title: Epigenetic and Expression Profiling of Tissue Selective Estrogen Compounds in Breast Epithelial Cell Lines (ChIP-seq)

summary: Current approaches to menopausal hormone therapy consist of compounds which provide tissue specific beneficial effects while working to minimize any increase in cancer risk associated with replacement therapy.  In this study, an examination of the genomic distribution of histone modification and expression changes in two breast epithelial cell lines, the normal MCF10A cell line and the MCF-7 breast cancer cell line, upon treatment with several of the current therapies used for treating post-menopausal symptoms is presented. Analysis of the observed changes upon treatment with either 17-estradiol (E2), a combined treatment with the top 10 most abundant Premarin chemical equivalents (EC10), the non-steroidal SERM, Bazadoxifene (BZA) and the TSEC; BZA combined with EC10 (Duavee) or BZA combined with E2 provides evidence of expression changes in genes involved in DNA replication, nucleosome assembly, RNA-splicing and processing, biological regulation and cell signaling, and many of the known steroid response genes.  There were distinct global changes in chromatin states observed that were specific to each of the breast epithelial cell type normal, basal MCF10A and ESR/PGR positive luminal breast cancer MCF7 cell lines and selective changes observed upon treatment with each of the treatment groups.  Specifically, we observed a global loss of H3K4ac upon treatment with BZA.  These changes were observed in both ESR1 bound and non-bound regions suggesting modifications in both promoters and enhancer regions.  The levels of H3K4ac upon treatment of BZA showed a decrease below basal non-treated H3K4ac levels in MCF7.  There was an increase observed with E2 and EC10 treatment in MCF7 that was mitigated by the addition of BZA to either steroid as a combined treatment.  This study provides a better understanding of the role of epigenetic modifications in endocrine responsiveness and provides evidence of an improved therapeutic profile for combined TSEC compounds for treating and managing the overall physiological effects of declining estrogen.

overall_design: 6 drug treatments in 2 cell lines with 2 biological replicates

first: Terri

last: Messier

first: Joseph

last: Boyd

first: Gary

middle: S

last: Stein


title: Adult nucleus accumbens single-nucleus RNA-seq dataset for "A dopamine-induced coordinated gene expression program regulates neuronal function and cocaine response"

status: Public on Jun 24 2020

city: Birmingham

first: Jeremy

last: Day

state: Alabama

address: 1825 University Blvd SHEL 910

department: Neurobiology

country: USA

institute: University of Alabama at Birmingham

type: Expression profiling by high throughput sequencing

summary: This dataset contains single-nucleus RNA sequencing results from adult rat brain (nucleus accumbens) and serves as the basis for characterization of transcriptional response to cocaine (20mg/kg, intraperitoneal injection). The goal of this experiment was to define the transcriptional response to cocaine across distinct cell types in the nucleus accumbens. Single-cell sequencing was carried out on FACS-sorted nuclei isolated from these experiments using the 10X Genomics Chromium single cell sequencing platform.

overall_design: This experiment contains 4 biological samples. Samples were adult rat nucelus accumbens brain tissue isolated from rats 1hr after treatment with either saline or cocaine. Tissue was lysed to allow for nuclei collection via centrifugation. FACS-sorted nuclei were subject to 10X Genomics GEM generation and barcoding, GEM-RT cleanup and cDNA amplication, and library construction using a Chromium Single Cell 3ʹ Reagent Kit (v3 chemistry). Libraries underwent directional, paired-end PolyA+ RNA-seq on an Illumina Next-seq 500. Each sample has multiple raw fastq files, corresponding to different sequencing lanes (e.g., L001, L002, etc) and different reads (e.g., R1, R2). Additional index files are also provided for each sample and each lane (I1).

first: Robert

middle: A

last: Phillps

first: Corey

middle: G

last: Duke

first: Jennifer

middle: J

last: Tuscher

first: Morgan

middle: E

last: Zipperly

first: Lara

last: Ianov


title: Striatal cultured neurons single-nucleus RNA-seq dataset for "A dopamine-induced coordinated gene expression program regulates neuronal function and cocaine response"

summary: This dataset contains single-nucleus RNA sequencing results from rat embryonic striatal neuronal cultures and serves as the basis for characterization of transcriptional response to dopamine (50µM), the Drd1 agonist SKF-38393 (1µM), and potassium chloride (25mM). The goal of this experiment was to define the transcriptional response to each of these stimulations across distinct cell types in cultured neurons. Single-cell sequencing was carried out on FACS-sorted nuclei isolated from these experiments using the 10X Genomics Chromium single cell sequencing platform.

overall_design: This experiment contains 4 biological samples. Samples were embryonic day 18 rat primary striatal neurons cultured for 11 days in vitro prior to treatment with vehicle (neurobasal media), dopamine (50µM), the Drd1 agonist SKF-38393 (1µM), and potassium chloride (25mM) for 1hr. Neurons were cultured on 12w plates (~250,000 neurons/well), treated with described stimuli for 1hr, washed, and harvested in lysis buffer prior to isolation of nuclei via centrifugation and FACS sorting. FACS-sorted nuclei were subject to 10X Genomics GEM generation and barcoding, GEM-RT cleanup and cDNA amplication, and library construction using a Chromium Single Cell 3ʹ Reagent Kit (v3 chemistry). Libraries underwent directional, paired-end PolyA+ RNA-seq on an Illumina Next-seq 500. Each sample has multiple raw fastq files, corresponding to different sequencing lanes (e.g., L001, L002, etc) and different reads (e.g., R1, R2). Additional index files are also provided for each sample and each lane (I1).


title: Dopaplex RNA-seq dataset for "A dopamine-induced coordinated gene expression program regulates neuronal function and cocaine response"

summary: This dataset contains whole-genome RNA sequencing results from rat embryonic striatal neuronal cultures and serves as the basis for characterization of multiplexed CRISPR/dCas9 gene activation of 16 genes induced by dopamine receptor activation. The goal of this experiment was to define the transcriptional response to multiplexed CRISPR-based upregulation of 16 dopamine-induced genes (Tent5b, Junb, Prox1, Nr4a2, Nptx1, Gsx1, Gadd45g, Btg2, Fosb, Nr4a1, Egr4, Fos, Sstr2, Gadd45b, Egr3, & Egr2), together termed "Dopaplex", as compared to a LacZ control sgRNA.

overall_design: This experiment contains 11 biological samples, each of which underwent directional, paired-end PolyA+ RNA-seq on an Illumina Next-seq 500. Samples were treated with Lacz sgRNA or Dopaplex sgRNAs (targeting the 16 genes outlined above), in addition to a dCas9-VPR fusion for CRISPR-based transcriptional activation. Datasets were obtained using RNA-seq from PolyA+ fractions fractions of RNA. Each sample has multiple files, corresponding to different sequencing lanes (e.g., L001, L002, etc) or different reads (e.g., R1, R2).

first: Katherine

last: Savell


title: Notch activity determines a common hepatocellular carcinoma subtype with unique molecular and clinicopathologic features [Venus]

status: Public on Sep 28 2020

city: New York

first: Yu-Jui

last: Ho

address: 1275 York Avenue

institute: Memorial Sloan Kettering Cancer Center

summary: Hepatocellular carcinoma (HCC) is the predominant primary liver cancer and an increasing cause of cancer-related deaths worldwide, due in part through increased obesity-associated nonalcoholic steatohepatitis (NASH) and associated fibrosis. The Notch signaling pathway is inappropriately “reactivated” in hepatocytes in obesity and plays an active role in the pathogenesis of NASH-associated fibrosis. We aimed to understand the involvement of Notch in the pathogenesis of mouse and human HCC.

overall_design: 6 samples of FACS-isolated hepatocytes (Venus-negative and -positive) from 3 different NASH diet-fed Notch-Venus reporter mice were analyzed.

first: Changyu

last: Zhu

first: Utpal

middle: B

last: Pajvani


title: RNA-seq datasets for enhancer RNA quantification in "Enhancer RNAs predict enhancer-gene regulatory links and are critical for enhancer function in neuronal systems"

status: Public on Jul 28 2020

summary: This dataset contains total RNA sequencing results from rat embryonic cortical, hippocampal, and striatal neuronal cultures treated with either vehicle (complete Neurobasal media) or potassium chloride (10mM) for 1hr prior to RNA extraction.

overall_design: This experiment contains 19 biological samples, each of which underwent directional, 75bp paired-end total RNA-seq on an Illumina NextSeq500 instrument. Samples were treated with neurobasal media (vehicle; samples V1C, V2C, V3C, V1H, V2H, V3H, V2S, V4S, V6S, V8S), or 10mM KCl (samples K1C, K2C, K3C, K1H, K2H, K3H, K1S, K5S, K7S) . The first letter of sample names denotes acute treatement (V = vehicle, K = KCl). The second digit denotes the unique replicate ID. The third digit denotes the region of cultured neuron (C = cortex, H = hippocampus, S = striatum). NEBnext rRNA depletion kit was used to remove ribosomal RNA to include both polyadenylated and non-polyadenylated RNA. Samples were sequenced across multiple lanes of the Illumina instrument, designated as L001, L002, L003, L004 in raw file names. Forward and reverse paired-end reads are denoted as "R1" or "R2" in raw file names.

first: Nancy

middle: V

last: Carullo

last: Phillips


title: RNA sequencing analysis of OTUD6B-knockdown KYSE30 cells

status: Public on Apr 01 2023

city: Shijiazhuang

first: li

last: lei

address: Jiankang Road 12

country: China

institute: the Fourth Hospital of Hebei Medical University

summary: Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are using RNA sequencing analysis to test the effects of OTUD6B-knockdown on the mRNA of KYSE30 cells.  Methods: KYSE30 cells were transfected with negative control or siOTUD6B for 48 hours in 1640 medium plus 10% serum. Three independent replicates were plated, transfected in parallel for each negative control and siOTUD6B. Results: Log-fold changes of mRNAs between negative control and siOTUD6B group were selected with a significance threshod of p<0.05. Conclusions: Our study determined the mRNA changes of OTUD6B-knockdown KYSE30 cells.

overall_design: KYSE30 cells mRNA profiles of NC (transfected with negative control siRNA) and siOTUD6B (transfected with OTUD6B siRNA) were generated by RNA sequencing.

first: Li

last: Lei


summary: The goal of this experiment was to determine how dopamine receptor activation in medium spiny neurons alters rapid transcriptional programs. To identify genes altered by dopamine, we used a rat primary striatal neuronal culture system in which striatal neurons were removed from the rat brain at embryonic day 18, cultured for 11 days in vitro using standard neuronal conditions, and treated neurons with either vehicle (neurobasal media) or dopamine (at a concentration of 1µM) for 1hr. This dataset contains PolyA+ RNA-seq results from this experiment, in which we identified 100 upregulated genes and 3 downregulated genes after dopamine stimulation using DESeq2 pipelines.

overall_design:
RNA-seq 1hr dopamine dataset for "A dopamine-induced coordinated gene expression program regulates neuronal function and cocaine response".
This experiment contains 6 biological samples, each of which underwent directional, paired-end PolyA+ RNA-seq on an Illumina HiSeq v4 instrument. Samples were treated with neurobasal media (vehicle; samples S37-1, S37-3, and S37-5), or 1µM dopamine (samples S37-2, S37-4, and S37-6) . Datasets were obtained using RNA-seq from PolyA+ fractions fractions of RNA.

first: Faraz

last: Sultan


title: Senescence rewires microenvironment sensing to facilitate anti-tumor immunity (scRNA-Seq)

status: Public on Jan 28 2023

summary: Cellular senescence involves a stable cell cycle arrest coupled to a secretory program that, in some instances, stimulates the immune clearance of senescent cells. Using an immune competent tumor model in which senescence triggers CD8 T cell-mediated tumor rejection, we show that senescence also remodels cell surface proteome to alter how they sense environmental factors, as exemplified by Type II interferon gamma (IFN-γ). Compared to proliferating cells, senescent cells upregulate IFN-γ receptor, become hypersensitized to microenvironmental IFN-γ, and more robustly induce antigen presenting machinery -effects also recapitulated in human tumor cells treated with senescence-inducing drugs. Disruption of the IFN-γ sensing by senescent cells blunts their immune-mediated clearance without disabling their characteristic secretory program or immune cell recruitment. Our results demonstrate that senescent cells have an enhanced ability to both send and receive environmental signals, and imply that each process is required for their effective immune surveillance.

overall_design: Characterization of single-cell transcriptional profiles of immune cells (CD45+) isolated directly by FACS from p53 off and p53 On (day 8) liver tumor-bearing mice to assess the effects of p53-restoration induced senescence in tumor cells on the immune compartments. 2-3 biological replicates (independent mice) were used per experimental condition.

first: Hsuan-An

last: Chen

first: Direna

last: Alonso-Curbelo

first: Scott

middle: W

last: Lowe


title: Senescence rewires microenvironment sensing to facilitate anti-tumor immunity

status: Public on Jan 27 2023

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

sample_organism:
Mus
Mus musculus

platform_organism:
Mus
Mus musculus

overall_design: Refer to individual Series


title: Specialized functions and sexual dimorphism explain the functional diversity of the myeloid populations during glioma progression

status: Public on Dec 06 2022

city: Warsaw

first: Pawel

last: Segit

address: Pasteur 3

country: Poland

institute: Nencki Institute of Experimental Biology, Polish Academy of Sciences

summary:
Cellular Indexing of Transcriptomes and Epitopes by sequencing (CITE-seq) used to dissect myeloid cell identities and states in murine experimental gliomas (GL261).

The study was supported by National Science Centre Poland research grant OPUS 2020/39/B/NZ4/0268

overall_design: CITE-seq sequencing analysis of 6 groups: female control (3 samples), female tumor D14 (14 days post implantation, 3 samples), female tumor D21 (21 days post implantation, 3 samples), male control (3 samples), male tumor D14 (14 days post implantation, 3 samples), male tumor D21 (21 days post implantation, 3 samples). Samples were hashed.

first: Natalia

last: Ochocka

first: Kamil

last: Wojnicki

first: Salwador

last: Cyranowski

first: Julian

last: Swatler

first: Karol

last: Jacek

first: Wieslawa

last: Grajkowska

first: Bozena

last: Kaminska


title: miRNA Microarray of cholesterol loaded and cholesterol depleted THP-1 macrophages

status: Public on Apr 14 2020

first: Coen

last: van Solingen

state: NY

address: 435 E30th street SB 704

department: Medicine/Cardiology

institute: NYU Grossman School of Medicine

summary:
THP-1 macrophages were treated with acLDL to load with cholesterol
THP-1 macrophages were treated with statin to deplete cholesterol

overall_design: SABiosciences miRNA PCR Assay for THP-1 macrophages treated with acLDL or statin.

first: Katey

last: Rayner

first: Kathryn

last: Moore


title: CAR T cells targeting uPAR are effective senolytics

status: Public on Apr 18 2020

first: Yu-Jui

last: Ho

summary: Cellular senescence is a stress-response program characterized by stable cell cycle arrest and a secretory program that modulates the tissue microenvironment.  Physiologically, senescence serves as a potent tumor suppressive mechanism that prevents the expansion of premalignant cells and plays a beneficial role in certain wound healing responses. Pathologically, the aberrant accumulation of senescent cells in response to chronic tissue damage produces an inflammatory milieu that contributes to diseases such as liver and lung fibrosis, atherosclerosis, diabetes and osteoarthritis.  Accordingly, experimental approaches to eliminate senescent cells from damaged tissues in mice can ameliorate symptoms of these pathologies and even promote longevity. Here we test the therapeutic concept that chimeric antigen receptor (CAR) T cells targeting senescent cells can serve as effective senolytics. We identify the urokinase plasminogen activator receptor (uPAR) as a cell surface protein that is broadly induced during senescence and demonstrate that uPAR-specific CAR T cells efficiently target senescent cells in vitro and in vivo. Administration of uPAR-targeting CAR T cells improves the survival of mice harboring lung adenocarcinoma cells treated with a senescence inducing drug combination, and eliminates fibrosis in mice treated with carbon tetrachloride or on a non-alcoholic steatohepatitis (NASH)-inducing diet. These results establish the potential of senolytic CAR T therapy for a range of senescence-associated diseases.

overall_design: Characterization of the transcriptional profiles of both murine therapy and oncogene induced senescence. For therapy induced senescence murine lung adenocarcinoma cell lines (KRASG12D,Tp53-/-) were cultured for 8 days with either DMSO or combined MEK and Cdk4/6 inhibitors to induce senescence. To study oncogene induced senescence hydrodynamic tail vein injection was performed to overexpress NRASG12V in murine hepatocytes and induce senescence.

first: Corina

last: Amor

first: Xiang

last: Li


title: Bulk DNA seq, GTO-seq, and RNA-seq with murin and human cell lines

status: Public on Mar 30 2020

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus


title: Single cell RNA sequencing of KPC1199 cell line [mouse scRNA-seq]

summary: In this study we describe a new method for GTO (Genomics Transcriptomics One-Tube) that allows for simeltaneous sequencing of RNA and DNA from a single cell in the same tube. This method relies on dual amplification. The first amplification is done immediately after the mRNA is transcribed to cDNA using SMART-Seq2 protocol (Picelli et al., 2014) to increase the level of cDNA to that comparable to gDNA. The second amplification with universal primers from SeqXE kit for whole genome amplification (from Sigma Aldrich) amplifies the cDNA and gDNA fragments to amounts required for library preparation. The reads generated from this mixed library are separated bioinformatically to exonic and nonexonic reads depending on where they align in the reference genome to generate the RNAseq and DNAseq data respectively. We present data generated from single in-vitro cells of multiple cell lines (A375, BT549, SKBR3, 315A) and also from single in-vivo cells isolated from tumors of an orthotopic tranplantation mouse model of pancreatic cancer (using KPC1199-EGFP cell line).

overall_design:
We first validated the robustness and high throughput capability of the GTO protocol in multiple cell lines. The key step to success of GTO is determining the optimum number of cycles for first amplification of cDNA. This step needs to be result in a balance between cDNA and gDNA reads so that the gene expression profiles generated are good while not comprimising on the CNA profile quality. This validation was done in SKBR3, A375 and BT549 cell lines. They were then compared to 315A control diploid cell line also processed by the GTO method. The scRNAseq and scDNAseq data generated by GTO was alsocompared to bulk RNAseq and DNAseq data generated in triplicates for each cell line by traditional methods. After the method was tested in cell lines, we next applied the method to mouse models to understand tumor biology. The mouse model we used here is an orthotopic transplantation model of pancreatic cancer in syngenic mice. EGFP positive KPC 1199 tumor cells were injected into the pancreas of adult wild-type mice to generate primary tumors in pancreas and metastases in liver, spleen, peritoneum and kidneys. These primary tumors and mets were then extracted and dissociated into single cell suspensions. The single cell suspensions were analyzed by flow cytometry with antibodies against CD45, Epcam and for endogeneous GFP to isolate single stromal and tumor cells. These cells were then processed by GTP protocol to analyze gDNA by CNV profiles and RNA through read counts from the same cell. To validate our data and also have controls, we generated scRNAseq gene expression data and scDNAseq CNA profiles from the 2D cells at the same passage number at which they were injected into the mice. The scRNAseq data was generated using Fluidigm C1 machine on theie medium IFC for mRNA seq according to user manual instructions. The scDNAseq data for CNA profiles was generated by sorting single nuclei into PCR plates and then using SeqXE kit from Sigma Aldrich for Whole genome amplification using their instructions again. The data analysis was done similar to scRNAseq or scDNAseq data from GTO.
single cell mRNA sequencing using Fluidigm C52

first: Naishitha

last: Anaparthy


title: Somatic tissue engineering in mouse models reveals an actionable role for WNT pathway alterations in prostate cancer metastasis

status: Public on Apr 28 2020

summary: To study genetic factors that influence the progression and therapeutic response of advanced prostate cancer, we developed a fast and flexible system that introduces genetic alterations relevant to human disease directly into the prostate glands of mice using tissue electroporation. These electroporation based genetically engineered mouse models (EPO-GEMM) recapitulate features of traditional germline models and, by modeling genetic factors linked to late stage human disease, can produce tumors that are metastatic and castration resistant.  Unexpectedly, a subset of particularly metastatic tumors acquired WNT pathway alterations, which are also associated with metastatic prostate cancer in humans. Harnessing features linked to the EPO-GEMM approach, we validate the WNT pathway as a key event in driving metastatic disease, a finding that we confirm in an orthogonal approach using mouse prostate organoids. Moreover, we show that tumors harboring WNT pathway alterations are sensitive to pharmacological WNT pathway inhibition. Thus, by leveraging the power of EPO-GEMMs, our studies reveal a functional role for WNT signaling in driving prostate cancer metastasis and validate the WNT pathway as an actionable therapeutic target in metastatic prostate cancer.

overall_design: For RNA-seq analysis of the transcriptional profiles of MPten and MP EPO-GEMM prostate tumors, as well as normal anterior lobes of prostates of wild-type (WT) C57BL/6, total RNA was extracted from bulk tissue using the RNeasy Mini Kit (Qiagen). Purified polyA mRNA was subsequently fragmented, and first and second strand cDNA synthesis performed using standard Illumina mRNA TruSeq library preparation protocols. Double stranded cDNA was subsequently processed for TruSeq dual-index Illumina library generation.

first: Josef

last: Leibold

first: Marcus

last: Ruscetti

first: Zhen

last: Cao

first: Timour

last: Baslan

first: Wassim

last: Abida

first: Min

last: Zhou

first: Judith

last: Feucht

first: Teng

last: Han

first: Francisco

middle: M

last: Barriga

first: Kaloyan

last: Tsanov

first: Leah

last: Zamechek

first: Amanda

last: Kulick

first: Sha

last: Tian

first: Nelson

middle: R

last: Salgado

last: Sanchez-Rivera

first: Elisa

last: de Stanchina

first: John

last: Wilkson

first: Cory

last: Abate-Shen

first: Philip

last: Watson

first: Lukas

last: Dow

first: Charles

middle: L

last: Sawyers


title: Single cell whole genome sequencing (WGS) of mouse KPC1199EGFP cell line [mouse WGS]

first: Yu-Jui

last: Ho

summary: We first validated the robustness and high throughput capability of the GTO protocol in multiple cell lines. The key step to success of GTO is determining the optimum number of cycles for first amplification of cDNA. This step needs to be result in a balance between cDNA and gDNA reads so that the gene expression profiles generated are good while not comprimising on the CNA profile quality. This validation was done in SKBR3, A375 and BT549 cell lines. They were then compared to 315A control diploid cell line also processed by the GTO method. The scRNAseq and scDNAseq data generated by GTO was alsocompared to bulk RNAseq and DNAseq data generated in triplicates for each cell line by traditional methods. After the method was tested in cell lines, we next applied the method to mouse models to understand tumor biology. The mouse model we used here is an orthotopic transplantation model of pancreatic cancer in syngenic mice. EGFP positive KPC 1199 tumor cells were injected into the pancreas of adult wild-type mice to generate primary tumors in pancreas and metastases in liver, spleen, peritoneum and kidneys. These primary tumors and mets were then extracted and dissociated into single cell suspensions. The single cell suspensions were analyzed by flow cytometry with antibodies against CD45, Epcam and for endogeneous GFP to isolate single stromal and tumor cells. These cells were then processed by GTP protocol to analyze gDNA by CNV profiles and RNA through read counts from the same cell. To validate our data and also have controls, we generated scRNAseq gene expression data and scDNAseq CNA profiles from the 2D cells at the same passage number at which they were injected into the mice. The scRNAseq data was generated using Fluidigm C1 machine on theie medium IFC for mRNA seq according to user manual instructions. The scDNAseq data for CNA profiles was generated by sorting single nuclei into PCR plates and then using SeqXE kit from Sigma Aldrich for Whole genome amplification using their instructions again. The data analysis was done similar to scRNAseq or scDNAseq data from GTO.

overall_design:
In this study we describe a new method for GTO (Genomics Transcriptomics One-Tube) that allows for simeltaneous sequencing of RNA and DNA from a single cell in the same tube. This method relies on dual amplification. The first amplification is done immediately after the mRNA is transcribed to cDNA using SMART-Seq2 protocol (Picelli et al., 2014) to increase the level of cDNA to that comparable to gDNA. The second amplification with universal primers from SeqXE kit for whole genome amplification (from Sigma Aldrich) amplifies the cDNA and gDNA fragments to amounts required for library preparation. The reads generated from this mixed library are separated bioinformatically to exonic and nonexonic reads depending on where they align in the reference genome to generate the RNAseq and DNAseq data respectively. We present data generated from single in-vitro cells of multiple cell lines (A375, BT549, SKBR3, 315A) and also from single in-vivo cells isolated from tumors of an orthotopic tranplantation mouse model of pancreatic cancer (using KPC1199-EGFP cell line).
single cell whole genome sequencing of KPC1199EGFp nuclei

first: Naishitha

last: Anaparthy


title: TNIK signaling imprints CD8+ T memory formation early after priming

status: Public on Feb 24 2020

city: Bern

first: Ramin

last: Radpour

address: Murtenstrasse 35

department: Medical Oncology

country: Switzerland

institute: Bern University Hospital

summary:
Purpose: The goal of this study is to compare NGS-derived transcriptome profiling (RNA-seq) of naïve/ effector (D6)/ memory (D80) p14 T cell [wild-type (WT) and Tnik-/- (KO)] isolated form spleen of naïve or adoptively transferred and LCMV-WE immunized mice.
Methods: Transcriptomic profiles of splenic naïve wild-type (WT) and Tnik-/- (KO) p14 T cells (8-week-old mice) or splenic effector (D6) and memory (D80) p14 T cells (post adoptive transfer and LCMV-WE immunization) were assessed in duplicates or triplicate by deep sequencing, using Illumina HiSeq 2500. qRT–PCR validation was performed using TaqMan and SYBR Green assays.
Results: We mapped about 30 million sequence reads per sample to the mouse genome (GRCm38 - mm10) and identified expressed transcripts in splenic WT and Tnik−/− p14 T cells. RNA-seq data confirmed stable expression of known housekeeping genes. Differentially expressed genes between the WT and Tnik−/− p14 T cells from naïve, D6 or D80 were identified with a fold change ≥1.5 and p-value <0.05. Hierarchical clustering and gene ontology analysis of differentially expressed genes uncovered genes that may contribute to regulation of stemness, transcriptional regulation, cell cycle regulation, survival and function of T cells.
Conclusions: Our study represents the first detailed transcriptome analysis of WT versus Tnik-/- p14 T cells in the context of acute viral infection, with biologic replicates, generated by RNA-seq technology. Our results show that TNIK regulates T cell fate. Evaluation of mRNA content in effector (D6) and memory (D80) p14 T cells revealed that TNIK-deficiency during T cell activation enhances proliferation, terminal differentiation and glycolysis of effector T cells, while compromising steady-state metabolic activity of memory T cells. We conclude that TNIK imprints memory formation early after CD8+ T cell priming.

overall_design: mRNA of naïve and adoptively transferred D6 or D80 wild-type (WT) and knock-out (KO) p14 Tcells isolated from spleen of naïve and LCMV-WE immunized mice, using Illumina HiSeq 2500.

first: Carla

last: Jaeger-Ruckstuhl

first: Carsten

last: Riether

first: Adrian

middle: F

last: Ochsenbein


title: Bulk DNA sequencing of A375, BT549, SKBR3 and 315A cell lines [cell line bulk DNA-seq]

overall_design:
In this study we describe a new method for GTO (Genomics Transcriptomics One-Tube) that allows for simeltaneous sequencing of RNA and DNA from a single cell in the same tube. This method relies on dual amplification. The first amplification is done immediately after the mRNA is transcribed to cDNA using SMART-Seq2 protocol (Picelli et al., 2014) to increase the level of cDNA to that comparable to gDNA. The second amplification with universal primers from SeqXE kit for whole genome amplification (from Sigma Aldrich) amplifies the cDNA and gDNA fragments to amounts required for library preparation. The reads generated from this mixed library are separated bioinformatically to exonic and nonexonic reads depending on where they align in the reference genome to generate the RNAseq and DNAseq data respectively. We present data generated from single in-vitro cells of multiple cell lines (A375, BT549, SKBR3, 315A) and also from single in-vivo cells isolated from tumors of an orthotopic tranplantation mouse model of pancreatic cancer (using KPC1199-EGFP cell line).
single cell whole genome sequencing of A375 DNA


title: Transcriptional profiling of p53 mutants in KP LUAD cells

status: Public on Mar 18 2020

summary: Transcriptional profiling of KP LUAD cells expressing a panel of p53 mutant cDNAs.

overall_design: 1 million KP LUAD cells were seeded in 10cm plates and immediately transduced with retroviral supernatants and 8ug/mL polybrene. Cells were grown for 48hrs before purifying using fluorescence activated cell sorting (FACS). All transductions were done in triplicate. Following transduction, stable GFP+ populations were purified by FACS on a FACSAria (BD Biosciences). 120 hours post-transduction, total RNA was isolated using the RNeasy Mini Kit (Qiagen) following standard manufacturer protocols.

first: Francisco

last: Sánchez-Rivera


title: GTO sequencing of Human Cell Lines [cell line GTO-seq]

overall_design:
In this study we describe a new method for GTO (Genomics Transcriptomics One-Tube) that allows for simeltaneous sequencing of RNA and DNA from a single cell in the same tube. This method relies on dual amplification. The first amplification is done immediately after the mRNA is transcribed to cDNA using SMART-Seq2 protocol (Picelli et al., 2014) to increase the level of cDNA to that comparable to gDNA. The second amplification with universal primers from SeqXE kit for whole genome amplification (from Sigma Aldrich) amplifies the cDNA and gDNA fragments to amounts required for library preparation. The reads generated from this mixed library are separated bioinformatically to exonic and nonexonic reads depending on where they align in the reference genome to generate the RNAseq and DNAseq data respectively. We present data generated from single in-vitro cells of multiple cell lines (A375, BT549, SKBR3, 315A) and also from single in-vivo cells isolated from tumors of an orthotopic tranplantation mouse model of pancreatic cancer (using KPC1199-EGFP cell line).
GTO sequencing to analyze RNA and DNA from single cell


title: GTO sequencing of Mouse Tumors In KPC transplantation model [mouse GTO-seq]

overall_design:
In this study we describe a new method for GTO (Genomics Transcriptomics One-Tube) that allows for simultaneous sequencing of RNA and DNA from a single cell in the same tube. This method relies on dual amplification. The first amplification is done immediately after the mRNA is transcribed to cDNA using SMART-Seq2 protocol (Picelli et al., 2014) to increase the level of cDNA to that comparable to gDNA. The second amplification with universal primers from SeqXE kit for whole genome amplification (from Sigma Aldrich) amplifies the cDNA and gDNA fragments to amounts required for library preparation. The reads generated from this mixed library are separated bioinformatically to exonic and nonexonic reads depending on where they align in the reference genome to generate the RNAseq and DNAseq data respectively. We present data generated from single in-vitro cells of multiple cell lines (A375, BT549, SKBR3, 315A) and also from single in-vivo cells isolated from tumors of an orthotopic tranplantation mouse model of pancreatic cancer (using KPC1199-EGFP cell line).
GTO sequencing to analyze RNA and DNA from single cell


title: Bulk RNA sequencing of A375, BT549, SKBR3 and 315A cell lines [cell line bulk RNA-seq]

overall_design: In this study we describe a new method for GTO (Genomics Transcriptomics One-Tube) that allows for simeltaneous sequencing of RNA and DNA from a single cell in the same tube. This method relies on dual amplification. The first amplification is done immediately after the mRNA is transcribed to cDNA using SMART-Seq2 protocol (Picelli et al., 2014) to increase the level of cDNA to that comparable to gDNA. The second amplification with universal primers from SeqXE kit for whole genome amplification (from Sigma Aldrich) amplifies the cDNA and gDNA fragments to amounts required for library preparation. The reads generated from this mixed library are separated bioinformatically to exonic and nonexonic reads depending on where they align in the reference genome to generate the RNAseq and DNAseq data respectively. We present data generated from single in-vitro cells of multiple cell lines (A375, BT549, SKBR3, 315A) and also from single in-vivo cells isolated from tumors of an orthotopic tranplantation mouse model of pancreatic cancer (using KPC1199-EGFP cell line).


title: Senescence-induced vascular remodeling creates therapeutic vulnerabilities in pancreas cancer

status: Public on Feb 03 2020

summary: KRAS mutant pancreatic ductal adenocarcinoma (PDAC) is characterized by a desmoplastic response that promotes hypovascularity, immunosuppression, and resistance to chemo- and immunotherapies. We show that a combination of MEK and CDK4/6 inhibitors that target KRAS-directed oncogenic signaling can suppress PDAC proliferation through induction of retinoblastoma (RB) protein-mediated senescence. In preclinical mouse models of PDAC, this senescence-inducing therapy produces a senescence-associated secretory phenotype (SASP) that includes pro-angiogenic factors that promote tumor vascularization, which in turn enhances drug delivery and efficacy of cytotoxic gemcitabine chemotherapy. In addition, SASP-mediated endothelial cell activation stimulates the accumulation of CD8+ T cells into otherwise immunologically “cold” tumors, sensitizing tumors to PD-1 checkpoint blockade. Therefore, in PDAC models, therapy-induced senescence can establish emergent susceptibilities to otherwise ineffective chemo- and immunotherapies through SASP-dependent effects on the tumor vasculature and immune system.

overall_design: For RNA-seq analysis of the transcriptional profiles of tumor cells from KPCmut transplant PDAC tumors treated for 10 days with vehicle or combined trametinib (1 mg/kg body weight), and palbociclib (100 mg/kg), total RNA was extracted from GFP+ tumor cells (sorted on a FACSAria (BD Biosciences)) using the RNeasy Mini Kit (Qiagen). Purified polyA mRNA was subsequently fragmented, and first and second strand cDNA synthesis performed using standard Illumina mRNA TruSeq library preparation protocols. Double stranded cDNA was subsequently processed for TruSeq dual-index Illumina library generation. For sequencing, pooled multiplexed libraries were run on a HiSeq 2500 machine on RAPID mode. Approximately 10 million 76bp single-end reads were retrieved per replicate condition. Resulting RNA-Seq data was analyzed by removing adaptor sequences using Trimmomatic (Bolger et al., 2014), aligning sequencing data to GRCh37.75(hg19) with STAR (Dobin et al., 2013), and genome wide transcript counting using HTSeq (Anders et al., 2015) to generate a RPKM matrix of transcript counts. Genes were identified as differentially expressed using R package DESeq2 with a cutoff of absolute log2FoldChange ≥ 1 and adjusted p-value < 0.05 between experimental conditions (Love et al., 2014). Functional enrichments of these differential expressed genes were performed with enrichment analysis tool Enrichr (Kuleshov et al., 2016) and the retrieved combined score (log(p-value) * z-score) was displayed.

first: Marcus

last: Ruscetti

last: Morris IV

first: Riccardo

last: Mezzadra

first: James

last: Russell

first: Josef

last: Leibold

first: Paul

last: Romesser

first: Janelle

last: Simon

last: Kulick

first: Myles

last: Fennell

first: Jinyang

last: Li

first: Robert

last: Norgard

last: Wilkinson

first: Direna

last: Alonso-Curbelo

first: Ramya

last: Sridharan

first: Daniel

last: Heller

first: Ben

middle: Z

last: Stanger

last: Sherr

first: Scott

last: Lowe


title: Vitamin B6 addiction in acute myeloid leukemia

status: Public on Jan 13 2020

first: Yu-Jui

last: Ho


title: Transcriptome upon PDXK knock-out in mouse and human leukemic cells

summary: To study transcriptomic changes upon PDXK knock-out in mouse and human leukemic cells

overall_design: For RNA-seq analysis of the transcriptional profiles of mouse MLLAF9-NrsG12D and human MOLM13 leukemic cells transduced with PDXK targeting sgRNA, total RNA was extracted from bulk tissue using the RNeasy Mini Kit (Qiagen). Purified polyA mRNA was subsequently fragmented, and first and second strand cDNA synthesis performed using standard Illumina mRNA TruSeq library preparation protocols. Double stranded cDNA was subsequently processed for TruSeq dual-index Illumina library generation.

first: Chichao

last: Chen


title: Genome-wide transcriptome profiling of human trabecular meshwork cells treated with TGF-β2.

status: Public on May 09 2022

city: Buncrana

first: Breedge

last: Conwell

address: Cromore rd

country: United Kingdom

institute: Ulster university

summary: we employed RNA-Seq to delineate the TGF-β2 induced changes in the transcriptome of normal primary human trabecular meshwork cells (HTM).

overall_design: Examine changes to gene expression in TGF-β2 treated human trabecular meshwork cells derived from patients

last: Callaghan

first: Karen

last: Lester

first: Brian

last: Lane

first: Xiaochen

last: Fan

first: Katarzyna

last: Goljanek-Whysal

first: David

last: Simpson

first: Carl

last: Sheridan

first: Colin

last: Willoughby


title: Analysis for transcriptome profiling changes in 4'-O-methylpyridoxine (OMP) treated leukemic cells

summary: To validate the transcriptome profiling changes upon pharmacologic inhibition of PDXK, a key enzyme in vitamin B6 metabolism pathway, mouse leukemic cells were treated with PDXK inhibitor OMP. mRNA was extracted and purified for RNA_seq.

overall_design: There are 3 replicates for vehicle or OMP treatment and 6 samples in total.

first: Bo

last: Li

first: Lingbo

last: Zhang

first: ChiChao


title: Analysis of methylation In childhood acute myeloid leukemia (AML)

status: Public on Jan 01 2020

city: Little Rock

first: Jason

middle: Eli

last: Farrar

state: AR

address: 1 Children's Way, #512-10

department: Pediatrics/Hematology-Oncology

institute: University of Arkansas for Medical Sciences

summary: The Illumina Human Methylation EPIC array was used to assess methylation status at initial diagnosis in bone marrow or peripheral blood specimens from children with acute myeloid leukemia.

overall_design: Disease state design

first: Timothy

last: Triche


title: Signature of CD45- metastatic niche cells at different stages of the metastatic disease

status: Public on Aug 28 2019

city: London

first: Probir

last: Chakravarty

address: 1Midland Road

institute: Francis Crick

summary: We analysed the signature of the non-immune cells from the metastatic niche and the distal lung using the breast tumour 4T1 cell line as a model of lung metastasis from breast cancer

overall_design: Syngenic Balb/c female mice were injected intravenously with 4T1 tumour cells and lung tissue were collected at day 5 and day 10. post-injection. RNA was isolated from CD45-Ter119- FACS sorted cells.

first: Luigi

last: Ombrato

first: Ilaria

last: Malanchi

first: Miriam

last: Llorian-Sopena


title: Alpha-ketoglutarate links p53 to cell fate during tumor suppression

status: Public on May 10 2019

first: Yu-Jui

last: Ho

summary: The tumor suppressor TP53 is mutated in the majority of human cancers, including over 70% of pancreatic ductal adenocarcinoma (PDAC). Wild-type p53 accumulates in response to cellular stress and regulates the expression of genes that alter cell fate and constrain tumorigenesis. p53 also modulates several cellular metabolic pathways, though it remains unclear whether particular p53-regulated metabolites contribute to tumor suppression or whether metabolic alterations driven by p53 mutation sustain cancer progression. Here, we show that restoring endogenous p53 function in cancer cells derived from a murine PDAC model driven by oncogenic Kras and a regulatable p53 short hairpin RNA (shRNA) rewires glucose and glutamine metabolism leading to the accumulation of α-ketoglutarate (αKG), an obligate substrate for several chromatin modifying enzymes. p53 induces transcriptional programs characteristic of premalignant differentiation, an effect that can be partially recapitulated by addition of cell permeable αKG. Similarly, enforcing αKG accumulation in p53-deficient PDAC cells though the inhibition of oxoglutarate (αKG) dehydrogenase (Ogdh), the enzyme that consumes αKG in the tricarboxylic acid cycle, reduces tumor-initiating capacity and promotes tumor cell differentiation. Decreases in 5-hydroxymethylcytosine (5hmC), an αKG-dependent chromatin modification, are associated with the appearance of p53 mutations in the transition from premalignant to de-differentiated malignant lesions, whereas increases in 5hmC accompany tumor cell differentiation triggered by either p53 restoration or Ogdh depletion. Together these data nominate αKG as an effector of p53-mediated tumor suppression whose accumulation in p53-deficient tumors can drive tumor cell differentiation and antagonize malignant progression.

overall_design: 6 samples were analyzed in duplicates of 3 conditions.  1. Control, KPsh cells grown on dox, treated with vehicle DMSO for 72 hours. 2. KPsh cells grown on dox, treated with 4mM cell permeable dimethyl-alpha ketoglutarate for 72 hours.  3. KPsh cells grown off dox for 8 days, treated with DMSO vehicle for 72 hours.

first: Lydia

last: Finley

first: Scott

last: Lowe

first: John

middle: P

last: Morris IV


title: KDM6A(UTX) is an epigenetic gatekeeper of mTORC1 signalling in cancer

status: Public on Jul 20 2022

summary: Large-scale genome sequencing efforts of human tumors identified epigenetic modifiers as one of the most frequently mutated gene class in human cancer. However, how these mutations drive tumor development and progression is largely unknown. Here, we identify the histone demethylase KDM6A as an important tumor suppressor in solid cancers, such as liver cancer and pancreatic cancer. KDM6A-deficient tumors show hyperactivation of mTORC1 signaling, whereas endogenous KDM6A re-expression in established KDM6A-deficient tumors diminishes mTORC1 activity by fostering the expression of crucial negative pathway regulators, such as DEPTOR, TSC1, and TSC2, resulting in tumor regression. Importantly, KDM6A expression in human tumors correlates with mTORC1 activity and KDM6A-deficient tumors exhibit increased sensitivity to mTORC1 inhibition. Hence, our results link KDM6A-dependent epigenetic remodeling to mTORC1 signaling and provide a potential therapeutic strategy for KDM6A-deficient tumors.

overall_design: Examination of 2 different genotypes of primary liver cancer cell lines, Myc_Utx (derived from murine KDM6A-deficient tumor) and Myc_p53 (as control, derived from murine p53 knockout tumor). Three biological replicates (tumor from different mice) were used in this study for each genotype. For Myc_Utx: Myc_Utx_2A, Myc_Utx_2B and Myc_Utx_2D cell lines were used. As reference/control, Myc_p53: Myc_p53_1, Myc_p53_2 and Myc_p53_3 were used.

first: Steffie

last: Revia

first: Darjus

last: Tschaharganeh


title: A gain-of-function p53 mutant oncogene promotes cell fate plasticity and myeloid leukemia through the pluripotency factor Foxh1

status: Public on May 08 2019

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Mutations in the TP53 tumor suppressor gene are common in many cancer types, including the acute myeloid leukemia (AML) subtype known as complex karyotype (CK) AML. Here, we identify a gain-of-function (GOF) p53 mutation that accelerates CK-AML initiation beyond p53 loss and, surprisingly, is required for disease maintenance. The p53R172H mutation (TP53R175H in humans) exhibits a neomorphic function by promoting aberrant self-renewal in leukemic cells, a phenotype that is present in hematopoietic stem and progenitor cells (HSPCs) even prior to their transformation. We identify the Forkhead box H1 transcription factor (Foxh1) as a critical mediator of mutant p53 function that binds to and regulates stem cell-associated genes and transcriptional programs. Our results identify a context where mutant p53 acts as a bona fide oncogene that contributes to the pathogenesis of CK-AML and suggests a common biological theme for TP53 gain-of-function in cancer.

overall_design:
RNA-sequencing of duplicated samples per condition were derived from idependent biological replicates and were then sequenced using Illumina Hiseq 2500
ChIP-sequencing of Foxh1 in p53R172H mutant murine AML cells

first: Evangelia

last: Loizou

first: Richard

last: Koche


title: Comparing CTEPH to Control and IPAH pulmonary arteries

status: Public on Apr 27 2019

city: Nashville

first: James

last: West

state: TN

address: 4200 E 9TH Ave, Box B133

department: Medicine

institute: Vanderbilt University

summary:
This sample set is cultured lymphocytes from the patients.

Samples surgically extracted from CTEPH pulmonary arteries were compared to post-transplant pulmonary arteries from IPAH and failed donor pulmonary arteries from controls
The goal was to determine if altered gene expression in CTEPH associated with development of chronic thrombi

overall_design: CTEPH samples were compared to IPAH and control

first: Anna

last: Hemnes


title: CD8+ T cells expand stem and progenitor cells in favorable but not adverse risk acute myeloid leukemia

status: Public on Apr 04 2019

first: Ramin

last: Radpour

summary: We performed a comprehensive transcriptomic profiling of BM derived leukemia stem cells (LSCs) and leukemia progenitor cells (LPCs) together with paired CD8+ T cells of AML patients from different molecular risk groups, as well as hematopoietic stem cells (HSCs) and progenitor cells (HPCs) with paired CD8+ T cells of controls. This analysis revealed that epigenetic alterations mainly via histone deacetylation reduced the expression of immune-related genes in bone marrow (BM)-infiltrating CD8+ in AML when compared to CD8+ T cells in normal BM and that silenced gene expression pattern correlated with an improved prognosis. Correlation network modeling indicated that CD8+ T cells regulate leukemia stem/progenitor cells (LSPCs) in favorable risk but not in adverse risk AML.

overall_design: We characterized the molecular signature of well-defined leukemia stem/progenitor cells and paired CD8+ T cells in patients with acute myeloid leukemia (AML) compared to control subjects.

first: Carsten

last: Riether

first: Adrian

last: Ochsenbein


title: CAR T cell trogocytosis and cooperative killing   regulate tumour antigen escape

status: Public on Mar 27 2019

summary: Chimeric antigen receptors (CARs) are synthetic receptors for antigen that reprogram T cell specificity, function and persistence. Patient-derived CAR T cells have demonstrated remarkable efficacy against a range of B-cell malignancies, and early trial results suggest activity in multiple myeloma. Despite high complete response rates, relapses occur in a large fraction of patients, some of which are antigen-negative and others antigen-low. Unlike mechanisms resulting in complete and permanent antigen loss, those leading to antigen-low tumour escape remain obscure. In murine leukaemia models, we show that CARs provoke reversible antigen loss through trogocytosis, an active process whereby target antigen is transferred to T cells, thereby decreasing target density on tumour cells and abating T cell activity by promoting fratricide killing and exhaustion. These mechanisms affect both CD28 and 4-1BB-based CARs, albeit differentially, depending on antigen density. They can be offset by cooperative killing and combinatorial targeting.

overall_design: Examination of mRNA CD19 expression in  NALM6 retrieved from relapsed mice treated with 19-BBζ CAR T cells (D0),  retrieved NALM6 after ex-vivo culture (D6), and  NALM6 culture in vitro (ctrl).

first: Mohamad

last: Hamieh

first: Michel

last: Sadelain


title: RNA-seq datasets for "A neuron-optimized CRISPR/dCas9 activation system for robust and specific gene regulation"

status: Public on Feb 25 2019

first: Jeremy

last: Day

summary: This dataset contains whole-genome RNA sequencing results from rat embryonic hippocampal neuronal cultures and serves as the basis for characterization of CRISPR/dCas9 gene activation in neuronal systems.

overall_design: This experiment contains 9 biological samples, each of which underwent directional, paired-end PolyA+ RNA-seq on an Illumina Next-seq 500. Samples were treated with Lacz sgRNA (LZ2, LZ4, & LZ5), Bdnf-I sgRNA (B16, B17, B18), or Bdnf-IV sgRNA (BIV11, BIV14, BIV15), in addition to a dCas9-VPR fusion. Datasets were obtained using RNA-seq from PolyA+ fractions fractions of RNA. Each sample has multiple files, corresponding to different sequencing lanes (e.g., L001, L002, etc) or different reads (e.g., R1, R2).

first: Katherine

last: Savell

first: Svitlana

last: Bach

first: Morgan

last: Zipperly

first: Jasmin

last: Revanna

first: Nicholas

last: Goska

first: Jennifer

last: Tuscher

first: Corey

last: Duke

first: Faraz

last: Sultan

first: Julia

middle: N

last: Burke

first: Derek

last: Williams

first: Lara

last: Ianov


title: Calibrated CAR activation potential directs alternative T cell fates and therapeutic potency

status: Public on Dec 17 2018

first: Yu-Jui

last: Ho

summary: CD19-specific CARs that comprise CD28 and CD3z signaling domains program highly performing effector functions that mediate potent tumor elimination, but they impart a relatively limited T cell lifespan. Increasing functional T cell persistence without reducing effector potency is therefore likely to further enhance the therapeutic success of 1928z CAR T cells. We demonstrate that the number and position of ITAMs in 1928z CAR T cells influence functional, phenotypic and transcriptional programs, resulting in profound effects on antitumor efficacy. Improved therapeutic potency of CAR T cells can thus be achieved by calibrating activation strength, thereby retaining memory functions and preventing exhaustion, without compromising effector functions. Our transcriptional analysis underscores the potential of ITAM dosage and position to direct different T cell fates. We were able to identify a novel CAR design, termed 1XX, which programs a favorable balance of effector and memory signatures, inducing increased persistence of highly functional CARs with the replicative capacity of long-lived memory cells and potent effector functions.

overall_design: In order to assess the different phenotypic and functional patterns of CARs encoding a single immunoreceptor tyrosine-based activation motif (ITAM), we compared the genome-wide transcriptional profiles of 1928z, 1XX and XX3 after CD19 antigen stimulation of TRAC-edited naïve T cells. Sorted naïve (TN), stem cell memory (TSCM) and effector (TEFF) CD8+ T cells served as controls.

first: Judith

last: Feucht

first: Jie

last: Sun

first: Justin

last: Eyquem


title: SRSF1 role in cellular gene expression and splicing

status: Public on Mar 20 2019

city: Boca Raton

first: Massimo

last: Caputi

state: Florida

address: 777 Glades Road

department: Biomedical Science

institute: Florida Atlantic University

summary: SRSF1 is an abundant RNA-binding protein with functions in mRNA splicing, stability, translation and transcription of cellular and viral genes. We analyzed the role that SRSF1 plays in cellular gene transcription and splicing by transfecting HEK293 cells with a SRSF1 expression plasmid and by analyzing the transcriptome of the transiently transfected cells 15 hrs and 48 hrs post transfection. We also explored the role of the SRSF1 domains by transfecting a deletion clone of SRSF1 carrying only the RNA Recognition Motifs 1 and 2 (RRM1,2) but not the Arg-Ser rich (SR) domain.

overall_design: Comparison of the transcriptome of HEK293 cells transfected with SRSF1,SRSF1  RRM12 and control expression vector at 15 and 48 hours post transfection

first: Evan

last: Clark

first: Sean

last: Paz


title: p53 Represses the Mevalonate Pathway to Mediate Tumor Suppression

status: Public on Oct 23 2018

summary: We have discovered that loss of wild-type p53 correlates with elevated expression of mevalonate pathway genes in murine liver cancer and in human tumors. Mechanistically p53 blocks activation of SREBP-2, the master transcriptional regulator of this pathway, by transcriptionally inducing the ABCA1 cholesterol transporter gene, which inhibits SREBP-2 maturation. In mice the increase in mevalonate gene expression occurs in premalignant p53-null hepatocytes at a stage when p53 is needed to actively suppress tumorigenesis. Either RNAi mediated suppression of key genes in the mevalonate pathway or pharmacological inhibition of its rate-limiting enzyme restricts the development of mouse hepatocellular carcinomas driven by p53 loss. Conversely, like p53 loss, ablation of ABCA1 promotes tumorigenesis in a murine model and is associated with increased SREBP-2 maturation. Our findings thereby demonstrate that repression of the mevalonate pathway is a crucial component of p53-mediated tumor suppression and outline the mechanism by which this occurs.

overall_design: 4 samples were analyzed in replicates of 2 conditions.  1. Control, p53 WT hepatocytes transduced with MYC transposons delivered by hydrodynamic tail vein injection and isolated 6 days after injection. 2. Experimental, p53 null hepatocytes transduced with MYC transposons delivered by hydrodynamic tail vein injection and isolated 6 days after injection.

first: Carol

last: Prives

first: Scott

last: Lowe

first: Sung-Hwan

last: Moon

first: Chun-Hao

last: Huang

first: Shauna

last: Houlihan


title: Macrophage reprogramming by negatively-charged membrane phospholipids controls infection

status: Public on Nov 02 2018

city: La Jolla

first: David

middle: Mario

last: Cauvi

state: California

address: 9500 Gilman Dr.

department: Surgery

institute: University of California, San Diego

summary: Extracellular vesicles (ECV) are heterogeneous membrane-enclosed structures containing proteins, nucleic acids and lipids that participate in intercellular communication by transferring their content to recipient cells. Although most of the attention has been directed at the biological effect of proteins and microRNA, the contribution of phospholipids present in ECV on cellular activation has not been extensively addressed. Here we investigated the biological effect of phosphatidylserine (PS) and phosphatidylcholine (PC), two phospholipids highly abundant in ECV. A transcriptomic analysis revealed that about 4,700 genes were specifically modified by exposing peritoneal macrophages to PS or PC liposomes in vivo. Among them, the expression of several chemokines and cytokines was highly upregulated by PS liposome treatment, translating into a massive neutrophil infiltration into the peritoneum capable of neutralizing a septic polymicrobial insult. Both the L and D stereoisomers of PS induced the same response, suggesting that the effect was related to the negative charge of the phospholipid head. We concluded that an increase in the internal negative charge of the cell triggers a signaling cascade activating an innate immune response capable of controlling infection.

overall_design: Total RNA isolated from peritoneal cells obtained by lavage from CD-1 mice intraperitoneally injected for 1 hour with POPC liposomes (400 µg at 1 mg/ml, n=3 mice), POPS liposomes (400 µg at 1 mg/ml, n=3 mice), or PBS (400 µl, controls n=3 mice)

first: Antonio

last: De Maio


title: NK cell-mediated cytotoxicity contributes to tumor control by a cytostatic drug combination

status: Public on Dec 01 2018

summary: Accumulating evidence suggests that molecularly targeted therapies, which were originally developed to target  the underlying cell autonomous genetic drivers of tumorigenesis, can provoke tumor specific immune  responses. Using immunocompetent mouse models of KRAS mutant lung adenocarcinoma, we show that a  combination of MEK and CDK4/6 inhibitors can induce natural killer (NK) cell immune surveillance that is  necessary for its full anti-tumor effect. Mechanistically, we show that the drug combination – but neither agent  alone – drives RB-mediated cellular senescence induction leading to potent cell cycle arrest and activation of  the immunomodulatory senescence-associated secretory phenotype (SASP). This, in turn, leads to an increase in  tumor-specific NK cell ligand expression and secretion of TNF-α that provokes NK cell-mediated targeting of  senescent tumor cells, culminating in tumor regressions and long-term survival in genetically engineered mouse  models of lung cancer. These studies identify a means and method of invoking a unique form of NK cell  mediated immune surveillance in lung tumors through molecularly targeted therapies that induce senescence.

overall_design: For RNA-seq analysis of transcriptional changes after drug treatment of human KRAS mutant lung and pancreas cancer cell lines, total RNA was extracted using the RNeasy Mini Kit (Qiagen) from cell lines following 8 day treatment with vehicle (DMSO), trametinib (25 nM), palbociclib (500nM), or both in combination. For RNA-seq analysis of transcriptional changes in Natural Killer (NK) cells in vivo, total RNA was extracted as described above from CD45+CD3-NK1.1+ NK cells sorted from the lungs of tumor bearing KP transplant mice following 1 week treatment with vehicle or combined trametinib (1 mg/kg body weight) or palbociclib (100 mg/kg body weight). PolyA mRNA was selected using beads coated with polyT oligonucleotides. Purified polyA mRNA was subsequently fragmented, and first and second strand cDNA synthesis performed using standard Illumina mRNA TruSeq library preparation protocols. Double stranded cDNA was subsequently processed for TruSeq dual-index Illumina library generation. For sequencing, pooled multiplexed libraries were run on a HiSeq 2500 machine on RAPID mode. Approximately 10 million 76bp single-end reads were retrieved per replicate condition. Resulting RNA-Seq data was analyzed by removing adaptor sequences using Trimmomatic (29), aligning sequencing data to GRCh37.75(hg19) with STAR (30), and genome wide transcript counting using HTSeq (31) to generate a RPKM matrix of transcript counts.

first: Marcus

last: Ruscetti

first: Josef

last: Leibold

first: Matthew

last: Bott

first: Myles

last: Fennell

first: Amanda

last: Kulick

first: Chi-Chao

last: Chen

first: Timour

last: Baslan

first: Nelson

last: Salgado

first: Francisco

last: Sanchez-Rivera

first: Sha

last: Tian

first: Paul

last: Romesser

first: John

middle: T

last: Poirier

first: Charles

last: Rudin

first: Elisa

last: de Stanchina

first: Eusebio

last: Manchado

last: Sherr


title: Genome-wide mapping and functional analysis of DNA methylation in clonal invasive crayfish

status: Public on Oct 11 2018

city: Heidelberg

first: Fanny

last: Gatzmann

address: Im Neuenheimer Feld 580

department: A130 Epigenetics

country: Germany

institute: DKFZ

type: Methylation profiling by high throughput sequencing

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

sample_organism:
Procambarus fallax
Procambarus virginalis

overall_design: Refer to individual Series


title: Genome-wide mapping and functional analysis of DNA methylation in clonal invasive crayfish [RNA-seq]

summary: The marbled crayfish (Procambarus virginalis) is a unique freshwater crayfish characterized by genetic uniformity, phenotypic variability, and substantial invasive potential. As invasion into different habitats occurs in the absence of genetic variation, epigenetic mechanisms have been suggested to mediate phenotypic adaptation. However, epigenetic regulation has not been analyzed in this organism yet. Here we show that the recently published P. virginalis draft genome sequence encodes a conserved DNA methylation system. Whole-genome bisulfite sequencing of multiple replicates and different tissues revealed a methylation pattern that is characterized by gene body methylation of housekeeping genes. Interestingly, this pattern was largely tissue-invariant, suggesting a function that is unrelated to cell-fate specification. Indeed, integrative analysis of RNA-seq datasets showed that gene body methylation correlated with stable gene expression, while unmethylated genes often showed a high degree of inter-individual expression variation. Our findings thus establish the methylome of an emerging model organism and suggest that methylation-dependent regulation of gene expression variability may facilitate the phenotypic adaptation and invasive spread of this animal.

platform_organism:
Procambarus fallax
Procambarus virginalis

overall_design: RNA-seq data of different tissues (hepatopancreas, abdominal musculature) from 6 different P. virginalis individuals and 2 different P. fallax individuals

first: Cassandra

last: Falckenhayn

first: Julian

last: Gutekunst

first: Katharina

last: Hanna

first: Guenter

last: Raddatz

first: Vitor

middle: C

last: Carneiro

first: Frank

last: Lyko


title: Expression analysis of genes modulated after knock-down of lncRNA CHROME.

status: Public on Sep 20 2018

first: Coen

last: van Solingen

summary: Thousands of long non-coding RNAs (lncRNAs) have been identified in the human genome, many of which are not conserved in lower mammals. The majority of these lncRNAs remain functionally uncharacterized and may have important implications in human physiology and disease. Here, we identify a primate-specific lncRNA, CHROME, which is increased in the plasma and atherosclerotic plaques of individuals with coronary artery disease compared to healthy controls. Using a loss-of-function approach, we show that CHROME functions as a competing endogenous RNA of microRNAs  and regulates the concentration and biological functions of target genes.

overall_design: We used three replicate samples of HEPG2 cells that were treated with shRNA for CHROME compated to three replicate control samples.

first: Elizabeth

last: Hennessy

first: Kathryn

last: Moore


title: A dynamic co-expression map of early inflorescence development in Setaria viridis provides a resource for gene discovery and comparative genomics

status: Public on Aug 17 2018

city: Saint Louis

first: Andrea

last: Eveland

state: MO

address: 975 North Warson Road, Donald Danforth Plant Science Center/ Eveland lab

institute: Donald Danforth Plant Science Center

summary: Setaria viridis is a small, rapidly growing grass species in the subfamily Panicoideae, a group that includes economically important cereal crops such as maize and sorghum. The S. viridis inflorescence displays complex branching patterns, but its early development is similar to that of other panicoid grasses, and thus is an ideal model for studying inflorescence architecture. Here we report detailed transcriptional resource that captures dynamic transitions across six sequential stages of S. viridis inflorescence development, from reproductive onset to floral organ differentiation. Co-expression analyses identified stage-specific signatures of development, which include homologs of previously known developmental genes from maize and rice, suites of transcription factors and gene family members, and genes of unknown function. This spatiotemporal co-expression map and associated analyses provide a foundation for gene discovery in S. viridis inflorescence development, and a comparative model for exploring related architectural features in agronomically important cereals.

overall_design: We profiled the transcriptome of S.viridis inflorescence primordia across six sequential developmental stages that captured key events from the reproductive transition to floral organ differentiation. Three to four biological replicates were used for each stage.

first: Jiani

last: Yang

first: Chuanmei

last: Zhu

first: Mathew

middle: S

last: Box

middle: A

last: Kellogg


title: Arid1a restrains Kras-dependent changes in acinar cell identity

status: Public on Jul 03 2018

first: Yu-Jui

last: Ho

summary: Mutations in members of the SWI/SNF chromatin remodeling family are common events in human cancer, but the mechanisms whereby disruption of SWI/SNF components alters tumorigenesis remain poorly understood.  To model the effect of loss of function mutations in the SWI/SNF subunit Arid1a in pancreatic ductal adenocarcinoma (PDAC) initiation, we directed shRNA triggered, inducible and reversible suppression of Arid1a to the pancreas of mice in the setting of oncogenic KrasG12D.  Arid1a cooperates with Kras in the adult pancreas as postnatal silencing of Arid1a following sustained KrasG12D expression induces rapid and irreversible reprogramming of acinar cells into mucinous PDAC precursor lesions. In contrast, Arid1a silencing during embryogenesis, concurrent with KrasG12D activation, leads to retention of acinar cell fate. Together, our results demonstrate Arid1a as a critical modulator of Kras-dependent changes in acinar cell identity, and underscore an unanticipated influence of timing and genetic context on the effects of SWI/SNF complex alterations in epithelial tumorigenesis.

overall_design: Identification of the transcritpional changes induced upon dox-inducible Arid1a knockdown in mutant Kras pancreatic epithelial cells: Arid1a-downregulated vs Arid1a-expressing mutant Kras-pancreatic epithelial cells were FACS-sorted from pancreata from p48Cre;RIK;LSL-KrasG12;TRE-shRNA mice (harbouring Arid1a shRNAs -6421 or 1803- or control Renilla shRNA -713-) at day 5 post- doxycycline treatment. Mice were placed on doxycycline diet at 5 weeks of age to induce expression of the indicated shRNAs. 3 biological replicates (independent mice) were used per shRNA condition (713, 6421, 1803).

first: Geulah

last: Livshits

first: Scott

last: Lowe


title: Brassinosteroids modulate meristem fate and differentiation of unique inflorescence morphology in Setaria viridis

status: Public on Dec 23 2017

summary: RNA-seq was performed to profile the transcriptomes of inflorescence primordia hand-dissected from the bristleless1-1 (bsl1-1)mutant in Setaria viridis compared to wild-type controls sampled under the same conditions. Bsl1 encodes a rate limiting enzyme in BR biosynthesis, which is the ortholog of D11 from rice. Mutants are characterized by a homeotic conversion of sterile bristles to spikelets in the inflorescence.

overall_design: Inflorescence primordia were hand-dissected from bsl1 mutant and wild-type control Setaria plants at 15 Days After Sowing (DAS). Plants were grown together in a controlled growth chamber. Inflorescence primordia were pooled (~10 representative primordia per pool) into biological replicates (3 for wild-type and 2 for mutant).  Transcriptomes were sequenced and compared for differentially expressed genes.


title: A multidimensional blood stimulation assay reveals immune alterations underlying systemic juvenile idiopathic arthritis

status: Public on Sep 18 2017

city: Dallas

first: Nicole

last: Baldwin

address: 3434 Live Oak St

institute: Baylor Research Institute

first: Alma-Martina

last: Cepika

first: Romain

last: Banchereau

first: Brandi

last: Cantarel

first: Marina

last: Ohouo

first: Kristina

last: Goller

first: Victoria

last: Cantrell

first: Emily

last: Ruchaud

last: Gatewood

first: Phuong

last: Nguyen

first: Esperanza

last: Anguiano

first: Sandra

last: Zurawski

first: Jeanine

last: Baisch

first: Marilynn

last: Punaro

first: Gerlinde

last: Obermoser

first: Jacques

first: Karolina

last: Palucka

first: Virginia

last: Pascual


title: Healthy adult and sJIA blood stimulation with innate immune ligands

status: Public on Sep 19 2017

first: Nicole

last: Baldwin

summary: The goal of this study was to characterize altered inducible immune networks in Systemic onset juvenile idiopathic arthritis (sJIA), an IL-1-driven autoinflammatory disease of unknown etiology. To this end, we developed a high-throughput assay that quantifies the transcriptional and protein-level responses of blood leukocytes to innate stimuli. Herein, we report transcriptional data from healthy adult blood stimulated with 16 different conditions, including TLR ligands, cytosolic receptor ligands and inflammatory cytokines. We further report blood transcriptional profiles from sJIA patients with various disease activity and treatment statuses, both ex vivo (baseline) and after in vitro stimulation with a subset of innate stimuli including heat-killed bacterial pathogens.

overall_design:
291 total samples, 31 donors and 17 stimuli (excluding media controls). All stimuli were not used with all donors. There is one technical replicate for donors BD1277, BD1350, BD1635 for all stimuli - the replicate was run in a later batch (larger barcode number).

Stimuli used were agonists for surface Toll-like receptors (CpG-C, R837,  R848, TLR8L, Flagellin, LPS), a protein kinase C activator (PMA/I: phorbol myristate acetate and ionomycin), cytokines (IFNalpha, IFNgamma, IL-17A/F,  IL-18, IL-1beta, TNFalpha), gram+ derived ligands (PGN, LTA), and  heat-killed Staph. aureus (HKSA) as well as heat-killed Staph enteric (HKSE).

Ligands were diluted in medium (RPMI 1640 with GlutaMax, Thermo Fisher  Scientific, Grand Island, NY, USA) and aliquoted into 2 mL polypropylene  96-well plates (Greiner Bio-One, Monroe, NC, USA). Blood was drawn in the  morning into the BD Vacutainers containing ACD anticoagulant (BD, Franklin  Lakes, NJ, USA). Time between blood draw and culture setup was less than  2h. Blood and appropriately diluted ligands were mixed in 1:1 ratio in  total volume of 1 mL/well and incubated for 6h at 37oC/5% CO2. After the culture, blood was mixed and  100 uL aliquot set aside for activation marker phenotyping and  intracellular cytokine staining (ICS) for IL-1b. The plate was then  centrifuged at 12,000 x g/15  and 400 mL of plasma aliquoted and stored at  -80oC until the Luminex analysis. The pellet was the  lysed with Tempus  solution (Tempus blood RNA tubes, Thermo Fisher Scientific) in 2:1 ratio  and stored at -20oC until the RNA extraction.

Please note that data were analyzed using either V1 manifest (pre-Oct 2010) or the V2 manifest (post-Oct 2010) as indicated in the sample title.

Sample Group abbreviation definitions:
sIU - SoJIA, Inactive, Untreated
sAU - SoJIA, Active, Untreated
sIU - SoJIA, Inactive, Anakinra

first: Elizabeth


title: A multidimensional blood stimulation assay reveals immune alterations underlying systemic juvenile idiopathic arthritis [RNA-Seq]

summary: Despite great scientific and technological advances, immune alterations predisposing to sporadic human inflammatory diseases remain mostly unknown. To fill this gap, we developed a strategy to evaluate blood leukocyte responses to innate stimuli, simultaneously at the transcriptional, cellular and secreted protein levels. The data were integrated using weighted gene co-expression network analysis. When applied to systemic juvenile idiopathic arthritis (sJIA), an autoinflammatory disease of unknown etiology, this approach identified gene sets associated with specific cytokine environments and activated leukocyte subsets. During the remission phase of disease and off treatment, sJIA patients displayed dysregulated responses to TLR4, TLR8 and TLR7 stimulation in blood. Isolated sJIA monocytes accumulated higher levels of intracellular IL-1β after stimulation, and underexpressed the IL-1 inhibitor AHR at baseline. In line with the recent demonstration that AHR downregulation in monocytes is linked to macrophage differentiation, we show that the differentiation of sJIA monocytes in vitro was skewed towards macrophages, away from dendritic cell phenotype. This might contribute to the increased incidence of macrophage activation syndrome in these patients. The integrated analysis of these high-dimensional data can thus help unravel underlying immune alterations predisposing to complex inflammatory diseases.

overall_design: 32 total samples, 4 control healthy in vitro, 6 control systemic JIA in vitro, 4 LPS healthy in vitro, 6 LPS systemic JIA in vitro, 5 healthy ex vivo, 7 systemic JIA ex vivo

first: Alma-Martina

last: Cepika

first: Romain

first: Elodie

last: Segura

first: Jinghua

last: Gu

first: Sebastian

last: Amigorena


title: Extra-coding RNAs regulate neuronal DNA methylation dynamics

status: Public on May 31 2016

first: Jeremy

last: Day


title: Extra-coding RNAs regulate neuronal DNA methylation and long-term memory formation [RNA-seq]

summary: This dataset contains whole-genome RNA sequencing results from cortical neuronal cultures and serves as the basis for characterization of extra-coding RNA species from neuronal systems.

overall_design: This experiment contains six biological samples, each of which underwent PolyA+ and PolyA- RNA-seq. Samples were either unstimulated (i.e., treated with media alone; samples V1 and V2), stimulated with 25mM potassium chloride for 1hr (K1, K2) or inactivated with tetrodotoxin for 1hr (T1, T2). Datasets were obtained using RNA-seq from PolyA+ fractions or PolyA- fractions of RNA. PolyA- fractions are denoted "ec". Thus, 12 samples are listed here due to the difference in RNA library preparation.

first: Katherine

middle: E

last: Savell

first: Nancy

last: Gallus

first: Esther

last: Song

first: Rhiana

last: Simon

first: Mary

middle: K

last: Osborn

first: Jordan

last: Brown

first: John

middle: J

last: O'Malley

first: J

middle: D

last: Sweatt


title: H2A.Z knockdown RNAseq

status: Public on Sep 09 2014

summary: This experiment seeks to ascertain the transcriptional changes in the adult mouse hippocampus (CA1 subregion) that occur following viral knockdown of the histone variant H2A.Z. We are especially interested in understanding the role of this histone variant in memory formation and memory maintenance in the adult central nervous system.

overall_design: This experiment includes 3 groups, each with 3 biological replicates. Samples S108, S109, and S110 are from controls infected with an AAV expressing a scrambled shRNA control. Samples A100, A101, A102, A104, A106, and A107 were infected with an AAV expressing an shRNA against H2A.Z. Samples A100, A101, and A102 were naive animals, whereas samples A104, A106, and A107 were trained in contextual fear conditioning.

first: Iva

middle: B

last: Zovkic

first: Brynna

last: Paulukaitis

first: Deepa

middle: M

last: Etikala


title: FASCIATED EAR4 encodes a bZIP transcription factor that controls shoot meristem size in maize

status: Public on Jan 21 2015

first: Andrea

middle: L

last: Eveland

summary: fasciated ear4 (fea4) is a semi-dwarfed mutant with fasciated ears and tassels, and greatly enlarged vegetative and inflorescence meristems.  Chromatin Immunoprecipitation-sequencing (ChIP-seq) and expression profiling by RNA-seq suggest that fea4 is required to regulate the auxin response and leaf differentiation programs in the periphery of the meristem, suggesting a new mechanism of meristem size regulation that is spatially and mechanistically distinct from the CLV-WUS model.

overall_design: To gain insight into transcriptional regulation by FEA4, we used RNA-seq to profile genome-wide expression changes in developing ear primordia of fea4 loss-of-function mutants compared to fea4/+ wild-type (wt) siblings.  To map genome-wide occupancy of FEA4 and define its putative transcriptional targets, we performed ChIP-seq using the pFEA4-YFP::FEA4 transgenic lines.

first: Michael

last: Pautler

first: David

last: Jackson


title: Extra-coding RNAs regulate neuronal DNA methylation and long-term memory formation [MBD-seq]

summary: This dataset contains whole-genome MBD (methylbinding domain) sequencing results from cortical neuronal cultures and serves as the basis for characterization of DNA methylation profiles from neuronal systems.

overall_design: This experiment contains three sequencing datasets from 2 biological samples. Two datasets originate from samples that underwent MBD-capture prior to whole-genome sequencing. A third dataset contains non-MBD-captured genomic DNA as a control.


title: Regulatory Modules Controlling Maize Inflorescence Architecture

status: Public on Nov 27 2013


title: Regulatory Modules Controlling Maize Inflorescence Architecture: ChIP-seq data

summary: In this study we investigated the developmental dynamics of genes targeted in vivo by the transcription factor RAMOSA1, a key regulator of determinacy, and revealed potential mechanisms for repressing branches in distinct stem cell populations in developing maize inflorescences.  To identify targets of RA1 and to distinguish direct vs. indirect interactions, we performed Chromatin Immunoprecipitation (ChIP)-seq and compared the results to gene expression data (RNA-seq datasets for Eveland et al., 2013, submitted).  We mapped genome-wide occupancy of RA1 and showed that it differently regulates modules of target genes based on spatiotemporal context.

overall_design: Plants expressing complementing RA1 transgenes tagged with HA or YFP were used in parallel experiments.  Ear and tassel primordia were collected and tag-specific antibodies were used to pull down RA1 bound to its target loci.  Genome-wide analysis of RA1 occupancy revealed thousands of putative binding sites (i.e. peaks significantly enriched (p < 1e-05) compared to input DNA).

first: Doreen

last: Ware

first: Kengo

last: Morohashi

first: Erich

last: Grotewold

first: Erik

middle: W

last: Vollbrecht


title: MicroRNA expression data from human breast cancer cell lines after demethylation treatment.

status: Public on Jan 24 2012

first: Ramin

last: Radpour

summary:
The contribution of aberrant DNA methylation and the downstream effects in tumorogenesis through silencing of tumor suppressor genes (TSGs) and microRNAs has been investigated. Since these epigenetic alterations can be reversed, we investigated the effects of the epigenetic therapy in breast cancer cell lines.
We used microarrays to investigate the global microRNA expression profile after demethylation treatment with 5-aza-2’-deoxycytidine (DAC) in breast cancer cell lines and identified distinct classes of early and late systematic stable or transient effects of the treatment.

overall_design: Three selected breast cell lines including MDA-MB231, SKBR3, BT549, HS578T, MCF7 and HB2 (a breast epithelial cell line as control) were subject for miRNA isolation before treatment, after treatment with DAC and at five point follow-ups (1st, 3rd, 5th, 7th and 10th passages) at “drug holiday” condition and hybridized on Affymetrix microarrays.

first: Frank

last: Staedtler

first: Martin

last: Schumacher

first: Nicole

last: Hartmann


title: MRNA expression data from human breast cancer cell lines after demethylation treatment.

summary:
The contribution of aberrant DNA methylation and the downstream effects in tumorogenesis through silencing of tumor suppressor genes (TSGs) and microRNAs has been investigated. Since these epigenetic alterations can be reversed, we investigated the effects of the epigenetic therapy in breast cancer cell lines.
We used microarrays to investigate the global gene expression profile after demethylation treatment with 5-aza-2’-deoxycytidine (DAC) in breast cancer cell lines and identified distinct classes of early and late systematic stable or transient effects of the treatment.

overall_design: Three selected breast cell lines including MDA-MB231 (a highly aggressive cell line), SKBR3 (a non-aggressive cell line) and HB2 (a breast epithelial cell line as control) were subject for RNA isolation before treatment, after treatment with DAC and at five point follow-ups (1st, 3rd, 5th, 7th and 10th passages) at “drug holiday” condition and hybridized on Affymetrix microarrays.


title: Expression data from human breast cancer cell lines after demethylation treatment

type:
Expression profiling by array
Non-coding RNA profiling by array

platform_organism:
Homo sapiens
synthetic construct


title: Regulatory Modules Controlling Maize Inflorescence Architecture: mRNA-seq data

summary: In this study we used the maize (Zea mays) inflorescence to investigate gene networks that modulate determinacy, specifically the decision to allow branch growth.  We characterized developmental transitions by associating spatiotemporal expression profiles with morphological changes resulting from genetic perturbations that disrupt steps in a pathway controlling branching.  These are the RNA-seq datasets used in this study.

overall_design: We profiled changes in gene expression during normal maize ear and tassel development and in developing maize ear primordia upon genetic perturbation of the RAMOSA branching pathway.  For the wild-type ear and tassel developmental series, greenhouse-grown B73 inbred plants were used.  10mm ears were collected and sectioned as follows from tip to base along the developmental gradient: tip 1mm sampled (tip; Inflorescence Meristem/Spikelet Pair Meristem), next 1mm discarded, next 1mm sampled (mid; Spikelet Meristem), next 2mm discarded, next 2 mm sampled (base; Floral Meristem), and immediately frozen in liquid nitrogen.  Sections from ~30 sampled ears were pooled for each of 2 biological replicates to represent tip, mid, and base stages.  Tassels were hand-dissected, measured, separated by stage: 1-2mm (stg1), 3-4mm (stg2), and 5-7mm (stg3), and immediately frozen in liquid N.  For each stage, ~20-30 tassels were pooled for each of 2 biological replicates.  For ramosa mutant series, segregating families (1:1) of ra1-R, ra2-R, and ra3-fea1 mutant alleles, all introgressed at least 6 times into the B73 inbred background, were grown at CSHL Uplands Farm.  Field-grown plants were genotyped and collected 6-7 weeks after germination (V7-V8 stage).  First and second ear primordia were immediately hand-dissected, measured, and frozen in liquid nitrogen.  For ra1, ra2 and ra3 mutants and wild-type controls, ears were pooled into two size classes: 1) 1mm class included a range of 0.7-1.5mm sized ears and nine ears were pooled for each of 2 biological replicates; 2) 2mm class included a range of 1.8-2.5mm sized ears and six ears were pooled for each of three biological replicates.  Wild-type samples were proportional mixtures of heterozygote siblings segregating in ra1, ra2, and ra3 populations.  Variability factors (e.g. ear size within class, ear rank on the plant, and time of collection) were distributed evenly across pooled samples.

first: Alexander

last: Goldshmidt

first: Robert

last: Schmidt


title: Digital gene expression profiling for maize development - 2mm ear libraries, B73 vs. ramosa3 mutant

status: Public on Oct 20 2010

summary: Genome-wide expression signatures detect specific perturbations in developmental programs and contribute to functional resolution of key regulatory networks.  In maize (Zea mays) inflorescences, mutations in the RAMOSA (RA) genes affect determinacy of axillary meristems and thus alter branching patterns, an important agronomic trait.  In this work, we developed and tested a framework for analysis of tag-based, digital gene expression (DGE) profiles using Illumina’s high-throughput sequencing technology and the newly-assembled B73 maize reference genome.  We also used a mutation in the RA3 gene to identify putative expression signatures specific to stem cell fate in determinacy of axillary meristems.  The RA3 gene encodes a trehalose-6-phosphate-phosphatase and may act at the interface between developmental and metabolic processes.  Deep sequencing of DGE libraries, representing three biological replicate ear samples from wild-type and ra3 plants, generated 27 million 20-21nt reads with frequencies spanning four orders of magnitude.  Unique sequence tags were anchored to 3´-ends of individual transcripts by DpnII and NlaIII digests, which were multiplexed during sequencing.  We mapped 86% of non-redundant signature tags to the maize genome, which associated with 37,117 working gene models and un-annotated regions of expression.  66% of maize genes were detected at ≥ nine reads in immature maize ears.  We used comparative genomics to leverage existing information from Arabidopsis and rice in functional analyses of differentially expressed maize genes.  Results from this study provide a basis for analysis of short-read expression data in maize and resolved specific expression signatures that will help define mechanisms of action for the RA3 gene.

overall_design: 3' tag-based DGE libraries were generated using Illumina 1G technology.  Libraries were prepared from 2mm ear samples and represent: 3 biological replicates of wild-type B73, 3 biological replicates of ramosa3 (ra3) mutants, and 1 technical replicate of a ra3 sample.  Each sample was sequenced in a single lane of a flow cell.  Each lane includes library preparations from both DpnII and NlaIII digests for that sample.  Sequences from DpnII digests start with GATC and those from NlaIII digests start with ATG.

first: Namiko

last: Satoh-Nagasawa

first: Sandra

last: Meyer

first: Mary

last: Beatty

first: Hajime

last: Sakai


title: Cross-regulation of amino acid synthesis and anaerobic electron transfer by MetR-mediated methionine signaling

status: Public on Jun 01 2026

city: Hachioji

first: Atsushi

last: Kouzuma

state: Tokyo

address: 1432-1 Horinouchi

country: Japan

institute: Tokyo University of Pharmacy and Life Sciences

summary: Microorganisms living in nutrient- and energy-limited environments must finely coordinate anabolic and catabolic pathways. However, the molecular mechanisms underlying this balance remain poorly understood. Here, we report a mechanism cross-regulating amino acid biosynthesis and anaerobic respiration in bacterial cells. Using the model bacterium Shewanella oneidensis MR-1, we demonstrated that methionine activates extracellular electron transfer activity at submillimolar concentrations. This regulation is mediated by the transcription factor MetR, a canonical regulator of methionine biosynthesis. Under methionine-limited conditions, MR-1 upregulates methionine biosynthesis genes while repressing genes involved in extracellular electron transfer (metal reduction) and other anaerobic respiratory processes. Conversely, methionine availability relieves this repression, enhancing extracellular electron transfer. Furthermore, the MetR-mediated regulation of extracellular electron transfer was observed in Aeromonas hydrophila. These findings reveal a novel physiological role for MetR, suggesting its involvement in the coordinated energy distribution between anabolic and catabolic pathways.

overall_design: The transcriptomic profiles of Shewanella oneidensis MR-1 cultivated under methionine-supplemented and non-supplemented conditions were compared, with three biological replicates prepared for each condition.

first: Erika

last: Yoshino

first: Kazuya

last: Watanabe


title: TET2 affects keratinocytes and psoriatic dermatitis by regulating MECOM and NTRK1 hydroxymethylation

city: Nantong

first: Xin

last: Wang

address: Affiliated Hospital of Nantong University, Medical School of Nantong University, 20 Xisi Road, Nantong 226001,P.R. China

summary: The ten-eleven translocation (TET) family dioxygenases TET2 oxidize 5-methylcytosine (5-mC) to 5-hmC in DNA. DNA hydroxymethylation is the most crucial epigenetic modifications in the occurrence and development of a variety of skin diseases. The goal is to determine the targets of TET2 knockdown including DNA hydroxymethylation in keratinocytes. We profiled a number of important signaling pathways and key factors that may be associated with psoriasis hydroxymethylation and may be potential biomarkers for psoriasis.

overall_design: Hydroxymethylated DNA Immunoprecipitation sequencing of control, TET2-knockdown human immortalized keratinocytes (HaCaT) cells.

first: Chang

last: Liu

first: Sheng

last: Yang

first: Qi

middle: Y

last: Shen

first: Yue

last: Zhu

first: Zi

last: He

first: Shuang

last: Cao


title: Highly multiplexed single cell chromatin regulator and accessibility profiling elucidates chromatin dynamics in human B cell development and oncogenesis, and reveals an adverse prognostic gene expression signature in B lymphoblastic leukemias

city: Palo Alto

first: Reema

last: Baskar

address: 3373 Hillview Ave

institute: Stanford University

summary: Developing B cells undergo successive stages of chromatin reorganization which modulate DNA accessibility to control V(D)J recombination and gene transcription. These epigenetic shifts are mediated by histone modifications (e.g., H3K4me3, H3K27me3), histone-modifying enzymes (e.g., MLL1, LSD1), and chromatin remodelers (e.g., CTCF, DNMT1). We combined highly-multiplexed, multi-omic approaches to capture this choreographed dance – from single cell changes in chromatin modulators, to alterations in histone modifications, to genome-wide chromatin accessibility. This epigenetic map of normal human B cell differentiation enabled us to peer into the early origins of B cell leukemias and lymphomas – specifically, stage-specific chromatin accessibility at sites of recurrent chromosomal translocations and oncogenic transcription factors (TF). It further identified a chromatin state characterized by relative depletion of chromatin structure modulators and high leukemic TF accessibility, enriched in a small cohort of B lymphoblastic leukemias and associated with adverse prognosis.

overall_design: Chromatin accessibility profiling of GM12878 cells after gating for high and low H3K27ac using InTAC-seq protocol


title: Axin1 and Axin2 cooperate to instruct mesodermal fates during mouse gastrulation

city: Cambridge

first: Bertie

last: Gottgens

email: bg200@cam.ac.uk, nkw22@cam.ac.uk

address: Jeffrey Cheah Biomedical Centre

institute: University of Cambridge

summary: How distinct mesodermal lineages - extraembryonic, lateral, intermediate, paraxial and axial – are specified from pluripotent epiblast during gastrulation is a longstanding open question. By investigating Axin, a negative regulator of the WNT/b-catenin pathway, we uncover new roles for WNT signaling in the determination and segregation of extraembryonic, posterior lateral plate, and intermediate mesoderm fates. We undertook two complementary approaches to spatially and temporally dissect the role of WNT signaling at gastrulation. We augmented detailed analysis of Axin1;Axin2 mutant mouse embryos, including single-cell and single-embryo transcriptomics, with in vitro Epiblast-Like Cell (EpiLC) differentiation assays, which allowed us to reveal how coordinated activities of the WNT/b-catenin and BMP/pSmad1/5/9 pathways activate mesoderm lineage-specific programs. Our findings lead us to propose that Axin1 and Axin2 act to prevent aberrant differentiation of epiblast cells into extraembryonic, intermediate, paraxial and axial mesoderm, by spatially and temporally regulating levels of WNT signaling.

overall_design: (15) E6.5 control and (10) E6.5 Axin1DEpi;Axin2-/-  double mutant embryos were single cell dissociated to generate the sc-RNA libraries. Control and mutant embryos were dissected at the same time. Control embryos were obtained by crossing FVB females (Jackson) to the same males used to obtain the Axin1DEpi;Axin2-/-  double mutant embryos (Sox2-Cre+; Axin1DEpi/+;Axin2+/- ).

first: Bart

last: Theeuwes

first: Luke

last: Harland

first: Anna-Katerina

last: Hadjantonakis

first: Berthold


title: Development and Validation of Targetable Prognostic Risk Score in Locally Advanced Nasopharyngeal Carcinoma [Nanostring]

city: Hong Kong

first: Huaping

last: Li

address: 21 Sassoon Rd, Pok Fu Lam

country: Hong Kong

institute: The University of Hong Kong

summary: A novel gene signature for risk stratification of patients with locally advanced NPC suggests potential therapies for high-risk patients.

overall_design: We identified 5 prognostic risk genes (SPP1, IL18BP, PALMD, WDR35, SOCS6) from the whole transcriptomic pattern of 99 advanced NPC FFPE samples by bootstrapping LASSO regression method. A prognostic risk prediction model was developed using Cox regression. Then we validated this prediction model with Nanostring cohort.

first: Chi-leung

last: Chiang

first: Jason Wing Hon

last: Wong


title: Gene expression profile at the single-cell level of Shiga toxin 2+ white cells in the blood

city: Nanjing

first: Xinlei

last: Sun

address: 163 Xianlin Road

institute: Nanjing University

summary: Shiga toxin (Stx)-induced hemolytic uremic syndrome (HUS) poses a grave, life-threatening complication for which a definitive treatment remains elusive. To exert its cytotoxic effect on renal cells, Stx must be delivered from infected intestines to the kidney as the passive toxin dissemination has been excluded. The mechanism underlying Stx delivery, however, remains unclear. Here we pinpoint monocytes as the primary carriers responsible for transporting Stx2 to the renal region. Through single-cell sequencing analysis of peripheral white blood cells that have bound to Stx2 and have been sorted by flow cytometry, we observe that nearly all monocytes exhibit binding to Stx2, whereas less than 10% of neutrophils are associated with Stx2, albeit with a lower affinity.

overall_design: We used the scRNA-seq to test the white blood cell combined with the Stx2,  for the identification of mediator cells delivering stx2.

last: Qu

first: Fujie

last: Shi

first: Ke

last: Zen


title: Highly multiplexed single cell chromatin regulator and accessibility profiling elucidates chromatin dynamics in human B cell development and oncogenesis, and reveals an adverse prognostic gene expression signature in B lymphoblastic leukemias (ATAC-Seq)

overall_design: Chromatin accessibility profiling (ATAC-seq) of BALL cell lines

first: Pablo

last: Domizi

first: Warren

last: Reynolds