title: Genome-wide maps of chromatin state in Germinal Center B cells.

status: Public on Dec 11 2013

city: Milano

first: Stefano

last: Casola

state: Select a State or Province

address: Via Adamello 16

institute: IFOM

summary: Protection against deadly pathogens requires the production of high-affinity antibodies by B cells. High-affinity antibody-forming cells are generated in germinal centers (GC) as a result of a developmental program that is commonly altered in B cell malignancies. Identification of regulators of the GC response is crucial to develop targeted therapies for GC B cell dysfunctions including lymphomas. The Histone H3 lysine-27 methyltransferase Enhancer of Zeste homolog 2 (EZH2) is highly expressed in GC B cells and often constitutively activated in GC-derived Non-Hodgkin lymphomas (NHL), but its function in these cells remains largely unknown. Herein, we show that Ezh2 inactivation in mouse GC B cells causes a profound impairment in GC responses, memory B cell formation and humoral immunity. Mechanistically, Ezh2 protected GC B cells against AID mutagenesis, facilitated cell-cycle progression independent of p16INK4a repression and silenced plasma cell determinants Blimp1 and Irf4. Likewise, EZH2 inhibition in GC-derived NHL cells induced tumor suppressor BLIMP1, which impaired tumor growth. In conclusion, in GC B cells Ezh2 sustains AID function and prevents terminal differentiation to allow antibody diversification and affinity maturation. Dysregulation of such mechanisms may facilitate lymphomagenesis and identifies EZH2 as possible therapeutic target in NHL and other GC-derived B cell diseases.

overall_design: In  this study, two independent primary samples of splenic GC B cells from C5/BL/6J mice were subjected to H3K27me3 and H3K4me3 ChIP-sequencing. A totale of 6 samples, including input samples,  were subjected to ultra-deep sequencing.

first: Marieta

last: Caganova

first: Chiara

last: Carrisi

first: Gabriele

last: Varano

first: Federica

last: Mainoldi

last: Zanardi

first: Pierre-Luc

last: Germain

first: Laura

last: George

last: Alberghini

first: Luca

last: ferrarini

first: Asoke

last: talukder

first: Maurilio

last: Ponzoni

first: Giuseppe

last: testa

first: Takuya

last: Nojima

first: Claudio

last: Doglioni

first: Daisuke

last: Kitamura

first: Kai-Michael

last: Toellner

first: I-hsin

last: Su


title: A mesodermal factor, T (Brachyury), specifies mouse germ cell fate by directly activating germline determinants

city: Kyoto

first: Yukihiro

last: Yabuta

state: Kyoto

address: Yoshida-Konoe-cho, Sakyo-ku

department: Anatomy and Cell Biology

country: Japan

institute: Kyoto University, Graduate school of medicine

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Germ cells ensure reproduction and heredity in metazoans.  Primordial germ cells (PGCs) in mice are induced in pluripotent epiblasts by BMP4 and WNT3, yet their mechanism of action remains unclear.  Here, using in vitro PGC specification system, we show that WNT3 induces many transcription factors associated with mesoderm in epiblast-like cells (EpiLCs) through b-CATENIN.  Among these, T (BRACHYURY), a classical and conserved mesodermal factor, was essential for robust activation of Blimp1 and Prdm14, two of the germline determinants.  T, but not SMAD1 or b-CATENIN/TCF1, binds distinct regulatory elements of both Blimp1 and Prdm14, and without BMP4 and WNT3, directly up-regulates these genes, consequently delineating downstream PGC program.  Without BMP4, a program induced by WNT3 prevents T from activating Blimp1 and Prdm14, demonstrating that BMP4 is permissive for PGC specification.  These findings establish a fundamental role of a mesodermal transcription factor in PGC specification, a potentially evolutionarily conserved mechanism across metazoans.

overall_design: Genome target search for transcription factors, SMAD1, T, and TCF1 on PGC-like cells

first: Shinya

last: Aramaki

first: Katsuhiko

last: Hayashi

first: Kazuki

last: Kurimoto

first: Hiroshi

last: Ohta

first: Hiroko

last: Iwanari

first: Yasuhiro

last: Mochizuki

first: Takao

last: Hamakubo

last: Shirahige

first: Mitinori

last: Saitou


title: Root transcriptomes for the 19 parents of the Arabidopsis thaliana MAGIC lines

city: Salt Lake City

first: Richard

middle: M

last: Clark

state: Utah

department: Department of Biology

country: USA

institute: University of Utah

type: Expression profiling by high throughput sequencing

summary: We produced RNA-Seq reads from messenger RNA isolated from root tissue for the 19 MAGIC founder accessions (inbred strains) of Arabidopsis thaliana (see Gan et al. 2011. Nature 477:419-23 for a description of the MAGIC genetic mapping resource). The read data was generated with biological replication (two replicates). The resulting RNA-Seq data provide a resource to assess root gene expression across different accessions of A. thaliana. Comparable RNA-Seq read data for the MAGIC founder accessions for aerial seedling tissue has previously been released under GEO series GSE30720 (Gan et al. 2011. Nature, 477:419-23).

overall_design: Examination of RNA expression in root for the 19 Arabidopsis thaliana accessions used as parents for the MAGIC lines (parental accessions are Bur-0, Can-0, Col-0, Ct-1, Edi-0, Hi-0, Kn-0, Ler-0, Mt-0, No-0, Oy-0, Po-0, Rsch-4, Sf-2, Tsu-0, Wil-2, Ws-0, Wu-0, Zu-0).

first: Joshua

middle: G

last: Steffen

first: Robert

middle: T

last: Greenhalgh


title: Beta arrestin1 associated genomic regions in nicotine induced Non-Small Cell Lung Carcinoma (NSCLC) cell line A549

status: Public on Dec 16 2013

city: Tampa

first: Jonathan

last: Nguyen

state: FL

department: Tumor Biology

institute: Moffitt Cancer Centre & Research Institute

summary: The multifunctional scaffolding protein β-arrestin-1 plays a vital role in mediating the proliferative effects of nicotine through nAChR signaling. β-arrestins were initially known as negative regulators of GPCR mediated signaling as they promote internalization and desensitization of GPCRs. However, new roles of β-arrestins in receptor trafficking and signaling have been discovered in recent years. They are known to regulate signaling through a number of receptors such as Notch, endothelin A receptor, frizzled, smoothened and the nicotinic cholinergic receptors.  Studies from our lab revealed that nAChR signaling induces the translocation of β-arrestin-1 to the nucleus, in a Src dependent manner, where it directly binds to the proliferative E2Fs.  Furthermore, the nuclear translocation of β-arrestin-1 results in recruitment of p300 to E2F1 regulated proliferative promoters facilitating histone acetylation and transcription of these promoters. Given the role of β-arrestin-1 in nicotine induced gene expression, we attempted to explore the global association of β-arrestin-1 to the genomic regions upon nicotine stimulation by ChIP sequencing. It was found that β-arrestin-1 is recruited on the promoters of many genes that regulate EMT as well as other regulatory pathways. In this assay, β-arrestin-1 was found to be associated with the promoter regions of genes such as ZNF768, ZNF131, CSF3R, HMGA2, TAL1, RCC1, NKX2-4 etc in response to nicotine stimulation.

overall_design: Serum starved/quiescent A549 cells and Nicotine stimulated A549 cells

first: Smitha

last: Pillai

first: Srikumar

last: Chellappan


title: Sequencing of milk fat, milk cell, and mammary tissue RNA from rhesus macaques

status: Public on Dec 12 2013

city: Davis

first: Danielle

last: Lemay

state: CA

address: 451 Health Sciences Dr

department: Genome Center

institute: University of California Davis

summary: Studies of normal human mammary gland development and function have mostly relied on cell culture, limited surgical specimens, and rodent models. Although RNA extracted from human milk has been used to assay the mammary transcriptome non-invasively, the transcriptome derived from the milk fat layer has not been compared with the mammary-derived transcriptome nor have sources of RNA been quantified in milk. In this study the effects of milk collection and processing on RNA quality and origin were assessed in humans and rhesus macaques. Total RNA in milk was quantitated in acridine orange-stained milk using an automated whole slide scanner and custom-built Globulator software. Total RNA extracted from milk fat, cells in milk, and mammary biopsies of lactating rhesus macaques were compared using RNA sequencing and analysis. Compared with human milk, milk from macaques contained similar amounts of RNA-containing cytoplasmic crescents, but more cells. Total RNA extracted from milk fractions was also evaluated for factors that affect RNA quality. Degradation of RNA extracted from human milk fat was positively correlated with geographic distance from collection site, storage time, and sample type. There were no differences in RNA degradation in macaque milk collected after 10 min or 4 hr accumulation, suggesting that degradation of RNA extracted from milk fat may not occur in the mammary gland. Using RNA-Seq, RNA extracted from macaque milk fat and cells in milk more accurately represented RNA from mammary epithelial cells (cells that produce milk) than did RNA from mammary tissue. Mammary epithelium-specific transcripts were more abundant in macaque milk fat whereas adipose or stroma-specific transcripts were more abundant in mammary tissue. Functional analyses confirmed the validity of milk as a source of RNA from mammary epithelial cells. Analysis of highly abundant putative microRNAs in macaque milk fat revealed a potentially novel non-coding RNA species that is conserved in humans. RNA extracted from the milk fat during lactation accurately portrayed the RNA profile of milk-producing mammary epithelial cells. However, this sample type clearly requires protocols that minimize RNA degradation.

overall_design: Transcript profiles from milk cells, milk fat, and mammary tissue from 6 lactating rhesus macaques at 30 and 90 days lactation; 34 samples run in triplicate


title: Cohesin is a key determinant of interphase chromosome domain architecture (HiC)

city: London

first: Suzana

last: Hadjur

address: 72 Huntley street

department: Cancer biology

country: United Kingdom

institute: Ucl

summary: To ensure proper gene regulation within constrained nuclear space, chromosomes facilitate access to transcribed regions, while compactly packaging all other information. Recent studies revealed that chromosomes are organized into megabase-scale domains that demarcate active and inactive genetic elements, suggesting that compartmentalization is important for genome function. Here we show that very specific long-range interactions are anchored by cohesin/CTCF sites, but not cohesin-only or CTCF-only sites, to form a hierarchy of chromosomal loops. These loops demarcate topological domains and form intricate internal structures within them. Post-mitotic nuclei deficient for functional cohesin exhibit global architectural changes associated with loss of cohesin/CTCF contacts and relaxation of topological domains. Transcriptional analysis shows that this cohesin-dependent perturbation of domain organization leads to widespread gene deregulation of both cohesin-bound and non-bound genes. Our data thereby support a role for cohesin in the global organization of domain structure and suggest that domains function to stabilize the transcriptional programs within them.

overall_design: Hi-C, ChIP-Seq and RNA-Seq experiments were conducted in mouse neural stem cells and mouse astrocytes

first: Sevil

last: Sofueva

first: Eitan

last: Yaffe

first: Wen-Ching

last: Chan

first: Dimitra

last: Georgopoulou

first: Matteo

last: Vietri-Rudan

first: Steven

last: Pollard

first: Gary

middle: P

last: Schroth

first: Amos

last: Tanay


title: Distinct Cellular Origins for Serotonin-Expressing and Enterochromaffin-like Cells in the Gastric Corpus

city: Worcester

first: Daniel

middle: R

last: Caffrey

state: MA

address: 364 Plantation St

institute: UMASS Medical School

summary: The alimentary tract contains a diffuse endocrine system comprising enteroendocrine cells that secrete peptides or biogenic amines to regulate digestion, insulin secretion, food intake, and energy homeostasis. Lineage analysis in the stomach revealed that a significant fraction of endocrine cells in the gastric corpus did not arise from neurogenin3-expressing cells, unlike enteroendocrine cells elsewhere in the digestive tract. We aimed to isolate enriched serotonin-secreting and enterochromaffin-like (ECL) cells from the stomach and to clarify their cellular origin. We used Neurod1 and Neurog3 lineage analysis, and examined differentiation of serotonin-producing and ECL cells in stomach tissues of Neurod1-cre;ROSAtdTom, Tph1-CFP, c-Kitwsh/wsh, and Neurog3Cre;ROSAtdTom mice, by immunohistochemistry. We used fluorescence-activated cell sorting to isolate each cell type for gene expression analysis. We performed RNA-seq analysis of ECL cells. Neither serotonin-secreting nor ECL cells of the corpus arose from cells expressing Neurod1. Serotonin-secreting cells expressed a number of mast cell genes, but not genes associated with endocrine differentiation; they did not develop in c-Kitwsh/wsh mice and were labeled with transplanted bone marrow cells. RNA-seq analysis of ECL cells revealed high expression levels of many genes common to endocrine cells including transcription factors, hormones, ion channels, and solute transporters but not markers of bone marrow cells.

overall_design: We used fluorescence-activated cell sorting to isolate Hdc+ cells from stomach corpus and performed RNA-seq for gene expression analysis to determine the origin of those cells.

first: Hui

middle: J

last: Li

last: Aiello

first: Andrew

middle: A

last: Leiter


title: Restriction of LINE1 activity by RNAi correlates with mouse ES cell differentiation

city: Paris Cedex 05

first: Nicolas

last: Servant

address: 26 rue d'ulm

country: France

institute: Institut Curie

type: Non-coding RNA profiling by high throughput sequencing

summary:
Long interspersed elements 1 (LINE-1 or L1) are retrotransposons that dominate the mouse genomic landscape, and are expressed in Embryonic Stem Cells (ESCs), germ cells, and during early development.
Based on clear precedents in plants and fission yeast, we investigated in this study a role for RNAi and other RNA degradation pathways in the regulation of L1 expression and mobilization.
We uncovered the existence of novel small (s)RNAs that map to active L1 elements. Some of these sRNAs have characteristics of cognate short-interfering RNA populations, while others display length heterogeneity that evokes a biogenesis through a RNA surveillance pathway, in a Dicer-independent manner.
We additionally found that genetic ablation of Dicer and the sRNA effector protein AGO2 has complex and profound consequences on L1 transcription and mobilization in ESCs, indicating that endogenous RNA interference (RNAi) pathway indeed maintain genomic integrity against L1 proliferation.
Finally, we investigated the implication of L1 retrotransposition during ESC differentiation and propose that the mobilization of L1 elements in Dicer mutant ESCs could partially explain the inability of these cells to differentiate.

overall_design: 2 samples examined: WT E14 and Dicer mutant mouse ESCs

first: Constance

last: Ciaudo

first: Florence

last: Jay

first: Iku

last: Okamoto

first: Chongjian

last: Chen

first: Emmanuel

last: Barillot

first: Edith

last: Heard

first: Olivier

last: Voinnet


title: Cohesin is a key determinant of interphase chromosome domain architecture (ChIP-seq)

first: Suzana

last: Hadjur

first: Sevil

last: Sofueva

first: Eitan

last: Yaffe

first: Wen-Ching

last: Chan

last: Georgopoulou


title: iMir: An integrated pipeline for high-throughput miRNA-Seq data analysis

city: Baronissi

first: Maria Rosaria

last: De Filippo

address: Via Allende 1

country: Italy

institute: University of Salerno

summary: We report a novel modular pipeline (iMir) for comprehensive analysis of miRNA-Seq data, from linker removal and sequence quality check to differential expression and biological target prediction, integrating multiple open source modules and resources linker together in an automated flow.

overall_design: Development of an integrated pipeline (iMir) for comprehensive analysis of miRNA-Seq experiment.

first: Alessandro

last: Weisz

first: Giorgio

last: Giurato


title: Cohesin is a key determinant of interphase chromosome domain architecture (RNA-seq)

first: Dimitra

first: Matteo

last: Vietri-Rudan

first: Steven

last: Pollard

first: Gary

last: Schroth

first: Amos

last: Tanay


title: Polycomb-dependent H3K27me1 and H3K27me2 characterization in mouse ES cells (E14 and E36 with H3K27me1/2 and H3)

city: Milan

first: Sri Ganesh

last: Jammula

institute: IFOM-IEO

summary: These data include the genome wide location analysis of H3K27me1/2/ac and H3K4me1/3  by ChIP in mouse ES cells.

overall_design: Precipitation of formaldehyde cross-linked chromatin prepared from mouse ES cell using specific antibodies against Histone H3 and its specific modifications. Rabbit IgG were used as control.

first: Karin

last: Ferrari

first: Ganesh

first: Diego

last: Pasini


title: Architecture of Epigenetic Reprogramming Following Twist1 Mediated Epithelial-Mesenchymal Transition

status: Public on Dec 10 2013

city: Illkirch

first: Gabriel

last: Malouf

address: 1 rue Laurent Fries

department: Cancer and functional genomics

institute: IGBMC

type: Methylation profiling by high throughput sequencing

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

overall_design: Refer to individual Series

first: Jean-Pierre

last: Issa

first: Yue

last: Lu

first: Sendurai

last: Mani

first: Joseph

middle: H

last: Taube


title: Architecture of Epigenetic Reprogramming Following Twist1 Mediated Epithelial-Mesenchymal Transition [DREAM-seq]

summary:
Purpose: to characterize epigenetic changes following Twist1 mediated Epithelial-Mesenchymal Transition in human

Methods: we characterized the epigenetic and transcriptome landscapes using whole genome transcriptome analysis by RNA-seq, DNA methylation by digital restriction enzyme analysis of methylation (DREAM) and histone modifications by CHIP-seq of H3K4me3 and H3K27me3 in immortalized human mammary epithelial cells relative to cells induced to undergo EMT by Twist1.

Results: EMT is accompanied by focal hypermethylation and widespread global DNA hypomethylation, predominantly within transcriptionally repressed gene bodies. At the chromatin level, the number of gene promoters marked by H3K4me3 increases by more than one fifth; H3K27me3 undergoes dynamic genomic redistribution characterized by loss at half of gene promoters and overall reduction of peak size by almost one-half. This is paralleled by increased phosphorylation of EZH2 at serine 21. Among genes with highly altered mRNA expression, 23.1% switch between H3K4me3 and H3K27me3 marks, and those point to the master EMT targets and regulators CDH1, PDGFRA and ESRP1. Strikingly, Twist1 increases the number of bivalent genes by more than two fold. Inhibition of the H3K27 methyltransferases EZH2 and EZH1, which form part of the PRC2 complex, results in blocking EMT and stemness properties.

Conclusion: Our findings demonstrate that the EMT program requires epigenetic remodeling by the Polycomb/Trithorax complexes leading to increased cellular plasticity which suggests that its inhibition will prevent EMT, and the associated breast cancer metastasis.

overall_design: DREAM profiles of human mammary epithelial cells before (HMLE_parental) and after Twist1 transfection (HMLE_Twist) were generated in monolayer (HMLE_Twist2D) and sphere culture by deep sequencing using Illumina GAIIx or Illumina hiseq2000. Furthermore, DREAM profile was also obtained in parental human mammary epithelial cells transfected with GFP


title: Discovery and Characterization of Super-Enhancer Associated Dependencies in Diffuse Large B-Cell Lymphoma

city: Boston

first: James

last: Bradner

address: 450 Brookline

department: Medical Oncology

institute: Dana-Farber Cancer Institute

summary: Diffuse Large B-Cell Lymphoma (DLBCL) is a biologically heterogeneous and clinically aggressive disease. Here, we explore the role of BET bromodomain proteins in DLBCL, using integrative chemical genetics and functional epigenomics. We observe highly asymmetric loading of BRD4 at enhancers, with approximately 33% of all BRD4 localizing to enhancers at 1.6% of occupied genes. These super-enhancers prove particularly sensitive to bromodomain inhibition, explaining the selective effect of BET inhibitors on oncogenic and lineage-specific transcriptional circuits. Functional study of genes marked by super-enhancers identifies DLBCLs dependent on OCA-B and suggests a strategy for discovering unrecognized cancer dependencies. Translational studies performed on a comprehensive panel of DLBCLs establish a therapeutic rationale for evaluating BET inhibitors in this disease.

overall_design: ChIP-Seq for various transcription factors and histone modifications in diffuse large B-cell lymphoma cells

first: M

last: McKeown

first: C

middle: Y

last: Lin

first: J

middle: E


title: Architecture of Epigenetic Reprogramming Following Twist1 Mediated Epithelial-Mesenchymal Transition [ChIP-seq]

overall_design:
ChIPseq profiles of human mammary epithelial cells before (HMLE_parental) and after Twist1 transfection (HMLE_Twist) were generated in monolayer (HMLE_Twist2D) and sphere culture by deep sequencing using Illumina GAIIx or Illumina hiseq2000
Please note that the processed data files were generated by pooling both replicate samples after confirming the high correlation of the two replicates.


title: Initiation of MLL-rearranged AML is orchestrated by C/EBPa

city: Copenhagen

first: Bo

middle: Torben

last: Porse

address: Ole Maaloes vej 5

department: BRIC/Finsen Laboratory

country: Denmark

institute: University of Copenhagen/Rigshospitalet

summary: Acute Myeloid Leukemia (AML) is associated with a number of genetic and epigenetic events that result in malignant transformation of hematopoietic cells. In particular, transcription factors essential for normal hematopoiesis and stem cell function are often found mutated leading to the formation of leukemic stem cells and the accumulation of immature blasts. Among them, translocations involving the mixed lineage leukemia (MLL) gene at chromosome band 11q23 are one of the most commonly events (~10 %) and is associated with poor prognosis in human leukemias. Whereas the downstream effects of MLL-fusion proteins are well established, the modes on which these effects are mediated are still unclear and whether MLL-fusion proteins are dependent on other transcriptional regulators or act alone remains elusive. To investigate this we searched gene expression profiles from patients with MLL-rearranged AML compared with normal hematopoietic progenitor cells for transcriptional regulators and found targets of C/EBPα to be up-regulated in the AML samples, suggesting that C/EBPα might collaborate with MLL-fusion proteins in the initial transformation process. We could show that transformation by MLL-fusion proteins is dependent on C/EBPα activity both in early progenitors as well as in GMPs. In contrast, C/EBPα was found to be indispensable in an already established leukemia. These finding led us to study the early transcriptional changes induced by MLL-ENL expression and we identified a combined C/EBPα / MLL-ENL transcriptional signature. Collectivly, our data shows that C/EBPα configure a proper chromatin state required for MLL-fusions to induce malignant transformation.

overall_design: Histone modification profiles (H3K4me3 and H3K27me3) in haematopoietic progenitor cells (preGM, wild type and Cebpa knock out), and C/EBPα binding in GMP cells

first: E

last: Ohlsson

first: B


title: Architecture of Epigenetic Reprogramming Following Twist1 Mediated Epithelial-Mesenchymal Transition [RNA-seq]

overall_design: RNAseq profiles of human mammary epithelial cells before (HMLE_parental) and after Twist1 transfection (HMLE_Twist) were generated in monolayer (HMLE_Twist2D) and sphere culture by deep sequencing using SOLID


title: Sequencing of small RNAs in healthy human serum from China

status: Public on Dec 21 2013

city: Nanjing

first: Zhen

last: Zhou

state: Jiangsu

address: 22th,Hankou Road

department: School of Lifesciences

country: China

institute: Nanjing University

summary: We applied Solexa sequencing technology to identify normal human serum microRNA genes. Sequencing result was refered to miRBase 15.0, including 904 human miRNAs.Using this method, we found that there were total number of 513 miRNAs in normal human serum and the total copy number of the serum miRNA was 3780436.

overall_design: 18-30 nt small RNAs from 43 healthy Chinese serum pool was sequenced in one Solexa lane

first: Limin

last: Li

first: Dihan

last: Zhu


title: Differential Methylation Of The  TRPA1 Promoter In Pain Sensitivity

status: Public on Dec 09 2013

first: Jordana

last: Bell

address: St Thomas' Hospital, Westminster Bridge Rd

department: Department of Twin Research and Genetic Epidemiology

institute: King's College London

type: Methylation profiling by genome tiling array


title: Lsh regulates LTR retrotransposon repression independent of Dnmt3b function (HELP-seq)

city: Edinburgh

first: Donncha

middle: S

last: Dunican

address: Western General Hospital

institute: IGMM / MRC HGU

summary: Background: Global DNA methylation contributes to genomic integrity by supressing repeat associated transposition events. Several chromatin factors are required in addition to DNA methyltransferases to maintain DNA methylation at intergenic and satellite repeats. Embryos lacking Lsh, a member of the SNF2 superfamily of chromatin helicases, are hypomethylated. The interaction of Lsh with the de novo methyltransferase, Dnmt3b, facilitates the deposition of DNA methylation at stem cell genes. We wished to determine if a similar targeting mechanism operates to maintain DNA methylation at repetitive sequences. Results: We used HELP-seq to map genome wide DNA methylation patterns in Lsh-/- and Dnmt3b-/- somatic cells. DNA methylation is predominantly lost from specific genomic repeats in Lsh-/- cells: LTR-retrotransposons, LINE-1 repeats and mouse satellites. RNA-seq experiments demonstrate that specific IAP (Intracisternal A-type particle) LTRs and satellites, but not LINE-1 elements, are aberrantly transcribed inLsh-/- cells. LTR hypomethylation in Dnmt3b-/- cells is moderate and hypomethylated repetitive elements (IAP, LINE-1 and satellite) are silent. Chromatin immunoprecipitation (ChIP) indicates that repressed LINE-1 elements gain H3K4me3, but H3K9me3 levels are unaltered in Lsh-/- cells, indicating that DNA hypomethylation alone is not permissive for their transcriptional activation. Mis-expressed IAPs and satellites lose H3K9me3 and gain H3K4me3 in Lsh-/- cells. Conclusions: Our study emphasizes that regulation of repetitive elements by DNA methylation is selective and context dependent. We propose a model where Lsh is specifically required at a precise developmental window to target de novo methylation to repeat sequences, which is subsequently maintained by Dnmt1 in somatic cells to enforce repeat silencing thus contributing to genomic integrity.

overall_design: Two pairs of genomic samples compared: WT and Lsh-/- DNA isolations from tail-tip fibroblasts; WT and Dnmt3b knockout DNA isolations from mouse embryonic fibroblasts.

first: Richard

last: Meehan


title: Evaluation of the effectiveness of semen collection and sperm purification methods for spermatozoa transcript profiling

city: Detroit

first: Stephen

last: Krawetz

state: MI

address: 275 E. Hancock

department: Center for Molecular Medicine and Genetics

institute: Wayne State University School of Medicine

summary: RNA-Seq technique was applied to investigate the effects of two semen collection methods (Pelleted vs Liquefied) and two sperm purification methods (SCLB vs PS) to the integrity of isolated RNAs at different perspectives.

overall_design: The same set of semen samples were applied to investigate the qualitative and quantitative effect of semen collection methods and sperm cell purification methods on sperm transcript profiling.

first: Shihong

last: Mao

first: Robert

last: Goodrich

first: Russ

last: Hauser


title: Lsh regulates LTR retrotransposon repression independent of Dnmt3b function


title: A RNA-seq-based gene expression profiling of radiation-induced tumorigenic mammary epithelial cells

city: Beijing

first: lina

last: ma

address: No.7 Beitucheng West Road, Chaoyang District

institute: Beijing Institute of Genomics,Chinese Academy of Sciences

summary: To understand tumorigenesis and cancer progression of mammary epithelium, we established a cell model combining over-expression of human telomerase reverse transcriptase gene (hTERT) and heavy-ion radiation from normal human mammary epithelial cells. We subsequently used RNA-seq method to acquire their transcriptomes from two characteristic cell lines, an immortal epithelial cell line (I_hMEC) and a tumorigenic epithelial cell line (T_hMEC), and to look for differentially expressed genes (DEGs) between immortalization and tumorigenicity. We identified 7,053 DEGs, of which 84 were not only highly expressed but also significantly regulated. We found that house-keeping (HK) genes and tissue-specific (TS) genes were regulated differently during tumorigenesis; HK genes tend to be activated but TS genes tend to be repressed. We looked into three important pathways in cancer development: p53 signaling, cell cycle, and apoptosis. Although both immortal and tumorigenic cells have infinite potential to replicate and can escape apoptosis, a great number of genes exhibited different modulation pattern between the two processes. We also found that a significant number of DEGs were involved in epigenetic modification of chromatins. In conclusion, these findings may provide novel biomarkers in studying tumorigenicity and further our understanding of cellular mechanism(s) in the transition from immortal to tumorigenic processes.

overall_design: 2 samples examined: immortalized human mammary epithelial cell (I_hMEC) and tumorigenic  human mammary epithelial cell (T_hMEC)

first: Jun

last: Yu

first: Yongliang

last: Zhao

first: Lina

last: Ma


title: Cohesin is a key determinant of interphase chromosome domain architecture

first: Suzana

last: Hadjur


title: Impact of genomic polymorphisms on the repertoire of human MHC class I-associated peptides

status: Public on Dec 08 2013

city: Montreal

first: Diana

middle: Paola

last: Granados

state: Quebec

address: C.P. 6128, succursale Centre-ville

department: Medecine

country: Canada

institute: IRIC

summary: We developed a novel approach combining next generation sequencing, bioinformatics and mass spectrometry to assess the impact of non-MHC polymorphisms on the repertoire of MHC I-associated peptides (MIPs). We compared the genomic landscape of MIPs eluted from B lymphoblasts of two MHC-identical siblings and determined that MIPs mirror the genomic frequency of non-synonymous polymorphisms but they behave as recessive traits at the surface level. Moreover, we showed that 11.7% of the MIP coding exome is polymorphic at the population level. Our method provides fundamental insights into the relation between the genomic self and the immune self and accelerates the discovery of polymorphic MIPs (also known as minor histocompatibility antigens), which play a major role in allo-immune responses.

overall_design: RNA-seq of human B lymphoblasts derived from peripheral blood mononuclear cells from 2 HLA-identical female siblings.

first: Dev

last: Sriranganadane

first: Tariq

last: Daouda


title: Genome-wide identification of regulatory elements in preSertoli cells based on DNaseI hypersensitivity

status: Public on Dec 07 2013

city: Chicago

first: Danielle

last: Maatouk

address: 303 E. Superior St., Lurie 7-250

institute: Northwestern University

summary: Identifying transcriptional networks active during development has been hindered by the slow pace at which regulatory elements are identified, and by the difficulty in purifying cells from developing tissues. We used DNaseI-seq to identify functional regulatory elements in purified fetal preSertoli cells and uncovered thousands of preSertoli-specific DNaseI hypersensitive sites (DHSs) that are likely to activate or repress genes required for Sertoli cell development and function. To demonstrate their functional significance, we validated the preSertoli-specific enhancer activity of a DHS located 50 kb upstream of the Wt1 gene. This genome-wide data will guide many future studies into gene-regulatory mechanisms that operate in the fetal gonad and may lead to the identification of non-coding mutations leading to DSDs.

overall_design: One biological replicate was analyzed for each of the eight mouse samples using DNase-seq. Three biological replicates were analyzed for RNA-seq. Two biological replicates for ChIP-seq.

first: Yoichiro

last: Shibata


title: Functional categorization of transcriptome in the species Symphysodon aequifasciatus Pellegrin 1904 (Perciformes: Cichlidae) exposed to benzo[a]pyrene and phenanthrene

city: Brisbane

first: Renato

last: de Souza Pinto Lemgruber

state: QLD

address: Building 75,Cnr College Rd & Cooper Rd

country: Australia

institute: University of Queensland

summary: This study aims to evaluate the transcriptome alterations, through cDNA libraries, associated with the effects of the combination of two PAHs, benzo[a]pyrene (0.5μg/L) and phenanthrene (50μg/L), present in crude oil, on specimens of Symphysodon aequifasciatus (discus fish) after 48h of exposure. The cDNA libraries were constructed according to the SOLiDTM SAGETM protocol for sequencing in the SOLiD v.3 Plus sequencer. The results were analyzed by bioinformatics and differentially expressed genes were categorized using the gene ontology program (AmiGO v.1.8). The functional categories (terms) found in the gene ontology and the gene network generated using STRING software were used to predict the adverse effects in the liver. In the present study, 27,127 genes (compared to Danio rerio database) were identified. Considering only those genes with a p-value less than or equal to 0.05 and a greater than or equal to two-fold change in expression across libraries, we found 804 genes, 438 down-regulated (54%) and 366 up-regulated genes (46%), in the experimental group compared to the control group. Out of this total, 327 genes were successfully categorized, 174 down-regulated and 153 up-regulated genes, using gene ontology tool. The final confidence gene network, analyzed in STRING, was composed by 199 nodes of proteins, 124 of them resulted in 274 interactions.  The results showed that even an acute exposure of 48 h caused metabolic change in response to environmental contaminants, resulting in changes of cell integrity, changes in oxidation-reduction processes, disturbances of intracellular signaling and changes in the immune response of discus fish. Also the gene network interactions have showed no central interplay cluster, exhibiting interconnected clusters interactions and sparsely connected sub-networks. These findings highlight that even an acute and sublethal exposure of PAHs can cause metabolism changes that may affect this fish species survival rates. Our findings using the SAGE-method and the SOLiD sequencer showed that this is a powerful tool for gene expression analysis in discus fish, a non-model organism.

overall_design: Transcriptome analysis of differentially expressed genes in liver in benzo[a]pyrene and phenanthrene-exposed and non-exposed fish (N=6 per treatment)

last: Lemgruber

first: Nislanha

last: Marshall

first: Andrea

last: Ghelfi

first: Daniel

middle: B

last: Fagundes

first: Adalberto

middle: L

last: Val


title: Expression profiling in the heart of wild-type and kctd10 mutant zebrafish larvae

status: Public on Dec 06 2013

first: Xiangjun

last: Tong

address: Yiheyuan Road No. 5

department: College of Life Sciences

institute: Peking University

summary: We sequenced mRNA of hearts from 50 wild-type and 50 kctd10 mutant embryos at 48 hpf.

overall_design: Examination of mRNA levels in the larvae hearts between the the wt and the kctd10 mutant.

first: Bo

last: Zhang


title: Lsh regulates LTR retrotransposon repression independent of Dnmt3b function (RNA-Seq)

first: Donncha

last: Dunican

overall_design: Two pairs of RNA samples compared: WT and Lsh-/- RNA isolations from tail-tip fibroblasts; WT and Lsh-/- RNA isolations from E13.5 mouse embryos.

first: Richard

last: Meehan


title: Transcriptomic analysis of LaeA and LaeB deletion strains in fungus Beauveria bassiana

status: Public on Dec 05 2013

city: Jinan

first: Yuqi

last: Qin

address: 27 Shanda Nan Lu

institute: Shandong University

summary: Transcriptomic analysis of LaeA-deletion and  overexpression LaeB in LaeA deletion strains in fungus Beauveria bassiana

overall_design: Examination of differential gene expressions by Beauveria bassiana wild type, LaeA-deletion and  overexpression LaeB in LaeA deletion strains  in fungus Beauveria bassiana

first: Nemat

last: Keyhani


title: Genetic and molecular basis of drug resistance and species-specific drug action in Schistosome parasites

city: SAN ANTONIO

first: Timothy

last: ANDERSON

state: TEXAS

department: Genetics

institute: Texas Biomedical Research Institute

summary: Oxamniquine resistance evolved in the human blood fluke (Schistosoma mansoni) in Brazil in the 1970s. We crossed parental parasites differing ~500-fold in drug response, determined drug sensitivity and marker segregation in clonally-derived F2s, and identified a single QTL (LOD=31) on chromosome 6. A sulfotransferase was identified as the causative gene using RNAi knockdown and biochemical complementation assays and we subsequently demonstrated independent origins of loss-of-function mutations in field-derived and laboratory-selected resistant parasites. These results demonstrate the utility of linkage mapping in a human helminth parasite, while crystallographic analyses of protein-drug interactions illuminate the mode of drug action and provide a framework for rational design of oxamniquine derivatives that kill both S. mansoni and S. haematobium, the two species responsible for >99% of schistosomiasis cases worldwide.

overall_design: mRNA profiles from adult worms resistant (HR, 3 replicates) and susceptible (LE, 2 replicates) to the oxamniquine drug were compared to identify differential expression in genes related to drug resistance

first: Claudia

last: Valentim

first: Donato

last: Cioli

first: Frédéric

middle: D

last: Chevalier

first: Xiaohang

last: Cao

first: Alexander

last: Taylor

first: Stephen

last: Holloway

first: Livia

last: Pica-Mattoccia

first: Alessandra

last: Guidi

first: Annalisa

last: Basso

first: Isheng

last: Tsai

first: Matthew

last: Berriman

last: Carvalho-Queiroz

first: Marcio

last: Almeida

first: Hector

last: Aguilar

first: Doug

last: Frantz

first: P

last: Hart

last: Anderson

first: Philip

last: LoVerde


title: An Integrated Model of the Transcriptome Landscape of HER2-Positive Breast Cancer

city: Rochester

first: Krishna

last: Kalari

state: MN

address: 200 1st ST SW

department: Department of Biomedical Statistics and Informatics

institute: Mayo Clinic

summary: The goal of our study is to build an integrated transcriptome landscape model for HER2 positive breast tumors and identify the crucial signaling pathways associated with HER2 tumors. Genomic features include, 685 genes that were differentially expressed only in HER2-positive tumors, 102 genes that were alternatively spliced in a pattern that is unique to HER2-positive tumors, and 303 genes that expressed single nucleotide sequence variants (eSNVs) that were unique to HER2-positive tumors. Network analysis was performed to integrate the genomic features into a transcriptome landscape model that identified 12 highly interconnected cellular processes that appear to be critical to the establishment and maintenance of HER2-positive tumors. We observed that integrin signaling was linked to lapatinib sensitivity in vitro and strongly associated with risk of relapse in the NCCTG N9831 adjuvant trastuzumab clinical trial dataset.

overall_design: We analyzed RNA-seq data from a survey panel consisting of 8 benign breast lesions, 8 ER+, 8 triple negative, and 8 HER2-positive primary breast tumors to identify genomic features that were uniquely associated with HER2-positive tumors

first: Brian

last: Necela

first: Xiaojia

last: Tang

first: Melissa

last: Lau

first: Jeanette

last: Eckel-Passow

first: S

middle: K

first: Zhifu

last: Sun

first: Saurabh

last: Baheti

first: Jennifer

last: Carr

first: Tiffany

last: Baker

first: Kevin

last: Thompson

first: Derek

middle: C

last: Radisky

first: Richard

middle: W

last: Joseph

first: Sarah

last: McLaughlin

first: High-seng

last: Chai

first: Stephan

last: Camille

first: David

last: Rossell

first: Yan

last: Asmann

first: Edith

last: Perez

first: E


title: Gene expression, single nucleotide variant and fusion transcript discovery in archival material from breast tumors

department: HSR-BSI

summary: Advantages of RNA-Seq over array based platforms are quantitative gene expression and discovery of expressed single nucleotide variants (eSNVs) and fusion transcripts from a single platform, but the sensitivity for each of these characteristics is unknown. We measured gene expression in a set of manually degraded RNAs, nine pairs of matched fresh-frozen, and FFPE RNA isolated from breast tumor with the hybridization based, NanoString nCounter, (226 gene panel) and with whole transcriptome RNA-Seq using RiboZeroGold ScriptSeq V2 library preparation kits. We performed correlation analyses of gene expression between samples and across platforms. We then specifically assessed whole transcriptome expression of lincRNA and discovery of eSNVs and fusion transcripts in the FFPE RNA-Seq data. For gene expression in the manually degraded samples, we observed Pearson correlation of >0.94 and >0.80 with NanoString and ScriptSeq protocols respectively. Gene expression data for matched fresh-frozen and FFPE samples yielded mean Pearson correlations of 0.874 and 0.783 for NanoString (226 genes) and ScriptSeq whole transcriptome protocols respectively. Specifically for lincRNAs, we observed superb Pearson correlation (0.988) between matched fresh-frozen and FFPE pairs. FFPE samples across NanoString and RNA-Seq platforms gave a mean Pearson correlation of 0.838. In FFPE libraries, we detected 53.4% of high confidence SNVs and 24% of high confidence fusion transcripts. Sensitivity of fusion transcript detection was not overcome by an increase in depth of sequencing up to 3-fold (increase from ~56 to ~159 million reads). Both NanoString and ScriptSeq RNA-Seq technologies yield reliable gene expression data for degraded and FFPE material. The high degree of correlation between NanoString and RNA-Seq platforms suggests discovery based whole transciptome studies from FFPE material will produce reliable expression data. The RiboZeroGold ScriptSeq protocol performed particularly well for lincRNA expression from FFPE libraries but detection of eSNV and fusion transcripts was less sensitive.

overall_design: We performed RNASeq on RNA from nine matched pairs of fresh-frozen and FFPE tissues from breast cancer patients. The goal was to test the RiboZeroGold ScriptSeq complete low input library preparation kit for degraded RNA samples.

first: Nadine

last: Norton

first: Daniel

last: Serie

first: Brian

middle: M

last: Necela

first: Aditya

last: Bhagwate

first: Jin

last: Jen

first: Bruce

last: Eckloff

first: Krishna

last: Kalari

first: Jenifer

last: Kachergus


title: Polycomb-dependent H3K27me1 and H3K27me2 regulate active transcription and enhancer fidelity

first: Sri Ganesh

last: Jammula

address: Via Adamello 16

summary: These data include the genome wide location of different histone modifications by ChIP sequencing in mouse ES cells, and RNA Seq data generated from wild type and EED KO mouse ES cells and knocked down for unrelated protein and Setd2 protein.

overall_design:
ChIP-Seq: Immuno-precipitation of formaldehyde cross-linked chromatin prepared from wild type mouse E14 ES cells, wild type E36 ES cells, EED KO E36 ES cells, wild type Embryoid bodies (Ebs), EED KO Embryoid bodies (Ebs EED KO) using specific antibody against different histone modifications.

RNA-Seq: Total RNA extracted from wild type E36 ES cells, EED KO E36 ES cells, wild type E36 Embryoid bodies (Ebs), EED KO Embryoid bodies (Ebs EED KO), E14 Ctrl KD,  E14 Setd2 KD.

first: Karin

last: Ferrari

first: Andrea

last: Scelfo

first: Sri

middle: G

first: Diego

last: Pasini


title: Ribosome profiling reveals pervasive and regulated stop codon readthrough  in Drosophila melanogaster

city: San Francisco

first: Joshua

middle: Griffin

last: Dunn

state: California

address: 1700 4th Street, BH404

department: Cellular & Molecular Pharmacology

institute: UCSF / HHMI

summary: Ribosomes can read through stop codons in a regulated manner, elongating rather than terminating the nascent peptide. Stop codon readthrough is essential to diverse viruses, and phylogenetically predicted to occur in a few hundred genes in Drosophila melanogaster, but the importance of regulated readthrough in eukaryotes remains largely unexplored. Here, we present a ribosome profiling assay (deep sequencing of ribosome-protected mRNA fragments) for Drosophila melanogaster, and provide the first genome-wide experimental analysis of readthrough. Readthrough is far more pervasive than expected: the vast majority of readthrough events evolved within D. melanogaster and were not predicted phylogenetically. The resulting C-terminal protein extensions show evidence of selection, contain functional subcellular localization signals, and their readthrough is regulated, arguing for their importance. We further demonstrate that readthrough occurs in yeast and humans. Readthrough thus provides general mechanisms both to regulate gene expression and function, and to add plasticity to the proteome during evolution.

overall_design: 12 samples of Drosophila ribosome profiling and poly(A)+ mRNA-seq, including technical replicates in S2 cells, and biological replicates of 0-2 hour embryos

first: Jonathan

last: Weissman


title: Differential Protein Occupancy Profiling of the mRNA Transcriptome

city: Berlin

first: Markus

last: Landthaler

address: Robert-Rössle-Straße 10

department: Berlin Institute for Medical Systems Biology

country: Germany

institute: Max-Delbrück-Center for Molecular Medicine

summary: Protein-RNA interactions are fundamental to core biological processes, such as mRNA splicing, localization, degradation and translation. We have developed a photoreactive nucleotide-enhanced UV crosslinking and oligo(dT) purification approach to identify the mRNA-bound proteome using quantitative proteomics and to display the protein occupancy on mRNA transcripts by next-generation sequencing (Baltz and Munschauer et al. 2012). Our current work focuses on streamlining and extending protein occupancy profiling on poly(A)-RNA. Our objectives are to identify previously unknown protein-bound transcripts and, more importantly, to assess global and local differences in protein occupancy across different biological conditions. To this end, we have implemented poppi, the first pipeline for differential analysis of protein occupancy profiles. We have applied our analysis pipeline to pinpoint changes in occupancy profiles of MCF7 cells against already published HEK293 cells [GSE38157].

overall_design: We generated protein occupancy cDNA libraries for two biological replicates. Briefly, we crosslinked 4SU-labeled MCF7 cells and purified protein-mRNA complexes using oligo(dT)-beads. The precipitate was treated with RNAse I to reduce the protein-crosslinked RNA fragments to a length of about 30-60 nt. To remove non-crosslinked RNA, protein-RNA complexes were precipitated with ammonium sulfate and blotted onto nitrocellulose. The RNA was recovered by Proteinase K treatment, ligated to cloning adapters, and reverse transcribed. The resulting cDNA libraries were PCR-amplified and next-generation sequenced.


title: Parallel analysis of RNA ends enhances global investigation of microRNAs and target RNAs of Brachypodium distachyon

city: NEWARK

first: Dong-Hoon

last: Jeong

state: DE

address: 15 Innovation Way

institute: University of Delaware

summary:
The wild grass Brachypodium distachyon has emerged as a model system for temperate grasses and biofuel plants. However, the global analysis of miRNAs, molecules known to be key for eukaryotic gene regulation, has been limited in B. distachyon to studies examining a few samples or that rely on computational predictions. Similarly an in-depth global analysis of miRNA-mediated target cleavage using Parallel Analysis of RNA Ends (PARE) data is lacking in B. distachyon.
B. distachyon small RNAs were cloned and deeply sequenced from 17 libraries that represent different tissues and stresses. Using a computational pipeline, we identified 116 miRNAs including not only conserved miRNAs that have not been reported in B. distachyon, but also non-conserved miRNAs that were not found in other plants. To investigate miRNA-mediated cleavage function, four PARE libraries were constructed from key tissues and sequenced to a total depth of approximately 70 million sequences. The roughly 5 million distinct genome-matched sequences that resulted represent an extensive dataset to analyze small RNA-guided cleavage events. Analysis of the PARE and miRNA data provided experimental evidence for miRNA-mediated cleavage of 264 sites in predicted miRNA targets. In addition, PARE analysis revealed that differentially expressed miRNAs in the same family guide specific target RNA cleavage in a correspondingly tissue-preferential manner.
B. distachyon miRNAs and target RNAs were experimentally identified and analyzed. Knowledge gained from this study should provide insights into the roles of miRNAs and the regulation of their targets in B. distachyon and related plants.

overall_design: Examination of various tissues and stresses  in Brachypodium by high throughput sequencing for small RNA profiling and PARE (Parallel Analysis of RNA Ends)

first: Pamela

middle: J

last: Green


title: Diet rapidly and reproducibly alters the human gut microbiome

first: Peter

middle: James

last: Turnbaugh

address: 513 Parnassus Ave

department: Microbiology and Immunology

institute: University of California San Francisco

summary: Long-term dietary intake influences the structure and activity of the trillions of microorganisms residing in the human gut, but it remains unclear how rapidly and reproducibly the human gut microbiome responds to short-term macronutrient change. Here we show that the short-term consumption of diets composed entirely of animal or plant products alters microbial community structure and overwhelms inter-individual differences in microbial gene expression. The animal-based diet increased the abundance of bile-tolerant microorganisms (Alistipes, Bilophila and Bacteroides) and decreased the levels of Firmicutes that metabolize dietary plant polysaccharides (Roseburia, Eubacterium rectale and Ruminococcus bromii). Microbial activity mirrored differences between herbivorous and carnivorous mammals, reflecting trade-offs between carbohydrate and protein fermentation. Foodborne microbes from both diets transiently colonized the gut, including bacteria, fungi and even viruses. Finally, increases in the abundance and activity of Bilophila wadsworthia on the animal-based diet support a link between dietary fat, bile acids and the outgrowth of microorganisms capable of triggering inflammatory bowel disease. In concert, these results demonstrate that the gut microbiome can rapidly respond to altered diet, potentially facilitating the diversity of human dietary lifestyles.

overall_design: RNA-Seq analysis of the human gut microbiome during consumption of a plant- or animal-based diet.

first: Lawrence

middle: A

last: David


title: Cellular generation of 5-Hydroxymethylcytosine by redox-active chemicals via an unprecedent non-enzymatic mechanism

first: Zechen

last: Chong

address: NO. 7 Beitucheng West Road

institute: Beijing Institute of Genomics

summary: We proposed that besides TET family dioxygenase oxidizing 5mC to 5hmC, there is a non-enzyme pathway which is due to hydroxyl radica l(OH) or OH-like species also involvement in demethylation of 5mC forming 5hmC. This pathway includes classical fenton reagents such as H2O2 and Fe2+, and more important redox-activity quinoid compounds, especially, tetrachloro-1,4-benzoquinone (TCBQ), which was reported producing hydroxyl radicals independent of transition metal

overall_design: Examination of 5hmC and transcriptome levels with TCBQ and DMSO in human MRC-5 cell lines

first: Yun-Gui

last: Yang

first: Hailin

last: Wang


title: Simultaneous transcriptional profiling of bacteria and their host cells by heterogeneous RNA-Seq (hRNA-Seq)

city: Baltimore

first: Sean

last: Daugherty

state: Maryland

address: 801 W. Baltimore St.

department: Institute for Genome Sciences

institute: University of Maryland School of Medicine

summary: We developed heterogeneous RNA-Seq (hRNA-Seq) to simultaneously capture prokaryotic and eukaryotic expression profiles of bacteria-infected cells. As proof of principle, hRNA-Seq was applied to Chlamydia-infected cells, successfully obtaining the transcriptomes of both Chlamydia and their host cells at 1 and 24 hours post-infection. Substantial transcription was found in the immediate-early period of infection for both Chlamydia and the host cell. We discovered possible chlamydial immune dampening strategies, and putative positive feedback mechanisms for Chlamydia-induced fibrotic scarring. In summary, hRNA-Seq helps to reveal the complex interplay between invading bacterial pathogens and their host mammalian cells and is immediately applicable to any bacteria/host cell interaction.

sample_organism:
Chlamydia trachomatis
Homo sapiens

platform_organism:
Chlamydia trachomatis
Homo sapiens

overall_design: Development of heterogeneous RNA-Seq (hRNA-Seq) to simultaneously capture prokaryotic and eukaryotic expression profiles of bacteria-infected cells.  This Series reports Chalmydia expression profiles only.

first: Michael

last: Humphrys

first: Todd

last: Creasy

first: Yezhou

last: Sun

first: Marcus

last: Chibucos

first: Anup

last: Mahurkar

first: Elliott

middle: F

last: Drabek

first: Umar

last: Farooq

first: Naomi

last: Sengalamay

first: Sandy

last: Ott

first: Huizhong

last: Shou

first: Patrik

last: Bavoil

first: Claire

last: Fraser

first: Garry

last: Myers


title: Age-related methylation changes are associated with altered transcriptional circuitry

status: Public on Dec 20 2013

city: Heidelberg

first: Guenter

last: Raddatz

address: Im Neuenheimer Feld 580

institute: German Center for Cancer Research

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

overall_design: Refer to individual Series


title: Polycomb-dependent H3K27me1 and H3K27me2 characterization in mouse ES cells (E36 with H3K27ac, H3K27me1, and H3K4me1/3)

city: Milan

first: Sri Ganesh

last: Jammula

institute: IFOM-IEO

summary: These data include the genome wide location analysis of H3K27me1/2/ac and H3K4me1/3  by ChIP in mouse ES cells.

overall_design: Precipitation of formaldehyde cross-linked chromatin prepared from mouse ES cell using specific antibodies against Histone H3 and its specific modifications. Rabbit IgG were used as control.

first: Karin

last: Ferrari

first: Sri

first: Diego

last: Pasini


title: Differential Methylation Of The TRPA1 Promoter In Pain Sensitivity: MeDIP-sequencing

status: Public on Dec 04 2013

city: London

first: Jordana

last: Bell

address: St Thomas' Hospital, Westminster Bridge Rd

institute: King's College London

summary: Chronic pain is a global public health problem, but the underlying molecular mechanisms are not fully understood. Here we examine genome-wide DNA methylation, first in 50 identical twins discordant for heat pain sensitivity and then in 50 further unrelated individuals. Whole blood DNA methylation was characterized at 5.2 million loci by MeDIP-sequencing and assessed longitudinally to identify differentially methylated regions associated with high or low pain-sensitivity (pain-DMRs). Nine meta-analysis pain-DMRs show robust evidence for association (false discovery rate 5%) with the strongest signal in the pain gene TRPA1 (P=1.2×10-13). Several pain-DMRs show longitudinal stability consistent with susceptibility effects, have similar methylation levels in brain, and altered expression in skin. Our approach identifies epigenetic changes in both novel and established candidate genes that provide molecular insights into pain and may generalize to other complex traits.

overall_design: MeDIP-sequencing in 100 individulas using a 2 stage design: paired-end MeDIP-seq in 50 monozygotic twins and single-end MeDIP-seq in 50 unrelated individuals.

middle: T


title: Evolution of Translation Regulation in budding Yeast

city: Pittsburgh

first: Charles

middle: Joel

last: McManus

state: Pennsylvania

address: 4400 Fifth Ave

department: Biological Sciences

institute: Carnegie Mellon University

summary: Comparison of translation efficiency in S. cerevisiae, S. paradoxus, and their F1 hybrid.

sample_organism: Saccharomyces cerevisiae x Saccharomyces paradoxus

overall_design: SRA submission number SRP028552; BioProject number PRJNA213844; Ribosome profiling was used to compare mRNA abundance, ribosome occupancy, and translation efficiency in two yeast species and their F1 hybrid.

first: Gemma

middle: E

last: May

first: Pieter

last: Spealman

first: Alan

last: Shteyman


title: Polycomb-dependent H3K27me1 and H3K27me2 characterization in mouse ES cells


title: Small and large RNA sequencing of a Kras G12D mouse model of lung adenocarcinoma

city: Seattle

first: Paul

last: Valdmanis

state: WA

address: 1705 NE Pacific St, HSB J-309

department: Medicine

institute: University of Washington

summary: Mice bearing a G12D activating mutation in Kras consistently develop lung adenocarcinomas in a manner analogous to humans.  By performing small and large RNA sequencing on KrasG12D tumors from F1 hybrid mice we were able to identify genes and microRNAs differentially expressed in these tumor samples.  Quantification of reads that cover single nucleotide polymorphisms that distinguish between the parental mouse strains enabled an analysis of allele specific expression and imprinting status in these tumors.

overall_design: mRNA and small RNA fractions of mouse lungs and lung adenocarcinomas were deep sequenced in triplicate

middle: N

first: Mark

last: Kay


title: Spatial Regulation Dominates Gene Function in the Ganglia Chain

city: Ramat Gan

first: Dror

last: Hibsh

address: Bar Ilan

department: Bioinformatics

country: Israel

institute: Bar Ilan University

summary: Spatial regulation analysis across multiple condition comparisons revealed distinct patterns of gene expression. We combined these transcriptome data with spatial CNS data to produce the spatio-transcripto map of the ganglia chain. The Hirudo Medicinalis set of transcripts generated here provides a resource for gene discovery and gene regulation within the nervous system. In addition, the strategy for de novo assembly of transcriptome data presented here may be helpful in other similar transcriptome studies.

overall_design: Examination of 3 different ganglia in 3 different leeches.


title: ChIP-seq analysis of PHF20 and Wdr5 binding sites in ESCs

city: Houston

first: Stephen

last: Ayers

state: Texas

address: 6565 Fannin Street F8-060

department: Genomic Medicine

institute: Methodist Hospital Research Institute

summary: Although reprogramming of somatic cells to generate inducible pluripotent stem cells (iPSCs) is associated with remarkable epigenetic changes, the role and mechanisms of epigenetic factors in this process remains poorly understood. Here we describe identification of Jmjd3 as a potent negative regulator of reprogramming. Jmjd3-deficient MEFs produced significantly more iPSC colonies than did wild-type cells, while ectopic expression of Jmjd3 markedly inhibited reprogramming. We further show that the inhibitory effects of Jmjd3 are produced through both histone demethylase-dependent and -independent pathways acting in concert. The latter pathway is entirely novel and involved Jmjd3 targeting of PHF20 for ubiquitination and degradation by recruiting an E3 ligase Trim26. Importantly, PHF20-deficient MEFs could not be converted to fully reprogrammed iPSCs, even with knockdown of Jmjd3, Ink4a or p21, indicating dominant effects of this protein on reprogramming. Our findings identify a previously unrecognized role of Jmjd3 in reprogramming and provide molecular insight into the mechanisms by which the Jmjd3-PHF20 axis controls the reprogramming process.

overall_design: This study was an examination of phf20 and wdr5 binding patterns in embryonic stem cells, using a ChIP-Sequencing methodology with antibodies for each of these factors in the cell lines indicated.

first: Rongfu

last: Wang

first: Wei

last: Zhao


title: Analysis of the regulation of circadian gene expression in cyanobacteria with RNA sequencing

city: Cambridge

first: Joseph

middle: Scott

last: Markson

address: 52 Oxford St, Room 445

department: Molecular and Cellular Biology

institute: Harvard Univ / Howard Hughes Medical Institut

summary: The cyanobacterium Synechococcus elongatus contains a circadian clock which coordinates circadian changes in gene expression of a large percentage of its genes. The response regulator RpaA has been implicated as an important regulator of many circadian genes, but the role of this protein in regulating changes in gene expression genome-wide is not known. We show that deletion of rpaA abrogates circadian gene expression genome-wide and arrests cells in a gene expression state highly similar to that of wildtype cells in the morning. Furthermore, we show that RpaA binds DNA in an circadian manner that is dependent on phosphorylation of the protein. To demonstrate the sufficiency of phosphorylated RpaA in driving global changes in gene expression, we used RNA sequencing to measure changes in gene expression elicited by a phosphomimetic of RpaA (RpaA D53E) and compared these changes to those that occur during a circadian cycle in wildtype cells. This analysis reveals that induction of RpaA D53E is sufficient to drive all circadian gene expression changes that happen from dawn to dusk in wildtype cells. Interestingly, the dynamics of gene expression elicited by RpaA D53E induction mirror those observed during a circadian cycle in wildtype cells, suggesting that the dynamics of circadian gene expression and hard-wired in the regulon downstream of RpaA.

sample_organism: Synechococcus elongatus PCC 7942 = FACHB-805

overall_design: Enriched mRNA was prepared from synchronized wildtype S. elongatus cells every four hours over a circadian cycle and sequenced using the Illumina TruSeq Stranded mRNA Sample Prep Kit and Illumina HiSeq technology. To test the role of phosphorylated RpaA in driving circadian gene expression, we generated a strain which we refer to as OX-D53E that lacks core clock components (ΔrpaA, ΔkaiBC) with an RpaA phosphomimetic (RpaA D53E) under the control of an IPTG-inducible promoter (Ptrc::rpaAD53E). We used the same methodology to measure gene expression in OX-D53E before and after induction of RpaA D53E. As a control, we also measured gene expression in the OX-D53E strain over time in the absence of IPTG. Also, we generated a strain similar to OX-D53E in which the only difference was that no gene was inserted downstream of the IPTG inducible promoter (OX-Empty). We measured gene expressio in OX-Empty before and after IPTG addition to test for off-target effects of IPTG.

middle: S

middle: R

last: Piechura

first: Anna

last: Puszynska

first: Erin

last: O'Shea


title: ChIP-Seq analysis of RpaA binding dynamics

summary: The response regulator RpaA is required for control of genome-wide gene expression by the cyanobacterial circadian clock.  RpaA is predicted to be a DNA binding protein based on sequence homology, but prior studies have been unable to detect binding in vitro or in vivo to a small panel of promoters.  We used ChIP-Seq to determine whether RpaA associates with DNA in vivo, and if so, with what dynamics.  We find that RpaA binds to over 100 location in the genome in a circadian manner, with strongest binding occuring around subjective dusk.  Analysis of these binding sites shows that RpaA directly regulates the expression of clock components to generate feedback on the core oscillator, and also regulates expression of a small set of circadian effectors that in turn orchestrate global expression rhythms.

platform_organism: Synechococcus elongatus PCC 7942 = FACHB-805

overall_design: Crosslinked samples were acquired every four hours from a turbidostatic wild-type (AMC408) cultures in constant light (LL) following entrainment with two light-dark (LD) cycles.  As a negative control, we acquired samples similarly from an ΔrpaA culture.  Chromatin immunoprecipitation was performed on each sample from wht wild-type and from a pool of all samples from the ΔrpaA culture.  Libraries were prepared from the ChIP samples and sequenced with Illumina technology.  For the wild-type, biological replicate samples were acquired at times of maximum and minimum RpaA binding.

middle: K


title: Plasmodium falciparum NF54 Transcriptome

status: Public on Dec 03 2013

city: Rockville

first: Ewen

last: Kirkness

address: 9704 Medical Center  Drive

institute: JCVI

summary: Transcriptomic Analysis of Cultured Sporozoites of P. falciparum

overall_design: RNA-seq reads from each of three developmental stages (2 replicates per sample) were mapped to the reference Plasmodium falciparum genome, and gene expression levels were calculated for each sample.

first: Elisabet

last: Caler

middle: L

last: Hoffman

first: Christopher

middle: D

last: Town


title: Genome-wide maps of H3K27me3 in WT (wild type) and Epi-df (mutant) rice

city: Beijing

first: Liguo

last: Zhang

address: South Zhongguancun Street

institute: Chinese Academy of Agricultural Sciences

summary: DNA methylation and histone H3 lysine 9 dimethylation (H3K9me2) are important epigenetic repression marks for silencing transposons in heterochromatin and regulating gene expression in plant development. However, the mechanistic relationship to other repressive marks, such as histone H3 lysine 27 trimethylation (H3K27me3), is unclear. OsFIE1 (Fertilization Independent Endosperm 1) encodes an Esc-like core component of the Polycomb repressive complex 2 (PRC2), which is involved in H3K27me3-mediated gene repression. Here, we identify a gain-of-function epi-allele (Epi-df) of rice OsFIE1; this allele exhibits a dwarf stature and various floral defects that are inherited in a dominant fashion. We found that Epi-df has no changes in its nucleotide sequence, but is hypo-methylated in the promoter and the 5' region of OsFIE1 and has reduced H3K9me2 and increased H3K4me3. In Epi-df, OsFIE1 was ectopically expressed and its imprinting status was disrupted. OsFIE1 interacted with rice E(z) homologs, consistent with its role in H3K27me3 repression. Ectopic expression of OsFIE1 in Epi-df resulted in alteration of H3K27me3 levels in hundreds of genes. Therefore, this work identifies a novel epi-allele involved in H3K27me3-mediated gene repression, that itself is highly regulated by histone H3K9me2, thereby shedding light on the link between two important epigenetic marks regulating rice development.

overall_design: We report the application of ChIP-Seq technology for high-throughput profiling of histone modifications in WT (wild type) and Epi-df (mutant). We demonstrate that the H3K27me3 status is perturbed at target genes and leads to mis-regulated expression in Epi-df.

first: Jianmin

last: Wan


title: Overexpression of Dimethylarginine Dimethylaminohydrolase 1 Attenuates Airway Inflammation in a Mouse Model of Asthma

city: Cincinnati

first: Mario

last: Medvedovic

state: OH

address: 3223 Eden Av. ML 56

department: Department of Environmental Health

institute: University of Cincinnati

summary: Levels of asymmetric dimethylarginine (ADMA), an endogenous inhibitor of nitric oxide synthase, are increased in lung, sputum, exhaled breath condensate and plasma samples from asthma patients. ADMA is metabolized primarily by dimethylarginine dimethylaminohydrolase 1 (DDAH1) and DDAH2. We determined the effect of DDAH1 overexpression on development of allergic inflammation in a mouse model of asthma. The expression of DDAH1 and DDAH2 in mouse lungs was determined by RT-quantitative PCR (qPCR). ADMA levels in bronchoalveolar lavage fluid (BALF) and serum samples were determined by mass spectrometry. Wild type and DDAH1-transgenic mice were intratracheally challenged with PBS or house dust mite (HDM). Airway inflammation was assessed by bronchoalveolar lavage (BAL) total and differential cell counts. The levels of IgE and IgG1 in BALF and serum samples were determined by ELISA. Gene expression in lungs was determined by RNA-Seq and RT-qPCR. Our data showed that the expression of DDAH1 and DDAH2 was decreased in the lungs of mice following HDM exposure, which correlated with increased ADMA levels in BALF and serum. Transgenic overexpression of DDAH1 resulted in decreased BAL total cell and eosinophil numbers following HDM exposure. Total IgE levels in BALF and serum were decreased in HDM-exposed DDAH1-transgenic mice compared to HDM-exposed wild type mice. RNA-Seq results showed downregulation of genes in the inducible nitric oxide synthase (iNOS) signaling pathway in PBS-treated DDAH1-transgenic mice versus PBS-treated wild type mice and downregulation of genes in IL-13/FOXA2 signaling pathway in HDM-treated DDAH1-transgenic mice versus HDM-treated wild type mice. Our findings suggest that decreased expression of DDAH1 and DDAH2 in the lungs may contribute to allergic asthma and overexpression of DDAH1 attenuates allergen-induced airway inflammation through modulation of Th2 responses.

overall_design: mRNA profiles of WT and DDAH1-transgenic mice treated with PBS or house dust mite (HDM).

first: Kayla

last: Kinker

first: Aaron

middle: M

last: Gibson

first: Stacey

last: Bass

first: Brandy

last: Day

first: Jingyuan

last: Deng

first: Gurjit

last: Khurana Hershey

first: Weiguo

last: Chen

first: Julio

last: Landero Figueroa


title: A Novel Gene Regulates Vesiculogenesis and Immunomodulation in Mycobacterium tuberculosis

city: New York

first: Olivier

last: Elemento

state: NY

address: 1305 York Avenue

institute: WEILL MEDICAL COLLEGE OF CORNELL UNIV

summary: After co-evolving with humans, its only major host, Mycobacterium tuberculosis (Mtb) restrains immune responses well enough to escape eradication, yet elicits enough immunopathology to ensure its transmission. Here, we provide evidence that this balance is regulated in part by a previously uncharacterized, cytosolic, membrane-associated protein with a novel structural fold, encoded by the Mtb gene rv0431. The protein acts by regulating the quantity and quality of Mtb-derived membrane vesicles bearing TLR2 ligands, including the lipoproteins LpqH and SodC. We propose that rv0431 be named virR (vesiculogenesis and immune response regulator). To our knowledge, VirR is the first bacterial protein identified to regulate vesiculogenesis.

overall_design: Transcriptome sequencing of mouse macrophages uninfected, infected with WT Mtb, or infected with rv0431 mutant Mtb.

first: Poonam

last: Rath

first: Carl

last: Nathan


title: Circadian control of global gene expression by the cyanobacterial master regulator RpaA

first: Joseph

last: Markson


title: In vivo transcriptional goverance of hair follicle stem cells by canonical Wnt regulators

first: Wen-Hui

last: Lien

address: 1230 York Ave Box#300

department: Mammalian cell biology and development

institute: The Rockefeller Unoversity

summary: Mouse hair follicles undergo synchronized cycles. Cyclical regeneration and hair growth is fueled by hair follicle stem cells (HFSCs). HFSCs regenerate hair in response to canonical Wnt signalling. We used RNA-seq to unfold genome-wide chromatin landscapes of β-catenin within the native HFSC-niche.

overall_design:
β-catenin control and cKO hair follicle stem cells (HFSCs) at the onset of anagen skins were FACS-purified for RNA-sequcencing.

Telogen quiescent hair follicle stem cells (HFSCs) were FACS-purified for ChIP-sequcencing.

Telogen>anagen activated bulge hair follicle stem cells (HFSCs) were FACS-purified for RNA-sequcencing.

first: Elaine

last: Fuchs


title: Global analyses of the effect of different cellular contexts on microRNA targeting (3P-Seq Mouse)

status: Public on Dec 02 2013

city: Seoul

first: Jin-Wu

last: Nam

address: 222 Wangsimni Seongdong-gu

country: South Korea

institute: Hanyang University

summary: PolyA, Position, Profiling (3P-seq) for mouse tissues and cell lines

overall_design: There is no biological replicates. For six mouse tissues, 3P-seq libraries were constructed in wild-type and miR-22 knockout mouse.

first: Olivia

last: Rissland

first: David

last: Koppstein

middle: P

last: Bartel


title: Independent Regulation of Metabolism but Coordinated Control of Tissue Development by Epidermal Specific Transcription Factors in Caenorhabditis elegans

first: Jiaofang

last: Shao

address: 6431 Fannin St

department: Department of Biochemistry and Molecular Biology

institute: McGovern Medical School, University of Texas Health Science Center

summary: We identified target genes for NHR-25 by ChIP-seq at L1 stage of C. elegans. Transcription factor genes were tagged with GFP and their expression examined at L1 stage. Since there are no direct target genes known for NHR-25 that can be used for assessment of enrichment efficiency by quantitative PCR (qPCR), we chose to repeat ChIP-seq experiment of another GFP tagged transcription factor, PHA-4 for which the ChIP-seq was performed during a pilot experiment of modENCODE project using the same transgenic strain and antibody (a gift from Tony Hyman lab).

overall_design: pha-4 and nhr-25 transgenic worm were studied in Fed L1 stage.

first: Zhongying

last: Zhao


title: Genome-wide regulatory analysis reveals T-bet controls Th17 lineage differentiation through direct suppression of IRF4

city: Liverpool

first: Aditi

last: Kanhere

address: Institute of Systems, Molecular and Integrative Biology

institute: University of Liverpool

summary: The complex relationship between Th1 and Th17 cells is incompletely understood. The transcription factor T-bet is best known as the master regulator of Th1 lineage commitment. However, attention is now focused on the repression of alternate T cell subsets mediated by T-bet, particularly the Th17 lineage. Specifically it has recently been suggested that pathogenic Th17 cells express T-bet and are dependent on IL-23. However, T-bet has previously been shown to be a negative regulator of Th17 cells. We have taken an unbiased approach to determine the functional impact of T-bet on Th17 lineage commitment. Genome-wide analysis of functional T-bet binding sites provides an improved understanding of the transcriptional regulation mediated by T-bet, and suggests novel mechanisms by which T-bet regulates T helper cell differentiation. Specifically, we show that T-bet negatively regulates Th17 lineage commitment via direct repression of the transcription factor interferon regulatory factor-4 (IRF4). An in vivo analysis of the pathogenicity of T-bet deficient T cells demonstrated that Th17 responses were augmented in the absence of T-bet, and we have defined a critical temporal window for T-bet function. The interaction of the two key transcription factors T-bet and IRF4 during the determination of T cell fate choice significantly advances our understanding of the mechanisms underlying the development of pathogenic T cells.

overall_design: ChIP-seq analysis of T-bet in WT and Tbet -/- mice.

first: M

last: Gökmen

first: Richard

last: Jenner

first: Graham

last: Lord


title: Global analyses of the effect of different cellular contexts on microRNA targeting (RNA-Seq)

summary: RNA-seqs followed by miRNA transfections (miR-124 and miR-155) into four different cell lines( HeLa, HEK293, Huh7, and IMR90).

overall_design: There are two biological replicates of RNA-seqs per each miRNA transfection per each sample and there are corresponding mock transfections.


title: Role of SWI/SNF in acute leukemia maintenance and enhancer-mediated Myc regulation (4C-seq)

city: Cold Spring Harbor

first: Christopher

last: Vakoc

address: 1 Bungtown Rd

institute: Cold Spring Harbor Lab

summary: Cancer cells frequently depend on chromatin regulatory activities to maintain a malignant phenotype. Here, we show that leukemia cells require the mammalian SWI/SNF chromatin remodeling complex for their survival and aberrant self-renewal potential. While Brg1, an ATPase subunit of SWI/SNF, is known to suppress tumor formation in several cancer types, we found that leukemia cells instead rely on Brg1 to support their oncogenic transcriptional program, which includes Myc as one of its key targets. To account for this context-specific function, we identify a cluster of lineage-specific enhancers located 1.7 megabases downstream of Myc that are occupied by SWI/SNF, as well as the BET protein Brd4. Brg1 is required at these distal elements to maintain transcription factor occupancy and for long-range chromatin looping interactions with the Myc promoter. Notably, these distal Myc enhancers coincide with a region that is focally amplified in 3% of acute myeloid leukemia. Together, these findings define a leukemia maintenance function for SWI/SNF that is linked to enhancer-mediated gene regulation, providing general insights into how cancer cells exploit transcriptional coactivators to maintain oncogenic gene expression programs

overall_design: Enhancer usually regulates its targets through physical contact/interaction. In order to study chromosome conformation of Myc locus and potential distal enhancer E1-E5 region in murine AML cells, we utilize the high resolution 4C-seq and analysis pipeline to search cis elements that physical interact with Myc and E1-E5 region through setting up two individual viewpoints in these two regions.

first: Junwei

last: Shi

first: Cinthya

last: Zepeda-Mendoza

last: Spector


title: Global analyses of the effect of different cellular contexts on microRNA targeting (3P-Seq Human)

summary: PolyA, Position, Profiling (3P-seq) for human cell lines

overall_design: There is no biological replicates.

first: Calvin

last: Jan


title: Global analyses of the effect of different cellular contexts on microRNA targeting

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

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Refer to individual Series


title: HDAC3-dependent regulation of H3K9 acetylation in IECs

city: Philadelphia

first: Kyoung Jae

last: Won

address: 3400 Civic Center Blvd

institute: University of Pennsylvania

summary: The development and severity of inflammatory bowel diseases (IBD) and other chronic inflammatory conditions can be influenced by host genetic and environmental factors, including signals derived from commensal bacteria.  However, the mechanisms that integrate these diverse cues remain undefined. Here we demonstrate that intestinal epithelial cells (IECs) isolated from IBD patients exhibit decreased expression of the epigenome-modifying enzyme histone deacetylase 3 (HDAC3). Further, genome-wide analyses of murine IECs that lack HDAC3 (HDAC3ΔIEC) revealed that HDAC3 deficiency resulted in dysregulated gene expression coupled with alterations in histone acetylation.  Critically, conventionally-housed HDAC3ΔIEC mice demonstrated loss of Paneth cells, impaired IEC function and alterations in the composition of intestinal commensal bacteria.  In addition, HDAC3ΔIEC mice exhibited significantly increased susceptibility to intestinal damage and inflammation, indicating that epithelial expression of HDAC3 plays a central role in maintaining intestinal homeostasis.  Strikingly, rederivation of HDAC3ΔIEC mice into germ-free conditions revealed that dysregulated IEC gene expression, Paneth cell homeostasis, and intestinal barrier function were largely restored in the absence of commensal bacteria.  Collectively, these data indicate that the HDAC3 is a critical factor that integrates commensal bacteria-derived signals to calibrate epithelial cell responses required to establish normal host-commensal relationships and maintain intestinal homeostasis.

overall_design: Analyses of histone acetylation in primary IECs from HDAC3FF (3 biologic replicates) and HDAC3ΔIEC (3 biologic replicates) mice were conducted utilizing ChIP-seq for H3K9Ac.

last: Artis

first: Theresa

last: Alenghat

first: Inchan

last: Choi

first: Kyoung-Jae


title: Role of SWI/SNF in acute leukemia maintenance and enhancer-mediated Myc regulation


title: A microRNA-directed program of cytotoxic CD8+ T cell differentiation

first: Tarmo

last: Äijö

address: 162 5th Avenue

department: Center for Computational Biology

institute: Flatiron Institute

summary: Acquisition of effector properties is a key step in the generation of cytotoxic T lymphocytes (CTLs). Here we show that inflammatory signals regulate Dicer expression in CTL, and that deletion or depletion of Dicer in mouse or human activated CD8+ T cells causes upregulation of perforin, granzyme and effector cytokines. Genome-wide analysis of miRNA changes induced by exposure of differentiating CTLs to IL-2 and inflammatory signals identifies miR-139 and miR-150 as components of a miRNA network that controls perforin, eomesodermin (Eomes) and IL-2Ra expression in differentiating CTLs and whose activity is modulated by IL-2, inflammation and antigenic stimulation. Overall our data show that strong IL-2R and inflammatory signals act through Dicer and miRNAs to control the cytolytic program and other aspects of effector CTL differentiation.

overall_design:
Comparison of control and Dicer knock-out CTLs differentiated in vitro;
Comparison of wild type CTLs differentiated in vitro with or without inflammatory stimuli;
Comparison of effector and memory precursor CTLs isolated from mice infected with LCMV-Armstrong

first: Sara

last: Trifari

first: Anjana

last: Rao


title: Role of SWI/SNF in acute leukemia maintenance and enhancer-mediated Myc regulation [ChIP-Seq]

overall_design: In order to understanding the lineage specific requirement of coactivaor, such as Brg1 and Brd4, in AML, we performed ChIP-seq with Brg1, Brd4 together with histone modification marks in murine MLL-AF9/NrasG12D AML cell line to search tissue specific cis regulation element that can be accounted for the leukemia specific dependence.

first: Warren

last: Whyte

last: Young


title: Intertwined pathways for Argonaute-mediated microRNA biogenesis in Drosophila

first: Jakub

middle: Orzechowski

last: Westholm

address: 1275 York Avenue, Box 252

department: Developmental Biology

institute: Memorial Sloan-Kettering Cancer Center

summary: Ago1-IP small RNA sequencing from S2R+ cells, with dsRNA knockdown of Dicer1 (and GFP control)

overall_design: One Dicer knockdown sample and one GFP knockdown control.

first: Eric

last: Lai


title: Genome-wide analyses of GATA6 occupancy and functions provide insights into its oncogenic mechanisms in human gastric cancer [ChIP-Seq]

city: Boston

first: Ramesh

last: Shivdasani

state: Massachusetts

address: Dana Bldg, Room 720, 450 Brookline Ave

department: Department of Medical Oncology

institute: Dana-Farber Cancer Institute

summary: To identifiy core GATA6 functions and transcriptional targets in human gastric cancer, including additional subservient transcriptional regulators via integrative analysis of GATA6 transcription factor occupancy, gene dependency, and tumor synexpression.

overall_design: Transcription factor binding site identification ChIP-Seq performed on 4 samples.

first: Rita

last: Sulahian


title: Role of Tet1 in genomic imprinting erasure

first: Li

last: Shen

address: 200 Longwood Ave

institute: HHMI/Boston Children's Hospital/Harvard Medical School

first: Shinpei

last: Yamaguchi

first: Yuting

last: Liu

first: Damian

last: Sendler

first: Yi

last: Zhang


title: Role of Tet1 in genomic imprinting erasure [RRBS-PGC]

summary: Genomic imprinting is an allele-specific gene expression system important for mammalian development and function. The molecular basis of genomic imprinting is allele-specific DNA methylation 2. While it is well known that the de novo DNA methyltransferases Dnmt3a/b are responsible for the establishment of genomic imprinting, how the methylation mark is erased during primordial germ cell (PGC) reprogramming remains a mystery. Here we report that Tet1 plays a critical role in the erasure of genomic imprinting. We show that despite their identical genotype, progenies derived from mating between Tet1-KO males and wild-type females exhibit a number of variable phenotypes including placental, fetal and postnatal growth defects, and early embryonic lethality. These defects are, at least in part, caused by the dysregulation of imprinted genes, such as Peg10 and Peg3, which exhibit aberrant hypermethylation in the paternal allele of differential methylated regions (DMRs). RNA-seq reveals extensive dysregulation of imprinted genes in the next generation due to paternal functional loss of Tet1. Genome-wide DNA methylation analysis of E13.5 PGCs and sperm derived from Tet1-KO mice reveals hypermethylation of DMRs of imprinted genes in sperm, which can be traced back to PGCs. Dynamics of methylation change in Tet1-affected sites suggested that Tet1 swipes remaining methylation including imprinted genes at late reprogramming stage. We also revealed that Tet1play a role in paternal imprinting erasure in females germline. Thus, our study establishes a critical function for Tet1 in the erasure of genomic imprinting.

overall_design: Genome-wide DNA methylation analysis of E13.5 PGCs from control and Tet1-KO mice


title: Role of Tet1 in genomic imprinting erasure [RRBS-sperm]

overall_design: Genome-wide DNA methylation analysis of sperm derived from control and Tet1-KO mice


title: A Cdx4-Sall4 regulatory module controls the transition from mesoderm formation to embryonic hematopoiesis

city: University Park

first: Shaun

last: Mahony

state: PA

address: 404 South Frear Bldg

department: Biochemistry & Molecular Biology

institute: Penn State University

summary: Deletion of caudal/cdx genes alters hox gene expression and causes defects in posterior tissues and hematopoiesis. Yet, the defects in hox gene expression only partially explain these phenotypes. To gain deeper insight into Cdx4 function, we performed ChIP-seq combined with gene expression profiling in zebrafish, and identified the transcription factor spalt-like 4 (sall4) as a Cdx4 target. ChIP-seq revealed that Sall4 bound to its own gene locus and the cdx4 locus. Expression profiling showed that Cdx4 and Sall4 co-regulate genes such as hox, scl, and lmo2 that initiate hematopoiesis. Combined cdx4/sall4 gene knock-down impairs erythropoiesis, and overexpression of the Cdx4 and Sall4 target genes scl and lmo2 together rescued the erythroid program. These findings suggest that auto- and cross- regulation of Cdx4 and Sall4 establish a stable molecular circuit in mesoderm that facilitates the activation of the blood-specific program as development proceeds.

overall_design: ChIP-seq was performed against Cdx4, Sall4, H3K27ac, and H3K4me3 in bud-stage zebrafish embryos. Input material was sequenced as controls.

first: Elizabeth

middle: J

last: Paik

first: Richard

last: White

first: Emily

last: Price

first: Bilguujin

last: Dorjsuren

first: Anthony

last: DiBiase

first: Christian

last: Mosimann

first: Alan

last: Davidson

first: David

last: Gifford

first: Leonard

middle: I

last: Zon


title: Role of Tet1 in genomic imprinting erasure [RNA-seq]

overall_design: Gene expression analysis of E9.5 embryos


title: Role of SWI/SNF in acute leukemia maintenance and enhancer-mediated Myc regulation (RNA-seq)

first: Christopher

last: Vakoc

overall_design: To profile the basal transcription level, we performed NSR and PolyA+ (illumine TruSeq) in a murine AML RN2 cell lines. To define the rapid downregulated genes in response to JQ1, BET bromodomian inhibitor, in RN2 cell, we performed RNA-seq in RN2 exposing to 250nM JQ1 for 48h time course.

first: Junwei

last: Shi

last: Kawaoka

first: Zhu

last: Zhu

first: Jude

last: Kendall

first: Michael

middle: A

last: Wigler


title: Evolutionary conservation of the eumetazoan gene regulatory landscape

city: Vienna

first: Michaela

last: Schwaiger

address: Althanstr. 14

country: Austria

institute: University of Vienna

summary: Bilaterian animals differ from other metazoans in their apparent bilateral symmetry and the development of a third germ layer. Both might have facilitated the evolution of the diverse and complex bilaterian body plans. The first cnidarian genome sequence revealed that despite their morphological simplicity, this sister group to all bilaterians shares an immense genomic complexity with vertebrates. This suggested that it might have been the complexity of gene regulation which increased during the evolution of bilaterians. We compared the gene regulatory landscape of cnidarians and bilaterians. To this end we generated the first genome-wide prediction of gene regulatory elements and profiled five epigenetic marks in a non-bilaterian animal, the cnidarian Nematostella vectensis. We found that the location of chromatin modifications relative to genes and distal enhancers is conserved among eumetazoans. Surprisingly, the genomic landscape of gene regulatory elements is highly similar between Nematostella and bilaterian model organisms. This suggests that complex regulation of developmental gene expression evolved in eumetazoans without a major increase in complexity in bilaterians.

overall_design: ChIP-seq of p300, RNA Pol2, and five histone modifications in Nematostella vectensis.

first: Anna

last: Schönauer

first: André

middle: F

last: Rendeiro

first: Carina

last: Pribitzer

first: Alexandra

last: Schauer

last: Gilles

first: Johannes

last: Schinko

first: Eduard

last: Renfer

last: Fredman

first: Ulrich

last: Technau


title: Sequence-specific mono-uridylation of mature miRNAs by Zcchc6 and Zcchc11

status: Public on Dec 01 2013

first: Peng

last: Du

state: MA

address: 1 Blackfan Circle

institute: Boston Children’s Hospital, Harvard Medical School

summary: Recent small RNA sequencing data has uncovered extensive modification of the 3’ end of mature microRNAs (miRNAs). This non-templated nucleotide addition can impact miRNA gene regulatory networks through the control of miRNA stability or by interfering with the repression of target mRNAs. The miRNA modifying enzymes responsible for this regulation remain largely uncharacterized. Here we describe the ability for two related terminal uridyl transferases (TUTases), Zcchc6 (TUT7) and Zcchc11 (TUT4), to 3’ mono-uridylate a specific subset of miRNAs involved in cell differentiation and Hox gene control. Zcchc6/11 selectively uridylate these miRNAs in vitro, and we biochemically define a bipartite sequence motif that is necessary and sufficient to confer Zcchc6/11 catalyzed uridylation. Depletion of these TUTases in cultured cells causes the selective loss of 3’ mono-uridylation of many of the same miRNAs. Interestingly, upon TUTase dependent loss of uridylation we observe a concomitant increase in non-templated 3’ mono-adenylation. Our results uncover the molecular basis for sequence specific miRNA mono-uridylation by Zcchc6/11, highlight the precise control of different 3’ miRNA modifications in cells, and have implications for miRNA regulation during development.

overall_design: small RNA profiles in TUTases knock-down and control HeLa cells were generated by Illumina deep sequencing

first: Richard

last: Gregory

first: James

last: Thornton


title: Epigenetic regulation of the MEG3-DLK1 microRNA cluster in human Type 2 diabetic islets

first: Jonathan

last: Schug

address: 12-156 Smilow Ctr Trans Research

department: School of Medicine

summary: Type 2 diabetes mellitus (T2DM) is a multi-factorial disease characterized by the inability of beta-cells in the endocrine pancreas to produce sufficient amounts of insulin to overcome insulin resistance in peripheral tissue. To investigate the function of miRNAs in T2DM, we sequenced the small RNAs of human islets cells from diabetic and non-diabetic organ donors and identified a cluster of miRNAs in an imprinted locus on human chromosome 14 to be dramatically down-regulated in T2DM islets. These miRNAs are highly and specifically expressed in human beta-cells. The down-regulation of this imprinted locus strongly correlates with increased methylation of its promoter in T2DM islets, providing evidence for an epigenetic modification that contributes to the pathogenesis of T2DM. Targets of the Chr 14q32 cluster of miRNAs were identified by high-throughput sequencing of cross-linked and immunoprecipitated RNA (HITS-CLIP) of Argonaute. We have also identified a unique class of sequences, termed chimeric reads, that represent an in vivo ligation of miRNAs and their targets while in complex with Argonaute, and which allow for the direct identification of miRNA:target relationships in vivo.

overall_design: There are three experiments in this submission. All are in human islets or islet cell types. The first is a  comparison of miRNA levels in sorted alpha versus beta cells. There is one replicate for this experiment. The second  experiment is to measure the expression of miRNAs in whole islets as a function of glucose levels. There are three  levels and one replicate for each condition. The third exeriment is a comparison of whole islets taken from human  donors that were suspected/confirmed Type 2 diabetic or considered controls. There are 3 controls and 4 T2D samples.

first: Vasumathi

first: Nuria

middle: C

last: Bramswig

first: Lindsay

middle: B

last: McKenna

first: Melinda

last: Penn

first: Ying

last: Chen

first: Kyoung-Jae

last: Won

first: Inchan

last: Choi

first: Anastassios

last: Vourekas

first: Nicholas

last: Hand

first: Chengyang

last: Liu

first: Ali

last: Naji

first: Joshua

last: Friedman

first: Klaus

middle: H

last: Kaestner


title: Modelling and rescuing neurodevelopmental defect of Down syndrome using induced pluripotent stem cells from monozygotic twins discordant for trisomy 21 [RNA-seq]

city: Geneva

first: Audrey

last: Letourneau

address: Rue Michel-Servet 1

department: Genetic Medicine and Development

country: Switzerland

institute: University of Geneva Medical School

summary: Down syndrome (trisomy 21) is the most common viable chromosomal disorder with intellectual impairment and several other developmental abnormalities. Here, we report the generation and characterization of induced pluripotent stem cells (iPSCs) derived from monozygotic twins discordant for trisomy 21 in order to eliminate the effects of the variability of genomic background. The alterations observed by genetic analysis at the iPSC level and at first approximation in early development illustrate the developmental disease transcriptional signature of Down syndrome. Moreover, we observed an abnormal neural differentiation of Down syndrome iPSCs in vivo when formed teratoma in NOD-SCID mice, and in vitro when differentiated into neuroprogenitors and neurons. These defects were associated with changes in the architecture and density of neurons, astroglial and oligodendroglial cells together with misexpression of genes involved in neurogenesis, lineage specification and differentiation. Furthermore, we provide novel evidence that dual-specificity tyrosine-(Y)-phosphorylation regulated kinase 1A (DYRK1A) on chromosome 21 likely contribute to these defects. Importantly, we found that targeting DYRK1A pharmacologically or by shRNA results in a considerable correction of these defects.

overall_design: mRNA-seq profiling of iPS cells (4 euploid and 3 trisomy 21) derived from fibroblasts of monozygotic twins discordant for trisomy 21

first: Youssef

last: Hibaoui

first: Iwona

last: Grad

last: Sailani

first: Sophie

last: Dahoun

first: Federico

last: Santoni

first: Stefania

last: Gimelli

first: Michel

last: Guipponi

first: Marie

last: Pelte

first: Frédérique

last: Béna

first: Stylianos

middle: E

last: Antonarakis

first: Anis

last: Feki


title: Next generation sequencing facilitates quantitative analysis of Caenorhabditis elegans N2 wild type, emr-1(gk119), lem-2(tm1582) and emr-1(RNAi) lem-2(tm1582) transcriptomes.

city: Seville

first: Peter

last: Askjaer

address: UPO Carretera de Utrera km 1

country: Spain

institute: Centro Andaluz de Biología del Desarrollo

summary: Laminopathies are caused by mutations in components of the nuclear envelope (NE). While most NE components are widely expressed, laminopathies affect only a subset of tissues. However, the understanding of the molecular mechanisms that explain this phenomenon is still elusive. Here we have performed RNA-Seq analysis in adult C. elegans nematodes comparing gene expression in wild type and single and double mutants of two components of the NE, EMR-1 and LEM-2. Our data confirm that EMR-1 and LEM-2 facilitate gene repression and that both proteins control the expression of mainly muscle and neuronal genes.

overall_design: mRNA profiles of wild type, emr-1(gk119), lem-2(tm1582) and emr-1(RNAi) lem-2(tm1582) young adult worms were generated by deep sequencing, in triplicate for the wild type and duplicates for the other backgrounds, using Illumina GAIIx.

first: Cristina

last: González-Aguilera