title: The miR-96 and RARG signaling axis governs androgen signaling and prostate cancer progression V

city: Buffalo

first: Mark

middle: D

last: Long

address: Elm & Carlton Streets

department: Bioinformatics & Biostatistics

institute: Roswell Park Comprehensive Cancer Center

summary:
Expression levels of retinoic acid receptor gamma (NR1B3/RARG, encodes RARG), are commonly reduced in prostate cancer (PCa). Therefore we sought to establish the cellular and gene regulatory consequences of reduced RARG expression, and determine RARG regulatory mechanisms. RARG shRNA approaches in non-malignant (RWPE-1 and HPr1-AR) and malignant (LNCaP) prostate models revealed that reducing RARG levels, rather than adding exogenous retinoid ligand, had the greatest impact on prostate cell viability and gene expression. ChIP-Seq defined the RARG cistrome which was significantly enriched at active enhancers associated with AR binding sites. Reflecting a significant genomic role for RARG to regulate androgen signaling, RARG knockdown in HPr1-AR cells significantly regulated the magnitude of the AR transcriptome. RARG down-regulation was explained by increased miR-96 in PCa cell and mouse models, and TCGA PCa cohorts. Biochemical approaches confirmed that miR-96 directly regulated RARG expression and function. Capture of the miR-96 targetome by biotin-miR96 identified that RARG and a number of RARG  interacting co-factors including TACC1 were all targeted by miR-96, and expression of these genes were prominently altered, positively and negatively, in the TCGA-PRAD cohort. Differential gene expression analyses between tumors in the TCGA-PRAD cohort with lower quartile expression levels of RARG and TACC1 and upper quartile miR-96, compared to the reverse, identified a gene network including several RARG  target genes (e.g. SOX15) that significantly associated with worse disease free survival (hazard ratio 2.23, 95% CI 1.58 to 2.88, p=0.015). In summary, miR-96 targets a RARG network to govern AR signaling, PCa progression and disease outcome.
RNA-seq: HPr1-AR non-malignant, immortalized cell line +/- shRARG, +/- 10nM DHT for 0,24,96 hours

overall_design: 18 samples total, 6 experimental conditions (each in triplicate); shCTL-0hr, shCTL_24hr, shCTL_96hr, shRARG-0hr, shRARG-24hr, shRARG-96hr

first: Dominic

last: Smiraglia

first: Moray

last: Campbell


title: The miR-96 and RARG signaling axis governs androgen signaling and prostate cancer progression VI

status: Public on Jul 20 2018

state: NY

country: USA

type: Genome binding/occupancy profiling by high throughput sequencing

summary:
Expression levels of retinoic acid receptor gamma (NR1B3/RARG, encodes RARG), are commonly reduced in prostate cancer (PCa). Therefore we sought to establish the cellular and gene regulatory consequences of reduced RARG expression, and determine RARG regulatory mechanisms. RARG shRNA approaches in non-malignant (RWPE-1 and HPr1-AR) and malignant (LNCaP) prostate models revealed that reducing RARG levels, rather than adding exogenous retinoid ligand, had the greatest impact on prostate cell viability and gene expression. ChIP-Seq defined the RARG cistrome which was significantly enriched at active enhancers associated with AR binding sites. Reflecting a significant genomic role for RARG to regulate androgen signaling, RARG knockdown in HPr1-AR cells significantly regulated the magnitude of the AR transcriptome. RARG down-regulation was explained by increased miR-96 in PCa cell and mouse models, and TCGA PCa cohorts. Biochemical approaches confirmed that miR-96 directly regulated RARG expression and function. Capture of the miR-96 targetome by biotin-miR96 identified that RARG and a number of RARG  interacting co-factors including TACC1 were all targeted by miR-96, and expression of these genes were prominently altered, positively and negatively, in the TCGA-PRAD cohort. Differential gene expression analyses between tumors in the TCGA-PRAD cohort with lower quartile expression levels of RARG and TACC1 and upper quartile miR-96, compared to the reverse, identified a gene network including several RARG  target genes (e.g. SOX15) that significantly associated with worse disease free survival (hazard ratio 2.23, 95% CI 1.58 to 2.88, p=0.015). In summary, miR-96 targets a RARG network to govern AR signaling, PCa progression and disease outcome.
ChIP-seq: RWPE-1-BAC-RARG-EGFP non-malignant, immortalized cell line infected with BAC-RARG-EGFP, +/- 10nM CD437 for 2 hours

overall_design: 18 samples total, 6 groups (each in triplicate); DMSO-input, DMSO-IgG, DMSO-GFP, CD437-input, CD437-IgG, CD437-GFP

middle: J


title: The miR-96 and RARG signaling axis governs androgen signaling and prostate cancer progression

type: Expression profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Genome-wide maps of EWS-FLI1 binding sites and histone modification in murine Ewing sarcoma cells

status: Public on Jul 25 2018

city: Tokyo

first: Takuro

last: Nakamura

address: 3-8-31 Ariake, Koto-ku

department: The Cancer Institute

country: Japan

institute: Japanese Foundation for Cancer Research

summary: We identified global DNA binding properties of EWS-FLI1 in mouse Ewing sarcoma. GGAA microsatellites were found as binding sites of EWS-FLI1 but with less frequency than that in human Ewing sarcoma, and genomic distribution is not conserved between human and mouse.

overall_design: Examination of binding sites of EWS-FLI1, histone H3K27Ac and H3K27me3 in two Ewing sarcoma cell lines.

first: Rikuka

last: Shimizu


title: Abnormal RNA stability in amyotrophic lateral sclerosis

city: Ann Arbor

first: Mats

last: Ljungman

state: Michigan

address: NCRC, B520 Room 1346 2800 Plymouth Rd.

institute: University of Michigan

summary: Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) share key features, including accumulation of the RNA binding protein TDP-43. TDP-43 regulatesRNA homeostasis, but it remains unclear whether RNA stability is affected in these disorders. We used Bru-seq and BruChase-seq to assessgenome-wide RNA stabilityinALS patient-derived cells,demonstratingprofound destabilization of ribosomal and mitochondrial transcripts. This pattern wasrecapitulatedbyTDP-43 overexpression, suggesting a primary role for TDP-43 in RNA destabilization, and in post-mortem samples from ALS and FTD patients. Proteomics and functional studies illustrated corresponding reductionsin mitochondrial components and compensatory increasesin protein synthesis. Collectively, these observations suggest that TDP-43 deposition leads to targeted RNA instability in ALS and FTD, ultimately causing cell death by disrupting energy production and protein synthesis pathways.

overall_design: Genome-wide assessment of RNA synthesis and stability was done using Bru-seq and BruChase-seq in fibroblasts and human induced pluripotent stem cells (iPSCs) derived from individuals with sporadic (sALS) and familial ALS due to C9orf72 mutations (C9ALS).  There are biological replicates for the iPSC samples.

first: E

middle: M

last: Tank

first: C

last: Figueroa-Romero

first: L

last: Hinder

first: K

last: Bedi

first: H

middle: C

last: Archbold

first: X

last: Li

last: Weskamp

first: N

last: Safren

last: Paez-Colasante

last: Pacut

first: S

last: Thumma

first: M

middle: T

last: Paulsen

last: Guo

first: J

last: Hur

middle: L

last: Feldman

last: Barmada


title: Non-invasive prediction of NAFLD severity: a comprehensive, independent validation of previously postulate serum microRNA biomarkers

status: Public on Jul 19 2018

city: Valencia

first: Joaquin

last: Panadero

state: Valencia

address: Avenida Fernando Abril Martorell, 106 Torre A

department: Genomics

country: Spain

institute: Instituto Investigación Sanitaria La Fe

type: Non-coding RNA profiling by high throughput sequencing

summary: Liver biopsy is currently the only reliable method to establish non‐alcoholic fatty liver disease (NAFLD) severity. However, this technique is invasive and occasionally associated with severe complications. Thus, non‐invasive diagnostic markers for NAFLD are needed. Former studies have postulated 18 different serum biomarker microRNAs with altered levels in NAFLD patients. In this study, we have re‐examined the predictive value of these serum microRNAs and found that only 6 of them (miR‐34a, ‐192, ‐27b, ‐122, ‐197 and ‐30c) are validated in our independent cohort as biomarkers associated with NAFLD severity. Among them, miR‐192, ‐27b and ‐122 are abundantly expressed in liver and confidently detected in serum, and display strong correlations with transaminases. The classification performance of validated miRNAs (and their ratios) for patients with non‐alcoholic steatohepatitis (NASH) is similar to that reached by AST, whereas for advanced fibrosis prediction, the miR‐27b/‐197 ratio demonstrated a good performance and an excellent sensitivity and, along with the FIB‐4 index, may constitute a potent non‐invasive predictive tool.

overall_design: 8 patients classified in SAF1 and SAF2 groups. SAF1 as control and SAF2 as case

first: Mireia

last: López-Riera

first: Isabel

last: Conde

first: Guillermo

last: Quintas

first: Ángela

last: Zaragoza

first: Judith

last: Pérez-Rokas

first: Mario

last: Salcedo

first: José

middle: V

last: Castell

first: Ramiro

last: Jover

first: Salvador

last: Benlloch

first: Laia

last: Pedrola Vidal


title: Bmal1 and Rev-erb⍺ cistromes in skeletal muscle

city: Munich

first: Nina

middle: Henriette

last: Uhlenhaut

state: Bavaria

address: Ingolstädter Landstraße 1,  Neuherberg

department: Molecular Endocrinology

country: Germany

institute: HelmholtzZentrum München - IDC

summary: We mapped the genome-wide binding profiles of BMAL1 and REV-ERB⍺ during peak protein expression of each factor (ZT4, and ZT8, respectively) by ChIP-Seq in gastrocnemius muscles from control C57BL/6J mice.

overall_design: Wildtype 5 month old male C57BL/6J mice maintained on 12:12 light:dark and standard chow diet were sacrificed by cervical dislocation and skeletal muscle was immediately collected and snap frozen in liquid nitrogen. ChIP was performed as previously described (Uhlenhaut et al. 2012). Sheared chromatin was immunoprecipitated either with anti-BMAL1 antibody (ab93806; LOT: GR279846-2), a mixture of anti-REV-ERB⍺ (#23678 and #27401 produced by Ron Evans), or anti-RNA Polymerase II (Biolegends #MMS-126R; clone 8WG16). Two replicates of BMALl1, RNA polymerase 2, and Rev-erb⍺ were made, for a total of 6 ChIP-seq samples + 1 input.

first: Kenneth

middle: A

last: Dyar

first: Ashfaq

last: Mir


title: Nascent transcription of E14, PWWP2AKO, and PWWP2A/B DKO

city: Oxford

first: Guifeng

last: Wei

address: South Parks Road

department: Department of Biochemistry

country: United Kingdom

institute: University of Oxford

summary: Transcriptional regulation by chromatin is a highly dynamic process directed through the recruitment and coordinated action of epigenetic modifiers and readers of these modifications. Using an unbiased proteomic approach to find interactors of H3K36me3, a modification enriched on active chromatin, here we identify PWWP2A and HDAC2 among the top interactors. PWWP2A and its paralog PWWP2B form a stable complex with NuRD subunits MTA1/2/3:HDAC1/2:RBBP4/7, but not with MBD2/3, p66α/β, and CHD3/4. PWWP2A competes with MBD3 for binding to MTA1, thus defining a new variant NuRD complex that is mutually exclusive with the MBD2/3-containing NuRD. In mESCs, PWWP2A/B is most enriched at highly transcribed genes. Loss of PWWP2A/B leads to increases in histone acetylation predominantly at highly expressed genes, accompanied by decreases in Pol II elongation. Collectively, these findings suggest a role for PWWP2A/B in regulating transcription through the fine-tuning of histone acetylation dynamics at actively transcribed genes.

overall_design: In order to explore the influence of PWWP2A/B on nascent transcription, we isolated the 4sU-labelled nascent transcripts, followed by deep sequencing. Three cell lines, E14, PWWP2A KO, and PWWP2A/B double knockout, and three biological replicates.

first: T

last: Zhang

first: G

first: N

last: Brockdorff


title: Mapping the genetic landscape of human cells

city: San Francsico

first: Max

last: Horlbeck

state: CA

address: 1700 4th Street

department: Cellular and Molecular Pharmacology

institute: UCSF

summary: Seminal yeast studies have established the value of comprehensively mapping genetic interactions (GIs) for inferring gene function. Efforts in human cells using focused gene sets underscore the utility of this approach, but the feasibility of generating large-scale, diverse human GI maps remains unresolved. We developed a CRISPR interference platform for large-scale quantitative mapping of human GIs. We systematically perturbed 222,784 gene pairs in two cancer cell lines. The resultant maps cluster functionally related genes, assigning function to poorly characterized genes, including TMEM261, a new electron transport chain component. Individual GIs pinpoint unexpected relationships between pathways, exemplified by a specific cholesterol biosynthesis intermediate whose accumulation induces deoxynucleotide depletion, causing replicative DNA damage and a synthetic-lethal interaction with the ATR/9-1-1 DNA repair pathway. Our map provides a broad resource, establishes GI maps as a high-resolution tool for dissecting gene function, and serves as a blueprint for mapping the genetic landscape of human cells.

overall_design: A library of constructs expressing two sgRNAs and a joint barcode corresponding to the sgRNA pair were cloned and transduced as a pool in K562 or Jurkat cells. Cells were passaged in standard culture conditions and T0 and endpoint samples were harvested. Genomic DNA was processed and sequenced to obtain a readout of sgRNA pair read counts and phenotypes were quantified by comparing representation at T0 and endpoint. Genetic interaction scores and functional gene clustering were derived from these phenotypes.

first: Jonathan

middle: S

last: Weissman

first: Luke

last: Gilbert


title: A selective gut bacterial bile salt hydrolase alters host metabolism

city: Boston

first: Lina

last: Yao

state: Massachusetts

address: 250 Longwood Ave

department: Department of Biological Chemistry and Molecular Pharmacology

institute: Harvard Medical School

summary: The human gut microbiota impacts host metabolism and has been implicated in the pathophysiology of obesity and metabolic syndromes. However, defining the roles of specific microbial activities and metabolites on host phenotypes has proven challenging due to the complexity of the microbiome-host ecosystem. Here, we identify strains from the abundant gut bacterial phylum Bacteroidetes that display selective bile salt hydrolase (BSH) activity. Using isogenic strains of wild-type and BSH-deleted Bacteroides thetaiotaomicron, we selectively modulated the levels of the bile acid tauro-b-muricholic acid in monocolonized gnotobiotic mice. B. thetaiotaomicron BSH mutant-colonized mice displayed altered metabolism, including reduced weight gain and respiratory exchange ratios, as well as transcriptional changes in metabolic, circadian rhythm, and immune pathways in the gut and liver. Our results demonstrate that metabolites generated by a single microbial gene and enzymatic activity can profoundly alter host metabolism and gene expression at local and organism-level scales.

overall_design: 6 biological replicates of mouse distal ileum samples from monocolonizing germfree (GF) mice with the wild-type or knock-out bacterial strains were used for Hiseq sequencing.

first: A

last: Devlin


title: Gene expression analysis in fly ovaries growing from somatic tkv-knockdown gonads driven by c587-GAL4

status: Public on Jul 18 2018

city: Taipei

first: Hweijan

last: Hsu

address: no 128, Sec 2,  Academia Rd. Nankang

department: Institute of Cellular Biology

country: Taiwan

institute: Academia Research

summary:
Purpose: The goal of this study is to analyze genes affected by Tkv knockdown in the developing ovarian soma using RNA-seq.
Ovarian mRNA prolife of one-day-old control flies and flies carring tkv-knockdown ovarian somatic cells duirng development by deep sequence, in duplicate, using Illumina NextSeq 500. The sequence reads that passed quality filters were analyzed at the transcript isoform level using TopHat followed by Cufflinks.

overall_design: Ovarian mRNA prolife of one-day-old control flies and flies carring tkv-knockdown ovarian somatic cells duirng development by deep sequencing, in duplicate, using Illumina NextSeq 500 sequencer.

first: Cheng-Yuan

last: Tseng


title: Translational control through differential ribosome pausing during amino acid limitation in mammalian cells

city: Seattle

first: Arvind

last: Subramaniam

state: WA

address: 1100 Fairview Ave. N., MS #A3-020

institute: Fred Hutchinson Cancer Research Center

summary: Limitation for amino acids is thought to regulate translation in mammalian cells primarily by signaling through the kinases mTORC1 and GCN2. We find that limitation for the amino acid arginine causes a selective loss of tRNA charging, which regulates translation through ribosome pausing at two of six arginine codons. Interestingly, limitation for leucine, an essential and abundant amino acid in protein, results in little or no ribosome pausing. Chemical and genetic perturbation of mTORC1 and GCN2 signaling revealed that their robust response to leucine limitation prevents ribosome pausing, while an insufficient response to arginine limitation led to loss of arginine tRNA charging and ribosome pausing. Codon-specific ribosome pausing decreased protein production and triggered premature ribosome termination without significantly reducing mRNA levels. Together, our results suggest that amino acids which are not optimally sensed by the mTORC1 and GCN2 pathways still regulate translation through an evolutionarily conserved mechanism based on synonymous codon usage.

overall_design: Ribosome profiling was performed in HEK293T, HCT116, or HeLa cells during limitation for leucine or arginine for 3 or 6 hours to determine the effect of limiting single amino acid levels of ribosome elongation kinetics at the cognate codons. The same cell lines grown in nutrient-rich conditions were used as a control. These experiments were repeated in HEK293T cells with 250 nM Torin1, in cells stably expressing a flag-tagged wild-type or Q99L mutant RagB-GTPase or hrGFP, and in a GCN2 knockout cell line to determine the role of the mTORC1 and GCN2 pathways.

first: Alicia

last: Darnell

middle: R

first: Erin

middle: K

last: O'Shea


title: Metabolism regulates muscle stem cell self renewal by connecting the microenvironment and histone acetylation

city: Melbourne

first: James

middle: Gordon

last: Ryall

state: VIC

address: Department of Physiology, The University of Melbourne

department: Physiology

country: Australia

institute: The University of Melbourne

summary: Skeletal muscle contains a resident population of somatic stem cells capable of both self-renewal and differentiation. The signals that regulate this important decision have yet to be fully elucidated. Here we use single cell RNAseq to identify the innate metabolic signature of muscle stem cells. We show that committed muscle progenitor cells exhibit an enrichment of glycolytic and TCA cycle genes and that extracellular monosaccharide availability regulates intracellular citrate levels and global histone acetylation. Muscle stem cells exposed to a reduced (or altered) monosaccharide environment demonstrate reduced global histone acetylation and transcription of myogenic determination factors (including myod1). Importantly, reduced monosaccharide availability was linked directly to increased rates of asymmetric division and muscle stem cell self-renewal. Our results reveal an important role for the extracellular metabolic environment in the decision to undergo self-renewal or myogenic commitment, suggesting local metabolite production may be a therapeutic target to improve muscle regeneration.

overall_design: Examination of single skeletal muscle stem cells that were either freshly isolated or cultured for 96hrs in growth media containing either high glucose (25mM), low glucose (5mM) or galactose (10mM)

first: C

middle: H

last: Ly

first: Jennifer

last: Trieu

first: Shian

last: Su

first: Luyi

last: Tian

first: Daniela

last: Zalcenstein

first: Shalin

last: Naik

first: Matthew

middle: E

last: Ritchie

first: Gordon

last: Lynch

middle: G


title: Solid phase chemistry to covalently and reversibly capture thiolated RNA

middle: Elizabeth

last: Duffy

state: MA

address: 200 Longwood Ave, Goldenson 405

department: Neurobiology

summary: Here, we describe an approach to enrich newly transcribed RNAs from primary mouse neurons using 4-thiouridine (s4U) metabolic labeling and solid phase chemistry. This one-step enrichment procedure captures s4U-RNA by using highly efficient methane thiosulfonate (MTS) chemistry in an immobilized format. Like solution-based methods, this solid-phase enrichment can distinguish mature RNAs (mRNA) with differential stability, and can be used to reveal transient RNAs such as enhancer RNAs (eRNAs) and primary microRNAs (pri-miRNAs) from short metabolic labeling. Most importantly, the efficiency of this solid-phase chemistry made possible the first large scale measurements of RNA polymerase II (RNAPII) elongation rates in mouse cortical neurons. Thus, our approach provides the means to study regulation of RNA metabolism in specific tissue contexts as a means to better understand gene expression in vivo.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: s4U metabolic labeling of RNA in K562 cells and mouse cortical neurons, followed by biochemical enrichment of labeled RNA with solid-phase activated disulfides, and RNA sequencing

first: Daniele

last: Canzio

first: Tom

last: Maniatis

last: Simon


title: Novel differences in renal gene expression in a diet-induced obesity model

city: Baltimore

middle: L.

last: Pluznick

state: Maryland

address: 725 N. Wolfe Street, WBSB 205

institute: Johns Hopkins University

summary: Obesity is a significant risk factor for both chronic kidney disease and end-stage renal disease. To better understand disease development, we sought to identify novel genes differentially expressed early in disease progression. We first confirmed that mice fed a high-fat (HF) diet exhibit early signs of renal injury including hyperfiltration. We then performed RNA-Seq using renal cortex RNA from C57BL6/J male mice fed either HF or control (Ctrl) diet. We identified 1,134 genes differentially expressed in the cortex on HF vs. Ctrl, of which 31 genes were selected for follow-up analysis. This included the 9 most upregulated, the 11 most downregulated, and 11 genes of interest (primarily sensory receptors and G proteins). Quantitative (q)RT-PCR for these 31 genes was performed on additional male renal cortex and medulla samples, and 11 genes (including all 9 upregulated genes) were selected for further study based on qRT-PCR. We then examined expression of these 11 genes in Ctrl and HF male heart and liver samples, which demonstrated that these changes are relatively specific to the renal cortex. These 11 genes were also examined in female renal cortex, where we found that the expression changes seen in males on a HF diet are not replicated in females, even when the females are started on the diet sooner to match weight gain of the males. In sum, these data demonstrate that in a HF-diet model of early disease, novel transcriptional changes occur that are both sex specific and specific to the renal cortex.

overall_design: Identification of differentially expressed genes in the renal cortex of 15 week old mice fed control vs high fat diet for 9 weeks

first: Victoria

last: Halperin Kuhns


title: Paneth cells respond to inflammation and SCF/c-Kit signaling by acquiring stem-like features and contributing to tissue regeneration

city: Rotterdam

first: Mark

last: Schmitt

address: Dr. Molewaterplein 40

department: Pathology

country: Netherlands

institute: Erasmus MC

summary: The experiment aims to analyze the differences in expressed genes in Paneth cells upon acute intestinal inflammation compared to Paneth cells from non-inflamed control mice

overall_design: 8-12 week old Lgr5-EGFP-IRES-CreERT2 mice were subjected to 3% DSS in the drinking water for seven days. Three days after removal of DSS, intestinal tissues were dissected, intestinal crypts isolated and single cell digested. Paneth cells were isolated from single cell suspension by FACS according to previously established gates (Roth et al., 2012).  RNA was isolated from isolated Paneth cells using TRIzolTM Reagent (ThermoFisher Scientific) according to the manufacturer’s instructions. RNA quality and quantity was evaluated on a 2100 Bio-analyzer (Agilent) using the Agilent RNA 6000 Pico Kit (Agilent Technologies). For RNA sequencing analysis, RNA samples were further processed by GATC-Biotech (Konstanz, Germany) according to the INVIEWTM Transcriptome Explore library preparation protocol (Illumina based). Water treated mice were used as controls.

first: Riccardo

last: Fodde

first: Wesley

middle: W

last: van de Geer

first: Harmen

last: van de Werken


title: Inhibition of the Aryl Hydrocarbon Receptor - Polyamine Biosynthesis Axis Suppresses Multiple Myeloma and prostate cancer progression

middle: D

last: Long

summary:
Polyamine inhibition for cancer therapy is, conceptually, an attractive approach but has yet to meet success in the clinical setting. The aryl hydrocarbon receptor (AHR) is the central transcriptional regulator of xenobiotic response. Our study revealed that AHR also positively regulated intracellular polyamine production via direct transcriptional activation of two genes (ODC1 and AZIN1) involved in polyamine biosynthesis and control, respectively. In multiple myeloma patients, AHR levels inversely correlated with survival, suggesting that AHR inhibition may be beneficial for treatment of this disease .We identified clofazimine, an FDA-approved anti-leprosy drug, as a potent AHR antagonist and a suppressor of polyamine biosynthesis. Experiments in a transgenic model of multiple myeloma (Vk*Myc mice) and in immunocompromised mice bearing multiple myeloma cell xenografts, revealed high efficacy of clofazimine comparable to that of bortezomib, a first-in-class proteasome inhibitor used for treatment of multiple myeloma. This study identified a previously unrecognized regulatory axis between AHR and polyamine metabolism and discovered clofazimine as an inhibitor of AHR and a potentially clinically-relevant anti-multiple myeloma agent.
RNA-seq: human multiple myeloma MM1S and human normal fibroblasts WI38 cells -/+ CLF 2-4uM for 24hrs; -/+ shAHR

overall_design: 18 samples total, 9 experimental conditions (each in duplicate): MM1S_pLKO_NT_2hrs ; MM1S_pLKO_NT_24hrs; MM1S_pLKO_CLF_2uM_24hrs; MM1S_shAHR2; MM1S_shAHR_3; WI38_pLKO_NT; WI38_pLKO_CLF_4uM; WI38_shAHR1; WI38_shAHR4

first: Anna

last: Bianchi-Smiraglia

first: Archis

last: Bagati

first: Emily

last: Fink

first: Hayley

last: Affronti

first: Brittany

last: Lipchick

first: Sudha

last: Moparthy

first: Spencer

last: Rosario

first: Shivana

middle: M

last: Lightman

first: Kalyana

first: David

last: Wolff

first: DongHyun

last: Yun

first: Zhannan

last: Han

first: Anthony

last: Polechetti

last: Roll

first: Ilya

middle: I

last: Gitlin

first: Katerina

last: Leonova

first: Aryn

last: Rowsam

first: Eugene

last: Kandel

first: Andrei

last: Gudkov

first: P

last: Bergsagel

first: Kelvin

middle: P

last: Lee

first: Dominic

middle: J

last: Smiraglia

first: Mikhail

last: Nikiforov


title: ATF4 couples MYC-dependent translational activity to bioenergetic demands during tumor progression

city: Philadelphia

first: Priyankara

last: Wickramasinghe

state: PA

address: 3601 Spruce Street

department: Bioinformatics

institute: The Wistar Institute

summary: Dysregulation of the proto-oncogene c-Myc (MYC henceforward) drives malignant progression, but also induces robust anabolic and proliferative programs leading to intrinsic stress. The mechanisms enabling adaptation to MYC-induced stress are not fully understood. We have uncovered an essential role for the transcription factor ATF4 in cell survival following MYC activation. MYC- upregulates ATF4 by activating GCN2 kinase through uncharged tRNAs.  Subsequently, ATF4 co-occupies promoter regions of over 30 MYC target genes, including those regulating amino acid biosynthesis/transport and protein synthesis.  ATF4 is essential for MYC-induced upregulation of the negative translational regulator and mTOR target 4E-BP1 and genetic or pharmacological inhibition of mTOR signaling rescues ATF4 deficient cells from MYC-induced stress. Acute deletion of ATF4 significantly delays MYC-driven tumor progression and increases survival in mouse models. Our results establish ATF4 as a cellular rheostat of MYC-activity, ensuring enhanced translation rates are compatible with survival and tumor progression.

overall_design: ATF4 and MYC CHIP-seq under control and MYC-activated conditions

first: Constantinos

last: Koumenis

first: Feven

last: Tameire


title: RNA-seq analysis identifies different transcriptomic types and developmental trajectories of primary melanomas

city: Leipzig

first: Edith

last: Willscher

address: Haertelstraße 16-18

institute: IZBI

summary: Recent studies revealed trajectories of mutational events in early melanomagenesis, but the accompanying changes in gene expression are far less understood. Therefore, we performed a comprehensive RNA-Seq analysis of laser-microdissected melanocytic nevi (n=23) and primary melanoma samples (n=57) and characterized the molecular mechanisms of early melanoma development. Using self-organizing maps, unsupervised clustering and analysis of pseudotime (PT) dynamics to identify evolutionary trajectories, we describe here two transcriptomic types of melanocytic nevi (N1 and N2) and primary melanomas (M1 and M2). N1/M1 lesions are characterized by pigmentation-type and MITF gene signatures, and a high prevalence of NRAS mutations in M1 melanomas. N2/M2 lesions are characterized by inflammatory-type and AXL gene signatures with an equal distribution of wild type and mutated BRAF and low prevalence of NRAS mutations in M2 melanomas. Interestingly, N1 nevi and M1 melanomas and N2 nevi and M2 melanomas, respectively, cluster together, but there is no clustering in a stage-dependent manner. Transcriptional signatures of M1 melanomas harbour signatures of BRAF/MEK inhibitor resistance and M2 melanomas harbour signatures of anti-PD-1 antibody treatment resistance. Transcriptomic signatures suggest that the epigenetic regulators SMARCA2 and SMARCA4 are differentially involved in the epigenetic programming of the two melanoma types. Pseudotime dynamics of nevus and melanoma samples reflect a switch-like immune-escape mechanism in melanoma development with downregulation of immune genes paralleled by an increasing expression of a cell cycle signature in late-stage melanomas. Taken together, the transcriptome analysis identifies gene signatures and mechanisms underlying development of melanoma in early and late stages with relevance for diagnostics and therapy.

overall_design: RNA-Seq analysis of laser-microdissected melanocytic nevi and primary melanoma samples. Some Melanoma samples are NRAS and/or BRAF mutated.

first: Manfred

last: Kunz

first: Henry

last: Löffler-Wirth

first: Michael

last: Dannemann

first: Gero

last: Doose

first: Janet

last: Kelso

first: Tina

last: Kottek

first: Birgit

last: Nickel

first: Lydia

last: Hopp

first: Jenny

last: Landsberg

first: Steve

last: Hoffmann

first: Thomas

last: Tüting

first: Paola

last: Zigrino

first: Cornelia

last: Mauch

first: Jochen

last: Utikal

first: Mirjana

last: Ziemer

first: Hans-Joachim

last: Schulze

last: Hölzel

first: Alexander

last: Roesch

first: Susanne

last: Kneitz

first: Svenja

last: Meierjohann

first: Anja

last: Bosserhoff

first: Hans

last: Binder

last: Schartl


title: Transcriptional profiling of mouse plasma cells (scRNA-Seq)

city: Tucson

first: Deepta

last: Bhattacharya

state: Arizona

address: 1656 E. Mabel Street, MRB 217

department: Department of Immunobiology

institute: University of Arizona

summary: CD138-high plasma cell subsets were analyzed by single cell RNA-sequencing to identify gene expression differences between long- and short-lived plasma cells. Five different CD138-high subsets were analyzed based upon differential B220 expression and uptake of the fluorescent glucose analog 2NBDG: splenic B220+2NBDG-, B220+2NBDG+, B220-2NBDG-, B220-2NBDG+, and bone marrow B220-2NBDG+ cells.

overall_design: Approximately 1000 cells were analyzed of each subset. Samples were double sorted from and pooled from three  8-12 week old C57Bl6/N male mice.

first: Wing

middle: Y

last: Lam


title: Transcriptional profiling of mouse plasma cells (population RNA-Seq)

summary: CD138-high plasma cell subsets were analyzed by RNA-sequencing to identify gene expression differences between long- and short-lived plasma cells. Five different CD138-high subsets were analyzed based upon differential B220 expression and uptake of the fluorescent glucose analog 2NBDG: splenic B220+2NBDG-, B220+2NBDG+, B220-2NBDG-, B220-2NBDG+, and bone marrow B220-2NBDG+ cells.

overall_design: Approximately 30,000 cells were analyzed of each subset. Three biological replicates of each subset were included. Samples were double sorted from 8-12 week old C57Bl6/N male mice.


title: Towards minimal residual disease-directed therapy in melanoma

city: Leuven

first: Sara

last: Aibar

address: Herestraat 49

department: Center for Brain & Disease Research

country: Belgium

institute: VIB-KU Leuven

summary: Many patients with advanced cancers achieve dramatic responses to a panoply of therapeutics, yet retain minimal residual disease (MRD), which ultimately results in relapse. To gain insights into the biology of MRD we applied single-cell RNA-sequencing to malignant cells isolated from BRAF-mutant patient-derived xenograft (PDX) melanoma cohorts exposed to concurrent RAF/MEK-inhibition.

overall_design: We performed single cell RNA and single cell DNA sequencing from a PDX melanoma model before and on treatment (BRAF&MEK inhibitors). Furthermore, we performed bulk RNAseq of in vitro cultures that were derived from our PDX model, which were treated with BRAF and MEK inhibitors and FACS sorted for GFRA2high and low cells.

first: Daniel

last: Brown

first: Florian

last: Rambow


title: Transcriptional profiling of mouse plasma cells

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

overall_design: Refer to individual Series


title: RNAs Interact with BRD4 to Promote Enhanced Chromatin Engagement and Transcription Activation

city: San Diego

first: Shannon

last: Lauberth

state: California

address: 9500 Gilman Drive #0322, Bonner Hall 3202

department: Biological Sciences

institute: UCSD

summary: In this study, we provide a previously unrecognized convergence between eRNAs and histone acetylation in regulating the chromatin interactions and transcription functions of BRD4.

overall_design: Examination of BRD4 binding prior to and following 16 hr TNF signaling in SW480 colon cancer cells.

first: Homa

last: Rahnamoun

first: Jihoon

last: Lee

first: Zhengxi

last: Sun

first: Hanbin

last: Lu

first: Kristen

last: Ramsey

first: Elizabeth

middle: A

last: Komives


title: Amlexanox targets the IKK-ɛ/TBK1/NF-κB signaling axis to suppress the epithelial-to-mesenchymal transition and prostate cancer metastasis

city: Shanghai

first: zhongzhong

last: ji

state: Shanghai China

address: Huashan Rd. 1940, build 3.  Shanghai,

country: China

institute: SJTU CHINA

summary: Tumor metastasis is the major cause of death for prostate cancer (PCa) patients. However, the treatment options for metastatic PCa are very limited. Epithelial-mesenchymal transition (EMT) has been reported to be an indispensable step for tumor metastasis and is suggested to associate with acquisition of cancer stem cell (CSC) attributes. Herein we establish a EMT reporter system based on firefly or renilla luciferase reporter driven by promoters of CDH1 and VIM genes. High-throughput drug screening from an approved drug library identifies Amlexanox, a commonly used drug to treat aphthous ulcers, to be a strong EMT inhibitor. Amlexanox significantly suppresses PCa cell migration, metastasis and tumor initiation capacity. Mechanistically, the inhibitory effect of Amlexanox on EMT and cell mobility is acted through targeting the IKK-ɛ/ TBK1/ NF-κB signaling pathway. Considering the known safety as well as the pharmacokinetic and pharmacodynamic profile of Amlexanox, our findings suggest a great potential of repositioning Amlexanox as a new anti-metastatic drug for PCa.

overall_design: Examination of the transcriptional difference between Amlexanox and vehicle treated PCa cells

first: Chaping

last: Cheng

first: Zhongzhong

last: Ji


title: PKR senses nuclear and mitochondrial signals by interacting with endogenous double-stranded RNAs

city: Seoul

first: Joha

last: Park

address: 1 Gwanak-ro, Gwanak-gu

country: South Korea

institute: Seoul National University

summary: Protein kinase RNA-activated (PKR) induces immune response by sensing viral double-stranded RNAs (dsRNAs). However, growing evidence suggests that PKR can also be activated by endogenously expressed dsRNAs. Here, we capture these dsRNAs by formaldehyde-mediated crosslinking and immunoprecipitation-sequencing and find that various noncoding RNAs interact with PKR. Surprisingly, the majority of the PKR-interacting RNA repertoire is occupied by mitochondrial RNAs (mtRNAs). MtRNAs can form intermolecular dsRNAs owing to bidirectional transcription of mitochondrial genome and regulate PKR and eIF2α phosphorylation to control cell signaling and translation. Moreover, PKR activation by mtRNAs is counteracted by PKR phosphatases, disruption of which causes apoptosis from PKR overactivation even in uninfected cells. Our work unveils dynamic regulation of PKR even without infection and establishes PKR as a sensor for nuclear and mitochondrial signaling cues in regulating cellular metabolism.

overall_design: PKR fCLIP was performed on S and M phase arrested HeLa cells using PKR & J2 antibodies. Size-matched input samples are also collected and sequenced for each IP sample.

first: Yoosik

last: Kim

first: V

middle: N


title: Single cell RNAseq integrates T cell phenotypes associated with Listeria infection in mice

status: Public on Jul 17 2018

city: Cambridge

middle: Bartholomew

last: Graham

address: 415 Main St.

department: Infectious Disease and Microbiome

institute: Broad Institute

summary: Mice were inoculated with Listeria by i.p. injection on days 0 and 11. On day 18, FSC(high)CD4(+)CD8(-)B220(-)MHCII(-) T cells were FACS-sorted from spleens for single cell RNAseq.

overall_design: 5' digital gene expression was measured by RNAseq

middle: B

first: Ramnik

last: Xavier


title: Inter-tumoral heterogeneity in SCLC is influenced by the cell-type of origin

city: Stanford

first: William

last: Greenleaf

address: 279 Campus Drive, Beckman Center B257

department: Genetics

institute: Stanford University

summary: We describe cell-of-origin as an important influence on suscebtability to changes in chromatin accessibility by Nfib

overall_design: We performed ATAC-seq on primary tumors and metastasis from SCLC mouse model (Rb/p53/p130/mTmG) transduced by Ad-CMV-Cre or Ad-CGRP-Cre

first: Sarah

last: Denny

first: Dian

last: Yang

first: Julien

last: Sage

first: Monte

last: Winslow


title: Spatial Recovery of the Hepatic Lobule Using Single Cell RNA Sequencing

city: Madison

first: James

last: Thomson

state: WI

address: 330 N. Orchard st.

department: Cellular and regenerative biology

institute: Morgridge institute for discovery

summary: Hepatocytes perform different metabolic functions depending on their proximity to the portal triad (periportal) or central vein (perivenous) in the liver lobule. To improve our understanding of the spatial and functional heterogeneity of hepatocytes, we used single cell RNA sequencing (scRNA-seq) and a novel bioinformatics method, Wave-Crest, to reorder hepatocytes along the periportal to perivenous axis. After isolating hepatocytes from an 8-week old male C57BL/6 mouse, performing scRNA-seq, and reordering the cells along the periportal to perivenous axis, Wave-Crest allowed us to identify novel genes differentially expressed along the axis. Using immunocytochemistry, we verified that select novel genes were expressed in the spatial pattern predicted by Wave-Crest. Hence, our study demonstrates a new approach for recovering spatial information from scRNA-seq data.

overall_design: Single cell RNA-seq was performed from primary mouse hepatocytes. A total of 149 cells were sequenced.

first: Morten

last: Seirup


title: RNA-seq analysis of PMA treament with or without LPS on human monocytic cell line THP1

city: Bruxelles

first: Pierre

last: Coulie

address: Avenue hippocrate 74, B1.74.04

department: GECE

institute: de Duve Institute

summary: Transcriptome analysis of the human monocytic cell line THP1 (ATCC TIB-202) treated or not with phorbol myristate acetate (PMA) +/- lipopolysaccharides-lipopolysaccharides binding protein (LPS-LBP)

overall_design: THP1 were treated or not with PMA +/- LPS-LBP

first: Charlotte

last: Six

first: Nathalie

last: Remy

first: Orian

last: Bricard

first: Tiphanie

last: Gomard


title: RNA-seq after siRNA targeting DDX24 applied to iHUVECs cell lines

city: Guangzhou

first: Pengfei

last: Pang

state: Guangdong

address: No. 600 Tianhe Road, Tianhe District

institute: The Third Affiliated Hospital,Sun Yat-Sen University

summary: DDX24 Mutations Associated with Vascular Malformations

overall_design: To detect the vascular-related functions of the causative gene, two short interfering (si)RNAs targeting DDX24 were designed by GenScript bioinformatics tools (https://www.genscript.com/tools/sirna-target-finder) and transfected into immortal human umbilical vein endothelial cells (iHUVECs). A vehicle comprising scrambled siRNA (RIBOBIO) was used as a control. The knockdown efficiency was determined by real-time quantitative PCR and western blotting. RNA sequencing analysis was performed using the siRNA-targeted HUVEC cells to evaluate the effects of the causative gene knockdown on gene expression. Genes involved in cell migration were validated by real-time quantitative PCR.

first: Xiaojun

last: Hu

first: Bin

last: Zhou

first: Junjie

last: Mao

first: Huanhuan

last: He

first: Hong

last: Shan


title: RNA-seq analysis of RPE cells from Sox9 cKO and wild-type control mice

city: Tel Aviv

first: Ran

last: Elkon

address: Ramat Aviv

department: Human Genetics

country: Israel

institute: Tel Aviv University

summary: RNA-seq analysis was applied in triplicates to RPE cells extracted at two developmental time-points (E15.5 and P5) from Sox9-cKO and wild-type control mice

overall_design: Transcriptome analysis of RPE cells, in triplicates, from two genotypes (Sox9-cKO and wild-type) at two developmental time-points (E15.5 and P5)

first: Yamit

last: Cohen-Tayar

first: Rani

first: Ruth

last: Ashery-Padan


title: Fine-tuning Mybl2 levels are required for proper MET process during somatic reprogramming [ATAC-Seq]

city: Birmingham

first: Carl

last: Ward

address: Edgbaston

institute: University of Birmingham

summary: Genome wide data investigating the role of Mybl2 in reprogramming

overall_design:
3 biological replicates of ATAC-seq in control and Mybl2  overexpression conditions at day 3 of reprogramming in doxycycline inducible MEFs.

Two biological replicates for controls at day 0 (two technical replicates of each).

Two biological replicates of Mybl2  overexpression at day 3 with no induction of reprogramming.

first: Paloma

last: Garcia


title: Osterix functions downstream of anti-Müllerian  hormone signaling to regulate Müllerian duct  regression

status: Public on Jul 16 2018

city: Houston

first: Rachel

middle: Diane

last: Mullen

state: TX

address: 1515 Holcombe Blvd

institute: MD Anderson Cancer Center

summary: The goal of this study was to identify potential AMH-induced genes and regulatory networks controlling  regression by RNA-Seq transcriptome analysis of differences in Müllerian Duct mesenchyme between males (AMH signaling on) and females (AMH signaling off) in purified fetal Müllerian Duct mesenchymal cells. This analysis found 82 genes up-regulated in males during MD regression and identified Osterix (Osx)/Sp7, a key transcriptional regulator of osteoblast differentiation and bone formation, as a novel downstream effector of AMH signaling during MD regression.

overall_design: Müllerian Duct mesenchymal cells mRNA profiles from 2-7 embryonic day 14.5  embryos were generated by deep sequencing, in triplicate, using Illumina HiSeq 2000.

first: Richard

middle: R

last: Behringer


title: miR-205 mediates adaptive resistance to MET inhibition via ERRFI1 targeting and raised EGFR signaling

city: Candiolo

first: Davide

last: Cora'

state: TO

department: Dept. of Oncology

country: Italy

institute: Piemonte Orientale University and IRCCS Candiolo Cancer Institute

summary: The onset of secondary resistance represents a major limitation to long term efficacy of target therapies in cancer patients. Thus, the identification of mechanisms mediating secondary resistance is key to the rational design of alternative therapeutic strategies for resistant patients. MiRNA profiling combined with RNA-seq in MET-addicted gastric and lung cancer cell lines led us to identify the miR-205/ERRFI1 (ERBB receptor feedback inhibitor-1) axis as a novel mediator of resistance to MET tyrosine kinase inhibitors (TKIs). In cells resistant to MET-TKIs, increased miR-205 expression determined the downregulation of the EGFR inhibitor ERRFI1, which, in turn, caused EGFR activation and MET-TKI resistance. MiR-205/ERRFI1 driven EGFR activation rendered MET-TKI resistant cells sensitive to combined MET/EGFR inhibition. As a proof of concept of the clinical relevance of this newly identified mechanism of adaptive resistance, we report that a patient with a MET amplified lung adenocarcinoma displayed deregulation of the miR-205/ERRFI1 axis in concomitance with the onset of clinical resistance to anti-MET therapy.

overall_design: RNA-Seq of human EBC-1 and GTL16 cell lines, in two experimental conditions: WT and resistant to crizotinib. Two biological replicates for each condition.

first: Cristina

last: Migliore

first: D

last: Corà

first: Silvia

last: Giordano


title: Shared Nucleotide Flanks Confer Transcriptional Competency to bZip Core Motifs

first: David

last: Steger

address: 12-193 SCTR 3400 Civic Center Blvd

department: Institute of Diabetes, Obesity, and Metabolism

institute: University of Pennsylvania

summary: We report highChIP-exo mapping of sequences occupied by CEBPa (mouse liver), CEBPb (human mesenchymal stem cells, naked human genomic DNA, mouse liver), and recombinant CEBPb-ATF4 heterodimer (naked human genomic DNA).

overall_design:
High resolution TFBS mapping for CEBPb and CEBPa using ChIP-exo

Grant:
R01-DK09854205
Transcriptional and Epigenomic Control of Adipose Tissue Development and Function NIDDK

first: Daniel

last: Cohen


title: Genomic DNA sequencing of budding yeast strains

city: Amsterdam

first: Tessy

last: Korthout

state: Noord-Holland

address: Plesmanlaan 121

department: Gene Regulation

institute: The Netherlands Cancer Institute

summary: We report DNA content of yeast strains that are growing asynchronously or treated with hydroxyurea (HU). We selected two strains from our Epi-Decoder library, that are wild-type but have BAR1 or RPO21 TAP-tagged. This reveals that HU-treated samples have peaks in DNA content representing stalled replication forks.

overall_design: Whole-genome DNA sequencing of two biological replicates with and without hydroxyurea

first: Deepani

last: Poramba


title: Regenerating hair cells in human vestibular sensory epithelia

city: London

first: Michael

last: Lovett

address: National Heart & Lung Institute, Guy Scadding Building

institute: Imperial College London

summary: To investigate alterations in gene expression after Ad2-ATOH1-GFP transduction of human supporting cells, we measured transcriptome changes by RNA-seq. We compared gentamicin-treated tissue (to kill the hair cells) transduced with Ad2-ATOH1-GFP (to promote hair cell production) cultured for 5 days, versus control tissues only treated with gentamicin (cultured for 2 days, 8 days and 14-18 days).

overall_design: Human utricle mRNA profiles after treatement with gentamycin alone or incombination with Atoh1 and untreated controls.

first: Anastasia

last: Filia


title: Effect of FOS and JUN expression in inmortalized human mesenchymal progenitor cells

first: Ander

last: Abarrategi

address: 44 Lincoln's Inn Fields

institute: The Francis Crick Institute

summary: Mesenchymal progenitor cells (MPCs) have been proposed as cells of origin for sarcomas, while c-Fos and c-Jun transcription factors have been proposed has a oncogenes in sarcoma tumors. Here we study the effects of the induction of c-fos or c-Jun expression in immortalized hMPCs. C-Fos expression in these cells not only generated modifications in cell cycle, but also morphological changes, reduced mobility capacity and impaired adipogenic and osteogenic differentiation potentials. Even more interesting, c-Fos expressing immortalized hMPCs generated tumors with chondrogenic phenotype when implanted in immunodeficient mice. In accordance with these results, c-Fos is expressed at protein level in many human bone tumors, specifically in chondrosarcomas (76% c-fos positive samples) and osteosarcomas (49% c-fos positive samples). c-Jun expression in immortalized hMPCs induces increased proliferation and cell transformation and, once implanted in immunodeficient mice, these cells generate fibroblastic osteosarcomas. Doble c-jun and c-fos expression in immortalized hMPCs also induces cell transformation and yields high grade pleomorphic osteosarcoma tumors when implanted in immunodeficient mice.

overall_design: c-DNA was cloned into pOTB7 plasmid, which carries Tomato gene as reporter fluorescent protein. Lentiviral particles were produced by transient calcium phosphate transfection of HEK-293 cells. hMPCs were transduced with those lentiviral particles. Tomato expression was assessed by flow cytometry and c-fos expression was confirmed via qPCR, western blot and immunofluorescence. Cells transduced with lentivirus coding only for fluorescent protein were used as controls. Three independent replicas are provided from each treatment: 3H 1, 3H 2, 3H 3 correspond to hMPCs transduced with control lentiviral particles. 3HF 1, 3HF 2 and 3HF 3 correspond to hMPCs tranduced with lentiviral particles designed for c-fos expression. 3HJ 1, 3HJ 2 and 3HJ 3 correspond to hMPCs tranduced with lentiviral particles designed for c-jun expression. 3HJF 1, 3HJF 2 and 3HJF 3 correspond to hMPCs tranduced with lentiviral particles designed for c-jun and c-fos expression.

first: Javier

last: García-Castro


title: High-Resolution Dissection of Conducive Somatic Cell Reprogramming to Naïve Ground State Pluripotency in Mbd3/NuRD and Gatad2a/NuRD Depleted Systems [smallRNA-Seq]

status: Public on Jul 15 2018

city: Rehovot

first: Noa

last: Novershtern

address: Weizmann Institute

department: Molecular Genetics

institute: Weizmann Institute of Science

summary: Reprogramming of somatic cells to induced pluripotent stem cells provoke immense interest both for clinical applications in regenerative medicine, and in understanding the regulation of cell identity. Previous characterization of the molecular events that underlie this process were limited by the low yield and by the stochastic dynamics of this process. Here we present an in-depth mapping of the molecular events that underlie an efficient and successful reprogramming, covering transcriptional, epigenetic and translational changes in a single day resolution.

overall_design: Mbd3f/-, Gatad2a-/- or wild-type (WT-1,WT-2) MEFs that carry DOX-inducible OKSM cassette were used,  and iPSC reprogramming was initiated by addition of DOX in FBS/LIF medium in 5% pO2 conditions. On day 3.5 after DOX initiation, medium was changed to LIF/KSR-based with the addition of two small molecule inhibitors for MEK/ERK and GSK3 signaling (2i). Cells were harvested every 24 hours until day 8, and processed for library preparation followed by high-throughput sequencing. Mbd3f/-, Gatad2a-/- and WT established iPSC line (after 3 passages or more), and Mbd3f/- or WT V6.5 mouse ESCs were used as positive controls. PolyA RNA-seq measured in each time point. WT-2 cell line is isogenic to Gatad2a-/-

first: Asaf

last: Zviran

first: Yoach

last: Rais

first: Nofar

last: Mor

first: Jacob

last: Hanna


title: Immune cells gene expression from rheumatoid arthritis and healthy donors

city: Fujisawa

first: Shinya

last: Tasaki

state: Kanagawa

address: 26-1, Muraoka-Higashi 2-chome

institute: Takeda Pharmaceutical Company Limited

summary: Sustained clinical remission (CR) without drug treatment has not been achieved in patients with rheumatoid arthritis (RA). This implies a substantial difference between CR and the healthy state, but it has yet to be quantified. We report a longitudinal monitoring of the drug response at multi-omics levels in the peripheral blood of patients with RA. Our data reveal that drug treatments alter the molecular profile closer to that of HCs at the transcriptome, serum proteome and immunophenotype level. Patient follow-up suggests that the molecular profile after drug treatments is associated with long-term stable CR. In addition, we identify molecular signatures that are resistant to drug treatments. These signatures are associated with RA independently of known disease severity indexes and are largely explained by the imbalance of neutrophils, monocytes, and lymphocytes. This high-dimensional phenotyping provides a quantitative measure of molecular remission and illustrates a multi-omics approach to understanding drug response.

overall_design: Transcriptome of sorted immune cells from patients with RA and healthy volunteers.

first: Hiroyoshi

last: Toyoshiba

first: Tsutomu

last: Takeuchi


title: Whole blood gene expression of rheumatoid arthritis

overall_design: Transcriptome of whole blood from RA patients with or without drug treatments (methotrexate, infliximab, and tocilizumab) and healthy volunteers. Serum proteins measured by SOMAscan and peripheral immune cell counts were also measured for the same blood samples and can be found at http://dx.doi.org/10.7303/syn8483403 .


title: KAP1 regulates ERVs in differentiated human cells and contributes to innate immune control

first: Helen

last: Rowe

address: 90 Gower Street

department: Infection and Immunity

institute: University College London

summary: Endogenous retroviruses (ERVs) have accumulated in vertebrate genomes and contribute to the complexity of gene regulation. KAP1 represses ERVs during development by its recruitment to their repetitive sequences through KRAB-zinc finger proteins (KZNFs), but little is known about the regulation of ERVs in differentiated cells. We observed that KAP1 repression of HERVK14C was conserved in differentiated human cells and performed KAP1 knockout to obtain an overview of KAP1 function. Our results show that KAP1 represses ERVs (including HERV-T and HERV-S) and ZNFs, both of which overlap with KAP1 binding sites and H3K9me3 in multiple cell types. Furthermore, this pathway is functionally conserved in primary peripheral blood mononuclear cells. Cytosine methylation that acts on KAP1-regulated loci is necessary to prevent an interferon response, and KAP1-depletion leads to activation of some interferon-stimulated genes. Finally, loss of KAP1 leads to a decrease in H3K9me3 enrichment at ERVs and ZNFs and an RNA-sensing response mediated through MAVS signaling. These data indicate that the KAP1-KZNF pathway contributes to genome stability and innate immune control in differentiated human cells.

overall_design: Dissection of which transposons and genes KAP1 regulates in differentiated human cells

first: Christopher

middle: H

last: Tie

first: Liane

last: Fernandes

first: Lucia

last: Conde

first: Luisa

last: Robbez-Masson

first: Rebecca

last: Sumner

first: Tom

last: Peacock

first: Maria

middle: T

last: Rodriguez-Plata

first: Greta

last: Mickute

first: Robert

last: Gifford

first: Greg

last: Towers

last: Herrero


title: The mediator subunit Med23 serves as a gatekeeper of the myeloid-primed state of hematopoietic stem cells (RNA-Seq)

city: new york

first: Xufeng

last: Chen

state: NY

address: 550 First Avenue, Smilow 1307

institute: NYU Langone Medical Center

summary: To ensure the rapid response to stimuli, some HSCs are specifically prepared (primed) for tasks involving activation and reconstitution. However, the key factors that regulate the primed state of HSCs are largely unknown. Here we report that Med23 controls the formation of myeloid-primed HSCs. Ablation of Med23 in hematopoietic system leads to lymphocytopenia. Moreover, Med23-deficient HSCs undergo myeloid-biased differentiation and lose the self-renewal capacity. Interestingly, Med23-deficient HSCs are in myeloid-primed state and are much easier to be activated in response to physiological stresses. Mechanistically, Med23 plays essential roles in maintaining stemness genes expression and suppressing myeloid lineage genes expression. Med23 is down-regulated in HSCs and Med23 deletion results in better survival under myeloablative stress. Altogether, our findings identified Med23 as a gatekeeper of the myeloid-primed state of HSCs, thus providing unique insights into the relationship among Med23-mediated transcriptional regulations, the myeloid-primed state of HSCs and HSC activation upon stresses.

overall_design: Med23-deficient HSCs Vs WT HSCs. For RNAseq, We have 3 replicates for Med23-deficient cases, including KO1_RNA, KO2_RNA, KO3_RNA, as well as 3 replicates for wild type as controls, WT1_RNA, WT2_RNA, WT3_RNAseq. For ATAC, we have 2 replicates for med23 knockout cases, including KO1_ATAC, KO2_ATAC, as well as 2 replicates for wild type as controls, WT1_ATAC, WT2_ATAC.

first: X

first: J

last: Zhao

last: Liu


title: Identification of an Early Unipotent Neutrophil Progenitor with Pro-Tumoral Activity in Mouse and Human Bone Marrow

city: La Jolla

first: Huy

middle: Q

last: Dinh

state: CA

address: 9420 Athena Circle

department: Division of Inflammation Biology

institute: La Jolla Institute

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.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.

overall_design: Expression profiling by high throughput sequencing for sorted immune cells in mouse bone marrow

first: Yanfang

last: Zhu

first: Catherine

middle: C

last: Hedrick


title: The mediator subunit Med23 serves as a gatekeeper of the myeloid-primed state of hematopoietic stem cells

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

overall_design: Refer to individual Series


title: Transcriptomic Analysis of Gonadal Adipose Tissue in Male Mice Exposed Perinatally to 2,2',4,4'- Tetrabromodiphenyl Ether (BDE-47)

city: Amherst

first: Alexander

last: Suvorov

address: 149A Goessman, 686 N.Pleasant Str

department: Environmental Health Sciences

institute: University of Massachusetts

summary: For the majority of lipophilic compounds adipose tissue is traditionally considered as storage depot and only rarely as a target organ. Meanwhile, abnormalities in adipose tissue physiology induced by chemical exposures may contribute to the current epidemic of obesity and metabolic diseases. Polybrominated diphenyl ethers (PBDEs) is a group of lipophilic flame retardants found in majority of human samples in North America. Their ability to alter physiology of adipose tissue is unknown. We exposed pregnant mice to 0.2 mg/kg body weight/day of BDE-47 perinatally. Transcriptomic changes in gonadal adipose tissue were analyzed in male offspring using RNA-seq approach with subsequent bioinformatic analysis. Genes of coagulation and complement cascade, de novo lipogenesis, and xenobiotic metabolism were altered in expression in response to BDE-47 exposure. The affected molecular network included the following hubs: PPARα, HNF1A and HNF4. These findings suggest that adipose tissue should be considered a target tissue for BDE-47, in addition to its role as a storage depot. This study also builds a background for a targeted search of sensitive phenotypic endpoints of BDE-47 exposure, including lipid profile parameters and coagulation factors in circulation. Additional studies are needed to investigate the role of PBDEs as an obesogen.

overall_design: CD-1 mice were exposed to vehicle (corn oil) or 0.2 mg/kg body weight/day of 2,2',4,4'- Tetrabromodiphenyl Ether from gestation day 8 till postpartum day 21. Gonadal adipose tissue was collected from male pups (4 exposed and 3 control) and analysed using RNA-seq approach.


title: Small RNAs are trafficked from the epididymis to developing mammalian sperm

city: Santa Cruz

first: Upasna

last: Sharma

address: 1156 High St

department: MCD Biology

institute: UCSC

summary: RNAs present in mature mammalian sperm are delivered to the zygote at fertilization, where they have the potential to affect early development. The biogenesis of the small RNA payload of mature sperm is therefore of great interest, as it may be a target of signaling pathways linking paternal conditions to offspring phenotype. Recent studies have suggested the surprising hypothesis that the small RNA payload carried by mature sperm may include RNAs that were not synthesized during testicular spermatogenesis, but that are instead delivered to sperm during the process of post-testicular maturation in the epididymis. To further test this hypothesis, we characterized small RNA dynamics during testicular and post-testicular germ cell maturation in mice, confirming and extending prior observations that testicular germ cell populations carry extremely low levels of tRNA fragments (tRFs), which only become highly abundant only after sperm have entered the epididymis. We examined the sperm RNA repertoire in greater detail, finding that the majority of 5’ tRNA fragments carry a 2’-3’ cyclic phosphate at their 3’ end, pointing to a role for RNaseA or T-family nucleases in tRNA cleavage in the male reproductive tract. The process of small RNA delivery to sperm can be recapitulated in vitro, as caput epididymosomes deliver small RNAs including tRFs and microRNAs to mature testicular spermatozoa. Finally, to definitively identify the tissue of origin for small RNAs in sperm, we carried out tissue-specific metabolic labeling of RNAs in intact mice, finding that mature sperm carry small RNAs that were originally synthesized in the somatic cells of the epididymis. Taken together, our data demonstrates that soma-germline small RNA transfer occurs in male mammals, most likely via vesicular transport from the epididymis to maturing sperm.

overall_design: Small RNA sequencing of various male reproductive tract tissues such as testis and epididymis, and sperm during testicular and post-testicular maturation

first: Fengyun

last: Sun

first: Colin

last: Conine

first: Shweta

last: Kukreja

first: Brian

last: Reichholf

first: Veronika

last: Herzog

first: Stefan

middle: L

last: Ameres

first: Oliver

last: Rando


title: Oxysterol signatures distinguish age-related macular degeneration from physiologic aging

city: Saint Louis

first: Jonathan

last: Lin

state: MO

address: 660 S. Euclid Ave, Campus Box 8096

institute: Washington University School of Medicine

summary: Macrophage aging is pathogenic in numerous diseases, including age-related macular degeneration. Although prior studies have explored the functional consequences of macrophage aging, less is known about its cellular basis or what defines the transition from physiologic aging to disease. The purpose of this experiment was to characterize the transcriptomic changes associated with macrophage aging.

overall_design: We prepared 24 samples (N=4/group with 3 treatments and 2 ages). We identified 4 outliers (Yut_1, Yac_1, Oac_1, Oox_1) after principal component analysis and hierarchical clustering, which we omitted from further analysis.

first: Abdoulaye

last: Sene


title: The mediator subunit Med23 serves as a gatekeeper of the myeloid-primed state of hematopoietic stem cells (ATAC-Seq)

first: Xufeng

last: Chen

overall_design: Med23-deficient HSCs Vs WT HSCs. For RNAseq, We have 3 replicates for med23 knockout cases, including KO1_RNA, KO2_RNA, KO3_RNA, as well as 3 replicates for wild type as controls, WT1_RNA, WT2_RNA, WT3_RNA. For ATAC, we have 2 replicates for med23 knockout cases, including KO1_ATAC, KO2_ATAC, as well as 2 replicates for wild type as controls, WT1_ATAC, WT2_ATAC.


title: Characterization of the deregulation of the RNAs associated to the RISC complex in skeletal muscle of DM1 patients (RISC-IP long RNA)

city: Milan

first: Fabio

last: Martelli

address: via morandi 30

institute: IRCCS-Policlinico San Donato

summary: DM1 (OMIM #160900) is a chronic, slowly progressing multisystemic disease, with symptoms that include loss of muscle strength, myotonia and excessive fatigue. DM1 is caused by a dynamic expansion of CTG repeats, ranging from 50 to several thousands, in the 3’ untranslated region of the DMPK gene. Characteristic molecular features of the disease have been associated with a toxic RNA gain of function of the CUG expansions. Expanded CUG repeats have been demonstrated to be toxic per se in several cell types and animal models disrupting gene transcription and pre-mRNA alternative splicing. Deep sequencing is here used to characterize the deregulation of the RNAs associated to the RISC complex in the skeletal muscle of DM1 patients.

overall_design: 2 conditions are available, Control and DM1. Each condition is represented by three samples from individual patients for a total of 6 samples. The samples contain long RNA-Seq data obtained by deep sequencing of the RNA enriched by immunoprecipitating the RISC complex.

first: Alessandra

last: Perfetti

first: Matteo

last: Carrara

first: Jose

last: Garcia-Manteiga


title: Characterization of the deregulation of the RNAs associated to the RISC complex in skeletal muscle of DM1 patients (Input small RNA)

overall_design: 2 conditions are available, Control and DM1. Each condition is represented by three samples from individual patients for a total of 6 samples. The samples contain small RNA-Seq data obtained by deep sequencing of the input RNA from the cell lysate. The samples used are identical to those submitted for immunoprecipitation.


title: Characterization of the deregulation of the RNAs associated to the RISC complex in skeletal muscle of DM1 patients


title: The Transcription factor Zeb2 ia required to maintain tissue-specific identities of macrophages [bulk RNA-seq]

status: Public on Jul 14 2018

city: Zwijnaarde

first: Liesbet

last: Martens

address: Technologiepark 927

department: VIB Inflammation Research Center

institute: VIB-University of Ghent

summary: Microarray, Bulk RNA Sequencing and Single cell RNA Sequencing of different murine tissue-resident macrophage populations to assess role of Zeb2 and LXRa

overall_design: Bulk RNA Seq = Replicates of ZEB2KO alveolar macrophages (sorted on CD11b expression) and ZEB2KO Kupffer cells (sorted on Tim4 expression) were compared with replicates of WT Alveolar Macs and Kupffer cells. This data was used to define core Alveolar Mac and Kupffer Cell profiles as well as core profies of other macrophages from previously published data.

first: Charlotte

last: Scott

first: Wouter

last: T'Jonck

first: Sofie

last: De Pricjk

first: Niels

last: Vandamme

first: Yvan

last: Saeys

first: Martin

last: Guilliams


title: The homeobox transcription factor HB9 induces senescence and blocks differentiation in hematopoietic stem and progenitor cells

city: Homburg (Saar)

first: Marc

last: Remke

state: Saarland

address: Kirrberger Straße 100, Gebäude 9

department: Pediatric Translational Oncology Lab

institute: Universitätsklinikum des Saarlandes

summary: The translocation t(7;12)(q36;p13) occurs in infants and very young children with AML and usually has a fatal prognosis. Whereas the transcription factor ETV6, located at chromosome 12p13, has largely been studied in different leukemia types, the influence of the translocation partner HB9 (chr. 7q36), is still unknown. This is particularly surprising as ectopic expression of HB9 is the only recurrent molecular hallmark of translocation t(7;12) AML. We investigated the influence of HB9 as a potential oncogene on cell proliferation and cell cycle in vitro, as well as on hematopoietic stem cell differentiation in vivo using murine and human model systems.   We show, that HB9 induces premature senescence in human HT1080 and murine NIH3T3 cells, providing for the first time evidence for an oncogenic potential of HB9. Furthermore, HB9-transduced primary murine hematopoietic stem and progenitor cells underwent a profound differentiation arrest and accumulated at the megakaryocyte/erythrocyte progenitor stage, resulting in a premalignant myeloid cell population in vivo. Concomitantly, HB9 expression upregulates erythropoiesis-related genes in primary human hematopoietic stem and progenitor cells, and enriches gene expression profiles for cell cycle and mitosis-related biological processes.   In summary, the novel findings of HB9 dependent premature senescence and perturbed hematopoietic differentiation shed light on the oncogenic properties of HB9 in translocation t(7;12) AML and offer novel targets for therapeutic intervention.

overall_design: CD34+ cells were transduced with either GFP or HB9

first: Daniel

last: Picard

first: Deborah

last: Ingenhag

first: Julia

last: Hauer


title: Multi-omics monitoring of drug response in rheumatoid arthritis.

first: Shinya

last: Tasaki

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

Sustained clinical remission (CR) without drug treatment has not been achieved in patients with rheumatoid arthritis (RA). This implies a substantial difference between CR and the healthy state, but it has yet to be quantified. We report a longitudinal monitoring of the drug response at multi-omics levels in the peripheral blood of patients with RA. Our data reveal that drug treatments alter the molecular profile closer to that of HCs at the transcriptome, serum proteome and immunophenotype level. Patient follow-up suggests that the molecular profile after drug treatments is associated with long-term stable CR. In addition, we identify molecular signatures that are resistant to drug treatments. These signatures are associated with RA independently of known disease severity indexes and are largely explained by the imbalance of neutrophils, monocytes, and lymphocytes. This high-dimensional phenotyping provides a quantitative measure of molecular remission and illustrates a multi-omics approach to understanding drug response.

overall_design:
Refer to individual Series

Transcriptome of whole blood and sorted immune cells from patients with or without drug treatments (methotrexate, infliximab, and tocilizumab) and healthy volunteers. Serum proteins measured by SOMAscan and peripheral immune cell counts were also measured for the same blood samples and can be found at http://dx.doi.org/10.7303/syn8483403 .


title: The Transcription factor Zeb2 ia required to maintain tissue-specific identities of macrophages [single-cell RNA-seq]

overall_design: SIngle cell RNA Seq = WT or Zeb2 KO macrophages from different tissues performed with the 10X Genomics platform. KOs were compared with WT controls and DE genes compared across tissues.


title: Expression data of A2058-TET2WT, A2058-TET2M, and Mock cells treated under high-g and normal-g

city: Boston

first: Di

last: Wu

address: 221 longwood Ave, EBRC (LMRC) 222A

department: Endocrinology

institute: The Brigham and Women's Hospital

summary:
Diabetes is a complex metabolic syndrome characterized by prolonged high blood glucose levels. It is known that diabetes is associated with an elevated risk of cancer, however, the underlying molecular mechanisms are largely unknown. In particular, it remains unclear as to how hyperglycemia may affect epigenetic checkpoints and tumor suppressor pathways, thus enabling oncogenic transformation. Here we show that long-term hyperglycemic conditions adversely impact the anti-tumor epigenetic mark DNA 5-hydroxymethylcytosine (5hmC) through direct regulation of the tumor suppressor and DNA 5mC hydroxymethylase, TET2. We identify TET2 as a novel substrate of the AMP-activated kinase (AMPK).
The microarray data is used to study genes that are differentially regulated by glucose through TET2.

overall_design: A2058-TET2WT, A2058-TET2M, and Mock cells were grown under high-g (4.5g/L) and normal-g (1g/L) DMEM medium for 7 days, with splitting every 2-3 days during growth. RNA was extracted and hybridized on Affymetrix microarrays.

last: Hu

first: Ruitu

last: Lv


title: Cancer associated mutants of eIF1A impair Rps3/Rps10 binding and enhance scanning of cell cycle genes

status: Public on Jul 13 2018

first: Dena

last: Leshkowitz

department: Bioinformatics Unit, Life Sciences Core Facilities


title: Cancer associated mutants of eIF1A impair Rps3/Rps10 binding and enhance scanning of cell cycle genes [Mars-seq]

summary: The Ribo-seq analysis demonstrated that eIF1A is predominantly essential for translation of genes with long 5'UTR genes including cell proliferation and cell cycle progression genes. eIF1A depletion causes broad stimulation of initiation in 5’UTRs at near-cognate AUG codons that diminshes the translation initiation fidelity

overall_design: Mars-seq analysis was performed either in contol or eIF1A knockdown MEFs

first: Urmila

last: Sehrawat

first: Shaked

last: Ashkanazi

first: Femke

last: Koning

first: Gil

last: Stelzer

first: Rivka

last: Dikstein


title: SAP97 Regulates Behavior and Expression of Schizophrenia Risk Enriched Gene Sets in Mouse Hippocampus

city: Philadelphia

first: Preetika

last: Gupta

address: 3400 Civic Center Blvd

summary: Analysis of DE genes between littermate control animals and animals lacking SAP97 specifically in the nervous system

overall_design: 4 control vs 4 cKO animals at 30 million read depth

first: Ogul

middle: E

last: Uner

first: Soumyashant

last: Nayak

first: Gregory

last: Grant

first: Robert

middle: G

last: Kalb


title: Chromatin run-on and sequencing maps the transcriptional regulatory landscape of glioblastoma multiforme

status: Public on Jul 30 2018

city: Ithaca

first: Charles

last: Danko

address: Hungerford Hill Rd

department: Baker Institute

institute: Cornell University

summary: The human genome encodes a variety of poorly understood RNA species that remain challenging to identify using existing genomic tools. We developed chromatin run-on and sequencing (ChRO-seq) to map the location of RNA polymerase using virtually any input sample, including samples with degraded RNA that are intractable to RNA-seq. We used ChRO-seq to map nascent transcription in primary human glioblastoma (GBM) brain tumors. Whereas enhancers discovered in primary GBMs resemble open chromatin in the normal human brain, rare enhancers activated in malignant tissue drive regulatory programs similar to the developing nervous system. We identified enhancers that regulate genes characteristic of each known GBM subtype, identified transcription factors that drive them, and discovered a core group of transcription factors that control the expression of genes associated with clinical outcomes. This study reveals the transcriptional basis of GBM and introduces ChRO-seq to map regulatory programs contributing to complex diseases.

overall_design: We devised ChRO-seq to map the location of RNA polymerases in samples that are difficult to analyze using existing genomic tools. We tested this new strategy using paired PRO-seq and ChRO-seq experiments in Jurkat T-cells. This GEO entry contains paired PRO-seq and ChRO-seq data.

first: Edward

last: Rice


title: A novel enhancer regulates MGMT expression and promotes temozolomide resistance in glioblastoma

city: New York

first: zhiguo

last: zhang

address: 1130 St. Nicholas Avenue

department: Pediatric and Genetics and Development

institute: Columbia University

summary: Temozolomide (TMZ) has been used for the treatment of glioblastoma (GBM) since last decade, but its treatment benefits are limited by acquired resistance, a process that remains incompletely understood. Here we report that a novel enhancer, located between the promoters of Ki67 and O6-methylguanine-DNA-methyltransferase (MGMT) genes, is activated in TMZ-resistant patient-derived xenograft (PDX) lines as well as in recurrent tumor samples. Activation of the enhancer correlates with increased MGMT expression, a major known mechanism for TMZ resistance. We show that forced activation of the enhancer in cell lines with low MGMT expression results in elevated MGMT expression. Deletion of this enhancer in cell lines with high MGMT expression leads to reduced levels of MGMT and Ki67, increased TMZ sensitivity and impaired proliferation. Together, these studies uncover a novel mechanism that regulates MGMT expression, confers TMZ resistance and potentially regulates tumor proliferation.

overall_design: To analyze how epigenetic landscapes are altered during the development of TMZ resistance, we performed H3K4me1, H3K4me3, H3K9me3, H3K9ac, H3K27ac and H3K36me3 ChIP-seq on GBM12-5199 and GBM12-3080 tissue samples. We also performed RNA-seq on GBM12-5199 and GBM12-3080 tissue samples.

first: Xiaoyue

last: Chen

first: Haiyun

last: Gan

first: Jeong

last: Lee

first: Gaspar

last: Kitange

first: Jann

last: Sarkaria

first: Zhiguo

last: Zhang


title: Transcriptome-wide analysis of HuD (ELAVL4) RNA interactome and post-transcriptional regulation in a motor neuron cell line

city: Povo

first: Toma

last: Tebaldi

state: Trento

address: via Sommarive n. 9

department: Centre for Integrative Biology

institute: University of Trento

summary:
The neuronal ELAV-like RNA-binding protein HuD (ELAVL4) plays important roles in multiple post-transcriptional regulatory processes, including RNA stability, transport and translation. Besides its functional role in neuronal plasticity, HuD has been implicated in peripheral axon injury recovery and motor neuronal function.  The characterization of HuD specific interactions has always been a challenging task due to the high similarity of sequence and structure with the other members of the ELAVL family, and the lack of specific antibodies. To selectively identify in vivo HuD binding sites, we adapted the CRAC protocol (cross-linking and analysis of cDNAs), originally developed for yeast, to be used with mouse motor neuron NSC-34 cells engineered with inducible tagged HuD. In parallel, to characterize the role of HuD in post-transcriptional regulation, we also profiled the transcriptome (total RNA, RNA-Seq), the translatome (polysomal RNA, POL-Seq) and the alternative polyadenilation (APA, 3’end mRNA sequencing) of HuD induced and control NSC-34 cells.
Keywords: HuD, Elavl4, Y3, Rny3, CRAC, RNA interactome, RNA binding, NSC-34, transcriptome profiling, translatome profiling, RNA-Seq, POL-Seq, motor neuron

overall_design: In this study, NSC-34 cell lines were stably transduced with an inducible vector expressing tagged HuD. To identify the RNA interactome of HuD, the CRAC experiment was performed in biological triplicate, and control experiments with an empty vector were performed in parallel, to filter out any aspecific signal. Transcriptome profiling (total RNA, RNA-Seq) and translatome profiling (polysomal RNA, POL-Seq) were performed in HuD induced and control NSC-34 cells, in biological duplicate. Alternative polyadenilation profiling (APA) was performed by 3’end mRNA sequencing of HuD induced and control NSC-34 cells, in biological triplicate.

first: Paola

last: Zuccotti

first: Daniele

last: Peroni

first: Tatiana

last: Dudnakova

first: Roberto

last: Bertorelli

first: Veronica

last: De Sanctis

last: Fassan

first: David

last: Tollervey

first: Alessandro

last: Quattrone


title: characterization of mesenchymal stem cell derived from N1-12 and 201B7 via mesoderm and neuroepithelia

city: kumamoto

first: Shinya

last: Eto

address: honjo 2-2-1

institute: Kumamoto univercity

summary:
We established new methods for differentiating hiPSCs into MSCs via mesoderm-like and neuroepithelium-like cells. We named MSCs via mesoderm-like as PSP-MSC, and MSCs via neuroepithelium-like cells.
To further characterize our PSP-MSCs and RA-Pα-MSCs, we compared their genome-wide expression patterns with those of human iPSCs and BM-MSCs (PRC-010; Bay bioscience) using DNA microarrays.

overall_design: Gene expression of N1-12, BM-MSC, MSC derived from N1-12 and 201B7 via mesoderm, MSC derived from N1-12 and 201B7 via neuroepithelia by Affymetrix Human Genome U133 Plus 2.0 Array

first: Takumi

last: Era


title: Functional CRISPR Screen Identifies AP1-associated Enhancer regulating FOXF1 to modulate Oncogene-Induced Senescence

city: Tel Aviv

first: Ran

last: Elkon

address: Ramat Aviv

institute: Tel Aviv University

summary: We carried out CRISPR-Cas9 functional screen focused on AP-1 bound enhancers that are activated upon oncogenic stress. The screen discovered Enh.AP1.OIS1 - an enhancer bound by AP1 that is required for activation of oncogene-induced senescence (OIS) response. We applied RNA-seq analysis to RASG12V-activated BJ cells transduced with either sgRNAs targeting AP-1 bound enhancer (sgRNA-69 or sgRNA71), or sgRNA targeting the target gene FOXF1 or non-targeting control sgRNA (sgNT)

overall_design: RNA-seq was applied to oncogenic RASG12V-induced BJ cells genetically manipulated using CRISPR-Cas9

first: Ruiqi

last: Han

first: Rani

first: Reuven

last: Agami


title: Ribosome profiling and transcriptome profiling of MCF-7 cells

summary:
We performed ribosome profiling (RIBO-seq) and transcriptome profiling (RNA-seq) to monitor RNAs associated with ribosome in the MCF-7 cell model
Keywords: ribosome profiling, translation, MCF-7

overall_design: We performed ribosome profiling (RIBO-seq) and transcriptome profiling (RNA-seq) on MCF-7 cells

last: Bernabò

first: Fabio

last: Lauria

first: Gabriella

last: Viero


title: miRNA changes induced by cyclic and chronic hypoxia in human Ecs

status: Public on Jul 12 2018

city: GDANSK

first: Rafal

last: Bartoszewski

address: Hallera 107

country: Poland

institute: Medical University of Gdansk

summary:
We report the results NGS based of miRNA profiling in human endothelial cells under chronic and cyclic hypoxia
Solid tumor microenvironments are often subjected to various levels of hypoxia. Although regulation of gene expression has been examined extensively, most studies have focused on chronic hypoxia. The tumor microenvironment, however, experience waves of hypoxia and reoxygenation that stimulates the expression of pro-angiogenic factors that promote blood vessel formation. In this study, we examined human umbilical vascular endothelial cells (HUVECs) under waves of intermittent (cyclic hypoxia) to determine how this process compares to chronic hypoxia, and more importantly, how this influences the microRNA profiles that potentially affect the posttranscriptional regulation of angiogenic genes. The rationale for these studies is that cancer cells subjected to cyclic hypoxia appear to have increased metastatic potential and endothelial cells exhibit a higher radiation resistance and greater migration potential. This indicates that the gene regulatory networks in cyclic hypoxia may be different from chronic hypoxia. Here we examined the consequences of cyclic hypoxia on miRNA gene expression and how these changes in miRNA expression could influence angiogenesis. Using Next Generation Sequencing, our results demonstrate that cyclic hypoxia has very different effects on the miRNA networks compared to chronic hypoxia, the in silico predicted effects on the certain mRNA target genes are more similar than might be expected. More importantly, these studies indicate that identifying potential miRNAs (including hsa-miR-19a-5p) as therapeutic targets for inhibiting angiogenesis and tumor progression will require this type of physiologically relevant analysis.

overall_design: mRNA profiles of HUVECs exposed to cyclic and chronic chypoxia were generated by deep sequencing.


title: Characterization of the deregulation of the RNAs associated to the RISC complex in skeletal muscle of DM1 patients (RISC-IP small RNA)

last: Martelli

overall_design: 2 conditions are available, Control and DM1. Each condition is represented by three samples from individual patients for a total of 6 samples. The samples contain small RNA-Seq data obtained by deep sequencing of the RNA enriched by immunoprecipitating the RISC complex.

first: Alessandra

last: Perfetti

first: Matteo

last: Carrara

first: Jose

middle: M

last: Garcia-Manteiga


title: Epigenetic regulation of brain region-specific microglia clearance activity

first: Pinar

last: Ayata

state: New York

address: 1425 Madison Ave

department: Neuroscience

institute: Icahn School of Medicine at Mt Sinai

summary: The rapid elimination of dying neurons and non-functional synapses in the brain is carried out by microglia, the resident myeloid cells of the brain. Here we show that microglia clearance activity in the adult brain is regionally regulated and depends on the rate of neuronal attrition. Cerebellar, but not striatal or cortical, microglia exhibited high levels of basal clearance activity, which correlated with an elevated degree of cerebellar neuronal attrition. Exposing forebrain microglia to apoptotic cells activated gene expression programs supporting clearance activity. We provide evidence that the Polycomb repressive complex 2 (PRC2) epigenetically restricts the expression of genes that support clearance activity in striatal and cortical microglia. Loss of PRC2 led to the aberrant activation of a microglia clearance phenotype, which triggers changes in neuronal morphology and behavior. Our data highlight a key role of epigenetic mechanisms in preventing microglia-induced neuronal alterations that are frequently associated with neurodegenerative and psychiatric diseases.

overall_design: Microglial RNAs or nuclei were isolated based on the eGFP-L10a expression of ribosomes and fluorescence of newly formed ribosomes in the microglia nucleoli, respectively

first: Ana

last: Badimon

first: Hayley

last: Strasburger

first: Mary Kaye

last: Duff

first: Sarah

last: Montgomery

first: Yong-Hwee

last: Loh

first: Anja

last: Ebert

first: Anna

last: Pimenova

first: Brianna

last: Ramirez

first: Andrew

last: Chan

first: Josefa

last: Sullivan

first: Immanuel

last: Purushothaman

first: Joseph

last: Scarpa

first: Alison

last: Goate

first: Meinrad

last: Busslinger

first: Li

last: Shen

first: Bojan

last: Losic

first: Anne

last: Schaefer


title: Turbulence activates platelet biogenesis to enable clinical scale manufacturing from human iPS cells

city: Kyoto

first: Koji

last: Eto

address: 53 Kawahara-cho, Shogoin Kawahara-cho, Sakyo-ku

department: CiRA

institute: Kyoto University

summary: To identify the genes whose expression levels are changed in VerMES and dish culture, we performed global gene expression analysis of immortalized megakaryocyte cell lines delived from hiPSC.

overall_design: RNAs were extracted from imMKCL (CL-7) and analyzed on Affymetrix microarrays.

first: Y

last: Ito

first: S

last: Nakamura

first: T

last: Shigemori

last: Yamamoto


title: GRHL2-dependent enhancer switching maintains a pluripotent stem cell transcriptional subnetwork after exit from naïve pluripotency [ESC & EpiLC]

city: Stanford

first: Amy

last: Chen

address: 279 Campus Drive West

institute: Stanford University

summary: During early development, pluripotent cells of the epiblast show extensive rewiring of enhancers with little associated change in gene expression. The mechanisms underlying and purpose of this rewiring are largely unknown. Here we identified a transcription factor, GRHL2, that is both necessary and sufficient to activate latent enhancers during the transition from naïve embryonic stem cells (ESC) to primed epiblast cells (EpiC). GRHL2 is necessary to maintain expression of its targets in EpiCs. However, these genes are already expressed at equivalent levels in ESCs, suggesting these genes switch enhancer usage during the transition. Identification of alternative enhancers driving these genes in ESCs uncovered an enrichment for the ESC-specific KLF transcription factors. While many KLF targets remain expressed in EpiCs, GRHL2 only regulates a specific subset promoting an epithelial program. These data suggest a model where a large naïve-specific transcriptional network is partitioned into smaller networks to uncouple their regulation in EpiCs, providing more flexibility in gene regulation during lineage specification.

overall_design: RNA-seq in wildtype embryonic stem cells (ESCs) and wildtype epiblast-like cells (EpiLCs)

first: Robert

last: Blelloch


title: GRHL2-dependent enhancer switching maintains a pluripotent stem cell transcriptional subnetwork after exit from naïve pluripotency [WT & KO]

overall_design: RNA-seq in wildtype and KLF2/KLF4 double knockout embryonic stem cells (ESCs)


title: Synovial fluid and blood neutrophils from rheumatoid arthritis patients and matched healthy controls

status: Public on Jul 11 2018

city: Parkville

first: CSL

last: Research

state: VIC

address: 30 Flemington Rd

institute: CSL Limited / bio21 Institute

summary: Synovial fluid and blood neutrophils from 7 rheumatoid arthritis patients and 5 matched healthy controls

overall_design: Experiment designed to compare gene signatures of neutrophils isolated from site of pathiology (synovial fluid (SF)) and from peripheral blood (PB) of RA patients.  Neutrophils were isolated from SF and PB from the same individuals (n = 3) to provide paired sample analysis.  Neutrophils were also isolated from PB from healthy donors as controls. There are two technical replicates for each donor and tissue.  The technical replicates were made by running the same library in two lanes.

first: Gabrielle

last: Goldberg

first: Milica

last: Ng

first: Simon

last: Chatfield

first: Jane

last: Murphy

first: Damian

last: D’Silva

first: Ee

middle: S

last: Pang

first: Yunshun

first: Monther

last: Alhamdoosh

first: Nicholas

last: Wilson

first: Ian

last: Wicks


title: Interleukin-1β promotes an atheroprotective distribution of smooth muscle cells and macrophages in late stage murine atherosclerotic lesion

city: Charlottesville

first: Gary

middle: K

last: Owens

state: VA

address: 415 Lane Road MR5 Building Room 1322

institute: University of Virginia

summary: Despite decades of research, our understanding of processes controlling the stability of late-stage atherosclerotic plaques remains poor. However, a prevailing hypothesis is that reducing inflammation may improve plaque stability. Indeed, the potent inflammatory cytokine, interleukin-1β (IL-1β), has been shown to be a key driver of atherosclerosis development. Importantly, the CANTOS Trial recently demonstrated that administration of an anti-IL-1β antibody to high-risk post-myocardial infarction (MI) patients reduced the incidence of recurrent nonfatal MI, but did not reduce the incidence of cardiovascular death or stroke. As such, although the CANTOS trial results providing exciting evidence that targeting inflammation can have clinical benefit for treating advanced atherosclerosis, extensive further investigation is needed to better understand the mechanisms by which IL-1β inhibition impacts established lesions. Therefore, we performed intervention studies on smooth muscle cell (SMC) lineage tracing mice with advanced atherosclerosis using anti-IL- 1 or IgG control antibodies. Surprisingly, we found no effect on lesion size but a profound shift in the composition of the fibrous cap characterized by reduced collagen and SMC content but an increase in macrophage number, which was primarily driven by opposite effects on the proliferation of these respective cell types. By generating SMC-specific and macrophage-selective Il1r1 KO Apoe -/- mice, we found that SMC-specific Il1r1 KO resulted in a ~60% reduction in lesion size and lesions that were nearly devoid of YFP + SMC whereas, myeloid-selective loss of IL1R1 had no effect on lesion size, or cell composition. This suggests that SMC are a primary cell type responding to IL-1β in atherosclerosis and that IL1 signaling in SMC is critical for their investment and retention within the fibrous cap. In addition, we found that inhibition of IL-1β promoted an expansion of the M2 macrophage population within the fibrous cap, and that these effects may be due in part to elevated levels of IL4. Taken together, results show that IL-1β can promote beneficial changes in late-stage murine atherosclerosis by promoting maintenance of a SMC/collagen-rich fibrous cap. Moreover, studies identify critical cell types and pathways that need to be considered when attempting to develop safer and more effective anti-inflammatory therapies for widespread treatment of atherosclerotic disease, including in moderate or low risk patients.

overall_design: mRNA profiles of 18 week fed Western diet wild type (WT) SMC lineage tracing with Il1B antibody injection or IgG control were generated by deep sequencing, four animals per group, using Illumina HiSeq 2000.

first: Delphine

last: Gomez

first: Richard

last: Baylis

first: Alexandra

last: Newman

first: Brittany

last: Durgin

first: Gabriel

middle: F

last: Alencar

first: Sheila

last: Francis

first: Emmanuel

last: Pinteaux

first: Gwendalyn

middle: J

last: Randolph

first: Matthias

last: Nahrendorf

first: Philip

last: Swirski

first: Herman

last: Gram

first: Paul

last: Ridker

first: Peter

last: Libby