title: Stage-specific dynamic reorganization of genome topology shapes transcriptional neighborhoods in developing human retinal organoids

first: Zachary

middle: A

last: Batz

address: 6 Center Dr Bldg 6, Rm 303

institute: NIH

summary: We have generated high-resolution Hi-C map of developing human retinal organoids to elucidate spatiotemporal dynamics of genomic architecture and its relationship with gene expression patterns. We demonstrate progressive and developmental stage-specific alterations in DNA topology and correlate these changes with transcription of cell type-restricted gene markers during retinal differentiation. Temporal Hi-C reveals a shift towards A compartment for protein-coding genes and B compartment for non-coding RNAs, displaying high and low expression respectively. Notably, retina-enriched genes are clustered near lost boundaries of topologically associated domains (TADs), and higher-order assemblages of TADs (i.e., TAD cliques) tend to localize in the active chromatin regions with binding sites for eye-field transcription factors. These genes show a gain of chromatin contacts at their transcription start site as organoid differentiation proceeds. Our study provides a global view of chromatin architecture dynamics associated with diversification of cell types during retinal development and serves as a foundational resource for in-depth functional investigations of retinal developmental traits.

overall_design: High resolution Hi-C at five time points in retinal organoid development (D60, D70, D90, D120, D200).

first: Zepeng

last: Qu

first: Nivedita

last: Singh

first: Claire

last: Marchal

first: Anand

last: Swaroop


title: Tissue-specific and tissue-agnostic effects of genome sequence variation modulating blood pressure

status: Public on Nov 01 2023

city: New York

first: Or

last: Yaacov

state: NY

department: Center of Human Genetics and Genomics

country: USA

institute: NYU School of Medicine

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Genome-wide association studies in large population cohorts have now mapped thousands of variants and loci for numerous polygenic traits and diseases. However, with some exceptions, mechanistic understanding of which precise variants affect which genes and in which tissues to modulate trait variation is still lacking. Here, we propose genomic analyses of any complex trait using gene expression and chromatin accessibility across multiple tissues, to identify the TFs and regulatory variants within active enhancers regulating specific genes in individual tissues to explain trait heritability. We apply these methods to blood pressure, a classical polygenic trait, to show that variant heritability contributions of kidney, adrenal, heart and arterial tissues are 3.7%, 5.6%, 6.9%, and 9.8%, respectively, from a total of ~500,000 regulatory variants in the four tissues. We demonstrate that these variants are enriched in enhancers binding specific TFs in each tissue. Our findings suggest that gene regulatory networks perturbed by common regulatory variants in a tissue relevant to a phenotype is the primary source of interindividual variation of BP. These studies provide an approach to scan each human tissue for its physiological contribution to a polygenic trait.

overall_design: Assay for transposase-accessible chromatin using sequencing (ATAC-seq) for human snap frozen tissue samples.

first: Dongwon

last: Lee

first: Prabhu

last: Mathiyalagan

first: Aravinda

last: Chakravarti


title: Validation of a QTL for Flavobacterium psychrophilum resistance in rainbow trout Oncorhynchus mykiss

city: Frederiksberg

first: Per

middle: Walter

last: Kania

address: Stigbojlen 7

department: Department of Veterinary and Animal Sciences

country: Denmark

institute: University of Copenhagen

summary:
Rainbow trout (1000 fish) was exposed to the bacterial pathogen F. psychrophilum by simple bath challenge without any pre-treatment with hydrogen peroxide.
Samples (fin clip for Affymetrix QTL analysis) were taken from 167 moribund fish during the course of infection.
When mortality/morbidity ended (day 40) we euthanized a total of 197 specimens of the remaining fish and took samples for DNA (QTL analysis) and assigned the status: Survivor.
For gene expression analysis we took samples from gill, spleen and liver between day 11 and 15 from fish with clinical signs (CS) and no clinical signs (NCS), whereas samples from survivors were taken at day 40.

overall_design:
Rainbow trout (1000 fish) from an outbred batch were used for infection and additionally 1,000 fish were kept as non-infected controls (3.3 ± 0.75 g; 6.9 ± 0.5 cm).
Samples for gene expression (gills, liver and spleen) were taken at three different time-points, at day 0 (before exposure), at day 11-14 (when the mortality rate peaked) and at day 40 (termination of experiment). At day 0, samples were taken from 15 uninfected control fish. At day 11 to 14 we sampled from 15 uninfected control fish, 15 infected fish showing clinical signs (CS) and 15 fish infected fish showing no clinical signs (NCS). The samples at day 40 was from 15 uninfected control and from 15 surviving rainbow trout. All samples were fixed in RNAlater (cat. no. R90901. Sigma-Aldrich, Denmark) and stored at 4°C for 24 h before being placed at -20°C until further analysis.

first: Asma

middle: M

last: Karami

middle: W

first: Kurt

last: Buchmann

first: Heidi

last: Mathiessen


title: The Triptryium wilfordii derivative celastrol has anti-fibrotic effects in systemic sclerosis [Spatial Transcriptomics]

city: Saskatoon

first: Andrew

last: Leask

state: Saskatchewan

address: 105 Wiggins

department: College of Dentistry

country: Canada

institute: University of Saskatchewan

summary: Objectives: Scleroderma (systemic sclerosis, SSc), as a prototypic inflammation-driven fibrotic disease, possesses connective tissue lesions populated with persistently activated myofibroblasts maintained by an mechanotranductive/pro-adhesive signaling loop. Drugs targeting this pathway are therefore of likely therapeutic benefit. The mechanosensitive transcriptional co-activator, yes activated protein-1 (YAP1), is activated in SSc fibroblasts. The terpenoid celastrol has recently been identified as a YAP1 inhibitor: however, if celastrol can alleviate SSc fibrosis is unknown. Methods: Human dermal fibroblasts from healthy individuals and patients with diffuse cutaneous SSc were treated with or without transforming growth factor b1 (TGFb1) in the presence or absence of celastrol. C57BL6J mice were subjected to the inflammatory-driven bleomycin-induced model of skin SSc, in the presence or absence of celastrol. RNA expression was assessed using RNAseq, real-time polymerase chain reaction and spatial transcriptomic analyses. Protein expression was determined using Western blot and enzyme-linked immunosorbent assay. Fibrosis was monitored by hematoxylin and eosin and trichrome staining, and indirect immunofluorescence analysis. Results: In dermal fibroblasts, celastrol impaired the ability of TGFβ1 to induce an SSc-like pattern of gene expression, including the induction of cellular communication network factor 2 (CCN2), collagen I and TGFβ1 protein. Celastrol alleviated the persistent fibrotic phenotype of dermal fibroblasts cultured from lesions of SSc patients. In the bleomycin-induced model of SSc dermatopathology, celastrol inhibited fibrosis and blocked nuclear localization of YAP in myofibroblasts. Conclusion: Our data are consistent with the hypothesis that compounds, such as celastrol, that antagonize the YAP pathway may be potential treatments for SSc skin fibrosis.

overall_design: To investigate the gene expression of the PBS, Bleomycin and Celastrol treated mouse skin tissue samples

first: Pratyusha

last: Chitturi

first: Xu

last: Shi-wen

first: Bahja

last: Ahmed Abdi

first: John

last: Nguyen

first: David

middle: E

last: Carter

first: Richard

middle: J

last: Stratton

first: Sarthak

last: Sinha

first: Rohit

last: Arora


title: Pre-existing interferon gamma conditions the lung to mediate early control of SARS-CoV-2

city: Bethesda

first: Sivaranjani

last: Namasivayam

address: 33 North Drive

department: NIAID

type: Expression profiling by high throughput sequencing

summary: Interferons (IFNs) are critical for anti-viral host defence. Type-1 and type-3 IFNs are typically associated with early control of viral replication and promotion of inflammatory immune responses; however, less is known about the role of IFNγ in anti-viral immunity, particularly in the context of SARS-CoV-2. We have previously observed that lung infection with attenuated bacteria M. bovis BCG achieved though intravenous (iv) administration provides strong protection against SARS-CoV-2 (SCV2) infection and disease in two mouse models. Assessment of the pulmonary cytokine milieu revealed that iv BCG induces a robust IFNγ response and low levels of IFNβ. Here we examined the role of ongoing IFNγ responses due to concurrent bacterial infection on SCV2 disease outcomes in two murine models. We report that IFNγ is required for iv BCG induced reduction in pulmonary viral loads and that this outcome is dependent on IFNγ receptor expression by non-hematopoietic cells. Further analysis revealed that BCG infection promotes the upregulation of ISGs with reported anti-viral activity by pneumocytes and bronchial epithelial cells in an IFNγ dependent manner, suggesting a possible mechanism for the observed protection. Finally, we confirmed the importance of IFNγ in these anti-viral effects by demonstrating that the recombinant cytokine itself provides strong protection against SCV2 challenge when administered intranasally. Together, our data show that a pre-established IFNγ response within the lung is protective against SCV2 infection, suggesting that concurrent or recent infections that drive IFNγ may limit the pathogenesis of this virus and supporting possible prophylactic uses of this cytokine in COVID-19 management.

overall_design: PBS iv, BCG iv inoculated mice as well as each group administred anti-IFNγ were infected with SARS-CoV-2 via the intra-nasal route. Lungs were harvested from the mice 3 days post infection and single cell suspenions were generated for scRNAseq analysis


title: Identification of genes that confer cancer stem cell properties in CTNNB1-mutated NSCLC cells.

city: Maebashi

first: Noriaki

last: Sunaga

state: Gunma

address: 3-39-15 Showa-machi

department: Department of Respiratory Medicine

country: Japan

institute: Gunma University Graduate School of Medicine

summary: mRNA seq analysis comparing the expression profiles of sphere-forming and adherent A427 cells carrying a homozygous CTNNB1 T41A mutation revealed that several stem cell-associated genes and pathways were upregulated in the spheroid cultures.

overall_design: The A427 NSCLC cell line, which grows as both spheroid and adherent cultures, was used in the current study. Gene expression was examined in sphere-forming and adherent A427 cells. Sphere-forming and adherent cells were separately collected and total RNA was extracted from the cells.

first: Sunaga

last: Noriaki

first: Kawabata-Iwakawa

last: Reika


title: Discovery of diagnostic biomarker and molecular mechanisms of placenta accreta spectrum

city: Sapporo

first: Tasuku

last: Mariya

state: Hokkaido

address: South 1 West 16, Chuo-ku

institute: Sapporo Medical University

summary:
Transcriptome analysis of FFPE placenta of placenta accreta spectrum, over increta.
Pre-diagnosis is essential to safely deal with placenta accrete spectrum, but definitive diagnostic markers have not yet been established. We conducted a transcriptome analysis of FFPE placenta to establish early diagnostic markers for placenta accreta.

overall_design: We analyzed 11 FFPE placenta (3 control, 2 increta and 6 percreta) using the GeneChipTM Human Gene 2.0 ST Arrays. Array data was processed by Transcriptome Analysis Console 4.0 software package . No techinical replicates were performed.

first: Shinichi

last: Ishioka


title: Cell-type differential targeting of Setdb1 prevents aberrant CTCF binding, chromatin looping, and cis-regulatory interactions

city: Hong Kong

first: Lut Fei

last: Tam

address: The Hong Kong University of Science and Technology, Clear Water Bay, Kowloon

department: LIFS

country: Hong Kong

institute: HKUST

summary: SETDB1 is a histone methyltransferase with essential roles in mouse development. It mediates transcriptional repression of genes and repetitive elements in different cell-types by depositing histone H3 lysine 9 trimethylation (H3K9me3). However, the function of differential H3K9me3 enrichment between cell- and tissue-types remains unclear. In this study, we demonstrate a global mutual exclusivity of H3K9me3 and CTCF across mouse tissues from different developmental time points. To investigate the mechanistic relationship, we analyzed SETDB1 depleted mouse embryonic stem cells and discovered that H3K9me3 prevents aberrant CTCF binding independently of DNA methylation and H3K9me2. Such sites are enriched with retrotransposons such as SINE B2 elements, which are not previously defined as targets of SETDB1. While the epigenetic modifier is known to silence transcriptional enhancers, its effects on higher-order chromatin structures have not been clearly defined. Overall, large chromatin structures including topologically associated domains and subnuclear compartments, remain intact. However, chromatin loops and local interactions are disrupted. Such perturbations lead to transcriptional changes by altering pre-existing chromatin landscapes, where specific genes have differential interactions with dysregulated cis-regulatory elements. This mode of transcriptional regulation is distinct from the direct repression of genic promoters or enhancers. Collectively, we find that cell-type specific targets of SETDB1 contributes to maintaining cellular identities through shaping genome architecture and transcriptomic networks by modulating CTCF binding, representing a novel function for SETDB1 and H3K9me3.

overall_design: ChIP-seq, ATAC-seq, Mnase-seq, and Hi-C were conducted for WT and Setdb1 KO mouse ESCs.

first: Phoebe Lut Fei

first: Ming Fung

last: Cheung

first: Lu Yan

last: Chan

first: Danny

last: Leung


title: A Diversity of Novel Type-2 Innate Lymphoid Cell Subpopulations Revealed During Tumour Expansion

status: Public on Oct 31 2023

city: Vancouver

first: Wilf

last: Jefferies

institute: UBC

summary: Type 2 innate lymphoid cells (ILC2s) perform vital functions in orchestrating humoral immune responses, facilitating tissue remodelling, and ensuring tissue homeostasis. Additionally, in a role that has garnered considerably less attention, ILC2s can also enhance Th1-related cytolytic T lymphocyte immune responses against tumours. Studies have thus far generally failed to address the mystery of how one ILC2 cell-type can participate in a multiplicity of functions. Here we utilized single cell RNA sequencing analysis to create the first comprehensive atlas of naïve and tumour-associated lung ILC2s and discover multiple unique subtypes of ILC2s equipped with developmental gene programs that become skewed during tumour expansion favouring inflammation, antigen processing, immunological memory and Th1-related anti-tumour CTL responses. The discovery of these new subtypes of ILC2s challenges current paradigms of ILC2 biology and provides an explanation for their diversity of function.

overall_design: Primary ILC2s isolated, purified, enriched and single-cell sequenced at different stages of tumour growth in WT C57BL/6 mice to investigate the immune surveillance dynamics during tumour progression through gene expression changes. Samples include tumour-carrying activated, tumour-carrying non-activated and naive non-activated as control.

first: Clara

last: Xia

first: Iryna

last: Saranchova

first: Pablo

last: de Lucía Finkel

first: Stephanie

last: Besoiu

first: Lonna

last: Munro

first: Cheryl

middle: G

last: Pfeifer

first: Anne

last: Haegert

first: Yen-Yi

last: Lin

first: Stéphane

last: Le Bihan

first: Colin

last: Collins

first: Wilfred


title: The H2Bub1-deposition complex is required for human and mouse cardiogenesis (ChIP-Seq)

city: Boston

first: Syndi

last: Barish

state: MA

address: 77 Avenue Louis Pasteur

institute: Harvard Medical School

summary: De novo variants affecting monoubiquitination of histone H2B (H2Bub1) are enriched in human congenital heart disease. H2Bub1 is required in stem cell differentiation, cilia function, post-natal cardiomyocyte maturation, and transcriptional elongation. However, how H2Bub1 affects cardiogenesis is unknown. We show that the H2Bub1-deposition complex (RNF20-RNF40-UBE2B) is required for mouse cardiogenesis and for differentiation of human iPSCs into cardiomyocytes. Mice with cardiac-specific Rnf20 deletion are embryonic lethal and have abnormal myocardium. We then analyzed H2Bub1 marks during differentiation of human iPSCs into cardiomyocytes. H2Bub1 is erased from most genes at transition from cardiac mesoderm to cardiac progenitor cells, but is preserved on a subset of long cardiac-specific genes. When H2Bub1 is reduced in iPSC-cardiomyocytes, long cardiac-specific genes have fewer full-length transcripts. This correlates with H2Bub1 accumulation near the center of these genes. H2Bub1 accumulation near the center of tissue-specific genes was also observed in embryonic fibroblasts and fetal osteoblasts. In summary, we show that normal H2Bub1 distribution is required for cardiogenesis and cardiomyocyte differentiation, and point to H2Bub1 regulating tissue-specific gene expression by increasing the amount of full-length transcripts.

overall_design: Examination of histone 2B monoubiquitination (H2Bub1) in 5 cell types in triplicate (induced pluripotent stem cells, mesoderm, cardiac mesoderm, cardiac progenitor, and cardiomyocyte). UBE2B-/- mutants (member of the H2Bub1-deposition complex) are examined at the cardiomyocyte stage in triplicate.

first: Martina

last: Brueckner


title: The H2Bub1-deposition complex is required for human and mouse cardiogenesis (RNA-Seq)


title: The H2Bub1-deposition complex is required for human and mouse cardiogenesis

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

overall_design: Refer to individual Series


title: Divergent immune microenvironments in two tumor nodules from a patient with mismatch repair-deficient prostate cancer.

city: Houston

first: Nadim

last: Ajami

state: TX

address: 1515 Holcombe Blvd

institute: MD Anderson Cancer Center

summary: Patients with prostate cancer (PC) generally do not respond favorably to immune checkpoint inhibitors, which may be due to a low abundance of tumor-infiltrating lymphocytes even when mutational load is high. Here, we identified a patient who presented with high-grade primary prostate cancer with two adjacent tumor nodules. While both nodules were mismatch repair- deficient (MMRd), exhibited pathogenic MSH2 and MSH6 alterations, had a high tumor mutational burden (TMB), and demonstrated high microsatellite instability (MSI), they had markedly distinct immune phenotypes. The first displayed a dense infiltrate of lymphocytes (“hot nodule”), while the second displayed significantly fewer infiltrating lymphocytes (“cold nodule”). Whole-exome DNA analysis found that both nodules shared many identical mutations, indicating that they were derived from a single clone. However, the cold nodule appeared to be sub-clonal relative to the hot nodule, suggesting divergent evolution of the cold nodule from the hot nodule. Whole-transcriptome RNA analysis found that the cold nodule demonstrated lower expression of genes related to antigen presentation (HLA) and, paradoxically, classical tumor immune tolerance markers such as PD-L1 (CD274) and CTLA-4. Immune cell deconvolution suggested that the hot nodule was enriched not only in CD8+ and CD4+ T lymphocytes, but also in M1 macrophages, activated NK cells, and gamma delta T cells compared to the cold nodule. This case highlights that MMRd/TMB-high PC can evolve to minimize an anti-tumor immune response, and nominates downregulation of antigen presentation machinery (HLA loss) as a potential mechanism of adaptive immune evasion in PC.

overall_design: Two adjacent prostate tumor nodules from the same patient (TIL-hot and TIL-cold) were sequenced for whole-transcriptome and whole-exome. A matched normal saliva sample was also sequenced for whole-exome.

first: Justin

middle: H

last: Hwang

first: Abderrahman

last: Day

first: Hannah

last: Bergom

first: Emmanuel

middle: S

last: Antonarakis


title: Inflammation and neutrophil extracellular traps in cerebral cavernous malformation

city: Uppsala

first: Ross

last: Smith

address: Skrindvägen 20

department: IGP

country: Sweden

institute: Uppsala University

summary: Cerebral Cavernous Malformation (CCM) is a brain vascular disease with various neurological symptoms. In this study, we describe the inflammatory profile in CCM and show for the first time the formation of neutrophil extracellular traps (NETs) in rodents and humans with CCM. Through RNA-seq analysis of cerebellum endothelial cells from wild-type mice and mice with anendothelial cell-specific ablation of the Ccm3 gene (Ccm3iECKO), we show that endothelial cells from Ccm3iECKO mice have anincreased expression of inflammation-related genes.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for cerebellar endothelial cells derived from two related mouse models of cerebral cavernous malformations, one model acute and the other chronic.  Comparisons are made between wild-type and Ccm3 gene knock-out mice in both models.

first: Anthony

middle: C

last: Yau

first: Maria

last: Globisch

first: Favour

last: Onyeogaziri

first: Lei

middle: L

last: Conze

middle: O

first: Suvi

last: Jauhiainen

first: Monica

last: Corada

first: Fabrizio

last: Orsenigo

first: Hua

last: Huang

first: Melanie

last: Herre

first: Anna-Karin

last: Olsson

first: Matteo

last: Malinverno

first: Veronica

last: Sundell

first: Behnam

middle: R

last: Jahromi

first: Mika

last: Niemelä

first: Aki

last: Laakso

first: Cecilia

last: Garlanda

first: Alberto

last: Mantovani

last: Lampugnani

first: Elisabetta

last: Dejana

first: Peetra

middle: U

last: Magnusson


title: Integrative genotyping of cancer and immune phenotypes by long-read sequencing

first: Livius

last: Penter

address: 450 Brookline Avenue

department: Medical Oncology

institute: Dana-Farber Cancer Institute

summary: Single-cell transcriptomics has become the definitive method for classifying cell types and states, and can be augmented with genotype information to improve cell lineage identification. Due to constraints of short-read sequencing, current methods to detect natural genetic barcodes often require cumbersome primer panels and early commitment to targets. Here we devise a flexible long-read sequencing workflow and analysis pipeline, termed nanoranger, that starts from intermediate single-cell cDNA libraries to detect cell lineage-defining features, including single-nucleotide variants, fusion genes, isoforms, sequences of chimeric antigen and TCRs. Through systematic analysis of these classes of natural ‘barcodes’, we define the optimal targets for nanoranger, namely those loci close to the 5’ end of highly expressed genes with transcript lengths shorter than 4 kB. As proof-of-concept, we apply nanoranger to longitudinal tracking of subclones of acute myeloid leukemia (AML) and describe the heterogeneous isoform landscape of thousands of marrow-infiltrating immune cells. We propose that enhanced cellular genotyping using nanoranger can improve the tracking of single-cell tumor and immune cell co-evolution.

overall_design: Single cell RNA sequencing was performed on single cell suspensions after ficoll density centrifugation using the 5' chemistry (10x Genomics).

first: Mehdi

last: Borji

first: Adi

last: Nagler

first: Catherine

last: Wu


title: PDGFRβ signaling cooperates with β-catenin to modulate c-Abl and biologic behavior of desmoid-type fibromatosis [microarray]

first: Bic

last: MSKCC

address: 1275 York Ave.

institute: Memorial SLoan-Kettering Cancer Center

summary:
Unsupervised clustering of desmoid tumors and normal mesenchymal tissues was performed using henes associated with HIF1 activity.  This accurately distiguished neoplastic tissues from normal controls
The study sought to identify genes differentially expressed in desmoid-type fibromatosis as opposed to normal mesenchymal tissues.  We noted that beta-catenin, the central driver in desmoid-type fibromatosis, appeared to regulate HIF1 signaling in in vitro studies.  Genes associated with HIF1 and angiogenesis pathways were then used to perform unsupervised clustering on desmoid tumors and normal mesenchymal tissues.  The genes accurately differentiated neoplastic and normal samples.

overall_design: Gene set enrichment analysis of desmoid cells treated with shRNA to knockdown CTNNB1 suggested that β-catenin altered HIF1 target expression and angiogenesis-related genes.  These genes were used to perform unsupervised clustering on desmoid tumors and normal tissue analyzed by Affymetrix U133A 2.0 gene expression arrays.

first: Aimee

last: Crago

first: Nicholas

middle: D

last: Socci


title: Single cell ATAC-sequencing analysis of human SUV39H1-knockout CAR T compared to control CAR T cells (mock) in a xenograft model of lung adenocarcinoma

city: Paris

first: Sebastian

last: Amigorena

address: 26 rue d'Ulm

country: France

institute: Institut Curie

summary: We have observed that the inactivation of SUV39H1 enhances 41BBz-CAR T cell long-term persistence, providing protection against tumor relapses and rechallenges in a xenograft mouse model of lung adenocarcinoma. The purpuse of this study was to profile chromatin accessibility of SUV39H1-deficient compared to mock 41BBz-CAR T cells by single-cell assay for transposase accessible chromatin (scATAC-seq) on FACS-sorted T cells eight days after CAR T cell infusion into the mice.

overall_design: NSG mice that received intravenous injections of tumor cells (A549) ectopically expressing CD19 that develop tumors in the lungs, were treated 21 days later with 0,9 million of 41BBz-CAR T cells (i.v.) that were SUV39H1-knockout (CRISPR/Cas9 technology) or controls CAR T cells (same CRISPR/Cas9 protocol with a sgRNA that does not have any complimentary sequence in the genome). CAR T cells were isolated from lungs at day 8 after infusion, T-cells were FACS sorted and subjected to Chromium Single Cell ATAC protocol (for each condition there were three biological replicates from pooled mice).

first: Sheila

last: López-Cobo

first: Jaime

last: Fuentealba

first: Paul

last: Gueguen

first: Pierre-Emmanuel

last: Bonté

first: Michael

last: Saitakis


title: single cell RNA-sequencing analysis of human SUV39H1-knockout CAR T compared to control CAR T cells (mock) in a xenograft model of lung adenocarcinoma

summary: We have observed that the inactivation of SUV39H1 enhances 41BBz-CAR T cell long-term persistence, providing protection against tumor relapses and rechallenges in a xenograft mouse model of lung adenocarcinoma. The purpuse of this study is to investigate the transcriptomic differences of lung-infiltrating SUV39H1 knock-out versus mock 41BBz-CAR T cells by single cell RNA sequencing at days 8 and 28 after CAR T cell infusion.

overall_design: NSG mice that received intravenous injections of tumor cells (A549) ectopically expressing CD19 that develop tumors in the lungs were treated 21 days later with 0,9 million of 41BBz-CAR T cells (i.v.) that were SUV39H1-knockout (CRISPR/Cas9 technology) or controls CAR T cells (same CRISPR/Cas9 protocol with a sgRNA that does not have any complimentary sequence in the genome). CAR T cells were isolated from lungs at day 8 and 28 after infusion, T-cells were FACS sorted and subjected to Chromium 10× Single-Cell 3' mRNA sequencing (three biological replicates for each condition and timepoint).


title: Nanostring analysis of human SUV39H1-knockout CAR T compared to control CAR T cells (mock) at different timepoint

summary: Failure of adoptive T cell therapies in cancer patients is linked to limited T cell expansion and persistence, even in memory-prone 41BB-(BBz)-based chimeric antigen receptor (CAR) T cells. In murine CD8+ T cells, SUV39H1 promotes differentiation and expansion of effector CD8+ T cells during acute infection by Listeria monocytogenes by silencing stemness and memory genes (Pace et al. Science, 2018). The purpuse of this study is to investigate the transcriptomic differences of SUV39H1 knock-out versus mock human 41BBz-CAR T cells by Nanostring at different cycles of restimulation.

overall_design: In the study presented here, four donors were used at three different timepoints: after CAR production (D0), or 7 days after one or three weekly restimulations (D7, D21)

first: Irena

last: Chacón


title: BRD9 determines the cell fate of hematopoietic stem cells by regulating chromatin state

city: Kobe

first: Weijia

last: Zang

address: Hyogo, Kobe, Chuo Ward, Minatojima Minamimachi, 6 Chome−３−7

department: Department of Hematology-Oncology

institute: Foundation for Biomedical Research and Innovation at Kobe

summary: Switch/sucrose nonfermentable (SWI/SNF) complexes are ATP-dependent chromatin remodeler complexes that play critical roles in timely and appropriate gene regulation by modulating chromatin architecture and DNA accessibility. SWI/SNF complexes can be grouped into three subcomplexes of differing sizes, canonical BAF (cBAF), polybromo BAF (PBAF), and newly identified noncanonical BAF (ncBAF). The most recently characterized ncBAF lacks the core BAF subunits ARID, BAF47, and BAF57, but includes unique subunits GLTSCR1/1L and BRD9, one of the bromodomain-containing proteins.We recently demonstrated the novel mechanism for the ncBAF disruption caused by mutations in a spiceosomal protein, SF3B1, especially in 65–83% for myelodysplastic syndromes (MDS) with ring sideroblasts as well as in >20% of mucosal/uveal melanomas, suggesting that the disturbed ncBAF may have some roles in the malignant hematopoiesis.Mechanistically, SF3B1 mutant recognizes an aberrant, deep intronic branchpoint within BRD9 and thereby induces the inclusion of an aberrant exon and subsequently profound degradation of BRD9 mRNA by triggering nonsense-mediated RNA decay (NMD). However, compared to the roles of cBAF, the functions of BRD9/ncBAF in normal and malignant hematopoiesis in vivo have been totally uncharacterized.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: The MOLM13 cell lines were cultured in RPMI with 10% FCS and 1% P/S to the logarithmic growth phase,  treated MOLM13 cells with DMSO and 50nM dBRD9 for 4 days and abstract RNA for RNA-seq.


title: Investigation of Rap1 binding occupancy in global H3-depletion condition [ChIP-Seq]

city: Pathumthani

first: Peerapat

last: Khamwachirapithak

address: Phahonyothin Road, Khlong Luang

country: Thailand

institute: Thammasat University

summary: We constructed the inducible H3-depleted and FLAG-tag of the repressor activator protein 1 (Rap1) yeasts to investigate the depleted effect of nucleosomal H3 on DNA-binding profiles of Rap1 and resulting transcriptional changes. The H3 inducible shut-off strain (named “DGS200.1” or “H3i” strain) was prepared from the wildtype YEF473A strain by knocking out the HHT1 gene (encoding H3), and replacing the native promoter of the HHT2 gene (the other H3-encoding gene) with an inducible Gal1 promoter. Conditional depletion of H3 was induced by culturing the H3i strain in the YPGal liquid medium (1% yeast extract, 2% peptone, 2% galactose) and transferring to the YPD medium (1% yeast extract, 2% peptone, 2% dextrose) for three hours. WT yeast were also cultured and switched the growth media in the same way as controls. DNA-binding profiles of H3 and Rap1 were determined using chromatin immunoprecipitation followed by next-generation sequencing (ChIP-seq), and the changes in transcriptional levels were investigated using RNA-seq.

overall_design: Genome-wide nucleosomal H3 binding was investigated using ChIP-seq to study effects of H3 depletion in WT and H3i (conditional H3 shut-off) yeasts

first: David

last: Guillaume-Schoepfer

first: Varodom

last: Charoensawan

first: Phillip

last: Wigge

first: Sarah

last: Teichmann


title: Disruption of SUV39H1-mediated H3K9 methylation sustains CAR T cell function [scRNA/CITE-seq]

status: Public on Nov 03 2023

first: Michel

last: Sadelain

address: 1275 York Avenue, Box 182

institute: Memorial Sloan Kettering Cancer Cener

summary: Single cell gene expression profile of WT and SUV39H1-edited CAR T cells at pre-infusion (Day 0), day 9 and day 16 post infusion in tumor (NALM6) bearing NSG mice

overall_design: Unedited and SUV39H1-edited human CAR T cells were sorted from bone marrow (at day 9 and 16) from tumor bearing  NSG mice. Pre-infusion single cell gene expression profiling of unedited and SUV39H1-edited human CAR T cells was also performed.

first: Nayan

last: Jain

first: Zeguo

last: Zhao

first: Chenling

last: Antelope

first: Yosi

last: Gozlan

first: Antonino

last: Montalbano

last: Brocks

first: Anton

last: Dobrin

last: Lopez


title: Next Generation Sequencing Facilitates Quantitative Analysis of HN12 Cell Transcriptome with ATAD3A loss

city: Atlanta

first: Liwei

last: Lang

state: GA

address: 1701 Uppergate Drive

institute: Emory University

summary: The goal of this study is to investigate the influence of ATAD3A expression on the transcriptome profiles of head and neck cancer

overall_design: Examination of mRNA profiles in head and neck cancer cells with or without ATAD3A knockout

first: Teng

last: Yong


title: BRD9 determines the cell fate of hematopoietic stem cells by regulating chromatin state [mouse_Brd9_H3K27Ac-ChIP-seq]

city: Suita

first: Masaki

last: Nomura

state: Osaka

address: 2-2, Yamadaoka

department: Department of Cancer Pathology

institute: Graduate School of Medicine, The University of Osaka

overall_design: 2.0 x 10E6 total BM cells of Mx1-Cre control, Mx1-Cre Brd9fl/fl mice transplanted via tail vein injection into lethally irradiated (two times 450 cGy) CD45.1+ recipient mice. Four weeks after BMT, the recipient mice received 20 mg/kg pIpC injection every other day for a total of 3 doses. Five months after pIpC injection, the recipient mice were sacrificed for sample preparation.

first: Daichi

last: Inoue

first: Koutarou

last: Nishimura

first: Muran

last: Xiao


title: BRD9  determines the cell fate of hematopoietic stem cells by regulating chromatin state [mouse_Brd9_ATAC-seq]

summary: Chromatin remodeling caused by SWI/SNF (BAF) complexes is essential for regulating cell fates. Of these, BRD9, an essential component of the non-canonical BAF (ncBAF) complex, is post-transcriptionally decayed in multiple types of cancer, especially in myelodysplastic syndrome (MDS). However, how BRD9 loss or ncBAF disruption alters the HSCs' integrity remains unknown. Here, we identify that Brd9 loss enhances chromatin accessibility, promoting myeloid-lineage skewing at the expense of B cell development in vivo, followed by MDS development. Brd9 significantly colocalizes with Ctcf, whose peaks are stoichiometrically enhanced by Brd9 loss, leading to increased chromatin loops with the larger chromatin domains intact. These Ctcf-associated chromatin loops newly occur in myeloid-related genes. These data uncover the unrecognized roles of BRD9/ncBAF in the cell fate specification of HSCs via fine-tuning chromatin loops and shed light on the mechanism of ncBAF-disrupted cancer.

overall_design: Four weeks after birth, the primary mice received 20 mg/kg pIpC injection every other day for a total of 3 doses. Five months after pIpC injection, the recipient mice were sacrificed for sample preparation.


title: BRD9  determines the cell fate of hematopoietic stem cells by regulating chromatin state [human_Brd9_ATAC-seq]

overall_design: The K562 cell lines were cultured in RPMI with 10% FCS and 1% P/S to the logarithmic growth phase,  treated K562 cells with DMSO and 100nM dBRD9 for 4 days, then compare CTCF binding in K562 cells after different treatment.


title: BRD9 determines the cell fate of hematopoietic stem cells by regulating chromatin state [human_Brd9_bulkRNA-seq]

overall_design: The K562 cell lines were cultured in RPMI with 10% FCS and 1% P/S to the logarithmic growth phase,  treated K562 cells with DMSO and 100nM dBRD9 for 4 days and RNA abstraction for RNA-seq.


title: An interactive resource of molecular signalling in the developing human haematopoietic stem cell (HSC) niche

city: Seattle

first: Edie

middle: I

last: Crosse

state: Washington

address: 1100 Fairview Ave N

institute: Fred Hutchinson Cancer Center


title: Investigation of Rap1 binding occupancy in global H3-depletion condition [RNA-seq]

first: Peerapat

last: Khamwachirapithak

overall_design: Transcriptomes of WT and H3i (conditional H3 shut-off) yeasts cultured in YPGal and YPD were performed in three biological replicates

first: David

last: Guillaume-Schoepfer

first: Varodom

last: Charoensawan

last: Wigge

last: Teichmann


title: An interactive resource of molecular signalling in the developing human haematopoietic stem cell (HSC) niche [scRNA-seq]

summary: The emergence of definitive human haematopoietic stem cells (HSCs) during Carnegie Stages (CS) 14-17 in the aorta-gonad-mesonephros (AGM) region is a complex and tightly regulated process. In a previous study we conducted spatial transcriptomic analysis of the human AGM region at the end of this period (CS16/17) and identified secreted factors involved in HSC development. Here, we extend our analysis to investigate the progression of dorso-ventral polarized signalling around the dorsal aorta over the entire period of HSC emergence. Our results reveal a dramatic increase in signalling complexity from the CS13 to CS14 transition, coinciding with the activation of endothelial-to-haematopoietic transition (EHT) in the ventral domain of the dorsal aorta. We further observe stage-specific changes in signalling complexity up to CS17. Signalling progression described here may underpin step-wise maturation of HSCs described in the mouse model. A data-rich bioinformatics resource presented here along with an interactive online interface enables in silico analysis of molecular interactions between spatially defined domains of the AGM region. This resource will be of particular interest for researchers studying mechanisms underlying human HSC development as well as those developing in vitro methods for the generation of clinically relevant HSCs from pluripotent stem cells.

overall_design:
10X single cell analysis of a carnegie stage (CS13) dorsal aorta from a human embryo.

Third-party reanalysis: CS13 dorsal aorta single cell data was processed using the Cell Ranger 2.1.0 (10x Genomics) analysis pipeline by aligning reads to GRCh38 human transcriptome (Ensembl). The ScanPy pipeline (Wolf, et al. 2018) was used to explore and integrate the dataset with other dorsal aorta single cell datasets spanning stages CS13 - CS15 from a publicly available resource (Zeng, et al. 2019, accession GSE135202 (including samples GSM3993422, GSM3993424, GSM3993425 and GSM3993426)). Cells with less than 200 genes and genes that were in less than 3 cells were filtered out. Cells with a percentage of mitochondrial genes >1 were also filtered out. Contaminating HB genes were also removed from the CS13 dorsal aorta dataset. The reads per cell were normalized and logarithmised. The variance effects of total counts per cell, cell cycle and mitochondrial genes were regressed out. Each dataset was then subset to include only genes with highly variable expression. The tool Scanorama (Hie, et al. 2019) was used to integrate datasets. Then nearest neighbours was computed using 10 principal components and the neighbourhood graph was embedded in two dimensions in a Uniform Manifold Approximation and Projection (UMAP) (Becht, et al. 2018). Clustering of sub-populations within the UMAP were made using the Leiden algorithm (Traag, et al. 2019). Following cluster identification by gene signature and any known identities from prior sorting strategies, the data was then subset into two parts to include hematopoietic and endothelial populations in one subset and stromal and epithelial populations in the second. Partition-based graph abstraction (PAGA) (Wolf, et al. 2019) was used to make lineage inferences from the neighbourhood graphs.

first: Alexander

last: Medvinsky

first: Anahi

last: Binagui-Casas

first: Sabrina

last: Gordon-Keylock

first: Sara

last: Tamagno

first: Didrik

last: Oloffson

first: Richard

last: Anderson


title: An interactive resource of molecular signalling in the developing human haematopoietic stem cell (HSC) niche [LCM-Seq]

overall_design: Laser capture microdissection of spatial domains around the carnegie stage (CS) 13 and 14 dorsal aorta in the developing human embryo with RNA-Seq libraries prepared from each region. N = 2 for each embryo and 6 spatial domains were taken for each stage.


title: Investigation of Rap1 binding occupancy in global H3-depletion condition [Rap1_ChIPseq]

overall_design: Genome-wide Rap1 binding was investigated using ChIP-seq to study effects of H3 depletion in WT and H3i (conditional H3 shut-off) yeasts

first: Phillip

first: Sarah


title: Investigation of Rap1 binding occupancy in global H3-depletion condition [H3MNase_ChIPseq]


title: Bcl6 is a subset defining transcription factor of Lymphoid Tissue inducer-like ILC3

city: Manchester

first: Matthew

last: Hepworth

address: AV Hill Building

country: United Kingdom

institute: University of Manchester

summary: Innate lymphoid cells (ILC) are tissue-resident effector cells with important roles in tissue homeostasis, protective immunity and inflammatory disease. Current nomenclature divides ILC into subsets based on the expression of master transcription factors and effector cytokine programs. In mucosal barrier tissues, group 3 ILC (ILC3) have been defined by the expression of the master transcription factor RORgt. However, ILC3 can be further subdivided into two major subsets – natural cytotoxicity receptor-expressing (NCR+) ILC3 and lymphoid tissue inducer (LTi)-like ILC3 which share type 3 effector modules but also exhibit significant ontological, transcriptional, phenotypic and functional heterogeneity. In particular, LTi-like ILC3 exhibit effector functions not typically associated with other RORgt-expressing lymphocytes, provoking the hypothesis that other master transcription factors may contribute to LTi-like ILC3 biology. Here we identify Bcl6 as an LTi-like ILC3 associated transcription factor in both mice and humans. Deletion of Bcl6 led to dysregulation of the LTi-like ILC3 transcriptional program and changes to subset-specific phenotypic markers and effector functions. Strikingly, loss of Bcl6 enhanced expression of the type 3 effector cytokines IL-17A and IL-17F in LTi-like ILC3, which was found to be in part dependent upon the commensal microbiota. Together these findings implicate Bcl6 as an ILC3 subset-defining transcription factor and part of a network that confers phenotype and function on LTi-like ILC3. Our study further provides a missing link to redefine analogous immune modules in innate and adaptive lymphocyte responses.

overall_design: Comparative gene expression analysis from RNA-seq data obtained from sorted LTi-like ILC3 and NCR+ ILC3 from the mLN and small intestine of Rorc-Cre Bcl6-fl/fl mice and Bcl6-fl/fl controls.

first: Roser

last: Tachó-Piñot


title: T. denticola response to CBD

city: Louisville

middle: Albert

last: Scott

state: KY

address: 501 S Preston St

department: OIID

institute: University of Louisville

summary: Transcriptome of T. denticola ATCC 35405 exposed to cannabadiol (5 ug/ml) vs solvent control (methanol)

overall_design: 3 control cultures vs 3 test  cultures, log phase RNA extraction

first: Jinlian

last: Tan


title: RIPK1 plays a crucial role in maintaining regulatory T cell homeostasis by inhibiting both RIPK3 and FADD-mediated cell death

city: Shanghai

first: Xiaoxue

last: Deng

address: 320 Yueyang Road, Shanghai, China

country: China

institute: University of Chinese Academy of Sciences

summary: Regulatory T (Treg) cells are a critical for maintaining immune balance in various physiological and pathological conditions. However, the mechanisms underlying Treg homeostasis remain incompletely understood. In this study, we demonstrated that RIPK1 is crucial for Treg cell survival and homeostasis. We generated mice with Treg cell-specific ablation of Ripk1 and found that these mice developed fatal systemic autoimmunity due to a dramatic reduction in the number of Treg cell caused by excessive cell death. Unlike conventional T cells, Treg cells with Ripk1 deficiency were only partially rescued from cell death by blocking the FADD pathway. However, simultaneous removal of both Fadd and Ripk3 completely restored the number of Ripk1-deficient Treg cells by blocking cell death. Our results demonstrated that RIPK1 plays a critical role in regulating Treg cell survival by controlling both necroptosis and apoptosis. These findings provide new insights into the mechanisms of Treg cell homeostasis and suggest potential therapeutic targets for autoimmune diseases.

overall_design: Spleen and peripheral lymph nodes were harvested in Foxp3-YFP-Cre and Ripk1fl/fl Foxp3-YFP-Cre mice. For RNAseq, We have four samples, WT rTreg, WT aTreg, cKO rTreg, cKO aTreg.

first: Haikun

last: Wang

first: Haibing

last: Zhang

first: Yuejuan

last: Zheng

first: Lingxia

first: Yunze

last: Zhai


title: RNA sequencing analysis of stable medulloblastoma DAOY, D425, and CHLA01 cells expressing vector only (V/O) or miR-211

city: St. Petersburg

first: Rudrmani

last: Pokhrel

state: United States

address: 600 Fifth Street S.

department: Oncology

institute: Johns Hopkins All Children’s Hospital

summary: To investigate the role of miR-211 in medulloblastoma, next-generation sequencing analysis of RNA extracted from DAOY, D425, and CHLA01 cells expressing vector only (V/O) or miR-211 were performed.

overall_design: Medulloblastoma mRNA profiles were generated by deep sequencing, using Illumina NextSeq 550.

first: Menglang

last: Yuan

first: Keisuke

last: Katsushma

first: Rudramani

first: Bongyong

last: Lee

first: Ranjan

last: Perera


title: Investigation of Rap1 binding occupancy in global H3-depletion condition

first: Peerapat

last: Khamwachirapithak

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

overall_design: Refer to individual Series


title: Study of differential chromatin accessibility within primary breast epithelial cells from healthy donor and its TONSL overexpressing counterpart

city: Indianapolis

first: Harikrishna

last: Nakshatri

state: IN

address: 980 W. Walnut St. R3C218C

department: General Surgery

institute: Indiana University School of Medicine

summary: To evaluate the probable role of TONSL during tumorigenesis we overexpressed TONSL  in primary breast epithelial cells and studied differential chromatin accessibility.

overall_design: KTB103 primary and TONSL overexpressing KTB103 cells were subjected to ATAC-seq using the previously established protocol (Oncogene 40:1332-1346) . Assays were done in biologic triplicates with ~50,000 cells.

first: H


title: Tumor derived MDA-MB-436 pLKO versus  MDA-MB-436 shTONSL cells-enriched gene expression

summary: To study TONSL regulated genes in neoplastically transformed cells, we established tumor derived MDA-MB-436 cell lines in which TONSL is been knocked down by shRNA.

overall_design: To study transcriptomic changes upon TONSL knockdown in TONSL-amplified cell lines, RNA from TMD436 shControl (PLKO), TMD436 shTONSL Clone 1, TMD436 shTONSL Clone 2 and TMD436 shTONSL Clone 3 in triplicates was subjected to RNA-seq. Three different shRNA targeting vectors were used from Sigma-Aldrich: TMD436 shTONSL Clone 1 (TRCN0000016858), TMD436 shTONSL Clone 2 (TRCN0000016861) and TMD436 shTONSL Clone 3 (TRCN0000016862).


title: Hormone-induced enhancer assembly requires an optimal level of hormone receptor multivalent interactions

city: San Antonio

first: Zhijie

middle: Jason

last: Liu

state: TEXAS

address: 7703 Floyd Curl Drive

department: Department of Molecular Medicine

institute: Universality of Texas Health Science Center at San Antonio


title: FMRP phosphorylation modulates neuronal translation through YTHDF1

city: Chicago

first: Chuan

last: He

state: Illinois

address: 929 E 57th St, Room E319B

institute: University of Chicago

summary: Translation activation of local synaptic mRNA is critical to learning and memory. Despite extensive studies on how phosphorylations of ribosome units and translation factors enable fast response to exogenous stimuli, our knowledge on molecular pathways utilized by RNA binding proteins (RBPs) to control translation of specific mRNAs remains incomplete. We have previously found that YTHDF1 regulates depolarization-induced protein synthesis by promoting translation of N6-methyladenosine (m6A)-modified transcripts. Here we report an unexpected mechanism that the stimuli-induced neuronal translation is mediated by phosphorylation of a YTHDF1-binding protein FMRP. Phosphorylation of FMRP serine 499 induced by neuronal depolarization alters the condensing behavior of prion-like protein YTHDF1. Unphosphorylated FMRP sequesters YTHDF1 away from the translation initiation complex, whereas the stimulation-induced FMRP phosphorylation releases YTHDF1 to form translational active condensates with the ribosome to activate translation of YTHDF1 target transcripts. In fragile X syndrome (FXS) models characterized with low FMRP expression, we observed YTHDF1-mediated hyperactive translation, which notably impacts FXS pathophysiology. Developmental defects in an FXS forebrain organoid model could be reversed by a selective small-molecule inhibitor of YTHDF1 which acts by suppressing its condensation in neurons. We characterized transcriptome-wide translation alterations with the inhibitor treatment in organoids and identified targets that explain alleviated FXS pathology. Our study thus reveals FMRP and its phosphorylation as an important regulator of the activity-dependent translation during neuronal development and stimulation, and identifies YTHDF1 as a potential therapeutic target for FXS in which developmental defects caused by FMRP depletion could be reversed through YTHDF1 inhibition.

overall_design:
[Dataset 1] 16 samples. Duplicates for mRNA-Seq and riboLace ribosome-protected fragments samples in isolated primary neurons from P1-3 wildtype, Fmr1 knockout mouse treated by YTHDF1 inhibitor SAC or DMSO control
[Dataset 2] 16 samples. Duplicates for mRNA-Seq and riboLace ribosome-protected fragments samples in cultured control or FXS human forebrain organoids treated by SAC or DMSO

first: Zhongyu

last: Zou


summary: Transcription factors (TFs) activate enhancers to drive cell-specific gene expression in response to signals, but our understanding of enhancer assembly in signaling events is incomplete. Here we show that Androgen Receptor (AR), a steroid hormone signaling-regulated transcription factor, forms phase-separated condensates in response to androgen to activate enhancers. We demonstrate that the intrinsically disordered NTD of AR drives condensate formation and that NTD deletion or aromatic residue mutation reduces AR self-association and abolishes AR transcriptional activity. AR NTD can be substituted by some IDRs from other proteins for AR condensation capacity and transactivation function. Extending the polyQ tract within AR NTD strengthens AR phase separation and also leads to impaired transcriptional activity. PolyQ expansion does not affect AR binding on enhancers, but instead impairs enhancer assembly. These results suggest that AR phase separation mediates enhancer assembly, and an optimal level of AR condensation is required for its proper function in mediating AR-AR homotypic and AR-cofactor heterotypic interactions to regulate transcription in response to signals. Our study supports that alteration of the fine-tuned protein condensation might underlie AR-related human pathologies, therefore providing novel molecular insights for potential therapeutic strategies to treat prostate cancer and other AR-involved diseases by targeting AR condensation.

overall_design: Assay for Transposase-Accessible Chromatin followed by high throughput sequencing (ATAC-seq) for checking chromatin accessibilities for LNCaP cells with different treatments or with the rescue tests for different AR cDNAs.

first: Zhang

last: Zhao

first: Liu

last: Zhijie


title: Identification of LGR6-associated genes in NSCLC

city: Maebashi

first: Noriaki

last: Sunaga

address: 3-39-15 Showa-machi

institute: Gunma University Graduate School of Medicine

summary: mRNA seq analysis comparing the expression profiles of siRNA-mediated LGR6-silenced and LGR6-retained NSCLC cell lines harboring CTNNB1 exon 3 mutations identified several LGR6-related genes that have been associated with cancer development and stemness properties.

overall_design: HCC15 and A427 NSCLC cell lines were used in the current study. Gene expression was examined in siRNA-mediated LGR6-silenced NSCLC cells and LGR6-retained (control siRNA-treated) NSCLC cells. Cells were transfected with siRNAs targeting LGR6 and were collected at 48 h post-transfection, followed by total RNA extraction from the cells.

first: Sunaga

last: Noriaki

first: Kawabata-Iwakawa

last: Reika


title: BRD9 determines the cell fate of hematopoietic stem cells by regulating chromatin state [scRNA-Seq]

first: Masaki

last: Nomura

overall_design: Four weeks after birth, the primary mice received 20 mg/kg pIpC injection every other day for a total of 3 doses. Eight weeks after pIpC injection, the primary mice were sacrificed for sample preparation.

first: Muran

last: Xiao

first: Daichi

last: Inoue

first: Omar

last: Abdel-Wahab


title: BRD9 determines the cell fate of hematopoietic stem cells by regulating chromatin state [ChIP-Seq II]

overall_design: 2.0 x 10E6 total BM cells of Mx1-Cre control, Mx1-Cre Brd9fl/fl mice transplanted via tail vein injection into lethally irradiated (two times 450 cGy) CD45.1+ recipient mice. Four weeks after BMT, the recipient mice received 20 mg/kg pIpC injection every other day for a total of 3 doses. Five months after pIpC injection, the recipient mice were sacrificed for sample preparation.

first: Shinji

last: Kondo


title: BRD9 determines the cell fate of hematopoietic stem cells by regulating chromatin state


title: BRD9 determines the cell fate of hematopoietic stem cells by regulating chromatin state [bulk RNA-Seq I]


title: BRD9 determines the cell fate of hematopoietic stem cells by regulating chromatin state [bulk RNA-Seq II]

overall_design: 1.0 x 10E3 total BM leukemic cells of Mx1-Cre Brd9fl/fl mice which transduced with MLL-AF9 cDNA transplanted via tail vein injection into lethally irradiated (two times 450 cGy) CD45.1+ recipient mice. The next day after BMT, the recipient mice received 20 mg/kg pIpC or PBS injection every other day for a total of 3 doses. 10 days after pIpC/PBS injection, the recipient mice were sacrificed for sample preparation.

first: Weijia

last: Zang


title: TONSL is an immortalizing oncogene of the chromosome 8q24.3 amplicon and new therapeutic target in breast cancer


title: BRD9 determines the cell fate of hematopoietic stem cells by regulating chromatin state [ChIP-Seq I]


title: BRD9 determines the cell fate of hematopoietic stem cells by regulating chromatin state [ATAC-Seq II]


title: Outward-oriented sites within clustered CTCF boundaries are key for intraTAD chromatin interactions and gene regulation

first: Xiao

last: Ge

address: No. 800 Dongchuan Road

institute: Shanghai JiaoTong University


title: BRD9 determines the cell fate of hematopoietic stem cells by regulating chromatin state [ATAC-Seq I]

overall_design: Four weeks after birth, the primary mice received 20 mg/kg pIpC injection every other day for a total of 3 doses. Five months after pIpC injection, the recipient mice were sacrificed for sample preparation.


title: Metabolically Healthy Obesity is not a Myth: A Case Report

city: Izumo

first: Jun

last: Yoshino

state: Shimane

address: 89-1 Enya-cho

institute: Faculty of Medicine, Shimane University

summary: People with obesity who do not have the metabolic syndrome or components of the metabolic syndrome have been characterized as having metabolically healthy obesity (MHO). However, the existence of MHO has been questioned because people with MHO are at greater risk of developing diabetes and fatal cardiovascular disease than people who are lean and healthy. A 25 year-old woman with rigorously defined MHO (based on normal oral glucose tolerance, insulin sensitivity (assessed by using the hyperinsulinemic-euglycemic clamp procedure), plasma triglyceride and HDL-cholesterol, intrahepatic triglyceride content and carotid intima-media thickness [CIMT]) was evaluated at baseline (BMI=37.7 kg/m2) and 5 years later, after gaining 30.8 kg (32%) in weight (BMI=49.6 kg/m2). The increase in weight was comprised of an 8.8 kg (20%) increase in FFM, 22.0 kg (42%) increase in total body fat, 8.1 kg (37%) increase in leg fat mass, 57% increase in subcutaneous abdominal fat and a 78% in intra-abdominal fat. Weight gain did not have adverse effects on fasting plasma glucose, oral glucose tolerance, beta-cell function, insulin sensitivity, plasma triglyceride, intrahepatic triglyceride content and CIMT. Adipose tissue expression of genes involved in extracellular matrix formation did not change. Adipose tissue expression of several inflammation-related genes increased by more than 30%, but was not associated with a corresponding increase in plasma cytokine concentrations, with the exception of an increase in plasma IL-6. The present case study demonstrates that some people with obesity are resistant to the adverse cardiometabolic effects of excess adiposity and marked weight gain.

overall_design: In this study, we evaluated the effect of weight gain (32%) on subcutaneous dipose tissue gene expression profile by analyzing subcutaneous adipose tissue samples obtained before and after weight gain.

first: Sarah

last: Farabi

first: Gordon

last: Smith

first: Samuel

last: Klein


title: BRD9 determines the cell fate of hematopoietic stem cells by regulating chromatin state [human_Brd9_CTCF-ChIP-seq]

summary: Switch/sucrose nonfermentable (SWI/SNF) complexes are ATP-dependent chromatin remodeler complexes that play critical roles in timely and appropriate gene regulation by modulating chromatin architecture and DNA accessibility. SWI/SNF complexes can be grouped into three subcomplexes of differing sizes, canonical BAF (cBAF), polybromo BAF (PBAF), and newly identified noncanonical BAF (ncBAF). The most recently characterized ncBAF lacks the core BAF subunits ARID, BAF47, and BAF57, but includes unique subunits GLTSCR1/1L and BRD9, one of the bromodomain-containing proteins.We recently demonstrated the novel mechanism for the ncBAF disruption caused by mutations in a spiceosomal protein, SF3B1, especially in 65–83% for myelodysplastic syndromes (MDS) with ring sideroblasts as well as in >20% of mucosal/uveal melanomas, suggesting that the disturbed ncBAF may have some roles in the malignant hematopoiesis.Mechanistically, SF3B1 mutant recognizes an aberrant, deep intronic branchpoint within BRD9 and thereby induces the inclusion of an aberrant exon and subsequently profound degradation of BRD9 mRNA by triggering nonsense-mediated RNA decay (NMD). However, compared to the roles of cBAF, the functions of BRD9/ncBAF in normal and malignant hematopoiesis in vivo have been totally uncharacterized.

overall_design: The K562 cell lines were cultured in RPMI with 10% FCS and 1% P/S to the logarithmic growth phase,  treated K562 cells with DMSO and 100nM dBRD9 for 4 days, then compare CTCF binding in K562 cells after different treatment.

first: Koutarou

last: Nishimura


title: Gene expression profiles of wild type and Zbtb20-KO cochleae at P10

first: zhifang

last: xie

address: 1665 Kongjiang Road

department: Ministry of Education-Shanghai Key Laboratory of Children's Environmental Health

institute: Xinhua Hospital, Shanghai Jiaotong University School of Medicine

summary: The goal of this study is to identify differentially expressed genes in Zbtb20 deleted cochleae.

overall_design: Cochlear mRNA profiles of P10 wild type (WT) and ZB20KO mice

first: Zhifang

last: Xie

first: Weiping

last: Zhang


title: Outward-oriented sites within clustered CTCF boundaries are key for intraTAD chromatin interactions and gene regulation [RNA-seq]

summary: CTCF plays an important role in 3D genome organization by adjusting insulation at TAD boundaries, where clustered CBS (CTCF-binding site) elements are often arranged in tandem array with a complex divergent or convergent orientation. Here using cPcdh and HOXD loci as a paradigm, we look into the clustered CTCF TAD boundaries and find that, counterintuitively, outward-oriented CBS elements are crucial for inward enhancer-promoter interactions as well as for gene regulation. Specifically, by combinatorial deletions of a series of putative enhancer elements in vivo or CBS elements in vitro, in conjunction with chromosome conformation capture and RNA-seq analyses, we show that deletions of outward-oriented CBS elements weaken the strength of long-distance intraTAD promoter-enhancer interactions and enhancer activation of target genes. Our data highlight the crucial role of outward-oriented CBS elements within the clustered CTCF TAD boundaries and have interesting implications on the organization principles of clustered CTCF sites within TAD boundaries.

overall_design: RNA-seq of P0 old wild type (WT) and several mouse line mice and HEK293T cells were seperately generated by deep sequencing using Illumina Novaseq 6000 or HiSeq 2500

first: X

first: H

last: Huang

first: K

last: Han

first: W

last: Xu

first: Z

last: Wang

first: Q

last: Wu


title: Comparing the transcriptomic profiles of anti-viral candidates knockout cells

status: Public on Oct 30 2023

city: Cleveland

first: Daniel

last: McGrail

state: Ohio

address: 9500 Euclid Avenue

department: Center for Immunotherapy and Precision Immuno-Oncology

institute: Cleveland Clinic

summary: Host anti-viral factors are essential for controlling SARS-CoV-2 infection but remain largely unknown due to the biases of previous large-scale studies toward pro-viral host factors. To fill in this knowledge gap, we perform a genome-wide CRISPR dropout screen and integrate analyses of the multi-omics data of the CRISPR screen, genome-wide association studies, single-cell RNA-Seq, and host-virus proteins or protein/RNA interactome. This study uncovers many host factors that are missed by previous studies, including the components of V-ATPases, ESCRT, and N-glycosylation pathways that modulate viral entry and/or replication. The cohesin complex is also identified as an anti-viral pathway, suggesting an important role of three-dimensional chromatin organization in mediating host-viral interaction. Furthermore, we discover another anti-viral regulator KLF5, a transcriptional factor involved in sphingolipid metabolism, which is up-regulated and harbored genetic variations linked to COVID-19 patients with severe symptoms. Anti-viral effects of three identified candidates (DAZAP2/VTA1/KLF5) are confirmed individually. Molecular characterization of DAZAP2/VTA1/KLF5-knockout cells highlights the involvement of genes related to the coagulation system in determining the severity of COVID-19. Together, our results provide wealthy resources for understanding the host anti-viral network during SARS-CoV-2 infection and may help develop new countermeasure strategies.

overall_design: A549 derived KO cell lines and related negative control cell line were preseeded and cultured for 24 hrs prior to RNA isolation and stranded RNAseq.


title: Integrative multi-omic profiling of adult mouse brain endothelial cells and potential implications in Alzheimer’s disease

city: Guangzhou

first: Yage

last: Nie

address: No.74 Zhongshan Rd.2, Guangzhou

institute: Sun Yat-sen University

summary: The blood-brain barrier (BBB) is primarily manifested by a variety of physiological properties of brain endothelial cells (ECs), but the molecular foundation for these properties remains incompletely clear. Here, we generate a comprehensive molecular atlas of adult brain ECs using acutely purified mouse ECs and integrated multi-omics. Using RNA-seq and proteomics, we identify the transcripts and proteins selectively enriched in brain ECs and demonstrate that they are partially correlated. Using single-cell RNA-seq, we dissect the molecular basis of functional heterogeneity of brain ECs. Using integrative epigenomics and transcriptomics, we determine that TCF/LEF, SOX, and ETS families are top-ranked transcription factors regulating the BBB. We then validate the identified brain EC-enriched proteins and transcription factors in normal mouse and human brain tissue, and assess their expression changes in mice with Alzheimer’s disease. Overall, we present a valuable resource with broad implications for the BBB regulation and treatment of neurological disorders.

overall_design: Bulk-RNA-seq and ATAC-seq were performed on mouse brain, lung and liver endothelial cells. Single-cell RNA-seq was performed on adult mouse brain endothelial cells.

first: Min

last: Yu

first: JiaWen

last: Yang

first: Shilun

first: Rui

last: Li

first: Varsha

last: Rao

first: Xiaoyan

last: Hu

first: Cheng

last: Fang

first: Simeng

first: Dengpan

last: Song

first: Fuyou

last: Guo

first: Michael

middle: P

last: Snyder

first: Howard

middle: Y

last: Chang

first: Calvin

last: Kuo

first: Jin

first: Junlei


title: ETV4-Dependent Transcriptional Plasticity Maintains MYC Expression and Results in IMiD Resistance in Multiple Myeloma [ChIP-seq]

first: Benjamin

last: Barwick

address: 1365 Clifton Rd. NE WCI-C 4th Floor Benches 36-37

department: Hematology and Medical Oncology

summary: Immunomodulatory Drugs (IMiDs) are a backbone therapy for multiple myeloma (MM). Despite their efficacy, most patients develop resistance and the mechanism(s) are not fully defined. Here, we show that IMiD responses are directed by IMiD-dependent degradation of IKZF1 and IKZF3 that bind to enhancers necessary to sustain the expression of MYC and other myeloma oncogenes. IMiD treatment universally depleted chromatin-bound IKZF1, but eviction of P300 and BRD4 co-activators only occurred in IMiD-sensitive cells. IKZF1-bound enhancers overlapped other transcription factor-binding motifs, including ETV4. ChIP-seq showed that ETV4 bound to the same enhancers as IKZF1, and ETV4 CRISPR/Cas9-mediated ablation resulted in sensitization of IMiD-resistant MM. ETV4 expression is associated with IMiD resistance in cell lines, poor prognosis in patients, and upregulated at relapse. These data indicate that ETV4 alleviates IKZF1 and IKZF3 dependency in MM by maintaining oncogenic enhancer activity and identify transcriptional plasticity as a previously unrecognized mechanism of IMiD resistance.

overall_design: RNA-seq of multiple myeloma cell lines treated with lenalidomide as compared to control or with IKZF3 knock-down by Dox-inducible shRNAs

first: Paola

last: Neri

first: Nizar

last: Bahlis


title: Gene expression profiles of basophil progenitors from mouse bone marrow

city: Daejeon

first: Jiyeon

last: Park

address: Daehak-ro 291

country: South Korea

institute: KAIST

summary: To find genes governing basophil development or functional maturation, we analyzed the gene expression profiles of basophil progenitors by RNA-sequencing.

overall_design: BaPs, MAR1HI, MAR1INT cells were sorted from mouse bone marrow and subject to whole transcritome analysis. For each cell types, biological duplicates were analyzed

first: Yuri

last: Cho

first: Dongchan

first: Suk-Jo

last: Kang


title: NF-kB and Circadian Enhancers Link β-cell Function with Anti-IL1B Therapy [ChIP-Seq]

last: Weidemann

state: IL

address: 303 E. Superior St.

department: Medicine

institute: NORTHWESTERN UNIVERSITY

summary: Crosstalk between lineage-determining and signal-dependent transcription factors controls chromatin plasticity within heterogeneous tissues, yet how alterations in these interactions within single cells contribute to disease remains unknown. Here, by profiling chromatin activity in single nuclei of human pancreatic islets, we identified 19 clusters of cells, including β-cell subpopulations defined by differences in accessibility at non-coding cis-regulatory elements (CREs) for the lineage-determining factor pancreatic duodenal homeobox factor 1 (PDX1). Single cells with a high density of PDX1 CREs were enriched in accessible enhancer landscapes driving gene networks controlling nutrient sensing, insulin exocytosis, and circadian rhythms. In contrast, cells with reduced opening at PDX1 CREs had increased accessibility at regulatory elements for pro-inflammatory genes controlled by NF-kB and IL-1β. Genetic deficiency of Pdx1 in mice led to increased chromatin occupancy by the classical NF-κB subunit p65 and glucose intolerance that was more pronounced at the time of day when mice were awake and feeding. Within murine and human islets, PDX1 formed long-range repressive contacts with canonical regions of p65-mediated transcription. Pharmacological inhibition of signaling through the IL-1β receptor, an abundant β-cell target of p65, enhanced glucose-dependent insulin secretion in Pdx1-deficient β cells. Together, our single-cell epigenomic analyses provide a rationale to antagonize NF-kB signaling as an insulinotropic therapy for β-cell failure and diabetes.

overall_design: Single-cell ATAC-sequencing, RNA-sequencing, ChIP-sequencing, HiChIP-sequencing, gene manipulation, metabolic studies


title: ETV4-Dependent Transcriptional Plasticity Maintains MYC Expression and Results in IMiD Resistance in Multiple Myeloma [ATAC-seq]

institute: Emory University


title: YAP/TAZ regulates immunomodulatory properties of mesenchymal stem cells

city: Hiroshima-city

first: Hiroki

last: Yoshii

state: Hiroshima-ken

address: 1-2-3 Kasumi Minami-ku

institute: Hiroshima University

summary: mRNA sequencing of mesenchymal stem cells transfected with YAP/TAZ siRNAs were treated with or without TNF-a for 24hr to profile gene expressions.

overall_design: The siRNAs for human YAP, TAZ or negative control siRNA were chemically synthesized by FASMAC (FASMAC, Kanagawa, Japan). MSCs were seeded in 24-well plastic culture plates and maintained for 48 hr. Then, siRNAs (10 pmol/ml) were transfected into the cells by using Lipofectamine RNAi-MAX (Invitrogen, Carlsbad, CA). After 48 hr cultivation, MSCs were stimulated with or without 10 ng/ml TNF-a for 24 hr. RNA extraction was conducted by using RNA-iso (Takara). The construction and sequencing of the cDNA library were carried out by the Beijing Genomics Institute using the DNBSEQ platform.  This project contains 12 samples of (3 from sicontrol-MSCs with non-treated, 3 from sicontrol-MSCs with TNF-a-treated, 3  from siYAP/TAZ-MSCs with non-treated, and 3 from siYAP/TAZ-MSCs with TNF-a-treated) RNA-seq data.

first: Kajiya

last: Mikihito

first: Toshinori

last: Ando


title: Cyclic stretch-induced exosomes from periodontal ligament cells promote osteoblasts osteogenic differentiation via the miR-181d-5p/TNF signaling pathway

first: Fang

last: Qu

state: Shanghai

address: No.639 Zhizaoju Road

department: Department of Prosthodontics

institute: Shanghai Ninth People’s Hospital

type: Non-coding RNA profiling by high throughput sequencing

summary: Objective: To investigate the effects of stretch-induced periodontal ligament cell (PDLC) exosomes on osteoblast differentiation, and to explore their regulatory role in mechanical force-related periodontal tissue remodeling.Design: After applying 20% stretch loading to human periodontal ligament cells, exosomes were extracted from the supernatant and co-cultured with osteoblasts to detect their effects on osteogenic differentiation. Meanwhile, the exosomes were sequenced by high-throughput microRNA sequencing for bioinformatic analysis and validation to explore exosome signaling pathways through miRNAs.Results: Stretch-induced PDLC exosomes could be taken up by osteoblasts and promoted osteogenic differentiation of osteoblasts, as demonstrated by the increased expression levels of osteogenesis-related factors and enhanced ALP staining. In the miRNA profile of differentially expressed PDLC exosomes under stretch loading, miRNA-181d-5p was up-regulated significantly. The expression levels of osteogenesis-related factors and ALP staining were also increased in osteoblasts transfected with miR-181d-5p, and this effect might be related to the inhibitory role of exosomal miR-181d-5p on tumor necrosis factor (TNF).Conclusions: Stretch-induced PDLC exosomes exhibited a promoting effect on osteogenic differentiation, which might result from the inhibition of TNF via exosomal miR-181d-5p.

first: Yi-Fan

last: Zhang

first: Ying-Ying

last: Wang

first: Xi-Meng

last: Cao

first: Ying-Yi

last: Shen

first: Zi-Ang

last: Wu

first: Ya-Qin

first: Chun

last: Xu


title: Epigenetic repression of Cend1 by LSD1 is essential for murine heart development

city: Xi'an

first: Luo

last: Yuru

state: Shannxi

address: 28 Xianning West Road

department: Health Science Center

institute: Xi'an Jiaotong University

summary: Epigenetic regulation of heart development remains incompletely understood. Here we show that lysine-specific demethylase 1 (LSD1), a histone demethylase, critically controls cardiomyocyte proliferation during heart development. Cardiomyocyte-specific deletion of Lsd1 in mice inhibited cardiomyocyte proliferation, causing severe growth defect of embryonic and neonatal heart. In vivo RNA-seq and in vitro functional studies identified Cend1, a cell cycle repressor, as a target suppressed by LSD1. Loss of Lsd1 resulted in elevated Cend1 transcription associated with increased active histone mark H3K4me2 at the Cend1 promoter regions. Cend1 knockdown fully relieved the cell cycle arrest and proliferation defect caused by LSD1 inhibition in primary rat cardiomyocytes. Consistently, genetic deletion of Cend1 rescued cardiomyocyte proliferation defect and embryonic lethality in Lsd1 null embryos. Mechanistically, LSD1 promoted cardiomyocyte cycling and proliferation via CEND1-mediated suppression of p53 pathway. In line with the observations in mice, LSD1 promoted the cell cycle of cardiomyocytes derived from human-induced pluripotent stem cells by repressing CEND1-p53 axis. Together, these findings reveal an epigenetic regulatory mechanism consisting of the LSD1-CEND1-p53 axis that controls cardiomyocyte proliferation essential for murine heart development.

overall_design: the E12.5 heart ventricles tissue of control and LSD1-CKO mouse were aqcuired for total RNA extraction. The RNA extraction method was described in manuscript.

first: Rui

last: Zhou

first: Shanshan

last: Li


title: ETV4-Dependent Transcriptional Plasticity Maintains MYC Expression and Results in IMiD Resistance in Multiple Myeloma [RNA-seq]

city: Atlanta

first: Benjamin

last: Barwick

first: Paola

last: Neri

first: Nizar

last: Bahlis


title: Disruption of SUV39H1-mediated H3K9 methylation sustains CAR T cell function [ATAC-Seq]

status: Public on Oct 27 2023

city: New York

first: Michel

last: Sadelain

address: 1275 York Avenue, Box 182

institute: Memorial Sloan Kettering Cancer Cener

summary: Comparing genome accessibility profiles (ATACseq) of WT and SUV39H1-edited CAR T cells

overall_design: Unedited and SUV39H1-edited human CAR T cells were sorted from bone marrow (at day 50) of NSG mice that had undergone multiple rounds of NALM6 rechallenge

first: Nayan

last: Jain

first: Zeguo

last: Zhao

first: Richard

last: Koche

first: Anton

last: Dobrin


title: Disruption of SUV39H1-mediated H3K9 methylation sustains CAR T cell function [RNA-Seq]

summary: Comparing transcriptional profiles (RNAseq) of WT and SUV39H1-edited CAR T cells


title: NF-kB and Circadian Enhancers Link β-cell Function with Anti-IL1B Therapy [RNA-Seq 2]

last: Weidemann


title: Gene expression profile at single cell level of totoal peritoneal cells from WT and HDC KO mouse

city: Seoul

first: Kwang

last: Kim

address: 50 Yonsei-ro

institute: Yonsei University College of Medicine

summary: Histamine is required for full differentiation of macrophage. We used single cell RNA sequencing(sc-RNA seq) to characterize the impairment of macrophage differentiation in HDC KO at molecular level.

overall_design: The total peritoneal cells from mice were isolated using 10-ml syringe with ice cold PBS, and cells were prepared for scRNA-seq on a LUNA-FL Automated Fluorescence Cell Counter

first: Ki

middle: T

last: Nam

first: Soo

last: Cho


title: Spatial transcriptomics reveals expression of novel genes during the remodelling of the embryonic human heart

city: Newcastle upon Tyne

first: Deborah

last: Henderson

state: Tyne and Wear

address: International Centre for Life

department: Biosciences

institute: Newcastle University

summary: Congenital heart defects are very common, with around 2% of the population having a valve anomaly. During development of the heart, and specifically the heart valves, cells change from distinct undifferentiated populations of progenitors into specialized cells organized with precise spatial locations within complex tissue structures. Knowledge of both the spatial location and the transcriptional profile of the progenitors and differentiated cells is necessary to understand the differentiation process and the formation of the complex 3-dimensional architecture of the valves. Disruption to these processes leads to congenital valve defects and these are associated with significant morbidity (valve disease is the most common cause of cardiac surgery in the over-50s), particularly during aging. Here we utilized a spatial transcriptomic approach (Stahl et al, 2016; DeLaughter et al, 2016; Vickovic et al, 2019) to derive data about gene expression in the arterial valves of the human embryo, during the sculpting phases of their development.  We identified a number of genes that have not been previously linked to valve development and are strongly expressed in the remodelling valve leaflets. Examination of the mouse model for one of these, RBP1, shows previously undescribed defects in the great arteries and valve anomalies in mutant animals, highlighting that these genes are good candidate genes for causing BAV. Thus, we have developed new methodologies for ST data analysis and show that they have the ability to identify previously unknown developmentally important genes. ST is thus an exciting tool for novel gene discovery in the developing human embryo.

overall_design:
Spatial transcriptomics of 2 embryonic human hearts. Consecutive tissue sections were taken from each sample CS19 (5 sections) and CS16 (4 sections)
CS = Carnegie stage.

first: Rachel

last: Queen

first: Janet

last: Kerwin

first: Lorraine

last: Eley

first: Moira

last: Crosier

first: Jasmin

middle: E

last: Turner

first: Hannah

last: Soetjoadi

last: Baldock

first: Jonathon

last: Coxhead

first: Alex

last: Laude

first: Simon

last: Cockell

first: Bill

last: Chaudhry

first: Steven

last: Lisgo


title: Engineering irradiated tumor-derived microparticles as personalized vaccines to enhance antitumor immunity

city: Wuhan

first: Yu

last: Tian

address: NO.156,Wujiadun,Jianghan District

institute: Cancer center,Union hospital, Tongji medical colloge,Huazhong university of science and technology

summary: The inadequate activation of antigen-presenting cells, the entanglement of T cells, and the highly immunosuppressive conditions in the tumor microenvironment (TME) are important factors that limit the effect of cancer vaccines. Studies have shown that individualized and broad antigens can fully activate anti-tumor immunity and inhibiting the function of TGF-β can facilitate T cell migration to tumor sites. Based on our previous study, we introduced a new vaccine strategy by engineering irradiated tumor cell-derived microparticles (RT-MPs), which have both individualized and broad antigens, to induce broad antitumor effects and cause immunogenic death. Encouraged by the proinflammatory effects of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein and the high affinity between TGF-βR2 and TGF-β, we developed RT-MPs with the SARS-CoV-2 spike protein and TGF-βR2. We found that this spike protein and high TGF-βR2 expression induces the innate immune response and ameliorates the immunosuppressive TME, thereby promoting T cell activation and infiltration, and ultimately inhibiting tumor growth. In addition, when combined with anti-programmed death 1 (anti-PD-1) the engineered RT-MPs were able to generate an immune memory response and eliminate subcutaneous tumors. Our study provides a novel strategy for producing an effective personalized anti-tumor vaccine for clinical application.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for bone marrow derived macrophages which treated with RT-MPs or RT-MPs@S

first: Yajie

last: Sun

first: Shuhui

last: Wu

first: Ai

last: Huang


title: SUV39H1 Ablation Enhances Long-Term CAR-T Function in Solid Tumors.

city: Paris

first: Sebastian

last: Amigorena

address: 26 rue d'Ulm

institute: Institut Curie

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

overall_design: Refer to individual Series


title: Natural killer cell proliferation requires canonical IRE1 function during viral infection

city: Zwijnaarde

first: Sophie

last: Janssens

address: Technologiepark 71

department: Inflammation Research Center

country: Belgium

institute: VIB/UGhent


title: Inhibition of hepsin in human breast cancer explants

city: Helsinki

first: Topi

last: Tervonen

address: Haartmaninkatu 8

country: Finland

institute: University of Helsinki

summary: Human breast tumor explants were treated with the hepsin inhibitory antibody Ab25 (5μM, 48h).

overall_design: The raw data files in this submission are not de-multiplexed. All sample metadata are included in the EGFR_sample_information.xlsx file linked to the foot of this record.

first: Juha

last: Klefström


title: Targeting aberrant sialylation and fucosylation in prostate cancer cells using potent metabolic inhibitors

first: Jennifer

last: Munkley

state: Tyne & Wear

summary:
To identify signalling pathways altered by treating prostate cancer cells with the metabolic inhibitors P-SiaFNEtoc, Fucotrim I and Fucotrim II, we treated CWR22Rv1 cells with P-SiaFNEtoc, Fucotrim I and Fucotrim II for 72 hours and their respective vehicle control with DMSO.
We then performed differential gene expression, Gene Ontology and GSEA analyses using data obtained from RNA-seq of the control and treated cell lines.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for vehicle control CWR22Rv1 cells vs CWR22Rv1 cells treated with metabolic inhibitors.

first: Margarita

last: Orozco-Moreno

first: Eline

middle: A

last: Visser

first: Kirsty

last: Hodgson

first: Agnes

last: Hipgrave Ederveen

first: Kayla

last: Bastian

first: Emily

last: Archer Goode

first: Özden

last: Özturk

first: Johan

middle: F

last: Pijnenborg

first: Nienke

last: Eerden

first: Sam

middle: J

last: Moons

first: Emiel

last: Rossing

first: Ning

last: Wang

first: Noortje

last: de Haan

first: Christian

last: Büll

first: Thomas

last: Boltje


title: Natural killer cell proliferation requires canonical IRE1 function during viral infection (RNA-Seq)

first: Sophie

last: Janssens

summary: The unfolded protein response (UPR) aims to restore ER homeostasis under conditions of high protein folding load, a function primarily serving secretory cells. Additional, non-canonical UPR functions have recently been unraveled in immune cells. We addressed the function of the inositol-requiring-enzyme 1 (IRE1) signaling branch of the UPR in NK cells in homeostasis and microbial challenge. Cell-intrinsic compound deficiency (DKO) of IRE1 and its downstream transcription factor XBP1 in NKp46 + NK cells, did not affect basal NK cell homeostasis, or overall outcome of viral MCMV infection. However, mixed bone marrow chimeras revealed a competitive advantage in the proliferation of IRE1 sufficient Ly49H + NK cells after viral infection. CITE-Seq analysis confirmed strong induction of IRE1 early upon infection, concomitant with the activation of a canonical UPR signature. Therefore, we conclude that cell-intrinsic IRE1/XBP1 activation is required for NK cell proliferation early upon viral infection, as part of a canonical UPR response.

overall_design: Comparative gene expression profiling analysis of RNA-seq data between WT, SKO (Xbp1-KO) and DKO (Ire1Xbp1-KO) splenic NK cells.

first: Jessica

last: Vetters

first: Mary

last: Van Helden

first: Clint

last: De Nolf


title: Genome-wide CRISPR/Cas9 knockout screen for novel regulators of mechanical stress on NLRP3 inflammasome activation

status: Public on Oct 26 2023

city: Illkirch-Graffenstaden

first: Zhirong

last: ZHANG

address: 1 Rue Laurent fries

department: Stem celll and development

institute: IGBMC

summary: Immune cells sense the microenvironment to fine-tune their inflammatory responses. Patients with cryopyrin associated periodic syndrome (CAPS), caused by mutations in the NLRP3 gene, present auto-inflammation and its manifestation is largely dependent on environmental cues. However, the underlying mechanisms are poorly understood. Here, we uncover that KCNN4, a calcium-activated potassium channel, links PIEZO-mediated mechanotransduction to NLRP3 inflammasome activation. Yoda1, a PIEZO1 agonist, lowers the threshold for NLRP3 inflammasome activation. PIEZO-mediated sensing of stiffness and shear stress increases NLRP3-dependent inflammation. Myeloid-specific deletion of PIEZO1/2 protects mice from gouty arthritis. Activation of PIEZO1 triggers calcium influx, which activates KCNN4 to evoke potassium efflux promoting NLRP3 inflammasome activation. Activation of PIEZO signaling is sufficient to activate the inflammasome in cells expressing CAPS-causing NLRP3 mutants via KCNN4. Finally, pharmacologic inhibition of KCNN4 alleviates auto-inflammation in CAPS patient cells and in CAPS-mimicking mice. Thus, PIEZO-dependent mechanical inputs augment inflammation in NLRP3-dependent diseases including CAPS.

overall_design: This is a pooled genome-wide CRISPR/Cas9 knockout screen for novel regulators downstream mechanical stress to promote NLRP3 inflammasome activation. THP-1 cells stably expressing Cas9 protein were infected with the pooled Brunello lentivirus library. After puromycin selection for 3 days, cells were further expanded. To ensure a representation of minimal 500 cells per sgRNA, at least 4.0×107 cells were used in each group and primed with LPS in suspension for 3 hours and treated with vehicle (control group) or R837 plus Yoda1 (treated group). When pyroptotic cell death reached ~85% in the treated group, R837 and Yoda1 were washed out and the survivals were expanded. When cell number reached 4.0 × 107 cells, these cells were repeatedly treated with R837 plus Yoda1 to reach reached ~85% of pyroptotic cell death. R837 and Yoda1 were washed out and the survivals were further expanded to reach 4.0 × 107 cells. Genomic DNA from both control- and treated groups was isolated using the phenol/chloroform extraction method. Fragments containing sgRNA were amplified from genomic DNA by PCR and subjected to deep sequencing. We sequenced control-treated cells and survivals of R837 plus Yoda1-treated cells for each of the two replicates and performed MAGeCK analysis to identify the top hits.

first: Li

last: Ran

first: Tao

last: Ye

last: Zhang


title: Transcriptional modulation by PRMT5 inhibition with EPZ015666 (GSK3235025)

first: Daniel

last: McGrail

summary: Aberrant protein arginine methylation has been observed in multiple cancer types, making it an attractive drug target. Proteins can undergo asymmetric arginine methylation by type I protein arginine methyltransferases (PRMTs), predominately by PRMT1 and to a lesser extent PRMT4, or symmetric arginine methylation by type II PRMTs, predominately PRMT5. Here, we performed targeted proteomics following inhibition of PRMT1, PRMT4, and PRMT5 across cancer cell lines. We found that inhibition of both type I and type II PRMTs suppressed levels of total and phosphorylated ATR protein in cancer cell lines, and down-regulated expression of the ATR gene. Loss of ATR from PRMT inhibition resulted in defective DNA replication stress response activation in following exogenous replication stress. Since PARP inhibitors are known to induce replication stress, we next combined PRMT inhibition with PARP inhibition and found inhibition of PRMT1 or PRMT5 greatly exacerbated PARP inhibitor induced DNA damage. Based on this observation, we assessed the combination of PARP and PRMT inhibition in a panel of cell lines. While inhibition of both type I and type II PRMTs were synergistic with PARP inhibition in both cells with intact and deficient homologous recombination, type I PRMT inhibition resulted in higher toxicity in non-malignant cells. Therefore, we validated the synergy of combined PARP/PRMT5 inhibition in primary patient-derived organoids. Finally, we demonstrate that the combination of PARP and PRMT5 inhibition improves overall survival in both BRCA-mutant and wild-type patient-derived xenograft models without any detectable hematological toxicities typically associated with PARPi combination therapies. Taken together, these results demonstrate that PRMT5 inhibition may be a well-tolerated approach to improve tumor sensitivity to PARP inhibition. .

overall_design: Cells were treated with either DMSO vehicle control or 5 micromolar EPZ015666 for 3 days or 5 days prior to RNA isolation and stranded RNAseq.


title: Fungal metabolism and free amino acid content may predict nitrogen transfer to the host plant in the ectomycorrhizal relationship between Pisolithus spp. and Eucalyptus grandis

city: Richmond

first: Jonathan

middle: Michael

last: Plett

state: NSW

address: Bourke St. Entrance, Building L9

department: Hawkesbury Institute for the Environment

country: Australia

institute: Western Sydney University

summary: Ectomycorrhizal (ECM) fungi are crucial for tree nitrogen (N) nutrition, however, mechanisms governing N transfer from fungal tissues to the host plant are not well understood. ECM fungal isolates, even from the same species, vary considerably in their ability to support tree N nutrition resulting in a range of often unpredictable symbiotic outcomes. In this study, we used isotopic labelling to quantify the transfer of N to the plant host by isolates from the ECM genus Pisolithus known to have significant variability in colonisation and transfer of nutrients to a host. We considered the metabolic fate of N acquired by the fungi and found that the percentage of plant N acquired through symbiosis significantly correlated to the concentration of free amino acids present in the ECM extra-radical mycelium. Transcriptomic analyses complemented these findings with isolates having high amino acid content and N transfer showing increased expression of genes in amino acid transport and catabolic pathways. These results suggest that fungal N metabolism drives transfer to the host plant in this interaction and that relative N transfer may be possible to predict through basic biochemical analyses.

sample_organism:
Eucalyptus grandis
Pisolithus microcarpus

overall_design: In total 43 samples were analysed by RNA-sequencing of which 22 samples included 3-4 biological replicates each of 1 timepoint of symbiosis between Eucalyptus grandis and Pisolithus microcarpus (2 week post contact). The remaining samples included biological replicates of six genotypes of P. microcarpus grown axenically as a control and three biological replicates of axenically grown E. grandis. All organisms and symbiotic compartment was performed on ½ strength Modified Melin Norkans Medium (with 0.1g/L of glucose).

middle: M

first: Krista

first: Ian

last: Anderson


title: Natural killer cell proliferation requires canonical IRE1 function during viral infection (CITE-Seq)

overall_design: Mixed bone marrow chimeras were generated as follows: CD45.1 animals were sublethally irradiated, followed by replenishment of the hematopoietic system with a 50/50 mixture of bone marrow cells isolated from CD45.1.2 (wild type) and CD45.2 (NK-DKO-/-) animals. At least 8 weeks after successful reconstitution, animals were infected with MCMV Smith strain or left untreated (i.e. mock infected with DMEM). On days 1,5 and 4,5 post-infection, spleens were isolated and distinct congenically marked NK cells were sorted for CITE-Seq analysis. Uninfected animals were included on day 1,5 post-infection.


title: Dynamic metabolism of endothelial triglycerides protects against atherosclerosis

city: New Haven

first: Carlos

last: Fernandez-Hernando

address: 10 Amistad St

institute: Yale University

summary: Blood vessels are continually exposed to circulating lipids and elevations of ApoB containing lipoproteins cause atherosclerosis. Lipoprotein metabolism is highly regulated by lipolysis, largely at the level of the capillary endothelium lining metabolically active tissues. How large blood vessels, the site of atherosclerotic vascular disease, regulate the flux of fatty acids (FA) into triglyceride (TG) rich lipid droplets (LD) is not known. Here, we show that deletion of the enzyme, adipose triglyceride lipase (ATGL) in the endothelium, leads to neutral lipid accumulation in vessels and impairs endothelial dependent vascular tone and nitric oxide synthesis to promote endothelial dysfunction. Mechanistically, the loss of ATGL leads to endoplasmic reticulum stress-induced inflammation, thereby promoting EC dysfunction. Consistent with this mechanism, deletion of endothelial ATGL markedly increases lesion size in a model of atherosclerosis. Together, these data demonstrate that the dynamics of FA flux through LD impacts EC homeostasis and consequently large vessel function during normal physiology and in a chronic disease state

overall_design: Endothelial specefic ATGL knockout (ATGL ECKO) and control (Atgl f/f) mice were used in this study to explore the role of lipid droplet breakdown (rate limited by ATGL) on endothelial function. Mice used for single cellRNAseq were injected with AAV8-PCSK9 to raise total cholesterol levels and fed a western type diet for four-weeks. Aorta were harvested from these mice for single cell sequencing.

first: Nabil

last: Boutagy

first: Ana

last: Gamez-Mendez

first: Joseph

middle: W

last: Fowler

first: Hanming

first: Bal

middle: K

last: Chaube

first: Enric

last: Espluges

first: Sungwoon

last: Lee

first: Daiki

last: Horikami

first: Jiasheng

first: Kathryn

last: Citrin

first: Abhishek

last: Singh

first: Brian

middle: G

last: Coon

first: Yajaira

last: Suarez

first: William

middle: C

last: Sessa


title: The role of GCNT1 mediated O-glycosylation in aggressive prostate cancer.

first: Jennifer

last: Munkley

summary:
To identify signalling pathways altered by GCNT1 in prostate cancer cells, we established PC3 stable cell lines in which GCNT1 was overexpressed.
We then performed differential gene expression, Gene Ontology and GSEA analyses using data obtained from RNA-seq of the control and OE cell lines.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for Lenti-ORF negative control PC3 cells vs PC3 cells with GCNT1 overexpression.

first: Kirsty

last: Hodgson

first: Margarita

last: Orozco-Moreno

first: Emma

last: Scott

first: Rebecca

last: Garnham

first: Karen

last: Livermore

first: Huw

last: Thomas

first: Michelle

last: Lawson

first: Yuhan

last: Zhou

first: Jiepei

last: He

first: Abel

last: Bermudez

first: Fernando

last: Jose Garcia Marques

first: Kayla

last: Bastian

first: Gerald

last: Hysenaj

first: Emily

last: Archer Goode

first: Rakesh

last: Heer

first: Sharon

last: Pitteri

first: Ning

last: Wang

first: David

last: Elliott


title: Baseline CD11b+ CD115+ Co-housed vs specific pathogen free monocytes

city: Minneapolis

first: Thomas

last: Griffith

state: MN

address: 2231 6th Street SE

department: Department of Urology

institute: University of Minnesota

summary: The immune system of SPF mice is dominated by naïve phenotype immune cells, especially within the T cell compartment, and resembles that of neonatal humans (Beura et al., 2016; Reese et al., 2016). Sequential infection of SPF-housed laboratory mice with common experimental pathogens, such as MHV, MCMV, Listeria monocytogenes, LCMV, and influenza A virus (Berton et al., 2022; Reese et al., 2016), or cohousing (CoH) SPF mice with pet store mice carrying multiple pathogenic and commensal bacteria, viruses, and/or fungi (Beura et al., 2016) induces immune system alterations and maturations that more closely resemble the adult human immune system. Such microbial exposure drastically alters the composition and function of the immune system of laboratory mice, which can significantly influence the overall outcome (and survival) to subsequent infection. Here we report CD11b+ CD115+ monocytes sorted from female specific pathogen-free (SPF) C57BL/6N (B6) mice co-housed with petstore mice for 60 days are transcriptionally different than SPF CD11b+ CD115+ monocytes

overall_design: We performed gene expression profiling analysis using data obtained from RNAseq of sorted CD11b+ CD115+ monocytes from the spleens of four 60 day co-housed (with petstore mice)mice  and three specific pathogen free (SPF) mice

first: Matthew

middle: D

last: Martin

first: Cara

last: Skon-Hegg

first: Molly

last: Ingersoll

middle: S