title: PRC2 loss amplifies Ras-driven transcription and confers sensitivity to BRD4-based therapies

first: Thomas

last: De Raedt

address: 77 Avenue Louis Pasteur

institute: Harvard Medical School -  Brigham Women's Hospital

overall_design: Refer to individual Series

first: Karen

last: Cichowski


title: Hematopoietic stem/progenitor cells HELP-tagging cytosine methylation data

status: Public on Jul 01 2014

city: BRONX

first: Fabien

last: Delahaye

state: NY

address: 1300 Morris Park Avenue

department: Genetics

country: USA

institute: AECOM

type: Methylation profiling by high throughput sequencing

summary: Using HELP-Tagging, we defined the cytosine methylation profiles of hematopoietic stem/progenitor cells (CD34+) from umbilical cord blood in 29 healthy newborns.  We found substantial variation of DNA methylation to occur in human CD34+ hematopoietic stem and progenitor cells from different individuals. Empirical annotation of the genome of this cell type reveals the variability to be targeted to candidate promoters and to candidate enhancer sequences of genes expressed at lower levels. The variability of DNA methylation at candidate enhancers was low for housekeeping genes but high for genes associated with leukocyte differentiation. The enrichment of DNA methylation variability at loci  with intermediate methylation values, occurring at apparently “poised” enhancers, suggests cell subpopulation heterogeneity between individuals as the basis for the epigenomic variability observed. The “meta-epigenome” present even in purified cells complicates the design and interpretation of epigenome-wide association studies, but also allows insights into the gene expression characteristics and the number of cell types comprising the cell population analyzed. One Sentence Summary: Variability in epigenomic patterns between different individuals occurs predominantly at promoters and enhancers, reflecting cell subpopulation heterogeneity.

overall_design: 29 HpaII test

first: N

middle: A

last: Wijetunga

first: Yong

last: Zhao

first: Aaron

last: Golden

first: Jessica

middle: C

last: Mar

first: Francine

middle: H

last: Einstein

first: John

middle: M

last: Greally


title: Transcriptome profiling reveals roles of meristem regulators and polarity genes during multicellular fruit trichome development in cucumber (Cucumis sativus L.).

city: Shanghai

first: Renyi

last: Liu

state: Shanghai

address: 3888 Chenhua Road

department: Bioinformatics

country: China

institute: Shanghai Center for Plant Stress Biology

type: Expression profiling by high throughput sequencing

summary: Trichomes are the hair-like structures that are widely present on the surface of aerial organs and function in plant defense against biotic and abiotic stresses. Previous studies focus on the single cell trichomes in Arabidopsis and cotton, or multicellular glandular trichomes in tomato, but the developmental process and molecular mechanisms controlling multicellular non-glandular trichome development are largely neglected. Here, we extensively characterized the fruit trichome (spine) development in wild type cucumber and in a tiny branched hair (tbh) mutant that contains a spontaneous mutation and has hairless foliage and smooth fruit surface. Our data indicated that cucumber trichome was multicellular and non-glandular, with no branches or endoreduplication. Further, the major feature of cucumber trichome development was spine base expansion. Transcriptome profiling through Digital Gene Expression indicated that meristem-related genes and transcription factors were implicated in the fruit spine development, and polarity regulators were upregulated during spine base expansion. qRT-PCR verified the reliability of our RNA-SEQ data, and in situ hybridization confirmed the enriched expression of meristem regulators CUP-SHAPED COTYLEDON3 (CUC3) and STM (SHOOT MERISTEMLESS) , as well as the abaxial identity gene KANADI (KAN) in cucumber fruit spine. Together, our results suggest a distinct regulatory pathway involving meristem genes and polarity regulators in multicellular trichome development in cucumber.

overall_design: Using Digital Gene Expression technology to compare the genome-wide gene expression profiles in the fruit spines of wild type cucumber and the tbh mutant, as well as the fruit spines on fruits of 0.5cm and 1.6cm long, repectively. Two biological repelicates were generated for each tissue.

first: Chunhua

last: Chen

first: Meiling

first: Jiang

last: Li

first: Xiaofeng

first: Huazhong

last: Ren

first: Xiaolan

last: Zhang


title: Differential RNA-sequencing of Campylobacter jejuni fur perR mutant

city: Norfolk

first: Arnoud

last: van Vliet

address: Colney Lane

department: Gut Health and Food Safety Programme

country: United Kingdom

institute: Institute of Food Research

summary: We report the use of differential RNA-sequencing for the determination of the primary transcriptome of the fur perR mutant of Campylobacter jejuni NCTC 11168. This allows for the genome-wide determination of transcription start sites.

overall_design: Campylobacter jejuni NCTC 11168 fur perR mutant was grown to late log phase, RNA was purified and used for differential RNA-sequencing by 454 sequencing with barcoded libraries, and used for determination of genome-wide transcription start sites

first: Rebecca

last: Handley

first: Francis

last: Mulholland

first: Mark

last: Reuter

first: Vinoy

middle: K

last: Ramachandran

first: Heather

last: Musk

first: Leah

last: Clissold

first: Nick

middle: E

last: Le Brun


title: Direct ChIP-bisulfite sequencing reveals a role of H3K27me3 mediating aberrant hypermethylation of promoter CpG islands in cancer cells

city: Shenzhen

first: Desheng

last: Gong

address: No.7 PengFei road

institute: Agricultural Genomes Institute at Shenzhen

type: Genome binding/occupancy profiling by high throughput sequencing

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


title: Transcriptional profiling of staurosporine-induced cell death in wild type versus Δczt-1 in Neurospora crassa

city: Berkeley

first: António Pedro

last: Gonçalves

address: 341A Koshland Hall

department: Plant and Microbial Biology

institute: University of California, Berkeley

summary:
Purpose: Compare the transcriptional profile of staurosporine-treated cells in Neurospora crassa wild type and Δczt-1 (ΔNCU09974) cells
Methods: Conidial suspensions were obtained and 1 x 106 cells/ml incubated in minimal medium for 6 hours (26ºC, 140 rpm, constant light) followed by the addition of staurosporine (or DMSO) and growth for 1 more hour. Cells were harvested using 0.45 μm filters and immediately frozen in liquid nitrogen. Total RNA was isolated by the Trizol-Phenol-Chloroform method. After digestion of 25 μg RNA with TURBO DNAse (Life Technologies), mRNA was purified using Dynabeads oligo(dT) magnetic beads (Life Technologies). The mRNA was chemically fragmented using the Ambion RNA fragmentation kit (Life Technologies). First and second strand cDNA synthesis was achieved using appropriate kits (Life Technologies). The illumina TruSeq kit was employed to generate the cDNA libraries with indexing adapters essentially following the manufacturer’s protocol. After purification of the libraries with AMPure XP beads (Roche), the quality of the libraries was analysed in a Agilent 2100 Bioanalyzer. The cDNA libraries were sequenced in a illumina HiSeq2000 and single reads of 50 bp were obtained. Sequencing data was handled essentially with Tophat, Cufflinks and Cuffdiff. Expression levels are presented as Fragments/Reads Per Kilobase of transcript per Million mapped reads (FPKM/RPKM).
Results: Transcriptional profiling of staurosporine-treated cells wild type by RNA-sequencing showed that genes encoding the machinery for protein synthesis are enriched among the genes repressed by the drug. Functional category enrichment analyses also show that genes encoding components of the mitochondrial respiratory chain are downregulated by staurosporine, whereas genes involved in endoplasmic reticulum activities are upregulated. In contrast, a staurosporine-treated czt-1 deletion strain is unable to repress the genes for the respiratory chain and to induce the genes related with the endoplasmic reticulum, indicating a role for CZT-1 on the regulation of activity of these organelles.
Conclusions: This transcriptional profiling study is framed in a comprehensive study on the role of the novel transcription factor CZT-1 (NCU09974) in Neurospora crassa.

overall_design: Transcriptional profile of wild type and Δczt-1 cells staurosporine-treated cells in Neurospora crassa was compared using illumina HiSeq2000 and single reads of 50 bp.

first: A

middle: P


title: Direct ChIP-bisulfite sequencing reveals a role of H3K27me3 mediating aberrant hypermethylation of promoter CpG islands in cancer cells (ChIP-BS)

summary: The model describing that aberrant CpG island (CGI) methylation leads to transcription repression of tumor suppressor genes and thereby is implicated in tumor progression has been established in many cancers. However, recent studies indicated aberrantly hypermethylated genes in multiple cancers are already repressed in pre-cancerous tissues despite their promoters are hypomethylated. Here, we hypothesized that the occurrence of CGI promoter hypermethylation in cancers are associated with Polycomb-repressive complex and the associated H3K27me3 mark in pre-cancerous tissues. By using a ChIP-BS-seq technology that examines methylation of the DNA fragments precipitated by the antibodies to histone modifications, we provided direct evidences showing that genes highly enriched with H3K27me3 marks both in cancer and normal cells became aberrantly hypermethylated in CGI promoters in cancer cells in comparison with normal cells. Furthermore, we confirmed that these genes consistently were significantly hypermethylated in TCGA primary cancer in comparison with normal tissues. Thus, we provided direct evidences supporting that the presence of H3K27m3 may serve as a guide to promoter hypermethylation. This will spur future work on epigenetic signature of combined histone and DNA methylation that could define a cancer’s epigenetic abnormalities, therefore helping distinguish subtypes of cancers and aiding future diagnosis and therapeutics of cancers.

overall_design: We applied ChIP-BS technology to examine H3K27me3 marks, which are catalyzed by the SET domain histone methyltransferase EZH2 and have a repressive function with 50bp pair-end sequencing. found H3K27me3 marks were enriched preferentially at CpG islands, (+/-500) transcription start sites (TSSs) and exons in two GC cell lines (BGC-823 and AGS). In YH cells, H3K27me3 marks were only preferentially enriched at CpG islands. In contrast, Hela cells presented a reverse pattern with highest H3K27me3 enrichment in intergenic regions. To confirm this result in Hela cells, we performed two independent replicates of ChIP-Seq and ChIP-BS-seq. Cause of useful was relative small. we still sequenced one 100bp pe reads replicate for H3K4me3 and two replicate for H3K27me3 ChIP-BS-seq.

first: Fei

last: Gao

first: Guanyu

last: Ji

first: Zhaowei

first: Xu

last: Han

first: Mingzhi

last: Ye

first: Zhimei

last: Yuan

first: Huijuan

last: Luo

first: Xiaojun

last: Huang

first: Huanming

last: Yang

first: Xiuqing


title: Identification and expression profiling of microRNAs in brain, liver and gonads of marine medaka (Oryzias melastigma) by Illumina small RNA Sequencing

city: Notre Dame

first: Yunjuan

last: Bao

state: Indiana

address: 1234 Notre Dame Avenue

institute: University of Notre Dame

type: Non-coding RNA profiling by high throughput sequencing

summary: We performed miRNA transcriptome analysis of mature medaka tissues using high-throughput next generation sequencing technology. The small RNA libraries were contructed from brain, liver and gonads in mature male and female medaka and a total of more than 128 million sequencing reads were generated from the six small RNA libraries. By comparison to known miRNA database, a total of 223 known miRNA types were identified with more than 50% expressed in brain. The expression profiling showed that almost half of the miRNAs are tissue-specific, with only 55 miRNA types from 34 families common to all tissues. A small number of miRNAs are also gender-specific. The experimental validation was performed for five represented miRNA candidates and four of them are confirmed.

overall_design: Investigation of miRNA expression profiles from three tissues (brain, liver and gonads) of mature male and female medaka.

first: Karen

middle: Y

last: Lau

first: Jessie


title: Epigenetic remodeling and deregulation of SAPAP4/DLGAP4 is linked with early-onset cerebellar ataxia

city: Copenhagen

first: Sheroy

last: Minocherhomji

state: Copenhagen N

address: Blegdamsvej 3B, Building 18.1

department: Nordea Centre for Healthy Aging, Faculty Of Health Sciences Department of Cellular and Molecular Medicine

country: Denmark

institute: University of Copenhagen

summary: Epigenetic effects at the breakpoints of disease-associated balanced chromosomal rearrangements are not fully understood. To elucidate and characterize epigenetic effects of such a rearrangement, we studied the familial translocation t(8:20)(p12;q11.23) inherited across five generations together with  early-onset non-progressive cerebellar ataxia. This translocation disrupts DLGAP4 and a DLGAP4 CpG island at 20q11.23, a locus encoding a brain-specific DLGAP4 mRNA (isoform a) and a gene-desert at 8p12. We find describe translocation-dependent, monoallelic epigenetic changes at DLGAP4 affecting its expression in cis. We found hypermethylation of the disrupted DLGAP4 CpG island and a delay in replication timing of this locus in multiple carriers. Hypermethylation of CpGs at the disrupted DLGAP4 CpG island is re-established in the somatic cells of each translocation carrier after being erased in their germ cells. Brain-specificity of DLGAP4 mRNA isoform a is maintained in carriers. However, this rearrangement leads to increased mono-allelic expression of another DLGAP4 mRNA isoform b from the truncated DLGAP4 locus. Furthermore, there is a translocation dependent enrichment of the conserved histone variant H2A.Z at the undisrupted DLGAP4 CpG island. Functional analyses reveal a novel intronic non-coding RNA, transcribed from the DLGAP4 promoter region and the DLGAP4 mRNA isoform a, both of which are and disrupted by the translocation, to be and highly expreed in the human brain. Using in situ hybridization we found this ncRNA to be bi-directionally expressed in human and mouse cerebellum, relative to elevated expression of DLGAP4 mRNA isoform a only in Purkinje neurons. These data provide a basis for the understanding of the roles epigenetic regulatory mechanisms transmitted through germ cells have in controlling expression of disrupted gene loci highly expressed in the human brain.

overall_design: Examination of hypermethylation (MBD-Seq using a his-MBD2B antibody) and hypomethylation (MRE-Seq using HpaII restriction enzyme digestion) in translocation t(8;20)(p12;q11.23) carriers and normal controls. ChIP-Seq using H2A.Z, H3K4me3 and H3K27me3 antibodies in EBV-transformed peripheral blood lymphocytes obtained from translocation carriers and controls.

first: Yuan

last: Mang

first: Niels

last: Tommerup

first: Asli

last: Silahtaroglu


title: Direct ChIP-bisulfite sequencing reveals a role of H3K27me3 mediating aberrant hypermethylation of promoter CpG islands in cancer cells (RNA-seq)

first: Desheng

summary: Aberrant DNA hypermethylation of CpG island (CGI) promoters are associated with transcriptional repression of many tumor suppressor genes and lead to tumor progression in many cancers. Most recently, one research group observed that aberrantly hypermethylated genes in multiple cancers are already repressed, but their promoters are maintained in a hypomethylated state in pre-cancerous tissues.Their studies didn't provide a clue to explain by what mechanisms those genes were repressed in pre-cancerous tissues. Another research group found that many genes with de novo promoter hypermethylation in colon cancer were among the subset of genes "bivalently" marked in embryonic stem cells and adult stem/progenitor cells by repressive Polycomb group proteins (PcG), which are known for maintaining low, but poised, transcription.These observations provide a clue that CGI promoter hypermethylation in cancers is associated with PcG target genes in pre-cancerous tissues.we took advantage of ChIP-BS-seq technology and applied it to examine H3K27me3 and H3K4me3 profiles for one normal lymphoblastoid cell line (YH) and three cancer cell lines including one cervical cancer cell line (Hela) and two gastric cancer (GC) cell lines (BGC-823 and AGS).

overall_design: We aplied ChIP-BS technology to examine H3K27me3 marks, which are catalyzed by the SET domain histone methyltransferase EZH2 and have a repressive function with 50bp pair-end sequencing. found H3K27me3 marks were enriched preferentially at CpG islands, (+/-500) transcription start sites (TSSs) and exons in two GC cell lines (BGC-823 and AGS). In YH cells, H3K27me3 marks were only preferentially enriched at CpG islands. In contrast, Hela cells presented a reverse pattern with highest H3K27me3 enrichment in intergenic regions. To confirm this result in Hela cells, we performed two independent replicates of ChIP-Seq and ChIP-BS-seq. Cause of useful was relative small. we still sequenced one 100bp pe reads replicate for H3K4me3 and two replicate for H3K27me3 ChIP-BS-seq.

first: Fei

last: Zhang


title: Characterization of the small RNA populations of developing Arabidopsis thaliana pollen.

city: Leicester

first: David

last: Twell

state: Leicestershire

address: Department of Biology

institute: University of Leicester

summary: Small RNA diversity and function has been widely characterized in various tissues of the sporophytic generation of the angiosperm model Arabidopsis thaliana. In contrast, there is limited knowledge about small RNA diversity and their roles in developing male gametophytes. We thus carried out small RNA sequencing on RNA isolated from four stages of developing Arabidopsis thaliana pollen.

overall_design: Spores from 4 stages of pollen development (UNM: Uninucleate microspore – BCP: Bicellular pollen – TCP: Tricellular pollen – MP: Mature pollen) were isolated using a percoll gradient-based method (Honys and Twell, 2004) and the small RNA fraction for each sample was isolated and sequenced by Illumina technology. Reference: Honys, D. and Twell, D. (2004) Transcriptome analysis of haploid male gametophyte development in Arabidopsis. Genome Biol. 5/11/R85.

first: Gael

last: LeTrionnaire

first: Said

last: Hafidh

first: Ralf

last: Schmid

first: Robert

last: Grant-Downton

first: Sofia

last: Kourmpetli

first: Hugh

last: Dickinson


title: c-Jun promotes cell migration and drives expression of the motility factor ENPP2 in soft tissue sarcomas [RNA-Seq]

status: Public on Jun 30 2014

city: Boston

first: Stefano

last: Sioletic

state: Ma

address: 450 Brookline Ave, Boston,

department: Medical Oncology

institute: Dana Farber Cancer Institute

summary: We assayed the effect of c-Jun overexpression on gene expression in the three DDLPS cell lines using RNA-Seq (Illumina).

overall_design: 141, LPS12 and 510 has been overexpressed with c-Jun or control c-DNA and results were analyzed in high-througput sequencing metadata.

first: Lan

last: Hu

first: Eric

last: Snyder


title: Transcriptome sequencing at early stage infection of the wheat pathogen Zymoseptoria tritici reveals chromosomal differences in transcription patterns and host specific gene expression.

city: Marburg

first: Ronny

last: Kellner

address: Karl-von-Frischstrasse 10

country: Germany

institute: MPI for Terrestrial Microbiology

summary: We have applied whole transcriptome profiling to infer genetic determinants of pathogenicity and host specialization in Z. tritici. Our data includes RNAseq data from early infection stages of a compatible (wheat) and a non-compatible host (Brachypodium distachyon). Overall transcription of AC genes is remarkably lower than genes on core chromosomes (CC) and only 40% of the genes are transcribed. We identify 31 AC and 1069 CC genes showing plant specific expression. In addition 21 CC genes are only upregulated in wheat supporting functional relevance in host specificity. We further explore the genomic composition and distribution of unique and paralogous genes in Z. tritici focusing on the evolutionary origin of AC genes. In contrast to previous studies we show that ACs mainly encode unique genes. Phylogenetic analyses suggest that rare duplication events in the Z. tritici genome precede diversification of Zymoseptoria species and demonstrate that ACs have been maintained in the genome of Zymoseptoria over long evolutionary times.

overall_design: Examination of gene expression at 3 different growth condition of the wheat pathogen Z. tritici.

first: Eva

last: Stukenbrock


title: Aorta- and liver-specific ERalpha-binding patterns and gene regulation by estrogen

first: Gavin

middle: R.

last: Schnitzler

state: MA

address: 800 Washington St. Box 80

department: Molecular Cardiology Research Institute

institute: Tufts Medical Center

summary: Estrogen has vascular protective effects in premenopausal women and in women under 60 receiving hormone replacement therapy. However, estrogen also increases risks of breast and uterine cancers and of venous thromboses linked to upregulation of coagulation factors in the liver. In mouse models, the vasoprotective effects of estrogen are mediated by the estrogen receptor alpha (ERa) transcription factor. Here, through next generation sequencing approaches, we show that almost all of the genes regulated by 17-b-estradiol (E2) differ between mouse aorta and mouse liver, and that this is associated with a distinct genomewide distribution of ERa on chromatin. Bioinformatic analysis of E2-regulated promoters and ERa binding site sequences identify several transcription factors that may determine the tissue specificity of ERa binding and E2-regulated genes, including the enrichment of NFkB, AML1 and AP-1 sites in the promoters of E2 downregulated inflammatory genes in aorta but not liver. The possible vascular-specific functions of these factors suggests ways in which the protective effects of estrogen could be promoted in the vasculature without incurring negative effects in other tissues. Our results also highlight the likely importance of rapid signaling of membrane-associated ERa to cellular kinases (altering the activities of transcription factors other than ER itself) in determining tissue specific transcriptional responses to estrogen.

overall_design: The aortas or liver fragments of wild-type C57/BL6 mice were incubated ex vivo with 10nM E2 or ethanol vehicle for 4 hours before harvesting for RNA collection. Each condition was performed with two biological replicates, and each replicate contained aortas or liver fragments from 4 mice.

first: Francesca

last: Gordon

first: Caroline

last: Vallaster

first: Thomas

last: Westerling

first: Lakshmanan

last: Iyer

first: Myles

last: Brown

middle: R


title: SIRT1 affects DNA methylation of polycomb group protein target genes (PGCTs), a hotspot of the epigenetic shift observed in ageing.

city: Newcastle

first: Simon

last: Cockell

address: Framlington Place

institute: Newcastle University

summary: Beneficial effects of SIRT1 on healthspan are likely to be pleiotropic and may include effects on DNA methylation.  We demonstrated recently that manipulating SIRT1 in human cells affected DNA methylation of a panel of test genes, and that genes with expression modified by dietary restriction corresponded with genes that underwent changes in DNA methylation during ageing.  Here we tested the hypothesis that genes particularly susceptible to SIRT1-induced effects on DNA methylation across the genome map to genes for which DNA methylation changes during ageing.  We increased or reduced SIRT1 expression in human intestinal (Caco-2) and vascular endothelial (HuVEC) cells by transient transfection with an expression construct or with siRNA respectively.  Effects on DNA methylation were measured by enriching for the methylated faction then either sequencing (HuVEC) or hybridising to a human promoter microarray (Caco-2).  Effects using these two different cell lines and techniques for analysis were remarkably consistent.  Genes with a DNA methylation status affected by SIRT1 manipulation were enriched for those that undergo age-dependent changes in DNA methylation, thus supporting our hypothesis.  Polycomb group protein target genes (PCGTs), which are suppressed by epigenetic mechanisms in stem cells and have been shown previously to correspond with loci particularly susceptible to age-related changes in DNA methylation, were over-represented within the set of genes showing altered DNA methylation in response to SIRT1 manipulation in both cell lines.  We thus propose that effects of SIRT1 to extend healthspan include influences on the DNA methylation status of genes affected during ageing, in particular PCGTs.

overall_design: MBD-Sequencing to ascertain effects of SIRT1 over & under expression on methylation, in presence and absence of TNF-alpha. One sample per condition.

first: Luisa

last: Wakeling

first: Laura

middle: J

last: Ions

first: Gail

last: Jenkins

first: Linda

last: Wainwright

first: Jill

last: McKay

first: Dianne

last: Ford


title: Genome-wide Profiling of SRC-2 (NCOA2) binding in mouse prostate [ChIP-Seq]

city: Houston

first: Hui-Ju

last: Lee

state: Texas

address: One Baylor Plaza

department: Molecular & Cellular Biology

institute: Baylor College of Medicine

summary: Using genetically engineered mice, overexpressing SRC-2, specifically in the prostate epithelium of PTEN heterozygous mice accelerates PTEN mutation induce tumor progression and develops a metastasis-prone cancer. Here we used ChIP-Seq analysis to identify genome-wide SRC-2 binding sites in mouse prostate.

overall_design: Examination of genome-wide SRC-2 binding in mouse prostate by ChIP-seq analysis.  Samples were collected from pooled dorsal-lateral prostates of 7 months old-PTEN flox/+; Rosa26-SRC-2 OE/+ mice.  Flash frozen tissues were then sent to Active Motif, Inc. for chromatin extraction and followed by immunoprecipitation using anti-SRC-2 antibody (A300-346A, Bethyl Lab., Inc.).

first: Sophia

last: Tsai

first: Ming-Jer


title: Discovery of a "White-Gray-Opaque" Tristable Phenotypic Switching System in Candida Albicans: Roles of Non-Genetic Diversity in Host Adaption

city: Beijing

first: guanghua

last: huang

address: NO.1 West Beichen Road, Chaoyang District

institute: Institute of Microbiology

summary: The capacity of the commensal yeast Candida albicans to grow in several forms, referred to as phenotypic plasticity, is critical for its survival, and abilities to thrive and cause infection in the human host. In this study, we report a novel phenotype of C. albicans, referred as the “gray” phenotype. The gray cell type, together with the previously discovered “white” and “opaque” cell types, forms a tristable phenotypic switching system. The three phenotypes differ in cellular and colony appearance, global transcriptional profiles, secreted aspartyl proteinase (Sap) activities and virulence in different infection models. The addition of the gray cell type to the phenotypic transition systems may enhance fitness and confers an adaptive advantage for C. albicans in the host. We further demonstrate that two key regulators of white-opaque switching, Wor1 and Efg1, are not required for maintenance of the gray phenotype, suggesting that a different regulatory circuitry may be involved in the regulation of gray cell formation. Our study provides an example of multiple stable and heritable switching systems, indicating that the regulation of morphological forms to adapt to environmental changes could be much more elaborate than previously thought.

overall_design: total RNA profiles of 24h old white, gray and opaque cell types

first: Guanghua

last: Huang

first: Li

last: Tao

first: Han

last: Du


title: H3K27ac profiling in control and DSS-treated colonic epithelium

city: Piscataway

first: Michael

last: Verzi

state: NJ

address: 145 Bevier Road

institute: Rutgers, the State University of New Jersey

summary: We determined changes in enhancer chromatin that occur during colonic inflammation, found that dynamic chromatin regions are enriched for HNF4A binding motirfs, and then measured HNF4A binidng by ChIP-seq in each condition.

overall_design: Examination of H3K27ac histone modification in normal and DSS-treated colon, examination of HNF4A binding by ChIP-seq in normal and DSS-treated colon epithelium

first: Sanjay

last: Chahar

first: Mike


title: Aorta- and liver-specific ERalpha-binding patterns and gene regulation by estrogen (ChIP-seq)

first: Gavin

last: Schnitzler

overall_design: The aortas or liver fragments of wild-type C57/BL6 mice were incubated ex vivo with 10nM E2 or ethanol vehicle for 45 minutes before formaldehyde fixation, harvesting of chromatin & ChIP with anti- mouse estrogen receptor alpha antibodies. Each condition was performed with two biological replicates, and each replicate contained aortas or liver fragments from 5 mice.

first: Francesca

last: Gordon

first: Caroline

last: Vallaster

first: Thomas

last: Westerling

first: Lakshmanan

last: Iyer

first: Myles


title: Alveolar Macrophage Gene Expression in Human Pulmonary Fibrosis

city: Bethesda

first: Eric

last: Billings

state: MD

address: 10 Center Drive

institute: NHLBI

summary: Gene expression profiles for patients affected with Sporadic and Familial Pulmonary Fibrosis.

overall_design: Case/control study of alveolar macrophage response in 15 IPF and 8 familial pulmonary fibrosis patients; and 45 normal volunteers and 16 unaffected relatives.

first: Ivan

middle: O

last: Rosas


title: Gene expression profiling of breast cancer cells with knockdown of PTEN

first: Chad

last: Creighton

state: TX

address: One Baylor Plaza, Mail Stop: BCM305

department: Biostatistics, Ducan Cancer Center

summary: Activation of the PI3K pathway in estrogen receptor α (ER)-positive (+) breast cancer is associated with reduced ER expression and activity, luminal B subtype, and poor outcome.  PTEN is a negative regulator of the PI3K pathway typically lost in ER-negative (-) breast cancer.  To clarify the effect of PTEN down-regulation on the response of ER+/HER2- breast cancer to endocrine therapy, we established reduced PTEN cell models using inducible knockdown.  We found that only moderate PTEN reduction is sufficient to enhance PI3K signaling, generate a gene signature associated with luminal B subtype, and cause endocrine resistance.  Combining endocrine therapy with mTOR, AKT, or MEK inhibitors improves antitumor activity, but the efficacy varies by type of endocrine therapy and the specific inhibitor.  Fulvestrant plus an AKT inhibitor is the most potent combination when PTEN is reduced, inducing apoptosis and tumor regression.  This combination deserves further study in patients with PI3K pathway activation.

overall_design: Gene expression profiles (by RNA-seq) were taken of a cell model with or without reduced PTEN (using inducible knockdown).

first: Xiaoyong

last: Fu

first: Rachel

last: Schiff


title: RNA-seq Analysis Reveal Global Differential Transcritomes of Mesorhizobium huakuii 7653R between Bacteroids and Free-living Cells

city: Wuhan

first: Youguo

last: Li

state: Hebei

address: No.1,Shizishan Street · Hongshan District

department: College of Life Science and Technology

institute: Huazhong Agricultural University

summary:
Mesorhizobium huakuii 7653R is an α-proteobacterium that occurs either in a nitrogen-fixing symbiosis with its host plant, A. sinicus, or free-living in the soil.
We performed RNA-Seq on free-living cells grown in rich media and in bacteroids isolated from root nodules to understand how M. huakuii 7653R responds to alterations in its environment and to the physiological changes that occur during bacteroid differentiation.
Understand how M. huakuii 7653R responds to alterations in its environment and to the physiological changes that occur during bacteroid differentiation.

overall_design: Examination of mRNA levels in free-living cells and bacteroids at 32 days postinoculation

first: Jieli

last: Peng

first: Shanming

last: Wang


title: NFATc1 integrates STAT3 inflammatory signals to promote Kras(G12D)-driven carcinogenesis in the pancreas.

city: Giessen

first: Marek

last: Bartkuhn

state: Hessen

address: Aulweg 132

department: Biomedical Informatics and Systems Medicine

institute: Justus-Liebig-University Giessen

summary: Inflammatory transcription networks have been linked with the development of pancreatic ductal adenocarcinoma (PDAC). Here, we demonstrate that NFATc1 is both necessary and sufficient to drive progression of KrasG12D-initiated PDAC, particularly in the context of inflammation. Significantly, nuclear NFATc1 accelerates PDAC development in KrasG12D mice, whereas conditional NFATc1 deletion or pharmacological inhibition attenuates inflammation-mediated carcinogenesis. Mechanistically, NFATc1 induces STAT3 expression, complex formation and signal integration in PDAC. Genome-wide ChIP-sequencing and expression analysis in cells derived from c.n.NFATc1;KrasG12D mice identified combinatorial NFATc1/STAT3 binding at chromatin enhancer sites and subsequent regulation of key molecules involved in oncogenic signaling, growth and inflammation. Together, this study supports the relevance of inflammatory transcription factor networks in pancreatic carcinogenesis and provides a theoretical platform for therapeutic targeting of NFATc1 nucleoprotein complexes in PDAC.

overall_design: NFATc1 ChIP followed by high throughput sequencing in primary murine pancreatic cancer cells (referred to as NKC cells) derived from transgenic mice with pancreas-specific constitutive activation of NFATc1 and KrasG12D mutation in the presence or absence of STAT3 shRNA; 2 ChIP samples (scramble DNA and shSTAT3 DNA) and 1 unenriched input control from the same chromatin pool.

first: Sandra

last: Baumgart

first: Elmar

last: Wolf

first: Volker

last: Ellenrieder


title: the analysis of piRNA shows the function of Slicer activity of AGO3 in piRNA biogenesis.

first: Haidong

last: Huang

address: 1 Beichen West Road, Chaoyang District

institute: Institute of Zoology, Chinese Academy of Sciences

summary: Piwi-interacting RNAs (piRNAs) silence transposons in animal germ cells. In Drosophila, the reciprocal “Ping-Pong” cycle of piRNA-directed RNA cleavage, catalyzed by the PIWI proteins Aubergine (Aub) and Argonaute3 (AGO3) through their Slicer activity, is believed to expand the population of antisense piRNAs in response to transposon expression. Whether and how the Slicer activity of AGO3/Aub promotes the process of the secondary piRNA amplification remain unclear. Here we generated transgenic flies that could express AGO3 Slicer mutant forms to ellucidate the Slicer activity of AGO3.

overall_design: small-RNA libraries from 3 samples of D. mel. Ovaries.

first: Yujing


title: RNA-sequencing reveals allelic expression imbalance in the diploid pathogen Candida albicans

city: Aberdeen

first: Sophie

last: Shaw

state: Aberdeen City

address: 23 St. Machar Drive

department: Centre for Genome Enabled Biology and Medicine

institute: University of Aberdeen

summary: The diploid fungal pathogen Candida albicans is a highly heterozygous organism, with numerous non-synonymous substitutions often seen within two alleles. RNA-sequencing of the wild-type strain SC5314 has revealed 233 genes with significant levels of allelic expression imbalance. Overall percentage protein identity comparisons were significantly lower in these differentially expressed alleles. This suggests that two different, perhaps functionally divergent, proteins are being expressed at significantly different quantities by the two alleles of a single gene. Previously, gene expression levels have been correlated with structural factors such as GC content, ORF length and codon usage. Here, these factors were first correlated with overall gene expression data to decipher the relationship they have with gene expression in Candida albicans. These relationships were then used to assess the contribution of these factors to allelic expression imbalance. GC content and codon usage did not differ significantly in differentially expressed alleles whereas ORF length was found to be significantly lower in the allele with lowest expression. This surprising result goes against the overall trend observed between length and gene expression. Differences in GC content and ORF length between alleles correlated strongly with percentage protein identity, suggesting an indirect link between these factors and allelic expression imbalance.

overall_design: One sample (SC5314: wild-type strain) assessed in triplicate and compared to the reference diploid genome

last: Clarke

first: Mark

last: Ramsdale

first: Konrad

last: Paszkiewicz

first: Alex

last: Moorhouse


title: Insights into the invasiveness of triple negative breast cancer from genome-wide profiling of transcription factor AP-1 (ChIP-seq)

city: Stockholm

first: Chunyan

last: Zhao

address: Hälsovägen 7-9

department: Biosciences and Nutrition

country: Sweden

institute: Karolinska Institute

summary: Triple negative breast cancer (TNBC) is an aggressive clinical phenotype, and accounts for 15% to 20% of all breast cancers. The molecular determinants of malignant cell behaviors in TNBC remain largely unknown. We find that the AP-1 transcription factor component, Fra-1, is overexpressed in basal-like breast tumors, and its expression level has high prognostic significance. Depletion of Fra-1 or its heterodimeric partner c-Jun inhibits the proliferative and invasive phenotypes in TNBC cells. To gain insights into the transcriptional regulatory networks of AP-1 in TNBC cells, we combine genome-wide ChIP-seq with loss-of-function transcriptome analyses. We observe dysregulation of direct targets of the Fra-1/c-Jun heterodimer involved in cell proliferation, cell adhesion, and cell-cell contact. Intriguingly, we find that AP-1 mediates downregulation of E-cadherin through direct transcriptional induction of ZEB2. This work sheds light on the mechanisms and pathways by which TNBC acquires invasiveness and proliferative propensity.

overall_design: Examination of genome-wide binding sites of AP-1 transcription factor components, Fra-1 and c-Jun, in BT549 breast cancer cell line.

first: Yichun

last: Qiao

first: Karin

last: Dahlman-Wright

first: Philip

last: Jonsson

first: Cecilia

last: Williams

first: Li

last: Xu

first: Indranil

last: Sinha


title: Id2 and Id3 maintain the regulatory T cell pool to suppress inflammatory disease

status: Public on Jun 29 2014

city: La Jolla

first: Kazuko

last: Miyazaki

state: CA

address: 9500 Gilman Drive

department: Division of Biological Sciences

institute: University of California, San Diego

summary: Regulatory T (Treg) cells suppress the development of inflammatory disease, but our knowledge of transcriptional regulators that control this function remains incomplete. Here we show that expression of Id2 and Id3 in Treg cells was required to suppress development of fatal inflammatory disease. We found that T cell antigen receptor (TCR)-driven signaling initially decreased the abundance of Id3, which led to the activation of a follicular regulatory T (TFR) cell–specific transcription signature. However, sustained lower abundance of Id2 and Id3 interfered with proper development of TFR cells. Depletion of Id2 and Id3 expression in Treg cells resulted in compromised maintenance and localization of the Treg cell population. Thus, Id2 and Id3 enforce TFR cell checkpoints and control the maintenance and homing of Treg cells.

overall_design: Treg mRNA profiles in lymph node from 3-week-old Id2fl/flId3fl/fl;Foxp3Cre/Cre (Id2 Id3 double-knockout) and control mice are generated by deep sequencing.

first: Masaki

first: Shuwen

last: Chen

first: Manami

last: Itoi

first: Marina

last: Miller

middle: F

last: Lu

first: Nissi

last: Varki

first: Aaron

middle: N

last: Chang

first: David

last: Broide

first: Cornelis

last: Murre


title: Analysis Reveal Global Differential Transcritomes of Mesorhizobium huakuii 7653R between Bacteroids and Free-living Cells

first: Youguo

last: Li

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

overall_design: Refer to individual Series


title: Long-range chromatin interactions in mouse embryonic stem cells identified by ChIA-PET [H3K27me3 ChIP-Seq]

status: Public on Jun 27 2014

city: Cambridge

first: Richard

last: Young

address: 9 Cambridge Center

institute: Whitehead Institute for Biomedical Research

summary: The pluripotent state of embryonic stem cells (ESCs) is produced by active transcription of cell identity genes and repression of genes encoding lineage-specifying developmental regulators. Here we use large ESC cohesin ChIA-PET datasets and other genomic data to identify the local chromosomal structures at both active and repressed genes across the genome. The results show that super-enhancer driven cell identity genes generally occur within large loops that are connected through CTCF-CTCF interaction sites occupied by cohesin.

overall_design: H3K27me3 ChIP-seq data from wild type murine embryonic stem cells V6.5 were generated by deep sequencing using Illumina Hi-Seq 2000.

first: Warren

last: Whyte


title: Long-range chromatin interactions in mouse embryonic stem cells identified by ChIA-PET


title: Rapid transcriptional response to physiological neuronal activity in vivo revealed by transcriptome sequencing

first: Yarden

last: Katz

department: Dept. of Biology and Whitehead Institute

institute: MIT

summary: mRNA-seq of olfactory bulbs from odor-deprived and odor-stimulated mice (RNA-seq done in triplicates)

overall_design: Control mice were kept in a clean air environment (using an activated carbon filter). Stimulated mice were exposed to a cocktail of odorants for 30 minutes ("30 min" time point), or for 30 minutes followed by a period of 90 minutes of clean air ("120 min" time point.) Mice were sacrificed immediately at end of time point, and olfactory bulbs were dissected out for RNA extraction and RNA-FISH. Three adult mice (6-8 weeks of age or older) were used in each condition.


title: c-Jun promotes cell migration and drives expression of the motility factor ENPP2 in soft tissue sarcomas

first: Stefano

last: Sioletic


title: Insights into the invasiveness of triple negative breast cancer from genome-wide profiling of transcription factor AP-1

first: Chunyan

last: Zhao


title: Next generation sequencing based transcriptomic analysis of bile stress response in Bifidobacterium longum BBMN68

first: Haoran

last: An

address: 17th Qinghua East Road

institute: China Agricultural University

summary: Bifidobacterium longum is one of the natural inhabitants in human gastrointestinal tract. In order to colonize and exert particular functions in the gut, it has to be tolerant to the physiological concentrations of bile salts. In this work, we used RNA-Seq transcriptomics based on the Next-Generation Sequencing to investigate the global response to bile in B. longum BBMN68, a potential probiotic strain isolated from a healthy centenarian. In the presence of 0.75 g liter-1 ox-bile, the transcription of 236 genes were regulated (≥ 3-fold, p < 0.001). Function analysis and Gene Ontology suggested that the bile stress response of B. longum BBMN68 covers almost all biological processes, including bile stress resistance, general stress response, central metabolic process, transmembrane transport, gene expression, cell proliferation and interaction with the host. Remarkably, 3 two-component systems and 11 transcription factors were up- or down-regulated by bile, and target genes of the regulators were identified by bacterial one-hybrid system and bioinformatics methods, resulting in a putative regulatory network that controls bile stress response in B. longum for the first time. This study significantly develops our understanding on bile stress response and brings new insight to the regulatory mechanism in bifidobateria.

overall_design: Whole mRNA profiles of B. longum BBMN68 growing in the absence (CK) and presence (OG) of ox-bile were generated using AB SOLiD technology and differentially expressed genes were anylyzed.

first: Yanling

last: Hao


title: In vivo LPS responses in murine splenic CD8 and CD11b DC subsets

city: Camperdown

middle: Graeme

last: Hancock

state: NSW

address: Building 93, RPAH, Missenden Road

country: Australia

institute: Centenary Institute

summary: Background: Dendritic cells (DCs) are critical for regulating CD4 and CD8 T cell immunity, controlling Th1, Th2, and Th17 bias, generating inducible Tregs, and inducing tolerance. Multiple DC subsets have been identified in the mouse that are thought to have evolved to control these different immune outcomes. However, how these subsets differentially respond to inflammatory and/or tolerogenic signals in order to accomplish their divergent functionality remains unclear.     Results: We analysed the responses of murine, splenic CD8 and CD11b DC subsets to in-vivo stimulation with lipopolysaccharide using RNA-Seq and systems biology approaches and observed responses are highly subset-specific. We reanalysed multiple datasets from the literature and show that these subset responses are obscured when analysing signaling at the population level. We show that the subset-specificity is due to the unique regulation of distinct TLR4 pathway modulators that ‘fine-tune’ a common TLR4 cascade rather and not due to major differences in signaling pathways or transcription factors.     Conclusions: We propose the Pathway Modulation Model wherein common signaling pathways are regulated by unique sets of modulators allowing for distinct immune responses in closely related DC subsets. We extend these observations using analagous datasets from the literature and show that our model provides a global mechanism for generating cell subset-specific signaling in multiple subpopulations in mouse and man.

overall_design: Splenic CD8 and CD11b DC subsets from LPS stimulated (10 pooled animals) and Control (5 pooled animals) mice were analysed by RNA-Seq.

middle: G

first: Elena

last: Shklovskaya

first: Thomas

middle: V

last: Guy

first: Reza

last: Falsafi

first: Chris

middle: D

last: Fjell

first: William

last: Ritchie

first: Robert

first: Barbara

last: Fazekas de St Groth


title: Core promoter factor TAF9B controls neuronal gene expression

city: Berkeley

first: Francisco

last: Herrera

address: 475D Li Ka Shing Center

department: Molecular and Cell Biology

institute: University of California, Berkeley

summary: Purpose: Characterize the role of the coactivator subunit TAF9b during differentiation of embryonic stem cells into motor neurons as well in mouse newborn spinal column tissues.

overall_design: RNA-seq comparing WT and TAF9B KO mouse ES cells differentiated into motor neurons. RNA-seq comparing WT and TAF9B KO mouse newborn spinal column tissues. ChIP-seq mapping TAF9b and RNA Pol II binding sites in in vitro differentiated motor neurons.


title: Developmental Profile of DNA Methylation in the Developing Heart

city: ny

first: mingyan

last: lin

address: 1925 morris park ave

institute: aecom

summary: Background: To better understand the role DNA methylation plays in regulating gene expression in the developng heart and furthermore the role it plays in heart development we performed  genome wide DNA methylation profiling of embryonic hearts at embryonic day (E)11.5 and E14.5 using methyl sensitive tiny fragment enrichment coupled with massive parallel sequencing by using the methyl-sensitive restriction enzyme HpyCH4IV, recognition site 'ACGT'. Results: We found that global methylation remains stable at analyzed 'ACGT' sites (1.64 million site) in developing hearts between E11.5 and E14.5. However, differential methylation was identified at individual loci enriched at genes involved in heart development suggesting a role for DNA methylation in the developing heart.

overall_design: Used Methyl Sensitive Tiny Fragment Enrichment/Massive Parallel Sequencing (MSFE/MPS) to assay methylation at 'ACGT' sites throughout the genome and generate a developmental profile of DNA methylation in the embryonic heart and to identify differences between developing mouse hearts at E11.5 and E14.5.

first: Alyssa

middle: A

last: Chamberlain

first: Bin

last: Zhou


title: RNA-sequencing of Culex pipiens molestus and Culex pipiens quinquefasciatus

status: Public on Jun 26 2014

first: Chunxiao

last: Li

address: 20 Dongdajie, Fengtai District

institute: Beijing Institute of Microbiology and Epidemiology

summary: Culex pipiens molestus and Cx. p. quinquefasciatus are the members of Culex pipiens Complex, but they display relatively large differences in behavior and physiological responses. We compared the genes of these mosquitoes to identify those that were differentially expressed in each subspecies. Such genes could play important roles in subspecies-specific blood feeding or oviposition behavior.

sample_organism:
Culex quinquefasciatus
Culex pipiens molestus

overall_design: Culex pipiens molestus and Cx. p. quinquefasciatus females were undertaken Illumina RNA sequencing.

first: Tongyan


title: VDR primary targets by genome-wide transcriptional profiling

status: Public on Jun 25 2014

city: Rome

first: Matteo

last: Pallocca

address: Via Fermo Ognibene

country: Italy

institute: Italian National Cancer Institute "Regina Elena"

summary: We carried out a genome-wide investigation of the primary transcriptional targets of 1α,25(OH)2D3 in breast epithelial cancer cells using RNA-Seq technology. We identified early transcriptional targets of 1α,25(OH)2D3 involved in adhesion, growth regulation, angiogenesis, actin cytoskeleton regulation, hexose transport, inflammation and immunomodulation, apoptosis, endocytosis and signaling. Furthermore, we found several transcription factors to be regulated by 1α,25(OH)2D3 that subsequently amplify and diversify the transcriptional output driven by 1α,25(OH)2D3 leading finally to a growth arrest of the cells. Moreover, we could show that 1α,25(OH)2D3 elevates the trimethylation of histone H3 lysine 4 at several target gene promoters. Our present transcriptomic analysis of differential expression after 1α,25(OH)2D3 treatment provides a resource of primary 1α,25(OH)2D3 targets that might drive the antiproliferative action in breast cancer epithelial cells.

overall_design: ChIP-Seq for trimethylated histone H3K4 with SKBr3 cells treated for 2h with 100nM 1α,25(OH)2D3 or vehicle as control.

first: Frauke

last: Goeman

first: Francesca

last: De Nicola

first: Paolo

last: D’Onorio De Meo

first: Berardino

last: Elmi

first: Tiziana

last: Castrignanò

first: Graziano

last: Pesole

first: Sabrina

last: Strano

first: Giovanni

last: Blandino

first: Maurizio

last: Fanciulli

first: Paola

last: Muti


title: DOCK8 regulates protective immunity by controlling the function and survival of RORγt+ ILCs

city: Seattle

first: Akhilesh

last: Singh

state: WA

department: Center for Immunity and Immunotherapies

institute: Seattle Childrens Research Institute

summary: Retinoic-acid receptor-related orphan receptor-γt-positive (RORγt+) innate lymphoid cells (ILCs) produce interleukin (IL)-22 and IL-17, which are critical for protective immunity against enteric pathogens. The molecular mechanism underlying the development and survival of RORγt+ ILCs is not thoroughly understood. Here we show that Dedicator of cytokinesis 8 (DOCK8), a scaffolding protein involved in cytoskeletal rearrangement and cell migration, is essential for the protective immunity against Citrobacter rodentium. A comparative RNA sequencing-based analysis reveals an impaired induction of antimicrobial peptides in the colon of DOCK8-deficient mice, which correlates with high susceptibility to infection and a very low number of IL-22-producing RORγt+ ILCs in their GI tract. Furthermore, DOCK8-deficient RORγt+ ILCs are less responsive to IL-7 mediated signaling, more prone to apoptosis and produce less IL-22 due to a defect in IL-23-mediated STAT3 phosphorylation. Our studies reveal an unsuspected role of DOCK8 for the function, generation and survival of RORγt+ ILCs.

overall_design: Control and DOCK8 KO mice were infected with 2X109 CFU of Citrobacter rodentium and day 8 post infection mice were sacrificed and their colons were harvested (n=5) . Total RNA was purified from the infected colons with RNeasy mini kit (Qiagen). RNA sequencing was performed (pooled RNA sample from five mice in each group) at Genomic Core Facility Southwestern Medical Center, University of Texas.

first: Mohamed

last: Oukka


title: Helminth Infection Reactivates Latent γ-herpesvirus Via Cytokine Competition at a Viral Promoter

first: Maxim

middle: N.

last: Artyomov

state: MO

address: 660 S. Euclid Avenue, Campus Box 8118

department: Immunology&Pathology

institute: Washington University in St.Louis

summary: Mammals are co-infected by multiple pathogens that interact through unknown mechanisms. We found that helminth infection, characterized by the induction of the cytokine interleukin-4 (IL-4) and the activation of the transcription factor Stat6, reactivated murine gammaherpesvirus infection in vivo. IL-4 promoted viral replication and blocked the antiviral effects of interferon-g (IFNg) by inducing Stat6 binding to the promoter for an important viral transcriptional transactivator. IL-4 also reactivated human Kaposi's sarcoma associated herpesvirus from latency in cultured cells. Exogenous IL-4 plus blockade of IFNg reactivated latent murine gammaherpesvirus infection in vivo, suggesting a ‘two-signal’ model for viral reactivation. Thus chronic herpesvirus infection, a component of the mammalian virome, is regulated by the counterpoised actions of multiple cytokines on viral promoters that have evolved to sense host immune status.

overall_design: All of the RNA from virus-pos cells and 50 ng of RNA from virus-neg cells was prepared for RNA-seq using ScriptSeq v2 RNA-seq library preparation kit (Epicentre). Index Primers (Epicentre) were added and samples underwent Duplex-Specific thermostable nuclease (DSN) (Evrogen) treatment to remove ribosomal RNA. Samples were pooled and sequenced on HiSeq.


title: Global identification of growth-related long non-coding RNAs in breast muscle of chicken

status: Public on Jun 24 2014

city: Guangzhou

first: Xuenong

last: Zhu

state: Guangdong

address: Wushan street 483

department: College of Animal Science

institute: South China Agricultural University

summary: Long non-coding RNAs (lncRNAs) can have potential roles in development of tissues and organs. We selected breast muscle of fast-growing White Recessive Rock chicken (WRR) and slow-growing Xing Hua chicken (XH) to identify lncRNA transcripts by LncRNA-Seq. This study identified 21,993 novel lncRNAs. Among 7,339 differentially expressed lncRNAs, 723 up-regulated and 6,616 down-regulated lncRNAs were found in WRR compared with XH. Of them, five novel lncRNA were antisense transcripts for growth-related genes CACNA1D (unigene 14689_all), IL4I1 (unigene 15355_all), LEF-1 (unigene 19525_all) and FABP1 (unigenes 17536_all and 17537_all) respectively. Meanwhile, 12 other novel lncRNAs were found in the intron or downstream of some known growth-related genes (IGF1, IGF2BP2, IGF2BP3, CACNA1D, IL4I1, LEF-1 and FABP1). In addition, 4,043 SSRs and 200,049 SNPs were identified. Our data revealed the global lncRNA expression pattern in muscle tissue, and contributed a useful genomic resource towards studying the effects of lncRNAs in regulating chicken growth.

overall_design: Two RNA pool for WRR and XH strains

first: Haiping

last: Xu

first: Qinghua

last: Nie

first: Xiquan

last: Zhang


title: Mapping of Rec12 oligonucleotides in Schizosaccharomyces pombe

city: New York

first: Bic

last: MSKCC

state: NY

address: 1275 York Ave.

institute: Memorial SLoan-Kettering Cancer Center

summary: Meiotic recombination facilitates accurate pairing and faithful segregation of homologous chromosomes by forming physical connections (crossovers) between homologs. Developmentally programmed DNA double-strand breaks (DSBs) generated by Spo11 protein (Rec12 in fission yeast) initiate meiotic recombination. Until recently, attempts to address the basis of the highly non-random distribution of DSBs on a genome-wide scale have been limited to 0.1–1 kb resolution of DSB position. We have assessed individual DSB events across the Schizosaccharomyces pombe genome at near-nucleotide resolution by deep-sequencing the short oligonucleotides connected to Rec12 following DNA cleavage. The single oligonucleotide size-class generated by Rec12 allowed us to effectively analyze all break events. Our high-resolution DSB map shows that the influence of underlying nucleotide sequence and chromosomal architecture differs in multiple ways from that in budding yeast. Rec12 action is not strongly restricted to nucleosome-depleted regions but is nevertheless spatially biased with respect to chromatin structure. Furthermore, we find strong evidence across the genome for differential DSB repair previously predicted to account for crossover invariance (constant cM/kb in spite of DSB hotspots). Our genome-wide analyses demonstrate evolutionarily fluid factors contributing to crossover initiation and its regulation.

overall_design: Three samples total: one sample sequenced by 454 and two technical replicates (independent adaptor ligations from material purified from one culture) sequenced by SOLiD

first: Kyle

last: Fowler

first: Mariko

last: Sasaki

first: Neta

last: Milman

first: Scott

last: Keeney

first: Gerald

last: Smith


title: Small RNAs expression of three hepatocellular carcinoma cell lines with different organ-tropism.

city: Shanghai

first: Wei zhong

last: Wu

address: Fenglin Rd 180, Xuhui district

department: Zhongshan Hospital

institute: Fudan University

summary: In this study we investigated the small RNAs expression profiling of three HCC cell lines with different organ-tropism.The parent cell line has low mestastasis ability in nude mice model. Subclone 1 has higher metastasis ability specific to lung, and subclone 2 has dual metastasis ability to lung and celiac lymph node. We aimed to explore differentially expressed small RNAs involved in process of HCC metastasis and organ-specific metastasis, and identify their biological functions.

overall_design: Examination of  different small RNAs expression among the 3 cell types.

first: W

middle: Z

first: J

middle: L

last: Wan

first: Z

middle: H

last: Tao


title: Genome-wide analysis of next generation sequencing for Lsh+/+ and Lsh-/- mouse embryonic fibroblasts

city: Frederick

last: Durum

address: Building 560, Room 31-71

department: CCR

institute: National Cancer Institute

summary: To examine the relationship of reduced CG methylation and gene expression in Lsh KO MEFs, we computed mean CG methylation levels at promoter regions of protein-coding genes. About 60% of TSS regions of protein-coding genes display a difference of CG methylation values greater than 0.3 (WT CG methylation minus KO CG methylation) indicating that Lsh deletion has widespread effects at promoter regions. RNA-seq analysis detects similar transcript steady state levels in WT and KO samples.  To determine the relationship of Pol II binding and CG methylation reduction in KO MEFs, Pol II Chip-seq was performed. Protein coding genes were ranked according their CG methylation differences between WT MEFs and KO MEFs. The greatest loss of CG methylation is found at promoter with low CG density. Pol II association is inversely related to the number of CpG sites within promoter regions. KO MEFs show less Pol II association at CG rich promoter regions. However, RNA-seq reads are indistinguishable comparing WT and KO samples, suggesting similar transcriptional efficiency in the absence of Lsh. To explore other molecular mechanisms that may preserve low transcription activity or repression at CG hypomethylated promoter regions, we examined H3K27me3 and H3K4me3 modifications by ChIP-seq. Genome wide computation of histone modifications at 5kb tiles shows no increase of H3K27me3 level in KO MEFs. When we ranked 5kb tiles based on CG methylation differences between WT and KO, we observed alterations in H3K27me3 distribution, while H3K4me3 modifications are unremarkable. Regions with moderate CG methylation reduction exhibit concomitant decreases in H3K27me3.

overall_design: mRNA profiles and Genome-wide maps of H3K27me3, H3K4me3 and Pol II in wildtype (WT) and Lsh KO primary MEFs.

first: Weishi

last: Yu

first: Yixing

last: Han

first: Jianke

last: Ren

first: David

last: Landsman

first: Carl

last: McIntosh

first: Ryan

last: Lister

first: Iris

first: Eunice

last: Lee

first: Victorino

last: Briones

first: Minoru

last: Terashima

first: Robert

last: Leighty


title: A specific missense mutation in GTF2I occurs at high frequency in thymic epithelial tumors

city: Washington

first: Krithika

last: Bhuvaneshwar

state: DC

address: 2115 Wisconsin Ave NW

institute: Georgetown University

summary: Next generation sequencing of 28 thymic epithelial tumors (TETs) revealed a high frequency of GTF2I missense mutation (chr7:74146970T/A) in A thymomas, a  relatively indolent subtype. The GTF2I mutation was confirmed in 82% of A and 74% of AB thymomas in a series of 274 TETs but was rare in aggressive subtypes, where recurrent mutations of known cancer genes were identified. Therefore, GTF2I mutation correlated with a better survival. GTF2I Beta and Delta isoforms were expressed in TETs and both mutant isoforms were able to stimulate cell proliferation in vitro. Thymic carcinomas presented a higher number of mutations than thymomas (average 43.5 and 18.4, respectively). Recurrent mutations of known cancer genes, including TP53, CYLD, CDKN2A, BAP1 and PBRM1 were identified in thymic carcinomas. These findings will complement the diagnostic work up of these rare tumors, and also help the development of a molecular classification, and assessment of prognosis and treatment strategies.

overall_design: Tumor samples of 286 patients were collected from 4 different institutions: National Cancer Institute (Bethesda MD), Pisa University Hospital (Pisa, Italy), Padua University Hospital (Padua, Italy) and IRCCS Istituto Clinico Humanitas (Rozzano, Italy).

first: Giuseppe

last: Giaccone

first: Iacopo

last: Petrini


title: RNA-seq of control and talpid2 facial prominences

city: Evanston

first: Carole

last: LaBonne

state: IL

address: 2220 Campus Drive

department: Molecular Biosciences

institute: Northwestern University

summary: We report the RNA profiles of both control and talpid2 frontonasal, maxillary, and mandibular prominences of the chick face at Hamburger and Hamilton (HH) stage 25. For more details please see: "The cellular and molecular etiology of the craniofacial defects in the avian ciliopathic mutant, talpid2."

overall_design: Facial prominences (frontonasal, maxillary, and mandibular) from 8 control and 8 talpid2 HH 25 embryos were harvested, pooled, and RNA-seq was preformed on samples.

first: Ching-Fang

last: Chang

first: Elizabeth

last: Schock

last: O'Hare

first: Jerry

last: Dodgson

first: Hans

last: Cheng

first: William

middle: M

last: Muir

first: Richard

middle: E

last: Edelman

first: Mary

last: Delany

first: Samantha

last: Brugmann


title: The Vasa homolog RDE-12 engages target mRNA and a secondary-Argonaute WAGO-1 to promote RNAi in C. elegans

city: Worcester

first: Craig

last: Mello

address: 368 Plantatoin Street, Suite AS5-2047

department: Program in Molecular Medicine

institute: University of Massachusetts Medical School

summary: Argonaute proteins (AGOs) are key nuclease effectors of RNA interference (RNAi) [1]. Although purified AGOs can mediate a single round of target-RNA cleavage in vitro, accessory factors are required for siRNA loading and to achieve multiple-target turnover [2, 3]. To identify AGO co-factors we immunoprecipitated the C. elegans AGO WAGO-1, which engages amplified small RNAs during RNAi [4]. These studies identified a robust association between WAGO-1 and a conserved Vasa ATPase-related protein RDE-12. rde-12 mutants are deficient in RNAi including viral suppression, and fail to produce amplified secondary siRNAs and certain endogenous siRNAs (endo-siRNAs). RDE-12 co-localizes with WAGO-1 in germline P-granules and to peri-nuclear cytoplasmic foci in somatic cells. These findings and our genetic studies suggest that (i) RDE-12 is first recruited to target mRNAs by upstream AGOs (RDE-1 and ERGO-1) where it promotes small-RNA amplification and/or WAGO-1 loading, and that (ii) downstream of these events, RDE-12 forms an RNase-resistant (target mRNA-independent) complex with WAGO-1 that may scan for additional target mRNAs.

overall_design: Examine small RNA population changes in rde-12 mutants

first: Masaki

last: Shirayama

last: Stanney

first: Weifeng

last: Gu

first: Meetu

last: Seth


title: Chromatinized PKC-q directly regulates inducible genes in epithelial to mesenchymal transition and breast cancer stem cells

status: Public on Jun 22 2014

city: Bruce

first: Kristine

last: Hardy

state: ACT

address: University of Canberra

institute: University of Canberra

summary: Epithelial to mesenchymal transition (EMT) is activated during cancer invasion and metastasis, enriches for cancer stem cells (CSCs), and contributes to therapeutic resistance and disease recurrence. Signal transduction kinases play a pivotal role as chromatin-anchored proteins in eukaryotes. Here we report for the first time that protein kinase C-theta (PKC-q) regulates EMT by acting as a critical chromatin-anchored switch for inducible genes via TGF-β and the key inflammatory regulatory protein, NFkB. Chromatinized PKC-q exists as an active transcription complex and is required to establish a permissive chromatin state at signature EMT genes. Genome-wide analysis identifies a unique cohort of inducible PKC-q-sensitive genes that are directly tethered to PKC-q in the mesenchymal state. Collectively, we show that crosstalk between signaling kinases and chromatin is critical for eliciting inducible transcriptional programs that drive mesenchymal differentiation and CSC formation, providing novel mechanisms to target using epigenetic therapy in breast cancer.

overall_design: 2 biological samples were analysed, Immunoprecipitated and total input samples were obtained from each biological treatment.  2 Technical replicates were performed (samples from the sample lib prep were run on two different lanes).

first: Anjum

last: Zafar

first: Sudha

last: Rao


title: Regulation of inducible genes in epithelial to mesenchymal transition by chromatinized PKC-theta

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

overall_design: Refer to individual Series


title: PDX1 Binding in human embryonic stem cell derived pancreatic progenitor cells

status: Public on Jun 21 2014

city: Singapore

first: Shawn

last: Hoon

address: 61 Biopolis Street

country: Singapore

institute: Agency for Science Technology and Research


title: PDX1 Binding in human embryonic stem cell derived pancreatic progenitor cells [ChIP-seq]

summary: PDX1 binds and regulates numerous genes involved in human pancreatic development but also binds hepatic genes

overall_design: Examination of PDX1 regulated genes in pancreatic derived cell types

first: Adrian

middle: K

last: Teo


title: Long-range chromatin interactions in mouse embryonic stem cells identified by ChIA-PET [ChIA-PET]

last: Young

state: MA

address: 9 Cambridge Center

institute: Whitehead Institute for Biomedical Research

summary: The pluripotent state of embryonic stem cells (ESCs) is produced by active transcription of cell identity genes and repression of genes encoding lineage-specifying developmental regulators. Here we use large ESC cohesin ChIA-PET datasets and other genomic data to identify the local chromosomal structures at both active and repressed genes across the genome. The results show that super-enhancer driven cell identity genes generally occur within large loops that are connected through CTCF-CTCF interaction sites occupied by cohesin.

overall_design: Smc1 ChIA-PET data from wild type murine embryonic stem cells V6.5 were generated by deep sequencing using Illumina Hi-Seq 2000.

first: Jill

last: Dowen

first: Zi

last: Fan

first: Brian

last: Abraham

first: Lyndon

last: Zhang

first: Denes

last: Hnisz

first: Tong

middle: I

last: Lee

first: Gang

last: Ren

first: Keji

last: Zhao


title: Transcriptome profiling of tomato fruits of a FRUITFULL-suppressed line and rin mutant by next generation RNA sequencing

status: Public on Jun 23 2014

city: Tsukuba

first: Yasuhiro

last: Ito

state: Ibaraki

address: 2-1-12 Kannondai

department: Food Biotechnology Division

country: Japan

institute: National Food Research Institute

summary: The tomato MADS-box FRUITFULL (FUL) homologs, FUL1 and FUL2, interact with the main ripening regulator RIPENING INHIBITOR (RIN). To clarify their role in fruit ripening, we generated FUL1/FUL2-suppressed transgenic lines by RNAi. We found that five transgenic lines bearing fruits that did not ripen normally: lycopene accumulation and increase of ethylene production were severely inhibited. We then performed next generation RNA sequencing (RNA-Seq) analysis of the fruits of a FUL1/FUL2-suppressed line (TF18) with those of the wild type (Ailsa Craig cultivar; AC) and rin mutant. The comparison of RNA-Seq data among them indicated that FUL1/FUL2-suppression significantly affected the expression of a larger portion of ripening-induced and -repressed genes than the rin mutation did. Moreover, the effect of FUL1/FUL2-suppression was observed not only in the fruits harvested at the wild type ripening age [45 days after pollination (DAP)] but also in those at the pre-ripening age (35 DAP). This suggests that the FUL homologs play an essential role in the regulation of fruit development and ripening, the role which covers a wider range of biological processes than RIN does. Differentially expressed genes (DEGs) between the wild type and TF18 fruits included known ripening-related genes such as ACS2 and ACS4 involved in ethylene production and PSY1 in carotenoid biosynthesis, consistent with the phenotype of TF18 fruits described above. The DEGs also included many direct RIN target genes, which supports the hypothesis that the FUL homologs regulate fruit ripening in a form of MADS-box complex with RIN.

overall_design: mRNA profiles of wild type (Ailsa Craig cultivar), rin mutant and FUL1/FUL2-suppressed tomato fruits harvested at 35DAP and 45 DAP were generated by next generation sequencing, in triplicate, using Illumina Hiseq2000.

last: Fujisawa

first: Yoko

last: Shima

first: Toshitsugu

last: Nakano


title: MicroRNA expression profile in mouse cone photoreceptors at P60 [miRNA-seq]

status: Public on Jun 20 2014

city: Basel

first: Tim

last: Roloff

address: Maulbeerstrasse 66

department: Functional Genomics

country: Switzerland

institute: FMI

summary: The outer segments of cones serve as light detectors for daylight color vision, and their dysfunction leads to human blindness conditions. We show that the cone-specific disruption of DiGeorge Syndrome Critical Region Gene 8 (DGCR8) in adult mice led to the loss of miRNAs and the loss of outer segments, resulting in photoreceptors with significantly reduced light responses. Using next-generation sequencing of RNA from isolated wild type P60 cones, we determine the most highly expressed miRNAs as candidates for controlling outer segment maintenance. The expression pattern of miRNAs was highly uneven, with a single miRNA, miR-182, representing 64% of all miRNA expressed in cones. Re-expression of miR-182 and miR-183 (third most abundant miRNA) prevented outer segment loss. These miRNAs were also necessary and sufficient for the formation of inner segments, connecting cilia and short outer segments, as well as light responses in stem-cell-derived retinal cultures. Our results show that miR-182- and miR-183-regulated pathways are necessary for cone outer segment maintenance in vivo and functional outer segment formation in vitro.

overall_design: microRNA profile in cone photoreceptors from P60 D4-cre/Ai9 tdTomato mice representing wild type control.

first: Jacek

last: Krol

first: Volker

last: Busskamp

first: Botond

last: Roska

first: Witold

last: Filipowicz


title: MiRNAs 182 and 183 are necessary to maintain adult cone photoreceptor outer segments and visual function


title: Expression data of wild type and C-DGCR8 KO cones at different timepoints [RNA-seq]

summary: We have analyzed gene expression in cone photoreceptors isolated from wild type and C-DGCR8 (DiGeorge Syndrome Critical Region Gene 8) KO mice at five different time points to get a mechanistic inside into the altered molecular pathways after microRNAs depletion.

overall_design: Cones were isolated by FACS from wild type and C-DGCR8 KO mice expressing Cre recombinase postnatally specifically in cones (D4-Cre) and bearing conditional null Dgcr8 allele. Experiments were performed in duplicates from different time points (P30, P40, P50, P60, and P90). Total RNA was extracted and next-generation RNA sequencing was performed.


title: RNA-seq analysis of the effect of kanamycin and the ABC transporter AtWBC19 on Arabidopsis thaliana seedlings reveals changes in metal content

city: Atlanta

first: Mentewab

last: Ayalew

state: GA

address: 350 Spelman Lane

department: Biology Department

institute: Spelman College

summary: Plants are exposed to antibiotics produced by soil microorganisms but little is known about their responses at the transcriptional level. Likewise, few endogenous mechanisms of antibiotic resistance have been reported. The Arabidopsis thaliana ATP Binding Cassette (ABC) transporter AtWBC19 (ABCG19) is known to confer kanamycin resistance, but the exact mechanism of resistance is not well understood. Here we examined the transcriptomes of control seedlings and wbc19 mutant seedlings using RNA-seq analysis. Exposure to kanamycin indicated changes in the organization of the photosynthetic apparatus, metabolic fluxes and metal uptake. Elemental analysis showed a 60% and 80% reduction of iron uptake in control and wbc19 mutant seedlings respectively, upon exposure to kanamycin. The drop in iron content was accompanied by the upregulation of the gene encoding for FERRIC REDUCTION OXIDASE 6 (FRO6) in mutant seedlings but not by the differential expression of other transport genes known to be induced by iron deficiency. In addition, wbc19 mutants displayed a distinct expression profile in the absence of kanamycin. Most notably, the expression of several zinc ion binding proteins, including ZINC TRANSPORTER 1 PRECURSOR (ZIP1), was increased, suggesting abnormal zinc uptake. Elemental analysis confirmed a 50% decrease of zinc content in wbc19 mutants. Thus, the antibiotic resistance gene WBC19 appears to also have a role in zinc uptake.

overall_design: Examination of transcriptome in control and wbc19 mutant seedlings with or without exposure to kanamycin.

first: Kinnari

last: Matheson

first: Morayo

last: Adebiyi

first: Shanice

last: Robinson

first: Brianna

last: Elston


title: EXPRSS: an Illumina based high-throughput expression-profiling method to reveal transcriptional dynamics - I

city: Norwich

first: Ghanasyam

last: Rallapalli

address: Norwich Research Park

institute: The Sainsbury Laboratory

summary:
Background
Next Generation Sequencing technologies have facilitated differential gene expression analysis through RNA-seq and Tag-seq methods. RNA-seq has biases associated with transcript lengths, lacks uniform coverage of regions in mRNA and requires 10–20 times more reads than a typical Tag-seq. Most existing Tag-seq methods either have biases or not high throughput due to use of restriction enzymes or enzymatic manipulation of 5’ ends of mRNA or use of RNA ligations.

Results
We have developed EXpression Profiling through Randomly Sheared cDNA tag Sequencing (EXPRSS) that employs acoustic waves to randomly shear cDNA and generate sequence tags at a relatively defined position (~150-200 bp) from the 3′ end of each mRNA. Implementation of the method was verified through comparative analysis of expression data generated from EXPRSS, NlaIII-DGE and Affymetrix microarray and through qPCR quantification of selected genes. EXPRSS is a strand specific and restriction enzyme independent tag sequencing method that does not require cDNA length-based data transformations. EXPRSS is highly reproducible, is high-throughput and it also reveals alternative polyadenylation and polyadenylated antisense transcripts. It is cost-effective using barcoded multiplexing, avoids the biases of existing SAGE and derivative methods and can reveal polyadenylation position from paired-end sequencing.

Conclusions
EXPRSS Tag-seq provides sensitive and reliable gene expression data and enables high-throughput expression profiling with relatively simple downstream analysis.

overall_design: Five weeks old Arabidopsis (Col-0) leaf discs treated with water or flg22 for 60min; mRNA profiles were generated by deep sequencing on Illumina GAIIx using EXPRSS and NlaIII-DGE protocols, in quadruplicate.

first: Jonathan

last: Jones


title: RNA-seq analysis reveals endogenous aryl hydrocarbon receptor regulation is highly associated with eicosanoid synthesis and tumor necrosis factor activity in MCF-7 cancer cells

city: Huntington

first: James

last: Denvir

state: WV

address: One John Marshall Drive

department: Biochemistry and Microbiology

institute: Marshall University School of Medicine

summary:
Background: The Aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor that is activated by xenobiotic chemicals that function as AHR ligands.  The response to xenobiotic AHR ligands is toxicity and the induction of drug metabolizing enzymes. The impact of AHR knockdown on gene expression and pathways in human breast cancer cells in absence of xenobiotic AHR ligands has not been investigated on a genome-wide scale.
Methods: MCF-7 cells were used as a model of human breast cancer.  The AHR was silenced with short interfering RNA against AHR (siAHR).  RNA-sequencing coupled with Ingenuity pathway analysis (IPA) was used to determine the impact of AHR knockdown on gene expression and pathways in the absence of xenobiotic AHR ligands.  Western blot analysis and recombinant tumor necrosis factor (TNF) was used to investigate the impact of AHR knockdown on TNF induction of MNSOD expression.
Results: We found that the AHR is transcriptionally active in MCF-7 breast cancer cells in the absence of xenobiotic AHR ligands.  In total, the expression of 634 genes was significantly changed in AHR knockdown cells compared to controls at a false discovery rate of < 10%. The analysis confirmed that drug metabolizing enzymes were AHR targets; however, we found that AHR also promoted the expression of genes that were not directly related to the metabolism of xenobiotics, such as those involved in lipid and eicosanoid synthesis.  Gene pathway analysis of the AHR regulated gene dataset predicted TNF activity to be reduced in AHR knockdown cells.  Our finding that AHR knockdown inhibited TNF-stimulated increases in MNSOD expression confirmed this IPA prediction.
Conclusions: This is the first gene expression profiling study of AHR knockdown breast cancer cells.  Several known and novel AHR targets were identified.  The results suggest that endogenous AHR regulation impacts eicosanoid synthesis by regulating gene expression.  The IPA prediction of reduced TNF activity in AHR knockdown cells was confirmed by showing that TNF-induced increases in MNSOD expression was inhibited in AHR knockdown cells.   The requirement of AHR for MNSOD activation is novel and provides a new link by which AHR may impact reactive oxidative species (ROS) signaling.

overall_design: Expression profiles by mRNA sequencing were generated for human MCF-7 cells transfected with cRNAi (6 replicates) or AHR-siRNA (5 replicates) for 36hr. Sequencing was performed on an Illumina HiSeq 1000 using a 2x100 base paired-end strategy.

first: Travis

middle: B

last: Salisbury

first: Justin

last: Tomblin

first: Donald

last: Primerano

first: Goran

last: Boskovic

first: Jun

last: Fan

first: Jackie

last: Fletcher

first: Nalini

last: Santanam

first: Estil

last: Hurn

first: Gary

last: Morris


title: EXPRSS: an Illumina based high-throughput expression-profiling method to reveal transcriptional dynamics - II

overall_design: Five weeks old Arabidopsis (Col-0, ein2-5, jar1-1 and npr1-1) leaf discs treated with water or flg22 and a time course of samples were collected; mRNA profiles were generated by deep sequencing on Illumina GAIIx using EXPRSS protocols.

first: Eric

last: Kemen

first: Daniel

last: MacLean

first: Cécile

last: Segonzac

first: Graham

last: Etherington

first: Kee

last: Sohn


title: Haploinsufficiency of an RB-E2F1-Condensin II complex causes replication stress and contributes to mesenchymal cancers

status: Public on Jun 19 2014

city: London

first: Fred

last: Dick

state: ON

address: Cancer Research Labs 790 Commissioners Road East

country: Canada

institute: Western University


title: EXPRSS: an Illumina based high-throughput expression-profiling method to reveal transcriptional dynamics

first: Ghanasyam

last: Rallapalli


title: High-throughput sequencing of genome-wide translocations from stimulated wild-type or ATM-deficient B cells

status: Public on Jun 18 2014

city: Boston

first: Jiazhi

last: Hu

address: 300 Longwood Ave

department: PCMM

institute: Children's Hospital Boston

summary: High-throughput genome-wide translocation sequencing (HTGTS) is a robust approach to identify genome-wide translocation junctions. We performed HTGTS to study the fate of introduced c-myc DSBs in mouse splenic B cells activated for activation cytidine deaminase (AID)-dependent class switch recombination (CSR).  We found frequent translocations of c-myc DSBs to AID-initiated DSBs in IgH switch regions in wild-type (WT) and ATM-deficient B cells.  However, c-myc also translocated frequently to newly generated DSBs within a 35-megabase region downstream of IgH in ATM-deficient, but not WT, CSR-activated B cells.  Moreover, we found such DSBs and translocations in activated B cells that did not express AID or undergo CSR.  These findings indicate that ATM deficiency leads to formation of chromosome 12 dicentrics via RAG-initiated IgH DSBs in progenitor B cells and that these dicentrics can be propagated developmentally into mature B cells where they generate new DSBs downstream of IgH via breakage-fusion-bridge cycles.

overall_design: Preparation of libraries from WT or ATM-deficient activated by a-CD40/IL4 or RP105.

first: Frederick

middle: W

last: Alt

first: Robin

last: Meyers


title: Inducible PAX5 expression in a human B lymphoblastic leukemia cell line

city: Parkville

first: Gordon

last: Smyth

state: Vic

address: 1G Royal Pde

department: Bioinformatics

institute: Walter and Eliza Hall Institute of Medical Research

summary: Hypomorphic mutations of the transcription factor PAX5 occur in one third of B-progenitor acute lymphoblastic leukemias (B-ALLs). To identify PAX5-regulated genes in B-ALL, here we employ inducible expression of PAX5 in a human B-ALL cell line (REH) that harbors a loss-of-function mutation in PAX5. In this model, inducing PAX5 expression is associated with competitive disadvantage.

overall_design: Comparison of REH cell lines with Dox-inducible expression of PAX5-IRES-GFP, or control GFP alone. GFP positive cells were isolated by FACS.

first: Julian

last: Jude

first: Johannes

last: Zuber

first: Yifang


title: Haploinsufficiency of an RB-E2F1-Condensin II complex causes replication stress and contributes to mesenchymal cancers [γH2AX localization]

summary: Genome instability is a characteristic of malignant cells, however, evidence for its contribution to tumorigenesis has been enigmatic. In this study we demonstrate that a complex containing the retinoblastoma protein, E2F1, and Condensin II localizes to major satellite repeats at pericentromeres. In the absence of this complex, this genomic region fails to properly replicate and H2AX phosphorylation at pericentromeric repeats is increased. Our data indicates that these lesions contribute to defective chromosome segregation in the ensuing mitosis. Surprisingly, loss of even one copy of the retinoblastoma gene was sufficient to reduce recruitment of Condensin II to pericentromeres and cause this phenotype. Furthermore, we determined that human RB1 mutation status in cancers of mesenchymal origin correlated with copy number variation, chromosomal gains and losses, as well as chromothriptic rearrangements. Importantly, the magnitude of these chromosomal abnormalities was indistinguishable between RB1+/- and RB1-/- genotypes. Lastly, using gene-targeted mice we determined that mutation of just one copy of the murine Rb1 gene causes sarcomas and lymphomas with increased chromosome copy number variation. Our study connects replication stress with a number of common chromosomal abnormalities in cancer through a dosage sensitive complex present at pericentromeric repeats.

overall_design: γH2AX localization was studied in four different genotypes: Rb+/+, RbΔL/+, RbΔL/ΔL, and Rb-/-. As a control, input from the ChIP experiment was also sequenced. The γH2AX antibody used was supplied by Millipore (catalog number 05-636, and lot number 2116620).

first: Courtney

last: Coschi

first: Aren

last: Marshall


title: High temporal resolution of mRNA expression patterns during the early macrophage response to LPS [RNA_Seq]

city: Mannheim

first: Johanna

middle: Daniela

last: Schott

address: Franz-Volhard-Straße 6

department: Mannheim Institute for Innate Immunoscience

institute: Heidelberg University

summary: When macrophages encounter pathogens, they transiently induce an orchestrated cascade of pro- and anti-inflammatory genes. To obtain a precise picture of transcriptome-wide mRNA expression patterns, we performed RNA-Seq of total RNA at a high temporal resolution during the first two hours of macrophage activation. We systematically analyzed the contribution of translational regulation to the early phase of macrophage activation. While the expression of most cytokines is pre-dominanatly regulated by changes in mRNA levels, de-repression of translation was found to permit expression of many feedback inhibitors of the inflammatory response.

overall_design: Expression profiles of LPS-treated Raw264.7 cells (0, 15, 30, 45, 60, 75, 90 and 120 min after stimulation) were generated by deep sequencing using Illumina HiSeq 2000.

first: Sonja

last: Reitter

first: Janine

last: Philipp

first: Heiner

last: Schaefer

first: Georg

last: Stoecklin


title: Translational regulation of specific mRNAs controls feedback inhibition and survival during macrophage activation


title: Gene Expression Analysis of Cancer-Associated Fibroblast (CAF) compared to Normal Fibroblast (NF) [RNA-seq]

city: Houston

first: Valerie

last: LeBleu

state: TX

address: 1881 East Road

institute: MDACC

summary: Analysis of differentially expressed genes in CAF associated with PDAC vs NF. Genetically engineered mice with spontaneous pancreas cancer were generated. Their genotype is Ptfa-cre/+:LSL KrasG12D/+;Tgfrb2flox/flox. Cancer associated fibroblasts were expanded in vitro from the tumors of these mice (CAF). Normal fibroblasts (NF) were also expanded from normal pancreas of mice. The experiement consists in comparing the expression profile of CAF vs. NF.

overall_design: Expanded cultures of fibroblasts in vitro from pancreas tumor and normal pancreas tissue were lyzed in TRIzol and RNA extracted from them.

first: Raghu

last: Kalluri


title: An Integrated Approach to Reconstructing Genome-scale Transcriptional Regulatory Networks

city: Madison

first: Timothy

middle: J

last: Donohue

state: WI

address: 1552 University Avenue

department: Bacteriology

institute: University of Wisconsin - Madison

sample_organism:
Cereibacter sphaeroides
Cereibacter sphaeroides 2.4.1


title: Expression profiling of mouse bone marrow pre-B cells

summary: Triplicate RNA-seq expression analysis of bone marrow pre-B cells isolated from mice, to demonstrate repertoire at the IgH locus

first: Grace

last: Liu

first: Ross

last: Dickins


title: Haploinsufficiency of an RB-E2F1-Condensin II complex causes replication stress and contributes to mesenchymal cancers [CAP-D3 localization]

first: Fred

last: Dick

overall_design: CAP-D3 localization was studied in two different genotypes: Rb+/+ and RbΔL/ΔL. As a control, input from the ChIP experiment was also sequenced. The CAP-D3 antibody used was raised against a GST fusion protein containing amino acids 1243-1506 (Coschi et al., 2010).

first: Frederick


title: EXPRSS: an Illumina based high-throughput expression-profiling method to reveal transcriptional dynamics - III

first: Ghanasyam

last: Rallapalli

overall_design: Four-week-old Arabidopsis (No-0 and slh1) plants were grown at 28°C and transferred to a 21°C growth chamber. Time course samples were collected after the temperature shift; mRNA profiles were generated by deep sequencing on Illumina MiSeq using EXPRSS protocol.

first: Eric

last: Kemen

first: Daniel

last: MacLean

first: Alexandre

last: Robert-Seilaniantz

first: Cécile

last: Segonzac

first: Graham

last: Etherington

first: Kee

last: Sohn

first: Jonathan

last: Jones


title: Correcting Direct Effects of Ethanol on Translation and Transcription Machinery Confers Ethanol Tolerance in Bacteria [Terminator_Readthrough_RNAseq]

first: Rembrandt

last: Haft

address: 1550 Linden Dr. Room 5445

department: Great Lakes Bioenergy Research Center

institute: University of Wisconsin

summary: The molecular mechanisms of ethanol toxicity and tolerance in bacteria, while important for biotechnology and bioenergy applications, remain incompletely understood. Genetic studies have identified potential cellular targets for ethanol and revealed multiple mechanisms of tolerance, but it remains difficult to separate direct and indirect effects of ethanol. We used adaptive evolution to generate spontaneous ethanol-tolerant strains of Escherichia coli, then characterized the mechanisms of toxicity and resistance associated with select mutations. Evolved alleles of metJ, rho, and rpsQ were sufficient to recapitulate much of the observed ethanol tolerance, implicating translation and transcription as key processes affected by ethanol. We found that ethanol induces mistranslation errors during protein synthesis, and that the evolved rpsQ allele protects cells by rendering the ribosome hyper-accurate. Ribosome profiling and RNAseq analyses of the ethanol-tolerant strain versus the wild type established that ethanol negatively affects transcriptional and translational processivity. Ethanol-stressed cells exhibited ribosomal stalling at internal AUG codons, which may be ameliorated by the adaptive inactivation of the MetJ repressor of methionine biosynthesis genes. Ethanol also caused aberrant intragenic transcription termination for mRNAs with low ribosome density, which was reduced in a strain with the adaptive rho mutation. Furthermore, ethanol inhibited transcript elongation by RNA polymerase in vitro. We propose that ethanol-induced inhibition and uncoupling of mRNA and protein synthesis are major contributors to ethanol toxicity in E. coli, and that adaptive mutations in metJ, rho, and rpsQ protect central dogma processes in the presence of ethanol.

overall_design: RNA-seq comparison of wild-type and mutant strains to assess readthrough of Rho-dependent transcriptional terminators

first: Jason

last: Peters

first: Robert

last: Landick


title: DNA Methylation Identifies Epigenetic Variation across Embryo and Endosperm in Maize (Zea may)

status: Public on Jun 17 2014

city: Ji'nan

first: Xingjun

last: Wang

state: Shandong

address: 202#, Gongye Bei Road

department: Biotechnology Research Center

institute: Shandong Academy of Agricultural Sciences

summary: Epigenetic modification plays important roles in plant and animal development. DNA methylation can impact the transposable element (TE) silencing, gene imprinting and regulate gene expression.Through a genome-wide analysis, DNA methylation peaks were respectively characterized and mapped in maize embryo and endosperm genome. Distinct methylation level across maize embryo and endosperm was observed. The maize embryo genome contained more DNA methylation peaks than endosperm. However, the endosperm chloroplast genome contained more DNA methylation peaks to compare with the embryo chloroplast genome. DNA methylation regions were characterized and mapped in genome. More CG island (CGI) shore are methylated than CGI in maize suggested that DNA methylation level is not positively correlated with CpG density. The DNA methylation occurred more frequently in the promoter sequence and transcriptional termination region (TTR) than other regions of the genes. The result showed that 99% TEs we characterized are methylated in maize embryo, but some (34.8%) of them are not methylated in endosperm. Maize embryo and endosperm exhibit distinct pattern/level of methylation. The most differentially methylated two regions between embryo and endosperm are High CpG content promoters (HCPs) and high CpG content TTRs (HCTTRs). DNA methylation peaks distinction of mitochondria and chloroplast DNA were less than the nucleus DNA. Our results indicated that DNA methylation is associated with the gene silencing or gene activation in maize endosperm and embryo. Many genes involved in embryogenesis and seed development were found differentially methylated in embryo and endosperm. We found 17 endosperm-specific expressed imprinting genes were hypomethylated in endosperm and were hypermethylated in embryo. The expression of a maize DEMETER -like (DME-like) gene and MBD101 gene (MBD4 homolog) which direct bulk genome DNA demethylation were higher in endosperm than in embryo. These two genes may be associated with the distinct methylation level across maize embryo and endosperm.The methylomes of maize embryo and endosperm was obtained by MeDIP-seq method. The global mapping of maize embryo and endosperm methylation in this study broadened our knowledge of DNA methylation patterns in maize genome, and provided useful information for future studies on maize seed development and regulation of metabolic pathways in different seed tissues.

overall_design: Examination of DNA methylated modifications in 2 maize tissues.

first: Pengfei


title: Timed regulation of Piwi-interacting RNAs expression during rat liver regeneration

city: Oslo

first: Adnan

last: Hashim

state: Oslo

address: NCMM - Forskningsparken, House 1, 3rd Floor - Gaustadalléen 21

department: NCMM (Centre for Molecular Medicine Norway)

country: Norway

institute: UiO (University of Oslo)

summary: Small noncoding (snc) RNAs represent a growing family of transcripts that regulate key cellular processes, including mRNA degradation, translational repression and transcriptional gene silencing. Among these, the PIWI-interacting RNAs (piRNAs), a major class of sncRNAs initially identified in the germline of a variety of species, are now being found to be functionally active also in somatic cells. However, whether the Piwi/piRNA pathway is associated with fundamental biological processes, such as cell cycle progression, remains elusive. Here we investigated the possibility that piRNAs are expressed in liver and modulated during regenerative proliferation of this organ. To this aim, smallRNA-Seq was applied to identify and quantitate expression of these RNAs in rat liver before, during and after the wave of cell proliferation that follows partial hepatectomy (PH). Q-PCR analysis revealed the presence in rat liver of two PIWI (PIWI-Like) subfamily members (PIWIL2/HILI and, to a much lower level, PIWIL4/HIWI2) and other components of the piRNA biogenesis pathways, suggesting that this is present and functional in hepatocytes. Indeed, ~1400 piRNAs originally identified in rat and other mammalian germline cells are expressed in adult rat liver, including 72 that show timed changes in expression during cell cycle progression. Most piRNAs  are up-regulated 24-48h after hepatectomy, a timing that corresponds to cell transition through the S phase, and return to basal levels by 168 h, when organ regeneration is completed and hepatocytes reach quiescence. These results indicate that the piRNA pathway is active in somatic cells and, more important, that it is subject to regulation during physiological processes, such as cell proliferation, when piRNAs may exert their regulatory functions on the cell genome and transcriptome.

overall_design: smallRNA-Seq was applied to identify and quantitate expression of RNAs in rat liver before and after partial hepatectomy (PH).

first: Alessandro

last: Weisz

first: Francesca

last: Rizzo


title: Gene Expression Analysis of Cancer-Associated Fibroblast (CAF) compared to Normal Fibroblast (NF)

first: Valerie

last: LeBleu

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

overall_design: Refer to individual Series


title: Genome-wide identification of CRISPR/Cas9 off-targets in human genome

first: xie

last: zhan

summary: By a robust unbiased ChIP-seq approach, we demonstrated that CRISPR/Cas9 had crRNA-specific off-target binding activities in human genome. However, most of those binding off-targets could not be efficiently cleaved both in vivo and in vitro which suggested the cleavage off-target activity of CRISPR/Cas9 in human genome is very limited. We provided a valuable tool to further investigate the molecular mechanism of CRISPR/Cas9 and to optimize its in vivo targeting

overall_design: sgRNA binding sites were identified with ChipSeq by using GFP antibody (there are 2 replicates for egfa-t1 sgRNA,emx1 sgRNA and control without sgRNA in Hek293T cells, one egfa-t1 sgRNA,emx1 sgRNA and control without sgRNA in HeLaS3 cells)

first: Yu

last: Zhang

first: Jinzhi

last: Duan

first: Guangqing

last: Lu