk RNA-seq. Total CD8αα+ αβ IELs were isolated from small intestine of steady state Bcl6fl/flCd8cre, Prdm1fl/flCd8cre and Tcf7fl/flCd8cre mice and their littermate WT control (Bcl6fl/fl, Prdm1fl/fl and Tcf7fl/fl) mice for bulk RNA-seq.

first: Shiyuan

last: Xie

first: Xiaohong

last: Zhao

first: Chen

last: Dong


title: Expression data from Heatstroke patients

city: Thuwal

first: Maria

middle: de los Angeles

last: Gomez

address: 4700 King Abdullah University of Science and Technology, Thuwal 23955-6900, Kingdom of Saudi Arabia

department: Bioengineering

country: Saudi Arabia

institute: KAUST

summary: This study examined transcriptomic responses to heat exposure in peripheral blood mononuclear cells from 19 patients with heat stroke and 19 controls.

overall_design: We enrolled 19 non-heat stroke (NHS) pilgrims, friends, or relatives of heat stroke patients living in the same environmental heat as a control group. We obtained blood samples from control subjects upon enrolment and heat stroke patients on arrival to the cooling unit, before cooling (T0) and after cooling (T1).

first: Abderrezak

last: Bouchama

first: Saeed

last: Al Mahri


title: Transcriptomic profiling of human hepatocellular carcinoma biopsy samples using bulk RNA-seq

city: Suita

first: Shuhei

last: Yamamoto

state: Osaka

address: 2-2 Yamadaoka, Suita, Osaka

country: Japan

institute: Osaka University, Graduate School of Medicine

summary: Bulk RNA sequencing was performed on human hepatocellular carcinoma (HCC) tumor biopsy specimens to characterize transcriptomic features of tumor tissues. Tumor samples were obtained from patients receiving systemic therapy, and gene expression profiles were generated for comparative analysis. This dataset provides a transcriptomic resource for studies of molecular features in human HCC.

overall_design: Tumor biopsy samples were collected from patients with hepatocellular carcinoma prior to and/or following systemic therapy. Total RNA was extracted from tumor tissues and subjected to bulk RNA sequencing to generate gene-level expression profiles for comparative transcriptomic analysis.

first: Takahiro

last: Kodama

first: Akifumi

last: Kuwano

first: Kenta

last: Motomura


title: scRNA-seq of mouse liver tumors: early treatment, late treatment, and control

summary: Single-cell RNA sequencing was performed on mouse liver tumors to investigate the immune microenvironment under early and late treatment conditions. This dataset provides a resource to study treatment-dependent changes in immune cell composition.

overall_design: Single-cell RNA sequencing was performed on mouse liver tumors to investigate the immune microenvironment under different treatment conditions. Nine samples were analyzed, including three from early treatment (ST), three from late treatment (LT), and three vehicle-treated controls (Control). The dataset enables comparison of immune cell composition and gene expression profiles across treatment groups.


title: RNA sequencing analysis of cells infected with chimeric EV-A71 strains in which the 3C protease region was swapped with other strains of EV-A71

first: Daisuke

last: Motooka

address: 3-1, Yamadaoka

institute: NGS core facility, Research Institute for Microbial Diseases, Osaka University

summary: The EV-A71 genome has accumulated mutations since the virus was first detected in 1969. To characterize the polymorphism of the EV-A71 3C protease, we performed RNA sequencing using chimeric EV-A71 strains carrying the 3C protease from a clinical strain. The chimeric EV-A71 was generated using a reverse genetic system and inoculated into RD-S cells.  Gene expression profiling analysis of RD-S cells infected with the chimeric EV-A71 strains was performed.

overall_design: The 3C protease coding sequence in the EV-A71 genome (strain SK-EV-006) was swapped to the sequence of the clinical strain of 3C protease. The 3C protease sequence from the clinical strains 1095/Japan 97 or 7F-AUS-6-99 was used. The RD-S cells were infected with the chimeric EV-A71 at an MOI of 1.0, extracted total RNA at 16 h post-infection, and analyzed by RNA sequencing.

first: Junki

last: Hirano

first: Yoshiharu

last: Matsuura


title: Targeting the Non-Homologous End-Joining Pathway Sensitizes MDM2-Amplified Liposarcoma to Doxorubicin-Induced Senescence, p53-Mediated Senescence.

city: Amsterdam

first: Daniel

last: Peeper

state: Outside the US or Canada

address: Plesmalaan 121

department: Molecular Oncology and Immunology

country: Netherlands

institute: Netherlands Cancer Institute

summary:
Introduction Dedifferentiated liposarcoma (DDLPS) is a rare cancer defined by amplification of MDM2 and CDK4. Conventional chemotherapy (Doxorubicin) and targeted inhibition of MDM2 and CDK4 show sporadic responses, but most tumors display primary resistance. We used an unbiased approach to identify therapeutic strategies sensitizing to these DDLPS therapies.
Methods Three parallel genome-wide CRISPR-Cas9 knockout screens were conducted in DDLPS cells to sensitize to palbociclib (CDK4 inhibitor), nutlin-3a (MDM2 inhibitor) or Doxorubicin. Top screen hits were validated and characterized in both in vitro and in vivo models, while clinical data were used to corroborate molecular findings.
Results We uncovered pathways related to G1/S transition (CDK2, CKS1B, E2F3 and CCNE1) and Non-Homologous End-Joining (NHEJ; TDP2, PRKDC and XRCC4), inactivation of which sensitized to palbociclib and Doxorubicin, respectively. Following validation of both pathways, we focused on mechanistic characterization of Doxorubicin sensitization by genetic perturbation of TDP2 or pharmacological inhibition of DNA-PKcs using peposertib. Synergistic cell cycle arrest and senescence were induced by prolonged administration of low-dose doxorubicin. Senescent cells were triggered to undergo apoptosis by subsequent senolytic treatment with Bcl2 inhibitor navitoclax. Despite the amplification of MDM2, senescence proved dependent on p53. Consistent with this, TCGA and DepMap data suggest p53 activity in DDLPS.
Conclusion These findings provide a rationale for targeting the NHEJ pathway to enhance the efficacy of low-dose Doxorubicin in DDLPS, highlighting a potential therapeutic strategy exploiting p53-dependent cell cycle arrest and senescence. Furthermore, we provide, to our knowledge, the first evidence of maintained baseline p53 activity in MDM2-amplified DDLPS.

overall_design: DDLPS cell lines (LPS853, T778) containing sgTDP2 or sgCtrl were treated with Doxorubicin (4nM, 24h) +/- peposertib (0.5uM)

first: Thijs

last: Jalving

first: Carine

last: Dufau

first: Juan

last: Simon Nieto

first: Daniela

last: D'empaire Altimari

first: Susan

last: Van Hal-van Veen

first: Elisabeth

last: Jin

first: Nils

last: Visser

first: Hester

last: van Boven

first: Winette

last: van der Graaf

first: Winan

last: van Houdt


title: The Liver Regulates Ectopic Calcification in Pseudoxanthoma Elasticum

city: Los Angeles

first: Feiyang

last: Ma

state: CA

address: 610, Charles Young Dr East, TLSB 3000C

department: Molecular Biology Institute

institute: UCLA

summary: Pseudoxanthoma Elasticum (PXE) is a rare disease caused by loss of function of the gene Abcc6 and characterized by ectopic calcification of multiple tissues including the heart and vasculature. The physiological reasons underlying ectopic calcification in PXE remain unclear. We created a robust model of ectopic cardiac calcification where animals globally deficient in the gene Abcc6 developed extensive mineralization of the heart within 3 days of cardiac injury. To obtain insight into the underlying mechanisms of ectopic cardiac calcification in PXE, we generated animals with cardiomyocyte specific deletion of Abcc6 but did not observe any calcification, suggesting that Abcc6 was acting in a myocyte non-autonomous manner to cause calcification. Inflammation is known to play a key role in mediating ectopic calcification of the cardiovascular system but bone marrow transplantation experiments from wild type animals into Abcc6 deficient animals did not rescue the calcific phenotype. However, tissue specific deletion of Abcc6 in the liver was sufficient to cause post injury cardiac calcification, suggesting a role of the liver in driving ectopic calcification of distant tissues. Metabolomics and gene expression analysis of animals globally deficient or with liver specific deletion of Abcc6 demonstrated deficiencies in nucleotide metabolism and pathways regulating cellular energetics. We show that such metabolic abnormalities are associated with profound abnormalities in cellular respiration in the injured cardiac tissue. Functional abnormalities in cellular respiration in the injured heart were similar in animals globally deficient in Abcc6 or with liver specific deletion demonstrating that hepatic Abcc6 expression regulates cellular respiration in the injured heart. Calcification of the heart in Abcc6 deficient states was not secondary to induction of an osteogenic gene expression program but occurred rapidly secondary to dystrophic calcification with initial mineralization deposits noted within dead cardiac muscle. Treatment with clodronate or etidronate, bisphosphonates that prevent the growth of mineralization, was sufficient to rescue the phenotype of ectopic cardiac calcification in Abcc6 deficient states. Taken together, these observations highlight the role of the liver in regulating target tissue metabolic and mitochondrial function in causing ectopic calcification in Abcc6 deficient states.

overall_design: 3 days post cryo injury WT and ABCC6 KO mice scar regions of the heart were harvested and digested. Digested live cells were collected by flow cytometry and analyzed by sc-RNA seq.

first: Yijie

last: Wang

first: Arjun

last: Deb


title: An oncogenic KRAS-driven secretome involving TNFα promotes niche preparation prior to pancreatic cancer onset [co-culture scRNA-seq]

city: Ulm

first: Alexander

last: Kleger

address: Albert-Einstein-Allee 23, 89071 Ulm

department: Institute of Molecular Oncology and Stem Cell Biology

country: Germany

institute: University Hospital Ulm

summary:
Background: Pancreatic ductal adenocarcinomas (PDACs) are highly lethal and aggressive with oncogenic KRAS being the main oncogenic driver of the disease. PDACs have been extensively profiled at advanced stages, and in advanced disease the tumor microenvironment is a major determinant that critically shapes patient outcomes. Since the molecular events occurring prior to invasive growth remain poorly understood, we aimed to investigate changes in the precancerous epithelium and its surrounding niche.
Methods: We acquired time-resolved, single-cell transcriptomic (scRNAseq), and accessible-chromatin data from human pluripotent stem cell-derived pancreatic duct-like organoids (PDLO) inducibly expressing KRASG12D and from various niche cells.
Results: Analysis of the pure epithelium already revealed key signatures of matrix remodeling and inflammation-related signaling upon few days of KRASG12D expression. Machine learning captured KRASG12D-dependent transcriptomic classifiers with high prediction accuracy and niche preparatory relevance. Various co-culture approaches followed by scRNAseq and functional validation, including T-cell microfluidics, demonstrated that the KRASG12D-induced PDLO-secretome activates pancreatic stellate cells (PaSCs) and protects precancerous organoids from T cell infiltration. Additional, in silico approaches reconstructed a virtual pancreatic (pre)cancerous space to profile cell-cell interactions between PDLOs and niche cells. TNFα emerged as a top-ranked ligand and was functionally validated to mediate T-cell shielding and PaSC activation. Cyst fluid from 80 prospectively sampled Intraductal Papillary Mucinous Neoplasm (IPMNs) –well-known cystic PDAC precursor lesions– showed a stepwise TNFα rise across LGD (low-grade), HGD (high-grade), and IC (invasive cancer).
Conclusion: Our study reveals that oncogenic KRAS orchestrates niche-preparatory programs that precede PDAC formation and highlight a T cell exclusion program governed by epithelial-derived TNFα.

overall_design: Single-cell RNA-sequencing (scRNA-seq) of human stem-cell derived pacnreatic duct-like organoids (PDLOs) after treatment with doxycycline to induce KRASG12D expression, either in monoculture, bi-culture with PBMC-derived T cells from healthy donors (biTcells) or bi-culture with human pancreatic stellate cells (biStellate).

first: Chantal

last: Allgoewer

first: Markus

last: Breunig


title: Sex-associated changes to synovial macrophages in the aging joint

first: Deborah

middle: R

last: Winter

address: 240 E. Huron Street, M300

institute: Northwestern University

summary: Single-cell analysis of aging synovial macrophage heterogeneity between sexes

overall_design: CITE-seq on CD45+CD11b+Ly6g-SiglecF-Cd64+ cells isolated from hind joints of young and old, male and female mice.


title: Mapping the regulatory landscape of sheep pituitary gland associated with puberty at single-cell resolution

first: Shanglai

last: Li

address: Nanjing

institute: Nanjing agricultural university

summary: The pituitary gland is a remarkably dynamic organ with roles in gonadotropin secretion, and it is composed of numerous cell populations. Defining the roles of pituitary cell in hormone secretion has been challenging, the gene expression patterns of different hormone-producing cells of the pituitary in large animal species are poorly understood, and a cell atlas for ewe is lacking. Single-cell sequencing methods can comprehensively characterize the diversity of pituitary cell types by directly measuring multiple molecular signatures in thousands to millions of individual cells. We created a single-cell transcriptomic atlas of the anterior pituitary gland in ewes at the pre-puberty (3-month-old) and post-puberty (6-month-old) stages, identifying 30,335 cells and ten distinct cell types. The research identified several signaling pathways enriched in different pituitary cells, including the TGFβ, SPP1, and NPR2 pathways. Additionally, we isolated and categorized the gonadotropes for further analysis. This allowed us to identify potentially important genes involved in gonadotropin secretion. We also compared pituitary RNA-seq data from four different developmental stages and discovered several genes associated with gonadotropin secretion, including SCG2 and NPW. Besides, in vitro experiments confirmed that SCG2 was predominantly expressed in gonadotropes and promotes the synthesis and secretion of follicle-stimulating hormone. By comparing pituitary data from humans and ewes, we found 79 conserved genes in gonadotrope cells and 479 in immune cells. Our study systematically mapped out the gene expression profile of the anterior pituitary, pinpointing key genes involved in gonadotropin secretion in ewes. Additionally, we created a cell atlas that could provide crucial molecular information for ewes' reproduction.

overall_design: Single-cell sequencing of pituitary glands from 3-month-old and 6-month-old Hu sheep

first: Yanli

last: Zhang


title: DDX53 binds a wide range of RNA molecules in human TCam-2 cells (eCLIP data)

city: Poznan

first: IHG PAS

last: Bioinfo Team

state: Greater Poland

address: Strzeszynska 32

department: Bioinformatics Team

country: Poland

institute: Institute of Human Genetics, Polish Academy of Sciences

summary: DDX53 (DEAD-box helicase 53, known also as CAGE) is an intronless gene on the X chromosome, which expression shows strong testis and brain specificity. Additionally, it belongs to the group of cancer-testis (CT) antigens, with most studies to date focusing on its role in cancer. Previous reports of rare variants within the DDX53 in infertile men have provided a rationale for our study of DDX53 in the context of human spermatogenesis. By using the human seminoma cell line (TCam-2) as an in vitro male germline model, we aimed to investigate the function and molecular targets of DDX53 protein. Our eCLIP and RNA-seq data show that DDX53 protein binds a wide repertoire of RNA targets, drives transcriptome changes in human cells, and is involved in alternative splicing of RNA transcripts. Further, we examined the localization of DDX53 using confocal microscopy, revealing that it is mainly located in the cytoplasm, but also in the nucleus of TCam-2 cells. Our data indicate that DDX53 protein acts as a regulator of RNA metabolism in human cells. Collectively, we show that DDX53 participates in transcriptome regulation (including splicing) in male germ cells and exhibits transcriptome-wide RNA interactions, but its wider biological role remains to be clarified.

overall_design: Two biological replicates of TCam-2 cells expressing the DDX53-FLAG were used in eCLIP-seq performed by Eclipse BioInnovations (USA). Cells were crosslinked, then lysed, RNase I treated, and protein-RNA complexes were immunoprecipitated with anti-FLAG antibody coupled to magnetic beads. The eCLIP experiment was performed by Eclipse BioInnovations’ standard RBP-eCLIP pipeline.

first: Natalia

last: Rozwadowska

first: Agata

last: Barszcz

first: Erik

last: Dassi


title: ATAC-seq Profiling of Adult Mouse Cardiomyocytes Treated with AAV9-cTnT-Rbm22 in MI Model

first: Zhenzhen

last: Zhan

address: Jimo Road 150

department: Research Center for Translational Medicine

institute: Shanghai East Hospital, Tongji University School of Medicine,

summary: Purpose: The purpose of this study was to identify chromatin alterations regulated by RBM22 in adult mouse cardiomyocytes (AMCMs) after myocardial infarction (MI). We performed ATAC sequencing on AMCMs isolated from mice post-MI that were infected with either AAV9-cTnT-GFP (control viral vector, AAV9-Control) or AAV9-cTnT-Rbm22-GFP (AAV9-Rbm22). Methods: 8-week-old C57BL/6J mice underwent permanent ligation of the left anterior descending (LAD) coronary artery, followed immediately by intramyocardial injection of either AAV9-Control or AAV9-Rbm22 at the infarct border zone. Adult cardiomyocytes were isolated post-MI using the Langendorff perfusion method. Results: We identified differentially altered chromatin accessibility regions between adult cardiomyocytes injected with the indicated viral vectors post-MI.

overall_design: Assay for Transposase-Accessible Chromatin with sequencing (ATAC-seq) was performed to profile chromatin accessibility in adult cardiomyocytes isolated from the infarcted hearts of mice injected with AAV9-Control or AAV9-Rbm22 viruses.

first: Xingguang

last: Liu

first: Xuewen

last: Duan


title: A High-Fiber Plant-Based Diet Delays Progression to Myeloma in Precursor Disorders

first: Juan-Jose

last: Garces

address: 1275 York Ave

institute: Memorial Sloan Kettering Cancer Center

summary: Consumption of a western diet and high body mass index (BMI) are risk factors for progression from pre-malignant phenotypes to multiple myeloma, a hematologic cancer. In the NUTRIVENTION trial (NCT04920084), we administered a high-fiber plant-based diet to patients with myeloma precursor states and elevated BMI. The intervention was feasible, improved quality of life and modifiable risk factors: metabolic (BMI, insulin resistance), microbiome (diversity, composition), and immune (inflammation, monocyte subsets). Disease-progression trajectory improved in two participants and was stable in others. In Vk*MYC+/- mice modeling the myeloma-precursor state, a high-fiber diet delayed disease progression through improved microbiome composition and increased short-chain fatty acid production that reinvigorated anti-tumor immunity and inhibited tumor growth. These effects resulted from fiber consumption and were independent from calorie restriction-induced weight loss.

overall_design:
Viably frozen paired bone marrow mononuclear cells from baseline and W52 from 13 participants (1001, 1003, 1004, 1006, 1009, 1011, 1012, 1013, 1014, 1015, 1017, 1022 and 1023) were sorted by fluorescence-activated cell sorting (FACS) for live CD45+ single cells (CD45: BD Bioscience, catalog# 563204; DAPI: Millipore Sigma, catalog# 508741) and submitted for single-cell sequencing in 0.04% bovine serum albumin in PBS. They were processed using the Chromium Next GEM Single Cell Multiome Reagent Kit A (catalog no. 1000282) and ATAC Kit A (catalog no. 1000280). We followed the Chromium Next GEM Single Cell Multiome ATAC + Gene Expression Reagent Kits User Guide with the difference that single-cell suspension was prepared as described (DOGMA-seq protocol with Digitonin buffer). Briefly, cells were treated with a digitonin lysis buffer (20 mM Tris-HCl pH 7.4, 150 mM NaCl, 3 mM MgCl2, 0.01% digitonin and 2 U/µl of RNase inhibitor) for 5 min on ice, followed by adding 1 mL of chilled wash buffer (20 mM Tris-HCl pH 7.4, 150 mM NaCl, 3 mM MgCl2, and 1 U/µl of RNAse inhibitor) and inversion before centrifugation at 500g for 5 min at 4 °C. The supernatant was discarded, and cells were resuspended in digitonin wash buffer, followed by counting using trypan blue and DAPI on a Countess II FL Automated Cell Counter. In addition, samples for each patient were pooled together on one lane of 10X Chromium (using Hash Tag Oligonucleotides - HTO). Final libraries were sequenced on an Illumina NovaSeq S4 platform (R1 – 28 cycles, i7 – 8 cycles, R2 – 90 cycles).

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

first: Urvi

last: Shah

first: Laura

last: Cogrossi

first: Francesca

last: Castro

first: Anna

last: Policastro

first: Andriy

last: Derkach

first: Teng

last: Fei

first: Susan

last: DeWolf

first: Matteo

last: Grioni

first: Sofia

last: Sisti

first: Jenna

last: Blaslov

first: Peter

last: Adintori

first: Kinga

last: Hosszu

first: Devin

last: McAvoy

first: Mirae

last: Baichoo

first: Justin

last: Cross

first: Jenny

last: Paredes

first: Aishwarya

last: Anuraj

first: Sandeep

last: Raj

first: Charlotte

last: Pohl

first: Paola

last: Zordan

first: Victoria

last: Zinsmeyer

first: Ruben

last: Ramos

first: Marco

last: Lorenzoni

first: Brianna

last: Gipson

first: Kylee

last: Maclachlan

first: Ana

last: Gradissimo

first: Leonardo

last: Boiocchi

first: Nathan

last: Aleynick

first: Camilla

last: Marchigiani

first: Erica

last: Salehi

first: Richard

last: Koche

first: Ronan

last: Chaligne

first: Torin

last: Block

first: Neha

last: Korde

first: Carlyn

last: Tan

first: Malin

last: Hultcrantz

first: Hani

last: Hassoun

first: Gunjan

first: Michael

last: Scordo

first: Oscar

last: Lahoud

first: David

last: Chung

first: Heather

last: Landau

first: Jonathan

last: Peled

first: Nicola

last: Clementi

first: Marta

last: Chesi


title: Disease-associated mutations on the STAT5B SH2 domain regulate cholesterol and lipid metabolism in liver

city: Bethesda

first: Hye Kyung

last: Lee

state: MD

address: 8 CENTER DR RM 107

department: National Institute of Diabetes and Digestive and Kidney (NIDDK)

institute: National Institutes of Health (NIH)

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

overall_design: Refer to individual Series


title: DDX53 drives transcriptome changes in human TCam-2 cells (RNA-seq data)

first: IHG PAS

last: Bioinfo Team

overall_design: The RNA-seq experiment was done in duplicate and performed using TCam-2 cells transduced with DDX53-FLAG, GFP-FLAG and non-transduced WT. Cells transduced with GFP-FLAG and non-transduced WT TCam-2 cells were used as negative controls in order to perform analysis of differentially expressed and differentially spliced genes upon DDX53 overexpression. RNA-seq data from non-transduced WT TCam-2 cells and TCam-2 transduced with GFP-FLAG are deposited in the GEO database under accession code GSE294696.

first: Natalia

last: Rozwadowska

first: Agata

last: Barszcz

first: Katarzyna

last: Tutak

first: Joanna

last: Zyprych-Walczak

first: Erik

last: Dassi

first: Erkut

last: Ilaslan


title: Disease-associated mutations on the STAT5B SH2 domain regulate cholesterol and lipid metabolism in liver [RNA-Seq liver]

summary: Growth hormone (GH) signaling through STAT5B is a central regulator of hepatic metabolism, yet the functional consequences of disease-associated STAT5B variants remain poorly understood. Here, we generated mice carrying STAT5BY665F (gain-of-function) and STAT5BY665H (loss-of-function) alleles using CRISPR/Cas9-mediated base editing to dissect their impact on metabolic regulation. STAT5BY665F mutants developed hepatic lipid accumulation, hypercholesterolemia, and enhanced insulin sensitivity, whereas STAT5BY665H mice displayed reduced body weight and impaired insulin responsiveness. Transcriptomic analyses revealed that STAT5BY665F strongly activated lipid, cholesterol, and immune transcriptional programs, while STAT5BY665H failed to induce these pathways. ChIP-seq demonstrated extensive STAT5BY665F enhancer occupancy at metabolic and immune loci, contrasting with the minimal chromatin engagement of STAT5BY665H. Beyond the liver, STAT5BY665F broadly reprogrammed adipose tissue gene expression, activating lipid metabolism and immune regulatory networks, whereas STAT5BY665H exerted more restricted effects. Notably, transcriptional and chromatin remodeling by STAT5B variants displayed sex-dependent differences, with male livers showing more pronounced reprogramming than females. Together, these findings illustrate how alterations in STAT5B activity that alter enhancer activation led to altered metabolic function in liver and adipose tissue.

overall_design: Total RNA-seq was performed on mouse liver tissue.


title: Disease-associated mutations on the STAT5B SH2 domain regulate cholesterol and lipid metabolism in liver [RNA-seq]

overall_design: Total RNA-seq was performed on mouse adipose tissue.


title: PI3K and MAPK signaling nodes as divergent drivers of phenotypic plasticity in cancer-associated fibroblasts in colorectal cancer [scRNA-Seq]

city: Ghent

first: sandor

last: Dedeyne

address: C. Heymanslaan 10

country: Belgium

institute: University Ghent

summary: Cancer-associated fibroblasts (CAFs) exhibit phenotypic heterogeneity with each functional state playing critical roles in tumor progression. Notably, subtypes like inflammatory CAF (iCAF), characterized by increased chemokine/cytokine secretion, and myofibroblast-like CAF (myCAF), characterized by enhanced extracellular matrix (ECM) deposition and increased actomyosin contractility, can undergo phenotypic switching in response to cues from the tumor microenvironment (TME) and/or therapeutic interventions. However, the signaling pathways associated with their diverse phenotypes remain poorly understood. Through the analysis of single-cell RNA sequencing analysis of human colorectal cancer (CRC) we identified that the PI3K/mTOR and MAPK/ERK signaling pathways, among other pathways, are linked to the formation of myCAF and iCAF subtypes, respectively.In addition,we identified that PI3K/mTOR inhibition promotes the formation of iCAF through compensatory FGF-2 secretion and stimulation of the FGFR1-JAK2-STAT3 pathway; iCAF-derived chemokines/cytokines consequently enhance tumor spheroid growth and neutrophil infiltration.

overall_design: single-cell RNA sequencing data of patient-derived tumor fragments (PDTFs), derived from a peritoneal metastasis lesion from colorectal adenocarcinoma, treated with Trametinib, Everolimus or DMSO control.

first: Zihan

last: Xia

first: Felix

last: De Vuyst

first: Olivier

last: De Wever


title: MGMT crosspaths with cGAS/STING pathway to regulate innate immune response in macrophage

city: Bangkok

first: Tanapat

last: Palaga

address: Payathai Road, Pathumwan

department: Microbiology

country: Thailand

institute: Chulalongkorn University

summary: Transcriptomic profiling demonstrated that stimulation of bone marrow–derived macrophages (BMMs) with the STING agonist DMXAA induced a substantial number of differentially expressed genes (DEGs). Comparative analysis between DMXAA-treated MGMT knockout (KO) and wild-type (WT) BMMs identified a total of 202 DEGs, of which 75 were significantly upregulated genes and 127 were significantly downregulated genes.To elucidate the biological processes underlying these transcriptional alterations, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis was performed on the whole-transcriptome dataset. This analysis revealed enrichment of pathways associated with cellular metabolic activity. Notably, pentose and glucuronate interconversions represented the most positively enriched pathway in DMXAA-stimulated MGMT KO BMMs, whereas linoleic acid metabolism was the most negatively enriched pathway relative to WT cells. Furthermore, Gene Set Enrichment Analysis (GSEA) was conducted using the Hallmark gene set collection. The anaysis showed that the most significantly enriched gene signatures in DMXAA-treated MGMT KO BMMs were associated with IL-6_JAK_STAT3 signaling and hypoxia, underscoring the activation of key inflammatory and stress-response programs under conditions of MGMT deficiency.

overall_design: Bone marrow-derived macrophages (BMMs) from WT and MGMT-deficient mice were incubated with 10 mg/ml of STING agonist DMXAA for 24 hours. RNA was extracted from BMMs using RNeasy kit. Total RNA was analyzed by RNA-seq to identify DEGs and pathway enrichment.

first: Pornrat

last: Kongkavitoon

first: Atsadang

last: Boonmee


title: PI3K and MAPK signaling nodes as divergent drivers of phenotypic plasticity in cancer-associated fibroblasts in colorectal cancer

overall_design: RNA-seq profiling of CT5.3hTERT colorectal CAFs following 24-hour PI3K/mTOR-axis inhibition


title: Intermediate cryopreservation and transcriptomic profiling as strategy to identify optimal donors for functional NK cell expansion for AML immunotherapy

city: Brno

first: Ioanna

last: Papatheodorou

address: Studentska

department: Center for Translational Medicine

country: Czech Republic

institute: International Clinical Research Center (ICRC), St.Anne’s Hospital

summary: NK cells obtained from some of the donors show decreased cytotoxicity towards selected cell lines and/or weak expansion rate. For the manufacturing of immunotherapeutic product and favourable results of clinical studies, it is important to predict these properties of NK cells. Here we show gene expression signature that distinguishes donors whose NK cells show superior cytotoxicity and expansion potential. Based on these data, we propose a donor selection strategy guided by this predictive signature.

overall_design: NK cells were isolated from PBMC that were isolated from buffycoats obtained from healthy donors by magnetic separation and stimulated overnight with IL-2 (500IU/ml) in NK MACs medium with 5% human serum. RNA samples were collected for RNAseq and remaining cells were further expanded for two weeks. RNAseq data after isolation were correlated to functional properties of expanded NK cells in order to setup criteria for highly cytotoxic NK cells and donor selection

first: Eva

last: Mašínová

first: Veronika

last: Švubová

first: Kateřina

last: Kuglerová

first: Kamila

last: Bendíčková

first: Marcela

last: Hortová Kohoutková

first: Jan

last: Vydra

first: Petr

last: Lesný

last: Frič


title: Newly Evolved Endogenous Retroviruses Prime the Ovarian Reserve for Activation

city: Davis

first: Satoshi

middle: H

last: Namekawa

address: One Shields Avenue

department: Department of Molecular and Cellular Biology

institute: University of California, Davis


title: Bulk RNA-Seq gene expression profiles from 13 TCGA cancer types and normal tissues for tumor classification

city: Usti nad Labem

first: Sergii

last: Babichev

address: Tolsteho 1200/9

institute: Babichev Sergii

summary: This dataset includes bulk RNA-Seq gene expression profiles of 6310 samples from 13 cancer types and normal tissues, obtained from The Cancer Genome Atlas (TCGA) and processed using a standardized pipeline. The raw count matrix, initially containing 60,660 genes, was filtered to retain only valid ENTREZ gene identifiers matched via the org.Hs.eg.db annotation package. Genes with zero variance across samples were removed. Normalization was performed by transforming expression values into log2 Counts Per Million (CPM) using the edgeR::cpm() function with log transformation. To ensure all values were positive and suitable for downstream modeling, a global shift was applied by adding the absolute minimum value. Subsequently, Gene Ontology (GO) analysis was performed across all three categories (Biological Process, Molecular Function, Cellular Component) using the TopGO package. Significant genes were selected, followed by Benjamini–Hochberg p-value correction, and validated using both Fisher’s and Kolmogorov–Smirnov tests. The resulting expression matrix consists of 18,564 functionally relevant genes across 6310 samples and forms the basis for clustering, classification, and metric evaluation within hybrid modeling frameworks for cancer diagnostics.

overall_design: Gene expression data were retrieved from The Cancer Genome Atlas (TCGA) using the Genomic Data Commons (GDC) portal via the TCGAbiolinks package in R. For each of the 13 cancer types, data were queried with the following parameters: data.category = "Transcriptome Profiling", data.type = "Gene Expression Quantification", and workflow.type = "STAR - Counts". Only primary tumor and normal tissue samples were included. The raw count matrices were filtered to retain valid ENTREZ gene identifiers, followed by the removal of genes with zero variance across samples. Log2 Counts Per Million (CPM) were computed using the edgeR::cpm() function, and a global shift was applied to ensure all values were positive. Subsequently, gene selection was performed using Gene Ontology (GO) analysis across all three ontologies (BP, MF, CC). The final matrix (6310 samples × 18,564 genes) was used for downstream clustering and classification tasks.


title: Effects of attenuating astrocytic Ca2+ activity in the medial prefrontal cortex with iβARK on transcriptional profiles in wild-type mice.

city: Guangzhou

first: Yamei

last: Tang

state: Guangdong

address: 107 Yan Jiang West Road

department: Department of Neurology

institute: Sun Yat-Sen Memorial Hospital

summary: We injected adeno-associated viruses (AAV2/5) carrying an inhibitory peptide derived from β-adrenergic receptor kinase 1 (iβARK), tagged with mCherry for visualization, into the medial prefrontal cortex (mPFC) of wild-type mice. This approach selectively attenuated Gq-coupled GPCR-induced Ca²⁺ activity in astrocytes. Control mice received AAV2/5 expressing mCherry alone. Subsequently, we performed RNA sequencing on mPFC tissue samples to compare transcriptomic profiles and investigate the potential functional consequences of perturbing Gq-GPCR signaling.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for WT-mCherry and WT-iβARK.

first: Zhan

last: Zhang

first: Sitai

last: Chen

first: Wei-Jye

last: Lin


title: Newly Evolved Endogenous Retroviruses Prime the Ovarian Reserve for Activation [RNA-seq]

summary: Female mammals are born with a finite pool of non-growing oocytes (NGOs) housed in primordial follicles, forming the ovarian reserve that determines reproductive lifespan. Mechanisms underlying the reserve’s long-term maintenance and subsequent follicular activation remain elusive. Using total RNA sequencing and de novo transcriptome assembly across perinatal oogenesis in mice, we captured the full oocyte transcriptome. We show that NGOs establish extensive accessible chromatin at gene regulatory elements—including promoters and enhancers—with formation of many regions driven by newly evolved endogenous retroviruses. In NGOs, epigenetic priming for follicular activation involves pre-loading RNA polymerase II and transcription factors TCF3 and TCF12 at these sites, counteracted by repression via Polycomb Repressive Complex 1-mediated H2AK119 ubiquitylation. This transposable element (TE)-mediated epigenetic priming underlies the ovarian reserve’s long-term maintenance and poises oocytes for activation. Our findings establish the ovarian reserve as an actively primed population, with newly evolved TEs orchestrating genome-wide transcriptional activation.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for WT perinatal oocytes.

first: Yasuhisa

last: Munakata


title: Species-Specific Roles of SP1 in Bovine and Human Embryonic Genome Activation and Early embryonic Development

city: hangzhou

first: Hao

last: Jin

address: Nongshenghuan Building E, Zhejiang University, Hangzhou, Zhejiang

institute: Zhejiang University

summary: Embryonic genome activation (EGA) marks the onset of embryonic program and enables the transition toward the first lineage specification. However, the molecular features of EGA and the transcription factors (TFs) orchestrating this process remain unclear. Here, by performing single-cell RNA-seq on bovine embryos, we reveal that major EGA is asynchronously initiated among blastomeres at the 8-cell stage. Integrative analyses reveal distinctive protein accumulation compared to transcription and translation activation during bovine EGA. Furthermore, we investigate the role of SP1, a TF activated at the minor EGA stage, with motifs enriched in accessible chromatin during major EGA stage in bovine and human embryos. SP1 deficiency leads to morula arrest in bovine and impairs EGA in human embryos. Multi-omics analysis demonstrates that SP1 promotes early lineage gene expression by modulating nearby chromatin states in bovine and directly targets key EGA genes in human embryos. Together, our study delineates the dynamics of bovine EGA and uncovers the conserved and species-specific roles of SP1 in regulating EGA and early development in mammals.

sample_organism:
Homo sapiens
Bos taurus

platform_organism:
Homo sapiens
Bos taurus

overall_design:
To investigate the dynamic expression during bovine embryonic genome activation, we performed scRNA-seq of bovine embryos at GV, MII, zygote, 2-cell (2C), 4-cell (4C), early/late 8-cell (E/L8C), and 16-cell (16C) stages.
To investigate the role of SP1 during bovine early embryo development, we performed RNA-seq, SP1 CUT-Tag, H3K27ac ChIP-seq, and ATAC-seq in control, SP1 KD and SP1 KO embryos at 8C, 16C and morula stage,
To investigate the role of SP1 during human early embryo development, we performed RNA-seq, SP1 CUT-Tag in control, SP1 KD embryos at 8C stage,

first: Lieying

last: Xiao

first: Kun


title: Histone lactylation-mediated metabolic remodeling in vascular smooth muscle cells aggravates aortic aneurysm and dissection via promoting lactate accumulation

city: Shanghai

first: Liwei

last: Liu

address: 180 Fenglin Road

department: Department of Cardiology,Zhongshan Hospital

institute: Shanghai Institute of Cardiovascular Diseases,Fudan University

summary: The phenotypic switch of vascular smooth muscle cells (VSMCs) is a crucial pathogenesis in aortic aneurysm and dissection (AAD), metabolic remodeling from oxidative phosphorylation (OXPHOS) to glycolysis involved in the process. Histone lactylation expression was analyzed in aorta of aortic aneurysm (AA) patients and AAD mice, as well as the AngII(angiotensin II)-treated VSMCs. CUT&Tag was used to explore the downstream target gene for H4K16 lactylation (H4K16la) between AngII-induced VSMCs and Control. The finding was to explore the impact of histone lactylation (H4K16la) on the progress of AAD.

overall_design: CUT&Tag analysis of H4K16 lactylation in human aortic vascular smooth muscle cells.

first: Aijun

last: Sun

first: Junbo

last: Ge


title: Enhancer Dynamics and Spatial Organization Drive Anatomically Restricted Cellular States in the Human Spinal Cord [snRNA-seq]

city: Madison

first: Abbas

middle: Haider

last: Rizvi

state: WI

address: 1500 Highland Ave, Room 715 Room 715

department: Neuroscience

institute: University of Wisconsin-Madison

summary: Here, we report the spatial organization of RNA transcription and associated enhancer dynamics in the human spinal cord at single-cell and single-molecule resolution. We expand traditional multiomic measurements to reveal epigenetically poised and bivalent active transcriptional enhancer states that define cell type specification. Simultaneous detection of chromatin accessibility and histone modifications in spinal cord nuclei reveals previously unobserved cell-type specific cryptic enhancer activity, in which transcriptional activation is uncoupled from chromatin accessibility. Such cryptic enhancers define both stable cell type identity and transitions between cells undergoing differentiation. We also define glial cell gene regulatory networks that reorganize along the rostrocaudal axis, revealing anatomical differences in gene regulation. Finally, we identify the spatial organization of cells into distinct cellular organizations and address the functional significance of this observation in the context of paracrine signaling. We conclude that cellular diversity is best captured through the lens of enhancer state and intercellular interactions that drive transitions in cellular state. This study provides fundamental insights into the cellular organization of the healthy human spinal cord.

overall_design: single nucleus RNA-seq from the C6, T4, and L4 segments of healthy adult human spinal cord resected from a tissue donor.


title: Newly Evolved Endogenous Retroviruses Prime the Ovarian Reserve for Activation [ATAC-seq]

overall_design: Comparative chromatin profiling analysis of ATAC-seq data for WT perinatal oocytes.


title: Disease-associated mutations on the STAT5B SH2 domain regulate cholesterol and lipid metabolism in liver [ChIP-seq]

first: Hye Kyung

last: Lee


title: Comparative clinical transcriptome of vir genes in severe Plasmodium vivax malaria

city: Bangalore

first: Genotypic

last: technology

state: Karnataka

address: 259, Apoorva 4th cross,80 feet Road,RMV 2ND STAGE

country: India

institute: Genotypic Technology

summary: Vir/pvpir genes, a multigene family in Plasmodium vivax that are a part of a larger  superfamily of genes called the pir (Plasmodium interspersed repeat) genes have been  reported earlier to be possibly involved in cytoadherence and evasion of splenic clearance.  Plasmodium vivax, historically characterized as a "benign" malaria parasite, has been  associated with clinical outcomes including hepatic dysfunction, renal failure, and cerebral  malaria in India and several global regions. It constitutes an economic burden and presents a  public health challenge alongside other Plasmodium species. Here, we present a part of  global transcriptomic studies by custom designed microarray, that compare the transcriptome  of the parasite responsible for severe Plasmodium vivax manifestations, specifically hepatic  dysfunction and cerebral malaria from India, with an emphasis on the vir/pvpir genes, some  of which are reported to play a role in cytoadherence. 23 patients with Plasmodium vivax  malaria (Uncomplicated=6, Hepatic dysfunction=12 and Cerebral malaria=5) were subjected  to microarray hybridization and the data so obtained showed a wide range of vir/pvpir  subtelomeric subfamilies have been differentially expressed. Upregulation has been seen in  24 vir/pvpir genes in cerebral malaria samples (n=5) and 28 genes in hepatic dysfunction  samples (n=12) belonging to different subfamily in at least 50% of the patient samples. Out  of the upregulated vir/pvpir genes in cerebral malaria manifestation, members of vir  subfamily E and pvpir H are maximum in number whereas in hepatic dysfunction  manifestation, members of vir subfamily E and C comprise a significant proportion.

overall_design: Total RNA from 23 clinical isolates of Plasmodium vivax was isolated from malaria patients with differing clinical conditions (n=23). Each of these samples was hybridized individually on custom-designed 8*15K arrays printed on the Agilent platform for their transcriptional profiling.

first: Saurabh

last: Singh

first: Pon Arunachalam

last: Boopathi

first: Priyanka

last: Roy

first: Ashis

last: Das


title: Cell autonomous and non-autonomous effects of constitutive Yap1 activation in distal nephron segments

city: Cincinnati

first: Hee-Woong

last: Lim

state: Ohio

address: 3333 Burnet Ave. MLC 7024

department: Division of Biomedical Informatics

institute: Cincinnati Children's Hospital Medical Center

summary: The distal convoluted tubule (DCT) and connecting tubule (CNT) are critical for fine-tuning electrolyte reabsorption and maintaining renal homeostasis. While the Hippo pathway effector Yap1 is known to regulate renal development, its specific function in distal nephron segments remains poorly defined. To investigate Yap1 function in the distal nephron, we generated a Cre-inducible Yap1 gain-of-function allele (Col1a1-YAP5SA) and a distal nephron-specific Slc12a3-IRES-Cre to express constitutively active Yap1 in the distal nephron segments. Lineage tracing analysis showed that the Slc12a3Cre targets both DCT and CNT, suggesting that CNT cells arise from Slc12a3+ DCT cells. Constitutive activation of Yap1 in DCT and CNT results in increased proliferation and abnormal migration into adjacent nephron segments. This was accompanied by disrupted expression of DCT/CNT marker genes, loss of apicobasal polarity, and compromised junctional architecture. Unexpectedly, the mutant kidneys showed the downregulation of proximal tubule marker genes and Ren1 and the upregulation of inflammation and injury markers. These findings demonstrate that sustained Yap1 activation in the distal nephron segments impairs epithelial identity and structure, and also induces non–cell-autonomous effects in other nephron segments, particularly the proximal tubule. This highlights the importance of tightly regulated Hippo-Yap signaling in maintaining epithelial integrity and proper nephron segmentation.

overall_design: RNA-seq profiling of Yap1 gain of function mutant and control kidneys at postnatal day 3.

first: Joo-Seop

last: Park

first: Eunah

last: Chung

first: Christopher

last: Ahn


title: Newly Evolved Endogenous Retroviruses Prime the Ovarian Reserve for Activation [CUT&Tag]

first: Satoshi

last: Namekawa

overall_design: Comparative chromatin profiling analysis of CUT&Tag data for WT perinatal oocytes.

first: Yasuhisa

last: Munakata


title: Enhancer Dynamics and Spatial Organization Drive Anatomically Restricted Cellular States in the Human Spinal Cord [STABseq]

overall_design: Single nuclei were profiled for simultaneous histone state and chromatin accessibility from the T4 and L4 segments of healthy adult human spinal cord resected from organ donors.


title: ATAC-seq to Profile the Chromatin Accessibility in RBM22-silencing and Control Primary Cardiomyocytes

first: Zhenzhen

last: Zhan

summary: Purpose: The purpose of this study is to identify chromatin alterations regulated by RBM22. We performed ATAC-sequencing in primary cultured neonatal mouse cardiomyocytes transfected with control siRNA or Rbm22 siRNA. Methods: Cardiomyocytes were isolated from 1-day-old C57BL/6J mice and cultured in DMEM with 10% fetal bovine serum at 37°C in an atmosphere with 5% CO2. Cardiomyocytes were stained to confirm the expression of c-TnT. Cells with purity >97.5% were used for subsequent experiments. Cardiomyocytes were transfected with control siRNA or Rbm22 siRNA for 48 hours. Results: We identified regions of chromatin that were altered between cardiomyocytes transfected with the indicated siRNAs.

overall_design: Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq) was performed to profile chromatin accessibility in primary cultured neonatal mouse cardiomyocytes transfected with control siRNA or Rbm22 siRNA.

first: Xingguang

first: Xuewen

last: Duan


title: Persistent T cell Activation and Cytotoxicity against Glioblastoma Following Single Oncolytic Virus Treatment in a Clinical Trial

city: Boston

first: Maxime

last: Meylan

state: MA

address: 450 Brookline Avenue

department: Cancer Immunology Virology

institute: Dana-Farber Cancer Institute

summary: A recent first-in-human clinical trial (clinicaltrials.gov NCT03152318) demonstrated that survival in glioblastoma (GBM) patients following rQNestin34.5v.2 oncolytic virus treatment was associated with immune activation signatures.  Here we provide in situ evidence for ongoing T cell-mediated cytotoxicity against tumor cells at late timepoints following single treatment, with deep and persistent T cell infiltration into tumor regions. Shorter distances between cleaved caspase-3+ tumor cells and granzyme B+ T cells were associated with longer progression free survival following treatment. Pre-existing tumor-infiltrating T cells expanded locally upon treatment, correlating with longer overall patient survival.  T cells with an early activation program closely interacted with tumor cells and were strongly enriched upon treatment.  Viral remnants were restricted to necrotic regions, while T cells infiltrated deeply into live tumor regions.  These data demonstrate that single oncolytic virus treatment can expand pre-existing T cell clones and trigger persistent T cell-mediated immunity against GBM.

overall_design: Xenium spatial transcriptomics analysis of 16 FFPE tissue sections (8 paired pre/post treatment) from recurrent GBM patients treated with a single intratumoral injection of rQNestin34.5v.2 (CAN-3110) oncolytic virus in clinical trial NCT03152318. Pre-treatment biopsies and post-treatment surgical specimens collected at time of progression/recurrence. Specimens analyzed with Xenium Analyzer using a 480-gene panel (380 commercial IO genes + 100 custom probes).

first: Ye

last: Tian

first: Lijian

last: Wu

first: Alexander

last: Ling

first: Mario

last: Suvà

first: Antonio

last: Chiocca

first: Kai

last: Wucherpfennig


title: Enhancer Dynamics and Spatial Organization Drive Anatomically Restricted Cellular States in the Human Spinal Cord [scRNA-seq]

first: Abbas

last: Rizvi

overall_design: single nucleus RNA-seq from the T4 and L4 segments of healthy adult human spinal cord resected from organ donors.


title: RBM22 CUT&Tag Profiling of Regenerative (P1) and Non-Regenerative (P8) Neonatal Mouse Cardiomyocytes

summary: Purpose:  To uncover the molecular mechanisms underlying RBM22-mediated neonatal heart regeneration, we performed Cleavage Under Targets and Tagmentation (CUT&Tag) analysis on regenerative and non-regenerative wild-type (WT) mouse cardiomyocytes over a 7-day period. Methods:  Cardiomyocytes were isolated from postnatal day 1 (P1, regenerative) and postnatal day 8 (P8, non-regenerative) mice, and cultured in DMEM supplemented with 10% fetal bovine serum at 37°C in a 5% CO₂ atmosphere. Cells were stained to confirm the expression of c-TnT, and only those with purity >97.5% were used for subsequent experiments.Results: We carried out a CUT&Tag assay using antibody against RBM22 and found the global influence of RBM22 in regulating genes in transcription regulatory regions of key genes involved in proliferative cardiomyocyte effector function.

overall_design: Cleavage Under Targets and Tagmentation (CUT&Tag) analysis for RBM22-binding DNA  in P1 (regenerative) and P8 (non-regenerative) wild-type (WT) mouse cardiomyocytes.

last: Liu


title: Single-cell RNA sequencing data from human CAR-T cell samples

city: Tsukiji

first: Yuki

last: Okuhiro

state: Tokyo-to

institute: National cancer center research institute

summary: Raw RNA sequencing data derived from human CAR-T cell samples are deposited in this repository. The datasets include bulk single-cell RNA-seq reads generated for research purposes.

overall_design: CD3+ T cells purified from PBMCs of healthy individuals were activated with anti-CD3 and CD28 monoclonal antibodies and lentivirally transduced with the anti-CD19-BBz CAR gene with DMSO or ponatinib (100 nM) in RPMI-1640 supplemented with 10% Fetal bovine serum (FBS) and 4 mM L-glutamine (Thermo Fisher Scientific; Cat. #25030-081). Recombinant human IL-7 (R&D System, #207-IL) and IL-15 (R&D System, #247-ILB) were added at 10 ng/mL. Beads were removed on day 4 using a magnet, and cells were rested for a further 4 days. CD4+ CAR-T cells and CD8+ CAR-T cells were isolated by Fluorescence-Activated Cell Sorting (FACS). Finally, samples were prepared by mixing CD4+ CAR-T cells and CD8+ CAR-T cells at a 1:9 ratio and analized by scRNA-seq.

first: Yasuhiro

last: Kojima

first: Hiroyoshi

last: Nishikawa


title: Enhancer Dynamics and Spatial Organization Drive Anatomically Restricted Cellular States in the Human Spinal Cord [ATAC-seq]

overall_design: single nucleus ATAC-seq from the T4 and L4 segments of healthy adult human spinal cord resected from organ donors.


title: Next Generation Sequencing Facilitates Quantitative Analysis of  RBM22-silencing and Control Primary Neonatal Mouse Cardiomyocytes Transcriptomes

summary: Purpose:The purpose of this study is to identify genes that are either activated or silenced in cardiomyocytes treated with siRNA targeting RBM22 (si-Rbm22) compared to control siRNA (si-Control). Gene expression differences between the two samples were identified using transcriptome profiling (RNA-seq) analysis. Methods: Primary neonatal mouse cardiomyocytes were isolated from neonatal C57BL/6J mice at postnatal day 1.Cardiomyocytes  were stained to confirm the  expression of c-TnT. Cells with purity >97.5% were used for subsequent experiments. Neonatal cardiomyocytes in primary culture were transfected with either siRNA targeting Rbm22 (si-Rbm22) or a non-targeting control siRNA (si-Control) for 48 h, of which RNA profiles were generated by deep sequencing using Illumina. Results: We mapped about 10 million sequence reads per sample to the mouse genome and identified numerous genes with significant mRNA variation between cardiomyocytes transfected with the indicated siRNAs.

overall_design: mRNA profiles of neonatal mouse cardiomyocytes transfected with control siRNA or Rbm22 siRNA were generated by deep sequencing.


title: Temporal constraints for developmental cell competition in mosaic gastruloids

city: Barcelona

first: Joshua

middle: D

last: Frenster

address: Carrer del Doctor Aiguader, 88

country: Spain

institute: Universitat Pompeu Fabra

summary: In early mammalian embryogenesis, the selective elimination of suboptimal cells is critical for developmental integrity. Cell competition is a non-autonomous quality control in which “winner” cells eliminate viable but suboptimal “loser” cells based on their relative difference in fitness. Due to its central role in fitness perception, loss of p53 results in the emergence of “supercompetitor” cells, which stand at the apex of cell competition and induce apoptosis in neighboring wild-type (WT) cells. Here, we investigate cell competition dynamics using mosaic 3D mouse gastruloids, an embryonic stem cell (ESC)-based in vitro model of gastrulation, composed of defined numbers of WT and p53-KO cells. In mosaic gastruloids, even low numbers of p53-KO cells robustly outcompete WT cells, and introduction of as few as two p53-KO cells is sufficient to measurably impair neighboring WT cell growth. Cell competition in gastruloids is independent of proliferation rates, nutrient availability, or reactive oxygen species, and not influenced by Nodal and ERK signaling. However, we observe that Wnt and BMP signaling protect from cell competition, which is exclusively driven by intrinsic apoptosis, as indicated by Bcl2-mediated complete rescue of WT cells. During gastruloid development, cell competition is temporally restricted to a window of 48–96 hours after aggregation, mirroring embryonic days E5.5–E7.5 in the mouse. Heterochronic mosaic gastruloid experiments demonstrate that relative differences in pluripotency levels are neither necessary nor sufficient to cause supercompetition, but that cell competition is contingent on both competitors residing within the developmental window permissive to competition. Neither pluripotent mosaic 3D aggregates, nor 3D EpiGastruloids, which model more advanced developmental processes, display any competition, supporting the hypothesis that developmental cell competition is specific to the onset of gastrulation. Our findings offer insights into the mechanisms of cell fitness evaluation in mammalian embryogenesis and establish gastruloids as a powerful 3D model for investigating developmental stage-specific cell competition.

overall_design: Mouse E14tg2A embryonic stem cells were cultures in serum+LIF conditions. Three clones were generated, one clone referred to as "F" carried an H2B-mCherry transgene, a clone "W" carrying an H2B-emiRFP670 transgene, and a daughter clone "K" which had the same H2B-emiRFP670 transgene and additionally a p53-KO by +1 frameshift insertion in exon 4 of the Trp53 gene. Each one of these clones was sequenced in pluripotent conditions before gastruloid formation. (o h). Then these clones were used for the generation of mosaic 3D gastruloids consisting of either 150 "F" cells with 150 "W" cells, or 150 "F" cells with 25 "K" cells. Mosaic gastruloids were dissociated and sorted into their respective clone of origin groups by FACS using their mCherry or emiRFP670 fluorophore markers. Gastruloids were harvested at 48h, 72h, 96h, 120h, 144h after gastruloid aggregation to generate a timecourse. Sorted cells were lysed and RNA was extracted and used for library preparation for RNA-TruSeq mRNA- sequencing on a NovaSeq6000-S4 with 2x150- 50 M read/sample conditions.

first: Pau

last: Pascual-Mas

first: Alfonso

last: Martinez Arias


title: Transcriptome-wide analysis of Sense and Natural Antisense Transcripts (NATs) in Plasmodium vivax clinical isolates from uncomplicated or complicated diseases manifestation.

first: Genotypic

last: technology

summary: We developed a custom-designed 8×60K Plasmodium vivax microarray to enable genome-wide expression profiling of both nuclear and mitochondrial transcripts from field isolates. Probe selection was guided by a combination of bioinformatic prediction and experimental validation to ensure optimal detection of parasite transcripts in clinical samples. The array features 60-mer oligonucleotide probes in situ synthesized on glass slides using Agilent technology. It includes validated probes representing 5,565 nuclear genes and 19 mitochondrial genes in sense orientation, along with 5,566 and 19 genes, respectively, in antisense orientation capturing dual-stranded expression. In most cases, multiple probes were designed in both orientations against each transcript. Validation using 14 clinical samples (including both uncomplicated and severe (Hepatic Dysfunction) cases, demonstrated a high sensitivity, with ≥ 90% transcript detection. Multiple probes targeting the same gene showed consistent expression profiles across samples. Hybridization results were further validated against previous transcriptomic data and qRT-PCR assays, confirming the array's reliability. Uniquely designed for P. vivax, this sensitive and reproducible microarray provides a powerful functional genomics platform to generate high-confidence expression data and identify differentially regulated genes for both sense and Natural Antisense transcripts (NATs) from field isolates, including both the nuclear and mitochondrial transcriptomes.

overall_design: Total RNA from 14 clinical isolates of Plasmodium vivax was isolated from malaria patients with Hepatic Dysfunction manifestations (n=14). Each of these samples was hybridized individually on custom-designed 8*60K arrays printed on the Agilent platform for their sense and antisense transcriptional profiling

first: Priyanka

last: Roy

first: Ashis

last: Das


title: Pro-inflammatory signaling originates from epicardium-derived fibroblasts in a PKP2 deficient model of arrhythmogenic cardiomyopathy

city: Rochester

first: Cameron

last: Baker

address: 601 Elmwood Ave

institute: University of Rochester

summary: Introduction: Arrhythmogenic Cardiomyopathy (AC) is an inherited disease that is caused by desmosome protein mutation with plakophilin 2 (PKP2) mutation being most common. AC is known to cause sporadic ventricular arrhythmias, myocyte damage, and subepicardial fibrosis. Given the origin of fibrosis from epicardium, we wanted to understand how epicardium-derived cells (EDCs) contribute to the AC pathogenesis. Hypothesis: EDCs propagate the pro-inflammatory signaling that contributes to AC pathogenesis. Methods: We developed transgenic mice that lack PKP2 in cardiomyocytes (PKP2-cKO) or in both cardiomyocyte and epicardial cells (PKP2-ceKO) via the tissue-specific expression of tamoxifen-inducible Cre recombinase. Tamoxifen injected mice expressing Cre in cardiomyocytes, but with no floxed PKP2 gene, was used as a control. EDCs were traced with Cre-dependent Green Fluorescence Protein (GFP). Non-myocyte populations were isolated 21 days post-tamoxifen injection for single cell RNA-sequencing (scRNA-seq) using 10X Genomics platform. Perfused hearts were separated into left and right ventricle for qRT-PCR. Immunohistochemistry was used for cardiac structure and cellular composition. Echocardiography was performed to measure cardiac physiology.  Results: The scRNA-seq analysis identified an epicardium-derived fibroblast population that secretes pro-inflammatory cytokines, including CCL2, CCL7, Thbs1, and PTX3. While pro-inflammatory EDCs are found in both PKP2-cKO and PKP2-ceKO mice, they were most numerous in PKP2-ceKO mice. Importantly, macrophages and B cells accumulated in both PKP2-cKO and PKP2-ceKO mice compared to controls. qRT-PCR confirmed that the expression of genes encoding fibrosis markers (POSTN, Col1A1, Col1A2, and Col3A1), pro-inflammatory fibroblast markers (CCL2, CCL7, Thbs1, PTX3), macrophage markers (F4/80, Cx3cr1, Spp1, Trem2, and CCR2), and B cell markers (CD19 and CD79b) were significantly enriched in both PKP2-cKO and PKP2-ceKO mice compared to control. Our expression and histologic data also revealed an exaggerated pro-inflammatory response in PKP2-ceKO mice, that progresses from the right to bi-ventricular predominance. However, echocardiography showed no significant difference in cardiac function between PKP2-cKO and PKP2-ceKO, and B cell depletion did not alter disease progression. Conclusion: A subset of epicardium-derived fibroblasts in PKP2 KO mice secretes pro-inflammatory cytokines that contributes to cardiomyocyte damage and cardiac fibrosis in AC.

first: Daniel

last: Han

first: Alan

last: Brooks

first: John

last: Ashton

last: Delmar

first: Eric

last: Small


title: Transcriptomes of endothelial cells from mesenteric adipose tissue from wildtype and Myct1ecKO mice

city: Lausanne

first: Tania

last: Wyss

address: Rue du Bugnon 25A

country: Switzerland

institute: SIB Swiss Institute of Bioinformatics

summary: Single cell RNA sequencing of mesenteric adipose endothelial cells from adult willdtype and Myct1ecKO mice

overall_design: Wildtype and Myct1ecKO (Myct1 fl/fl; Pdgfb-CreERT2) mice (C57BL6) were induced with tamoxifen at 3 weeks old. When they reached 14 weeks old, mesenteric adipose tissue was collected and endothelial cells (CD45-CD31+) were sorted by FACS. Sorted cells from 2 replicates per genotype were subjected to 10x Genomics single-cell RNA seq analysis.

first: Anna

last: Köck

first: Amélie

last: Sabine

first: Tatiana

middle: V

last: Petrova


title: Enhancer Dynamics and Spatial Organization Drive Anatomically Restricted Cellular States in the Human Spinal Cord [snATAC-seq]

first: Abbas

last: Rizvi

overall_design: single nucleus ATAC-seq from the C6, T4, and L4 segments of healthy adult human spinal cord resected from a tissue donor.


title: Circadian regulator PER1 inhibits osteoclastogenesis by activating inflammatory genes

city: Minneapolis

first: Nobuaki

last: Kikyo

state: MN

address: University of Minnesota, MTRF, Room 2-216, 2001 6th St SE

institute: University of Minnesota

summary: Disruption of circadian rhythms predisposes shift workers to many chronic conditions, including osteoporosis. However, the effects of disrupted circadian rhythms on bone remodeling remain largely unknown. Here, we show that one of the core circadian regulators PER1 inhibits osteoclastogenesis by upregulating genes involved in inflammation. The conditional knockout of Per1 in osteoclasts and related cells resulted in decreased bone mass in the femurs of mice, along with increased osteoclasts and decreased osteoblasts. Osteoclastogenesis was also promoted by Per1 depletion in vitro with 16 downregulated inflammatory genes. Seven of these genes were known to promote or inhibit osteoclastogenesis depending on the stage of osteoclastogenesis and the presence or absence of infection. Knockdown of Nlrp3, Tlr8, or Tlr9 in the group of genes promoted osteoclastogenesis, mirroring the effects of Per1 knockout and offering a mechanistic explanation for the Per1-mediated inhibition of osteoclastogenesis. These results were not observed following the knockout of a paralog Per2. Per1 knockout mice maintain general circadian rhythms, unlike arrhythmic Per1;Per2 double knockout mice. This gives credence to Per1 as a selective target for therapeutic interventions without disrupting the circadian rhythms. This study uncovered a molecular link between a circadian regulator and osteoclastogenesis in the broader context of inflammatory reactions. Our findings may be mechanistically relevant to inflammatory bone diseases influenced by circadian rhythms, such as rheumatoid arthritis and osteoarthritis, as well as other bone diseases predisposed by chronic circadian disruption.

overall_design: Monocytes were harvested from the bone marrow of 8-12 weeks old mice and induced to differentiate into osteoclasts with M-CSF and RANKL. The cells were harvested 4 days later for RNA-seq.

first: Nobuko

last: Katoku-Kikyo

first: Elizabeth

last: K. Vu

first: Ismael

last: Y. Karkache

first: Samuel

last: Mitchell

middle: W

last: Bradley


title: Selective CDK7 Inhibition Sensitizes Tumors to Topoisomerase I Inhibitor-Antibody Drug Conjugates by Blocking The Transition of The RNA polymerase II Initiation Complex into Elongation [ChIP-seq]

city: Nam-gu, Pohang-si

first: Hyerim

last: Jung

state: Gyeongsangbuk-do

address: 47, Jigok-ro

department: Dept. of Life Sciences

country: South Korea

institute: Pohang University of Science & Technology (POSTECH)

summary: This study investigates the transcriptional impact of Q901, a highly selective CDK7 inhibitor in clinical development. Q901 primarily disrupted MYC and E2F-dependent transcription program, downregulating cell cycle control and DNA damage repair pathways. CDK7 binding at the promoter-proximal regions was dramatically stabilized by Q901, leading to reduced occupancy of MYC, E2F, and RNA Polymerase II (RNAPII). These findings offered a novel therapeutic strategy to enhance cancer susceptibility to TOP1-DNA protein crosslinks (TOP1-DPCs) induced by TOP1 inhibitors. Resistance to TOP1 inhibitors arises through activation of DNA repair pathway when elongating RNAPII encounters TOP1-DPCs. By suppressing RNAPII transition from initiation to elongation and DNA repair pathways, Q901 stabilizes TOP1-DPCs and sensitizes tumor to TOP1 inhibitors. Preclinical studies demonstrated enhanced tumor suppression when combining Q901 with TOP1 inhibitor-based antibody-drug conjugates (TOP1i-ADCs), highlighting its potential as a therapeutic option for cancers resistant to TOP1i-ADC therapy.

overall_design: ChIP-seq of pan pol II, Ser5P, Ser2P, CDK7, MYC, E2F1, and TOP1 in MCF-7 cells with Q901 or TPT treatment.

first: Tae-Kyung

last: Kim


title: RNA sequencing datasets generated from murine CD8-positive T cells

first: Yuki

last: Okuhiro

summary: This repository contains raw bulk and single-cell RNA sequencing reads generated from mouse T lymphocyte samples. Libraries were prepared and sequenced using standard protocols.

overall_design: RNA-seq profiling of murine naive CD8+ T cells from splenocytes of C57BL/6 mice after 48 hours of stimulation with anti-CD3 and anti-CD28 monoclonal antibodies in the presence of either DMSO (control) or 10 nM ponatinib.

first: Yasuhiro

last: Kojima

first: Hiroyoshi

last: Nishikawa


title: High-throughput linear amplification for single-cell template strand sequencing with sci-L3-Strand-seq

city: Los Angeles

first: Yi

last: Yin

address: 695 Charles E. Young Dr. S.

department: Human Genetics

institute: University of California, Los Angeles

summary: Despite the many advances in single cell genomics, detecting structural rearrangements in single cells, particularly error-free sister-chromatid exchanges, remains challenging. Here we describe sci-L3-Strand-seq, a combinatorial indexing method with linear amplification for DNA template strand sequencing that cost-effectively scales to millions of single cells, as a platform for mapping mitotic crossover (CO) and resulting genome instability events. We provide a computational framework to fully leverage the throughput, as well as the relatively sparse but multifaceted genotype information within each cell that includes strandedness, digital counting of copy numbers, and haplotype-aware chromosome segmentation, to systematically distinguish seven possible types of mitotic CO outcomes. We showcase the power of sci-L3-Strand-seq by quantifying the rates of error-free and mutational COs in thousands of cells, enabling us to explore enrichment patterns of genomic and epigenomic features. The throughput of sci-L3-Strand-seq also gave us the ability to measure subtle phenotypes, opening the door for future large mutational screens. Furthermore, mapping clonal lineages provided insights into the temporal order of certain genome instability events, showcasing the potential to dissect cancer evolution. Altogether, we show the wide applicability of sci-L3-Strand-seq to the study of DNA repair and structural variations.

overall_design: We performed replication template strand sequencing of single cells using sci-L3-Strand-seq for mouse diploid Patski, human diploid BJ-5ta, human haploid HAP1, and HAP1 LIG4- knockout cells.

first: Peter

last: Chovanec

first: Trevor

last: Ridgley