title: Effect of Vnut depletion on gene expression during CD4+ T cell differentiation into Th1 cells

city: Beijing

first: Ligong

last: Chen

address: Shuangqing Road No.30

institute: tsinghua university

summary: Extracellular ATP (eATP), a well-recognized danger signal and immune activator, is implicated in the activation, differentiation and function of T cells, directly or indirectly. Yet, how T cells release ATP themselves and its effects remain poorly investigated. Here we found vesicular nucleotide transporter (VNUT)—critical for vesicular ATP storage/release—is highly expressed in Th1 cells and selectively restricts their differentiation and effector functions. Mechanistically, VNUT facilitates lysosomal ATP import and its extracellular release upon T cell receptor engagement. This eATP then activates the purinergic receptor P2X7R and downstream SRC kinase, triggering a signaling cascade involving heightened Ca2+ influx and hyper-phosphorylation of JNK & FOXO3a, which ultimately impairs Eomes-directed IFN-γ production in Th1 cells. Genetic/pharmacological of VNUT inhibition significantly potentiates Th1 effector functions against Listeria infection and transplanted tumors. These findings identify VNUT as a critical checkpoint in limiting Th1 immunity, coupling vesicular ATP transport to transcriptional control via the P2X7R-JNK-FOXO3a-Eomes axis, offering a target for treating infection and cancer.

overall_design: RNA-seq profiling of wild-type and Vnut knockout Th1 cells at the differentiation in four days

first: Bolong

last: Wu

first: Linlin

last: Sheng


title: STAT3 regulates pathogen control and clearance in macrophages by reprogramming inflammation-induced microbial defense response

status: Public on Nov 22 2025

city: Houston

first: Xiaofeng

last: Zheng

state: TX

address: 7007 Bertner Ave

department: Bioinformatics & Computational Biology

country: USA

institute: UT MD Anderson Cancer Center

type: Expression profiling by high throughput sequencing

summary: STAT3 is pivotal in controlling hyperinflammation and governing effector macrophage responses to inflammation, yet the precise mechanisms by which STAT3 mediates pathogen control and clearance remain unclear. Herein, we performed RNA sequencing in bone marrow derived macrophages (BMDMs) to investigate STAT3-mediated transcriptional regulation of LPS-induced inflammation. Genome-wide profiling revealed significant enrichment of type-I interferon (IFN-I) associated genes and the IFN Gamma response pathway in Stat3-deficient BMDMs in homeostasis. We observed significant reduction in the expression of pathogen response genes and secretion of cytokines and chemokines associated with pathogen clearance in Stat3-deficient BMDMs upon LPS stimulation. Using BM chimeric mice with 20% Stat3-deficient hematopoietic cells infected with Citrobacter rodentium, we show that STAT3 is essential for preventing bacterial dissemination to tissues and regulating production of cytokines and chemokines related to immune cell recruitment and bacterial control. Stat3-deficient BMDMs infected with C. rodentium showed impaired ability to kill intracellular bacteria and demonstrated dampened expression of pathogen response genes. Further, IL-6 blockade subdued the  intrinsic IFN signature in Stat3-deficient macrophages and rescued aberrant expression of  pathogen response genes upon LPS stimulation, suggesting STAT3 restrains an aberrant autocrine IL-6 signal that deregulates macrophage-mediated pathogen control. Our data identifies novel functions for STAT3 in orchestrating an optimal microbial defense response, by promoting bacterial control and clearance in macrophages.

overall_design: To investigate the roles for STAT3 in regulation of macrophages in homeostasis and infection, we performed RNA-sequencing (RNA-seq) analyses of bone marrow-derived marophages (BMDMs) derived from CreER (wild-type controls) and CreER Stat3f/f (Stat3-deficient) bone marrow cells. BMDMs were treated with PBS or lipopolysaccharide (LPS) at two timepoints and three biologically independent samples for each treatment group and genotype were processed for RNA-seq.

first: Bhakti

last: Patel

first: Jing

last: Wang

first: Stephanie

last: Watowich


title: The sequencing results of biosensor-guided genome-wide mutagenesis using a transposon

city: Gwangju

first: YongHee

last: Han

address: 77 Yongbong-ro

department: School of Biological Sciences and Technology

country: South Korea

institute: Chonnam National University

summary: The economic viability of bio-based chemical production relies on microbial strain improvement, which optimizes titer, yield, and productivity of the process. This improvement necessitates redirecting cellular metabolism towards target molecule synthesis, requiring not only heterologous gene introduction but also endogenous genomic modifications to reconfigure metabolic networks originally optimized for cellular growth. However, identifying engineering targets is complex due to the intricate nature of metabolic networks and the cumulative, interactive effects of multiple genes on compound production. Consequently, there is a critical need for technologies capable of elucidating unpredictable engineering targets and synergistic gene combinations that enhance product formation in microbial strains. This study presents an innovative, integrated methodology named Tn-MAGE for rapidly identifying unpredictable engineering targets and synergistic gene combinations to enhance microbial strain performance in compound production. Our approach synergistically combines two key strategies: (1) integration of Tn-seq with biosensor-assisted ALE in a single batch culture, and (2) utilization of MAGE for combinatorial knockout library creation coupled with biosensor-assisted high-throughput screening. This integrated Tn-MAGE methodology offers a powerful tool for discovering unpredictable targets across the genome and their combinations, facilitating the development of high-performing strains for bio-based production of valuable compounds. The versatility of this approach suggests its potential applicability in various metabolic engineering contexts, promising significant advancements in strain improvement strategies.

overall_design: We cultivated the E. coli BL21*(DE3)ΔsucCΔfumC strain with a production plasmid harboring naringenin biosynthetic pathways, a naringenin sensor plasmid containing the tetA gene as a reporter, and a single-copy plasmid carrying Himar1C9. The naringenin sensor couples naringenin production with cell growth, enabling naringenin-overproducing strains to become the dominant population under tetracycline conditions. When transposition occurs, and thereby nonessential genes for naringenin production or genes in competing metabolic pathways are properly knocked out, strains can enhance naringenin production, becoming enriched in the population. Using next-generation sequencing, we identified knockout gene candidates beneficial for naringenin overproduction and validated their effects.

first: Jaeseong

last: Hwang

first: Yong Hee

first: Sang Woo

last: Seo

first: Sungho

last: Jang

first: Gyoo Yeol

last: Jung

first: Ina

last: Bang

first: Joon Young

last: Park

first: Donghyuk

last: Kim


title: Cellular and molecular mechanisms of Resmetirom in metabolic dysfunction-associated steatohepatitis (MASH): A transcriptomic profiling study

status: Public on Nov 21 2025

city: Hefei

first: Chenyang

last: He

address: 443 Huangshan Road

country: China

institute: University of Science and Technology of China

summary: This study demonstrates that the thyroid hormone receptor-β (THR-β) agonist Resmetirom, the first FDA-approved anti-MASH drug, exerts potent therapeutic effects in a human relevant dietary mouse model of metabolic dysfunction–associated steatohepatitis (MASH) induced by fructose, palmitate, and cholesterol (FPC). Resmetirom treatment markedly improved hepatic steatosis, inflammation, and fibrosis, accompanied by reduced systemic inflammatory markers and hepatocyte death. Transcriptomic profiling revealed that Resmetirom profoundly reprograms hepatic gene expression, particularly suppressing pathways involved in lipid metabolism, inflammation, and multiple forms of programmed cell death, including apoptosis, pyroptosis, and necroptosis. These findings highlight a previously unrecognized role of THR-β activation in regulating hepatocyte survival and inflammation, providing mechanistic insight into the multifaceted hepatoprotective effects of Resmetirom in MASH.

overall_design: A diet-induced metabolic dysfunction–associated steatohepatitis (MASH) model using a fructose, palmitate, and cholesterol (FPC) diet. Mice were maintained on the FPC diet for 12 weeks, with therapeutic administration of Resmetirom (Rem) (5 mg/kg/day) initiated after 4 weeks of feeding and continued for an additional 8 weeks via intraperitoneal injection. The vehicle consisted of 0.5% carboxymethyl cellulose-sodium (CMC-Na) and 0.4% Tween-80.

first: Zhihua

first: Jianping

last: Weng

first: Suowen

last: Xu


title: Whole transcriptomics data under different extracellular matrix stiffness in NB cells

first: Miao

last: Chen

address: 37 Xueyuan Road, Haidian District

institute: Beihang University

summary: Nuclear pore complex (NPC), the gatekeepers of macromolecules across the nuclear envelope (NE), undergoes dynamic changes pivotal in physiology and pathology.  Our study established ECM stiffness as a key driver of NB cell migration through a novel lamin A/C/E2F4/PLK1 axis, which governed post-mitotic NPC assembly and nucleocytoplasmic transport function. Targeting NPC-mediated transport may represent a promising therapeutic strategy for NB.

overall_design: Analyzing whole transcriptomic profiles across different stiffness conditions aims to investigate how substrate stiffness influences gene expression and signaling pathways in neuroblastoma cells.

first: Guozhu

last: Sun

first: Chen

last: Miao

first: Wenhao

last: Tian

first: Jialong

last: Cui

first: Zhongqian

last: Liu

first: Qingping

last: Yao

first: Kai

last: Huang


title: Transcriptional profiling of Slc7a7-deficient bone marrow derived macrophages subjected to in vitro LDL loading

city: Kuopio

first: Tiit

last: Örd

address: Neulaniementie 2

country: Finland

institute: A.I. Virtanen Institute for Molecular Sciences, University of Eastern Finland

summary: Atherosclerotic coronary artery disease (CAD) development encompasses changes in immune cell populations, including macrophages. In atherosclerotic plaques, macrophages engulf LDL, becoming lipid-laden foam cells. Human and mouse atherosclerosis single-cell RNA-Seq datasets revealed Slc7a7 as a gene expressed specifically in macrophages, particularly in the lipid-associated macrophage subpopulation. Here, we investigate the gene expression profiles of Slc7a7-deficient (Slc7a7-/-) and Slc7a7+/+ bone marrow-derived macrophages exposed to either acetylated or oxidized LDL in vitro.

overall_design: Bulk RNA-Seq of Slc7a7 knockout or control mouse bone marrow-derived macrophages (BMDM) treated with acetylated LDL (AcLDL) or oxidized LDL (OxLDL).

first: Susanne

last: Bodoy

first: Judith

last: Giroud Gerbetan

first: Manuel

last: Palacin

first: Minna

middle: U

last: Kaikkonen


title: Staphylococcal Enterotoxin A Shapes Monocyte Transcription and Macrophage Polarization: Implications for Immune Responses in Infection and Inflammation

city: Stockholm

first: David

last: Brodin

address: Hälsovägen 7-9

department: Department of Biosciences and Nutrition

country: Sweden

institute: Karolinska Institute

summary: Staphylococcal enterotoxins (SE) crosslink the MHC-II on antigen-presenting cells (APC) with the T-cell receptor, inducing a polyclonal T-cell response. Although APCs are the initial targets of SE and are critical in shaping subsequent T-cell activation, the effects of SE on APC function remain poorly understood. This study investigates the immunomodulatory effects of staphylococcal enterotoxin A (SEA) on monocytes and their differentiation into monocyte-derived dendritic cells (moDC) or macrophages (MDM). Transcriptomic analyses of human monocytes via RNA sequencing revealed SEA-induced enrichment of gene pathways associated with inflammation, infection and dermatitis, effects that were amplified in the presence of T-cells. Phenotypic and functional characterization showed that SEA-primed monocytes differentiated into MDM with an altered polarization, deviating from classical M1/M2 pathways. SEA-primed MDM exhibited downregulayion of key markers including HLA-DR, CD80, CD86 and PD-L1. Functional assays demonstrated that SEA-primed MDM pushed hyperinflammatory T-cell responses, with significantly enhanced proliferation and IFN-γ secretion. In contrast, moDC retained robust antigen-presenting capabilities even upon SEA-priming. These findings provide mechanistic insights into SEA-mediated immune modulation, illustrating how SEA reprograms MDM functions and amplify proinflammatory T-cell responses. This advances our understanding of superantigen-driven immune interactions, offering a foundation for developing therapeutic strategies to mitigate superantigen-mediated immune conditions.

overall_design: RNA-seq profiling of human monocytes either unstimulated or stimulated with SEA or S. aureus cell-free supernatants for 24h

first: Claudia

last: Arasa

first: Khaleda

last: Rahman Qasi

last: Mata Forsberg

first: Eva

last: Sverremark-Ekström


title: Human fallopian tube epithelial organoids with TP53 mutation recapitulate features of serous tubal intraepithelial carcinoma (STIC) -LP-WGS

city: Stanford

first: Bo

last: Yu

state: CA

address: 1291 Welch Road, Biomedical Innovations Building Room 2700 Room 2700

department: OB/GYN

institute: Stanford University

summary:
Human fallopian tube epithelial organoids with TP53 mutation recapitulate features of serous tubal intraepithelial carcinoma (STIC)
Judith Kraiczy, Bo Yu
Gynecologic Oncology, Volume 203, 2025, Pages 198-208, ISSN 0090-8258,
https://doi.org/10.1016/j.ygyno.2025.10.038.

ABSTRACT
Objective Serous tubal intraepithelial carcinoma (STIC) is the immediate precursor lesion for high-grade serous ovarian carcinoma (HGSOC) and harbors universal TP53 mutations. The lack of an appropriate in vitro model for STIC presents a major challenge in studying its pathogenesis. We aimed to develop a human in vitro model that mimics STIC lesions. Methods Using CRISPR-Cas9 gene editing, we generated human fallopian tube epithelial organoids with TP53 loss-of-function mutations (TP53-/- FTOs). We characterized TP53-/- FTOs on a cellular and molecular level using immunofluorescence confocal imaging, copy number variation (CNV) analysis, and RNA sequencing. Results TP53-/- FTOs recapitulated key features of STIC lesions. They exhibited increased proliferation and nuclear abnormalities, including nuclear enlargement and atypical mitotic figures. Copy number variation analysis revealed aneuploidy in some TP53-/- FTOs. Compared to unedited controls, TP53-/- FTOs demonstrated significant transcriptomic changes, including the downregulation of DNA repair genes and upregulation of epithelial-mesenchymal transition (EMT) pathways. Similar to STIC lesions, TP53-/- FTOs showed a marked reduction in ciliated cells and ciliogenesis-associated gene expression. Conclusions These findings suggest that p53 loss in FTOs promotes a proliferative and genomically unstable state that is conducive to carcinogenesis. The TP53-/- FTO model we have generated provides a valuable tool for studying early events in ovarian carcinogenesis and for developing new strategies for the early detection and prevention of ovarian cancer.

overall_design: Low-pass whole genome sequencing of human fallopian tube organoids with TP53 loss-of-function mutations (TP53-/- FTOs) and controls, as well as longitudinal samples of these organoids at a later passage

last: Kraiczy


title: Postnatal maternal care impacts hypothalamic Esrrg gene expression, co-expression profiles, and the DNA methylome in prenatal bisphenol-exposed rats [BiSulfite-seq]

city: Austin

first: Frances

middle: Anne

last: Champagne

state: Texas

address: 108 E Dean Keeton St

department: Psychology

institute: University of Texas at Austin

type: Methylation profiling by high throughput sequencing

summary: Background: Environmental exposures co-occurring during early life have a profound influence on neurodevelopment. Our previous work in rats suggests that postnatal maternal care modulates the effects of prenatal exposure to bisphenols, an estrogenic endocrine disrupting chemical, on offspring neurodevelopment. Elevated postnatal maternal licking/grooming and prenatal bisphenol exposure have known opposing effects on estrogen receptor alpha (Esr1) expression in the medial preoptic area (MPOA) of the hypothalamus, which could impact expression of estrogen-responsive genes. Based on this previous work, we hypothesized that postnatal maternal licking/grooming would mitigate the effects of prenatal bisphenol exposure on Esr1 expression and estrogen-responsive genes in the developing MPOA. In addition, we hypothesized that there would be interactive effects of prenatal bisphenol exposure and postnatal maternal licking/grooming on DNA methylation, particularly nearby estrogen responsive elements.  Results: Our results indicated a significant interaction between prenatal bisphenol exposure and maternal postnatal licking/grooming on estrogen-related receptor gamma (Esrrg) expression in female pups. These interactions were also evident in co-expression gene profiles in female pups; the majority of which were enriched for estrogen-responsive genes. Finally, DNA methylation analyses indicated that adding postnatal maternal licking/grooming as a covariate influenced the number of differentially methylated regions for prenatal bisphenol-exposed male and female pups. These differentially methylated regions were enriched for binding sites for transcription factors that are known to interact with estrogen receptors, suggesting some secondary effects on postnatal gene regulation.  Conclusions: These results suggest a novel biological mechanism in which postnatal maternal care can mitigate the negative neurodevelopmental impacts of prenatal bisphenol exposure.

overall_design: Reduced Representation Bisulfite Sequencing of postnatal 10 medial preoptic area of rat pups prenatally exposed to corn oil (control), 50 ug/kg/day BPA, 50 ug/kg/day Mixed Bisphenols (BPA, BPS, BPF), and 150 ug/kg/day Mixed Bisphenols (BPA, BPS, BPF). Maternal licking/grooming from postnatal days 1-5 were analyzed as a covariate. All analyses were stratified by offspring sex.

first: Samantha

middle: C

last: Lauby

first: Dennis

last: Wylie

first: Hannah

middle: E

last: Lapp

first: Melissa

last: Salazar

first: Amy

last: Margolis

middle: A


title: Human fallopian tube epithelial organoids with TP53 mutation recapitulate features of serous tubal intraepithelial carcinoma (STIC)- RNA-sequencing

summary:
Human fallopian tube epithelial organoids with TP53 mutation recapitulate features of serous tubal intraepithelial carcinoma (STIC),
Judith Kraiczy, Bo Yu,
Gynecologic Oncology, Volume 203,2025, Pages 198-208, ISSN 0090-8258,
https://doi.org/10.1016/j.ygyno.2025.10.038.

SUMMARY
Objective Serous tubal intraepithelial carcinoma (STIC) is the immediate precursor lesion for high-grade serous ovarian carcinoma (HGSOC) and harbors universal TP53 mutations. The lack of an appropriate in vitro model for STIC presents a major challenge in studying its pathogenesis. We aimed to develop a human in vitro model that mimics STIC lesions. Methods Using CRISPR-Cas9 gene editing, we generated human fallopian tube epithelial organoids with TP53 loss-of-function mutations (TP53-/- FTOs). We characterized TP53-/- FTOs on a cellular and molecular level using immunofluorescence confocal imaging, copy number variation (CNV) analysis, and RNA sequencing. TP53-/- FTOs recapitulated key features of STIC lesions. They exhibited increased proliferation and nuclear abnormalities, including nuclear enlargement and atypical mitotic figures. Copy number variation analysis revealed aneuploidy in some TP53-/- FTOs. Compared to unedited controls, TP53-/- FTOs demonstrated significant transcriptomic changes, including the downregulation of DNA repair genes and upregulation of epithelial-mesenchymal transition (EMT) pathways. Similar to STIC lesions, TP53-/- FTOs showed a marked reduction in ciliated cells and ciliogenesis-associated gene expression. Conclusions These findings suggest that p53 loss in FTOs promotes a proliferative and genomically unstable state that is conducive to carcinogenesis. The TP53-/- FTO model we have generated provides a valuable tool for studying early events in ovarian carcinogenesis and for developing new strategies for the early detection and prevention of ovarian cancer.

overall_design: RNA sequencing of human fallopian tube organoids with TP53 loss-of-function mutations (TP53-/- FTOs) and controls


title: A non-toxic, abscisic acid-based anchor-away system for efficient nuclear depletion of budding yeast proteins

city: Edinburgh

first: Sander

last: Granneman

address: Mayfield Road, Kings Buildings, Waddington building, room 3.06

department: Centre for Synthetic and Systems Biology

country: United Kingdom

institute: University of Edinburgh

summary: With the aim of overcoming some of the problems associated with yeast genetic depletion systems, we developed a modified anchor-away (AA) system that relies on a chemically induced dimerization system involving the plant phytohormone abscisic acid (ABA). An advantage of employing this CID in an anchor-away system is that ABA is inexpensive, non-toxic to humans and it is easy to reverse the effect of ABA by simply washing out the hormone. Here we demonstrate that the ABA CID can be effectively employed as an AA system in Saccharomyces cerevisiae. We tested the ABA-AA with three proteins that accumulate at very different levels in the cell. We found that even for a highly abundant and essential proteasome component we were still able to get a ~50% decrease in cell growth upon ABA treatment. Compared to the rapamycin AA system, higher concentrations of the chemical is needed to observe a phenotypical effect. Nevertheless, we show that even at these high concentrations, ABA does not noticeably impair cellular growth nor does it cause major changes in gene expression in exponentially growing yeast. We propose that the ABA anchor-away system is highly complementary to existing tools but has no toxicity issues. Because of its simplicity, we anticipate it will be broadly useful.

overall_design: Developing a new anchor-away system for removing yeast proteins from the nucleus.

first: Hanne

last: Grosemans


title: Multi-omics analysis identifies a microbiota–bile acid–TLR signaling axis driving bladder injury in interstitial cystitis

city: Chengdu

first: Lede

last: Lin

state: Sichuan

address: No.37 Guoxue Lane, Wuhou District, Chengdu, Sichuan, P. R. China

institute: West China Hospital of Sichuan University

summary: Hunner-type interstitial cystitis/bladder pain syndrome (HIC) is a complex and clinically challenging chronic disorder characterized by bladder-centered symptoms, including persistent pelvic pain and urinary urgency frequency, with elusive pathogenesis involving poorly defined multi-system interactions and systemic implications. To investigate upstream drivers of urothelial injury, we employed an integrative multi-omics approach, incorporating metagenomic sequencing, targeted metabolomics of urine and serum, and single-cell RNA sequencing. Our analysis identified Enterococcus avium, Megasphaera micronuciformis, and Fusobacterium nucleatum as the significantly enriched species in HIC patients, with bile acid metabolism appearing as a prominently altered pathway. Metabolomic profiling revealed the elevated levels of taurochenodeoxycholic acid (TCDCA) and tauroursodeoxycholic acid, while single-cell transcriptomic data pointed to TLR3 signaling as a key mediator of urothelial injury. Cross-omics correlation proposed a mechanistic axis involving E. avium, TCDCA, and TLR3 activation. Functional validation using fecal microbiota transplantation, E. avium transplantation, and TCDCA administration demonstrated that this axis contributes to the urothelial barrier disruption and inflammatory responses via TLR3. Intervention with a TLR3 inhibitor demonstrated therapeutic potential. These findings shed light on the pathogenesis of HIC, highlighting the potential role of the gut-bladder axis in disease development. This may contribute not only to the discovery of novel biomarkers but also to the development of personalized treatment strategies targeting specific molecular and microbial pathways.

overall_design: Single-cell RNA sequencing was performed from bladders of 10 Hunner-type IC/BPS patients and 7 PSUI controls.

first: Liao

last: Peng

first: Jia-wei

last: Chen

first: Yuan-zhuo

first: Xing-peng

last: Di

first: Le-de

first: Bo-ya

last: Li

first: Chi

last: Zhang

first: Wei

last: Wang

first: Xiao-shuai

last: Gao

first: Yu-cheng

last: Ma

first: Si-hong

last: Shen

first: Hai-rui

first: Xiao-fang

last: Xu

first: Xiao

last: Zeng

first: Hong

first: Qun

last: Sun

first: Tao

last: Jin

first: De-yi

last: Luo


title: Distinct contributions of Etv2+ and Flk1+ progenitors to endothelial, hematopoietic, and cardiac lineages [RNA-seq]

city: St. Louis

first: Dereck

last: Alleyne

state: MO

address: 425 South Euclid Avenue

department: Department of Pathology and Immunology

institute: Washington University School of Medicine

summary: The ETS family transcription factor ETV2, VEGFA and its receptor FLK1 are essential for hematopoietic, vascular and cardiac development. Here, we combine dual Etv2 and Flk1 lineage tracing with molecular profiling to define how mesoderm progenitors are allocated to hematopoietic, endothelial, cardiomyocyte and smooth muscle lineages. We demonstrate that hematopoietic, endothelial, and cardiac valves arise from dual Etv2+ and Flk1+ lineages and that Etv2+ and Flk1+ mesoderm contributing to the hemangiogenic fate is molecularly distinct from those generating muscle. Mechanistically, we show that ETV2 cooperates with the BAF chromatin remodeling complex to establish accessibility at ETV2 target loci. Loss of Baf155 expression reduces chromatin accessibility at ETV2 target loci and impairs hemangiogenic lineage specification. This work defines lineage relationships and the molecular circuitry underlying hemangiogenic specification during cardiovascular development.

overall_design: RNA-seq profiling of day 4 embryoid bodies (EBs) derived from wildtype, iEtv2 and T-GFP;Etv2-tdTomato mouse ES cells. Etv2 overexpression was induced by treating day 2.5 EBs with 2ug/ml doxycline. EBs were sorted by FACS to sort PDGFRalpha+FLK1+ (DP) and PDGFRalpha-FLK1+ (SP) mesodermal cells.

first: Alleyne

last: Dereck

first: Kim

last: Minseo

first: Wu

last: Jun

first: Kwon

last: Yoojung

last: Ye-Ram

first: Ul Kabir

last: Ashraf

first: Ishahak

last: Matthew

first: Millman

last: Jeffrey R.

first: Fan

last: Changxu

first: Lee

last: Hyung Joo

first: Krchma

last: Karen

first: Xing

last: Xiaoyun

first: Lavine

last: Kory

first: Wang

last: Ting

first: Choi

last: Kyunghee


title: Distinct contributions of Etv2+ and Flk1+ progenitors to endothelial, hematopoietic, and cardiac lineages

type: Genome binding/occupancy profiling by high throughput sequencing

overall_design: ATAC-seq profiling of day 4 embryoid bodies (EBs) derived from wildtype, Etv2 overexpressing and Etv2KO mouse ES cells. Etv2 overexpression was induced by treating day 2.5 EBs with 2ug/ml doxycline. EBs were sorted by FACS to sort PDGFRalpha+FLK1+ (DP) and PDGFRalpha-FLK1+ (SP) mesodermal cells.


title: TGFα is Required for Hair Follicle Function During Aging and its Loss Leads to Progressive Alopecia

city: Charlestown

first: Alexander

last: Marneros

state: MA

address: 13th Street

institute: Massachusetts General Hospital

summary: Gene expression changes in the epidermis of mice lacking Tgfa

overall_design: Comparative expression analysis of total RNA by RNA-Seq. Bulk RNA-seq of isolated epidermis from back skin of 22 months-old WT and Tgfa KO mice.

middle: G


title: Postnatal maternal care impacts hypothalamic Esrrg gene expression, co-expression profiles, and the DNA methylome in prenatal bisphenol-exposed rats [3' Tag-seq]

first: Frances

last: Champagne

overall_design: 3' Tag-seq of postnatal 10 medial preoptic area of rat pups prenatally exposed to corn oil (control), 50 ug/kg/day BPA, 50 ug/kg/day Mixed Bisphenols (BPA, BPS, BPF), and 150 ug/kg/day Mixed Bisphenols (BPA, BPS, BPF). Maternal licking/grooming and nest attendance observed from postnatal days 1-5 were analyzed as potential moderators. All analyses were stratified by offspring sex.

first: Samantha

last: Lauby

first: Dennis

last: Wylie

first: Hannah

last: Lapp

first: Melissa

last: Salazar

first: Amy

last: Margolis


title: Acute Kidney Injury Following Fatty Liver Ischemia-Reperfusion Injury

city: Indianapolis

first: Wenjun

last: Zhang

state: IN

address: 1044 W. Walnut Street, R4 W116

institute: Indiana University School of Medicine

summary: To understand the roles of ferroptosis in subsequent acute kidney injury (AKI) following hepatic ischemia-reperfusion injury (hIRI) in steatosis livers. The hIRI was induced in mice fed either a high-fat, high-sucrose diet (HFD) or a normal diet (ND)  to mimic the AKI commonly observed clinically after fatty liver transplantation. RNA sequencing was conducted to investigate the underlying mechanisms.  We found that apoptosis and inflammation are the prominent kidney injury mechanisms following fatty liver IRI. Although ferroptosis may not be directly involved in the renal injury, anti-ferroptosis intervention mitigates AKI, supporting the concept that ferroptosis-mediated liver injury may serve as the primary upstream trigger in this context.

overall_design: The 10-week-old C57BL/6 mice (n=3 in each group) were fed with either a control diet (ND) or a high-fat diet (HFD) for 12 weeks before being subjected to 1hr hepatic ischemia-reperfusion injury (IRI) or a sham procedure.  Total 4 groups: HFD_IRI, HFD_sham, ND_IRI, and ND_sham. The mouse liver and kidney tissue were harvested in 6 hrs after ischemia injury.

first: Zachary

last: Rokop

first: Chandan

middle: K

last: Sen

first: Chandrashekhar

last: Kubal

first: Hongyu

last: Gao


title: Eosinophils enhance granuloma-mediated control of persistent Salmonella infection

first: Daniel

last: Arve-Butler

address: 299 Campus Drive, FAIRCHILD BUILDING D33

summary: Salmonella enterica can persist asymptomatically within tissues for extended periods. This is achieved through intricate host-pathogen interactions in immune cell aggregates called granulomas, wherein Salmonella establish favorable cellular niches to exploit while the host limits its expansion and tissue dissemination. Here, using a mouse model of persistent Salmonella infection, we identify a host-protective role for eosinophils in control of Salmonella Typhimurium (STm) infection within the mesenteric lymph nodes (MLN), the main lymphoid tissue of STm persistence. Combining spatial transcriptomics and experimental manipulations, we found that monocytes and macrophages responding to STm infection recruited eosinophils in a C-C motif chemokine ligand 11 (CCL11)-dependent manner and enhanced their activation. The protein major basic protein, primarily expressed by eosinophils, was associated with altered macrophage polarization and bacterial control. Thus, eosinophils play a vital role in restraining Salmonella exploitation of granuloma macrophages at a key site of bacterial persistence.

overall_design: Sixteen Xenium in situ spatial transcriptomics samples from mouse mesenteric lymph node (MLN): Uninfected controls (UI1–UI2), WT infected at 4 wpi (WT1–WT8), and DblGATA1 infected at 4 wpi (GATA1–GATA6). Raw Xenium outputs (cell_feature_matrix.zarr.zip, cells.csv.gz, transcripts.parquet) deposited per sample. Processed outputs include an integrated Seurat object (RDS), per-cell cluster table (CSV), and marker tables (CSV).

middle: S

last: Butler

first: Blanda

last: Di Luccia

first: Jose

last: Vilches-Moure

first: Denise

middle: M

last: Monack


title: Early life microbiota skew long-term gene expression and epigenome of bone marrow hematopoietic precursors

city: Toronto

first: Wilder

last: Scott

state: ON

address: 2075 Bayview Ave

country: Canada

institute: SRI

summary: Asthma is the most prevalent chronic childhood disease in developed nations, yet its etiology remains incompletely understood. Emerging evidence suggest a crucial role for the gut microbiota in shaping immune development and future susceptibility to atopic disease. We previously demonstrated that short-chain fatty acids (SCFAs) as fermentation products of normal gut bacteria, regulate an innate type 2 immune axis that serves to dampen susceptibility to allergic lung inflammation. Here, we show that neonatal depletion of SCFA-producing gut bacteria via antibiotics robustly alters Lin⁻Sca-1⁺c-Kit⁺ (LSK) hematopoietic stem and progenitor cell (HSPC) output, resulting in a transplantable atopic immune phenotype:  bone marrow (BM) transplants from these mice generates recipients with increased lung B cell IgE expression, elevated serum levels of IgE, dowstream increased IgE bound to basophils and exacerbated allergic lung inflammation following papain challenge. Notably, neonatal depletion of SCFA bacteria also enhanced susceptibility to chemotherapy (5-fluorocuracil) -induced myelosuppression, marked by impaired neutrophil recovery and increased DNA damage in long-term HSCs. Importantly, these antibiotic effects were reversed by supplementation of antibiotic-treated mice with exogenous SCFA. Single-cell RNA sequencing (scRNA-seq) of LSK HSPCs revealed distinct shifts in gene expression following vancomycin treatment, including increased proliferation-associated signatures with stress-adaptive features. Epigenetic profiling further revealed differential histone acetylation in HSPCs, consistent with an SCFA-mediated epigenetic regulatory mechanism. Collectively, these findings establish an epigenetic link between gut microbiota composition, hematopoiesis, and long-term immune function, providing a mechanistic basis for microbiota-driven atopic disease susceptibility and hematopoietic dysfunction.

overall_design: Approximately 1 x 10^4 viable HSPCs (live-gatedCD45+Lin-Sca1+cKit+) were sorted (FACSAria II, BD Biosciences, San Jose, CA) from BM of 8-week-old male mice (3 mice per group) reared on vancomycin, vancomycin with BAP or regular drinking water. Sorted cells were encapsulated into droplets, and transcript libraries were prepared using Single Cell 3′ reagent kits (10x Genomics) and then sequenced using a 75 cycle NextSeq500. Samples were aligned using the mm10 reference transcriptome and the standard Cell Ranger count pipeline. Data analysis and visualization was performed using the R package Seurat

first: Ahmed

last: Kabil

first: Alissa

last: Cait

first: Michael

middle: R

last: Hughes

first: Kelly

last: McNagny


title: Transcriptomic changes upon doxorubicin treatment in hCMEC/D3

city: New Brunswick

first: Chadni

last: Patel

state: NJ

address: 195 Little Albany Street

institute: Rutgers Cancer Institute

summary: Current chemotherapy regimens have significantly improved overall survival for children with cancer. However, these curative treatments are associated with detrimental side effects such as chemotherapy-induced cognitive impairment (CICI), or “chemobrain.” Measurable deficits in cognitive function may persist years after chemotherapy treatments, significantly reducing quality of life. Specifically, doxorubicin (DOXO), a commonly used chemotherapeutic agent in curative regimens for children with cancer, plays a pivotal role in the development of CICI, even though it does not cross the blood brain barrier (BBB). We propose to address the poorly understood mechanism of DOXO-related CICI by studying changes induced by DOXO in BBB integrity in vitro, using human cerebral microvascular endothelial cells (hCMEC/D3). Our findings provide critical insights on how DOXO impacts the BBB and builds a foundation for developing preventative measures and therapeutic interventions that may improve the quality of life for patients.

overall_design: RNA-sequencing was performed to gain insights regarding the gene expression changes induced by doxorubicin (DOXO) treatment in human cerebral microvascular endothelial cells (hCMEC/D3). The hCMEC/D3 were treated 500 nM of doxorubicin (DOXO) or PBS, for 24 hours.

first: Peter

middle: D

last: Cole


title: Bulk RNA-seq time-course of human iPSc differentiation to thymic epithelial progenitors

city: Nantes

first: Matthieu

last: Giraud

address: 30 bd Jean Monnet

department: CRTI

country: France

institute: INSERM _ Université de Nantes

summary: Bulk RNA-seq was performed across a time-course of human iPSc differentiation toward thymic epithelial progenitors (TEPs). Twenty-nine experimental libraries spanning eleven time-points (days 0, 1, 2, 3, 7, 9, 10, 14, 15, 17, 22) plus four primary thymic epithelial cell (TEC) controls revealed progressive down-regulation of pluripotency genes and stepwise activation of transcriptional modules corresponding to anterior endoderm, third pharyngeal pouch endoderm, and thymic epithelial fate.

overall_design: Human induced pluripotent stem cells (iPSc; line LON71) were differentiated toward thymic epithelial progenitors (TEPs) using our defined iPSc-to-TEP protocol. RNA was extracted with the RNeasy Micro Kit (Qiagen) at eleven time-points: D0 (n = 3), D1 (n = 1), D2 (n = 1), D3 (n = 4), D7 (n = 5), D9 (n = 6), D10 (n = 4), D14 (n = 2), D15 (n = 1), D17 (n = 1), D22 (n = 1). Four primary pediatric thymic epithelial cell (TEC) samples served as references. All samples were sequenced using 3′ RNA-seq (DGE-seq).

first: Nathan

last: Provin


title: Circulating exosomal microRNA profiling of diabetic patients with and without Metabolic Dysfunction-Associated Steatohepatitis

city: Wuhan

first: Wenxi

last: He

address: No.1095, Jiefang Ave.

department: Department of Pharmacy

institute: Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology

type: Non-coding RNA profiling by high throughput sequencing

summary: This study was designed to identify non-invasive biomarkers for the diagnosis of MASH in patients with type 2 diabetes. The progression from simple steatosis to MASH in diabetic patients represents a critical clinical challenge, as it significantly increases the risk for cirrhosis and hepatocellular carcinoma. Current diagnostic methods are invasive and unsuitable for broad screening. We hypothesized that circulating exosomes contain a specific microRNA (miRNA) signature that can distinguish between diabetic patients with and without MASH. To test this, we isolated serum exosomes from both patient groups and performed small RNA sequencing to comprehensively profile their miRNA content. The goal of this dataset is to provide a resource of differentially expressed exosomal miRNAs that can serve as a foundation for the development of novel, non-invasive diagnostic tools for MASH in this high-risk population.

overall_design:
The experiment was designed as a case-control study to compare the exosomal miRNA profiles between two clinically defined groups.
Group 1 (Control): Consisted of 5 biological replicates from diabetic patients without MASH (DIA).
Group 2 (Case): Consisted of 5 biological replicates from diabetic patients with confirmed MASH (DIA+MASH).
Serum was collected from all 10 subjects. Exosomes were isolated from the serum of each individual patient. Total RNA was then extracted from each of the 10 exosome preparations, and individual small RNA sequencing libraries were constructed and sequenced. Each sample represents an independent biological replicate.

first: He

last: Zhu

first: Xiaoyu


title: IL-9 signaling redirects CAR T cell fate toward CD8+ memory and CD4+ cycling states, enhancing anti-tumor efficacy

city: Philadelphia

first: Regina

middle: M.

last: Young

state: Pennsylvania

address: 3400 Civic Center BLVD

department: CCI

institute: University of Pennsylvania

summary: Il9 signalling CAR-T phenotypes were explored by co-expressing IL9R on CAR-Ts and exposing them to antigen for 10 days followed by scRNA to elucidate the mechanism leading to increased expansion, persistence, and tumor infiltration

overall_design: Following chromic antigen exposure, IL9R co expressing CAR-T tells either exposed to IL9 or mock control underwent 10X scRNA sequencing

first: Sofía

last: Castelli

first: Wesley

middle: V

last: Wilson

first: Amanda

last: Finck

first: Sebastian

middle: J

last: Atoche

first: Mikko

last: Siurala

first: Carl

middle: H

last: June


title: Extracellular vesicles secreted from human myotubes of T2D and NGT donors

city: Oslo

first: Stian

last: Christiansen

address: Folke Bernadottes Vei 13D

country: Norway

institute: OsloMet

summary: Type 2 diabetes (T2D) is associated with increased morbidity, mortality, and substantial healthcare costs. Peripheral insulin resistance, involving interconnected dysregulation of multiple organs, is considered a major driver of T2D. Extracellular vesicles (EVs) are suggested as mediators of this dysregulation based on their properties in intercellular communication. Given the role of skeletal muscle in glucose metabolism, the content of skeletal muscle-derived EVs may provide insights into mechanisms of T2D.  To examine this, myotubes from severely obese female T2D donors and matched women with normal glucose tolerance (NGT) were cultured. Small EVs (sEVs) were isolated by differential centrifugation and filter columns and characterized by nanoparticle tracking analysis, flow cytometry, and transmission electron microscopy. The micro-RNA (miR) content of sEV was analyzed via Affymetrix GeneChip miRNA 4.0, while proteins were detected by LC-MS/MS.

overall_design: Small extracellular vesicles (sEVs) were isolated from primary human skeletal muscle cell cultures established from biopsies of 12 female donors (6 with type 2 diabetes, 6 with normal glucose tolerance). Cultures were differentiated into myotubes, conditioned media collected, and sEVs isolated and characterized by nanoparticle tracking analysis, transmission electron microscopy, and flow cytometry. sEV protein cargo was analyzed by nanoLC-MS/MS (Q Exactive, Thermo), and data were processed in MaxQuant with LFQ normalization. In parallel, sEV RNA was isolated and profiled using Affymetrix GeneChip miRNA 4.0 arrays, followed by RMA normalization and log2 transformation. One sample in the NGT group was excluded after quality control, yielding n=5 (NGT) and n=6 (T2D) for miRNA profiling, and n=5 per group for proteomic analysis. Differential expression was assessed with t-tests and multiple testing correction.

middle: F

first: Kari

middle: B

last: Haug

first: Misbah

last: Hussain

first: Abdille

last: Hussein

first: Berit

last: Brusletto

first: Reidun

last: Øvstebø

first: Oliwia

last: Witczak

first: Vigdis

last: Aas


title: iPSc-to-thymic epithelial progenitor differentiation under multifactorial DoE optimization and transcriptomic profiling

first: Matthieu

last: Giraud

summary: This study investigates the differentiation of human induced pluripotent stem cells (iPSc) into thymic epithelial progenitors (TEPs) through a combinatorial experimental design (Design of Experiments, DoE). Bulk RNA-seq was performed across 96 differentiation conditions covering key developmental transitions: definitive endoderm (DE), anterior foregut endoderm (AFE), third pharyngeal pouch endoderm (3PPE), and thymic epithelial progenitors (TEP). The multifactorial screen leveraged Plackett-Burman designs and transcriptomic similarity to in vivo pharyngeal development atlases to identify optimal differentiation cues.

overall_design: RNA was extracted at the AFE, 3PPE and TEP stages of directed differentiation from iPSc across 96 conditions generated from three Plackett-Burman designs (36+36+24 runs), corresponding respectively to the DE_to_AFE, the AFE_to_3PPE and the 3PPE_to_TEP transitions. Each condition combined low/high concentrations of pathway modulators (BMP, WNT, FGF, TGFβ, SHH, etc.) administered at defined time windows. Samples were sequenced using 3’ RNA-seq (DGE-seq) and analyzed via bulk transcriptomics to evaluate proximity to gene signatures from developmental single-cell atlases.

first: Nathan

last: Provin


title: NRDE2 interacts with transcription-associated and splicing factors and modulates gene expression and splicing

city: Montpellier

first: Rosemary

last: Kiernan

address: 141 rue de la cardonille

institute: UMR9002 CNRS-UM

summary: To investigate the function of Nuclear RNAi-Defective 2 (NRDE2) in gene expression, we performed RNA-seq following its transient knock-down. Differential expression analysis showed that loss of NRDE2 modulated the expression of thousands of genes.

overall_design: HeLa cells tranduced with shRNAs targeting NRDE2 (shNRDE2) or a non-targeting control (shControl) were harvested 5 days post-transduction. Total RNA was extracted using TRIzol. Paired end sequencing of polyA+ selected mRNA (20M clean reads per sample, 125 bp reads) was carried out.

first: Marina

last: Srbic

first: Raoul

last: Raffel

first: Alexandre

last: Heurteau

first: Xavier

last: Contreras


title: Selective elimination of donor bacteria enables global profiling of plasmid gene expression at early stages of conjugation

city: Gif sur Yvette

first: Elena

last: ESPINOSA

address: 1, Avenue de la Terrasse

institute: CNRS

summary: Conjugative plasmids are a major driving force for the dissemination of antimicrobial resistance. During conjugation, plasmid DNA is transferred from the donor cell as a single-stranded (ss) linear molecule. Recent researches highlighted intriguing DNA reactions at the early stage of conjugation that are important for plasmid establishment in the recipient cell, including prompt expression of anti-defense genes. However, genomics-based approaches to investigate plasmid establishment have been challenging, because identical, fully established plasmids in donor cells mask transconjugant-specific signals. To overcome this limitation, we developed a new EDTA method which exploits a donor mutant hypersensitive to hypoosmotic shock. It allows unprecedently quick and efficient Elimination of Donor population for Transconjugant Accumulation. RNAseq analysis revealed highly selective and robust induction of plasmid genes during early stage of conjugation. We experimentally identified +1 sites of 6 possible operons that are matched with ssDNA promoter predictions. We also showed gene expression program is altered in different recipient cells in which plasmid establishment is perturbed. As the EDTA method is straightforward and broadly applicable, it will further understanding of the plasmid establishment processes in new host cell, not only gene expression but also ss to double-stranded DNA conversion and plasmid circularization.

sample_organism:
Sinorhizobium meliloti
Escherichia coli

platform_organism:
Sinorhizobium meliloti
Escherichia coli

overall_design: RNA-seq of recipient cells after 30 minutes or 60 minutes conjugation of pESBL and F plasmid

first: Meng

last: Wen

last: Fraikin

first: Emma

last: Mettouchi

first: Christian

last: Lesterlin

last: Espinosa

first: Yoshiharu

last: Yamaichi


title: Mouse sperm: Control vs Cd treatment

city: Hefei

first: Jin

last: Zhang

address: Anhui Medical University, No. 81 Meishan Road, Shushan District, Hefei City, Anhui Province, China

institute: public health

type:
Expression profiling by array
Non-coding RNA profiling by array

summary: Compare the RNA m6A methylation profiles of sperm from control-treated mice and cadmium (Cd)-treated mice. The aim is to investigate the impact of the common environmental toxicant Cd on RNA m6A modification levels in sperm, and whether such alterations impair the healthy development of offspring

overall_design: Two-condition experiment, control-treated mice vs. Cd-treated mice. Biological replicates: 3 control replicates, 3 Cd replicates.

first: Yong-Wei

last: Xiong


title: Rb1 deficiency induces synthetic lethality with ATR and PKMYT1 co-inhibition in breast cancer

city: Houston

first: Khandan

last: Keyomarsi

address: 1515 Holcombe Blvd

department: Experimental Radiation Onc

institute: University of Texas - MD Anderson Cancer Center

summary: Targeting cell cycle checkpoints has emerged as a promising strategy in cancer therapy, yet single-agent inhibitors often fail due to compensatory mechanisms. Here, we demonstrate that co-inhibition of ATR (RP-3500) and PKMYT1 (RP-6306) induces synthetic lethality in Rb1-deficient breast cancers by disrupting both S/G2 and G2/M checkpoints, resulting in replication stress, premature mitotic entry, and DNA damage accumulation. In vitro, Rb1-deficient breast cancer cell lines exhibited marked apoptosis and loss of clonogenic potential, whereas Rb1-proficient models remained largely resistant to combination treatment. Genetic manipulation of Rb1 confirmed this dependency: Rb1 knockdown sensitized resistant models, and overexpression conferred protection. In vivo, patient-derived xenograft (PDX) models recapitulated these findings. Rb1-deficient tumors underwent complete regression, while Rb1-proficient tumors showed limited response with RP-3500/RP-6306 treatment. Combination therapy was well tolerated in vivo, without significant toxicity or weight loss. Biomarker analysis revealed increased γH2AX and reduced Ki67 staining exclusively in Rb1-deficient PDX models, underscoring the specificity of this response. Mechanistically, Rb1 loss impaired double-strand DNA repair by attenuating homologous recombination and non-homologous end joining, leading to replication fork collapse, chromosomal instability, and mitotic catastrophe. Proteogenomic analysis identified JNK/p38 stress response pathway activation as a key driver of apoptosis following ATR/PKMYT1 inhibition in Rb1-deficient cells. Clinically, analysis of stage IV breast cancer patient datasets revealed that Rb1-low tumors display reduced DNA repair pathway activity and are enriched in triple-negative and CDK4/6 inhibitor-resistant luminal breast cancers. These findings establish Rb1 loss as a functional biomarker for ATR/PKMYT1-targeted therapy, offering a precision treatment strategy for advanced breast cancers.

overall_design: Breast cancer cell lines MDA-MB-231, BT-549, MCF-7, MCF-7 PalbociclibResistant (MCF7PR) were collect at either baseline (DMSO) or under treatment 30 nM RP6306 (MYTi), 10 nM RP3500 (ATRi) or in combination (COMB) for 48 hours and harvest for total RNA and RNA sequencing. Each group havs at least three biological replicates.

first: Xiao-Ting

last: Jiang

first: Mei-Kuang

last: Chen

first: Jiwang

last: Lee

first: Xiayu

last: Rao

first: Jing

last: Wang


title: LowNitrogen and beneficial bacterial effects on Arabidopsis grown on root and shoot.

first: Véronique

last: BRUNAUD

address: avenue des sciences

institute: INRAE

summary: projet 2023_15_qs_sa187-what are the genes upregulated by SA187 in Arabidopsis thaliana under low N conditions.-comparison of plants inoculated or not with a beneficial bacterium under high and low N conditions

overall_design: 24 samples with 3 biological replicats. tissue comparison,treated vs untreated comparison

first: José

last: Caius

first: Christine

last: Paysant-LeRoux

first: Axel

last: De Zelicourt

first: Amina

last: Ilyas


title: The deubiquitinase OTUD1 orchestrates cisplatin chemosensitivity of non-small cell lung cancer through destabilizing RAD23B/XPC

city: tianjin

first: meixi

last: wang

address: Tianjin Sports Institute North Road Huanhu West Road

institute: Tianjin Medical University Cancer Hospital

summary: Lung cancer, especially the subgroup non-small cell lung cancer (NSCLC), continues to be one of the most commonly diagnosed cancers and the leading cause of cancer-related deaths worldwide. Cisplatin has long been a cornerstone of chemotherapy and has improved the prognosis for NSCLC patients. However, its overall efﬁcacy remains unsatisfactory, and patients ultimately develop drug resistance. Uncovering the underlying mechanism and identifying potential target to enhance cisplatin chemosensitivity is urgent. In this study, we uncovered that OTU deubiquitinase 1 (OTUD1) plays an important role in orchestrating cisplatin chemosensitivity of NSCLC. We found that promoter methylation resulted in downregulation of OTUD1 and the downregulated OTUD1 signiﬁcantly associates with cisplatin resistance and poor prognosis in NSCLC. Overexpression of OTUD1 enhances cisplatin sensitivity in vitro and in vivo. Mechanistically, OTUD1 promoted the degradation of RAD23B-XPC complex, which is the critical factor for nucleotide excision repair to remove cisplatin-induced DNA adducts, leading to cisplatin-induced cell death. OTUD1 cleaves the K63-linked ubiquitin chain of RAD23B and XPC, and enhances PRKN mediated K48-linked ubiquitination of RAD23B-XPC and the subsequent proteasomal degradation. The ﬁndings of this study highlighted that OTUD1 could be a potential therapeutic target for NSCLC.

overall_design: To investigate the mechanism of cisplatin (CDDP) resistance in non-small cell lung cancer, we have constructed a cisplatin resistant cell line. For RNA-seq, comparative gene expression profiling analysis of RNA-seq data for parent and CDDP resistant H460 cells.


title: Harnessing the E3 ligase SPOP for targeted degradation of the NUP98::KDM5A fusion oncoprotein

city: Vienna

first: Thomas

last: Eder

address: Veterinaerplatz 1

department: Department for Biological Sciences and Pathobiology

country: Austria

institute: University of Veterinary Medicine, Vienna

summary: NUP98-rearranged (NUP98-r) acute myeloid leukemia (AML) is associated with poor clinical outcomes and represents a major therapeutic challenge due to the lack of strategies for the direct elimination of NUP98-fusion oncoproteins. While targeted degradation of cancer-driving oncoproteins is an attractive strategy, the molecular mechanisms controlling NUP98 fusion oncoprotein stability are not known. Here, we used CRISPR/Cas9 screening to identify the E3 ligase SPOP as a direct regulator of NUP98 oncofusion stability and a novel tumor suppressor in NUP98-r AML. SPOP loss caused a selective increase of NUP98 fusion oncoprotein levels, and promoted leukemia cell proliferation. Leveraging this specificity, we show that induced proximity of SPOP and the NUP98::KDM5A fusion oncoprotein via a bioPROTAC approach induced full clearance of the fusion oncoprotein, and drove terminal differentiation and apoptosis of NUP98-r leukemia cells in vitro and in vivo. Altogether, our study identifies the SPOP E3 ligase as a direct regulator of NUP98 oncofusion stability and presents a blueprint for redirecting the ubiquitin proteasome system to selectively target cancer driving fusion oncoproteins.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design:
HEK293T cells: NUP98::KDM5A-expressing or wild-type HEK293T cells were subjected to 3' mRNA-sequencing to study effects of the introduction of NUP98::KDM5A.
GFP-NUP98::KDM5A-driven AML cells: Cells were transduced with SPOP-bioPROTAC or empty vector constructs. Doxycycline was added to cells to induce expression of the constructs. On days 3 and 6 post induction, cells were sent for RNA sequencing to assess changes to the transcriptional landscape upon bioPROTAC-mediated NUP98::KDM5A degradation.

first: Ecem

last: Kirkiz

first: Gabriel

last: Kaufmann

first: Simone

last: Bergqvist

first: Pablo

last: Fernandez Pernas

first: Laura

last: Quell

first: Melanie

last: Allram

first: Gabriele

last: Manhart

first: Wencke

last: Walter

first: Torsten

last: Haferlach

first: Florian

last: Grebien


title: High-Throughput Transcriptomic Profiling for Bioactivity Screening: Mapping Molecular Targets and Potencies for PFAS chemicals in HepaRG Cells

city: Research Triangle Park

first: Imran

last: Shah

state: NC

address: Mail Code: D143-02

department: Office of Research and Development

institute: US Environmental Protection Agency

summary: This study used high-throughput transcriptomics (HTTr) to evaluate the gene expression effects of 133 per- and polyfluoroalkyl substances (PFAS) in HepaRG cells. The PFAS chemicals were selected using a category-based prioritization approach developed by the U.S. Environmental Protection Agency (EPA) to ensure representation of structurally diverse compounds with relevance to human exposure and regulatory interest (Patlewicz et al., 2019; https://doi.org/10.1289/EHP4555). Differentiated HepaRG cells were exposed to each PFAS in concentration-response format (0.1 to 100 µM) for 24 hours. Transcriptomic changes were measured using the TempO-Seq platform (BioSpyder Technologies), enabling expression profiling of approximately 20,000 human genes. The resulting gene expression data were used to characterize the concentration-dependent molecular responses to PFAS exposure using connectivity mapping and to derive transcriptomic points of departure (tPODs). This dataset contributes to a broader effort to screen the bioactivity of environmental chemicals using HTTr in human liver-derived cells. It supports the use of transcriptomic profiling to identify bioactive PFAS, elucidate affected molecular pathways, and inform chemical hazard assessment.

overall_design:
Differentiated HepaRG cells were exposed to ~133 chemicals in an 8-point concentration-response format (0.1–100 µM) for 24 hours, and transcriptomic responses were measured using TempO-Seq.

***************************************************************
The table below lists SRA accessions for raw data already submitted to SRA under PRJNA1254200.
***************************************************************

first: Shah

last: Imran

first: Chambers

last: Bryant

first: Haggard

last: Derik

first: Word

last: Laura

first: Bundy

last: Joseph

first: Everett

last: Logan

first: Harrill

last: Joshua

first: Judson

last: Richard


title: Lung microenvironment effects on T cells following Francisella tularensis vaccination

city: Port Saint Lucie

first: Emily

last: Speranza

state: FL

address: 9801 SW Discovery Wy

department: Florida Research and Innovation Center

institute: Cleveland Clinic Lerner Research Institute

summary: Pulmonary infections often fail to produce long-lived immune memory and the underlying mechanism(s) remain unclear. We undertook a multiomics approach to compare adaptive immunity in two bacterial infections: Bordetella pertussis and Francisella tularensis. Both infections require CD4+ T cells, but B. pertussis results in long-lived memory responses whereas F. tularensis does not. Single cell RNA sequencing and flow cytometry revealed CD4+ T cells favored proliferation immediately after secondary B. pertussis infection but stagnated during F. tularensis challenge. Instead, rapid IFN-γ production by F. tularensis-elicited T cells induced IDO1 expression in endothelial cells, depleting pulmonary tryptophan and increasing kynurenine. This metabolic shift correlated with poor T cell proliferation and retention. Additionally, IDO1 knock-out resulted in larger T cell pools and improved survival during F. tularensis infection. Imaging confirmed the spatial association of immune cell centers and IDO1-expressing macrovascular endothelial cells underscoring the microenvironment’s impact on T cell function. Our study highlights the utility of multiomics in understanding cellular interactions within complex tissues.

overall_design: To generate immune animals, mice were intranasally inoculated with 1x105 CFU Bp or 150 CFU of LVS in 25 µL PBS delivered to a single nare after anesthetizing with ketamine/xylazine. Where indicated, mice were intranasally challenged with 1x106 CFU Bp or 25 CFU of Ftt in 25 µL PBS to a single nare after being anesthetized with ketamine/xylazine. Single cell suspensions were generated by mincing and then digested with Liberase TM (Sigma-Aldrich) for 45 minutes at 37°C. Red blood cells were lysed with ACK lysis buffer (Life Technologies). A TC20 automated cell counter (Bio-Rad) was used to determine the total number of viable cells using trypan blue exclusion. Samples were processed through the 10X Genomics V3.1 processing and prep of single-cell libraries

first: Lydia M

last: Roberts

first: Leanne

last: Arakkal

first: Tara

last: Wehrly

first: Claire

last: Poore Fonseka

first: Pavlina

last: Laskova

first: Benjamin

last: Schwarz

first: Eric

last: Bohrnsen

first: Ronald N

last: Germain

first: Catharine M

last: Bosio


title: CD205+ iPSc-derived thymic epithelial progenitor cells

first: Matthieu

last: Giraud

summary: We employed an ultra-sensitive bulk RNA sequencing approach to study thymic epithelial progenitor cells (TEPs) derived from iPSc differentiation and sorted by FACS based on their CD205 expression levels.

overall_design: iPSc were differentiated using our DoE based protocol. The CD205+ fraction of the resulting TEPs was analyzed using SMARTER bulk RNA-seq

first: Nathan

last: Provin


title: Detaching cells from human iPSc-derived thymic organoids (hTOs) seeded with primary early thymic progenitors (ETPs) at late phase of maturation (D28)

summary: We have employed a single-cell sequencing approach using 10x Genomics scRNA-seq to study hematopoietic cell generation from hTOs (at the late phase of differentiation/maturation at D28) obtained through reaggreagation of iPSc-derived thymic epithelial progenitors (TEPs) with primary ETPs.

overall_design: At D28 of organoid formation, cells were harvested by soft mechanical dissociation to obtain the CD45+ cell fraction and analyzed using scRNA-seq


title: Isolated TEC and DC from human iPSc-derived thymic organoids (hTOs) seeded with primary early thymic progenitors (ETPs) at late phase of maturation (D28)

summary: We employed an ultra-sensitive bulk RNA sequencing approach to study thymic epithelial cells (TECs), sorted by FACS based on their HLA-DR expression levels, and dendritic cells (DCs), both isolated from hTOs at the late stage of differentiation (day 28). These hTOs were generated by reaggregating iPSc-derived thymic epithelial progenitors (TEPs) with primary ETPs.

overall_design: At D28 of organoid formation, cells were isolated to obtain the CD45-EPCAM+ HLA-DRhigh and HLA-DRlow fractions (TEChi and TEClo), as well as the CD45+EPCAMlo fraction (DC), analyzed using SMARTER bulk RNA-seq


title: Human iPSc-derived thymic organoids (hTOs) seeded with primary early thymic progenitors (ETPs) at early phase of maturation (D7)

summary: We have employed a single-cell sequencing approach to using 10x Genomics scRNA-seq to study the differentation/maturation of thymic epithalial cell progenitors (TEPs) obtained via our iPSc differentitation protocol based on design of experiment (DoE) in the early phase of hTO formation at D7after reaggregation with primary ETPs.

overall_design: At D7 of organoid formation, hTOs were dissociated and EPCAM+CD45- cells isolated by FACS sorting and analyzed using scRNA-seq


title: Multi-omics Reveals CCL3 Driving Neuronal Sensitization in Chronic Pruritus of Unknown Origin

city: Bethesda

first: Lu

last: Zhang

state: Maryland

address: 4721 Rosedale Avenue, Apt 407

department: Department of Dermatology

institute: The First Affiliated Hospital, Sun Yat-sen University

summary:
Chronic pruritus is a pathological condition associated with numerous systemic, dermatologic, and neurologic disorders. Chronic pruritus of unknown origin (CPUO) represents a distinct clinical subtype characterized by persistent itch without an identifiable underlying cause. While emerging evidence implicates neuroimmune dysregulation in chronic pruritus, the specific mechanisms driving CPUO remain poorly understood.
Objective: To characterize the neuroimmune mechanisms underlying CPUO, with a focus on identifying potential diagnostic markers. Single-cell RNA sequencing of peripheral blood cells from patients and healthy controls was conducted. Single-cell transcriptomic analysis revealed dysregulated monocytes and hyperactive natural killer (NK) cells overexpressing CCL3.

overall_design: RNA-seq profiling of peripheral blood cells from 4 CPUO patients and 4 matched healthy controls

first: Lerong

last: Lun

first: Fang

last: Wang


title: HMGN1 and HMGN2 are recruited to acetylated and histone variant H2A.Z-containing nucleosomes to regulate chromatin state and transcription

city: Chapel Hill

first: Riya

last: Gohil

address: 250 Bell Tower Drive, 3314 Genome Sciences Building

institute: University of North Carolina at Chapel Hill

summary: The High Mobility Group Nucleosome-binding (HMGN) proteins are small abundant nuclear proteins that directly bind nucleosomes and constitute a major component of chromatin. HMGN proteins specifically localize to enhancers and actively transcribed genes across the genome, however the roles of HMGN proteins in regulating chromatin structure and transcription remain poorly understood. To investigate the localization and function of HMGN proteins on chromatin, we engineered mouse embryonic stem cells (mESCs) lacking HMGN1 and/or HMGN2 (Hmgn1-/- mESCs, Hmgn2-/- mESCs, and Hmgn1-/-Hmgn2-/- mESCs) and profiled gene expression and localization of architectural proteins. ChIP-sequencing was performed for the RAD21 subunit of cohesin and CTCF to assess the functional implications of loss of HMGN1 and HMGN2 on cohesin and CTCF localization.

overall_design: ChIP-seq was performed in wildtype mESCs (as a control) and Hmgn1-/-Hmgn2-/- mESCs in duplicate for RAD21 and CTCF.

first: Jill

last: Dowen

first: Zhihan

last: Gao

first: Rebecca

last: Lewis

first: Nathaniel

middle: T

last: Burkholder

first: Brian

last: Strahl


title: Maternal acute SARS-CoV-2 infection impairs preimplantation embryo development and reprograms early offspring hematopoietic system

city: Shanghai

first: Yuxuan

last: Zheng

address: 825 Zhangheng Rd.

institute: Fudan University

summary: The COVID-19 pandemic has raised significant concerns regarding its impact on reproductive health, particularly in the context of assisted reproductive technology (ART). This study investigates the effects of maternal acute SARS-CoV-2 infection on preimplantation embryo development and early offspring hematopoietic system. We analyzed embryo development and neonatal immune responses in women undergoing ART during acute SARS-CoV-2 infection. Our findings reveal that maternal SARS-CoV-2 infection significantly impairs blastocyst formation and quality, with molecular evidence of developmental delay in embryos. Single-cell RNA sequencing (scRNA-seq) of blastocysts and umbilical cord blood (UCB) cells demonstrated dysregulated mitochondrial function, impaired mTORC1 signaling, and altered immune responses in neonates from infected mothers. Additionally, DNA methylation analysis revealed hypermethylation of immune-related genes in embryos from infected mothers. These results highlight the potential long-term impacts of maternal SARS-CoV-2 infection on embryonic development and offspring immune health, emphasizing the need for further research to balance patient safety and reproductive outcomes during the pandemic.

overall_design:
From December 17, 2022, to January 6, 2023, a total of 123 patients underwent ovarian stimulation cycles in our ART clinic. Among these, 28 patients were assigned to the SARS-CoV-2-positive (SARS-CoV-2 infected, IS) group, defined by a positive nucleic acid test or antigen test for SARS-CoV-2 within 24 hours prior to oocyte retrieval and a body temperature below 37°C during the procedure. The remaining 95 patients were initially classified as non-infected; however, 9 were excluded due to unsuccessful oocytes retrieval or cryopreservation of oocytes, resulting in 86 patients in the SARS-CoV-2 negative group (non-infected, NI). All participants in IS and NI groups underwent either intracytoplasmic sperm injection (ICSI) or in vitro fertilization (IVF). Good-quality blastocysts were transferred back into the maternal uterus to achieve pregnancy, while additional blastocysts were cryopreserved for future use. With the patients’ informed consent, the cryopreserved blastocysts and neonatal umbilical cord blood (UCB) were collected for experimental purposes in this study.

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The raw sequence data reported in this study have been deposited in the Genome Sequence Archive in National Genomics Data Center, China National Center for Bioinformation / Beijing Institute of Genomics, Chinese Academy of Sciences (GSA-Human: HRA010475) that are publicly accessible at https://ngdc.cncb.ac.cn/gsa-human.
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first: Meiling

first: Di

last: Liu

first: Songmao

last: Li

first: Jiansheng

first: Fanghao

last: Guo

first: Haibin

last: Zhu

first: Li

last: Sun

first: Yu

last: Yan

first: Yanquan

first: Rui

last: Qiao

first: Haixia

last: Ding

first: Qing

first: Mengxi

first: Yongjian

last: Ma

first: Zhiwei

first: Wen


title: Metabolic Adaptations Rewire CD4 T Cells in a Subset-Specific Manner in Human Critical Illness with and without Sepsis

city: Nashville

first: Matthew

middle: Tyler

last: Stier

state: TN

address: 1161 21st Avenue South

department: Internal Medicine

institute: Vanderbilt University Medical Center

summary: Bioenergetic dysfunction is an archetypal finding in critical illness, particularly among patients with severe sepsis. A central feature of sepsis is the shift towards pathologic immunosuppression, especially among CD4 T cells, predisposing to secondary infections and ineffectual organ recovery. However, it remains unknown whether metabolic abnormalities contribute to CD4 T cell dysregulation in critically ill sepsis patients. We collected PBMC from medical and surgical ICU patients at Vanderbilt University Medical Center or community-matched healthy controls. We performed single cell RNA-sequencing with 10x Genomics droplet-based technology on a subset of 9 healthy controls, 19 ICU patients without sepsis, and 19 ICU patients with sepsis to generate a 644,147 cell map of critical illness.

overall_design:
Peripheral blood mononuclear cells (PBMCs) were collected from critically ill patients in the medical and surgical intensive care units (ICUs) at Vanderbilt University Medical Center 48-72 hours after admission to the ICU. This includes crtiically ill patients with sepsis (CI-Sep) and non-septic critically ill patients (CI-NS). Non-acutely ill healthy control participants (NHC) were recruited from the community including bedside surrogates of enrolled ICU patients. Cryothawed PBMC were analyzed by scRNA-seq.

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Raw files for human/patient samples are available in dbGaP (accessions phs004377.v1.p1 and phs004258.v1.p1) for controlled access to the personally identifiable sequence data. All subjects with age >89 have been assigned an age of 90 consistent with current HHS policy regarding de-identification of protected health information.
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first: Paul

last: Lindau

first: Jeffrey

last: Rathmell


first: Riya

last: Gohil

summary: The High Mobility Group Nucleosome-binding (HMGN) proteins are small abundant nuclear proteins that directly bind nucleosomes and constitute a major component of chromatin. HMGN proteins specifically localize to enhancers and actively transcribed genes across the genome, however the roles of HMGN proteins in regulating chromatin structure and transcription remain poorly understood. To investigate the localization and function of HMGN proteins on chromatin, we engineered mouse embryonic stem cells (mESCs) lacking HMGN1 and/or HMGN2 (Hmgn1-/- mESCs, Hmgn2-/- mESCs, and Hmgn1-/-Hmgn2-/- mESCs) and profiled gene expression and localization of architectural proteins. Gene expression was profiled in WT mESCs, Hmgn1-/- mESCs, Hmgn2-/- mESCs, and Hmgn1-/-Hmgn2-/- mESCs via RNA-sequencing.

overall_design: RNA-seq was performed in wildtype mESCs, Hmgn1-/- mESCs, Hmgn2-/- mESCs, and Hmgn1-/-Hmgn2-/- mESCs in triplicate replicates.

first: Jill

last: Dowen

first: Zhihan

last: Gao

first: Rebecca

first: Nathaniel

first: Brian


title: Tumor-localized CD40 agonism with MP0317, an FAPxCD40 DARPin, reprograms the tumor microenvironment - results of a Phase 1 monotherapy study

city: Zürich

first: Ana Maria

last: Florescu

address: Wagistrasse 14

country: Switzerland

institute: Molecular Partners

summary: This Phase 1 dose-escalation study evaluated MP0317 (FAP x CD40 DARPin) in adults with advanced solid tumors (n=46; 0.03–10 mg/kg IV weekly or every-3-weeks). Primary outcome measure was safety; secondary/exploratory outcome measures included antitumor activity, pharmacokinetics and pharmacodynamics. Most treatment-related adverse events were of Grade 1–2 (95%) and a maximum tolerated dose was not reached. One patient achieved an unconfirmed partial response and 14 patients stable disease. MP0317 serum pharmacokinetics confirmed extended half-life properties; terminal half-life estimates  increased with dose and ranged from 21.8–120 hours. Paired tumor biopsies confirmed colocalization of MP0317 with FAP and CD40. Activation of the CD40 pathway in the tumor microenvironment was shown by increased infiltration of antigen-presenting, plasma, and T follicular helper cells, dendritic cell maturation, IFN signaling, and circulating immune  markers. Collectively, these data confirm a favorable safety profile for MP0317 and support further clinical evaluation in combination with complementary immunotherapies.

overall_design:
RNAseq analysis of patient , biopsies pre and post treatement with increasing doses of  MP0317, a tumor localized CD40 agonist

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Raw data can not be uploaded due to patient privacy concerns.
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title: Systemic chemotherapy in metastatic TNBC polarizes effector T cell differentiation

city: Incheon

first: Hye-Yeon

last: Ju

address: 119 Academy-ro Yeonsu-gu, Incheon Republic of Korea

institute: Incheon National University

summary: In this study, the dynamic changes in the immune landscape in metastatic triple-negative breast cancer (mTNBC) were assessed and compared with healthy donors using single-cell RNA sequencing (scRNA-seq) analysis. This study is the first to provide potential diagnostic and therapeutic targets of East Asian chemotherapy-treated mTNBC with regard to effector T cells.

overall_design: Peripheral blood mononuclear cells (PBMC) of two patients with mTNBC and two healthy donors were obtained from a clinical study. ScRNA-seq libraries were prepared using the Chromium Single Cell 3′ v3.1 platform (10X Genomics) according to the manufacturer’s protocol.

first: Hyundong

last: Yoon

first: Seungpil

last: Jung

first: Serk In

last: Park

first: Tae Kon

last: Kim

first: Mi-Ryung

last: Han


title: Reorganization of H3K9me2-modified chromatin regions during mouse embryonic development

first: Emily

last: Shields

address: 3400 Civic Center Blvd

institute: University of Pennsylvania

summary: Nuclear architecture – the three-dimensional organization of chromatin in the nucleus – is a critical factor in the regulation of gene expression. The repressive histone modification histone H3 lysine 9 dimethyl (H3K9me2) is enriched at the nuclear periphery and demarcates two types of large genomic regions: lamina-associated domains (LADs); and H3K9me2-only domains (KODs) which have minimal contact with the lamina. LADs have been extensively studied and contain repressed lineage-specific genes. KODs, first identified in pluripotent stem cells, were enriched for transcriptional enhancers. By mapping genome-wide H3K9me2-modified and Lamin B1-associated chromatin in developing embryonic mouse cells, we identify cell type-specific configurations of both LADs and KODs. Our analyses reveal that LAD reorganization, which occurs during cell differentiation, correlates with lineage-specific gene expression changes, while KOD reorganization is linked to changes in lineage-specific transcriptional enhancer activity. Interestingly, cell type-specific local depletion of H3K9me2 in KODs was found to correlate with peaks of H3K27ac at active lineage-specific enhancers. We also note that ultra-long regulatory regions are frequently marked by KODs, suggesting a role for KODs in long-range gene regulation. We propose a model in which KODs organize and maintain cell type-specific enhancers in a repressed state to allow for tissue- and stage-specific gene activation.

overall_design: Cut&Run profiling H3K9me2 or LaminB in mES, E11.5 heart, or E11.5 forebrain

first: Diana

last: Fulmer

first: Andrey

last: Poleshko

first: Jonathan

last: Epstein

first: Cheryl

last: Smith


title: METTL3 contributes to osimertinib resistance in non-small cell lung cancer cell lines by regulating CDC25A and AURKB mRNA stability

city: Kanazawa

first: Shin-ichi

last: Horike

state: Ishikawa

address: 13-1 Takaramachi

department: Research Center for Experimental Modeling of Human Disease

country: Japan

institute: Kanazawa University

summary: Osimertinib, a third-generation EGFR tyrosine kinase inhibitor, is used as a first-line therapy for non-small cell lung cancer (NSCLC) harboring EGFR mutations. Nevertheless, the emergence of acquired drug resistance poses a significant challenge, leading to poor clinical outcomes. METTL3, a key methyltransferase responsible for N6-methyladenosine (m6A) modification of RNA, has been implicated in cancer development and progression across various cancer types. In this study, we explored the role of METTL3 in acquired resistance to osimertinib and assessed its potential as a therapeutic target. Using METTL3 knockdown EGFR-mutated NSCLC cell lines, we found that downregulation of METTL3 suppressed the acquisition of osimertinib resistance. Microarray analysis and qRT-PCR revealed that CDC25A and AURKB are downstream target genes of METTL3, with METTL3 facilitating the stabilization of their mRNAs. Downregulation of these METTL3 target genes also attenuated osimertinib resistance. Furthermore, we evaluated the effects of combining osimertinib with either a METTL3 inhibitor or a CDC25A inhibitor, both of which increased drug efficacy by promoting apoptosis. Collectively, this study highlights the involvement of METTL3 in the initial acquisition of osimertinib resistance and the therapeutic potential of targeting the METTL3-mediated regulatory pathway.

overall_design: Comparison between non-targeting control cells and METTL3 knocked-down cells. There are two biological replicates for each condition.

first: Ryusuke

last: Suzuki

first: Minoru

last: Terashima

first: Akihiko

last: Ishimura

first: Makiko

last: Meguro-Horike

first: Yuka

last: Kubota

first: Sasithorn

last: Wanna-udom

first: Seiji

last: Yano

first: Takahisa

last: Takino

first: Takeshi


title: Injured epithelial cell states critically impact kidney allograft survival after T-cell-mediated rejection

city: Hannover

first: Christian

last: Hinze

address: Carl-Neuberg-Str. 1

department: Department of Nephrology and Hypertension

country: Germany

institute: Hannover Medical School

summary: Background: T-cell mediated rejection (TCMR) remains a significant challenge after kidney transplantation and is associated with reduced allograft outcome. Previous research highlighted the critical role of TCMR-induced renal epithelial injury. Yet, the detailed cellular origin of these injury responses and the associated clinical implications remain poorly understood. Methods: To induce acute TCMR, we used mouse models of allogeneic (C57BL/6 to BALB/c and BALB/c to C57BL/6) kidney transplantation and syngeneic controls (C57BL/6 to C57BL/6 and BALB/c to BALB/c). Molecular changes were analyzed 7 days post-transplant using single-nucleus RNA sequencing and spatial transcriptomics. Results were compared with snRNA-seq data from three human TCMR biopsies and three stable allografts without rejection. The clinical impact of TCMR-induced epithelial injury was evaluated using marker gene sets on bulk transcriptomic data from 1292 kidney allografts, including 95 TCMR samples, with allograft outcome. Results: Mouse kidneys from allogeneic transplants exhibited all hallmark histological features of TCMR. Single-nucleus RNA sequencing revealed TCMR-induced injured cell states and significant gene expression changes particularly in proximal tubules (PT) and thick ascending limbs (TAL). Spatial transcriptomics showed a heterogeneous spatial distribution of these injured cell states and proximity to leukocytes. Cross-species analysis confirmed similar injured PT and TAL cell states in human TCMR. Signatures of a severely injured TCMR-induced TAL cell state correlated strongly with allograft survival in a large kidney transplant cohort specifically after TCMR. Conclusion: This is the first study showing kidney allograft outcome with distinct injured epithelial cell states. We found a particularly strong correlation between kidney allograft survival and TAL injury in TCMR, which occurred with reduced proximity to leukocytes, suggesting potential epithelial injury driven by non-immune mechanisms.

overall_design: The presented data comprise single nucleus RNA sequencing data from mouse kidneys of syngeneic (C57BL/6 to C57BL/6 and BALB/c to BALB/c) and allogeneic (C57BL/6 to BALB/c and BALB/c to C57BL/6) transplantation, spatial transcriptomics data of the same and single nucleus RNA sequencing data from human kidney biopsies of stable kidney allografts and samples with T cell-mediated rejection.

first: Anna

last: Pfefferkorn

first: Lorenz

last: Jahn

first: Patrick

last: Gauthier

first: Janna

last: Leiz

first: Sadia

last: Safraz

first: Vera

last: Kuhlow

first: Izabela

last: Plumbom

first: Svjetlana

last: Lovric

first: Jessica

last: Schmitz

first: Jan

last: Bräsen

first: Irina

last: Scheffner

first: Michael

last: Fähling

first: Igor

last: Sauer

first: Felix

last: Aigner

first: Janine

last: Altmüller

first: Thomas

last: Conrad

first: Kai

last: Schmidt-Ott

first: Wilfried

last: Gwinner

first: Philipp

last: Halloran

first: Muhammad

last: Ashraf


title: BiTE-secreting T cells rationally combine with PD-1 blockade and promote remodeling of the local immune landscape to enhance response durability in ovarian cancer

city: Buffalo

first: Mark

last: Long

state: NY

address: Elm & Carlton Streets

department: Bioinformatics & Biostatistics

institute: Roswell Park Comprehensive Cancer Center

summary: scRNA-seq (5' GEX) was used to examine the peritoneal TME in IE9-mp1-hFRa cells bearing animals treated with FRB-T cell + anti-PD1 therapies. We observed unique TME composition associated with acute and durable responses to combination therapy that was disrupted in progressive disease. Further we found that providing an AD-OVA immunization boost along the course of therapy enhanced favorable TME composition and overall tumor control.

overall_design: 5x106 IE9-mp1-hFRa cells (IP in 500µl PBS) were injected to establish tumors, with adoptive T cell transfer (ACT) delivered 5 days later. Mice received 1.0x106 FR-B T cells or an equivalent number of Control T cells delivered by loco-regional IP infusion, followed by 2 doses of aPD1 or IgG control. On Day 12 (7 days post ACT), mice were injected IP with 5ml PBS and the peritoneal wash (corresponding to the OC TME) collected from 4 mice/group. Recovered cells were washed in PBS and immediately stained with Zombie NIR, prepared in PBS for sorting of viable cells. Stained cells were sorted using a BD FACSAria II, collecting 30,000 live cells per mouse, which were then pooled within cohorts (120,000 live cells/group). For later time points, mice receiving  FR-B T cell + anti-PD-1 treated mice were categorized as having Progressive Disease (PD; defined as animals with ascites accumulation and abdominal circumference ≥ 8.0 cm) or Durable Response (DR; defined as mice having no ascites accumulation) on Day 38. Ascites and/or peritoneal washes were collected from PD and DR animals (n=4/group), subjected to ACK lysis to remove red blood cells, counted and pooled equally within each cohort (50,000 cells/mouse) to reach 200,000 total live cells/group.  Similarly, ascites or peritoneal wash from mice treated with FR-B T cells + PD1 or FR-B T cells+ PD1+ Ad-OVA Vax were collected 14 days post vaccination (day 28) were collected from 4 mice/group, subjected to ACK lysis, counted, and equally pooled to reach 120,000 live cells/group. Immediately following preparation, cells were provided to the Roswell Park Genomics Shared Resource (GSR) for further processing and single cell sequencing (10x Genomics).

first: Long

last: Mark

first: McGray

last: AJ Robert

first: Chiello

last: Jessie L


first: Christian

last: Hinze

summary: Background: T-cell mediated rejection (TCMR) remains a significant challenge after kidney transplantation and is associated with reduced allograft outcome. Previous research highlighted the critical role of TCMR-induced renal epithelial injury. Yet, the detailed cellular origin of these injury responses and the associated clinical implications remain poorly understood.  Methods: To induce acute TCMR, we used mouse models of allogeneic (C57BL/6 to BALB/c and BALB/c to C57BL/6) kidney transplantation and syngeneic controls (C57BL/6 to C57BL/6 and BALB/c to BALB/c). Molecular changes were analyzed 7 days post-transplant using single-nucleus RNA sequencing and spatial transcriptomics. Results were compared with snRNA-seq data from three human TCMR biopsies and three stable allografts without rejection. The clinical impact of TCMR-induced epithelial injury was evaluated using marker gene sets on bulk transcriptomic data from 1292 kidney allografts, including 95 TCMR samples, with allograft outcome.  Results: Mouse kidneys from allogeneic transplants exhibited all hallmark histological features of TCMR. Single-nucleus RNA sequencing revealed TCMR-induced injured cell states and significant gene expression changes particularly in proximal tubules (PT) and thick ascending limbs (TAL). Spatial transcriptomics showed a heterogeneous spatial distribution of these injured cell states and proximity to leukocytes. Cross-species analysis confirmed similar injured PT and TAL cell states in human TCMR. Signatures of a severely injured TCMR-induced TAL cell state correlated strongly with allograft survival in a large kidney transplant cohort specifically after TCMR.  Conclusion: This is the first study showing kidney allograft outcome with distinct injured epithelial cell states. We found a particularly strong correlation between kidney allograft survival and TAL injury in TCMR, which occurred with reduced proximity to leukocytes, suggesting potential epithelial injury driven by non-immune mechanisms.

overall_design: Xenium spatial transcriptomics data from 7 mouse kidney transplantations including 2 syngeneic controls and 5 allogeneic transplantations with acute rejection.

first: Anna

last: Pfefferkorn

first: Lorenz

last: Jahn

first: Patrick

last: Gauthier

first: Janna

last: Leiz

first: Sadia

last: Safraz

first: Vera

last: Kuhlow

first: Izabela

last: Plumbom

first: Svjetlana

last: Lovric


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

overall_design: Refer to individual Series


title: Establishment of U2OS osteosarcoma cell line with GREM1 knockdown II

city: Beijing

first: hanyuchen

last: han

address: 27 Taiping Rd

institute: Beijing Institute of biotechnology

summary: To explore the downstream regulatory pathway of GREM1, we established a U2OS osteosarcoma cell line with GREM1 knockdown

overall_design: Comparative gene expression profiling analysis of RNA seq data between GREM1 knockdown U2OS cell lines and control U2OS cell lines

first: Yuchen

last: Han


title: Genomic and transcriptomic analysis of Japanese melanoma reveals candidate biomarkers for immune checkpoint inhibitor responders.

city: Tokyo

first: Norio

last: Tanaka

address: Ariake 3-8-31, Koto

institute: Japanese Foundation Cancer Research

summary: Background Immune checkpoint inhibitor (ICI) has greatly improved the prognosis of advanced melanoma. Whereas the efficacy in Japanese patients has been found to be lower than Caucasian, the genomic and transcriptomic features associated with response to ICI of Japanese melanomas remain to be elucidated. Patients and methods Total 129 tumour samples from 78 melanoma patients, who received therapeutic regimens with or without ICI treatment, were collected at 13 institutions in Japan. We performed exome and RNA sequencing and investigated the association of genomic and transcriptomic factors with clinical efficacy of ICI therapy. Time-course data were also analysed. This is the first and largest genomic cohort study for Japanese melanoma, in which tumour samples were prospectively analysed. Results Number of somatic SNVs of Japanese melanomas is lower than that of TCGA Caucasian data due to the biased distribution of WHO subtypes. Driver subtypes of BRAF, NRAS and NF1 was less prevalent but Triple Wildtype predominantly existed in the Japanese cohort. Whereas exome-wide survey revealed no significant association of mutated genes with ICI response, by transcriptomic analysis we identified inflammation-associated genes including several chemokines and cytokines were highly expressed in responders. Follicular helper T cells estimated by immune-cell composition analysis were found significantly enriched in responders (p = 0.0422). Through time-course transcriptome analysis, in addition to several cytotoxic T-cell genes as previously reported, MARCO on tumour-associated macrophages was found induced by ICI treatment in responders (p = 0.0040). The protein expression of these genes was confirmed by immunohistochemical and multiplex immunofluorescent analyses. Some of these genes can be useful biomarkers for prediction of ICI response in the treatment of Japanese melanoma. Conclusions Through prospective genomic and transcriptomic analyses of Japanese melanoma samples, candidate biomarkers for ICI treatment were identified.

overall_design:
mRNA libraries were generated using the TruSeq Stranded mRNA Library Prep Kit (Illumina, San Diego, CA, USA), following the manufacturer’s instructions. Sequencing of captured DNA was conducted with an in-house HiSeq2500 or HiSeq2000 (Illumina) or outsourced to Macrogen Japan Corp (Aomi, Tokyo).  ***Raw data are not available for this series due to patient privacy concerns. Furthermore, the raw data will not be available through controlled-access resources such as dbGaP or EGA. The IRB prohibits public access to the raw data.

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Submitter's institutional review board prohibits public disclosure of raw data due to concerns about patient privacy.
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title: Human-Mouse Chimeric Brain Models to Study Human Glial-Neuronal and Macroglial-Microglial Interactions [scRNA-seq]

city: Piscataway

first: Ziyuan

last: Ma

address: 604 Allison Rd

department: Cell Biology and Neuroscience

institute: Rutgers University

summary: Human-mouse chimeric brain models, generated by transplanting human induced pluripotent stem cell (hiPSC)-derived neural cells, are valuable for studying the development and function of human neural cells in vivo. Understanding glial-glial and glial-neuronal interactions is essential for unraveling the complexities of brain function and developing treatments for neurological disorders. To explore these interactions between human neural cells in vivo, we co-engrafted hiPSC-derived neural progenitor cells together with primitive macrophage progenitors into the neonatal mouse brain. This approach creates human-mouse chimeric brains containing human microglia, macroglia (astroglia and oligodendroglia), and neurons. Using super-resolution imaging and 3D reconstruction techniques, we examine the dynamics between human neurons and glia, and observe human microglia pruning synapses of human neurons, and often engulfing neurons themselves, as well as the interactions between human oligodendrocytes and neurons. Single-cell RNA sequencing analysis of the chimeric brain uncovers a close recapitulation of the human glial progenitor cell population, along with a dynamic stage in astroglial development that mirrors the processes found in the human brain. Furthermore, cell-cell communication analysis highlights significant neuronal-glial and macroglial-microglial interactions, especially the interaction between adhesion molecules neurexins and neuroligins.  This innovative co-transplantation model opens up new avenues for exploring the complex pathophysiological mechanisms underlying human neurological diseases. It holds particular promise for studying disorders where glial-neuronal interactions and non-cell-autonomous effects play crucial roles.

overall_design: Two hMAN (human microglia-astrocyte-neuron co-transplanted) Rag2−/− IL2rγ−/− hCSF1KI chimeric mice were anesthetized and perfused with cold 1x Dulbecco's Phosphate-Buffered Saline (DPBS). Brains were then dissected and dissociated using the Adult Brain Dissociation Kit (Miltenyi Biotec) according to the manufacturer’s instructions. Human cells were isolated using the Mouse Cell Depletion Kit (Miltenyi Biotec), following the manufacturer’s protocols. The isolated cell suspension from the two mice were subsequently loaded into the Chromium Controller respectively for single-cell RNA sequencing (scRNA-seq) library preparation, utilizing the Chromium Next GEM Single Cell 3’ Kit v3.1 and Dual Index Kit TT Set A, in accordance with the manufacturer’s guidelines. Quality control of the scRNA-seq libraries was performed using an Agilent 2100 Bioanalyzer, and the libraries were sequenced on a NovaSeq 6000 system using S4 flow cells.

first: Jin

last: Mengmeng

first: Ma

last: Ziyuan

first: Jiang

last: Peng


title: Human-Mouse Chimeric Brain Models to Study Human Glial-Neuronal and Macroglial-Microglial Interactions  [Bulk RNA-seq]

overall_design: Cultured primitive neural progenitor cells (pNPCs) were used for bulk RNA-sequencing experiments.


title: ATAC-seq and RNA-seq analysis of Eriocheir sinensis muscle in inter-molt stage

city: Tianjin

first: Tong

last: Hao

state: China

department: College of Life Sciences

institute: College of Life Sciences, Tianjin Normal University

summary: The chromatin accessibility and gene expression of muscle in E. sinensis in inter-molt (C) stage were sequenced by Assay of Transposase Accessible Chromatin sequencing (ATAC-seq) and RNA-seq, respectively. The data was used to analysis the differentially expressed genes (DEGs) in muscle from post-molt to inter-molt by combining ATAC-seq and RNA-seq.

overall_design: Muscle of healthy crabs in inter-molt (C stages) were collected and used for ATAC-seq and RNA-seq analysis. In each stages, three muscle samples were collected as the experimental replications. ATAC-seq was performed with Illumina Hiseq platform with E. sinensis genome (accession number PRJNA555707) as the reference genome. RNA-seq was performed with Illumina novaseq platform with E. sinensis genome (accession number PRJNA555707) as the reference genome.

first: Jingjing

last: Li

first: Jinsheng

last: Sun


title: Inhibition of ADSS2-mediated de novo AMP biosynthesis resensitizes acute myeloid leukemia to BH3 mimetics [scRNA-seq]

city: Monrovia

first: Ling

address: 1218 S 5TH AVE, MONROVIA

department: Hem Malignancies Trans Sci

institute: City of Hope National Medical Center

summary: De novo purine synthesis is required to promote tumor growth; however, its role in therapy resistance remains elusive. Here, through a dynamic BH3-priming based CRISPR-Cas9 screen, we found that deletion of ADSS2, which encodes for an enzyme functioning in de novo AMP synthesis, re-sensitizes drug resistant acute myeloid leukemia (AML) cells to BH3 mimetics. Single-cell sequencing and metabolomics analyses reveal that high ADSS2 activity in leukemia samples including those with TP53 aberrations inversely correlates with venetoclax responsiveness. Further, we developed anADSS2 antagonist, exhibiting synergism with BH3 mimetics in preclinical AML models. Mechanistically, sensitization to BH3 mimetics mediated by ADSS targeting was associated with downregulated AMPK activity, which governs mitochondrial homeostasis. AMPK activity in resistant cells promotes mitophagy to eliminate damaged mitochondria upon BH3 mimetics treatment. These data demonstrate that AMP synthesis governs venetoclax resistance, and that combining ADSS targeting with BH3 mimetic treatment could represent a promising approach against AML.

overall_design: We engrafted NSGS mice with primary AML blasts(P9), then treated mice with Venetoclax/Azacitidine  or vehicle until resistance emerged. We next harvested and purified human cells from BM for analysis.

first: Xin

last: He

first: Wei

last: Chen