title: Clonal differences underlie variable responses to sequential and prolonged treatment

city: Philadelphia

first: Dylan

middle: L

last: Schaff

state: PA

address: 421 Curie Blvd, BRB II/II rm 445

department: Bioengineering

institute: University of Pennsylvania

summary: Cancer cells exhibit dramatic differences in gene expression at the single-cell level which can predict whether they become resistant to treatment. Treatment perpetuates this heterogeneity, resulting in a diversity of cell states among resistant clones. However, it remains unclear whether these differences lead to distinct responses when another treatment is applied or the same treatment is continued. In this study, we combined single-cell RNA-sequencing with barcoding to track resistant clones through prolonged and sequential treatments. We found that cells within the same clone have similar gene expression states after multiple rounds of treatment. Moreover, we demonstrated that individual clones have distinct and differing fates, including growth, survival, or death, when subjected to a second treatment or when the first treatment is continued. By identifying gene expression states that predict clone survival, this work provides a foundation for selecting optimal therapies that target the most aggressive resistant clones within a tumor.

overall_design: WM989 cells were tagged with unique, transcribed barcodes, and allowed to double so there were many cells with each barcode. The cells were split three ways and samples were treated for four weeks with dabrafenib and trametinib (DabTram), cisplatin and holiday, or CoCl2 before scRNAseq and barcode sequencing. The three populations were then split three ways again and treated for four weeks with dabrafenib and trametinib (DabTram), cisplatin and holiday, or CoCl2 before a second round of scRNAseq and barcode sequencing.

first: Aria

last: Fasse

first: Phoebe

last: White

first: Robert

last: Vander Velde

first: Sydney

last: Shaffer


title: NRF2-independent iron-triggered signaling via MAPK p38-ETS1 regulates LSEC Bmp6 mRNA expression

status: Public on Jan 24 2024

city: Warsaw

first: Michal

last: Mikula

address: Roentgena 5

department: Experimental Oncology

country: Poland

institute: Maria Skłodowska-Curie National Research Institute of Oncology

type: Expression profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Unfolded protein Response is a major hurdle in direct neuronal reprogramming of human astrocytes [Bulk RNA-seq]

city: Planegg-Martinstied

first: Giacomo

last: Masserdotti

address: Groshaderner Strasse 9

department: ISF

country: Germany

institute: Helmholtz Zentrum

summary: Mitochondria account for essential cellular pathways, from ATP production to nucleotide metabolism, and their deficits lead to neurological disorders and contribute to the onset of age-related diseases. Direct neuronal reprogramming aims at replacing neurons lost in such conditions, but almost nothing is known about the impact of mitochondrial dysfunction in human cell direct reprogramming. Here we explore the defects and how to improve the neuronal reprogramming of iPSC-derived astrocytes carrying mutations in NDUFS4 gene, important for Complex I and associated to Leigh Sydrome. This led to the identification of the unfolded protein response as a major hurdle in the direct neuronal conversion of not only patient, but also control astrocytes and fibroblasts. Our data highlight mitochondria-ER stress-mediated inhibition of translation as a key hurdle in direct neuronal reprogramming, emphasizing how disease modelling using patient cells can contribute to unravel novel pathways in human glia cell reprogramming.

overall_design:
human IPSCs from control or patients with NDUFS4 mutations were differentiated into proliferating and non-proliferating astrocytes. Three control and 3 patients lines were analyzed. Per each line, 3 biological replicates were collected at 3 stages: IPSC, proliferating astrocyte and non-proliferating astrocytes.

***************************************************************
Raw data will be deposited in GHGA when the database is available
***************************************************************

first: Giovanna

last: Sonsalla

first: Pawel

last: Smialowski

first: Holger

last: Prokisch


title: Hypoxic tumors are sensitive to FLASH radiotherapy

city: Epalinges

first: Marie-Catherine

last: Vozenin

address: Rue de la Corniche, 9A

department: Radiation Oncologie

country: Switzerland

institute: CHUV

summary:
Purpose: Tumor hypoxia is a major cause of treatment resistance, especially to radiation therapy at conventional dose rate (CONV), and we wanted to assess whether hypoxia does alter tumor sensitivity to FLASH.

Methods and materials: We engrafted several tumor types (glioblastoma [GBM], head and neck cancer, and lung adenocarcinoma) subcutaneously in mice to provide a reliable and rigorous way to modulate oxygen supply via vascular clamping or carbogen breathing. We irradiated tumors using a single 20-Gy fraction at either CONV or FLASH, measured oxygen tension, monitored tumor growth, and sampled tumors for bulk RNAseq and pimonidazole analysis. Next, we inhibited glycolysis with trametinib in GBM tumors to enhance FLASH efficacy.

Results: Using various subcutaneous tumor models, and in contrast to CONV, FLASH retained antitumor efficacy under acute hypoxia. These findings show that in addition to normal tissue sparing, FLASH could overcome hypoxia-mediated tumor resistance. Follow-up molecular analysis using RNAseq profiling uncovered a FLASH-specific profile in human GBM that involved cell-cycle arrest, decreased ribosomal biogenesis, and a switch from oxidative phosphorylation to glycolysis. Glycolysis inhibition by trametinib enhanced FLASH efficacy in both normal and clamped conditions.

Conclusions: These data provide new and specific insights showing the efficacy of FLASH in a radiation-resistant context, proving an additional benefit of FLASH over CONV.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for U-87 MG human glioblastoma tumors grafted to mouse flank at 24 hours post-radiotherapy (20 Gy electrons at ultra-high-dose-rate and coventional dose rates, under different oxygen tensions: hypoxia, physioxia, and hyperoxia) and 1 week post-recurrence.

first: Ron

middle: J

last: Leavitt

first: Pierre

last: Montay-Gruel

first: Benoit

last: Petit


title: Single-cell RNA-sequencing from stroma in a mouse submandibular salivary gland following 14-day ligation or mock surgery

status: Public on Jan 23 2024

city: Albany

first: Melinda

last: Larsen

state: NY

address: 1400 Washington Ave., LSRB 10

department: Dept. of Biological Science

country: USA

institute: University at Albany, SUNY

summary: The submandibular salivary gland stroma makes up only a small portion of the total salivary gland and the stromal response to salivary gland injury has been understudied. We used single-cell RNA-sequencing (scRNAseq) to analyze which cell types are present in ductal ligated and mock surgery salivary glands, how the cell type abundance is altered during injury, and how the transcriptome of those cells is being altered. This will allow us to examine which cell types are important contributors to recovery from salivary gland ductal ligation injury.

overall_design: Submandibular salivary glands were dissociated into single cells followed by two rounds of magnetic activated cell sorting (MACs) to deplete for epithelial cells expressing EpCAM and red blood cells expressing Ter119. The samples then went through one round of dead cell removal using Miltenyi dead cell removal kit.

first: Amber

last: Altrieth

first: James

last: Kenney

first: Deirdre

last: Nelson

first: Emily

last: Suarez

first: Victoria

last: Gellatly

first: Sergo

last: Gabunia


title: Association of inflammatory mediators with mitochondrial DNA mutations in geriatric COVID-19 patients

city: Ancona

first: Tiziana

last: Casoli

address: Via Birarelli 8

department: Scientific Technological Area

country: Italy

institute: INRCA

type: Genome variation profiling by genome tiling array

summary: This study aimed to evaluate single nucleotide substitutions in mtDNA and analyze their correlation with inflammatory biomarkers in elderly COVID-19 patients. A total of 30 COVID-19 patients and 33 older adult controls (aged over 65 years) were enrolled. mtDNA was extracted from buffy coat samples and sequenced using a chip-based resequencing system (Affymetrix MitoChip v2.0) which detects both homoplasmic and heteroplasmic mtDNA mutations, and allows the assessment of low-level heteroplasmy. Serum concentration of IL-6, IFN-α, TNF-α and IL-10 were determined in patients by a high-sensitivity immunoassay. We found a higher burden of total heteroplasmic mutation in COVID-19 patients compared to controls with a selective increment in ND1 and COIII genes. Low-level heteroplasmy was significantly elevated in COVID-19 patients, especially in genes of the respiratory complex I. Both heteroplasmic mutation burden and low-level heteroplasmy were associated with increased levels of IL-6, TNF-α, and IFN-α.

overall_design: The study population comprised elderly individuals with COVID-19 (n=30, experimental group) and age-matched controls (n=33). Blood samples were collected from individuals who were admitted to the IRCCS INRCA Hospital, Ancona, Italy, and were enrolled in either the Report-Age COVID (experimental patients) or Report-Age (control subjects) projects, respectively. The aim of the Report-Age COVID project is to provide a deeper understanding of COVID-19 disease in elderly patients (≥ 65 years). The experimental subjects were COVID-19 positive cases as confirmed by the detection of SARS-CoV-2 RNA in nasal/oropharyngeal swabs while controls were COVID-19 negative elderly subjects. Demographic and anamnestic data, biochemical and hematological variables, information on treatments, comorbidities, and survival were collected in a retrospective manner and anonymized before release.

first: Anna

middle: R

last: Bonfigli

first: Mirko

last: Di Rosa


title: Perlecan (HSPG2) signalling promotes structural, contractile, and metabolic development during the maturation stage of hPSC-CM differentiation

city: London

first: Tegan

last: Gilmore

address: 1 Midland Road

department: Bioinformatics & Biostatistics

country: United Kingdom

institute: Francis Crick Institute

summary: Perlecan (HSPG2), a multifunctional heparan sulphate proteoglycan (HSPG), is a key player in extracellular matrix maturation and stabilisation. Structurally perlecan is similar to agrin, another HSPG known to contribute to cardiomyocyte phenotype switching by reverting hypertrophy. Although perlecan is crucial for cardiac development, its role in cardiomyocyte phenotypic switching is unknown. Here we show perlecan expression increases over time during the differentiation of pluripotent stem cells to cardiomyocytes (hPSC-CMs). Haploinusuffient hPSCs (HSPG2+/-) differentiate efficiently towards cardiomyocytes with minimal transcriptomic differences seen early in differentiation, but wide-ranging changes seen in late-stage cardiomyocytes. These differences are associated with structural, contractile, metabolic, and ECM genes. In keeping, late-stageHSPG2+/-hPSC-CMs display reduced maximal metabolic capacity and increased extracellular acidification rate, characteristics of reduced maturity. Moreover, engineered heart tissues produced withHSPG2+/-hPSC-CMs exhibit increased ECM remodelling, reduced tissue thickness and force generation. Wild-type hPSC-CMs grown on a substrate coated with a perlecan peptide showed increased cardiomyocyte nucleation typical of hypertrophic growth, suggesting that perlecan plays the opposite role of agrin by promoting cellular maturation rather than hyperplasia and proliferation. These data show that perlecan promotes cardiomyocyte maturity, possibly through hypertrophic growth. Targeting perlecan-dependent signalling may, therefore, reverse the phenotypic switch common to many forms of heart failure, by improving cardiomyocyte functionality.

overall_design: Comparative gene expression profiling analysis and comparative transcript analysis of RNA-seq data for human pluripotent stem cell cardiomycocyte lines

first: Benjamin

middle: B

last: Johnson

first: Lorenza

middle: I

last: Tsansizi

first: Terri

last: Holmes

first: Katherine

last: Hampton

first: Nguyen

middle: T

last: Vo

first: Aishah

last: Nasir

first: Chris

last: Denning

first: Cathy

last: Merry

first: John

last: Whitelock

first: Linda

last: Troeberg

first: Stuart

last: Rushworth

first: Andreia

middle: S

last: Bernardo

middle: G

last: Smith


title: Phase separation of YAP-MAML2 differentially regulates the transcriptome

city: San Francisco

first: Xiaokun

last: Shu

state: CA

address: 555 Mission Bay Boulevard South, CVRI,RM484

institute: University of California, San Francisco

summary: SPARK-OFF tag does not perturb the core transcriptional function of YAP-MAML2.~2260 DEGs with ≥ 1.5-fold change in transcript level, including ~1270 up-regulated genes and ~990 down-regulated genes can be identified from YAP-MAML2 overexpression HEK293T cells.We identified 88 DEGs (p-value < 0.01, |Log2FC| ≥ 0.58, FDR < 0.1) that are regulated by YAP-MAML2 phase separation with ≥ 1.5-fold change in transcript levels, including 44 up-regulated genes and 44 down-regulated genes

overall_design: analyze the transcriptome difference in HEK293T cells which overexpress condensed YAP-MAML2 or diffusive YAP-MAML2. To control YAP-MAML2 phase separation, a rapamycin induced tool was applied to the cells. The control group is the effect on rapamycin alone. HEK293T express eGFP alone


title: Serotonin transporter-dependent histone serotonylation in placenta contributes to the neurodevelopmental transcriptome [ChIP-seq]

city: New York

first: Jennifer

middle: C

last: Chan

state: New York

address: 1470 Madison Ave 9-201

department: Neuroscience

institute: Icahn School of Medicine at Mount Sinai

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Brain development requires appropriate regulation of serotonin (5-HT) signaling from distinct tissue sources across embryogenesis. At the maternal-fetal interface, the placenta is thought to be an important contributor of offspring brain 5-HT and is critical to overall fetal health. Yet, how placental 5-HT is acquired, and the mechanisms through which 5-HT influences placental functions, are not well understood. Recently, our group identified a novel epigenetic role for 5-HT, in which 5-HT can be added to histone proteins to regulate transcription, a process called H3 serotonylation. Here, we show that H3 serotonylation undergoes dynamic regulation during placental development, corresponding to gene expression changes that are known to influence key metabolic processes. Using transgenic mice, we demonstrate that placental H3 serotonylation largely depends on 5-HT uptake by the serotonin transporter (Sert/Slc6a4). Sert deletion robustly reduces enrichment of H3 serotonylation across the placental genome, and disrupts neurodevelopmental gene networks in offspring brain tissues. Thus, these findings suggest a novel role for H3 serotonylation in coordinating placental transcription at the intersection of maternal physiology and offspring brain development.

overall_design: This study examined male and female placental tissues across embryogenesis at E9.5, E12.5 and E17.5 (H3K4me3Q5ser ChIP-seq and bulk RNA-seq), compared E12.5 WT vs Sert KO, Tph1 KO, and Oct3 KO placenta tissues (H3K4me3 and H3K4me3Q5ser ChIP-seq), and examined associated offspring brains at E12.5 (bulk RNA-seq)

first: Natalia

last: Alenina

first: Ashley

middle: M

last: Cunningham

first: Aarthi

last: Ramakrishnan

first: Li

last: Shen

first: Michael

last: Bader

first: Ian

last: Maze


title: Serotonin transporter-dependent histone serotonylation in placenta contributes to the neurodevelopmental transcriptome [RNA-seq]


title: Macrophage-enriched Sectm1a promotes efficient efferocytosis to attenuate ischemia/reperfusion-induced cardiac injury

city: Cincinnati

first: Guo-Chang

last: Fan

state: ohio

address: 231 Albert Sabin Way

department: Pharmacology & Systems Physiology

institute: University of Cincinnati

summary: Efficient clearance and degradation of apoptotic cardiomyocytes by macrophages (termed efferocytosis) are critical for inflammation resolution and restoration of cardiac function after myocardial ischemia/reperfusion (I/R). Here, we define secreted and transmembrane protein 1a (Sectm1a), a cardiac macrophage-enriched gene, as a modulator of macrophage efferocytosis in I/R hearts. Upon myocardial I/R, Sectm1a-KO mice exhibit impaired macrophage efferocytosis, leading to massive accumulation of apoptotic cardiomyocytes, cardiac inflammation, fibrosis, and consequently, exaggerated cardiac dysfunction. By contrast, therapeutic administration of recombinant SECTM1A protein significantly enhances macrophage efferocytosis and improves cardiac function. Mechanistically, SECTM1A can elicit autocrine effects on the activation of glucocorticoid-induced TNF receptor (GITR) at the surface of macrophages, leading to the upregulation of liver X receptor alpha (LXRα) and its downstream efferocytosis-related genes and lysosomal enzyme genes. Our study suggests that Sectm1a-mediated activation of Gitr/LXRα axis could be a promising approach to enhance macrophage efferocytosis for the treatment of myocardial I/R injury.

overall_design: To dissect the potential mechanism underlying the established link between Sectm1a and macrophage efferocytosis, we re-analyzed our previous RNA-sequencing data generated from (wilde type) WT and Sectm1a knockout (KO) macrophages (MФs). RNA files of bone marrow derived MФs from WT and Sectm1a KO mice were generated. Directional polyA RNA-sequencing was performed by the Genomics, Epigenomics and Sequencing Core (GESC) at the University of Cincinnati. To fully capitalize the gene signature of the whole transcriptome, especially to capture lower amplitude signals (e.g., lower fold changes) from expression variation that would otherwise fall below the signal-to-noise significance cutoff, we first performed unbiased gene set enrichment analysis (GSEA) using whole MФ transcriptomes. The volcano plot was created using the R package “EnhancedVolcano” and showed that there were 2117 significantly changed genes (FDR<0.1) out of 12987 genes used as input. We then performed gene ontology (GO) enrichment analysis on differentially expressed genes and focused on GO pathways related to efferocytosis. Our analysis results demonstrated that ablation of Sectm1a in MФs significantly reduced the expression of several sets of genes that are associated with the regulation of endocytosis and phagocytosis, phagocytotic engulfment, apoptotic cell clearance, and lysosome function.

first: Xiaohong

last: Wang

first: Wa

last: Du

first: Yutian

last: Li


title: The comparison for the transcript levels of chloroplast-encoded genes between wild type (Col-0) and pbf8 mutants

city: Taian

first: Aihong

last: Zhang

state: Shandong

address: 61#,Daizong Street

country: China

institute: Shandong Agricultural University

summary: We found that thylakoid-anchored protein PBF8 is a key regulator for Photosystem I (PSI) biogenesis. To explore the role of PBF8 in regulating chloroplast gene expression, we performed the RNA-seq to compare the the transcript levels of chloroplast-encoded genes between wild type (Col-0) and pbf8 mutants. To this end, we isolated the total RNA form 12-day-old wild type and pbf8 seedlings grown on the MS medium under long-day conditions (14 h light, 10 h dark) at 22 ºC and with a light intensity of 80 µmol m-2 s-1. The rRNAs were deleted using the Ribo-Zero Kit (Epicentre). The resulting rRNA-depleted RNA was used for preparing the sequencing library with NEBNext Single Cell/Low input library Prep Kit. The libraries were pooled and sequenced on an Illumina Nova 6000 system with 150-bp pair-end reads. Finally, our results show that the transcript accumulation for chloroplast-encoded PSI subunit and assembly factor genes between the wild type (Col-0) and pbf8 samples, suggesting PBF8 may not affect the transcript levels of chloroplast-encoded PSI subunits and assembly factors in chloroplasts.

overall_design:
To investigate the role of PBF8 in regulating chloroplast gene expression, we obtained the pbf8 T-DNA insertion mutant in which PBF8 gene has been knocked out.
We then performed gene expression profiling analysis using the data form RNA-seq of wild type and pbf8 seedlings.
Comparative profiling analysis of chloroplast-encoded genes of RNA-seq data for wild type and pbf8 seedlings.

first: Congming

last: Lu

first: Yi


title: Class A capsid assembly modulator apoptotic elimination of hepatocytes with high HBV core antigen level in vivo is dependent on de novo core protein translation

city: Beerse

first: Nádia

last: Conceição-Neto

address: Turnhoutseweg 30

country: Belgium

institute: Janssen R&D

summary: Treatment with hepatitis B virus (HBV) capsid assembly modulators that induce the formation of aberrant HBV core protein structures (CAM-A) leads to programmed cell death, apoptosis, of HBV-infected hepatocytes and subsequent reduction of HBV antigens, which differentiates CAM-A from other CAMs. The effect is dependent on the de novo synthesis and high levels of core protein.

overall_design: Treatment with CAM-A up to 28 days. 3 mouse livers collected at days 7, 8, 9, 10, 11, 12, 13, 14, 16, 18, 21, 22, 23, 24, 28 days post treatment. Treated and placebo groups.

first: Jan

last: Berke

first: Sarah

last: Sauviller


title: Improving prime editing with an endogenous small RNA-binding protein

status: Public on Jan 22 2024

city: Princeton

first: Brittany

last: Adamson

state: NJ

address: 144 Carl Icahn Laboratory, South Drive

department: Molecular Biology

institute: Princeton University

type: Non-coding RNA profiling by high throughput sequencing

summary: Prime editing enables the precise modification of genomes through reverse transcription of template sequences appended to the 3′ ends of CRISPR–Cas guide RNAs. To identify cellular determinants of prime editing, we developed scalable prime editing reporters and performed genome-scale CRISPR-interference screens. From these screens, a single factor emerged as the strongest mediator of prime editing: the small RNA-binding exonuclease protection factor La. Further investigation revealed that La promotes prime editing across approaches (PE2, PE3, PE4 and PE5), edit types (substitutions, insertions and deletions), endogenous loci and cell types but has no consistent effect on genome-editing approaches that rely on standard, unextended guide RNAs. Previous work has shown that La binds polyuridine tracts at the 3′ ends of RNA polymerase III transcripts. We found that La functionally interacts with the 3′ ends of polyuridylated prime editing guide RNAs (pegRNAs). Guided by these results, we developed a prime editor protein (PE7) fused to the RNA-binding, N-terminal domain of La. This editor improved prime editing with expressed pegRNAs and engineered pegRNAs (epegRNAs), as well as with synthetic pegRNAs optimized for La binding. Together, our results provide key insights into how prime editing components interact with the cellular environment and suggest general strategies for stabilizing exogenous small RNAs therein.

overall_design:
To study how La affects (e)pegRNAs, we transfected either targeting or nontargeting (e)pegRNA plasmids into K562 PEmax parental cells and La-knockout clonal cell line 4, constructed small RNA libraries from total RNA and performed paired-end sequencing. To study if PE7 affects gene expression as compared to PEmax or PE7 mutant, we transfected K562 cells with plasmids expressing PEmax, PE7 or PE7 mutant and pegRNA plasmid encoding HEK3 1TtoA or PRNP 6GtoT, constructed mRNA libraries from total RNA and performed single end sequencing.
Related to GSE255003.

first: Jun

last: Yan

first: Paul

last: Oyler-Castrillo

first: Purnima

last: Ravisankar

first: Carl

last: Ward

first: Sébastien

last: Levesque

first: Yangwode

last: Jing

first: Danny

last: Simpson

first: Anqi

last: Zhao

first: Hui

first: Weihao

first: Laine

last: Goudy

first: Ralf

last: Schmidt

first: Sabrina

last: Solley

first: Luke

middle: A

last: Gilbert

first: Michelle

first: Daniel

middle: E

last: Bauer

first: Alexander

last: Marson

first: Lance

last: Parsons

first: Britt


title: The self-reactive FVIII T cell repertoire in healthy individuals relies on a short set of epitopes and public clonotypes

city: Gif-sur-Yvette

first: Valeria

last: Porcheddu

address: Université de Paris-Saclay, CEA, INRAE, Département Médicaments et Technologies pour la Santé, SIMoS

country: France

institute: CEA Paris-Saclay

summary: Non-mutated FVIII-specific CD4 T cell epitopes have been recently found to contribute to the development of inhibitors in patients with hemophilia A (HA), while auto-reactive CD4 T cells specific to FVIII circulate in the blood of healthy individuals at a frequency close to the foreign protein Ovalbumin. Thus, although FVIII is a self-protein, the central tolerance raised against FVIII appears to be low. In this study, we conducted a comprehensive analysis of the FVIII CD4 T cell repertoire in twenty-nine healthy donors. Sequencing of the CDR3β TCR region from isolated FVIII-specific CD4 T cells revealed a limited usage and pairing of TRBV and TRBJ genes, as well as a mostly hydrophobic composition of the CDR3β region according to their auto-reactivity. FVIII repertoire is dominated by a few clonotypes, with only 13 clonotypes accounting for half of the FVIII response. Through a large-scale epitope mapping of the full-length FVIII sequence, we identified 18 immunodominant epitopes located in the A1, A3, C1, and C2 domains and covering half of the T-cell response. These epitopes exhibited a broad specificity for HLA-DR or DP molecules or both. T-cell priming with this reduced set of peptides revealed that highly expanded clonotypes specific to these epitopes were responsible individually for up to 32% of the total FVIII repertoire. These FVIII T cell epitopes and clonotypes were shared among HLA-unrelated donors tested and previously reported HA patients. Our study highlights the role of the auto-reactive T cell response against FVIII in HA and its similarity to the response observed in healthy individuals. Thus, it provides valuable insights for the development of new tolerance induction and deimmunization strategies

overall_design:
We aimed to characterise the T cell response to rFVIII in vitro, using chicken Ovalbumin (Ova) as a non-self-protein benchmark. Given the low frequency of FVIII-specific T cells in healthy donors, we enriched the CD4 population in vitro with 3-once-per-week rounds of stimulation using autologous DCs loaded with either rFVIII, Ova or KLH as positive control. We analysed their specificity through IFN-γ ELISPOT assay and positive T cell lines for each antigen were sorted, using activation-induced markers CD134/CD137 double positive staining, from 4 healhty donors for TCR CDR3β sequencing.
After an extensive epitope mapping on 16 healthy donors, we identified 18 immudominant and immunoprevalent FVIII peptides which are inducing the majority of the CD4 T cell response to FVIII in healthy individuals. In order investigate how representative the response to this 18 sequences was of the total FVIII repertoire, we amplified in vitro the CD4 T cell population from 5 healthy donors for both rFVIII and the pool of 18 FVIII peptides. We then sorted positive T cell lines for both conditions in order to perform TCR CDR3β sequencing and a comparative analyses of the response.

first: Gautier

last: Lhomme

first: Rémi

last: Giraudet

first: Evelyne

last: Correia

first: Bernard

last: Maillère


title: Serotonin transporter-dependent histone serotonylation in placenta contributes to the neurodevelopmental transcriptome

first: Jennifer

last: Chan


title: Molecular Analysis of Murine KitK641E Melanoma Progression

city: Boston

first: Hensin

last: Tsao

state: MA

address: 70 Blossom St, Edwards 202

institute: Massachusetts General Hospital

summary: Acral and mucosal melanomas (AMMs) are often driven by mutations in the KIT receptor tyrosine kinase, with nearly 40% harboring alterations in the KIT locus. Despite advances in the knowledge of KIT-mutated melanomas, little is known about the molecular reprogramming that occurs during KIT-mediated melanoma progression due to the rarity of AMMs and the lack of comprehensive biological tools and models. To this end, we used a novel murine model that allows us to ascertain the molecular underpinnings of the stages of cancer progression—transformation, tumorigenesis, immune engagement, and tumor escalation. We found dramatic increases in biosynthetic demands associated with the transformation stage, including DNA and RNA metabolism, leading to replication stress. Tumorigenesis was closely linked to neuronal and axonal development, likely necessary for invasion into the host. Immune engagement highlighted early immune excitation and rejection pathways, possibly triggered by abrupt neoantigen exposure. Finally, tumor escalation pathways proved consistent with immune evasion, with immune-related pathways becoming significantly downregulated. To our knowledge, this is the first time these critical milestones needed for KIT-driven-melanoma tumor formation have been studied at the molecular level using isogenically-matched and phenotypically-defined cells.

overall_design: To investigate Kit mutation induced melanoma transformation, we developed a series of cells lines to model the stages of cancer progression—transformation, tumorigenesis, immune engagement, and tumor escalation. RNAseq analysis of seven lines to obtain gene expression profiles, from which comparative gene expression profiles analysis was conducted.

first: Emily

last: Everdell

first: Zhenyu

last: Ji

first: Ching

middle: N

last: Njauw


title: RNAseq analysis of B6-Mx1(r) and D2-Mx1(r) mice infected with PR8 virus, map to mm11

city: Münster

first: Klaus

last: Schughart

address: Von-Esmarch-Str 56

department: Virology

institute: University Münster

summary: Strong differences in the immune response between the two mouse strains were found

overall_design:
Gene expression changes in the lung was studied by RNAseq
Lungs were collected from mice, RNA was isolated and sequenced.


title: T cell states, repertoire and function in classical Hodgkin lymphoma revealed through single-cell analyses

city: Chicago

first: Xiufen

last: Chen

state: IL

address: 900 E. 57th street KCBD 6230A

institute: University of Chicago

summary: The classical Hodgkin lymphoma (cHL) environment is comprised of a rich and varied immune cell infiltrate that is co-opted to support the survival and growth of malignant Hodgkin-Reed-Sternberg (HRS) cells. Several lines of evidence support the notion that, despite the immune suppressive environment that characterizes this disease, HRS cells are actively surveilled by endogenous T cells. For instance, HRS cells recurrently acquire genetic lesions associated with impaired antigen presentation and PD-L1 upregulation. Moreoever, cHL is particularly vulnerable to anti-PD-1/PD-L1 immunotherapy, which indicates that T cells can recognize and eliminate HRS cells when dominant immune checkpoints are disabled. However, the T cells that drive responses to anti-PD-1 therapy in cHL have not been defined. Furthermore, there is a paucity of data linking phenotypic and functional T cell states with clonality at the single cell level in cHL. To address this knowledge gap, we performed paired single cell RNA and T cell receptor sequencing (scRNA/TCR-seq) on 14 cHL and 5 reactive lymphoid tissue (RLT) specimens. Recurrent and significant clonal expansion within effector and exhausted CD8+ T cell and regulatory T (TREG) cell clusters was uniquely observed in most cHL specimens. Surprisingly, CD8+ T cells exhibiting transcriptional features of exhaustion were relatively uncommon in cHL specimens. Multi-plex flow cytometric analysis revealed maintained effector cytokine secretion by most conventional lymphoma-infiltrating T cells upon ex vivo restimulation, which argued against a profound T cell dysfunctional state in cHL, and suggested that lymphoma-iniltrating T cells, when removed from the immune suppressive cHL environment, are capable of exerting effector functions. Finally, these results raise new questions about the nature of the T cells that mediate response to PD-1 blockade therapy in patients with cHL.

overall_design: Single cell RNA and TCR sequencing analysis of 14 Hodgkin lymphoma samples and 5 reactive lymph tissue samples.

first: Jovian

last: Yu


title: Dual targeting of the androgen receptor and PI3K/AKT/mTOR pathways in prostate cancer elicits improved anti-tumor efficacy and apoptosis

city: Berlin

last: Haendler

address: Muellerstr. 178

institute: Bayer AG

summary: Both the PI3K/AKT/mTOR and androgen receptor (AR) signaling pathways play essential roles in prostate cancer. We compared the impact of PI3K/AKT/mTOR pathway inhibitors with different selectivity profiles on in vitro cell proliferation and apoptosis induction, and observed the strongest effects in androgen-sensitive prostate cancer models. Combination treatment with the AR inhibitor darolutamide led to enhanced apoptosis in these cell lines, with most pronounced effects of the pan-PI3K inhibitor copanlisib. Transcriptomic analysis performed in the androgen-sensitive VCaP cell line revealed that gene expression was more affected by the co-treatment with darolutamide and copanlisib, compared to single agents. A comprehensive reversal of the androgen response and the mTORC1 transcriptional programs was observed. The combination treatment also markedly induced DNA damage. An in vivo efficacy study performed using LuCaP 35, an androgen-sensitive patient-derived prostate cancer model, indicated superior efficacy after combined treatment with copanlisib and darolutamide. Immunohistochemistry analysis of treated tumors furthermore showed increased apoptosis. Altogether these data demonstrate that blocking the PI3K/AKT/mTOR and AR pathways with potent inhibitors has superior anti-tumor efficacy and induces apoptosis in androgen-sensitive prostate cancer models.

overall_design: RNA-seq of VCaP cells treated by DMSO control, synthetic androgen R1881, AR inhibitor sarolutamide, the pan-PI3K inhibitor copanlisib and their combination for 24 and 48 hours.

first: Ekaterina

last: Nevedomskaya


title: SMARCAL1 is a dual regulator of innate immune signaling and PD-L1 expression that promotes tumor immune evasion.

first: Alberto

last: Ciccia

address: 1130 St. Nicholas avenue, ICRC Room 407B

institute: The Trustees of Columbia University in the City of New York

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

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Refer to individual Series


title: SMARCAL1 is a dual regulator of innate immune signaling and PD-L1 expression that promotes tumor immune evasion [RNA-seq]

summary: Genomic instability can trigger cancer-intrinsic innate immune responses that promote tumor rejection. However, cancer cells often evade these responses by overexpressing immune checkpoint regulators, such as PD-L1. Here, we identify the SNF2-family DNA translocase SMARCAL1 as a factor that favors tumor immune evasion by a dual mechanism involving both the suppression of innate immune signaling and the induction of PD-L1-mediated immune checkpoint responses. Mechanistically, SMARCAL1 relieves endogenous DNA damage and suppresses cGAS-STING-dependent signaling during cancer cell growth. Simultaneously, it cooperates with the AP-1 family member JUN to maintain chromatin accessibility at a transcriptional regulatory element in the PD-L1 gene, thereby promoting PD-L1 expression in cancer cells. SMARCAL1 loss hinders the ability of tumor cells to induce PD-L1 in response to genomic instability, enhances anti-tumor immune responses and sensitizes tumors to immune checkpoint blockade in a mouse melanoma model. Collectively, these studies uncover SMARCAL1 as a promising target for cancer immunotherapy.

overall_design: Gene expression profiling analysis using data from RNA-seq and plate-seq generated from SMARCAL1-proficient and -deficient MDA-MB-436 cells

first: Giuseppe

last: Leuzzi

first: Alessandro

last: Vasciaveo

first: Andrea

last: Califano


title: Cis inhibition of NOTCH1 through JAGGED1 sustains embryonic Hematopoietic stem cell fate [RNA-seq]

city: Barcelona

first: Anna

last: Bigas

state: Barcelona

address: Dr. Aiguader 88

department: Cancer Research

country: Spain

institute: Institut Hospital del Mar d'Investigacions Mèdiques

summary:
Hematopoietic stem cells (HSCs) develop from the hemogenic endothelium (HE) in the aorta- gonads-and mesonephros (AGM) region and reside within Intra-aortic hematopoietic clusters (IAHC) along with hematopoietic progenitors (HPC). The signalling mechanisms that distinguish HSCs from HPCs are unknown. Notch signaling is essential for arterial specification, IAHC formation and HSC activity, but current studies on how Notch segregates these different fates are inconsistent.
We now demonstrate that Notch activity is highest in a subset of, GFI1+, HSC-primed HE cells, and is gradually lost with HSC maturation. We uncover that the HSC phenotype is maintained due to increasing levels of NOTCH1 and JAG1 interactions on the surface of the same cell (cis) that renders the NOTCH1 receptor from being activated. Forced activation of the NOTCH1 receptor in IAHC activates a hematopoietic differentiation program. Our results indicate that NOTCH1-JAG1 cis-inhibition preserves the HSC phenotype in the hematopoietic clusters of the embryonic aorta.

overall_design: We performed RNAseq in AGM GFI1+IAHC (CD31+cKIT+GFI1+) FACS sorted after compound E treatment overnight. The following morning, sample was washed out and separated in sets of 300-400 cells for  incubation for 4 hours in methylcellulose with the addition EU with DMSO,  10mM of compound E or recombinant Fc-JAG1.   A total of three washed out, two CompoundE and two Fc-JAG1 samples  were analysed.

first: Roshana

last: Thambyrajah

first: Maria

last: Maqueda

first: Wen

last: Hao Neo

first: Kathleen

last: Imbach

first: Yolanda

last: Guillen

first: Daniela

last: Grases

first: Zaki

last: Fadlullah

first: Stefano

last: Gambera

first: Francesca

last: Matteini

first: Xiaonan

last: Wang

first: Fernando

last: Calero

first: Manel

last: Esteller

last: Florian

first: Eduard

last: Porta

first: Rui

last: Benedito

first: Berthold

last: Göttgens

first: Georges

last: Lacaud

first: Lluis

last: Espinosa


title: NRF2-independent iron-triggered signaling via MAPK p38-ETS1 regulates LSEC Bmp6 mRNA expression [II]

status: Public on Jan 25 2024

city: Warsaw

first: Michal

last: Mikula

address: Roentgena 5

institute: Maria Skłodowska-Curie National Research Institute of Oncology

summary: BMP6 is an iron-sensing cytokine produced by liver sinusoidal endothelial cells (LSECs). The transcription factor NRF2 was proposed to induce Bmp6 expression in response to iron-triggered oxidative stress, and NRF2 knock-out mice were shown not capable of regulating Bmp6 in response to massive diet-imposed iron overload. However, LSEC Bmp6 expression levels in the hemochromatosis mouse model reflect hepatocytic, not endothelial iron content. Hence, it is not fully resolved how iron signals translate into alterations of Bmp6 mRNA levels. To further explore the mechanisms of Bmp6 regulation, we employed female mice aged 10-11 months that are hallmarked by hepatocytic, but not LSEC iron accumulation and decreased transferrin saturation, hinting at the absence of systemic iron overload. We found that LSECs of aged mice exhibit increased Bmp6 levels as compared to young controls, but do not show oxidative stress or a transcriptional signature characteristic of activated NFR2-mediated signaling in FACS-sorted LSECs. We further observed that primary murine LSECs derived from both wild-type and NRF2 knock-out mice induce Bmp6 in response to acute iron exposure. By analyzing RNA sequencing data of FACS-sorted LSECs from aged versus young mice, as well as upon acute injections of iron citrate in young mice, we identified ETS1 as a candidate transcription factor involved in Bmp6 regulation. By conducting siRNA-mediated knock-down and small-molecule treatments in primary LSECs, we show that Bmp6 transcription is regulated in an NRF2-independent manner via ETS1, which is activated downstream of  p38 MAP kinase-mediated signaling. This knowledge expands the understanding of Bmp6 transcriptional control in both iron-triggered oxidative stress and under the conditions uncoupled to LSECs reactive oxygen species levels.

overall_design: We analyzed in total 7 versus 7 RNA samples derived from FAC-sorted LSECs obtained from young (8-10 weeks old) and aged (10-11 months old) female C57BL6J mice, respectively. The experiemnt was performed in two biological replicates (3 vs 3, and 4 vs 4).

first: Gabriela

last: Zurawska

first: Aneta

last: Jończy

first: Zuzanna

last: Sas

first: Izabela

last: Rumieńczyk

last: Kulecka

first: Katarzyna

last: Piwocka

first: Tomasz

last: Rygiel

first: Michał

last: Mleczko-Sanecka


title: Single-cell transcriptome Atlas reveals embryonic endothelial heterogeneity

city: Jamaica Plain

first: Xiaoran

last: Zhang

address: 14 Perkins Square, Apt 11

department: Cardiology

institute: Boston Children's Hospital

summary: Mature endothelial cells (ECs) are heterogeneous, with subtypes defined by tissue origin and by position within the vascular bed. Here, we performed scRNA-seq with mouse embryonic ECs and identified 19 subclusters, including Etv2+Bnip3+ early EC progenitors. Most of these subtypes were grouped by their vascular-bed types, while ECs from brain, heart and liver were grouped by their tissue origins. Compared to arterial ECs (aECs), embryonic venous (vECs) and capillary ECs (cECs) shared less markers with their adult counterparts. cECs showed some venous characteristics. One cEC cluster with both venous and capillary features served as a branch point for aEC and vEC lineages. aECs and vECs showed distinct transcriptional regulatory networks.

overall_design: Cdh5(PAC)-CreERT2 mice were bred with R26mTmG mice. After tamoxifen induction, ECs were enriched from E9.5, E12.5 and E15.5 embryos by magnetic-activated cell sorting (MACS), then scRNA-seq was performed with droplet-based tool (10x Genomics).

first: Jian

last: Chen

first: Daniel

last: DeLaughter

first: William

last: Pu


title: SMARCAL1 is a dual regulator of innate immune signaling and PD-L1 expression that promotes tumor immune evasion [ATAC-seq]

first: Alberto

last: Ciccia

overall_design: Chromatin accessibility analysis using ATAC-seq data generated from  SMARCAL1-proficient and -deficient MDA-MB-436 and B16F10 cells.

first: Giuseppe

last: Leuzzi

first: Xiao

first: Chao

last: Lu


title: Complex transcription factor networks and sex hormones synergically regulate sexually dimorphic gene expression in PiC neurons

city: Guangzhou

first: Boxing

last: Li

address: Zhongshan Erlu 74

institute: Sun Yat-sen University

summary: Investigate cellular and molecular control of sexually dimorphic behaviours in PiC neurons

overall_design: 4-5 EGFP-labeled cells in PiC retrograded from PrL or MeA were collected in one tube to perform RNA-seq.

first: Shunchang

last: Fang

first: Zicheng

last: Wei

first: Hongyang

first: Yuxin

last: Qin


title: SMARCAL1 is a dual regulator of innate immune signaling and PD-L1 expression that promotes tumor immune evasion [CUT&RUN]

overall_design: Cleavage under targets and Release using nuclease (Cut-and-Run), for the chromatin profiling of SMARCAL1, JUN, H3K4me3 and H3K24me3 proteins in MDA-MB-436 cell lines treated as indicated.

first: Tessa

last: Firestone

first: Allison

last: Hickman

first: Bryan

last: Venters

first: Michael-Christopher

last: Keogh

first: Lu

last: Sun


title: PFG controls gene expression in female gametocytes cooperatively with AP2-FG [PFG_ChIP-seq]

city: tsu

first: masao

last: yuda

state: mie

address: edobashi 2-174

country: Japan

institute: mie university

summary:
AP2-FG is a female-specific transcription factor (TF) that plays an essential role in female gametocyte development. AP2-FG activates hundreds of genes by binding to a female-specific ten-base cis-acting element. Here, we report that in the rodent malaria parasite Plasmodium berghei, another female-specific TF designated as a partner of AP2-FG (PFG), controls gene expression in female gametocytes cooperatively with AP2-FG.
Transcriptional mechanisms were analyzed in Plasmodium berghei female gametocytes.

overall_design: PFG_ChIP-seq, PFG_ChIP-seq in AP2-FG(-),  AP2-FG_ChIP-seq in PFG(-)

first: Masao

last: Yuda

first: Izumi

last: Kaneko

first: Shiroh

last: Iwanaga

first: Tsubasa

last: Nishi

first: Yuho

last: Murata


title: Myeloid Zfhx3 Deficiency Protects Against Influenza A Virus-induced Lung Injury and Mortality Increased by Hypercapnia

status: Public on Jan 21 2024

first: Marina

middle: Susana

last: Casalino-Matsuda

address: 303 E Superior

department: Medicine

institute: Northwestern University

summary: Zfhx3, human ortholog of the Drosophila Zfh2 and first known mediator of hypercapnic immune suppression in vivo, is expressed in human and mouse MØs and its myeloid deletion reversed the effects of hypercapnia in gene expression in murine alveolar macrophages and protects against influenza A virus proliferation, lung injury and mortality increased by hypercapnia in infected mice.

overall_design: Transcriptomic profiles of flow sorted alveolar epithelial type 2 (AT2) cells from wild type and myeloid Zfhx3 deficient C57Bl/6 mice exposed to room air or 10% CO2 for 7 days were generated by standard illumina RNA-Seq protocols.

first: Hiroaki

last: Matsuda

first: Peter

last: Sporn


title: PFG controls gene expression in female gametocytes cooperatively with AP2-FG


title: Epigenetic regulation of CD38/CD48 by KDM6A mediates NK cell response in multiple myeloma

first: Jiye

last: Liu

address: 450 Brookline Ave

department: Medical Oncology

institute: Dana-Farber Cancer Institute


title: Cis inhibition of NOTCH1 through JAGGED1 sustains embryonic Hematopoietic stem cell fate [scRNA-seq]

first: Anna

last: Bigas

overall_design: We performed single cell RNA seq with sorted AGM E11.5 and E10.5 CD31+ cKit+ GFI1+ with a total of 860 cells in order to define HSCs at a molecular level.

first: Roshana

last: Thambyrajah

first: Maria

last: Maqueda

first: Wen

last: Hao Neo

first: Kathleen

last: Imbach

first: Yolanda

last: Guillen

first: Daniela

last: Grases

first: Zaki

last: Fadlullah

first: Stefano

last: Gambera

first: Francesca

last: Matteini

first: Xiaonan

last: Wang

first: Fernando

last: Calero

first: Manel

last: Esteller

last: Florian

first: Eduard

last: Porta

first: Rui

last: Benedito

first: Berthold

last: Göttgens

first: Georges

last: Lacaud

first: Lluis

last: Espinosa


title: Monocytes release pro-cathepsin D to drive blood-to-brain transcytosis in diabetes

city: Shenbei

first: Weidong

last: Zhao

state: Liaoning

address: 77 Puhe Road, Shenbei New District

department: Department of Developmental Cell Biology

institute: China Medical University

summary: Microvascular complications are the major outcome of type 2 diabetes mellitus progression and the underlying mechanism remains to be determined. To explore the contribution of circulating monocytes to the microvascular lesions, high-throughput RNA sequencing was conducted and we found high-level expression of cathepsin D (CTSD) in the monocytes of type 2 diabetes mellitus patients. The transgenic mice expressing human CTSD in the monocytes showed increased brain microvascular permeability resembling the diabetic microvascular phenotype, accompanied with cognitive deficit. Mechanistically, we found the monocytes release non-enzymatic pro-CTSD to upregulate caveolin expression in brain endothelium triggering caveolae-mediated transcytosis, without affecting the paracellular route of brain microvasculature. The circulating pro-CTSD activated the caveolae-mediated transcytosis in brain endothelial cells via its binding with low density lipoprotein receptor-related protein 1 (LRP1). Importantly, genetic ablation of CTSD in the monocytes exhibited protective effect against the diabetes-enhanced brain microvascular transcytosis and the diabetes-induced cognitive impairment. Our findings thus uncover the novel role of circulatory pro-CTSD from monocytes in the pathogenesis of cerebral microvascular lesions in diabetes. The circulatory pro-CTSD is thus a potential target for the intervention of microvascular complications in diabetes.

overall_design: To evaluate the transcriptional changes in the circulating monocytes of T2DM patients compared with healthy controls

first: Dan

first: Zeng-Kang

last: Huang

first: Yu

last: Liang

first: Wei-Dong


title: Effect of metabolite intervention in mice muscle gene expression

city: San Diego

first: REYNA

last: HERNANDEZ-BENITEZ

state: California

address: 5510 MOREHOUSE DR, FLOOR 3

department: SDI

institute: ALTOS LABS INC.

summary:
To investigate the effects of 1C-metabolite cocktail on the muscle quadriceps and determine the changes in gene expression. Particularly looking at potential insights of what the metabolite intervention induces in vivo.
We then performed gene expression profiling analysis using data obtained from RNA-seq of 5 biological replicates conditions including control  and  supplemented mice with 1C-MIM during 3 months

overall_design: Comparative gene expression profiling analysis of RNA-seq data for mice control and supplemented

first: Reyna

last: Hernandez-Benitez

first: Juan

last: Izpisua-Belmonte


title: Human induced pluripotent stem cells are resistant to human cytomegalovirus infection primarily at the attachment level due to the reduced expression of cell-surface heparan sulfate

status: Public on Jan 19 2024

city: Hamamatsu-shi

first: Hideya

last: Kawasaki

state: Shizuoka-ken

institute: Hamamatsu University School of Medicine

summary: Cytomegalovirus (CMV), a type of herpes virus, is the predominant cause of congenital anomalies due to intrauterine infections in humans. Adverse outcomes related to intrauterine infections with human CMV (HCMV) vary widely depending on factors, such as fetal infection timing, infection route, and viral virulence. The precise mechanism underlying HCMV susceptibility remains unclear. In this study, we compared the susceptibility of neonatal human dermal fibroblast cells (NHDFCs) and human induced pluripotent stem cells (hiPSCs) derived from NHDFCs, which are genetically identical to HCMV, using immunostaining, microarray, in situ hybridization, quantitative PCR, and scanning electron microscopy. These cells were previously used to compare CMV susceptibility, but the underlying mechanisms were not fully elucidated. HCMV susceptibility of hiPSCs was significantly lower in the earliest phase. No shared gene ontologies were observed immediately post-infection between the two cell types, using microarray analysis. Early-stage expression of HCMV antigens and the HCMV genome was minimal in immunostaining and in in situ hybridization in hiPSCs. This strongly suggests that HCMV does not readily bind to hiPSC surfaces. Scanning electron microscopy performed using the NanoSuit method confirmed the scarcity of HCMV particles on hiPSC surfaces. The zeta potential and charge mapping of the charged surface in NHDFCs and hiPSCs exhibited minimal differences when assessed using zeta potential analyzer and scanning ion conductance microscopy; however, the expression of heparan sulfate (HS) was significantly lower in hiPSCs compared with that in NHDFCs. Thus, HS expression could be a primary determinant of HCMV resistance in hiPSCs at the attachment level.

overall_design: Control cells (NHDFCs and hiPSCs) and HCMV-infected cells (NHDFCs and hiPSCs) 30 min post infection at an MOI of 10.

first: Takahiko

last: Hariyama

first: Isao

last: Kosugi

first: Shiori

last: Meguro

first: Futoshi

last: Iwata

first: Kosuke

last: Shimizu

first: Yasuhiro

last: Magata

first: Toshihide

last: Iwashita


title: Dose escalation of Wnt stimulation during the exit from pluripotency identifies tranilast as a small molecule regulator of cardiac mesoderm differentiation (ATAC-Seq)

city: Birsbane

first: Nathan

middle: J.

last: Palpant

state: QLD

address: 306 Carmody Road

country: Australia

institute: University of Queensland

summary: Wnt signalling is a critical determinant of cell lineage development. This study used Wnt dose-dependent induction programs to gain insights into molecular regulation of stem cell differentiation. We performed single cell RNA-sequencing of cells responding to a dose-escalation protocol with Wnt agonist CHIR-99021 during the exit from pluripotency to identify cell types and genetic activity driven by Wnt stimulation. Results of activated gene sets and cell types were then used to build a multiple regression model that predicts the efficiency of cardiomyocyte differentiation. Cross-referencing Wnt-associated gene expression profiles to the Connectivity Map database, we identified a novel property of the small molecule drug, tranilast. We found that tranilast synergistically activates Wnt signalling to promote cardiac lineage differentiation which we validate in vitro and in vivo. Our study provides an integrated workflow that links experimental datasets, prediction models and small molecule databases to identify novel drug-like compounds that control cell differentiation.

overall_design: WTC-11 cells differentiated under different CHIR concentrations +/- tranilast were collected on day 2 of differentiation for Omni-ATAC-seq.We then analysed chromatin accessibility and motif enrichment to understand the mechanisms of tranilast.


title: Myeloid Zfhx3 Deficiency Protects Against Hypercapnia-induced Suppression of Host Defense Against Influenza A Virus

first: Marina

last: Casalino-Matsuda

overall_design: Transcriptomic profiles of flow sorted alveolar macrophages cells from wild type and myeloid Zfhx3 deficient C57Bl/6 mice exposed to room air or 10%  CO2 for 7 days were generated by standard illumina RNA-Seq protocols.

first: Hiroaki

last: Matsuda

first: Peter

last: Sporn


title: Effect of metabolites supplementation on C2C12 gene expression

summary:
To investigate the effects of 1C-metabolite cocktail on C2C12 differentiated myofibers in vitro and determine the changes in gene expression. Particularly looking at potential insights of what the metabolite intervention induces in vitro
We then performed gene expression profiling analysis using data obtained from RNA-seq of 3 different conditions including control myofibers untreated, myofibers treated with 1C-MIM, and myofibers treated with a scramble metabolite supplementation

overall_design: Comparative gene expression profiling analysis of RNA-seq data for C2C12 differentiated myofibers and the conditions in which were supplemented


title: The unique dual targeting of AGO1 by two types of PRMT enzymes promotes phasiRNA loading in Arabidopsis thaliana

city: Perpignan

first: Jacinthe

last: Azevedo Favory

address: 58 avenue Paul Alduy

institute: CNRS

summary: Arginine / R methylation (R-met) of proteins is a widespread post-translational modification (PTM), deposited by a family of protein arginine / R methyl transferase enzymes (PRMT). Regulations by R-met are involved in key biological processes deeply studied in metazoan. Among those, post-transcriptional gene silencing (PTGS) can be regulated by R-met in animals and in plants. It mainly contributes to safeguard processes as protection of genome integrity in germ lines through the regulation of piRNA pathway in metazoan, or response to bacterial infection through the control of AGO2 in plants. So far, only PRMT5 has been identified as the AGO / PIWI R-met writer in higher eukaryotes. We uncovered that AGO1, the main PTGS effector regulating plant development, contains unique R-met features among the AGO / PIWI superfamily, and outstanding in eukaryotes. Indeed, AGO1 contains both symmetric (sDMA) and asymmetric (aDMA) R-dimethylations and is dually targeted by PRMT5 and by another type I PRMT in Arabidopsis thaliana . We showed also that loss of sDMA didn’t compromise AtAGO1 subcellular trafficking in planta. Interestingly, we underscored that AtPRMT5 specifically promotes the loading of a set of phasiRNA in AtAGO1. All our observations bring to consider this dual regulation of AtAGO1 in plant development and response to environment, and pinpoint the complexity of AGO1 post-translational regulation.

overall_design: To investigate the impact of atprmt5 mutations on total sRNA accumulation and AtAGO1 sRNA loading in reproductive tissues. To do so, we carried out anti-AGO1 immunoprecipitations performed on close buds of A. thaliana, using Col0, atprmt5-2 and atprmt5-6 mutants. A fraction of inputs (total extracts) and all the IP products are used for sRNA extraction. The experiment includes biological duplicates.

first: Clément

last: Barre-Villeneuve

first: Marie-Christine

last: Carpentier

first: Michèle

last: Laudié

first: Lauriane

last: Kuhne

first: Thierry

last: Lagrange


title: PFG controls gene expression in female gametocytes cooperatively with AP2-FG [AP2-FG_DIP]

first: masao

last: yuda

overall_design:
AP2-FG_DIP
The recombinant AP2 domain of AP2-FG was prepared as a GST-fused protein. This recombinant GST-fused AP2 domain of AP2-FG and genomic DNA fragments were incubated at RT for 30 min in 100 μL of binding/washing buffer. The mixture was then incubated with 50 μL glutathione sepharose resin (Cytiva) at RT with rotation. Resin was washed three times with 150 μL of binding/washing buffer, and bound protein was eluted with 10 mM glutathione solution. Input genomic DNA fragments were also sequenced as a control.

first: Masao

last: Yuda

first: Izumi

last: Kaneko

first: Shiroh

last: Iwanaga

first: Tsubasa

last: Nishi

first: Yuho

last: Murata


title: PFG controls gene expression in female gametocytes cooperatively with AP2-FG [PFG_RNA-seq]

summary: AP2-FG is a female-specific transcription factor (TF) that plays an essential role in female gametocyte development. AP2-FG activates hundreds of genes by binding to a female-specific ten-base cis-acting element. Here, we report that in the rodent malaria parasite Plasmodium berghei, another female-specific TF designated as a partner of AP2-FG (PFG), controls gene expression in female gametocytes cooperatively with AP2-FG.

overall_design: Transcriptional mechanisms were analyzed in Plasmodium berghei female gametocytes.


title: The histone acetylation reader ENL is required for oncogenic transcription driven by super-enhancer and represents a synergistic vulnerability for BET inhibition [ChIP-Seq II]

status: Public on Jan 18 2024

city: Shenzhen

first: Xing-sheng

last: Shu

state: Guangdong

address: No 3688, Nanhai Avenue

department: Medical School

institute: Shenzhen University

summary: The histone acetylation reader ENL is identifed as a key factor controlling super-enhancer-driven transcription. Therefore, we constructed a targeted degradation system of endogenous ENL protein and investigated the roles of ENL in regulating oncogenic transcription through epigenetic reprogramming.

overall_design: The ENL gene was taggged by a auxin-inducible degron (AID) in HCT116 colon cancer cells stably expressing E3 ligase AtAFB2. The HCT116 ENL-AID cells was treated with or without 500 um auxin (IAA) for 3 hours and an additional group of IAA treated cells were subjected to PBS washout and subsequent culture in normal medieum for another 48 hours . The resulting cells were harvested for analysis as described below.

first: Xingsheng


title: SHIFTR enables the unbiased identification of proteins bound to specific RNA regions in live cells

city: Würzburg

first: Alexander

last: Gabel

address: Josef-Schneider-Str. 2 / D15

institute: Helmholtz Institute for RNA-based Infection Research

summary: RNA-protein interactions determine the cellular fate of RNA and are central to regulating gene expression outcomes in health and disease. To date, no method exists that is able to identify proteins that interact with specific regions within endogenous RNAs in live cells. Here, we develop SHIFTR (Selective RNase H-mediated interactome framing for target RNA regions), an efficient and scalable approach to identify proteins bound to selected regions within endogenous RNAs using mass spectrometry. Compared to state-of-the-art techniques, SHIFTR is superior in accuracy, captures minimal background interactions and requires orders of magnitude lower input material. We establish SHIFTR workflows for targeting RNA classes of different length and abundance, including short and long non-coding RNAs, as well as mRNAs and demonstrate that SHIFTR is compatible with sequentially mapping interactomes for multiple target RNAs in a single experiment. Using SHIFTR, we comprehensively identify interactions of cis-regulatory elements located at the 5ʹ and 3ʹ-terminal regions of the authentic SARS-CoV-2 RNA genome in infected cells and accurately recover known and novel interactions linked to the function of these viral RNA elements. SHIFTR enables the systematic mapping of region-resolved RNA interactomes for any RNA in any cell type and has the potential to revolutionize our understanding of transcriptomes and their regulation.

overall_design: We mapped direct protein-RNA interaction sites of the human proteins LARP4 and ZAP, as well as the viral protein NSP9 in SARS-CoV-2 infected cells using enhanced crosslinking and immunoprecipitation (eCLIP) in combination with RNA-sequencing.

first: Mathias

last: Munschauer


title: Next Generation Sequencing of Transcriptomes of PM2.5-exposed Mice

city: Shanghai

first: Ting

last: Xu

address: No. 1239 Road Siping

institute: Tongji University

summary:
Purpose: The goal of this study is to investigate the effects of fine particulate matter (PM2.5) exposure on mouse olfactory bulb using next generation sequencing (NGS).
Methods: After 28 days of 3 mg/kg/3 day and 10 mg/kg/3 day PM2.5 exposure, olfactory bulb mRNA profiles of 8-week-old wild-type (WT) male C57BL/6 mice were generated by deep sequencing, in 3-4 repeats, using Illumina NovaSeq 6000. For each sample, clean reads were obtained that mapped to mm10 using HISAT2 (hierarchical indexing for spliced alignment of transcripts) v2.0.477.
Results: Our study revealed that PM2.5 treatment caused significant effects on the gene expression profilling of mouse olfactory bulb. Overall, the sequencing identified 34,745 transcripts, and two kinds of treatments obtained 60 and 138 differently expressed genes (DEGs) respectively, with a criteria of fold change >2 and q-value <0.5. Most biological events that DEGs involved were inflammation relevant.
Conclusions: Our study revealed that PM2.5 treatment caused significant effects on the gene expression profiling of mouse olfactory bulb.

overall_design: Olfactory bulb mRNA profiles of 8 week old wild type (WT), male C57BL/6J mice


title: Effect of Embigin on LTA1 induced-CD4+ TRM cells.

city: New Orleans

first: Jay

last: Kolls

state: LA

address: 1430 Tulane Ave

institute: Tulane University

summary:
To investigate the function of Embigin on LTA1 induced-CD4+TRM cells, we immunized CD4cre x Embfl/fl mice with OmpX and LTA1 at Day0 and Day21, and isolated CD4+ cells from Embfl/fl CD4cre-(WT) and Embfl/fl CD4cre+(KO) mice from the lung by magnetic beads.
We then performed gene expression profiling analysis using data obtained from RNA-seq of CD4+cells from 5 WT and 5 KO mice.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for CD4+cells from OmpX+LTA1 immunzied Embfl/fl CD4cre-(WT) and Embfl/fl CD4cre+(KO) mice.

first: Kejing

last: Song


title: Dose escalation of Wnt stimulation during the exit from pluripotency identifies tranilast as a small molecule regulator of cardiac mesoderm differentiation (scRNA-Seq)

first: Nathan

last: Palpant

overall_design: Barcoded WTC-11 cells differentiated under different CHIR concentrations were harvested on day 2 and pooled together. Single live cells were collected and FACS sorted for a single scRNA-seq run.


title: RNF20 contributes to epigenetic immunosuppression through CDK9-dependent LSD1 stablization

city: lexington

first: YADI

last: WU

state: KY

address: 741 S.Limestone

institute: University of kentucky

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

overall_design: Refer to individual Series


title: The histone acetylation reader ENL is required for oncogenic transcription driven by super-enhancer and represents a synergistic vulnerability for BET inhibition [RNA-Seq II]

summary: The histone acetylation reader ENL is identifed as a key factor controlling super-enhancer-driven transcription. We thus perturbed BRD4, a well-known super-enhancer regulator, protein expression with a well-validated BRD4 PROTAC (proteolysis targeting chimeric) compound dBET1 and examined whether BRD4 was required for proper function of ENL in regulation gene transcription.

overall_design: The ENL-AID HCT116 cells were treated with or without 5μm dBET1 for 6 hours . The resulting cells were harvested for analysis as described below.


title: The role of the C. albicans transcriptional repressor NRG1 during filamentation and disseminated candidiasis is strain-dependent

status: Public on Jan 20 2024

city: Iowa City

first: Rohan

middle: S.

last: Wakade

state: Iowa

address: 25, South Grand Avenue, 2020 MedLabs, Department of Pediatrics

department: Department of Pediatrics

institute: University of Iowa

summary: Clinical isolates of the human fungal pathogen Candida albicans show significant variation in their ability to undergo in vitro filamentation. In this study, we show that Nrg1, a key repressor of filamentation and filament specific gene expression in standard laboratory strain, has strain dependent functions, especially during infection.

overall_design: In this experiment we characterize the expression profile of Candida albicans clinical isolates and their respective nrg1 mutant strains by NanoString. The probe set contains 186 environmentally responsive gene sets. 3 biological replicates (expection is 94015, in which only 2 biological replicates were used) were used to examine the expression of gene of interest.

first: Melanie

last: Wellington

first: Damian

last: Krysan


title: RNF20 contributes to epigenetic immunosuppression through CDK9-dependent LSD1 stablization (ChIP-Seq)

summary: Cyclin-dependent kinase 9 (CDK9) plays a critical role in transcription initiation and is essential for maintaining gene silencing at heterochromatic loci. Inhibition of CDK9 increases sensitivity to immunotherapy, but the underlying mechanism remains unclear. We now report that RNF20 stabilizes LSD1 via K29-mediated ubiquitination, which is dependent on CDK9-mediated phosphorylation. This CDK9- and RNF20-dependent LSD1 stabilization is necessary for the demethylation of histone H3K4, then subsequent repression of endogenous retrovirus (ERVs), and an interferon response, leading to epigenetic immunosuppression. Moreover, we found that loss of RNF20 sensitizes cancer cells to the immune checkpoint inhibitor anti-PD-1 in vivo, and that this effect can be rescued by the expression of ectopic LSD1. Our findings are supported by the observation that RNF20 levels correlate with LSD1 levels in human breast cancer specimens. This study sheds light on the role of RNF20 in CDK9-dependent LSD1 stabilization, which is crucial for epigenetic silencing and immunosuppression. Our findings explore the potential importance of targeting the CDK9-RNF20-LSD1 axis in the development of new cancer therapies.

overall_design: ChIP-seq analysis of H3K4me2 to compare differences between control group and shRNF20 knockdown group

first: Yadi

last: Wu


title: Gene Expression profile at the single cell level of quiescent and activated muscle stem cells (MuSC) from Extraocular muscle (EOM) and Tibialis Anterior (TA) muscle

city: Paris

first: Glenda

middle: Evangelina

last: Comai

address: 25-28 rue du Dr Roux

institute: Institut Pasteur

summary:
Skeletal muscle stem cells (MuSCs) are recognised as functionally heterogeneous. Cranial MuSCs are reported to have greater proliferative and regenerative capacity when compared with those in the limb. A comprehensive understanding of the mechanisms
underlying this functional heterogeneity is lacking. Here, we have used clonal analysis, live imaging and scRNA-seq to identify crucial features that distinguish extraocular muscle (EOM) from limb muscle stem cell populations. A MyogenintdTom reporter showed that the increased proliferation capacity of EOM MuSCs correlates with deferred differentiation and lower expression of the myogenic commitment gene Myod. Unexpectedly, EOM MuSCs activated in vitro expressed a large array of extracellular matrix components
typical of mesenchymal non-muscle cells. Computational analysis underscored a distinct co-regulatory module, which is absent in limb MuSCs, as the driver of these features. The EOM transcription factor network, with Foxc1 as key player, appears to be hardwired to EOM identity as it persists during growth, disease and in vitro after several passages. Our findings shed light on how high-performing MuSCs regulate myogenic commitment by remodelling their local environment and adopting properties not generally associated with myogenic cells.

overall_design: MuSCs from Tg:Pax7-nGFP mice were isolated by Fluorescence-based active cell sorting (FACS) based on GFP fluorescence. Quiescent MuSCs were immediatly processed for single-cell RNA sequencing and activated MuSCs were put in culture for 4 days before sequencing.

first: Daniela

last: Di Girolamo

first: Maria

last: Benavente-Diaz

first: Alexandre

last: Grimaldi

first: Priscilla

last: Thomas Lopes

first: Melania

last: Murolo

first: Brendan

last: Evano

first: Stamatia

last: Gioftsidi

first: Vincent

last: Laville

first: Sebastien

last: Mella

first: Mao

last: Kuriki

first: Shahragim

last: Tajbakhsh


title: Ancestral allele of DNA polymerase gamma modifies antiviral tolerance

city: Helsinki

first: Yilin

last: Kang

state: UUSIMAA

address: Biomedicum Helsinki 1 Haartmaninkatu 8 (C526)

country: Finland

institute: University of Helsinki

summary: Mitochondria are arising as critical modulators of antiviral tolerance by releasing mtDNA/mtRNA fragments to cytoplasm upon infection, activating virus sensors and type-I interferon response. The relevance of these mechanisms for mitochondrial diseases remains understudied. Here, we investigated mitochondrial recessive ataxia syndrome (MIRAS), caused by a common European founder mutation in DNA polymerase gamma (POLG1). The patients homozygous for the MIRAS variant p.W748S show exceptionally variable ages-of-onset and symptoms, indicating unknown modifying factors contributing to disease manifestation. We report that the mitochondrial DNA (mtDNA) replicase POLG1 has a role in antiviral defence mechanisms to double-stranded DNA and positive-strand RNA virus infections (HSV-1, TBEV, SARS-CoV-2) and its p.W748S variant dampens innate immune responses. Our patient and knock-in mouse data show that p.W748S compromises mtDNA replisome stability, causing mtDNA depletion, aggravated by virus infection. Low mtDNA and mtRNA release to cytoplasm and slow interferon response in MIRAS allow an early replicative advantage for viruses, leading to augmented pro-inflammatory response, subacute loss of GABAergic neurons, liver inflammation and necrosis. A population databank of ~300,000 Finns demonstrates enrichment of immunodeficient traits in p.W748S carriers. Our evidence suggests that POLG1 defects compromise antiviral tolerance, triggering epilepsy and liver disease. The finding has important implications to mitochondrial disease spectrum, including epilepsy, ataxia, and parkinsonism.

overall_design:
To investigate the in vivo response of MIRAS to viral infection, we generated a MIRAS-knock-in mouse model and subjected the mice to TBEV infection. These animals carry a homozygous knock-in mutation and the accompanying polymorphic variant homologous to the human ancestral MIRAS allele (p.W726S+E1121G in mice and p.W748S+E1143G in humans).
We performed RNA sequencing on the mouse brain (cerebral cortex) isolated from the control (wildtype) and MIRAS mice at uninfected (baseline) condition and 4 days post TBEV infection (n=5 mice per condition).
Comparative gene expression analysis of RNA-seq data for control vs MIRAS mouse cerebral cortex at baseline versus Day 4 after TBEV infection

first: Jussi

last: Hepojoki

first: Rocio

last: Maldonado

first: Takayuki

last: Mito

first: Mügen

last: Terzioglu

first: Tuula

last: Manninen

first: Ravi

last: Kant

first: Sachin

last: Singh

first: Alaa

last: Othman

first: Rohit

last: Verma

first: Kirmo

last: Wartiovaara

first: Lauri

last: Kareinen

first: Nicola

last: Zamboni

last: Nyman

first: Anders

last: Paetau

first: Anja

last: Kipar

first: Olli

last: Vapalahti

first: Anu

last: Suomalainen


title: The histone acetylation reader ENL is required for oncogenic transcription driven by super-enhancer and represents a synergistic vulnerability for BET inhibition [RNA-seq]

first: Xing-sheng

last: Shu

first: Xingsheng


title: RNA helicase Ighmbp2 regulates THO complex to ensure cellular mRNA homeostasis

first: Florian

last: Erhard

address: Versbacher Str. 7

department: Institut für Virologie und Immunbiologie

institute: Julius-Maximilians-Universität Würzburg

summary: RNA helicases constitute a large protein family implicated in cellular RNA homeostasis and disease development. Here we show that the RNA helicase Ighmbp2, linked to the neuromuscular disorder SMARD1 (DSMA1) associates with polysomes and impacts on translation of cellular mRNAs containing short, GC-rich and highly structured 5’UTRs. Absence of Ighmbp2 causes ribosome stalling at the start codon of target mRNAs, leading to their reduced translation efficiency. The main mRNA targets of Ighmbp2-mediated regulation encode for components of the THO complex that links mRNA production and nuclear export. Accordingly, failure of Ighmbp2 regulation of the THO complex causes perturbations of the cellular transcriptome and its encoded proteome. Ablation of essential THO complex subunits phenocopies these perturbations. Thus, Ighmbp2 is an upstream regulator of the THO complex that affects cellular mRNA homeostasis. Of note, Ighmbp2 dependent regulation of the THO complex is also observed in astrocytes derived from SMARD1 patients, suggesting that de-regulated mRNA metabolism contributes to SMARD1 etiology and offers new avenues for developing alternative therapeutics to treating SMARD1.

overall_design: To investigate the effect of Ighmbp2 on translation, we performed RNASeq and ribosome profiling after knockdown of Ighmbp2. Additionally, to confirm ability of Ighmbp2's targets THOC to phenocopy Ighmbp2 deficiency, we performed knockdown of THOC1 or THOC2 or both and compared the transcriptome of these cells to control and Ighmbp2 knockdown conditions.

last: Hirmer

first: Utz

last: Fischer


title: Gene expression signatures in patients with posttraumatic stress disorder (PTSD) and healthy controls

city: Tokyo

first: Hiroaki

last: Hori

address: 4-1-1, Ogawahigashi, Kodaira

department: Department of Behavioral Medicine

institute: National Institute of Mental Health, National Center of Neurology and Psychiatry

summary: To investigate gene expression signatures in PTSD, we used microarray and performed blood-based transcriptome profiling. Our analyses identified key genes and molecular pathways that can be centrally involved in the etiology of PTSD.

overall_design: The blood sampling was performed around noon (before lunch), between 11:30 AM and 12:30 PM. Gene expression in blood was measured for 32 women with PTSD and 16 healthy control women.


title: Keystone pathobionts associated with colorectal cancer promote oncogenic reprograming

city: Ithaca

first: Joshua

last: Jones

address: 289 Kimball Hall

institute: Cornell University

summary: Fusobacterium nucleatum (Fn) and enterotoxigenic Bacteroides fragilis (ETBF) are two pathobionts consistently enriched in the gut microbiomes of patients with CRC compared to healthy counterparts and frequently observed for their direct association within tumors. Although several molecular mechanisms have been identified that directly link these organisms to features of CRC in specific cell types, their specific effects on the epithelium and local immune compartment are not well-understood. To fill this gap, we leveraged single-cell RNA sequencing (scRNA-seq) on wildtype mice and mouse model of CRC. We find that Fn and ETBF exacerbate cancer-like transcriptional phenotypes in transit-amplifying and mature enterocytes in a mouse model of CRC. We also observed increased T cells in the pathobiont-exposed mice, but these pathobiont-specific differences observed in wildtype mice were abrogated in the mouse model of CRC.  Although there are similarities in the responses provoked by each organism, we find pathobiont-specific effects in Myc-signaling and fatty acid metabolism. These findings support a role for Fn and ETBF in potentiating tumorigenesis via the induction of a cancer stem cell-like transit-amplifying and enterocyte population and the disruption of CTL cytotoxic function.

overall_design: Mouse intestinal cells underwent an a multi-enzyme dissociation, yielding a single cell suspension. We used this single-cell suspension for single cell RNA sequencing to analyze the host response to F. nucleatum and enterotoxigenic B. fragilis

first: Qiaojuan

last: Shi

first: Rahul

last: Nath

first: Ilana

last: Brito


title: ChIP-seq of pro-B cells to test the role of phase separation in EBF1 pioneering function

city: Freiburg

first: Nikolay

last: Zolotarev

address: Stübeweg, 51

department: Cellular and Molecular Immunology

institute: Max Planck Institute of Immunobiology and Epigenetics

summary: EBF1 is a transcription factor crucial for early B cell development. The phase separation ability of the EBF1 C-terminal domain (CTD) is essential for the transcription activation and pioneering function of EBF1. We identified the specific tyrosine residues in the CTD required for phase separation. We transduced pro-B cells with EBF1 wt, deletion of the entire CTD (dC), or mutation of these tyrosines (YmA). We performed ChIP-seq, ATAC-seq, and RNA-seq to see the effect of disrupting phase separation ability on DNA binding, chromatin opening, and target gene expression. Our findings confirm that phase separation is important for the EBF1 function.

overall_design: EBF1 and BRG1 ChIP-seq in EBF1 wt, dC, or YmA pro-B cells


title: RNA-seq of pro-B cells to test the role of phase separation in EBF1 pioneering function

overall_design: RNA-seq (ploy-A+) in EBF1 wt, dC, or YmA pro-B cells


title: RNF20 contributes to epigenetic immunosuppression through CDK9-dependent LSD1 stablization (RNA-Seq)

first: YADI

last: WU

overall_design: RNA-seq analysis of gene expression in human breast cancer cell line MDA-MB-231, comparing control group with RNF20 knockdown.

first: Yadi

last: Wu


title: Features associated with enhanced proliferation define lung tumors that respond to dual immune checkpoint and radiation therapy

city: New York

first: Olivier

last: Elemento

state: NY

address: 1305 York Ave

department: Biochemistry

institute: Weill Cornell Medicine

summary: In a randomized phase II trial, patients with early-stage non-small cell lung cancer were were treated with either two preoperative cycles of the anti-PD-L1 antibody durvalumab alone or combined with immunomodulatory doses of stereotactic radiation. We conducted an analysis of the gene expression profiles of treatment-naive tumors from these patients and identified a 140-gene signature that was correlated with a positive response to the dual therapy. This signature showed enrichment in genes associated with cell proliferation.

overall_design: RNA-seq studies of resected tumors to identify biomarkers of response in a clinical trial of immune checkpoint blockade therapy in the treatment of non-small cell lung cancer.

first: Nasser

middle: K

last: Altorki

first: Timothy

last: McGraw

first: Vivek

last: Mittal

first: Alain

last: Borczuk

first: Bhavneet

last: Bhinder


title: ATAC-seq data

city: Stanford

first: Feng

last: Pan

state: CA

address: 265 Campus Drive

department: Pathology

institute: Stanford University School of Medicine

summary:
We applied CRISPR gene-editing to induce t(4;11) chromosomal translocations in primary human umbilical cord blood (UCB) hematopoietic stem and progenitor cells (HSPCs) to faithfully model leukemias with the KMT2A-AFF1 fusion gene.
We then performed gene expression profiling analysis using data obtained from RNA-seq of different primary cells and human cell lines.

overall_design: ATAC-seq profiles of CRISPR-engineered MLL-AF4 BCP-ALL, patient samples, human cell lines.


title: ATAC-seq of pro-B cells to test the role of phase separation in EBF1 pioneering function

overall_design: ATAC-seq in EBF1 wt, dC, or YmA pro-B cells


title: Tetrahedral framework nucleic acids loaded siCCR2 restrains M1 macrophage polarization to ameliorate pulmonary fibrosis in chemoradiation-induced murine model

city: Chengdu

first: Mu

last: Yang

state: Sichuan Province

address: No. 55, Section 4, Renmin South Road

department: Centre for Translational Research in Cancer

institute: Sichuan Cancer Hospital & Institute, University of Electronic Science and Technology of China

summary: Idiopathic pulmonary fibrosis (IPF) is a chronic lethal disease in the absence of demonstrated efficacy for preventing progression. Although macrophage-mediated alveolitis is determined to participate myofibrotic transition during disease development, the paradigm of continuous macrophage polarization is still under-explored due to lack of proper animal-models. Here, by integrating 2.5 U/kg intratracheally Bleomycin administration and 10 Gray thorax irradiation at day 7, we generated a murine model with continuously alveolitis-mediated fibrosis, which mimics most of the clinical features of our involved IPF patients. In combination with data from scRNA-seq of patients and murine IPF model, a decisive role of CCL2/CCR2 axis in driving M1 macrophage polarization was revealed. And M1 macrophage was further confirmed to boost alveolitis in leading myofibroblast activation. Multiple sticky-end tetrahedral Framework Nucleic Acids conjunct with quadruple ccr2-siRNA (FNA-siCCR2) was synthesized in targeting M1 macrophages. FNA-siCCR2 successfully blocked macrophage accumulation in pulmonary parenchyma of IPF murine model thus prevent myofibroblast activation and led to disease remitting. Overall, our studies lay the groundwork to develop novel IPF murine model, reveal M1 macrophages as potential therapeutic target, and establish new treatment strategy by using FNA-siCCR2, which are highly relevant to clinical scenarios and translational researches in the field of IPF.

overall_design: The lung sample (n=1) from IPF patient was harvested and dissociated into single cells suspensions by mixed digestion buffer. Cells were collected after filtering through a 70 μm and 40 μm strainer. The viability and concentration of cells were calculated in a Counting Star system. Single-cell suspensions were loaded on the Chromium Single Cell Controller (10X genomics) for analysis, following the manufacturer’s instructions.

first: Lu

last: Guo

first: Chen

last: Li

first: Xiaorong

last: Feng

first: Xiaofei

last: Wang


title: Effect of traumatic brain injury on gene expression in bone.

status: Public on Jan 17 2024

city: Berlin

first: Denise

last: Jahn

state: Berlin

address: Augustenburger Platz 1

department: Julius Wolff Institut

institute: Charité - Universitätsmedizin Berlin

summary: It is clinically known, that patients often develop osteoporosis following traumatic brain injury (TBI). To enable a better understanding of the underlying mechanism, we developed a murine TBI model. Mice received a controlled cortical impact inducing a TBI. Expression analysis of the femur 3 days following the injury should reveal the regulated genes. We observed a strong downregulation of osteoblast genes, suggesting an inhibition of bone formation. Furthermore, the beta2-adrenoreceptor was downregulated, what is supposed to be induced by the increased adrenergic signaling in these mice.

overall_design: Mice received a controlled cortical impact inducing a TBI or served as uninjured controls. RNA was isolated from the femur 3 days following the injury.

first: Ellen

last: Otto

first: Paul

last: Köhli

first: Johannes

last: Keller

first: Serafeim

last: Tsitsilonis

first: Andranik

last: Ivanov

first: Mireille

last: Tchouto

first: Dieter

last: Beule


title: Human Otic progenitor cell models of congenital hearing loss applied to to Zika virus and cytomegalovirus infections

city: Cambridge

first: Charles

middle: Arthur

last: Whittaker

state: MA

institute: Koch Institute

summary: Congenital hearing loss is a common chronic condition affecting children in both developed and developing nations. In many cases, congenital hearing loss is ultimately attributed to viral infection, most often by cytomegalovirus (CMV), but also in Congenital Zika Syndrome (CZS). The mechanisms by which CMV and ZIKV virus cause these cranial developmental defects have not been elucidated. Inner ear development has been particularly difficult to study, given the inaccessibility and scarcity of the tissue in animal models or on human autopsy; however, it is now possible to culture stem-cell derived otic progenitor cells (OPCs). Here we describe successful in-vitro infection of OPCs with either CMV or ZIKV. We find that ZIKV infection rapidly and strongly induces the expression of type I IFN and inflammatory genes, while simultaneously decreasing otic cell viability in culture that is at least in part attributable to apoptosis. In contrast, CMV infection did not appear to elicit either of these effects and instead demonstrated a clear dysregulation of the expression of many key genes and pathways associated with inner ear development and function, including Cochlin, NGFR, SOX11 and TGF-β signaling. These findings suggest that ZIKV and CMV infections cause congenital hearing loss through very different pathways; that is, by killing progenitor cells in the case of ZIKV infection, and via disruption of critical developmental pathways in the case of CMV infection. In addition to demonstrating differential hCMV and ZIKV pathogenesis mechanisms in OPCs, this study highlights the advantages of otic progenitor cell models for the study of congenital hearing loss induced by viral infection.

overall_design: Zika and CMV infection of human stem-cell derived otic progenitor cells compared with Mock and heat-inactivated viral infection samples at different time points.

first: Ocwieja

last: Karen

first: Jeong

last: Minjin

first: Harding

last: Alfred

first: Zhang

last: Yichen

first: Jala

last: Nairuti

first: Stankovic

last: Konstantina

first: Gehrke

last: Lee


title: The histone acetylation reader ENL is required for oncogenic transcription driven by super-enhancer and represents a synergistic vulnerability for BET inhibition [ChIP-seq]

first: Xing-sheng

last: Shu

institute: Shenzhen University

summary: The histone acetylation reader ENL is identifed as a key factor controlling super-enhancer-driven transcription. Therefore, we constructed a targeted degradation system of endogenous ENL protein and investigated the roles of ENL in regulating oncogenic transcription through epigenetic reprogramming.

overall_design: The ENL gene was taggged by a auxin-inducible degron (AID) in HCT116 colon cancer cells stably expressing E3 ligase AtAFB2. The HCT116 ENL-AID cells was treated with or without 500 um auxin (IAA) for 3 hours and an additional group of IAA treated cells were subjected to PBS washout and subsequent culture in normal medieum for another 48 hours . The resulting cells were harvested for analysis as described below.

first: Xingsheng


title: KDM5C-mediated BRD4 recruitment is essential for active states of enhancers and BET inhibitor sensitivity [ChIP-seq]

city: Wuhan

first: Yulong

last: Qiang

address: 185 Donghu Road

institute: Wuhan University

summary:
A series of Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) experiments were conducted to investigate the effect and mechanism of KDM5C on the recruitment of BRD4 to chromatin, and the effect of this process on histone modification and gene expression.
We then performed ChIP-seq for BRD4, KDM5C, H3K27ac, H3K4me3, and H3K4me1 in HeLa and its derivatives cell lines.

overall_design: ChIP-seq for BRD4 in CaSki, HeLa parental, sgKDM5C, KDM5C_WT, KDM5C_H514K to investigate the effect of KDM5C on BRD4 peaks intensity. ChIP-seq for KDM5C, BRD4, H3K27ac, H3K4me3, H3K4me1 in HeLa parental and sgKDM5C cell lines to investigate the effect of KDM5C depletion on histone modification. ChIP-seq for H3K4me3 in HeLa parental, shBRD4, sgKDM5C, KDM5C_WT, KDM5C_H514K cell lines to explore the effect of BRD4 and KDM5C on H3K4me3 peak width. ChIP-seq for BRD4 in HeLa cells treated with DMSO, or JQ1, or compound 3b, or in combination to detect the synergistic effect of the two drugs on BRD4 peaks intensity.

first: Jiachen

last: Fan

first: Feng

last: Li


title: KDM5C-mediated BRD4 recruitment is essential for active states of enhancers and BET inhibitor sensitivity [RNA-seq]

summary:
To investigate the effect of the KDM5C depletion on transcription, we established HeLa cell lines in which KDM5C has been knocked off.
To explore the effect of synergistic lethal action of KDM5C PROTAC drug and BETi on transcription, we treated HeLa cells with JQ1, or compound 3b, or in combination.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for HeLa parental and sgKDM5C cell lines, as well as parental and three different treatment cells (JQ1, compound 3b, and in combination).


title: Effect of knocking out RNVU1-18 on gene expression

city: Suzhou

first: Fan

last: Zhang

address: 100 Chongwen Road

institute: Suzhou Institute of Systems Medicine

summary: Test transcripts in WT and RNVU1-18-/- A549 cells

overall_design: Gene expression profiling analysis of RNA-seq data for WT and RNVU1-18-/- A549 cells

first: Siying

last: Liu


title: Retinoids and EZH2 inhibitors cooperate to orchestrate cytotoxic effects on bladder cancer cells

city: İZMİR

first: Serap

last: Erkek Özhan

state: BALÇOVA

address: İzmir Biyotıp ve Genom Merkezi (IBG), Dokuz Eylül Üniversitesi Sağlık Yerleşkesi, Mithatpaşa Caddesi No:58/5

country: Turkey

institute: Izmir Biomedicine and Genome Center

summary: Emerging evidence has highlighted the importance of targeting EZH2 in bladder cancer owing to the highly mutated nature of bladder cancers harboring mutations in chromatin regulatory genes opposing Polycomb-mediated repression, with enhanced expression of EZH2 contributing to pathogenesis. Here we report that coordinated targeting of EZH2 and the retinoic acid signaling pathway caused the cytotoxic effects on bladder cancer cells by inducing a synergistic reduction in proliferative potential that was associated with increased apoptosis and cell cycle arrest in a cooperative manner. Moreover, combined treatment caused the modulation of the expression of certain genes towards an anti-oncogenic profile, as reflected by the stimulation of marker genes associated with apoptosis and differentiation. To investigate the effects on gene regulatory networks associated with fenretinide and GSK-126 treatments at a genome-wide scale, we performed gene expression profiling analysis using data obtained from RNA-seq of T24 cells. We further portrayed a molecular mechanism whereby EZH2 maintains H3K27me3-mediated repression of certain genes associated with unfolded protein response and some metabolic processes. This work also characterized an apoptotic program centered on the master transcriptional regulators C/EBPβ and CHOP. Our findings encourage the design of novel treatments employing retinoids coupled with EZH2 inhibitors in bladder carcinoma.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for T24 cells. Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for histone modifications H3K27Me3 in T24 cells.

first: Gizem

last: Ozgun

first: Tutku

last: Yaras

first: Gokhan

last: Karakulah

last: Ozhan


title: Unstranded RNA sequencing data

last: Pan

overall_design: Transcription profiles of CRISPR-engineered MLL-AF4 BCP-ALL, patient samples, human cell lines.


title: The histone acetylation reader ENL is required for oncogenic transcription driven by super-enhancer and represents a synergistic vulnerability for BET inhibition

city: Shenzhen

address: No 3688, Nanhai Avenue

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

overall_design: Refer to individual Series


title: The single-cell transcriptome program of nodule development cellular lineages in Medicago truncatula

city: Gainesville

first: Matias

last: Kirst

address: 367 Newins-Ziegler Hall, PO Box 110410

institute: University of Florida

summary: Medicago truncatula engages in root nodule symbiosis by developing a de novo plant organ (known as nodule) in its roots in response to the infection by rhizobia. These nodules are de novo plant organs that provide an optimal environment for the rhizobia to fix nitrogen in exchange for photosynthates. The establishment of root nodule symbioses (RNS) requires the coordination of two distinct processes: bacterial infection and nodule organogenesis. In this study we used single-cell RNA-seq to investigate the first hours of the establishment of the root nodule symbiosis aiming to identify the transcriptional mechanisms governing this process.

overall_design: Two genotypes of M. truncatula were used, a wild-type Jemalong A17 a hypernodulating sunn-4 mutant. Root segments were isolated at 0h (control), and at 24h, 48h and 96h after the infection with S. meliloti. Nuclei extraction was performed, followed by Fluorescence-activated cell sorting (FACS). Nuclei were then used to generate single-cell libraries using 10x Chromium technology. Sequencing was performed using Illumina platforms.

first: Carolina

last: Novaes

first: Christopher

last: Dernivis

first: Daniel

last: Conde

first: Henry

middle: W

last: Schimidt

first: Jade

last: Boyd

first: Jean-Michel

last: Ané

first: Julia

last: Frugoli

first: Kelly

last: Balmant

first: Noé

last: Perron

first: Paolo

last: Triozzi

first: Sanhita

last: Chakraborty

first: Sara

last: Knaack

first: Sushmita

last: Roy

first: Wendell

middle: J

last: Pereira


title: Next Generation Sequencing and Quantitative Analysis of mRNAs in Wild-Type Caenorhabditis elegans Animals Consuming Kombucha Tea-assocaited Microbes

city: Chapel Hill

first: Robert

middle: Houston

last: Dowen

state: NC

address: 321 Fordham Hall, CB7100

department: Cell Biology and Physiology

institute: University of North Carolina at Chapel Hill

summary: Kombucha Tea (KT), a fermented tea with roots in traditional Chinese medicine, has surged in worldwide popularity due to its purported health benefits. KT contains a symbiotic culture of yeast and bacterial species, many of which are considered human probiotics. The molecular basis of the health benefits of KT has yet to be thoroughly explored in any animal model. We establishC. elegansas a model to query the molecular interactions between Kombucha-associated microbes (KTM) and the host. We find that worms have an established gut microbiome after consuming a KTM-exclusive diet that mirrors the microbial community found in the fermenting culture. Remarkably, animals consuming KTMs display strikingly reduced lipid levels, yet develop and reproduce similarly toE. coli-fed animals. Critically, consumption of a non-fermenting mix of KT microbial isolates (Kombucha microbe mix) resulted in elevated fat accumulation, suggesting that KTMs do not impair nutrient absorption. To identify the host metabolic pathways altered by KTMs, we performed mRNA-seq on KTM-fed animals, finding widespread changes in lipid metabolism genes. Specifically, we found that three lysosomal lipase genes are significantly upregulated in these animals. These lipases, LIPL-1-3, have been previously shown to promote lipophagy via catabolism of lipid droplets. Consistently, KTM-fed animals display reduced levels of triglycerides and smaller lipid droplet sizes. We propose that KTM-fed animals exhibit a fasting-like metabolic state, even in the presence of sufficient nutrient availability, possibly through induction of lipophagy. Elucidating the host metabolic response to KT consumption will provide unprecedented insight into how this popular fermented beverage may impact human health and inform its use in complementary healthcare plans.

overall_design: The mRNAs of wild-type day 1 adult animals reared on E. coli, Acetobacter tropicalis, or Kombucha-associated microbes were profiled by high-throughput sequencing, in triplicate.

middle: H


title: CFTR-dependent anion secretion in porcine pancreatic ductal epithelium

city: Rotterdam

first: Kalyan

last: Dulla

address: Postbus 2040

country: Netherlands

institute: Erasmus MC

summary:
To investigate pancreatic, ductal CFTR function, organoids from porcine pancreas were cultured.
RNA was extracted from pancreatic organoids at passage 3 and 5 after organoid establishment.

overall_design: RNA expression of pancreatic organoids at passage 3 and 5

first: Marcel

last: Bijvelds