title: Novel nucleosomal particles containing core histones and linker DNA but no histone H1

city: Bethesda

middle: Johannes

last: Clark

address: 6 Center Drive Bldg 6A Rm 2A02

department: DDB

institute: NICHD, NIH

summary: Eukaryotic chromosomal DNA is assembled into regularly spaced nucleosomes, which play a central role in gene regulation by determining accessibility of control regions. The nucleosome contains ~147 bp of DNA wrapped ~1.7 times around a central core histone octamer. The linker histone, H1, binds both to the nucleosome, sealing the DNA coils, and to the linker DNA between nucleosomes, directing chromatin folding. Micrococcal nuclease (MNase) digests the linker to yield the chromatosome, containing H1 and ~160 bp, and then converts it to a core particle, containing ~147 bp and no H1. Sequencing of nucleosomal DNA obtained after MNase digestion (MNase-seq) generates genome-wide nucleosome maps that are important for understanding gene regulation. We present an improved MNase-seq method involving simultaneous digestion with exonuclease III, which removes linker DNA. Remarkably, we discovered two novel intermediate particles containing 154 or 161 bp, corresponding to 7 bp protruding from one or both sides of the nucleosome core. These particles are detected in yeast lacking H1 and in H1-depleted mouse chromatin. They can be reconstituted in vitro using purified core histones and DNA. We propose that these "proto-chromatosomes" are fundamental chromatin subunits, which include the H1 binding site and influence nucleosome spacing independently of H1.

sample_organism:
Saccharomyces cerevisiae
Mus musculus
Synthetic plasmid

overall_design: Comparison of budding yeast nucleosome sequences obtained using micrococcal nuclease (MNase-seq) and MNase + exonuclease III (ExoIII) (MNase-ExoIII-seq): wild type cells and hho1-null cells. Nucleosome sequences from native chromatin and H1-depleted chromatin from mouse liver. Nucleosome sequences from a plasmid reconstituted into nucleosomes using recombinant yeast histones or native chicken erythrocyte histones.

first: Hope

last: Cole

first: Josefina

last: Ocampo

first: Tara

last: Burke

first: Tatiana

last: Nikitina

first: V

last: Nagarajavel


title: Genome-wide map of H3K4me3 in HEK293 cells after PRDM9 expression

status: Public on Sep 21 2015

city: Bar Harbor

first: Christopher

middle: Lee

last: Baker

state: ME

address: 600 Main Street

department: Research

country: USA

institute: The Jackson Laboratory

type: Genome binding/occupancy profiling by high throughput sequencing

summary: PRDM9 is a histone methyltransferase expressed in meiotic germ cells that determines the location of genetic recombination hotspots through binding of its allele-specific DNA binding domain. Here we characterize the genome-wide chromatin modification for two human PRDM9 alleles (A and C) in human cell lines. HEK293 cells were transfected with both alleles and an empty vector control. Resulting chromatin was subjected to H3K4me3 ChIP followed by high-throughput sequencing. We find that different PRDM9 allele largely modified chromatin in entirely different genomic regions in somatic cells determined by the protein's zinc-finger DNA binding domains. Many of the allele-specific peaks overlap sites of meiotic double-strand breaks found in vivo in human germ cells suggesting that transient expression of PRDM9 in somatic cells can reflect binding in vivo.

overall_design: Identify PRDM9-dependent H3K4me3 sites by comparing modified chromatin after expression of different human PRDM9 alleles in HEK293 cells.

middle: L

first: Kenneth

last: Paigen


title: De novo transcriptome assembly and comparative analysis elucidate complicated mechanism regulating Astragalus chrysochlorus response to selenium stimuli

status: Public on Sep 20 2015

city: Istanbul

first: Ozgur

last: Cakir

address: Vezneciler

country: Turkey

institute: Istanbul University

type: Expression profiling by high throughput sequencing

summary: We assembled the transciptome of A. chrysochlorus by Illumina Hi-seq sequencing, and analyzed the differentially expressed genes in A. chrysochlorus responsive to Se exposure

overall_design: Examination of Control callus cultures and Se treated callus cultures

first: Neslihan

middle: T

last: Kara

first: Şule

last: Arı

first: Baohong

last: Zhang


title: Genome wide ChIP-seq analysis of human TOP2B occupancy in MCF7 breast cancer epithelial cells

city: Newcastle

first: Simon

last: Cockell

address: Framlington Place

country: United Kingdom

institute: Newcastle University

summary: TOP2B is involved in transcriptional initiation in response to nuclear hormone ligands and plays a role in transcriptional elongation. Whole genome TOP2B ChIP-seq was carried out on human MCF7 cells in the presence and absence of the nuclear hormone estradiol. Three peak calling methods were used and the peaks identified by at least two methods were analyzed further. Approximately half of the peaks fell either within a gene or within 5Kb of a transcription start site. The coincidence of TOP2B peaks and gene promoters was analyzed; TOP2B peaks were less frequently associated with promoters in estradiol treated than in control cells, suggesting a role of TOP2B in repression of transcription or a transient role in estradiol induced transcriptional changes.

overall_design: Whole genome TOP2B ChIP-seq was carried out on human MCF7 cells in the presence (30 mins exposure) and absence of the nuclear hormone estradiol. "Input" control samples were also sequenced for background detection and comparison.

first: Catriona

middle: M

last: Manville

first: Kayleigh

last: Smith

first: Zbyslaw

last: Sondka

first: Holly

last: Rance

first: Ian

middle: G

last: Cowell

first: Ka

middle: C

last: Lee

first: Nicholas

middle: J

last: Morris

first: Kay

last: Padget

first: Graham

middle: H

last: Jackson

first: Caroline

middle: A

last: Austin


title: Next Generation RNA-Sequencing data of Hematopoietic stem cells and CML stem cells

status: Public on Sep 18 2015

city: Dongjak-Gu

first: Junil

last: Kim

state: Seoul

address: 369 Sangdo-Ro

department: Department of Bioinformatics

country: South Korea

institute: SOONGSIL UNIVERSITY

summary: To investigate why dipeptides accumulate in immature CML cells, we examined upstream gene expression patterns. We isolated the most primitive long-term stem cells, short-term stem cells, and KLS- progenitor cells from healthy littermate control and CML-affected mice and performed gene expression profiling using next-generation RNA-sequencing.

overall_design: Gene expression profiles of the most primitive long-term (LT) stem cells (CD150+CD48-CD135-KLS+ cells), short-term (ST) stem cells (CD150-CD48-CD135- KLS+ cells), and KLS- progenitor cells from healthy littermate control and CML-affected mice

first: Kazuhito

last: Naka


title: Ligand-dependent genomic function of glucocorticoid receptor in triple-negative breast cancer

city: Stanford

first: Xun

last: Lan

state: CA

address: 318 Campus Dr. Room S240

department: Genetics

institute: Stanford University

summary: Glucocorticoids (GC) have been widely used as coadjuvants in the treatment of solid tumors, but GC treatment may be associated with poor pharmacotherapeutic response and/or prognosis. The genomic action of GC in these tumors is largely unknown. Here we find that dexamethasone (Dex, a synthetic GC) regulated genes in triple-negative breast cancer (TNBC) cells are associated with drug resistance. Importantly, these GC-regulated genes are aberrantly expressed in TNBC patients and associated with unfavorable clinical outcomes. Interestingly, in TNBC cells, Compound A (CpdA, a selective GR modulator) only regulates a small number of genes not involved in carcinogenesis and therapy resistance. Mechanistic studies using a ChIP-exo approach reveal that Dex- but not CpdA-liganded glucocorticoid receptor (GR) binds to a single glucocorticoid response element (GRE), which drives the expression of pro-tumorigenic genes. Our data suggest that development of safe coadjuvant therapy should consider the distinct genomic function between Dex- and CpdA-liganded GR.

overall_design: To study GR-regulated genes and define GRE in human genome, RNA-seq and GR ChIP-exo are performed in MDA-MB-231 cells before/after dex and CpdA stimulation. Each experiment includes two replicates.

first: Qianben

last: Wang

first: Zhong

last: Chen


title: Expression analysis of TCRgd+LAP+ versus TCRgd+LAP- cells

status: Public on Sep 17 2015

city: Boston

first: Rafael

middle: Machado

last: Rezende

state: MA

address: 77 Avenue Louis Pasteur

department: Neurology

institute: Brigham and Women's Hospital

summary: We used RNA-Seq to identify differential gene expression in LAP+ vs. LAP- Gamma-delta T-cells. We report a signature of 407 genes that were enriched in TCRgd+LAP+ vs. TCRgd+LAP- cells with p<0.05. Among the up-regulated genes, we found increased expression of genes related to antigen presentation, including MHC class II molecules (H2-Aa, H2-Ab1, H2-Eb1 and H2-Eb2), CD40 and CD86

overall_design: RNA-Seq of TCRgd+LAP+ versus TCRgd+LAP- cells harvested from mouse spleen and Peyer's patches; began with 20 mice for two independent experiments (10 mice each); samples were pooled and for each experiment resulted in one LAP- and one LAP+ sample for RNA-SEQ, or four samples in total; The two LAP+ were combined, resulting in three samples in total for RNA-Seq.

first: Andre

middle: P

last: da Cunha

first: Chantal

last: Kuhn

first: Stephen

last: Rubino

first: Hanane

last: M’Hamdi

first: Galina

last: Gabriely

first: Tyler

last: Vandeventer

first: Shirong

last: Liu

first: Ron

last: Cialic

first: Natalia

last: Rosa

last: Oliveira

first: Jellert

last: Gaublomme

first: Nikolaus

last: Obholzer

first: James

last: Kozubek

first: Nathalie

last: Pochet

first: Ana

last: Faria

first: Howard

last: Weiner


title: Small RNA and High Molecular Weight RNA Deep Sequencing Analysis of miR-17~92 Mimics Transfected HeLa cells

city: La Jolla

first: Changchun

last: Xiao

state: California

address: 10550 North Torrey Pines Road

department: Immunology & Microbial Science

institute: The Scripps Research Institute

type: Non-coding RNA profiling by high throughput sequencing

summary: Transient transfection of miRNA mimics are widely used to alter microRNA expression in cells. The goal of this study is to perform small RNA deep sequencing analysis of HeLa cells transfected with miR-17~92 cluster and address potential source of non-specific effects caused by this approach.

overall_design: HeLa cells were cultured as adherent cells. Equal molar concentration of miRNA mimics of miR-17~92 (100nM, 16.7nM each) were transfected and harvedsted at 6h post-transfection for small RNA deep sequencing analysis.

first: Hyun-Yong

last: Jin

first: Mohsen

last: Sabouri‐Ghomi


title: Single-cell transcriptome profiling for metastatic renal cell carcinoma patient-derived cells [RNA-seq]

city: Seoul

first: Kyu-Tae

last: Kim

address: Irwon-Ro 81

department: Samsung Genome Institute

institute: Samsung Medical Center

summary: Clear cell renal cell carcinoma (ccRCC) initiated from the renal epithelium is the most prevalent histological type of adult kidney cancers. Dissecting intratumoral heterogeneity (ITH) of ccRCC has leveraged to extend our knowledge on how primary tumors harboring driver mutations evolve and spread to other sites. The cellular fractions within and across the primary (pRCC) and metastatic RCC (mRCC) are heterogeneous in both their genetic and biological features determining the variability in clinical aggressiveness and sensitivity to the therapy. To achieve sustainable therapeutic benefit with targeted agents in mRCC, the effective target should focus on signaling pathways that are related to driver mutations occurred early in the clonal evolution of the disease and thus should be common to primary tumor and metastatic sites. Considering that extensive genetic heterogeneity may result in drug response variability among patients and treatment resistance, the tailored strategies for metastatic RCC is urgently needed. Here, we analyze single-cell RNA-seq (scRNA-seq) data from a matched primary RCC (pRCC) and lung metastasis (mRCC) to dissect ITH at the highest resolution to date with the objective of discovering the better therapeutic regimen.

overall_design: In order to identify successful clonal propagation from patient to PDX samples and understand pathogenesis from primary to metastatic RCC, we performed whole-exome sequencing (WES, n=4) and matched aCGH (n=4) on bulk tumor samples. And we utilized single-cell RNA sequencing (scRNA-seq) to model and dissect functional heterogeneity acroass primary and metastatic RCC tumors. We checked whether of capturing live one cell, not more cells, in microfluidics by fluorescent microscopic observation. To construct RNA sequencing libraries, we performed further quality controls including adequate quantities and qualities of amplified transcriptomes respectively from single cells. Tumor cells from the parental mRCC (n=34), PDX-mRCC (n=36) and PDX-pRCC (n=46) were finally analyzed in this study after filtering out poor quality cells.

first: Hye

middle: W

last: Lee

first: Hae-Ock

first: Kyeung

last: Joo

first: Woong-Yang

last: Park


title: Single-cell transcriptome profiling for metastatic renal cell carcinoma patient-derived cells

type: Genome variation profiling by genome tiling array

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

overall_design: Refer to individual Series


title: Novel IDH1 Mutant Inhibitors for Treatment of Acute Myeloid Leukemia (ERRBS)

city: Bronx

first: Britta

last: Will

state: NEW YORK

address: 1300 Morris Park Ave

department: Cell Biology

institute: Albert Einstein College of Medicine

type: Methylation profiling by high throughput sequencing

summary: Neomorphic mutations in isocitrate dehydrogenase 1 (IDH1) are driver mutations in acute myeloid leukemia (AML) and other cancers. We report the development of new allosteric inhibitors of mutant IDH1. Crystallographic and biochemical results demonstrated that compounds of this chemical series bind to an allosteric site and lock the enzyme in a catalytically inactive conformation, thereby enabling inhibition of different clinically relevant IDH1 mutants. Treatment of IDH1 mutant primary AML cells uniformly led to a decrease in intracellular 2-HG, abrogation of the myeloid differentiation block and induction of granulocytic differentiation at the level of leukemic blasts and more immature stem-like cells, in vitro and in vivo. Molecularly, treatment with the inhibitors led to a reversal of the DNA cytosine hypermethylation patterns caused by mutant IDH1 in AML patients’ cells. Our study provides proof-of-concept for the molecular and biological activity of novel allosteric inhibitors for targeting different mutant forms of IDH1 in leukemia. To obtain insight into the molecular mechanism of the novel IDH1 mutant allosteric inhibitor, primary AML cells were treated with either GSK321 IDH1 active inhibitor or Controls (DMSO or GSK990 inactive inhibitor) followed by DNA extraction for ERRBS analysis.

overall_design: Primary IDH1 mutant acute myeloid leukemia (AML) mononuclear (MNC) cells were treated in suspension cultures in differentiating media for 6 days with 3 microM GSK990 or GSK321 and an equal volume of DMSO, Followed ERRBS analysis after DNA extraction.

first: Ulrich

last: Steidl

first: Ujunwa

last: Okoye-Okafor

first: Boris

last: Bartholdy


title: Loss of Ring1B catalytic activity causes a pronounced reduction in H3K27me3 deposition yet minimally disrupts the expression of target genes

city: Edinburgh

first: Rob

middle: S

last: Illingworth

state: Midlothian

address: Centre for Regenerative Medicine

department: Centre for regenerative Medicine

institute: The University of Edinburgh


title: Genome-wide profiling of nucleosome sensitivity and chromatin accessibility in D. melanogaster [MNase]

first: Razvan

middle: V.

last: Chereji

state: Maryland

address: 6 Center Drive, Room 2A14

department: NICHD

institute: National Institutes of Health

summary: Nucleosome structure and positioning play pivotal roles in gene regulation, DNA repair and other essential processes in eukaryotic cells. Nucleosomal DNA is thought to be uniformly inaccessible to DNA binding and processing factors, such as MNase. Here, we show, however, that nucleosome accessibility and sensitivity to MNase varies. Digestion of Drosophila chromatin with two distinct concentrations of MNase revealed two types of nucleosomes: sensitive and resistant. MNase-resistant nucleosome arrays are less accessible to low concentrations of MNase, whereas MNase-sensitive arrays are degraded by high concentrations. MNase-resistant nucleosomes assemble on sequences depleted of A/T and enriched in G/C containing dinucleotides. In contrast, MNase-sensitive nucleosomes form on A/T rich sequences represented by transcription start and termination sites, enhancers and DNase hypersensitive sites. Lowering of cell growth temperature to ~10°C stabilizes MNase-sensitive nucleosomes suggesting that variations in sensitivity to MNase are related to either thermal fluctuations in chromatin fiber or the activity of enzymatic machinery. In the vicinity of active genes and DNase hypersensitive sites nucleosomes are organized into synchronous, periodic arrays. These patterns are likely to be caused by “phasing” nucleosomes off a potential barrier formed by DNA-bound factors and we provide an extensive biophysical framework to explain this effect.

overall_design: Mnase-seq, Mnase-ChIP-seq of Drosophila melanogaster embryo and S2 cells chromatin

middle: V

first: Yuri

last: Moshkin

first: Victor

last: Guryev

first: Alexandre

last: Morozov


title: RNA-Seq of Single Prostate CTCs Implicates Non-Canonical Wnt Signaling in Antiandrogen Resistance

first: Ben

middle: S.

last: Wittner

address: 149 13th Street

department: Center for Cancer Research

institute: Massachusetts General Hospital

summary: Prostate cancer is initially responsive to androgen deprivation, but the effectiveness of androgen receptor (AR) inhibitors in recurrent disease is variable. Biopsy of bone metastases is challenging, hence sampling circulating tumor cells (CTCs) may reveal drug resistance mechanisms. We established single cell RNA-sequencing profiles of 77 intact CTCs isolated from 13 patients (mean 6 CTCs/patient) using microfluidic enrichment. Single CTCs from each individual display considerable heterogeneity, including expression of AR gene mutations and splicing variants. Retrospective analysis of CTCs from patients progressing on AR inhibitor, compared with untreated cases indicates activation of noncanonical Wnt signaling (P=0.0064). Ectopic expression of Wnt5a in prostate cancer cells attenuates the antiproliferative effect of AR inhibition, while its suppression in drug-resistant cells restores partial sensitivity, a correlation also evident in an established mouse model. Thus, single cell analysis of prostate CTCs reveals heterogeneity in signaling pathways that could contribute to treatment failure.

overall_design: A total of 221 single candidate prostate CTCs were isolated from 18 patients with metastatic prostate cancer with bone metastases and 4 patients with localized prostate cancer. Of these, 133 cells (60%) had RNA of sufficient quality for amplification and next generation RNA sequencing, and 122 (55%) had >100,000 uniquely aligned sequencing reads. In addition to candidate CTCs, we also obtained comprehensive transcriptomes for 12 bulk primary prostate cancers (macrodissected for >70% tumor content), 30 single cells derived from four different prostate cancer cell lines, and 5 patient-derived leukocyte controls. The leukocytes were readily distinguished by their expression of hematopoietic lineage markers and served to exclude any CTCs with potentially contaminating signals. Strict expression thresholds were used to define lineage-confirmed CTCs, scored by prostate lineage-specific genes (PSA, PSMA, AMACR, AR) and standard epithelial markers (KRT7, KRT8, KRT18, KRT19, EpCAM). 28 cells were excluded given the presence of leukocyte transcripts suggestive of cellular contamination or misidentification during selection, and 17 cells were excluded given low expression of both prostate lineage-specific genes and 5 standard epithelial markers. The remaining 77 cells, defined as lineage-confirmed CTCs, displayed expression of either prostate lineage-specific or epithelial genes, and low expression of leukocyte-specific genes, consistent with their tumor of origin.

first: David

last: Miyamoto

first: Yu

last: Zheng

first: Richard

first: Huili

last: Zhu

first: Katherine

last: Broderick

first: Rushil

last: Desai

first: Douglas

middle: B

last: Fox

first: Brian

last: Brannigan

first: Julie

last: Trautwein

first: Kshitij

last: Arora

last: Dahl

first: Lecia

last: Sequist

first: Matthew

middle: R

last: Smith

first: Ravi

last: Kapur

first: Chin-Lee

last: Wu

first: Toshi

last: Shioda

first: Sridhar

last: Ramaswamy

last: Ting

first: Mehmet

last: Toner

first: Shyamala

last: Maheswaran

first: Daniel

last: Haber


title: Transcription factor Oct1 and its coactivator OCA-B are selectively required for CD4 memory T cell formation and function

city: Salt Lake City

first: Dean

last: Tantin

state: UT

address: 15 North Medical Drive East

department: Department of Pathology

institute: University of Utah

summary: Epigenetic changes are crucial for the generation of immunological memory1-4. Failure to generate or maintain these changes will result in poor memory responses. Similarly, augmenting or stabilizing the correct epigenetic states offers a potential method of enhancing immune memory. Yet the transcription factors that regulate these processes are poorly defined, as are the chromatin modifying complexes they recruit and the chromatin modifications they control. Using pathogen infection models and three different mouse models, including a new conditional allele, we find that the widely expressed transcription factor Oct15, and its cofactor OCA-B6,7, are selectively required the in vivo generation of functional CD4 memory. In vitro, both proteins are also required to maintain a poised state at the Il2 target locus in resting but previously stimulated CD4 T cells, and to generate robust Il2 expression upon restimulation. OCA-B is also required for the robust re-expression of other known targets including Il17a, and Ifng. We identify an underlying mechanism involving OCA-B recruitment of the histone lysine demethylase Jmjd1a8 to targets such as Il2 and Ifng. The findings pinpoint Oct1 and OCA-B as unanticipated mediators of CD4 T cell memory.

overall_design: Examination of 4 different conditions in 2 genotypes

first: Arvind

last: Shakya

first: Alon

last: Goren

first: Alex

last: Shalek

first: Cody

last: German

first: Jeremy

last: Snook

first: Vijay

last: Kuchroo

first: Nir

last: Yosef

first: Raymond

last: Chan

last: Williams


title: Oct1 and OCA-B are Selectively Required for CD4 Memory T Cell Formation and Function


title: Oct1 and OCA-B are Selectively Required for CD4 Memory T Cell Formation and Function (ChIP-seq)

summary: Epigenetic changes are crucial for the generation of immunological memory. Failure to generate or maintain these changes will result in poor memory responses. Similarly, augmenting or stabilizing the correct epigenetic states offers a potential method of enhancing immune memory. Yet the transcription factors that regulate these processes are poorly defined, as are the target genes they control and they chromatin-modifying complexes they recruit. Using model pathogens and three different mouse models, we find that the widely expressed transcription factor Oct1 and its cofactor OCA-B are selectively required for the in vivo generation of functional CD4 memory. In vitro, both proteins are required to maintain a poised state at the Il2 target locus in resting but previously stimulated CD4 T cells, and to generate robust Il2 expression upon restimulation. Gene expression profiling indicates that OCA-B is also required for the robust re-expression of multiple other targets including Ifng and Il17a. ChIPseq identify multiple differentially expressed direct targets. We identify an underlying mechanism involving OCA-B recruitment of the histone lysine demethylase Jmjd1a to targets such as Il2 and Ifng. The findings pinpoint Oct1 and OCA-B as unanticipated mediators of CD4 T cell memory.

overall_design: Examination of transcription factor occupancy in CD4 T cells upon rest and restimulation.

first: Matthew


title: Genome-wide binding of GATA1 and TAL1 determined at high resolution

city: University Park

first: Bongsoo

last: Park

state: PA

address: 453 North Frear Building

institute: Penn State University

summary: Erythroid development and differentiation from multiprogenitor cells to red blood cells requires precise transcriptional regulation. Key erythroid transcription factors, GATA1 and TAL1, co-operate, along with other proteins, to regulate many aspects of this process. How GATA1 and TAL1 are positionally organized with respect to each other and their cognate DNA binding site across the mouse genome remains unclear. We applied high resolution ChIP-exo to GATA1 and TAL1 to study their positional organization across the mouse genome during GATA1-dependent maturation. Two complementary methods, MultiGPS and peak-pairing, were used to determine high confidence binding locations by ChIP-exo. We identified ~10,000 GATA1 and ~15,000 TAL1 locations, which were essentially confirmed by ChIP-seq. Of these, ~4,000 locations were bound by both GATA1 and TAL1. About three-quarters of these were tightly linked (<40 bp away) to a partial E-box located 7-8 bp upstream of a WGATAA motif. Both TAL1 and GATA1 generated distinct characteristic ChIP-exo peaks around WGATAA motifs, that reflect on their positional arrangement within a complex. We show that TAL1 and GATA1 form a precisely organized complex at a compound motif consisting of a TG 7-8 bp upstream of a WGATAA motif across thousands of genomic locations.

overall_design: Genome wide analysis of GATA1 and TAL1 in G1E and G1E-ER4 cells using ChIP-exo experiments

first: Garam

last: Han

first: Vinesh

last: Vinayachandran

first: Alain

last: Bataille

first: Ka

middle: Y

last: Chan-Salis

first: Cheryl

last: Keller

first: Maria

last: Long

first: Shaun

last: Mahony

first: Ross

last: Hardison

first: B

middle: F

last: Pugh


title: RNAseq analysis of impact of PNN on gene expression and alternative splicing in corneal epithelial cells

status: Public on Sep 16 2015

city: Gainesville

first: Stephen

last: Sugrue

address: 1600 SW Archer Road

institute: University of Florida

summary: The specialized corneal epithelium requires differentiated properties, specific for its role at the anterior surface of the eye, thus tight maintenance of the differentiated qualities of the corneal epithelial is essential.  Our studies have focused on pinin (PNN), an exon junction component (EJC) that has dramatic implications on corneal epithelial cell differentiation and may act as a stabilizer of the corneal epithelial cell phenotype. Our studies revealed that PNN is involved in both transcriptional repression complexes and the spliceosomal complexes, placing PNN at the fulcrum between chromatin and mRNA splicing.  Transcriptome analysis of PNN-knockdown cells revealed clear and reproducible alterations in transcript profiles and splicing patterns of a subset of genes that would significantly impact the epithelial cell phenotype.  Here, we further investigate PNN’s role in the regulation of gene expression and alternative splicing (AS) in a corneal epithelial context.  We used human corneal epithelial cells (HCET cells) that carry doxycycline-inducible PNN-knockdown shRNA vector and performed RNA-seq to determine differential gene expression and differential AS events. Multiple genes and AS events were identified as differentially expressed between PNN-knockdown and controls cells. Genes up-regulated by PNN-knockdown included a large proportion of genes that are associated with processes associated with enhanced cell migration and ECM remodeling including: MMPs, ADAMs, HAS2, LAMA3, CXCRs and UNC5C.  Genes down-regulated in response to PNN depletion included: IGFBP5. FGD3, FGFR2, PAX6, RARG and SOX10. AS events in PNN compared to controls was also more likely to be detected, and uregulated in PNN-knockdowns.  In particular, 60% of exon skipping events detected in only one condition were detected in PNN-knockdowns and of the shared exon skipping events, 92% of those differentially expressed were more frequent in the PNN-knockdown. This suggests that in the absence of PNN the epithelial cells are dramatically transformed in the amount and composition of isoforms and that PNN plays a crucial role in the selection of which isoforms differentiating cells produce.   Many of the genes affected by PNN-knockdown are known to affect epithelial phenotype. This window into the complexity of RNA splicing in the corneal epithelium implies that PNN exerts broad influence over the regulation and maintenance of epithelial cell phenotype.

overall_design: We used HCET cells that carry doxycycline-inducible PNN knockdown shRNA vector and performed RNA-seq to determine differential gene expression and differential alternative splicing events.

first: D

last: Akin

first: J

last: Newman

first: L

middle: M

last: McIntyre

first: S


title: Healthy donor PBMC RNA-seq with or without interferon-alpha stimulation

city: South San Francisco

first: Tiffany

last: hung

address: 1 DNA Way

department: Human Genetics

institute: Genentech

summary: Determine the gene expression profile in peripheral blood monocytes isolated from 3 healthy donors +/- 6 hours of interferon-alpha treatment.

overall_design: 3 healthy donor PBMCs +/- interferon-alpha.

last: Hung

first: Timothy

last: Behrens

first: Christina

last: Chaivoropol

first: Ward

last: Ortmann


title: SLE lupus RNA-seq

summary: RNA-seq of systemic lupus erythematosus (SLE) whole blood and healthy controls to determine the gene expression changes in these patients.

overall_design: RNA-seq of PAXgene blood from SLE and healthy donors.

first: Michael

last: Townsend

first: Tushar

last: Bhangale


title: Ro60 iCLIP

summary: Autoantibodies target the RNA-binding protein Ro60 in systemic lupus erythematosus (SLE) and Sjögren's syndrome. However, whether Ro60 and its associated RNAs contribute to disease pathogenesis is unclear. We catalogued the Ro60-associated RNAs in human cell lines.

overall_design: iCLIP in 2 cell lines (GM12878, K562).


title: Ro60-knockout cells

summary: RNA-seq of Ro60-null GM12878 cell lines in order to determine the gene expression changes resulting from loss of Ro60.

overall_design: 3 separate clones of Ro60(Trove2)-null cells derived from zinc finger nuclease targeting of exon 2, two wildtype biological replicates, +/- IFNa for 6 hours.


title: Gene expression profiling of control cohesin knockdown LSK cells

city: New York

first: Jasper

last: Mullenders

state: NY

address: 550 1st ave

department: Pathology

institute: NYU School of Medicine

summary: We performed RNA-sequencing in LSK cells (Lin(neg)/c-Kit(+)/Sca-1(+)) from shRNA mice carrying an shRNA for Renilla, Smc1a or Stag2.

overall_design: RNA-sequencing control (Renilla) and cohesin (Smc1a and Stag2) knockdown cells.

first: Iannis

last: Aifantis


title: Gene expression profiling from cohesin knockdown cells obtained grown in methylcellulose

summary: We performed RNA-sequencing in c-Kit+ cells that were infected with retroviruses expressing shRNAs for Renilla, Rad21, Smc1a, Smc3 or Stag2. These cells were grown in methylcellulose (M3434) for either one passage (P1) or replated for five passages (P5).

overall_design: RNA-sequencing control (Ren) and cohesin (Rad21, Smc1a, Smc3 and Stag2) knockdown cells.


title: Loss of cohesin in vivo alters adult stem cell homeostasis leading to myeloproliferative neoplasms


title: The Ro60 Autoantigen Binds Endogenous Retroelements and Regulates Inflammatory Gene Expression


title: Genome-wide profiling of nucleosome sensitivity and chromatin accessibility to MNase in D. melanogaster [RNA-seq]

first: Razvan

last: Chereji

overall_design: RNA-seq S2 cells Drosophila melanogaster

first: Yuri

last: Moshkin

first: Victor

last: Guryev

first: Alexandre

last: Morozov


title: ATAC-sequencing in control and Stag2 knockdown cells

summary: We performed ATAC-sequencing in LSK cells (Lin(neg)/c-Kit(+)/Sca-1(+)) from shRNA mice carrying an shRNA for either Renilla or Stag2.

overall_design: ATAC-sequencing control (Renilla) and Stag2 knockdown cells.


title: Tolerance mechanisms towards high-temperature and -acid stress conditions during itaconic acid biosynthesis in Aspergillus terreus

city: Jinan

first: Mei

last: Chen

state: Shandong

address: No.3501,Daxue Road

department: Qilu University of Technology

country: China

institute: Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences

summary: A. terreus LYT10 is an industrial strain for itaconic acid production, in which the biosynthesis of itaconic acid and glucose conversion rate were affected by temperature and initial concentration of itaconic acid  in industrial production. RNA-seq was used to identify the key regulators related to tolerance mechanism toward various stress conditions.

overall_design: A total of 4 samples were analzyed. The mycelia cultivated in itaconic acid production medium (IPM) on a rotary shaker at 220 rpm and 37°C for 36 h  was considered as a reference. IPMs with 5 g/L and 40 g/L itaconic acid (pH 3.25)  were used for high-acid culture respectively. High-temperature condition was possesed at 42°C for 36 h in IPM.

first: Xuenian

last: Huang

first: Jianjun

last: Li

first: Xuefeng

last: LU


title: Regulation of gene expression during the onset of ligninolytic oxidation by Phanerochaete chrysosporium on colonized wood

city: Madison

first: Dan

last: Cullen

state: WI

address: One Gifford Pinchot Dr

institute: UW/FPL

summary: The biodegradation of lignocellulose requires the disruption of its lignin, which shields the metabolically assimilable polysaccharides in this recalcitrant natural composite. Although a variety of microorganisms can attack lignocellulose, white rot basidiomycetes are uniquely efficient at this process, cleaving the recalcitrant intermonomer linkages of lignin via extracellular oxidative mechanisms and mineralizing many of the resulting fragments to carbon dioxide via intracellular processes. Considerable progress has been made in understanding this process in the model white rot fungus Phanerochaete chrysosporium, which expresses important components of its ligninolytic system in response to nutrient limitation, as part of its secondary metabolism. Biochemical and genetic evidence point to an important role in P. chrysosporium for secreted lignin peroxidases (LiPs), manganese peroxidases (MnPs), and H2O2-producing oxidases, which are thought to work together to cleave lignin into low molecular weight fragments. However, many aspects of ligninolysis by P. chrysosporium remain poorly understood. Although a definitive picture of the entire ligninolytic system in P. chrysosporium is not yet attainable, transcriptome analyses of the fungus grown on wood can provide useful clues. With the advent of the initial genome assembly and annotations (v1.0 and v2.1), microarray-based transcriptome analysis allowed examination of transcript levels of P. chrysosporium genes when grown in ball-milled wood and in defined growth media. This approach provided useful insights but was limited to 10048 v2.1 targets and complicated by the unpredictable manner in which the fungus responds to unnatural carbon sources in submerged basal salts media. A complete, fully coordinated ligninolytic system is likely not expressed by P. chrysosporium on ball-milled wood, because a potential route for regulatory feedback has been eliminated: the cellulose and hemicellulose in this substrate is readily accessible to enzymes, and thus ligninolysis is not essential for growth. An alternative approach is to compare levels of gene expression just before and after the onset of secondary metabolism and extracellular substrate oxidation by P. chrysosporium as it utilizes solid wood as its carbon source. If this can be done, and decay of the substrate is also confirmed, then the genes undergoing marked changes in expression during the metabolic transition can be identified with greater confidence. Although not all such genes are expected to have roles in biodegradation, this strategy may identify interesting candidates for future investigation. Here we report RNAseq-based transcriptomes to characterize changes in gene expression that occur during the transition to ligninolytic metabolism.

overall_design: Phanerochaete chrysosporium was inoculated onto thin sections of wood. RNA was purified from colonized material after 40 and 96 hours. Single read 100 bp Illumina runs were performed.


title: The Mediator complex subunit MED23 couples H2B mono-ubiquitination to transcriptional control and cell fate determination

city: Shanghai

first: Gang

last: Wang

address: Yueyang Road 320

department: Institute of Biochemistry and Cell Biology, State Key Laboratory of Cell Biology,

institute: Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences

summary: The Mediator complex orchestrates multiple transcription factors with the Pol II apparatus for precise transcriptional control. However, its interplay with the surrounding chromatin remains poorly understood. Here, we analyze differential histone modifications between WT and MED23-/- (KO) cells and identify H2B mono-ubiquitination at lysine 120 (H2Bub) as a MED23-dependent histone modification. Using tandem affinity purification and mass spectrometry, we find that MED23 associates with the RNF20/40 complex, the enzyme for H2Bub, and show that this association is critical for the recruitment of RNF20/40 to chromatin. In a cell-free system, Mediator directly and substantially increases H2Bub on recombinant chromatin through its cooperation with RNF20/40 and the PAF complex. Integrative genome-wide analyses show that MED23 depletion specifically reduces H2Bub on a subset of MED23-contolled genes. Importantly, MED23-coupled H2Bub levels are oppositely regulated during myogenesis and lung carcinogenesis. In sum, these results establish a mechanistic link between the Mediator complex and a critical chromatin modification in coordinating transcription with cell growth and differentiation.

overall_design: To examine the enrichment of H2B ubiquitination, Pol II, H3K4me3, H3K79me3 in WT and KO MED23 MEF cells, we performed H2Bub ChIP-seq, Pol II ChIP-seq, H3K4me3 ChIP-seq and H3K79me3 ChIP-seq assays. 10 high-throughput sequencing data were deposited and WT, KO input data were controls for peak calling.

first: Xiao

last: Yao

first: Xing

last: Fu


title: genes regulated by mitoflash

city: guangzhou

first: zhongfu

last: ying

state: guangdong

address: kaiyuan

department: stem cell research

institute: chinese academy of sciences

summary: We found mitoflash has function in reprogramming. So we examined the genes expression after activation of mitoflash during somatic cell reprogramming. There are two ways to improve the mitoflash. First we overexpressed CypD, a mPTP component, to improve the mitoflash. Second we used the durgs, mastoparan and paraquat to increase the mitoflash. The MEF cells were reprogrammed, and treatment with mastoparan, paraquat or overexpressed with CypD. After activation of mitoflash at different times, the cells were collected for RNA-Seq.

overall_design: MEF cells were reprogrammed and expressed with CypD and Flag (as control) for 4 days. The cells were collected for RNA-Seq (Samples: Falg-D4, CypD-D4). MEF cells were reprogrammed and treated with mastoparan and paraquat for 8 days. The cells were collected for RNA-Seq (Samples: control-D8, mastoparan-D8 and paraquat-D8).

first: yao

last: deyang

first: chen

last: qinyu


title: small RNA Next Generation Sequencing of human primary macrophages treated with staurosporine

city: Valbonne - Sophia-Antipolis

first: Kevin

last: Lebrigand

address: 660 route des lucioles

country: France

institute: IPMC/CNRS

summary: Human primary macrophages were treated  with 1 µM staurosporine (STS) and profiled for small RNAs.

overall_design: Human primary macrophages were treated  with 1 µM staurosporine (STS) for 0, 4 and 6h.

first: M

last: Trabucchi

first: L

middle: E

last: Zaragosi

first: P

last: Barbry

first: K


title: Atherogenic Stimuli Trigger Ago2-hnRNP A1-Dependent Processing of RNYs Regulating Apoptosis and Inflammatory Response in Macrophages

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: PER2 synchronizes mitotic expansion and decidual transformation of human endometrial stromal cells

city: Coventry

first: Jan

last: Brosens

address: University Hospital Coventry and Warwickshire

department: Warwick Medical School

institute: University of Warwick

summary: Implantation is dependent on synchronized interactions between the conceptus and surrounding decidual cells but the involvement of clock genes in this process is not well understood. Circadian oscillations are predicated on transcriptional-translational feedback loops, which balance the activities of the transcriptional activators CLOCK and BMAL1 and repressors encoded by PER and CRY genes. Here we show that loss of PER2 expression silences circadian oscillations in decidualizing human endometrial stromal cells (HESCs). Downregulation was preceded by reduced CLOCK binding to a noncanonical E-box enhancer in the PER2 promoter and occurred between 12 - 24 h after exposure to a deciduogenic stimulus. RNA sequencing revealed that premature inhibition of PER2 by siRNA knockdown leads to a grossly disorganised decidual response. Gene ontology analysis highlighted a preponderance of cell cycle regulators amongst the 1,121 genes perturbed upon PER2 knockdown. Congruently, PER2 inhibition abrogated mitotic expansion of differentiating HESCs by inducing cell cycle block at G2/M. Analysis of mid-luteal endometrial biopsies revealed an inverse correlation between PER2 transcript levels and the number of miscarriages in women suffering reproductive failure. Thus, PER2 synchronizes mitotic expansion of HESCs with a periodic decidual gene expression; uncoupling of these events may cause persistent pregnancy failure.

overall_design: Endometrial mRNA profiles of paired control (siRNA-NT) and siRNA-PER2 were generated by deep sequencing, in triplicate using Illumina

first: Joanne

last: Muter

first: Emma

last: Lucas

first: Yi-Wah

last: Chan

first: Jonathan

last: Moore


title: small RNAnext Generation Sequencing of bone marrow-derived macrophages treated in M-CSF withdrawal medium

summary: The goal of this study is to investigate the presence of novel small RNAs in apoptotic macrophage

overall_design: Method: cultured mouse (bl6 line) bone marrow were differentiated in for 7 days in conditioned medium, in preence of Macrophage-Colony Stimulating Factor (M-CSF). M-CSF also stimulates the survive of macrophages by activating PI3K/AKT pathway. In accordance to previous publication (Lombardo, E., Alvarez-Barrientos, A., Maroto, B., Bosca, L., and Knaus, U.G, 2007, TLR4-mediated survival of macrophages is MyD88 dependent and requires TNF-alpha autocrine signalling, J Immunol 178, 3731-3739) apoptosis was induced in culturaed bone marrow-derived macrophages upon 36 hr of M-CSF withdrawal. Total RNA was isolated and a small RNA library was made using "Small RNA sequencing kit" (Illumina), according to the manufacturer’s instructions.

first: Michele

first: Bogdan

last: Tanasa


title: Lineage Reprogramming of mouse fibroblasts into induced cardiac progenitor cells by defined factors

first: Pratik

middle: Arvind

last: Lalit

state: Wi

address: 1111 highland Ave, WIMR2 8th floor

department: Medicine

institute: UW-Madison

summary: Mouse adult cardiac fibroblasts were infected with doxycycline inducible, 11 cardiac factors. Addition of doxycycline resulted in appearance of proliferative cells that had high nuclear-to-cytoplasmic ratio. We called these cells iCPCs. iCPC were stably reporgrammed and could be maintined in proliferative state without doxycycline. Here we compared the transcriptome of iCPC with their parental cells; adult cardiac fibroblasts. We found that iCPCs have upregulated genes involved in cardiovascular development and downregulated fibroblasts specific genes. iCPCs did not expressed endoderm, ectoderm or pluripotency associated genes indicating that they were cardiac mesoderm restricted progenitors.

overall_design: We isolated RNA from low passage iCPCs, high passage iCPCs and cardiac fibroblasts and performed RNASeq analysis

first: Timothy

last: Kamp


title: On-target and Off-target Activities of RAG Are Restricted by Topological Chromatin Domains

status: Public on Sep 15 2015

first: Jiazhi

last: Hu

address: 300 Longwood Ave

department: PCMM

institute: Children's Hospital Boston

summary: RAG initiates V(D)J recombination in developing lymphocytes by generating "on-target" DNA double-stranded breaks at bona fide recombination signal sequence (RSS) pairs.  We now employ RAG-generated DNA breaks in endogenous or ectopically-inserted RSS pairs as bait to identify huge numbers of RAG "off-target" sites. These off-target DNA breaks occur across convergent CTCF-binding element (CBE)-flanked loop domains containing the bait RSS pairs. Such off-target cleavage occurs at the simple CAC motif that defines the RSS cleavage-site and shows orientation-dependence best explained by a two-dimensional tracking mechanism.  Deletion of the CBE-based IGCR1 IgH regulatory element disrupts antibody IgH recombination domains and, correspondingly, alters distributions of RAG on- and off-targets across the IgH locus. RAG off-targets frequently involved in chromosomal translocations occur in convergent RSS pairs at enhancer regions within a loop. Our findings reveal how RAG is developmentally focused and re-focused in lymphocytes and implicate mechanisms by which chromatin domains harness biological processes within them.

overall_design: We performed high-throughput genome-wide translocation sequencing (HTGTS) in progenitor B cells or cell lines to study signatures of RAG on and off-targeting activity using RAG-generated breaks in endogenous or ectopically-inserted paired bona fide RSSs as bait. Sequenceing was dong by Illumina Miseq.

first: Frederick

last: Alt

first: Yu

last: Zhang


title: Epigenetic profiling of Juvenile Idiopathic Arthritits (JIA) patients

city: Utrecht

first: Janneke

last: Peeters

address: Heidelberglaan 100

country: Netherlands

institute: UMC Utrecht

summary: With H3K27ac chromatin immunoprecipitation we identified a disease-specific, inflammation-associated, (super-)enhancer signature in JIA patient synovial fluid-derived CD4+ memory/effector T cells. This signature consists of unique pathogenesis-associated epigenetic changes which extend beyond general T cell activation-induced alterations, indicating that disease-specific (super-)enhancers contribute to JIA pathogenesis. In addition, our analysis shows that there is a profound enrichment of JIA-related SNPs in (super-)enhancers in JIA patients compared to controls, illustrating the importance of these non-coding regions for disease pathogenesis.

overall_design: H3K27ac ChIP-sequencing of CD4+ memory/effector T cells derived from peripheral blood (PB) from healthy controls (HC) and  PB or synovial fluid (SF) from JIA patients.

first: J

last: . van Loosdregt

middle: G

first: S

middle: J

last: Vervoort


title: In vivo recruitment analysis and a mutant strain without any group 2 σ factors reveal roles of different σ factors in cyanobacteria

city: Turku

first: Taina

last: Tyystjarvi

address: Pharmacity/Itäinen Pitkäkatu 4 C, 6th floor

department: Biochemistry

country: Finland

institute: University of Turku

summary: In eubacteria, replacement of one σ factor in the RNA polymerase (RNAP) holoenzyme by another one changes the transcription pattern. Cyanobacteria are eubacteria characterized by oxygenic photosynthesis and they typically encode numerous group 2 σ factors that closely resemble the essential primary σ factor. A mutant strain of the model cyanobacterium Synechocystis sp. PCC 6803 without functional group 2 σ factors (named as ΔsigBCDE) could not acclimate to heat, high salt, or bright light stress but in standard conditions ΔsigBCDE grew only 9% slower than the control strain. One-fifth of the genes in ΔsigBCDE were differently expressed compared to the control strain in standard growth conditions and several physiological changes in photosynthesis, and pigment and lipid compositions were detected. To directly analyze the σ factor content of RNAP holoenzyme in vivo, a His-tag was added to the γ subunit of RNAP in Synechocystis and RNAPs were collected. The results revealed that all group 2 σ factors were recruited by RNAP in standard conditions, but recruitment of SigB and SigC increased in heat stress, SigD in bright light, SigE in darkness and SigB, SigC and SigE in high salt, explaining the poor acclimation of ΔsigBCDE to these stress conditions.

overall_design: Cells from cyanobacteria Synechocystis sp. PCC 6803 named as control strain (CS) and a mutant strain without any functional group 2 sigma factors, ΔsigBCDE, were harvested (A730=1, 40 mL) directly from standard growth conditions (continuous illumination at the PPFD of 40 µmol m-2s-1, 32°C, ambient CO2). From three to four independent experiments were performed at each conditions.

first: Liisa

last: Gunnelius

first: Kaisa

last: Hakkila

first: Hans

middle: C

last: Matthijs


title: Epigenetic profiling and RNA-sequencing of Juvenile Idiopathic Arthritits (JIA) patients


title: Genome-wide maps of nuclear lamina interactions in single human cells. (Hi-C, KBM7 haploid)

status: Public on Sep 24 2015

city: Worcester

first: Bryan

last: Lajoie

address: 368 Plantation St.

department: Program in Systems Biology

institute: UMMS

summary: Mammalian interphase chromosomes interact with the nuclear lamina (NL) through hundreds of large Lamina Associated Domains (LADs). We report a method to map NL contacts genome-wide in single human cells. Analysis of ~400 maps reveals a core architecture of gene-poor LADs that contact the NL with high cell-to-cell consistency, interspersed by LADs with more variable NL interactions. The variable contacts are more sensitive to a change in genome ploidy than the consistent contacts. Single-cell maps indicate that NL contacts involve multivalent interactions over hundreds of kilobases. Moreover, we observe extensive intra-chromosomal coordination of NL contacts, even over tens of megabases. Such coordinated loci exhibit preferential interactions as detected by Hi-C. Finally, single-cell gene expression and chromatin accessibility analysis shows that loci with consistent NL contacts are expressed at lower levels and are more consistently inaccessible than loci with lower contact frequencies. These results highlight fundamental principles of single cell chromatin organization.

overall_design: Hi-C Data

first: Ludo

last: Pagie


title: Genome-wide mapping of H3K27Me3 in mouse rib chondrocytes

city: Boston

first: Garyfallia

last: Papaioannou

address: Thier 1101, 50 Blossom Street

institute: MGH

summary: We performed ChiP sequencing analysis for H3K27Me3 in DNA from mouse rib chondrocytes.

overall_design: Chromatin immunoprecipitation for H3K27Me3 in mouse rib chondrocytes after overnight culture.Chromatin Immunoprecipitation was performed using Cell Signaling Technology SimpleChIP Plus Enzymatic Chromatin IP Kit (Magnentic Beads) #9005 Sequencing using Next Generation Sequencing (Otogenetics), mapping of reads using DNA Nexus mapper and region calling using Quest software (DNA Nexus)

first: Tatsuya

last: Kobayashi


title: A new phosphate-starvation response in fission yeast requires the endocytic function of myosin I

city: Cambridge

first: Felix

last: Krueger

address: Granta Park

department: Bioinformatics

institute: Altos Labs


title: RNA-sequencing of cells derived from the site of inflammation of Juvenile Idiopathic Arthritis patients

summary: Through RNA sequencing of CD4+ Tmemory/effector cells derived from the synovium of JIA patients and healthy controls, we analyzed the JIA gene expression signature. Treatment of autoinflammatory site-derived patient T cells with the BET-inhibitor JQ1 inhibited immune-related super-enhancers and preferentially reduced disease-associated gene expression, including cytokine-related processes.

overall_design: RNA-sequencing of CD4+ memory/effector T cells derived from Healthy Controls (HC) and JIA patients upon JQ1 treatment.

last: van Loosdregt


title: Genome-wide profiling of nucleosome sensitivity and chromatin accessibility in D. melanogaster

city: Bethesda

first: Razvan

last: Chereji

address: 6 Center Drive, Room 2A14

institute: National Institutes of Health


title: A novel phosphate-starvation response in fission yeast requires the endocytic function of Myosin I (ChIP-seq)

summary: S. pombe Myosin I (MyoI) plays a key role in cytoskeletal functions, in particular it is part of the endocytotic machinery. Here we show a novel role for MyoI in regulating the response to low phosphate stress. Deletion of MyoI led to an increased proliferation in low phosphate media when compared to a control strain; mutations in other proteins involved in the early steps of endocytosis, such as ARP2 and Sla2/END4, led to the same phenotype. We show that once cells are deprived of phosphate they undergo clear morphological changes, slow down growth but retain viability. This is reminiscent of a quiescence response, and it is completely abolished when MyoI is absent. Transcriptomic analysis revealed a wide perturbation of gene expression upon phosphate starvation, with rapid induction of stress-regulated genes. However, Æmyo1 cells displayed little or negligible effects at gene expression level when they were phosphate-starved. Using a combination of super resolution microscopy and genetic approaches we pinpointed the specific function of MyoI in endocytosis as critical for the response to low phosphate stress. Furthermore, introduction of point-mutated MyoI (with ablated endocytic function) into a ÆmyoI background failed to restore phosphate stress response. Taken together, our data indicate how endocytosis, and in particular MyoI, play a pivotal role in mediating cellular response to nutrients, bridging the external environment and internal molecular functions of the cells.

overall_design: ChIP-seq analysis of Myo1-GFP genomic binding in control or low phosphate condition

first: Edoardo

last: Petrini

first: Victoire

last: Baillet

first: Jake

last: Cridge

first: Cassandra

last: Hogan

first: Cindy

last: Guillaume

first: Huiling

last: Ke

first: Elisa

last: Brandetti

first: Hashem

last: Koohy

first: Mikhail

last: Spivakov

first: Patrick

last: Varga-Weisz


title: Distinct processes and transcriptional targets underlie CDX2 requirements in intestinal stem cells and differentiated villus cells

first: Ramesh

last: Shivdasani

address: 450 Brookline Ave. Dana 720D

department: Medical Oncology

institute: Dana Farber Cancer Institute

summary:
To define target genes of the intestine-restricted transcription factor (TF) CDX2 in intestinal stem cells, we performed chromatin immunoprecipitation coupled with high-throughput sequencing (ChIP-Seq).
We used RNA-sequencing to profile gene expression changes during cell differentiation from mouse intestinal stem cells to mature villus cells, as well as genes perturbed in intestinal stem cells upon loss of Cdx2. We find thousands of genes that change in expression during cell differentiation, including known stem cell and mature markers.  Upon loss of Cdx2, hundreds of genes are up and down-regulated in intestinal stem cells, some of which are also bound by CDX2 nearby and constitute candidate direct target genes.

overall_design: CDX2 ChIP-Seq analysis of isolated mouse intestinal stem cells.  RNA seq analysis of control mouse villus cells, control intestinal stem cells and Cdx2-deleted intestinal stem cells.

first: Adrianna

middle: K

last: San Roman

first: Alessio

last: Tovaglieri

first: David

middle: T

last: Breault

middle: A


title: Characterization of the platelet transcriptome by RNA sequencing in patients with acute myocardial infarction.

city: Framingham

first: Andrew

middle: Danner

last: Johnson

state: MASSACHUSETTS

address: The Framingham Heart Study

department: Cardiovascular Epidemiology and Human Genomics Branch

institute: National Heart, Lung and Blood Institute

summary: Recent technological advances have made transcriptome sequencing (RNA-seq) possible in cells with low RNA copy number including platelets. Resulting studies have used RNA-seq in platelets isolated from healthy individuals to characterize the platelet transcriptome. However, platelets, possibly through gene expression changes, contribute to the etiology of and response to cardiovascular disease and events. To address this, we performed the largest human platelet RNA-seq analysis to date in 34 platelet samples: 16 ST-segment elevation myocardial infarction (STEMI), 16 non-STEMI (NSTEMI), and 2 controls.

overall_design: RNA-seq of platelet samples from 34 individuals: 16 with ST-elevation myocardial infarction (STEMI), 16 with non-STEMI, and 2 non-myocardial infarction controls

first: John

middle: D

last: Eicher

first: Yoshiyuki

last: Wakabayashi

first: Olga

last: Vitseva

first: Nada

last: Esa

first: Yanqin

last: Yang

first: Jun

last: Zhu

first: Jane

last: Freedman

last: McManus


title: A novel phosphate-starvation response in fission yeast requires the endocytic function of Myosin I (RNA-seq)

first: Felix

last: Krueger

overall_design: RNA-seq transcriptomic analysis in control and ∆myo1 S.pombe strains comparing gene expression profiles at 4 and 10 hours after phosphate depletion.

first: Edoardo

last: Petrini

first: Victoire

last: Baillet

first: Jake

last: Cridge

first: Cassandra

last: Hogan

first: Cindy

last: Guillaume

first: Huiling

last: Ke

last: Brandetti


city: Tokyo

first: Daizo

last: Koinuma

address: Hongo 7-3-1, Bunkyo-ku

country: Japan

institute: University of Tokyo

summary: Transforming growth factor (TGF)-beta induces apoptosis of many types of cancer cells and acts as a tumor suppressor. We found lower expression of TGF-beta type II receptor (TbRII) in most of SCLC cells and tissues than in normal lung epithelial cells and normal lung tissues, respectively. In vitro cell growth and in vivo tumor formation were suppressed by TGF-beta-mediated apoptosis when the wild-type TbRII was overexpressed in SCLC cells. We therefore determined Smad2 and Smad3 (Smad2/3) binding sites in a SCLC cell line H345 stably expressing exogenous TbRII (H345-TbRII) to identify target genes of TGF-beta.

overall_design: Smad2 and Smad3 binding sites in H345-TbRII cells were determined by ChIP-seq (one sample analysis, without replicates).

first: Fumihiko

last: Murai

first: Shogo

last: Ehata

first: Kohei

last: Miyazono


title: DNA Methylation Dynamics of Germinal Center B Cells Are Mediated by AID

first: Matt

last: Teater

address: 445 E 69th St

institute: Weill Cornell Medical College

summary: Changes in DNA methylation are required for the formation of germinal centers (GC), but the mechanisms of such changes are poorly understood.  Activation-induced cytidine deaminase  (AID) has been recently implicated recently in DNA demethylation through its deaminase activity coupled with DNA repair.  We investigated the epigenetic function of AID in vivo in germinal center B cells (GCB) isolated from wild type (WT) and AID-deficient (Aicda-/-) mice.  We determined that the transit of B cells through the GC is associated with marked locus-specific loss of methylation and increased methylation diversity, both of which are lost in Aicda-/- animals.  Differentially methylated cytosines (DMCs) between GCB and naïve B cells (NB) are enriched in genes that are targeted for somatic hypermutation (SHM) by AID and these genes form networks required for B cell development and proliferation. Finally, we observed significant conservation of AID-dependent epigenetic reprogramming between mouse and human B cells.

overall_design: ERRBS and RNA-seq of wild type and Aicda knockout murine naive and germinal center B cells.  ERRBS of human naive and germinal center B cells

first: Olivier

last: Elemento

first: Rita

last: Shaknovich


title: ChIP of BET proteins in iBET resistance [ChIP-seq]

city: Melbourne

first: Mark

last: Dawson

state: VIC

address: 305 Grattan Street

country: Australia

institute: Peter MacCallum Cancer Centre

summary: Bromodomain and Extra Terminal protein (BET) inhibitors are first-in-class targeted therapies that deliver a new therapeutic paradigm by directly targeting epigenetic readers1,2. Early clinical trials have shown significant promise especially in acute myeloid leukaemia (AML)3; therefore the evaluation of resistance mechanisms, an inevitable consequence of cancer therapies, is of utmost importance to optimise the clinical efficacy of these drugs. Using primary murine stem and progenitor cells immortalised with MLL-AF9, we have used an innovative approach to generate 20 cell lines derived from single cell clones demonstrating stable resistance, in vitro and in vivo, to the prototypical BET inhibitor, I-BET. Resistance to I-BET confers cross-resistance to chemically distinct BET inhibitors such as JQ1, as well as resistance to genetic knockdown of BET proteins. Resistance is not mediated through increased drug efflux or metabolism but is demonstrated to emerge from leukaemia stem cells (LSC). Resistant clones display a leukaemic granulocyte-macrophage progenitor (L-GMP) phenotype (Lin-, Sca-, cKit+, CD34+, Fc³RII/RIII+) and functionally exhibit increased clonogenic capacity in vitro and markedly shorter leukaemia latency in vivo. Chromatin bound BRD4 is globally reduced in resistant cells, however expression of key target genes such as MYC remains unaltered, highlighting the existence of alternative mechanisms to regulate transcription. We demonstrate that resistance to BET inhibitors is in part a consequence of increased Wnt/²-catenin signaling. Negative regulation of this pathway results in differentiation of resistant cells into mature leukaemic blasts, inhibition of MYC expression and restoration of sensitivity to I-BET in vitro and in vivo. Finally, we show that the sensitivity of primary human AML cells to I-BET correlates with the baseline expression of Wnt/²-catenin target genes. Together these findings provide novel insights into the biology of AML, highlight the potential therapeutic limitations of BET inhibitors and identify strategies that may overcome resistance and enhance the clinical utility of these unique targeted therapies.

overall_design: Comparison of iBET resistant and sensitive cell lines

first: Fong

last: Chun


title: Genome wide expression analysis of BET inhibitor resistance [RNA-seq]


title: iBET resistance

overall_design: Refer to individual Series


title: Characterization of the complete genome sequence of Barley yellow striate mosaic virus reveals an overprinted ancillary gene encoding a small hydrophobic protein

city: Beijing

first: Xian-Bing

last: Wang

state: Beijing

address: Yuanmingyuan West Road

institute: China Agricultural University

summary: In the recent years, RNA silencing has been studied extensively to be a conserved regulatory process in plants. In the antiviral silencing, the intermediate double-stranded RNA form during the replication of RNA viruses were recognized and processed into abundant of overlapping viral siRNA (viRNAs). Accordingly, the cloned viRNAs could be conversely assembled into some contigs of viruses, which is recently exploited for identifying new viruses and their genome sequences.To obtain rapidly the complete genome sequence of BYSMV, we carried out deep sequencing of small RNAs from healthy and BYSMV infected wheat, respectively. Thirteen contigs were assembled from the overlapping viRNAs only present in the infected wheat but not in the healthy wheat. The results of BLAST showed that ten contigs shared about 96% identity with the reported L gene of BYSMV isolate Zanjan-1.

overall_design: Viral assembly from the BYSMV infected wheat plants to obtain the full lengh genome and characterise the viral siRNAs

first: Jingrong

last: Zhu


title: DNMT1-associated long non-coding RNA regulate global gene expression and DNA methylation in colon cancer

city: Cleveland

first: Ahmad

last: Khalil

state: OH

address: 2109 Adelbert Rd

department: Genetics and Genome Sciences

institute: Case Western Reserve University

summary: The investigation of the association of long non-coding RNAs (lncRNAs) with DNMT1 by RIP-seq reveals that DNMT1 interacts with DACOR1. We identified the genomic occupancy sites of DACOR1 through ChIRP-seq, which we found to significantly overlap with known differentially methylated regions (DMRs) in colon tumors.  Induction of DACOR1 in colon cancer cell lines significantly reduced their ability to form colonies in vitro, suggesting a growth suppressor function. Consistent with the observed phenotype, induction of DACOR1 led to the activation of tumor-suppressor pathways and attenuation of cancer-associated metabolic pathways. Notably, DACOR1 induction resulted in down-regulation of Cystathionine β-synthase (CBS), which is known to lead to increased levels of S-adenosyl methionine (SAM) – the key methyl donor for DNA methylation.

overall_design: The DNA methyltransferase DNMT1 was immunoprecipitated and all associated RNAs were identified by RNA-seq and subsequent bioinformatic analyses. Samples were prepared for DNA sequencing through the ChIRP-seq protocol which uses antisense DNA oligos to bind to DACOR1 and pull down associated DNA in complex. We sequenced one sample with DACOR1 specific probes and non-specific DNA probes as control. Three samples were collected for each control lentivirus transduced V852 colon cancer line and DACOR1 lentivirus transduced V852. Samples were processed for RNA-sequencing and analyzed through differential expression analysis.


title: RNA-sequencing of mRNAs from control and CAP-D3 deficient Salmonella infected HT-29 cells

first: Michelle

middle: Suzanne

last: Longworth

address: 9500 Euclid Ave NE20

department: Molecular Genetics

institute: Cleveland Clinic Foundation

summary:
BACKGROUND & AIMS- More frequent interaction of bacteria with the colonic epithelium is associated with ulcerative colitis (UC).  The identities of all proteins which promote bacterial clearance in colonic epithelial cells are unknown.  Previously, we discovered that dCAP-D3 (Chromosome Associated Protein-D3), regulates responses to bacterial infection.  We examined whether CAP-D3 promotes bacterial clearance in human colonic epithelium.
METHODS- Clearance of Salmonella or adherent-invasive Escherichia coli LF82 was assessed by gentamycin protection assays in HT-29 and Caco-2 cells expressing CAP-D3 shRNA.  CAP-D3 levels in colonic epithelial cells from healthy and UC patient tissues were analyzed by immunoblot.  RNA-sequencing identified bacterially-induced CAP-D3 target genes.  The role of CAP-D3 target genes in bacterial clearance was analyzed by gentamycin protection assays, immunofluorescent staining, and by using pharmacologic inhibitors.
RESULTS- CAP-D3 expression was reduced in colonic epithelial cells from UC patients with active disease. Reduction of CAP-D3 expression inhibited autophagy and decreased intracellular bacterial clearance.  The components of the heterodimeric SLC7A5/SLC3A2 amino acid transporter were identified as CAP-D3 target genes; their levels increased in infected, CAP-D3 deficient cell lines and in cells from UC patients.  In HT-29 cells, this resulted in earlier SLC7A5 recruitment to Salmonella-containing vacuoles, increased mTOR activity, and enhanced bacterial survival.  Inhibition of SLC7A5/SLC3A2 or mTOR activity rescued the bacterial clearance defect in CAP-D3 deficient cells.
CONCLUSIONS- CAP-D3 attenuates amino acid transporter transcription to promote bacterial autophagy in colon epithelial cells.  CAP-D3 protein levels are decreased in patients with active UC, suggesting that CAP-D3 is a potential therapeutic target to restore mucosal homeostasis in UC patients.

overall_design: Three RNA samples from 3 independent experiments including timepoints taken at 0, 0.5 and 7 hours post-infection were analyzed on a bioanalyzer for quality; one of the 0.5 hour post-infection samples was excluded at this time due to poor RNA purity.  Directional, cDNA libraries made from cellular mRNAs were generated from the other 8 samples and sequenced (paired-end sequencing of 100 bp reads) in the Genomics Core at the University of Chicago on an Illumina HiSeq2000.

middle: S


title: TMPyP4 promotes cancer cell migration at low doses, but induces cell death at high doses

status: Public on Sep 14 2015

city: Guangzhou

first: Yong

last: Zhao

state: Guangdong

address: No. 135, Xingang Xi Road

institute: Sun Yat-Sen University

summary: The RNA-seq results reveal TMPyP4 and TPyP4 could promote cancer cell migration.

overall_design: Examination of the gene expression of the following cells:A549-untreated and TMPyP4-treated or TPyP4-treated.

first: Xiao-Hui

last: Zheng

first: Hai-Ying

last: Liu

first: Xin

last: Nie

first: Yi-Min

last: Fang

first: Qi

first: Li-Xin

last: Xia


title: Transcriptome profiling of human neural progenitor cells and neurons with DISC1 interruption

city: Santa Barbara

first: Kenneth

middle: S.

last: Kosik

address: UC Santa Barbara

department: NRI

institute: UCSB

summary:
Purpose: Genetic and clinical association studies have identified disrupted-in-schizophrenia 1 (DISC1) as a candidate risk gene for major mental illness. DISC1 is interrupted by a balanced chr(1;11) translocation in a Scottish family, in which the translocation predisposes to psychiatric disorders. We investigate the consequences of DISC1 interruption in human neural cells using TALENs or CRISPR-Cas9 to target the DISC1 locus. We sought to compare the gene expression profiles of human neural progenitor cells (NPCs) and neurons with interruption of the DISC1 gene in exon 2 (affecting all known coding transcripts) or exon 8 (near the site of the Scottish translocation, affecting longer transcripts).
Methods: Wild-type and DISC1-targeted iPSCs (wild-type = "WT", exon 8 single allelic frameshift mutant = "ex8_wm", exon 8 biallelic frameshift mutant = "ex8_mm", exon 2 biallelic frameshift mutant = "ex2mm") were differentiated to NPCs and neurons using an embryoid aggregate method. NPC or neuronal cultures were used for RNA harvest and subsequent paired-end stranded sequencing of >50M reads/sample and 3-6 biological replicates per group.
Results: We find that a subset of genes related to neuronal differentiation and development are dysregulated with DISC1 disruption at the NPC timepoint, whereas expression of genes related to neuronal function and signaling are altered at the neuronal timepoint. This study implicates DISC1 as a regulator of neuronal development.

overall_design: mRNA profiles of wild-type and DISC1-targeted human iPSC-derived neural progenitor cells (day 17) and neurons (day 50) by paired-end sequencing, with 3-6 biological replicates, using Illumina HiSeq

first: Priya

last: Srikanth

first: Tracy

middle: L

last: Young-Pearse

first: Matthew

last: Lalli

first: Hongjun

last: Zhou


title: EGFR Mutation Promotes Glioblastoma Through Epigenome and Transcription Factor Network Remodeling

city: Shanghai

first: Feng

address: 197 Ruijin Er Lu

department: National Research Center for Translational Medicine (Shanghai)

institute: Ruijin Hospital, Shanghai Jiaotong University School of Medicine

summary: Epidermal Growth Factor Receptor (EGFR) gene amplification and mutations are the most common oncogenic events in Glioblastoma (GBM), but the mechanisms by which they promote aggressive tumor growth are not well understood. Here, through integrated epigenome and transcriptome analyses of cell lines, genotyped clinical samples and TCGA data, we show that EGFR mutations remodel the activated enhancer landscape of GBM, promoting tumorigenesis through a SOX9 and FOXG1-dependent transcriptional regulatory network in vitro and in vivo. The most common EGFR mutation, EGFRvIII, sensitizes GBM cells to the BET-bromodomain inhibitor JQ1 in a SOX9, FOXG1-dependent manner. These results identify the role of transcriptional/epigenetic remodeling in EGFR-dependent pathogenesis and suggest a mechanistic basis for epigenetic therapy.

overall_design: ChIP-Seq for H3K27ac, H3K4me1, and H3K4me3, and RNA-seq for Glioblastoma (GBM) cells and/or tissues with or without EGFRvIII mutation.

first: Gary

last: Hon

first: Genaro

middle: R

last: Villa

first: Kristen

last: Turner

first: Shiro

last: Ikegami

first: Huijun

last: Yang

first: Zhen

last: Ye

first: Bin

last: Li

first: Samantha

last: Kuan

first: Ah

middle: Y

last: Lee

first: Ciro

last: Zanca

first: Bowen

last: Wei

first: Greg

last: Lucey

first: David

last: Jenkins

first: Wei

last: Zhang

first: Cathy

last: Barr

first: Frank

middle: B

last: Furnari

first: Timothy

last: Cloughesy

first: William

middle: H

last: Yong

first: Timonthy

last: Gahman

first: Andrew

last: Shiau

first: Webster

last: Cavenee

first: Bing

last: Ren

first: Paul

last: Mischel


title: Foxd3 promotes the exit from naïve pluripotency and prevents germline specification through enhancer decommissioning

city: Koeln

first: Milos

last: Nikolic

state: Nordrhein-Westfalen

address: Robert Koch Str. 21

country: Germany

institute: Center for Molecular Medicine Cologne

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


title: Pol-IV dependent transcripts in Arabidopsis

status: Public on Sep 13 2015

city: Los Angeles

first: Steve

last: Jacobsen

state: CA

address: 4045 Terasaki Life Sciences Building

department: Department of Molecular, Cell and Developmental Biology

institute: University of California, Los Angeles

summary: small RNA, PATH and mRNA libraries were made to characterize Col and various small RNA biogenesis mutants.

overall_design: Genome-wide RNA profiling was done by Illumina TruSeq sample preparation kits followed by high-throughput sequencing with Illumina HiSeq 2500 platform.

first: Jixian

last: Zhai


title: RNA-seq analysis reveals the role of red light in resistance against Pseudomonas syringae pv. tomato DC3000 in tomato plants

city: Hangzhou

first: Youxin

last: Yang

address: Zijingang Campus, 866 Yuhangtang Road

department: Department of Horticulture

institute: Zhejiang University

summary:
Background: Plants attenuate their responses to a variety of bacterial and fungal pathogens, leading to higher incidences of pathogen infection at night. However, little is known about the molecular mechanism responsible for the light-induced defence response; transcriptome data would likely facilitate the elucidation of this mechanism.
Results: In this study, we observed diurnal changes in tomato resistance to Pseudomonas syringae pv. tomato DC3000 (Pto DC3000), with the greatest susceptibility before midnight. Nightly light treatment, particularly red light treatment, significantly enhanced the resistance; this effect was correlated with increased salicylic acid (SA) accumulation and defence-related gene transcription. RNA-seq analysis revealed that red light induced a set of circadian rhythm-related genes involved in the phytochrome and SA-regulated resistance response. The biosynthesis and signalling pathways of multiple plant hormones (auxin, SA, jasmonate, and ethylene) were co-ordinately regulated following Pto DC3000 infection and red light, and the SA pathway was most significantly affected by red light and Pto DC3000 infection. This result indicates that SA-mediated signalling pathways are involved in red light-induced resistance to pathogens. Importantly, silencing of nonexpressor of pathogensis-related genes 1 (NPR1) partially compromised red light-induced resistance against Pto DC3000. Furthermore, sets of genes involved in redox homeostasis (respiratory burst oxidase homologue, RBOH; glutathione S-transferases, GSTs; glycosyltransferase, GTs), calcium (calmodulin, CAM; calmodulin-binding protein, CBP), and defence (polyphenol oxidase, PPO; nudix hydrolase1, NUDX1) as well as transcription factors (WRKY18, WRKY53, WRKY60, WRKY70) and cellulose synthase were differentially induced at the transcriptional level by red light in response to pathogen challenge.
Conclusions: Taken together, our results suggest that there is a diurnal change in susceptibility to Pto DC3000 with greatest susceptibility in the evening. The red light induced-resistance to Pto DC3000 at night is associated with enhancement of the SA pathway, cellulose synthase, and reduced redox homeostasis.

overall_design: Four treatments including control, three biological replicates each treatment

first: You-Xin

first: Meng-Meng

last: Wang

first: Yan-Ling

last: Yin

first: Eugen

last: Onac

first: Guofu

last: Zhou

first: Sheng

last: Peng

first: Xiao-Jian

last: Xia

first: Kai

last: Shi

first: Jing-Quan

last: Yu

first: Yan-Hong


title: Expression profiling of centroacinar cells from adult zebrafish pancreas

status: Public on Sep 12 2015

city: Baltimore

first: Rebecca

last: Beer

address: 733 N Broadway Rm 460

department: Institute of Genetic Medicine

institute: Johns Hopkins University

summary: We sequenced mRNA from two preparations of isolated Notch-responsive ductal pancreas cells and compared transcript expression to all other non-Notch-responsive cells from each sample to charactarize zebrafish centroacinar cells.

overall_design: Determination of gene expression levels in centroacinar cells and non-centroacinar cells from adult pancreas.

first: Michael

last: Parsons


title: Foxd3 promotes the exit from naïve pluripotency and prevents germline specification through enhancer decommissioning [RNA-Seq]

summary: Following implantation, mouse epiblast cells transit from a naïve to a primed state in which they are competent for both somatic and primordial germ cell (PGC) specification. Using mouse embryonic stem cells (mESC) as an in vitro model to study the transcriptional regulatory principles orchestrating peri-implantation development, here we show that the transcription factor Foxd3 is necessary for the exit from naïve pluripotency and the progression to a primed pluripotent state. During this transition, Foxd3 acts as a repressor that dismantles a significant fraction of the naïve pluripotency expression program through the decommissioning of active enhancers associated with key naïve pluripotency and early germline genes. Subsequently, Foxd3 needs to be silenced in primed pluripotent cells to allow the reactivation of relevant genes required for proper PGC specification. Our findings uncover a wave of activation-deactivation of Foxd3 as a crucial step for the exit from naïve pluripotency and subsequent PGC specification.

overall_design: mRNA profiles were generated by RNA-seq in duplicates for each of the following mESC lines: Foxd3fl/fl;Cre-ER  mESC maintained in "Serum+LIF" (SL) treated with TM for three days (SL Foxd3-/-); untreated Foxd3fl/fl;Cre-ER SL mESC (SL Foxd3fl/fl); tetON Foxd3 SL mESC treated with Dox for three days; WT SL mESC treated with Dox for three days;  Foxd3fl/fl;Cre-ER mESC maintained in "2i+LIF" (2i) treated with TM for three days (2i Foxd3-/-); untreated Foxd3fl/fl;Cre-ER 2i mESC (2i Foxd3fl/fl).

first: Patricia

last: Respuela

first: Alvaro

last: Rada-Iglesias


title: Systematic Identification of Factors Crucial for Provirus Silencing in Embryonic Stem Cells (RNA-seq)

city: Singapore

first: Chadi

last: El Farran

address: 61 Biopolis Drive

country: Singapore

institute: Institute of Molecular and Cell Biology

summary: Our study reports the first genome-wide atlas of functional nodes that mediate proviral silencing in ESCs. It provides evidences for the comprehensive, interconnected and multi-layered genetic/epigenetic machineries by which ESCs maintain the repressive state of provirus and ERVs.

overall_design: RNA-seq analysis to determine the regulated endogenous retroviruses by Histone chaperones (Chaf1a/b), sumoylation factors (Sumo2/ Ube2i/Sae1/Uba2/Senp6) and chromatin modifiers (Trim28/Eset/Atf7ip)

first: Bin

middle: X

last: EL Farran

first: Hong

last: Guo

first: Tao

first: Hai

last: Fang

first: Yuin-Han

last: Loh


title: EGF-Mediated Transcriptional Coregulation of EpICD-LEF1 Alters Nanomechanical Properties of Endometrial Cancer Cells for Promoting Cell Migration

city: San Antonio

first: Tim

middle: Hui-Ming

last: Huang

state: TX

address: 8403 Floyd Curl Drive

department: Molecular Medicin

institute: The University of Texas Health Science Center at San Antonio

summary: Epithelial cell adhesion molecule (EpCAM), a membrane protein known to modulate cell-cell adhesion, is also a signaling molecule internalized into the nucleus for transcriptional regulation. Here we demonstrate that activated EGF/EGFR is a signaling factor to drive the proteolysis of EpCAM. Cleavage of the extracellular fragment EpEX results in topographic fading of cell-surface EpCAM detected by antibody-conjugated cantilevers of atomic force microscope (AFM). As a result, internalization of the cytoplasmic domain EpICD forms a transcription factor complex with LEF1 that regulates gene transcription for enhanced cell-mobility functions. Comprehensive probing of cell surface using AFM tip (without antibody) reveals increased elasticity and non-stickiness of these cells, promoting epithelial to mesenchymal transition. While EpCAM cleavage may contribute to the loss of cell-surface adhesiveness, its internalized EpICD additionally regulates targets for promoting cell migration. Thus, this EGF/EGFR-modulated action on structural EpCAM and regulatory EpICD can enhance invasion potential of transformed cells.

overall_design: RL95-2 were stimulated with EGF for 0,12,24,and 48 hr.Immunoprecipitation was carried out using antibodies against EpCAM and Lef-1, sequenced by Illumina HiSeq 2000

first: Ya-Ting

last: Hsu

first: Hui-Ming

first: Yao

last: Wang


title: Systematic Identification of Factors Crucial for Provirus Silencing in Embryonic Stem Cells (ChIP-seq)

summary: Our study reports the first genome-wide atlas of functional nodes that mediate proviral silencing in ESCs.  It provides evidences for the comprehensive, interconnected and multi-layered genetic/epigenetic machineries by which ESCs maintain the repressive state of provirus and ERVs.

overall_design: ChIP-seq analysis of Chaf1a, Trim28, Sumo2 and Zfp809 to demonstrate the mechanism of the silencing of Endogenous retroviruses

last: Yang


title: Linking Transcriptomic Organization to the Regulatory Landscape of Cyanobacteria through Context-Based Network Analysis

status: Public on Sep 11 2015

city: Boston

first: Ryan

last: McClure

address: 650 Albany St.

institute: Boston University

summary: Construction of a global gene co-expression network to determine gene relatedness and expand regulatory pathways in the model cyanobacterium Synechococcus 7002.

overall_design: These 5 conditions represent part of a data set of RNA-seq conditions that were examined with the Context-Liklihood of Relatedness (CLR) program to generate a gene co-expression network.  Network structure was used group genes by transcriptomic context and thus function and link regulators with putative targets

first: Christopher

last: Overall

first: Jason

middle: E

last: McDermott

first: Eric

last: Hill

first: Lye

middle: M

last: Markillie

first: Lee

last: McCue

first: Ronald

last: Taylor

first: Marcus

last: Ludwig

first: Donald

last: Bryant

first: Alex

last: Beliaev


title: Comprehensive analyses of microRNA gene evolution in paleopolyploid soybean genome

first: Zhixi

last: Tian

address: No.1 Building, No.1 West Beichen Road,Chaoyang District

institute: Institute of Genetics and Developmental Biology, Chinese Academy of Sciences

summary: miRNA genes are thought to undergo quick birth and death processes in genomes and the emergence of a MIRNA-like hairpin provides the base for functional miRNA gene formation. However, the factors affecting the formation of an active miRNA gene from a MIRNA-like hairpin within a genome remain unclear. We performed a genome-wide investigation of MIRNA-like hairpin accumulation, expression, structural changes and relationships with annotated genomic features in the paleopolyploid soybean genome. Our results showed that adjacent gene and transposable element content, rates of genetic recombination at location of emergence, along with its own gene structure divergence greatly affected miRNA gene evolution. Further investigation suggested that miRNA genes from different duplication sources followed distinct evolutionary trajectories and that the accumulation of MIRNA-like hairpins might be an important factor in causing long terminal repeat retrotransposons to lose activity during genome evolution.

overall_design:
In order to identify miRNAs at a genome-wide level in soybean, 23 small RNA libraries from root, shoot, leaf, flower and seed at different developmental stages, includng gernination, trefoil, flowering, seed development and plant senescene were constructed.
23 samples, no replicates.


title: Regional centromeres in Candida lusitaniae lack pericentromeric heterochromatin

city: Houston

first: Lisha

last: Zhu

address: 7000 fannin st

institute: UT Health Science Center at Houston


title: Regional centromeres in Candida lusitaniae lack pericentromeric heterochromatin (RNA-seq)

summary: We sequenced mRNA from wild type and Hst1 knock out strains of Candida lusitaniae to generate the gene expression profiles and studied the differentially expressed genes between the two conditions.

overall_design: RNA profiles of wild type (WT) and Hst1 knockout of Candida lusitaniae were generated by deep sequencing

first: Laura

middle: N

last: Rusche


title: Regional centromeres in Candida lusitaniae lack pericentromeric heterochromatin (ChIP-seq)

summary: The goal of this study is to identifiy centromere locations of Candida lusitaniae through ChIP-Seq analysis. We applied ChIP-Seq on two centromere proteins: Cse4 and Mif2. After peak calling, we identified one peak in each of the 8 supercontigs of Candida lusitaniae which may represent the potential centromere location.

overall_design: Identification of  centromere location of Candida lusitaniae using two centromere proteins: Cse4 and Mif2


title: Identification of conserved and novel microRNAs during tail regeneration in the Mexican Axolotl

city: Minneapolis

first: Micah

last: Gearhart

state: MN

address: 6-160 Jackson Hall, 321 Church St.

department: Genetics, Cell Biology and Development

institute: University of Minnesota

summary: The Mexican axolotl (Ambystoma mexicanum) is one member of a select group of vertebrate animals that has retained the amazing ability to regenerate multiple body parts. In addition to being an important model system for regeneration, the axolotl is also a leading model system for developmental biologists. Many genes used in development have been identified to be reused again during regeneration, however how this molecular circuitry is controlled during regeneration is unknown. In recent years microRNAs have been identified as key regulators of gene expression during development, in many diseases and also in regeneration. Here we have used deep sequencing combined with qRT-PCR to identify microRNAs that are involved in regulating regeneration in axolotl. This approach has enabled us to identify well known families of microRNAs and in addition to identify putative novel microRNAs that differentially regulated in the regenerating tissue. These findings suggest that microRNAs may play key roles in managing the spatial and temporal expression of genes important for ensuring that the correct tissues are regenerated.

overall_design: small RNA Sequencing (2 samples) in Ambystoma Mexicanum

first: Karen

last: Echeverri


title: Differential Toxicity of Nuclear RNA Foci Versus Dipeptide Repeat Proteins in a Drosophila Model of C9ORF72 FTD/ALS

city: Worcester

first: Fen-Biao

last: Gao

state: Massachusetts

address: 55 Lake Avenue North

department: Department of Neurology

institute: University of Massachusetts Medical School

summary: Nuclear RNA foci are often associated with mis-regulation of RNA processing in repeat expansion diseases. Here we report transcriptomic analysis of flies carrying expanded G4C2 repeats, the most common genetic cause of FTD/ALS. By obtaining an average of 32 million reads per library, we show that the presence of numerous nuclear G4C2 repeat RNA foci does not cause extensive changes in RNA processing in this model organism.

overall_design: Comparision of the transcriptomic profile of flies expressing 5R or 160R from two different chromosomes

first: Helene

last: Tran

first: Jill

last: Moore

first: Zhiping

last: Weng


title: Induction of Interleukin-9-producing Mucosal Mast cells Promotes Susceptibility to IgE-mediated Experimental Food Allergy

city: Cincinnati

first: Yui-Hsi

last: Wang

state: Oh

address: 3333 Burnet Ave

institute: CCHMC

summary: Experimental IgE-mediated food allergy depends on intestinal anaphylaxis driven by interleukin (IL)-9. However, the primary cellular source of IL-9 and the mechanisms underlying the susceptibility to food-induced intestinal anaphylaxis remain unclear. Herein, we have reported the identification of multifunctional IL-9-producing mucosal mast cells (MMC9s) that can secrete prodigious amounts of IL-9 and IL-13 in response to IL-33, and mast cell protease-1 (MCPt-1) in response to antigen and IgE complex crosslinking, respectively. Repeated intragastric antigen challenge induced MMC9 development that required T cells, IL-4, and STAT6 transcription factor, but not IL-9 signals. Mice ablated of MMC9 induction failed to develop intestinal mastocytosis, which resulted in decreased food allergy symptoms that could be restored by adoptively transferred MMC9s. Finally, atopic patients that developed food allergy displayed increased intestinal expression of Il9 and MC-specific transcripts. Thus, the induction of MMC9s is a pivotal step to acquire the susceptibility to IgE-mediated food allergy.

overall_design: dUTP mRNA-Seq profiles of indicated hematopoietic cell lineages were generated on Illumina HiSeq2500.   Hematopoietic cells were  isolated from Balb/C mice that developed food allergy and bone marrow-derived mast cells were generated from naïve Balb/C mice

first: Wang

last: Yui-Hsi


title: Histone Deacetylase 3 is required for efficient T cell development

city: Nashville

first: Scott

last: Hiebert

state: TN

address: 2220 Pierce Ave

institute: Vanderbilt University

summary: Hdac3 is an important target of HDAC inhibitors used in the treatment of cutaneous T cell lymphoma.  In order to gain an understanding of Hdac3 function in T cells,we deleted Hdac3 from early mouse thymocytes using LCK-Cre. Hdac3 deletion resulted in a loss of single positive thymocytes due to a defect in positive selection at the double  positive (DP) stage of thymocyte development. To better characterize this defect, we sorted the DP1 and DP2 populations to for gene expression profiling.

overall_design: Total RNA was extracted from DP1 (GFP+CD4+CD8+CD5loTCRblo) or DP2 (GFP+CD4+CD8+CD5hiTCRbint) thymocytes isolated by FACS from Hdac3+/+ or Hdac3F/F LCK-Cre+ animals. Libraries were constructed from rRNA-depleted total RNA pools to identify altered gene expression in DP populations following Hdac3 deletion.

first: Kristy

last: Stengel

first: Alyssa

last: Summers


title: Systematic Identification of Factors Crucial for Provirus Silencing in Embryonic Stem Cells

first: Chadi

last: El Farran

overall_design: Refer to individual Series


title: Foxd3 promotes the exit from naïve pluripotency and prevents germline specification through enhancer decommissioning [ChIP-Seq]

first: Milos

last: Nikolic

overall_design: Genome-wide binding profiles for Foxd3 were investigated in mouse embryonic stem cells (mESC). A mESC line (FH-Foxd3 mESC line) expressing exogenous Foxd3 tagged with Flag and HA epitope (FH-Foxd3) at nearly endogenous levels was generated. ChIPs were performed against FH-Foxd3 using anti-HA or anti-Flag antibodies.

first: Patricia

last: Respuela

first: Alvaro

last: Rada-Iglesias


title: Next Generation Sequencing Facilitates Quantitative Analysis of dendritic cell Transcriptomes during LPS response

city: shang hai

first: Qian

last: Zhang

address: xiangyin road 800

institute: Second military medical university

summary:
Purpose:The purpose of this study is to detect activated or silenced genes during LPS-induced dendritic cell. Gene expression differences between two samples could be found using transcriptome profiling (RNA-seq) analysis.
Methods:Mouse dendritic cells were generated from bone marrow cells in  RPMI-1640 medium with recombinant mouse GM-CSF and IL-4, immature DCs were obtained before or after LPS stimulation. Immature DCs were sorted respectively based on marker CD86 and Iab(MHCII) using flowcytrometer. DC mRNA profiles were generated by deep sequencing,using Illumina.
Results: We mapped about 10 million sequence reads per sample to the mouse genome, identified hundreds of genes with significant mRNA variation during LPS stimulation.

overall_design: DC mRNA profiles immature BMDCs were generated by deep sequencing

first: Xuetao

last: Cao

first: Zhang

last: Qian


title: Transcriptome of hepatocellular carcinoma using CAGE

status: Public on Sep 10 2015

city: Yokohama

first: Kosuke

last: Hashimoto

state: Kanagawa

address: 1-7-22 Suehiro-cho, Tsurumi-ku

department: IMS

institute: RIKEN

summary: An increasing number of non-coding RNAs (ncRNAs) are implicated in various human diseases including cancer; however ncRNA transcriptome of hepatocellular carcinoma (HCC) remains largely unexplored. We use CAGE (Cap Analysis of Gene Expression) to comprehensively map transcription start sites (TSSs) across different etiologies of human HCC as well as mouse HCC, with particular emphasis on ncRNAs distant from protein-coding genes. We find thousands of significantly up-regulated distal ncRNAs in HCC tumors compared to their matched non-tumors, which are as many as protein-coding genes. Moreover, we identify many LTR retroviral promoters activated in HCC tissues and expressed in a subfamily-specific manner, which account for approximately 20% of the up-regulated distal ncRNAs. The transcripts derived from LTRs, determined by 3' RACE, are multi-exon nuclear ncRNAs typically 0.5-2kb in length. This study sheds light on ncRNA transcriptome of human and mouse HCC.

overall_design: Expression profiles using CAGE for 37 mouse HCC. The human data are archived at dbGaP (phs000885.v1.p1). An umbrella BioProject has been created to associate the GEO and dbGaP BioProjects: PRJNA278792

first: Jamila

last: Faivre

first: Piero

last: Carninci


title: Expression of milk miRNA during the lactation cycle of the marsupial tammar wallaby (Macropus eugenii)

city: Waurn Ponds

first: christophe

last: lefevre

state: vic

address: pigdons road

department: School of medicine

institute: Deakin University

summary: High throughput sequencing of miRNAs collected from tammar milk at different time points of lactation showed high levels of miRNA secreted in milk and allowed the identification of differentially expressed milk miRNAs during the lactation cycle as putative markers of mammary gland activity and functional candidate signals to assist growth and timed development of the young. Comparative analysis of miRNA distribution in milk and blood serum suggests that milk miRNAs are primarily expressed from mammary gland rather than transferred from maternal circulating blood, likely through a new putative exosomal secretory pathway.

overall_design: 8 profiles were produced. Duplicates of day175

first: Vengamanaidu

last: Modepalli

first: Amit

last: Kumar

first: Lyn

last: Hinds

first: Julie

last: Sharp

first: Kevin

last: Nicholas


title: Human stem cell based models of neuronal migration provide insight into neurological disease pathogenesis and potential treatment

city: New York

first: Joseph

last: Gleeson

state: NY

address: 1230 York Avenue

institute: Rockefeller University

summary: Neuronal migration defects (NMDs) are among the most common and severe brain abnormalities in humans. Lack of disease models in mice or in human cells has hampered the identification of underlying mechanisms. From patients with severe NMDs we generated iPSCs then differentiated neural progenitor cells (NPCs). On artificial extracellular matrix, patient-derived neuronal cells showed defective migration and impaired neurite outgrowth. From a cohort of 107 families with NMDs, sequencing identified two homozygous C-terminal truncating mutations in CTNNA2, encoding αN-catenin, one of three paralogues of the α-catenin family, involved in epithelial integrity and cell polarity. Patient-derived or CRISPR-targeted CTNNA2- mutant neuronal cells showed defective migration and neurite stability. Recombinant αN-catenin was sufficient to bundle purified actin and to suppress the actin-branching activity of ARP2/3. Small molecule inhibitors of ARP2/3 rescued the CTNNA2 neurite defect. Thus, disease modeling in human cells could be used to understand NMD pathogenesis and develop treatments for associated disorders.

overall_design: 2 biological replicates per individual (2 iPSC clone differentiations), excluding 1263A, which has one sample

middle: G

first: Ashleigh

last: Schaffer