title: Recurring heat waves affect DNA methylation to reduce respirational losses, enhancing heat tolerance Arabidopsis thaliana

city: Antwerpen

first: Jesper

middle: Ruben

last: van Dijk

state: Antwerpen

address: Universiteitsplein 1, Building C

country: Belgium

institute: University of Antwerp

type: Methylation profiling by high throughput sequencing


title: Recurring heat waves affect DNA methylation to reduce respirational losses, enhancing heat tolerance Arabidopsis thaliana [RNA-seq]

status: Public on Jun 01 2025

department: Biology

type: Expression profiling by high throughput sequencing

summary: we investigated the effect of exposing 4 successive generations of Arabidopsis thaliana to severe heatwave conditions. We show that heat-primed offspring has increased survival rates and seed production after exposure to heat wave conditions. Bisulfite sequencing allowed us to link this to altered DNA methylation, including a global hypomethylation of the mitochondrial DNA. mRNA sequencing showed that priming affected genes related to the mitochondrial energy system and oxidative stress responses.

overall_design: Bisulfite sequencing and mRNA sequencing on plants exposed to heat stress in previous generations, as well as the controls

middle: R

first: Gaurav

last: Zinta

first: Hamada

last: AbdElgawad

first: B

last: Cuypers

first: Katrien

last: Sprangers

first: Marios

middle: N

last: Markakis

first: K

last: Laukens

first: Ivan

middle: A

last: Janssens

first: Han

last: Asard

first: Gerrit

middle: T

last: Beemster


title: Recurring heat waves affect DNA methylation to reduce respirational losses, enhancing heat tolerance Arabidopsis thaliana [Bisulfite-seq]


title: The long non-coding RNA, miRNA, and mRNA landscapes of cementoblasts during cementogenesis

city: Beijing

first: yiping

last: huang

address: 22 zhongguancun south street

department: School of stomatology

country: China

institute: Peking University

summary: To screen and analyze the differentially expressed lncRNAs, miRNAs, and mRNAs in cementogenic differentiation of cementoblasts.

overall_design: OCCM-30 cells were induced in cementogenic medium for 0, 7, and 14 days. Total RNA was extracted and submitted to RNA sequencing to screen the differentially expressed lncRNAs, miRNAs, and mRNAs.

first: yingying

last: zhang


title: Dissection of the T-cell infiltrate in mouse pancreatic tumors reveals an extensive and diverse tumor-reactive T-cell repertoire

city: Heidelberg

first: Rienk

last: Offringa

state: Baden-Wuerttemberg

address: Im Neuenheimer Feld 280

department: Division of Molecular Oncology of Gastrointestinal Tumors

country: Germany

institute: DKFZ Heidelberg

summary:
Background: Pancreatic ductal adenocarcinoma (PDAC) is refractory to cytostatic and immunotherapeutic treatment regimens, including immune checkpoint inhibition (ICI). The latter is due to the limited mutational load of most of these tumors, in combination with the immunosuppressive hurdles imposed by the tumor microenvironment. Here we present an in-depth analysis of the T-cell response at the single-cell level in an orthotopic PDAC mouse model that recapitulates the therapy resistance of the human disease. In parallel, we investigated an ICI-sensitive variant of this model expressing an immunogenic neo-antigen.

Methods: Single-cell RNA and T-cell receptor (TCR) from tumor-infiltrating lymphocytes (TILs) of 14 tumor-bearing mice were combined with functional in vitro screening of TCR specificity against tumor. Transcriptomic differences between tumor-reactive (TR) and non-tumor-reactive (NTR) T-cells were harnessed to define gene signatures for the purpose of predicting the specificity of additional TCRs.

Results: Our efforts revealed extensive tumor-reactive and bystander CD8+ and CD4+ T-cell repertoires, not only in the immunogenic but also in the ICI-refractory tumors. By integrating the functional screening and single-cell transcriptome data sets, we delineated gene signatures that distinguish between tumor-reactive and bystander T-cells in the mouse model as well as in human cancers. The TCRs in the murine models detect a range of tumor-associated antigens beyond mutanome-encoded neoepitopes and retain functional anti-tumor activity in vitro.

overall_design: Transplantable tumor cell lines PDA30364 and PDA-OVA, which were generated from a genetically engineered mouse model on Elas-tTA/TetO-Cre Kras+/LSL-G12D Trp53+/LSL-R172H mouse line background, were implanted orthotopically into pancreas of immune-competent C57BL/6 mice. Mice were subjected to control treatment or treatment with MEK inhibitor GDC-0623 in combination with agonist anti-CD40 antibody treatment. After two cycles of treatment, tumors were harvested from mice, dissociated into single-cells for the sorting of tumor-infiltrating T-cells. T-cells were analyzed for single-cell RNA and VDJ expression using 10xGenomics for discrimination of tumor-reactive T-cells from bystander T-cells.

first: Hannes

last: Kehm


title: FAM3A plays a key role in protecting against tubular cell pyroptosis and acute kidney injury through PI3K/AKT/NRF2-dependent pathway

city: Guangzhou

first: Xiaolong

last: Li

address: 1838 North Guangzhou Ave

department: Nanfang Hospital

institute: Southern Medical University

summary: Acute kidney injury (AKI) is in a high prevalence worldwide, but there is no therapeutic strategies nowadays. Renal tubular cell death plays a key role in the initiation and progression of AKI. However, the underlying mechanisms have not been elucidated. In this study, we found that tubular cell pyroptosis was primarily responsible to AKI progression, which is highly related with ATP depletion. FAM3A is a mitochondrial protein responsible for ATP synthesis. Herein, we found that FAM3A was decreased in AKI mice, especially in those pyroptotic tubular cells. FAM3A also decreased in clinical patients with AKI, and negatively correlated with inflammation and tubular cell injury. Gene ablation of FAM3A strongly induced NLRP3 inflammasome, Caspase-1, GSDMD, IL-1β, and IL-18, the pyroptotic markers in tubular epithelial cells, as well as triggered inflammation and mitochondrial dysfunction. Conversely, overexpression of FAM3A greatly ameliorated pyroptosis and inflammation, and reversed fatty-acid oxidation function in tubular cells. Mechanically, FAM3A activated PI3K/AKT/NRF2 pathway, which improved mitochondrial dysfunction and prevented cell pyroptosis. To activate NRF2 could strongly inhibit tubular cell pyroptosis, inflammation, mitochondrial dysfunction, and renal function decline in FAM3A knockout mice. Hence, our study provides the new mechanisms for AKI progression. We also demonstrated that FAM3A is a potential therapeutic target for treating AKI.

overall_design: To verify the potential mechanism by which FAM3A protects AKI, we established IRI models and performed RNA-seq in WT and FAM3ACKO mice. We compared the gene expression differences between the two groups and performed GO and GSEA enrichment analysis

first: Feifei

last: Yuan


title: Genome-Wide CRISPR-Cas9 screening identifies BRAF inhibitor as a sensitizer of dasatinib in treating T-acute lymphoblastic leukemia

city: guangzhou

first: Chen

last: Jun-Zhu

state: State...

address: 广州市越秀区中山二路58号中山大学附属第一医院

institute: Sun Yat-Sen University

summary: T-cell acute lymphoblastic leukemia is an aggressive hematologic tumor with fewer treatment strategies. Dasatinib, an FDA-approved tyrosine kinase inhibitor applied in chronic myelogenous leukemia and acute lymphoblastic leukemia with Philadelphia chromosome-positive has been utilized in T-ALL for cure but the clinical outcomes are not satisfied, calling for further investigation on the mechanism and potential combinations to overcome resistance. In this study, we performed a genome-wide CRISPR-Cas9 screening and identified the MAPK pathway as an essential regulator of sensitivity to dasatinib. Then we confirmed that the inhibition of the MAPK pathway by dabrafenib could sensitize T-ALL cells to dasatinib in vitro and reveal the underlying mechanism by RNA sequencing. Together, we put forward a promising combining strategy for T-ALL.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for Jurkat cells with different treatments (vehicle, dasatinib, and the combination of dasatinib and dabrafenib).

first: Jun-Zhu

last: Chen

first: Xue-Qun

last: Luo


title: DNMT1-repressed NSun2 Expression Wires DNA Cytosine Methylation and mRNA Cytosine Methylation to Modulate Osteosarcoma Apoptosis

first: liu

last: cihang

address: Xueyuan Road

institute: Peking University Health Science Center

summary:
Background: Cellular apoptosis is a central mechanism the chemotherapy leveraged to treat human cancers. Cytosine methylations installed on both DNA and mRNA are documented to regulate apoptosis independently. However, the interplay or crosstalk between them in cellular apoptosis has not yet been explored.
Methods: Cisplatin and doxorubicin were used to induce cell apoptosis in three differed types of osteosarcoma cell lines, respectively. Bisulfite-PCR, MS-PCR, and MeDIP experiments were employed to evaluate promoter cytosine methylation, and MeRIP were performed to investigate mRNA cytosine methylation. MeRIP-seq revealed the common target genes regulated by DNMT1 and NSun2. Particular chimeric reporter constructs were generated and dual luciferase assays dissected corresponding regulations via DNA and/or mRNA methylation. Cell viability assay, apoptosis analysis by flow cytometry, and tumor xenograft model were utilized to determine osteosarcoma drug resistance.
Results: Promoter methylation by DNMT1 dialogues with mRNA methylation by NSun2 to coordinately regulate osteosarcoma cell apoptosis. DNMT1 was induced in osteosarcoma cell apoptosis triggered by chemotherapeutic drugs, whereas NSun2 expression was dampened. DNMT1 was found to represses NSun2 expression by methylating NSun2 promoter. Moreover, DNMT1 and NSun2 regulate anti-apoptotic genes AXL, NOTCH2, and YAP1 through DNA and mRNA methylation, respectively. Upon exposure to cisplatin or doxorubicin, DNMT1 elevation drastically reduced these anti-apoptotic gene expressions via enhanced promoter methylation coupled with NSun2 ablation-mediated attenuation of mRNA methylation, thus rendering osteosarcoma to apoptosis.
Conclusions: Our findings establish a crosstalk of importance between DNA and RNA cytosine methylations in determining osteosarcoma resistance to apoptosis during chemotherapy, shedding new light on future treatment of osteosarcoma, and add additional layers to the control of gene expression at different epigenetic levels.

overall_design: Two input RNA from U6 and T1, and 2v2 co-immunoprecipitated RNA from those samples respectively

first: Dongxing

last: Shao

first: Cihang

last: Liu


title: Perturb-Seq in pluripotent stem cells

city: San Francisco

first: David

last: Wu

state: California

address: 35 Medical Center Way

country: USA

institute: UCSF

summary: CRISPRi Perturb-Seq of coding genes and lncRNAs in pluripotent stem cells

overall_design: Perturb-Seq

first: Daniel

last: Lim


title: Transcriptional profiles of HSCs from control and ABX-treated mice in steady-state and regenerative conditions

city: Bronx

first: Dachuan

last: Zhang

state: NY

address: 1301 Morris Park Avenue, Price Center 113

department: Cell Biology

institute: Albert Einstein College of Medicine

summary: Hematopoietic stem cells (HSCs) are largely quiescent and metabolically inactive under steady state, but can be activated to rapidly proliferate and produce progeny in response to inflammation, hematopoietic stress and in diseases. The microbiota has recently emerged as an important regulator of hematopoiesis, with significant contributions to the production of multipotent progenitors (MPPs) and myeloid cells. However, whether the microbiota directly regulates HSC functions remain unclear. Here using antibiotics (ABX)-induced microbiota depletion model and RNA sequencing, we showed that microbiota depletion induced dramatic differences in HSC transcriptome between control and ABX-treated mice under steady state. Differences in HSC transcriptome persisted under stress condition, despite fewer pathways were clustered after 5-FU treatment.

overall_design: We analyzed the transcriptional profiles of HSCs from control and ABX-treated mice with or without 5-FU treatment by RNA sequencing.

first: Qiaozhi

last: Wei

first: Paul

middle: S

last: Frenette


title: Cage-rearing-induced duck transcriptome regulation study by RNA-seq and differential expression analysis

city: Nanchang

first: Biao

address: Zhimin street, Jingkai district

institute: Jiangxi Agricultural University

summary: In order to investigate the effect of cage-rearing on waterfowl, we collected the hypothalamus and pituitary tissues of 113-days-old ducks after caged for 3 days, and same tissues of ducks in same batch raised in water-floor system. The transcriptomes were sequenced and the expression level of genes were compared. Sequencing analysis showed that a total 506 and 342 genes were differentially expressed in the hypothalamus comparison and pituitary comparison, respectively.

overall_design: Examination of gene expression profiles  in hypothalamus and pituitary tissues of ducks from cage-rearing system and water-floor combination system.

first: Sanfeng


title: LRRFIP2 modulates the response to hypoxia during embryonic cardiogenesis

city: Paris

first: Franck

last: Letourneur

address: 22 rue mechain

country: France

institute: INSERM

summary: We identified LRRFIP2 as a novel negative cofactor of HIF-1 Indeed, we showed that the absence of Lrrfip2 expression in a mouse KI model leads to an increase in many HIF-1 target genes including AldoC, Bnip3 and Ndufa4l2 in embryonic CM during development. Overactivation of HIF-1 in Lrrfip2-null mice led to structural and functional mitochondrial dysfunction, decreased ROS levels and ultimately early CM maturation.

overall_design: Transcriptomic Affimetrix analysis performed on embryonic hearts

first: Laura

middle: B

last: DRISS

first: Christophe

last: HOUBRON

first: Philippe

last: DAUBAS

first: Pascal

last: MAIRE


title: RNA-Seq Analysis of p53 Knockdown validated Human SU. 86.86  Pancreatic Cancer cells

city: Birmingham

first: Narendra

last: Wajapeyee

state: AL

address: 720 20th Street South, Kaul 540A

department: Department of Biochemistry and Molecular Genetics

institute: University of Alabama at Birmingham

summary: The purpose of this study is to understand the differential gene expression analysis in p53 knockdown human SU.86.86 pancreatic cancer cells

overall_design: Total RNA was extracted from all cells, followed by poly-A selection and RNA-sequencing using Illumina NextSeq 500 platform, with three biological replicates for each transformed sample.

first: Raj

last: Kumar


title: YAP1-TFE3 mediates endothelial to mesenchymal transition in epithelioid hemangioendothelioma

status: Public on May 31 2025

city: West Lafayette

first: Majid

last: Kazemian

state: IN

address: 201 S. University Steet

department: Biochemistry and Computer Science

institute: Purdue University

summary: Epithelioid Hemangioendothelioma (EHE) is an ultra-rare vascular sarcoma with a high metastatic rate and a lack of reliable prognostic markers. Because of its rarity, EHE is an understudied disease with limited research tools and few effective treatment options. A thorough understanding of the molecular underpinnings that drive EHE tumorigenesis and facilitate metastasis is essential to discovery of druggable targets that could improve patient outcomes. To better understand this EHE biology we generated transgenic endothelial EHE cell models and observed robust transformation of YAP1-TFE3 (YT) expressing cells resulting in a pro-migratory, anti-apoptotic phenotype. We identified endothelial to mesenchymal transition (EndMT) as a contributing process to this transformation as evidenced by the loss of enothelial gene expression and strong increases in EndMT biomarkers Acta2 and Tgm2. To understand the underlying drivers of this process, we utilized mutant versions of YT and pharmacologic inhibitors to determine that the DNA-binding and dimerization bHLH-LZ domain of YT was necessary for EndMT to occur while canonical TEAD activation was dispensable. This project sheds light on molecular mechanisms of EHE which directly contribute to endothelial transformation.

overall_design: To characterize the transcriptional landscape of EHE, transgenic MS1 murine endothelial cell lines were generated to express either YAP1-TFE3 or an RFP control under a doxycycline inducible promoter. RNA was extracted 3 days post-doxycycline induction (1µg/mL). RNA-seq was performed to identify differential gene expression in response to YAP1-TFE3 expression.

first: Jason

last: Hanna

first: Ant

last: Murphy


title: Regulation of PRDM1 (PRDBF-1/BLIMP1) isoform expression in T cells by an Inflammatory Bowel Disease -associated polymorphism

city: West Hollywood

first: AGCT

last: Core

state: CA

address: 8687 Melrose Ave, PDC B230

department: BMS

institute: Cedars Sinai Medical Center

summary: This study aimed to determine if a particular IBD-associated PRDM1 SNP (rs6911490) could be implicated in regulating human CD4+ T cell responses and ultimately IBD pathogenesis.

overall_design: Whole genome transcriptome analysis in stimulated CD4+ TE/M cells to compare the transcriptome change between UC patients carrying or lacking the rs6611490 risk allele.

first: Rebecca

last: Porritt

first: Ana

last: Maretti-Mira

first: Truc

last: Nguyen

first: Michael

last: Couse

first: Michelle

last: Li

first: Rivkah

last: Gonsky

first: Richard

last: Deem

first: Yizhou

last: Wang

first: Jie

last: Tang

first: Talin

last: Haritunians

first: Dermot

last: McGovern

first: Stephan

last: Targan

first: Gislâine

last: Martins


title: Expression data from BJ-hTERT cells expressing shRNA control vector or shRNA against p53.

city: Jerusalem

first: Batsheva

last: Kerem

address: Edmond J Safra Campus

department: Genetics

country: Israel

institute: The Hebrew University

summary:
p53 is a major tumor suppressor gene that is frequently inactivated in human cancers. One prominent suppressive function of p53 is executing cell cycle arrest, senescence and apoptosis upon stress. Importantly, p53 loss is a prerequisite for cancer development, as is evident in Li-Fraumeni patients and p53 knockout mice. Here we investigated the effect of p53-loss, without induction of additional stress, on DNA replication and genomic instability. We revealed that p53 deficiency leads to enhanced proliferation without upregulating essential pathways requited to support normal DNA replication. This uncoordinated S-phase entry leads to replication-induced genomic instability. Interestingly, DNA replication analyses revealed a novel and non-classical form of replication stress induced by p53-loss, characterized by reduced fork rate progression without the compensation of dormant origin activation. Altogether, the results reveal a non-canonical tumor suppressive role of p53 in regulating DNA replication which safeguards genomic stability, underlying cancer predisposition induced by p53-loss.
In order to understand the molecular basis for shp53 non-classical form of replication stress and replication-induced genomic instability, we performed unbiased whole transcriptome analysis of BJ-hTERT control cells in comparison to BJ-hTERT cells expressing shRNA against p53.  Our results suggest that shp53 cells fail to upregulate essential pathways required for normal DNA replication.

overall_design: We analized two sets of BJ-hTERT cells infected with an empty shRNA vector and shp53 vector, each.

first: Maayan

last: Roniger


title: Therapeutic Potential of Human Amniotic Membrane-derived Mesenchymal Stem Cell Conditioned Medium in Combating Oxidative Stress and Age-Related Female Infertility

status: Public on May 30 2025

city: Seoul

first: Kihae

last: Ra

address: Gwan-ak ro 1

department: Research Institute for Veterinary Science

country: South Korea

institute: Seoul National University

summary: Oxidative stress is a crucial factor accelerating age-related functional deterioration of reproductive organ and fertility. Recently, human amniotic membrane-derived mesenchymal stem cells (AMSCs) have emerged as a promising source with notable potential to reduce oxidative damage and support tissue regeneration. This research investigates the impact of intravenous administration of human amniotic membrane-derived mesenchymal stem cell conditioned medium (AMSC-CM) on oxidative stress and reproductive competence in aged female mice. Antioxidative effects of human AMSC-CM was found in the reproductive organs of aged mice at both RNA and protein levels. Human AMSC-CM positively regulated age-dependent changes in reproductive hormones, comparable to those observed in younger mice. RNA sequencing analysis revealed alterations in ovarian and uterine gene expression in aged mice showing that AMSC-CM treatment promotes the expression of genes essential for anti-aging, energy metabolism, and female reproductive processes. These findings highlight the potential of human AMSC-CM as a therapeutic strategy with anti-aging and antioxidative effects against age-related female infertility.

overall_design: RNA sequencing was performed on the ovaries and uterine tissues collected from the mice in the ‘old control’ (OC) and ‘old treatment’ (OT) groups


title: Natriuretic Peptides Modulate Monocyte-Derived Langerhans Cell Differentiation and Promote a Migratory Phenotype

city: Debrecen

first: Attila

middle: Gábor

last: Szöllősi

address: Egyetem tér 1.

department: Department of Immunology

country: Hungary

institute: University of Debrecen

summary: Introduction: The interaction between the nervous and immune systems is crucial for maintaining homeostasis and can influence disease progression in inflammatory skin diseases, such as atopic dermatitis (AD). Sensory neurons in the skin can secrete neuropeptides that modulate immune cell activity, including Langerhans cells (LCs), the primary antigen-presenting cells in the epidermis. This study investigates the effects of neuropeptides on the differentiation of monocyte-derived LCs (moLCs), specifically the neuropeptides with the most profound effect, i.e. atrial- and B-type natriuretic peptides (ANP and BNP, respectively).

overall_design: Transcriptomic profiling of human monocyte-derived Langerhans cells differenitated in the presence of B-type natriuretic peptide or vehicle control (nuclease-free water).

middle: G

first: Dorottya

last: Horváth


title: Gene expression profile of embryonic stem cell-derived TS-like cells by single cell RNA-sequencing

city: Kyoto-shi

first: Masatoshi

last: Ohgushi

state: Kyoto

address: 53 Shogoin Kawahara-cho

country: Japan

institute: Kyoto University

summary: Human embryonic stem cells (ESCs) were differentiated into GATA3-expressing extraembryonic cells (GATA3+ ExE cells) by exposure to chemical inhibitors targeting ACTIVIN/NODAL (A83-01, 1uM) and FGF signaling (PD03714, 100nM). Using these GATA3-expressing cells as a starting material, we established trophoblast stem-like cells (TSLCs) following the established protocol for deriving trophoblast stem cells (TSCs) from the villous trophoblast of human first-trimester placenta. To understand the transcriptional properties of individual TSLCs, we obtained the gene expression profile from single cells.

overall_design: The TSLCs derived from ESCs were cultured for several months with 18 passages.

first: Ohgushi

last: Masatoshi

first: Kaori

last: Honda

first: Rina

last: Takagi

first: Mototsugu

last: Eiraku


title: Whole Genome Bisulfite Sequencing of Col, h2a.w7, HP3 and h2a.w7 x HP3 Arabidopsis thaliana floral buds

city: Los Angeles

first: Zhenhui

last: Zhong

address: 610 Charles E Young Dr East

department: Department of Molecular, Cell and Developmental Biology

institute: University of California, Los Angeles

summary: Whole Genome Bisulfite Sequencing of floral buds of Col, h2a.w7, HP3 and h2a.w7 x HP3 Arabidopsis thaliana plants.

overall_design: The experiment is designed to determine DNA methylation variations of Col, h2a.w7, HP3 and h2a.w7 x HP3 Arabidopsis thaliana floral buds.

first: Pallas

last: Kuo

first: Andrew

last: Tock

first: Xuexia

last: Liu

first: Stephanie

last: Topp

first: Ian

last: Henderson

first: Christophe

last: Lambing


title: Single cell RNA sequencing data obtained from aortic specimens of Fbn1G234D/G234D mouse model

city: Tsukuba

first: Hiromi

last: Yanagisawa

address: 1-1-1 Tennodai

department: TARA center

institute: University of Tsukuba

summary: We generated a novel genetic mouse model of fatal aortic dissection by introducing a pathogenic variant of Fbn1 (Fbn1G234D/G234D), identified in a patient with familial aortic dissection, into mice using the CRISPR/Cas9 system. To investigate the cellular heterogeneity underlying the progression of aortic dissection, we performed single cell RNA sequencing on aortic tissues obtained from Fbn1G234D/G234D (AD) and wild-type mice (WT).

overall_design: Following anesthesia, aortic tissues—including the ascending aorta and aortic arch up to the left subclavian artery—were collected from 3- and 5-week-old wild-type (WT) and Fbn1G234D/G234D mutant (AD) mice. The tissues were enzymatically digested to generate single-cell suspensions, which were then processed using the Chromium™ Single Cell Platform (10x Genomics) according to the manufacturer’s protocol. A total of eight single-cell suspensions were prepared for the study. For 3-week-old mice, the first batch included pooled samples from WT (n=2) and AD (n=2), and the second batch included another set of pooled WT (n=2) and AD (n=2). Similarly, for 5-week-old mice, the first batch consisted of WT (n=2) and AD (n=2), and the second batch included WT (n=3) and AD (n=3).

first: Kenichi

last: Kimura

first: Eri

last: Motoyama

first: Keiichi

last: Asano

first: Erna

last: Raja

first: Ryutaro

last: Ishii


title: Co-targeting of epigenetic regulators and BCL-XL improves efficacy of immune checkpoint blockade therapy in solid tumors

city: Pamplona

first: Ruben

last: Pio

state: Navarra

address: Pio XII Avenue, 55

department: Solid Tumors

country: Spain

institute: Cima Universidad de Navarra

summary: We have used a single-cell sequencing approach to study the changes induced in the immune cells present in the tumor microenvironment of Lewis lung cancer (LLC) mouse tumors treated with a novel therapeutic strategy based on a dual inhibitor of G9a and DNMTs (CM272), a Bcl-XL inhibitor (A1331852) and anti-PD-1 (RMP1-14).

overall_design: Single-cell RNAseq analysis of CD45+/CD11b-/Ly6G- cells  sorted from Lewis lung carcinoma (LLC) tumors treated with vehicle, CM2727 (dual inhibitor of G9a and DNMTs), RMP1-14 (anti-PD-1), and/or A1331852 (anti-Bcl-XL)

first: Yaiza

last: Senent

first: Patxi

last: San Martin-Uriz

first: Ibon

last: Tamayo


title: Single-cell and single-nucleus transcriptome analysis of stripe pattern formation in the embryo of the spider Parasteatoda tepidariorum

city: Takatsuki

first: Yasuko

last: Akiyama-Oda

state: Osaka

address: Murasaki-cho 1-1

institute: JT Biohistory Research Hall

summary: Segmented body plans are a hallmark of arthropods, yet their underlying developmental processes vary widely. We applied single-cell and single-nucleus RNA sequencing of Parasteatoda tepidariorum embryos at late stage 5 and stage 7, a critical period of emergence of segmental units along the anterior-posterior (AP) axis. Single-nucleus RNA sequencing yielded high-quality quantitative data, and subsequent clustering analysis reconstructed an axial alignment of ectoderm cells, reflecting the differing cell states along the segmenting AP axis. This enabled us to generate genome-wide quantitative gene expression profiles along the reconstructed axis.  Employing unbiassed statistical methods, we comprehensively identified pattern elements. These data lay the groundwork for exploring the origins of developmental diversity for arthropod body plans.

overall_design:
Cells and nuclei were isolated from whole embryos at an early germ-band stage (stage 7) and analyzed using single-cell and single-nucleus RNA-seq. The data GSM6069861 in the series GSE201705, single-nucleus RNA-seq data at a late germ-disc stage (late stage 5), were reanalyzed to generate filtered and normalized count matrices.

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The table below lists GEO accessions reused/reanalyzed for this study.
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first: Takanori

last: Akaiwa

first: Hiroki

last: Oda


title: Based on multi-omics analysis, the mechanism of cannabidiol improving SNL mouse model of pain with depression was analyzed

city: Nanchong

first: Guohui

last: Jiang

address: 234 Fujiang Road

department: Institute of Neurological Diseases

institute: North Sichuan Medical College

summary: Neuropathic pain (NP) has a high incidence in the general population, is closely related to anxiety disorders, and has a negative impact on the quality of life. As a natural product, Cannabidiol (CBD) has been widely studied for its potential therapeutic effects on symptoms such as pain and depression (DP). However, the mechanism by which CBD improves neuropathic pain with depression is not fully understood. Therefore, we used multi-omics studies (transcriptomics combined with proteomics) to deeply explore the intersection genes of CBD treatment of NP and DP. Secondly, the core targets were screened by protein-protein interaction (PPI) network, gene ontology (GO), Kyoto Encyclopedia of genes and Genomes (KEGG) analysis, molecular docking and molecular dynamics simulation. Behavioral tests were performed to evaluate the effects of CBD intervention on pain and depression behavior in mice with spinal nerve ligation (SNL), and a dose-response curve was drawn. After determining the optimal intervention dose, the core targets and signaling pathways were verified by Western blotting (WB) and quantitative polymerase chain reaction (qPCR). Finally, we investigated the underlying mechanism of CBD by Nissl staining, immunofluorescence (IF) and transmission electron microscopy (TEM). This study aims to provide further theoretical and experimental basis for the discovery of CBD as a potential therapeutic agent.

overall_design: The experiments were divided into three groups: sham-operated group, SNL model group and CBD (5mg/kg, ip) intervention group. After reaching the predicted time point, the hippocampal tissues of each group were removed for sequencing

first: Ziyi

last: Shen

first: Ming

last: Tang


title: Proliferative arrest induces neuron differentiation and innate immune responses in control and infectious CJD rat septal neurons

city: New Haven

first: Rolando

last: Garcia-Milian

state: CT

address: 333 Cedar St

department: CWML

institute: Yale University

summary: Rat postmitotic septal neurons (SEP) engineered to conditionally proliferate at 33oC were arrested at 37.5oC for weeks without cytotoxic effects. Nine independent cDNA libraries were interrogated to broadly evaluate elevations in b-interferon (b-IFN) and prion protein (PrP) found previously by RT/qPCR in arrested uninfected and CJD infected cells. RNA-seq revealed a variety of upregulated innate immune responses in control and CJD infected cells. Proliferating uninfected, and latently infected (CJ-) cells, showed rare RNA-seq differences. However, major quantitative changes became evident post-arrest. In uninfected cells these included anti-proliferative DNA replication/cell xxNeuregulin-1, GDF6, SFRP4 and PrP. Recruitment of many IFN innate immune pathways, complement, cytokines and clusterin were present in both sets but re-arrested cells with high infectivity (CJ+) escaped replication controls and suppressed neuronal differentiation. Moreover, IFN anti-viral stimulated networks (OAS, cytokines, IFN- and metalloproteinases) were also enhanced among the 342 unique CJ+ transcripts, e.g., Il17, ISG15, CXCL10 and RSAD2 (viperin). These data show 1) defensive innate immune molecules and/or pathways are produced by neurons and participate in normal neuronal differentiation; 2) CJD infectious agents that survive in an undetectable latent state can epigenetically prime cells to produce a strong anti-viral response; 3) downregulation of a plethora of proliferative pathways is also imprinted in cells exposed to infection. Several innate-immune transcripts shared with CJ+ myeloid microglia from brain may be physiologically stimulated to identify latent CJD infections that can be clinically silent for >30 years.

overall_design: Rat postmitotic septal neurons (SEP) engineered to conditionally proliferate at 33oC were arrested at 37.5oC for weeks, cDNA libraries were interrogated to broadly evaluate elevations in b-interferon (b-IFN) and prion protein (PrP) in proliferating uninfected, and latently infected (CJ-), and re-arrested cells with high infectivity (CJ+)

first: Laura

last: Manuelidis


title: Epigenomic alterations in the postnatal period of calves persistently infected with bovine viral diarrhea virus

city: Fort Collins

first: Thomas

middle: R.

last: Hansen

state: Colorado

address: 3107 Rampart Road

department: Biomedical Sciences

institute: Colorado State University

summary: Bovine Viral Diarrhea Virus (BVDV) is a globally prevalent pathogen that causes severe detriment within the commercial cattle industry. BVDV is a vertically transmissible virus that can cross the placenta, infecting both fetus and dam. Infection occurring before the development of the fetal adaptive immune response, prior to approximately 125 days of gestation, results in a persistently infected (PI) calf. The PI calf is unable to produce BVDV specific antibodies, is immunotolerant, and sheds virus consistently. PI fetuses display alterations of the methylome and proteome at day 245 of gestation corresponding to pathologies of the immune system, bone, brain, and heart. It was hypothesized that epigenetic alterations observed in the prenatal period are a remnant of fetal programming due to fetal BVDV infection and persist into the postnatal period. To test this hypothesis, white blood cells were isolated from whole blood collected from 5 PI and 5 control heifers at 4 months of age and subjected to reduced representation bisulfite sequencing. Analysis of the methylome at 4 months of age indicated that 5,349 (64%) of the identified differentially methylated CpG sites (DMSs) were hypomethylated and 3,018 (36%) of DMSs were hypermethylated. Genes in which DMSs were identified were associated with the immune system, hematopoiesis, and the cardiac system. Complete blood count and flow cytometry data corroborate abnormalities of the immune response. Data presented here introduces a new perspective on the predisposition of PI cattle to fatal secondary infections.

overall_design: Reduced-representation bisulfite sequencing (RRBS-Seq ) for five randomly selected 4 month old calves known to be seronegative for BVDV and five age-matched calves known to be persistently infected with BVDV. Jugular vein was collected, and PMBC were collected for DNA analysis.

first: Jessica

last: Kincade

first: Dilyara

last: Murtazina

first: Hanah

middle: M

last: Georges

first: Carolina

middle: L

last: Gonzalez-Berrios

first: Jeanette

middle: V

last: Bishop

first: Terry

middle: E

last: Engle

first: Marcela

last: Henao-Tamayo

first: Jordan

last: Eder

first: Erin

last: McDonald

first: Darcy

last: Deines

first: Hana

last: Van Campen


title: Genome wide expression analysis of high passage growth arrested BJ fibroblasts maintained or withdrawn from rapamycin maintenance.

city: La Jolla

first: Tom

last: Suter

address: 9500 Gilman Dr

department: Department of Medicine

institute: UCSD

summary: BJ fibroblasts were grown in the presence of rapamycin until reaching growth arrest at high passages, at which point cells were maintained in or withdrawn from rapamycin.  RNA was then harvested for RNA-seq

overall_design: BJ fibroblasts were grown to growth arrest in the presence of rapamycin maintenance.  Cells were then either maintained in rapamycin or withdrawn from rapamycin for two weeks.  Experiments, excepting control treatment, were done in duplicates.  RNA-seq tag density was calculated over gene bodies, and each withdrawn samples was compared to its paired maintained sample to determine fold change in gene epression.


title: Genome wide analysis of the effect of p65 inhibition upon the epigenomic landscape in high passage growth arrested cells being withdrawn from rapamycin

type: Genome binding/occupancy profiling by high throughput sequencing

summary: BJ Fibroblasts were maintained in rapamycin and were passaged until reaching growth arrest at very high passages.  These cells were then maintained in or withdrawn from rapamycin, during which time they were given either control treatments or the p65 inhibitor TPCA-1.

overall_design: BJ Fibroblasts were maintained in rapamycin and were passaged until reaching growth arrest at very high passages (>PD75).  These cells were then maintained in or withdrawn from rapamycin for 2 weeks, during which time they were given either treated with control or the p65 inhibitor TPCA-1.  Cells were then harvested for ChIP-seq of enhancer marking histone modification H3K27ac, or the transcription factor p65.  ChIP-seq tag density was then compared between rapamycin withdrawal and maintained samples, as well as between treatment and control samples.


title: Examining changes in the chromosomal interaction network in replicative senescent, rapamycin treatment, and p65 inhibition

summary: Analysis of HiC genome wide chromosomal interaction data in the context of replicative senescence, rapamycin treatment, the withdrawal of rapamycin, and with p65 inhibition in rapamycin withdrawal.  This was done in order to determine what changes in chromosomal architecture occur during replicative senescence, and what impact rapamycin treatment or withdrawal or p65 inhibition has upon these changes.

overall_design: BJ fibroblasts were grown in the presence or absence of rapamycin, and harvested at PD31 or PD66 (when the non-rapamycin treated BJ fibroblasts were senescent).  A set of BJ fibroblasts treated with rapamycin were also grown to high passage (PD75+) growth arrest (HPGA), and were then either maintained in rapamycin, or withdrawn from rapamycin in the presence or absence of p65 inhibitor.  in situ HiC was then performed upon each sample, and all possible pairwise combinations of samples were compared to determine whether significant changes in A/B domains were observed between conditions.


title: Genome wide expression analysis of senescent BJ fibroblasts in the presence or absence of various SMAD2/3 regulating molecules

summary: BJ fibroblasts were grown to replicative senescence, and were then treated with control or SMAD2/3 regulating ligands.  RNA was then harvested for RNA-seq.

overall_design: BJ fibroblasts were grown to replicative senescence.  Cells were then given treatment with control or SMAD2/3 regulating ligands Activin A (100ng/ml), TGFB2 (1ng/ml), Inhibin (100ng/ml)+TGFB2 (1ng/ml), or Inhibin (100ng/ml).  Experiments, excepting control treatment, were done in duplicates, and each treatment was compared with controls to determine a fold change in transcription for genes.


title: Genome wide expression analysis of high passage growth arrested BJ fibroblasts maintained or withdrawn from rapamycin maintenance, in the presence or absence of various SMAD2/3 regulating molecules

summary: BJ fibroblasts were grown in the presence of rapamycin until reaching growth arrest at high passages, at which point cells were maintained in or withdrawn from rapamycin, while also being treated with control or SMAD2/3 regulating ligands.  RNA was then harvested for RNA-seq

overall_design: BJ fibroblasts were grown to growth arrest in the presence of rapamycin maintenance.  Cells were then either maintained in rapamycin or withdrawn from rapamycin for two weeks, along with treatment with control or SMAD2/3 regulating ligands Activin A (100ng/ml), TGFB2 (1ng/ml), Inhibin (100ng/ml)+TGFB2 (1ng/ml), or Inhibin (100ng/ml).  Experiments, excepting control treatment, were done in duplicates, and each treatment was compared with controls to determine a fold change in transcription for genes.


title: Genome wide expression analysis of the effect of senescence associated transcription factor knockdown upon the transcription of senescent cells.

summary: Senescence associated transcription factors were stably knocked down in senescent BJ fibroblasts, which were then profiled by RNA-seq.

overall_design: BJ fibroblasts were grown to replicative senescence, at which point they were stably transduced with lentivirus expressing shRNAs targetting the senescence associated transcription factors p65, NFIA, NFIB, NFIC, NFIX, SMAD2, SMAD3, or GFP control.  Samples were then harvested for RNA-seq.  RNA-seq tag density was calculated over each knockdown sample and compared with control.


title: Genome wide expression analysis of high passage growth arrested BJ fibroblasts maintained or withdrawn from rapamycin maintenance, upon control or transcription factor knockdown

summary: shRNAs targetting p65, NFIA, or GFP (control) were expressed in High Passage Growth Arrested BJ fibroblasts maintained in rapamycin, which were then maintained or withdrawn from rapamycin and harvested for RNA-seq

overall_design: shRNAs targetting p65, NFIA, or GFP (control) were expressed in High Passage Growth Arrested BJ fibroblasts maintained in rapamycin, which were then maintained or withdrawn from rapamycin (10nM) for 14 days and then harvested for RNA-seq.  RNA-seq tag densities over gene bodies were compared between rapamycin withdrawal and maintained samples, as well as between withdrawal transcription factor and withdrawal control samples.  Multiple control shRNA replicates were collected, and comparisons were made between samples with the same "Rep" number.


title: Genome wide expression analysis of the impact of JMJD6 knockdown upon the senescent transcription program.

summary: RNA-seq was performed in senescent BJ fibroblasts which had undergone stable JMJD6 or control knockdown since early young passages.

overall_design: shRNAs targetting JMJD6 or control non-silencing sequences were stably transduced into young, non-senescent BJ fibroblasts.  These lines were then cultured until reaching replicative senescence, at which point they were harvested for RNA-seq.  Two separate JMJD6 targetting shRNAs were used, each into a separate cell line.  RNA-seq tag density over gene bodies in JMJD6 knockdown cells were compared to the density in the control senescent sample.


title: Genome wide expression analysis of young vs senescent BJ fibroblasts

summary: Young-quiescent and senescent BJ fibroblasts were harvested for RNA-seq analysis in the presence or absence of doxycycline treatment.

overall_design: BJ fibroblasts transduced with doxycycline inducible empty vector control retroviruses were grown in the absence or continued presence of doxycycline (20nm), and young-quiescent or senescent cells were collected for RNA-seq.  Senescent and young doxycycline treated samples were compared for changes in genes expression, and senescent and young doxycycline untreated samples were compared separately.


title: Genome wide expression analysis of the impact of rapamycin and DOT1L inhibitor treatment upon the progression of replicative senescence

summary: BJ fibroblasts were grown in the continued presence of either rapamycin, the DOT1L inhibitor EPZ-5676, rapamycin + EPZ-5676, or control treatment, and collected for RNA-seq at young-quiescent, intermediate-quiescent, or senescent cell states.

overall_design: BJ fibroblasts were grown in the continued presence of either rapamycin (10nM), the DOT1L inhibitor EPZ-5676 (500nM), rapamycin (10nM) + EPZ-5676 (500nM), or control treatment, and collected for RNA-seq at young-quiescent, intermediate-quiescent, or senescent cell states.  RNA-seq tag density across gene bodies was determined for each sample and compared between different cell stages of senescence as well as between treatment and control samples.


title: Genome wide expression analysis of high passage growth arrested BJ fibroblasts maintained or withdrawn from rapamycin maintenance

summary: BJ fibroblasts were grown in the presence of rapamycin until reaching growth arrest at high passages, at which point cells were maintained in or withdrawn from rapamycin, and RNA was then harvested for RNA-seq

overall_design: BJ fibroblasts were grown to growth arrest in the presence of rapamycin maintenance.  Cells were then either maintained in rapamycin or withdrawn from rapamycin for two weeks, and harvested for RNA-seq.  The experiment was replicated, with the withdrawal samples from the first replicate compared to the maintained samples from the first replicate, and the second withdrawal samples compared with the second maintained samples.


title: Genome wide mapping of open chromatin in senescence cells upon knockdown of senescence associated transcription factors

summary: ATAC-seq was performed upon senescent BJ fibroblasts upon controls shRNA or knockdown of p65, SMAD2 or SMAD3, or NFI family members, in order to determine the role of each factor upon senescence associated enhancers.

overall_design: Senescent BJ fibroblasts were collected after 2 weeks knockdown of listed transcription factors, and ATAC-seq was performed upon each sample.  Knockdown samples were compared with control GFP knockdown, in terms of the ATAC-seq tag counts at senescence associated enhancers.


title: Genome wide mapping of histone mark and transcription factor binding in the context of experiments exploring the enhancer program of senescence II

summary: Analysis of histone and transcription factor epigenomic landscape of BJ fibroblasts from young to replicative senescent

overall_design: BJ Fibroblasts were collected at young, intermediate early, and intermediate late, and senescent stages, at which point chromatin was harvested and prepared for ChIP-seq against a variety of histone marks and transcription factors.


title: Genome wide mapping of the effects of SMAD2/3 ligand treatment upon the distribution of senescence associated transcription factors

summary: Analysis of SMAD2/3 and NFI binding in replicative senescent BJ fibroblasts upon treatment with SAMD2/3 regulating ligands.

overall_design: Senescent BJ fibroblasts were treated with SMAD2/3 regulating ligands including Activin A (100ng/ml), TGFB2 (1ng/ml), Inhibin (100ng/ml)+TGFB2 (1ng/ml), or Inhibin (100ng/ml), or control, and subsets of these cells were collected for ChIP-seq with SMAD2/3 and NFI.  Tag densities of ligand treated samples at senescence associated enhancers were compared with those of control treated samples.


title: Genome wide mapping of the effect of SMAD2/3 regulating treatments upon the epigenomic landscape in high passage growth arrested cells being withdrawn from rapamycin

summary: BJ Fibroblasts were maintained in rapamycin and were passaged until reaching growth arrest at very high passages.  These cells were then maintained in or withdrawn from rapamycin, during which time they were given either control treatments or treatments with factors regulating SMAD2/3 activity.

overall_design: BJ Fibroblasts were maintained in rapamycin and were passaged until reaching growth arrest at very high passages (>PD75).  These cells were then maintained in or withdrawn from rapamycin for 2 weeks, during which time they were given either treated with control or the SMAD2/3 regulating ligands Activin A (100ng/ml), TGFB2 (1ng/ml), Inhibin (100ng/ml)+TGFB2 (1ng/ml), or Inhibin (100ng/ml).  Cells were then harvested for ChIP-seq of enhancer marking histone modifications H3K27ac or H3K4me2, or the transcription factors p65, SMAD2/3, or NFI.  ChIP-seq tag density was then compared between rapamycin withdrawal and maintained samples, as well as between treatment and control samples.


title: Genome wide mapping of histone mark and transcription factor binding in the context of replicative senescence in IMR90 cells.

summary: Analysis of histone and transcription factor epigenomic landscape of IMR90s from young to replicative senescent

overall_design: IMR90s were collected at young-quiescent and senescent stages, at which point chromatin was harvested and prepared for ChIP-seq against a variety of histone marks and transcription factors.  ChIP-seqs of histone marks H3K27ac and H3K4me2 were performed in duplicate, and, in all experiments, senescent samples were compared against young samples for each factor.


title: The effect of rapamycin upon the enhancer landscape in the context of replicative senescence

summary: ChIPs of the histone modifications H3K27ac and H3K4me2, which mark enhancers and promoters, were examined during the course of replicative senescence in BJ fibroblasts that were grown in the presence or absence of rapamycin.

overall_design: BJ fibroblasts were cultured in the presence or absence of rapamycin and were collected at young-quiescent, intermediate-quiescent, and senescent stages, at which point chromatin was harvested and prepared for ChIP-seq the histone marks H3K27ac and H3K4me2.  The ChIP-seq tag density of various enhancer regions  were compared between different cell stages as well as between rapamycin treatment and non-treatment.  These fold changes in tag density were compared between marks to determine which enhancers showed consistent changes in activation in both histone modifications


title: Genome wide mapping of nascent transcription in the progression of replicative senescence

summary: GRO-seq was performed upon BJ fibroblasts that were young-quiescent, intermediate passage-quiescence, or senescent

overall_design: BJ Fibroblasts were collected at young-quiescent, intermediate-quiescent, and senescent stages, cells were prepared for GRO-seq in order to determine nascent transcription, and fold change of tags representing nascent transcripts were quantified upon genes and compared between cell states.


title: Genome wide mapping of the effect of JMJD6 knockdown upon H4R3me2s in the context of replicative senescence

summary: Native ChIP-seq of H4R3me2s was performed in young-quiescent and senescent BJ fibroblasts that had been treated with control or JMJD6 targetting shRNA

overall_design: Bj fibroblasts lines stably expressing either non-silencing or JMJD6 targetting shRNAs were generated.  These lines were harvested at young-quiescent or senescent cell stages, and native ChIPs against H4R3me2s were performed in each cell state/knockdown-line.  Tag density of the H4R3me2s ChIP-seq data was compared between knockdown vs control lines, and between young and senescent cells.


title: Genome wide mapping of histone mark and transcription factor binding in the context of experiments exploring the enhancer program of senescence I


title: Genome wide mapping of JMJD6 binding in the context of experiments exploring the enhancer program of senescence

summary: Analysis of JMJD6 binding relative to the epigenomic landscape of BJ fibroblasts from young to replicative senescent.

overall_design: BJ Fibroblasts with doxycycline inducible JMJD6 tagged with either TY1 or HA, were collected for ChIP-seq after acute doxycycline treatment, at young-quiescent and senescent stages.  Tagged JMJD6 binding was then compared between young and senescent cells at senescence associated enhancer subsets.


title: Meningeal lymphatics-microglia axis regulates synaptic physiology [acz.vehicle]

status: Public on May 29 2025

city: St. Louis

first: Jonathan

last: Kipnis

state: MO

address: 4515 McKinley Ave

institute: Washington University in St. Louis

summary: Meningeal lymphatics serve as the primary outlet for cerebrospinal fluid, and their dysfunction is associated with various neurodegenerative conditions. Previous studies have demonstrated that dysfunctional meningeal lymphatics evoke behavioral deficits, but the neural mechanisms underlying those behavioral changes remained elusive. Here, we show that prolonged impairment of meningeal lymphatics alters the balance of cortical excitatory and inhibitory synaptic inputs by reducing inhibitory synapses, accompanied by deficits in novelty recognition tasks. These synaptic and behavioral alterations are mediated by microglia, which exhibit transcriptomic, morphological, and functional alterations as a result of lymphatic dysfunction. Notably, microglial expression of Il6 increases, thereby mediating the reduction in inhibitory synapses via neuronal signaling. Interestingly, improving the function of meningeal lymphatics in aged mice restores the numbers of functional inhibitory synapses and cortical network activity. Our findings suggest that dysfunctional meningeal lymphatics adversely impact cortical circuitry through a microglia−IL-6-dependent mechanism, providing a potential target for the treatment of aging-associated cognitive decline.

overall_design: The subject C57BL/6J male mice were cardio-perfused with 30 ml of 5 U/ml Heparin-PBS after seven days of treatment of idiopathic intracranial hydrocephalus patient-equivalent dose of acetazolamide treatment (200 mg ACZ/kg mouse) twice daily. Brains were harvested, located in heparin-PBS and cut through the sharp razor roughly at the AP=+1.00 mm. The anterior part of the brain faces down to the bottom at the section point and is cut horizontally half and half to remove the olfactory tract and striatum; only the upper chunk was used as a PFC sample.  5 PFC samples were pulled per group, and minced with the sharp razor in RPMI 1640 medium with 2% FBS and digested by collagenase VIII and DNase (in 5 ml RPMI 1640 + 2 % FBS) for 45 minutes and titrated with P1000 pipettes once at 30 minutes after digestion starts, and one more at 45 minutes after.  Samples were strained through a 70-um strainer, and 5 ml RPMI 1640 + 10% FBS medium was added to stop digestion and spun down in 450g for 5 minutes. Pellets were resuspended in 14% BSA and spun down in 800 g for 10 minutes with slow acceleration/declaration. Supernatnace was removed carefully, and the pellet was resuspended in MACS buffer (0.5% BSA, 2 mM EDTA containing PBS), followed by spinning down (450g, 5 mins). 50 ul of CD45-bead (Miltenyi, 130-052-301) was added with 450 ul MACS buffer, and CD45+ cells were prepared according to the manufacturer's protocol.

first: Kyungdeok

last: Kim

first: Daviti

last: Abramishvili

first: Siling

last: Du

first: Zachary

last: Papadopoulos

first: Jay

last: Cao

first: Igor

last: Smirnov

first: Jean-Leon

last: Thomas

first: Marco

last: Colonna


title: Immune-responsive gene-1: The mitochondrial Key to Th17 Cell Pathogenicity in CNS Autoimmunity

status: Public on May 28 2025

city: detroit

first: Shailendra

last: Giri

state: MI

address: 2799 W grand boul

department: Neurology Research

institute: henry ford hospital

summary: Pathogenic Th17 cells play crucial roles in CNS autoimmune diseases such as multiple sclerosis (MS), but their regulation by endogenous mechanisms remains unknown. Through RNA-seq analysis of primary brain glial cells, we identified immuno-responsive gene 1 (Irg1) as one of the highly upregulated gene under inflammatory conditions. Validation in the spinal cord of animals with experimental autoimmune encephalomyelitis (EAE), an MS model, confirmed elevated Irg1 levels in myeloid, CD4, and B cells in the EAE group raising the concern if Irg1 is detrimental or protective. Irg1 knockout (KO) mice exhibited severe EAE disease, increased mononuclear cell infiltration, and increased levels of triple-positive CD4+ T cells expressing IL17a, GM-CSF, and IFNγ. A lack of Irg1 in macrophages elevates Class II expression, promoting the polarization of myelin-primed CD4+ T cells into pathogenic Th17 cells via the NLRP3/IL-1β axis. Adoptive transfer in Rag-1 KO and single-cell RNA sequencing highlighted the crucial role of Irg1 in shaping pathogenic Th17 cells. Moreover, bone marrow chimeras revealed that immune cells lacking Irg1 maintained pathogenic and inflammatory phenotypes, suggesting its protective role in autoimmune diseases, including MS.

overall_design:
For the generation of bone marrow chimeras, host (CD45.1) animals were treated with three doses of busulfan (20 mg/ml), and on the fourth day, the host received an i.v. injection of 10 million BM cells [50:50 mixture of CD45.1 (Wt) and CD45.2 (Irg1 KO)]. Using flow cytometry, we sorted CD45.1+ and CD45.2+ cells from BILs isolated via a Percoll gradient from the brain and spinal cord after EAE induction. BILs were stained with primary antibodies against mouse PerCP-Cy5.5-CD45.1 and BV421-CD45.2 (BioLegend) and sorted on a BD FACSAria™ III. Furthermore, the sorted populations were re-run to check the purity of the sorted cells. Next, we used an automated cell counter (Bio-Rad) to determine the viability of CD45+1+ and CD45.2 cells via trypan blue and found that >98% of the cells were viable. Then, RNA was harvested from the sorted cells and processed for RNA sequencing.
Three mice per group (WT and Irg1 knockout) were pooled for analysis. Experimental autoimmune encephalomyelitis (EAE) was induced using MOG peptide in combination with Pertussis toxin and monitored for 20 days. Following euthanasia, spinal cords were harvested, and single-cell suspensions were prepared using Accumax enzymatic dissociation followed by Percoll gradient purification. A total of approximately 200,000 cells per group were processed using the Fluent pipeline. Two single-cell RNA-seq libraries per group were generated using the Fluent PIPseq T20 3' capture and lysis kit.
The timed pregnant rat was purchased from Charles River Laboratory. The brains of 2–3-day-old female and male pups were used to prepare mixed glial cell cultures. We followed our previously published protocol to prepare primary mixed glial cell cultures (3). After 10 days of culture, the cells were treated with LPS (0.5 µg/ml) + IFN (20 ng/ml) to induce inflammatory conditions in our study. After 8 h of treatment, the cells were processed for RNA isolation and RNA-seq

first: Mohammad

last: Nematullah

first: Mena

last: Fatma

first: Guoli

last: Zhou

first: Bharat

last: Prajapati

first: Faraz

last: Rashid

first: Kameshwar

last: Ayasolla

last: Ahmed

first: Ruicong

last: She

first: Sajad

last: Mir

first: Insha

last: Zahoor

first: Nasrul

last: Hoda

first: Ramandeep

last: Rattan


title: Genome wide mapping of H3K27ac in IMR90 oncogene induced senescence

first: Tom

last: Suter

summary: Analysis of H3K27ac ChIP-seq in IMR90 cells in oncogene induced senescence versus control.

overall_design: IMR90 cells were treated with either control or H-Ras restrovirus to induce oncogene induced senescence in the latter condition.  H3K27ac ChIP-seq was then performed upon each condition, and tag density at enhancers was compared between the control and oncogene induced senescent conditions.


title: Polyglutamylation of microtubules drives neuronal remodeling

city: Munich

first: Monika

last: Leischner-Brill

address: Arcisstr. 21

department: School of Medicine and Health

institute: Technical University of Munich

summary: Developmental remodeling shapes neural circuits via activity-dependent pruning of synapses and axons. The cytoskeleton is critical for this process, as microtubule loss via enzymatic severing is an early step of pruning across many circuits and species. However, how microtubule-severing enzymes, such as spastin, are activated in specific neuronal compartments remains unknown. Here, we reveal that polyglutamylation, a posttranslational tubulin modification that is enriched in neurons, plays an instructive role in developmental remodeling by tagging microtubules for severing. Motor neuron-specific gene deletion of enzymes that add or remove tubulin polyglutamylation—TTLL glutamylases vs. CCP deglutamylases—accelerates or delays neuromuscular synapse remodeling in a neurotransmission-dependent manner. This mechanism is not specific to peripheral synapses but also operates in central circuits, e.g., the hippocampus. Thus, tubulin polyglutamylation acts as an activity-dependent rheostat of remodeling and shapes neuronal morphology and connectivity.

overall_design: RNA sequencing following ribotagging from murine spinal cord at different postnatal ages (day 5, 7, 9, 11, 14). HA-tag on ribosomes was induced by crossbreeding RiboTag (also known as Rpl22tm1.1Psam, JAX #011029), ChAT-Cre (Chattm2(cre)Lowl, JAX #006410), and Spast flox mice (Brill et al., Neuron, 2016).

first: Antoneta

last: Gavoci

first: Thomas

last: Misgeld

last: Brill


title: Field Performance, Alkaloid Quantities, and Transcriptome Profiles of Nearly Isogenic Tobacco Genotypes Possessing Recessive Allele Combinations at the Nic1, Nic2, and Myc2a Loci

city: Raleigh

first: Ramsey

last: Lewis

state: North Carolina

address: 101 Derieux Pl

department: Crop and Soil Sciences Department

institute: North Carolina State University

summary: Tobacco with modified genetics controlling alkaloid accumulation has been of interest because of the possibility of mandated lowering of nicotine levels in combustible cigarettes by regulatory authorities. Transcription factors coded by the Nic1, Nic2, and Myc2a loci act as positive regulators of genes directly involved in alkaloid accumulation. Different combinations of alternative alleles at these loci were assembled in homozygous nearly isogenic lines (NILs). Recessive nic1/nic2 alleles were found to have greater influence on alkaloid reduction as compared to a mutant myc2a allele. When combined into single genotypes, these alleles operated in an additive manner to further reduce alkaloid levels in field and greenhouse experiments. This was at the expense of reduced cured leaf quality, however. An RNA-seq experiment was carried out to investigate global changes in root expression due to different genotypes at these three loci. Up to 681 differentially expressed genes (DEGs) were identified between the studied NILs, with expression of most DEGs being downregulated by recessive alleles. In general, the mutant myc2a allele appeared to suppress a subset of previously characterized alkaloid biosynthetic genes with relatively weaker effect as compared to the nic1/nic2 alleles. There was little evidence that root expression of Nic1 or Nic2 genes was affected by allelic variability at the Myc2a locus, or vice versa. The list of DEGs influenced by genotypes at these three loci may contain candidate genes coding for currently uncharacterized enzymes involved with tobacco alkaloid accumulation.

first: Rui

last: Shi


title: Genomic and functional characterization of the tumor-killing capacity of Imiquimod-activated plasmacytoid dendritic cells

city: Vienna

first: Iros

last: Barozzi

address: Borschkegasse 8a

country: Austria

institute: Medical University Vienna

summary: Plasmacytoid dendritic cells (pDCs) are a subset of DCs that act as important modulators of anti-tumor immune responses. pDCs have been associated with poor prognosis of tumor patients. Despite this, upon TLR7 activation, pDCs have also been shown to acquire tumor killing capacities, and we previously provided evidence on the role of activated pDCs in eliminating tumor cells in vivo, independently of adaptive immunity. In this study we investigated the mechanism by which pDCs acquire tumor-killing capacities following TLR7/8 triggering by Imiquimod (IMQ). By combining pathway perturbation and analysis of transcriptome, surfaceome, secretome, and killing function, we identified the MAPKs and NF-kB pathways as important downstream mediators of the tumor-killing ability of these cells. JNK inhibition reduces killing by secreted factors whereas p38 or NF-kB inhibition enhances cell mediated killing. Analysis of surface and secreted proteins revealed differential control of expression and secretion by IFN-I/-II, MAPK and NF-kB, while indicating that a complex signaling network is necessary to shape the cytotoxic phenotype of activated pDCs. Our integrated analysis converged on a new pDCs killing gene signature that is predictive of survival in cohorts of melanoma patients. These newly identified markers will be instructive for the rationale design and interpretation of further single-cell and functional approaches aimed at studying the role of pDCs in the tumor microenvironment. These might pinpoint novel strategies to improve cancer management and treatment, by modulating the pDCs pro- vs anti-tumorigenic functions.

overall_design: Total RNA-seq of murine Plasmacytoid dendritic cells (pDCs) either untreated, or treated with either vehicle alone (Limulus amebocyte lysate; LAL) or with Imiquimod (IMQ), for 6 or 16 hours.

first: T

last: Gastaldi

first: P

last: Novoszel

first: M

last: Sanlorenzo

first: C

last: de Sà Fernandes

last: Mohr

first: D

last: Krauss

first: B

last: Blauensteiner

last: Holcmannn

first: I

last: Sibilia


title: Characterization of a centrosme loss - induced tumorigenic signature in prostate epithelial cells

city: Tucson

first: Jiawen

last: Yang

state: Arizona

address: 1515 N Campbell Ave

department: University of Arizona Cancer Center

institute: The University of Arizona

summary: Unlike many cancers, prostate cacner (PCa) lacks hallmark mutations in key oncogenes and tumor-suppressorgenes. Instead, PCa exhibit extensive genomic rearrangements and chromosomal instability (CIN). Previously, we found that cell within early-grade human primary prostate adenocarcinomas (PRAD) frequently lack centrosomes, and this frequency correlates with tumor grade. We demonstrated that transient removal of centrosomes within non-tumorigenic human prostate epithelial cells (hPrEC) induces CIN and was, strikingly, sufficient to transform subpopulations of cells capable of producing xenograft tumors in mice. To unravel the molecular mechanisms underlying this path to tumorigenesis, we isolated RNA from parental hPrECs, clonal lines subjected to transient centrosome loss, and xenograft tumor cells and performed bulk RNA-seq. This allowed us to characterize the transcriptomic profile induced by centrosome loss.

overall_design: To induce centrosome loss in prostate epithelial cells, we applied the PLK4 inhibitor centrinone to PrEC Hahn parental cells for 15 days, followed by monoclonal expansion in normal culture media without centronone. We selected lines with elevated levels of MYC and ERG and xenografted them into male NSG mice. Among all the lines injected into NSG mice, CN9 exhibited the highest tumorigenicity level, with 9 out of 12 injection sites forming tumors. Xenograft tumors derived from CN9 were then collected and cultured on petri dishes. Subsequently, we performed gene expression profiling analysis using RNA-seq data from parental cells (PrEC Hahn cells), centrosome loss clone (CN9), and xenograft tumor cells (CN1_1, CN1_2, CN2_2).

first: Gregory

middle: C

last: Rogers

first: Emily

last: Loertscher


title: Host-pathogen interaction profiling of nontypeable Haemophilus influenzae and Moraxella catarrhalis coinfection of bronchial epithelial cells.

status: Public on May 27 2025

city: Baltimore

first: Adonis

last: D'Mello

state: Maryland

address: Health Science Facility III, 670 W Baltimore St 3rd floor

institute: IGS- University of Maryland Baltimore

summary: Chronic obstructive pulmonary disease (COPD) is a progressive inflammatory lung disease and the third leading cause of death globally. Nontypeable Haemophilus influenzae (NTHi) and Moraxella catarrhalis (Mcat) are common pathogens in individuals with COPD. Acquisition of NTHi or Mcat can cause acute exacerbations of COPD. NTHi and/or Mcat can also persist for months in the lower airways and lead to chronic inflammation. We hypothesized that infection by NTHi or Mcat, singly or during coinfection, requires regulation of specific bacterial and host cell pathways. We investigated this phenomenon using an in vitro cell culture model consisting of lung carcinoma H292 cell lines, infected with NTHi, Mcat, or both species. Samples were fractionated into “apical fluid”, containing free floating bacteria, and adhered/invaded bacteria on or within H292 cells. We used transcriptomic profiling with RNA-seq and various bioinformatic analyses to disentangle host-pathogen interactions in epithelial cell infection from the perspective of each species. Several biological pathways were differentially regulated across all conditions (31, NTHi; 22, Mcat; and 169, Human). NTHi transcriptomic profiles differed during single and coinfection; examples included downregulation of iron-sulfur metabolism (IscR regulon) and differential regulation of quorum sensing in coinfection compared to single infection.  Mcat was comparatively less affected by the presence of NTHi during coinfection.  H292 epithelial cells responded broadly to all infections with distinct responses to single and coinfection. Enriched host pathways included influenza/interferon/Wnt and proinflammatory responses. These findings suggest common and distinct processes involved in NTHi and/or Mcat induced COPD pathogenesis and have implications for therapeutic intervention.

sample_organism:
Moraxella catarrhalis
Haemophilus influenzae
Homo sapiens

overall_design: To investigate the host-pathogen interactions of COPD pathogens, nontypable Haemophilus influenzae (NTHi) and Moraxella catarrhalis, with human epithelial cells NCI-H292. We generated single infections and coinfections of H292 cells, fractioned samples into apical fluid or adherent/invaded cells, as well as bacterial controls. We then performed comparative gene expression profiling analysis using multi-species RNA-seq.

first: Hervé

last: Tettelin

first: Timothy

middle: F

last: Murphy

first: Melinda

last: Pettigrew


title: A spatial transcriptomic atlas of acute neonatal lung injury across development and disease severity

city: Phoenix

first: Nicholas

last: Banovich

state: AZ

address: 445 North Fifth Street

department: Integrated Cancer Genomics

institute: TGen - Translational Genomics Research Institute

summary: A molecular understanding of lung organogenesis requires delineation of the timing and drivers of spatial-temporal cellular movements that ultimately form and support a surface capable of gas exchange. While the advent of single-cell transcriptomics has allowed for the discovery and identification of transcriptionally distinct populations present during lung development, the spatiotemporal dynamics of these transcriptional shifts remain undefined.. With imaging-based spatial transcriptomics, we analyzed the gene expression patterns in 17 human infant lungs at varying stages of development and lung injury, creating a spatial transcriptomics atlas of ~1.2 million cells. We applied computational clustering approaches to identify shared molecular similarities among this cohort, establishing a framework that will generate hypotheses about how tissue architecture and molecular spatial relationships are coordinated during normal development and disrupted in disease. Recognizing that all preterm birth represents an injury to the developing lung, we have moved away from the conventional paradigm of classifying an infants as “disease” or “control” in favor of a linear regression approach that accounted for the routinely collected object measures of gestational age, life span, and disease severity. Within this new framework, we have identified cell type patterns across these variables that would likely be overlooked when using a binary conventional “diseased vs. control” comparison. Together, these data represent a resource for the lung research community, supporting discovery-based inquiry and identification of targetable molecular mechanisms in both normal and arrested human lung development.

overall_design:
We profiled 17 infant lung samples that were collected from the Human Infant Lung Repository at Vanderbilt University with a gene probeset of 343 genes using the 10X Genomics Xenium platform. Sample names were generated by appending ‘PDL’ and then ranged from ‘001’ - ‘017’ (ex. PDL001). This dataset is comprised of individuals that range from the canalicular stage of lung development to the alveolar, with variable levels of disease pathologies. We took a 3x3 μ square of each tissue and arranged them in a tissue micro-array. We then took two adjacent serial sections and ran them concurrently, generating two technical replicates per sample.

UPDATE: [Jun-26-2025] The transcript.csv.gz files were replaced.

first: Saahithi

last: Mallapragada

first: Ruqian

last: Lyu

first: Arianna

last: Williams-Katek

first: Brandon

middle: K

last: Fischer

first: Niran

last: Hadad

first: Evan

middle: D

last: Mee

first: Shawyon

middle: P

last: Shirazi

first: Anne

last: Hilgendorff

first: Laurie

last: Eldredge

first: Gail

middle: H

last: Deutsch

first: Davis

middle: J

last: McCarthy

first: Jonathan

last: Kropski

first: Jennifer

last: Sucre


title: ONECUT2 reprograms neuroendocrine fate and is an actionable therapeutic target in small cell lung cancer [DMS53]

first: Mirja

last: Rotinen

state: Navarre

address: Avenida Barañain s/n

department: Health Sciences

institute: Public University of Navarre

summary: Small cell lung cancer (SCLC) is a highly aggressive malignancy with extremely poor prognosis. SCLC cells exhibit high plasticity and can progress from neuroendocrine (NE) to non-NE phenotypes. This dynamic evolution promotes treatment resistance and relapses, representing a challenge for targeted therapies in this elusive disease. Here we identify the transcription factor ONECUT2 (OC2) as a driver of plasticity in SCLC, leading to non-NE transcriptional states. OC2 is highly expressed in SCLC tumors compared to normal lung tissue and its expression is associated with heightened clinical stage and lymph node metastasis. We show that OC2 is a repressor of ASCL1, the NE master regulator transcription factor. In addition, OC2 upregulates non-NE programs through activation of c-MYC and NOTCH signaling. We also demonstrate that OC2 is required for growth and survival of SCLC cells and that it can be targeted with a small-molecule inhibitor that acts synergically with cisplatin, providing a novel therapeutic strategy for OC2 active SCLC tumors.

overall_design: We performed RNA-Seq after OC2 enforced expression (OE vs C) in the DMS53 cell line, to investigate whether OC2 plays a role in SCLC cell plasticity.


title: ONECUT2 reprograms neuroendocrine fate and is an actionable therapeutic target in small cell lung cancer [NCI-H510A]

overall_design: We performed RNA-Seq after an induction/off-phase/induction of OC2 expression in the NCI-H510A cell line, to investigate whether OC2 plays a role in SCLC cell plasticity.


title: Functional genomics assays, including ChIP-seq, ATAC-seq, RNA-seq, GRO-seq, and in situ Hi-C, in the context of replicative senescence

first: Tom

last: Suter

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

overall_design: Refer to individual Series


title: Tape Strip Profiling of Checkpoint Inhibitor-Associated Dermatitis Highlights Pan-T-Cell Activation: A Pilot Study

status: Public on May 26 2025

city: New York

first: Madeline

last: Kim

address: 1 Gustave Levy Place

institute: Icahn School of Medicine at Mount Sinai

summary: This pilot study examined gene expression in lesional and non-lesional tape strip samples obtained from 5 patients with immune-checkpoint inhibitor (ICI) eczematous reactions/ICI-Ecz, 3 patients with ICI lichen planus-type reactions/ICI-LP, and 8 patients with atopic dermatitis (AD), as well as from 9 demographically matched healthy controls. Differential expression was defined with |fold-change/FCH|>2 and false discovery rate/FDR<0.1. Significant upregulation in Th1 and Th2, notably in IL4R, was observed in both ICI groups. JAK/STAT upregulation was additionally observed in ICI-Ecz and AD.

overall_design: RNA-seq was performed on lesional and non-lesional tape strip samples obtained from 5 patients with immune-checkpoint inhibitor (ICI) eczematous reactions/ICI-Ecz, 3 patients with ICI lichen planus-type reactions/ICI-LP, and 8 patients with atopic dermatitis, as well as from 9 demographically matched healthy controls. Differential expression was defined with |fold-change/FCH|>2 and false discovery rate/FDR<0.1.

first: Camille

last: Powers

first: Annie

last: Chang

first: Benjamin

last: Hu

middle: R

last: Block

first: Austin

last: Piontkowski

first: Jeremy

last: Orloff

first: Jade

middle: N

last: Young

first: Yeriel

last: Estrada

first: Digpal

last: Gour

first: Emma

last: Guttman-Yassky

last: Gulati


title: Diet modulates the protective effects of dimethyl fumarate mediated by the immunometabolic neutrophil receptor HCA2

status: Public on May 25 2025

city: Lübeck

first: Kristian

last: Händler

address: Ratzeburger Allee 160

department: Human Genetics

institute: UKSH

summary: Monomethyl fumarate (MMF) and its prodrug dimethyl fumarate (DMF) are currently the most widely used agents for the treatment of multiple sclerosis (MS). However, not all patients benefit from DMF. We hypothesized that the variable response of patients may be due to their diet. In support of this hypothesis, mice subjected to experimental autoimmune encephalomyelitis (EAE), a model of MS, did not benefit from DMF treatment when fed a lauric acid-rich (LA) diet, in contrast to normal chow (NC) or, more so, a high-fiber (HFb) diet. DMF lacked efficacy in the LA diet fed group despite similar resorption and preserved effects on plasma lipids. When mice were fed the permissive HFb diet, the protective effect of DMF treatment depended on hydroxycarboxylic receptor 2 (HCA2) which is highly expressed in neutrophil granulocytes. Indeed, deletion of Hca2 in neutrophils abrogated DMF protective effects. Diet had a profound effect on the transcriptional profile of neutrophils and modulated their response to MMF. In summary, DMF required HCA2 on neutrophils as well as permissive dietary effects for its therapeutic action. Translating the dietary intervention into the clinic may improve MS therapy.

overall_design: RNA-Seq of neutrophils isolated from mice fed LA, NC, or HFb diets (5 mice per diet). Isolated neutrophils were incubated with either MMF (100 µM) or PBS as a control for 3 hours.

first: Joanna

last: Kosinska

first: Julian

last: Assmann

first: Julica

last: Inderhees

first: Sven

last: Geisler

first: Axel

last: Küstner

first: Hauke

last: Busch

first: Jörg

last: Heeren

first: Vincent

last: Prevot

first: Ruben

last: Nogueiras

first: Misa

last: Hirose

first: Malte

last: Spielmann

first: Stefan

last: Offermanns

first: Nina

last: Wettschureck

first: Markus

last: Schwaninger


title: Combinatorial therapy to target melanoma resistant to immune checkpoint blockade

city: Cincinnati

first: Nathan

last: Salomonis

state: OH

address: 3333 Burnet Avenue

department: Biomedical Informatics

institute: Cincinnati Children's Hospital

summary: Immune checkpoint blockade (ICB) has revolutionized melanoma therapy, but drug resistance represents a significant limitation. Here we utilize a platform incorporating transcriptomic profiling, high-throughput drug screening (HTDS) and murine models to demonstrate the pre-clinical efficacy of cobimetinib and regorafenib (termed Cobi+Reg) for ICB-resistant melanoma. RNA-Sequencing (RNA-Seq) analysis of ICB-resistant melanomas demonstrated activation of several targetable pathways. HTDS targeting these pathways identified several effective combinations in ICB-resistant patient-derived xenograft (PDX) models. Cobi+Reg emerged as the most promising regimen, with efficacy against distinct molecular melanoma subtypes and following progression on ICB in immunocompetent models. RNA-Seq analysis of Cobi+Reg-treated tumors demonstrated upregulation of antigen presentation machinery, with concomitantly increased activated T cell infiltration. Combining Cobi+Reg with ICB was superior to either treatment in vivo. This analytical platform has identified several effective combinations, presenting Cobi+Reg as a rational therapeutic strategy either following resistance to or combined with ICB for advanced melanoma.

overall_design: Few effective therapeutic options exist following progression on ICB in melanoma. Here, the biology of ICB-resistant melanoma was exploited in order to identify therapeutic vulnerabilities, resulting in the identification of drug combinations that form the basis for rational clinical trial design in the setting of advanced melanoma resistant to ICB.


title: ONECUT2 reprograms neuroendocrine fate and is an actionable therapeutic target in small cell lung cancer.

city: Pamplona

first: Mirja

last: Rotinen


title: Optimized culturing yields high success rates and preserves molecular heterogeneity, enabling personalized screening for high-grade gliomas

city: Rotterdam

first: Martine

last: Lamfers

address: Dr. Molewaterplein 40

department: Neurosurgery

country: Netherlands

institute: Erasmus MC


title: Nat10 regulates the homeostasis of pluripotency and 2C state in mESCs by N4-acetylcytidine mRNA modification [ChIP-seq]

status: Public on May 24 2025

city: Tianjin

first: Guofeng

last: Feng

address: 94 Weijin Road

institute: Nankai University

summary: Mouse embryonic stem cells (mESCs) were isolated from the inner cell mass (ICM), which can prevent an excessive number of cells from transitioning to the 2-cell state and retain only 1%–5% of cells transiently expressing 2C genes such as Zscan4 and ERVs, and 95%–99% of normally expressing pluripotent transcription factors such as Oct4 and Nanog. The homeostasis state is always regulated by epigenetics especially heterochromatin modifications. However, it is still unclear what a specific factor can regulate this homeostasis state. Here we report that N-acetyltransferase 10 (Nat10), the writer for N4-acetylcytidine (ac4C) modification of mRNA, can serve as a new heterochromatin associated protein which can regulate the homeostasis of 2C genes and the expression of pluripotency genes in mESCs through a dual role. Mechanistically, Nat10 deficiency decreased ac4C modification on Oct4 and Esrrb mRNAs, resulting in decreased pluripotency. At the same time, ac4C modification on the heterochromatin component Kap1 mRNA is also reduced, which indirectly leads to a decrease in the binding of the H3K9me3 histone modification to the 2C gene, thereby removing the repression of the 2C genes and allowing the 2C genes to be expressed at high levels. These findings elucidate a novel role of Nat10 in mESCs and reveal new mechanisms linking the 2C program and pluripotency to ac4C modifications in mESCs.

overall_design: About 2×107 of Nat10 CON and Nat10 KD mESCs were first fixed with 1% paraformaldehyde, and then lysed and sonicated to achieve the majority of DNA fragments at 100–500bp. DNA fragments were then enriched by immunoprecipitation with 5 μg H3K9me3 antibody (ab8898, Abcam) and Dynabeads M280 (Life Technologies), rocked at 4 ℃ overnight. The immunoprecipitated material was eluted from the beads by heating for 30 min at 65 ℃. Then, samples were incubated at 65 ℃ for overnight to reverse crosslinking. The samples were then extracted with phenol: chloroform: isoamyl alcohol (25: 24: 1, pH > 7.8) followed by chloroform, ethanol precipitated in the presence of glycogen, and re-suspended in ddH2O. ChIP enriched DNA was used for library construction by Novogene Corporation. Each library was subsequently sequenced, resulting in ~20 million reads (125 bp).


title: Lineage recording in monoclonal gastruloids reveals heritable modes of early development

city: Seattle

first: Chengxiang

last: Qiu

state: WA

address: William H. Foege Hall, 3720 15th Ave NE

department: Genome Sciences

institute: University of Washington

summary: Mammalian stem cells possess a remarkable capacity for self-organization, a property that underlies increasingly sophisticated in vitro models of early development. However, even under carefully controlled conditions, stem cell-derived models exhibit substantial “inter-individual” heterogeneity. Focusing on gastruloids, a powerful model of the early posterior embryo, we sought to investigate the origins of this heterogeneity. To this end, we developed a scalable protocol for generating gastruloids that are monoclonal, i.e. derived from a single mouse embryonic stem cell (mESC). Single cell transcriptional profiling of monoclonal gastruloids revealed extensive inter-individual heterogeneity, with some hardly progressing, others resembling conventional gastruloids but biased towards mesodermal or neural lineages, and yet others bearing cell types rare or absent from conventional polyclonal gastruloids. To investigate this further, we leveraged DNA Typewriter to record the cell lineage relationships among the mESCs from which monoclonal gastruloids originate. Early in the expansion of “founder” mESCsーi.e. prior to induction of the resulting aggregates to form gastruloidsーwe observe clear examples of fate bias or fate restriction, e.g. sister clades that exhibit markedly different cell type compositions. In a separate experiment with DNA Typewriter, we find that founder mESCs that are more closely related are more likely to give rise to monoclonal gastruloids with similar cell type compositions. Our results suggest that fluctuations in the intrinsic states of mESCs are heritable, and shape their descendants’ fates across many cell divisions. Our study also showcases how DNA Typewriter can be used to reconstruct high-resolution, monophyletic cell lineage trees in stem cell models of early development.

overall_design: In this study, we cultured monoclonal gastruloids from mouse embryonic stem cells. Briefly, dissociated mESCs are FACS-sorted followed by density controlled seeding onto a layer of MEF feeder cells. After single cells expand into monoclonal colonies, they are lifted off intact through collagenase IV treatment and gentle agitation, and transferred to a nonadherent plate where they give rise to spherical aggregates. The aggregates are then transferred again to an individual well where they undergo induction via 24-hr exposure to CHIR, a Wnt agonist. Monoclonal gastruloids were then pooled for single-cell profiling and sequencing using either sci-RNA-seq3 or 10X technology.

first: Sam

last: Regalado

first: Junhong

last: Choi

first: Sanjay

last: Kottapalli

first: Beth

last: Martin

first: Wei

last: Chen

first: Hanna

last: Liao

first: Haedong

last: Kim

first: Xiaoyi

last: Li

first: Jean-Benoît

last: Lalanne

first: Nobu

last: Hamazaki

first: Silvia

last: Domcke

first: Jay

last: Shendure


title: Optimized culturing yields high success rates and preserves molecular heterogeneity, enabling personalized screening for high-grade gliomas [scRNA-seq]

summary: Robust and reliable and in vitro models are essential in the discovery of new treatment options for high-grade glioma (HGG), however, establishing successful patient-derived cell cultures has posed significant challenges. We established glioma stem-like cell (GSC) cultures from 114 consecutive HGG specimens, obtained via traditional surgical resection and/or ultrasonic aspiration, using fully-dissociated single cells (single cell-derived, SCD) and partially dissociated tissue fragments (3D-derived, 3DD). Copy number profiling assessed genetic similarities, while single-cell RNA sequencing evaluated transcriptomic heterogeneity. Proof-of-concept personalized drug screening was performed using a panel of approved anti-cancer agents. Higher success rates in establishing GSC cultures were obtained from ultrasonic aspirates (SCD and 3DD) and 3DD surgical samples compared to SCD cultures from surgical samples. Combining these approaches in parallel yielded a 96% success rate. In rare cases, copy number variations in original tumor tissue were lost in culture, leading to discernible changes in cell morphology. Single-cell sequencing revealed greater heterogeneity of transcriptomic clusters in ultrasonic aspiration-derived cultures compared to resection-derived cultures. Our study's protocol supported the screening of 20 anti-cancer agents within a clinically-relevant timeframe in 16 of 18 consecutive HGG samples. Our optimized protocol therefore provides a reliable tool for generating HGG cell cultures which capture the tumors’ molecular characteristics and supports precision medicine applications.

overall_design: Single cell RNA sequencing was performed on a 3DD-GSC culture and a 3D-UA-GSC culture derived from two primary HGG patients. For each patient, cells were collected from each culture type for analysis at passages P1/P2, P5, P11 and P16.

first: Cassandra

last: Posthoorn-Verheul

first: Federica

last: Fabro

first: Ioannis

last: Ntafoulis

first: Chelsea

last: den Hollander

first: Iris

middle: S

last: Verploegh

first: Rutger

last: Balvers

first: Trisha

last: Kers

first: Judith

last: van der Burg

first: Kate

last: Feller

first: Miao-Ping

last: Chien

first: Clemens

last: Dirven

first: Sieger

last: Leenstra


title: Genome wide mapping of NFI and SMAD2/3 binding in senescence

first: Tom

last: Suter

summary: ChIP-seq of NFI or SMAD2/3 was performed in young-quiescent or senescent BJ fibroblasts

overall_design: BJ Fibroblasts were young-quiescent or senescent stages, at which point chromatin was harvested and prepared for ChIP-seq against either NFI or SMAD2/3.  Experiments were replicated, and, in each case, senescent and young cells were compared within each replicate.


title: Nat10 regulates the homeostasis of pluripotency and 2C state in mESCs by N4-acetylcytidine mRNA modification [RNA-Seq]

first: Guofeng

last: Feng

overall_design: mRNA was purified from total RNA using poly-T oligo-attached magnetic beads. Fragmentation was carried out using divalent cations under elevated temperature in NEB Next First Strand Synthesis Reaction Buffer (5×). First strand cDNA was synthesized using formed using DNA Polymerase I and RNase H. Remaining overhangs were converted into blunt ends via exonuclease/polymerase activities. After adenylation of 30 ends of DNA fragments, NEB Next Adaptors with hairpin loop structure were ligated to prepare for hybridization. To select cDNA fragments of preferentially 150–200 bp in length, the library fragments were purified with AMPure XP random hexamer primer and M-MLV Reverse Transcriptase (RNase H). Second strand cDNA synthesis was subsequently per-system (Beckman Coulter, Beverly, USA). Then 3 μL USER Enzyme (NEB, USA) was used with size-selected and cDNA adaptor ligated at 37 ℃ for 15 min followed by 5 min at 95 ℃ prior to PCR. PCR was performed with Phusion High-Fidelity DNA polymerase, Universal PCR primers and Index Primer. At last, PCR products were purified using AMPure XP system and library quality assessed on the Agilent Bioanalyzer 2100 system. Cluster of the index-coded samples was performed on a cBot Cluster Generation System using TruSeq PE Cluster Kit (Illumina) according to the manufacturer’s instructions. After cluster generation, the library preparations were sequenced on an Illumina Hiseq platform.


title: CHD7 Regulates Definitive Endodermal and Mesodermal Development from Human Embryonic Stem Cells [RNAseq]

city: Storrs

first: Laijun

last: Lai

state: Connecticut

address: 1390 Storrs Road

institute: University of Connecticut

summary: CHD7 encodes an ATP-dependent chromodomain helicase DNA binding protein; mutations in this gene lead to multiple developmental disorders, including CHARGE (Coloboma, Heart defects, Atresia of the choanae, Retardation of growth and development, Genital hypoplasia, and Ear anomalies) syndrome. How the mutations cause multiple defects remains largely unclear. Embryonic definitive endoderm (DE) generates the epithelial compartment of vital organs such as the thymus, liver, pancreas, and intestine. In this study, we used clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 technique to delete the CHD7 gene in human embryonic stem cells (hESCs) to generate CHD7 homozygous mutant (CHD7-/-), heterozygous mutant (CHD7+/-), and control wild-type (CHD7+/+) cells. We then investigated the ability of the hESCs to develop into DE and the other two germ layers, mesoderm and ectoderm. We found that deletion of CHD7 led to reduced capacity to develop into DE and mesoderm in a dose-dependent manner. To determine the underlying mechanisms, we compared global gene expression and chromatin accessibility among the hESC-DE cells by RNA sequencing (RNA-seq) and the assay for transposase-accessible chromatin with sequencing (ATAC-seq). We found that loss of CHD7 led to significant changes in the expression and chromatin accessibility of genes associated with several pathways. We identified 40 genes that were highly down-regulated in both the expression and chromatin accessibility in CHD7 deleted hESC-DE cells.  Our results provide new insights into the mechanisms by which CHD7 mutations cause multiple congenital anomalies.

overall_design: We used clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 technique to delete the CHD7 gene in human embryonic stem cells (hESCs) to generate CHD7 homozygous mutant (CHD7-/-), heterozygous mutant (CHD7+/-), and control wild-type (CHD7+/+) cells. We then investigated the ability of the hESCs to develop into definitive endoderm (DE) and the other two germ layers, mesoderm and ectoderm in vitro. We also compared global gene expression among the hESC-DE cells by RNA sequencing (RNA-seq).


title: Cytotoxic NK cells impede response to checkpoint immunotherapy in melanoma with an immune-excluded phenotype

city: Leuven

first: Jean-Christophe

last: Marine

address: Herestraat

institute: VIB-KULeuven

summary: The inability of lymphocytes to infiltrate the tumor nest drastically limits immune checkpoint blockade (ICB) responsiveness. Analyzing the immune landscape of matched pre- and early on-treatment biopsies of melanoma patients undergoing ICB therapy, we observed a significant increase in cytotoxic NK cells in early on-treatment biopsies from non-responders. Spatial OMICs revealed that while NK cells colocalized with CD8 T cells within the tumor bed in responding lesions, they were excluded from the tumor parenchyma in non-responding lesions. Strikingly, depletion of NK cells using a NK1.1 antibody or the FDA-approved drug Daratumumab in a unique melanoma mouse model exhibiting an immune-excluded phenotype unleashed immune infiltration of the tumor core and tumor clearance upon ICB exposure. Mechanistically, we show that NK cells are actively recruited to immune-excluded areas upon ICB exposure via the chemokine receptor CX3CR1 to suppress tumor infiltration and antitumor function of CD8 T cells, possibly by promoting ferroptosis.

overall_design: 2.5 × 10^⁵ NRAS;Ink4a cells were injected subcutaneously into female mice, and 2.5 × 10^⁵ YUMM5.2 cells into male mice (n≥5 per cohort). Tumor volume was monitored, and treatments included anti-PD-1 (200 μg) or IgG2a control (twice weekly, IP). NK cell depletion was performed using anti-NK1.1 (100 μg, starting 3 days before tumor injection, every five days). Mice were sacrificed when control tumors reached ~500 mm^³, except for extended anti-PD-1 + anti-NK1.1 treatment in NRAS;Ink4a.

first: Joanna

last: Pozniak

first: Niccolo

last: Roda


title: DNA Crosslink-induced Transcriptional Changes in Mouse Embryonic Stem Cells

city: Ithaca

first: John

last: Schimenti

state: NY

address: T9 014A Veterinary Research Tower, Box 17

institute: Cornell University

summary: Genome maintenance is of the utmost importance in stem cells, as mutations can be propagated and cause defects in derivative tissues. Many stem cell types display low mutation rates, with embryonic stem cells (ESCs) being a notable example. The bases for this property are unclear but may be achieved by optimization of various processes including high-fidelity DNA repair, cell cycle checkpoint controls, and hypersensitivity to genotoxic insults that trigger cell death. Here, we investigate the transcriptional basis of the unique responses of mouse ESCs (mESCs) to replication stress (RS) using MMC. We find that whereas mESCs lack a strong G2/M checkpoint, they maintain a spindle assembly checkpoint (SAC).

overall_design: Embryonic stem cells (V6.5) of the indicated genotypes (Fancm=fancm-/-, WT=Wild type) were treated with 150nM mitomycin C (MMC) or a vehicle control (NT) for 24 hours before harvest and RNA extraction. Each genotype/condition was repeated for 3 biological replicates.

first: Ryan

last: James

first: Jerry

last: Wang

first: Siddhanth

last: Bhatt

first: Sophie

last: Shadid

first: Daryl

last: Phuong