title: Tff1-expressing Tregs in lung prevents exacerbation of Bleomycin-induced pulmonary fibrosis (RNA-Seq)

city: Suita

first: Daisuke

last: Motooka

state: Osaka

address: 3-1, Yamadaoka

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

summary: Bleomycin (BLM) induces lung injury, leading to inflammation and pulmonary fibrosis. Regulatory T cells (Tregs) maintain self-tolerance and control host immune responses. However, little is known about their involvement in the pathology of pulmonary fibrosis. Here we show that a unique Treg subset that expresses trefoil factor family 1(Tff1) emerges in the BLM-injured lung. These Tff1-expressing Tregs (Tff1-Tregs) were induced by IL-33. Moreover, although Tff1 ablation in Tregs had no impact, selective ablation of Tff1-Tregs using an intersectional genetic method promoted pro-inflammatory features of macrophages in the injured lung and exacerbated the fibrosis. Taken together, our study revealed the presence of a unique Treg subset expressing Tff1 in BLM-injured lungs and their critical role in the injured lung to ameliorate fibrosis.

overall_design: GFP+CD4+ T cells were sorted from lung and spleen of BLM-administrated FDG-mice at day 15.

first: Masaaki

last: Okamoto

last: Okuzaki

first: Masahiro

last: Yamamoto


title: Molecular Signatures of CB-6644 inhibition of the RUVBL1/2 complex in Multiple Myeloma [ATAC-seq]

status: Public on Aug 19 2024

city: Morgantown

first: Gangqing

last: Hu

state: West Virginia

address: 64 Medical Center Drive

department: MicroBiology, Immunology, and Cell Biology

country: USA

institute: West Virginia University

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Multiple Myeloma is the second most hematological cancer. RUVBL1 and RUVBL2 forms a subcomplex of many chromatin remodeling complexes implicated in cancer progress. As an inhibitor specific to the RUVBL1/2 complex, CB-6644 exhibits remarkable anti-tumor activity in xenograft models of Burkitt’s lymphoma and multiple myeloma (MM). In this work, we defined transcriptional signatures corresponding to CB-6644 treatment in MM cells and determined underlying epigenetic changes in terms of chromatin accessibility.CB-6644 upregulated biological processes related to interferon response and downregulated those linked to cell proliferation in MM cells. Transcriptional regulator inference identified ATF4/CEBP and E2Fs as regulators for downregulated genes and MED1 and MYC as regulators for upregulated genes. CB-6644-induced changes in chromatin accessibility occurred mostly in non-promoter regions. Footprinting analysis identified transcription factors implied in modulating chromatin accessibility in response to CB-6644 treatment, including ATF4/CEBP and IRF4. Lastly, integrative analysis of transcription responses to various chemical compounds of the molecular signature genes from public gene expression data identified CB-5083, a p97 inhibitor, as a synergistic candidate with CB66-44 in MM cells, but experimental validation refuted this hypothesis.

overall_design: To characterized the regulome (chromatin accessibility) signatures of CB-6644 treatment in MM cells. We inferred the underling epigenetic regulatory mechanisms by analyzing chromatin accessibility using Omni-ATAC in MM.1S, treated with 120nm of CB-6644 (72 hours) and compared to DMSO-treated cells.

first: Weijun

last: Yi

first: Sebastian

middle: A

last: Dziadowicz

first: Rachel

last: Mangano

first: Lei

last: Wang

first: Joseph

last: McBee

first: Steven

middle: M

last: Frisch

first: Lori

last: Hazlehurst

first: Donald

last: Adjeroh


title: Genome-wide analysis of the transcriptional regulation by ERRα and ERRγ in SH-SY5Y cells

city: Tokyo

first: Kaoru

last: Sato

address: 35-2 Sakae-cho, Itabashi-ku

country: Japan

institute: Tokyo Metropolitan Institute of Gerontology

type: Expression profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Alternative splicing events driven by altered levels of Gemin5 undergo translation

city: Madrid

first: Encarnacion

last: Martinez-Salas

state: Madrid

address: Nicolás Cabrera 1

country: Spain

institute: Centro de Biologia Molecular Severo Ochoa, CSIC-UAM

summary: We show that upregulation of Gemin5 can be detrimental as it modifies the steady state of mRNAs and enhances alternative splicing (AS) events of genes involved in a broad range of cellular processes. RNA-Seq analyses of the mRNA associated with polysomes in cells with high levels of Gemin5 revealed that a significant fraction of the differential AS events was enriched in polysomes.

overall_design: In independent biological samples of HEK293 cells expressing Gemin5 (G5) and control cells (Ct), individual cell lysates were subjected to polysomal fractionation. In each case, the mRNAs present in cytoplasmic cell lysates (Input) and polysomes (POL) were extracted and analyzed by RNAseq.

first: Rosario

last: Francisco-Velilla

first: Salvador

last: Abellan

first: JuanAntonio

last: Garcia Martin

first: JuanCarlos

last: Oliveros


title: Premature aging effects on COVID-19 pathogenesis: new insights from mouse models

status: Public on Aug 28 2024

city: Guangzhou

first: Haoyu

last: Wu

address: Kai Yuan Avenue 190

country: China

institute: GIBH

summary: Aging is identified as a significant risk factor for severe coronavirus disease-2019 (COVID-19), often resulting in profound lung damage and mortality. Yet, the biological relationship between aging, aging-related comorbidities, and COVID-19 remains incompletely understood. This study aimed to elucidate the age-related COVID19 pathogenesis using a Hutchinson-Gilford progeria syndrome (HGPS) mouse model with humanized ACE2 receptors. Pathological features were compared between young, aged, and HGPS hACE2 mice following SAR-CoV-2 challenge. We demonstrated that young mice display robust interferon response and antiviral activity, whereas this response is attenuated in aged mice. Viral infection in aged mice results in severe respiratory tract bleeding, likely contributing a higher mortality rate. In contrast, HGPS hACE2 mice exhibit milder disease manifestations characterized by minor immune cell infiltration and dysregulation of multiple metabolic processes. Comprehensive transcriptome analysis revealed both shared and unique gene expression dynamics among different mouse groups. Collectively, our studies evaluated the impact of SARS-CoV-2 infection on progeroid syndromes using a HGPS hACE2 mouse model, which holds promise as a valuable tool for investigating COVID-19 pathogenesis in individuals with premature aging.

overall_design:
To investigate the pathological outcomes of lungs in HGPS hACE2 mice upon SARS-CoV-2 infection, we infected young (2 month old), aged (14 month old), and HGPS (2 month old) hACE2 mice with the original strain of the coronavirus from Wuhan (WIV04) , and harvest RNA samples of lungs at 1, 3, 5, 7 day-post-infection (dpi).
we then performed RNA-seq, and compared the transcriptome changes of lungs between young, old, and HGPS hACE2 mice.

first: Wu

last: Haoyu

first: Hong

last: Guangliang

first: Chen

last: Jiekai


title: Targeting SWI/SNF ATPases reduces cell invasiveness and plasticity in core transcription factors-addicted neuroblastoma

city: New Brunswick

first: Xiyuan

last: Zhang

state: NJ

address: 195 Little Albany St.

department: Pediatric Hematology/Oncology

institute: Rutgers Cancer Institute

summary: Neuroblastoma (NB) comprises mesenchymal (MES) and adrenergic (ADRN) subtypes, and the cell identity is determined by core transcription factors (TFs). However, the binding mechanism of these TFs to DNA and the epigenetic mechanisms governing NB plasticity remain unclear. In this study, we investigated the impact of targeting SWI/SNF ATPases with SMARCA2/4 dual degraders on NB cells. Our results revealed that depletion of SWI/SNF ATPases compacted cis-regulatory elements, diminished enhancer activity, and displaced core TFs (MYCN, HAND2, PHOX2B, and GATA3) from DNA, suppressing transcriptional programs linked to plasticity and invasiveness. These findings underscore the pivotal role of SWI/SNF ATPases in driving NB progression, positioning them as promising therapeutic targets.

overall_design: Patient-derived xenograft (PDX) NB cells exhibit intertumoral heterogeneity, comprises MES-type and ADRN-type cells. SJNBL012407_X1 is a PDX cell line, which was maintained in neural stem-cell culture media (SCM) as spheres. To investigate the impact of depleting SWI/SNF ATPases on the emergence of heterogenous NB subpopulations, we cultured the SJNBL012407_X1 cells in 10% FBS containing media overnight, then treated the cells with DMSO (as a vehicle control) or 250 nM ACBI1 (SMARCA2/4 dual degrader)  for 4 days. We used single-cell RNA sequencing (scRNA-seq) to analyze the diversity of PDX cells cultured in stem cell media, FBS-containing media that treated with or without ACBI1.

first: Zhihui

last: Liu

first: Carol

middle: J

last: Thiele


title: The transcriptional signatures of Caenorhabditis elegans dauer larvae

status: Public on Aug 15 2024

city: Buffalo

first: Johnny

last: Cruz Corchado

state: NY

address: Elm & Carlton Streets Buffalo,L3-L125

department: Cell Stress

institute: Roswell Park Comprehensive Cancer Institue

summary: Both plasticity and robustness are pervasive features of developmental programs.  The dauer in Caenorhabditis elegans is an alternative to the third larval stage of the nematode and is an example of phenotypic plasticity. The dauer is an arrested, hypometabolic state that undergoes dramatic changes in gene expression compared to conspecifics that continue development, and can be induced by several adverse environments or genetic mutations that act as independent and parallel inputs into the larval developmental program. However, given the different genetic or environmental triggers that can induce dauer, gene expression in dauer larvae could be invariant or vary depending on the larvae’s route into dauer entry; this question has not been examined. Here we use RNA-sequencing to characterize gene expression in dauer larvae induced to arrest development in response to different stimuli.  By assessing the variance in the expression levels of all genes and computing the Spearman's rank-order correlation of gene expression within several Gene Ontologies (GO) and gene networks, we find that the expression patterns of most genes, except for those that act in specific defense and metabolic pathways, are strongly correlated between the different dauer larvae, suggestive of transcriptional robustness.  We speculate that the transcriptional robustness of core dauer pathways allows for the buffering of variation in the expression of genes involved in their response to the environment, allowing the different dauers to be better suited to survive in and exploit different niches.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for  C. elegans dauer of the following gennotypeswild-type dauers (N2; Bristol) ;daf-2 (e1370) III,daf-7(e1372) III ;ilc-17.1 (syb5296) X  ;cep-1 (cep-1oe) [unc-119 (ed4); gtIs1 [CEP-1::GFP + unc-119 (+)]

last: Cruz

first: Abhishiktha

last: Godthi

first: Veena

last: Prahlad


title: Translation efficiency in the pancreatic tumor in response to ketogenic diet and targeting eIF4E phosphorylation

status: Public on Aug 14 2024

city: San Francisco

first: Haojun

last: Yang

state: CA

address: 1450 3rd Street

institute: UCSF

summary:
To investigate the translatome in the pancreatic tumor inresponse to ketogenidc diet and the combination of ketogenic diet with an inhibitor targeting eIF4E phosphorylation.
We generated pancreatic tumor xenograftmice model, treated mice with regular chow, ketogenic diet or ketogenic diet with eFT508, an inhibitor targeting eIF4E phosphorylation. Then we performed Poly-RNAseq on the tumor isolated from mice upon different treatments.

overall_design: We isolated the tumor isolated from T3M4 injected nude mice on different treatments. We then seperated highly translated mRNA and low translated mRNA using sucrose gradient and perform comparative gene expression profiling analysis of RNA-seq data for those high tanslated and low translated mRNA.

first: Davide

last: Ruggero


title: Cancer-associated DNA Hypermethylation of Polycomb Targets Requires DNMT3A Dual Recognition of histone H2AK119 Ubiquitination and the Nucleosome Acidic Patch

city: New York City

first: Kristjan

middle: H.

last: Gretarsson

state: New York

institute: Columbia University

type: Methylation profiling by high throughput sequencing


title: Towards the DNA methylation haplotype map of 11 common solid cancers

status: Public on Aug 23 2024

city: Shanghai

first: Leiqin

address: 320 Yueyang Road

institute: CAS Center for Excellence in Molecular Cell Science (CEMCS)

summary: Aberrant DNA methylation is an epigenetic hallmark of cancer, known to play an essential role in cancer initiation, progression and drug resistance. Traditional analyses focus on mean methylation that measures the aggregated signal across a group of cells and neglect intra-sample heterogeneity. Sequencing-based techniques such as whole genome bisulfite sequencing (WGBS) are now widely used to measure DNA methylation on each read at single-nucleotide resolution. A single read fragment stems from a single chromosome of a single cell and represents a DNA methylation haplotype (mHap). In this study, we have profiled 110 fresh frozen tumor samples that cover 11 most common solid cancer types using WGBS and constructed a comprehensive DNA methylation haplotype map.

overall_design:
We have focused on 11 most common solid cancer types, i.e., head and neck squamous cell carcinoma (HNSC), Thyroid carcinoma (THCA), Non-small cell lung cancer (NSCLC), Breast carcinoma (BRCA), Esophageal carcinoma (ESCA), liver hepatocellular carcinoma (LIHC), Stomach adenocarcinoma (STAD), Pancreatic carcinoma (PAAD), Colon adenocarcinoma (COAD), Ovarian serous adenocarcinoma (OV), Cervical and endocervical cancers (CESC). These cancer types cumulatively represent 85% of all cancer cases worldwide. Whole-genome bisulfite sequencing (30x) was performed on 9 to 11 fresh-frozen tumor samples for each cancer type, resulting in 110 WGBS samples covering 11 cancer types.
**Raw data not provided due to patient privacy concerns**

first: Zhiqiang

first: Yuyang

last: Hong

first: Jiantao

last: Shi


title: Translation efficiency in the mouse liver upon 24h chow or ketogenic diet fed

summary:
To investigate the translatome in the ketogenic diet fed condition and the role of phosphoralated eIF4E in it
We performed polysome sequencing on chow fed and 24h ketogenic diet fed wild type mice and chow fed eIF4E S209A/S209A mice

overall_design: We separated highly translated mRNA and low translated mRNA using sucrose gradient and performed comparative gene expression profiling analysis of RNA-seq data for those high translated and low translated mRNA from fed and fasted WT livers, and fed eIF4E S209A/S209A livers


title: Translation efficiency in the mouse liver upon 24h fasting

summary:
To investigate the translatome in the fasted condition and the role of phosphoralated eIF4E in it
We performed polysome sequencing on fed and fasted wild type mice and fasted eIF4E S209A/S209A mice

overall_design: We seperated highly translated mRNA and low translated mRNA using sucrose gradient and perform comparative gene expression profiling analysis of RNA-seq data for those high tanslated and low translated mRNA from fed and fasted WT livers, and fasted eIF4E S209A/S209A livers


title: HDAC6 Inhibition Disrupts HDAC6-P300 Interaction Reshaping the Cancer Chromatin Landscape (ATAC-Seq)

city: Rome

first: daniele

last: santoni

address: via dei taurini 19

country: Italy

institute: IASI - CNR

summary: Histone deacetylases (HDACs) are crucial regulators of gene expression, DNA synthesis, and cellular processes, making them essential targets in cancer research. HDAC6, specifically, influences protein stability and chromatin dynamics. Despite HDAC6's potential therapeutic value, its exact role in gene regulation and chromatin remodeling needs further clarification. This study examines how HDAC6 inactivation influences lysine acetyltransferase P300 stabilization and subsequent effects on chromatin structure and function in cancer cells. We employed the HDAC6 inhibitor ITF3756, siRNA, or CRISPR/Cas9 gene editing to inactivate HDAC6 in different epigenomic backgrounds. Constantly, this inactivation led to significant changes in chromatin accessibility, particularly increased acetylation of histone H3 lysines 9, 14, and 27 (ATAC-seq and H3K27Ac ChIP-seq analysis). Transcriptomics, proteomics, and gene ontology analysis revealed gene changes in cell proliferation, adhesion, migration, and apoptosis. Significantly, HDAC6 inactivation altered P300 ubiquitination, stabilizing P300 and leading to downregulating genes critical for cancer cell survival. Our study highlights the substantial impact of HDAC6 inactivation on the chromatin landscape of cancer cells and suggests a role for P300 in contributing to the anticancer effects. The stabilization of P300 with HDAC6 inhibition proposes a potential shift in therapeutic focus from HDAC6 itself to its interaction with P300. This finding opens new avenues for developing targeted cancer therapies, improving our understanding of epigenetic mechanisms in cancer cells.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for H3K27ac in B16F10 cells WT vs KO_HDAC6.

first: Michela

last: Gottardi Zamperla

first: Barbara

last: Illi

first: Veronica

last: Barbi

first: Chiara

last: Cencioni

first: Daniele

last: Santoni

first: Stella

last: Gagliardi

first: Maria

last: Garofalo

first: Gabriele

last: Zingale

first: Irene

last: Pandino

first: Diego

last: Sbardella

first: Lina

last: Cipolla

first: Simone

last: Sabbioneda

first: Antonella

last: Farsetti

last: Ripamonti

first: Gianluca

last: Fossati

first: Christian

last: Steinkühler

first: Carlo

last: Gaetano

first: Sandra

last: Atlante


title: Multi-omics profiling of longitudinal samples reveals early genomic changes in follicular lymphoma

status: Public on Aug 20 2024

city: Oslo

first: Ankush

last: Sharma

address: 3rd Floor,  Ullernchausseen 70

department: Department of Cancer Immunology, Institute for Cancer Research

country: Norway

institute: Oslo University Hospital

summary: Follicular lymphoma (FL) is the most common indolent type of B-cell non-Hodgkin lymphoma. Advances in treatment have improved overall survival, but early relapse or transformation to aggressive disease is associated with inferior outcome. To identify early genetic events and track tumor clonal evolution, we performed multi-omics analysis of 94 longitudinal biopsies from 44 FL patients; 22 with transformation (tFL) and 22 with relapse without transformation (nFL). Deep whole-exome sequencing confirmed recurrent mutations in genes encoding epigenetic regulators (CREBBP, KMT2D, EZH2, EP300), with similar mutational landscape in nFL and tFL patients. Calculation of genomic distances between longitudinal samples revealed complex evolutionary patterns in both subgroups. CREBBP and KMT2D mutations were identified as genetic events that occur early in the disease course, and cases with CREBBP KAT domain mutations had low risk of transformation. Gains in chromosomes 12 and 18 (TCF4), and loss in 6q were identified as early and stable copy number alterations. Identification of such early and stable genetic events may provide opportunities for early disease detection and disease monitoring. Integrative analysis revealed that tumors with EZH2 mutations exhibited reduced gene expression of numerous histone genes, including histone linker genes. This might contribute to the epigenetic dysregulation in FL.

overall_design:
To investigate differential expression of genes between follicular lymphoma patient samples harbouring mutations in genes encoding epigentic regulators (CREBBP, KMT2D, EZH2) vs. patient samples without such mutations. We included sorted tonsillar B cell subsets (naive, memory and GC B cells) from four healthy donor tonsils as comparisons.
***Submitters state that raw data will not be submitted to a repository due to GDPR ***

first: Baoyan

last: Bai

first: Jillian

middle: F

last: Wise

first: Daniel

last: Vodák

first: Sigve

last: Nakken

first: Yngvild

middle: N

last: Blaker

first: Marianne

last: Brodtkorb

first: Vera

last: Hilden

first: Gunhild

last: Trøen

first: Weicheng

last: Ren

first: Susanne

last: Lorenz

first: Michael

middle: S

last: Lawrence

first: Ola

last: Myklebost

first: Eva

last: Kimby

first: Qiang

middle: P

last: Hammarström

first: Leonardo

last: Meza-Zepeda

first: Klaus

last: Beiske

first: Erlend

middle: B

last: Smeland

first: Eivind

last: Hovig

first: Ole

middle: C

last: Lingjærde

first: Harald

last: Holte

first: June

middle: H

last: Myklebust


title: RNA-seq analysis of ABA-treated Nicotiana benthamiana seedlings

status: Public on Aug 13 2024

city: Valencia

first: Pedro L

last: Rodriguez

address: CPI-UPV, C/ de l'Enginyer Fausto Elio, s/n, Algirós

institute: IBMCP (CSIC-UPV)

summary: Analysis of abscisic acid (ABA) perception and signaling in crops is complicated by the multigenic nature of the PYR/PYL/RCAR ABA receptor family and the reported functional redundancy, which is a formidable challenge, particularly in polyploid plants. Therefore, few molecular genetics studies have been performed in crop species to study the contribution of ABA signaling to balance plant growth and adaptation to the environment. Our data reveal ABA-responsive genes in Nicotiana benthamiana, an allotetraploid biotech crop, and provide a molecular framework  for gene expression analyses aimed at optimizing plant growth and environmental adaptation in agriculture.  To obtain the genome-wide transcriptional effect of ABA in Nb WT plants, we performed RNA-seq studies as described in experimental procedures. Differential gene expression analysis between mock- and ABA-treated seedlings was done with DESeq2 and genes with an absolute value of log2 fold change (log2FC) > 1 or (log2FC) < -1 and p-adjusted value (padj) < 0.05 were selected

overall_design:
N. benthamiana seedlings (10 d.a.g.) were transferred from plates to flashes with 2 ml of liquid Murashige and Skoog (MS medium) for ten days in a growth chamber at 22ºC under long-day conditions (16 h light/8 h dark). Before treatment, the medium was refreshed and supplemented with either 0.1% DMSO (mock) or 10 μM ABA. Samples were collected after 3 hours of incubation, and the experiment was conducted with three independent biological replicates.
We performed genome-wide expression profiling analysis using data obtained from RNA-seq of the three biological replicates
Comparative analysis of transcriptional profiles using RNA-seq data for mock and ABA treatments (WT genotype and pent1, LAB strain)

first: Jonatan

last: Illescas-Miranda

first: Javier

last: Forment

first: Pedro

middle: L


title: Ageing-Related Changes to H3K4me3, H3K27ac and H3K27me3 in Purified Mouse Neurons

status: Public on Aug 21 2024

city: Hobart

first: Phillippa

middle: Claire

last: Taberlay

state: Tasmania

address: 17 Liverpool St

department: College of Health and Medicine

country: Australia

institute: University of Tasmania

summary: Neurons are central to lifelong learning and memory, but ageing disrupts their morphology and function, leading to cognitive decline. Although epigenetic mechanisms are known to play crucial roles in learning and memory, neuron-specific genome-wide epigenetic maps into old age remain scarce, often being limited to whole-brain homogenates and confounded by glial cells. Here, we mapped H3K4me3, H3K27ac, and H3K27me3 in mouse neurons across their lifespan. This revealed stable H3K4me3 and global losses of H3K27ac and H3K27me3 into old age. We observed patterns of synaptic function gene deactivation, regulated through the loss of the active mark H3K27ac, but not H3K4me3. Alongside this, embryonic development loci lost repressive H3K27me3 in old age. This suggests a loss of a highly refined neuronal cellular identity linked to global chromatin reconfigu-ration. Collectively, these findings indicate a key role for epigenetic regulation in neurons that is inextricably linked with ageing.

overall_design: H3K4me3 and H3K27ac ChIP-seq from 3, 6, 12, and 24m old Mus musculus forebrain neuronal nuclei sorted by FACS. Biological replicates (2-4 samples) for H3K4me3 and H3K27ac, and total input controls are included as fastq files, and analysed bigwigs.

first: Katherine

last: Giles

first: Andrew

last: Phipps

first: Jake

last: Cashion

first: Shannon

last: Huskins

first: James

last: Vickers

first: Timothy

middle: R

last: Mercer

first: Mark

middle: D

last: Robinson

first: Adele

last: Woodhouse


title: Molecular Profiling Defines Three Subtypes of Synovial Sarcoma

city: Solna

first: Felix

last: Haglund de Flon

state: Stockholm

address: Solnavägen 1

department: Oncology-Pathology

country: Sweden

institute: Karolinska Institute

summary: Synovial Sarcomas (SS) are characterized by the presence of the SS18::SSX fusion gene, which protein product induce chromatin changes through remodeling of the BAF complex (BRG1/BRM-associated factor complex), which leads to widespread alterations in gene expression that drive tumor development and progression. However, there is no established molecular model which explain the variation in clinical behavior of SS. To elucidate the genomic events that drive phenotypic diversity in SS, we performed RNA and targeted DNA sequencing on 91 tumors from 55 patients. Our results were verified by proteomic analysis, public gene expression cohorts and single-cell RNA sequencing. Transcriptome profiling identified three distinct SS subtypes resembling the known histological subtypes: SS subtype I and was characterized by hyperproliferation, evasion of immune detection and a poor prognosis. SS subtype II and was dominated by a vascular-stromal component and had a significantly better outcome. SS Subtype III was characterized by biphasic differentiation, increased genomic complexity and immune suppression mediated by checkpoint inhibition, and poor prognosis despite good responses to neoadjuvant therapy. Chromosomal abnormalities were an independent significant risk factor for metastasis. KRT8 was identified as a key component for epithelial differentiation in biphasic tumors, potentially controlled by OVOL1 regulation. Our multi-omics analysis revealed that the biological and genetic variability between these subtypes, including key transcriptome patterns and secondary genomic events such as copy number alterations, were associated with long-term outcomes. Our findings explain the histological grounds for SS classification and indicate that a significantly larger proportion of patients have high risk tumors (corresponding to SS subtype I) than previously believed.

overall_design: Investigate gene expression patterns of primary and metastatic synovial sarcomas from human patients.

first: Yi

last: Chen


title: Effect of treatment with 5uM of the type I PRMT inhibitor MS023 dissolved in DMSO for 3 days and of a stably transduced PRMT1 specific dox-inducible knockdown over 4 days in ccRCC cell line models RCC243 and 786-0

status: Public on Aug 16 2024

city: Toronto

first: Laurie

last: Ailles

state: ON

address: 101 College St.

department: Princess Margaret Cancer Centre

country: Canada

institute: University Health Network

summary: To investigate the effect of pharmacological inhibition of the type I PRMTs in ccRCC cell lines. We treated one patient derived cell line (RCC243) and one commercially available line (786-0) in duplicate with 5uM MS023 for 3 days vs. 0.1% DMSO alone. We then performed gene expression profiling analysis using data obtained from RNA-seq. Each cell line was treated as a biological replicate. Similarly, to investigate the genetic knockdown of PRMT1 using the same cell lines, stably transduced dox-inducible PRMT1 targeting shRNA's.  Cell lines were treated with or with out 1ug/mL dox for 4 days in triplicate.  Gene expression profiling analysis was then performed using data obtained from RNA-seq.

overall_design:
Comparative gene expression profiling analysis of RNA-seq data for RCC243 and 786-0 treated vs untreated with 5uM MS023 for 3 days - Each cell line treated in duplicate and data pooled for analysis.
Comparative gene expression profiling analysis of RNA-seq for RCC243 and 786-0 stably transduced with dox-inducible PRMT1 targeting shRNAs - Each cell line was dox induced (1ug/mL) in triplicate and comparison was made to non-induced controls. Cell line data pooled for analysis.

first: Joseph

last: Walton


title: Tff1-expressing Tregs in lung prevents exacerbation of Bleomycin-induced pulmonary fibrosis (scRNA-Seq)

first: Daisuke

last: Motooka

summary: Bleomycin (BLM) induces lung injury, leading to inflammation and pulmonary fibrosis. Regulatory T cells (Tregs) maintain self-tolerance and control host immune responses. However, little is known about their involvement in the pathology of pulmonary fibrosis. Here we show that a unique Treg subset that expresses trefoil factor family 1(Tff1) emerges in the BLM-injured lung. These Tff1-expressing Tregs (Tff1-Tregs) were induced by IL-33. Moreover, although Tff1 ablation in Tregs had no impact, selective ablation of Tff1-Tregs using an intersectional genetic method promoted pro-inflammatory features of macrophages in the injured lung and exacerbated the fibrosis. Taken together, our study revealed the presence of a unique Treg subset expressing Tff1 in BLM-injured lungs and their critical role in the injured lung to ameliorate fibrosis.

overall_design: Living CD45+ cells were sorted from naïve lung or BLM-injured lung (day 15) of WT mice. The sorted samples from three individual mice under identical conditions were pooled together, ensuring an equal cell count across the combined sample.

first: Masaaki

last: Okamoto

last: Okuzaki

first: Masahiro

last: Yamamoto


title: Impact of Chromatin Immunoprecipitation Crosslinking Conditions on Selective Genomic Interactions by NF‐κB Family Members

status: Public on Aug 11 2024

city: Los Angeles

first: Stephen

last: Smale

state: Ca

address: 6-730 MRL 675 Charles E. Young Drive South

department: MIMG

institute: University of California Los Angeles

summary: The ability to capture protein: DNA interactions in a quantitative, high-resolution, and temporal manner in the cell context is crucial for elucidating the mechanisms underlying transcriptional regulation.  Chromatin immunoprecipitation followed by sequencing (ChIP-seq) is a widely used technique that enables the study of histone modifications and genome-wide interactions of transcription factors and chromatin regulators. Although ChIP-seq generates a vast amount of information, crosslinking conditions need to be carefully evaluated to minimize the capture of non-specific interactions. In this study, we conduct an in-depth analysis to evaluate the impact of crosslinking conditions on ChIP-seq results for two NF-kB family members, RelA and c-Rel. We titrate two commonly used chemical crosslinkers, formaldehyde and DSG. We demonstrate that while raising the crosslinking concentrations increases the total number of binding sites and the strength of the binding signal, high crosslinking conditions lead to an abundance of interactions detected at locations that lack anticipated motifs or annotations to potential NF-kB target genes. Our findings emphasize the importance of a comprehensive evaluation of ChIP-seq results and a cautious evaluation of chemical crosslinking conditions. Moreover, we compare ChIP-seq to CUT&TAG, a newer technique that does not require crosslinking. This comparison revealed strongly overlapping genomic interactions between the two methods. However, both methods display evidence of a large number of non-specific signals that exhibit little overlap between the two methods.

overall_design: To investigate the effect of crosslinking on the outcome of ChIP-seq, we performed ChIP-seq for cRel and RelA with varying concentrations of both DSG and PFA.

first: Allison

last: Daly

last: Schiffman


title: Characterization of ACTN4 as a novel antiviral target against SARS-CoV-2

city: Wuhan

first: zhu

last: miao

address: jin long street

institute: Wuhan institution of virology

summary: this study unveil the pivotal role of ACTN4 in SARS-CoV-2 infection, offering novel insights into the intricate interplay between the virus and host cells, and reveal two potential candidates for future anti SARS-CoV-2 drug development.

overall_design: Total RNAs were extracted from Huh7 cells with or without SARS-CoV-2 infection.After treating with 20 mM ZnCl2, 10% RNA fragments was saved as “Input”, while the rest was proceeded to m6A immunoprecipitation (IP) using anti m6A Abs.The stranded RNA sequencing library was constructed by KC DigitalTM Stranded mRNA Library Prep Kit for Illumin,following with enrichment, quantification and finally sequencing on DNBSEQ T7 sequencer with PE150 model.


title: Postprandial exercise regulates tissue-specific triglyceride uptake through bifurcated regulation of tissue LPL by Angiopoietin-like proteins [RNA-seq]

status: Public on Aug 08 2024

first: Yan

last: Wang

state: China

address: Luojiashan, wuchang, wuhan

institute: College of Life Sciences, Wuhan University

summary: We performed RNA-seq in the liver of wild-type (WT) mice in response to a single bout aerobic exercise. Exercise induced the downregulation of insulin signaling pathway.

overall_design: Before exercise, mice were synchronized with food intake for three days by daytime feeding (9:00 a.m. to 15:00 p.m.) and night time fasting (15:00 p.m. to 9:00 a.m.). On each day, three hours after food intake (12:00 p.m.), mice were acclimated to the treadmill by putting in their respective lanes with the belt unmoving and shock grids on for 10 minutes. Then the mice were forced to run for 5 minutes at 10 m/min. On day 4, a single bout aerobic exercise for 3hours was performed at postprandial period. Mice from 2 different conditions (Sedentary, Exercise) were subjected to RNA sequencing.


title: Embryo Mortality with an Impaired Interferon-Tau Release Mounts a Massive T-Helper Immune Response in Lactating Holstein-Friesian cows

status: Public on Aug 09 2024

city: Miles City

first: Carolina

middle: Lucila

last: Gonzalez-Berrios

state: Montana

address: 243 Fort Keogh Road

institute: USDA-ARS

summary:
Embryo mortality (EM) contributes to infertility, but its exact mechanism is poorly understood. It was hypothesized that bovine EM pregnancies have impaired conceptus-derived interferon-tau (IFNT) release and action and are associated with altered transcriptome responses.
The objective was to discover transcriptome response in EM tissues (endometrium [ENDO], corpus luteum [CL] and peripheral blood mononuclear cells [PBMC]) in lactating Holstein-Friesian cows.
Two experiments (E1, E2) in day 16 pregnant (exposed to semen; E1:n=13, E2:n=15) or non-pregnant (NP; not exposed to semen; E1:n=7, E2:n=7) cows were completed.
Uterine flushings (UF) and tissues were collected.
Pregnant cows were also classified based on conceptus morphology and appearance for EM (E1:n=5, E2:n=6) or normal (N) conceptuses (E1:n=8, E2:n=9).
Conceptuses and tissues were RNA sequenced and analyzed.
The N conceptuses were longer (P<0.05) than EM conceptuses. The IFNT protein concentrations in UF were greater in N compared to EM and NP cows in E1 but not for E2.
The ENDO conjugated ISG15 concentrations were greater in N (E1) than EM and NP cows but N (E2) were only greater than NP not EM cows.
The major up-regulated canonical pathway in EM conceptuses was T helper 1 (Th1) and Th2.
The ENDO had a massive increase in interferon stimulated genes in N and EM compared to NP cows.
Estradiol-associated genes were up-regulated in EM compared to N ENDO for E2.
The PBMC in E1 reflected up-regulation of genes associated Th1 immune responses in EM compared to N cows.
Luteolytic pathways were upregulated in EM CL compared to N cows.
This disruption of maternal recognition of pregnancy in EM pregnancies entails a massive T helper immune response within the conceptus, estradiol re-modelling of the ENDO, abnormal immune system in PBMC, luteolysis cascade in CL and, potentially, loss of pregnancy.

overall_design: Comparison of the gene expression profiles within conceptus and maternal tissues (endometrium, corpus luteum and peripheral blood mononuclear cells) in normal pregnancies compared to pregnancies with embryo mortality occurring at the preimplantation stage (day 16).

first: Leticia

last: Sinedino

first: Bethany

middle: E

last: Liebig

first: Hanah

last: Georges

first: Jeanette

middle: V

last: Bishop

first: Hana

last: Van Campen

first: Milton

middle: G

last: Thomas

first: Thomas

last: Hansen


title: Defining Cell Type-specific Immune Responses in Allergic Contact Dermatitis by Single-cell Transcriptomics

city: Xiamen

first: Lingjuan

last: Zhang

address: Xiangan South Street

institute: Xiamen University

summary: In brief, skin resident cells were isolated from ear skin of mice and 10x Genomics single cell RNAseq was applied to identify distinct cell populations. Low quality cells and outliers were discarded, and only ～27600 viable cells were used for downstream analysis. Unsupervised clustering and gene expression were visualized with the Seurat on R studio, and assignment of cell clusters was based on expression of validated marker genes. Subsequent in-depth analysis was assisted by GENE DENOVO Inc (Guangzhou, China). Overall, We found that hapten application in pre-sensitized mice resulted in rapid infiltration of IFNγ-producing T cells, IL4- and IL13-producing basophils, and IL1β-producing neutrophils and macrophages. Sc-RNAseq identified a cluster of dermal fibroblasts enriched with IFNγ pathway signature and expressed high level of Cxcl9/10. In vitro, IFNγ–primed dFBs shifted the T cell Th1/Th2 polarization balance towards the Th1 phenotype through a CXCR3-dependent mechanism. Topical application of birch sap extract selectively blocked T cell-dFB interaction, suppressed Th1 cell activation and alleviated skin inflammation in the ACD model.

overall_design: Ear skin biopsies were collected from 2 groups, Healthy control and DNFB-induced ACD  topically applied  DNFB-induced ACD 7~8 weeks old ICR mice.

first: Ling-juan


title: Gene expression profiling of human cell

city: Bethesda

first: Pamela

middle: Gehron

last: Robey

state: MD

address: 30 Convent Dr

department: NIDCR

institute: NIH

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

overall_design: Refer to individual Series


title: The Human Circular RNA CircHMGCS1 Sponges miR-4521 to Aggravates Diabetes-Induced Vascular Endothelial Dysfunction Through Upregulating ARG1

city: hangzhou

first: Ming

state: zhejiang

address: Zijingang Campus of Zhejiang University,Yuhangtang Road No.388

institute: School of Biosystems Engineering and Food Science

type: Non-coding RNA profiling by high throughput sequencing

summary: Cardiovascular diseases pose a significant global health burden. Our study investigates the pivotal role of circular RNAs (circRNAs) and MicroRNAs (miRNAs) in diabetes-induced endothelial dysfunction, a critical aspect of cardiovascular diseases. CircHMGCS1 (hsa_circ_0008621) emerges as a specific circRNA that effectively targets miR-4521. The upregulation of CircHMGCS1 acts as a miR-4521 sponge, facilitating the expression of arginase 1 and accelerating the progression of diabetes-induced endothelial dysfunction. Furthermore, intricate associations among circRNA, miRNA, and mRNA are revealed, illuminating the underlying molecular mechanisms involved in diabetes-induced vascular endothelial dysfunction. These findings significantly advance our understanding of cardiovascular diseases, providing potential therapeutic targets for intervention.

overall_design: In order to simulate the regulatory effect of diabetes on non-coding RNA in vascular endothelium in vitro, We used Human Umbilical Vein Endothelial Cells (HUVECs) to culture them in high fat/high glucose (PAHG) medium for 24h, and The medium contained 25 mM glucose and 250 μM saturated free fatty acid (FFA) palmitate were repeated three times, followed by total RNA extraction for RNA-SEQ analysis

first: Wei

last: Chen


title: The Circular RNA HMGCS1 Sponges miR-4521 to Aggravate Diabetes-Induced Vascular Endothelial Dysfunction Through Upregulating ARG1

summary: Type 2 diabetes is recognized as one of the primary contributors to cardiovascular diseases, with vascular endothelial dysfunction (VED) being a crucial mechanism underlying its development. In this study, we present the aberrant expressions of CircHMGCS1 and miR-4521 in diabetes-induced VED. Overexpression of CircHMGCS1 or silencing of miR-4521 expedited the onset of diabetes and aggravated VED. Mechanistically, CircHMGCS1 upregulated ARG1 by sequestering miR-4521, resulting in the inhibition of vascular NO secretion, enhanced adhesion molecules expression of VCAM1, ICAM1, ET-1, and increased ROS generation. Consequently, these events accelerated the impairment of vascular endothelial function. These findings underscore the physiological roles of CircRNA and miRNA in cardiovascular diseases triggered by diabetes and suggest that modulating the expression of CircHMGCS1 and miR-4521 could serve as a potential strategy for preventing endothelial cell dysfunction and the development of diabetes-associated cardiovascular diseases.


title: Host-Microbe Interactions in the Nasal Cavity of Dogs with Chronic Idiopathic Rhinitis

city: Fort Collins

first: Steven

last: Dow

state: CO

address: 300 W Drake Rd

department: Clinical Sciences

institute: Colorado State University

summary: Chronic rhinitis (CR) is a frustrating clinical syndrome in dogs and our understanding of the disease pathogenesis in is limited.  Increasingly, host-microbe interactions are considered key drivers of clinical disease in sites of persistent mucosal inflammation such as the nasal and oral cavities. Therefore, we applied next generation sequencing tools to interrogate abnormalities present in the nose of dogs with CR and compared immune and microbiome profiles to those of healthy dogs.  Host nasal cell transcriptomes were evaluated by RNA sequencing, while microbial communities were assessed by 16S rRNA sequencing. Correlation analysis was then used to identify significant interactions between nasal cell transcriptomes and the nasal microbiome and how these interactions were altered in animals with CR. Notably, we observed significant downregulation of multiple genes associated with ciliary function in dogs with CR, suggesting a previously undetected role for ciliary dysfunction in this syndrome. We also found significant upregulation of immune genes related to the TNF-a and interferon pathways. The nasal microbiome was also significantly altered in CR dogs, with overrepresentation of several potential pathobionts. Interactome analysis revealed significant correlations between bacteria in the genus Porphyromonas and the upregulated host inflammatory responses in dogs with CR, as well as defective ciliary function which was correlated with Streptococcus abundance. These findings provide new insights into host-microbe interactions in a canine model of CR and indicate the presence of potentially causal relationships between nasal pathobionts and the development of nasal inflammation and ciliary dysfunction.

overall_design: Nasal swabs were collected under general anesthesia by gently swabbing the distal third of the nasal cavity using 6-inch PurFlock Ultra, sterile flocked collection devices (Puritan Medical Products, Guilford, ME). Swabs (2 swabs obtained per each nostril) were placed in 15 ml conical tubes (Corning Inc. Corning, NY) containing RNA later (ThermoFisher Scientific, Waltham, MA) and stored at 4C prior to RNA and DNA extraction. To extract RNA from nasal cells, swabs were vortexed for 1 min at high speed to dislodge cells, PBS was then added to RNA later at a ratio of 1:5. Cells were pelleted, then processed for RNA extraction using Qiagen RNeasy micro kit (Qiagen, Hilden, Germany) following manufactures instructions or DNA extraction as described below.

first: Lyndah

last: Chow

first: Zhe


title: BIO 300 attenuates whole blood transcriptome changes in mice exposed to total-body radiation

city: Minneapolis

first: Artur

last: Serebrenik

state: MN

address: 7760 France Ave S, Suite 920

institute: Humanetics Corporation

summary: Development of radiation medical countermeasures under the U.S. Food and Drug Administration Animal Rule requires the capability to translate an effective animal to human drug dose. One method of human dose translation is using a biomarker and determining drug doses that modulate the biomarker to the desired level. BIO 300 Oral Powder (BIO 300) is a prophylactic radiation medical countermeasure that is currently being developed following the Animal Rule. The present study aimed to identify biomarkers that can be used for human dose conversion by conducting transcriptomics of whole blood collected from BIO 300-treated CD2F1 mice in the presence and absence of total-body irradiation (TBI). Mice were treated with vehicle or 50, 100 or 200 mg/kg BIO 300, twice daily, for 6 days. Whole blood samples were collected 24 and 48 h after the last dose of the drug. Animals were also treated with vehicle or 200 mg/kg BIO 300 for 6 days prior to 9.2 Gy TBI 24 h after the last dose, with blood collection 24 h after irradiation. RNA sequencing demonstrated 100 – 200 mg/kg doses caused significantly more differential gene expression at 48 h post-drug dose compared to 50 mg/kg. Pathway analysis of the transcriptome profile from vehicle-treated/radiation-exposed mice revealed that many inflammatory signaling pathways were activated in these animals. Signaling pathways enriched in BIO 300-treated/TBI mice were involved in cellular stress and immune response and were predicted to be inhibited. We also identified pathways that were in inverse states (activation/inhibition) in vehicle and BIO 300-treated mice with TBI. There were two pathways specifically activated in vehicle-treated animals with TBI and inhibited in BIO 300-treated animals (with or without TBI), pathogen-induced cytokine storm signaling and S100 family signaling. In comparison to previously published human data, pulmonary fibrosis idiopathic signaling and wound healing signaling were enriched in healthy volunteers treated with BIO 300 and BIO 300-treated mice with TBI. In all, four signaling pathways were identified that were activated in vehicle-treated animals exposed to total body radiation but inhibited in irradiated animals or healthy humans treated with BIO 300. These pathways should be explored to identify potential biomarkers of BIO 300 that can be used for human dose translation.

overall_design: To investigate gene expression changes in unirradiated and irradiated mice treated with vehicle or one of three doses (low, med, high) of BIO 300. Changes in BIO 300-treatment groups were compared relative to vehicle groups. Each group included six mice that were terminal bled to collect whole blood for RNA sequencing.

first: Oluseyi

last: Fatanmi

first: Stephen

last: Wise

first: Sarah

last: Petrus

first: Michael

last: Kaytor

first: Vijay

last: Singh


title: Direct mapping of G-quadruplex anchored 3D chromatin structure via ViCAR

status: Public on Aug 12 2024

city: Cambridge

first: Somdutta

last: Dhir

address: Robinson Way

department: CRUK-CI

country: United Kingdom

institute: University of Cambridge

summary: We developed ViCAR (viewpoint HiCAR), a technique that uses antibody-based capture of genome features and their interacting chromatin domains. ViCAR uncovers numerous cell-type-specific G4-dependent enhancer-promoter interactions and is easily extended to other features such as histone marks. ViCAR represents a practical and powerful tool to explore the relationship between epigenetic marks and 3D genome interactomes.

overall_design: ViCAR to capture genome architecture associated with G4s and histone marks in K562 and hESCs.

first: S

middle: M

last: Flynn

last: Balasubramanian


title: Multi-omics profiling of retinal pigment epithelium reveals enhancer-driven activation of RANK-NFATc1 signaling in traumatic proliferative vitreoretinopathy [RNA-seq]

city: Tianjin

first: Yumei

last: Lu

address: No22.Qixiangtai Rd.,Heping Dist

institute: Tianjin Medical University

summary: During the progression of proliferative vitreoretinopathy (PVR) following ocular trauma, previously quiescent retinal pigment epithelium (RPE) transition into a state of rapid proliferation, migration, and secretion. The elusive molecular mechanisms behind these changes have hindered the development of effective pharmacological treatments, presenting a pressing clinical challenge. In this study, by monitoring the dynamic changes in chromatin accessibility and various histone modifications, we charted the comprehensive epigenetic landscape of RPE in traumatic PVR. Coupled with transcriptomic analysis, we reveled a robust correlation between enhancer activation and the upregulation of the PVR-associated gene programs. Furthermore, by constructing transcription factor regulatory networks, we identified the aberrant activation of enhancer-driven RANK-NFATc1 pathway as PVR advanced. Importantly, we demonstrated that intraocular interventions, including nanomedicines inhibiting enhancer activity, gene therapies targeting NFATc1 and antibody therapeutics against RANK pathway, effectively mitigate PVR progression. Together, our findings elucidate the epigenetic basis underlying the activation of PVR-associated genes during RPE cell fate transitions and offer promising therapeutic avenues targeting epigenetic modulation and the RANK-NFATc1 axis for PVR management.

overall_design: RNA-seq was performed on retinal pigment epithelium cells from normal and PVR mice.

first: Mengyu

last: Liao


title: Multi-omics profiling of retinal pigment epithelium reveals enhancer-driven activation of RANK-NFATc1 signaling in traumatic proliferative vitreoretinopathy [ATAC-seq]

overall_design: ATAC-seq was performed on retinal pigment epithelium cells from normal and PVR mice.


title: Multi-omics profiling of retinal pigment epithelium reveals enhancer-driven activation of RANK-NFATc1 signaling in traumatic proliferative vitreoretinopathy [ChIP-seq]

overall_design: ChIP-seq was performed on retinal pigment epithelium cells from normal and PVR mice.


title: Type I interferon signaling pathway enhances immune-checkpoint inhibition in KRAS mutant lung tumors [scRNA-seq]

city: Madrid

first: Laura

last: Serrano-Ron

address: Calle Emilio Ortuño 35 1B

department: Bioinformatics Unit

institute: CNIO

summary: Lung cancer is the leading cause of cancer mortality worldwide. KRAS oncogenes are responsible for at least a quarter of lung adenocarcinomas, the main subtype of lung cancer. After four decades of intense research, selective inhibitors of KRAS oncoproteins are finally reaching the clinic. Yet, their effect on overall survival is limited due to the rapid appearance of drug resistance, a likely consequence of the high intratumoral heterogeneity characteristic of these tumors. In this study, we have attempted to identify those functional alterations that result from KRAS oncoprotein expression during the earliest stages of tumor development. Such functional changes are likely to be maintained during the entire process of tumor progression regardless of additional co-occurring mutations. Single-cell RNA sequencing analysis of murine alveolar type 2 cells expressing a resident Kras oncogene revealed impairment of the type I interferon pathway, a feature maintained throughout tumor progression. This alteration was also present in advanced murine and human tumors harboring additional mutations in the p53 or LKB1 tumor suppressors. Restoration of type I interferon (IFN) signaling by IFN-beta or constitutive active stimulator of interferon genes (STING) expression had a profound influence on the tumor microenvironment, switching them from immunologically “cold” to immunologically “hot” tumors. Therefore, enhancement of the type I IFN pathway predisposes KRAS mutant lung tumors to immunotherapy treatments, regardless of co-occurring mutations in p53 or LKB1.

overall_design: We sorted alveolar type II (AT2) cells four weeks after the expression of oncogenic KRAS(G12V) and compared to KRAS wild type AT2 control cells by scRNAseq to detect early alterations in lung tumorigenesis.

first: Fernando

last: Fernández-García

first: Mariano

last: Barbacid

first: Monica

last: Musteanu


title: Gene expression profiling of human induced pluripotent stem cell (hiPSC)-derived chondrospheroids

first: Pamela

last: Robey

summary: Differentiated chondrocytes from human induced pluripotent stem cells (hiPSCs) have the potential to permanently restore damaged cartilage in arthritic joints, yet chondrocyte hypertrophy is a major barrier for translational therapy.  We developed a serum-free hiPSC differentiation strategy that inhibited, then activated BMP signaling in a purified SOX9+ subpopulation that naturally detaches from monolayer cultures (termed “chondrospheroids”).  Transcriptional profiles of chondrospheroids at days 14, 28, and 42 were measured using bulk RNA sequencing, with hiPSC and sclerotome cells included as controls.

overall_design:
Three chondrospheroid replicates derived from the NCRM NL5 hiPSC line at three specific timepoints of differentiation (days 14, 28, and 42) were processed for bulk RNA sequencing.  Sclerotome cells (day 6) and hiPSCs (day 0) were included as controls.
contributor: NIDCD/NIDCR Genomics and Computational Biology Core

middle: J

last: Gadomski

first: Byron

middle: W

last: Mui

first: Raphael

last: Gorodetsky

first: Sriram

last: Paravastu

first: Joseph

last: Featherall

first: Li

last: Li

first: Abigail

last: Haffey

first: Jae-Chun

last: Kim

first: Sergei

middle: A

last: Kuznetsov

first: Kathryn

last: Futrega

first: Astar

last: Lazmi-Hailu

first: Randall

middle: K

last: Merling

first: Daniel

last: Martin

first: Andrew

last: McCaskie


title: Time- and cell-specific activation of BMP signaling restrains chondrocyte hypertrophy

summary: Differentiated chondrocytes from populations of human bone marrow stromal cells including skeletal stem cells (hBMSCs/SSCs) and induced pluripotent stem cells (hiPSCs) have the potential to restore damaged cartilage, yet chondrocyte hypertrophy is a barrier for translational therapy. However, hBMSCs/SSCs attached to a hyaluronic acid-coated fibrin microbead scaffold (HyA-FMBs) produce cartilage that resists hypertrophy. We show that HyA-FMBs initially suppress Bone Morphogenic Protein (BMP) signaling early in chondrogenic differentiation of hBMSCs/SSCs, then restore BMP signaling in a stable chondrogenic population enriched for Insulin-like Growth Factor Binding Protein 5 and Matrix Gla Protein. We subsequently developed a serumfree hiPSC differentiation strategy that inhibited, then activated BMP signaling in a purified subpopulation derived from ?chondrospheroids? that exhibited stable expression of Type II Collagen, Aggrecan (ACAN), and Lubricin (PRG4) and minimal e  xpression of Type X Collagen. Overall, timed, and selective BMP signaling restrains chondrocyte hypertrophy, supporting the potential of chondrospheroid-derived cells for clinical utility in osteoarthritis.

overall_design:
Gene expression profiling of human bone marrow stromal cells/skeletal stem cells (hBMSCs/SSCs) attached to fibrin microbead scaffolds coated with hyaluronic acid (HyA-FMBs).
Differentiated chondrocytes from human bone marrow stromal cells including skeletal stem cells (hBMSCs/SSCs) have the potential to permanently restore damaged cartilage in arthritic joints, yet chondrocyte hypertrophy is a major barrier for translational therapy.  With chondrogenic differentiation, hBMSCs/SSCs undergo hypertrophy in vitro and mineralization in vivo, leading to inferior fibrocartilage and bone formation.  However, hBMSCs/SSCs attached to a fibrin microbead scaffold coated with hyaluronic acid (HyA-FMBs) produce hyaline-like cartilage for up to 28 weeks in vivo.  Therefore, the goal of the present study was to examine the signaling pathways that govern the development of hypertrophic-resistant chondrocytes using the HyA-FMB model system.  hBMSCs/SSCs attached to HyA-FMBs formed organoids during chondrogenic differentiation in vitro, and these organoids were digested at days 1, 3, 5, and 10 for single-cell RNA sequencing.  hBMSCs/SSCs attached to HyA-FMBs were also transplanted, and tissues were harvested after 8 weeks for single-cell RNA sequencing.
Passage 2 hBMSC/SSCs from three donors were used in these studies.  Organoids generated from hBMSCs/SSCs during days 1, 3, 5, and 10 of chondrogenic differentiation were dissociated and analyzed by scRNAseq using 10x Genomics platform.  Naïve hBMSCs/SSCs (no chondrogenic differentiation) were included as controls. Transplanted hBMSCs/SSCs attached to HyA-FMBs were also dissociated after 8 weeks for comparison.

Gene expression profiling of human induced pluripotent stem cell (hiPSC)-derived chondrocytes supplemented with TGFb and BMP ligands.
Human induced pluripotent stem cell (hiPSC)-derived sclerotome differentiated in chondrogenic medium with TGFb forms organoids that retain strong Collagen I expression, scattered Aggrecan expression, and absence of PRG4 expression.  However, hiPSC-derived sclerotome differentiated with BMP2 and GDF5 (i.e., in the formation of chondrospheroids) forms cartilage tissues with homogenous Aggrecan and Collagen 2 expression with uniform PRG4 expression, which remains expressed upon transplantation at the joint site in NSG mice and SRG rats.  To confirm the role of BMP activation in strengthening this chondrogenic phenotype, we performed single cell RNA sequencing of tissues exposed to TGFb alone or supplemented with BMP ligands.

contributor: NIDCD/NIDCR Genomics and Computational Biology Core

first: Stephen

middle: S


title: Type I interferon signaling pathway enhances immune-checkpoint inhibition in KRAS mutant lung tumors [cell lines]

first: Laura

last: Serrano-Ron

overall_design: We performed RNAseq of KP and KL murine lung tumor cell lines 48 hours after inducing the expression of IFN-beta or constitutively active STING (V154M mutation) and compared to non-expressing cells.

first: Fernando

last: Fernández-García

first: Mariano

last: Barbacid


title: Prdm1 Positively Regulates Liver Group 1 ILCs Cancer Immune Surveillance and Preserves Functional Heterogeneity [RNA-Seq]

first: Youwei

last: Wang

state: --- Select One ---

address: 92 Weijin Road

department: Medical College

institute: Tianjin University

summary: Group 1 innate lymphoid cells (ILCs) comprise conventional natural killer (cNK) cells and type 1 innate lymphoid cells (ILC1s). The main functions of liver cNK cells and ILC1s not only include directly killing target cells but also regulating local immune microenvironment of the liver through the secretion of cytokines. Uncovering the intricate mechanisms by which transcriptional factors regulate and influence the functions of liver cNK cells and ILC1s, particularly within the context of liver tumors, presents a significant opportunity to amplify the effectiveness of immunotherapies against liver malignancies. Using Ncr1-drived conditional knockout mouse model, our study reveals the regulatory role of Prdm1 in shaping the composition and maturation of cNK cells. Although Prdm1 did not affect the killing function of cNK cells in an in vivo cytotoxicity model, a significant increase in cancer metastasis was observed in Prdm1 knockout mice. Interferon-gamma (IFN-γ), granzyme B, and perforin secretion decreased significantly in Prdm1 deficient cNK cells and liver ILC1s. scRNA sequencing data also provided evidences that Prdm1 maintains functional subsets of cNK cells and liver ILC1s and facilitates communications between cNK cells, liver ILC1s and macrophages. The present study unveiled a novel regulatory mechanism of Prdm1 in cNK cells and liver ILC1s, showing promising potential for developing innovative immune therapy strategies against liver cancer.

overall_design: RNA-seq of sorted control or Prdm1 knockout mouse splenic NK cells.

first: Jitian

last: He

first: Le

last: Gao

first: Peiying


title: Type I interferon signaling pathway enhances immune-checkpoint inhibition in KRAS mutant lung tumors [LCM]

overall_design: We performed laser capture microdissection (LCM) of small lung adenomas driven by oncogenic KRAS alone (K) or in combination with loss of p53 (KP) or LKB1 (KL) tumor suppressors.We also isolated normal adjacent tissue (NL). We then subjected the samples to RNAseq.

first: Fernández-García

last: Fernando

first: Barbacid

last: Mariano

first: Musteanu

last: Monica


title: Comprehensive transcriptomic analysis reveals stress physiology behind differential gene expression in vitrified–warmed and fresh mouse blastocysts

status: Public on Aug 07 2024

city: Taichung

first: Chi-Ying

last: Lee

address: No.30-6, Sec. 1, Changping Rd., Beitun Dist., Taichung 406, Taiwan

country: Taiwan

institute: Taiwan IVF center, Lee's women's hospital

summary: Blastocyst vitrification has improved embryo transfer methods, leading to higher implantation success rates and better pregnancy outcomes in subsequent frozen embryo transfer cycles. In this study, we used mouse blastocysts as a model to simulate the transcriptional changes caused by vitrifying human blastocysts and to further investigate their effects. Utilizing a human vitrification protocol, we implanted both vitrified and fresh embryos into mice and observed a higher implantation success rate for the vitrified embryos. Transcriptomic analysis revealed that vitrified-warmed blastocysts exhibited differential gene expression primarily associated with thermogenesis, chemical carcinogenesis-reactive oxygen species, oxidative phosphorylation, immune response, and MAPK-related signaling pathways. To validate the next-generation sequencing results, we performed reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR), which confirmed increased expression of genes such as Cdk6 and Nfat2, and decreased expression of genes such as Dkk3 and Mapk10. Among these pathways, immune responses and MAPK-related signaling pathways indicate that embryos need to communicate with the external environment, likely through microRNAs. We used gene-microRNA interaction predictions to anticipate potential embryonic microRNA expressions and conducted an analysis of embryonic microRNA expression. Twelve microRNAs exhibited expression patterns consistent with the predicted results, potentially affecting uterine epithelial cell adhesion, trophectoderm development, invasive capacity, or immune responses. Our findings suggest that vitrification induces transcriptomic changes in mouse blastocysts, and even small changes in gene expression can increase implantation success.

overall_design: Embryos from B6CBF1 mice were cultured in human tubal fluidfor 3days. Embryo transfer was performed 2.5 days following mating. Six vitrified–warmed blastocysts were transferred to the right uterine horn in the same recipient. The same number of fresh blastocystswere transferred to the left uterine horn of pseudopregnant recipients. The mice were euthanized4days after embryo transfer.

first: Han-Ni

last: Tsai

first: En-Hui

last: Cheng

first: Tsung-Hsien

first: Pin-Yao

last: Lin

first: Chun-I

first: Maw-Sheng


title: The diadenosine tetraphosphate hydrolase ApaH is crucial for Pseudomonas aeruginosa pathogenicity

city: Rome

first: Francesco

last: Imperi

state: Rome

address: Viale Marconi, 446

department: Department of Scienze

institute: Roma Tre University

summary: The role of the intracellular signaling molecule diadenosine tetraphosphate (Ap4A) has not so far been investigated in Pseudomonas aeruginosa. To fill this gap, we performed RNA sequencing (RNA-seq) analysis to compare the transcriptome of the reference strain P. aeruginosa PAO1 and an isogenic deletion mutant (ΔapaH) which is deficient in the Ap4A hydrolysing enzyme ApaH and accumulates high intracellular levels of Ap4A.

overall_design: 2 different groups with 3 biological replicates each. Group A: wild type strain. Group B: apaH mutant strain.

first: Matteo

last: Cervoni

first: Davide

last: Sposato

first: Paolo

last: Visca

first: Livia

last: Leoni

first: Giordano

last: Rampioni


title: Multi-omics profiling of retinal pigment epithelium reveals enhancer-driven activation of RANK-NFATc1 signaling in traumatic proliferative vitreoretinopathy.

first: Yumei

last: Lu


title: Transcriptomic analysis of the acuK and the acuM deletion mutants of Talaromyces marneffei

city: Sri Phum, Mueang

first: Tanaporn

last: Wangsanut

state: Chiang Mai

address: 110 Inthawarorot Rd

country: Thailand

institute: Chiang Mai University

summary: Identification of global target genes that their expression levels are dependent on the AcuK and the AcuM transcription factors from T. marneffei. Results provided information that the AcuK and AcuM are global regulators important for adaptation and defense against harsh environmental conditions, such as the response to iron limitation, utilization of alternative carbon sources, and antioxidation.

overall_design: Conidia of T. marneffei ATCC18224, ∆acuK and ∆acuM strains were inoculated at a final concentration of 108 cell/mL in 250 mL synthetic dextrose (SD) broth (1%glucose , 7 uM ferrous sulfate) and incubated at 25°C for 36 hours for the generation of mycelium mass. The cultures were harvested by centrifugation at 4500 rpm for 10 min. Approximately 2 g wet weight was homogenized in a TRIzol reagent (Invitrogen, Life Technologies, Carlsbad, CA, USA) by using a bead beater (Biospec, Bartlesville, OK, USA). Total RNA was isolated following the manufacturer’s protocol. The RNA was digested with 1 U/µL DNase I to eliminate the residual DNA and purified by isopropanol precipitation. RNA concentrations were determined with a spectrophotometer (Nanodrop 2000: Thermo Scientific, Waltham, MA, USA). RNA integrity and purity were tested by agarose gel electrophoresis. Library preparation and RNA sequencing were performed by a service from Novogene Co., Ltd. (Hong Kong).

first: Monsicha

last: Pongpom


title: Establishment of Pig Embryonic Stem Cells using Simplified Serum-free media and Adaptation of Feeder-free conditions

city: Cheongju

first: Sang-Hwan

last: Hyun

state: Please select your state

address: 1 Chungdae-ro, Seowon-gu

department: Veterinary Medical Center and College of Veterinary Medicine

country: South Korea

institute: Chungbuk national university

summary: Most porcine embryonic stem cells (pESC) derived from in vitro blastocysts Still require complex media compositions and feeder layers which make difficulty to adapt their broad and routine use. We derived pESC using simplified culture media (FIW) consisting of small molecules FGF2, IWR-1, and WH-4-023 in serum-free medium. We established several pESC-FIW lines capable of single cell passaging with short cell doubling time (about 12 hours). These cells expressed pluripotency markers, OCT4, SOX2, and NANOG, as well as cell surface markers, SSEA1, SSEA4, and TRA-1-60. pESC-FIW showed stable proliferation rate and normal karyotype even after long-term culture over 50 passages. Established pESC showed formative characteristics based on the negative expression for the naive marker KLF4 and primed marker T, whereas strong expression for the formative marker OTX2 with high levels of related genes. Transcriptome analysis showed that pESC-FIW had similar cellular identities to reported pESC maintained in complex media formulations and had characteristics of gastrulating epiblast cells. In addition, pESC-FIW could be maintained for multiple passages in a feeder-free condition on fibronectin coated plate using mTeSR™, a commercial medium for feeder-free culture. These results indicate that the canonical WNT (IWR-1) inhibition and the SRC inhibition (WH-4-023) are sufficient to establish pESC capable of single cell passaging and feeder-free expansion under the serum-free condition. Easy to maintain-pESC can be a tool for gene editing application useful for agriculture and biomedicine, and lineage commitment studies.

overall_design:
To examine cellular identity of pESC-FIW, we prepared RNA-seq libraries in duplicate from three lines, two PA-ESC lines (PA_1_pESC_FIW and PA_3_pESC_FIW) and one IVF-ESC line (IVF_1_pESC_FIW).
First, we performed a comparative transcriptome analysis of the bulk RNA-seq data of pESC-FIW and those of other pESC lines from previous reports
We further compared the transcriptome with pig pre-implantation embryos

first: Hyerin

last: Choi

first: Dongjin

last: Oh

first: Mirae

last: Kim

first: Joohyeong

first: Lian

last: Cai

first: Jawad

last: Ali

first: Haomiao

last: Zheng

first: Eunhye


title: Prdm1 Positively Regulates Liver Group 1 ILCs Cancer Immune Surveillance and Preserves Functional Heterogeneity

first: Youwei

last: Wang

overall_design: Liver CD45+ cells were FACS sorted from control or Prdm1 knockout mouse and analyzed using scRNA-seq

first: Jitian

last: He

first: Le

last: Gao

first: Peiying


title: Persistent changes in nociceptor translatomes govern hyperalgesic priming in mouse models

status: Public on Aug 06 2024

city: Dallas

first: Theodore

last: Price

state: TX

address: 800 W Campbell Rd

institute: The University of Texas at Dallas

summary: Hyperalgesic priming is a model system that has been widely used to understand plasticity in painful stimulus-detecting sensory neurons, called nociceptors. A key feature of this model system is that following priming, stimuli that do not normally cause hyperalgesia now readily provoke this state. We hypothesized that hyperalgesic priming occurs due to reorganization of translation of mRNA in nociceptors. To test this hypothesis we used paclitaxel treatment as the priming stimulus and translating ribosome affinity purification (TRAP) to measure persistent changes in mRNA translation in Nav1.8+ nociceptors. TRAP sequencing revealed 161 genes with persistently altered mRNA translation in the primed state. We identified Gpr88as upregulated, and Metrn as downregulated. We confirmed a functional role for these genes wherein a GPR88 agonist causes pain only in primed mice, and established hyperalgesic priming is reversed by Meteorin. Our work demonstrates that altered nociceptor translatomes are causative in producing hyperalgesic priming.

overall_design: Paclitaxel (Sigma Aldrich, Y0000698) was dissolved in a 1:1 solvent mixture of Kolliphor EL (Sigma-Aldrich) and ethanol. Female Nav1.8-TRAP mice were administered paclitaxel diluted in sterile Dulbecco’s phosphate buffered saline (DPBS; Thermo Scientific) or vehicle solvent intraperitoneally at a dosage of 4 mg/kg every other day, resulting in a cumulative dosage of 16 mg/kg. The animals were tested fro mechanical hypersensitivity until they reached back to baseline measurement. The animals were euthanized, and all dorsal root ganglion were dissected and trap sequencing was performed. The input and the immunoprecipitate fractions were bulk sequenced.

first: Ishwarya

last: Sankaranarayanan

first: Diana

middle: T

last: Ferreira


title: Unravelling tRNA Fragments in DENV Pathogenesis: Insights from RNA Sequencing

status: Public on Aug 10 2024

city: New Delhi

first: Bhupendra

middle: Kumar

last: Verma

state: Delhi

address: Ansari Nagar

department: Biotechnology

country: India

institute: All India Institute of Medical Sciences

summary: tRNA-derived small RNA including tRNA-derived fragments (tRFs) and tRNA halves (tiRNAs) plays significant roles in the various molecular mechanisms that underlie certain human diseases. of the generation of tRFs/tiRNAs and their potential roles during Dengue virus (DENV) infection is not yet known.  Here, we have performed small RNA sequencing to identify the generation and alterations in tRF expression profiles of DENV infected Huh7 cells. Our results suggest that 733 tiRNAs/tRFs were found to be differentially expressed during DENV infection. Interestingly, 3’tRF population were found to be upregulated and i-tRF population were found to be downregulated. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) were performed to analyze the impact of differentially expressed tRFs on DENV pathogenesis. Our results suggest the significant involvement in transcriptional regulation via RNA polymerase II promoter and metabolic pathways. Overall, our study contributes significantly to our understanding of the roles played by tRFs in the complex dynamics of DENV infection.

overall_design: we conducted a comprehensive RNA sequencing analysis to decipher the distinctive patterns of tRFs/tiRNAs in DENV-infected Huh7 cell lines relative to mock-infected controls. For RNA sequencing, triplicate samples were provided of Mock infected Huh7 cells, 0hr DENV infected Huh7 cells (i.e. 2 hrs post adsorption) and48hrs DENV ijfected Huh7 cells.

first: Deeksha

last: Madhry

first: Varsha

last: Meena

first: Kiran

last: Kumari

first: Riya

last: Roy


title: Transcriptional analysis of efferocytosis in mouse skin wounds

city: NEW HAVEN

first: Will

last: Krause

state: CT

address: 20 Winchester Ave., Unit one, Unit, Unit one

department: Molecular, Cellular, and Developmental Biology

institute: Yale University

summary: Defects in apoptotic cell clearance, or efferocytosis, can cause inflammatory diseases and prevent tissue repair due in part, to a key role of efferocytosis in inducing a pro-repair transcriptional program in phagocytic cells like macrophages. While the cellular machinery and metabolic pathways involved in efferocytosis have been characterized, the precise efferocytic response of macrophages is dependent on the identity and macromolecular cues of apoptotic cells, and the complex tissue microenvironment in which efferocytosis occurs. Here, we find that macrophages undergoing active efferocytosis in mid-stage mouse skin wounds in vivo display a pro-repair gene program, while efferocytosis of apoptotic skin fibroblasts in vitro induces an immature/inflammatory transcription response. These data provide a resource for understanding how the skin wound niche influences macrophage efferocytosis and will be useful for future investigations that define the role of efferocytosis during tissue repair

overall_design: In order to better understand efferocytosis function in the skin, we used both an in-vivo and in-vitro model of efferocytosis, followed by FACS isolation of Eff+ and Eff- cells. In-vivo samples were macrophages isolated from day 3 wounds on LysMCreER/mTmG mice. In-vitro samples were bone marrow derived macrophages treated with labled apoptotic fibroblasts and sorted by FACS.

first: Diane

last: King

first: Valerie

last: Horsley


title: Immunologic features of nontuberculous mycobacterial pulmonary disease based on spatially resolved whole transcriptomes

city: Seoul

first: Jaemoon

last: Koh

state: Seoul

address: Jongnogu 101 Seoul National University hospital, B1, Department of Pathology

institute: Seoul National University Hospital

summary: The immunologic features of nontuberculous mycobacterial pulmonary disease (NTM-PD) are largely unclear. This study investigated the immunologic features of NTM-PD using digital spatial profiling techniques. Lung tissues obtained from six patients with NTM-PD between January 1, 2006, and December 31, 2020, at Seoul National University Hospital were subjected to RNA sequencing. Cores from the peribronchial and fibrotic stromal areas were stained with CD3, CD68, and DNASyto13, and gene expression at the whole-transcriptome level was quantified using PCR amplification and Illumina sequencing. Lung tissues from four patients with bronchiectasis collected during the same period were used as controls. The RNA sequencing results were validated using immunohistochemistry (IHC) in another cohort (30 patients with NTM-PD and 15 patients with bronchiectasis). NTM-PD exhibited distinct gene expression patterns in T cells and macrophages. Gene set enrichment analysis revealed that pathways related to antigen presentation and processing were upregulated in NTM-PD, particularly in macrophages. Macrophages were more prevalent and the expression of genes associated with the M1 phenotype (CD40 and CD80) was significantly elevated. Although macrophages were activated in the NTM-PD group T cell activity was unaltered. Notably, expression of the costimulatory molecule CD28 was decreased in NTM-PD. IHC analysis showed that T cells expressing Foxp3 or TIM-3, which facilitate the regulatory functions of T cells, were increased. From these, NTM-PD exhibits distinct immunologic signatures characterized by the activation of macrophages without T cell activation.

overall_design: RNA sequencing of lung tissue from patients underwent surgical resection because of nontuberculous mycobacterial lung disease or bronchiectasis using Illumina TruSeq Stranded mRNA Library prep and sequenced on Illumina HiSeq 2500.

first: Nakwon

last: Kwak

first: Sehui

last: Kim

first: Joong-Yub

first: Jae-Joon

last: Yim


title: Effect of Calorie Restriction in the Congenic Mating Types of Cryptococcus neoformans

city: Stony Brook

first: Natalia

last: Kronbauer de Oliveira

state: NY

address: 100 Nicolls Rd

institute: Stony Brook University

summary: Cryptococcus neoformans (Cn) is an opportunistic fungal microorganism that causes life-threatening meningoencephalitis. Calorie restriction is an intervention that extends the lifespan of Cn. The mating types of Cn have striking differences in prevalence in both environmental and clinical settings. This is hypothesized to be due to differences in stress responses between mating types. Using RNAseq, we investigated how the two mating types respond to the stress of starvation during calorie restriction.

overall_design: For this analysis, KN99alpha and KN99a were grown overnight in SM media containing 2% glucose and calorie restriction (CR) media containing 0.05% glucose. The RNA was isolated using a QIAGEN Rneasy kit. Then the transcriptome was analyzed for differentially expressed genes.

last: Oliveira

first: Somanon

last: Bhattacharya

first: Bettina

middle: C

last: Fries


title: Spatially clustered type I interferon responses at injury borderzones

city: La Jolla

first: Kevin

middle: Robert

state: CA

address: 9500 Gilman Dr.

department: Medicine

institute: UCSD

summary: Sterile inflammation after myocardial infarction (MI) is classically credited to myeloid cells interacting with dead cell debris in the infarct zone (IZ). Here, we show that borderzone cardiomyocytes are the dominant initiators of a novel type I interferon (IFN) response in the infarct borderzone zone (BZ). Using spatial transcriptomics analysis of mice and humans, we find that MI induces colonies of interferon induced cells (IFNICs) expressing interferon stimulated genes (ISGs) decorating the BZ, where cardiomyocytes experience mechanical stress, nuclear rupture, and escape of chromosomal DNA. Cardiomyocyte-selective deletion of interferon regulatory factor 3 (Irf3) abrogated IFNIC colonies, whereas mice lacking Irf3 in fibroblasts, macrophages, neutrophils, or endothelial cells; Ccr2-deficient mice; or plasmacytoid dendritic cell-depleted mice did not. IFNs blunted the protective matricellular programs and contractile function of BZ fibroblasts, and increased vulnerability to pathologic remodeling. In mice that died after MI, IFNIC colonies were immediately adjacent to sites of ventricular rupture, while mice lacking IFNICs were protected from rupture and exhibited improved survival. Together, these results reveal a pathologic BZ niche characterized by a cardiomyocyte-initiated innate immune response. We suggest that selective inhibition of Irf3 activation in non-immune cells of the borderzone could limit ischemic cardiomyopathy while avoiding broad immunosuppression.

overall_design: Gene expression profiling of 10x Genomic Chromium single nuclei RNA-Seq or Visium spatial transcriptomics from the left ventricular myocardium or short axis sections of healthy and injured adult male and female C57BL6 and transgenic mouse hearts (10-12 weeks). Injuries include MI (permanent ligation of the LAD coronary artery), I/R-30 (30 minutes of ischemia followed by reperfusion), and traumatic needle injection.

first: V

last: Ninh

first: D

last: Calcagno

first: J

last: Yu

first: Bo

last: Zhang

first: N

last: Taghdiri

first: R

last: Sehgal

last: Mesfin

first: C

last: Chen

first: K

last: Kalhor

first: A

last: Toomu

last: Duran

first: E

last: Adler

last: Hu

middle: L

last: Christman

first: Z

last: Fu

first: B

last: Bintu


title: Spatially clustered type I interferon responses at injury borderzones [scRNA-Seq]

last: King

summary: Sterile inflammation post-myocardial infarction was thought to be driven by myeloid cells, however, we show that borderzone cardiomyocytes are the critical initators of the Type I Interferon response through spatial, single-cell, and single-nuclei transcriptomics.

overall_design: Nuclei and cells were isolated from 12-14-week-old adult wild-type C57BL/6J, IFNAR-/-, and IRF3-/- mice. For whole cell suspensions, freshly harvested hearts were enzymatically digested prior to filtering and flow cytometry sorting. Nuclei were isolated from fressh frozen tissue, minced and lysed, then filtered priored to collection.

middle: D

middle: R


title: Pyrimidine Triones as Potential Activators of p53 Mutants

status: Public on Aug 05 2024

city: Irvine

first: Fiona

last: Law

address: 839 Health Sciences Road

institute: UCI

summary: p53 is a crucial tumor suppressor in vertebrates that is frequently mutated in human cancers. Most mutations are missense mutations that render p53 inactive in suppressing tumor initiation and progression. Developing small molecule drugs to convert mutant p53 into an active, wild-type-like conformation is a significant focus for personalized cancer therapy. Prior research indicates that reactivating p53 suppresses cancer cell proliferation and tumor growth in animal models. Early clinical evidence with a compound selectively targeting p53 mutants with substitutions of tyrosine 220 suggests potential therapeutic benefits of reactivating p53 in patients. This study identifies and examines the UCI-1001 compound series as a potential corrector for several p53 mutations. The findings indicate that UCI-1001 treatment in p53 mutant cancer cell lines inhibits growth and reinstates wild-type p53 activities, including DNA binding, target gene activation, and induction of cell death. Cellular thermal shift assays, conformation-specific immunofluorescence staining, and differential scanning fluorometry suggest that UCI-1001 interacts with and alters the conformation of mutant p53 in cancer cells. These initial results identify pyrimidine trione derivatives of the UCI-1001 series as candidates for p53 corrector drug development.

overall_design: RNA samples were extracted from TOV-112D cells treated with UCI- 250 1001, UCI-1014, and DMSO using the RNeasy kit (Qiagen). Library generation and sequencing 251 was performed by Novogene Corporation Inc. Sequencing reads were quantified and aligned to 252 the human genome GRCh38 using kallisto v0.48.0. Estimated counts of transcripts from the 253 same Ensembl gene ID were added together and the sums were converted to integers. Differential 254 expression analysis was performed using DESeq2 v1.38.3.


title: Lipid Associated Macrophages' Promotion of Fibrosis Resolution during MASH regression requires TREM2

first: Souradipta

last: Ganguly

address: 9500 Gilman Drive

institute: University of California, San Diego

summary: While macrophage heterogeneity during Metabolic dysfunction-associated steatohepatitis (MASH) has been described, the fate of these macrophages during MASH regression is poorly understood. Comparing macrophage heterogeneity during MASH progression vs regression, we identified specific macrophage sub- populations that are critical for MASH/fibrosis resolution. We elucidated the restorative pathways and gene signatures that define regression associated macrophages (RAM) and establish the importance of TREM2+ macrophages during MASH regression. Liver resident Kupffer cells are lost during MASH and are replaced by four distinct monocyte derived macrophage sub-populations. Trem2 is expressed in two macrophage sub- populations: (i) monocyte-derived macrophages occupying the Kupffer cell niche (MoKC) and (ii) lipid-associated macrophages (LAM). In regression livers, no new transcriptionally distinct MF sub-population emerged. However, the relative macrophage composition changed during regression compared to MASH. While MoKC was the major macrophage sub-population during MASH, they decreased during regression. LAM was the dominant macrophage sub-type during MASH regression and maintained Trem2 expression. Both MoKC and LAM were enriched in disease resolving pathways. Absence of TREM2 restricted the emergence of LAMs and formation of hepatic crown like structures (hCLS). TREM2+ macrophages are functionally important not only for restricting MASH-fibrosis progression, but also for effective regression of inflammation and fibrosis. TREM2+ MF are superior collagen degraders. Lack of TREM2+ macrophages also prevented elimination of hepatic steatosis and inactivation of HSC during regression, indicating their significance in metabolic co-ordination with other cell- types in the liver. TREM2 imparts this protective effect through multifactorial mechanisms, including improved phagocytosis, lipid-handling and collagen degradation.

overall_design: Foz/Foz mice were put on a western diet (WD) for 12 weeks for the development of MASH-fibrosis.  MASH regression was induced by switching Western diet to normal chow for 8 weeks. Total non parenchymal cells (NPC) was isolated and subjected to scRNAseq

first: Sara Brin

last: Rosenthal

first: Kei

last: Ishizuka

first: Ty

last: Troutman

first: Theresa

last: Rohm

first: Naser

last: Khader

first: German Aleman

last: Muench

first: Yasuyo

last: Sano

first: Sebastiano

last: Archilei

first: Pejman

last: Soroosh

first: Jerrold

last: Olefsky

first: Ariel

middle: E

last: Feldstein

first: Tatiana

last: Kisseleva

first: Rohit

last: Loomba

first: Christopher

last: Glass

first: David

last: Brenner

first: Debanjan

last: Dhar


title: Type I interferon signaling induces melanoma cell-intrinsic PD-1 and its inhibition antagonizes immune checkpoint therapy

city: Boston

first: Shannan

last: Ho Sui

address: 401 Park Drive

institute: Harvard

summary: This study focuses on cancer cell-intrinsic programmed cell death (PD-1) functional regulation by examining induction of PD-1 in melanoma cells via type I interferon receptor signaling. Treatment of mouse melanoma cells with IFN-alpha or IFN-beta led to chromatin opening at and STAT1 and IFN regulatory factor 9 (IRF9) binding to a PDCD1 enhancer motif.

overall_design: Serum-starved murine YUMM4.1 melanoma cells were treated with or without recombinant mouse IFN-alpha (60 ng/ml) or IFN-beta (60 ng/ml) for 3 hrs. Cells were harvested, washed twice in ice-cold PBS, centrifuged at 700 g for 5 min at 4°C, and cell suspensions (~1 x 106, > 90% viability) cryopreserved according to GENEWIZ ATAC-seq sample preparation guidelines (Azenta Life Sciences, South Plainfield, NJ). ATAC-seq library preparation and DNA sequencing were conducted at Azenta Life Sciences. In brief, cell samples were thawed, washed, and genomic DNA contamination removed by DNase I (Thermo Fisher Scientific) treatment. Cell number and viability were quantified on a Countess Automated Cell Counter (Thermo Fisher Scientific). Cells were lysed, cytosolic fractions removed, nuclei treated with Tn5 transposase (Illumina, Hayward, CA) for 30 min at 37°C, and tagmented DNA was purified using the MinElute PCR Purification Kit (Qiagen). Subsequently, tagmented DNA was barcoded using the Nextera Index Kit v2 (Illumina), PCR-amplified, and cleaned using solid-phase reversible immobilization (SPRI) beads to generated purified DNA libraries. Sequencing libraries were clustered on a flow cell, loaded onto an Illumina HiSeq instrument (model 4000 or equivalent) according to the manufacturer’s instructions and samples sequenced using a 2x150 bp Paired End (PE) configuration. Image analysis and base calling were conducted using Control Software (CS). Raw sequence bcl data files were converted to fastq files and de-multiplexed using bcl2fastq v2.17 conversion software (Illumina). One mismatch was allowed for index sequence identification.


title: Fibrosis-related transcriptome unveils a distinctive matrix remodeling pattern in penetrating but not in stricturing ileal Crohn's disease

city: Vizela

first: Marta

middle: Patrícia

last: Ferreira

state: Braga

address: Rua da junuqeira, Santo Adrião

country: Portugal

summary: Using Nanostring technology and comparative bioinformatics, we analyzed the expression of 760 fibrosis-related genes in 36 ileal surgical specimens, 12 B2(Penetrating) and 24 B3(structuring), the latter including 12 cases with associated stricture(s) (B3s) and 12 without (B3o).

overall_design: nCounter® Fibrosis Consortium Panel was runned in 36 ileal surgical specimens

first: Helena

last: Sousa

first: Irene

last: Gullo

first: Ana

last: Rocha

first: Carla

last: Oliveira

first: Fátima

last: Carneiro

first: Fernando

last: Magro


title: Heterozygous knockout of Synaptotagmin13 phenocopies ALS features and TP53 activation in human motor neurons

status: Public on Aug 02 2024

city: Blaubeuren

first: Alberto

last: Catanese

address: Albert-Einstein-Allee 11

country: Germany

institute: Ulm University, Anatomy and Cell Biology

summary: Spinal motor neurons (MNs) represent a highly vulnerable cellular population, which is affected in fatal neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS) and spinal muscular atrophy (SMA). Despite the advances in our understanding of the genetic causes of these pathologies, the molecular determinants of MN vulnerability are still largely unknown. In this work, we aimed at clarifying the specific contribution of a member of the protein family synaptotagmins to MNs sufferance. We indeed combined multi-omics and deep learning strategies with human iPSC-derived MN to highlight Synaptotagmin 13 (SYT13) as a candidate gene contributing to MN vulnerability when its levels are detrimentally reduced. By using CRISPR-Cas9 technology, we investigated whether the heterozygous loss of SYT13 might be sufficient to trigger a neurodegenerative phenotype resembling those observed in ALS and SMA. SYT13+/- hiPSC-derived MNs displayed a progressive manifestation of typical neurodegenerative hallmarks such as loss of synaptic contacts and accumulation of aberrant aggregates. By comparing the transcriptome of SYT13-deficient cells to SYT13+/+ ones, we found a significant impairment in biological mechanisms involved in motoneuron specification and spinal cord differentiation. In addition, we identified a striking correlation of the Syt13+/- transcriptome with an ALS signature generated using RNAseq data from human MNs and post mortem spinal cord samples. This significant overlap converged toward a shared expression of pro-apoptotic and pro-inflammatory genes, which are controlled by the transcription factor TP53. Our data show for the first time that the heterozygous loss of a single member of the synaptotagmins family, SYT13, is sufficient to trigger a series of abnormal alterations leading to cellular sufferance in human MNs, thus revealing novel insights into the selective vulnerability of this cell population.

overall_design:
SYT13 WT and SYT13+/- hiPSCs were differentiated for 5 weeks into spinal motor neurons using a small molecules-based protocol (PMCID: PMC8261490)
We then performed RNAseq analysis of SYT13 WT and SYT13+/- motor neurons

first: Johannes

last: Lehmann


title: Transcriptomic landscape of seedstick in Arabidopsis thaliana funiculus after fertilisation

status: Public on Aug 04 2024

city: Porto

first: Sílvia

last: Coimbra

address: Rua do Campo Alegre s/n

department: Biology

institute: LAQV/REQUIMTE, Faculty of Sciences, University of Porto

summary: In Angiosperms, the continuation of plant species is intricately dependent on the funiculus multifaceted role in nutrient transport, mechanical support, and dehiscence of seeds. SEEDSTICK (STK) is a MADS-box transcription factor involved in seed size and abscission, and one of the few genes identified as affecting funiculus growth. Given the importance of the funiculus to a correct seed development, allied with previous phenotypic observations of stk mutants, we performed a transcriptomic analysis of stk funiculi from floral stage 17, using RNA-sequencing, to infer on the deregulated networks of genes. The generated dataset of differentially expressed genes was enriched with cell wall biogenesis, cell cycle, sugar metabolism and transport terms, all in accordance with stk phenotype observed in funiculi from floral stage 17. We selected eight differentially expressed genes for transcriptome validation using qPCR and/or promoter reporter lines. Those genes were involved with abscission, seed development or novel functions in stk funiculus, such as hormones/secondary metabolites transport, for transcriptome validation using qPCR and/or promoter reporter lines. Overall, the analysis performed in this study allowed delving into the STK-network established in Arabidopsis funiculus, fulfilling a literature gap. Simultaneously, our findings reinforced the reliability of the transcriptome, making it a valuable resource for candidate genes selection for functional genetic studies in the funiculus. This will enhance our understanding on the regulatory network controlled by STK, on the role of the funiculus and identified processes and new candidate genes that will enable a better understanding on the role of this sporophytic structure and how seed development may be affected by itthem.

overall_design:
Stage 17 funiculi (according to Smyth et al. 1990) were used for mRNA extraction in both stk and wt samples, in duplicate.

Grants:
. "National funding" (Grant ID UIDB/50006/2020, Fundacão para a Ciencia e Tecnologia and Ministerio da Ciencia, Tecnologia e Ensino Superior)
. "PhD scholarship" (Grant ID SFRH/BD/143579/2019, Fundacao para a Ciência e Tecnologia)
. "PhD scholarship" (Grant ID LCF/BQ/DR20/11790010, "la Caixa" Foundation (ID 100010434))
. "Grant-in-Aid for Early-Career Scientists" (Grant ID 18K14729, Japanese Society for the Promotion of Science)
. "Grant-in-Aid for Early-Career Scientists" (Grant ID 20K15817, Japanese Society for the Promotion of Science)
. "Grant-in-Aid for Transformative Research Areas" (Grant ID 22H05677, Japanese Society for the Promotion of Science)
. "Grant-in-Aid for Transformative Research Areas" (Grant ID 23H04740, Japanese Society for the Promotion of Science)
. "Fellowship" (Grant ID 22H04980, Japanese Society for the Promotion of Science)
. "Fellowship" (Grant ID 22K21352, Japanese Society for the Promotion of Science)

first: Maria

first: Jessy

last: Silva

first: Hidenori

last: Takeuchi

first: Takamasa

last: Suzuki

first: Tetsuya

last: Higashiyama


title: Essential functions of RNA helicase Vasa in Drosophila spermatogenesis, from maintenance of germline stem cells to passage through meiosis between Drosophila melanogaster and Drosophila simulans [smRNA-seq]

city: Moscow

first: Alexei

middle: A.

last: Kotov

address: 26 Vavilov Street

country: Russia

institute: Koltzov Institute of Developmental Biology of the Russian Academy of Sciences

summary: DEAD-box RNA helicase Vasa is required for gonad development and fertility in multiple animals. In Drosophila, Vasa performs essential functions in oogenesis, including the maintenance of germline stem cells (GSCs), piRNA silencing of mobile elements, translation regulation, and primordial germ cell specification. Despite its evident significance, the mechanistic basis of Vasa action and its precise role in spermatogenesis become incomprehensible. Several papers affirm the fertility of males carrying vasa mutations. However, it is also shown that Vasa is essential for piRNA-mediated repression of Stellate genes needed for the maintenance of male fertility.Here we found that loss-of-function vasa mutations led to a rapid decline in GSC maintenance in the testes, a severe loss of total germ cell content, and a strong decrease in male fertility over time. With the aid of analysis of small RNA libraries, we revealed that collapse of piRNA biogenesis in the absence of vasa expression. Despite that, we did not reveal increasing cell death events in the early germ cells of vasa mutant testes. The introduction of the transgene rhino copy, encoding a nuclear component of the piRNA pathway, in vasa mutant background allowed us to rescue premeiotic stages of spermatogenesis, including GSC maintenance and the development of spermatogonia and spermatocytes. However, the progression of spermatocytes through meiosis and the fertility of the rhino transgene-rescued males were disrupted by strong Stellate gene derepression owing to the absence of corresponding piRNAs. We have shown that Vasa functions in spermatogenesis are essential at two separate developmental stages: in GSCs for their maintenance and in spermatocytes for the repression of Stellate genes.

overall_design: To study the contribution of Vasa in Stellate silencing in more detail, we prepared and sequenced on the Illumina platform whole-transcriptome libraries and 18–29 nt small RNAs isolated from the testes of vasa mutants, control heterozygous males, and males with the transgenic construct rhi-GFP in the background of vasa mutations.