title: Decoding RNA binding protein's binding preferences using a general generative model

city: Qingdao

first: yuhua

last: jiao

address: No. 72 Binhai Road, Jimo District

institute: shandong university

summary: Current computational approaches struggle to detect the binding preference of novel or data-scarce RNA binding proteins (RBPs). Here we present Protein Guided Ribonucleic Acid Generator (ProRiboGen), an integrated generative diffusion model that predicts RBPs binding preferences directly from their protein sequences. ProRiboGen leverages insights from protein language models and CLIP-seq interaction data, generating RNA sequences that align with the binding preferences of target RBPs. We demonstrated its capacity for recognizing RNA-binding domains and cross-species generalization ability. Using this approach, we experimentally identified binding motifs for five human and zebrafish RBPs in the absence of CLIP-seq data. Furthermore, its binding site prediction was validated in vivo, revealing a previously uncharacterized intrinsic Cas13d-RNA interaction across the transcriptome, suggesting a potential link to off-target and bystander effects. This study advances our understanding of RBP-RNA interaction and highlights the model’s potential in RNA post-transcriptional regulation.

overall_design: To systematically analyze transcriptome-wide Cas13d binding in vivo, POND-seq was utilized, a novel technique for identifying RNA targets bound to RBPs,  in which EPN24-Cas13d fusion proteins form secreted nanocages that encapsulate bound RNAs.

first: Zhaoyi

last: Chen

first: Yuhua

last: Jiao

first: Lufeng

last: Hu

first: Jiale

last: He

first: Kangyi

last: Li

first: Shaozheng

last: Yin

first: Yangming

last: Wang

first: Ming

last: Shao

first: Lei

last: Sun


title: IL-36γ Armored CAR T Cells Reprogram Neutrophils to Induce Endogenous Antitumor Immunity

status: Public on Mar 01 2026

city: BUFFALO

first: Eduardo

last: Cortes

state: NY

address: ELM AND CARLTON STREETS

department: Biostatistics and Bioinformatics

country: USA

institute: ROSWELL PARK CANCER INSTITUTE

type: Expression profiling by high throughput sequencing

summary: Chimeric antigen receptor (CAR) T cells are ineffective against solid tumors due to obstacles of antigen heterogeneity and the immunosuppressive tumor microenvironment (TME). Previous efforts focused on enhancing cytotoxicity and persistence of CAR T cells, while the feasibility of improving their therapeutic efficacy by leveraging the modulatory effects of CAR T cells on host anti-tumor immunity remains unclear. Here, we report that IL-36γ armored CAR T cells eradicated primary solid tumors and enabled rejection of rechallenged antigen-negative tumors. IL-36γ armored CAR T cells favorably modulated the TME and reprogrammed unique neutrophil subsets with tumoricidal ability and antigen-(cross)presenting functions, resulting in the induction of endogenous T cells recognizing tumor antigens beyond CAR-targeted antigens. Our study demonstrates that neutrophil engagement by CAR T cells is a critical step in the establishment of the cancer-immunity cycle and introduces a broadly applicable method to overcome key barriers to adoptive cell therapies for solid tumors.

overall_design: Human bone marrow neutrophils were isolated by positive selection using EasySep protocols according to the manufacturer’s instructions. After treatment under different conditions for 6 hours, the treated neutrophils were collected for RNA extraction.

first: Yihan

last: Zuo

first: Renier

last: Brentjens


title: HERC5-HSP60 ISGylation Axis Sustains Mitochondrial Homeostasis in Glioblastoma Stem Cells to Promote Tumor Progression

city: Nanjing

first: zhiting

address: Longmian Avenue No.101

country: China

institute: Nanjing Medical University

summary: Glioblastoma (GBM), the most aggressive primary central nervous system (CNS) malignancy, exhibits rapid progression, therapy resistance, and inevitable recurrence—hallmarks primarily driven by glioblastoma stem cells (GSCs). Within the tumor microenvironment, dynamic crosstalk between GSCs and cytokines creates a niche for GSC maintenance. While interferon-gamma (IFN-γ) plays a well-established role in antitumor immunity, we reveal its paradoxical function in promoting the proliferation and self-renewal of GSCs.  In this study, we show that IFN-γ induces the expression of HECT and RLD domain-containing E3 ubiquitin ligase 5 (HERC5) in GSCs, which functions as an ISG15 E3 ligase. HERC5 promotes mitochondrial stability by mediating the ISGylation of heat shock protein 60 (HSP60), a key mitochondrial chaperone. Genetic depletion of HERC5 leads to HSP60 degradation, mitochondrial dysfunction, and marked suppression of tumor growth, significantly prolonging survival in orthotopic GBM mouse models. These findings identify the HERC5–HSP60 axis as a critical mediator of GSC adaptation to cytokine-induced stress, highlighting HERC5 as a potential therapeutic target to disrupt GSC-driven tumorigenesis.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for MGG8 and 3028 GSCs and its knockdown derivatives (shHERC5).

first: Zhiting

first: Jiancheng

last: Gao

first: Daqi

first: Linping

last: Yan

first: Yuhan

last: Jiang

first: Danling

last: Gu

first: Zhe

last: Zhu

first: Markus

last: Siegelin

first: Junlei

last: Yang

first: Gaoyuan

last: Cui

first: Zhumei

last: Shi

first: Xiuxing

first: Kun

first: Qian

last: Zhang

first: Minhong

last: Pan


title: Intrinsically Disordered Regions Form Nucleoli and Cajal Bodies While Fostering RNA Modification

city: Gosselies

first: Denis

middle: LJ

last: Lafontaine

state: Hainaut

address: Av Profs Jeneer & Brachet, 12

country: Belgium

institute: Universite libre de Bruxelles

summary: One of the densest compartments in the cell is the dense fibrillar component (DFC) of the nucleolus, consisting mainly of nascent ribosomal RNA (rRNA), small nucleolar ribonucleoproteins (snoRNPs) and their chaperone Nopp140. How this biomolecular condensate is formed and what underlies its structure is poorly understood like that of most liquid-liquid phase separated condensates. Although we established that Nopp140 is important for the cohesiveness of the DFC and for rRNA modification, it is not known how this is achieved. Here we demonstrate that Nopp140 concentrates intrinsically disordered and nuclear localization signal (NLS)-rich protein regions (IDRs), including a newly identified RNA polymerase I C-terminal domain (CTD) of the RNA polymerase I associated factor PAF49. Altogether, this network forms the DFC, a liquid-liquid phase separated biomolecular condensate that fosters rRNA modification. This mechanism ensures near 100 percent modification efficiency at some 200 nucleotides in every one of the 10 million or so rRNAs per cell.

overall_design: RiboMethSeq sequencing data from PAF49, PAF49-GFP, PAF49delta1/2IDR-GFP, and PAF49deltaIDR-GFP samples used to make the Fig.4J

first: Koceila

last: Meznad

first: Manisha

last: Deogharia

first: Ludivine

last: Wacheul

first: Christiane

last: Zorbas

first: Thomas

last: Meier


title: Single-Cell Transcriptomics Unravels the Early Immune Landscape of Renal Allograft Rejection and Nominates Ccl3-Ccr5 as a Therapeutic Target

city: Xi'an

first: Lijun

state: Shaanxi

address: 127 Changle West Road

institute: Department of Urology, Xijing Hospital, Air Force Medical University,

summary:
Background Acute rejection is a primary determinant of long-term allograft survival, and the early-stage immune cell dynamics with local intercellular communication during this process require further elucidation.
Methods We performed single-cell RNA sequencing (scRNA-seq) on CD45+ immune cells isolated from rat renal allografts during the early phase of acute rejection (days 0, 1, 3, and 7). Using unsupervised clustering, functional enrichment analysis, cellular trajectory inference, and intercellular communication network mapping, we delineated the immune cell dynamics and local communication networks at single-cell resolution. Our findings were subsequently validated through multiplex immunofluorescence and therapeutic intervention experiments.
Results Macrophages constituted the dominant immune population during acute rejection. Sub-clustering analysis revealed a rapid expansion of the Isg15+Mac subset by post-transplant day 1, which persisted at elevated levels thereafter. Functional enrichment and trajectory inference demonstrated the pro-inflammatory properties of Isg15+Mac, implicating this subset in acute rejection. Cell-cell communication analysis identified Ccl3-Ccr5 ligand-receptor interactions between Isg15+Mac and T cells. Multiplex immunofluorescence confirmed abundance of Isg15+Mac within the allografts. Moreover, We alleviated the acute rejection after kidney transplantation using the FDA-approved Ccr5 blocker Maraviroc.
Conclusions Our study establishes an in-depth, early-stage immune landscape of renal transplantation, uncovering the Isg15+Mac subset as a critical driver of acute rejection via the Ccl3-Ccr5 axis. And also indicating that Maraviroc may potentially be a therapeutic candidate for transplant rejection.

overall_design: Untransplanted kidneys served as the control group. For the experimental groups, transplanted kidneys were sampled at 1, 3, and 7 days post - transplantation, with Wistar rats as donors and SD rats as recipients. CD45+ immune cells were sorted from these samples for single - cell sequencing. No drug treatment was administered to the experimental groups, and each group (both experimental and control groups) had a sample size of n = 3.


title: Targeting Menin in T-Lineage Acute Lymphoblastic Leukemia [menin_treatments]

city: Chicago

first: Caner

last: Saygin

state: IL

address: 900 E 57th Street, KCBD 7123B

institute: The University of Chicago

summary: The efficacy of menin inhibition was evaluated in 14 primary T-ALL samples with diverse genetic backgrounds using ziftomenib. Sensitivity was not correlated with HOXA expression or KMT2A rearrangements, and in vivo treatment significantly reduced tumor burden in xenograft models without notable toxicity. Transcriptomic and proteomic analyses confirmed on-target activity, including downregulation of menin targets and activation of differentiation pathways. Phosphoproteomic profiling identified phospho-MEF2C (S222) as a key marker of sensitivity, regulated by CDK and MAPK signaling. Combination treatment with ziftomenib and CDK1/2 or ERK1/2 inhibitors showed synergistic effects, suggesting a mechanistic link between menin inhibition and p-MEF2C–driven T-ALL vulnerability.

overall_design: Primary human T-ALL blasts were collected from the peripheral blood or bone marrow of patients under informed consent, in accordance with protocols approved by the University of Chicago Institutional Review Board. Mononuclear cells were isolated using Ficoll-Paque PLUS density gradient centrifugation and cryopreserved in fetal bovine serum (FBS) containing 10% dimethyl sulfoxide (DMSO). For in vitro assays, cells were thawed and cultured in RPMI-1640 medium supplemented with 10% FBS, 2 mM L-glutamine, and an insulin–transferrin–selenium supplement at 37°C and 5% CO2 in the gas phase. Cells were treated with 2.5 mM ziftomenib, and total RNA was extracted on days 4 and 7 using TRIzol reagent (Thermo Fisher Scientific), following the manufacturer’s instructions. Untreated baseline samples (day 0) were also collected, and all conditions were processed in technical replicates.

first: Kathryn

last: Shimamoto

first: Diren A

last: Karaoglu


title: Targeting Menin in T-Lineage Acute Lymphoblastic Leukemia [MEF2CKD]

overall_design: Karpas-45 cells were thawed and cultured in RPMI-1640 medium supplemented with 10% FBS and 1% penicillin-streptomycin-glutamine at 37°C and 5% CO₂. Cells were transduced with lentiviral vectors expressing a GFP-linked shRNA targeting MEF2C (target sequence: CCCAATGAATTTAGGAATGAA). Lentivirus was generated in HEK293T cells via co-transfection of the shRNA plasmid with packaging plasmids psPAX2 and pMD2.G using PEI. Viral supernatants were collected at 48 and 72 hours post-transfection and concentrated by ultracentrifugation. Transductions were performed by spinoculation. After 24 hours, cells were washed and cultured in fresh medium. GFP-positive cells were sorted by FACS at 120 hours post-transduction.


title: Targeting Menin in T-Lineage Acute Lymphoblastic Leukemia [MEF2COE]

overall_design: KARPAS-45 cells were thawed and cultured in RPMI-1640 medium supplemented with 10% FBS and 1% penicillin-streptomycin-glutamine at 37°C and 5% CO₂. Cells were transduced with lentiviral vectors expressing MEF2C under the control of the EF1A promoter and linked to mCherry (MEF2C-OE-mCherry). Lentivirus was generated in HEK293T cells via co-transfection of the overexpression plasmid with packaging plasmids psPAX2 and pMD2.G using PEI. Viral supernatants were collected at 48 and 72 hours post-transfection and concentrated. Transductions were performed by spinoculation. After 24 hours, cells were washed and cultured in fresh medium. mCherry-positive cells were sorted by FACS at 120 hours post-transduction.


title: CXCL16-driven CD4+ T cells orchestrate immunosurveillance against MHC-I-deficient tumors

city: Shanghai

first: Yunyi

last: Zhou

address: 1954 Huashan Road, Xuhui District

institute: Shanghai Jiao Tong University

summary: MHC-I loss is a prevalent mechanism for immune evasion and resistance to immunotherapy. However, the mechanisms by which MHC-I loss shapes the tumor microenvironment (TME) and influences the interactions between MHC-I-deficient tumors and immune cells, ultimately affecting tumor growth, remain largely unknown. Here, we found that MHC-I/B2M loss increased tumor growth rates in the MC38, AKR and LLC1 tumor models, but surprisingly led to regression of Hepa1-6 tumors. The effect of MHC-I/B2M loss on tumor growth was associated with the changes in immune infiltrates. CD4+ T cell, NK cells and macrophages were required to suppress the growth of MHC-I-deficient Hepa1-6 tumors, with CD4+ T cells appearing essential for maintaining of the tumoricidal phenotype of monocytes/macrophages and for recruiting NK cells and monocytes/macrophages. Moreover, the differential infiltration of CD4+ T cells was attributed to the upregulation of CXCL16 expression in Hepa1-6 upon loss of MHC-I/B2M, and the downregulation of CXCL16 due to MHC-I deficiency in other tumor models. CXCL16 exhibited a potent antitumor effect on MHC-I-deficient tumors by recruiting CD4+ T cells. In summary, CXCL16-driven CD4+ T cells are central regulators of antitumor immunity against MHC-I-deficient tumors.

overall_design: To investigate the mechanisms of MHC-I loss effecting tumor growth and tumor microenvironment, we established 4 MHC-I-deficient tumor cell lines (MC38, AKR, LLC1, and Hepa1-6) in which B2m has been knocked out by sgRNA. The non-targeting sgRNA was used for control. We then performed gene expression profiling analysis using data obtained from RNA-seq of 4 paired different cell lines.

first: Ping

last: Chen

first: Zhixue

last: Wang

first: Liming

last: Gui

first: Zhengheng

last: Zheng

first: Weiqiang

last: Gao

first: Bin

last: Ma


title: RNA Sequencing of Mouse Liver Post Caloric Resrtiction and Anti-Anging Interventions

first: Alireza

last: Khoddam

address: 303 E Superior St 8-400

department: Cardiology

institute: Northwestern University

summary: Caloric restriction (CR) extends lifespan in mammals but the stringent 20% reduction in food intake is challenging to sustain; therefore, validating pharmacological mimics is of high interest. It has been previously reported that a SERPINE1 mutation in an Amish cohort links to a 10-year lifespan increase and improved metabolism. SERPINE1 encodes for Plasminogen Activator Inhibitor-1 (PAI-1) which is a clinical predictor of vascular stiffness, hypertension, and diastolic dysfunction. Moreover, PAI-1 amplifies accumulation of senescent cells as a component of the senescence associated secretory phenotype (SASP). Here, we compared hepatic transcriptomic profiles in mice after 8 weeks of CR, rapamycin, metformin, or the PAI-1 inhibitor TM5614 versus an ad libitum control. CR induced extensive changes (1087 genes downregulated, 1145 upregulated), while TM5614 showed the closest mimicry (211 down, 218 up), sharing ~50% of CR’s differentially expressed genes, including downregulated Il11ra1 and upregulated Per2, tied to longevity and metabolism. Rapamycin and metformin showed minimal overlap with CR. Gene Ontology analysis revealed CR and TM5614 downregulated metabolic pathways, and both countered gene-length-dependent transcription decline, an emerging measure of an aging transcriptomic landscape. TM5614 presents as a CR mimic, offering an alternative to CR to extend healthspan.

overall_design: Liver transcriptomes were analyzed from 30 C57BL/6J mice following 8 weeks of intervention beginning at 21 weeks of age. Mice were assigned to one of five groups: ad libitum control (n=6), 30% caloric restriction (CR) (n=3), TM5614 (20 mg/kg/day) (n=3), rapamycin (42 mg/kg/day) (n=3), or metformin (37.5 mg/kg/day) (n=3), with compounds incorporated into chow. At the end of the study, whole-liver RNA was extracted for sequencing (reported here in the GEO Submission).

first: Benjamin

last: Weidemann

first: Mesut

last: Eren

first: Lisa

last: Wilsbacher


title: Establishment of Human Formative Pluripotent Stem Cell Like Cells Exhibiting Amniotic Differentiation Potentials [WGBS]

city: Beijing

first: Qizhi

address: Chaoyang District

institute: Chinese Academy of Sciences

type: Methylation profiling by high throughput sequencing

summary: Human embryos undergo pivotal morphogenetic remodelling shortly after implantation. The understanding of this crucial stage is severely impeded by the scarcity of embryonic samples and ethical constraints. Pluripotent stem cells with the competence for somatic and germline differentiation serve as in vitro models of epiblast. In this study, we established human formative pluripotent stem cells-like cell (hfPSC-LCs) from naive human embryonic stem cells (hESCs), conventional hESCs, human induced pluripotent stem cells (hiPSCs), as well as human blastocysts using the three dimensional (3D) Matrigel culture system. Similar to pre gastrula stage epiblast hfPSC-LCs self-organize into self-renewing colonies with an apical lumen and exhibit several hallmarks of formative pluripotency, consistent with the properties observed in mouse fPSCs. Notably, single cells of hfPSC-LCs could differentiate into amnion-like precursor cells (hALPCs) which are transcriptionally and morphologically similar to the bona fide amnion. Meanwhile, hfPSC-LCs directly respond to primordial germ cell (PGC) induction signals, generating PGC-like cells (PGCLCs) either as single - cell aggregates or intact colonies, with an efficiency of approximately 50%. Chromatin accessibility analysis revealed that the differentiation capacity of hfPSC-LCs for gametes and amnion lineages might correlate with the accessible chromatin architecture of PGC and amnion associated genes. Loss of 3D-Matrigel niche disrupts formative pluripotency in both mouse and human, manifesting as downregulated formative markers and compromised differentiation capacity. Collectively, our findings establish hfPSC-LCs as a 3D model for investigating formative pluripotency of humans, thereby probably addressing a critical gap in the understanding of human pluripotency transitions.

overall_design:
To decipher the multi-omics landscape of hfPSCs cultured in three-dimensional Matrigel, we conducted bulk RNA-seq, single-cell RNA-seq (scRNA-seq), ATAC-seq, and whole-genome bisulfite sequencing (WGBS).
The hfPSC-LCs were cultured in medium (50 ml) consisted of: 50 ml E8 complete medium, 2 µM IWP2 (Merck, Cat# 681671), 1% HSA (Sigma, Cat# A1653) and 10 µM Rho-associated kinase inhibitor Y27632 (TOCRIS, Cat# 1254) or 10% CloneR (STEM CELL, Cat# 05888). hfPSC-LCs were established from H9 and MGT3 cell line. The WGBS samples associated with this culture method are labeled hfPSCs_H9_DNA methylation and hfPSCs_MGT3_DNA methylation.

first: Xiaoxiao

first: Yu

last: Wu


title: Glucocorticoid and mineralocorticoid receptors jointly promote vascular development in kidney  organoids

city: BAR HARBOR

first: CGDS

last: Data

state: ME

address: 159 OLD BAR HARBOR RD

institute: MDI Biological Laboratory

summary: To examine the co-development of vasculature and renal epithelial tissue, we employed a human pluripotent stem cell-derived kidney organoid system. We found that cooperative signaling through the glucocorticoid receptor and mineralocorticoid receptor via hydrocortisone enabled rich endothelial cell differentiation and vessel formation. Bulk RNA sequencing analysis revealed that hydrocortisone perturbs an angiogenic transcriptional program early in development and promotes instead a pro-endothelial survival transcriptional program, with upregulation of angiopoietin 1 at both the mRNA and protein level. Additionally, we saw that hydrocortisone does not seem to significantly affect gene expression of canonical nephrogenic genes compared to our controls, suggesting its effect is largely restricted to endothelial cell differentiation. Our results show that kidney organoids offer a unique platform to study developmental signals that drive endothelial cell differentiation and vessel formation.

overall_design: Bulk RNA-seq profiling of hydrocortisone-treated and dmso-treated (control) kidney organoid cultures at day 9 of differentiation

first: Cory

middle: P

last: Johnson

first: Hannah

middle: M

last: Somers

first: Sophie

middle: E

last: Craig

first: Heath

last: Fuqua

first: Lynne

last: Beverly-Staggs

first: Kailee

last: Tanaka

first: Sydney

last: Brown

first: Charles

middle: H

last: Toulmin

first: Joel

last: Graber

first: Melissa

last: Maginnis

first: Hermann

last: Haller


title: Establishment of Human Formative Pluripotent Stem Cell Like Cells Exhibiting Amniotic Differentiation Potentials [bulk RNA-seq]

last: Wang

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design:
To decipher the multi-omics landscape of hfPSC-LCs cultured in three-dimensional Matrigel, we conducted bulk RNA-seq, single-cell RNA-seq (scRNA-seq), ATAC-seq, and whole-genome bisulfite sequencing (WGBS).
The primary mfPSCs (mfPSCs_P0) were transformed from mouse embryonic stem cells (CMTI-1) in N2B27 medium supplemented with 1% KSR, 20 ng/ml Activin A, 12 ng/ml bFgf2, and 5 μM XAV939 in Matrigel. In RNA-seq samples formatted as mfPSCs_P0_24-72h_1/2_R1/2.fq.gz, '24-72h' denotes the time interval of 24 to 72 hours post-passage; ’1/2’ indicates two biological replicates; ’R1/R2’ represents the paired-end sequencing reads.
Conventional embryonic stem cells (hESCs-H9/MGT3) were maintained in commercial E8 medium (Cat# A15169) on Matrigel (Cat# 356230/354230/356231) coated dishes at 37 °C and 5% CO2. The hESCs were passaged mechanically every 5–7 days by a brief PBS (HUANKE, Cat #HK2109.21) wash followed by dissociation using EDTA (Cat# 15575-038). hESCs_H9_R1/R2.fq.gz and hESCs_MGT3_R1/R2.fq.gz represent conventional embryonic stem cells.
The hfPSC-LCs/hifPSCs were cultured in medium (50 ml) consisted of: 50 ml E8 complete medium, 2 µM IWP2 (Merck, Cat# 681671), 1% HSA (Sigma, Cat# A1653) and 10 µM Rho-associated kinase inhibitor Y27632 (TOCRIS, Cat# 1254) or 10% CloneR (STEM CELL, Cat# 05888). hfPSC-LCs were established from H9 and MGT3 cell line. hifPSCs were obtained from human induced pluripotent stem cells (hiPSCs). The RNA-seq samples associated with this culture method are labeled hfPSCs_H9, hfPSCs_MGT3, hifPSCs
Human amnion-like stem cell (hALSCs) were established from hfPSC-LCs based on the following procedures. Firstly, hfPSC-LCs were dissociated into single cells using 50% TrypLE Express and seeded in Matrigel. Then they were cultured in amnion-like stem cell medium. Human amnion-like stem cell medium (50 ml) including: 50 ml N2B27 basal medium, 1 mM PD0325901 (LC Laboratories, Cat# P-9688) and 1 mM A83-01 (Selleck, Cat# S7692). For human amnion-like stem cells (hALSCs) cultured using the above method, the corresponding RNA-seq samples are hALSCs_1_R1/R2.fq.gz and hALSCs_2_R1/R2.fq.gz
hESCs_sus were hESCs cultured in a suspension manner in E8 medium supplemented with 5uM Y27632. Suspension-cultured hESCs are labeled as hESCs_sus_2_R1/R2.fq.gz and hESCs_sus_1_R1/R2.fq.gz.
Following stable 3D culture, mfPSCs were adapted to Matrigel-coated 2D plates for 1-2 passages (3D to 2D_P1/P2) in mfPSCs medium. In the filenames: 3D_to_2D_P1_1/2_R1/R2.fq.gz denote the first generation of cells cultured under this protocol. And 3D_to_2D_P2_1/2_R1/R2.fq.gz  denote the second generation of cells cultured under this protocol. ’1/2’ indicates two biological replicates.


title: Establishment of Human Formative Pluripotent Stem Cell Like Cells Exhibiting Amniotic Differentiation Potentials [ATAC-seq]

type: Genome binding/occupancy profiling by high throughput sequencing

overall_design:
To decipher the multi-omics landscape of hfPSC-LCs cultured in three-dimensional Matrigel, we conducted bulk RNA-seq, single-cell RNA-seq (scRNA-seq), ATAC-seq, and whole-genome bisulfite sequencing (WGBS).
The hfPSC-LCs were cultured in medium (50 ml) consisted of: 50 ml E8 complete medium, 2 µM IWP2 (Merck, Cat# 681671), 1% HSA (Sigma, Cat# A1653) and 10 µM Rho-associated kinase inhibitor Y27632 (TOCRIS, Cat# 1254) or 10% CloneR (STEM CELL, Cat# 05888). The ATAC-seq samples associated with this culture method are labeled hfPSCs_1_ATAC and hfPSCs_2_ATAC.


title: Establishment of Human Formative Pluripotent Stem Cell Like Cells Exhibiting Amniotic Differentiation Potentials [scRNA-seq]

overall_design:
To decipher the multi-omics landscape of hfPSC-LCs cultured in three-dimensional Matrigel, we conducted bulk RNA-seq, single-cell RNA-seq (scRNA-seq), ATAC-seq, and whole-genome bisulfite sequencing (WGBS).
The hfPSC-LCs were cultured in medium (50 ml) consisted of: 50 ml E8 complete medium, 2 µM IWP2 (Merck, Cat# 681671), 1% HSA (Sigma, Cat# A1653) and 10 µM Rho-associated kinase inhibitor Y27632 (TOCRIS, Cat# 1254) or 10% CloneR (STEM CELL, Cat# 05888). The scRNA-seq samples associated with this culture method are labeled hfPSCs_single cell_1, hfPSCs_single cell_2, hfPSCs_single cell_3 and hfPSCs_single cell_4.


title: Establishment of Human Formative Pluripotent Stem Cell Like Cells Exhibiting Amniotic Differentiation Potentials

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

overall_design: Refer to individual Series


title: BRD4 signaling maintains the differentiated state of β cells [scATAC-seq]

city: Jinan

first: Jia

last: Song

state: Select...

address: No.107 Wenhua Xi Road, Jinan,Shandong Province

institute: Qilu Hospital of Shandong University

summary: In diabetes, pancreatic β cells degenerate from their mature differentiated state to a dedifferentiated state. BRD4 plays a pivotal role during embryogenesis and cancer development, but its function in modulating β-cell differentiation remains unknown. In this study, multiple models including calorie restriction db/db mouse, long-term and acute conditional knockout mouse and human islet organoids were adopted to assess BRD4 function in β cells. 222 young patients with diabetes were also recruited for whole exome sequencing (WES) to screen for BRD4 mutations. Our study showed that BRD4 expression was significantly reduced in human diabetic β cells while significantly increased after calorie restriction in the diabetic mouse. β cell differentiation was impaired after long-term and acute Brd4 knockout. BRD4 knockdown in human islet organoids results in the loss of differentiation and reduction of insulin synthesis. We found that p.R749C can significantly affect BRD4 signaling and might be a causative mutation contributing to diabetes development in patients. Our study also showed that ATF5 is a direct target of the BRD4 pathway in β cells. Targeting BRD4-mediated regulatory networks may hold promise for developing novel therapeutic strategies to maintain the differentiated state of β cells.

overall_design: The islets of Brd4 conditional knockout (PBKO) and control mice were subjected to scATAC-seq.

first: Xinguo

last: Hou

first: Fuqiang

last: Liu

first: Yujing

last: Sun


title: BRD4 signaling maintains the differentiated state of β cells [scRNA-seq]

overall_design: db/db mice were given a limited food. The islets of Db/db, Db/m, dbR2 and dbR4 mice were subjected to scRNA-seq.


title: RNA sequencing data of leptomeningeal/perivascular CAMs and MG following LPS stimulation 8 weeks after repopulation

city: Freiburg

first: Maximilian

last: Fliegauf

address: Breisacherstraße 113

department: Institute for Neuropathology

country: Germany

institute: Uniklinikum Freiburg

summary: Perivascular and leptomeningeal macrophages are non-parenchymal macrophages summarized as central nervous system (CNS)-associated macrophages (CAMs) that mediate immune responses at brain boundaries. Both, CAMs and juxtaneuronal microglia are derived from prenatal yolk sac (YS) precursors, long-living and maintain their populations by homeostatic self-renewal without input from the periphery. Whereas microglia have been shown to be repopulated by CNS endogenous remnants of the same lineage following depletion, the renewal biology of CAMs is still poorly understood. Here, by combining multilineage myeloid fate mapping, bulk and single-cell profiling and high resolution confocal imaging, we show that the repopulation is strikingly different between CAMs and microglia. In contrast to microglia, CAMs do not renew exclusively cell-autonomously, but transiently utilize CCR2+Ly-6C+ monocytes after niche induction in an integrin-dependent manner. Remarkably and unlike repopulated microglia, replenished monocyte-derived CAMs remain transcriptionally and functionally distinct from their YS-derived counterparts. Finally, we established a protocol that allows to selectively exchange CAMs modulating disease response without functionally affecting parenchymal microglia. These new insights into the biology of the CNS immune system offer completely new therapeutic avenues for diverse neuroinflammatory and neurodegenerative diseases.

overall_design: Three WT or three Ccr2-/- mice per group either were left untreated or were treated with Vehicle (CTL) or with BLZ945 and 8 weeks later, all treated mice were injected with LPS. 12h later, mice were perfused and CAMs as well as MG were sorted for the generation of bulk transcriptomics data. In addition, three Ccr2CreERT2 R26tdT mice per group were treated with BLZ945 and 8 weeks later injected with either PBS or LPS. 12h later, mice were perfused and CAMs (tdTpos and tdTneg) as well as MG were sorted for the generation of bulk transcriptomics data. RNA expression levels between the different groups were compared. For details reagrding the analysis see methods section of the publication.

first: Marco

last: Prinz

first: Lukas

last: Amann


title: Cuproptosis and disulfidptosis converge to empower PD-L1 checkpoint therapy via Cadict-induced PD-L1 translation

city: Guangzhou

first: Shaoqing

last: Huang

address: No. 58, Zhongshan 2nd Road, Guangzhou

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

summary: To elucidate the mechanism by which Cadict and Cu-Cys exert cytotoxic effects on colorectal cancer cells, we performed transcriptome analysis using RNA-seq to investigate the signaling pathways affected by Cadict and Cu-Cys.

overall_design: Total RNAs from MC38 cells were extracted with TRIzol reagent and sequenced on the Illumina Novaseq 6000 sequencing instrument (BGI, China). There were three experimental groups (CTL vs Cu-Cys vs Cadict) with 3 replicates for each.


title: RNA sequencing data of whole tissue or CAMs isolated from WT mice after ischemic stroke

overall_design: WT mice were treated for seven consecutive days either with Vehicle or BLZ945 (200 mg/KG BW). Eight weeks the treatment, mice were subjected to a thromboembolic MCAo model of ischemic stroke. At 1d after stroke induction, whole tissue sections of contralateral and ipsilateral hemispheres were isolated for bulk RNAseq. In addition, leptomeningeal/perivascular CNS-associated macrophages (CAMs) were sorted from contralateral and ipsilateral hemisphere for bulk RNAseq.

first: Damien

last: Levard

first: Marina

last: Rubio


title: synNotch CAR-Macrophages promote persistent antitumour activity through enhanced adaptive immunity [scRNA-Seq human]

city: Changchun

first: Jiayi

last: chen

state: 吉林Jilin

address: School of Life Sciences

institute: Jilin University

summary: Chimeric antigen receptor-macrophage (CAR-M) possess the ability to migrate into the immunosuppressive tumour microenvironment (TME) of solid tumours and mediate phagocytosis. However, their efficacy is limited by challenges in precisely controlling tumour-specific immune activation and sustaining adaptive antitumour responses due to systemic off-target risks and inefficient antigen presentation. To overcome these limitations, we engineered an antigen prime-and-kill circuit, wherein tumour-specific synthetic Notch (syN) receptors locally induce the production of a kill antigen. This circuit enhances the precision of CAR-M while establishing self-sustaining antigen-presentation networks, amplifying tumour-specific T-cell adaptive immunity without eliciting systemic cytokine toxicity. To enable in situ CAR-M generation, we developed a macrophage-derived exosome-based delivery system to introduce syN-CAR genes into tumour-associated macrophages (TAMs), thereby generating syN-CAR-M within the TME. These engineered CAR-M actively sought and engulfed cancer cells while promoting a robust adaptive antitumour immune response in the TME in a murine tumour model. Our findings highlight that in situ induction of syN-CAR-M triggers a potent, tumour-specific adaptive immune response, offering a novel strategy for CAR-M-based immunotherapy and advancing precision cancer treatment by enhancing efficacy while minimizing systemic toxicity.

overall_design: RNA-seq profiling of lung tumour tissues after treatment with PBS or syN-CAR-exo.


title: RNA sequencing data of leptomeningeal/perivascular CAMs 8 weeks after repopulation. [scRNA-Seq]

overall_design: Two male and two female Mrc1CreERT2 R26tdT mice per group were treated with CTL or BLZ945 for seven consecutive days. Eight weeks after the treatment, Lineage- CD45int-hi CD11b+ were isolated from the mice. Cells isolated from each of the tow animals of the same sex and treatment group were pooled and used for the generation of in total four single-cell RNA-sequencing libraries.


title: A LINE-1 retrotransposon promotes SEMA3C expression as a cis-regulatory enhancer to sustain breast cancer stem cell survival

first: Qidong

last: Xia

state: Guangdong

address: No.107, Yangjiangxi Road, Guangzhou

department: Breast Tumor Center

institute: Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University

summary: Transposable elements (TEs), which are under tight epigenetic control, have been co-opted as cis-regulatory elements to regulate gene expression in development and cancer. Among them, long interspersed element 1 (LINE1) retrotransposons are the most abundant and exhibit high activity in embryonic stem cells.To investigate the precise roles of LINE1 in breast cancer stem cells (BCSCs) , we performed total RNA-seq analysis of non-CSCs and CSCs enriched from EO771 cells.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for non-CSCs and CSCs.

first: shicheng

last: Su


title: Indirect identification of genomic G-quadruplexes via a small protein probe that specifically recognizes C-rich single-stranded DNA

city: Changsha

first: Kewei

last: Zheng

address: Lushan Road

department: School of Biomedical Sciences

institute: Hunan University

summary: Detecting intracellular genomic G-quadruplexes (G4s) is crucial for understanding their biological functions. Although various G4 recognition probes have been developed, there remains a need for new G4 detection technologies to create detailed and reliable genomic G4 maps. In this study, we developed a small protein (CK13) that specifically recognizes the complementary C-rich single-stranded DNA (ssDNA) released during the formation of G4. Based on CK13 and CUT&Tag technology, we identified tens of thousands of C-rich ssDNA sites within human genomic DNA. These sites contain the vast majority of G4 sites detected by G4 probes, indicating that CK13 can well confirm the results of traditional G4 probes. Since CK13’s binding to C-rich ssDNA is minimally influenced by G4-binding proteins, it produces strong signals at the sites where intracellular G4-binding proteins are present. This indicates that, beyond free G4 structures, CK13 can also detect G4s occupied by G4-binding proteins within cells. Our findings demonstrate that C-rich ssDNA complementary to G4 can serve as an indirect marker for G4 formation, offering a promising approach to further explore the regulatory roles of G4s and their interacting proteins.

overall_design: Cleavage under targets and tagmentation (CUT&Tag) for G4P,CK13,NCL,and Top1 in HeLa, HEK293T, and HCT116 cells.

first: Juan-nan

last: Chen

first: Mei-lin

last: Xie

first: Jiang-yu

last: Yan

first: Jing

first: Ke-wei


title: BRD4 signaling maintains the differentiated state of β cells

first: Jia

last: Song

overall_design: The islets of Brd4 conditional knockout (PBKO) and control mice were subjected to scRNA-seq

first: Xinguo

first: Fuqiang


title: Cranial placode differentiation defect in individuals born without a nose

city: Singapore

first: Vanitha

middle: Venkoba

last: Rao

address: National University of Singapore Department of Biological Sciences  14 Science Drive 4 Block S1A, #05-05

department: Department of Biological Sciences

country: Singapore

institute: National University of Singapore

summary: Bosma arhinia microphthalmia syndrome (BAMS) is a rare congenital disorder characterized by the absence of a nose, along with eye and reproductive anomalies. BAMS is caused by heterozygous missense variants in SMCHD1, an epigenetic regulator. Despite uncovering its genetic basis, the cellular basis of the disease has remained elusive. The embryonic development of the nose involves direct contributions from both the neural crest and cranial placodes. Here we differentiated patient-derived induced pluripotent stem cells towards the cranial placode lineage and found that they exhibited significant differentiation defects. Combined transcriptome and DNA methylome analyses further revealed dysregulation in cell adhesion but without overt apoptosis. Together our research suggests that BAMS is caused by impaired differentiation to cranial placode cells and changes in cell adhesion, offering new insights into the cellular pathology of this enigmatic syndrome.

overall_design: RNA-seq of Cranial Placodal cells (Day 12) differentiated from Control and BAMS patient derived iPSC

last: Venkoba Rao

first: Shifeng

last: Xue


title: Spatial  transcriptomic comparison between IPF and donor lung distal explant tissue

city: los angeles

first: Changfu

last: Yao

state: Ca

address: 8700 Beverly Blvd. AHSP Room A9401

department: Lung and Regenerative Medicine Institutes

institute: cedars-sinai medical center

summary: Pulmonary fibrosis remains a significant public health concern due to an increasing incidence and limited treatment options. Alveolar type 2 epithelial cell (AT2) senescence causes stem cell exhaustion and progenitor failure, leading to ineffective and aberrant alveolar repairs and unresolved fibrosis. AT2 senescence regulatory molecules require additional studies. The spatial transcriptomics study we present here is aiming to comparing end stage IPF explant tissue with donor distal tissue to better understand the molecular changes associated with diseases.

overall_design: 3 donor explant samples vs 2 IPF explant samples are subjected to 10x visium spatial analysis

first: Tanyalak

last: Parimon

first: Peter


title: scRNAseq on mouse lungs of bleomycin injured aged mice

summary: Pulmonary fibrosis remains a significant public health concern due to an increasing incidence and limited treatment options. Alveolar type 2 epithelial cell (AT2) senescence causes stem cell exhaustion and progenitor failure, leading to ineffective and aberrant alveolar repairs and unresolved fibrosis. AT2 senescence regulatory molecules require additional studies. We demonstrate that the overexpression of syndecan-1 on IPF-AT2 cells is positively associated with the AT2 composite transcriptomic senescence score (SenMayo Senescence Score) and cellular senescence proteins. These findings imitate those of young and aged bleomycin-injured wild-type (WT) mice. The roles of syndecan-1 in regulating AT2 cellular senescence are corroborated in human and mouse lung epithelial cell lines. Functionally, the AT2 alveolospheres from bleomycin-injured Sdc1–/– mice had higher colony-forming efficiency (CFE) and normal differentiation, whereas WT alveolospheres were less proliferative and aberrant differentiation. Using precision-cut lung slices (PCLS), bleomycin-injured WT lungs had lower AT2 cells and a lower secretory surfactant protein C than Sdc1–/– PCLS, indicating the detrimental role of excess syndecan-1.  We further showed that the transcriptomic p53 activity score is higher in IPF and bleomycin-injured WT AT2 cells compared to Sdc1–/– AT2 cells. Additionally, syndecan-1 activated p53 by inducing acetylation at lysine379. Our work indicates that syndecan-1 reprograms AT2 cells to become senescent during lung fibrogenesis and likely through the p53 signaling pathway.

overall_design: 18-24 months old wild type and Sdc1 knockout mice were subjected to bleomycin injury. Lung tissue are reovered at different timepoints including day 14, 28 and 56.


title: Activity-dependent ribosome profiling reveals the landscape of canonical and non-canonical translation in brain tissue [Ribo and RNA-Seq]

city: Wako

first: Nayan

last: Suryawanshi

state: Saitama

address: Hirosawa 2-1

department: Center for Brain Science

country: Japan

institute: RIKEN

summary: Identification hundreds of activity-induced mORFs and uORFs, a subset of which served as robust markers for activity status

overall_design: Ribosome footprints count analysis to indentify novel ORFs using data obtained from RNA-seq (Ribo-seq)

first: Yi-Kai


title: synNotch CAR-Macrophages promote persistent antitumour activity through enhanced adaptive immunity

first: Jiayi


title: RNA sequencing data of leptomeningeal/perivascular CAMs 8 weeks after repopulation.

first: Maximilian

last: Fliegauf

overall_design: tdTpos and tdTneg leptomeningeal/perivascular CNS-associated macrophages (CAMs) were isolated from 4 individual Mrc1CreERT2 R26tdT mice per treatment group and used for the generation of bulk transcriptomics data. RNA expression levels between the different groups were compared. CTL = tdT+ CAMs from Vehicle treated mice; neg = tdT- CAMs from BLZ945 treated mice; pos = tdT+ CAMs from BLZ945 treated mice

first: Marco

last: Prinz

first: Lukas

last: Amann


title: synNotch CAR-Macrophages promote persistent antitumour activity through enhanced adaptive immunity [scRNA-Seq]

last: chen


title: NSUN7 knockdown increases the sensitivity of tamoxifen-resistant MCF-7 cells to 4-hydroxytamoxifen

city: La Jolla

first: Andrei

last: Chernov

state: CA

address: 9500 Gilman Dr

department: Anesthesiology

institute: University of California San Diego

summary: Tamoxifen is the commonly prescribed drug for estrogen receptor positive breast cancer treatment. Recently dysregulation in RNA 5-methylcytosine (m5C) modification and its regulating enzymes have been suggested as potential contributors in chemotherapeutic resistance in various cancers, however their role in determining tamoxifen resistance in breast cancer remains unknown. In this study we analyzed the expression of m5C modulating enzymes recognized as writers (NSUN2-7), readers (YBX-1 and ALYREF) and eraser (TET3), and ELISA based assay to check global methylation status in tamoxifen-resistant breast cancer cell line (MCF-7 Tam1) compared to the sensitive MCF-7 cell line. We found that global m5C status and NSUN7 writer enzyme expression was significantly higher in MCF-7 Tam1 cells. Furthermore,  Small interfering RNA mediated NSUN7 depletion significantly restored the sensitivity and resulted in an approximate 50% decrease in IC50 value compared to the control. Furthermore, NSUN7 deficient cells showed a significant reduction in migration and colony formation ability. RNA sequencing and Gene set enrichment analysis showed that NSUN7 targeted genes were enriched in pathways and processes regulating cell cycle, cell migration and proliferation. Further screening of differentially expressed genes revealed that NSUN7 deficiency caused dysregulation in experimentally determined genes associated with drug resistance particularly tamoxifen resistance. The hub genes identified in NSUN7-deficient tamoxifen-resistant cell line showed significant association with the disease prognosis in terms of overall and disease-free survival. Hence, NSUN7 is a potential therapeutic target for combating tamoxifen resistance in breast cancer cells. Moreover, it has a critical role in determining metastatic characteristics in breast cancer which makes it a candidate prognostic marker.

overall_design: Since NSUN7 writer enzyme expression was significantly higher in MCF-7 Tam1 cells, therefore, to investigate its role in tamoxifen resistance and cancer progression, we knocked down its expression using siRNA. We performed gene expression profiling analysis using data obtained from RNA-seq of scrambled/control-siRNA and NSUN7 targeted siRNA treated MCF-7 Tam1 cells each having two replicates. Comparative gene expression profiling of RNA-seq data of control treated MCF-7 Tam1 cells and its KD derivatives followed by gene set enrichment analysis was performed.

first: Sana

last: Mahmood

middle: V

first: Sehrish

last: Khan

first: Ramamurthy

last: Chitteti

first: Salma

middle: S

last: Salihah

first: Sohail

middle: A

last: Qureshi

first: Hemal

last: Patel

first: Asma

last: Gul


title: NOC4L coordinates neuronal and pharyngeal arch development by regulating ribosome biogenesis-related processes

city: Zhan Jiang

first: Xiaoyu

last: Li

address: 57 People's Avenue

institute: Affiliated Hospital of Guangdong Medical University

summary: Bowen-Conradi syndrome (BCS) is an autosomal-recessive disease marked by lethality and characterized by neurodevelopmental delays, microcephaly, and craniofacial abnormalities, yet remains mechanistically underexplored due to the absence of robust biological models. To address this gap, we generated the first vertebrate model of BCS through CRISPR-Cas9-mediated noc4l knockout in zebrafish. Our model recapitulates key clinical features observed in BCS patients, including microcephaly, micrognathia, and developmental delay. Histopathology revealed diminished cell proliferation and elevated apoptosis within cephalic and pharyngeal arch regions of mutants. Molecular characterization revealed that rRNA processing defects impaired ribosome biogenesis, reducing 40S and 80S subunits and polysomes, ultimately leading to suppression of protein synthesis. Mechanistically, translatomic profiling identified PPARγ signaling as a significantly downregulated pathway in noc4l mutants. Rosiglitazone-induced PPARγ activation partially rescued cranial and pharyngeal cartilage defects, while also alleviating neurodevelopmental deficits. This work establishes impaired translational machinery as the pathogenic core of BCS and highlights PPARγ agonism as a viable therapeutic strategy, with rosiglitazone demonstrating preclinical efficacy in rescuing disease-relevant phenotypes. Our findings provide both a pathophysiological framework for BCS and a platform for testing translation-targeted interventions.

overall_design: We cut the head of the siblings and the noc4l -/- mutants with 1-ml syringe needle. Total RNA of the head from embryos at 5 dpf were isolated using TRIzol. RNA-Seq and Ribo-seq were performed in the Gene Denovo Biotechnology (Guangzhou, China). Differentially expressed genes (DEGs) were detected using DESeq2. Reactome enrichment analysis and GO enrichment analysis were performed to elucidate the functional implications of DEGs. DEGs with a false discovery rate (FDR) of less than 0.05 were selected for enrichment analysis.


title: Cancer-associated SF3B1 mutation suppress DNA damage repair by disrupting circRNA biogenesis

city: Chang Chun

first: Xin

last: Hu

address: 2699 Qianjin road,china

institute: Jilin university

type: Non-coding RNA profiling by high throughput sequencing

summary: Cancer-associated SF3B1 mutation leads to alternative splicing of hundreds of genes that affect cancer progression in multiple pathways. It has been reported that SF3B1 mutation interrupt with DNA damage repair independent of the BRCA1/RAD51 pathway. Here, we report that SF3B1 mutation compromise DNA homologous recombination (HR) repair by disrupting circATP9B biogenesis to repair the later stages of Rad51 loading. Mechanistically, circATP9B specifically binds to MYH9, and accelerate MYH9 protein degradation at the post-transcriptional level. circATP9B interferes with the motivity of DNA double-strand damage (DSB) sites by obstructing the assembly of nuclear actin and leads to persistent DSB sites in heterochromatin. It was further determined that the up-regulation of circATP9B enhanced the sensitivity of tumor cells to the PARP inhibitors, indicating that circATP9B interrupt with DNA damage repair. Taken together, these data shed light on the mechanisms by which SF3B1 interferes with DNA damage repair, providing new perspectives for treating tumors with SF3B1 mutation.

overall_design: In an effort to investigate whether SF3B1 mutation interferes with circRNA biogenesis, we performed RNA-seq of Hela and HEK293T cells with or without SF3B1 mutation to identify promising circRNA candidates.

first: Rui

last: Qian

first: Youzhong

last: Wan


title: Analyze the transcriptional signature of primary human renal progenitor cells (hRPCs) [scRNA-Seq]

city: Firenze

first: Giulia

last: Carangelo

address: Viale Pieraccini 6

country: Italy

institute: University of Florence

summary: We found that hRPC differentiation capacity  contributed to the sexual dimorphism of kidney disorders . To study the mechanisms controlling these differences we characterized the transcriptional signature of hRPC

overall_design: Transcriptional signature of primary human renal progenitor cells

first: Conte

last: Carolina

first: Carangelo

last: Giulia

first: Mazzinghi

last: Benedetta

first: Romagnani

last: Paola


title: Transcriptomics analysis of an EMS-induced bacterial blight resistant line of rice

city: Hyderabad

first: Ramesh

middle: Venkata

last: Sonti

state: Telangana

address: Habsiguda

country: India

institute: CSIR-Centre for Cellular and Molecular Biology

summary: An induced-mutant line derived from the elite rice cultivar, Samba Mahsuri, was identified to exhibit broad-spectrum resistance to BB. Using next-generation gene mapping approaches, we identified a genomic interval in chromosome 6 to be linked to BB resistance. Analysis of the SNPs in the locus and subsequent linkage analysis indicated that a missense SNP located in the second exon of the Mitogen-Activated Protein Kinase 6 (OsMAPK6) could be the causal mutation. A detailed examination of OsMAPK6 gene sequences in over 4000 rice genomes revealed that the mutant allele identified in this study is novel to rice germplasm. The candidate SNP causes the substitution of an invariant Serine residue with a Proline residue (S84P), thus likely affecting the protein function. Global transcriptome, metabolome, biochemical, and molecular analyses suggested that BB42 exhibit autoimmunity.

overall_design: SM and BB42 plants were inoculated with the Xoo isolate IXO1221 during the maximum tillering stage. One-centimeter-long leaf samples from the site of clipping were collected at the time of infection as control (0 hour/Mock) and 24-hour post infection (Xoo) from both SM and BB42. The samples were collected from three independent plants and considered as biological replicates. The samples were flash-frozen immediately after collection and stored at -80℃ until further processing. RNA was isolated from the samples using RNeasy Plant RNA Isolation Kit (Qiagen, Germany) with on-column DNA digestion using RNase-Free DNase Set (Qiagen, Germany). Purified RNA was quantified using QubitTM RNA HS Assay Kit (Thermo Fisher Scientific, USA). RNA integrity was assessed using agarose gel electrophoresis and BioAnalyzer 2100 (Agilent Technologies, USA). 1 µg of intact total RNA samples were used for library preparation using Illumina TruSeq Stranded Total RNA with Ribo-Zero Plant (Illumina, USA). Sequencing was performed on Illumina’s NovaSeq6000 platform with cycling conditions to obtain 2x100bp reads on an S2 flow cell.

first: Gokulan

last: CG

first: Hitendra


title: Metabolic Atlas of Early Human Cortex Identifies Regulators of Cell Fate Transitions

city: Stanford

first: Aparna

last: Bhaduri

address: 269 Campus Drive

institute: Stanford University

summary: Characterization of cell type emergence during human cortical development, which enables unique human cognition, has focused primarily on anatomical and transcriptional characterizations. Metabolic processes in the human brain that allow for rapid expansion, but contribute to vulnerability to neurodevelopmental disorders, remain largely unexplored. We performed a variety of metabolic assays in primary tissue and stem cell derived cortical organoids and observed dynamic changes in core metabolic functions, including an unexpected increase in glycolysis during late neurogenesis. By depleting glucose levels in cortical organoids, we increased outer radial glia, astrocytes, and inhibitory neurons. We found the pentose phosphate pathway (PPP) was impacted in these experiments and leveraged pharmacological and genetic manipulations to recapitulate these radial glia cell fate changes. These data identify a new role for the PPP in modulating radial glia cell fate specification and generate a resource for future exploration of additional metabolic pathways in human cortical development.

overall_design: 10X GEMX single-cell (sc) and single-nuclei (sn) RNA-sequencing of cortical organoids subjected to a metabolic perturbation. These perturbations were 1) glucose depletion followed by snRNA-seq 2) pharmacologic inhibition of the pentose phosphate pathway followed by snRNA-seq or 3) knockdown of pentose phosphate pathway enzymes

first: Jessenya

last: Mil

first: Jose

last: Soto


title: Mesenchymal stromal cells induce MYC repression in activated T cells

city: Atlanta

first: Edwin

last: Horwitz

state: Georgia

address: 1570 Haygood Drive NE

institute: Emory University

summary: We utilized 10X Genomics Multiome technology to obtain single cell gene expression and chromatin accessibility profiles to identify the genetic and epigenetic changes which underlie the suppressive effects of yMSCs on alloreactive T cells.

overall_design: Naïve T cells from 5 anonymous human donors activated for 6 days by allogeneic DCs from a sixth donor. Cells in experiemental group exposed to yMSC on the bottom of a transwell insert for 48 hours before being placed back into fresh media. snRNAseq and snATACseq libraries generated with 10x Multiome at every timepoint for longitudinal computational analysis.


title: NtispH as a Key Regulator of Glandular Trichome Development and Terpenoid Metabolism in Tobacco (Nicotiana tabacum)

city: GuiYang

first: Zeyang

last: Song

state: GuiZhou

address: West Campus: Jiaxiu South Road, Huaxi District.

institute: Guizhou University

summary: The comparative transcriptome analysis of leaves overexpressing NtispH and WT revealed that NtispH is a crucial regulator of glandular trichome development and terpenoid metabolism.

overall_design: To elucidate the pivotal role of NtispH in glandular trichome development, RNA-seq analysis was conducted on the leaves of positive overexpression materials and wild-type (WT) tobacco plants grown under identical conditions

first: Xinyue

last: Ye

first: Jie

last: Yang


title: Fish on Antidepressants: Venlafaxine Contamination in Our Waterways

city: Waterloo

first: Louis

middle: Joseph

last: Pfeifer

state: ON

address: 200 University Ave W

department: Biology

country: Canada

institute: University of Waterloo

summary: Wastewater treatment plant effluents (WWTPE) represent a pervasive source of pharmaceutically active compounds in surface waters, with the serotonin and norepinephrine reuptake inhibitor venlafaxine (VEN) frequently detected at elevated concentrations in receiving waters worldwide. VEN is a chiral pharmaceutical administered as a racemic (50:50) mixture of R- and S-enantiomers, which exhibit distinct pharmacological activities and can be selectively degraded or enriched in effluent depending on treatment processes. However, little is known about the stereoselective toxicological effects of VEN in fish.  This work investigates the effects of VEN exposure on Etheostoma caeruleum (rainbow darter) using transcriptomic profiling.

overall_design: Wild  Etheostoma caeruleum (rainbow darter) were collected from the Grand River, ON, Canada, and brought to the lab. Fish were allowed to acclimate and depurate for two weeks before being exposed to 1 µg/L of either the R- or S-enantiomer of venlafaxine for 14 days. Liver tissue was analyzed for transcriptomic responses to each enantiomer.

middle: J

first: Paul

last: Craig


title: Single-Cell RNA Sequencing Unveils Macrophage Involvement in Aortic Valve Stenosis Progression

city: Kobe

first: Takuo

last: Emoto

state: Hyogo

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

department: Division of Cardiovascular Medicine, Department of Internal Medicine

institute: Kobe University Graduate School of Medicine

summary: Introduction: Calcific aortic stenosis (AS) is an age-related progressive disease with increasing global prevalence. Although chronic inflammation is a key factor in its pathogenesis, the detailed immune cell profiles and their interactions with non-immune cells remain poorly understood. This study aimed to characterize the immune cell landscape in calcific AS at a single-cell level and evaluate the potential of anti-inflammatory therapy. Methods: Human aortic valve samples were obtained from four patients with non-calcified aortic regurgitation (AR) and 11 patients with AS undergoing surgical aortic valve replacement. Single-cell 3’ gene expression sequencing (scRNA-seq (3’)) was performed on CD45+ immune cells, while single-cell RNA sequencing Flex (scRNA-seq (Flex)) and bulk RNA sequencing were conducted on unsorted cells. Additionally, an anti-TNFα antibody experiment was performed in a murine AS model induced by aortic valve wire injury. Results: In myeloid cells, LYVE1 resident macrophages were significantly decreased, whereas classical monocytes expressing pro-inflammatory genes and C1Q macrophages were increased in AS valves compared to AR valves. These macrophages exhibited strong interactions with valve interstitial cells, particularly with chondrogenic subtypes expressing COMP and SPARC. Anti-TNFα treatment in the murine model significantly reduced aortic valve calcification without hematopoietic complications. Conclusions: This study is the first to integrate scRNA-seq (3’) and scRNA-seq (Flex) analyses of human aortic valves, revealing single cell-level characteristics of immune cells and their interactions with non-immune cells in calcific AS. These findings suggest that anti-inflammatory therapy could be a therapeutic target for calcific AS.

overall_design: scRNA-seq of surgically extracted human aortic valve tissues (aortic regurgitation vs. aortic stenosis), which includes scRNA-seq (3') for CD45 sorted immune cells and scRNA-seq (Flex) for non-sorted cells. Bulk RNA-seq was also conducted for human aortic valve tissues. AS model mice induced by aortic valve wire injury were treated either TNFa antibody or IgG control, then murine aortic valve tissues were evaluted using spatial transcriptomics (visium HD).

first: Tomoyo

last: Hamana


title: IGF2BP3 plays a critical role in the growth heterogeneity of colorectal adenoma cells by regulating MYC expression [RIP-seq]

city: Kyoto

first: Masahiro

last: Inoue

address: Shimoadachi-cho 46, Sakyou-ku

institute: Kyoto University, Graduate School of Medicine

summary: Background & Aims: Colorectal adenoma is a major precancerous lesion in colorectal cancer (CRC), and the adenoma-carcinoma sequence is a well-known multistep progression to CRC caused by the accumulation of genetic mutations. Accumulating evidence suggests that intra-tumor heterogeneity in CRC is influenced not only by genetic alterations, but also by non-genetic mechanisms. However, in adenomas, the non-genetic mechanisms remain largely unknown. Methods: Organoids were isolated from colorectal adenoma tissue. Single cells derived from patient-derived adenoma organoids were evaluated for their proliferative potential. The growth patterns of single-cell-derived spheroids were divided into two groups (D-/S-pattern) and the molecular mechanisms underlying the differences in growth patterns were examined. Results: Organoids were established from 31 colorectal adenomas from 19 patients. Some adenomas showed heterogeneity in the proliferative potential of single cells, as observed in CRC, and this heterogeneity was regulated by non-genetic mechanisms. IGF2BP3 was identified as a differentially expressed gene between organoids with different growth patterns and was found to be a key regulator of the high proliferative potential of adenomas. IGF2BP3 positively regulated MYC expression at the transcriptional level and negatively regulated it at the translational level. This promoted high proliferative potential with high levels of oxidative phosphorylation, while allowing cells to avoid MYC-induced cell death. IGF2BP3 affected the tumorigenicity of mouse adenomas in vivo. Conclusion: Intra-tumor heterogeneity in growth potential is acquired at the precancerous stage in the adenoma-carcinoma sequence of colorectal carcinogenesis, and IGF2BP3 plays an important role in regulating MYC levels. These findings provide new insights into the non-genetic regulation of adenomas during CRC development.

overall_design:
RIP-seq profiling of patient-derived adenoma organoids (D-pattern) to identify IGF2BP3-associated mRNAs

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

last: Sunami

first: Roberto

last: Coppo


title: IGF2BP3 plays a critical role in the growth heterogeneity of colorectal adenoma cells by regulating MYC expression [mouse RNA-seq]

overall_design: RNA-seq profiling of intestinal organoids derived from Apc-knockout mice with and without IGF2BP3 overexpression


title: Immune cell sequencing after allogeneic hematopoietic cell transplantation [scRNA-seq]

first: Cameron

middle: Scott

last: Bader

state: California

address: 269 Campus Drive, CCSR Room 2250

department: BMT and Cell Therapy

summary: Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a curative therapy for hematologic malignancies. The primary non-relapse complication after allo-HSCT is graft-versus-host disease (GVHD). The use of T regulatory (Treg) cells to prevent GVHD has emerged as a promising allogeneic T cell immunotherapy in the form of Orca-T. However, the precise differences in immune states which may influence clinical outcomes after Orca-T compared with unmanipulated peripheral blood stem cell (PBSC) grafts remain unexplored. Using peripheral blood specimens longitudinally collected between 3 weeks and one year after leukemia treatment, we examined single-cell mRNA sequencing (scRNA-seq) analysis of 16 HLA-matched patients receiving either Orca-T or unmanipulated PBSC grafts.

overall_design: Patient whole PBMC were collected 16-21 days after allogeneic hematopoetic cell transplantation, sorted into iNKT, CD4+CD25- Tcon, CD4+CD25+CD127low Treg, or CD19+ B cells, multiplexed, and captured for targeted transcriptomic and VDJ scRNA-seq using the BD Rhapsody platform and the Human Immune Response Panel.


title: scRNA-seq analysis of liver constituent cells from human FALD and normal livers

city: Suita

first: Seiya

last: Kato

state: Osaka

address: 2-2, Yamadaoka

department: Gastroenterology and Hepatology

institute: Osaka University

summary: scRNA-seq was performed on Fontan-associated liver disease (FALD) and normal livers using 10x Genomics Fixed RNA Profiling. The expression of connective tissue growth factor (CTGF) was upregulated in the EC cluster in the FALD livers. The scRNA-seq analysis also showed that the integrin signaling pathway was activated and the expression of YAP/TAZ target genes was upregulated in the EC cluster in the FALD livers.

overall_design: Frozen surgical liver tissue was cut into pieces approximately 1 mm3, fixed with formalin using a Chromium Next GEM Single-Cell Fixed RNA Sample Preparation Kit (10x Genomics), and then dispersed into single cells using gentleMACS (Miltenyi Biotec) according to the manufacturer’s instructions.

first: Hayato

last: Hikita

first: Tetsuo

last: Takehara


title: Immune cell sequencing after allogeneic hematopoietic cell transplantation

overall_design: Patient whole PBMC were collected 16-21 days after allogeneic hematopoetic cell transplantation and captured for whole transcriptome scRNA-seq.


title: RNA-seq analysis of liver sinusoidal endothelial cells exposed to hydrostatic pressure stimulation

summary: Primary murine LSECs (mLSECs) were exposed to hydrostatic pressure (HP) stimulation. The expression of YAP/TAZ target genes such as CTGF and basement membrane-related genes such as COL4A1was upregulated in mLSECs exposed to HP stimulation.

overall_design: After incubation on a Collagen I-coated plate overnight, the cells were serum-starved for 6 hours and then exposed to periodic HP stimulation for repeated cycles (0.01 Hz) between 101 and 135 kPa (absolute) for 8 hours using our original HPS system. The system was maintained at 37°C in a CO2 incubator (5% CO2).


title: Mixed molecular subtypes coexist in estrogen receptor heterogenous primary breast cancers

city: Boston

first: Fangyuan

last: Chen

state: MA

address: 450 Brookline Avenue

institute: Dana-Farber Cancer Institute

summary: Breast cancers show intratumor heterogeneity in estrogen receptor (ER) expression, but it is unclear whether cancer cells of different molecular subtypes (i.e. Luminal A, Luminal B, HER2-enriched, or Normal) can coexist in the same tumor. We performed spatial transcriptomic profiling of ER-negative, ER-low, and ER-high tumor cell regions of breast cancers that were 10-60% ER positive on routine immunohistochemistry using the GeoMxTM platform targeting 18,676 genes. We found that Luminal A and B molecular subtypes were intermixed in these tumors but there were no Basal-like cell populations. The ER-negative tumor regions were Luminal B-like and showed significantly lower expression of ESR1 and endocrine therapy sensitivity gene signatures but higher expression of inflammation and immune related genes, and also had higher recurrence scores indicating an endocrine-resistant but more chemotherapy sensitive phenotype. We also found that ESR1 strongly positive cells were enriched after preoperative chemotherapy in clinical trial tissues. These findings explain the worse clinical outcome of ER heterogenous breast cancers and suggest benefit from combined endocrine and chemotherapy strategies.

overall_design: We defined tumor cells regions with variable ER expression for spatial transcriptomic profiling using immunofluorescent (IF) ER and cytokeratin (CK) staining of a tissue microarray (TMA) containing 30 breast cancers. Each ROI corresponds to a case, and each ROI is segmented into 4 distinct AOIs, or tumor regions, by combining the CK and ER IF filters. The AOIs included stroma and 3 distinct epithelial compartments with ER-High, ER-Low, or ER- negative (ER-Neg) cancer cell populations within a cancer. Transcriptional profiling was performed separately for each AOI using the GeoMxTM technology and the GeoMx Human Whole Transcriptome Atlas probe set that targets 18,676 genes.

first: Julia

last: Foldi

first: Kaitlyn

last: Xiong

first: Matthew

last: Liu

first: Charles

last: Robbins

first: Haiying

last: Zhan

first: Sneha

last: Burela

first: Jiawei

last: Dai

first: Philipp

last: Karn

first: Matteo

last: Dugo

first: Giampaolo

last: Bianchini

first: Adrian

last: Lee

first: Steffi

last: Oesterreich

first: David

middle: L

last: Rimm

first: Lajos

last: Pusztai


title: Identification of the zinc finger transcription factor SlZFP2 as a new regulator of locular tissue morphogenesis

city: Villenave d'Ornon

first: martine

last: lemaire-chamley

address: 71 Avenue Edouard Bourlaux

department: FDFE

country: France

institute: INRAE

summary: In tomato (Solanum lycopersicum L.) fruit, locular tissue (LT) is a unique jelly-like structure that differentiates from the central axis of the fruit following ovule fertilization. LT is essential for seed development and dispersal, preventing early germination and initiating fruit ripening. In this work, we studied a “gel-less” mutant and identified the underlying mutation in the coding sequence of the C2H2 zinc finger transcription factor (TF) SlZFP2. Histological, cytological and molecular characterization of zfp2-CRISPR/Cas9 knockout lines revealed an early and significant impact of zfp2 disruption on cell division and endoreduplication in LT. Further, model-based analysis of cellular data revealed that the cell division was the main altered process explaining the zfp2 mutant phenotype. RNA-seq analysis of young LT further highlighted global transcriptional changes of cell cycle-related genes between WT and zfp2 mutants, providing new insights into the molecular players involved in locular tissue differentiation. This multi-faceted approach uncovers SlZFP2 as a novel regulator of LT morphogenesis and reveals its main action in regulating cell division. While LT developmental processes are poorly investigated and only rare regulators have been described yet, this work on SlZFP2 establishes groundwork for future studies aimed at deciphering the complex regulatory networks governing fruit tissue development.

overall_design: To better understand the early changes in the morphogenesis program of LT cells in zfp2-c lines, we performed a RNA-seq analysis on LT dissected from 6 DPA fruits of SlZFP2 crispr lines (c2.5, c2.11) and WT. Three biological replicates were performed for each genotype

first: Gabriel

last: Hoang

first: Joana

last: Jorly

first: Dario

last: Constantinescu

first: Pascal GP

last: Martin

first: Stéphanie

last: Gadin

first: Jean-Philippe

last: Mauxion

first: Cécile

last: Brès

first: Virginie

last: Garcia

first: Nathalie

last: Gonzalez

first: Christophe

last: Rothan

first: Nadia

last: Bertin

first: Lucie

last: Fernandez-Lochu

first: Martine

last: Lemaire-Chamley


title: ZBTB7B Inhibits Glioma Tumorigenicity by Upregulating GPR17 and CXCL10

city: DaLian

first: YiDing

last: Liu

address: wwde

institute: Dalian Medical University

summary: Background: We utilized a CRISPR-Cas9 screening library to examine the functions of tumor-related genes. Among these, we identified the transcription factor ZBTB7B, which overexpressed in breast cancer cells and significantly inhibited breast cancer growth in the brains of nude mice. Based on this finding, we hypothesized that ZBTB7B might exert similar effects on glioma, a possibility not yet explored in the literature. This study aims to evaluate the effects and mechanisms of ZBTB7B in glioma progression. Methods:The U118 and the GL261 cell line , both overexpressing ZBTB7B, were generated via lentiviral transduction. The impact of ZBTB7B on glioma growth was assessed using in vitro cell culture and in vivo intracranial inoculation. Downstream targets regulated by ZBTB7B were identified through transcriptome sequencing, PCR arrays, ELLA assays, and Western blotting. Cell apoptosis and the cell cycle were analyzed via flow cytometry. Multicolor immunofluorescence staining was performed to evaluate the expression and localization of ZBTB7B in human and mouse glioma tissues. Findings:Overexpression of ZBTB7B significantly inhibited glioma cell survival both in vitro and in vivo. Mechanistically, ZBTB7B upregulated the expression of GPR17, which suppressed PKA phosphorylation, promoted the generation of reactive oxygen species, and ultimately induced apoptosis. Additionally, ZBTB7B regulated the chemokine CXCL10, facilitating the recruitment of CD4+ and CD8+ T cells to the tumor periphery. Human glioma tissue microarray analysis revealed ZBTB7B expression was significantly reduced or absent in grade II to IV glioma samples. Interpretation:ZBTB7B exhibits significant tumor-suppressive effects and holds promise as a potential molecular target for glioma therapy.

overall_design: We performed RNA-sequencing on a total of 12 samples, with 6 samples overexpression ZBTB7B in U118 cells and GL261 cells, and the other 6 serving as vector groups.

first: Yan-Ke

last: Chen

first: Lin-Mei

last: Zhang


title: Macrophage-Mediated IL1beta/IL23 Signaling Regulates gamma delta (gammadelta) T Cell Immunity Against Non-Tuberculous Mycobacteria [scRNA-seq]

city: Shanghai

first: Xiaoqian

last: Hu

address: No. 130, Dongan Road, Xuhui District, Shanghai

institute: Fudan University

summary: Mechanistic Insights. Our study reveals the crucial role of gammadelta T cells in non-tuberculous mycobacteria (NTM) infection. We observed a significant increase and activation of gammadelta T cells in mice infected with MAB or with MAB infection combined with pulmonary fibrosis. Depletion of gammadelta T cells worsened the infection, while transfer of gammadelta T cells reversed this effect. Mechanistically, we found that MAB infection stimulates macrophages to produce IL-1beta and IL-23, which promotes the expansion of gammadelta T17 cells. MAB can also directly activate gammadelta T cells, leading to the clearance of MAB through an IL-17A-dependent pathway. Our findings suggest that gammadelta T cells represent a potential therapeutic target for NTM infections.

overall_design: For mouse single-cell sequencing, the lung of infected and uninfected mice were harvested and converted into single-cell suspension.


title: Transcription factor binding and individual genetic risk for valproate teratogenicity

city: Melbourne

first: Alison

middle: May

last: Anderson

state: VIC

address: Commercial Road

department: Neuroscience

country: Australia

institute: Monash University

summary: Valproate (VPA) use during pregnancy is associated with a wide range of birth defects and adverse neurodevelopmental outcomes, but not all exposed children are affected and there is evidence for a genetic predisposition. We hypothesised that genomic variants that impact on the binding affinity of transcription factors (TFs) are integral to VPA-associated teratogenicity and a plausible explanation for variance in interindividual risk. We interrogated maternal exomes from women recruited through international epilepsy genomics consortia. The variant burden within genes associated with 32 different birth defect types was higher for those exposed to VPA as compared to other antiseizure medications (OR 1·73 [95% CI 1·40 to 2·14], p = 2·25E-07). Variants in women exposed to VPA were predicted to impact the binding affinity of 359 TFs and network analysis of encoded proteins indicated that a master regulator, EP300, interacts with 42% (151/359) of all variant sensitive TFs. We then profiled coexpression between EP300 and other TFs in differentiating neurons derived from human embryonic stem cells (hESCs) exposed to VPA at 300µM and 700 µM, or unexposed, and a reference map generated using public data. We found strong overlap in EP300-TF coexpressed pairs between the reference and all comparison groups (99%,900/911) but only 32% (134/422) of pairs observed in unexposed cells were evident following VPA exposure, and over half of all pairs (489/911) were observed in VPA-exposed cells only. Our findings suggest that VPA-induced disruption of EP300-related regulation is common across birth defect types and that genetic variation can modify subsequent transcriptional dysregulation, explaining why only some pregnancies are affected. The results have implications for the development of genetic risk biomarkers and safer drugs.

overall_design: RNA-seq profiling from human embryonic stem cells (hESC) exposed to valproate (VPA) at 300uM and 700uM or unexposed

first: Piero

last: Perucca

first: Elena

last: Vianca

first: Daniel

last: Sandvik

first: Ana

last: Antonic-Baker

first: Roland

last: Krause

first: Dana

last: Jazayeri

last: Hitchcock

first: Janet

last: Graham

first: Marian

last: Todaro

first: Torbjörn

last: Tomson

first: Dina

last: Battino

first: Emilio

first: Meritxell

last: Martinez Ferri

first: Anne

last: Rochtus

first: Lieven

last: Lagae

first: Maria

last: Canevini

last: Zambrelli

first: Ellen

last: Campbell

first: Aleksei

last: Rakitin

first: Bobby

last: Koeleman

first: Ingrid

last: Scheffer

first: Samuel

middle: F

last: Berkovic

first: Patrick

last: Kwan

first: Sanjay

last: Sisodiya

first: John

last: Craig

first: Frank

last: Vajda

first: Terence

last: O’Brien


title: Transcriptomic Analysis of WT and IGF2BP2 KO Macrophages Reveals Metabolic Alterations

city: Saarbruecken

first: Jessica

last: Hoppstaedter

address: Campus C2 3

department: Pharmacy

institute: Saarland University

summary:
This dataset contains RNA-Seq data from wild-type (WT) and IGF2BP2 knockout (KO) bone marrow-derived macrophages (BMMs) under basal and IL-4-stimulated conditions. Loss of IGF2BP2 led to significant transcriptional changes, particularly in genes regulating metabolism, mitochondrial function, and cell migration. Pathway analysis revealed a metabolic shift toward glycolysis and reduced migratory capacity.
Details: Int J Biol Sci 2026; 22(6):2754-2773. doi:10.7150/ijbs.122142

overall_design: Myeloid-specific IGF2BP2 KO mice were generated by crossing B6.Cg-Igf2bp2tm1.1Thor/J mice (The Jackson Laboratory), which carry loxP sites flanking exons 4 and 5 of the Igf2bp2 gene, with 29P2-Lyz2tm1(cre)Ifo/J mice (The Jackson Laboratory). Control wild-type (WT) mice were littermates that did not carry floxed alleles and either expressed or lacked Cre recombinase. Bone marrow-derived macrophages (BMMs) were isolated from WT and IGF2BP2 KO mice and differentiated with M-CSF for six days. Cells were either left untreated (M0) or stimulated with IL-4 (20 ng/mL) for 8 hours to induce M2 polarization. RNA was extracted, and libraries were prepared using poly(A)-enriched mRNA. Sequencing was performed on an Illumina NextSeq 500 platform.

first: Hanna

last: Schymik

first: Gilles

last: Gasparoni

first: Jörn

last: Walter

last: Hoppstädter

first: Alexandra

middle: K

last: Kiemer


title: Femtosecond laser microgrooves combined with CGF promote implant transgingival integration through Smad3/JunD/TIMP-1 pathway and its immune regulatory mechanism

city: 济南

first: 之涵

last: 崔

address: 山东省济南市文化西路四十四号山东大学趵突泉校区

institute: 山东大学

summary: In implant repair, early soft tissue healing can form a stable barrier to prevent implant loosening failure caused by bacterial invasion. The adhesion between the epithelium surrounding the implant and the surface of the titanium implant is achieved through hemidesmosomes. The connective tissue interface is composed of fibroblasts and collagen fibers with poor vascular structure and rich scar tissue like structure, which are parallel to the surface of titanium implants. In natural teeth, the collagen fibers on the surface of the dentin are vertical. The surface structure and coverage of biological factors at the neck ring of the implant are important factors affecting the soft tissue adhesion around the implant. The microgroove structure can alter the fiber pathway at the junction of the abutment and soft tissue, and regulate the phenotypic transformation of macrophages; Concentrated growth factor (CGF) derived from blood can promote the healing of gingival soft tissue after surgery and regulate immune response. However, the combined effect of microgrooves and CGF has not been studied yet. We use femtosecond laser to etch micro grooves on the neck ring of the implant, and infiltrate CGF into the neck ring of the implant micro groove through immersion, so that the micro groove structure works in combination with CGF. The implant is implanted into the alveolar bone of beagle dogs, and the color, shape, texture, and gingival margin position of the gingiva are checked for relevant clinical data measurement. Then, the implant is harvested together with the surrounding soft and hard tissues to preserve the ability of soft tissue attachment and recognition of biological width, for histological preparation and analysis. This study delves into the phenomenon of promoting implant transgingival integration, anchoring collagen fibers vertically, and making connective tissue adhesion more stable and firm, as well as its immune regulatory mechanism. This study constructed an implant with a micro groove neck ring and concentrated growth factor (CGF) derived from blood, clarifying the relationship between the binding of the micro groove structure in the neck of the implant and the sealing effect of the connective tissue around the implant. It revealed that the micro groove structure and CGF can promote M2 polarization of macrophages in the gingival connective tissue, and clarified that the morphology of the micro groove in the neck of the implant combined with CGF can promote the phenomenon of implant transgingival binding.

overall_design: Comparative gene expression profiling analysis of RNA seq data of gingival tissue around different types of implants, including smooth neck ring implants, micro groove neck ring implants, and micro groove neck ring combined with CGF implants.

first: Huiqiang

last: Sun

first: Zhihan

last: Cui

first: Yingbei

last: Yang

first: Xiaoxue

last: Wang

first: Yuting

last: Pei

first: Lili

last: Zhang


title: Transcriptomic Responses to Pharmaceutical-Laden Wastewater Effluent Exposure in Etheostoma spp. from the Grand River

first: Louis

last: Pfeifer

summary: Aquatic organisms, such as fish, are in constant contact with various anthropogenic stressors present in their environments, including contaminants from wastewater treatment plant effluents (WWTPE). WWTPE represents a pervasive source of pharmaceutically active compounds  in surface waters  This study investigates the effects of WWTPE exposure on Etheostoma spp. (darters) using hepatic transcriptomic profiling. Wild darters—Etheostoma caeruleum (rainbow darter), Etheostoma blennioides (greenside darter), Etheostoma flabellare (fantail darter), and Etheostoma nigrum (Johnny darter)—were collected upstream and downstream of the Waterloo municipal WWTP effluent outfall (Grand River, ON).

sample_organism:
Etheostoma caeruleum
Etheostoma flabellare
Etheostoma nigrum
Etheostoma blennioides

overall_design: mRNA-seq profiling of liver tissue from populations of rainbow, fantail, johnny, and greenside darters collected upstream and downstream of the Waterloo Municipal Wastewater Treatment Plant effluent outfall (Grand River, ON, Canada).

first: Paul


title: Effects of circAASS overexpression in human HK2 cells

city: guangzhou

first: peng

last: wang

address: 1838 North Guangzhou Avenue

institute: Southern Medical University

summary: The mechanism underlying circRNA-mediated maintenance of mitochondrial function remains largely unknown. Our study found circAASS was downregulated in renal cortex from mice suffered ischemic reperfusion (IR)-induced acute kidney injury (AKI). Functional study demonstrated that circAASS regulated mitochondrial homeostasis in tubular epithelial cells (TECs). Therefore, to reveal the mechanisms underlying the anti-injury effects of circAASS in TECs. circAASS was overexpressed in hypoxia/reperfusion treated- human immortalized cell line HK2 cells. RNA sequencing was performed and the dysregulated mRNAs were found.

overall_design: RNA-seq profiling of HK2 cells transfected with circAASS-overexpressing plasmids or empty vector (control plasmids) at 24 hours post hypoxia/reperfusion treatment.

first: Peng

first: Tongtong

last: Ma


title: Impact of Mek1 and Mek2 deletion on the transcriptome of spleen B cell subpopulations

city: Québec

first: Jean

last: Charron

state: QC

address: 2260, boul. Henri-Bourassa

department: Biologie moléculaire, Biochimie médicale & Pathologie

institute: CRCHU de Québec-Université Laval

summary: MEK1 and MEK2, the only known activators of ERK, are attractive therapeutic candidates for autoimmune diseases. The deletion of Mek1 in mouse hematopoietic cells was performed using the Mek1 conditional allele (Map2k1tm1Chrn) and the Vav1-icre deleter mouse line (Commd10Tg(Vav1-icre)A2Kio) in the Mek2 heterozygous null background (Map2k2tm1Chrn). Mek1flox/flox Mek2+/- Tg+/Vav1-icre mice (named thereafter 1Mek2) die prematurely of lymphoproliferative disorders associated with spontaneous B cell activation and the accumulation of plasma cells, age-associated B cells and germinal center B cells (Houde, N., et al. (2022). Cell Rep 38(2): 110223). However, how MEK signaling finely regulates immune cell activation is only partially understood. To address this question, we applied bulk RNA-seq assays to characterize the transcriptional impact of the specific loss of MEK protein in B cell subpopulations using the 1Mek2 mutants.

overall_design: To investigate the global transcriptional impact resulting from the loss of Mek1 and Mek2 gene function in B cells, we applied bulk RNA sequencing (RNA-seq) on splenic B cell subpopulations. We extracted splenocytes from 3 to 5 months wild-type and Mek1flox/flox Mek2+/- Tg+/Vav1-icre mice (Houde, N., et al. (2022). Cell Rep 38(2): 110223.). Following splenocyte isolation, B cell subpopulations were purified by cell sorting: plasma cell (P; B220+CD138+), immature (Immature; B220+CD93+CD138-), germinal center (GC; B220+CD93-CD138-GL7+FAS+), follicular (F; B220+CD93-CD138-GL7-FAS-CD21+CD23+), marginal zone B cell (M; B220+CD93-CD138-GL7-FAS-CD21+CD23-).

first: Nicolas

last: Houde


title: scRNA-seq analysis of liver nonparenchymal cells from the mice after pIVCL and sham surgery

first: Seiya

last: Kato

summary: We performed scRNA-seq analysis on murin livers after partial inferior vena cava ligation (pIVCL), a mouse model of congestive hepatopathy. We identified connective tissue growth factor (CTGF) as one of the most upregulated genes in pericentral (zones 2 and 3) LSECs. The scRNA-seq analysis also showed that the integrin signaling pathway was activated and the expression of YAP/TAZ target genes was upregulated in pericentral LSECs.

overall_design: After liver perfusion with pronase and collagenase, hepatocytes were removed via low-speed centrifugation, and dead cells and erythrocytes were removed via density gradient centrifugation.

first: Hayato

last: Hikita

first: Tetsuo

last: Takehara


title: Mechanism of Corynebacterium striatum-mediated inhibition of Staphylococcus aureus-induced epithelial barrier injury and inflammation activation in chronic rhinosinusitis [RNA-Seq]

city: 广州

first: Zhou

last: Jiewen

address: 广东省广州市越秀区中山大学北校区

institute: 中山大学附属第一医院

summary: Microbes are an important trigger of chronic rhinosinusitis (CRS). Staphylococcus aureus is a CRS-related pathogen that disrupts the epithelial barrier and activates mucosal inflammation. Given the ineffectiveness of antimicrobial therapy in CRS, we explored a potential treatment based on the interaction of bacteria within the dysbiotic microenvironment of the nasal mucosa. Microbiome analyses of clinical isolates from CRS patients and healthy controls were employed to identify Corynebacterium striatum as a potential inhibitor of pathogenic S. aureus. A co-infection culture model using human nasal epithelial cells (hNECs) was constructed to investigate the inhibition of S. aureus-induced disruption of the host epithelium by C. striatum. Further analysis of the inhibition of S. aureus by C. striatum was carried out using a bacteria co-culture model.

overall_design: The bacterial cultures used for infection experiments were prepared when hNECs were differentiated and matured. Briefly, cryopreserved bacterial strains were recovered by streaking on LB agar plates. A single colony was then picked and inoculated into LB liquid medium for overnight shake cultivation (37℃, 220 rpm). After centrifugation and two washes with sterile phosphate-buffered saline (PBS), the bacterial suspension was quantified to an OD600 of 0.4 and applied onto the hNECs at a volume of 10 µL per insert, which precisely covered the entire surface of the hNEC layer. Both bacterial strains were treated in the same manner. To simulate symbiosis in the human nasal cavity, C. striatum was pre-inoculated and allowed to colonize the hNECs for 48 hours before infection with S. aureus for another 12 hours. The cells and media were then collected for experimental analysis.

first: Nianzhen

last: Zheng

first: Min

last: Li

first: Jiewen

last: Zhou


title: An Interleukin 9-ZBTB18 axis promotes germinal center development of memory B cells [ATAC-seq]

city: Beijing

first: xiaocui

last: luo

address: Haidian District

institute: Tsinghua university

summary: Memory B cell (MBC) development from germinal centers (GCs) entail profound changes in cell cycling, localization and survival. Here we examined the mechanisms that induce the memory program, focusing on interleukin (IL)-9, given its importance for normal recall antibody responses. Using adoptive transfer and radiation chimera models, we found that T cell-derived IL-9 was required for MBC development and function. In contrast, B cells deficient in IL-9 generated functionally normal MBCs that support antibody recall normally. IL-9 induced expression of the transcriptional repressor ZBTB18 in GC memory precursor cells and MBCs. ZBTB18 was dispensable for naïve B cell activation and GC formation but required for development of GC-derived MBCs. ZBTB18 directly repressed expression of a suite genes encoding cyclin and cyclin-dependent kinases, pro-apoptotic genes Bid and Casp3, and the GC-retaining factor S1pr2. Lack of IL-9-mediated instruction or intrinsic programming by ZBTB18 impaired GC-derived MBC development and antibody recall. Thus, an IL-9-ZBTB18 axis instructs development of functional B cell memory from GCs.

overall_design: Mice were immunized intraperitoneally with 100 μg NP-KLH (conjugate ratios20, Biosearch Technologies) mixed with 1 μg lipopolysaccharide (LPS, Sigma) in alum (Thermo Scientific). Single-cell suspension of splenocytes was stained in MACS buffer (PBS supplemented with 1% FBS and 5 mM EDTA) containing Fc blocker (supernatant from 2.4G2 cell culture) for 20 min on ice before surface staining with primary antibodies in appropriate concentrations. 5×104 naïve B cells or GCMP cells were sorted from Cd79acre/+ mice and Cd79acre/+Zbtb18fl/fl mice, respectively. Cells were collected by gentle centrifugation (500x g, 5 minutes) at 4 °C, and then resuspended in lysis buffer (10 mM Tris.Cl, 10 mM NaCl, 3 mM MgCl2, 0.1% (v/v) Igepal CA-630) and incubated on ice for 10 min. Nuclei were collected and resuspended into 50 μl tagmentation mix, which contains 10 μl 5×TTBL (Vazyme), 5 μl TTE Mix V50 and 35 μl ddH2O. Samples were incubated at 37 °C for 30 min. After tagmentation, fragmented DNA was isolated by using VAHTS DNA Clean Beads. ATAC libraries were amplified for 12 PCR cycles using TruePrepTM Index Kit V2 for Illumina (Vazyme #TD202). 0.55×VAHTSTM DNA Clean Beads were used to exclude the High–molecular weight DNA, followed by positive selection with 1×beads. High-quality DNA libraries were sequenced on the Illumina HiSeq2000.

first: Hai

last: Qi

first: Xiaocui

last: Luo

first: XiaoXiao

last: Hou

first: Yifeng

first: Ye

first: Shangcheng

last: Yu


title: Cancer-associated synonymous mutations selectively target the p53 DNA damage response [RNA-seq]

city: Umeå

first: Lixiao

last: Wang

state: Sverige

address: Medicinsk biovetenskap, M21, Försörjningsvägen 1

department: Department of Medical Biosciences

country: Sweden

institute: Umeå University

summary: Recent technical advances have facilitated studies on changes in mRNA structures in response to signaling pathways, but how mRNA structures relate to the function of the encoded protein remains poorly understood. Synonymous mutations (SMs) possess comparable oncogenic potential to missense mutations. Here we have taken advantage of two Cancer Associated SMs at codon 34 (CASM34) in the p53 mRNA to demonstrate the impact of the RNA structure on protein activity in response to DNA damage. In-cell RNA structural probing illustrates how CASM34s prevent DNA damage-induced p53 mRNA folding. Genome-wide transcriptomics show that CASM34 alters the expression of genes in the p53 DNA-damage response pathway. DNA ChIP-seq analysis reveals that p53 expressed from CASM34 has reduced promoter binding and reduced induction of p53 downstream target genes PUMA and 14-3-3-σ but not p21CDKN1A. These results demonstrate that CASM34s selectively target p53 activity towards downstream targets in the DNA damage response pathway by interfering with p53 mRNA folding.

overall_design: RNA-seq profiling of p53-WT and CASM34 mutant expressed in H1299 p53 null cells

first: Sivakumar

last: Vadivel Gnanasundram


title: Mechanism of Corynebacterium striatum-mediated inhibition of Staphylococcus aureus-induced epithelial barrier injury and inflammation activation in chronic rhinosinusitis

address: 广东省广州市越秀区中山大学北校区

sample_organism:
Staphylococcus aureus
Corynebacterium striatum

overall_design: To explore the competition between C. striatum and S. aureus, a bacterial co-culture model was established. Briefly, the two bacterial strains were recovered and amplified in LB liquid medium. The bacterial cells were then harvested, washed, and quantified to an OD600 of 1.0 in PBS. Next, each bacterial suspension was diluted 100-fold in LB liquid medium and incubated on a shaker at 37℃ and 220 rpm. Referring to the growth curves of the two strains and aiming to simulate the natural colonization state of C. striatum, it was cultured for 9 hours, while S. aureus was cultured for only 3 hours to simulate the invasion condition. Subsequently, equal volumes of the two bacterial cultures were fully mixed for 12 hours of shake co-cultivation, then collected for experimental analysis. As an experimental control, each bacterial culture was mixed with an equal volume of LB liquid medium (Figure 3B). For colonies counting, the cultures after co-cultivation were diluted and plated onto solid LB medium.


title: Essential role of METTL3 in mitochondrial function and epigenetic regulation during preimplantation development in mice

first: Shinnosuke

last: Honda

address: Kitashirakawa Oiwake-cho, Sakyo-ku, Kyoto-shi

institute: Kyoto University

summary: We obtained transcriptomic profiles in mouse morula stage embryos (72 h post-insemination (hpi)) which had been microinjected with non-targeting or Mettl3-targeting antisense oligonucleotides (ASOs) at the one-cell stage (3–6 hpi).

overall_design: In vitro produced mouse morula stage embryos were processed to RNA-Seq. The experiments were duplicated using ten embryos per group.

first: Dan

last: Liu

first: Xinyao

last: Chen

first: Yuwei

last: Gao

first: Haoxue

first: Naojiro

last: Minami

first: Shuntaro

last: Ikeda


title: IGF2BP3 plays a critical role in the growth heterogeneity of colorectal adenoma cells by regulating MYC expression

first: Masahiro

last: Inoue

summary:
Background & Aims: Colorectal adenoma is a major precancerous lesion in colorectal cancer (CRC), and the adenoma-carcinoma sequence is a well-known multistep progression to CRC caused by the accumulation of genetic mutations. Accumulating evidence suggests that intra-tumor heterogeneity in CRC is influenced not only by genetic alterations, but also by non-genetic mechanisms. However, in adenomas, the non-genetic mechanisms remain largely unknown.  Methods: Organoids were isolated from colorectal adenoma tissue. Single cells derived from patient-derived adenoma organoids were evaluated for their proliferative potential. The growth patterns of single-cell-derived spheroids were divided into two groups (D-/S-pattern) and the molecular mechanisms underlying the differences in growth patterns were examined.   Results: Organoids were established from 31 colorectal adenomas from 19 patients. Some adenomas showed heterogeneity in the proliferative potential of single cells, as observed in CRC, and this heterogeneity was regulated by non-genetic mechanisms. IGF2BP3 was identified as a differentially expressed gene between organoids with different growth patterns and was found to be a key regulator of the high proliferative potential of adenomas. IGF2BP3 positively regulated MYC expression at the transcriptional level and negatively regulated it at the translational level. This promoted high proliferative potential with high levels of oxidative phosphorylation, while allowing cells to avoid MYC-induced cell death. IGF2BP3 affected the tumorigenicity of mouse adenomas in vivo.   Conclusion: Intra-tumor heterogeneity in growth potential is acquired at the precancerous stage in the adenoma-carcinoma sequence of colorectal carcinogenesis, and IGF2BP3 plays an important role in regulating MYC levels. These findings provide new insights into the non-genetic regulation of adenomas during CRC development.
The gene expression profiles of S-pattern (CP1-5, CP3-5, and CP9-1) and D-pattern (CP1-1, CP3-2, and CP9-2) patient-derived adenoma organoid lines were compared using microarray analysis.

overall_design: Five hundred organoids for each patient-derived adenoma line were cultured for 6 days. The gene expression profiles of three S-pattern (CP1-5, CP3-5, and CP9-1) and three D-pattern (CP1-1, CP3-2, and CP9-2) patient-derived adenoma organoid lines were analyzed.

first: Tomohiko

last: Sunami

first: Roberto

last: Coppo