title: Expression profile of HER2-overexpressing CHO cells treated with trastuzumab and pertuzumab

city: Edmonton

first: Babak

last: Nami

state: Alberta

address: 8-35 Medical Sciences Building, 114 St.

department: Medical Genetics

institute: University of Alberta

summary: CHO-K1 (wildtype) and CHO-K6 (stablely overexpressing human HER2) cells were terated with 10 ug/ml transtuzumab or pertuzumab and their combination for 24 h and then the whole gene expression was analyzed by cDNA array.

overall_design: CHO-K1 and CHO-K6 were plated in 60 mm dishes in RPMI-1640 medium containing 1% FBS and cultured for 24 h. Afterwards, the old medium discarded and fresh RPMI-1640 supplemented with 10% FBS and containing 10 ug/ml of trastuzumab or pertuzumab or their combination added to the cells and the cell were cultured for 24 h. After the treatment period, the total RNA was isolated using TRIzol reagent (Invitrogen, Waltham, USA) according to standard protocol. Samples for microarray hybridization were prepared according to the Affymetrix Manual Target Preparation for GeneChip® Whole Transcript (WT) Expression Arrays (#902130, Affymetrix Inc., Santa Clara, CA, USA). Amount 100 ng of total RNA were used to make double-stranded cDNA following cRNA synthesis. After purification of cRNA, 15 ug of each cRNA was reverse transcribed into sense-strand (ss) cDNA when unnatural dUTP residues were incorporated. ss cDNA were purified and 5.5 ug of each ss cDNA was fragmented using uracil DNA glycosylase (UDG) and apurinic/apyrimidinic endonuclease 1 (APE 1) at the unnatural dUTP which breaks the DNA strand. Fragmented ss cDNA were terminal labeled with biotin and 3ug of each sample was hybridized to Affymetrix GeneChip® CHO Gene 2.0 ST array (Format 100) for 16 hours at 45C with rotation at 60 rpm in an Affymetrix GeneChip® Hybridization Oven 645. After hybridization, the arrays were washed and stained in an Affymetrix GeneChip® Fluidics Station FS450 and the fluorescent signals were measured with an Affymetrix GeneChip® Scanner 3000 7G. Row data was analyzed by Affymetrix Transcriptome Analysis Console (TAC) 3.0 software (Affymetrix Inc., Santa Clara, CA, USA)

first: Hamid

last: Maadi

first: Zhixiang

last: Wang


title: High-fat diet induces the modulation of hepatic nuclear receptors and their target genes in C57BL/6 mice

status: Public on Aug 28 2018

city: Bloomington

first: Xilin

last: Li

state: Indiana

address: 2719 E 10th street Rm230

country: USA

institute: Indiana University

type: Expression profiling by high throughput sequencing

summary: We reported the hepatic gene expression profiling in mice fed with a high fat diet compared to the controls. We found that the modulation of pathways related to peroxisome proliferator-activated receptors, cytochrome P450s, and ATP-binding cassette transporters in high fat diet mice. Particularly, constitutive androstane receptor and pregnane X receptor signature genes Cyp2b10 and Cyp3a11 were significantly increased in high fat diet mice compared to controls. Liver X receptor mediated Abcg5 and Abcg8 were also found increased by HFD treatment.

overall_design: Examing the liver transcriptome of C57BL/6 mice fed by a high fat diet and a low-fat control diet using deep sequencing, with 4 biological replicates in each group

first: Douglas

last: Rusch

first: Zemin

first: James

middle: E

last: Klaunig


title: Quantitative Analysis of Wild Type,Ctnnb1 deltaEx3 and Ctnnb1 deltaEx2-6 Spinal Cord Transcriptomes

city: Hangzhou

first: Zhong-Min

last: Dai

state: Zhejiang

address: 16 Xuelin Street, Xiasha

country: China

institute: Hangzhou Normal University

summary: The goals of this study are to analyze NGS-derived spinal cord transcriptome profiling (RNA-seq) of wild type (WT), Nestin-Cre mediated Ctnnb1 Ex3 deletion (gain of function, dE3) and Ex2-6 deletion (loss of function, cKO) at embryonic day 13.5 (E13.5). Spinal cord RNA profiles were generated by deep sequencing, using Illumina Hiseq. The sequence reads that passed quality filters were analyzed at the transcript isoform level with HISAT2 and RSeQC.

overall_design: Spinal cord RNA profiles of E13.5 wild type (WT), Ctnnb1-dE3 and Ctnnb1-cKO mice were generated by deep sequencing, using Illumina Hiseq 2500.

first: Shuhui

last: Sun


title: Perturbed maintenance of transcriptional repression on the inactive X-chromosome in the mouse brain after XIST deletion

city: Seattle

first: Eric

middle: Jan

last: Foss

state: WA

address: 1100 Fairview Ave N

department: Clinical Research Division

institute: Fred Hutchinson Cancer Research Center

type: Methylation profiling by high throughput sequencing

summary: The long noncoding RNA, Xist, is critical for initiation and establishment of X- chromosome inactivation during embryogenesis in mammals but it is unclear whether its continued expression is required for maintaining X inactivation in vivo.  By using an inactive X chromosome-linked MeCP2-GFP reporter, which allowed us to enumerate reactivation events in the mouse brain even when they occur in very few cells, we found that deletion of Xist in the brain after establishment of X chromosome inactivation leads to reactivation in 2-5% of neurons and in a smaller fraction of astrocytes. In contrast to global loss of both H3 lysine 27 tri-methylation (H3K27m3) and histone H2A lysine 119 monoubiquitylation (H2AK119ub1) we observed upon Xist deletion, alterations in CpG methylation were subtle, and this was mirrored by only minor alterations in X- chromosome-wide gene expression levels.  Our results demonstrate that Xist is required for maintaining tight transcriptional repression on the inactive X-chromosome and that partial loss of X-dosage compensation in the absence of Xist in the brain is well tolerated.

overall_design: high throughput sequencing-based assessment of mRNA and DNA methylation levels in mice deleted for XIST after transcriptional silencing on the X chromosome has been established; 5 RNA seq samples and 3 LpnPI DNA samples from mice deleted for XIST; 3 RNA seq and 2 LpnPI DNA samples from control mice with intact XIST

first: Robin

middle: L

last: Adrianse

first: Kaleb

last: Smith

first: Tonibelle

last: Gatbonton-Schwager

first: Smitha

middle: P

last: Sripathy

first: Uyen

last: Lao

middle: J

first: Ruben

middle: G

last: Boers

first: Joachim

middle: B

first: Joost

last: Gribnau

first: Antonio

last: Bedalov


title: RNA-sequencing of primary intestinal epithelial cells from wild-type mice

city: Berkeley

first: Gregory

last: Barton

state: CA

department: Molecular and Cell Biology

institute: UC Berkeley

summary: This dataset consists of RNA-sequencing of isolated mouse intestinal epithelial cells. Intestinal epithelial cells were isolated from C57BL/6 wild-type mice in our SPF facility at  4 weeks and 8 weeks of age. The goal of this study was to understand the patterns of gene expression in these cells under homeostatic conditions.

overall_design: Intestinal epithelial cells were isolated from the small intestines and colons of wild-type mice and RNA was prepared for RNA-sequencing. For each tissue, IECs were isolated from 2 mice aged 4 weeks and 2 mice aged 8 weeks.

first: Jacques

last: Deguine

first: April

last: Price


title: Discovery of a Pro-Tumoral Clonogenic Unipotent Neutrophil Progenitor in Mouse and Human Bone Marrow

city: La Jolla

first: Catherine

last: Hedrick

state: California

address: 9420 Athena Circle

department: Division of Inflammation Biology

institute: La Jolla Institute for Allergy and Immunology

summary: Neutrophils are short-lived immune cells that play important roles in a variety of diseases. The oligopotent Granulocyte Monocyte Progenitors (GMP) in the bone marrow give rise to monocytes and all granulocytes. Although several monocyte progenitors have been identified in mouse bone marrow, the unipotent neutrophil progenitors are still not well-defined. Here, we use Cytometry by Time-of-Flight (CyTOF) and Single-cell RNA-Sequencing (scRNA-Seq) methodologies to identify a committed unipotent early-stage neutrophil progenitor in adult mouse bone marrow. Importantly, we also discovered a similar unipotent, committed neutrophil progenitor (hNeP) that is present in healthy human bone marrow. Both mouse and human progenitors demonstrate unipotent neutrophil potency in vivo. Study of the identified mouse (NeP) and human (hNeP) neutrophil progenitors in cancer revealed that both NeP and hNeP significantly increased tumor growth when transferred into murine cancer models, including a humanized model.  Further, we discovered that the hNeP was present in the blood of human patients recently diagnosed with melanoma, and could be readily identified by flow cytometry, suggesting that this human neutrophil progenitor could be used as a biomarker for early cancer discovery.  The discovery of this early committed unipotent neutrophil progenitor in humans will allow for development of important new therapeutic targets for regulation of neutrophil levels and function in disease, particularly in cancers, where neutrophils play a significant role.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Neutrophil progenitor isolated from mouse and human fresh bone marrow without treatment were sequeced to explore heterogeneities; 3' Transcriptome sequencing (10X genomics)

first: Yanfang

last: Zhu

first: Lindsey

last: Padgett

first: Huy

last: Dinh

first: Paola

last: Marcovecchio

first: Amy

last: Blatchley

first: Runpei

last: Wu

first: Erik

last: Ehinger

first: Cheryl

last: Kim

first: Zbigniew

last: Mikulski

last: Seumois

first: Ariel

last: Madrigal

first: Pandurangan

last: Vijayanand

middle: C


title: To investigate the decay constants (half-lives) of transcript isoforms generated by alternative polyadenylation in proliferating and quiescent cells

city: Los Angeles

first: Hilary

last: Coller

address: 610 Charles E Young Drive E

department: Molecular, Cell and Developmental Biology

institute: University of California, Los Angeles

summary:
Purpose: Alternative polyadenylation (APA) can result in the generation of transcripts that terminate at different locations within the gene, which can result in different 3' UTRs.   3´ UTRs are known to contain RNA stabilizing and destabilizing motifs that provide a platform for binding of  RNA binding proteins. Changes in 3´ UTRs for the different APA-generated isoforms of a gene could allow for inclusion or exclusion of these RNA stability elements. The goal of this study is determine the stability (half-lives based on transcript decay) of APA isoforms in proliferating and quiescent cells.
Methods: The proliferating and quiescent (7-day contact inhibited) cells were treated with actinomycin D to stop transcription and the cells were harvested at different time points (0. 2, 4, 8 hours) for RNA isolation and global sequencing of 3´ ends. The adaptors and polyA sequenced were trimmed from each read.  The reads were aligned to the human genome (hg19) using TopHat (v2.0.14). The aligned read files were used to determine the counts of APA isoforms of each gene using the code provided by Gruber et al. (PMID: 27382025). The counts of the APA isoforms were fit to an exponential decay model to obtain half-lives. The entire workflow was performed for two biological replicates: 12-1 and 12-3 strains of human dermal fibroblasts.
Results:  In two different fibroblast strains (12-1 and 12-3), we found that isoforms terminating at distal polyadenylation sites were more stable than isoforms terminating at proximal polyadenylation sites in quiescent, but not proliferating, fibroblasts.
Conclusions: The transition from shorter to longer isoforms in quiescent cells is associated with stabilization of the transcripts.

first: Mithun

last: Mitra

first: Lois

last: Nersesian

first: Wei

last: Wang

first: Jessica

last: Buckles

middle: A


title: Differential gene expression analysis between proliferating and quiescent human dermal fibroblasts

summary:
Purpose: Quiescence is a state or reversible cell cycle arrest. A previous study using microarrays (Coller et al., PMID: 16509772) revealed gene expression changes between quiescent and proliferating human dermal fibroblasts and defined a "quiescece program" of genes that change in abundance when fibroblasts were introduced into quiescence by one of three methods. The goal of this study is to perform high throughut RNA sequencing to determine global changes in gene expression between proliferating and quiescent human dermal fibroblasts.
Methods: Three different biological replicates (corresponding to two fibroblast strains, 10-5 and 12-1) were collected in proliferating and contact--inhibited (quiescent) conditions. RNA extracted from these cells were used for library preparation. The libraries were sequenced on an Illumina HiSeq 2000 instrument.
Results: Among the 19,673 genes monitored, 1,993 genes (10.1%) changed in expression two-fold or more, demonstrating widespread changes in gene expression with contact inhibition-induced quiescence. Fifty-two percent of these genes were upregulated in 7dCI compared with proliferating fibroblasts, and 48% were downregulated in 7dCI fibroblasts.
Conclusions: There are widespread changes in gene expression as fibroblasts transition between proliferating and quiescent states.

first: Elizabeth

last: Johnson


title: Asymmetric partition of parental histone (H3-H4)2 tetramers onto replicating DNA strands

city: New York

first: zhiguo

last: zhang

state: NY

address: 1130 St. Nicholas Avenue

department: Pediatric and Genetics and Development

institute: Columbia University

type: Genome binding/occupancy profiling by high throughput sequencing

summary: In eukaryotic cells, inheritable changes in gene expression in response to environmental and developmental stimuli is associated with changes in histone modifications and relies on the passage of these changes into daughter cells during cell division, a process that remains elusive. Here, we show that parental histone (H3-H4)2 tetramers, the primary carrier of epigenetic modifications, are assembled into nucleosomes onto both replicating leading and lagging strands, with a preference for lagging strands of DNA replication forks. This asymmetric distribution of parental (H3-H4)2 is exacerbated in cells lacking Dpb3 and Dpb4, two subunits of DNA polymerase Pol ε. Dpb3-Dpb4 binds (H3-H4)2 and participates in the transfer of parental (H3-H4)2 tetramers onto leading strands of DNA replication forks. Cells lacking Dpb3 and Dpb4 exhibits defects in epigenetic inheritance. These results reveal a previously undocumented mechanism of histone segregation and a direct role for Pol ε in this poorly understood process.

overall_design: We synchronized yeast cells (Wile type and other mutant cells) at G1 and released into early S phase in the presence of BrdU, and hydroxyurea (HU). We then performed BrdU immunoprecipitation using anti-BrdU antibodies following single-strand DNA library preparation and sequencing (ssSeq). we also performed protein ChIP followed by single-strand DNA sequencing (ChIP-ssSeq) for Dpb3, Dpb4, H3K56ac, H3K4me3, T7-tag, and HA-tag. The sequencing tag was mapped to both Watson (red) and Crick (blue) strands of the reference genome.

first: Chuanhe

last: Yu

first: Haiyun

last: Gan

first: Albert

last: Cardona Serra

first: Zhiguo

last: Zhang


title: Genome wide analysis of SAHA (vorinostat) treatment in primary mouse cortical neurons upon miR-203 overexpression

city: Irvine

first: Vivek

last: Swarup

state: CALIFORNIA

address: 3400 Bio Sci 3

department: Neurobiology and Behavior

institute: University of California Irvine

summary: Purpose: The goal of this study was to assess gene expression changes upon SAHA treatment in cells overexpressing miR-203.

overall_design: Primary cortical cultures were established using E15 cortical cultures from C57BL/6J mice. At DIV0, cells were infectd with either miR-203 (high titre - 1MOI) or intermediate titre (0.5MOI) or control (high 1MOI) lentivirus andalso treated with different dose of SAHA. At DIV8, total RNA was isolated using NucleoSpin RNA XS kit (Takara). Libraries were prepared using Standard illumina stranded mRNA-seq protocol.

first: Daniel

middle: H

last: Geschwind


title: RNA-sequencing from flagellin-stimulated intestinal organoids

first: Gregory

last: Barton

summary: This dataset consists of RNA-sequencing of intestinal organoids at rest and after stimulation with the TLR5 ligand flagellin for 4 hours. The goal of this study was to understand the differential patterns of gene expression induced upon stimulation with a TLR ligand in small intestine organoids, colon organoids, and organoids that had been skewed to generate a higher proportion of Paneth cells.

overall_design: Analysis was performed on RNA isolated from 3 separate experimental replicates of both unstimulated and flagellin-stimulated SI organoids, 4 separate experimental replicates of unstimulated and flagellin-stimulated colon organoids, and 6 separate experimental replicates of unstimulated and flagellin-stimulated Paneth cell-skewed organoids

first: Jacques

last: Deguine

first: April

last: Price


title: Dissection of memory response B cell populations by single cell RNA-seq

city: Darlinghurst

first: Tri

middle: Giang

last: Phan

state: New South Wales

address: 384 Victoria St

department: Immunology Division

country: Australia

institute: Garvan Institute

summary: SWHEL B cells were adoptively transferred and recipient mice were immunised with hen egg lysozyme (HEL) to establish anti-HEL memory B cells. Immune mice were then re-immunised with HEL and single donor-derived antigen-specific B cells sorted for RNA sequencing on day 5 of the secondary antibody response.

overall_design: Single-cell resolution transcriptional profiles of 144 memory response (Day 5 post-recall) SWHEL-Tomato B cells from three mice. Single-cell samples were isolated by FACS and processed in 9 batches. Samples were split into two pools of multiplexed samples. Each pool was run on a single Illumina HiSeq 2500 lane, averaging 3 million reads per cell.

first: Akira

last: Nguyen

first: Imogen

last: Moran


title: Effect of methione restriction in the liver of WT and Lmna G609G KI mice

status: Public on Aug 27 2018

city: Oviedo

first: Jose

last: Freije

state: Asturias

address: C/ Fernando Bongera s/n

department: Bioquimica y Biologia Molecular

country: Spain

institute: Universidad de Oviedo

summary: Dietary intervention constitutes a feasible approach for modulating metabolism and improving healthspan and lifespan. Methionine restriction (MR) delays the appearance of age-related diseases and increases longevity in normal mice. However, the effect of MR on premature aging remains to be elucidated. Here, we describe that MR extends lifespan in two different mouse models of Hutchinson-Gilford progeria syndrome (HGPS) by reversing the transcriptome alterations in inflammation and DNA-damage response genes present in this condition. Further, MR improves the lipid profile and alters the levels of bile acids, both in wild-type and in progeroid mice. Notably, treatment with the bile acid cholic acid improves healthspan and lifespan in vivo. These results suggest the existence of a metabolic pathway involved in the longevity extension achieved by MR and support the possibility of dietary interventions for treating progeria.

overall_design: 12 liver samples (4 groups with 3 replicates per group) extracted upon 6 hours of fasting

first: C

last: Barcena

first: P

middle: M

last: Quiros

first: J


title: The NORAD lncRNA assembles a topoisomerase complex critical for genome stability

city: Cambridge

first: Mathias

last: Munschauer

state: MA

address: 415 Main Street

institute: The Broad Institute of MIT and Harvard

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

overall_design: Refer to individual Series


title: Comprehensively profiling the chromatin architecture of tissue restricted antigen expression in thymic epithelial cells

city: Oxford

first: Adam

last: Handel

address: John Radcliffe Hospital

department: Weatherall Institute of Molecular Medicine

country: United Kingdom

institute: University of Oxford

summary: Thymic epithelial cells (TEC) effect crucial roles in thymopoiesis including the control of negative thymocyte selection. This process depends on their capacity to express promiscuously genes encoding tissue-restricted antigens. This competence is accomplished in medullary TEC (mTEC) in part by the presence of the transcriptional facilitator AutoImmune REgulator, AIRE. AIRE-regulated gene transcription is marked by repressive chromatin modifications, including H3K27me3.  When during TEC development these chromatin marks are established, however, remains unclear. Here we use a comprehensive ChIP-seq dataset of multiple chromatin modifications in different TEC subtypes to demonstrate that the chromatin landscape is established early in TEC differentiation. Much of the chromatin architecture found in mature mTEC was found to be present already over earlier stages of mTEC lineage differentiation. This was reflected by the fact that a machine learning approach accurately classified genes as AIRE-induced or AIRE-independent both in immature and mature mTEC. Moreover, analysis of TEC specific enhancer elements identified candidate transcription factors likely to be important in mTEC development and function. Our findings indicate that the mature mTEC chromatin landscape is laid down early in mTEC differentiation, and that AIRE is not required for large-scale re-patterning of chromatin in mTEC.

overall_design: Single cell RNA-seq, ATAC-seq and ChIP-seq on thymic epithelial cell subsets (cTEC, mTEClo and mTEChi) from wild-type mice or from FOXN1flag homozygous mice (see Žuklys S et al. Nature Immunology 2016:17(10):1206-1215 for details)

first: Georg

last: Holländer


title: IL-2/CD25: A Long-Acting Fusion Protein That Promotes Immune Tolerance by Selectively Targeting the IL-2 Receptor on Regulatory T Cells

city: Miami

first: Thomas

middle: R

last: Malek

state: FL

address: 1600 NW 10th Ave, RMSB, Room 3119

department: Microbiology and Immunology

institute: University of Miami

summary: Low-dose IL-2 represents an immunotherapy to selectively expand regulatory T cells (Tregs) to promote tolerance in patients with autoimmunity. In this article, we show that a fusion protein (FP) of mouse IL-2 and mouse IL-2Rα (CD25), joined by a noncleavable linker, has greater in vivo efficacy than rIL-2 at Treg expansion and control of autoimmunity. Biochemical and functional studies support a model in which IL-2 interacts with CD25 in the context of this FP in trans to form inactive head-to-tail dimers that slowly dissociate into an active monomer. In vitro, IL-2/CD25 has low sp. act. However, in vivo IL-2/CD25 is long lived to persistently and selectively stimulate Tregs. In female NOD mice, IL-2/CD25 administration increased Tregs within the pancreas and reduced the instance of spontaneous diabetes. Thus, IL-2/CD25 represents a distinct class of IL-2 FPs with the potential for clinical development for use in autoimmunity or other disorders of an overactive immune response.

overall_design: Splenic murine Tregs mRNA profiles of IL-2/CD25 (FP) or control PBS injected mice were generated 72 hrs post injection by deep sequencing, in quadruplicate (FP) or triplicate (PBS), using NextSeq 500 with a High Output Kit 150-cycle flow cell (Illumina). Reads from RNA-seq were mapped to the Mus musculus genome GRCm38 using STAR (ver.2.5.0) aligner 44. Raw counts were generated on Ensembl gene (GENCODE M13) with featureCounts (ver.1.5.0) 45. Differential expressed genes in Tregs between the IL2/CD25 and PBS injected mice were identified using DESeq2 46, and determined according to threshold of false discovery rate (FDR) <0.05

first: Natasha

last: Ward

first: Aixin

last: Yu

first: Alejandro

last: Moro

first: Yuguang

last: Ban

first: Xi

last: Chen

first: Sunnie

last: Hsiung

first: James

last: Keegan

first: Jaren

last: Arbanas

first: Martine

last: Loubeau

first: Anil

last: Thankappan

first: Aaron

last: Yamniuk

first: Jonathan

last: Davis

first: Mary

last: Struthers


title: The NORAD lncRNA assembles a topoisomerase complex critical for genome stability [RNA-seq]

first: Mathias

last: Munschauer

summary: Thousands of long non-coding RNAs (lncRNAs) have been identified in the human genome, but specific biological functions and biochemical mechanisms have been discovered for only about a dozen lncRNAs. One specific lncRNA, Non-coding RNA Activated by DNA Damage (NORAD), has recently been shown by genetic deletion to be required for maintaining genomic stability, but its molecular mechanism is unknown. Here, we combine RNA antisense purification (RAP) and quantitative mass spectrometry to identify proteins that directly interact with NORAD in living cells. We show that NORAD interacts with proteins involved in DNA replication and repair in steady-state cells and localizes to the nucleus upon stimulation with replication stress or DNA damage. In particular, NORAD interacts with RBMX (an emerging component of the DNA-damage response) and encodes the strongest RBMX-binding site in the transcriptome. We demonstrate that NORAD controls the ability of RBMX to assemble a ribonucleoprotein complex, which we term NORAD-Activated Ribonucleoprotein Complex 1 (NARC1), containing known suppressors of genomic instability: topoisomerase I (TOP1), ALYREF and the PRPF19/CDC5L complex. Cells depleted of NORAD or RBMX display an increased frequency of chromosome segregation errors, reduced replication-fork velocity and altered cell cycle progression phenotypes that are mechanistically linked to TOP1 and PRPF19/CDC5L function. Expression of NORAD in trans can rescue defects caused by NORAD depletion, but rescue is significantly impaired when the RBMX-binding site in NORAD is deleted. Our results demonstrate that the interaction between NORAD and RBMX is important for NORAD function and that NORAD is required for the assembly of a previously unknown topoisomerase complex (NARC1) that contributes to maintaining genomic stability. Moreover, we uncover a novel function for lncRNAs in modulating the ability of an RNA-binding protein to assemble a higher-order ribonucleoprotein complex.

overall_design: We examined gene expression changes and alternative splicing events in wildtype and NORAD depleted cells using RNA sequencing.

first: Eric

middle: S

last: Lander


title: The NORAD lncRNA assembles a topoisomerase complex critical for genome stability [CLIP-seq]

overall_design: We mapped direct protein-RNA interaction sites using crosslinking and immunoprecipitation (CLIP) for various RNA binding proteins and generated size-matched input libraries to estimate the average protein occupancy of background proteins migration at the same molecular weight range as the target protein.


title: Survey of Human Fetal Kidney Development Through Single Cell RNA Sequencing Analysis

city: BeiJing

first: Yidong

address: Yiheyuan Road

institute: Peking University

summary: Human kidney is a major metabolic organ, which plays a crucial role in regulating homeostasis of human body. However, gene expression characterizations of human fetal kidney development is still not fully explored. Here, we used single-cell RNA-seq to analyze more than 3,000 individual renal cells of human fetal kidneys covering over four months of development in vivo.We found co-expression of self-renewal and differentiation genes in cap mesenchyme, progenitors differed from that of mouse as they persistently express SIX1 throughout the renal morphogenesis. Besides, we recapitulated the transcription factors and signaling pathways potentially important for segmentation of nephron tubule. Furthermore, we explored the human specific features of the heterogeneous collecting duct epithelium. We also dissected the metabolism signatures of fetal kidney as well as the extracellular matrix composition of glomerulus mesangium. Eventually, we identified novel markers for renal tubules. Our study highlights the gene expression features of human fetal kidney development, which will contribute to dissecting the mechanisms of renal dysplasia and congenital nephrotic disease.

overall_design: Here we sequenced 3,543 single cells and 3,023 of them passed the screening criteria, which we set the expressed gene number and transcripts in each cell should respectively above 1000 and 5000.And then we constructed the gene expression landscape throughput kidney organogenesis.

first: F

last: Tang

first: Y

first: P

last: Wang


title: sgRNA cassette sequencing from H226 cells with doxycycline-inducible p63-targeting shRNA construct, treated with doxycycline for 14 days

city: Aurora

first: Matthew

middle: D

last: Galbraith

state: CO

department: Department of Pharmacology; Linda Crnic Institute for Down Syndrome

institute: University of Colorado Anschutz Medical Campus

summary: Genome-wide CRISPR/Cas9-based screen to identify anti-proliferative genes during ΔNp63α depletion

overall_design: H226 cells transduced with the GeCKOv2 A and B libraries were treated to induce p63 knockdown for 14 days to screen for genes affecting proliferation upon depletion of p63.

first: Christopher

last: Abraham

first: Ahwan

last: Pandey

first: Kelly

last: Sullivan

first: Joaquín

last: Espinosa


title: sgRNA cassette sequencing and RNA-seq from H226 cells expressing doxycycline-inducible Control (non-targeting) and p63-targeting shRNAs


title: Global proteome remodeling during ER stress involves Hac1-driven expression of long undecoded transcript isoforms

status: Public on Aug 26 2018

first: Gloria

last: Brar

address: 626 Barker Hall

department: MCB

summary: Cellular stress response pathways often require transcription-based activation of gene expression to promote cellular adaptation. However, whether general mechanisms exist for stress-responsive gene down-regulation is less clear. A recently defined gene regulatory mechanism enables both up- and down-regulation of protein levels for distinct gene sets by the same transcription factor (TF) via coordinated induction of canonical mRNAs and long undecoded transcript isoforms (LUTIs). We analyzed deep, parallel gene expression datasets to determine whether this mechanism contributes to the conserved Hac1-driven branch of the unfolded protein response (UPRER). Indeed, we found Hac1-dependent protein down-regulation that accompanied the well-characterized up-regulation of ER-related proteins that typifies UPRER activation. Proteins down-regulated by Hac1-driven LUTIs include those with electron transport chain (ETC) function.  Aerobic respiration also appears dampened during the UPRER, and abrogated ETC function improves the fitness of UPRER-activated cells, suggesting functional importance of LUTI regulation during the UPRER. We conclude that the UPRER involves large-scale proteome remodeling, mediated in part by Hac1-induced LUTIs, and that this mechanism enables coordination of up- and down-regulation of gene expression during this stress response.

overall_design: Sequencing of mRNA molecules and ribosome footprints to identify regulated transcript isoform changes that result in apparent translational regulation. In short, two major experiments are included here, one for parallel mRNA-seq and ribosome profiling of cells with or without UPR induction by either DTT or Tm, with or without deletion of HAC1. The other experiment investigates the examines the same type of measurements for cells that are either WT or depleted for Hac1 and treated with DTT.

first: Kelsey

last: Van Dalfsen


title: Pitx2 maintains mitochondrial function during regeneration to prevent myocardial fat deposition

last: Hill

address: 75 Aimes Street

institute: Broad Institute of MIT and Harvard

summary: Here, we performed single nuclear RNA-seq (snRNA-seq) of control and Pitx2 deficient cardiac tissue 3 weeks post myocardial infarction. Next, unsupervised graph-based clustering of the combined snRNA-Seq data set mapped to both introns and exons, comprising 7848 cells. Overall, we identified nine transcriptionally distinct clusters representing all the major cardiac cell types, including cardiac fibroblasts (FB), cardiomyocytes (CM), endothelial cells (EC), vascular smooth muscle cells (SMC), macrophages (Mφ), epicardial cells (EpiC), endocardial cells (EndoC), lymphatic endothelial cells (LEC), and mural cells or pericytes (PeC). Moreover, two distinct populations of fibroblasts, designated FB-1 and FB-2, were also identified.

overall_design: Cardiac tissue dissociated, nuclei were isolated via density gradient centrifugation, and then ran through the 10X Chromium device to generate snRNA-seq libraries that were sequenced on an Illumina NextSeq 500.

first: Lele

last: Li

first: James

middle: F

last: Martin


title: Study of mitotic chromatin shows involvement of histone modifications in bookmarking and reveals nucleosome deposition patterns

city: Jerusalem

first: Itamar

last: Simon

state: APO/FPO IRAQ / AFGHANISTAN / MIDDLE EAST

address: Hadassah Ein Kerem

department: Microbiology and Molecular Genetics

country: Israel

institute: Hebrew University medical school

summary: Here we compared the histone modification landscapes of mitotic and interphase cells to understand how histone modifications play a role in mitotic bookmarking. We found that the localization of histone modifications is maintained during mitosis, with methylation levels remaining the same, and acetylation levels decreasing. We found that at nucleosome depleted regions (NDRs), there is a nucleosome present during mitosis comprising a distinctive set of histone modifications, revealing a mitotic deposition and deacetylation of nucleosomes at NDRs.

overall_design: Examination of RNA pol II and six histone modifications during mitosis and interphase.

first: Inbal

last: Shamir

first: Shashi

last: Gandhi

first: Elisheva

last: Javasky

first: Alon

last: Goren


title: Neural progenitors derived from Tuberous Sclerosis Complex patients exhibit attenuated PI3K/AKT signaling and delayed neuronal differentiation

city: Piscataway

first: Ronald

middle: P.

last: Hart

state: NJ

address: 604 Allison Rd Rm B430

department: Cell Biology & Neuroscience

institute: Rutgers University

summary: Tuberous Sclerosis Complex (TSC) is a disease caused by autosomal dominant mutations in the TSC1 or TSC2 genes, and is characterized by tumor susceptibility, brain lesions, seizures and behavioral impairments. The TSC1 and TSC2 genes encode proteins forming a complex (TSC), which is a major regulator and suppressor of mammalian target of rapamycin (mTOR) in complex 1 (mTORC1), a signaling complex that promotes cell growth and proliferation. TSC1/2 loss of heterozygosity (LOH) and the subsequent complete loss of TSC regulatory activity in null cells causes mTORC1 dysregulation and TSC-associated brain lesions or other tissue tumors. However, it is not clear whether TSC1/2 heterozygous brain cells are abnormal and contribute to TSC neuropathology. To investigate this issue, we generated induced pluripotent stem cells (iPSCs) from TSC patients and unaffected controls, and utilized these to obtain neural progenitor cells (NPCs) and differentiated neurons in vitro. These patient-derived TSC2 heterozygous NPCs were delayed in their ability to differentiate into neurons. Patient-derived progenitor cells also exhibited a modest activation of mTORC1 signaling downstream of TSC, and a marked attenuation of upstream PI3K/AKT signaling. We further show that pharmacologic AKT inhibition, but not mTORC1 inhibition, causes a neuronal differentiation delay, mimicking the patient phenotype. Together these data suggest that heterozygous TSC2 mutations disrupt neuronal development, potentially contributing to the disease neuropathology, and that this defect may result from dysregulated AKT signaling in neural progenitor cells.

overall_design: Two replicates each of TSC#1 and CON#1 NPC cell RNA were prepared for sequencing library preparation and seqeuencing.

first: Avery

last: Zucco

first: Valentina

last: Dal Pozzo

first: Alina

last: Afinogenova

first: Orrin

last: Devinsky

first: Gabriella

last: D'Arcangelo


title: Poly(A)+ RNA-seq from H226 cells expressing doxycycline-inducible Control (non-targeting) and p63-targeting shRNAs

first: Matthew

last: Galbraith

summary: To determine the impact of ΔNp63α knockdown on steady-state mRNA levels, we performed poly(A)-enriched RNA-seq analysis of lung squamous cell carcinoma line H226 (inducible shControl and shp63) in the presence of 1µg/mL doxycycline to induce shRNA expression.

overall_design: Poly(A)+ RNA for two independent biological replicates was purified  from H226 cells (inducible shControl and shp63) incubated treated for six days with 1 µg/mL doxycycline. a TruSeq Stranded mRNA Library Prep Kit (Illumina). Libraries were sequenced on an Illumina HiSeq 2000 system at the University of Colorado Cancer Center Genomics and Microarray Core facility. Reads were aligned (TopHat2) to the Human reference genome (GRCh37/hg19) and gene-level counts (HTseq-count) were used for differential expression analysis (DESeq2).

first: Christopher

last: Abraham

first: Ahwan

last: Pandey

first: Joaquin

last: Espinosa


title: Expression data from mouse sarcoma tumor cell lines

city: marseille

first: Thien-Phong

last: Vu Manh

address: 163 avenue de Luminy

country: France

institute: CIML

summary:
Vanin1, a regulator of vitamin B5 metabolism, is expressed by sarcoma tumors. We evaluated its impact on sarcoma growth by using sarcoma cell lines derived from p16p19Vnn1-deficient mice and further transduced with an oncogenic RasV12 oncogene (R tumors) in the presence or not of a catalytically active (VR tumors) or mutated (VdR tumors) Vnn1 isoform.
We used microarrays to detail the global programme of gene expression associated with the growth potential of various tumor cell lines grafted in Nude mice

overall_design: Tumor cell lines were injected in Nude mice under normal diet (nd) and harvested between 2 and 3 weeks of growth. After RNA extraction, analysis of the transcriptome was perfomed using an Affymetrix platform at the Strasbourg

first: Caroline

last: Giessner

first: Virginie

last: Millet

first: Philippe

last: Naquet


title: Effect for target mRNA degradation by methylated microRNAs

status: Public on Aug 25 2018

city: Suita

first: Ayumu

last: Asai

address: 2-2 Yamadaoka

country: Japan

institute: Osaka University

summary: To confirme the function of methylated micro RNAs (miR-17 and let-7a), we transfected methylated or non-methylated microRNAs to DICER KO HCT116 cell line. mRNA were isolated at 48 houes after transfection. This data show that the function were different between methylated or non-methylated microRNAs.

overall_design: Methyated or non-methyated microRNAs were transfected in dicer KO HCT116 cell lines.

first: Masamitsu

last: Konno

first: Jun

last: Koseki

first: Hideshi

last: Ishii


title: Genome wide analysis of SAHA (vorinostat) treatment in human iPSC-derived neurons from Tau A152T patients and controls

first: Vivek

last: Swarup

summary: Purpose: The goal of this study was to assess gene expression changes upon SAHA treatment in neurons derived from patients with A152T Tau mutation

overall_design: iPSC derived neurons were treated with SAHA at different dosage for several days

first: Daniel

last: Geschwind


title: RNA-Seq Analysis of Spinal Cord Tissues from hPFN1G118V Transgenic Mouse Model of ALS at Presymptomatic and End Stages of Disease

city: Little Rock

first: Kiaei

last: Mahmoud

state: AR

address: 4301 W. Markham St., Slot 611 (BioMed 1, Rm B-306A)

institute: University of Arkansas for Medical Sciences

summary: Amyotrophic Lateral Sclerosis (ALS) is a fatal neurodegenerative disease that leads to the loss of motor neurons. The molecular mechanisms of motor neuron degeneration are largely unknown and there are currently no effective therapies to treat this disease. Cases of familial ALS have been linked to mutations in the profilin1 gene (PFN1) and here we utilize the in vivo mouse model of PFN1-related motor neuron disease, hPFN1G118V. We conducted whole transcriptome profiling of spinal cords of mutant transgenic hPFN1G118V mice and their wildtype transgenic hPFN1WT controls at 50 days, a presymptomatic stage and the earliest known time point at which aggregation of PFN1G118V occurs, and at end stage of disease, beginning around 180 days. The overall transcriptome profiles of spinal cord were highly similar while end stage hPFN1G118V mice had gene expression that clustered away from hPFN1WT and presymptomatic hPFN1G118V mice. Differential expression analysis revealed that end stage hPFN1G118V mice had 890 differentially expressed genes (747 up and 143 down) when compared to pre-symptomatic hPFN1G118V mice, and they had 836 differentially expressed genes (742 up and 94 down) when compared to their age-matched hPFN1WT controls. 50 day old hPFN1G118V mice were not significantly different from their age matched hPFN1WT controls. This is the first study that has utilized next generation RNA-sequencing to measure gene expression in pre-symptomatic and end-stage hPFN1G118V mice, and may lead to identification of molecular features that could become therapeutic targets for ALS.

overall_design: RNA-sequencing of spinal cords of transgenic hPFN1G118V mice and transgenic hPFN1WT mice at presymptomatic and end stage of disease

last: Barham

last: Fil

first: Yasir

last: Rahmatallah

first: Galina

last: Glazko

first: Mahmoud

last: Kiaei

first: Stephanie

last: Byrum


title: Genome wide mapping of polyadenylation sites in proliferating and contact-inhibited cells and cells with knockdown of cleavage and polyadenylation factors

first: Hilary

last: Coller

summary:
Purpose: Quiescence is a state of reversible cell cycle exit. Levels of polyadenylation factors decreases when proliferating cells become quiescent.  The goals of this study are to determine the differential use of polyadenylation sites (changes in alternative polyadenylation) in quiescent vs. proliferating cells and also upon knockdown of polyadenylation factors.
Methods: Two biological replicates of human dermal fibroblasts (12-1 and 12-3) were used for polyadeylation-site enriched RNA-seq on an Illumina HiSeq 2500 to compare quiescent vs. proliferating cells and polyadenylation factor knockdown vs. control cells. The reads were aligned to the human genome (hg19) uisng Tophat (2.0.14). The resulting bam files were used as an input to a python script provided by Gruber et al. (PMID: 27382025) to determine the counts for each polyadenylation site.
Results: We observed a shift toward greater use of distal polyadenylation sites when the fibroblasts entered quiescence. We observed significant overlap between the genes that shift to greater distal site use with quiescence and CstF-64 or CPSF73 knockdown.
Conclusions: The shift to greater distal site use with quiescence may reflect in part the reduced levels of cleavage and polyadenylation factors.

overall_design: Perform polyadenylation site-enriched RNA-Seq on: (1) two biological replicates of proliferating and quiescent (contact-inhibited) cells, and (2) two biological replicates of control and polyadenylation factor (CstF64, CPSF73 or CFIm25) knockdown cells.

first: Mithun

last: Mitra

first: Vinay

last: Swamy

first: Wei

last: Wang

first: Jessica

last: Buckles


title: Single-cells transcriptomes of mouse developing forelimbs

status: Public on Aug 24 2018

city: Lausanne

first: Marion

last: LELEU

address: EPFL-SV-ISREC-UPDUB- Station 19

department: School of Life Sciences

country: Switzerland

institute: EPFL

summary:
In order to try and evaluate Hoxd transcript heterogeneity during limb development, we produced single-limb cell transcriptomes of different origins, to see whether the apparently homogenous behavior in Hox gene transcriptional program as observed upon large-scale analyses was confirmed at the cellular level. We report here thatHoxd genes transcripts are present in various combinations in different limb cells and discuss the importance of these results in our understanding of how Hoxd genes are regulated and how their global functions are achieved in these structures.
Single-cells transcriptomes from Hoxd11::GFP forelimbs with micro-dissection of autopod and zeugopod at embryonic day 12 (E12.5).

overall_design: Dissociated single-cells were obtained from eight Hoxd11::GFP forelimbs micro-dissected at E12.5.  Cells with the highest level of GFP fluorescence (top 20%) were sorted using an Astrios cell sorter with a 100-μ m nozzle. 75bp reads were uniquely mapped to the latest Mus Musculus reference genome (mm10) and the ERCC sequences using bowtie2 (REF: doi:10.1038/nmeth.1923) in local mode.

first: P

last: Fabre

first: M

last: Leleu

first: D

last: Duboule


title: RNA-seq Analysis of Msx2fl/fl(Msx2 WT) and Le-Cre+/- Msx2fl/fl(Msx2 CKO) Lens Transcriptomes

city: Shenyang

first: ziyan

last: yu

state: Liaoning

address: 11 Xinhua Road

institute: China  Medical University

summary:
Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare NGS-derived lens transcriptome profiling (RNA-seq) to microarray and quantitative reverse transcription polymerase chain reaction (qRT–PCR) methods and to evaluate protocols for optimal high-throughput data analysis
Methods: Lens RNA was extracted from Msx2 WT and Msx2 CKO mice at postnatal day 60 (P60) using RNAeasy™ Animal RNA Isolation Kit. The freshly frozen RNA solution was prepared for RNA sequencing on the same day. The RNA-seq was carried out using Illumina HiSeq sequencing (Novogene Bioinformatics Institute, Beijing, China). The insert size of the library was tested by Agilent 2100 Bioanalyzer to ensure library quality before Illumina HiSeq sequencing. The company used STAR software to compare RNA-seq sequencing data, HTSeq to analyze the quantitative of gene levels, RSEM to analyze transcript levels quantitatively, ClusterProfiler software to analyze differential gene enrichment, rMATS software to analyze alternative splicing, GATK software to analyze mutation sites and SnpEff software to analyze annotation of the variation sites. After mutation site detection and annotation, statistical analysis was done by Ts/Tv and Hom/Het, respectively.
Results: RNA-sequencing results showed that 1,911 differentially expressed genes (DEGs) were detected. Among them, the expression of 1,586 genes were upregulated and the rest downregulated.

overall_design: Lens mRNA profiles of 60-day old Msx2 WT and Msx2 CKO mice were generated by deep sequencing, in once, using Illumina HiSeq sequencing.

first: Wenting

last: Yu

first: Ziyan

first: Jiangyue

last: Zhao


title: Myocardial-specific ablation of Jarid2 leads to dilated cardiomyopathy in mice

status: Public on Aug 23 2018

city: Madison

first: Youngsook

last: Lee

state: WI

address: 1111 Highland Ave

department: CRB

institute: University of Wisconsin-Madison

summary: Analysis of heart from myocardil specific deletion of Jarid2 by crossing Jarid2 floxed mice with aMHC-Cre mice (Jarid2-aMHC). Jarid2-aMHC mice regult in dilated cardomyopathy and reveal premature death. To identify differentially expressed genes in Jarid2-aMHC hearts, we performed RNA-seq analysis on mutant hearts and control heart at postnatal day 10 (P10) and 7 months of ages (7M).

overall_design: Examination of 2 different stages in control and mutant hearts

first: Eunjin

last: Cho


title: Single Cell RNAseq Uncovers a Robust Transcriptional Response to Morphine by Glia

city: St. Louis

first: Robi

state: MO

address: 4515 McKinley Ave

institute: Washington University School of Medicine

summary: Molecular and behavioral responses to opioids are thought to be primarily mediated by neurons, although there is accumulating evidence that other cell types play a prominent role in drug addiction. To investigate cell-type-specific opioid responses, we performed single-cell RNA sequencing of the nucleus accumbens of mice following acute morphine treatment.  Differential expression analysis uncovered unique morphine-dependent transcriptional responses by oligodendrocytes and astrocytes. Further analysis using RNAseq of FACS-purified oligodendrocytes revealed a large cohort of morphine-regulated genes. Importantly, the affected genes are enriched for roles in cellular pathways intimately linked to oligodendrocyte maturation and myelination, including the unfolded protein response. Altogether, our data illuminate the morphine-dependent transcriptional response by oligodendrocytes and offer mechanistic insights into myelination defects associated with opioid abuse.

overall_design: For single-cell experiments, we perfoed Drop-seq on 4 biological replicates each of the nucleus accumbens from saline- or morphine-treated mice. For bulk RNAseq experiments, we performed FACS to purify oligodendrocytes from the entire brains of 6 mice (3 mock, 3 morphine), then isolated RNA and subjected to RNA sequencing.

first: Denis

last: Avey


title: The RNA-binding protein ROD1/PTBP3 cotranscriptionally defines AID-loading sites to mediate antibody class switch in mammalian genomes

city: Beijing

first: Yuanchao

last: Xue

address: Datun Road #15

department: Institute of Biophysics

institute: Chinese Academy of Sciences

summary: Activation-induced cytidine deaminase (AID) mediates class switching by binding to a small fraction of single-stranded DNA (ssDNA) to diversify the antibody repertoire. The precise mechanism for highly selective AID targeting in the genome has remained elusive. Here, we report an RNA-binding protein, ROD1 (also known as PTBP3), that is both required and sufficient to define AID-binding sites genome-wide in activated B cells. ROD1 interacts with AID via an ultraconserved loop, which proves to be critical for the recruitment of AID to ssDNA using bi-directionally transcribed nascent RNAs as stepping stones. Strikingly, AID-specific mutations identified in human patients with hyper-IgM syndrome type 2 (HIGM2) completely disrupt the AID interacting surface with ROD1, thereby abolishing the recruitment of AID to immunoglobulin (Ig) loci. Together, our results suggest that bi-directionally transcribed RNA traps the RNA binding protein ROD1, which serves as a guiding system for AID to load onto specific genomic loci to induce DNA rearrangement during immune responses.

overall_design: ROD1/PTBP3 CLIP-seq was performed with naïve splenic B cells and LPS-activated B cells, and two biological replicates were labeled as Rep1 and Rep2. Two biological repeats of AID CLIP-seq in LPS-activated B cells were labeled as Rep1 and Rep2; in addition, two AID CLIP-seq libraries were also generated upon the depletion of ROD1/PTBP3. For comparison, IgG antibodies were applied to generate two replicates of CLIP-seq libraries for negative control purposes. To examine whether ROD1/PTBP3 ablation alter the global nascent RNA levels, two biological repeats of GRO-seq were performed on LPS-activated B cells.

first: Juan

last: Chen

first: Zhaokui

last: Cai


title: Transcriptional analysis of whole blood and PBMCs from human HOIP deficiency patients

city: Bethesda

first: Daniel

last: Kastner

state: MD

address: 9000 Rockville Pike

department: NHGRI

institute: NIH

summary: LUBAC (linear ubiquitin assembly complex) consists of HOIP, HOIL1 and SHARPIN. HOIP deficiency has been characterized by immunodeficiency, autoinflammation and amylopectinosis. To explore the molecular mechanism of the autoinflammation, we performed a transcriptome-wide analysis using whole blood and unstimulated PBMCs of a HOIP deficiency patient.

overall_design: RNAseq using the whole blood and PBMCs of the HOIP deficiency patient, healthy unrelated controls and the patient's parent (heterozygous carrier of a mutation in HOIP)

first: Hirotsugu

last: Oda

first: Ivona

last: Aksentijevich


title: Dietary antigens influence the antigen presentation pathway of small intestine epithelium

city: Seoul

first: Tae-Young

last: Roh

state: --- Select One ---

address: 52 Ewhayeodae-gil, Seodaemun-gu

department: Life Sciences

country: South Korea

institute: Ewha Womans University

summary: Epithelial cells of the small intestine are designed to absorb the nutrients from the diet, so we tested if the absence of dietary macromolecules and protein antigens had an impact on the physiological condition of IECs. We generated RNAseq data from small intestine IECs isolated from mice free of any conventional non-self antigens for TCRab T cells. A comparison with RNAseq data generated from IECs isolated from SPF or GF mice showed clear differences between the transcriptomes of IECs isolated from the AF, SPF or GF mice.

overall_design: mRNA were purified from small intestinal epithelial cells of SPF, GF or AF mice and sequenced using Nextseq 500 (Illumina).

first: Jisun

last: Jung

first: Kwang

last: Kim

first: Hyun-Ja

last: Ko

first: Jeongeun

last: Park

first: Seokjin

last: Ham

first: Charles

last: Surh


title: Genome-wide maps of chromatin state in primary MLL-AF9 AML

first: Yinon

last: Ben-Neriah

address: Ein Kerem

department: The Lautenberg Center for Immunology

institute: Hebrew University-Hadassah Medical School

summary: Here we describe the development of CKIα inhibitors, which co-target the transcriptional kinases CDK7 and CDK9, thereby augmenting CKIα-induced p53 activation and its anti-leukemic activity. Oncogene-driving super-enhancers (SEs) are highly sensitive to CDK7/9 inhibition. We identified multiple newly-gained SEs in primary mouse AML cells and demonstrate that the inhibitors abolish many SEs and preferentially suppress the transcription elongation of SE-driven oncogenes. We show that blocking CKIα together CDK7 and/or CDK9 synergistically stabilize p53, deprive leukemia cells of survival and proliferation-maintaining SE-driven oncogenes, and induce apoptosis.

overall_design: H3K27Ac in MLL-AF9 driven mouse AML cells are compared to normal mouse bone cells to identify oncogene-driving super-enhancers (SEs). We further study H3K27Ac and pSer2 RNA polymerase II CTD enrichment in primary MLL-AF9 driven AML cells upon treatment with a small molecule inhibitor (A51) and Control (DMSO).

first: Avanthika

last: Venkatachalam

first: Waleed

last: Minzel

first: Eitan

last: Winter


title: Evf2 ultraconserved enhancer long non-coding RNA (UCE lncRNA)-dependent UCE interactions (4Cseq), and cohesin binding, histone methylation and acetylation (ChIPseq) in mouse E13.5 ganglionic eminences

city: Chicago

first: Jhumku

last: Kohtz

state: Illinois

address: 303 E Chicago

department: Neurology

institute: Northwestern University

summary: In this study we used mice lacking Evf2 (Evf2TS/TS) and mice expressing a truncated form of Evf2 (Evf1TS/TS) to determine  UCE lncRNA epigenetic and chromosome toplogical control.  We used 4Cseq to investigate how Evf2 regulates UCE interactions along chromosome 6 (where Evf2 is expressed).  We used ChIpseq to compare histone methylation profiles from Evf2TS/TS and Evf1TS/TS.  In addition, we used ChIPseq to determine Evf2-depedent regulation of cohesin subunit binding (SMC1 and SMC3) and histone H3K27acetylation.  Together, these data support that Evf2 UCE lncRNA controls chromosome topology over multi-megabse distances, through cohesin binding and effects on histone methylation and acetylation.  Also included is the ChIPseq profile of Dlx binding sites in SW (outbred strain of mice) from E13.5 GE.

overall_design: We use ChiPseq and 4Cseq and bioinformatic analytics to determine significant differences between mice expressing the Evf2 UCE lncRNA and mice lacking the Evf2 UCE lncRNA, and mice expressing a truncated form of Evf2.

first: Ivelisse

last: Cajigas

first: Abhiit

last: Chakraborty

first: Ferhat

last: Ay


title: Elucidating the regulation of the Yeast Metabolic Cycle through the integration of gene expression and chromatin status

city: Boston

first: Salvador

last: Casani

address: 450 Brookline Avenue

department: Cancer Biology

institute: Dana Farber Cancer Institute

summary: YMC has been characterized in different molecular levels, proving to be a good model to study crosstalk between histone modification and gene regulation. In this study, we used different statistical methodologies to integrate histone modifications and gene expression data

overall_design: H3K18ac histone modification ChIP-Seq in 16 time-points across the Yeast Metabolic Cycle

first: Victor

last: Sanchez-Gaya

first: Salva

first: Manuel

last: Ugidos

first: Zheng

last: Kuang

first: Jane

last: Mellor

first: Ana

last: Conesa

first: Sonia

last: Tarazona


title: A comprehensive examination of the molecular and pathological changes of the colonic injury response

city: New York

first: Akanksha

last: Verma

state: New York

address: 1300, York Avenue

department: Department of Physiology and Biophysics

institute: Weill Cornell Medical College

summary: Gene expression changes in the murine colon were determined at early and late stages following colonoscopic-guided pinch biopsy by comparing normal mucosa of sham treated mice to wound beds

overall_design: RNA seq analysis was carried out on colonic mucosa of mice 1 hour, 6 hours, 3 days or 6 days in normal tissue following  sham treatment or the wound bed following pinch biopsy

first: David

last: Montrose

first: Xi

middle: K

last: Zhou

first: Erin

last: McNally

first: Erika

last: Sue

first: Hanhan

last: Wang

first: Ryohei

last: Nishiguchi

first: Olivier

last: Elemento

first: Kenneth

middle: W

last: Simpson

first: Peiying

last: Yang

first: Timothy

last: Hla

first: Andrew

last: Dannenberg


title: Cell non-autonomous regulation of chromosome segregation

city: Cambridge

first: Charles

middle: Arthur

last: Whittaker

institute: Koch Institute

summary: Chromosome segregation has been assumed to be a cell-autonomous process. We tested this assumption by comparing chromosome segregation fidelity in epithelial cells in various contexts and discovered that these cells have increased chromosome missegregation outside of their native tissue. Using organoid culture systems, we show that tissue architecture, specifically integrin function, is required for accurate chromosome segregation in epithelia. We find that tissue architecture enhances the correction of merotelic microtubule-kinetochore attachments, and this is critically important for maintaining chromosome stability in the polyploid liver. Our data lead to the surprising conclusion that chromosome segregation in epithelia is a cell non-autonomous process. We propose that disruption of tissue architecture could underlie the chromosome instability that characterizes and drives epithelial cancers. Moreover, our findings highlight the importance of context for fundamental cellular processes and caution against the exclusive reliance on cell culture systems for deciphering and manipulating mammalian biology.

overall_design: Three biological replicates of mammary epithelial cells were seeded on collagen-coated dishes to form monolayers or embedded in Matrigel to form spheroids and cultured for 48 or 96 hours.

first: Kristin

last: Knouse

first: Kristina

middle: E

last: Lopez

first: Marc

last: Bachofner

first: Angelika

last: Amon


title: Accurate recycling of parental histones reproduces the histone modification landscape during DNA replication III

city: Copenhagen

first: Anja

last: Groth

address: Blegdamsvej 3B

country: Denmark

institute: Novo Nordisk Foundation Center for Protein Research

summary: Chromatin organisation is disrupted genome wide during DNA replication. On newly synthesized DNA, nucleosomes are assembled from new naïve histones and old modified histones. It remains unknown whether the landscape of histone post-translational modifications (PTMs) is copied during DNA replication or the epigenomeis perturbed. Here we develop Chromatin Occupancy after Replication, ChOR-seq, a technology that combines chromatin immunopreciptation of histone marks and purification of newly replicated DNA by streptavidin pull-down followed by next generation sequencing. We use this technology to address propagation of epigenetic states across cell division and reveal that accurate recycling of modified parental histones ensures that positional information of histone marks is faithfully reproduced on daughter strands and inherited to daughter cells.

overall_design: Analysis of H3K4me3, H3K36me3 and H3K79me3 distribution and abundance after replication and across the cell cycle by ChOR-seq in HeLa S3 cells. All experiments were done in duplicates. Replicated chromatin was labelled in early S. Chromatin of Drosophila melanogater S2 cells were used as spike-in for normalization purpose.

first: Nazaret

last: Reverón-Gómez

first: Cristina

last: González-Aguilera

first: Kathleen

last: Stewart-Morgan


title: Novel human full-thickness three-dimensional non-keratinized mucous membrane model for pharmacological studies in wound healing

city: Aachen

first: Sebastian

last: Huth

address: Pauwelsstr. 30

country: Germany

institute: Uniklinik RWTH Aachen

summary: Efforts are increasingly aiming to develop in vitro models that can provide effective alternatives to in vivo experiments. The main aim of this study was the establishment of an in vitro model of the non-keratinized mucous membrane that can be used as a standardized tool to evaluate biological and therapeutic effects of pharmaceuticals for mucosal wound healing. Performing histological and immunofluorescence analyses with known differentiation markers we proved that our model mimics the two distinctive layers of the mucous membrane – the stratified squamous epithelium and the lamina propria. In our study we used our model to investigate molecular effects of a dexpanthenol-containing ointment that is widely used in the wound treatment of the oral mucosa. For that purpose our model exhibits a unique feature in that dexpanthenol and proliferation enhancing additives that may interfere with our studies are not required for the maintenance of the model culture. After setting standardized lesions with a CO2 laser, topical treatment with the dexpanthenol-containing ointment enhanced wound closure in our non-keratinized mucous membrane model compared to placebo and untreated controls. Furthermore, microarray analysis revealed that the treatment of our laser wounded model with the dexpanthenol-containing ointment evoked an upregulated expression of various genes related to accelerated wound healing. Overall, we verified that our mucous membrane model can be utilized in future to monitor ex vivo effects of various topical therapies on mucosa morphology, physiology, and gene expression. Our findings confirm the potential of the non-keratinized mucous membrane model as an in vitro tool for the replacement of pharmacological in vivo studies regarding mucosal wound healing.

overall_design: Six samples were analyzed: Comparison between 3D mucous membrane models after laser injury and treatment with the dexpanthenol-containing (n=3)  vs. placebo treated controls (n=3).

first: Laurenz

last: Schmitt

first: Yvonne

last: Marquardt

first: Ruth

last: Heise

first: Verena

last: von Felbert

first: Philipp

last: Amann

first: Timm

last: Steiner

first: Frank

last: Hölzle

first: Jens

last: Baron


title: Intrinsic Xenobiotic Resistance of The Intestinal Stem Cell Niche

city: Singapore

first: David

last: Virshup

address: 8 College Road

country: Singapore

institute: Duke-NUS Medical School

summary: To characterize intestinal myofibroblasts that may contribute to the stromal compartment of the intestinal stem cell niche and identify possible causes of intrinsic xenobiotic resistance

overall_design: Gene expression profile of 4 samples of isolated intestinal crypts compared to 4 samples of freshly isolated intestinal myofibroblasts

first: Yuemin

last: Chee

first: Babita

last: Madan


title: Accurate recycling of parental histones reproduces the histone modification landscape during DNA replication II

summary: Chromatin organisation is disrupted genome wide during DNA replication. On newly synthesized DNA, nucleosomes are assembled from new naïve histones and old modified histones. It remains unknown whether the landscape of histone post-translational modifications (PTMs) is copied during DNA replication or the epigenomeis perturbed. Here we develop Chromatin Occupancy after Replication, ChOR-seq, a technology that combines chromatin immunopreciptation of histone marks and purification of newly replicated DNA by streptavidin pull down followed by next generation sequencing. We use this technology to address propagation of epigenetic states across cell division and reveal that accurate recycling of modified parental histones ensures that positional information of histone marks is faithfully reproduced on daughter strands and inherited to daughter cells.

overall_design: Analysis of H3K27me3 and total H3 distribution and abundance after replication and across the cell cycle by ChOR-seq in HeLa S3 cells. All experiments were done in duplicates. Replicated chromatin was labelled in Mid S.


title: Non-inflammatory tumor microenvironment of Diffuse Intrinsic Pontine Glioma (DIPG)

city: Stanford

first: Michelle

last: Monje

state: CA

address: 265 Campus Drive G3035

institute: Stanford University

summary: Diffuse intrinsic pontine glioma (DIPG) is a universally fatal malignancy of the childhood central nervous system, with a median overall survival of 9-11 months. We have previously shown that primary DIPG tissue contains numerous tumor-associated macrophages, and substantial work has demonstrated a significant pathological role for adult glioma-associated macrophages. However, work over the past decade has highlighted many molecular and genomic differences between pediatric and adult glioblastomas (GBM). Thus, we directly compared inflammatory characteristics of DIPG and adult GBM. We found that the leukocyte (CD45+) compartment in primary DIPG tissue samples is predominantly composed of CD11b+ macrophages, with very few CD3+ T-lymphocytes. In contrast, T-lymphocytes are more abundant in adult GBM tissue samples. RNA sequencing of macrophages isolated from primary tumor samples revealed that DIPG- and adult GBM-associated macrophages both express gene programs related to ECM remodeling and angiogenesis, but DIPG-associated macrophages express substantially fewer inflammatory factors than their adult GBM counterparts. Examining the secretome of glioma cells, we found that patient-derived DIPG cell cultures secrete markedly fewer cytokines and chemokines than patient-derived adult GBM cultures. Concordantly, bulk and single-cell RNA sequencing data indicates low to absent expression of chemokines and cytokines in DIPG. Together, these observations suggest that the inflammatory milieu of the DIPG tumor microenvironment is fundamentally different than adult GBM. The low intrinsic inflammatory signature of DIPG cells may contribute to the lack of lymphocytes and non-inflammatory phenotype of DIPG-associated microglia/macrophages. Understanding the glioma subtype-specific inflammatory milieu may inform the design and application of immunotherapy-based treatments.

overall_design: RNA-seq of primary isolated microglia/macrophages from early post-mortem DIPG tissue samples, pediatric normal cortex, and adult GBM tissue samples. Libraries were sequenced on Illumina NextSeq 500, 1x75.

first: Grant

middle: L

last: Lin


title: Accurate recycling of parental histones reproduces the histone modification landscape during DNA replication

first: Anja

last: Groth

overall_design: Analysis of H3K27me3 and total H3 distribution and abundance after replication and across the cell cycle by ChOR-seq in HeLa S3 cells. All experiments were done in duplicates. Replicated chromatin was labelled in mid S. Chromatin of Drosophila melanogater S2 cells were used as spike-in for normalization purpose.

first: Nazaret

last: Reverón-Gómez


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

overall_design: Refer to individual Series


title: The effect of permethrin resistance on Aedes aegypti transcriptome following ingestion of Zika virus infected blood

status: Public on Aug 22 2018

city: GAINESVILLE

first: FAHONG

last: YU

address: 2033 Mowry Rd

department: ICBR

institute: UNIVERSITY OF FLORIDA

summary: Aedes aegypti (L.) is the primary vector of many emerging arboviruses. Insecticide resistance among mosquito populations is a consequence of the application of insecticides for mosquito control. We used RNA-sequencing to compare transcriptomes between permethrin resistant and susceptible strains of Florida Ae. aegypti in response to Zika virus infection. A total of 2,459 transcripts were expressed at significantly different levels between resistant and susceptible Ae. aegypti. Gene ontology analysis placed these genes into 7 categories of biological processes. The 863 transcripts were expressed at significantly different levels between two strains (up/down regulated) more than 2-fold. Quantitative real-time PCR analysis validated Zika-infected response, and suggested a highly overexpressed P450, with AAEL014617 and AAEL006798 as potential candidates for the molecular mechanism of permethrin resistance in Ae. aegypti.  Our findings indicated that most detoxification enzymes and immune system enzymes altered their gene expression between the two strains of Ae. aegypti in response to Zika virus infection. Understanding the interactions of arboviruses with resistant mosquito vectors at the molecular level allows for the possible development of new approaches in mitigating arbovirus transmission. This information sheds light on Zika-induced changes in the insecticide resistance of Ae. aegypti with implications for mosquito control strategies.

overall_design: Ae. aegypti larvae were collected from Key West (24.55°N, 81.78°W), Florida, USA and maintained at the Florida Medical Entomological Laboratory (FMEL) in Vero Beach, FL since 2011. The parental collection of Ae. aegypti from the field was initially tested for permethrin resistance, then subjected to permethrin selection for 15 generations (see below) and again assayed for resistance (referred to as the resistant strain). The Orlando strain of Ae. aegypti was collected from Orlando (28.53°N, 81.37°W), Florida, USA and reared in the Mosquito and Fly Research Unit, Center for Medical, Agricultural and Veterinary Entomology, ARS-USDA in Gainesville, FL since 1952. The Orlando strain is recognized as a permethrin susceptible strain of Ae. Aegypti. Four-day-old female adults were fed defibrinated bovine blood containing either ZIKV (treatment) or blood lacking virus (control). The ZIKV infected and control mosquitoes from two Ae. aegypti strains were harvested for a time course study. Individual mosquitoes were dissected into body and legs and tested to confirm susceptibility to infection and disseminated infection rates, respectively. Ten mosquitoes (12 hours and 7 days post ZIKV infection) were pooled for each sample for RNA sequencing.

first: L

last: Zhao

first: B

last: Alto

first: D

middle: Y

last: Shin

first: F

last: Yu


title: Chromatin accessibility analysis of adipose progenitors (AP).

city: Pittsburgh

first: Yang

last: Zhang

state: PA

address: 100 Technology Dr.

department: Medicine

institute: University of Pittsburgh

summary: We hypothesized that core transcriptional regulatory circuitries analysis could be used in conjunction with gene expression profiling data to nominate specific TFs for additional functional studies of AP cell state. To accomplish this analysis, we utilized the assay of transposase-accessible chromatin coupled to massively parallel sequencing (ATAC-seq) to identify putative enhancers in APs.  The AP preparation involved adipose tissue digestion, and negative selection of the stromal vascular fraction (depletion of CD31+ endothelial cells and Lineage positive cells).

overall_design: 4 samples were anlyzed. 2 groups of adipose progenitors were isolated from subcutaneou(SAT) and visceral (VAT) adipose tissue from 16 wks C57BL/6 mice,  in duplicate.

first: Alexander

last: Federation

first: Jonathan

last: Brown

first: Matthew

last: Steinhauser


title: Transcriptomic analysis of wild type and Nr4a1-/- adipose progenitors.

summary: The project aims to identify differentially expressed genes in adipose progenitors that were freshly isolated from wild-type or Nr4a1-/- mice. The AP preparation involved adipose tissue digestion, and negative selection of the stromal vascular fraction (depletion of CD31+ endothelial cells and Lineage positive cells.

overall_design: 16 samples were anlyzed. 4 groups of adipose progenitors were isolated from subcutaneou(SAT) and visceral (VAT) adipose tissue from Nr4a1 wildtype(Nr4a1+/+)  and knockout(Nr4a1-/-) mice. Each group has 4 biological replicates.


title: RNA Sequencing of Tgd17 progenitors

city: Worcester

first: Nicholas

last: Spidale

address: 368 Plantation St.

department: Pathology

institute: University of Massachusetts Medical School

summary: Tgd17 cells are critical immune effector cells that exit the thymus committed to peripheral IL-17 responses, but whether the development of cells into this lineage is pre-programmed or driven by instructive TCR signals is unresolved.  Using newly generated reporter mice for a gene necessary for Tgd17 cell development, Sox13, we identified progenitor cells (known as DN1d cells) that expressed high levels of Sox13 prior to expression of the TCR.  To interrogate whether DN1d cells could be a candidate Tgd17-biased progenitor, whole transcriptome RNA sequencing was performed on these progenitors as well as canonical T cell progenitors thought to generate all T cell subsets and the proposed progeny Tgd17 cells. The data demonstrated that DN1d cells most closely resemble Tgd17 cells at a transcriptome level, while canonical T cell progenitors were very different from both DN1d cells and Tgd17 cells.  Further studies demonstrated that DN1d cells generated Tgd17 cells in developmental assays, and that the transcriptional signature of DN1d cells is established and maintained independently of TCR expression or signaling but is instead dependent on HMG transcription factors SOX13 and TCF1.  Thus, T cell lineage identity can be pre-programmed at the progenitor level and is not necessarily specified by specific TCR signals.

overall_design: Seven discrete populations of thymocytes, including 2-3 biological replicates for each population, were assessed by RNA-Seq.  While population comparisons were performed in an unbiased manner, the "ImmV2" population may be considered a reference population to which the other progenitor cell populations were being compared.

first: Nitya

last: Jain

first: Joonsoo

last: Kang


title: Systematic Identification of Transcription Factors Driving Cell Fate Conversion using CRISPR Activation Screens

first: Lei

middle: Stanley

last: Qi

address: 443 Via Ortega

department: Bioengineering

summary: In order to characterize CRISPRa induced and cDNA trans-differentiated neurons

overall_design: mRNA profiles of CamES, Mef, induced neuron, and trans-differentiated neuron cells were generated by deep sequencing, in replicate, using Illumina HiSeq sequencing systems.

first: Yanxia

last: Liu

first: Chen

first: Timothy

last: Daley


title: Generating a RAS expression signature in neuroblastoma

city: Amsterdam

first: Richard

last: Volckmann

state: Noord-Holland

address: van der Boechorststraat 6a

department: Laboratory of Experimental Oncology and Radiobiology

country: Netherlands

institute: msterdam UMC and University of Amsterdam

summary: Mutations affecting the RAS-MAPK pathway frequently occur in relapse neuroblastoma tumors, which suggests that activation of this pathway is associated with a more aggressive phenotype.  To explore this hypothesis we generated several model systems to define a neuroblastoma RAS-MAPK pathway signature. We could show that activation of this pathway in primary tumors indeed correlates with poor survival and is associated with known activating mutations in ALK and other RAS-MAPK pathway genes. From integrative analysis we could show that mutations in PHOX2B, CIC and DMD are also associated with an activated RAS-MAPK pathway. Mutation of PHOX2B and deletion of CIC in neuroblastoma cell lines induces activation of the RAS-MAPK pathway. This activation was independent of phosphorylated ERK in the CIC knock out systems. Furthermore, deletion of CIC causes a significant increase in tumor growth in vivo. These results show that the RAS-MAPK pathway is involved in tumor progression, and establish CIC as a powerful tumor suppressor that functions downstream of this pathway in neuroblastoma.

overall_design: We made several model system in which RAS-MAPK pathway activation is induced/inhibited. NRAS is overexpressed and NF1 knocked out using CRISPR in the cell line NMB. SJNB8, a cell line which contains a KRAS mutation and has an active pathway, was treated with the MEK inhibitor Trametinib to inihbit pathway activation.

first: Jan

middle: J

last: Molenaar

first: Thomas

last: Eleveld


title: Telomeric epigenetic response mediated by Gadd45a regulates stem cell aging and lifespan

city: Hangzhou

first: Li

last: Shen

state: Zhejiang Province

address: 866 Yuhangtang Road

institute: Life Sciences Institute, Zhejiang University

summary: Progressive attrition of telomeres triggers DNA damage response (DDR) and limits the regenerative capacity of adult stem cells during mammalian aging. Intriguingly, the telomere integrity is not only determined by telomere length but also by epigenetic status of telomeric/sub-telomeric regions. However, the functional interplay between telomere shortening-induced DDR and epigenetic modification in aging is unknown. Here we show that deletion of Gadd45a improves the maintenance and function of intestinal stem cells (ISCs), and prolongs lifespan of late generation of telomerase deficient mice (G3Terc-/-). Mechanistically, Gadd45a facilitates the generation of a permissive chromatin status for DDR signaling transduction through the demethylating of CpG islands specifically at sub-telomeric regions of short telomeres. Deletion of Gadd45a restores the heterochromatin compaction in the sub-telomeric regions and thereby ameliorating the DDR initiation at short telomeres of G3Terc-/- ISCs. Treatment with a small molecule inhibitor of BER alleviates DDR and improves the maintenance and function of G3Terc-/- ISCs. Taken together, our study discloses a potential therapeutic approach to enhance stem cell function and prolong lifespan by targeting epigenetic modulating molecules.

overall_design: RNA-seq comparing mouse primary embryo fibroblast generated from WT, Gadd45a-/-, G3Terc-/- and G3-dKO mice.

first: Daojun

last: Diao

first: Hu

last: Wang

first: Tangliang

last: Li

first: Zhencan

last: Shi

first: Xiaoqing

last: Jin

first: Tobias

last: Sperka

first: Xudong

last: Zhu

first: Meimei

first: Fan

last: Yang

first: Yusheng

last: Cong

first: Qimin

last: Zhan

first: Jing

last: Yan

first: Zhangfa

last: Song

first: Zhenyu

last: Ju


title: Health-relevant phenotypes in the offspring of mice given CAR activators prior to pregnancy

city: Mainz

first: Piyush

last: More

address: Langenbeckstrasse 1

department: Department of Pharmacology

institute: University Medical Center, Johannes Gutenberg University of Mainz

summary: The hepatic induction in response to drugs and environmental chemicals affects drug therapies and energy metabolism. We investigated whether the induction is transmitted to the offspring. We injected 3-day and 6-week-old F0 female mice with the activator of the nuclear receptor CAR (constitutive androstane receptor, NR1I3) TCPOBOP and mated them 1-6 weeks afterwards. We detected in the offspring long-lasting alterations of CAR-mediated drug disposition, energy metabolism, and lipid profile. The transmission to F1 was mediated by TCPOBOP transfer from the F0 adipose tissue via milk, as revealed by embryo transfer, cross-fostering experiments, and liquid chromatography-mass spectrometry analyses. The important environmental pollutant PCB153 activated CAR in the F1 generation similarly to TCPOBOP. Our findings indicate that chemicals accumulating and persisting in adipose tissue may exert liver- mediated, health-relevant effects on F1 offspring simply via physical transmission with milk. Such effects may occur even if treatment has been terminated far ahead of conception. This should be considered in assessing developmental toxicity and in the long-term follow-up of offspring of mothers exposed to both approved and investigational drugs, and to chemicals known or suspected of accumulation in adipose tissue.

overall_design: Female F0 mice used to obtain F1 offspring were induced with TCPOBOP (3 mg/kg) three days after birth. We considered 6-day-old and 3-month-old mice of the F0 and of the F1 generation for mRNA-sequencing. For each group the RNA of 2 to 6 mice was pooled into one sample. We then sequenced TCPOBOP treated mice as well as their F1 offspring together with the respective corn oil controls.

first: Leszek

last: Wojnowski

first: Marianne

last: Mathäs


title: Single-cell analysis reveals stochastic regulation of type I IFN production by plasmacytoid dendritic cells and identifies host-derived environmental cues as amplifier of type I IFN production

city: Utrecht

first: Judith

middle: Anne

last: Vivié

state: Utrecht

address: Uppsalalaan 8

department: Single Cell Sequencing Facility

institute: Hubrecht Institute

summary: Type I interferon (IFN) is a key driver of immunity to infections and cancer. Plasmacytoid dendritic cells (pDCs) are uniquely equipped to produce large quantities of type I IFN but the mech-anisms that control this process are poorly understood. Here, we developed a droplet-based microfluidic platform and investigated type I IFN production in human pDCs at the single-cell level. We show that type I IFN but not TNF alpha production is limited to a small subpopulation of individually stimulated pDCs and controlled by stochastic gene regulation. Combining single-cell cytokine analysis with single-cell RNA-seq profiling reveals no evidence for a pre-existing subset of type I IFN-producing pDCs. Intriguingly, by modulating the droplet microenvironment, we demonstrate that vigorous pDC population responses are driven by a type I IFN amplification loop. Our study highlights the significance of stochastic gene regulation and suggests strategies to dissect the characteristics of immune responses at the single-cell level.

overall_design: Examination of the transcriptional profiles from single human plasmacytoid dendritic cells that were stimulated with CpG-C

first: Florian

last: Wimmers

last: Vivie

first: Rob

last: Woestenenk

first: Jurjen

last: Tel


title: Chemical Screening Identifies Enhancers of Mutant Oligodendrocyte Survival and Unmasks a Distinct Pathological Phase in Pelizaeus-Merzbacher Disease [RNA-seq]

city: Cleveland

first: Paul

last: Tesar

state: Ohio

address: 10900 Euclid Ave.

department: Genetics & Genome Sciences

institute: Case Western Reserve University

summary: Pelizaeus-Merzbacher disease (PMD) is a fatal X-linked disorder caused by loss of myelinating oligodendrocytes and consequent hypomyelination. The underlying cellular and molecular dysfunctions are not fully defined, but therapeutic enhancement of oligodendrocyte survival could restore functional myelination in patients. Here we generated pure, scalable quantities of iPSC-derived oligodendrocyte progenitor cells (OPCs) from a severe mouse model of PMD, Plp1jimpy. Temporal phenotypic and transcriptomic studies defined an early pathological window characterized by endoplasmic-reticulum (ER) stress and cell death as OPCs exit their progenitor state. High-throughput phenotypic screening identified a compound, Ro 25-6981, which modulates the ER stress response and rescues mutant oligodendrocyte survival in jimpy, in vitro and in vivo, and in human PMD oligocortical spheroids. Surprisingly, increasing oligodendrocyte survival did not restore subsequent myelination, revealing a second pathological phase. Collectively, our work shows that PMD oligodendrocyte loss can be rescued pharmacologically and defines a need for multifactorial intervention to restore myelination.

overall_design: RNAseq profiles comparing wild-type and jimpy oligodendrocyte progenitor cells and differentiating oligodendrocytes

first: Matthew

last: Elitt


title: Chemical Screening Identifies Enhancers of Mutant Oligodendrocyte Survival and Unmasks a Distinct Pathological Phase in Pelizaeus-Merzbacher Disease [scRNA-seq]

overall_design: Single cell RNAseq profiles comparing wild-type and jimpy differentiating oligodendrocyte cultures

first: Chen

last: Weng

first: Fulai

last: Jin

first: Yan

last: Li


title: Patients with human resected pancreatic cancer

city: Brussels

first: Sylvain

last: Brohée

address: Boulevard de Waterloo 127

country: Belgium

institute: Institut Jules Bordet

summary: This cohort included 118 consecutive and unselected patients who underwent, between September 1998 and September 2010, curative intent surgery for PDA (Brussels Dataset).  Archived tumor specimens were available for the entire study population. Exclusion criteria included: preoperative chemotherapy or chemoradiotherapy (n=12), macroscopically incomplete resection (R2) (n=6), or tumor histology other than ductal adenocarcinoma (n=19). We also excluded patients who died of postoperative complications within 30 days following surgery (n=1) because they are non-informative for this kind of translational study.

overall_design: Of the available samples, gene mRNA expression data was produced from 118 samples. All samples were re-evaluated to ensure tumour was present in the specimen. RNA was extracted from formalin-fixed, paraffin-embedded (FFPE) primary  tumour tissue.

first: Francesco

last: Puleo

first: Raphaël

last: Maréchal

first: Pieter

last: Demetter

first: Debora

last: Fumagalli

first: Jean-Baptiste

last: Bachet

first: Laurine

last: Verset

first: Eric

last: Quertinmont

first: Jacques

last: Devière

first: Denis

last: Franchimont

first: Christos

last: Sotiriou

first: Jean-Luc

last: Van Laethem


title: PRMT2 links histone H3R8 asymmetric dimethylation to oncogenic activation and tumorigenesis of glioblastoma

city: Tianjin

first: xudong

last: Wu

address: Qixiangtai Road

institute: Tianjin medical university

summary: Here we show that PRMT2 is highly expressed in GBM, which is correlated with poor prognosis. The silencing of PRMT2 inhibits GBM cell growth and glioblastoma stem cell self-renewal in vitro and suppreses orthotopic tumor growth. Transcriptome analysis demonstrated that PRMT2 depletion leads to significant deregulation of genes mainly associated with cell cycle progression and pathways in cancer. In agreement with previously published results, we show that PRMT2 is responsible for H3R8 asymmetric methylation (H3R8me2a) and that H3R8me2a enrichment at promoters and enhancers is closely correlated with known active histone marks. Furthermore, we show that PRMT2-mediated H3R8me2a is required for the maintenance of target gene expression and that the catalytic activity of PRMT2 is required for its protumorigenic functions. Taken together, this study demonstrates that PRMT2 acts as a transcriptional co-activator for oncogenic gene expression programs in GBM pathogenesis and provides a rationale for PRMT2 targeting in aggressive gliomas.

overall_design: To examine changes in histone-modification profiles and gene expression after knockdown of PRMT2 in human U87 cells or human primary glioblastoma cells TPC1115.

first: Feng

last: Dong

first: Xudong


title: Next-generation sequencing of control (scramble) and Myo1c knockdown human podocytes, with and without TGF-β treatment

status: Public on Aug 21 2018

city: Charleston

first: Deepak

last: Nihalani

state: SC

address: 70 President St.

department: Medicine/Nephrology

institute: Medical University of South Carolina

summary:
Purpose: Next-generation sequencing (NGS) was used to identify cellular pathways and genes through systems-based analysis. The goals of this study are to identify NGS-derived transcriptome profiling (RNA-seq) in control and Myo1c knockdown human podocytes upon TGFβ stimulation. These high-throughput data were further validated through qRT–PCR methods to confirm the cellular pathways and genes affected due to genetic knockdown of Myo1c and TGF-β stimulation.
Methods: Human control and Myo1c knockdown podocytes were differentiated for 14 days by thermoswitching from 33⁰C to 37⁰C and removal of growth factors, insulin-transferrin-selenium from the medium. These podocytes were incubated overnight in RPMI medium with 0.1% FBS and stimulated with 5ng/ml of TGF-β in the same medium for a period of 48 hours. Control and TGF-β stimulated podocytes were processed for RNA isolation and submitted to Novogene for RNA-Seq. All the experiments were performed in triplicates.
Conclusions: Our study is first to describe the detailed analysis of TGF-β stimulated Myo1c knockdown podocyte transcriptomes using the RNA-seq technology. A comparative analysis of the differential expression profile was obtained between control vs. Myo1c knockdown podocytes, and control vs. Myo1c knockdown podocytes that were stimulated with TGF-β. Complex genetic network and genes effected due to Myo1c knockdown and TGF-β stimulation will provide a platform to define biological pathways in podocytes where Myo1c participates.

overall_design:
Human podocytes mRNA profiles in control (scramble) and Myo1c knockdown cells were generated by deep sequencing, in triplicate, using Illumina.
Contributor: Novogene Corporation

first: Ehtesham

last: Arif


title: A reference collection of cell line and xenograft models of proneural, classical and mesenchymal GBM [xenograft]

city: Brisbane

first: Brett

middle: William

last: Stringer

address: 300 Herston Road

institute: QIMR Berghofer Medical Research Institute

summary:
Low-passage, serum-free cell lines cultured from patient tumour tissue are the gold-standard for preclinical studies and cellular investigations of glioblastoma (GBM) biology, yet entrenched, poorly-representative cell line models are still widely used, compromising the significance of much GBM research. We submit that greater adoption of these critical resources will be promoted by the provision of a suitably-sized, meaningfully-described reference collection along with appropriate tools for working with them. Consequently, we present a curated panel of 12 readily-usable, genetically-diverse, tumourigenic, patient-derived, low-passage, serum-free cell lines representing the spectrum of molecular subtypes of IDH-wildtype GBM along with their detailed phenotypic characterisation and a bespoke set of lentiviral plasmids for bioluminescent/fluorescent labelling, gene expression and CRISPR/Cas9-mediated gene inactivation. The cell lines and all accompanying data are readily-accessible via a single website, Q-Cell (qimrberghofer.edu.au/q-cell/) and all plasmids are available from Addgene. These resources should prove valuable to investigators seeking readily-usable, well-characterised, clinically-relevant, gold-standard models of GBM.
Gene expression in 12 matched patient tumour / derived cell line / resultant xenograft samples was measured using Illumina HumanHT-12_V4_0_R2_15002873_B direct hybridisation beadchips.

overall_design: This experiment measures RNA expression in GBM cell line-initiated xenografts.


title: A reference collection of cell line and xenograft models of proneural, classical and mesenchymal GBM


title: A reference collection of cell line and xenograft models of proneural, classical and mesenchymal GBM [cell line]

overall_design: This experiment measures RNA expression in patient-derived GBM cell lines.


title: A reference collection of cell line and xenograft models of proneural, classical and mesenchymal GBM [tumor tissue]

overall_design: This experiment measures RNA expression in patient tumour tissue.


title: Expression data from human CD4 T cells differentiated under inflammatory conditions

city: Kyoto

first: Hiroyuki

last: Yoshitomi

address: 53, Kawahara-Cho, Shogoin,

institute: Institute for Frontier Life and Medical Sciences, Kyoto University

summary: To identify transcriptio factors responsible for CXCL13 production by human CD4+ T cells, we differentiated CXCL13-producing CD4+ T cells in vitro under TGF-β-positive inflammatory conditions and conducted transcriptome analysis.

overall_design: Compare RNA exression between CXCL13-negative condition (IL-12) and CXCL13-positive conditions (TGFb+IL1b/TGFb+IL6)

first: Shio

last: Kobayashi


title: Single cell changes in DBZ or DMSO treated cells derived from Apc-mutant organoid cultures

city: Helsinki

first: Jenny

last: Hogstrom

address: Haartmaninkatu 8

country: Finland

institute: University of Helsinki

summary: We made intestinal organoid cultures from Villin-CreERT2;Apcflox/flox mice. Organoids were subjected to 4-OH-Tam treatment, whereafter Apc mutant organoids were selected in R-Spondin free medium. DBZ or DMSO treatment was started on day 3. On day 8, organoids were dissociated to obtain single cells and analyzed using the Chromium Single-cell 3'RNA-sequencing system

overall_design: We analyzed in total 2 samples (1 DBZ and 1 DMSO), both samples contained 3000 cells

last: Högström

first: Kari

last: Alitalo


title: Transcriptome analysis of human endometrial stromal cells with NR2F2 (COUP-TFII) knockdown

city: Tainan

first: Shaw-Jenq

last: Tsai

address: 1 , University Road

department: Department of Physiology, College of Medicine

country: Taiwan

institute: National Cheng Kung University

summary: To explore the roles of COUP-TFII during the pathogenesis of endometriosis, the human eutopic stromal cells were transfected with siRNA oligonucleotide against COUP-TFII, and total RNA were harvested and proceeded to microarray analysis.

overall_design: 2 treatments with 3 biological replicates

first: Kuei-Yang

last: Hsiao


title: A novel transcriptional network for the Androgen Receptor in human epididymis epithelial cells [RNA-Seq]

first: Ann

last: Harris

state: OH

address: 10900 Euclid Ave

country: USA

institute: Case Western Reserve Uni

summary: The androgen receptor (AR) has a pivotal role in regulating gene expression in the male reproductive system. Due to the involvement of AR in prostate cancer, its role is best studied in the prostate gland epithelium and prostate cancer cell lines. Here we investigate the transcriptional program of AR in normal human epididymis epithelial (HEE) cells. After AR stimulation of caput HEE cells with the synthetic androgen R1881, AR targets were revealed with RNA-sequencing. Next, AR occupancy genome-wide was determined in control or R1881-stimulated HEE cells by chromatin immunoprecipitation and deep sequencing (ChIP-seq). The results identify about 200 genes that are differentially expressed (DEGs) in HEE cells after AR activation. Some of these DEGs show occupancy of AR at their promoters or cis-regulatory elements suggesting direct regulation. However there is little overlap in AR-associated DEGs between HEE and prostate epithelial cells. Inspection of over-represented motifs in AR ChIP-seq peaks identified CAAT-enhancer binding protein beta (CEBPB) and Runt-related transcription factor 1 (RUNX1) as potential co-factors, with no evidence for FOXA1, which is an important co-factor in the prostate epithelium.  CEBPB and RUNX1 ChIP-seq in HEE cells showed that both these factors often occupied AR-binding sites, though rarely simultaneously.  Further analysis at a single AR-regulated locus (FK506-binding protein 5, FKPB5) suggests that RUNX1 may inhibit AR occupancy, while CEBP appears to be a co-activator. These data suggest a novel AR transcriptional network governs differentiated functions of the human epididymis epithelium.

overall_design: Examination of AR cistrome in primary human epididymis cells with or without R1881 treatment

first: Rui

last: Yang

first: James

middle: A

last: Browne

first: Scott

last: Eggener

first: Shih-Hsing

last: Leir


title: A novel transcriptional network for the Androgen Receptor in human epididymis epithelial cells [ChIP-Seq]


title: A novel transcriptional network for the Androgen Receptor in human epididymis epithelial cells

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

overall_design: Refer to individual Series