title: Argonaute CLIP defines a deregulated miR-122 bound transcriptome that correlates with patient survival in human liver cancer (Mouse CLIP-Seq)

city: New York

first: Joseph

last: Luna

state: NY

address: 1230 York Ave #64

institute: The Rockefeller University

summary: MicroRNA-122, an abundant and conserved liver-specific miRNA, regulates hepatic metabolism and functions as a tumor suppressor, yet systematic and direct biochemical elucidation of the miR-122 target network remains incomplete. To this end, we performed Argonaute crosslinking immunoprecipitation (Ago-CLIP) sequencing in miR-122 knockout and control mouse livers, as well as in matched human hepatocellular carcinoma (HCC) and benign liver tissue to identify miRNA target sites transcriptome-wide in two species. We observed a majority of miR-122 binding on 3’-UTRs and coding exons followed by extensive binding to other genic and non-genic sites. Motif analysis of miR-122 dependent binding revealed a novel G-bulged motif in addition to canonical motifs. A large number of miR-122 targets were found to be species-specific. Upregulation of several common mouse and human targets, most notably BCL9, predicted survival in HCC patients. These results broadly define the molecular consequences of miR-122 downregulation in hepatocellular carcinoma.

overall_design: AGO CLIP-seq libraries were generated from four miR-122 KO mice and five floxed littermate controls, all at six weeks of age.

first: Juan

last: Barajas

first: Kun-yu

last: Teng

first: Hui-Lung

first: Michael

last: Moore

first: Charles

last: Rice

first: Robert

middle: B

last: Darnell

first: Kalpana

last: Ghoshal


title: Argonaute CLIP defines a deregulated miR-122 bound transcriptome that correlates with patient survival in human liver cancer (Mouse RNA-Seq)

status: Public on Mar 28 2017

middle: M

country: USA

type: Expression profiling by high throughput sequencing

last: Sun

middle: J


title: Next Generation Sequencing Facilitates Quantitative Analysis of Ern1f/f and Ern1f/fLyz2-Cre macrophage Transcriptomes

city: Wuhan

first: Yong

last: Liu

state: Hubei

address: Wuhan 430072, China

country: China

institute: Wuhan University

summary:
Purpose: IRE1 are indicated to play important role in LPS-induced macrophage activation. The goals of this study are to investigate the effects of IRE1a abrogation in macrophages on Interleukin-4-induced alernative activation of macrophages.
Methods: Total cellular RNAs of M2-polarized BMDMs by IL-4 treatment were isolated and subjected to commercial RNA-seq analysis. RNA-seq data were uniquely mapped to mm10 reference genome by TopHat v1.4.1. Expression values were assigned to gene level by Cufflinks v1.3.0.
Results: Map ratios of all RNA-seq samples were above 80%. Merged expression values of all samples included 10,863 genes. Principle component analysis showed distinguished patterns between groups.
Conclusions: Our study represents detailed analysis of transcriptomes of IL-4-activated macrophages, with biologic replicates, generated by RNA-seq technology. Based on the analysis of our RNA-seq data, increased expression levels of marker genes of macrophage alternative activation were found in IRE1a-abrogated macrophages copared to control cells. Our results demonstrates the loss of ER stress sensor IRE1a in macrophages enhanced IL-4-induced alternative activation.

overall_design: Macrophage mRNA profiles of 8-week old Ern1f/f and Ern1f/fLyz2-Cre mice were generated by deep sequencing, in duplicate, using Illumina HiSeq 2000

first: Bo

last: Shan

first: Chi

last: Xu


title: The RNA helicase DDX39B regulates IL7R alternative splicing reducing the risk of Multiple Sclerosis

city: Galveston

first: Mariano

last: Garcia-Blanco

address: 301 University Blvd

institute: The University of Texas Medical Branch

summary:
Purpose: The goal of this study is to investigate the role of DDX39B in RNA splicing
Methods: RNA-Seq splicing analysis of HeLa cells with experimentally induced DDX39B expression knockdown

overall_design: DDX39B expression is reduced in HeLa cells by siRNAs. Effects on alternative RNA splicing are compared between knockdown and control cells.

first: Gaddiel

last: Galarza-Muñoz

first: Farren

last: Briggs

first: Irina

last: Evsyukova

first: Geraldine

middle: O

last: Schott-Lerner

first: Edward

last: Kennedy

first: Tinashe

last: Nyanhete

first: Liuyang

last: Wang

first: Laura

last: Bergamaschi

first: Steven

middle: G

last: Widen

first: Georgia

middle: D

last: Tomaras

first: Dennis

middle: C

last: Ko

first: Shelton

middle: S

last: Bradrick

first: Lisa

middle: F

last: Barcellos

first: Simon

last: Gregory

middle: A


title: DHX9 suppresses spurious RNA processing defects originating from the Alu invasion of the human genome [XL9 DHX9 FLASH CLIP-seq]

city: Freiburg

first: Asifa

last: Akhtar

address: Stuebeweg 51

department: Chromatin Regulation

country: Germany

institute: Max Planck Institute of Immunobiology and Epigenetics

summary: Transposable elements increase genetic diversity thus making them an important part of evolution and gene regulation in all organisms that carry these sequences. Bulk of our nascent transcriptome is comprised of transposable elements that have the propensity to form strong secondary structures. It is essential to resolve such strong secondary structures to maintain normal cellular function. Here, we show that the major nuclear RNA helicase DHX9/RHA interacts and remodels embedded Alu retrotransposable elements in the human transcriptome and B1 retrotransposable elements in the mouse transcriptome. To understand the function of DHX9 we used FLASH (Fast cloning of RNA After some Sort of affinity purification for High-throughput sequencing) to identify the in-vivo targets of human DHX9.

overall_design: FLASH (Fast cloning of RNA After some Sort of affinity purification for High-throughput sequencing) for mouse DHX9 and one control (IgG-Rb); two biological replicates each.

first: Ibrahim

last: Ilik

first: Daniel

last: Maticzka


title: Argonaute CLIP defines a deregulated miR-122 bound transcriptome that correlates with patient survival in human liver cancer (Human CLIP-Seq)

first: Joseph

last: Luna

first: Juan

last: Barajas

first: Kun-yu

last: Teng

first: Hui-Lung

last: Sun

first: Michael

last: Moore

first: Charles

last: Rice

first: Robert

last: Darnell

first: Kalpana

last: Ghoshal


title: RNAseq of ∆yacP (∆rae1) mutant and plasmid complemented strain

city: Paris

first: Ciaran

last: Condon

address: 13 rue Pierre et Marie Curie

country: France

institute: CNRS

summary: To identify the substrates of YacP (Rae1) by determine which RNAs are affected in a ∆yacP mutant and plasmid complemented strain

overall_design: 4 strains were sequenced in triplicate: SSB1002 (W168 wild-type), CCB375 (W168 ∆yacP), CCB604 (W168 ∆yacP + pDG-yacP) + IPTG, CCB604 (W168 ∆yacP + pDG-yacP) - IPTG

first: Magali

last: Leroy

first: Caroline

last: Proux

first: Jean-Yves

last: Coppée


title: Multi-tissue DNA methylation age predictor in mouse

city: Cambridge

first: Felix

last: Krueger

address: Granta Park

department: Bioinformatics

country: United Kingdom

institute: Altos Labs

type: Methylation profiling by high throughput sequencing

summary: Background: DNA-methylation changes at a discrete set of sites in the human genome are predictive of chronological and biological age. However, it is not known whether these changes are causative or a consequence of an underlying ageing programme. It has also not been shown whether this ‘epigenetic clock’ is unique to humans or conserved in other animals such as the experimentally tractable mouse.   Results: We have generated a comprehensive set of whole genome base-resolution methylation maps from multiple mouse tissues spanning a wide range of ages. A large number of CpG sites show significant tissue independent correlations with age and allowed us to develop a multi-tissue predictor of age in the mouse (‘mouse epigenetic clock’). The predictor, which estimates age based on DNA methylation at 644 unique CpG sites, is highly accurate (mean absolute error of 4.09 weeks) and has similar properties to the recently described human epigenetic clock. Using publicly available datasets from various biological manipulations in mice, we found that the mouse clock also measures biological age. While females and males showed no significant differences in predicted DNA methylation age, ovariectomy resulted in significant age acceleration in females. Furthermore we found significant differences in age-acceleration dependent on the lipid content of the maternal or offspring diet.  Conclusions: Here we identify and characterise a highly accurate epigenetic predictor of age in mice, the ‘mouse epigenetic clock’. This clock will be instrumental for understanding the biology of ageing and will allow modulation of its ‘ticking’ rate  and resetting the clock in vivo to study the impact on biological age.

overall_design: RRBS-seq of different tissues from mice at different ages

first: Thomas

last: Stubbs

first: Ferdinand

last: von Meyenn

first: Anne

last: Katrien-Stark

first: Wolf

last: Reik


title: DHX9 suppresses spurious RNA processing defects originating from the Alu invasion of the human genome [XL1 DHX9 FLASH CLIP-seq]

first: Asifa

last: Akhtar

overall_design: FLASH (Fast cloning of RNA After some Sort of affinity purification for High-throughput sequencing) for human DHX9 using two different antibodies (DHX9-mAb and DHX9-Rb) and one control (IgG-Rb); two biological replicates each.

first: Ibrahim

last: Ilik

first: Daniel

last: Maticzka


title: DHX9 suppresses spurious RNA processing defects originating from the Alu invasion of the human genome [XL8 DHX9 FLASH CLIP-seq]


title: The m6A Demethylase ALKBH5 Maintains Tumorigenicity of Glioblastoma Stem-Like Cells by Sustaining FOXM1 Expression and Cell Proliferation Programing

city: Chicago

first: zhike

last: lu

state: IL

address: 5801 South Ellis Avenue

department: department of chemistry

institute: University of Chicago

summary: The dynamic and reversible N6-methyladenosine (m6A) RNA modification installed and erased by N6-methyltransferases and demethylases regulates gene expression and cell fate. Here, we show that the m6A demethylase ALKBH5 is highly expressed in glioblastoma stem-like cells (GSCs). Silencing ALKBH5 suppresses the proliferation of patient-derived GSCs in vitro and in vivo. Integrated transcriptome and m6A-seq analyses revealed altered expression of select ALKBH5 target genes, including FOXM1, a critical transcription factor for the ALKBH5-dependent cell cycle gene expression. ALKBH5 binds to and demethylates FOXM1 nascent transcripts, leading to enhanced FOXM1 expression. Further, a long noncoding RNA antisense to the FOXM1 (FOXM1-AS) interacts with ALKBH5 and FOXM1 nascent transcripts and promotes their interaction. Depleting ALKBH5 and FOXM1-AS disrupted GSC tumorigenesis through the FOXM1 axis. Our work uncovers a novel function for ALKBH5 in maintaining GSC tumorigenicity and provides insight into critical roles of the m6A RNA methylation in human brain tumor.

overall_design: m6A-seq and anti-m6A RIP-seq of mRNA from control and shALKBH5 cells

first: Boxuan

last: Zhao

first: Chuan

last: He


title: RNA profiling of four DLD1 colorectal cancer cell derivatives that recapitulate EMT/MET transitions during metastasis.

city: Munich

first: Matjaz

last: Rokavec

address: Thalkirchner Str. 36

department: Institute of Pathology

institute: Ludwig-Maximilians-University Munich

summary: RNA profiling of a cellular model of colorectal cancer progression, which is based on four defined derivates of a CRC cell line that resulted from EMT/MET transitions and phenotypically recapitulate the metastatic cascade: epithelial-like primary tumor, mesenchymal-like tumor cells growing in soft agar or as colonospheres, and mouse xenografts of the latter resulting in lung-metastasis comprised of epithelial-state cells.

overall_design: Colorectal cancer cell line DLD1 and its derivatives were maintained in McCoy`s 5A Medium (Invitrogen) containing 10% fetal bovine serum (FBS, Invitrogen), 100 units/ml penicillin and 0.1 mg/ml streptomycin.

first: Heiko

last: Hermeking


title: Testing the Ret and Sema3d genetic interaction in mouse enteric nervous system development

city: Houston

first: Ashish

last: Kapoor

state: TX

address: 1825 Pressler St. Rm SRB 530C

institute: University of Texas Health Science Center at Houston

summary: For most multigenic disorders, clinical manifestation (penetrance) and presentation (expressivity) are likely to be an outcome of genetic interaction between multiple susceptibility genes.  Here, using gene knockouts in mice we evaluated genetic interaction between loss of Ret and loss of Sema3d, two Hirschsprung disease (HSCR) susceptibility genes.  We intercrossed Ret and Sema3d double null heterozygotes to generate mice with the nine possible genotypes and assessed survival by counting various genotypes, myenteric plexus development by acetylcholinesterase (AchE) staining and embryonic day 12.5 (E12.5) gut transcriptome by RNA-sequencing.  Survival rates of Ret wildtype, null heterozygote and null homozygote mice at E12.5, birth and weaning were not influenced by the genotypes at Sema3d locus and vice-versa.  Loss of myenteric plexus was observed only in all Ret null homozygotes, irrespective of the genotypes at Sema3d locus, and Sema3d null heterozygote and homozygote mice had normal gut innervation.  As compared to wildtype mice gut gene expression, loss of Ret in null homozygotes led to differential expression of ~300 genes, whereas loss of Sema3d in null homozygotes had no major consequence and there was no evidence supporting major interaction between the two genes influencing gut transcriptome.  Overall, given the null alleles and phenotypic assays used, we did not find evidence for genetic interaction between Ret and Sema3d affecting survival, myenteric plexus formation or gut transcriptome.

overall_design: poly-A RNA-seq in embryonic day 12.5 mouse gut from 3 wildtype males, 3 wildtype females, 3 Ret null homozyogote males, 3 Ret null homozyogote females, 3 Sema3d null homozyogote males, 3 Sema3d null homozyogote females, 3 Ret-Sema3d double null homozyogote males, 3 Ret-Sema3d double null homozyogote females

first: Dallas

middle: R

last: Auer

first: Dongwon

last: Lee

first: Aravinda

last: Chakravarti


title: Cellular model of colon cancer progression reveals signatures of mRNAs, miRNA, lncRNAs and epigenetic modifications associated with metastasis.

type: Genome binding/occupancy profiling by high throughput sequencing

summary: H3K4me3, H3K27me3, and H3K79me3 profiling of a cellular model of colorectal cancer progression, which is based on four defined derivates of a CRC cell line that resulted from EMT/MET transitions and phenotypically recapitulate the metastatic cascade: epithelial-like primary tumor, mesenchymal-like tumor cells growing in soft agar or as colonospheres, and mouse xenografts of the latter resulting in lung-metastasis comprised of epithelial-state cells.

overall_design: H3K4me3, H3K27me3, and H3K79me3

first: David

last: Horst


title: DHX9 suppresses spurious RNA processing defects originating from the Alu invasion of the human genome [hnRNPC FLASH CLIP-seq]

overall_design: FLASH (Fast cloning of RNA After some Sort of affinity purification for High-throughput sequencing) for human hnRNPC and one control (IgG); two biological replicates each.


type: Methylation profiling by genome tiling array

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

overall_design: Refer to individual Series


title: DHX9 suppresses spurious RNA processing defects originating from the Alu invasion of the human genome [RNA-Seq]

summary:
Transposable elements increase genetic diversity thus making them an important part of evolution and gene regulation in all organisms that carry these sequences. Bulk of our nascent transcriptome is comprised of transposable elements that have the propensity to form strong secondary structures. It is essential to resolve such strong secondary structures to maintain normal cellular function. Here, we show that the major nuclear RNA helicase DHX9/RHA interacts and remodels embedded Alu retrotransposable elements in the human transcriptome and B1 retrotransposable elements in the mouse transcriptome.
To understand the effect of loss of DHX9 in human transcriptome we performed knockdown of DHX9 in HEK293 cells using two different siRNAs, followed by polyA-plus and polyA minus RNA extraction and sequencing.

overall_design:
We perfomed knockdown of human RNA heliase-A (also known as DHX9 ) in HEK293 cells using 2 different siRNAs, followed by polyA-plus and polyA-minus RNA sequencing.
siRNAs used:
control: silencer select siRNA control #2 (https://www.thermofisher.com/order/catalog/product/4390846)
DHX9 KD_1: s4019 (http://www.ncbi.nlm.nih.gov/probe/16735410) chr1: 182860161-182860180 (hg38)
DHX9 KD_2: s4020 (http://www.ncbi.nlm.nih.gov/probe/16735422) chr1: 182858158-182858177 (hg38)
siRNA were used at a final concentration of 5nM, transfected to HEK293 cells with RNAiMAX.
Total RNA was isolated 3d after transfection. For polyA(+) samples, TruSeq library prep protocol was used (starting with ~1µg total RNA)
For polyA(-) samples, we depleted polyA(+) RNA from total RNA with oligo-dT(+) beads, twice. The rest was first ribodepleted with RiboZero kit, which then went through the TruSeq protocol.
Sequencing parameters : 150x2 bp reads, full run of the NextSeq500 machine.

last: Iilk

first: Vivek

last: Bhardwaj


title: Argonaute CLIP defines a deregulated miR-122 bound transcriptome that correlates with patient survival in human liver cancer

first: Joseph

last: Luna

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus


title: The role of Tmub1 protein in the regulation of hepatic cell transcription

status: Public on Mar 27 2017

city: Chongqing

first: Xu

last: jianhua

address: Daping

institute: TMMU

summary: Tmub1 is involved in liver regeneration and functions as a bridging protein in tumor cell proliferation. To investigate the transcriptional profile and potential biological processes affected by Tmub1 expression in normal rat hepatocytes, microarray and bioinformatics experiments were used to identify 127 mRNAs differentially expressed between Tmub1-overexpression, Tmub1-knockdown and normal BRL-3A cells (fold change ≥ 2.5, P < 0.05).

overall_design: Tmub1-overexpression, Tmub1-knockdown, 2 Negative Control and normal  BRL-3A cells (fold change ≥ 2.5, P < 0.05). Five independent.

first: Jianhua

last: Xu

first: Ping

last: Chen


title: let-7 regulates radial migration of newborn neurons through positive regulation of autophagy

city: Lund

first: Rebecca

last: Petri

address: Sölvegatan 17

country: Sweden

institute: Lund University

type: Non-coding RNA profiling by high throughput sequencing

summary: During adult neurogenesis, newly formed olfactory bulb (OB) interneurons migrate radially to integrate into specific layers of the OB. Despite the importance of this process, the intracellular mechanisms that regulate radial migration remain poorly understood. Here we find that microRNA (miRNA) let-7 regulates radial migration by modulating autophagy in new-born neurons. Using Argonaute2-immunoprecipitation, we performed global profiling of miRNAs in adult-born OB neurons and identified let-7 as a highly abundant miRNA family. Knockdown of let-7 in migrating neuroblasts prevented radial migration and led to an immature morphology of newly formed interneurons. This phenotype was accompanied by a decrease in autophagic activity. Overexpression of Beclin-1 or TFEB in new-born neurons lacking let-7 resulted in re-activation of autophagy and restored radial migration. Thus, these results reveal a miRNA-dependent link between autophagy and adult neurogenesis with implications for neurodegenerative diseases where these processes are impaired.

overall_design: Small RNA sequencing data from Argonaute2-RIPseq on newborn neurons of the olfactoy bulb from LV.GFP-AGO2 and sham-injected control mice.

first: Karolina

last: Pircs

first: Marie

last: Jönsson

first: Malin

last: Åkerblom

first: Per

last: Ludvik Brattås

first: Thies

last: Klussendorf

first: Johan

last: Jakobsson


title: Epigenetically repressing human cytomegalovirus lytic infection and reactivation from latency in THP-1 model by targeting H3K9 and H3K27 histone demethylases

city: Shanghai

first: Shanshan

last: Zhu

address: 320 Yue-Yang Road

institute: CAS-MPG Partner Institute for Computational Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences

summary: Human Cytomegalovirus (hCMV) infects a broad range of the population and establishes life-long latency in the infected individuals. Periodically the latently infected virus can reactivate and becomes a significant cause of morbidity and mortality in immunocompromised individuals. Upon reactivation, the repressive chromatin is remodeled to an active form, allowing viral lytic gene transcription, initiated by the expression of viral Immediate Early (IE) genes. During this process, a number of histone modification enzymes, including histone demethylases (HDMs), play important roles in driving IE expression, but the mechanisms involved are not fully understood. To get a better understanding of these mechanisms, we profiled this well-established CMV experimental latency-reactivation model based on TPA (12-O-Tetradecanoylphorbol-13-acetate) induced THP-1 differentiation, which recapitulates many key features of CMV latency and reactivation in vivo, including the important role of viral chromatin in controlling viral IE gene expression.

overall_design: Transcritome profiling of THP-1 cell lines with or without TPA induction

first: Yu

last: Liang


title: Transcriptional response to hepatitis C virus infection and interferon alpha treatment in the human liver

city: Lyon

first: Anamaria

last: Necsulea

address: 43 Bd. du 11 Novembre 1918

department: Laboratoire de Biométrie et Biologie Evolutive

institute: CNRS, Université Claude Bernard Lyon 1

summary: Hepatitis C virus (HCV) is widely used to investigate host-virus interactions and cellular responses to infection have been extensively studied in vitro. In human liver, interferon (IFN) stimulated gene expression can mask direct transcriptional responses to virus infection. To better characterize the direct effects of HCV infection in vivo, we analyze the transcriptomes of HCV-infected patients lacking an activated endogenous IFN system. We show that the expression changes observed in these patients predominantly reflect immune cell infiltrates rather than changes in cell-intrinsic metabolic pathways. We also investigate the transcriptomes of patients with endogenous IFN activation, which paradoxically cannot eradicate viral infection. We find that most IFN-stimulated genes (ISGs) are induced by both the endogenous IFN system and by recombinant IFN therapy, but with significantly higher induction levels in the latter. We conclude that the innate host immune response in chronic hepatitis C is too weak to clear the virus.

overall_design: In this study, we aimed to disentangle the direct and indirect effects of HCV infection on cellular transcriptional profiles, by performing a detailed characterization of the gene expression changes associated with HCV infection, endogenous IFN system activation and pegIFNa treatment in the human liver. With this objective, we generated and analyzed high-throughput transcriptome sequencing profiles from liver biopsies derived from different categories of HCV-infected and non-infected patients, prior to and during treatment. First, to unveil HCV-induced cell-autonomous effects and to separate them from IFN-induced changes in the transcriptome, we selected liver biopsies from patients with chronic hepatitis C (CHC) without hepatic ISG induction, and compared them with un-infected control biopsies. Second, we examined the transcriptomic changes associated with the endogenous activation of the IFN system. Finally, we analyzed the gene expression changes resulting from pegIFNa/ribavirin treatment, by comparing transcriptome data from liver biopsies obtained before treatment and at different time points during the first week of therapy.

first: Tujana

last: Boldanova

first: Aleksei

last: Suslov

first: Markus

middle: H

last: Heim


title: Time-course analysis of microglia-specific RNA-seq in two severities of spinal cord injury

city: Geneva

first: Celine

last: Delucinge

address: 1 rue Michel Servet

department: iGE3 Genomics Platform

country: Switzerland

institute: University of Geneva

summary: RNA-seq analysis of microglia following spinal cord injury demonstrated that microglia response is driven by time after injury but not lesion severity.

overall_design: Microglia were purified by FACS in non-injured CX3CR1+/eGFP mice and at 3 time points following moderate and severe spinal cord lesion. Gene profiles were then compared.

first: Harun

middle: N

last: Noristani

first: Florence

middle: E

last: Perrin


title: Histone gene replacement reveals a post-transcriptional role for H3K36 in maintaining metazoan transcriptome fidelity

city: Chapel Hill

first: Michael

last: Meers

state: NC

address: 250 Bell Tower Dr.

institute: UNC Chapel Hill

summary: To study the function of this residue independent from the enzymes that modify it, we used a “histone replacement” system in Drosophila to generate a non-modifiable H3K36 lysine-to-arginine (H3K36R) mutant. Post-translational modification of histone H3K36 is neither required to suppress cryptic transcription initiation nor to include alternative exons in Drosophila; instead it promotes expression of active genes by stimulating polyadenylation.

overall_design: Examination of poly-A RNA-seq, total nuclear RNA-seq, Start-seq, and ATAC-seq from HWT and K36R rescue larvae

first: A

last: Matera


title: DHX9 suppresses spurious RNA processing defects originating from the Alu invasion of the human genome [uvCLAP CLIP-seq]

first: Asifa

last: Akhtar

summary: Transposable elements increase genetic diversity thus making them an important part of evolution and gene regulation in all organisms that carry these sequences. Bulk of our nascent transcriptome is comprised of transposable elements that have the propensity to form strong secondary structures. It is essential to resolve such strong secondary structures to maintain normal cellular function. Here, we show that the major nuclear RNA helicase DHX9/RHA interacts and remodels embedded Alu retrotransposable elements in the human transcriptome and B1 retrotransposable elements in the mouse transcriptome. To understand the function of DHX9 we used uvCLAP (UV crosslinking and affinity purification) to identify the in-vivo targets of human and mouse DHX9 and its drosophila ortholog MLE.

sample_organism:
Drosophila melanogaster
Homo sapiens
Mus musculus

overall_design: UV crosslinking and affinity purification (uvCLAP) for 16 proteins in 3 organisms.

first: Ibrahim

last: Ilik

first: Daniel

last: Maticzka


title: Transcriptome analysis of human immortilized astrocytes reprogrammed into dopaminergic neurons

city: Stockholm

first: Sten

last: Linnarsson

address: Scheeles väg 1

department: Medical Biochemistry and Biophysics

institute: Karolinska Institutet

summary: We directly reprogram human astrocytes (hIA) in vitro into induced dopaminergic neurons (iDAs). This process requires three transcription factors, NEUROD1, ASCL1 and LMX1A as well as one microRNA, mir218. This combined protocol gives rise to human astrocyte-derived iDAs with appropriate midbrain markers as defined by analyzing markers derived from human embryonic ventral midbrain samples at 7 and 9 weeks of age.

overall_design: Four replicates of the uninduced astrocyte cell line, four replicates of astrocytes induced into dopaminergic cells, and two samples each of human embryonic midbrain tissue at 7 and 9 weeks of age

first: Pia

last: Rivetti di Val Cervo

first: Ernest

last: Arenas


title: WT1 expression in breast cancer disrupts the epithelial/mesenchymal balance of tumour cells and correlates with the metabolic response to docetaxel

city: Midlothian

first: Peter

last: Hohenstein

address: Easter Bush Campus

department: The Roslin Institute

institute: University of Edinburgh

summary: The Wilms' Tumour gene 1 (WT1), encodes for a complex protein with transcription factor activity which is essential in mammals throughout life. We provide a complete study of WT1 expression across different breast cancer subtypes as well as isoform specific expression analysis. Using in vitro cell lines, clinical samples and publicly available gene expression datasets, we demonstrate that WT1 plays a role in regulating the epithelial-mesenchymal balance of breast cancer cells and that WT1-expressing tumours are mainly associated with a mesenchymal phenotype. WT1 gene expression also correlates with CYP3A4 levels and is associated with poorer response to taxane treatment.

overall_design: RNA profiles of breast cancer cells (MDA-MB-157) were generated by deep sequencing on the Illumina HiSeq 2000 platform. Untreated MDA-MB-157 cells, MDA-MB-157 cells transduced with a lacZ control vector, and MDA-MB-157 cells transduced with a lentiviral vector carrying a Wt1 shRNA were sequenced (titled untreated, lacZ and Wt1 respectively).

first: Mara

last: Artibani

first: Stuart

last: Aitken


title: Bach2 regulates AID-mediated immunoglobulin gene conversion and somatic hypermutation in DT40 B cells

city: Turku

first: Paulina

last: Budzynska

address: Kiinamyllynkatu 13

country: Finland

institute: University of Turku

summary: The transcription factor Bach2 is required for germinal center formation and somatic hypermutation (SHM) of immunoglobulins, both central to an efficient antibody-mediated immune response. Activation-induced cytidine deaminase (AID) initiates SHM and CSR in germinal centers and has potential to induce human B cell lymphoma. To understand the role of Bach2 in AID-mediated immunoglobulin gene diversification processes, we established a Bach2-deficient DT40 B cell line. We show that in addition to allowing SHM, Bach2 drives immunoglobulin gene conversion (GCV), an important AID-dependent antibody gene diversification process. We demonstrate that Bach2 promotes GCV by increasing the expression of AID. Importantly, we found that the regulation of AID is independent of Blimp-1 and that Bach2-deficient cells have altered expression of several genes regulating AID expression, stability and function. These results demonstrate that Bach2 has a previously unappreciated role in the production of high-affinity antibodies.

overall_design: Gene expression profiles of Clone 18 (Wild type) and Bach2 knockout (Bach2KO)  chicken DT40 B cells were generated by deep sequencing, in triplicate, using Illumina HiSeq2500 instrument and paired-end sequencing chemistry with 100bp read length

first: Minna

middle: K

last: Kylaniemi


title: Global analysis of gene expression reveals mRNA superinduction is required for the inducible immune response to a bacterial pathogen

city: Seattle

first: Kevin

last: Barry

state: WA

address: 1100 Fairview Ave N

institute: Fred Hutchinson Cancer Center

summary: The inducible innate immune response to infection requires a concerted process of gene expression that is regulated at multiple levels. Most global analyses of the innate immune response have focused on transcription induced by defined immunostimulatory ligands, such as lipopolysaccharide. However, the response to pathogens involves additional complexity, as pathogens interfere with virtually every step of gene expression. How cells respond to pathogen-mediated disruption of gene expression to nevertheless initiate protective responses remains unclear. We previously discovered that a pathogen-mediated blockade of host protein synthesis provokes the production of specific pro-inflammatory cytokines. It remains unclear how these cytokines are produced despite the global pathogen-induced block of translation. We addressed this question by using parallel RNAseq and ribosome profiling to characterize the response of macrophages to infection with the intracellular bacterial pathogen Legionella pneumophila. Our results reveal that mRNA superinduction is required for the inducible immune response to a bacterial pathogen.

overall_design: Transcriptional and translational responses of bone marrow macrophages to infection with Legionella pneumophila were measured by RNAseq and ribosome profiling following standard protocols.

first: Nicholas

middle: T

last: Ingolia

first: Russell

last: Vance


title: Novel RNA biomarkers of prostate cancer revealed by RNA-seq analysis of formalin-fixed samples obtained from Russian patients

city: Moscow

first: Anastasia

last: Nikitina

address: Malaya Pirogovskaya

department: Department of Molecular Biology and Genetics

country: Russia

institute: Federal Research and Clinical Center of Physical-Chemical Medicine of FMBA of Russia

summary: Due to heterogeneous multifocal nature of prostate cancer (PCa), there is currently a lack of biomarkers that stably distinguish it from benign prostatic hyperplasia (BPH), predict clinical outcome and guide the choice of optimal treatment. In this study, RNA-seq analysis was applied to formalin-fixed paraffin-embedded (FFPE) tumor and matched normal tissue samples collected from Russian patients with PCa and BPH. We identified 3384 genes differentially expressed (DE) (FDR < 0.05) between tumor tissue of PCa patients and adjacent normal tissue as well as both tissue types from BPH patients. Overexpression of four of the genes previously not associated with PCa (ANKRD34B, NEK5, KCNG3, and PTPRT) was validated by RT-qPCR. Furthermore, the enrichment analysis of overrepresented microRNA and transcription factor (TF) recognition sites within DE genes revealed common regulatory elements of which 13 microRNAs and 53 TFs were thus linked to PCa for the first time. Moreover, 8 of these TFs (FOXJ2, GATA6, NFE2L1, NFIL3, PRRX2, TEF, EBF2 and ZBTB18) were found to be differentially expressed in this study, making them not only candidate biomarkers of prostate cancer but also potential therapeutic targets.

overall_design: Whole transcriptome profiling of tumor tissue and matched adjacent normal tissue from 15 patients with PCa and 2 with BPH.

first: Elena

last: Sharova

first: Tatiana

last: Butusova

first: Svetlana

last: Danilenko

last: Kostryukova


title: Cbx3 Maintains Lineage Specificity During Neural Differentiation [ChIP-seq]

city: Los Angeles

first: Siavash

last: Kurdistani

state: CA

address: 615 Charles E Young Dr South

department: Biological Chemistry

institute: UCLA

summary: Cbx3 (HP1γ) that is a member of the heterochromatin protein 1 familiy enriched in gene bodies of pluripotent ESCs but promoters of differentiated pre-iPSCs. To determine whether Cbx3 becomes enriched at promoters to influence gene regulation during differentiation, we converted ESCs to NPCs in monolayer culuture and performed genomewide ChIP-seq of Cbx3. Our results shows that Cbx3 enriches at the promoters of genes upon differentiation of ESCs to NPCs and stablizes the binding of Mediator subunit Med26 to pre-initiation complex (PIC) in NPCs.

overall_design: Examine the enrichment of Cbx3 and Pol II in wild-type ESC-derived NPCs, and the enrichment of Med26, Cdk8 and Pol II in NPC transfected with control siRNA targeting luciferase or siRNA mediated knockdown of Cbx3 by ChIP-seq analysis.

first: Chengyang

last: Huang

first: Trent

last: Su

first: Michael

last: Carey


title: CD161+ Tconv and Treg share transcriptome and display a migratory phenotype which is modified at the inflamed site

city: London

first: Chantal

last: Duurland

address: 30 Guilford Street

department: Infection, Inflammation and Rheumatology

institute: UCL Great Ormond Street Institute of Child Health

summary: Regulatory T cells (Treg) are a cornerstone of immune regulation. Recent evidence indicates that human Treg show plasticity in specific settings. A subpopulation of Treg expressing CD161, a marker associated with T helper (Th)17 cells, have T effector -like characteristics and are enriched at sites of autoimmune inflammation. Here we used RNAseq to confirm the effector-like signature of CD161+ Treg and demonstrated a shared transcriptional signature between CD161+ Treg and CD161+ conventional T cells (Tconv). Pathway analysis suggested that CD161+ T cells have a migratory phenotype, expressing high levels of CCR9 and integrin α4β7, markers associated with gut homing. In response to all-trans retinoic acid, CD161+ T cells expressed higher levels of CCR9 and integrin α4β7 than CD161- T cells. Our data suggest that blood CD161+ T cells may have adopted gut homing properties upon retinoic acid exposure. In contrast to their peripheral counterparts, CD161+ T cells from the site of autoimmune arthritis have a diminished gut homing phenotype and blunted response to retinoic acid. In health, the TCRβ repertoires of CD161+ and CD161- T cells showed limited overlap whereas there is clear overlap in T cell clones from synovial fluid of autoimmune arthritis patients. We therefore propose that CD161+ and CD161- T cells are largely distinct populations in the healthy immune system but that the inflamed site creates an environment where CD161 levels in T cells can be altered, potentially contributing to disease pathogenesis.

overall_design:
Transcriptional profilling of CD161+ and CD161- Tconv, and CD161+ and CD161- Tconv from blood of healthy adult controls. The raw files provided are BAM files, and the two associated raw files are merged into one BAM file for each sample.
Note: Sample 2 CD161+ Treg was excluded due to low cDNA yield

middle: L

first: Chrysothemis

last: Brown

first: Ryan

last: O'Shaughnessy

first: Lucy

last: Wedderburn


title: Cbx3 Maintains Lineage Specificity During Neural Differentiation


title: Cbx3 Maintains Lineage Specificity During Neural Differentiation [RNA-seq]

summary: Cbx3 (HP1γ) that is a member of the heterochromatin protein 1 family play important roles in development and differentiation. To determine the regulatroy mechanisms of Cbx3 during neural differentiation from ESCs to NPCs, we performed RNA-seq analysis of ESCs or ESC-derived NPCs depleted for Cbx3 or Cbx3-assocatied Mediator subunit Med26.

overall_design: ESCs or ESC-derived NPCs were transfected with control siRNA targeting to luciferase or siRNA mediated knockdown of Cbx3 or Med26. RNAs were extracted from control or knockdown group and subjected to library preparation and deep sequencing.


title: Graded responses to variable TCR signaling are encoded in the affinities of AICE-containing enhancers responding to BATF and IRF4

city: Chiba-shi

first: Arifumi

last: Iwata

state: Chiba

address: 1-8-1 Inohana, Chuou-ku

department: Department of Allergy and Clinical Immunology

country: Japan

institute: Chiba University

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

overall_design: Refer to individual Series


title: Temporal-spatial Establishment of Initial Niche for the Primary Spermatogonial Stem Cell formation is Determined by an ARID4B Regulatory Network (ChIP-Seq)

city: Washington

first: Ray-Chang

last: Wu

state: DC

address: 2300 I St, NW

department: Biochemistry and Molecular Medicine

institute: George Washington University

summary: The primary spermatogonial stem cells (SSCs), which arise from gonocytes during neonatal development, serve as a foundational self-renewing reservoir to ensure continuous production of spermatozoa throughout adulthood. The transformation of gonocytes into SSCs takes place in a niche established by Sertoli cells. To date, the factors that guide Sertoli cells to establish the initial stem cell niche remain largely unknown. Using Sertoli cell-specific Arid4b knockout (Arid4bSCKO) mice, we demonstrated that ablation of ARID4B resulted in failure to establish a niche for the SSC formation. We performed ChIP-Seq analysis to identify target genes of ARID4B in testes.

overall_design: A pool of 108 testes was dissected from wild type male mice at P1.5 for ChIP, library preparation, sequencing, and initial data analysis. ChIP assays were performed using the anti-ARID4B antibody. Input DNA was used for control.

first: Mei-Yi

first: Rainer

middle: B

last: Lanz


title: Quality of TCR signaling determined by differential affinities of enhancers for the composite BATF–IRF4 transcription factor complex [ChIP-Seq]

summary: Variable strengths of signaling via the T cell antigen receptor (TCR) can produce divergent outcomes, but the mechanism of this remains obscure. The abundance of the transcription factor IRF4 increases with TCR signal strength, but how this would induce distinct types of responses is unclear. We compared the expression of genes in the TH2 subset of helper T cells to enhancer occupancy by the BATF–IRF4 transcription factor complex at varying strengths of TCR stimulation. Genes dependent on BATF–IRF4 clustered into groups with distinct TCR sensitivities. Enhancers exhibited a spectrum of occupancy by the BATF–IRF4 ternary complex that correlated with the sensitivity of gene expression to TCR signal strength. DNA sequences immediately flanking the previously defined AICE motif controlled the affinity of BATF–IRF4 for direct binding to DNA. Analysis by the chromatin immunoprecipitation–exonuclease (ChIP-exo) method allowed the identification of a previously unknown high-affinity AICE2 motif at a human single-nucleotide polymorphism (SNP) of the gene encoding the immunomodulatory receptor CTLA-4 that was associated with resistance to autoimmunity. Thus, the affinity of different enhancers for the BATF–IRF4 complex might underlie divergent signaling outcomes in response to various strengths of TCR signaling

overall_design: ChIP-seq or ChIP-exo for 3 transcription factors (BATF, BATF3 and IRF4) in 8 different types of CD4 T cells cultured under TH2 conditions

first: Theresa

last: Murphy

first: Kenneth

middle: M


title: Temporal-spatial Establishment of Initial Niche for the Primary Spermatogonial Stem Cell formation is Determined by an ARID4B Regulatory Network (RNA-Seq)

summary: The primary spermatogonial stem cells (SSCs), which arise from gonocytes during neonatal development, serve as a foundational self-renewing reservoir to ensure continuous production of spermatozoa throughout adulthood. The transformation of gonocytes into SSCs takes place in a niche established by Sertoli cells. To date, the factors that guide Sertoli cells to establish the initial stem cell niche remain largely unknown. Using Sertoli cell-specific Arid4b knockout (Arid4bSCKO) mice, we demonstrated that ablation of ARID4B resulted in failure to establish a niche for the SSC formation. We performed RNA-Seq analysis to examine the gene expression profile of the Arid4bSCKO testes in comparison with that of control testes.

overall_design: We extracted RNA from testes of the control and Arid4bSCKO mice at postnatal day 1.5 of age. Each genotypic group consisted of three pools of testes, and each pool contained four testes. Purified RNA was processed for RNA-Seq analysis using Illumina HiSeq 2500.


title: Temporal-spatial Establishment of Initial Niche for the Primary Spermatogonial Stem Cell formation is Determined by an ARID4B Regulatory Network


title: A comprehensive analysis of gene expression patterns in wild-type and Batf2-/- bone marrow-derived macrophages (BMMφ)

city: Suita

first: Daisuke

last: Okuzaki

state: Osaka

address: Yamadaoka 3-1

department: Immunology Frontier Research Center

institute: Osaka univ.

summary: We previously identified TLR-independent expression of 4933430F08Rik, encoding Batf2, in T. cruzi-infected bone marrow-derived dendritic cells (BMDCs) (Kayama et al., 2009). To determine the functions of Batf2 in innate immune responses, we performed a comprehensive gene expression analysis in wild-type and Batf2-/- bone marrow-derived macrophages (BMMφ). RNA-seq analysis showed that 98 genes are upregulated in Batf2-/- BMMφ stimulated with LPS following IFN-γ treatment, when compared with that in wild-type cells. Among these genes, we focused on Il23a, encoding IL-23p19, because IL-23 is able to promote expression of Il17a in Th17 cells.

overall_design: mRNA of wild-type and Batf2-/- BMMφ were subjected to deep sequencing profiling using Illumina HiSeq 2000.

first: Shoko

last: Kitada

first: Hisako

last: Kayama

first: Ritsuko

last: Koga

first: Masao

last: Kobayashi

first: Yasunobu

last: Arima

first: Masaaki

last: Murakami

first: Kiyoshi

last: Takeda


title: Transposable element targeting by piRNAs in Laurasiatherians with distinct TE activities

status: Public on Mar 26 2017

city: Mississippi State

first: Michael

middle: W

last: Vandewege

state: Mississippi

address: 32 Creelman St

institute: Mississippi State University

summary: Transposable elements (TEs) are self-mobilizing elements that make up a large fraction of mammalian genomes. PIWI proteins and PIWI interacting RNAs (piRNAs) are a part of a system aimed at controlling and preventing TE proliferation. We examined the TE landscape and piRNA repertoires from three non-model Laurasiatherian mammals (dog, horse and big brown bat) with differing TE complements and proliferation patterns to address questions about the evolutionary relationships between piRNAs and TEs. We found that the genomic abundance of new TEs may not match TE transcription. We speculate that a higher rate of SINE targeting by piRNAs in the horse genome contributed to the reduced SINE insertion rate.

sample_organism:
Canis lupus familiaris
Equus caballus
Eptesicus fuscus

overall_design: Two testicular tissue sample from three taxa. Six samples total

first: Roy

last: Platt

first: David

last: Ray

first: Federico

last: Hoffmann


title: Context-dependent miR-204 and miR-211 affect the biological properties of amelanotic and melanotic melanoma cells

city: Pisa

first: Romina

last: D'Aurizio

state: PI

address: via Moruzzi 1

department: IIT

country: Italy

institute: CNR


title: Next Generation Sequencing Analysis of Wild Type(WT) and Gdf9-Cre;Kat8 fl/fl(Kat8 Gdf9 cKO)  Oocyte Transcriptome

status: Public on Mar 25 2017

city: Hefei

first: Shi

last: Yin

address: Huangshan Road 443

institute: The University of Science and Technology of China

summary:
Purpose: The goal of this study is to compare oocyte transcriptome profiling in WT and Kat8 Gdf9 cKO mice
Methods: Oocyte mRNA profiles of 2 weeks old WT and Kat8 Gdf9 cKO mice were generated by deep sequencing using Illumina HiSeq 2500.
Results: Using an optimized data analysis workflow, we mapped about 54-63 million sequence reads per sample to the mouse genome (build mm10) and identified 18,468 transcripts in the oocytes of WT and Kat8 Gdf9 cKO mice. Approximately 5% of the transcripts showed differential expression between the WT and Kat8 Gdf9 cKO oocyte, with a fold change ≥1.5 and p value <0.05.
Conclusions: Our study represents the first detailed analysis of oocyte transcriptomes after kat8 deletion, which is generated by RNA-seq technology.Our results show that Kat8 affects numerous gene expression in mouse oocytes.

first: Xiaohua

last: Jiang


title: Context-dependent miR-204 and miR-211 affect the biological properties of amelanotic and melanotic melanoma cells [miRNA-Seq]

summary:
Despite increasing amounts of experimental evidence depicting the involvement of non-coding RNAs in cancer, the study of BRAFV600E-regulated genes has thus far focused mainly on protein-coding ones. Here, we identify and study the microRNAs that BRAFV600E regulates through the ERK pathway.
By performing small RNA sequencing on A375 melanoma cells and a vemurafenib- resistant clone that was taken as negative control, we discover miR-204 and miR-211 as the miRNAs most induced by vemurafenib. We also demonstrate that, although belonging to the same family, these two miRNAs have distinctive features. miR-204 is under the control of STAT3 and its expression is induced in amelanotic melanoma cells, where it acts as an effector of vemurafenib’s anti-motility activity by targeting AP1S2. Conversely, miR-211, a known transcriptional target of MITF, is induced in melanotic melanoma cells, where it targets EDEM1 and consequently impairs the degradation of TYROSINASE (TYR) through the ER-associated degradation (ERAD) pathway. In doing so, miR-211 serves as an effector of vemurafenib’s pro-pigmentation activity. We also show that such an increase in pigmentation in turn represents an adaptive response that needs to be overcome using appropriate inhibitors in order to increase the ef cacy of vemurafenib.
In summary, we unveil the distinct and context-dependent activities exerted by miR-204 family members in melanoma cells. Our work challenges the widely accepted “same miRNA family = same function” rule and provides a rationale for a novel treatment strategy for melanotic melanomas that is based on the combination of ERK pathway inhibitors with pigmentation inhibitors.

overall_design: The expression levels of microRNAs in A375 parental cells and the vem-resistant A375 C2 clone were measured by sequencing of small RNAs upon 48h of treatment with DMSO or 2uM vemurafenib. Two independent replicates were analyzed for each condition.

first: Marianna

last: Vitiello

first: Andrea

last: Tuccoli

first: Laura

last: Poliseno


title: Global transcriptional analysis of different HT1080 cell phenotypes induced by cell proliferation and density

city: San Francisco

first: Minsuk

last: Kwak

address: 2340 Sutter St

institute: University of California San Francisco

summary: Following uncontrolled proliferation, a subset of primary tumor cells acquires additional traits/mutations to trigger phenotypic changes that enhance migration and are hypothesized to be the initiations of metastasis. This study reveals a novel adaptive mechanism that harnesses synergistic paracrine signaling via IL-6/8, which is amplified by cell proliferation and cell density, to directly promote cell migration. This effect occurs in metastatic human sarcoma and carcinoma cells - but not in non-metastatic cells-, through the downstream signaling of WASF3 and Arp2/3. Our RNA-seq based global transcriptional analysis showed that the transcriptional phenotype of high-density cells that emerges due to proliferation resembles that of low-density cells treated with a combination of Il-6/8. Simultaneous inhibition of IL-6/8 receptors significantly decreases the expression of WASF3 and Arp2/3 in a mouse xenograft model and consequently reduces metastasis.

overall_design: Population-level RNA sequencing on five different HT1080 cell phenotypes induced by varying microenvironment. HT1080 cells were embedded in type I 3D collagen matrices in increasing cell densitieis from 10 cells/mm3 to 150 cells/mm3. RNA-seq performed on matrix embedded cells with cell densities of 10 cells/mm3 (low-density) and 50 cells/mm3 (high-density). Recombinant IL-6, IL-8 (both R&D systems), or both reconstituted in DPBS were added to matrix embedded cells with a cell density of 10 cells/mm3 (low-density with IL-6/IL-8/IL6+IL8). One collagen matrices without cells was sequenced and analyzed as a negative control. All data is obtained from single sequencing experiment. RNA-seq performed on these cell populations. All populations were incubated for 24h at 37°C, 5% CO2. The collagen matrices were exposed to cell lysis buffer and mechanically broken down using a syringe. The lysates of matrices were centrifuged and the supernatant was separately collected. Total RNA isolation was performed with RNA MiniPrep Kit (Zymo Research). Since the cells were embedded in the collagen matrices at the low cell density, the RNA-seq protocol (modified from Pan et al PNAS 2013) was implemented to capture uniformly distributed, full-length coverage sequences from low-quality and low-quantity cells. The cDNA libraries were sequenced using 50 bp single-ended reads. Data was processed using Bowtie2 for read alignment and RSEM for quantification of gene expression.

first: Hasini

last: Jayatilaka

first: Jonathan

middle: J

last: Chen

first: Daniele

last: Gilkes

first: Rong

last: Fan

first: Denis

last: Wirtz


title: Single-cell analysis reveals lineage segregation and pluripotency state transition in early post-implantation mouse embryos

city: Shanghai

first: Ying

last: Jin

address: 320 Yue Yang Road

department: Institute of Health Sciences

institute: Shanghai JiaoTong University

summary: Here we report the transcriptome data of 236 single cells freshly isolated from mouse embryos on days 5.5 and 6.5. Initial transcriptome data from 124 single cells yielded signature genes for the epiblast, visceral endoderm, and extra-embryonic ectoderm and revealed a unique distribution pattern of fibroblast growth factor (Fgf) ligands and receptors. Further analysis indicated that early-stage epiblast cells do not segregate into lineages of the major germ layers. Instead, some cells began to diverge from epiblast cells, displaying molecular features of the pre-mesendoderm by expressing higher levels of mesendoderm markers and lower levels of Sox3 transcripts. In addition, we identified a late stage of the day 6.5 embryo in which mesoderm and DE cells emerge, with many of them coexpressing Oct4 and Gata6. Analysis of single-cell RNA-seq data from 112 cells of the late-stage day 6.5 embryos revealed differentially expressed signaling genes and networks of transcription factors that might underlie the segregation of the mesoderm and DE lineages. Moreover, we discovered a subpopulation of mesoderm cells that possess molecular features of the extraembryonic mesoderm. This study provides fundamental insight into the molecular basis for lineage segregation in post-implantation mouse embryos.

overall_design: Examination of 108 EPI cells, 8 VE cells and 8 EXE cells from 3 pregastrulation embryos (E5.5 and early-stage E6.5), and 112 cells from 3 early gastrulation embryos (late-stage E6.5). The lineages of cells were determined by Pricipal component analysis or diffusion map analysis using expression data of all genes or known markers.


title: siRNA RNA-seq from K562 (ENCSR287SHU)

status: Public on Mar 24 2017

city: Stanford

first: ENCODE

last: DCC

address: 300 Pasteur Dr

institute: ENCODE DCC

summary:
Control siRNA against no target in K562 cells followed by RNA-seq.

For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and http://www.genome.gov/Pages/Research/ENCODE/ENCODE_Data_Use_Policy_for_External_Users_03-07-14.pdf


title: Comparison of Wild type and Pofut1-deleted skeletal muscle in young and old mice

city: Columbus

first: Paul

last: Martin

state: OH

address: 700 Childrens Dr.

department: Center for Gene Therapy

institute: Nationwide Childrens Hospital

summary: Pofut1 is an essential gene that glycosylates proteins containing EGF-like repeats, including Notch Receptors (NotchRs).  Work in mice and in Drosophila has shown that O-fucosylation by Pofut1 is required for NotchR ligands to bind to and activate NotchRs.  As such, Pofut1 deletion in skeletal myofibers allows for an analysis of potential functions and molecular changes of Pofut1 in skeletal muscle that derive from its expression in skeletal myofibers. In this study we compared gene expression profiles between quadriceps muscles in young (2 month) and old (17 month) mice where Protein O-fucosyltransferase 1 (Pofut1) was deleted specifically in skeletal myofibers via use of a human skeletal alpha actin Cre transgene (Scre) and a loxP flanked Pofut1 gene (SCreFF) and mice which bore the only the Scre transgene but did not have floxed Pofut1 alleles (SCre++).

overall_design: A total of eighteen samples in three groups were analyzed for this study. The first group is Pofut1flox/flox quadriceps. The second group is Skeletal α actin cre/Pofut1 wild type quadriceps. The third group is skeletal α actin cre/Pofut1flox/flox quadriceps. For each group, three of the samples are replicates from young (2 month) mice, and three of the samples are replicates from old (17 month) mice.

first: Deborah

last: Zygmunt


title: Alignment of the Transcriptome with Individual Variation in Animals Selectively Bred for High Drinking-In-the-Dark (HDID).

city: Portland

first: Priscila

last: Darakjian

state: OR

address: 3181 SW Sam Jackson Park Road, L470

department: Behavioral Neuroscience

institute: Oregon Health and Science University

overall_design: S27 HDID-1 selected line began with a “within family” selection design but was switched to mass selection at S5. All animals were on reversed light-dark cycle. Animals underwent the standard 4-day DID trial with three 2 hour exposures to a single bottle of 20% ethanol and a final day of 4 hour exposure. The DID trial always began 3 hours into the dark phase. BECs were obtained at the end of the 4th hour. A subgroup (N=20) of the animals were retested 3 weeks later. The remaining animals, 18 males and 17 females were sacrificed 3 weeks later and used for gene expression analyses. All animals were sacrificed between 10 AM and 2 PM. Mice were euthanized, brains removed and immediately frozen on dry ice.  Frozen brains were slightly thawed and dissected by hand under RNAse-free conditions. Using the optic chiasm as the rostral marker, a 2 mm coronal slice of brain tissue was isolated. Beginning at the medial ventral aspect of the striatum and recognizing that the striatum has a partial cone shape, the dissection moved dorsal 1 mm, followed by a cut to the lateral boundary of the striatum, with a final cut following the lateral-ventral boundary. The isolated tissue was immediately placed into 1 ml of Trizol (Invitrogen, Carlsbad, CA).Among animals at risk for excessive ethanol consumption such as the HDID selected lines, there is considerable individual variation in the amount of ethanol consumed and the associated blood ethanol concentrations (BECs). For the HDID lines, this variation occurs even though the residual genetic variation associated with the DID phenotype has been largely exhausted and thus is most likely associated with epigenetic factors. Here we focus on the question of whether the genes associated with individual variation in HDID-1 mice are different from those associated with selection (risk) (Iancu et al. 2013). Thirty-three HDID-1 mice were phenotyped for their BECs at the end of a standard DID trial, were sacrificed 3 weeks later and RNA-Seq was used to analyze the striatal transcriptome. The data obtained illustrated that there is considerable overlap of the risk and variation gene sets; both focus on the fine tuning of NMDA mediated synaptic plasticity.

first: Hitzemann

last: Robert

first: Oberbeck

last: Denesa

first: Iancu

last: Ovidiu

first: McWeeney

last: Shannon

first: Spencer

last: Stephanie

first: Schlumbohm

last: Jason

first: Metten

last: Pamela

first: Crabbe

last: John


title: The methylome of a cartilaginous fish, Callorhinchus milii, reveals conservation of epigenetic regulation across jawed vertebrates

city: Dunedin

first: Tim

last: Hore

address: 270 Great King Street

department: Department of Anatomy

country: New Zealand

institute: University of Otago

summary:
Background
Methylation of CG dinucleotides constitutes a critical system of epigenetic memory in bony vertebrates, where it modulates gene expression and suppresses transposon activity. The genomes of studied vertebrates are pervasively hypermethylated, with the exception of regulatory elements such as transcription start sites (TSSs), where the presence of methylation is associated with gene silencing. This system is not found in the sparsely methylated genomes of invertebrates, and establishing how it arose during early vertebrate evolution is impeded by a paucity of epigenetic data from basal vertebrates.
Methods
We perform whole-genome bisulfite sequencing to generate the first genome-wide methylation profiles of a cartilaginous fish, the elephant shark Callorhinchus milii. Employing these to determine the elephant shark methylome structure and its relationship with expression, we compare this with higher vertebrates and an invertebrate chordate using published methylation and transcriptome data.
Results
Like higher vertebrates, the majority of elephant shark CG sites are highly methylated, and methylation is abundant across the genome rather than patterned in the mosaic configuration of invertebrates. This global hypermethylation includes transposable elements and the bodies of genes at all expression levels. Significantly, we document an inverse relationship between TSS methylation and expression in the elephant shark, supporting the presence of the repressive regulatory architecture shared by higher vertebrates.
Conclusions
Our demonstration that methylation patterns in a cartilaginous fish are characteristic of higher vertebrates imply the conservation of this epigenetic modification system across jawed vertebrates separated by 465 million years of evolution. In addition, these findings position the elephant shark as a valuable model to explore the evolutionary history and function of vertebrate methylation.

overall_design: Whole-genome bisulfite sequencing was used to generate genome-wide methylation profiles from one male and one female liver sample. Low-coverage sequencing was also performed on additional liver, spleen and pancreas samples.

first: Julian

last: Peat

first: Oscar

last: Ortega-Recalde

first: Olga

last: Kardailsky

first: Timothy


title: Mechanisms of colitis-accelerated colon carcinogenesis and its prevention with the combination of aspirin and curcumin: Transcriptomic analysis using RNA-seq

city: Piscataway

first: Ronald

middle: P.

last: Hart

state: NJ

address: 604 Allison Rd Rm B430

department: Cell Biology & Neuroscience

institute: Rutgers University

summary: Colorectal cancer (CRC) remains the leading cause of cancer-related death in the world. Aspirin (ASA) and curcumin (CUR) are widely investigated chemopreventive candidates for CRC. However, the precise mechanisms of their action and their combinatorial effects have not been evaluated. The purpose of the present study was to determine the effect of ASA, CUR, and their combination in azoxymethane/dextran sulfate sodium (AOM/DSS)-induced colitis-accelerated colorectal cancer (CAC). We also aimed to characterize the differential gene expression profiles in AOM/DSS-induced tumors as well as in tumors modulated by ASA and CUR using RNA-seq. Diets supplemented with 0.02% ASA, 2% CUR or 0.01% ASA + 1% CUR were given to mice from 1 week prior to the AOM injection until the experiment was terminated 22 weeks after AOM initiation. Our results showed that CUR had a superior inhibitory effect in colon tumorigenesis compared to that of ASA. The combination of ASA and CUR at a lower dose exhibited similar efficacy to that of a higher dose of CUR at 2%. RNA isolated from colonic tissue from the control group and from tumor samples from the experimental groups was subjected to RNA-seq. Transcriptomic analysis suggested that the low-dose combination of ASA and CUR modulated larger gene sets than the single treatment. These differentially expressed genes were situated in several canonical pathways important in the inflammatory network and liver metastasis in CAC. We identified a small subset of genes as potential molecular targets involved in the preventive action of the combination of ASA and CUR. Taken together, the current results provide the first evidence in support of the chemopreventive effect of a low-dose combination of ASA and CUR in CAC. Moreover, the transcriptional profile obtained in our study may provide a framework for identifying the mechanisms underlying the carcinogenesis process from normal colonic tissue to tumor development as well as the cancer inhibitory effects and potential molecular targets of ASA and CUR.

overall_design: 10 RNA samples (5 experimental groups with duplicates) were sequenced using Illumina NextSeq 500 instrument.

first: Yue

last: Guo

middle: P

first: A

last: Kong


title: De novo assembly of Aedes aegypti using Hi-C yields chromosome-length scaffolds

city: Houston

last: Dudchenko

address: 1 Baylor Plaza

institute: Baylor College of Medicine

type:
Other
Third-party reanalysis

summary: The Zika outbreak, spread by the Aedes aegypti mosquito, highlights the need to create high-quality assemblies of large genomes in a rapid and cost-effective fashion. Here, we combine Hi-C data with existing draft assemblies to generate chromosome-length scaffolds. We validate this method by assembling a human genome, de novo, from short reads alone (67X coverage, Sample GSM1551550). We then combine our method with draft sequences to create genome assemblies of the mosquito disease vectors Aedes aegypti and Culex quinquefasciatus, each consisting of three scaffolds corresponding to the three chromosomes in each species. These assemblies indicate that virtually all genomic rearrangements among these species occur within, rather than between, chromosome arms. The genome assembly procedure we describe is fast, inexpensive, accurate, and can be applied to many species.

sample_organism:
Aedes aegypti
Culex quinquefasciatus
Homo sapiens

platform_organism:
Aedes aegypti
Culex quinquefasciatus

overall_design: We use DNA proximity ligation (Hi-C) to create a genome assembly with chromosome-length scaffolds for the mosquito Aedes aegypti, principal vector of the Zika virus.

first: Erez

last: Aiden


title: Microglia  self-renewal comprises context-dependent random or clonal expansion.

city: Freiburg

first: Ori

last: Staszewski

address: Breisacher Str. 64

department: Institute of Neuropathology

institute: University Medical Center Freiburg

summary: Microglia constitute a highly specialized network of tissue resident immune cells that is important for the control of tissue homeostasis and the resolution of virtually all diseases of the central nervous system (CNS). However, how this dissemination  is established and maintained in vivo and its kinetics of these processes are poorly understood. Here we established a new multicolor fluorescence fate mapping system to monitor microglia dynamics during steady state  and disease. Our findings that microglia establish a stable network over time albeit with regional differences and remarkably high turnover rates challenge their postulated longevity. Microglia self-renewal constitutes a random process  without evidence of defined progenitor cells. Under pathological conditions this randomness shifts to clonal microglia expansion that is finally resolved by both cell apoptosis and egression for re-establishment of the stable microglia network. Our data reveal new insights on how microglia ensure their complex distribution throughout the healthy and diseased CNS .

overall_design: Total RNA was column purified using the RNeasy Mini Kit (QIAGEN) from the ipsilateral and contralateral ventral pons (containing the facial nucleus, and parts of the gigantocellular reticular nucleus, intermediate reticular nucleus, parvicellular reticular nucleus, alpha, and spinal trigeminal nucleus, oral) of CX3CR1GFP/wt mice that underwent unilateral facial nerve axotomy at 8 weeks of age. A rodent brain matrix (RBM-2000C, ASI Instruments) was used to ensure consistent cutting of brain tissue. Total RNA from the contralateral ventral pons of un-operated 8-week-old CX3CR1GFP/wt was used as baseline control (Day 0 post nerve transection) for the analysis. Three replicates were used per time point (Day 0, 7, 14 and 30 post axotomy). Illumina HiSeq deep sequencing was carried out at a genomics core facility (Center of Excellence for Fluorescent Bioanalytics, KFB, University of Regensburg, Germany) for 50 bases single-read sequencing that generate about 30 million reads per sample. Data analysis was performed using Bioconductor (Huber et al., 2015). Sequences were aligned using the Rsubread aligner (Liao et al., 2013) with default settings using the Gencode mouse assembly (version M7) as reference. Gene counts were generated using the featureCounts program included in the Rsubread library after sequence alignment (Liao et al., 2013). Counts were filtered for subsequent analysis. The sum of all counts over all samples was calculated. Genes with a sum of counts of 0 were removed from the data. All genes at or below the 32nd percentile of the sum of gene counts were also excluded from the remaining genes. Differential gene expression was tested for using limma (Ritchie et al., 2015), which utilizes parts of the edgeR (Robinson et al., 2010) code for RNAseq analysis, as described in the limma documentation (Ritchie et al., 2015). Genes were recognized as differentially expressed when having a BH (Benjamini-Hochberg) adjusted P < 0.05. The most significantly differentially expressed genes were identified based on a log2 (fold change) ≥ 1.7 as suggested by the SEQC consortium (Consortium 2014). Heat maps were generated with the heatmap3 Bioconductor package (Zhao et al., 2014). Pathway and Functional analysis was performed using Ingenuity Pathway Analysis (IPA, Qiagen). Subclustering of gene expression profiling utilized the Short Time-series Expression Miner (STEM) (Ernst et al., 2005; Ernst and Bar-Joseph 2006).

first: Tuan-Leng

last: Tay


title: Functional assessment of CTCF sites at cytokine-sensing mammary enhancers in mammary gland [4C-seq]

city: Bethesda

first: Michaela

last: Willi

address: 9000 Rockville Pike

department: NIDDK

institute: NIH

summary: In mammary gland, deletion of the border CTCF site separating the Csn1s1 mammary enhancer from neighboring genes resulted in the activation of Sult1d1 at a distance of more than 95 kb but not the more proximal and silent Sult1e1 gene. Loss of this CTCF site led to de novo interactions between the Sult1d1 promoter and several enhancers in the casein locus.

overall_design: Examination of chromatin interactions of Sult1d1 promoter in wild-type and CTCF site mutant mammary gland

first: Hye

last: Lee

first: Chaochen

last: Wang


title: Functional analysis of CTCF sites associated with cytokine-sensing mammary-specific enhancers in the complex casein locus

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


title: Identification of genes induced by NOTCH1 in a chronic lymphocytic leukaemia (CLL) cell line and tracking of these genes in primary CLL patients

city: New York

first: Riccardo

last: Dalla-Favera

address: 1130 St Nicholas Avenue

institute: Columbia University

summary: NOTCH1 is mutationally activated in ~15% of cases of chronic lymphocytic leukaemia (CLL), but its role in B-cell development and leukemogenesis is not known. Here, we report that the active intracellular portion of NOTCH1 (ICN1) is detectable in ~50% of peripheral blood CLL cases lacking gene mutations. We identify a ‘NOTCH1 CLL gene expression signature’ in CLL cells, and show that this signature is significantly enriched in primary CLL cases expressing ICN1, independent of NOTCH1 mutation. NOTCH1 target genes include key regulators of B-cell proliferation, survival and signal transduction physiology. In particular, we show that MYC is a direct target of NOTCH1 via B-cell specific distal regulatory elements, thus implicating this oncogene in the pathogenesis of the disease.

overall_design: RNA-Seq analysis

first: Giulia

last: Fabbri

first: Antony

last: Holmes


title: The sea urchin larval response to immune challenge with Vibrio diazotrophicus

first: Katherine

middle: Michele

last: Buckley

address: 800 22nd St NW

department: Department of Biological Sciences

institute: The George Washington University

summary: Seawater exposure to the gram negative marine bacterium Vibrio diazotrophicus induces a robust cellular response in sea urchin larvae that includes the migration of pigment cells to the gut epithelium, changes in cell behavior and altered gut morphology (Ho et al., 2016; PMID 27192936). To investigate the transcriptional underpinnings of this response, whole transcriptome sequencing was performed on mRNA isolated from larval samples collected at 0, 6, 12 and 24 hr of exposure to V. diazotrophicus. The morphological simplicity of the sea urchin larva provides a systems-level model for identifying biologically relevant transcriptional state changes in response to dysbiosis in the gut lumen.

overall_design: Sea urchin larvae were sampled at four time points in response to bacterial exposure

first: Jonathan

last: Rast


title: Transcriptional regulatory dynamics drive coordinated metabolic and neural response to social challenge in mice.

city: Urbana

first: Lisa

last: Stubbs

address: 1206 West Gregory Drive | MC-195

department: IGB

institute: University of Illinois Urbana-Champaign

overall_design: Refer to individual Series


title: Identification of NOTCH1 bound genes in chronic lymphocytic leukaemia (CLL) cells

overall_design: ChIP-Seq analysis of NOTCH1 bound regions and histone marks in CLL cells


title: Functional analysis of CTCF sites associated with cytokine-sensing mammary-specific enhancers in the complex casein locus [ChIP-seq]

summary: The zinc finger protein CTCF has been invoked in establishing functional boundaries between genes and thereby controlling enhancer activities. However, there is limited genetic evidence to support such a concept. We have now addressed this question in a locus containing five mammary-specific genes controlled by enhancers. We have identified four CTCF binding sites in the casein locus, two at the outside boundaries and two associated with a super-enhancer. Individual deletions of these sites from the mouse genome did not alter expression of the five casein genes and Odam. However, deletion of the border CTCF site separating the Csn1s1 enhancer from non-mammary genes resulted in the activation of Sult1d1 at a distance of more than 95 kb but not the more proximal Sult1e1 gene. Loss of this CTCF site led to de novo interactions between the Csn1s1 enhancer and the Sult1d1 promoter but not with the silent Sult1e1 gene. Our study suggests that most CTCF sites associated with cell-specific enhancers and super-enhancers have no measurable in vivo activity. Only the loss of one CTCF site led to the induction of a juxtaposed active non-target promoter. Our study also demonstrated that cell-specific enhancers are unable to activate juxtaposed silent non-target promoters.

overall_design: ChIP-seq for mutant mice ΔA were done for CTCF, GR, H3K27ac, Pol2, SMC1 and STAT5A in mammary tissue at lactation day 1 (L1). ChIP-seq for ΔC was done for CTCF in mammary tissue at L1.

first: Hye-Kyung

first: Chul-Min

last: Yang

first: Chengyu

last: Liu

first: Lothar

last: Hennighausen


title: Open chromatin profiling of human postmortem brain infers functional roles for non-coding schizophrenia loci

first: Panos

last: Roussos

address: 1470 Madison Ave

department: Psychiatry

institute: Icahn School of Medicine at Mount Sinai

summary: Open chromatin provides access to DNA binding proteins for the correct spatiotemporal regulation of gene expression. Mapping chromatin accessibility has been widely used to identify the location of cis regulatory elements (CREs) including promoters and enhancers. CREs show tissue- and cell-type specificity and disease-associated variants are often enriched for CREs in the tissues and cells that pertain to a given disease. To better understand the role of CREs in neuropsychiatric disorders we applied the Assay for Transposase Accessible Chromatin followed by sequencing (ATAC-seq) to neuronal and non-neuronal nuclei isolated from frozen postmortem human brain by fluorescence-activated nuclear sorting (FANS). Most of the identified open chromatin regions (OCRs) are differentially accessible between neurons and non-neurons, and show enrichment with known cell type markers, promoters and enhancers. Relative to those of non-neurons, neuronal OCRs are more evolutionarily conserved and are enriched in distal regulatory elements. Transcription factor (TF) footprinting analysis identifies differences in the regulome between neuronal and non-neuronal cells and ascribes putative functional roles to a number of non-coding schizophrenia (SCZ) risk variants. Among the identified variants is a Single Nucleotide Polymorphism (SNP) proximal to the gene encoding SNX19. In vitro experiments reveal that this SNP leads to an increase in transcriptional activity. As elevated expression of SNX19 has been associated with SCZ, our data provides evidence that the identified SNP contributes to disease. These results represent the first analysis of OCRs and TF binding sites in distinct populations of postmortem human brain cells and further our understanding of the regulome and the impact of neuropsychiatric disease-associated genetic risk variants.

overall_design: Open chromatin profiles in neuronal and non-neuronal nuclei from the frontopolar prefrontal cortex of 8 control samples

first: John

last: Fullard


title: Transcriptional regulatory dynamics drive coordinated metabolic and neural response to social challenge in mice. [RNA-Seq data set]

summary: Agonistic encounters with conspecifics are powerful effectors of future behavior that evoke strong and durable neurobiological responses. We recently identified a deeply conserved “toolkit” of transcription factors (TFs) that respond to social challenge across diverse species in coordination with distinct conserved signatures of energy metabolism and developmental signaling. To further characterize this response and its transcriptional drivers in mice, we examined gene expression and chromatin landscape in the hypothalamus, frontal cortex, and amygdala of socially challenged and control animals over time. The data revealed a complex spatiotemporal pattern of metabolic, neural, and developmental transcriptomic signatures coordinated with significant shifts in the accessibility of distally located regulatory elements. Transcriptional regulatory network and motif analyses revealed an interacting network of TFs correlated with differential gene expression across the tissues and time points we assayed, including the early-acting and conserved regulator of energy metabolism and development, ESRRA. Cell-type deconvolution analysis attributed the early metabolic activity implicated by our transcriptomic analysis primarily to oligodendrocytes and the developmental signal to neurons, and we confirmed the presence of ESRRA in both oligodendrocytes and neurons throughout the brain. To assess the role of this nuclear receptor as an early trigger of this coordinated response, we used chromatin immunoprecipitation to map ESRRA binding sites to a set of genes involved in metabolic regulation and enriched in challenge-associated differentially expressed genes. Together, these data support a rich model linking metabolic and neural responses to social challenge, and identify regulatory drivers with unprecedented tissue and temporal resolution.

overall_design: Territory-holding resident mice were males from the C57BL/6J strain co-housed with females to establish a territory. Intruder mice were males from the BALB/C strain. Animals were housed in a 12L:12D animal room until the resident-intruder paradigm was undertaken. Before behavior work, male C57BL/6J animals were cohoused with members of the same sex for two weeks, housed alone for a week, and then housed with a single C57BL/6J female for a week to establish a territory. Thus, before behavior work, the animals were allowed to habituate to our animal facility for four weeks. Three hours before testing, females were removed from the resident males’ cages. Immediately before the trial, residents’ cages were inserted into a blank-walled chamber. For experimental mice, we introduced unfamiliar intruder BALB/cJ male mice. Intruders were contained within a stainless steel wire ~1cm mesh cage to prevent animals from making contact and injuring one another. Control animals were exposed to the same cage, but containing a paper cup instead of an intruder mouse. The cages were removed in both intruder and control conditions after five minutes. After exposure to the intruder or control stimulus, resident animals were allowed to sit in a dark and quiet place for either 30 minutes, 60 minutes, or 120 minutes. Residents were then immediately euthanized by cervical dislocation. As soon as animals were euthanized, we extracted three brain regions of interest from our animals: frontal cortex, hypothalamus, and amygdala. This yielded tissue samples from which RNA was extracted. The RNA samples were pooled to generate libraries for sequencing.  For control mice there were 5 replicates for all combinations of time after stimulus (30, 60, 120 minutes) and brain region (frontal cortex, hypothalamus, amygdala) except for hypothalamus from control mice after 30 minutes (3 replicates) and for frontal cortex from control mice after 120 minutes (6 replicates). For experimental mice there were 5 replicates for all combinations of time after stimulus (30, 60, 120 minutes) and brain region (frontal cortex, hypothalamus, amygdala) except for frontal cortex from experimental mice after 120 minutes (6 replicates).

first: Michael

middle: C

last: Saul

first: Christopher

last: Seward


title: Dynamic RNA-protein interactions underlie the zebrafish maternal-to-zygotic transition

city: New Haven

first: Karla

last: Neugebauer

state: CT

address: 333 Cedar St

department: Molecular Biophysics and Biochemistry

institute: Yale University

summary: We adapted the iCLIP method to zebrafish embryos to investigate activities of Hnrnpa1 during the maternal-to-zygotic transition

overall_design: 3 and 4 biological replicates from zebrafish embryos before (preZGA) and during (ZGA) zygotic genome activation, respectively, after UV crosslinking. Negative control samples include 3 IgG replicates of UV crosslinked samples for both developmental stages examined

first: Vladimir

last: Despic


title: Transcriptional expression level of E. coli at 37 ℃ in LB media

city: Daejeon

first: Byung-Kwan

last: Cho

address: 291 Daehak-ro

country: South Korea

institute: KAIST

summary: In order to understand the impact of fitness effect on genome evolution, we measured gene expression level by directional RNA-seq of E. coli grown at 37 ℃ in LB media because it is known that gene expression level has correlation with sequence evolution rate (Koonin & Wolf, 2006).

sample_organism: Escherichia coli str. K-12 substr. MG1655

overall_design: Directional RNA-seq (dUTP method by TruSeq®) were generated  in duplicates by using Illumina MiSeq

first: Yoo-Bok

first: Donghui

last: Choe

first: Suhyung


title: Transcriptional regulatory dynamics drive coordinated metabolic and neural response to social challenge in mice. [ChIP-Seq data set]


title: ChIP-seq for SMARCE1 and SMARCC1 in non-invasive and invasive HMLE-Twist-ER cells

status: Public on Mar 23 2017

city: Cambridge

first: Dexter

middle: X

last: Jin

state: MA

address: 9 Cambridge Center

institute: Whitehead Institute

summary: We find that in invasive HMLE-Twist-ER cells, SMARCE1 binds to ILF3 and is localized to ILF motifs. In contrast, ILF motifs were not enriched at SMARCE1-bound sites in noninvasive HMLE-Twist-ER cells or at SMARCC1-bound sites.

overall_design: Performing ChIP-Seq on HMLE-Twist-ER cells +/- 125 nM 4-OHT

first: Ethan

middle: S

last: Sokol

first: Yu-Xiong

last: Feng


title: CRISPR-Cas9 epigenome editing enables high-throughput screening for functional regulatory elements in the human genome [SKBR3 DNase]

city: Durham

first: Charles

middle: A

last: Gersbach

state: North Carolina

address: 101 science drive rm 2323

institute: Duke University

summary: Large genome mapping consortia and thousands of genome-wide association studies have identified non-protein coding elements in the genome as a having a central role in tissue development, cell-type specification, response to environmental or pharmacologic signals, and susceptibility to most common diseases. However, decoding the function of the millions of putative regulatory elements discovered in these studies remains a primary challenge. New CRISPR/Cas9-based epigenome editing technologies have enabled the precise perturbation of the activity of specific regulatory elements. Here we describe CRISPR/Cas9-based Epigenomic Regulatory Element Screening (CERES) for high-throughput screening of regulatory element activity within the native genomic context. We perform both loss- and gain-of-function screens with complementary epigenome editing tools to identify known and unknown regulatory elements of medically relevant genes in human cells. The high-throughput functional annotation of putative regulatory elements by CERES constitutes a new platform for screening biological mechanisms that cannot be perturbed by traditional methods.

overall_design: Pooled gRNA screens with dCas9 based epigenome editing tools for regulatory element discovery.

first: Tyler

last: Klann

first: Joshua

last: Black

first: Malathi

last: Chellappan

first: Alexias

last: Safi

first: Lingyun

last: Song

first: Isaac

last: Hilton

first: Gregory

last: Crawford

first: Timothy

last: Reddy


title: CRISPR-Cas9 epigenome editing enables high-throughput screening for functional regulatory elements in the human genome [CERES]


title: Probing the Canonicity of the Wnt/Wingless Signaling Pathway [STARR-seq]

city: Vienna

first: Alexander

last: Stark

address: Campus-Vienna-Biocenter 1

country: Austria

institute: The Research Institute of Molecular Pathology (IMP)

summary: The hallmark of canonical Wnt signaling is the transcriptional induction of Wnt target genes by the beta-catenin/TCF complex. Several studies have proposed alternative interaction partners for beta-catenin or TCF, but the relevance of potential bifurcations in the distal Wnt pathway remains unclear. Here we study on a genome-wide scale the requirement for Armadillo (Arm, homolog of beta-catenin) and Pangolin (Pan, Drosophila’s TCF) in the Wnt/Wingless (Wg)-induced transcriptional response of Drosophila Kc cells. Using somatic genetics, we demonstrate that both Arm and Pan are absolutely required for mediating activation and repression of target genes. Furthermore, by means of STARR-sequencing we identified Wnt/Wg-responsive enhancer elements and found that all responsive enhancers depend on Pan. Together, our results confirm the dogma of canonical Wnt/Wg signaling and argue against the existence of distal pathway branches in this system.

overall_design: STARR-seq was performed in wild type Drosophila Kc167 cells and Pangolin mutated Drosophila Kc167 cells treated with CHIR99021 or DMSO in two replicates.

first: Daria

last: Shlyueva


title: Ribosome footprinting of fly heads

city: Waltham

first: Sebastian

last: Kadener

address: 415 South Street

institute: Brandeis University

summary: We applied ribosomal footprinting experiment followed by RNA-seq  in order to detect circular RNA associated with ribosomes in fly heads.

overall_design: Ribosome profiling of Drosophila fly heads. Two samples were treated with low detergent buffer and the other two with high detergent buffer.


title: Blockade of IFN-alpha by antibody or CYM-5442 reveals a unique transcriptional gene signature in autoimmune CD8+ T cells and pancreatic islets

city: San Diego

first: Jaroslav

last: Zak

address: 10550 North Torrey Rd

department: Immunology and Microbial Sciences

institute: The Scripps Research Institute

summary: Blockade of interferon (IFN)-alpha but not IFN-beta signaling using either an antibody or a selective S1PR1 agonist, CYM-5442, prevented T1D in the mouse Rip-LCMV T1D model. First, treatment with antibody or CYM-5442 limited the migration of autoimmune “antiself” T cells to the external boundaries around the islets and prevented their entry into the islets so they could not be positioned to engage, kill and thus remove insulin-producing β cells. Second, CYM-5442 induced an exhaustion signature in antiself T cells by upregulating the negative immune regulator receptor genes Pdcd1, Lag3, Ctla4, Tigit and Btla, thereby limiting their killing ability. By such means, insulin production was preserved and glucose regulation maintained, and a novel mechanism for S1PR1 immunomodulation described.

overall_design: Purified pancreatic islets or P14 CD8+ T cells from the spleens of Rip-LCMV mice treated with IgG isotype control, anti-IFN-alpha or CYM-5442

first: Brett

last: Marro


title: CRISPR-Cas9 epigenome editing enables high-throughput screening for functional regulatory elements in the human genome

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

Large genome mapping consortia and thousands of genome-wide association studies have identified non-protein coding elements in the genome as a having a central role in tissue development, cell-type specification, response to environmental or pharmacologic signals, and susceptibility to most common diseases. However, decoding the function of the millions of putative regulatory elements discovered in these studies remains a primary challenge. New CRISPR/Cas9-based epigenome editing technologies have enabled the precise perturbation of the activity of specific regulatory elements. Here we describe CRISPR/Cas9-based Epigenomic Regulatory Element Screening (CERES) for high-throughput screening of regulatory element activity within the native genomic context. We perform both loss- and gain-of-function screens with complementary epigenome editing tools to identify known and unknown regulatory elements of medically relevant genes in human cells. The high-throughput functional annotation of putative regulatory elements by CERES constitutes a new platform for screening biological mechanisms that cannot be perturbed by traditional methods.


title: Systematic Functional Perturbations Uncover a Prognostic Genetic Network Driving Human Breast Cancer [ChIP-Seq]

city: Boston

first: Kenneth

last: Ross

address: 450 Brookline Ave., Rm M640

department: Pediatric Oncology

institute: Dana-Farber Cancer Institute

summary: Gene-expression patterns of primary breast cancers aid clinicians in predicting the risk of metastatic disease. Some prognostic signatures have recently been prospectively validated, highlighting their clinical value. Such classifiers conceivably comprise biomarker genes that, in fact, functionally contribute to the oncogenic and metastatic properties of the tumors, but this has not been investigated systematically. We previously reported that the transcription factor Fra-1 not only has an essential role in breast cancer, but also drives the expression of a highly prognostic gene set. Here, we systematically perturbed the function of 31 individual Fra-1-dependent poor-prognosis genes and examined their impact on breast cancer growth in vivo. Because of the considerable number of genes in this gene set, we anticipated that the contribution of single genes to breast cancer progression would be limited. In contrast, we find that stable shRNA depletion of each of nine individual signature genes strongly inhibits breast cancer growth and aggressiveness. Several factors within this nine-gene set regulate each other's expression, suggesting that together they form a network. The nine-gene set is regulated by estrogen, ERBB2 and EGF signaling, all established breast cancer factors. We also uncover three transcription factors, MYC, E2F1 and TP53, which act alongside Fra-1 at the core of this network. ChIP-Seq analysis reveals that a substantial number of genes are bound, and regulated, by all four transcription factors. The nine-gene set retains significant prognostic power and includes several potential therapeutic targets, including the bifunctional enzyme PAICS, which catalyzes purine biosynthesis. Depletion of PAICS largely cancelled breast cancer expansion, exemplifying a prognostic gene with breast cancer activity. Our data uncover a core genetic and prognostic network driving human breast cancer. We propose that pharmacological inhibition of components within this network, such as PAICS, may be used in conjunction with the Fra-1 prognostic classifier towards personalized management of poor prognosis breast cancer.

overall_design: ChIP-Seq experiment to examine binding of transcription factors in metastatic breast cancer cell line (MDA-MB-231 LM2)

first: Tristan

last: Gallenne

first: Nils

last: Visser

first: Sridhar

last: Ramaswamy

first: Daniel

last: Peeper


title: Systematic Functional Perturbations Uncover a Prognostic Genetic Network Driving Human Breast Cancer [RNA-Seq]

overall_design: RNA-Seq experiment to examine mRNA levels in metastatic breast cancer cell line (MDA-MB-231 LM2) after treatment with shRNA for scrambled control or shRNA for transcription factors for FOSL1 (Fra-1), E2F1, TP53, MYC, or EZH2 (two biological replicates for each gene and control)


title: A family of double-homeodomain transcription factors promotes zygotic genome activation in placental mammals [RNA-seq]

city: Lausanne

first: Evarist

last: Planet

state: Vaud

address: Route Cantonale

institute: EPFL

summary: In metazoans, the fertilized embryo is transcriptionally mostly silent for a few cell divisions, until release of a first major wave of embryonic transcripts by so-called zygotic genome activation (ZGA). Maternally provided ZGA-triggering factors have been identified in Drosophila melanogaster and Danio rerio but their mammalian homologues are still undefined. Here, we reveal that DUX family transcription factors are key to this process in human and mouse. First, human Dux4 and murine Dux are expressed prior to ZGA in both species. Second, both orthologues bind to and activate the promoters of ZGA genes. Third, Dux knockdown hinders the progression of mouse embryos beyond the 2-cell stage. Fourth, it prevents the cycling of mouse embryonic stem (ES) cells through a 2-cell-like state whereas, conversely, Dux overexpression induces the accumulation of these ZGA genes-expressing ES cells. We conclude that DUX proteins are master regulators of mammalian ZGA.

overall_design: Transcriptome profiling of control and KAP1 KD mouse ES containing a MERVL-GFP reporter construct and depleted for the Dux transcription factor

first: Alberto

last: De Iaco


title: A family of double-homeodomain transcription factors promotes zygotic genome activation in placental mammals [ChIP-seq_d2]

overall_design: ChIP-sequencing analysis for endogenous Trim28 in mESCs and for H3K9me3 in mESC depleted of Trim28 or control


title: A family of double-homeodomain transcription factors promotes zygotic genome activation in placental mammals [ChIP-seq_d1]

overall_design: ChIP-sequencing analysis of DUX4 ectopically expressed in human embryonic stem cells


title: Liver miRNA profile of PiZ mice

city: Napoli

first: Annamaria

last: Carissimo

address: via Castellino

institute: Tigem

summary: Transgenic PiZ mice have been genetically engineered to express ATZ and have been a valuable experimental model for liver disease due to AAT polymerization. ATZ accumulates in these mice within the ER of hepatocytes in a nearly identical manner to livers of affected patients.   MiRNAs play a key role in a wide range of biological processes and regulate gene expression mainly at the translational level. To search for miRNA with altered expression in AAT liver disease, we analyzed by miRNA next generation sequencing (NGS) the livers of 6-week-old PiZ mice and strain-, age-, and gender-matched wild-type mouse controls. This analysis revealed 70 miRNAs with differential expressions.

overall_design: Liver miRNA profiles of 6 week-old wild-type (WT) and PiZ mice were generated in dupicate by next generation sequencing

first: Pasquale

last: Piccolo

first: Nicola

last: Brunetti-Pierri


title: Global rewiring of cis-regulatory units upon lineage commitment of human embryonic stem cells

first: Steven

middle: William

last: Wingett

state: Cambs

address: Francis Crick Avenue, Cambridge Biomedical Campus

department: Cell Biology

institute: MRC Laboratory of Molecular Biology

summary: Lineage commitment is characterised by orchestrated changes in gene expression and chromatin state. Long-range elements such as enhancers coordinate lineage-specific transcriptional programmes by engaging in DNA looping interactions with target promoters. However, the target genes of most long-range elements remain unknown, hindering an integrated understanding of cis-regulatory gene control. Here, we generate a high-resolution atlas of chromosomal interactions involving ~22,000 gene promoters in human pluripotent and lineage-committed cells, identifying putative target genes for known and predicted enhancer elements. We jointly consider promoters and their associated interacting regions as “cis-regulatory units” that potentially integrate and stabilise regulatory inputs from individual elements. We reveal extensive dynamics of cis-regulatory units upon lineage commitment, including the acquisition and loss of promoter interactions, as well as chromatin state changes at preformed interactions. Finally, we show that reconfiguration of cis-regulatory units associates with changes in target gene expression. Our results therefore provide a high-resolution view of promoter interactome dynamics during lineage commitment and provide insights into the mechanisms of developmental transcriptional regulation.

overall_design: Two human ESC CHi-C biological replicates were generated (comprising 1 and 2 technical replicates) and a human ESC Hi-C dataset. Two human NEC CHi-C biological replicates were generated and a human NEC Hi-C dataset.   Two RNA-Seq biological replicates for both human ESC and NEC were generated.

first: Paula

last: Freire-Pritchett

first: Stefan

last: Schoenfelder

first: Csilla

last: Várnai

first: Jonathan

last: Cairns

first: Amanda

last: Collier

first: Raquel

last: Garcia

first: Cameron

last: Osborne

first: Peter

last: Fraser

last: Rugg-Gunn

first: Mikhail

last: Spivakov


title: A family of double-homeodomain transcription factors promotes zygotic genome activation in placental mammals.

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

overall_design: Refer to individual Series