title: Resetting epigenetic memory by reprogramming of histone modifications in mammals

city: Beijing

first: Bingjie

last: Zhang

state: Beijing

address: 30 Shuangqing Rd

institute: Tsinghua University

summary: Polycomb group proteins and the related histone modification H3K27me3 can maintain the silencing of key developmental regulators and provide cellular memory. However, how such epigenetic state is reprogrammed and inherited between generations is poorly understood. Using an ultra-sensitive approach STAR ChIP-seq, we investigated H3K27me3 across 14 developmental stages along mouse gametogenesis and early development. Interestingly, highly pervasive H3K27me3 was found in regions depleted of transcription and DNA methylation in oocytes. Unexpectedly, we observed extensive loss of promoter H3K27me3 at Hox and other developmental genes upon fertilization. This is accompanied by global erasure of sperm H3K27me3 but inheritance of distal H3K27me3 from oocytes. The resulting allele-specific H3K27me3 patterns persist to blastocysts before being converted to canonical forms in postimplantation embryos, where both the H3K4me3/H3K27me3 bivalent promoter marks are restored at developmental genes. Together, these data revealed widespread resetting of epigenetic memory and striking plasticity of epigenome during gametogenesis and early development.

overall_design: Mouse preimplantation embryos were obtained from crosses of C57BL/6N and PWK. A newly developed ChIP-seq approach (STAR ChIP-seq) and RNA-seq were performed in these embryos at variou stages in preimplantation development.

first: Hui

first: Bo

last: Huang

first: Wei

last: Xie


title: Whole genome transcriptome analysis of serotype M1 Group A Streptococcus (Streptococcus pyogenes) wild type and isogenic FabT mutants grown at 35C and 40C.

status: Public on Sep 14 2016

city: Houston

first: Stephen

middle: B.

last: Beres

state: TX

address: 6670 Bertner Ave.

department: Pathology and Genomic Medicine

country: USA

institute: Houston Methodist Research Institute

type: Expression profiling by high throughput sequencing

summary: The goal was to perform global transcriptome analysis on isogenic wild type 2221 and FabT deletion mutant M1 strains to investigate how inactivation of the fabT gene affects gene expression.

overall_design:
The transcriptomes of 2 M1 GAS strains, one wild type MGAS221 strain and one isogenic MGAS2221 fabT deletion mutant strain, were compared at two phases of growth, mid-exponential and early-stationary, and at two temperatures, 35C and 40C, using 3 biologial replicates, to identify transcriptome changes. The fatty acid biosynthesis transcriptional regulator gene, fabT, was the most frequently mutated gene of the MGAS2221 geneome in isolates recovered from a NHP necrotizing fasciitis model of infection.
HMRI-DID (Houston Methodist Research Instittute, Department of Infectious Disease)

first: James

middle: M

last: Musser

middle: B

first: Jesus

last: Eraso


title: Interdependence of PRC1 and PRC2 for recruitment to Polycomb Response Elements

city: Umeå

first: Yuri

last: Schwartz

address: Byggnad 6L, NUS

country: Sweden

institute: Umeå University

type: Genome binding/occupancy profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Next Generation Sequencing of 23116 MT (low Arid1a expression) vs AB-C1 and AB-C2 (high Arid1a expression) Transcriptomes

city: Ithaca

first: John

last: Schimenti

state: New York

address: Veterinary Research Tower T9 007

department: Molecular Biology and Genetics

institute: Cornell University

summary: The goal of this study was to identify important genetic pathways that are altered in mammary tumor cells upon over-expression of the tumor suppressor gene Arid1a. The results of this experiment revealed that Arid1a helps regulate key cell-cycle checkpoint and growth regulatory pathways, either directly or indirectly. This helped explain in part the significant decrease in cell proliferation and tumor growth phenotypes observed both in vitro and in vivo, when comparing the same samples analyzed here by RNA-seq (untransduced replicates vs. add-back clonal lines).

overall_design: Whole transcriptome comparison of mammary tumor cells derived from Chaos3 mouse model (23116 MT- control) vs. add-back clones overexpressing Arid1a (AB-C1 & AB-C2 - exp). Control and experimental samples were run in duplicate.

first: Nithya

last: Kartha


title: Inhibition of DNA methylation promotes breast tumor sensitivity to netrin-1 interference [RNA-Seq]

city: Lyon

first: Robert

last: Dante

address: 28, rue Laennec

department: Department of Tumoral Escape

country: France

institute: Cancer Research Center – Lyon

summary: Results provide the animal proof of concept that inhibition of DNA methylation can sensitize solid tumors to antibodies mediating tumor cell apoptosis.

overall_design: RNAseq of untreated cells, or DAC treated HMLER and MDA-MB-231 cells.

first: Melodie

last: Grandin

first: Mathot

last: Pauline

first: Guillaume

last: Devailly

first: Yannick

last: Bidet

first: Akram

last: Ghantous

first: Clementine

last: Favrot

first: Benjamin

last: Gibert

first: Nicolas

last: Gadot

first: Isabelle

last: Puisieux

first: Zdenko

last: Herceg

first: Jean-Guy

last: Delcros

first: Agnès

last: Bernet

first: Patrick

last: Mehlen


title: Inhibition of DNA methylation promotes breast tumor sensitivity to netrin-1 interference

type: Methylation profiling by high throughput sequencing


title: Inhibition of DNA methylation promotes breast tumor sensitivity to netrin-1 interference [MeDIP-Seq]

overall_design: Methylated DNA precipitation (MeDP) ) followed by hight-throughput sequencing.


title: Identification of mRNAs with reduced ribosomal loading upon knock-down of translation factor DAP5 from hESCs.

city: Rehovot

first: Gilgi

last: Friedlander

address: Hertzel st

country: Israel

institute: Weizmann Institute of Science

summary: We have generated stable human ESCs (H9) expressing control or DAP5-targeting shRNA. Polysome profiles reveal no major changes in overall translation. PolyA+ RNA and RNA accociated with heavy polysomal fractions were purified in biological duplicates and sequenced using Illumina HiSeq 2000 instrument. We identified 122 potential mRNA targets of DAP5 translation that display reduced ribosomal loading, and hence reduced translation, in the absence of DAP5.

overall_design: Total mRNA and heavy polylsomal fractions from shNT and shDAP5 expressing hESCs, each in duplicate, was deep sequenced.

first: Yael

last: Yoffe

first: Ester

last: Feldmesser

first: Shani

last: Bialik

first: Adi

last: Kimchi


title: ChIP-Seq analysis of H3K4me2 and Pol II in Tetrahymena

city: Ann Arbor

first: Yifan

last: lIU

state: Michigan

address: 1301 Cathrine street

institute: University of Michigan

summary: This study describes the epigenetic profiling of H3K4me2 and Pol II in growth stage of Tetrahymena thermophila.

overall_design: ChIP-Seq analysis of Pol II and H3K4me2 occupancy.

last: Liu


title: Global Regulation of Alternative RNA Splicing by the SR-Rich Protein RBM39

status: Public on Sep 13 2016

first: Mai

last: Sanyue

address: 27 Taiping Rd, Haidian District

country: China

institute: Beijing Institute of Biotechnology

summary: RBM39 is extensively involved in alternative splicing of RNA and helps regulate transcript levels. RBM39 may modulate alternative splicing similarly to U2AF65 by either directly binding to RNA or recruiting other splicing factors, such as U2AF65.

overall_design: RNA extracted from wild-type and RBM39-deficient human breast adenocarcinoma MCF-7 cells was subjected to RNA-Seq, and alternative splicing events in the two cell types were analyzed and compared.


title: MNase-seq of Tetrahymena thermophial macronucleus (MAC) and micronucleus (MIC)

summary: In this study, we have employed MNase-Seq to reveal distinct nucleosome distribution patterns in morphologically and functionally differentiated MIC and MAC from asexually dividing Tetrahymena cells.

overall_design: Comparing the nucleosome occupancy features in Tetrahymena MAC and MIC.


title: Capture-C reveals preformed chromatin interactions between HIF-binding sites and distant promoters

city: Oxfors

first: James

last: Platt

address: Roosevelt Drive

country: United Kingdom

institute: University of Oxford

summary: Hypoxia inducible factor (HIF) directs an extensive transcriptional cascade that transduces numerous adaptive responses to hypoxia. Pan-genomic analyses, using chromatin immunoprecipitation and transcript profiling, have revealed large numbers of HIF-binding sites that are generally associated with hypoxia-inducible transcripts, even over long chromosomal distances. However, these studies do not define the specific targets of HIF-binding sites and do not reveal how induction of HIF affects chromatin conformation over distantly connected functional elements. To address these questions we deployed a recently developed chromosome conformation assay that enables simultaneous high-resolution analyses from multiple viewpoints. These assays defined specific long-range interactions between intergenic HIF-binding regions and one or more promoters of hypoxia-inducible genes, revealing the existence of multiple enhancer-promoter, promoter-enhancer and enhancer-enhancer interactions.

overall_design: We utilised Capture-C to study chromatin looping from distal HIF binding sites (enhancers) and the promoters of HIF regulated genes. Studying the dynamics in normoxia and hypoxia and across different cell types.

first: David

last: Mole


title: Genome-wide positioning of bivalent mononucleosomes

city: BALTIMORE

first: hari

last: easwaran

state: MD

address: 1650 Orleans Street, CRB1, Room 530

department: Oncology

institute: Johns Hopkins University

summary: Background: Bivalent chromatin refers to overlapping regions containing activating histone H3 Lys4 trimethylation (H3K4me3) and inactivating H3K27me3 marks. Existence of such bivalent marks on the same nucleosome has only recently been suggested. Previous genome-wide efforts to characterize bivalent chromatin have focused primarily on individual marks to define overlapping zones of bivalency rather than mapping positions of truly bivalent mononucleosomes. Results: Here, we developed an efficacious sequential ChIP technique for examining global positioning of individual bivalent nucleosomes. Using next generation sequencing approaches we show that although individual H3K4me3 and H3K27me3 marks overlap in broad zones, bivalent nucleosomes are focally enriched in the vicinity of the transcription start site (TSS). These seem to occupy the H2A.Z nucleosome positions previously described as salt-labile nucleosomes, and are correlated with low gene expression. Although the enrichment profiles of bivalent nucleosomes show a clear dependency on CpG island content, they demonstrate a stark anti-correlation with methylation status. Conclusions: We show that regional overlap of H3K4me3 and H3K27me3 chromatin tend to be upstream to the TSS, while bivalent nucleosomes with both marks are mainly promoter proximal near the TSS of CpG island-containing genes with poised/low expression. We discuss the implications of the focal enrichment of bivalent nucleosomes around the TSS on the poised chromatin state of promoters in stem cells.

overall_design: Development of sequential ChIP-seq protocols to map nuceosomes with the bivalent H3K4me3 and H3K27me3 marks

first: Hari

last: Easwaran

first: Subhojit

last: Sen


title: Nucleosome Occupancy and Methylome Sequencing (NOMe-seq) analysis using SureSelectXT Methyl-Seq Target Enrichment System

city: Augusta

first: Huidong

last: Shi

state: GA

department: Georgia Cancer Center

institute: Augusta University

summary: Chromatin accessibility plays a key role in epigenetic regulation of gene activation and silencing. Open chromatin regions allow regulatory elements such as transcription factors and polymerases to bind for gene expression while closed chromatin regions prevent the activity of transcriptional machinery.  Nucleosome occupancy and methylome sequencing (NOMe-seq) has been developed for simultaneously profiling of chromatin accessibility and DNA methylation on single molecules. In this study, we combined the principle of NOMe-seq with targeted bisulfite sequencing method to analyze the genome-wide nucleosome occupancy and chromatin accessibility in the promoter and enhancer regions of over 20,000 genes. In addition, we developed CAME, a seed-extension based approach that identifies chromatin accessibility from NOMe-seq. Our results show that our method not only can precisely identify chromatin accessibility but also outperforms other methods.

overall_design: Colon cancer cell line, HCT116, was treated GpC methyltransferase (M.CviPI) to methylate all GpC sites in the open chromain. The genomic DNA was then analyzed by targeted bisulfite sequencing. Oligo capture probes are designed to target 84Mb sequences covering 3.7 million CpGs including CpG islands, Cancer, tissue specific DMRs, Gencode promoters, DMRs or regulatory feature in CpG islands, shores, and shelves ±4kb, DNase I hypersensitive sites, Refseq genes, Ensembl regulatory features. The CAME package was used for analyzing the targeted NOMe-seq data. The efficiency and effectiveness of CAME were demonstrated through comparisons with other existing techniques on both simulated and real data.

first: Eun

last: Lee

first: Jeong-Hyeon

last: Choi


title: Identification Of Candidate Anti-Cancer Molecular Mechanisms Of Compound Kushen Injection Using Functional Genomics

city: Adelaide

middle: L.

last: Adelson

state: SA

address: North Terrace

department: Genetics & Evolution

country: Australia

institute: The University of Adelaide

type: Non-coding RNA profiling by high throughput sequencing

summary: Compound Kushen Injection (CKI) has been clinically used in China for over 15 years to treat various types of solid tumours.  However, because such Traditional Chinese Medicine (TCM) preparations are complex mixtures of plant secondary metabolites, it is essential to explore their underlying molecular mechanisms in a systematic fashion.  We have used the MCF-7 breast cancer cell line as an initial in vitro model to identify CKI induced changes in gene expression.  Cells were treated with CKI for 24 and 48 hours at two concentrations (1.0 and 2.0 mg/mL), and 5-Fluorouracil (5-FU) was used to treat cells as a positive control.  Cell proliferation and apoptosis activity were measured with XTT and Caspase-3 assays respectively.  Transcriptome data of cells treated with CKI or 5-FU for 24 and 48 hours were acquired using high-throughput Illumina RNA-seq technology. In this report we show that CKI inhibited MCF-7 cell proliferation and induced apoptosis in a dose-dependent fashion.  We integrated and applied a series of transcriptome analysis methods, including gene differential expression analysis, pathway over-representation analysis, de novo identification of long non-coding RNAs (lncRNA) as well as co-expression network reconstruction, to identify candidate anti-cancer molecular mechanisms of CKI.  Multiple pathways were perturbed and the cell cycle was identified as the potential primary target pathway of CKI in MCF-7 cells.  CKI may also induce apoptosis in MCF-7 cells via a p53 independent mechanism.  In addition, we identified novel lncRNAs and showed that many of them might be expressed as a response to CKI treatment.  Overall, we have comprehensively investigated the utility of transcriptome analysis with high-throughput sequencing to characterise the molecular response of cancer cells to a TCM drug, and provided a practical guideline for future molecular studies of TCM.

overall_design: High-depth paired-end RNA-seq from MCF-7 cell line. Each sample contains 3 biological replicates.

first: Zhipeng

last: Qu

middle: L


title: microRNA profiling from kidney cortex samples of Macaca fascicularis (Cynomolgus Monkey)

city: Chilly Mazarin

first: Veeranagouda

last: Yaligara

address: Sanofi R&D

institute: Sanofi

summary: MicroRNAs (miRNAs) present in tissues and biofluids are emerging as sensitive and specific safety biomarkers. MiRNAs have not been thoroughly described in M. fascicularis, an animal model used in pharmaceutical industry especially in drug safety evaluation. Here we investigated the miRNAs in M. fascicularis. For Macaca mulatta, a closely related species of M.fascicularis, 619 stem-loop precursor miRNAs (pre-miRNAs) and 914 mature miRNAs are available in miRBase version 21. Using M. mulatta miRNAs as a reference list and homology search tools, we identified 604 pre-miRNAs and 913 mature miRNAs in the genome of M.fascicularis. In order to validate the miRNAs identified by homology search we attempted to sequence miRNAs expressed in kidney cortex from M. fascicularis. MiRNAs expressed in kidney cortex may indeed be released in urine upon kidney cortex damage and be potentially used to monitor drug induced kidney injury. Hence small RNA sequencing libraries were prepared using kidney cortex tissues obtained from three naive M. fascicularis and sequenced. Analysis of sequencing data indicated that 432 out of 913 mature miRNAs were expressed in kidney cortex tissues. Assigning these 432 miRNAs to pre-miRNAs revealed that 273 were expressed from both the -5p and -3p arms of 150 pre-miRNAs and 159 miRNAs expressed from either the -5p or -3p arm of 176 pre-miRNAs. Mapping sequencing reads to pre-miRNAs also facilitated the detection of twenty-two new miRNAs. To substantiate miRNAs identified by small RNA sequencing, 313 miRNAs were examined by RT-qPCR. Expression of 262 miRNAs in kidney cortex tissues ware confirmed by TaqMan microRNA RT-qPCR assays. Analysis of kidney cortex miRNA targeted genes suggested that they play important role in kidney development and function. Data presented in this study may serve as a valuable resource to assess the renal safety biomarker potential of miRNAs in Cynomolgus monkeys.

overall_design: Examination of miRNAs in kidney cortex tissues obtained from three naïve M. fascicularis

first: Pierrick

last: Rival

first: Catherine

last: Prades

first: Claire

last: Mariet

first: Jean-François

last: Léonard

first: Jean-Charles

last: Gautier

first: Xiaobing

last: Zhou

first: Jufeng

last: Wang

first: Bo

last: Li

first: Marie-Laure

last: Ozoux

first: Eric

last: Boitier


title: ChIP-seq analysis of the C. elegans CEBP-1 protein

city: La Jolla

first: Kyung Won

last: Kim

state: CA

address: 9500 Gilman Drive

department: Section of Neurobiology, Division of Biological Sciences

institute: University of California, San Deigo

summary:
Purpose: The goal of this study is to identify genes selectively associated with C. elegans CEBP-1.
Methods: We generated transgenic animals expressing FLAG-tagged CEBP-1 in a cebp-1(tm2807) mutant background (cebp-1(0); juIs418 [Pcebp-1::FLAG::CEBP-1::cebp-1 3’UTR]) and then immunoprecipitated FLAG-CEBP-1-associated DNA fragments using anti-FLAG antibodies (M2 anti-FLAG magnetic beads; Sigma). We collected mixed stage worms grown at 20˚C on NGM plates followed by 2% formaldehyde and sonicated the samples as described (Mukhopadhyay et al., 2008). We next generated ChIP-seq DNA libraries. Briefly, both ChIPed DNA and input genomic DNA were ligated to specific adaptors and amplified by barcode primers followed by sequencing on the Illumina HiSeq-2000 platform. We performed two independent ChIP-seq experiments, with parallel genomic DNA controls prepared from the same strain. We conducted peak-calling using CLC genomics workbench 6.0 (CLCbio). To define genes associated with the peaks, we used the annotation of transcription start site (TSS) and transcription end site (TES) from WS220 and annotated the peak if it overlapped the gene or the 3 kb upstream of the TSS. We then manually confirmed the peaks and associated genes using UCSC browser and update to WS252.
Results: We found 209 CEBP-1 ChIP-seq peaks in the genome that were associated with 212 coding genes. Through motif discovery tools,  we also found that CEBP-1 binds conserved DNA motifs.
Conclusions: The conservation of C.elegans CEBP-1 binding motif supports functional parallels between C. elegans CEBP-1 and vertebrate C/EBPs.

overall_design: CEBP-1 ChIP and input sample

first: Kyung

first: Yishi

last: Jin


title: Micro-C XL: assaying chromosome conformation at length scales from the nucleosome to the entire genome

city: Berkeley

first: Xavier

last: Darzacq

address: 475D Li Ka Shing Center

department: Molecular and Cell Biology

institute: University of California, Berkeley

summary: Structural analysis of chromosome folding in vivo has been revolutionized by Chromosome Conformation Capture (3C) and related methods, which use proximity ligation to identify chromosomal loci in physical contact. We recently described a variant 3C technique, Micro-C, in which chromatin is fragmented to mononucleosomes using micrococcal nuclease, enabling nucleosome-resolution folding maps of the genome. Here, we describe an improved Micro-C protocol using long crosslinkers, termed Micro-C XL, which exhibits greatly increased signal to noise, and provides further insight into the folding of the yeast genome. We also find that signal to noise is much improved in Micro-C XL libraries generated from relatively insoluble chromatin as opposed to soluble material, providing a simple method to physically enrich for bona-fide long-range interactions. Micro-C XL maps of the budding and fission yeast genomes reveal both short-range chromosome fiber features such as chromosomally-interacting domains (CIDs), as well as higher-order features such as clustering of centromeres and telomeres, thereby addressing the primary discrepancy between prior Micro-C data and reported 3C and Hi-C analyses. Interestingly, comparison of chromosome folding maps of S. cerevisiae and S. pombe revealed widespread qualitative similarities, yet quantitative differences, between these distantly-related species. Micro-C XL thus provides a single assay suitable for interrogation of chromosome folding at length scales from the nucleosome to the full genome.

sample_organism:
Schizosaccharomyces pombe
Saccharomyces cerevisiae

overall_design: Chromatin is fragmented by Mnase, subsequenct nucleosomal end repair, and a modified two-step method for purfiying ligation products. Using Illumina paired-end sequencing maps Micro-C library and generates nucleosome resolution contact maps.

first: Oliver

middle: J

last: Rando

first: Tsung-Han

middle: S

last: Hsieh


title: The genome wide transcriptional profiling of periodontitis-associated fibroblasts identified DLX5 and RUNX2 as central regulatory elements [RNA-seq]

city: Los Angeles

first: Masafumi

last: Horie

address: 2011 Zonal Ave

institute: University of Southern California

summary: RNA-seq of normal gingival fibroblasts

overall_design: RNA-seq of normal gingival fibroblasts

first: Akira

last: Saito

first: Yoko

last: Yamaguchi

first: Mitsuhiro

last: Ohshima


title: Differentially Expressed Gene Transcripts Using RNA Sequencing from the Blood of Immunosuppressed Kidney Allograft Recipients

city: Minneapolis

first: Casey

last: Dorr

state: MN

address: 701 Park Avenue

institute: Minneapolis Medical Research Foundation

summary: We performed RNA sequencing (RNAseq) on peripheral blood mononuclear cells (PBMCs) to identify differentially expressed gene transcripts (DEGs) after kidney transplantation and after the start of immunosuppressive drugs. RNAseq is superior to microarray to determine DEGs because it’s not limited to available probes, has increased sensitivity, and detects alternative and previously unknown transcripts. DEGs were determined in adult kidney recipients, without clinical acute rejection (AR), treated with antibody induction, calcineurin inhibitor, mycophenolate, with and without steroids. Blood was obtained pre-transplant (baseline), week 1, months 3 and 6 post-transplant. PBMCs were isolated, RNA extracted and gene expression measured using RNAseq. Principal components (PCs) were computed using a surrogate variable approach. DEGs post-transplant were identified by controlling false discovery rate (FDR) at < 0.01 with at least a 2 fold change in expression from pre-transplant. The top 5 DEGs with higher levels of transcripts in blood at week 1 were TOMM40L, TMEM205, OLFM4, MMP8, and OSBPL9 compared to baseline. The top 5 DEGs with lower levels at week 1 post-transplant were IL7R, KLRC3, CD3E, CD3D, and KLRC2(Striking Image) compared to baseline. The top pathways from genes with lower levels at 1 week post-transplant compared to baseline, were T cell receptor signaling and iCOS-iCOSL signaling while the top pathways from genes with higher levels than baseline were axonal guidance signaling and LXR/RXR activation. Gene expression signatures at month 3 were similar to week 1. DEGs at 6 months post-transplant create a different gene signature than week 1 or month 3 post-transplant. RNAseq analysis identified more DEGs with lower than higher levels in blood compared to baseline at week 1 and month 3. The number of DEGs decreased with time post-transplant. Further investigations to determine the specific lymphocyte(s) responsible for differential gene expression may be important in selecting and personalizing immune suppressant drugs and may lead to targeted therapies.

overall_design: RNAseq data are from blood specimens from kidney transplant patients as a time course  before and after kidney transplant from living donors. There are 96 samples analyzed in total from 34 patients; however, only 32 patients with baseline data and pre-transplant RNA. None of the patients in the study developed acute rejection within the 6 month study period. Time points include: pre-transplant, 1 week following transplant, 3 and 6 months following transplant.

first: Baolin

last: Wu

first: Weihua

last: Guan

first: Amutha

last: Muthusamy

first: Kinjal

last: Sanghavi

first: David

middle: P

last: Schladt

first: Jonathan

last: Maltzman

first: Steven

middle: E

last: Scherer

first: Marcia

last: Brott

first: Arthur

last: Matas

first: Pamala

middle: A

last: Jacobson

first: William

last: Oetting

first: Ajay

middle: K

last: Israni


title: Genome-wide transcriptome analyses of E. coli treated with ribosome-targeting antibiotics

city: Chicago

first: Mee-Ngan

middle: F.

last: Yap

state: IL

address: 320 E Superior St

department: Microbiology-Immunology

institute: Northwestern University Feinberg School of Medicine

summary: Steady state RNA levels were measured using total RNA-seq.

overall_design: RNA levels in antibiotic-treated E. coli cells were compared to levels in untreated cells.

first: Ekaterina

last: Dzyubak

first: Mee

middle: N


title: Allelic reprogramming of the histone modification H3K4me3 in early mammalian development

status: Public on Sep 15 2016

city: Beijing

first: Bingjie

last: Zhang

address: 30 Shuangqing Rd

institute: Tsinghua University

summary: Histone modifications are considered as fundamental epigenetic regulators that control many crucial cellular processes 1. However, whether these marks can be passed on from gametes to the next generation in mammals is a long-standing question that remains unclear. By developing a highly sensitive approach, STAR ChIP-seq, we provided a panoramic view of the landscape of H3K4me3, a histone hallmark for transcription initiation 2, from developing gametes to postimplantation embryos. Interestingly, we found that upon fertilization, extensive reprogramming occurred on the paternal genome, as few H3K4me3 peaks were observed in zygotes before they reappeared after major zygotic genome activation at the late 2-cell stage. On the maternal genome, we unexpectedly found a non-canonical form of H3K4me3 (ncH3K4me3) in full-grown and mature oocytes, which exists as broad peaks at promoters and a large number of distal loci. Such broad H3K4me3 peaks are in striking contrast to typical sharp H3K4me3 peaks restricted in CpG rich regions of promoters. Interestingly, ncH3K4me3 in oocytes overlaps almost exclusively with partially methylated DNA domains (PMDs). It is then inherited in preimplantation embryos before being erased in the late 2-cell embryos, when canonical H3K4me3 starts to be established. The removal of ncH3K4me3 requires zygotic transcription but is independent of DNA replication mediated passive dilution. Finally, downregulation of H3K4me3 in full-grown oocytes by overexpressing an H3K4me3 demethylase KDM5B is associated with defects in genome silencing. Taken together, these data unveiled inheritance and highly dynamic reprogramming of the epigenome in early mammalian development.

sample_organism:
Danio rerio
Mus musculus

platform_organism:
Danio rerio
Mus musculus

overall_design:
Mouse preimplantation embryos were obtained from crosses of C57BL/6N and PWK. A newly developed ChIP-seq approach (STAR ChIP-seq) and RNA-seq were performed in these embryos at variou stages in preimplantation development.

Please note that [1] 'PN5 zygote histone H3' sample represents input files for PN5_K4me3 sample, using H3 antibody.
[2] both separate (for each replicate sample, i.e. sample supplementary file) and combined processed bed file (for all replicates, i.e. Series supplementary file) have been provided.

first: Hui

last: Zheng

first: Bo

last: Huang

first: Wenzhi

last: Li

first: Wei

last: Xie


title: RNA-Seq analysis of human medulloblastoma cell lines with SOX9 overexpression

city: Uppsala

first: Fredrik

middle: Johansson

last: Swartling

address: Dag Hammarskjölds väg 20

department: Immunology, Genetics and Pathology

institute: Uppsala University

summary: SOX9 is a master transcription factor that regulates development and stem cell programs. However, its potential oncogenic activity and regulatory mechanisms that control SOX9 protein stability are poorly understood. Here we show that SOX9 is a substrate of FBW7, a tumor suppressor and a SCF (Skp1-Cul1-F-box)-type ubiquitin ligase. FBW7 recognizes a conserved degron surrounding threonine 236 (T236) in SOX9 that is phosphorylated by GSK3 kinase. Specifically, FBW7-alpha targets T236-phosphorylated SOX9 for ubiquitylation and proteasomal degradation. Further, we demonstrate that FBW7 inactivation stabilizes SOX9 protein promoting migration, metastasis and treatment resistance in medulloblastoma, one of the most common childhood brain tumors. Notably, expression of mutationally stabilized SOX9-T236/240A in medulloblastoma cells coincides with activation of pro-metastatic genes. FBW7 is frequently downregulated in all medulloblastoma subgroups and mutated specifically in SHH-driven medulloblastoma. In cases where FBW7 mRNA levels are low, SOX9 protein is significantly elevated and this phenotype is associated with metastasis at diagnosis and poor patient outcome. Finally, pharmacological inhibition of PI3K/Akt/mTOR activity destabilizes SOX9 in a GSK3/FBW7-dependent manner, rendering medulloblastoma cells sensitive to cytostatic treatment.

overall_design: Triplicates of MB002 cells transduced with EV, SOX9-WT or SOX9-T236/240A.

first: Aldwin

last: Rahmanto

first: Vasil

last: Savov

first: Olle

last: Sangfelt


title: Rapid, dynamic activation of Müller glial stem cell responses in zebrafish

first: Pamela

last: Raymond

state: MI

address: 830 N. University

department: Molecular, Cellular, and Developmental Biology

summary: Purpose: Zebrafish neurons regenerate from Müller glia following retinal lesions. Genes and signaling pathways important for retinal regeneration in zebrafish have been described, but our understanding of how Müller glial stem cell properties are regulated is incomplete. Mammalian Müller glia possess a latent neurogenic capacity that might be enhanced in regenerative therapies to treat degenerative retinal diseases. Methods: To identify transcriptional changes associated with stem cell properties in zebrafish Müller glia, we performed a comparative transcriptome analysis from isolated cells at 8 and 16 hours following an acute, photic lesion, prior to the asymmetric division that produces retinal progenitors. Results: We report a rapid, dynamic response of zebrafish Müller glia, characterized by activation of pathways related to stress, NF-kappa B signaling, cytokine signaling, immunity, prostaglandin metabolism, circadian rhythm, and pluripotency, and an initial repression of Wnt signaling. When we compared publicly available transcriptomes of isolated mouse Müller glia from two retinal degeneration models, we found that mouse Müller glia showed evidence of oxidative stress, variable responses associated with immune regulation, and repression of pathways associated with pluripotency, development, and proliferation. Conclusions: Categories of biological processes/pathways activated following photoreceptor loss in regeneration-competent zebrafish Müller glia, which distinguished them from mouse Müller glia in retinal degeneration models, included: cytokine signaling (notably NF-kappa B), prostaglandin E2 synthesis, expression of core clock genes, and pathways/metabolic states associated with pluripotency. These regulatory mechanisms are relatively unexplored as potential mediators of stem cell properties likely to be important in Müller glial cells for successful retinal regeneration.

overall_design: Transcriptional profiles of 0, 8, and 16 hour post-lesion zebrafish Müller glia (in triplicate) were generated by high-throughput sequencing in an Illumina GAIIx.

first: Christopher

last: Sifuentes

first: Jung-Woong

last: Kim

first: Anand

last: Swaroop


title: RNA-seq profiling of mouse PGC (Mus musculus)

city: Taipei

first: Pao-Yang

last: Chen

address: 128 Sec. 2, Academia Rd, Nankang,

department: Institute of Plant and Microbial Biology

country: Taiwan

institute: Academia Sinica

summary: PGCs undergo two distinct stages of demethylation before reaching a hypomethylated ground state at E13.5.  Stage 1 occurs between E7.25- E9.5 in which PGCs experience a global loss of cytosine methylation.  However, discreet loci escape this global loss of methylation and between E10.5-E13.5, stage 2 of demethylation takes place.  In this stage these loci are targeted by Tet1 and Tet2 leading to the loss of the remaining methylation and resulting in the epigenetic ground state.  Our data shows that Dnmt1 is responsible for maintaining the methylation of loci that escape stage 1 demethylation, and that it functions in a UHRF1 independent manner.  Our data further demonstrates that when these loci lose methylation prior to stage 2 it results in early activation of the meiotic program, which leads to precocious differentiation of the germ line resulting in a decreased pool of PGCs in the embryo and subsequent infertility in adult mice.

overall_design: Examination of transcription of Mouse PGCs

first: Joseph

last: Hargan-Calvopina

first: Sara

last: Taylor

first: Helene

last: Cook

first: Zhongxun

last: Hu

first: Serena

last: Lee

first: Ming-Ren

last: Yen

first: Yih-Shien

last: Chiang

first: Amander

middle: T

last: Clark


title: Genome-wide distribution of H2A-S40Gc in mouse trophoblast stem cells

city: Imabari-shi

first: Koji

last: Hayakawa

state: Ehime

address: 1-3 Ikoino-oka

department: Department of Toxicology, Faculty of Veterinary Medicine

country: Japan

institute: Okayama University of Science

summary: We report genome-wide distribution of O-GlcNAcylated H2A at serine 40 (H2A-S40Gc) in mouse trophoblast stem cells (TSCs). We found that H2A-S40Gc was mainly located at genic area, positively correlated with the gene expression, and varied the localization during their differentiation. This study using ChIP-seq analysis provides genomic distribution of newly O-GlcNAc histone modification.

overall_design: ChIP-seq analysis of H2A-S40Gc in TSCs and differentiated TSCs (dTSCs).

first: Kunio

last: Shiota


title: Expression profiling identifies Sertoli and Leydig cell genes as Fsh targets in adult zebrafish testis

city: Bergen

first: Diego

last: Crespo

address: Nordnesgaten 50

department: Reproduction and Developmental Biology

country: Norway

institute: Institute of Marine Research

summary: Spermatogonial stem cells are quiescent, undergo self-renewal or differentiating divisions, thereby forming the cellular basis of spermatogenesis. This cellular development is orchestrated by follicle-stimulating hormone (FSH), through the production of Sertoli cell-derived factors, and by Leydig cell-released androgens. Here, we investigate the transcriptional events induced by Fsh in a steroid-independent manner on the restart of zebrafish (Danio rerio) spermatogenesis ex vivo, using testis from adult males where type A spermatogonia were enriched by estrogen treatment in vivo. Under these conditions, RNA sequencing preferentially detected differentially expressed genes in somatic/Sertoli cells. Fsh-stimulated spermatogonial proliferation was accompanied by modulating several signaling systems (i.e. Tgf-β, Hedgehog, Wnt and Notch pathways). In silico protein-protein interaction analysis indicated a role for Hedgehog family members potentially integrating signals from different pathways during fish spermatogenesis. Moreover, Fsh had a marked impact on metabolic genes, such as lactate and fatty acid metabolism, or on Sertoli cell barrier components. Fish Leydig cells express the Fsh receptor and one of the most robust Fsh-responsive genes was insulin-like 3 (insl3), a Leydig cell-derived growth factor. Follow-up work showed that recombinant zebrafish Insl3 mediated pro-differentiation effects of Fsh on spermatogonia in an androgen-independent manner. Our experimental approach allowed focusing on testicular somatic genes in zebrafish and showed that the activity of signaling systems known to be relevant in stem cell systems was modulated by Fsh, providing promising leads for future work, as exemplified by the studies on Insl3.

overall_design: 12 samples in total were analyzed: 6 biological replicates from control testis samples and 6 biological replicates from Fsh-treated testis samples (all co-incubated with trilostane).

first: Luiz

middle: H

last: Assis

first: Tomasz

last: Furmanek

first: Jan

last: Bogerd

first: Rüdiger

middle: W

last: Schulz


title: High-throughput single-base resolution mapping of 2’-O-Methylated residues

city: Groningen

first: Danny

last: Incarnato

state: Netherlands

address: Nijenborgh 7

department: Molecular Genetics

country: Netherlands

institute: University of Groningen

summary: Functional characterization of the transcriptome requires tools for the systematic investigation of RNA post-transcriptional modifications. 2’-O-Methylation of the ribose moiety is one of the most abundant post-transcriptional modifications of the RNA. We describe here a high-throughput method that enables fast and accurate mapping at single-base resolution, and quantitation, of 2’-OMe modified residues. Our approach expands the actual repertoire of methods for transcriptome-wide mapping of RNA post-transcriptional modifications.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Mapping 2’-O-Methylated residues in Hela cells and E14 mouse embryonic stem cells.

first: Francesca

last: Anselmi

first: Edoardo

last: Morandi

first: Francesco

last: Neri

first: Mara

last: Maldotti

first: Stefania

last: Rapelli

first: Caterina

last: Parlato

first: Giulia

last: Basile

first: Salvatore

last: Oliviero


title: RNA-Seq of eye tissues from Monkey eye and retina samples

status: Public on Sep 12 2016

city: Cleveland

first: Xiaodong

last: Bai

address: 10900 Euclid Avenue

department: Center for RNA Molecular Biology

institute: Case Western Reserve University

summary: We report RNA-Seq experiments of eye and retinal tissues from different ocular compartments in the Monkey

overall_design: Examine ocular tissue from different regions of the Monkey eye and retina

first: Debarshi

last: Mustafi

first: Krzysztof

last: Palczewski


title: RNA-Seq of retinal tissue from a Human Eye

summary: We report RNA-Seq experiments of retinal tissue from Homo Sapiens

overall_design: Examine retinal tissue from human


title: RNA-Seq of eye tissues from Ground Squirrel eye and retina samples

summary: We report RNA-Seq experiments of eye and retinal tissues from Ground Squirrel (Ictidomys tridecemlineatus)

overall_design: Examine ocular tissue from Ictidomys tridecemlineatus


title: ChIPseq and RNAseq to profile IFNg response in human primary monocytes

city: New York

first: Lionel

last: Ivashkiv

state: NY

address: 535 East 70th Street

institute: Hospital for Special Surgery

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

overall_design: Refer to individual Series


title: RNAseq to profile IFNg response in human primary monocytes

summary: We did transcriptome profiling for monocytes treated with or without IFNg to characterize IFNg response.

overall_design: Human primary monocytes were cultured for 24 hours with or without IFNg, harversted and prepared for RNA for RNAseq.

first: Yu

last: Qiao

first: Eugenia

last: Giannopoulou

first: Tuo

last: Zhang


title: The human olfactory transcriptome

city: Rehovot

first: Tsviya

last: Olender

address: 234 Herzl St.

institute: The Weizmann Institute

summary:
We report the first RNAseq study of human olfactory epithelium, aimed at obtaining a whole transcriptome overview of the sensory tissue. Our work reveals nearly 200 olfactory-enriched non-receptor transcripts, 80% of which have not yet been implicated in chemosensory function, thus providing a platform for future discovery.
Our study further allowed us to quantitate the expression levels of most (90%) of the ~400 intact human olfactory receptor (OR) genes. The expression levels of different paralogous members of the OR repertoire span a 1000-fold range, suggesting a strong imbalance among different odorant specificities. Further, we obtained convincing hints that different human beings show different cross-repertoire expression patterns. Having successfully assembled transcripts for 100 OR genes, we observed prevalent genome-encoded mutations that render some transcripts inactive despite the fact that the protein coding region is intact, an unreported mechanism for OR pseudogenization.
In sum, our transcriptome study offers important observations on genes that underlie olfactory function and provides a basis for significant future work.

overall_design: total RNAseq of human olfactory epithelium

first: Ifat

last: Keydar

first: Jayant

last: Pinto

first: Pavlo

last: Tatarskyy

first: Anna

last: Alkelai

first: Ming-Shan

last: Chien

first: Diego

last: Restrepo

first: Hiroaki

last: Matsunami

first: Yoav

last: Gilad

first: Doron

last: Lancet


title: Plasma Exosome microRNAs are Indicative of Breast Cancer

city: Oklahoma city

first: Bethany

last: Hannafon

state: OK

address: 975 NE 10th Street

department: Obstetrics and Gynecology

institute: University of Oklahoma Health Sciences Center

summary:
Introduction: microRNAs are promising candidate breast cancer biomarkers due to their cancer-specific expression profiles. However, efforts to develop circulating breast cancer biomarkers are challenged by the heterogeneity of microRNAs in the blood. To overcome this challenge, we aimed to develop a molecular profile of microRNAs specifically secreted from breast cancer cells. Our first step towards this direction relates to capturing and analyzing the contents of exosomes, which are small secretory vesicles that selectively encapsulate microRNAs indicative of their cell of origin. To our knowledge, circulating exosome microRNAs have not been well evaluated as biomarkers for breast cancer diagnosis or monitoring.
Methods: Exosomes were collected from the conditioned media of human breast cancer cell lines, mouse plasma of patient-derived orthotopic xenograft models (PDX), and human plasma samples. Exosomes were verified by electron microscopy, nanoparticle tracking analysis, and western blot. Cellular and exosome microRNAs from breast cancer cell lines were profiled by next-generation small RNA sequencing. Plasma exosome microRNA expression was analyzed by qRT-PCR analysis.
Results: Small RNA sequencing and qRT-PCR analysis showed that several microRNAs are selectively encapsulated or highly enriched in breast cancer exosomes. Importantly, the selectively enriched exosome microRNA, human miR-1246, was detected at significantly higher levels in exosomes isolated from PDX mouse plasma, indicating that tumor exosome microRNAs are released into the circulation and can serve as plasma biomarkers for breast cancer. This observation was extended to human plasma samples where miR-1246 and miR-21 were detected at significantly higher levels in the plasma exosomes of 16 breast cancer patients as compared to the plasma exosomes of healthy control subjects. Receiver Operating Characteristic (ROC) curve analysis indicated that the combination of plasma exosome miR-1246 and miR-21 levels is a better indicator of breast cancer than their individual levels.
Conclusions: Our results demonstrate that certain microRNA species, such as miR-21 and miR-1246, are selectively enriched in human breast cancer exosomes and significantly elevated in the plasma of breast cancer patients. These findings indicate a potential new strategy to selectively analyze plasma breast cancer microRNAs indicative of the presence of breast cancer.

overall_design: The small RNA in the cells and exosomes of the normal mammary epithelial cell line (MCF10A) and two breast cancer cell lines (MCF7 and MDA-MB-231) were analyzed by Next Gen RNA Sequencing.

first: Yvonne

middle: D

last: Trigoso

first: Cameron

last: Calloway

first: Daniel

last: Zhao

first: David

last: Lum

first: Joe

first: Ken

last: Blick

first: Alana

last: Welm

first: William

middle: C

last: Dooley

first: Wei-Qun

last: Ding


title: Differential cytokine sensitivities of mammary-specific and universal STAT5 enhancers depends on Stat5 autoregulation

city: Bethesda

first: Michaela

last: Willi

address: 9000 Rockville Pike

department: NIDDK

institute: NIH

summary: Cytokines utilize the transcription factor STAT5 to control cell-specific and universal genes. In general, the magnitude of cell-restricted gene activation greatly exceeds that of universal genes, with a mechanistic explanation yet to be supplied. Genome-wide studies have identified putative STAT5-based mammary-specific enhancers and universal STAT5-controlled regulatory elements, an opportunity to investigate mechanisms underlying their differential response to cytokines. We have now interrogated the integrity and function of both categories of regulatory elements using biological and genetic approaches. During lactation, STAT5 occupies mammary-specific and universal cytokine-responsive elements. Following lactation, prolactin levels decline and STAT5-dependent enhancers at mammary-specific genes are decommissioned with 24 hours while universal regulatory complexes remain intact. These differential sensitivities are linked to STAT5 concentrations and the mammary-specific Stat5 autoregulatory enhancer. In its absence, mammary-specific enhancers, but not universal elements, fail to be fully established. Upon termination of lactation, STAT5 binding to a subset of mammary enhancers is substituted by STAT3. No STAT3 binding was observed at the most sensitive STAT5 enhancers, suggesting that upon hormone withdrawal their chromatin becomes inaccessible. This study, for the first time, provides molecular insight into the differential sensitivities of mammary-specific and universal cytokine-sensing enhancers.

overall_design: ChIP-seq for STAT5A, GR, H3K27ac, STAT3 in wild type mammary tissues at lactation day one (L1), involution 12 hours (I12) and involution 24 hours (I24). ChIP-seq for STAT5A and H3K27ac of mice heterozygous for Stat5 (Stat5+/-), homozygous for the Stat5 enhancer mutation (Stat5ΔE/ΔE) and mice carrying one Stat5-null allele and one mutant allele (Stat5ΔE/-) at L1. ChIP-seq for STAT5A and H3K27ac of mice homozygous for the Stat5 enhancer mutation (Stat5ΔE/ΔE) at I12 and I24.

first: Kyung

last: Yoo

first: Chaochen

last: Wang

first: Zlatko

last: Trajanoski

first: Lothar

last: Hennighausen


title: RNA-Seq of eye tissue from Nile rat eye samples

first: Xiaodong

last: Bai

summary: We report RNA-Seq experiments of eye tissues from Nile rat (Arivacanthis niloticus)

overall_design: Examine ocular tissue from Arivacanthis niloticus

first: Debarshi

last: Mustafi

first: Krzysztof

last: Palczewski


title: RNA-seq analysis to identify the genes regulated by p53-SET interplay

first: Wei

last: Gu

address: 1130 Saint Nicholas Ave

department: ICG

institute: Columbia University

summary: To identify the genes regulated by SET in a p53-dependent manner, the endogenous SET was depleted by siRNA in control or p53 knockout U2OS cells. Total RNA from each sample was extracted and the whole profile of gene expression was analyzed by RNA-seq analysis.

overall_design: U2OS cells were transfected with control CRISPR/Cas9 construct or SET-specific targeting CRISPR/Cas9 construct. After selection and clone identification, we got a pair of cell lines: 1) control cell line which expresses p53 as normal; 2) p53 knockout cell line which is completely loss of p53 expression. Next, control siRNA or SET-specific siRNA was transfected into these cell lines for 96 hours. Total RNA was extracted for further RNA-seq analysis.

first: Donglai


title: RNA-Seq of tissues from Mouse eye and retina samples

summary: We report RNA-Seq experiments of eye and retinal tissues from WT and RHO KO mice

overall_design: Examine ocular tissue from different mouse genotypes


title: Expression analysis after PNLDC1 silencing in E14 cells

first: Markus

last: Hafner

address: 50 South Drive

department: NIAMS

summary: Next generation sequencing  using the Ion Torrent PGM platform after PNLDC1 silencing  in mouse embryonic stem cells E14 using esiRNA

overall_design: Differential expression analysis of E14 cells after PNLDC1 silencing

first: Dimitrios

last: Anastasakis

first: Ilias

last: Skeparnias

first: Athanasios-Nasir

last: Shaukat

first: Katerina

last: Grafanaki

first: Constantinos

last: Stathopoulos


title: Fetal Brain lesions after subcutaneous inoculation of Zika virus in a pregnant nonhuman primate

first: Michael

last: Gale

state: Minnesota

address: 689 23rd Ave SE

department: Microbiology and Immunology

institute: University of Minnesota

summary: ZIKV was inoculated (strain FSS13025, Cambodia 2010) subcutaneously at five separate locations on the forearms, each with 107 plaque-forming units (PFU) into a healthy pregnant pigtail macaque at 119 days gestation (~28 weeks human pregnancy) to test directly whether ZIKV causes fetal brain injury.

overall_design: Brain tissues from dam and fetus at necropsy (day 43 post-inoculation) were sequenced. A pigtail macaque brain sample from the Non-Human Primate Reference Transcriptome Project (NHPRTR.org) was used as a control

first: Kristina

last: Waldorf

last: Gale Jr.

first: Jennifer

last: Stencel-Baerenwald

first: Richard

last: Green

last: Tisoncik-Go


title: Identification and characterization of warm-sensitive thermoregulatory neurons in the preoptic area

city: San Francisco

first: Chan Lek

last: Tan

state: California

address: 1550 Fourth St. Room 346

department: Physiology

institute: University of California


title: Cell-specific gene expression profiling of warm-sensitive neurons

summary: TRAP-seq gene expression profiling. GFP-L10a is expressed in BDNF+ or PACAP+ in preoptic area.   Bdnf+ or Pacap+ cells in the preoptic area are labelled with GFP-L10a fusion protein expression to study cell-type specific gene expression. We used adult BDNF-Cre (custom generated, discussed in submitted paper) and Pacap-Cre mice (Allen Brain Institute, gift Hongkui Zeng) and injected Cre-depndent adenoassociated virus (AAV2-EF1a-DIO-EGFP-L10a, custom made) into the ventromedial preoptic area.

overall_design: Comparison of Immunoprecipitation and Input fraction mRNA

first: Chan

first: Zachary

last: Knight


title: Identification of microRNAs and their targets in Vernicia fordii seeds by high-throughput sequencing

city: Porto Alegre

first: Rogerio

last: Margis

state: RS

address: Av. Bento Goncalves 9500

department: Centro de Biotecnologia

country: Brazil

institute: Universidade Federal do Rio Grande do Sul

summary: The tung (Vernicia fordii) oil is a valuable industrial oil and has been considered to the production of biodiesel. Because of the poor agronomic traits of this crop, efforts have been made to transfer genes related to the production of its oil to higher yielding plants. However, the mechanisms highlighting gene expression in this plant is poorly understood. MicroRNAs are endogenously encoded small RNAs that play a key role in diverse plant biological processes. In order to identify and characterize miRNAs and their targets in V. fordii we generated by deep sequencing a sRNA and a mRNA library from mature seeds. Using sequence homology we identified 138 conserved plant miRNAs, which were distributed in 34 conserved miRNAs families. Further computation analysis allowed the prediction of secondary structures for 18 conserved and 14 novel miRNAs, as well as several miRNA variants (isomiRNAs). The predicted miRNA target genes are involved in a broad range of physiological functions, including growth and development, stress resistance and also lipid metabolism. This study presents the first identification of V. fordii miRNAs and contributes to understand the function of these miRNAs.

overall_design: microRNA identification in a mature seed library of Vernicia fordii by deep sequencing (Illumina HiSeq2000).

first: Vanessa

middle: V

last: Galli

first: Ana

last: Körbes

first: Frank

middle: F

last: Guzman

first: Rogério

middle: R

first: Rafael

last: Messias

first: Sérgio

last: Silva


title: RNA-Seq of tissues from Rat eye and retina samples

first: Xiaodong

last: Bai

summary: We report RNA-Seq experiments of eye and retinal tissues from Rattus Norvegicus

overall_design: Examine ocular tissue from Rattus Norvegicus

first: Debarshi

last: Mustafi

first: Krzysztof

last: Palczewski


title: mRNA sequencing of epidermal growth factor (EGF) treatment in 96, 108 and 120 hours of Stylonychia lemnae.

status: Public on Sep 11 2016

city: wuhan

first: Wei

last: Miao

state: hubei

address: donghu south road #7

institute: Institute of hydrobiology, CAS

summary: The goals of this study to find the differential expressed genes in epidermal growth factor (EGF) treated Stylonychia lemnae cells.

overall_design: Compare the expression of epidermal growth factor (EGF) treatment and control in Stylonychia lemnae cells.

first: Ying

last: Chen


title: CAGE sequences from three developmental states in the microcrustacean Daphnia pulex.

city: Tempe

first: R. Taylor

last: Raborn

state: AZ

address: 727 E Tyler St

department: Center for Mechanisms of Evolution

institute: Arizona State University

summary:
We report the application of CAGE (Cap Analysis of Gene Expression) on collections of Daphnia pulex individuals representing three major developmental states.

This submission comes from a project of Michael Lynch and was funded by a grant from the National Institutes of Health entitled 'Population Genomics of Daphnia pulex' (Project Number: 1R01GM101672-01A1).

overall_design: Characteriziation of the TSS landscape and transcriptionally-active putative promoters in three developmental states: adult parthenogenic (asexual) females, pre-ephipial (sexual) females and adult males.

first: R

first: Ken

last: Spitze


title: Imprinted DNA methylation reconstituted at a non-imprinted locus

city: Ithaca

first: Paul

last: Soloway

address: 237 Tower Road

institute: Cornell

summary:
Background: In mammals, tight regulation of cytosine methylation is required for embryonic development and cellular differentiation. The trans-acting DNA methyltransferases that catalyze this modification have been identified and characterized; however, these proteins lack sequence specificity, leaving the mechanism of targeting unknown. A cis acting regulator within the Rasgrf1 imprinting control region (ICR) is necessary for establishment and maintenance of imprinted methylation at the locus. Here we investigate whether 3kb of sequence from the Rasgrf1 ICR is sufficient to direct appropriate imprinted methylation and target-gene expression patterns when ectopically inserted at the Wnt1 locus.
Results: The Rasgrf1 ICR at Wnt1 lacked somatic methylation when maternally transmitted, and was fully methylated upon paternal transmission, consistent with its behavior at the Rasgrf1 locus. It was unmethylated in the female germline and was enriched for methylation in the male germline, though not to the levels seen at the endogenous Rasgrf1 allele. Wnt1 expression was not imprinted by our construct, likely due to additional sequences being required for this function.
Conclusion: We have identified sequences that are sufficient for partial establishment and full maintenance of the imprinted DNA methylation patterns. Because full somatic methylation can occur without full gametic methylation, we infer that somatic methylation of the Rasgrf1 ICR is not simply a consequence of maintained gametic methylation.

overall_design:
Targeted bisulfite sequencing of 20 tail, 6 sperm, 2 oocyte, 2 extraembryonic tissue, 2 head, 2 spine, and 2 visceral organs DNA samples from Wnt1DR animals. Unconverted sequence for both the normal and the nested PCR  included for easy analysis.
Unconverted reference sequence: wnt1405_250bp_OG.txt
Unconverted reference for ooctye: wnt1271_250bp_OG.txt

first: David

last: Taylor


title: Identification of warm-sensitive neuron markers using PhosphoTRAP

first: Chan Lek

last: Tan

summary: RNA-Seq profiling identifies transcripts enriched in preoptic neurons activated by warm temperature challenge

overall_design: Animals are subject to warm temperatures during which ribosomal S6 protein is phosphorylated selectively in active neurons. Transcripts associated with phosphorylated ribosome is isolated by immunoprecipitation. Comparison of Immunoprecipitation and Input fraction mRNA

first: Chan

first: Zachary


title: mRNA profiling of mouse embryonic cells with activin or SB431542 treatment

status: Public on Sep 10 2016

city: Leiden

first: Peter

last: ten Dijke

address: Einthovenweg 20

institute: Leiden University Medical Center

summary: We analysed the mRNA expression profiles of control and VprBP knockdown mouse embryonic stem cells (mESCs) with or without Activin treatment or TGF-β/Activin inhibitor SB431542 challenge for 4 days during differentiation

overall_design: Comparation of mRNA expression profile between control and VprBP knockdown mESCs upon Activin/SB431542 treatment furing 4 days differentiation

first: Yihao

last: Li


title: ChIPseq to profile IFNg response in human primary monocytes

first: Lionel

last: Ivashkiv

summary: We characterized the regulation of 2 histone marks by IFNg in human monocytes

overall_design: Human primary monocytes were cultured for 24 hours with or without IFNg. ChIP was performed and the DNA product was subject to ChIPseq

first: Yu

last: Qiao

first: Eugenia

last: Giannopoulou

first: Yushan


title: Function of MLL4 in mouse embryonic stem cells and somatic cell reprogramming

first: Kai

last: Ge

address: 50 South Dr Rm 4154

summary: Enhancers control cell type-specific gene expression and direct cell fate transition. Enhancers are marked by H3K4me1/2. MLL4 (KMT2D) is a major enhancer H3K4me1/2 methyltransferase. Here we show in embryonic stem cells (ESCs), MLL4 associates with, but is dispensable for the maintenance of, active enhancers of ESC identity genes. As a result, MLL4 is dispensable for cell identity gene expression and self-renewal in ESCs. In contrast, MLL4 is essential for activation of de novo enhancers and induction of cell identity genes during ESC differentiation. Similarly, MLL4 is dispensable for maintaining fibroblast cell identity but is essential for reprogramming into induced pluripotent stem cells. These results indicate that while MLL4 is dispensable for maintaining cell identity, it controls cell fate transitions by orchestrating de novo enhancer activation.

overall_design:
MLL3-/-;MLL4flox/fox and MLL3-/-;MLL4-/- mouse embryonic stem cells(ESCs) are generated. ChIP-Seq of histone modificaitons, MLL4 and RNA-Seq are performed in undifferentiated ESCs and day4 differentiated embryoid bodies (D4EB).
MLL3-/-;MLL4flox/fox and MLL3-/-;MLL4-/- immortalized mouse embryonic fibroblasts(MEFs) are reprogrammed with lentiviral infection of OSKM. RNA-Seq are performed in MEFs and day20 programmed cells.

first: Chaochen

last: Wang

first: Shiliyang

last: Xu

first: Ji-Eun

last: Lee

first: Binbin

last: Lai

first: Weiqun

last: Peng


title: RNA-seq analysis of the transcriptional response to blue and red light in the extremophilic red alga, Cyanidioschyzon merolae

city: Ann Arbor

first: mehmet

last: tardu

address: 930 N University Ave

department: Chemistry

institute: University of Michigan

summary: Light is one of the main environmental cues that affects the physiology and behavior of many organisms. The effect of light on genome-wide transcriptional regulation has been well-studied in green algae and plants, but not in red algae. Cyanidioschyzon merolae is used as a model red algae, and is suitable for studies on transcriptomics because of its compact genome with a relatively small number of genes. In addition, complete genome sequences of the nucleus, mitochondrion, and chloroplast of this organism have been determined. Together, these attributes make C. merolae an ideal model organism to study the response to light stimuli at the transcriptional and the systems biology levels. Previous studies have shown that light significantly affects cell signaling in this organism, but there are no reports on its blue light- and red light-mediated transcriptional responses. We investigated the direct effects of blue and red light at the transcriptional level using RNA-seq. Blue and red light were found to regulate 35% of the total genes in C. merolae. Blue light affected the transcription of genes involved protein synthesis while red light specifically regulated the transcription of genes involved in photosynthesis and DNA repair. Blue or red light regulated genes involved in carbon metabolism and pigment biosynthesis. Overall, our data showed that red and blue light regulate the majority of the cellular, cell division, and repair processes in C. merolae.

overall_design: mRNA profiles of wild type C. merolae 10D cells were generated under blue light, red light and complete darkness conditions by deep sequencing, in two biological replicates, using Illumina MiSeq platform.

first: Mehmet

last: Tardu

first: Ugur

middle: M

last: Dikbas

first: Ibrahim

last: Baris

last: Kavakli


title: Gene expression data from NRAS-driven CNS-PNET zebrafish brain tumors and normal brain.

city: Salt Lake City

first: Rodney

last: Stewart

state: UT

address: 2000 Circle of Hope Dr

department: Oncological Sciences

institute: University of Utah

summary: Zebrafish CNS-PNET tumors were generated by activating NRAS in oligoneural precursor cells. Gene expression in the zebrafish brain tumors and normal zebrafish brain was analyzed by RNA-seq.

overall_design: RNA-seq was performed on 7 zebrafish brain tumors and 8 normal brain samples on Illumina HiSeq 2000 using 50 Cycle Single-Read Sequencing v3 kit.

first: Katarzyna

last: Modzelewska


title: Transcriptome comparison of oocytes treated with RNAi against Btg4 or with control siRNAs

city: Cambridge

first: Michał

last: Pasternak

state: - None -

address: Francis Crick Avenue

institute: MRC Laboratory of Molecular Biology

summary: The comparison of trancriptomes was part of the study by Pasternak et al. The goal was to check if BTG4 regulates mRNA polyadenylation during mouse oocyte meiosis. To test this we compared the abundancies of the polyadenylated trancripts in control and Btg4-depleted oocytes.

overall_design: 3 samples of 50 oocytes were collected for both groups

first: Balaji

last: Santhanam

first: Schuh

last: Melina


title: RNA-seq analysis of seasonal and individual variation in blood transcriptomes of healthy captive bottlenose dolphins

city: Johns Island

first: Jeanine

last: Morey

state: SC

address: 3419 Maybank Hwy, Ste B

department: Conservation Medicine

institute: National Marine Mammal Foundation

summary: Four captive bottlenose dolpins housed in Kona, Hawaii, were serially sampled at approximately monthly intervals over the course of one year, in order to establish baseline information on the content and variation of the dolphin blood transcriptome.

overall_design: Illumina based RNA-seq analyses were carried out on globin-depleted RNA from blood using the Ensembl dolphin genome as a guide.  The data set was interrogated for transcripts exhibiting circa-annual rhymthicity, differential expression by season or sex, or correlation to hematological parameters

middle: S

first: Frances

last: Van Dolah

first: Michelle

last: Campbell


title: Chd8 mediates cortical neurogenesis through transcriptional regulation of cell cycle and Wnt signaling genes

first: Li-Huei

last: Tsai

state: Massachusetts

department: Picower Institute for Learning and Memory

institute: MIT

summary: De novo mutations in CHD8 are strongly associated with autism spectrum disorder (ASD), however the underlying mechanisms remain elusive. Here we report that Chd8 knockdown during cortical development results in defective neural progenitor proliferation and differentiation that ultimately manifests in abnormal neuronal morphology and behaviors in adult mice. Transcriptome analysis revealed that on the one hand Chd8 stimulates the transcription of cell cycle genes, while on the other it precludes the induction of neural specific genes by regulating the expression of PRC2 complex components. Furthermore, knockdown of Chd8 disrupts the expression of key transducers of Wnt signaling, and enhancing Wnt signaling rescues the transcriptional and behavioral deficits caused by Chd8 knockdown. We propose that these roles of Chd8 and the dynamics of its expression during development help negotiate the fine balance between neural progenitor proliferation and differentiation. Together, these observations provide new insights into the etiology of ASD.

overall_design: In utero electroporation of cortical radial glial cells with either control or Chd8 shRNA. Cortices from 3-5 embryos were pooled per biological replicates (three biological replicates for Chd8 sh1, and two biological replicates for Chd8 sh2), followed by FAC sorting of GFP positive cells.

first: Omer

last: Durak

first: Fan

last: Gao


title: ChIP-seq of E4F1 in mouse embryonic stem cells (mESCs)

city: Montpellier

first: Claude

last: Sardet

address: 208 rue des Apothicaires

department: Inserm U1194 - Université de Montpellier - Institut Régional du Cancer de Montpellier (ICM)

institute: Institut de Recherche en Cancérologie de Montpellier (IRCM)

summary: E4F1 is a ubiquitously expressed zinc-finger protein of the Gli-Kruppel family that was first identified, more than 30 years ago, as a cellular target of the adenoviral oncoprotein E1A13S (Ad type V), required for transcriptional regulation of adenoviral genes. In order to decipher E4F1 cellular target genes, we performed chromatin immunoprecipitation of endogenous E4F1 in mouse embryonic stem cells. Both input and immunoprecipitated DNA were exhaustively sequenced and mapped on the mouse genome (mm9). Peak detection has been achieved by combining two peak calling algorithms. Intersection of the two E4F1 peak lists on each cell line were considered as E4F1 chromatin bound regions.

overall_design: Genome-wide mapping of E4F1 binding in mouse embryonic stem cells.

first: Olivier

last: Kirsh

first: Geneviève

last: Rodier


title: Transcriptomic assessment of PCB-resistance in Atlantic killifish (Fundulus heteroclitus) embryos from a marine Superfund site

status: Public on Sep 09 2016

city: Woods Hole

first: Sibel

last: Karchner

state: MA

address: 45 Water St

department: Biology

institute: Woods Hole Oceanographic Institution

summary: RNA from PCB-126 exposed 10 days post-fertilization (dpf) embryos were sequenced to characterize the chemical exposure response of killifish from a PCB-resistant population (NBH, New Bedford Harbor, MA, Superfund site) as compared to a reference (SC, Scorton Creek, MA) population.  Since the genome sequence was not available, we also performed shot-gun sequencing of RNA from 1-15 dpf embryos sampled every day from both populations to serve as a transcriptome assembly.  The results suggest that the NBH fish possess a gene regulatory defect that is not limited to a few target genes.  We detected multiple genes that were differentially expressed in these two populations.  This study was the first application of pyrosequencing technology to combine transcriptome characterization and gene expression profiling in a marine animal.

overall_design: We performed 454 RNA-sequencing of 6 killifish embryo samples. Four of these were from a pool of 10 days post-fertilization (dpf) embryos from Scorton Creek (Sandwich, MA) and New Bedford Harbor (New Bedford, MA) populations, exposed to either DMSO or PCB126. The remaining two samples were pooled untreated embryos sampled every day from 1 to 15 dpf from the same two populations.

middle: I

first: Mark

last: Hahn


title: CCAN Assembly Configures Composite Binding Interfaces to Promote Cross-Linking of Ndc80 Complexes at the Kinetochore

city: Vienna

first: Stefan

last: Westermann

address: Dr. Bohr-gasse 7

country: Austria

institute: Institute of Molecular Pathology

summary: We report that the binding of Cnn1 (CENP-T homolog in yeast) occurs at the core of the yeast centromere and depends on the Ctf3 complex.

overall_design: The centromere localization of Cnn1 at the centromere in the absence of Ctf3 protein and comparison of its localization with the centromere localization of Cse4

first: Gulsah

last: Pekgoz Altunkaya

first: Francesca

last: Malvezzi


title: Transcriptional Maintenance of Pancreatic Acinar Identity, Differentiation and Homeostasis by PTF1A [E18.5 RNA-Seq]

city: Dallas

first: Galvin

last: Swift

state: TX

address: 6000 Harry Hines Blvd

department: Molecular Biology

institute: U TX Southwestern Medical Center

summary: RNA-seq analysis of RNA from embryonic day 18.5 pancreas

overall_design: pancreas mRNA profiles of E18.5 C57Bl/6 mice were generated by deep sequencing using an Illumina Hiseq 2500.

first: Chinh

last: Hoang

first: Ana

last: Azevedo-Pouly

first: Raymond

middle: J

last: MacDonald


title: Three-dimensional retinal organoids from mouse pluripotent stem cells mimic in vivo development with enhanced stratification and rod photoreceptor differentiation

first: Matthew

last: Brooks

state: MD

address: 6 Center Dr Bldg 6, Rm 303

department: NEI

institute: NIH

summary: Generation of three-dimensional (3D) organoids with optic cup like structures from pluripotent stem cells has created opportunities for investigating mammalian retinal development in vitro. However, retinal organoids in culture do not completely reflect the developmental state and in vivo architecture of the rod-dominant mouse retina. To assess rod photoreceptor differentiation in retinal organoids, we took advantage of Nrl-GFP mice that show rod-specific expression of GFP directed by the promoter of leucine zipper transcription factor NRL. Using embryonic and induced pluripotent stem cells (ESCs and iPSCs, respectively) derived from the Nrl-GFP mouse, we were successful in establishing long-term retinal organoid cultures (up to day 35) using modified culture conditions (called High Efficiency Hypoxia Induced Generation of Photoreceptors in Retinal Organoids, or HIPRO). We demonstrate efficient differentiation of pluripotent stem cells to retinal structures, with over 70% of embryoid bodies forming optic vesicles at Day (D)7, >50% producing optic cups by D10, and a majority of these surviving until at least D35. The HIPRO organoids include distinct inner retina neurons in somewhat stratified architecture and mature Müller glia spanning the entire retina. Almost 70% of the cells in retinal organoids are rod photoreceptors that exhibit elongated cilia. Transcriptome profiles of GFP+ rod photoreceptors, purified from organoids at Day 25-35, demonstrate a high correlation to gene profiles of purified rods from mouse retina at postnatal day (P) 2 to 6, indicating their early state of differentiation. Our 3D retinal organoids thus closely mimic in vivo retinogenesis and provide an efficient in vitro model to investigate photoreceptor development and modeling disease pathology.

overall_design: ESC clones were isolated from the inner cell mass (ICM) of 3.5-day blastocysts of Nrlp-GFP transgenic mice on feeders of mouse embryonic fibroblast (MEF). ESCs and iPSCs (passage 15-25) were differentiated using a modified SFEBq protocol. Flow sorted cells of GFP+ photoreceptors from 18, 21, 25, 28, and 35 days post differentiation initiation were collected for expression profiling.  RNA-seq profiles of high quality total RNA (RNA Integrity number ≥7.5) from were obtained using Illumina platform. After trimming Illumina adapters, defined in TrueSeq3-PE-2.fa file, by Trimmomatic (version 0.36) from paired end data, each sample was quantified against Ensemble release 84 cDNA FASTA file by Kallisto (version 0.42.5) via a custom pipeline in Perl. The quantifications were then normalized and summarized in gene level by tximport with the option “countFromAbundance=lengthScaledTPM”. The count values from tximport were converted to count per million (CPM) via edgeR.

first: Holly

middle: Y

last: Chen

first: Koray

last: Kaya

first: Lijin

last: Dong

first: Anand

last: Swaroop


title: RNA sequencing of human fibroblasts after SUPT4H1 siRNA treatment

city: Boston

first: Nicholas

last: Kramer

address: 77 Avenue Louis Pasteur

department: Genetics

institute: Harvard Medical School

summary: Asses the transcriptome-wide changes associated with depletion of transcription elongation factor, SUPT4H1, in human fibroblasts. An expanded hexanucleotide repeat in C9orf72 is the most common genetic cause of amyotrophic lateral sclerosis and frontotemporal dementia (c9FTD/ALS). Therapeutics (e.g., antisense oligonucleotides) are being developed to target RNAs containing the expanded repeat sequence (GGGGCC); however, this approach is complicated by the presence of antisense strand transcription of expanded GGCCCC repeats. We found that targeting the transcription elongation factor, Spt4, selectively decreased production of both sense and antisense expanded transcripts, as well as their translated dipeptide repeat (DPR) products, and also mitigated degeneration in animal models. Inhibition of SUPT4H1, the human ortholog of Spt4, similarly decreased production of sense and antisense RNA foci as well as DPR proteins in c9ALS patient fibroblasts and neurons from patient-derived induced pluripotent stem cells, consistent with our finding that cerebellar DPR protein levels correlate with SUPT4H1 expression in c9FTD/ALS patients. Therapeutic targeting of a single factor to eliminate pathological features of c9FTD/ALS offers advantages over approaches that require targeting sense and antisense repeats separately.

overall_design: 2 independent fibroblast lines: 2 replicate non-targeting siRNA treatments and 2 replicate SUPT4H1 siRNA treatments per line. Grouping by treatment for differential expression analysis (4 control siRNA samples and 4 SUPT4H1 siRNA samples)

first: Aaron

last: Gitler

first: Julien

last: Couthouis


title: A novel synthetic jumonji H3K27 demethylase inhibitor GSK-J4 exert immunosuppressive activities

city: Ansan

first: Young Gyu

last: Chai

state: Gyeonggi-do

address: 55 Hanyangdaehak-ro

department: Molecualr & Life Science

country: South Korea

institute: Hanyang Univ.

summary: Using RNA-seq, we report that jumonji H3K27 demethylase inhibitor, GSK-J4, exerts potent anti-inflammatory effects on LPS-stimulated BV-2 microglial cells.

overall_design: Examination of effects of LPS-stimulated BV2 cells with or without GSKJ4 treatment, were generated by deep sequencing on an Illumina HiSeq 2000(101 cycles PE lane).

first: Jin Choul


title: Follicular CXCR5-expressing CD8 T cells curtail chronic viral infection [RNA-seq]

city: Chongqing

first: Ran

last: He

address: 30th Gaotanyan street

institute: Third Military Medical University

summary: In mice chronically infected with lymphocytic choriomeningitis virus (LCMV), we defined a subset of exhausted CD8+ T cells abundantly expressing chemokine receptor CXCR5. These CXCR5+ CD8+ T cells were preferentially localized in B cell follicles, expressing less inhibitory receptors while exhibiting more potent cytolytic activity compared to the CXCR5- subset. Furthermore, we identified Id2-E2A axis as the regulator for the generation of this subset. In line with mouse LCMV chronic infection, we also identified a CXCR5+ CD8+ T cell subset in human HIV patients, which negatively correlated with viral load. Moreover, when adoptively transferred to chronically infected recipients, CXCR5+ subset showed greater therapeutic potential than CXCR5- subset.

overall_design: RNA sequenceing of two CD8 T cell subsets in chronic infection

first: Gang

last: Wei

first: Lilin

last: Ye

first: Ting

last: Ni

first: Qiang

last: Bai

last: Shen

first: Miao

last: Han


title: Unprecedented life- and healthspan extension and genome preservation by diet restriction in DNA repair deficient progeroid Ercc1∆/- mice [RNA-seq]

city: Rotterdam

first: Akos

last: Gyenis

address: Wytemaweg 80

institute: ERASMUS MC

summary: In this study, we investigate the anti-aging response induced by dietary restriction (DR) on gene expression level. For this, we carried out Ribosomal RNA depleted Total RNA sequencing in 16 weeks old Ercc1∆/- ad libidum (AL), DR and wt mice.

overall_design: Total RNA was extracted from fresh liver samples from 16 weeks old Ercc1∆/- AL, DR and wt mice. Ribosomal RNA was depleted from the extracts by using RiboMinus kit (Ambion) then sequenced  according to the Illumina TruSeq v3 protocol on HiSeq2000 platform.

first: Joris

last: Pothof

first: Wilbert

middle: P

last: Vermeij

first: Martijn

middle: E

last: Dollé

first: Jan

last: Hoeijmakers


title: Context-specific inhibition of translation by ribosomal antibiotics targeting the peptidyl transferase center

status: Public on Sep 08 2016

city: Nashville

first: James

last: Marks

state: Tennessee

address: 638A Robinson Research Building

institute: NIAMS/NIH

summary: We report the context-specific activity of two peptidyl transferase targeting antibiotics, chloramphenicol and linezolid. By generating ribosome profiling data in the presence or absence of either chloramphenicol or linezolid we mapped the relative change of ribosome density induced by these antibiotics. We find that both inhibitors preferentially arrest ribosomes that carry either an alanine, serine, or threonine in the penultimate (-1) position of the nascent peptide chain. Additionally ribosomes that carry a glycine in either the P site (0) or A-site (+1) counteract the inhibitory activity of both inhibitors. The context-specific action of chloramphenicol illuminates the operation of the mechanism of inducible resistance that relies on programmed drug-induced translation arrest. In addition, our findings expose the functional interplay between the nascent chain and the peptidyl transferase center.

overall_design: Ribosome profiling was performed on BWDK (-tolC) cells treated with a high concentration of either inhibitor and compared to a no antibiotic control.

first: Krishna

last: Kannan

first: Emily

last: Roncase

first: Cédric

last: Orelle

first: Amira

last: Kefi

first: Dorota

last: Klepacki

first: Nora

last: Vázquez-Laslop

first: Alexander

last: Mankin


title: Comparison between immune cells from mice challenged with Bacille Calmette Guérin (BCG)

city: Urbana

first: Sandra

last: Rodriguez Zas

state: IL - Illinois

address: 1207 W. Gregory Dr.

institute: University of Illinois at Urbana

summary: Microglia and macrophages are immune cells that differ in the inflammatory response to immune challenge.  The purpose of this study was to  profile mRNA levels (transcriptome) to better understand differences between immune cells 7 days after BCG challenge

overall_design: total RNA samples were obtained from 12 mice 7 days after  and immune cell type and were subjected to paired-end RNA sequencing


title: MiR-196b is epigenetically silenced during the pre-malignant stage of lung carcinogenesis

first: Adam

last: Gower

address: 72 East Concord Street, E632

department: Department of Medicine

institute: Boston University School of Medicine

summary: MicroRNA silencing by promoter hypermethylation may represent a mechanism by which lung cancer develops and progresses, but the microRNAs involved during malignant transformation are unknown. We previously established a model of pre-malignant lung cancer wherein we treated human bronchial epithelial cells (HBEC) with low doses of tobacco carcinogens. Here, we demonstrate that next-generation sequencing of carcinogen-transformed HBECs treated with the demethylating agent 5-aza-2'deoxycytidine revealed miR-196b and miR-34c-5p to be epigenetic targets. Bisulfite sequencing confirmed dense promoter hypermethylation indicative of silencing in multiple malignant cell lines and primary tumors. Chromatin immunoprecipitation studies further demonstrated an enrichment in repressive histone marks on the miR-196b promoter during HBEC transformation. Restoration of miR-196b expression by transfecting transformed HBECs with specific mimics led to cell cycle arrest mediated in part through transcriptional regulation of the FOS oncogene, and miR-196b re-expression also significantly reduced the growth of tumor xenografts. Luciferase assays demonstrated that forced expression of miR-196b inhibited the FOS promoter and AP-1 reporter activity. Finally, a case-control study revealed that methylation of miR-196b in sputum was strongly associated with lung cancer (OR = 4.7, p<0.001). Collectively, these studies highlight miR-196b as a tumor suppressor whose silencing early in lung carcinogenesis may provide a selective growth advantage to pre-malignant cells. Targeted delivery of miR-196b could therefore serve as a preventive or therapeutic strategy for the management of lung cancer.

overall_design: Nine tumor-derived cell lines (H1568, H1975, H2023, H2170, H23, CALU6, SW900, H1993, and H2085), HBEC1MBT, and HBEC2MBT were treated with vehicle or 5-aza-2’deoxycytidine and subjected to miRNA NGS.

first: Carmen

last: Tellez

first: Daniel

last: Juri

first: Kieu

last: Do

first: Maria

middle: A

last: Picchi

first: Teresa

last: Wang

first: Gang

last: Liu

first: Avrum

last: Spira

first: Steven

last: Belinsky


title: Transcriptomic characters of different pregnancy outcomes in CpG-induced nonobese diabetic mice and wild-type controls

city: Shanghai

first: Fei

last: Su

state: Shanghai

address: 800 Dongchuan Road

department: School of Life Science and Biotechnology

institute: Shanghai Jiao Tong University

summary: rna-seq was performed to reveal the profiles of differentially expressed genes in the maternal-fetal interface of wild-type and nod mice with or without CpG-induced intrauterine infection

overall_design: mRNA profiles of wild type (WT) and NOD mice were generated by deep sequencing without replicate

first: Xiaorui


title: The ERα Transcriptome

city: Los Angeles

first: Calvin

last: Pan

address: 675 Charles Young Dr South

institute: UCLA

summary: Impaired estrogen receptor α (ERα) action promotes obesity and metabolic dysfunction in humans and mice; however, the mechanisms underlying these phenotypes remain unknown. Considering that skeletal muscle is a primary tissue responsible for glucose disposal and oxidative metabolism, we established that reduced ERα expression in muscle is associated with glucose intolerance and adiposity in women and female mice. To test this relationship, we generated muscle-specific ERα knockout (MERKO) mice. Impaired glucose homeostasis and increased adiposity were paralleled by diminished muscle oxidative metabolism and bioactive lipid accumulation in MERKO mice. Aberrant mitochondrial morphology, overproduction of reactive oxygen species, and impairment in basal and stress-induced mitochondrial fission dynamics, driven by imbalanced protein kinase A–regulator of calcineurin 1–calcineurin signaling through dynamin-related protein 1, tracked with reduced oxidative metabolism in MERKO muscle. Although muscle mitochondrial DNA (mtDNA) abundance was similar between the genotypes, ERα deficiency diminished mtDNA turnover by a balanced reduction in mtDNA replication and degradation. Our findings indicate the retention of dysfunctional mitochondria in MERKO muscle and implicate ERα in the preservation of mitochondrial health and insulin sensitivity as a defense against metabolic disease in women.

overall_design: Quadriceps mRNA profiles of 20-week old wild type (WT) and muscle-specific ERα knockout (MERKO) mice were generated by deep sequencing, in triplicate, using Illumina HiSeq 2000.

first: Andrea

last: Hevener


title: N-Myc induces an EZH2-mediated transcriptional program driving Neuroendocrine Prostate Cancer

city: New York

first: Etienne

last: Dardenne

state: NY

address: 1300 York Avenue

department: Pathology

institute: Weill Cornell Medecine

summary: The transition from castration resistant prostate adenocarcinoma (CRPC) to neuroendocrine prostate cancer (NEPC) is emerging as an important mechanism of treatment resistance. NEPC are associated with over-expression and gene amplification of MYCN (encoding N-Myc). N-Myc is a bona fide driver oncogene in several rare tumor types, but its role in prostate cancer progression is not well established.  Integrating a novel genetically engineered mouse model and human prostate cancer transcriptome data, we show that N-Myc over-expression leads to the development of poorly differentiated, invasive prostate cancer that is molecularly similar to human NEPC tumors which includes an abrogation of AR signaling and induction of Polycomb Repressive Complex 2 signaling and that N-Myc interacts with AR and this interaction depends on Enhancer of Zeste Homolog 2. Altogether, our data shows that N-Myc drives the neuroendocrine phenotype in prostate cancer.

overall_design: RNA sequencing of mice prostates or human cell lines expressing N-Myc; ChIp-Seq of H3K27me3 +/- N-Myc in human cell line; RNA sequencing of human cell line expressing N-Myc after EZH2 knockdown or treatment with GSK343.

first: David

last: Rickman

first: Kaitlyn

last: Gayvert


title: Accessory subunits are integral for assembly and function of human mitochondrial complex I

city: Clayton

first: Traude

last: Beilharz

state: VIC

address: Wellington Rd

department: Biomedicine Discovery Institute

institute: Monash University

summary: For analysis of mRNA expression levels, total RNA was harvested from each cell-line in replicate with Trizol™ (Thermo scientific). Total RNA was purified using Direct-zol™ columns according to the manufacturers specifications (Zymo Research). For cDNA synthesis 1 μg of total RNA was process as the T12VN-PAT assay (Jänicke et al., RNA 2012), except that this was adapted for multiplexing on the Illumina MiSeq instrument. We refer to this assay as mPAT for multiplexed PAT. The approach is based on a nested-PCR that sequentially incorporates the Illumina platform’s flow-cell specific terminal extensions onto 3’ RACE PCR amplicons. First, cDNA was generated using the anchor primer mPAT Reverse, next this primer and a pool of 50 gene-specific primers were used in 5 cycles of amplification. Each gene-specific primer had a universal 5’ extension (see supplementary file primers) for sequential addition of the 5’ (P5) Illumina elements. These amplicons were then purified using NucleoSpin columns (Macherey-Nagel), and entered into second round amplification using the universal Illumina Rd1 sequencing Primer and TruSeq indexed reverse primers from Illumina. Second round amplification was for 14 cycles. Note, that each experimental condition was amplified separately in the first round with identical primers. In the second round, a different indexing primer was used for each experimental condition. All PCR reactions were pooled and run using the MiSeq Reagent Kit v2 with 300 cycles (i.e. 300 bases of sequencing) according to the manufacturers specifications. Data were analysed using established bioinformatics pipelines (Harrison et al., RNA 2015)

overall_design: Quantitation of a custom set of amplicons for the Accessory subunits of human mitochondrial complex I.

first: Michael

last: Ryan

first: Paul

last: Harrrison


title: ChIP-seq-based analysis of differential K27me3 coverage in neuroblastoma cell lines treated with epigenetic drugs

city: Heidelberg

first: Chunxuan

last: Shao

address: Im Neuenheimer Feld 280

country: Germany

institute: German Cancer Research Center

summary: The impact of drugs inhibiting DNA methylation (5-aza-2'-deoxycytodine, DAC) and EZH2 (EPZ-6438) on H3K27me3 coverage was analyzed in two neuroblastoma cell lines. Parallel analyses investigated associated changes in RNA expression and DNA methylation.

overall_design: The neuroblastoma cell lines Be(2)-C and IMR5-75 were treated with a combination of DAC and EPZ-6438. Controls were treated with solvent (DMSO). H3K27me3 ChIP seq was done to investigate treatment-related changes of this mark. In addition, H3K4me3 and H3K27ac ChIP seq was done in DMSO treated samples to identify putative regulatory regions.

first: Kai

middle: O

last: Henrich

first: Sebastian

last: Bender

first: Maral

last: Saadati

first: Moritz

last: Gartlgruber

first: Carl

last: Herrmann

first: Martha

last: Parzonka

first: Daniel

last: Dreidax

first: Matthias

last: Fischer

last: Duffy

first: Peter

last: Schmezer

first: Christoph

last: Plass

first: Thomas

last: Höfer

first: Axel

last: Benner

first: Stefan

last: Pfister

first: Frank

last: Westermann


title: ChIP-seq-based analysis of i) K27me3 promoter coverage and  ii) enhancer elements in neuroblastoma cell lines

summary: To investigate whether a set of differentially methylated/expressed genes, identified in neuroblastomas of differential outcome, co-localizes with K27me3 marks or enhancer elements, we performed H3K4me3, H3K4me1, H3K27ac and H3K27me3 ChIP seq in neuroblastoma cell lines.

overall_design: Enhancers were defined in neuroblastoma cell lines as overlapping H3K4me1 and H3K27ac peaks with a minimal distance of 2 kb to the closest H3K4me3 peak, a criterion imposed to rule out selection of (unannotated) promoters. Promoter-near K27me3 coverage was also analyzed in neuroblastoma cell lines. The effect of MYCN on K27me3 occupancy was investigated upon  modulation of MYCN levels.

first: David


title: AKAP95 regulates splicing through scaffolding RNAs and RNA processing factors

city: New York City

first: Alireza

last: Khodadadi-Jamayran

address: 550 1st Ave, MSB 304

department: Division of Advanced Research Technologies (DART)

institute: New York University, NYU Langone Medical Center

summary: Alternative splicing of pre-mRNAs significantly contributes to the complexity of gene expression in higher organisms, but the regulation of the splice site selection remains incompletely understood. We have previously demonstrated that a chromatin-associated protein, AKAP95 (AKAP8), has a remarkable activity in enhancing chromatin transcription. In this study, we have shown that AKAP95 physically interacts with many factors involved in transcription and RNA processing, and functionally regulates pre-mRNA splicing. AKAP95 directly promotes splicing in vitro and the inclusion of a specific exon of an endogenous gene FAM126A. The N-terminal YG-rich domain of AKAP95 is important for its binding to RNA processing factors including selective groups of hnRNP proteins, and its zinc finger domains are critical for pre-mRNA binding. Genome-wide binding assays revealed that AKAP95 bound preferentially to proximal intronic regions on a large number of pre-mRNAs in human transcriptome, and AKAP95 depletion predominantly resulted in reduced inclusion of many exons. AKAP95 also selectively coordinates with hnRNP H/F and U proteins in regulating alternative splicing events. We have further shown that AKAP95 directly interacts with itself. Taken together, our results establish AKAP95 as a novel and mostly positive regulator of pre-mRNA splicing and a possible integrator of transcription and splicing regulation, and support a model that AKAP95 regulates pre-mRNA splicing via through scaffolding RNAs and RNA processing factors and facilitating the splice site communication.

overall_design: Samples 1-8 are RNA-immunoprecipitation (RIP)-seq to determine AKAP95 binding to the transcriptome. Samples 9-15 are mRNA-seq to determine effect of AKAP95 knockdown in human 293 cells (9-11) or mouse ES cells (12-15).

first: Jing

last: Hu

first: Hao

last: Jiang


title: Dpy30 Is Critical for Maintaining the Identity and Function of Adult Hematopoietic Stem Cells

summary: As the major histone H3K4 methyltransferases in mammals, the Set1/Mll complexes play important roles in animal development and are increasingly associated with diseases including hematological malignancies. The role of H3K4 methylation activity of these complexes, however, remains elusive in fate determination of hematopoietic stem and progenitor cells (HSCs and HPCs). Here we address this question by generating a conditional knockout mouse for Dpy30, which is a common core subunit of all Set1/Mll complexes and facilitates genome-wide H3K4 methylation in cells. Dpy30 loss in the adult hematopoietic system results in severe pancytopenia but striking accumulation of HSCs and early HPCs that are defective in multilineage reconstitution, suggesting a differentiation block. In mixed bone marrow chimeras, Dpy30-deficient HSCs cannot differentiate or efficiently upregulate lineage-regulatory genes, and eventually fail to sustain for long term with significant loss of HSC signature gene expression. Our molecular analyses reveal that Dpy30 directly regulates H3K4 methylation and expression of key transcriptional factors, cofactors, and chromatin modulators involved in HSC function. Collectively, our results establish a critical role of Dpy30 and the H3K4 methylation activity of the Set1/Mll complexes for maintaining the identity and function of adult hematopoietic stem cells.

overall_design: Samples 1-4 are RNA-seq data to determine expression changes from LSK to MyePro cells in control versus Dpy30 KO background from chimeras; samples 5-8 are low-amount RNA-seq data to determine gene expression in control versus Dpy30 KO LT-HSCs from chimeras; and samples 9-14 are ChIP-seq data to determine H3K4me3 and H3K27me3 in control versus Dpy30 KO Lineage negative bone marrow cells.

first: Zhenhua

last: Yang

first: Kushani

last: Shah


title: RNA expression analysis of neuroblastoma cell lines treated with epigenetic drugs

first: Chunxuan

last: Shao

summary: The impact of drugs inhibiting DNA methylation (5-aza-2'-deoxycytodine, DAC) and EZH2 (EPZ-6438) on the neuroblastoma transcriptome was analyzed in two neuroblastoma cell lines. Parallel analyses investigated associated changes in histone modification and DNA methylation.

overall_design: The neuroblastoma cell lines Be(2)-C and IMR5-75 were treated with DAC and EPZ-6438 in combination or alone. Controls were treated with solvent (DMSO). RNA was isolated and sequenced.

first: Kai

last: Henrich

first: Sebastian

last: Bender

first: Maral

last: Saadati

first: Moritz

last: Gartlgruber


title: Global transcriptome analysis in the MYCN-amplified neuroblastoma cell line IMR5-75 upon inducible MYCN-knockdown

summary: Inducible MYCN-knockdown, followed by RNA-seq analysis in the MYCN-amplified neuroblastoma cell line IMR5-75, reveals profound time-dependent transcriptome changes.

overall_design: For modulation of MYCN levels, stable neuroblastoma cell models were used where MYCN can be  downregulated via vector-based hairpin RNA induction upon addition of 1µg/ml tetracycline (IMR5-75-shMYCN. From cells treated either with tetracycline or solvent (ethanol), RNA was isolated at time points 6 hours, 12 hours and 24 hours. Experiments were done in duplicates. RNA was sequenced.

first: Carl

last: Herrmann

first: Martha

last: Parzonka

first: Daniel

last: Dreidax

first: Matthias

last: Fischer

last: Duffy

first: Peter


title: RNA Sequencing Facilitates Quantitative Analysis of Primary Microglia Transcriptomes II

first: Young Gyu

last: Chai

summary: Using RNA-seq, we report here that primary microglia (PM) cells have a distinct transcriptomic signature and express a unique cluster of transcripts in response to 2hrs or 4 hrs with LPS.

overall_design: Examination of effects of LPS-stimulated PM microglial cells, were generated by deep sequencing on an Illumina HiSeq 2000(101 cycles PE lane).

first: Young-Gyu

first: Jin-Choul