title: Nicotinamide Mononucleotide Promotes Female Germline Stem Cell Proliferation by Activating the H4K16ac-Hmgb1-Fyn-PLD Signaling Pathway through Epigenetic Remodeling [Hi-C]

first: Ji

last: Wu

address: No. 800, Dongchuan Road

institute: Shanghai Jiao Tong University

summary: Nicotinamide mononucleotide (NMN), an endogenous nucleotide essential for various physiological processes, has an unclear role and regulatory mechanisms in female germline stem cell (FGSC) development. In this study, we demonstrate that NMN significantly enhances FGSC viability and proliferation. Quantitative acetylation proteomics revealed that NMN markedly increases the acetylation of histone H4 at lysine 16 (H4K16ac). Subsequent chromatin immunoprecipitation sequencing (ChIP-seq) and RNA sequencing (RNA-seq) identified high mobility group box 1 (Hmgb1) as a downstream target of H4K16ac, a finding further validated by ChIP-qPCR. Knockdown of Hmgb1 reduced FGSC proliferation by disrupting cell cycle progression, inducing apoptosis, and decreasing chromatin accessibility. High-throughput chromosome conformation capture (Hi-C) analysis showed that Hmgb1 knockdown induced A/B compartment switching, increased the number of topologically associating domains (TADs), and decreased chromatin loop formation in FGSCs. Notably, the chromatin loop at the promoter region of Fyn proto-oncogene (Fyn) disappeared following Hmgb1 knockdown. ChIP-qPCR and dual-luciferase reporter assays further confirmed the interaction between Hmgb1 and the Fyn promoter. Importantly, Fyn overexpression reversed the inhibitory effects of Hmgb1 knockdown on FGSC proliferation. Proteomic analysis suggested this rescue was mediated through the phospholipase D (PLD) signaling pathway, as Fyn overexpression selectively enhanced the phosphorylation of PLD1 at threonine 147 without affecting serine 561. Furthermore, treatment with 5-fluoro-2-indolyldechlorohaloamide, a PLD inhibitor, nullified the pro-proliferative effects of Fyn overexpression. In summary, our findings reveal that NMN promotes FGSC proliferation by activating the H4K16ac-Hmgb1-Fyn-PLD signaling pathway through epigenetic remodeling. These results deepen our understanding of FGSC proliferation and highlight potential therapeutic avenues for advancing FGSC applications in reproductive medicine.

overall_design: To investigate the effect of Hmgb1-knockdown on the development of female germline stem cells (FGSCs), we treated FGSCs with Hmgb1-knockdown lentivirus in vitro. Subsequently, we utilized Hi-C data from both the control group and the Hmgb1-knockdown group to conduct differential gene expression analysis.

first: Hong

last: Zhou

first: Yujie

last: Liu

first: Geng

last: Tian


title: Nicotinamide Mononucleotide Promotes Female Germline Stem Cell Proliferation by Activating the H4K16ac-Hmgb1-Fyn-PLD Signaling Pathway through Epigenetic Remodeling [ATAC-seq]

status: Public on Apr 23 2025

city: Shanghai

state: Shanghai

country: China

type: Genome binding/occupancy profiling by high throughput sequencing

overall_design: To investigate the effect of Hmgb1-knockdown on the development of female germline stem cells (FGSCs), we treated FGSCs with Hmgb1-knockdown lentivirus in vitro. Subsequently, we utilized ATAC-seq data from both the control group and the Hmgb1-knockdown group to conduct differential gene expression analysis.

middle: G


title: Acute TREM2 Blockade Impairs Remyelination by Reducing MAFB-High Microglia

city: Hangzhou

first: Jinchao

last: Hou

state: Zhejiang

address: Binsheng Road

department: Anesthesiology

institute: Zhejiang University

type: Expression profiling by high throughput sequencing

summary: We investigated the role of Triggering Receptor Expressed on Myeloid cells 2 (TREM2) in myelin regeneration in the brain.  TREM2 is a receptor found on microglia, which are crucial for clearing myelin debris and promoting remyelination. Previous studies in a mouse model of demyelination induced by the copper-chelating agent Cuprizone (CPZ) have shown that stimulation of TREM2 with a monoclonal antibody reduces demyelination, while deleting the Trem2 gene in mice impairs remyelination. Here we blocked TREM2 function acutely with an antibody during both the demyelination and remyelination phases and analyzed the impact of the antibody on myelination and gene expression in single cells. We found that blocking TREM2 depleted a specific population of microglia characterized by high expression of the transcription factor MAFB during remyelination. These MAFB-high microglia were crucial for myelin repair, and their depletion led to impairment of myelinating oligodendrocytes. Importantly, we identified MAFB+ microglia in acute and acute-chronic brain lesions from individuals with multiple sclerosis (MS), but not in inactive lesions. We conclude that TREM2 is essential for maintaining a population of MAFB-high microglia that are critical for myelin repair. This finding has important implications for understanding demyelinating diseases like MS and suggests that stimulating TREM2 could be a potential therapeutic strategy.

overall_design: To characterize the role of TREM2 blockade in the CNS during de- and re-myelination, eight-week-old male wild-type (WT) mice were divided into 5 experimental groups. A) Untreated: fed regular chow for 5 weeks. B) CPZ + control (Ctrl) IgG2a: fed 0.2% CPZ diet for 5 weeks to induce demyelination and injected intraperitoneally (i.p.) with Ctrl IgG2a during demyelination. C) CPZ + anti-TREM2: fed 0.2% CPZ diet and injected i.p. with anti-TREM2 during demyelination. D) Remyelination + Ctrl IgG2a: fed 0.2% CPZ diet initially, then returned to regular chow for 2 weeks to allow remyelination, and injected i.p. with Ctrl IgG2a during remyelination. E) Remyelination + anti-TREM2: fed 0.2% CPZ diet initially, then returned to regular chow for 2 weeks, and injected i.p. with anti-TREM2 during remyelination.

first: Roberta

last: Magliozzi

first: Yun

last: Chen

first: Junjie

first: John

last: Wulf II

first: Gregory

last: Strout

first: Xiangming

last: Fang

first: Marco

last: Colonna


title: Downregulation of Scd1 promotes cardiac reprogramming by enhancing mitochondrial metabolism through PGC1α [RNA-seq]

city: wuhan

first: yu

middle: zhang

last: yi

state: hubei

address: donghu street

department: wuhan

institute: wuhan university

summary: Direct cardiac reprogramming of fibroblasts into induced cardiac-like myocytes (iCMs) has emerged as a promising therapeutic strategy to remuscularize injured myocardium. However, the intrinsic metabolic demands that drive the iCM program and maturity remain unclear. Here, we induce efficient iCM production and maturation through metabolic pathway modulations. To explore the metabolic heterogeneity of iCMs, we performed single cell RNA sequencing (scRNA-seq) at reprograming day 14. Using single-cell metabolic flux estimation and flux balance analysis on day 14 iCM, we characterized the metabolic heterogeneity of iCMs and identified fatty acid oxidation (FAO) as a critical factor in iCM conversion. Mechanistically, we proved that Scd1 knockdown activates PGC1α/PPARβ transcription signal, which further promotes cardiac reprogramming, and induced cardiomyocytes demonstrated more frequent beating, calcium oscillation, and higher energy metabolism as evidenced by increased mitochondrial respiration and gene expression associated with FAO, which provides a new strategy and theoretical basis for cardiac regeneration and repair. Our findings collectively highlight FAO as a key determinant of iCM fate and offer new therapeutic avenues for advancing reprogramming strategies.

overall_design: To elucidate the similarities and differences between ZLN005-treated iCM or GW501516-treated iCM at reprogramming day 14 and primary cardiomyocytes.

first: Li

last: Wang

first: Zhenhua

last: Jia

first: Lilin

last: Xiang


title: STING mediates lysosomal quality control and recovery through its proton channel function and TFEB activation

city: Dallas

last: Yan

state: TX

address: 6000 Harry Hines Blvd

department: Immunology

country: USA

institute: University of Texas Southwestern Medical Center

summary: Lysosomes are essential organelles for cellular homeostasis. Defective lysosomes are associated with many human diseases, such as lysosomal storage disorders (LSD). How the cell detects lysosomal defects and then restores lysosomal function remain incompletely understood. Here, we show that STING mediates a common neuroinflammatory gene signature in three distinct lysosomal storage disorders, Galctwi/twi, Ppt1-/-, and Cln7-/-. Transcriptomic analysis of Galctwi/twi brain tissue revealed that STING also mediates the expression of a broad panel of lysosomal genes that are part of the CLEAR (Coordinated Lysosomal Expression and Regulation) signaling pathway, which is regulated by transcriptional factor EB (TFEB). Immunohistochemical and single-nucleus RNA-seq analysis show that STING regulates lysosomal gene expression in microglia in LSD mice. Mechanistically, we show that STING activation in both human and mouse cells leads to TFEB dephosphorylation, nuclear translocation, and expression of target lysosomal genes. In addition, STING-mediated TFEB activation requires its proton channel function, the V-ATPase-ATG5-ATG8 cascade, and is independent of immune signaling. Functionally, we show that the STING-proton channel-TFEB axis plays a role in facilitating lysosomal repair. Together, our data identify STING-TFEB as a lysosomal quality control and recovery mechanism that responds to both genetic and chemically induced lysosomal dysfunction.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: RNAseq profiling of the cerebrum of Galc+/+, Galctwi/twi, Galctwi/twi Sting-/- mice at 32 days of age. RNAseq profiling of mouse embryonic fibroblasts (MEFs) stimulated with DMXAA for 10 hours and BJ1 cells stimulated for 8 hours with diABZI. snRNAseq profiling of the cerebellum of Npc1+/+, Npc1-/- and Npc1-/-Sting1-/- mice at 6 weeks of age.

first: Zhen

last: Tang

first: Cong

last: Xing

first: Antonina

last: Araszkiewicz

first: Kun

last: Yang

first: Wanwan

last: Huai

first: Devon

last: Jeltema

first: Nicole

last: Dobbs

first: Yihe

last: Zhang

first: Lu

middle: O

last: Sun


title: Temozolomide Induces Glioblastoma Dedifferentiation into Highly Malignant Stem-like Cells Through Senescence-Associated Reprogramming via HIF1α/HIF2α Regulation [miRNA-seq]

city: Chongqing

first: liao

last: bin

address: 118 Xingguang Avenue

institute: Chongqing General Hospital, Chongqing University

type: Non-coding RNA profiling by high throughput sequencing

summary: A critical issue is that recurrent glioblastoma multiforme (GBM) after temozolomide (TMZ) exposure becomes more malignant, exhibiting higher invasion and stemness than the primary tumor. However, the detailed mechanism remains to be elucidated. While the majority of GBM cells succumb to TMZ treatment, some enter cell cycle arrest, adopt a senescence-associated secretory phenotype (SASP), and activate senescence-related signaling pathways. These cells later exit senescence, re-enter the cell cycle, and proliferate, forming aggregates with stemness characteristics, including high expression of stemness markers, colony formation, high invasion, migration, and chemotherapy resistance. Critically, these new aggregates promote the invasion, migration, and chemotherapy resistance of surrounding cells. Gene Set Enrichment Analysis (GSEA) and KEGG analysis of miRNA and mRNA sequences indicated that hallmark-hypoxia and HIF1-signaling pathways were activated. We verified that HIF1α and HIF2α levels changed before, during, and after TMZ treatment. Knocking out HIF1α and HIF2α in GBM cells and exposing them to TMZ resulted in fewer senescent cells and aggregates. This study clarifies how recurrent GBM becomes more malignant during and after TMZ treatment and highlights the regulatory roles of HIF1α and HIF2α, emphasizing that preventing senescence cell formation and inhibiting HIF1α and HIF2α expression are crucial for improving therapeutic outcomes.

overall_design: After continuous induction with 50 µM TMZ, U-87MG and U-118MG cells formed resistant cell lines, which were then compared with sensitive cell lines using miRNA-seq.

first: Bin

last: Liao

first: Pan

first: Sheng

last: Gong

last: Zhao


title: Metabolic Reprogramming into a Glycolysis Phenotype Induced by Extracellular Vesicles Derived from Prostate Cancer Cells

city: Cheonan-si

first: Dongsic

last: Choi

address: 31 Suncheonhyang 6-gil, Bongmyeong-dong

department: Department of Biochemistry

country: South Korea

institute: Soonchunhyang University, College of Medicine

summary: Most cancer cells adopt a less efficient metabolic process of aerobic glycolysis with high level of glucose uptake followed by lactic acid production, known as the Warburg effect. This phenotypic transition enables cancer cells to achieve increased cellular survival and proliferation in a harsh low-oxygen tumor microenvironment. Also, the resulting acidic microenvironment causes inactivation of the immune system such as T-cell impairment that favours escape by immune surveillance. While lots of studies have revealed that tumor-derived EVs can deliver parental materials to adjacent cells and contribute to oncogenic reprogramming, their functionality in energy metabolism is not well addressed. In this study, we established prostate cancer cells PC3-AcT resistant to cellular death in an acidic culture medium driven by lactic acid. Quantitative proteomics between EVs derived from PC-3 and PC-3AcT cells identified 935 confident EV proteins. According to cellular adaptation to lactic acidosis, we revealed 159 regulated EV proteins related to energy metabolism, cellular shape, and extracellular matrix. These EVs contained a high abundance of glycolytic enzymes. In particular, PC-3AcT EVs were enriched with apolipoproteins including apolipoprotein B100 (APOB). APOB on PC-3AcT EVs could facilitate their endocytic uptake depending on low density lipoprotein receptor of recipient PC-3 cells, encouraging increases of cellular proliferation and survival in acidic culture media via increased activity and expression of hexokinases and phosphofructokinase. The activation of recipient PC-3 cells can increase glucose consumption and ATP generation, representing an acquired metabolic reprogramming into the Warburg phenotype. Our study first revealed that EVs derived from prostate cancer cells could contribute to energy metabolic reprogramming and that the acquired metabolic phenotypic transition of recipient cells could favor cellular survival in tumor microenvironment.

overall_design: RNA-Seq profiling: 1) PC-3 cells with lactic acid; 2) PC-3 cells with lactic acid and PC-3AcT EV; 3) PC-3AcT cells with lactic acid. Please note that PC-3AcT cells established by continuously exposing PC-3 cells to 3.8 μM lactic acid with four serial passages during 15 days.

first: Chul Won

last: Seo


title: RESET: A reversible TCR-coupled antigen receptor with superior targeting sensitivity and pharmacologically controlled anti-tumor activity

city: Seattle

first: Jill

last: Rosenberg

state: WA

address: 188 E Blaine St

institute: Regeneron Pharmaceuticals

summary: Chimeric antigen receptor (CAR) T cells are effective cancer therapies, particularly in indications with high, stable, and tumor-specific antigen expression. Other settings may require improved targeting sensitivity, controllable targeting selectivity, and/or additional potency enhancements to achieve robust efficacy. Here, we describe a novel receptor architecture called RESET (rapamycin-enabled, switchable endogenous T-cell receptor) that combines: (i) cell surface antigen targeting; (ii) small-molecule regulation; and (iii) the signaling proficiency and inherent sensitivity of native T-cell receptors. RESET-T cells outperformed both constitutive and drug-regulated CAR-T cells and show hallmarks of TCR activation that suggest improved fidelity to native T-cell responses. Pharmacological control then increases safety through toggling T-cell activation between active and resting states and may mitigate T- cell exhaustion caused by continuous antigen exposure. This convergence of drug-regulated targeting and natural immune receptor signal transduction may better replicate the kinetics and physiology of a classical T-cell responses and potentiate more successful and safer immunotherapies.

overall_design: T cells from N=4 healthy donors were transduced with RESET, DARIC, CAR, TCR, or untransduced control were cultured either alone or with A549 cells engineered to overexpress the antigen of interest (CD33 or MAGEA4-HLA-A:0201) in the presence or absence of 1nM rapamycin for 18h. Following coculture, T cells were isolated from tumor cells using MACS sorting, tested for purity using flow cytometry, then RNA was isolated and evaluated by RNA-seq.

first: Jardin

last: Leleux

first: Olmo

last: Sonzogni

first: Rebecca

last: Walker

first: Anita

last: Venkitaraman

first: Sarah

last: Garrison

last: Jin

first: Philip

middle: D

last: Gregory

first: Jordin

last: Jarjour


title: Temozolomide Induces Glioblastoma Dedifferentiation into Highly Malignant Stem-like Cells Through Senescence-Associated Reprogramming via HIF1α/HIF2α Regulation [RNA-seq]

overall_design: After continuous induction with 50 µM TMZ, U-87MG and U-118MG cells formed resistant cell lines, which were then compared with sensitive cell lines using mRNA-seq.

last: Wang


title: Disordered maturation of mitochondrial matrix proteins in adipose tissue leads to local and systemic metabolic and inflammatory derangements in adipocyte-specific Mipep knockout mice

city: Noda

first: Yoshikazu

last: Higami

state: Chiba

address: 2641, Ymazaki

department: Faculty of Pharmaceutical Science

country: Japan

institute: Tokyo University of Science

summary: Most mitochondrial matrix proteins encoded in the nuclear genome are synthesized in the cytoplasm. These proteins subsequently undergo maturation through the cleavage of a signal sequence at the N-terminus by one or two mitochondrial signal peptidases, which is essential for their function within mitochondria. The present study demonstrates that adipocyte-specific knockout of one mitochondrial signal peptidase, mitochondrial intermediate peptidase (MIPEP), resulted in disordered mitochondrial proteostasis of MIPEP substrate proteins and their defective maturation. MIPEP deficiency in white and brown adipocytes suppressed the expression of adipocyte differentiation, lipid metabolism, and mitochondrial biogenesis genes. These alterations led to lipoatrophy in white adipose tissue and the whitening of brown adipose tissue. Additionally, it induced an atypical mitochondrial unfolded protein response and local inflammation in white and brown adipose tissue. Furthermore, it induced fatty liver and splenomegaly and caused systemic impairments in glucose metabolism and inflammation. These findings indicate that maturation defects of certain mitochondrial matrix proteins and subsequent proteostasis disorders in white and brown adipocytes cause chronic and systemic inflammatory and metabolic derangements.

overall_design:
To investigate the function of MIPEP, we established adipose-specific Mipep Knockout (aMKO) mice by crossing Mipepflox/flox mice with Adiponectin-Cre mice.
We then performed gene expression profiling analysis using data obtained from RNA-seq of genital white adipose tissue (gWAT) and brown adipose tissue (BAT) derived from control and aMKO mice (n =4 each).
Comparative gene expressin profiling analysis of RNA-seq data for control and aMKO mice in each gWAT and BAT.

first: Nozaki

last: Yuka

first: Kobayashi

last: Masaki

first: Fukuo

last: Tomoyoshi

first: Ishimatsu

last: Mamiko

first: Narita

last: Takumi

first: Taki

last: Kanari

first: Hirao

last: Yuto

first: Ayabe

last: Shota

first: Yokoyama

last: Miku

first: Otani

last: Yuina

first: Mizunoe

last: Yuhei

first: Matsumoto

last: Mami

first: Ohno

last: Nobuhiko

first: Kaifu

last: Tomonori

first: Okazaki

last: Shogo

first: Goitsuka

last: Ryo

first: Nakagawa

last: Yoshimi

first: Shimano

last: Hitoshi

first: Iwakura

last: Yoichiro

first: Higami

last: Yoshikazu


title: RHOB regulates megakaryocytes and erythroid differentiation by altering the cell cycle and cytoskeleton.

city: beijing

first: Xin

last: Yuan

address: haidian

institute: Beijing Institute of Radiation Medicine

summary: RHOB, a member of the Rho GTPase family, is a target of miR-1915-3p. miR-1915-3p has been proved by us to affect the differentiation of MEPs, promoting the differentiation of megakaryocytes and inhibiting the erythroid lineage, which may lead to an increase in the number of platelets and a decrease in the number of erythrocytes. Although RHOB has been confirmed to be involved in the process of proplatelets production in mice, its role in early hematopoietic differentiation, especially its impact on the megakaryocytic and erythroid differentiation of human HSCs and MEPs, has not yet been elucidated. In this study, we confirmed that the function of the RHOB gene was opposite to miR-1915-3p by knocking down the gene in progenitor cells and inducing them into different cells. Silencing of RHOB significantly decreased the erythroid surface markers, reduced the diameter of the cell and inhibited the expression of erythroid-related genes in human cells. Furthermore, RHOB knockdown in megakaryocytic-induced cells increased the megakaryocytic surface markers, enlarged cell and nuclear volume and promoted cell polyploidy and megakaryocyte-related gene expression. The results of RNA sequencing suggest that the RHOB may play a role in differentiation by influencing the cell cycle and the cytoskeleton. Based on that, we found RHOB could affect the expression of differential CDKs and cyclins to alter the proportions of G0/1 and S phase cells and influent the expression or aggregation of β-ACTIN during erythroid or megakaryocytic differentiation. Taken together, these results suggest that RHOB plays an important role in megakaryocytic and erythroid differentiation by cell cycle and cytoskeleton regulation.

overall_design: To investigate the function of RHOB in megakaryocyte and erythroid differentiation, we knocked down the expression of RHOB in K562 cells by siRNA transfection. We performed gene expression profiling analysis using data obtained from RNA-seq in K562 cell which cultured in growth medium (Ctl) and megakaryocyte differentiation medium (MK) for 3d. Comparative gene expression profiling analysis of negative control group (NC) and RHOB knockdown group (siRHOB).

first: Pengcong

last: Liu

first: Mingyi

last: Qu


title: Palmitic acid alters enhancers and super-enhancers near inflammatory and efferocytosis-associated genes in human monocytes

city: Duarte

first: Rama

last: Natarajan

state: California

address: 1500 E Duarte Rd

department: Department of Diabetes Complications and Metabolism

institute: Beckman Research Institute, City of Hope

summary: Free fatty acids like palmitic acid (PA) are elevated in obesity and diabetes and dysregulate monocyte and macrophage functions, contributing to enhanced inflammation in these cardiometabolic diseases. Epigenetic mechanisms regulating enhancer functions play key roles in inflammatory gene expression, but their role in PA-induced monocyte/macrophage dysfunction is unknown. We found that PA treatment altered the epigenetic landscape of enhancers and super-enhancers (SEs) in human monocytes. Integration with RNA-seq data revealed that PA-induced enhancers/SEs correlated with PA-increased expression of inflammatory and immune response genes, while PA-inhibited enhancers correlated with downregulation of phagocytosis and efferocytosis genes. These genes were similarly regulated in macrophages from mouse models of diabetes and accelerated atherosclerosis, human atherosclerosis, and by infectious agents. PA-regulated enhancers/SEs harbored SNPs associated with diabetes, obesity, and body mass index indicating disease relevance. We verified increased chromatin interactions between PA-regulated enhancers/SEs and inflammatory gene promoters, and reduced interactions at efferocytosis genes. PA-induced gene expression was reduced by inhibitors of BRD4, and NF-kB. PA treatment inhibited phagocytosis and efferocytosis in human macrophages. Together, our results show that PA-induced enhancer dynamics at key monocyte/macrophage enhancers/SEs regulate inflammatory and immune genes and responses. Targeting these PA-regulated epigenetic changes could provide novel therapeutic opportunities for cardiometabolic disorders.

overall_design: Chromatin Immunoprecipitation with H3K27ac  (Abcam, Cat # ab4729) followed by sequencing (ChIP-seq) in Human CD14+ monocytes treated with PA (200 µM) or vehicle BSA for 24 hours.

first: Vinay

middle: S

last: Tanwar

first: Marpadga

middle: A

last: Reddy


title: Positioning of sperm tail longitudinal columns depend on Nsun7, an RNA binding protein destabilizing elongated spermatid transcripts (BS-RNA-Seq)

city: Vienna

first: Sergey

last: Isaev

address: Spitalgasse, 4

department: Department of Neuroimmunology

country: Austria

institute: Medical University of Vienna

type: Methylation profiling by high throughput sequencing

summary: Spermatozoid’s flagella assemble in transcriptionally silent spermatids and thus depends on post-transcriptional regulation of gene expression. Mutations in Nsun7 gene are known to cause male infertility in human and mice. We identified m5C-specific Nsun7 RNA methyltransferase as a protein present in elongated spermatids and interacting with RNAs specific for this type of spermatozoid’s precursor cells. Inactivation of Nsun7 gene in mice leads to upregulation of its RNA interactors, thus indicating that Nsun7 downregulates a set of RNAs in the elongated spermatids. A physiologic consequence of Nsun7 gene knockout is male infertility, which is mechanistically explained by the observed mispositioning of longitudinal columns relative to the axonemal microtubular doublets leading to motility defect.

overall_design: To study the RNA targets of the m5C RNA methyltransferase Nsun7, bisulfite sequencing analysis was performed on elongated spermatids from wild-type and Nsun7 knockout mice. Each biological replicate included testis material of 3 mice. In order to obtain a fraction of elongated spermatids, cell sorting was performed.

first: Guseva

last: Ekaterina

first: Isaev

last: Sergey

first: Sergiev

last: Petr


title: MYC plus class IIa HDAC inhibiton potentiates mitochondrial dysfunction in non-small cell lung cancer [III]

city: Baltimore

first: Jina

last: Park

state: Maryland

address: 1650 Orleans St

department: Oncology

institute: Johns Hopkins University

summary: To evaluate potential therapeutic benefits from dual targeting of MYC and class IIa HDACs, we assessed the treatment efficacy of MYCi plus class IIa HDACi across 18 NSCLC cell lines, 10 of which demonstrated substantial reduction of cell viability upon combination treatment. RNA sequencing on treated cell lines identified large-scale transcriptional shifts facilitated by combination treatment, including suppression of MYC, cell cycle, and mitochondrial pathways. ATAC-sequencing showed chromatin structure changes in genomic regions involved in cell cycle progression and inflammatory signaling. Furthermore, G1/S arrest and elevated mitochondrial ROS levels in combination drug-treated cells were confirmed using flow cytometry. Concordant with the role of MYC as a key cell cycle regulator, MYC protein levels were decreased in the combination treatment group as compared to the vehicle or mono-treatment groups. In summary, we define here a novel drug paradigm combining MYC and class IIa HDAC inhibitors, which potentiates anti-tumor efficacy in NSCLC via MYC depletion and oxidative stress.

overall_design: To identify drug treatment-induced transcriptional changes in vivo, LLC1 tumor-bearing mice were treated with either DMSO, MYCi975 (50mg/kg), TMP195 (50mg/kg) or combination for 16 days and RNA-seq was performed with enriched tumor cells.

first: Michael

middle: J

last: Topper


title: MYC plus class IIa HDAC inhibiton potentiates mitochondrial dysfunction in non-small cell lung cancer [II]

overall_design: To identify drug treatment-induced transcriptional changes in vivo, LLC1 tumor-bearing mice were treated with either DMSO, MYCi975 (50mg/kg), TMP195 (50mg/kg) or combination for 16 days and RNA-seq was performed with single cell tumor suspension (bulk tumor).


title: MYC plus class IIa HDAC inhibiton potentiates mitochondrial dysfunction in non-small cell lung cancer [ATAC-seq]

overall_design: To identify drug treatment-induced chromatin structure changes, two NSCLC cell lines (H460 and H838) were treated with either DMSO, MYCi975, TMP195 or combination for 24 hours and subjected to ATAC-seq library preparation.


title: Positioning of sperm tail longitudinal columns depend on Nsun7, an RNA binding protein destabilizing elongated spermatid transcripts (CLIP-Seq)

overall_design: In order to identify the binding site of m5C RNA methyltransferase Nsun7, we performed UV crosslinking of Nsun7 with its RNA targets, followed by further co-immunoprecipitation of the obtained covalent complexes. To obtain a sufficient material, tissue from 30 wild-type and 30 HA-tagged Nsun7 mice was used.


title: Single nucleus RNA sequencing of hippocampal tissue from wildtype and SHIP1 conditional knockout mice at postnatal day 15.

status: Public on Apr 28 2025

city: Groningen

first: Bart

last: Eggen

state: Groningen

address: Antonius Deusinglaan 1

department: Department of Biomedical Sciences

country: Netherlands

institute: UMCG Groningen

summary: This study corresponds to the single nucleus RNA sequencing (snRNAseq) analysis done as part of the manuscript "Microglial SHIP1 controls synaptic pruning in the developing hippocampus via the complement system" by Matera et al. INPP5D, which encodes the lipid phosphatase SHIP1, is one of the most common genes associated with the risk of Alzheimer’s disease and is enriched in microglia in the central nervous system. SHIP1 has been found to be highly expressed in plaque-associated microglia. However, how it regulates microglial function and influences brain physiology has been poorly investigated. In this study, we generated a snRNAseq dataset from the hippocampi of wildtype (WT) and conditional SHIP1 knockout (KO) mice at postnatal day 15. By predicting cell-cell communication, we found decreased neuronal interactions in KO mice, in line with results of synaptic loss in the rest of the manuscript.

overall_design: Wildtype (WT) and conditional SHIP1 knockout (KO) mice were used in this study. KO mice were Inpp5d-floxed under a Cx3cr1 Cre-promoter. Recombination was induced by tamoxifen injection at postnatal days 3 and 4. Fresh frozen hippocampal tissues from one hemisphere were dissected from WT (n=4) and KO (n=4) mice at postnatal day 15. We used 2 male and 2 female pups per group. After performing single nucleus isolation, we enriched for lesser abundant cell types using a FANS strategy after staining for NeuN and Olig2. Nuclei were then loaded on 10x Genomics Chromium single cell chips. cDNA libraries were prepared and dual indexed using the 10x Genomics Chromium Single Cell 3' Reagents kit. Samples were then pooled in equimolar ratios and sequenced on a NextSeq2000 platform. Data were pre-processed with CellRanger7, and then analyzed using Seurat v4.3.

first: Janssen

middle: M

last: Kotah

first: Rosa

middle: C

last: Paolicelli

first: Alessandro

last: Matera


title: Progesterone signaling in oviductal epithelial cells modulates the immune response to support preimplantation embryonic development

city: Columbia

first: Wipawee

last: Winuthayanon

state: Missouri

address: 1030 Hitt Street

department: OB/GYN & Women's Health

institute: University of Missouri

summary: More than 60% of pregnancy loss occurs during the first trimester. As we lack a complete understanding of the environment within the oviduct (Fallopian tube in humans) and its role in regulating early pregnancy, it is difficult to provide diagnostic tools for oviductal origin of early pregnancy loss. To determine the role of progesterone (P4), we conditionally ablated the classical P4 receptor (Pgr) in oviductal epithelial cells using the Wnt7aCre/+;Pgrf/f (hereafter Pgrd/d) mouse model. Approximately 40% of embryos collected from Pgrd/d females at 3.5 days post-coitus were nonviable or developmentally delayed suggesting that epithelial PGR expression in the oviduct is required for normal embryonic development. Single-cell RNA-sequencing analysis showed an upregulation of genes involved in proinflammatory pathway (i.e., interleukin 22 (IL-22) and genes in IL-22 signaling) in secretory epithelial cells of Pgrd/d females as compared to controls. A loss of PGR in the epithelial cells resulted in an increase immune cell recruitment into the oviduct as compared to controls. Conversely, ovariectomized wild-type (WT) females treated with exogenous P4 showed a reduced expression of inflammatory response genes. Pharmacological inhibition of inflammation using non-steroidal anti-inflammatory drugs significantly reduced IL-22 levels in the oviducts and rescued embryo developmental rates in Pgrd/d females. Co-culture of WT zygotes with IL-22 significantly decreased the number of expanded blastocysts. Our findings indicate that P4 signaling through the classical PGR in oviductal epithelial cells is required for the immunoregulatory function and is indispensable for normal preimplantation embryonic development. These data may contribute to an understanding of the mechanisms underlying pre-implantation pregnancy failure, which can inform the development of targeted diagnostic tools and therapeutic interventions to improve reproductive outcomes when embryonic aneuploidy is not present.

overall_design: All animals were maintained at Washington State University and University of Missouri and were handled according to Animal Care and Use Committee guidelines using approved protocols #6147, 6151, 38927 and 38961. For scRNA-seq experiments, adult female mice (C57B6/J, 8–12 weeks old) were purchased from the Jackson (Jax) laboratory.

first: Emily

last: McGlade

first: Kalli

middle: K

last: Stephens

first: Jiude

last: Mao

first: Feyza

middle: N

last: Arguc

first: San-Pin

last: Wu

first: Sarayut

first: Haval

last: Shirwan

first: Esma

last: Yolcu

first: Mark

middle: I

last: Hunter

first: James

last: Pru

first: John

middle: P

last: Lydon

first: Francesco

last: DeMayo


title: ZIKV infection series in paediaitric brain tumour and NPC cell lines

city: Southampton

first: Rob

last: Ewing

state: Hampshire

address: University Rd, Southampton

department: School of Biological Sciences

country: United Kingdom

institute: University of Southampton

summary: Zika virus (ZIKV) is a mosquito-borne flavivirus consisting of historical African and epidemic-associated Asian lineages. The latter can access the central nervous system and causes microcephaly in the developing foetus through infection and depletion of neural stem and progenitor cells. The molecular mechanisms involved in this depletion response are grossly unknown for cells arising from CZS-affected children. Since 2017, the concept of employing ZIKV as oncolytic virotherapy against brain tumours has gained momentum. Oncolytic virotherapy exploits viruses that preferentially infect and destroy cancer cells via two distinct routes of therapeutic action. Following infection, intense viral replication induces oncolysis, releasing virions into the tumour microenvironment to infect neighbouring tumour cells. ZIKV induces an oncolytic event in infected pediatric brain tumour cells, but the molecular mechanisms involved in this response are grossly unknown. Here, we sought to investigate the molecular mechanisms underlying both the oncolytic and neuropathogenic responses of ZIKV infection in brain tumour and NPCs, respectively.

overall_design: The five cell lines were cultured under monolayer conditions and infected for 12-24 hours with Zika virus (Strain KU365779.1)

first: Matthew

last: Sherwood

first: Thiago

last: Mitsugi

first: Brandon

last: Coke

first: Yilu

last: Zhou


title: Human Multi-Regional Gastric Assembloids Favour Epithelial Cell Differentiation and Functional Parietal Maturation [paediatric patients]

city: Bologna

first: Camilla

last: Luni

address: Via Terracini 28

department: DICAM

country: Italy

institute: University of Bologna

summary: Patient-derived human organoids have the remarkable capacity to self-organise into more complex structures. However, to which extend the gastric organoids can recapitulate the human stomach functions remains unexplored. Here, we report how gastric region-specific organoids can self-assemble into complex multi-regional assembloids showing levels of differentiation unattainable with simpler models. The assembloid shows preserved fundus, body and antrum regionality and gastric-specific crosstalk pathways arise. Remarkably, the increased complexity and cross communication between the different gastric regions, allows for the arise of the elusive parietal cell type, responsible for the production of gastric acid, the essential function for food digestion. We showed functional response to drugs targeting ATPase H+/K+ and further maturation of the assembloid when transplanted in vivo. This advanced in vitro model, using patient-derived tissue-specific progenitors, successfully recapitulates the structural and functional characteristics of the human stomach, offering a promising strategy for studying developmental processes, tissue interactions, and disease mechanisms that were previously challenging to attain.

overall_design: To determine whether gastric stem cells have the ability to retain the identity of the region of origin, we isolated mucosal biopsies from the antrum, body and fundus regions of three paediatric patients. The organoids were derived according to previously established protocol (Giobbe et al., Nat. Comm., 2021) and cultured in gastric-specific medium. We designed a system to allow the self-aggregation of human gastric organoids in more complex multi-regional structures defined as assembloids. Briefly, full grown organoids were released from Matrigel, resuspended in collagen I gel, and cultured in floating conditions. We firstly defined the self-assembly conditions for each region (single-region assembloid, SRA) and then multi-regional, in the order fundus-body-antrum (multi-regional assembloid, MRA).

first: Brendan

last: Jones

first: Giada

last: Benedetti

first: Giuseppe

last: Calà

first: Ramin

last: Amiri

first: Lucinda

last: Tullie

first: Roberto

last: Lutman

first: Jahangir

last: Sufi

first: Lucy

last: Holland

first: Daniyal

last: Jafree

first: Monika

last: Balys

first: Glenn

last: Anderson

first: Ian

last: Simcock

first: Owen

last: Arthurs

first: Simon

last: Eaton

first: Nicola

last: Elvassore

first: Vivian

last: Li

first: Christopher J.

last: Tape

first: Kelsey DJ

first: Giovanni

last: Giobbe

first: Paolo

last: De Coppi


title: Positioning of sperm tail longitudinal columns depend on Nsun7, an RNA binding protein destabilizing elongated spermatid transcripts (RNA-Seq)

first: Sergey

last: Isaev

overall_design: The testis tissue of wild-type (n = 3) and Nsun7 knockout (n = 3) mice at the age of 3 months was collected for the whole transcriptome assay.

first: Guseva

last: Ekaterina

first: Isaev

last: Sergey

first: Sergiev

last: Petr


title: An unbiased genomewide screen uncovers 7 genes that drive hematopoietic stem cell fate from mouse embryonic stem cells

city: Barcelona

first: Anna

last: Bigas

state: Barcelona

address: Dr. Aiguader 88

department: Cancer Research

country: Spain

institute: Institut Hospital del Mar d'Investigacions Mèdiques

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

overall_design: Refer to individual Series

first: Luis

last: Galán-Palma

first: María

last: Maqueda

first: Eric

last: Canton


title: MYC plus class IIa HDAC inhibiton potentiates mitochondrial dysfunction in non-small cell lung cancer [I]

first: Jina

last: Park

overall_design: To identify drug treatment-induced transcriptional changes, two NSCLC responsive cell lines (H460, H1703) and two non-responsive cell lines (HCC827, H441) were treated with either DMSO, MYCi975, TMP195 or combination for 48 hours and RNA-seq was performed.

first: Michael

last: Topper


title: Clinical Data Investigation Identifies MARK3 as an Oncogenic Driver in Castration-Resistant Prostate Cancer

city: Sangareddy

first: Rajnikant

last: Raut

state: Telangana

address: IITH Road, Near NH65, Kandi

department: Biotechnology

country: India

institute: Indian Institute of Technology Hyderabad

summary: Castration-resistant prostate cancer (CRPC) represents an aggressive and fatal form of prostate cancer that emerges following resistance to androgen deprivation therapy. Despite the availability of various drugs that can enhance the quality and prolong the survival of CRPC patients, resistance to these therapies is frequently observed, making the disease increasingly difficult to treat. Alterations in the expression of kinases and phosphatases are key drivers of CRPC and present potential targets for more effective treatments. In this study, we have performed a comprehensive transcriptomic analysis of ~359 normal and CRPC patient samples from the cancer genome atlas to identify the differentially expressed kinases and phosphatases. We shortlisted the candidate genes based on their differential expression profiles, associations with patient survival, Gleason scores, and their impact on the fitness of prostate cancer cell lines. Our in-silico analysis identified microtubule affinity-regulating kinase 3 (MARK3) as a novel CRPC driver that is upregulated in CRPC patients, correlates with poor survival outcomes, and affects the fitness of CRPC cells. Pharmacological inhibition of MARK3 using PCC0208017, a MARK3 inhibitor, leads to reduced cell viability and migratory potential, as well as cell cycle arrest at G1 phase. Additionally, RNA sequencing analysis of 22Rv1 cells treated with the MARK3 inhibitor revealed that MARK3 influences genes involved in androgen response, epithelial-mesenchymal transition, mTOR and Myc signalling, underscoring its pivotal role in CRPC progression. Taken together, our results establish MARK3 as a novel and promising therapeutic target in CRPC.

overall_design: This experiment was performed using 22Rv1 cell line. Total 8 samples were subjected to the RNA sequencing with the Illumina NovaSeq 6000. Among them, 4 samples were treated with DMSO and 4 were treated with MARK3 inhibitor PCC0208017. Raw fastq files were processed and aligned to the Hg38 genome assembly and quantified with featureCounts. Count matrix was then utilized to perform differential gene expression analysis with DESeq2 package in R Studio.

first: Ashish

last: Misra

first: Manish

middle: V

last: Bais


title: MYC plus class IIa HDAC inhibiton potentiates mitochondrial dysfunction in non-small cell lung cancer [RNA-seq]

overall_design: To identify drug treatment-induced transcriptional changes, four NSCLC cell lines (H460, H838, H1703, H1755) were treated with either DMSO, MYCi975, TMP195 or combination for 24 hours and RNA-seq was performed.


title: A molecular brain atlas reveals cellular shifts during the repair phase of stroke

city: Los Angeles

first: Ruslan

last: Rust

state: CA

address: 1501 San Pablo Street

institute: University of Southern California

summary: Ischemic stroke triggers a cascade of pathological events that affect multiple cell types and often lead to incomplete functional recovery. Despite advances in single-cell technologies, the molecular and cellular responses that contribute to long-term post-stroke impairment remain poorly understood. To gain better insight into the underlying mechanisms, we generated a single-cell transcriptomic atlas from distinct brain regions using a mouse model of permanent focal ischemia at one month post-injury. Our findings reveal cell- and region-specific changes within the stroke-injured and peri-infarct brain tissue. For instance, GABAergic and glutamatergic neurons exhibited upregulated genes in signaling pathways involved in axon guidance and synaptic plasticity, and downregulated pathways associated with aerobic metabolism. Using cell-cell communication analysis, we identified increased strength in predicted interactions within stroke tissue among both neural and non-neural cells via signaling pathways such as those involving collagen, protein tyrosine phosphatase receptor, neuronal growth regulator, laminin, and several cell adhesion molecules. Furthermore, we found a strong correlation between mouse transcriptome responses after stroke and those observed in human nonfatal brain stroke lesions. Common molecular features were linked to inflammatory responses, extracellular matrix organization, and angiogenesis. Our findings provide a detailed resource for advancing our molecular understanding of stroke pathology and for discovering therapeutic targets in the repair phase of stroke recovery.

overall_design: The study was designed to generate a single-cell atlas of stroke-injured mouse tissue one month following permanent focal cerebral stroke. we used nine stroked adult male and female mice (3-5 months old) with a C57BL/6J background. We validated a successful stroke induction using Laser Doppler imaging and collected tissue from stroke, peri-infarct and intact cortex. We dissociated the tissue and isolated nuclei for subsequent snRNAseq analysis


title: Nutrient Starvation-induced Hda1C Rewiring: coordinated regulation of transcription and translation

city: Seoul

first: Tae-Young

last: Roh

state: --- Select One ---

address: 52 Ewhayeodae-gil, Seodaemun-gu

department: Life Sciences

institute: Ewha Womans University

summary: In yeast, Hda1 histone deacetylase complex (Hda1C) plays an important role in transcriptional regulation by modulating histone acetylation. We here explored the dynamics of Hda1C binding in nutrient-rich and -starved conditions. Chromatin immunoprecipitation sequencing revealed that starvation alters RNA Pol II and Hda1C binding to coding genes in a highly correlated manner. Interestingly, we discovered RNA Pol II transcription-independent recruitment of Hda1C to intergenic regions, particularly the upstream regulatory sequences (URS) of ribosomal protein (RP) genes, which are enriched with Rap1 binding sites. Under nutrient starvation, Rap1 contributes to the recruitment of Hda1C to these URS regions, where Hda1C deacetylates histones, thereby fine-tuning basal gene expression and delaying RP gene reactivation. Furthermore, Hda1C is also required for RNA Pol I transcription of rRNAs and RNA Pol III transcription of tRNA genes, especially in nutrient-limited conditions. Significantly, Hda1C mutants are sensitive to translation inhibitors and display altered ribosome profiles. Thus, Hda1C may coordinate transcriptional regulation within the nucleus with translation control in the cytoplasm and could be a key regulator of gene expression responses to nutrient stress.

overall_design: The chromatin immunoprecipitation sequencing (ChIP-seq) for RNA polymerases (Pol II and Pol III), Hda1, and histone acetylation (H3ac, H4ac) upon nutrient depletion in yeast. Changes in histone acetylation upon Hda1 deletion are analyzed.

first: Min Kyung

last: Lee

first: Byunghee

last: Kang

first: TaeSoo

last: Kim


title: The Long Non-coding RNA Lnc-RAINY Regulates Genes Involved in Radiation Susceptibility Through DNA:DNA:RNA triplex-forming Interactions and has Tumor Therapeutic Potential in Non-small Cell Lung Cancers [ATAC-seq]

city: Morgantown

first: Emily

last: Westemeier-Rice

state: WV

address: 1 Medical Center Drive

institute: West Virginia University

summary: Lung Cancer is the leading cause of cancer related deaths worldwide. Unfortunately, radiation resistance is a major problem facing lung cancer patients. Recently, we identified a group of long non-coding RNAs (lncRNAs) known as linc-SPRY3 RNAs, expressed from the Y-chromosome, that play a role in radiation sensitivity by decreasing tumor burden in vitro and in vivo after radiation. In this study, we found that the linc-SPRY3 RNAs are one large lncRNA that we named Radiation Induced Y-chromosome linked long non-coding RNA (lnc-RAINY). Through ATAC-Seq and Immunoprecipitation experiments, we show that lnc-RAINY interacts with DNA in a triple helix to change chromatin opening and gene expression. We also identified that lnc-RAINY regulates CDC6 and CDC25A expression affecting the induction of senescence, inhibition in cell migration, and cell cycle regulation. Furthermore, the administration of nanoparticle encapsulated lnc-RAINY into a lung cancer patient-derived xenograft model dramatically reduces tumor progression demonstrating its therapeutic potential.

overall_design: To understand the impact of lnc-RAINY on gene expression, we used 3 siRNAs against lnc-RAINY to identify differential gene expression after radiation, with or without the presence of lnc-RAINY.

first: Westemeier-Rice

last: Emily

first: Martinez

last: Ivan

first: Yi

last: Weijun

first: Hu

last: Gangqing


title: Lysine-specific demethylase 1 controls key OSCC preneoplasia inducer STAT3 through CDK7 phosphorylation during oncogenic progression and immunosuppression [Feline]

city: Boston

middle: V.

state: MA

address: W216, 700 Albany street

department: Translational Dental Medicine

institute: Boston University

summary: Oral Squamous Cell Carcinoma (OSCC) is a type of cancer that arises from preneoplastic precursors, which undergo genetic and epigenetic modifications. Lysine-specific demethylase 1 (LSD1) is a key OSCC promoter. LSD1 knockout or pharmacological inhibition in mouse OSCC reversed OSCC preneoplasia in our study. LSD1 inhibition attenuated the cell cycle, immunosuppression and promoted the infiltration of CD45+, TCRß+, CD4+, and CD8+ cells. Next, we tested the clinical candidate LSD1 inhibitor (Seclidemstat) in feline clinically spontaneous natural OSCC and found that it was safe and attenuated the STAT3 network. Mechanistic studies have shown that LSD1 reprograms preneoplasia to promote STAT3 signaling, cell cycle mediator CDK7, and immunosuppressive CTLA4 to OSCC progression. Interestingly, we found that the LSD1 inhibitor (SP2509) attenuated cyclin-dependent kinase 7 (CDK7) phosphorylation at T170 and eukaryotic initiation factor 4 B (Eif4B) phosphorylation at S422. We identified a new mechanism in OSCC preneoplasia that LSD1 promotes STAT3 mediated by CDK7 phosphorylation, leading to overall changes in the tumor microenvironment, leading to immunosuppression and progression to OSCC, which could have applications for understanding the mechanism as well as design strategies to control the progression of preneoplasia to OSCC.

overall_design: Total 1 feline patient with OSCC was recruited for this study. Tongue biopsies were collected before and after SP2577 treatment and subjected for RNA sequencing (n=3/condition).

first: Amit

last: Chakraborty

first: Sahay

last: Bikash

first: Takada

last: Marilia


title: Lysine-specific demethylase 1 controls key OSCC preneoplasia inducer STAT3 through CDK7 phosphorylation during oncogenic progression and immunosuppression [Mice]

overall_design: The 4NQO exposed mice were treated with either vehicle or SP2509 (n=3/condition) and subjected to RNA sequencing.

first: Chumki

last: Choudhury

first: Kisa

last: Iqbal


title: The Long Non-coding RNA Lnc-RAINY Regulates Genes Involved in Radiation Susceptibility Through DNA:DNA:RNA triplex-forming Interactions and has Tumor Therapeutic Potential in Non-small Cell Lung Cancers [RNA-seq]


title: An unbiased genomewide screen uncovers 7 genes that drive hematopoietic stem cell fate from mouse embryonic stem cells [scRNA-seq embryoid bodies]

first: Anna

last: Bigas

summary: Hematopoietic Stem Cells (HSCs) possess the ability to long-term reconstitute all the blood lineages and generate all blood cell types. As such, the in vitro generation of HSCs remains a central goal in regenerative medicine. Despite many efforts and recent advancements in the field, there is still no robust, reproducible and efficient protocol for generating bona-fide HSCs in vitro. This suggests that certain regulatory elements have yet to be uncovered. Here, we present a novel and unbiased approach to identifying endogenous components to specify HSCs from pluripotent stem cells. We performed a genome-wide CRISPR activator screening during mesodermal differentiation from mouse embryonic stem cells (mESCs). Following in vitro differentiation, mesodermal KDR+ precursors were transplanted into primary and secondary immunodeficient NSG mice. This approach led to the identification of seven genes (Spata2, Aass, Dctd, Eif4enif1, Guca1a, Eya2, Net1) that, when activated during mesoderm specification, induce the generation of hematopoietic stem and progenitor cells (HSPCs). These cells are capable of serial engraftment and multilineage output (erythroid, myeloid and T and B lymphoid) in vivo. Single-cell RNA sequencing further revealed that activating these seven genes biases the embryoid bodies towards intraembryonic development, instead of extraembryonic, increasing the number of mesodermal progenitors that can generate HSCs. Our findings underscore the importance of differentiation during the first germ layer specification to generate definitive blood stem cells.

overall_design: Mouse Embryoid Bodies at 144h were analyzed using 10x scRNA-seq considering 3xscrambled and 3xSADEiGEN samples, the latter generated after  CRISPR activation of a 7-genes set.

first: Luis

last: Palma

first: María

last: Maqueda

first: Eric

last: Canton


title: Mas1 Receptor Activation is Necessary and Sufficient to Transduce ACE2 Effect in PAH, but Ang(1-7) is Not the Ligand

city: Nashville

first: James

last: West

state: TN

address: 4200 E 9TH Ave, Box B133

department: Medicine

institute: Vanderbilt University

summary: Sugen-hypoxic Mice treated with ACE2, a MAS agonist AR234960, or a peptidase inhibitor Alt00

overall_design: Mice are put in 10% O2 for three weeks, and receive injections of the vegf inhibitor Sugen weekly, to induce pulmonary hypertension. They are simultaneously treated with either vehicle, ACE2, AR234960, or Alt00+Lisinopril


title: Downregulation of Scd1 promotes cardiac reprogramming by enhancing mitochondrial metabolism through PGC1α

city: wuhan

first: yu

last: yi

address: donghu street

institute: wuhan university

summary: Direct cardiac reprogramming of fibroblasts into induced cardiac-like myocytes (iCMs) has emerged as a promising therapeutic strategy to remuscularize injured myocardium. However, the intrinsic metabolic demands that drive the iCM program and maturity remain unclear. Here, we induce efficient iCM production and maturation through metabolic pathway modulations. We show that the knockdown of Stearoyl-CoA desaturase 1 (Scd1) facilitates cardiac reprogramming and functional maturation of iCM. Through Bulk-RNAseq analysis, we found that compared with control group, fibrosis and inflammatory response pathway were significantly down-regulated, while cardiac related features and fatty acid metabolism related genes were up-regulated in shscd1 group. We further observed the expression level of PGC1α which serves as a master regulator of mitochondrial biogenesis and function was significantly increased in shScd1 group from RNA-seq. Mechanistically, we proved that Scd1 knockdown activates PGC1α/PPARβ transcription signal, which further promotes cardiac reprogramming, and induced cardiomyocytes demonstrated more frequent beating, calcium oscillation, and higher energy metabolism as evidenced by increased mitochondrial respiration and gene expression associated with FAO, which provides a new strategy and theoretical basis for cardiac regeneration and repair

overall_design: To determine the molecular basis by which knockdown Scd1 enhances iCM reprogramming, we conducted RNA-seq using shScd1-iCMs and shNT-iCMs collected at reprogramming day 7.

first: Li

last: Wang

first: Zhenhua

last: Jia

first: Lilin

last: Xiang


title: Effect of monosodium urate (MSU) on gene expression of mouse bone marrow-derived dendritic cells (BMDCs) and splenic B cells

city: Seongnam

first: Eui Ho

last: Kim

state: Gyeongggi-do

address: 16 Daewangpangyo-ro 712 beon-gil, Bundang-gu

institute: Institut Pasteur Korea

summary: DCs and B cells are main target cells activated by uric acid (MSU, one of DAMP molecules). Ultimately, we would like to figure out signaling pathways significantly affected, and narrow down potential receptor candidates for uric acid (MSU).

overall_design: Comparison of each immune cell depending on stimulation (mainly [control vs MSU] or [LPS vs LPS+MSU]. If necessary, [control vs LPS+MSU])

first: Sun

last: Lee

first: Eui

middle: H


title: An unbiased genomewide screen uncovers 7 genes that drive hematopoietic stem cell fate from mouse embryonic stem cells [CRISPR screen]

overall_design: KDR+ cells from Ebs 144h were isolated and gDNA was extracted for gRNA library sequencing. In the case of gRNA library transplanted-mice, CD45+ BFP+ cells (gRNA library contains a BFP) were isolated and gDNA was extracted


title: Skeletal Muscle NAD Depletion Does Not Accelerate Muscle Aging

city: Copenhagen

first: Lars

middle: Roed

last: Ingerslev

address: Blegdamsvej 3B

department: NNF Center for Basic Metabolic Research

country: Denmark

institute: Copenhagen University

summary: NAD is a ubiquitous electron carrier essential for energy metabolism and the post-translational modification of numerous regulatory proteins. Perturbation of NAD metabolism is considered detrimental to health, with NAD depletion commonly thought to promote aging. However, to what extent normal NAD concentration can be decreased without deleterious repercussions is unclear. Here, we describe a mouse model where NAMPT-mediated NAD+ biosynthesis is disrupted in adult skeletal muscle. The resulting 85% decrease in muscle NAD+ abundance did not cause tissue degeneration or dysfunction, as reflected in its unchanged morphology, contractility, and exercise tolerance. The lack of defects was corroborated by intact mitochondrial respiratory capacity and unaffected muscle transcriptomic and proteomic profiles. Furthermore, lifelong NAD depletion did not accelerate muscle aging or impair whole-body metabolism. Collectively, these findings demonstrate that NAD depletion does not drive the age-related decline in skeletal muscle function.

overall_design: polyA RNA sequencing of Mus musculus skeletal muscle in wild-type (WT) and skeletal-muscle specific NAMPT knock-out (KO) animals

first: Sabina

last: Chubanava

first: Iuliia

last: Karavaeva

first: Amy

last: Ehrlich

first: Roger

last: Justicia

first: Astrid

middle: L

last: Basse

first: Ivan

last: Kulik

first: Danial

last: Ahwazi

first: Emilie

last: Dalbram

first: Samuel

last: Heaselgrave

first: Kajetan

last: Trost

first: Ben

last: Stocks

first: Ondřej

last: Hodek

first: Jesper

last: Havelund

first: Farina

last: Schlabs

first: Steen

last: Larsen

first: Carlos

last: Henriquez-Olguín

first: Daniela

last: Giustarini

first: Ranieri

last: Rossi

first: Zachary

last: Gerhart-Hines

first: Thomas

last: Moritz

first: Kei

last: Sakamoto

middle: E

last: Jensen

first: Nils

last: Færgeman

first: Gareth

last: Lavery

first: Atul

last: Deshmukh

first: Jonas

middle: T

last: Treebak


title: The effect of SETD1A knockout on gene expression in  ATM- or BRCA-deficient cells.

city: Birmingham

first: Martin

last: Higgs

address: Vincent Drive

department: Institute of Cancer and Genomic Sciences

institute: University of Birmingham

summary: Background: Cells deficient in DNA repair factors breast cancer susceptibility 1/2 (BRCA1/2) or ataxia-telangiectasia mutated (ATM) are sensitive to poly-ADP ribose polymerase (PARP) inhibitors. Building on our previous findings, we asked how the lysine methyltransferase SETD1A contributed to PARP inhibitor-mediated cell death and determined the mechanisms responsible.  Methods: We used cervical, breast, lung and ovarian cancer cells bearing mutations in BRCA1 or ATM and depleted SETD1A using siRNA or CRISPR/Cas9. We assessed the effects of the PARPi Olaparib on cell viability, homologous recombination, and DNA repair. We assessed underlying transcriptional perturbations using RNAseq. We also used data from The Cancer Genomics Atlas (TCGA) to investigate overall patient survival.  Results: Loss of SETD1A from both BRCA1-deficient and ATM-deficient cancer cells was associated with resistance to Olaparib, explained by an partial restoration of homologous recombination.  Mechanistically, SETD1A-dependent transcription of the crossover junction endonuclease EME1 correlated with sensitivity to Olaparib in these cells. Accordingly, when SETD1A or EME1 was lost, BRCA1 or ATM-mutated cells became resistant to Olaparib, and homologous recombination was partially restored.  Conclusions: Loss of SETD1A or EME1 may explain why patients develop resistance to PARP inhibitors in the clinic.

overall_design: To investigate the effect of SETD1A knockout on gene expression in backgrounds of HR-deficiency we treated Hela Kyoto inducible cas9 cells expressing SETD1A CRISPR gRNA with control, ATM or BRCA1 siRNA +/- doxycycline treatment. We performed gene expression analysis using data obtained from RNA-Seq experiments from three individual repeats.

first: Ellie

last: Sweatman


title: NGS PHRC OV

city: clermont ferrand

first: yoan

last: renaud

address: place henri dunand

country: France

institute: igred

summary: Sequencing mRNA of human cumulus cells obtained from patients with PCOS or CONTROL

first: Silvere

last: Baron

first: Yoan

last: Renaud


title: Attenuation of CD8 T Cell Exhaustion via miR-379-5p-Mediated Memory Promotion and Immune Checkpoint Inhibition

city: Taichung City

first: Shao-Chun

last: Wang

address: C1-6F., No.350, Jingmao 1st Rd.

department: Cancer Biology and Precision Therapeutics Center

country: Taiwan

institute: China Medical University

summary: MicroRNAs are epigenetic regulators of T cell maturation and exhaustion. However, the specific mechanisms by which miRNAs modulate T cell functions during tumor development are not well understood. Here, we demonstrate that miR-379-5p counteracts the exhaustion phenotype induced by chronic T cell stimulation, enhancing anti-tumor effector functions. MiR-379-5p is downregulated in exhausted T cells, negatively associated with exhausted tumor-infiltrating lymphocytes in advanced tumors, and positively correlated with favorable survival in breast cancer and other cancer types. MiR-379-5p directly targets the 3' untranslated region (3’-UTR) of TIM3 and TIGIT, suppressing their expression in CD8 T cells. Ectopic miR-379-5p expression directs differentiation to memory-like effector cells, enhancing cancer cell killing activity. Conversely, the nuclear receptor NR4A1 negatively regulates miR-379 in T cells, restoring immune checkpoint gene expression and mitigating the cancer-killing ability. OT-1 T cells expressing miR-379-5p have elevated cytotoxic killing against B16F10-OVA tumors in NOD-SCID mice. Importantly, autologous T cells isolated from breast cancer patients introduced with miR-379-5p significantly increase their killing activity against tumor organoids derived from the matched patients. Our findings identify miR-379-5p as an epigenetic tumor suppressor that promotes CD8+ T cell effector functions, offering promise for novel cancer immunotherapy strategies.

overall_design:
Whole blood was collected from healthy donors, and native CD8 T cells were purified from the isolated peripheral blood mononuclear cells (PBMCs). These CD8 T cells were then subjected to chronic stimulation with anti-CD3/anti-CD28 antibodies and IL-2 for 12 days to induce exhaustion.
To evaluate miRNA expression, small RNAs were isolated from both naïve and exhausted T cells and analyzed using next-generation sequencing (miRNA-seq). A comparative miRNA-seq analysis was performed on exhausted versus unstimulated T cells.
To evaluate gene expression, total RNA was isolated from both naïve and exhausted T cells and analyzed using next-generation sequencing (RNA-seq). A comparative RNA-seq analysis was performed on exhausted versus unstimulated T cells.
CD8 T cells were subjected to chronic stimulation with anti-CD3/anti-CD28 antibodies and IL-2 for 12 days to induce exhaustion. Then, the cells were transfected with miR-Scr or miR-379-5p, and cultured for 48 hours. To evaluate gene expression, total RNA was isolated and analyzed using next-generation sequencing (RNA-seq). A comparative RNA-seq analysis was performed on miR-379-5p versus miR-Scr treated T cells.

first: Wang

last: ShaoChun

first: Lin

last: YouZhe


title: Rapid and efficient generation of human iPSC-derived lymphatic endothelial cells for functional assays

city: Auckland

first: Justin

last: Rustenhoven

address: 133 Rangatira Road

department: Pharmacology

country: New Zealand

institute: The University of Auckland

summary: The lymphatic system plays a crucial role in fluid homeostasis, immune responses, and lipid metabolism.  Comprising specialized lymphatic endothelial cells (LECs), it enables interstitial fluid uptake and transit to lymph nodes. Investigations of lymphatic vasculature are typically performed in vivo due to the complex nature of this system. However, in vitro models, particularly those using induced pluripotent stem cells (iPSCs), offer a promising alternative by providing a renewable, animal-free, patient-specific, human cell source that can be easily manipulated. Here, we describe a robust and efficient 10-day protocol to differentiate human iPSCs into functional LECs, validated by immunostaining, flow cytometry, and RNA-sequencing. The iPSC-derived LECs (iLECs) exhibit comparable functionality to in vivo lymphatics, including their response to inflammatory and lymphangiogenic stimuli and ability to form 3D lumenized lymphatic networks. The protocol's reproducibility across multiple iPSC lines underscores its reliability and utility for studying lymphatic biology.

overall_design: To confirm a lymphatic endothelial cell fate of iPSC-derived lymphatic endothelial cells (LECs) we performed gene expression profiling from  three iPSC lines or matched iPSC-derived lymphatic endothelial cell differentiations.

first: B

last: Watkin

first: J


title: Impaired cardiac branched-chain amino acid metabolism in a novel model of diabetic cardiomyopathy

city: Kobe

first: Manabu

last: Nagao

state: Hyogo

address: 7-5-1 Kusunoki-cho, Chuo-ku

institute: Kobe University

summary: Metabolic alterations occur in the heart of patients with type 2 diabetes mellitus (T2DM). However, the relationships between branched-chain amino acid (BCAA) metabolism and the pathophysiology of diabetic cardiomyopathy (DbCM) remain unclear. Here, we sought to establish a novel model of DbCM and reveal its metabolic alterations, especially focusing on BCAA metabolism. Using adipocyte-specific 3′-phosphoinositide–dependent kinase 1 (PDK1)-deficient (A-PDK1KO) mice as a new model of DbCM, we demonstrated that cardiac BCAA catabolic enzymes were down-regulated in these mice, leading to the accumulation of BCAAs in the heart. Mechanistically, the accumulation of the BCAA leucine could cause cardiac hypertrophy via the activation of mammalian target of rapamycin complex 1 (mTORC1). A-PDK1KO mice mimic the cardiac phenotypes and metabolic alterations seen in human DbCM and exhibit impaired BCAA metabolism in the heart. This model may contribute to a better understanding of DbCM pathophysiology and the development of novel therapies for it.

overall_design: Heart tissues of adipocyte-specific 3′-phosphoinositide–dependent kinase 1 (PDK1)-deficient (A-PDK1KO) mice and PDK1 flox/flox mice (control).  The tissues were collected and snap frozen in liquid nitrogen, subsequently stored at -80℃ until further analysis.

first: Junko

last: Asakura

first: Tetsuya

last: Hosooka


title: Single-cell RNA-sequence of the resolution phase of lung inflammation in LPS-induced ARDS model

city: Bunkyo-ku

first: Kensuke

last: Miyake

state: Tokyo

address: 1-5-45, Yushima

department: Institute of Integrated Research

institute: Institute of Science Tokyo

summary: Acute respiratory distress syndrome (ARDS) is a serious lung failure with a high mortality rate and no effective pharmacological therapies. While the cellular and molecular mechanisms underlying the induction of ARDS have been extensively studied, those controlling its resolution remain poorly understood. Recent human cohort studies have implied the correlation between the severity of ARDS and the paucity of blood basophils. To examine the causative relationships between basophils and the pathogenesis of ARDS, we assessed the effect of basophil depletion in an LPS-induced mouse model of ARDS and explored the possible role for basophils in the induction and resolution of ARDS by using genetically engineered mice and single-cell RNA-sequencing (scRNA-seq) analyses.  Intratracheal LPS administration induced severe lung inflammation with massive neutrophil accumulation, followed by the recovery to homeostatic conditions. Basophil depletion impaired the recovery but not induction of lung inflammation, demonstrating the crucial role for basophils in the resolution phase of ARDS. scRNA-seq analysis identified that basophils accumulating in the lung were predominant sources of cytokine IL-4. Mice with basophil-specific IL-4 deficiency failed to resolve the lung inflammation. This was also the case in mice with neutrophil-specific IL-4 receptor deficiency. scRNA-seq analysis suggested that basophil-derived IL-4 acts on neutrophils to suppress their expression of anti-apoptotic genes and pro-inflammatory mediators. Thus, the present study demonstrated that basophils play a crucial role in the resolution phase of a mouse model of ARDS through the production of IL-4 which in turn acts on neutrophils to restrict lung inflammation.

overall_design: Cells isolated from inflamed lungs of WT and IL-4 KO mice on day 6 post-LPS inhalation were analyzed by scRNA-seq.

first: Seiko

last: Takasawa

first: Jun

last: Sugihara

first: Tomoya

last: Tateishi

first: Sho

last: Shibata

first: Sachiko

first: Hajime

last: Karasuyama

first: Yasunari

last: Miyazaki


title: The aryl sulfonamide indisulam inhibits acute megakaryoblastic leukemia (AMKL) by altering the alternative splicing of the transcription factor ZMYND8

city: suzhou

first: yang

last: yang

address: zhongnan road

institute: soochow university

summary: Acute megakaryoblastic leukemia (AMKL) is a subtype of acute myeloid leukemia with clinical heterogeneity. The poor prognosis of AMKL means that new therapies need to be developed to treat this disease. Studies have shown that the dysregulation of alternative splicing is associated with the occurrence and development of AMKL and the increase of chemotherapy resistance. indisulam, as a synthetic aryl sulfonamide, can effectively treat solid tumors by targeting the splicing factor RBM39. However, the effects of indisulam on AMKL and the underlying molecular mechanisms remain to be thoroughly investigated. We first screened AMKL for indisulam susceptibility in a public drug-susceptibility database. The results showed that AMKL was more sensitive to insulin than other tumor types. We then treated human AMKL cell lines (CMK, MEG01 and M07e) with indisulam to evaluate its efficacy and investigate its molecular targets. The results suggest that indisulam induces RBM39 degradation and inhibits tumor growth in a dose-dependent manner. RNA-seq and proteomic analyses revealed that indisulam selectively degraded RBM39 and resulted in mis-splicing of the transcription factor ZMYND8, which maintains AML survival and proliferation. We knocked down ZMYND8 and confirmed the critical role of ZMYND8 in maintaining AMKL survival and proliferation. In addition, we verified that the anti-tumor effect of indisulam on AMKL was DCAF15 dependent. By targeting DCAF15 using CRISPR/Cas9 system, we confirmed the DCAF15-dependent effect of indisulam in both in vivo and in vitro models. Taken together, our study suggests that indisulam has promising therapeutic potential for AMKL by specifically targeting the splicing factor RBM39 and is dependent on the expression of DCAF15. In addition, our study identified ZMYND8 as a novel downstream regulator of RBM39 and the RBM39-ZMYND8 axis is important for AMKL survival and proliferation.

overall_design: mRNA profiles of CMK cells treated with 5μmol indisulam or not for 24 h were generated by RNA sequencing, in double, using Illumina NovaSeq6000.

first: Ying

last: Yang

first: Jian

last: Pan


title: A Simplified Preparation Method for Single-Nucleus RNA-Sequencing Using Long-Term Frozen Brain Tumor Tissues

city: Heidelberg

first: Konstantin

last: Okonechnikov

state: Germany

address: Im Neuenheimer Feld 280

department: Pediatric Neurooncology

country: Germany

institute: German Cancer Research Center

summary: Due to an increasing need for advanced techniques to understand the heterogeneity of central nervous system tumors, we optimized a method to isolate intact nuclei from long-term frozen pediatric brain tumor tissues. The results demonstrated that the protocol is fast, simple and low-coust thus optinal for potential application for such cases.

overall_design:
A set of long-term frozen tissues from pediatric brain tumors were processed with a specific single nuclei protocol in order to evaluate the suitability of the protocol for studing tumor heterogeneity

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Submitter states that missing raw files are due to patient privacy concerns.
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first: Kati

last: Ernst


title: Effect of MITF depletion on gene expression in U2OS cells

city: Reykjavik

first: Daniel

last: Magnusson

address: Sturlugata 8

department: Biomedical

country: Iceland

institute: Háskóli Íslands

summary: Summary (abstract): The transcriptional factor MITF is a central regulator of melanocytes and plays an important role in melanoma. In melanocytes MITF regulates the transcription of large number of genes with approximately 500 genes as validated targets. Functionally, MITF target genes are quite diverse including genes involved in pigmentation, DNA damage response, mitotic regulation, and DNA replication. Considering the transcriptional regulation of genes involved in non-melanocyte specific processes and the fact that MITF is expressed across tissue types, raises the question of whether the role of MITF outside the melanocyte lineage might be underestimated. Our data shows that in the osteosarcoma cell line, significant number of genes involved in DNA replication, DNA repair and mitotic regulation are downregulated upon MITF depletion, suggesting that the protein might be vital to maintain genome stability in the cell line.

overall_design: All samples underwent RNAi treatmend, where control replicates were treated with a negative control (SiControl) and experimental replicates were were treated with an siRNA which targeted the MITF gene (SiMITF). Each sample except the ones marked UT (untreated) were also treated with 50ng/ml NCS (Neocarzinostatin)to induce Double stranded breaks.


title: Type II interleukin-4 receptor activation promotes tumor pro-gression in basal breast cancer cells via metabolic and epige-netic modulation

first: Demond

last: Williams

address: 715 CA Preston Research Building 2220 Pierce Avenue

department: Cancer Biology

summary: Interleukin-4 (IL4) is a Th2 cytokine that can signal through two different receptors, one of which – the type II receptor - is overexpressed by various cancer cells. Previously we have shown that type II IL4 receptor signaling increases proliferation and metastasis in mouse models of breast cancer, as well as increasing glucose and glutamine metabolism. Here we expand on those findings to determine mechanistically how IL4 signaling links glucose metabolism and histone acetylation to drive proliferation in the context of triple-negative breast cancer (TNBC). We used a combination of cellular, biochemical and genomics approaches to interrogate TNBC cell lines, which represent a cancer type where high expression of the type II IL4 receptor is linked with reduced survival. Our results indicate that type II IL4 receptor activation leads to increased glucose uptake, Akt and ACLY activation,and histone acetylation in TNBC cell lines. Inhibition of glucose uptake through deletion of Glut1 ablates IL4-induced proliferation. Additionally, pharmacological inhibition of histone acetyltransferase P300 attenuates IL4-mediated gene expression, and proliferationin vitro. Our work elucidates a role for type II IL4 receptor signaling in promoting TNBC progression, and highlights type II IL4 signaling as well as histone acetylation as possible targets for therapy.

overall_design: Investigate the effect of IL4 signaling through the type II Il4 receptor in triple-negative breast cancer

first: Barbara

last: Fingleton


title: Effect of HOXC6 deletion on gene expression in Ewing Sarcoma cell line RD-ES.

city: Wuxi City

first: Yonghua

last: Pang

state: Jiangsu Province

address: 101 Xingyuan North Road, North Street

institute: The 904th Hospital of the Joint Logistics Support Force of the People's Liberation Army of China

summary: The early diagnosis of Ewing sarcoma (ES) is crucial for improving patient prognosis. This study has identified new diagnostic biomarkers for ES and evaluated the in vitro effect of HOXC6 on the RD-ES cell line. Experimental results demonstrated that HOXC6 knockout inhibited ES cell proliferation and migration. To investigate the specific mechanisms by which HOXC6 knockout promotes RD-ES cell proliferation and migration, we conducted sequencing analysis on the HOXC6-knockout cells.

overall_design: To examine the precise influence of HOXC6 knockdown on proliferation and migration in the Ewing sarcoma RD-ES cell line, we established an shHOXC6 RD-ES cell line.

first: Pang

last: YongHua

first: Liang

last: JIaHui

first: Ren

last: zhiyao


title: Chronic BCR signaling generates and maintains age-associated B cells from anergic B cell

city: Suita

first: Daisuke

last: Motooka

state: Osaka

address: 3-1, Yamadaoka

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

summary: We report here that ABCs exhibited a unique phenotype in aging and lupus model mice, where BCR signaling is constitutively activated, and surface BCRs are internalized. Autoreactive anergic B cells exclusively differentiated into ABCs in a BCR-dependent manner in vivo. Additionally, anergic B cells showed enhanced ABC differentiation in vitro compared to naive B cells. This was suppressed by Nr4a1 expression, possibly due to the inhibition of several signaling pathways, including BCR.

overall_design: To investigate gene expression pattern of Nr4a1 overexpressed B cells in ABC inductive stimuli, we retoroviruslly transducted Nr4a1 in B cells stimulated with ABC inductive stimuli. Infected GFP+ B cells were purified by FACS sorting. CD43 naive B cells, mock transduced B cells were sampled by controls. These B cell subsets were subjected to generate cDNA libraries for RNA-seq analysis. We then performed comparative gene expression analysis to identify the characteristics of gene expression shaped in Nr4a1 overexpressed B cells.

first: Keisuke

last: Imabayashi

first: Yutaro

last: Yada

first: Airi

first: Motoki

last: Yoshimura

first: Takeshi

last: Iwasaki

first: Koichi

last: Akashi

first: Hiroaki

last: Niiro

first: Yoshihiro

last: Baba


title: LY6H is a specific marker of human pancreatic delta cells

city: Parkville

first: Jacqueline

last: Schiesser

address: 50 Flemington Road

country: Australia

institute: Murdoch Childrens Research Institute

summary:
The identification of cell surface markers specific to pancreatic islet cell subsets is important for both the study of islet biology and for investigating the pathophysiology of diseases in which these cell types are lost or damaged. Analysis of publicly available single-cell RNAseq data identified expression of LY6H specifically in the delta cells of the pancreas. This finding was confirmed using immunofluorescence analysis of histological sections of human pancreas, and flow cytometric analysis of human islet preparations.
We found that expression of LY6H was restricted to pancreatic delta cells in both control and diabetic donors. Furthermore, we demonstrate that antibodies against LY6H can be used to isolate live delta cell populations from preparations of human islets. This study identified LY6H as a novel cell surface marker of human pancreatic delta cells – a finding that will aid in the identification and characterisation of this important cell type.

overall_design: Isolated human pancreatic islets were stained with antibodies against LY6H and sorted by FACS using a BD Influx (BD Biosciences) or BD FACSAria Fusion (BD Biosciences). RNA extraction of FACS purified populations was performed using trizol extraction as directed by the manufacturer (ThermoFisher, 15596026).


title: Disrupted Minor Intron Splicing Activates Reductive Carboxylation-mediated Lipogenesis to Drive MASH Progression

city: Shanghai

first: Xuyun

last: Zhao

state: Shanghai

address: South Chongqing Road

department: Medical school

country: China

institute: Shanghai Jiaotong University

summary: Aberrant RNA splicing is tightly linked to diseases, including metabolic dysfunction-associated steatotic liver disease (MASLD). Here, we revealed that minor intron splicing, a unique and conserved RNA processing event, is largely disrupted upon the progression of metabolic dysfunction-associated steatohepatitis (MASH) in mice and humans. We demonstrated deficiency of minor intron splicing in the liver induces MASH transition upon obesity-induced insulin resistance and LXR activation. Mechanistically, inactivation of minor intron splicing leads to minor intron retention of Insig1 and Insig2, resulting in premature termination of translation, which drives proteolytic activation of SREBP1c. This mechanism is conserved in human patients with MASH. Notably, disrupted minor intron splicing activates glutamine reductive metabolism for de novo lipogenesis through the induction of Idh1, which causes the accumulation of ammonia in the liver, thereby initiating hepatic fibrosis upon LXR activation. Ammonia clearance or IDH1 inhibition blocks hepatic fibrogenesis and mitigates MASH progression. More importantly, the overexpression of Zrsr1 restored minor intron retention and ameliorated the development of MASH, indicating that dysfunctional minor intron splicing is an emerging pathogenic mechanism that drives MASH progression. Additionally, reductive carboxylation flux triggered by minor intron retention in hepatocytes serves as a crucial checkpoint and potential target for MASH therapy.

overall_design:
To investigate the gene expression in mouse liver regulated by Zrsr1/2, we established the Zrsr1/2 liver specific  knock out mice by injection  AAV expressing sgRNA targeting Zrsr1 and Zrsr2 (Zrsg) or AAV expressing GFP(control group) to Cas9 transgenic mouse through tail vein.
After AAV injection ,all mice were maintained  in 26℃ and fed with normal chow diet for 2 weeks, then were euthanasia and liver tissue were harvest and frozed immediately with liquid nitrogen.
we performed gene expression profile using data obtained from RNA-seq of liver from control and Zrsg mice.

first: Yinkun

last: Fu

first: Xin

last: Peng

first: Hongyong

last: Song


title: An unbiased genomewide screen uncovers 7 genes that drive hematopoietic stem cell fate from mouse embryonic stem cells [scRNA-seq bone marrow]

city: Barcelona

first: Anna

last: Bigas

address: Dr. Aiguader 88

institute: Institut Hospital del Mar d'Investigacions Mèdiques

summary: Hematopoietic Stem Cells (HSCs) possess the ability to long-term reconstitute all the blood lineages and generate all blood cell types. As such, the in vitro generation of HSCs remains a central goal in regenerative medicine. Despite many efforts and recent advancements in the field, there is still no robust, reproducible and efficient protocol for generating bona-fide HSCs in vitro. This suggests that certain regulatory elements have yet to be uncovered. Here, we present a novel and unbiased approach to identifying endogenous components to specify HSCs from pluripotent stem cells. We performed a genome-wide CRISPR activator screening during mesodermal differentiation from mouse embryonic stem cells (mESCs). Following in vitro differentiation, mesodermal KDR+ precursors were transplanted into primary and secondary immunodeficient NSG mice. This approach led to the identification of seven genes (Spata2, Aass, Dctd, Eif4enif1, Guca1a, Eya2, Net1) that, when activated during mesoderm specification, induce the generation of hematopoietic stem and progenitor cells (HSPCs). These cells are capable of serial engraftment and multilineage output (erythroid, myeloid and T and B lymphoid) in vivo. Single-cell RNA sequencing further revealed that activating these seven genes biases the embryoid bodies towards intraembryonic development, instead of extraembryonic, increasing the number of mesodermal progenitors that can generate HSCs. Our findings underscore the importance of differentiation during the first germ layer specification to generate definitive blood stem cells.

overall_design: Mouse Bone Marrow CD45 cells after 4.5month post-transplantation were isolated by Fluorescence-activated cell sorting (FACS) according to the presence or absence of BFP and analyzed using 10x scRNA-seq. BFP presence indicated their origin from mESCs after CRISPR activation of a 7-genes set (SADEiGEN). Both samples were generated from a pool of 3 mice.

first: Luis

middle: G

last: Palma

first: María

last: Maqueda

first: Eric

last: Canton


title: Matured hiPSC-derived cardiomyocytes possess dematuration plasticity

city: Dover

first: David

last: Zebrowski

state: DE

address: 1151 Walker Rd

country: USA

institute: GenKardia Inc.

summary: Human induced Pluripotent Stem Cell-derived cardiomyocytes (hiPSC-CMs) are increasingly used to identify potential factors capable of inducing endogenous cardiomyocyte proliferation to regenerate the injured heart. L-type calcium channel blockers have previously been identified as a class of factors capable of inducing matured hiPSC-CMs to proliferate. However, the mechanism by which L-type calcium channel blockers promote hiPSC-CM proliferation remains unclear. Here we provide evidence that matured hiPSC-CMs possess plasticity to undergo dematuration in response to certain pharmacological compounds. Consistent with primary cardiomyocyte maturation during perinatal development, we found that centrosome disassembly occurs in hiPSC-CMs during plate-based, temporal, maturation. A small molecule screen identified Nitrendipine, an L-type calcium channel blocker, and 1-NA-PP1, a Src kinase inhibitor, as factors capable of inducing centrosome reassembly in a subpopulation of hiPSC-CMs. Furthermore, centrosome-positive hiPSC-CMs were more likely to exhibit cell cycle activity than centrosome-negative hiPSC-CMs. In contrast, neither Nitrendipine or 1-NA-PP1 induced centrosome reassembly, or cell cycle activity, in neonatal rat ventricular myocytes (NRVMs). Differential bulk transcriptome analysis indicated that matured hiPSC-CMs, but not NRVMs, treated with Nitrendipine or 1-NA-PP1 undergo dematuration. ScRNA transcriptome analysis supported that matured hiPSC-CMs treated with either Nitrendipine or 1-NA-PP1 undergo dematuration. Collectively, our results indicate that matured hiPSC-CMs, but not primary NRVMs, possess plasticity to undergo dematuration in response to certain pharmacological compounds such as L-type calcium channel blockers and Src-kinase inhibitors. This study shows that once mature, hiPSC-CMs may not maintain their maturity under experimental conditions and thus may have implications for experimental systems where the state of hiPSC-CM maturation is relevant.

sample_organism:
Homo sapiens
Rattus norvegicus

platform_organism:
Homo sapiens
Rattus norvegicus

overall_design:
Bulk RNA prep and sequencing  P6-NRVMs and CDI-CMs were treated with Nitrendipine or 1-NA-PP1 for 2 days prior to RNA extraction. In addition, both P6-NRVMs and CDI-CMs during treatment with Nitrendipine or 1-NA-PP1 were cultured in the presence of araC. AraC was used to reduce overgrowth of contaminating cardiac fibroblasts (which generally amount to ~3% of cultured cells in primary cardiomyocyte isolation procedure). RNA was extracted from sub-confluent 10-cm plates using Qiagen RNeasy mini kit. RNA quality (RNA integrity number, RIN) and quantity was measured in a Bioanalyzer 2100 (Agilent) using the Agilent RNA 6000 Nano Kit (part number 5067-1511). The sequencing libraries were prepared using the NEBNext Ultra II Directional RNA Library Prep Kit for Illumina (E7760S, New England Biolabs), starting with 200 ng of total RNA. In brief, mRNA was isolated and fragmented using the NEBNext poly(A) mRNA magnetic isolation module (E7490S, New England Biolabs), with the first and second strands of cDNA synthesized and purified using AmPure XP SPRI beads (A63880, Beckman Coulter). Then adaptor ligation and size selection were performed according to the manufacturer’s protocol. Next, the adaptor ligated cDNA was PCR enriched to incorporate an Illumina compatible index sequence (NEBNext Oligos for Illumina, Dual Index Primers Set1 (E7600S, New England Biolabs). All libraries were purified using AmPure XP SPRI beads, and the size distribution of the libraries was analyzed at the Bioanalyzer 2100 using the Agilent High Sensitivity DNA Kit (part number 5067-4626). Subsequently, the quantification of the libraries was performed with the Qubit® 2.0 Fluorometer (ThermoFisher Scientific) and QubitTM dsDNA HS Assay Kits (Q32851, Invitrogen). Finally, all 19 libraries were pooled and diluted to 3.5 nM for sequencing on one lane of a Hiseq 3000 sequencer (Illumina), using a single read 50 bp and dual indexed sequencing strategy.

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Submitter states that missing raw files are due to file loss.
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first: Fang

last: Meng

first: Maxwell

last: Kwok

first: Yen

last: Hui

first: Ruofan

last: Wei

first: Alejandro

last: Hidalgo-Gonzalez

last: Walentinsson

first: Frederik

last: Bjerre

first: Qing-Dong

last: Wang

first: Ditte

last: Andersen

first: Ellen

last: Poon

first: Daniela

last: Später


title: Inhibition of USP28 sensitizes ovarian cancer to PARP inhibitor through mediating SOX9 deubiquitination

city: Jinan

first: Hanlin

last: Ma

address: 107 Wenhua Xi Road, Jinan, China.

institute: Qilu Hospital of Shandong University

summary: PARP inhibitor (PARPi) resistance presents a significant challenge in ovarian cancer treatment, necessitating the development of effective therapeutic strategies to overcome this resistance and improve patient outcomes. Our study demonstrated that elevated expression of SRY-box 9 (SOX9) contributes to olaparib resistance in ovarian cancer. Mechanistically, the deubiquitinating enzyme USP28 was identified as a novel interacting partner of SOX9. USP28 inhibited the ubiquitination and subsequent lysosomal degradation of SOX9, which is mediated by the E3 ubiquitin ligase FBXW7 during olaparib treatment. ChIP-Seq analysis revealed that SOX9 binds to the promoters of key DNA damage response (DDR) genes (SMARCA4, UIMC1, and SLX4), thereby regulating DDR processes in ovarian cancer. Additionally, USP28 promoted olaparib resistance by stabilizing SOX9 protein and enhancing DNA damage repair. Furthermore, the USP28 specific inhibitor AZ1 reduced SOX9 protein stability and increased the sensitivity of ovarian cancer cells to olaparib. In conclusion, targeted inhibition of USP28 promoted ubiquitination-mediated degradation of SOX9, thereby impairing DNA damage repair capabilities and sensitizing ovarian cancer cells to PARPi. These findings elucidate the underlying mechanisms of PARPi resistance in ovarian cancer and suggest the potential efficacy of combining USP28 inhibitors with PARPi to overcome this resistance.

overall_design: ChIP-seq was performed to identity SOX9 binding genes in UWB1.289 cells。

last: Han

first: Gonghua

last: Qi


title: The Hippo pathway and p27Kip1 cooperate to suppress sensory receptor regeneration in the organ of Corti and the retina.

city: Los Angeles

first: Ksenia

last: Gnedeva

address: 1501 San Pablo Street

institute: USC

summary: The mammalian auditory sensory organ, the organ of Corti, has no capacity for regenerating lost hair cells, leading to permanent hearing impairment. In this study, we demonstrate that auditory supporting cells resist Yap activation after Hippo inhibition and uncover the molecular mechanism maintaining quiescence in the adult cochlear supporting cells.

overall_design: Primary and cultured organs of Corti were dissociated into a single cell suspension and supporting cells were purified via fluorescence-activated cell sorting to prepare RNA-Seq libraries. For in vitro experiments, p1 organs of Corti were dissected from Lfng-eGFP mice and cultured in presence of DMSO or Lats Kinase inhibitor (LKI) for 5 days before profiling eGFP-positive supporting cells. For in vivo experiments, organs of Corti were dissected from P8  Lfng-CreERT2/Rosa26-LSL-tdT/Pou4f3-DTR mice, treated with tamoxifen at P2 to label supporting cells and treated with diphtheria toxin fat P4 to ablate hair cells. tdTomato-positive supporting cells with and without hair cell damage were profiled.

first: Gnedeva

last: Ksenia


title: Nicotinamide Mononucleotide Promotes Female Germline Stem Cell Proliferation by Activating the H4K16ac-Hmgb1-Fyn-PLD Signaling Pathway through Epigenetic Remodeling

first: Ji

last: Wu

address: No. 800, Dongchuan Road

institute: Shanghai Jiao Tong University

summary: Nicotinamide mononucleotide (NMN), an endogenous nucleotide essential for various physiological processes, has an unclear role and regulatory mechanisms in female germline stem cell (FGSC) development. In this study, we demonstrate that NMN significantly enhances FGSC viability and proliferation. Quantitative acetylation proteomics revealed that NMN markedly increases the acetylation of histone H4 at lysine 16 (H4K16ac). Subsequent chromatin immunoprecipitation sequencing (ChIP-seq) and RNA sequencing (RNA-seq) identified high mobility group box 1 (Hmgb1) as a downstream target of H4K16ac, a finding further validated by ChIP-qPCR. Knockdown of Hmgb1 reduced FGSC proliferation by disrupting cell cycle progression, inducing apoptosis, and decreasing chromatin accessibility. High-throughput chromosome conformation capture (Hi-C) analysis showed that Hmgb1 knockdown induced A/B compartment switching, increased the number of topologically associating domains (TADs), and decreased chromatin loop formation in FGSCs. Notably, the chromatin loop at the promoter region of Fyn proto-oncogene (Fyn) disappeared following Hmgb1 knockdown. ChIP-qPCR and dual-luciferase reporter assays further confirmed the interaction between Hmgb1 and the Fyn promoter. Importantly, Fyn overexpression reversed the inhibitory effects of Hmgb1 knockdown on FGSC proliferation. Proteomic analysis suggested this rescue was mediated through the phospholipase D (PLD) signaling pathway, as Fyn overexpression selectively enhanced the phosphorylation of PLD1 at threonine 147 without affecting serine 561. Furthermore, treatment with 5-fluoro-2-indolyldechlorohaloamide, a PLD inhibitor, nullified the pro-proliferative effects of Fyn overexpression. In summary, our findings reveal that NMN promotes FGSC proliferation by activating the H4K16ac-Hmgb1-Fyn-PLD signaling pathway through epigenetic remodeling. These results deepen our understanding of FGSC proliferation and highlight potential therapeutic avenues for advancing FGSC applications in reproductive medicine.

overall_design: To investigate the effect of nicotinamide mononucleotide (NMN) on the development of female germline stem cells (FGSCs), we treated FGSCs with NMN in vitro. Subsequently, we utilized RNA-seq data from both the control group and the NMN-treated group to conduct differential gene expression analysis.

first: Hong

last: Zhou

first: Yujie

last: Liu

first: Geng

last: Tian