title: Single cell RNA-sequencing of the preoptic area of hypothalamus from sleep deprivation and rebound sleep mice

city: Shanghai

first: Huateng

last: Cao

address: 320 Yue Yang Road

institute: Center for Excellence in Brain Science and Intelligence Technology (Institute of Neuroscience), Chinese Academy of Sciences

summary: Using single-cell transcriptomics to search for hypothalamic neurons representing and controllling sleep pressure, we identified a group of neurons expressing Gpr139  in the preoptic area (POA) selectively activated during rebound sleep (RS) after sleep deprivation (SD).

overall_design: Wildtype mice underwent 6 hours SD or SD followed 2 hours RS, brain tissues of the POA region were dissected and applied to scRNA-seq. Single-cell transcriptomic analyses of cluster markers and immediate early genes were performed to identify neuron population in the POA activated during rebound sleep.

first: Zhe


title: Charachterizing the Discordance Between ARID1A Protein and Genotype in Endometrioid Type Endometrial Tumors

status: Public on Apr 22 2026

city: Chapel Hill

first: Office of Genomics Research

last: UNC Lineberger Comprehensive Cancer Center

state: NC

address: 125 Mason Farm Road, CB #7599 5105-5106 Marsico Hall

country: USA

institute: University of North Carolina at Chapel Hill

summary: ARID1A is one of the most frequently mutated genes in endometrial cancer, with approximately 40% of patients harboring an ARID1A mutation. However, relatively little is known about how ARID1A protein loss shapes endometrial cancer pathogenesis. Mounting evidence from other malignancies suggests that ARID1A protein can be regulated post-translationally, independent of genotype. However, most studies in endometrial cancer evaluate genotype alone, overlooking the potential for alternative mechanisms of ARID1A loss. To address this gap, ARID1A protein expression and genotype were examined in endometrioid tumors and associated transcriptional changes were characterized. Evaluation of ARID1A protein in 71 human endometrioid tumors demonstrates that protein loss can occur regardless of ARID1A genotype. Retention or deficiency of ARID1A protein was not significantly related to variant allele frequency or location of mutation in human tumors with mutant ARID1A. A human endometrial cancer cell model suggests that ARID1A protein loss can occur through proteasomal degradation. Furthermore, ARID1A protein expression was found to be a predictor of worse overall survival in a TCGA cohort of ARID1A wild-type endometrioid tumors. Spatial transcriptomics of 16 human endometrioid tumors revealed that both genotype and protein expression of ARID1A play a role in shaping unique transcriptional signatures in endometrial cancer and can be used to predict patient prognosis. Suggesting evaluation of ARID1A should not be done solely by sequencing techniques.

overall_design: Sixteen formalin-fixed paraffin embedded endometrioid tumors with either wild-type(5) or mutant (11) ARID1A were analyzed using whole transcriptome digital spatial analysis. Tumors had varying levels of ARID1A protein expression with some experiencing retention of ARID1A throughout the tumor while others exhibited heterogenous or deficient ARID1A protein expression. Six regions of interest were chosen per tumor based on ARID1A protein expression, resulting in 51 ROIs which retained ARID1A protein expression and 44 which were ARID1A protein deficient.

first: Savannah

middle: E

last: LaBuda

first: Katherine

last: Spence

first: Rahul

middle: R

last: Niranjan

first: Isurika

last: Weeranshige

first: Macy

middle: L

last: Osborne-Frazier

first: Russell

last: Broaddus

first: Andrew

middle: B

last: Gladden


title: Pregnancy precipitates metabolic imbalance and accelerates death in an animal model of mitochondrial cardiomyopathy

city: New York

first: Nicole

last: Sayles

address: 407 East 61st Street

institute: Weill Cornell

type: Expression profiling by high throughput sequencing

summary: During pregnancy, the heart undergoes major physiological and metabolic changes to increase cardiac workload and the demand for energy production is especially elevated during the trial of labor. Normally, cardiac structure and metabolism revert to the pre-pregnancy state shortly after delivery. However, in some cases peripartum/postpartum cardiomyopathy (PPCM) occurs, which increases a person’s risk of major cardiac events following pregnancy. The molecular mechanisms underlying PPCM remain poorly understood. In this study, we investigate the transcriptional, metabolic, and bioenergetic profiles of postpartum (PP) hearts in a mouse model of cardiomyopathy caused by the pathogenic p.S55L mutation in the mitochondrial protein coiled-coil-helix-coiled-coil-helix domain containing 10 (CHCHD10). Heterozygote p.S55L mutant CHCHD10 mice develop acute heart failure during the immediate PP period. We observe cardiac remodeling, mitochondrial stress, and profound metabolic rewiring in PP mutant CHCHD10 hearts. Metabolic rewiring results decreased levels of heme and the depletion of key cofactors of energy metabolism, including NAD(H) and ADP. These findings suggest that mutant CHCHD10 hearts fail to meet the increased energy demands associated with the trial of labor due to the insufficient turnover rate of NAD+/NADH and ADP/ATP. We propose that this metabolic insufficiency drives PP mortality in mutant CHCHD10 mice. In support of this hypothesis, dietary supplementation with nicotinamide riboside and pterostilbene, a naturally derived polyphenol, increased PP survival and cardiac energy metabolites in mutant CHCHD10 mice. Our work provides novel insights into the molecular mechanisms of PP cardiomyopathy associated with mitochondrial stress and suggests potential beneficiary effects of dietary NAD(H) supplementation.

overall_design: RNAsequencing of heart tissue from naïve and postpartum (PP) WT and mutant (p.S55L) CHCHD10 mice.

middle: M

first: Gabriella

last: Casalena

first: Dazhi

last: Zhao

first: Ryan

middle: W

last: Dellinger

first: Holly

last: Holmes

first: Onorina

last: Manzo

first: Alexander

last: Galkin

first: Annarita

last: Di Lorenzo

first: Giovanni

last: Manfredi


title: HP1B and H3K9me3 Regulate Olfactory Receptor Choice and 2 Transcriptional Identity [ATAC-seq]

city: NYC

first: Martin

last: Escamilla

address: 3227 Broadway

institute: Columbia

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Diverse epigenetic regulatory mechanisms ensure and regulate cellular diversity. Among 18 others, the histone 3 lysine 9 me3 (H3K9me3) post translational modification participates 19 in silencing lineage-inappropriate genes. H3K9me3 restricts access of transcription factors 20 and other regulatory proteins to genes that control cell fate. Mouse olfactory sensory neu- 21 rons (OSN) select to express one olfactory receptor (OR) gene out of 2,600 possibilities. 22 This monoallelic and stochastic OR choice happens as OSNs differentiate and undergo 23 dramatic changes in nuclear architecture. OR genes from different chromosomes converge 24 into specialized nuclear bodies and chromatin compartments, as H3K9me3 and chromatin 25 binding proteins including heterochromatin protein 1 (HP1) are incorporated. In this 26 work, we have uncovered an unexpected role for HP1b in OR choice and neuronal iden- 27 tity that cannot be rescued by HP1a in vivo. With the use of a conditional knock-in mouse 28 model that replaces HP1b for HP1a, we observe changes in H3K9me3 levels and DNA 29 accessibility over OR gene clusters. These changes alter the expression paTerns that par- 30 tition the mouse olfactory epithelium into five OR expression zones, which results in a 31 reduced OR repertoire leading to a loss of olfactory sensory neuron diversity. We propose 32 that HP1b modulates the competition of OR-promoters for enhancers to promote receptor 33 diversity, by establishing repression gradients in a zonal fashion.

overall_design: atac-seq from FAC sorted cell populations from the Main olfactory epithelium. Mice Olfatory ephitelium was microdisected for Zone-1 (z1) and Zone-5 (z2) ORs. Cell extracs were prepere from a mix of mice: WT, Cbx (control) and Cbx.Foxg1CRE. OMNI-ATAC protocol and taguin was follow.

last: Escamilla del Arenal

first: Rachel

last: Duffié

first: Hani

last: Shayya

first: Valentina

last: Loconte

first: Axel

last: Ekman

first: Lena

last: Street

first: Carolyn

last: Larabell

first: Marko

last: Jovanovic

first: Stavros

last: Lomvardas


title: Transcriptomic and functional profiling of endothelial dysfunction induced by polystyrene nanoplastics

city: Bellaterra

first: Alba

last: Hernández

state: Barcelona

address: Campus UAB, s/n

department: Genetics and Microbiology

country: Spain

institute: Universitat Autònoma de Barcelona

summary: Micro- and nanoplastics (MNPLs) have emerged as pervasive environmental pollutants, raising growing concerns about their potential health risks. Their documented presence in the human bloodstream highlights the need to understand their interactions with endothelial cells and their role in cardiovascular diseases such as atherosclerosis. However, the full implications of these interactions remain largely unknown. To address this, we investigated the impact of three sizes (30, 50, and 100 nm) of polystyrene carboxylated nanoplastics (PS-C-NPLs) on human umbilical vein endothelial cells (HUVECs) at 100 µg/mL. By integrating in vitro assays with bioinformatic transcriptomic analyses, we employed a multimodal approach to comprehensively investigate the effects of PS-C-NPLs on endothelial cells. In vitro assessments included nanoplastics (NPLs) internalisation (flow cytometry, confocal microscopy, TEM), morphological and internal complexity changes (flow cytometry), genotoxicity (comet assay), and functional alterations, such as cholesterol biosynthesis (Filipin III staining), migration (wound healing assay), angiogenesis, and inflammation (IL-6 ELISA). Additionally, RNA sequencing provided transcriptomic insights into the cellular response to PS-C-NPLs, complementing the in vitro findings and revealing molecular pathways underlying the observed effects. All three PS-C-NPL sizes were rapidly internalised by HUVECs within 20 minutes, inducing significant alterations in cell morphology, internal complexity, and function. PS-C-NPLs caused genotoxic damage and disrupted cholesterol metabolism, migration, angiogenesis, and inflammatory responses. Notably, some effects exhibited a size-dependent trend, with similarities emerging between carboxylated polystyrene (PS-C) 50 and 100 nm NPLs, while the smallest 30 nm NPL showed slightly distinct responses. Transcriptomic analyses reinforced these findings, revealing shared pathways across all three PS-C-NPLs —linked to cholesterol metabolism, endothelial-to-mesenchymal transition, DNA damage, and inflammation — alongside size-specific molecular signatures. This study is the first to comprehensively link transcriptomic changes to size-dependent functional alterations in endothelial cells induced by PS-C-NPLs. Our findings demonstrate that PS-C-NPLs significantly impair endothelial cell function and integrity in a size-dependent manner, underscoring their potential cardiovascular risks.

overall_design: RNA-seq profiling of HUVEC acute exposure to different sizes of carboxylated PS-NPLs (PS-C 30, 50 and 100 nm) during 24h.

first: Joan

last: Martín-Pérez

first: Aliro

last: Villacorta

first: Javier

last: Gutiérrez-García

first: Raquel

last: Egea

first: Michelle

last: Morataya-Reyes

first: Mireia

last: Cassú-Casadevall

first: Irene

last: Barguilla

first: Ricard

last: Marcos

last: García-Rodriguez


title: PFKFB3 promotes kidney fibrosis through histone lactylation-mediated HIPK2 activation in diabetic kidney disease

city: shenyang

first: xiuli

last: Guo

address: No. 155 Nanjing North Street, Heping District, Shenyang, Liaoning Province

country: China

institute: The First Hospital of China Medical University

summary: Persistently enhancement of glycolytic activity has been increasingly implicated as a key contributor to the development of diabetic kidney disease (DKD). Lactate, the end product of glycolysis, can trigger histone lactylation, —an emerging epigenetic modification involved in regulating gene expression, yet its functional significance in DKD remains largely unclear. Here, we observed that 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (PFKFB3), a key glycolytic enzyme, was remarkably induced in tubular epithelial cells (TECs) in both DKD patients and mice model. Inhibition the expression of PFKFB3 mitigated kidney fibrosis, and preserved kidney function in DKD mice model.  Conversely, upregulation of PFKFB3 aggravates renal fibrogenesis and promotes the deterioration of renal pathology. Moreover, we demonstrated that reducing the lactate levels markedly alleviated renal fibrosis in DKD. Mechanistically, lactate generated via PFKFB3-driven glycolytic reprogramming selectively enhances H4K12 lactylation at the HIPK2 promoter, thereby activating its transcription and driving renal fibrotic progression. These findings suggest that tubular PFKFB3 enhancing HIPK2 expression by promoting H4K12la-mediated gene transcription. Thus, targeting the PFKFB3-mediated HIPK2 activation in renal tubular cells could be a novel strategy for DKD.

overall_design: HK-2 cells were collected, counted, and centrifuged at 600×g for 3 minutes at room temperature. Cells totaling 1 × 105 were gently pipetted and washed twice with 500 μL of wash buffer. Magnetic beads that were coated with Concanavalin A were prepared as per the instructions provided in the kit, and then 10 μL of the activated beads were incorporated into each sample, which was incubated at room temperature for 10 minutes thereafter. Following the kit instructions, cells were incubated sequentially with ConA beads, primary antibody (anti-H4K12La antibody, PTM-1411RM, PTMBio), secondary antibody (Goat anti-Rabbit IgG H&L, AB206-01-AA), and Hyperactive PG-TN5/PA-TN5 transposon, followed by fragmentation. The fragmented DNA was then extracted and amplified by PCR. The generation of CUT&Tag libraries was accomplished using the CUT&Tag-seq kit (904, Nanjing Vazyme Biotech Co., Ltd.). The quality of the library was assessed using LabChip Touch, and sequencing was performed on an Illumina NovaSeq platform. The sequencing data were subsequently analyzed by SeqHealth Tech Co. Ltd (Wuhan).

first: Mingkun

last: Xu

first: Linhang

last: Fu

first: Yulong

last: Zhang

first: Xiuli

first: Han

last: Wu

first: Sijing

last: Gao

first: Fei

last: Xiao


title: A comprehensive transcriptome analysis of the hearts of mice in lunar gravity [RNA-Seq]

city: Toyama

first: Jun-Dal

last: Kim

state: Toyama

address: 2630 Sugitani

department: Department of Complex Biosystem Research (CBR), Institute of National Medicine (INM)

country: Japan

institute: University of Toyama

summary: Despite developing cardiac atrophy and experiencing insufficient blood circulation in a microgravity environment, astronauts on long-term spaceflights did not exhibit significant cardiac dysfunction that may be due to the heart's unique robustness to withstand the space environment. However, the cardiac molecules associated with the robustness have not yet been identified. Here, we performed cardiac gene expression profiling using RNA sequencing to identify changes in gene expression in the hearts of mice housed on Earth and on the International Space Station under 1/6 g lunar gravity conditions for over 3 weeks.

overall_design: mRNA profiles of heart were generated by RNA sequencing using the NextSeq 500 (Illumina).

first: Akiyoshi

last: Fukamizu


title: An Engineered Viral RNA Degrader on Mitochondrial Surface that Mitigates RNA Virus Infection [E vs ECD]

city: Taipei

first: Wei-Ting

last: Liao

address: No. 7, Chung-Shan S. Rd., Chung-Cheng District

country: Taiwan

institute: NTU

summary: Infections by RNA viruses cause diseases. Host factor(s) that restrain viral propagation might offer opportunities for designing new anti-virus treatments. We used vesicular stomatitis virus (VSV) as a study model, and observed the synthesis of VSV RNAs at mitochondria /endoplasmic reticulum (mito/ER) spheres accompanied by the leakage of endonuclease G (ENDOG), a mitochondrial nuclease, to the cytosol. We provide evidence that ENDOG released from mitochondria is a host anti-viral factor by eliminating viral RNAs for replication. However, ENDOG outside mitochondria can be translocated to nuclei to cause nuclear DNA damages. We engineered an ENDOG expressed on mitochondrial outer membrane (MOM), namely MOM-ENDOG, to increase the accessibility to viral RNA transcripts synthesized at mitochondrial sites without damaging nuclear DNA (nDNA). Delivery of modified mRNA of wild-type but not catalytic-dead MOM-ENDOG markedly suppressed not only the propagation of VSV, but also Dengue, and Zika virus. Thus, this organelle-specific viral RNA degrader might be developed as a broad-spectrum anti-viral agent.

overall_design: RNA-seq profiling of A549 cells with MOM-GFP overexpression vs MOM-ENDOG overexpression

first: Zee-Fen

last: Chang

first: Chih-Wei

last: Chen

first: Tung

last: Chao

first: Yi-Han

first: Shih-Che

last: Weng

first: Shin-Hong

last: Shiao

first: Yun-Jung

last: Li

first: Kai-An

last: Cheng


title: A comprehensive translatome analysis of the hearts of mice in lunar gravity [Ribo-Seq]

summary: Despite developing cardiac atrophy and experiencing insufficient blood circulation in a microgravity environment, astronauts on long-term spaceflights did not exhibit significant cardiac dysfunction that may be due to the heart's unique robustness to withstand the space environment. However, the cardiac molecules associated with the robustness have not yet been identified. Here, we performed cardiac ribosome-protected fragments (RPFs) profiling using RNA sequencing to identify changes in gene expression in the hearts of mice housed on Earth and on the International Space Station under 1/6 g lunar gravity conditions for over 3 weeks.

overall_design: Cardiac ribosome-protected fragments (RPFs) profiles of mice were generated by ribosome profiling using the NextSeq 500 (Illumina).


title: Osteopontin links acquired resistance to hypoxia-inducing antiangiogenics with refractoriness to anti-PD-L1 agents in breast cancer

city: Madrid

first: Leonardo

middle: Daniel

last: Garma

state: Madrid

address: Melchor Fernández Almagro 3

institute: Spanish National Cancer Research Center

summary: Resistance to antiangiogenics is a major challenge in cancer therapy. These agents can either normalize or exacerbate tumor vascular abnormality and hypoxia. The mechanisms of resistance remain unclear in the latter setting. By integrating data from mouse models and clinical trials, we show that hypoxia-inducing anti-VEGF therapy upregulates programmed cell death ligand 1 (PD-L1), yet fails to sensitize tumors to PD-L1 blockade. Mechanistically, early hypoxic stress triggers epithelial osteopontin (Spp1) production, which recruits monocytes and skews macrophages toward M2 states, suppressing T-cell cytotoxicity. Pharmacologic Spp1 depletion impedes the development of hypoxia, M2 infiltration, restores T-cell activity and enables synergy between antiangiogenics and anti-PD-L1. Genetic dissection – tumor-epithelial Spp1 knockout grafts and bone-marrow chimeras generated by lethal irradiation and reconstitution with Spp1-/- or wild-type hematopoietic donors – show that myeloid Spp1 contributes only marginally compared with epithelial Spp1. These findings identify Spp1 as a central mediator of resistance to hypoxia-inducing antiangiogenics, contribute to a comprehensive model of antiangiogenic resistance, and support Spp1-targeted strategies to personalize immuno/antiangiogenic therapy according to tumor hypoxia.

overall_design: Splenocyte-derived cells from wild-type FVB mice were co-cultured with RAW 264.7 cells that were exposed to different experimental conditions: normoxia +/-Spp1 and hypoxia +/-Spp1 for 72h. First, Raw 264.7 cells were seeded in a 24-well plate (40,000 cells/well) and placed in a humidified 5% CO2 incubator at 37 °C. After 24h, fresh media with/without Spp1 (Biotechne, #441-OP-050/CF) at 1 g/mL were added to the corresponding wells, and cultures were maintained under normoxic (21%O2 and 5%CO2) or hypoxic (1%O2, 5%CO2 and 94%N2) incubator conditions. After 72h, primary splenocytes were obtained from wild-type FVB mice. Briefly, WT mice were sacrificed in a  CO2 chamber, and their spleens were harvested in cold PBS immediately after sacrifice. Splenocytes were mechanically dissociated using a syringe plunger. The cell suspension was filtered thought a 40µM strainer and centrifuged at 300g for 5 min at room temperature (RT). Red blood cells were lysed using RBC lysis buffer (Invitrogen, #00-4333-57). After  centrifugation, 300× g for 5 min, the cell pellet was resuspended in DMEM supplemented with 10% fetal bovine serum for counting. 1.5x106 splenocytes per well were co-cultured with RAW 264.7 cells exposed to different conditions. Cultures were maintained at 37ºC in a 5% CO2 atmosphere for 48 h. After 48h, 1x Brefeldin A (BioLegend, #420601) was added. Splenocytes were collected for FACS analysis and RNAseq analysis. Experiments were independently replicated at least three times. RNA was extracted using RNAeasy Mini Plus Kit (Qiagen, #74134) and quantified in LabChip GX Touch Nucleic Acid Analyzer (PerkinElmer). RNA quality was determined by Agilent's 2100 Bioanalyzer Lab-Chip technology. Only samples with RNA Integrity Number (RIN)>7 was used for sequencing. Sequencing libraries were prepared with the “QuantSeq 3′ mRNA-Seq Library Prep Kit” (Lexogen, #015). Libraries were sequenced in single-read format in a NovaSeq X instrument (Illumina). Raw RNA-seq reads were trimmed using bbduk to remove adapter sequences and polyA tails. The trimmed reads were aligned to the GRCh38 genome assembly using STAR 2.7.10b. The results from the alignment were quantified using the featureCounts tool from the Rsubread 2.0.3 package. Gene counts were normalized to counts per million reads (CPM) using the rnanorm Python library.

first: José Luis

last: Ruiz-Sepúlveda

first: Verónica

last: Jiménez-Renard

first: Rebeca

last: Jimeno

first: Miguel

last: Quintela-Fandino


title: CBFA2T3-GLIS2 fusion-mediated mSWI/SNF chromatin remodeler disruption generates cancer-specific dependencies in AMKL [Cut & Tag blood]

city: Seattle

first: Jay

last: Sarthy

state: WA

address: 1920 Terry Ave

department: Ben Towne Center for Childhood Cancer Research

institute: Seattle Children's Research Institute

summary: Transcription factor (TF) fusion oncoproteins are frequently associated with poor prognosis and remain challenging therapeutic targets. Here we show that the CBFA2T3-GLIS2 (C/G) fusion oncoprotein present in lethal pediatric acute megakaryocytic leukemias (AMKLs) co-opts mSWI/SNF chromatin remodeling complexes, reprogramming regulatory element architecture to sustain aberrant cell cycle progression. The C/G fusion binds to and redistributes mSWI/SNF complexes via the SMARCD1 SWIFT domain and targets complexes to oncogenic enhancers including those bound by ETS family TFs and the G1 cyclin CCND2, a C/G AML-specific dependency. Pharmacologic inhibition of mSWI/SNF-mediated G1 cell cycle phasing via FHD-286 suppresses C/G-driven gene expression programs, induces G1 arrest and apoptosis in cells, and reduces leukemic burden and extends survival in aggressive C/G AML in vivo. These findings define an oncogenic mSWI/SNF–G1 axis in C/G AML and establish a therapeutic framework for targeting chromatin dysfunction-mediated cell cycle aberrancy in high-risk pediatric leukemias.

overall_design: First, mSWI/SNF, histone modifications, and CBFA2T3::GLIS2 fusion protein chromatin binding (CUT&RUN) were profiled in CBF/GLIS+ AML cancer cell lines, as well as cord blood cells transduced with either GFP or CBFA2T3::GLIS2. Additional profiling of accessibility (ATAC-seq, in replicate) and transcription (RNA-seq, in replicate) were conducted in selected cell lines following treatment with DMSO (vehicle), Palbociclib, FHD-286, and combination treatment of Palbociclib and FHD-286.

first: Gabe

last: Boyle

first: Alexander

last: Ying


title: CBFA2T3-GLIS2 fusion-mediated mSWI/SNF chromatin remodeler disruption generates cancer-specific dependencies in AMKL [Cut&Tag WSU-AML]


title: Transcriptomic profiling reveals substrate- and shear stress_x0002_dependent maturation of human small intestinal epithelial cells

city: Berlin

first: Rainer

last: Glauben

address: Hindenburgdamm 30

department: Klinik für Gastroenterologie, Infektiologie und Rheumatologie

country: Germany

institute: Charite CBF

summary: Primary human intestinal epithelial cells (IECs) require microenvironments that reproduce various in vivo cues to maintain survival, differentiation, and function in vitro. In this study, we investigated how intestinal stem cell (ISC)-derived monolayers respond to biomimetic substrates and shear stress using 3D-printed hydrogels based on bioactive decellularized and methacrylated small intestinal submucosa (dSIS-MA) integrated into a custom millifluidic system. Extensive bulk RNA sequencing experiments revealed that, compared with Matrigel-coated tissue culture plastic, dSIS-MA hydrogels supported survival- and differentiation-related signaling, stabilized gene expression over time, and promoted absorptive lineage maturation while reducing crypt-associated signatures. By applying dynamic culture conditions to the hydrogel system, IECs underwent transcriptional remodeling, characterized by activation of metabolic and immune pathways. Longitudinal analysis further indicated that shear stress enhanced metabolic pathway–associated gene expression and promoted differentiation toward absorptive lineages. These findings establish dSIS-MA hydrogels with controlled fluid flow as a biomimetic in vitro model that supports survival, maturation of human IECs and enables transcriptional adaptation to defined biochemical and mechanical cues, supporting future applications in disease modelling, drug testing, and regenerative medicine.

overall_design: RNA-seq profiling of intestinal stem cell/organoid derived monolayers cultured on either Matrigel-coated tissue culture plastic or 3D-printed dSIS-MA hydrogels, under static or dynamic flow in a custom millifluidic system. All experiments were performed using two biological replicates and three to four technical replicates per condition.

first: Nadra

last: Alzain

first: Ahed

last: Almalla

first: Ketaki

last: Sandu

first: Lorenz

last: Gerbeth

first: Jörn

last: Ziegler

first: Catharina

last: Gutwein

first: Patrick

last: Weidner

first: Marie

last: Weinhart

first: Britta

last: Siegmund


title: CBFA2T3-GLIS2 fusion-mediated mSWI/SNF chromatin remodeler disruption generates cancer-specific dependencies in AMKL

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

overall_design: Refer to individual Series


title: CBFA2T3-GLIS2 fusion-mediated mSWI/SNF chromatin remodeler disruption generates cancer-specific dependencies in AMKL [Cut & Run]


title: Multi-omic analysis reveals nitric oxide dependent remodeling in classically activated macrophages and identifies negative regulation mediated by AKR1A1

city: Madison

first: Jing

last: Fan

state: WI

address: 330 N. Orchard St.

institute: Morgridge Institute for Research

summary: Nitric oxide (NO•) is an important signaling molecule in many biological processes, including immune response. During response to classical activation stimuli lipopolysaccharide (LPS) and interferon-γ (IFNγ), macrophages generate NO• via inducible nitric oxide synthase (iNOS). To comprehensively define the effects of NO•, we applied a multi-omic strategy integrating proteomics and transcriptomics to profile murine macrophages across conditions with or without LPS/IFNγ-activation, with or without iNOS expression or exogenous NO• donor treatment. The results revealed NO• has broad, yet selected and controlled, regulatory effects, playing a key role in coordinating the systematic remodeling during macrophage classical activation. Among the proteins that are most suppressed in a NO•-dependent manner, electron transport chain (ETC) is the most enriched. NO• drives complex-specific remodeling of ETC, causing selected downregulation of complex I, II, and IV, through a different combination of transcriptional and post-transcriptional mechanisms for each complex. Functionally, we found NO• is required, but not sufficient, for the strong suppression of cellular respiration upon macrophage activation. Among the most consistently upregulated proteins are many enzymes involved in redox defense. AKR1A1 was identified as a top hit. We found Akr1a1 induction requires both NO• and LPS/IFNγ stimulation. The S-nitroso-CoA reductase activity of AKR1A1 mitigates NO•-driven inhibition of pyruvate dehydrogenase complex by limiting the inhibitory modifications targeting its lipoyl cofactor. Knocking out Akr1a1 causes accelerated remodeling of TCA cycle, dysregulated immunoregulatory metabolite level, and altered functional gene expression and cytokine production at later stage of immune response. Thus, the NO•-dependent upregulation of AKR1A1 forms a negative regulatory loop to fine-tune NO•-mediated metabolic and functional remodeling during immune response. Together, this work provided a systems-level map of NO•-dependent regulation, revealed the crosstalk between NO• and immune signaling, and demonstrated mechanisms providing redox adaptation and precise control of NO•’s effects.

overall_design: To investigate how gene expression changes in macrophages with stimulation, wild-type or iNOS KO bone-marrow derived macrophages, or wild-type or Akr1a1 KO RAW264.7 cells were treated with continuous 48-hour LPS+IFNy stimulation. Then the stimulants were washed off and RNA was collected.

first: Nicholas

last: Arp

first: Uzziah

middle: S

last: Urquiza

first: Marcel

last: Morgenstern

first: Jonathan

middle: H

last: Schrope

first: James

middle: A

last: Votava

first: Steven

middle: V

last: John

first: Felicia

last: Deng

first: Jack

middle: J

last: Stevens

first: Anna

last: Huttenlocher

first: Joshua

last: Coon


title: Adipose tissue inflammation mediated by CCL19 overexpression exacerbates experimental periodontitis via elevated circulating saturated fatty acids and osteopontin in Western-diet-fed mice

city: Fukuoka

first: Takanori

last: Shinjo

state: fukuoka

address: Maidashi 3-1-1

department: Section of Periodontology

institute: Kyushu University

summary: Introduction: Individuals with obesity may be at a higher risk of developing severe periodontitis. We previously reported that C-C motif ligand 19 (CCL19) plays a pivotal role in adipose inflammation in obesity and that adipocyte-specific Ccl19 knock-in (CCL19-KI) mice exhibited greater inflammatory cell infiltration in visceral fat, increased weight gain, and abnormal glucose metabolism than wild-type (WT) mice under a 40% high-fat diet (HFD). In this study, we examined the susceptibility of mice fed diets with different fat contents to the severity of experimental periodontitis. Methods: Six-week-old male WT and CCL19-KI mice were fed either a normal diet (ND), 40% HFD, or 60% HFD for 8 weeks, after which ligature-induced periodontitis (LIP) was established. Two weeks after ligation, alveolar bone resorption, gingival inflammatory and osteoclastogenic gene expression, and serum free fatty acid (FFA) levels were compared among the groups. Additionally, RNA-sequencing (RNA-seq) was performed on epididymal white adipose tissue (eWAT) of each ligatured mouse to explore potential factors mediating periodontitis aggravation. Furthermore, in vitro studies using macrophages were conducted to investigate a possible mechanism in the progression of periodontitis. Results: Alveolar bone resorption, gingival inflammatory and osteoclastogenic gene expression, and serum FFA levels were significantly higher in ligated CCL19-KI mice than those in ligated WT mice under a 40% HFD. RNA-seq revealed that the OPN gene (Spp1) expression in eWAT was significantly upregulated in CCL19-KI mice fed an ND or 40% HFD compared with that in WT mice, which was consistent with serum OPN levels. Peritoneal macrophages from 40% HFD-fed CCL19-KI mice secreted significantly higher amounts of TNF-a than those from WT mice fed the same diet under Escherichia coli lipopolysaccharide (LPS) stimulation. Both palmitic acid priming and OPN significantly promoted osteoclastogenesis in RANKL-treated BMMs, and their combination further enhanced this effect. Conclusions: CCL19-modified adipose tissue inflammation may contribute to the severity of periodontitis via the upregulation of circulating saturated fatty acids and Spp1, resulting in enhanced TNF-a production in macrophages and increased osteoclastogenesis in BMMs under mild to moderate obesity.

overall_design: Normal diet or 40% HFD-fed or 60% HFD-fed WT and Adipoq-CCL19-KI mice (14 weeks old, feeding for 8 weeks) were induced ligature-induced experimental periodontitis for 2 weeks. Epididymal adipose tissue (eWAT) were collected and RNA was extracted from each mouse eWAT. RNA-seq was performed using eWAT RNA (n=3/group).

first: Naoaki

last: Ryo

first: Miyu

last: Shida

first: Kohei

last: Sato

first: Tatsuro

last: Zeze

first: Honoka

last: Otsuka

first: Yuki

last: Nishimura

first: Mio

last: Imagawa

first: Al-kafee

last: Ahmed

first: Gulinigeer

last: Dilimulati

first: Chikako

last: Hayashi

first: Akiko

last: Yamashita

first: Takao

last: Fukuda

first: Terukazu

last: Sanui

first: Misaki

last: Iwashita

first: Fusanori


title: Efficacy Studies of Exosomal Formulations of Celastrol against A549  Lung Cancer Cells

city: Louisville

first: Richa

last: Singhal

state: KY

address: 580 S Preston Street, Baxter II Building, Office 321B

department: Medicine

institute: University of Louisville

summary: In this study, the primary focus was to test the applicability of celastrol (CEL) against lung cancer using in vitro and in vivo models and to test wether loading CEL onto exosomes (Exo) help improve the cell uptake, tumor targetability, and bioavailability of CEL.

overall_design: In order to establish the mechanism of action of celastrol (CEL) at mechanistic level, A549 lung cancer cells were treated with 1 µM CEL for 48h and compared against the untreated control .

first: Aqil

last: Farrukh

first: Gupta

last: Ramesh C

first: Singhal

last: Richa

first: Moholkar

last: Disha Nagesh


title: Transneuronal cytokine delivery promotes functional recovery across spinal cord contusion severities via descending circuit plasticity

city: Cologne

first: Dietmar

last: Fischer

address: Paul-Schallück-Straße 8

department: Pharmacology

institute: University Hospital of Cologne

summary: Spinal cord injury (SCI) frequently leads to permanent motor and sensory deficits, with complete injuries causing total loss of function below the lesion and incomplete injuries preserving only partial connectivity. Intracortical delivery of an AAV2 vector encoding the designer cytokine hyper-interleukin-6 (AAV2-hIL-6) enhances recovery after complete SCI by transneuronally stimulating raphe nuclei. Here, we verified that AAV2-hIL-6 transneuronally activates subcortical neurons in the medulla and evaluated that this strategy enables recovery in clinically more relevant mouse models of mild, moderate, and severe contusion injury. Across all severities, AAV2-hIL-6 treatment significantly improved locomotor function compared to AAV2-GFP-treated controls. Although lesion size and neuronal loss correlated with contusion severity and were unaffected by the AAV2-hIL-6 treatment, it robustly increased the number and length of descending serotonergic axons in the lumbar cord. Selective ablation of serotonergic neurons abolished these gains, confirming their essential role in functional sensorimotor recovery. However, while AAV2-hIL-6 also reduced corticospinal tract (CST) axon retraction, it did not induce axon regeneration beyond the lesion, suggesting that CST regeneration was not required for recovery. Thus, intracortical AAV2-hIL-6 delivery drives circuit remodeling and functional restoration across contusion severities, highlighting its promise as a regenerative therapy for SCI with spared pathways.

overall_design: RNA-seq analysis of the medullary brainstem, including the raphe nuclei three weeks after intracortical stereotactic AAV2-GFP or AAV2-hIL-6 injections

first: Leibinger

last: Marco

first: Moskaliov

last: Igor

first: Ani

last: Chinonso-John

first: Halawani

last: Dalia

first: Fischer

last: Dietmar


title: SlMYB60 regulates cuticle formation in tomato

city: Milano

first: Raul

last: Pirona

address: Via Alfonso Corti 12

country: Italy

institute: IBBA-CNR

summary: The transcriptional regulator, SlMYB60 (Solyc10g081490.1.1) stands out as the tomato ortholog of AtMYB60, a well-characterized guard cell-specific transcription factor in Arabidopsis involved in integrating multiple stress signals in stomata. To investigate the role of SlMYB60, CRISPR-Cas9 editing was applied to two commercial tomato cultivars (Ailsa Craig and Red Setter), generating knockout alleles through multiple sgRNAs. T1 slmyb60-edited plants were assessed for stomatal aperture and drought response. Unexpectedly, mutant lines showed a slight but significative increase in stomatal pore opening in both leaves and cotyledons compared to wild type. Under dry-down conditions, leaf area, fresh weight, and relative water content were significantly affected by both genotype and treatment. RNA-seq analyses confirmed the strong downregulation of SlMYB60 in the edited lines and identified 186 differentially expressed genes (DEGs), many of which involved in cell wall or cuticle biosynthesis. Chlorophyll leaching assays indicated increased permeability in mutant lines, while electron microscopy revealed defects in epidermal cell surface structure and patterning. Notably, slmyb60 mutants exhibited thinner stomatal ledges compared to wild type, suggesting defects in stomatal architecture. Although modulating stomatal activity remains a promising strategy to improve drought resilience in tomato, our findings reveal that SlMYB60, unlike its Arabidopsis homolog, is not confined to guard cells and plays broader roles in epidermal development, indicating divergences in the MYB60 regulatory network between Arabidopsis and tomato.

overall_design: RNA was extracted from leaves of the wild type and the two mutants (Cr1 and Cr2) each with four biological replicates. Libraries were prepared from poly(A)-enriched mRNA fractions and sequecced on a NovaSeq 6000 platform using 150 bp paired-end reads.

first: Sara

last: Colanero

first: Beatrice

last: Landoni

first: Giulia

last: Castorina

first: Diana

last: Gervasoni

first: Manuela

last: Rigano

first: Elena

last: Baldoni

first: Damiano

last: Martignago

first: Lucio

last: Conti

first: Massimo

last: Galbiati


title: PTPRN2 hypomethylation and PHB2-modulated miR-153-3p maturation reveal dual epigenetic mechanisms linked to symptom variability in Myalgic Encephalomyelitis

city: Montreal

first: Alain

last: Moreau

state: QC

address: 3175 Côte-Ste-Catherine

country: Canada

institute: Azrieli Research Center of CHU Sainte-Justine

type: Methylation profiling by genome tiling array

summary:
Background: Myalgic encephalomyelitis (ME) is a chronic, debilitating condition increasingly linked to epigenetic changes. With its unclear pathophysiology and no validated diagnostic biomarkers, DNA methylation becomes of interest. Specifically, DNA methylation patterns in saliva, to study ME-related epigenetic changes.
Methods: Saliva samples from 54 ME patients and 21 sedentary healthy controls were analyzed by DNA methylation array. Symptom assessment was conducted using validated questionnaires (SF-36, MFI-20, and DSQ).
Results: A significant DNA hypomethylation at the CpG site cg19803194 (Bonferroni-corrected p=3x10-7) within the PTPRN2 gene body was identified. This hypomethylation was associated with cognitive impairments in both sexes, such as difficulties expressing thoughts and comprehension, commonly known as "brain fog," and as well as respiratory issues in male patients. The hypomethylation also corresponded with reduced circulating levels of miR-153-3p, an intronic microRNA of PTPRN2, which was associated with impaired memory recall in both sexes. Interestingly, the mitochondrial protein Prohibitin 2 (PHB2) emerged as a negative regulator of miR-153-3p maturation, likely through its physical interaction with immature forms of miR-153 within cells, which was attenuated in female patients due to increased extracellular export of PHB2.
Conclusions: These findings propose a potential epigenetic mechanism in ME involving PTPRN2 body hypomethylation and PHB2-modulated miR-153-3p maturation. While the directionality between gene-body methylation and expression remains a biological hypothesis, these results shedding light on possible pathways contributing to symptom variability and sex differences in ME severity.

overall_design: This is a prospective pilot study in which 54 consecutive ME patients diagnosed according to the Canadian Consensus Criteria were enrolled. We also recruited 21 sedentary healthy controls free from chronic pain conditions, mental disorders, recent infections (within 3 months). Saliva samples were collected using DNA Genotek Oragene kits according to the manufacturer's instructions. Genomic DNA was extracted from the saliva samples using the PureLink Genomic DNA Mini Kit following the provided protocol. This ensured efficient genomic DNA isolation for subsequent analyses. We employed a robust data processing and quality control pipeline for the Illumina Methylation EPIC microarray dataset (Illumina, San Diego, CA, USA), encompassing 75 saliva samples using the R Package Champ (Version 2.28.0) within R (Version 4.1.2) to ensure accuracy and reliability. First, the raw signal intensity data for each of the 75 samples were imported into Champ. Subsequently, these raw intensities were transformed to generate a beta value matrix. The beta values, which range from 0 to 1, reflect the methylation ratios of individual probes. To ensure data integrity, low-quality probes and samples were filtered out. Probes were removed based on the following criteria, including non-CpG probes, multi-hit probes, probes with Beadcount < 3, low-quality or failed probes with signal-background detection, P values exceeding 0.05 and probes residing on single-nucleotide polymorphism (SNP) sites or chromosomes X and Y. Simultaneously, samples containing more than 10% failed or low-quality probes were excluded from the analysis, in accordance with standard data quality practices.


title: Pleuromutilin Antibiotics Suppress Hepatocellular Carcinoma Growth

city: Shanghai

first: mingshan

last: zhou

address: Xizu Road Street

institute: Shanghai Cancer Institute

summary: Pleuromutilin antibiotics(Lefamulin, Tiamulin, et al) has application potential in malignancies treatment. To mechanistically investigate the inhibitory activity against HCC of Lefamulin and Tiamulin, we treated Hepa1-6 cells with 37 μg/ml Lefamulin or Tiamulin for 2 days. Then the total RNA was performed RNA-Seq analysis.

overall_design: RNA_seq of no treatment Hepa1-6 cells and following the treatment of 37 μg/ml Lefamulin or Tiamulin for 2 days

first: Min

last: Zhou

first: Mingshan


title: A ZBTB26–Integrator axis mediates targeted transcriptional activation

city: Guangzhou

first: dingdang

last: Yu

address: No. 106, Zhongshan 2nd Road, Yuexiu District

institute: Guangdong Provincial People's Hospital

summary: The Integrator complex is primarily known to attenuate transcription by inducing premature termination of paused RNA polymerase II (RNAPII) and restraining pause release through its endonuclease and phosphatase activities. However, the mechanisms that govern Integrator recruitment to chromatin and direct gene-specific transcriptional programs in response to external stimuli and diverse biological contexts remain poorly understood. In particular, the regulatory functions of the Integrator auxiliary or arm module (INTS10/13/4/15) are the least characterized. Here, we identify ZBTB26 as an interactor of the Integrator auxiliary module via INTS10 and INTS13, and show that it co-occupies select promoters and enhancers with Integrator and active histone markers across the genome. We demonstrate that ZBTB26 is required for the recruitment of Integrator to its target loci, including genes involved in stimulus response, development, and differentiation. Importantly, the ZBTB26-Integrator axis sustains the active state of specific promoters and enhancers and drives defined gene transcriptional programs. Our findings reveal a transcription factor-like mechanism of targeted gene regulation mediated by ZBTB26-Integrator binding, extending the functional paradigm of Integrator beyond its canonical catalytic roles.

overall_design: HEK293T cells were used to investigate the regulatory function of ZBTB26. FLAG- and HA- tagged ZBTB26 were stably overexpressed, and ChIP-seq performed using anti-FLAG and anti-HA antibodies to define ZBTB26 genomic binding sites.                                               To assess the functional consequences of ZBTB26 depletion, ZBTB26 KO and WT cells were subjected to ChIP-seq analysis of INTS10, INTS13, RNA polymerase II (total, Ser5-phosphorylated, and Ser2-phosphorylated forms), as well as histone modifications including H3K27ac, H3K4me1, and H3K4me3.                                                                                In parallel, transcriptional changes were evaluated using RNA-seq, while nascent transcription was assessed by TT-seq and PRO-seq in ZBTB26 KO and WT cells.

first: Dingdang

first: Ping

last: Zhu

first: Shibin

last: Hu


title: Histone H4 acetyl-methyllysine marks accessible chromatin that resists compaction

city: Orange

first: Matthew

middle: D

last: Simon

state: CT

address: West Campus, 100 West Campus Drive, Ste MIC312A

department: MBB

institute: Yale University

summary: Regulatory DNA often remains accessible even under conditions of widespread chromatin compaction. Specific factors and histone modifications mark these regions and help maintain local accessibility. Here, we examine the genomic landscape of acetyl-methyllysine (Kacme), a recently discovered histone post-translational modification. Across multiple systems, Kacme is highly enriched at sites of accessible chromatin, including active promoters, enhancers, silencers, and CTCF-binding sites. We find that Kacme is selectively retained at loci that resist condensation during mitosis, marks XIST and escapee regions on the inactive X chromosome in female cells, and demarcates the boundaries of broad heterochromatin domains. Using the KDM6A/6B inhibitor GSK-J4 to elevate H3K27me3 levels, we show that Kacme-marked insulator elements block heterochromatin spreading and protect adjacent genes from transcriptional repression. Taken together, our findings define the chromatin features associated with Kacme and support a model in which Kacme safeguards chromatin accessibility, particularly at loci that remain active despite compaction and silencing pressures.

overall_design: TimeLapse-seq, ATAC-seq, and CUT&RUN with Kacme and H4K5ac antibodies in HEK293T cells at select timepoints during mitosis and early-G1. ChIP-seq with Kacme, H4Kacme, and H4Kac antibodies in THP-1 cells +/- PMA (40 ng/mL, 72 h). CUT&RUN with Kacme in HEK293T cells treated with Triptolide (10 uM, 1h). TimeLapse-seq and CUT&RUN with H3K27me3 antibody in untreated HEK293T cells or HEK293T cells treated with GSK-J4 (5 uM, 6/24 h). CUT&RUN with CTCF antibody and ChIP-seq with H3K27ac antibody in wild-type HEK293T cells. All experiments performed in duplicate or triplicate. Inputs included.

first: Andreas

middle: P

last: Pintado-Urbanc

first: Charlie

last: Brown

first: Leah

last: Connor

first: Joshua

middle: T

last: Young

first: Yeil

last: Kim

first: Elizabeth

last: Black

first: Lilian

last: Kabeche


title: Novel driver gene MDC1 confers homologous recombination repair deficiency and genomic instability in chemoresistant relapsing ovarian cancer

city: Los Angeles

first: Michelle

last: Jones

state: CA

address: 8700 Beverly Blvd

department: Bioinformatics and Functional Genomics

institute: Cedars Sinai

summary: Mutational burdens and clonal compositions are established early and are maintained throughout recurrence. Using both next generation and ultra long read sequencing to analyze single nucleotide and structural variants (SVs) we discovered that although tumors from the same patient remained relatively stable, homologous recombination repair proficient (HRP) and homologous recombination repair deficient (HRD) tumors presented with distinct clonal profiles. SV signature analysis revealed three distinct classes: tumors defined by DNA losses, DNA gains, and copy number neutral changes. Each class displayed structural variation affecting distinct regions of the genome. Ultra long read sequencing validated most of the SVs identified in short read sequencing and identified additional SVs. A novel candidate driver gene from the HRP pathway, MDC1, was significantly mutated in patients with HRP tumors.

overall_design: RNA-seq profiling of two ovarian cancer cell lines and two normal fallopian tube lines with knockdown of MDC1 or a negative human control

first: Michael

last: Diaz


title: HP1B and H3K9me3 Regulate Olfactory Receptor Choice and 2 Transcriptional Identity [RNA-Seq]

city: NYC

first: Martin

last: Escamilla

address: 3227 Broadway

institute: Columbia

summary: Diverse epigenetic regulatory mechanisms ensure and regulate cellular diversity. Among others, the histone 3 lysine 9 me3 (H3K9me3) post translational modification participates in silencing lineage-inappropriate genes. H3K9me3 restricts access of transcription factors and other regulatory proteins to genes that control cell fate. Mouse olfactory sensory neurons (OSN) select to express one olfactory receptor (OR) gene out of 2,600 possibilities. This monoallelic and stochastic OR choice happens as OSNs differentiate and undergo dramatic changes in nuclear architecture. OR genes from different chromosomes converge into specialized nuclear bodies and chromatin compartments, as H3K9me3 and chromatin binding proteins including heterochromatin protein 1 (HP1) are incorporated. In this work, we have uncovered an unexpected role for HP1b in OR choice and neuronal identity that cannot be rescued by HP1a in vivo. With the use of a conditional knock-in mouse model that replaces HP1b for HP1a, we observe changes in H3K9me3 levels and DNA accessibility over OR gene clusters. These changes alter the expression paTerns that partition the mouse olfactory epithelium into five OR expression zones, which results in a reduced OR repertoire leading to a loss of olfactory sensory neuron diversity. We propose that HP1b modulates the competition of OR-promoters for enhancers to promote receptor diversity, by establishing repression gradients in a zonal fashion.

overall_design: Mouse olfactory epithelia were dissected and lysed using Trizol with standard protocols in an RNA clean area. DNase treatment was performed with Turbo DNA-free kit (Ambion, Cat. Number AM1907), sequencing libraries were prepared with Nugen NuQuant RNA-seq library system, QC was determined with Agilent Bioanalyzer, and sequenced on a HiSeq2500. Cut adapt was used to remove adapter sequences and reads were aligned to the mm10 genome with STAR. Samtools was used to select high mapping quality reads (-q 30). Normalization, calculation of FPKM (converted to TMP), and differential expression analysis was performed in R with DEseq2. For all RNA-seq data p-values refer to adjusted p-value (padj) calculated in DEseq2.

last: Escamilla del Arenal

first: Rachel

last: Duffié

first: Hani

last: Shayya

first: Valentina

last: Loconte

first: Axel

last: Ekman

first: Lena

last: Street

first: Carolyn

last: Larabell

first: Marko

last: Jovanovic

first: Stavros

last: Lomvardas


title: EpCAM Supports Exit from Pluripotency of Embryonic Stem Cells via Eomes

city: Munich

first: Kristian

last: Unger

address: Marchioninistr. 15

department: Department of radiation oncology

institute: LMU University Hospital

summary: Epithelial cell adhesion molecule (EpCAM) is a tumor-associated antigen that marks pluripotent embryonic stem cells (ESCs). Regulation of Epcam expression yields a spatiotemporal patterning during embryogenesis that is thoroughly mimicked in a 3D model of spontaneous differentiation of embryoid bodies (EBs). Here, we present a role of EpCAM in exit from pluripotency of murine ESCs (mESCs) to establish cardiomyocytes in EBs. Comparative transcriptomic analysis of wildtype and Epcam-knockout mESCs at strategic time points of spontaneous differentiation uncovered molecular deficiencies of Epcam-knockout ESCs in “Wnt signaling” and “Heart development”. Multi-level bioinformatic analyses revealed central lineage-defining transcription factors Eomes, Foxa2, and Gata6 as differentially expressed genes (DEGs) that are misregulated in Epcam-knockout mESCs. Gene expression association of Epcam with Eomes, Foxa2, and Gata6 was prominent at day three of spontaneous differentiation, representing primitive streak formation in EBs. Interrogation of public single-cell RNA sequencing (scRNAseq) datasets supported a co-expression of Epcam and Eomes at early stages of murine embryogenesis in epiblast, primitive streak, nascent mesoderm, extraembryonic ectoderm and endoderm. Newly generated scRNAseq of wildtype mESCs in spontaneous differentiation confirmed the formation of essential stages including epiblast, primitive streak, endo- and mesoderm cells, and cardiomyocytes. Expression and pseudotime analysis positioned Epcam expression slightly ahead of Eomes at the transition of early to late primitive streak, along with rising of Wnt signaling. Accordingly, conditional re-expression of Epcam or Eomes but not of Foxa2 or Gata6 complemented differentiation defects of Epcam-knockouts and confirmed an involvement of Wnt signaling in the EpCAM-dependent activation of Eomes. Hence, defective exit of pluripotency in Epcam-deficient ESCs is linked to Eomes regulation via Wnt signaling.

overall_design: Single cell RNA sequencing was performed using the BD Rhapsody HT single cell analysis system (BD Biosciences, Heidelberg, Germany) using adapted protocols. Wildtype murine embryonic stem cells E14TG2α were harvested in four consecutive passages to form embryoid bodies (EBs) for spontaneous differentiation in the absence of leukemia-inhibitory factor (LIF) for three, five, and seven days (D3, D5, D7) generating a total of n = 12 samples. EBs of each biological replicate were collected in separate reservoir tanks, transferred to 50 mL Falcon tubes, and sedimented via gravity before carefully removing supernatant. EBs were resuspended in 2 mL of TrypLE solution and incubated with constant mixing at 37°C for 1-2 minutes to generate single cell suspensions. Cells from each replicate and time point were separately centrifuged 5 min at 400 g, washed once with PBS with 1% FBS, and stained separately with 20 uL sample tag in 80 uL 2x AbSeq labeling MasterMix 45 min on ice. From here on, all steps were conducted on ice. Mouse immune sample tags are anti-CD45 monoclonal antibodies linked to unique identifier oligonucleotide sequences for sample identification. Labelled cells from four replicates (each 2,5 x 10E5 cells) were pooled for each individual time point to three samples (D3, D5, D7) and washed thrice in 2 mL of PBS with 1% FBS, centrifuged at 400 g for 5 min, and finally resuspended in 620 μL cold sample buffer. Staining cells with viability markers was achieved by adding 3.1 μL of 2 mM Calcein AM and 3.1 μL of 0.3 mM DRAQ7™ to the cell suspension in cold sample buffer (37°C in the dark, 5 min). Thereafter, 10 µL cells were counted using BD Rhapsody scanner 10 μL in an INCYTO™ disposable hemocytometer with the remaining cells kept on ice and protected from light. Counted cells were then processed according to BD Rhabsody protocol include all following steps: • Priming and treating BD Rhapsody 8 lane cartridge • Cell loading • Loading and washing BD Rhapsody™ Enhanced Cell Capture Beads • Lysing cells • Retrieving BD Rhapsody™ Enhanced Cell Capture Beads • Washing BD Rhapsody™ Enhanced Cell Capture Beads • Performing reverse transcription • Treating sample with Exonuclease I Libraries were prepared from captured RNA from single cells and were sequenced in collaboration with the group of Dr. Inti Alberto De La Rosa Velásquez (Helmholtz Center Munich, Germany). cDNA library sequencing was performed on an Illumina NovaSeq X Plus sequencer in a 10B200 flow as paired-end reads.

first: Ana Marija

last: Balaz

first: Jiahang

last: Song

first: Olivier

last: Gires


title: The immune response to a biodegradable polymer and corresponding collagen regeneration

first: Yanshuang

last: Zhang

address: No. 2005 Songhu Road

institute: Fudan university

summary: In this study, we used RNA sequencing to investigate the molecular mechanisms underlying site-specific tissue regeneration induced by an injectable thermosensitive PLGA-PEG-PLGA hydrogel (T-gel) in rats. We compared the transcriptomic profiles of skin tissue following intradermal or subcutaneous injection of T-gel, with saline injections at corresponding sites as controls. This is evidenced by a greater number of differentially expressed genes and a significant enrichment of extracellular matrix organization and collagen biosynthesis pathways in the intradermal group compared to the subcutaneous group.

overall_design: To elucidate the molecular basis for the site-dependent regenerative response to T-gel, we performed RNA sequencing on skin tissue samples from Sprague-Dawley rats 7 days post-injection, comparing four experimental groups (n=6 per group): intradermal (ID) injection of T-gel, ID injection of saline control, subcutaneous (SC) injection of T-gel, and SC injection of saline control. Total RNA was isolated from each sample, strand-specific mRNA libraries were constructed, and paired-end sequencing (2×150 bp) was performed on an Illumina HiSeq X Ten platform. Differential gene expression analyses were conducted to compare T-gel versus control at each site.

first: Tao

last: Sun

first: Ye

last: Fu

first: Yue

last: Yu

first: Runjia

last: Shen

first: Lin

first: Jiandong

last: Ding


title: A comprehensive transcriptome analysis of the livers of mice in microgravity.

city: Toyama

first: Jun-Dal

last: Kim

address: 2630 Sugitani

institute: University of Toyama

summary: This Japan Aerospace Exploration Agency project focused on elucidating the impacts of partial gravity (partial g) and microgravity (μg) on mice using newly developed mouse habitat cage units (HCU) that can be installed in the Centrifuge-equipped Biological Experiment Facility in the International Space Station. In the first mission, 12 C57BL/6 J male mice were housed under μg or artificial earth-gravity (1 g). After 35 days of habitation, all mice were returned to the Earth and processed. Here, we employed transcriptome sequencing (RNA-Seq) to detect alterations in gene expression profiles in the liver tissues among the MG, AG and GC (ground control) mice.

overall_design: mRNA profiles of liver were generated by RNA sequencing using the NextSeq 500 (Illumina).

first: Akiyoshi

last: Fukamizu


title: HP1B and H3K9me3 Regulate Olfactory Receptor Choice and 2 Transcriptional Identity [ChIP-seq]

first: Martin

last: Escamilla

overall_design: Native - ChIP (no PFA fixation) was perform on FAC sorted cell populations from the Main olfactory epithelium. Mice Olfatory ephitelium was microdisected for Zone-1 (z1) and Zone-5 (z2) ORs.

last: Escamilla del Arenal

first: Rachel

last: Duffié

first: Hani

last: Shayya

first: Valentina

last: Loconte

first: Axel

last: Ekman

first: Lena


title: HP1B and H3K9me3 Regulate Olfactory Receptor Choice and 2 Transcriptional Identity [CUT&Run]

summary: Diverse epigenetic regulatory mechanisms ensure and regulate cellular diversity. Among 18 others, the histone 3 lysine 9 me3 (H3K9me3) post translational modification participates 19 in silencing lineage-inappropriate genes. H3K9me3 restricts access of transcription factors 20 and other regulatory proteins to genes that control cell fate. Mouse olfactory sensory neu- 21 rons (OSN) select to express one olfactory receptor (OR) gene out of 2,600 possibilities. 22 This monoallelic and stochastic OR choice happens as OSNs differentiate and undergo 23 dramatic changes in nuclear architecture. OR genes from different chromosomes converge 24 into specialized nuclear bodies and chromatin compartments, as H3K9me3 and chromatin 25 binding proteins including heterochromatin protein 1 (HP1) are incorporated. In this 26 work, we have uncovered an unexpected role for HP1b in OR choice and neuronal iden- 27 tity that cannot be rescued by HP1a in vivo. With the use of a conditional knock-in mouse 28 model that replaces HP1b for HP1a, we observe changes in H3K9me3 levels and DNA 29 accessibility over OR gene clusters. These changes alter the expression paTerns that par- 30 tition the mouse olfactory epithelium into five OR expression zones, which results in a 31 reduced OR repertoire leading to a loss of olfactory sensory neuron diversity. We propose 32 that HP1b modulates the competition of OR-promoters for enhancers to promote receptor 33 diversity, by establishing repression gradients in a zonal fashion.

overall_design: Cut & Run using the Cutana kit from FAC sorted cell populations from the Main olfactory epithelium. Mice Olfatory ephitelium was microdisected for Zone-1 (z1) and Zone-5 (z2) ORs. Cell extracs were prepere from a mix of mice: WT, Cbx (control) and Cbx.Foxg1CRE. H3K9me3 ChIP was used a an analysis control

last: Street

first: Carolyn

last: Larabell

first: Marko

last: Jovanovic

first: Stavros

last: Lomvardas


title: PDS5 proteins control genome architecture by limiting the lifetime of cohesin-NIPBL complexes [ChIP-Seq]

city: Vienna

first: Roman

last: Stocsits

address: Campus-Vienna-Biocenter 1

country: Austria

institute: IMP - Research Institute of Molecular Pathology

summary: Cohesin-NIPBL complexes extrude genomic DNA into loops that are constrained by CTCF boundaries. This process has important regulatory functions and weakens the separation between euchromatic and heterochromatic compartments. Cohesin can also bind PDS5A or PDS5B, which do not support loop extrusion but are required for the formation of CTCF boundaries. How PDS5 proteins perform this function is unknown. Here we show by in vitro single-molecule imaging that PDS5 proteins stop loop extrusion by facilitating the dissociation of NIPBL from cohesin. Hi-C experiments suggest that this function is required for the establishment of CTCF boundaries in cells. In silico modelling indicates that PDS5 proteins enable the separation between compartments by limiting cohesin’s velocity and chromatin-residence time. The degree of this compartmentalization depends on the frequency with which chromatin is extruded relative to the time it takes for compartments to form. These results identify PDS5 proteins as key regulators of genome organization.

overall_design: The objective of this experiment was to investigate the role of cohesin and CTCF in positioning in different depletion conditions.

first: Wen

last: Tang

first: Gordana

last: Wutz

first: Jan-Michael

last: Peters


title: PDS5 proteins control genome architecture by limiting the lifetime of cohesin-NIPBL complexes [Hi-C]

sample_organism:
Gallus gallus
Homo sapiens

platform_organism:
Gallus gallus
Homo sapiens

overall_design: Hi-C to analyze the effect of mutations on chromatin architectures in different conditions


title: Influence of different pClik 5? plasmid-variants (native vs. high-copy) on mCherry expression in genome-reduced C. glutamicum CR099

city: Saarbruecken

first: Michael

last: Kohlstedt

address: Campus A1.5

institute: Saarland University

summary:
Whole transcriptome analysis of CR099 pClik 5? (= reference strain harbouring empty plasmid), CR099 pClik 5? repAmut (= strain harbouring empty high copy plasmid),
CR099 pClik 5? Ptuf mCherry (= reference strain harbouring mCherry-carrying plasmid), and CR099 pClik 5? repAmut Ptuf mCherry (= strain harbouring mCherry-carrying high copy plasmid)

overall_design:
CR099 strains harboring the native pClik 5? plasmid or its repAmut derivative, each either carrying or lacking the mCherry expression cassette, were cultivated in BHI medium and sampled in triplicates during mid-exponential growth
Experiments were conducted in mini-bioreactors (BioLector) under standardized cultivation conditions (1300 rpm, 30°C, 80% humidity).

first: Jens

last: Christmann

first: Christoph

last: Wittmann


title: HP1B and H3K9me3 Regulate Olfactory Receptor Choice and 2 Transcriptional Identity

last: Escamilla

overall_design: Single Cell RNA-Seq from FAC sorted cell populations from the Main olfactory epithelium. All mice were transgenic for gg8-tTA; tetOP2iresGFP or gg8-tTA constructs. Driven by gg8-tTA (Nguyen et al. 2007, PMID: 18045541).


title: An Engineered Viral RNA Degrader on Mitochondrial Surface that Mitigates RNA Virus Infection

city: Taipei

first: Wei-Ting

last: Liao

address: No. 7, Chung-Shan S. Rd., Chung-Cheng District

institute: NTU

summary: Infections by RNA viruses cause diseases. Host factor(s) that restrain viral propagation might offer opportunities for designing new anti-virus treatments. We used vesicular stomatitis virus (VSV) as a study model, and observed the synthesis of VSV RNAs at mitochondria /endoplasmic reticulum (mito/ER) spheres accompanied by the leakage of endonuclease G (ENDOG), a mitochondrial nuclease, to the cytosol. We provide evidence that ENDOG released from mitochondria is a host anti-viral factor by eliminating viral RNAs for replication. However, ENDOG outside mitochondria can be translocated to nuclei to cause nuclear DNA damages. We engineered an ENDOG expressed on mitochondrial outer membrane (MOM), namely MOM-ENDOG, to increase the accessibility to viral RNA transcripts synthesized at mitochondrial sites without damaging nuclear DNA (nDNA). Delivery of modified mRNA of wild-type but not catalytic-dead MOM-ENDOG markedly suppressed not only the propagation of VSV, but also Dengue, and Zika virus. Thus, this organelle-specific viral RNA degrader might be developed as a broad-spectrum anti-viral agent.

overall_design: RNA-seq profiling of A549 cells with MOM-GFP overexpression vs MOM-ENDOG overexpression

first: Zee-Fen

last: Chang

first: Chih-Wei

last: Chen

first: Tung

last: Chao

first: Yi-Han

first: Shih-Che

last: Weng

first: Shin-Hong

last: Shiao

first: Yun-Jung

last: Li

first: Kai-An

last: Cheng


title: Ovarian Tumor FAK Inhibition Releases Omega-3 Fatty Acids Stimulating GATA6 Peritoneal Macrophage CXCL13 Production Enhancing Immunotherapy II

city: La Jolla

first: Dwayne

middle: G

last: Stupack

address: 3855 Health Sciences

department: Obstetrics and Gynecology

institute: UC San Diego Moores Cancer Center

summary: Supernatants from murine KMF tumors lacking or expressing focal adhesion kinases were added to macrophage to determine the impact of each tumor line on macrophage gene expression.

overall_design: Peritoneal macrophages (PMs) were isolated and cultured in growth medium (RPMI-1640 supplemented with 10% fetal bovine serum and 1% penicillin-streptomycin). After 24 h, 1:1 volume of tumor cell CM and fresh PM growth medium were added. After 72 h, PMs were harvested, and total RNA was extracted

first: Xiao

first: David

last: Schlaepfer


title: 2,3-Bisphosphoglycerate mutase (BPGM) defines a stress-resilient metabolic state in clear cell renal cell carcinoma

city: Berlin

first: Robert

last: Labes

address: Charitéplatz 1

department: Institute of Translational Physiology

institute: Charité - Universitätsmedizin Berlin

summary: Background: Clear cell renal cell carcinoma (ccRCC) is characterized by profound metabolic reprogramming and limited responsiveness to diverse therapeutic stressors, including epigenetic modulation. How glycolytic enzymes contribute to metabolic stress tolerance in ccRCC remains incompletely understood. Methods: We investigated the role of the glycolytic enzyme 2,3-bisphosphoglycerate mutase (BPGM) in ccRCC using human tumor specimens, siRNA-mediated gene silencing, functional cell-based assays, and transcriptomic profiling. Epigenetic stress was induced using the histone deacetylase inhibitor Vorinostat as a tool compound. Results: BPGM expression was consistently elevated in human ccRCC tissue compared with adjacent normal kidney. ccRCC A498 cells exhibited high basal BPGM levels and limited sensitivity to Vorinostat, whereas BPGM depletion increased cellular stress responses and reduced proliferative capacity. Despite inducing similar phenotypic outcomes, BPGM silencing and Vorinostat treatment triggered markedly distinct transcriptional programs. While HDAC inhibition caused broad, unspecific gene deregulation, BPGM loss elicited a focused stress-associated response, including activation of unfolded protein response and ferroptosis-related gene signatures. Conclusions: Our data identify BPGM as a determinant of metabolic stress resilience in ccRCC. By shaping selective transcriptional stress responses rather than global epigenetic reprogramming, BPGM may contribute to the intrinsic robustness of renal cancer cells. Targeting metabolic stress adaptation pathways may therefore complement epigenetic strategies in ccRCC.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for A498 cells treated with vorinostat or under conditions with and without BPGM knockdown.

last: Faehling

first: Philipp

middle: N

last: Becker


title: Interleukin-27 remodels the bone marrow niche to suppress B-cell development and leukaemia progression in mouse models

first: duan

middle: cai

last: wen

address: 1678 dongfang road

institute: Shanghai Children's Medical Center

summary: Interleukin-27 (IL-27) is an immunoregulatory cytokine, but its role in B-cell haematopoiesis and B-cell acute lymphoblastic leukaemia (B-ALL) within the bone marrow (BM) niche remains unclear. IL-27 was delivered in vivo using adeno-associated virus. B-cell reconstitution, mixed BM chimeras, and an N-myc-driven B-ALL model were analysed by flow cytometry, transcriptional profiling, and survival studies. Group comparisons were assessed using Student’s t-test, and survival was evaluated by Kaplan-Meier analysis with log-rank tests. Sustained IL-27 expression selectively impaired B-cell reconstitution while preserving overall haematopoietic recovery, with marked reductions in common lymphoid progenitors and early B-cell subsets. IL-27 directly inhibited early B-lineage differentiation and concurrently remodelled the BM microenvironment by downregulating VCAM-1, ICAM-2, CXCL12, and IGF-1. These niche alterations were associated with reduced BM-resident B-ALL burden, enhanced chemotherapy efficacy, and improved survival in B-ALL-bearing mice. IL-27 showed no direct cytotoxicity toward B-ALL cells, supporting an indirect, niche-mediated mechanism. IL-27 constrains B-cell haematopoiesis and B-ALL progression through coordinated progenitor inhibition and BM niche remodelling, revealing a cytokine-driven strategy with the potential to enhance leukaemia therapy.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: To investigate how exogenous IL-27 remodels the bone marrow (BM) microenvironment and regulates B-cell development, mice were treated with AAV-IL-27 or control vectors. Bone marrow samples were collected at defined time points following treatment for transcriptomic analysis. Bulk RNA sequencing was performed on total BM cells, splenic B cells, and BM-derived B-ALL cells to evaluate IL-27–induced transcriptional changes associated with niche remodelling and B-cell developmental regulation.

first: In parallel

last: human mesenchymal stromal cells (hMSCs) were cultured in vitro in the presence or absence of IL-27 to assess direct transcriptional responses of stromal components to IL-27 stimulation. Cells were harvested for bulk RNA sequencing analysis. Independent biological replicates were included in all experimental groups to enable differential gene expression and pathway enrichment analyses between conditions.

first: Kaiming

first: Xuefeng

last: Bai

first: Caiwen

last: Duan


title: Niche-dependent modular regulation of the stem cell transcriptome separates cell identity and potential

city: Cambridge

first: Amelie

last: Raz

state: MA

address: 455 Main St

institute: Whitehead Institute for Biomedical Research

summary: Adult stem cells maintain tissue homeostasis, yet are themselves vulnerable to loss. One common mechanism to replace lost stem cells is dedifferentiation, in which progeny revert to stem cell identity. It is a paradox how stem cells and progeny retain the same stem cell potential while exhibiting distinct current identities of self-renewal, differentiation, and dedifferentiation. Here, we show that the Drosophila male germline lineage solves this paradox via two parallel and complementary mechanisms to separate potential and identity. First, differentiating progeny maintain stem cell potency by inheriting perdurant stem cell mRNAs without actively transcribing them. Second, two known niche signals (Bmp and Jak-Stat) activate distinct sets of targets, defining three identities (self-renewal, differentiation, and dedifferentiation) based on the combination of their on/off states. Together, this study reveals how a pool of dedifferentiation-competent progeny is maintained to regenerate stem cells as needed without resulting in stem cell overproduction, and resolves the puzzle of why most stem cell systems require multiple independent niche signals.

overall_design: This dataset was collected from testes of adult Drosophila males. Testes were dissected, dissociated in trypsin/collaganase solution, and subjected to capture on the 10X Chromium controller. Samples were sequenced on the NOVASEQ S4

first: Amelie A


title: Caspase-6 Controls Lipid and Energy Metabolism in Diet-Induced Obesity

city: San Antonio

first: Xiaoli

last: Sun

state: TX

address: 7703 Floyd Curl Dr

institute: University of Texas Health Science Center at San Antonio

summary: Caspases are cysteine proteases that regulate programmed cell death. While caspase-6, an executioner caspase, is known for its role in neurodegeneration and cell death, its broader physiological functions remain poorly understood. Our previous study revealed that caspase-6 drives liver injury and fibrosis in metabolic dysfunction-associate steatohepatitis. Here, we report that caspase-6 deficiency protects against high fat diet-induced obesity. Both global and adipocyte-specific caspase-6 knockout mice exhibit increased energy expenditure, reduced adiposity and inflammation, and improved glucose metabolism. Mechanistically, caspase-6 directly cleaves peroxisome proliferator-activated receptor gamma (PPARγ) and its cofactor specificity protein 1 (SP1), thereby suppressing adipose triglyceride lipase (ATGL) expression. Caspase-6 deficiency restores ATGL, enhancing lipolysis and elevating fatty acyl esters of hydroxy fatty acids (FAHFAs), which alleviate inflammation and enhance insulin sensitivity. These findings uncover a novel Casp6-PPARγ/SP1-ATGL axis in adipose tissue and establish caspase-6 as a potential therapeutic target for obesity and insulin resistance.

overall_design: RNA-seq profiling of epididymal (eWAT) and inguinal (iWAT) white adipose tissue, as well as brown (BAT) adipose tissue from wildtype or Caspase6 knockout (KO) mice fed a 60% high fat diet (HFD).

first: Peng

last: Zhao


title: Progression of Nipah virus infection in African green monkeys: role of macrophages and dendritic cells in early virus spread

city: Galveston

first: Jacquelyn

last: Turcinovic

address: Galveston National Laboratory, 301 University Blvd.

department: Microbiology and Immunology

institute: University of Texas Medical Branch

summary: Nipah virus (NiV) infection causes severe respiratory disease and encephalitis in humans and nonhuman primates with high case fatality rates.  While end-stage disease in primates has been well studied, particularly in African green monkeys (AGM), little is known about the development of NiV disease. Here, 12 AGM were infected with NiV and examined sequentially over 5 days to investigate the pathological and immunological events of NiV infection leading to death. DC-SIGN-positive immature dendritic cells in tonsil and alveolar macrophages were identified at 3 days postinfection (DPI) as early targets of NiV, implicating their important role in the spread of NiV to other tissues.  By 5 DPI, NiV spread to other tissues and cell types, including endothelium of medium caliber vessels and respiratory epithelium. Multifocal regions of fibrin deposition in small and medium caliber vessels and into alveolar spaces were also noted at 5 DPI. NiV antigen was not detected in cells of central nervous system tissues from any AGM at 3-5 DPI, indicating that neuroinvasion is a late event in untreated primates.  NiV infection elicited a biphasic host response characterized by early loss of anticoagulant factors followed by robust induction of interferon-stimulated cytokines, chemokines, and endothelial activation markers, revealing a tightly coupled inflammatory-coagulatory cascade underlying disease progression. RNA-seq analyses of tonsil and lung identified a robust, complement-driven immune response in tonsil but a muted, interferon-driven response in lung. The sequence of pathogenetic events identified here may provide insight into developing new therapeutic interventions and optimizing lead candidate antivirals.

overall_design: Tonsil and lung samples from 0, 3, 4, and 5 days postinfection. Each tissue's postinfection timepoints are compared to health baseline

first: Thomas

last: Geisbert


title: Lung microbiota coordinates alveolar macrophage and γδ T cell activation against Klebsiella pneumoniae via SCFA-FFAR2 axis

city: Taipei city

first: Ting-Chieh

last: Huang

state: Taipei City (TPE)

address: 8F., No. 161, Sec. 6, Minquan E. Rd., Neihu Dist.

department: Graduate Institute of Microbiology and Immunology

institute: National Defense Medical University

summary: Here, we show that intratracheal vancomycin-induced lung dysbiosis renders mice more susceptible to K. pneumoniae and dampens immune responses to LPS, characterized by reduced IL-17A production and impaired activation of IL-17A+ γδ T cells in the lung. This effect is dependent on decreased IL-1β secretion from alveolar macrophages (AMs), accompanied by restrained NF-κB and GPR signaling. Additionally, lung dysbiosis reduces microbiota-derived SCFAs in the lung, while FFAR2, their major receptor, is predominantly expressed on AMs. Furthermore, acetate activation of FFAR2 on AMs enhances NF-κB phosphorylation, inflammasome activation, and IL-1β production, promoting IL-17A+ γδ T cell activation. Moreover, acetate supplementation rescued lung dysbiosis-suppressed immune responses and host defense in a FFAR2-dependent manner on AMs. In conclusion, lung microbiota-derived SCFAs coordinate the activation of AMs and γδ T cells through FFAR2, suggesting that lung dysbiosis compromises the protective role of commensal bacteria in modulating innate immune network against K. pneumoniae infections.

overall_design: FACS-sorted AMs were isolated from the BALF of PBS or Van-treated WT C57BL/6 mice following LPS stimulation. The transcriptional profiling was evaluated by bulk RNA sequencing.

first: Jheng-Syuan

last: Shao


title: Synergistic blockade of cyclophilin A and pathogenic immunity minimizes pregnancy-context pathologies in response to the Zika virus

city: 厦门市

first: CHEN

last: JIAHAO

state: 陈嘉濠

address: 福建省福州市闽侯县南通镇苏坂村苏坂东路38号

institute: 厦门大学

summary: Zika virus (ZIKV) jeopardizes fetal health through both vertical transmission and placental disorders, while the regulatory mechanisms and therapeutic opportunities remain elusive. Here, we reveal cyclophilin A (CypA) as a critical host factor necessary for placental ZIKV replication, functioning independently of its isomerase activity and calcineurin-dependent pathway.

overall_design: Total RNA was extracted from mouse placentas using RNAprep Pure Cell Kit according to the manufacturer’s protocol. High-depth transcriptomic sequencing was conducted on the BGISEQ-500 platform (China, BGI). The obtained raw sequencing reads are aligned to the mouse reference genome (GRCm39) using HISAT2 alignment tools. Gene expression levels are quantified with featureCounts. Differential gene expression analysis was performed using DESeq2 (v1.32.0), with a significance threshold set at p-value < 0.05 and fold change > 1.5.


title: DEVELOPMENTAL LANDMARKS AND CELLULAR TRANSITIONS DURING EXTRAVILLOUS TROPHOBLAST CELL DIFFERENTIATION [scRNA-Seq]

city: Kansas City

first: Michael

last: Soares

state: KS

address: 3901 Rainbow Blvd

department: Pathology

institute: University of Kansas Medical Center

summary: Human trophoblast stem (TS) can be captured, maintained in vitro under specific conditions, and differentiated into extravillous trophoblast (EVT) cells. The regulatory mechanisms that govern the self-renewal and differentiation of human TS cells into EVT cells are largely unknown.  In this study, bulk RNA-sequencing (RNA-seq) and single cell RNA-seq (scRNA-seq) were performed on human TS cells maintained in the stem state and on cells progressing from the stem state into EVT cells (differentiation days 3, 6, and 8). Distinct bulk and single cell transcript profiles were identified for each day of analysis. Day 3 of EVT cell differentiation represented a striking transition point and was readily distinguished from stem state and days 6 and 8 of EVT cell differentiation. Analysis of scRNA-seq led to the identification of several unique cell populations. Trophoblast cell developmental state-specific regulons and trajectories. We elucidated functional roles of key regulators of EVT cell development: cyclin B1, CCAAT/enhancer-binding protein beta, and ADAM metallopeptidase with thrombospondin type 1 motif 20. Collectively, we have defined developmental landmarks and transitional cell populations during EVT cell differentiation. These findings provide a valuable resource and foundation for future investigations into regulatory mechanisms controlling TS cell differentiation into the EVT cell lineage

overall_design: Single-cell RNA sequencing (scRNA-seq) was performed on CT27 trophoblast stem (TS) cells (Okae et al.) maintained in the stem state, as well as on cells differentiating into EVT cells at days 3, 6, and 8. Two biological replicates were collected for each condition (D0, D3, D6, and D8). In addition, day 6 EVT samples were collected from three TS cell lines: CT27, CT29, and CT20.

first: Ayelen

last: Moreno-Irusta

first: Malay

middle: K

last: Basu

first: Esteban

last: Dominguez

last: Chen

first: Sawyer

last: Smith

first: Kaela

last: Varberg

first: Hiroaki

last: Okae

first: Takahiro

last: Arima


title: Enhanced immune response of human iPSC-macrophages via animal-free manufacturing processes

city: Hannover

first: Maximilian

last: Schinke

address: Nikolai-Fuchs-Str. 1

institute: Fraunhofer ITEM

summary: Studying human macrophages in vitro is crucial for understanding their immune functions. Recent interest in macrophages derived from induced pluripotent stem cells (iPSC-Mac) has led to various differentiation protocols, often using undefined animal serum like fetal bovine serum (FBS). This study compared FBS-based media to the defined medium X-VIVO™ 15 containing human serum albumin (hSA). Both produced functional macrophages, but those in hSA displayed a neutral activation state, enhanced cholesterol metabolism, and more significant changes in surface phenotype after stimulation. They also had a stronger response to co-stimulatory signals, making them better for testing immunomodulatory drugs. In contrast, FBS-grown macrophages were more adherent and pre-activated, showing reduced sensitivity upon stimulation, which may pose challenges for drug screening and immunomodulatory evaluations.

overall_design: Human iPSC-derived macrophages (iPSC-Mac) from one iPSC line were harvested at two different time points and terminally differentiated in either RPMI + 10% FBS or X-VIVO™ 15 medium for three days, in the presence of 50 ng/mL M-CSF. Subsequently, cells were harvested for RNA-Seq. As additional control served the freshly harvested iPSC-Mac prior to terminal differentiation.

first: Ingrid

last: Gensch

first: Tina

last: Rietschel

first: Nguyen

last: Ariane NH

first: Slevogt

last: Hortense

first: Lothar

last: Jaensch

first: Nico

last: Lachmann


title: Spatially resolved subtype progression reveals metabolic vulnerabilities in pancreatic ductal adenocarcinoma.

city: Göttingen

first: Lena-Christin

last: Conradi

address: Robert-Koch-Straße 40

department: Department of General, Visceral and Pediatric Surgery

institute: University Medical Center Göttingen

summary: Pancreatic ductal adenocarcinoma (PDAC) exhibits profound molecular heterogeneity and poor prognosis, necessitating novel tailored therapies. The basal and classical subtypes - driven by glycolysis versus lipid metabolism - have distinct prognostic implications. We mapped PDAC molecular subtype heterogeneity, capturing spatially-resolved gene expression signatures and generating a comprehensive high-resolution dataset of 42,035 spatial spots. Subtype assignments were validated via multiplex immunofluorescence and quantitative analyses in patient-derived organoids. Our analysis resolved cancer cell signatures, deconvoluted intra-tumoral heterogeneity, and delineated a classical-to-basal trajectory. We identified metabolically ‘hot’, high-grade tumor niches characterized by concurrent enrichment of glycolysis and lipogenesis across both subtypes, nominating them as subtype-agnostic therapeutic targets. Preclinical models demonstrated that despite the basal subtype’s glycolysis dependence, both classical and basal tumors are susceptible to glycolysis inhibition. This work challenges the dogma of subtype-specific therapeutic silos and demonstrates highly adaptable energetic niches as reservoirs to drive tumor progression.

overall_design:
RNA-seq profiling of 2 untreated PDX lines (classical PDX line generation F2 and F4; basal PDX line generation F1) generated from PDAC tissue from 2 different patients.

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


title: miRNA expression profiling of mouse hearts following myocardial infarction

city: Durham

first: Walter

last: Koch

address: 487 CARL Building, 213 Research Drive

department: Surgery

institute: Duke University School of Medicine

summary: Background: Pathological upregulation of G protein-coupled receptor kinase 2 (GRK2) is a hallmark of heart failure and contributes to maladaptive signaling, hypertrophic remodeling, and cardiomyocyte death. MicroRNAs (miRNAs) are key post-transcriptional regulators of cardiac stress responses, however whether GRK2 is subject to miRNA targeting has not yet been fully established.    Objective: We investigated potential miR-181a-mediated regulation of GRK2 in cardiomyocytes to better understand its effects on cardiac stress responses.    Methods: miRNA microarray profiling was performed on mouse hearts two weeks after myocardial infarction. Bioinformatic target prediction analysis identified candidate miRNAs that are predicted to bind GRK2 in the 3’ untranslated region (UTR). Direct binding was assessed using luciferase reporter assays, and miR-181a was selected as the miRNA of interest to further pursue mechanistic and functional validation. miR-181a was overexpressed or inhibited in neonatal rat ventricular myocytes (NRVMs) and exposed to several modes of cellular stress to induce hypertrophy, hypoxia, and accumulation of reactive oxygen species. GRK2 expression, hypertrophic remodeling, oxidative stress, cell viability, and cyclic AMP (cAMP) signaling were assessed using quantitative PCR, immunoblotting, fluorescence imaging, and biochemical assays.    Results: miR-181a directly targeted the GRK2 3′UTR and suppressed GRK2 expression at both mRNA and protein levels. miR-181a overexpression attenuated stress-induced hypertrophic gene expression, reduced cardiomyocyte cell size, decreased oxidative stress, improved survival under hypoxia, and enhanced cAMP production under β-AR stimulation. Conversely, inhibition of miR-181a resulted in sustained GRK2 expression, exacerbated hypertrophic signaling, and decreased cAMP production.    Conclusion: These findings identify miR-181a as a functional post-transcriptional regulator of GRK2 that limits maladaptive signaling, hypertrophic remodeling, and cardiomyocyte injury. miR-181a-mediated GRK2 inhibition represents a potential therapeutic strategy for mitigating pathological signaling in heart failure.

overall_design: Male C57BL/6J mice aged 9-12 weeks underwent sham or MI surgery. At 2 weeks post-MI, total RNA was extracted from the left ventricular (LV) tissue of the mouse myocardium.

first: Heidi

last: Cho

first: Melissa

last: Lieu

first: Erhe

last: Gao

first: Samuel

last: Slone


title: Restoration of Spermatogenesis is Dependent on Activation of a SPRY4-ERK Checkpoint Following Germline Stem Cell Damage

city: New York

first: Ying

last: Liu

state: NY

address: 1300 York Ave

institute: Weill Cornell Medical College

summary: Mammalian spermatogonial stem cells (SSCs) sustain male fertility through continuous self-renewal and differentiation, leading to the production of haploid spermatozoa throughout adulthood. However, SSCs are vulnerable to genotoxic drugs, and patients receiving chemotherapy face a high risk of germline instability and infertility. The molecular mechanisms and cellular pathways that choreograph SSC recovery after chemotherapeutic insult remain unknown. Previously, we identified SPRY4 as an ERK-dependent negative feedback regulator of growth factor signaling that is critical for preservation of stem cell activity in cultured mouse SSCs. Here, we demonstrate that following alkylating agent busulfan (BU)-induced injury in adult mice, germline-specific Spry4 gene deletion (Spry4G-KO) reduces stem cell regeneration with an enhanced genotoxic stress response and differentiation with rapidly enhanced nuclear ERK1/2 activity in undifferentiated (Aundiff) spermatogonia (including SSCs). Genes essential for stem cell maintenance, including Id1 and Cxcl12, were dysregulated by loss of Spry4. Furthermore, the MEK1/2 inhibitor PD0325901, but not mTORC1 inhibitor Rapamycin, was sufficient to promote spermatogonial proliferation in Spry4G-KO testis 10 days post-BU treatment. Notably, the restoration of both spermatogonia pool and fertility was delayed in adult Spry4G-KO males long-term after injury. In summary, germline-specific deletion of Spry4 results in hyper-activation of the MAPK/ERK pathway in Aundiff spermatogonia, reducing spermatogonial genome integrity, unleashing excessive spermatogenesis after germline damage, and ultimately impairing germline regeneration in adult males. Our study indicates an essential role for SPRY4-ERK signaling as a molecular checkpoint in securing SSC recovery upon chemotherapy drug-induced germline damage, revealing how stem cells normally withstand environmental stress.

overall_design: RNA-seq profiling of undifferentiated spermatogonia FACS-isolated from adult wild-typ and Spry4 G-KO mice at 10 days following busulfan and PD0325901 or vehicle (DMSO) treatment.

first: Tansol

last: Choi

first: Brad

last: Pearson

first: Ryan

last: Nachman

first: Whitney

last: Woo

last: Xu

last: Schreiner

first: Romulo

last: Hurtado

first: Marco

last: Seandel

first: Shahin

last: Rafii

first: Todd

last: Evans


title: DEVELOPMENTAL LANDMARKS AND CELLULAR TRANSITIONS DURING EXTRAVILLOUS TROPHOBLAST CELL DIFFERENTIATION

first: Michael

last: Soares

overall_design: Bulk RNA-sequencing (RNA-seq) were performed on CT27 Trophoblast stem cells (TS- Okae et.al.)  maintained in the stem state and on cells progressing  into EVT cells differentiation (differentiation days 3, 6, and 8). 3 biological replicates each conditions (D0, D3,D6 and D8)

first: Ayelen

last: Moreno-Irusta

first: Malay

last: Basu.

first: Esteban

last: Dominguez

first: Thomas

last: Chen

first: Sawyer

last: Smith

first: Kaela

last: Varberg

first: Hiroaki

last: Okae

first: Takahiro

last: Arima


title: Positive Feedback Regulation of Glycolysis by H4K5 Lactylation Promotes Malignant Progression of Hepatocellular Carcinoma [RNA-seq]

city: Bunkyo-Ku

first: Masahiro

last: Yamane

state: Tokyo

institute: Institute science  Tokyo

summary: [Background & Aims] Histone lactylation is a lactate-dependent epigenetic modification driven by glycolysis. Although elevated histone lactylation has been reported in various cancer, the biological relevance of histone H4 lactylation in HCC remains unclear. This study aimed to investigate the clinicopathological relevance of H4K5 lactylation (H4K5la) and its role in regulating lactate metabolism to promote HCC progression. [Methods] The expression levels of H4K5la, H3K18la, and PanKla were evaluated in HCC tissues by immunohistochemistry. RNA-seq and ChIP-PCR analyses were performed on HCC cells treated with lactate to identify the downstream target genes mediated by H4K5la. Functional validation of these targets was conducted through knockdown and overexpression experiments, and biological effects were assessed both in vitro and in vivo. [Results] Elevated levels of H4K5la, H3K18la, and PanKla were associated with a poor prognosis, with H4K5la identified as an independent prognostic factor of overall survival. Upon lactate treatment, five genes including glycolytic enzyme ENO2 were consistently identified in three HCC cell lines, showing enrichment of H4K5la at their promoter regions. ENO2-KD reduced intracellular lactate and H4K5la levels, as well as in vivo tumorigenicity in HCC cells, while ENO2 overexpression exerted opposite functions. These findings establish a positive feedback loop: H4K5la increases ENO2 expression, which in turn amplifies glycolysis and lactate production, further elevating H4K5la. Moreover, ENO2 expression was tightly correlated with H4K5la levels in HCC tissues. [Conclusions] High H4K5la expression is an independent prognostic biomarker in HCC. H4K5la promotes HCC aggressiveness by enhancing glycolysis and tumor growth, forming a positive feedback loop with ENO2. Targeting this pathway may represent a promising therapeutic strategy for HCC.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for the HLD, PLC/PRF/5, SNU475 with or without lactate treatment.

first: Shu

last: Shimada


title: Comprehensive Molecular and Functional Analysis of NUTM1-Rearranged Leukemia [methylation array]

city: Suita

first: Masaki

last: Nomura

state: Osaka

address: 2-2, Yamadaoka

department: Department of Cancer Pathology

institute: Graduate School of Medicine, The University of Osaka

summary: NUTM1-rearrangement (NUTM1-r) defines a significant subset of B-cell acute lymphoblastic leukemia (B-ALL), particularly in infants lacking KMT2A-rearrangements (KMT2A-r), yet its underlying molecular characteristics remain poorly understood. Here, we establish that NUTM1-r leukemia is a discrete entity characterized by a unique transcriptional and epigenetic landscape, notably featuring global DNA hypomethylation, irrespective of the 5' fusion partner. Functional interrogation of NUTM1 fusions reveals a dual oncogenic role: they drive commitment towards the B-lymphoid lineage while concurrently conferring potent leukemic stem cell properties. Strikingly, expression of a representative fusion, BRD9-NUTM1, is sufficient to induce serially-transplantable prepro-B-like leukemia in vivo, faithfully recapitulating the key molecular and immunophenotypic features of human NUTM1-r B-ALL. Mechanistically, NUTM1 fusions establish an aberrant chromatin state, marked by global enhancement of H3K27 acetylation and the creation of distinctive open chromatin regions that co-opt both B-lineage and stemness-related transcriptional programs, including those involving NF-κB and posterior HoxA genes. In stark contrast to resistant KMT2A-r leukemias, NUTM1-r leukemic cells exhibit a profound sensitivity to chemotherapy. This vulnerability is mechanistically linked to the leukemia's dependence on active transcription. Our findings delineate the unique molecular profile of NUTM1-r leukemias, revealing specific vulnerabilities that rationalize their favorable clinical outcomes and suggest opportunities for modified therapeutic strategies.

overall_design: BRD9-NUTM1 (BN1), MLL-AF9 (MA9) and MLL-ENL (ME) murine leukemic cells were sorted by FACS. DNA was extracted.

first: Koutarou

last: Nishimura

first: Weijia

last: Zang

first: Daichi

last: Inoue


title: Positive Feedback Regulation of Glycolysis by H4K5 Lactylation Promotes Malignant Progression of Hepatocellular Carcinoma [ChIP-seq]

overall_design: Chromatin immunoprecipitation (ChIP) assays were conducted using a SimpleChIP® Plus Sonication Chromatin IP Kit (CST) and ChIP-IT Express (Active motif) according to the manufacturer’s protocol. ChIP-seq reads underwent quality control (FastQC), trimming (Trimmomatifc), alignment (Bowtie2), and duplicate removal (Picard). Normalized coverage tracks were generated from BAM files. These tracks, along with called peaks, were visualized using the Integrative Genomics Viewer (IGV) to produce figures illustrating signal enrichment.