title: Role of miR-146a in neural stem cell differentiation and neural lineage determination: relevance for neurodevelopmental disorders

city: Paris

first: Lam Son

last: Nguyen

state: IDF

address: 24 Bld du Montparnasse

department: Institute Imagine

institute: INSERM U1163

summary:
Background: MicroRNAs (miRNAs) are small, non-coding RNAs that regulate gene expression at the post-transcriptional level. miRNAs have emerged as important modulators of brain development and neuronal function and are implicated in several neurological diseases. Previous studies found miR-146a upregulation is the most common miRNA deregulation event in neurodevelopmental disorders such as autism spectrum disorders (ASD), epilepsy and intellectual disability (ID). Yet, how miR-146a upregulation affects the developing fetal brain remains unclear.
Methods: We analyzed the expression of miR-146a in the temporal lobe of ASD children using Taqman assay. To assess the role of miR-146a in early brain development, we generated and characterized stably induced H9 human neural stem cell (H9 hNSC) overexpressing miR-146a using various cell and molecular biology techniques.
Results: We first showed that miR-146a upregulation occurs early during childhood in the ASD brain. In H9 hNSC, miR-146a overexpression enhances neurite outgrowth and branching and favors differentiation into neuronal like cells. Expression analyses revealed that ten percent of the transcriptome was deregulated and organized into two modules critical for cell cycle control and neuronal differentiation. Twenty known or predicted targets of miR-146a were significantly deregulated in the modules, acting as potential drivers. The two modules also display distinct transcription profiles during human brain development, affecting regions relevant for ASD including the neocortex, amygdala and hippocampus. Cell type analyses indicate markers for pyramidal and interneurons are highly enriched in the deregulated gene list. Up to 40% of known markers of newly defined neuronal lineages were deregulated, suggesting that miR-146a could participate also in the acquisition of neuronal identities.
Conclusion: Our results demonstrate the dynamic roles of miR-146a in early neuronal development and provide new insight into the molecular events that link miR146a overexpression to impaired neurodevelopment. This, in turn, may yield new therapeutic targets and strategies.

overall_design: For each cell line, we used technical triplicates. The samples are matched based on passage number.

first: Lam

first: Julien

last: Fregeac

first: Christine

last: Bole-Feysot

last: Cagnard

first: Anand

first: Jasper

last: Anink

first: Eleonora

last: Aronica

first: Olivier

last: Alibeu

first: Patrick

last: Nitschke

first: Laurence

last: Colleaux


title: Transcriptomic changes in EpCAM+ liver ductal cells cultured in organoid conditions

status: Public on Jun 27 2018

city: Cambridge

first: Charles

last: Bradshaw

address: Tennis Court Road

department: Gurdon Institute

country: United Kingdom

institute: University of Cambridge

type: Expression profiling by high throughput sequencing

summary: RNA profile of EpCAM+ sorted cells freshly isolated from healthy liver of  adult mice was performed upon culture in liver organoid culture conditions

first: Luigi

last: Aloia

first: Meritxell

last: Huch


title: RNA-seq of cells with TET1 knockout

status: Public on Jun 26 2018

city: Camden

first: Jozef

last: Madzo

state: NJ

address: 403 Haddon Avenue

department: Bioinformatics

country: USA

institute: Coriell Institute

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

overall_design: Refer to individual Series


title: Cerebellar gene expression in Δα6-Cre Crfr1 knock-out mice

status: Public on Jun 25 2018

city: Munich

first: Mareen

last: Engel

address: Kraepelinstr. 2-10

department: Stress Neurobiology and Neurogenetics

country: Germany

institute: Max-Planck-Institute of Psychiatry

summary: Corticotropin-releasing factor (CRF) and its type 1 receptor (CRFR1) play an important role in the behavioral and neuroendocrine responses to stressful challenges, facilitating or down-playing specific functions dependent on the region and goal involved. Despite the well-established expression CRFR1 in granular cells (GrCs), the role of this system in procedural motor performance and memory formation remains elusive. To investigate the role of CRFR1 expression in cerebellar GrCs, we conducted mRNA-Seq in a mouse model specifically depleted of CRFR1 in these cells (Δα6-Cre Crfr1).

overall_design: mRNA-Seq of 6 wild-type (WT) and 6 conditional knock-out (cKO) mouse cerebella.

first: Gili

last: Ezra-Nevo

first: Dena

last: Leshkowitz

first: Jan

last: Deussing

first: Alon

last: Chen


title: DNA methylation oscillations at CpG poor enhancers in primed embryonic stem cells [PBAT Bisfulfite-seq]

first: Felix

last: Krueger

address: Granta Park

institute: Altos Labs

type: Methylation profiling by high throughput sequencing

summary: Mouse embryonic stem cells (ESCs) primed for differentiation display dynamic heterogeneity characterised by stochastic switching between transcriptional states, and recent advances have highlighted the heterogeneous and dynamic nature of DNA methylation in these. Using single cell sequencing we report global oscillations in ESC DNA methylation that affect particularly CpG poor regions including distal enhancers. These oscillations are dependent on DNA methylation turnover by Dnmt3 and Tet enzymes and influence the probability of transcriptional state switching. Our observations suggest that regulated DNA methylation heterogeneity may contribute to lineage priming in cells poised for differentiation.

overall_design:
2i_release: A long-term culture of 2i/Lif mouse embryonic stem cells were transferred to serum/Lif culture conditions and triplicate samples were collected over a time course of 30 time points.

NanogGFP: NanogGFP (TNGA) mouse embryonic stem cells were sorted into Nanog-high and Nanog-low subpopulations. After 2 days of recapturing, a second cell sort was performed yielding 4 populations of cells. Data are from triplicate experiments.

first: Steffen

last: Rulands

first: Heather

middle: J

last: Lee

first: Stephen

last: Clark

first: Christof

last: Angermueller

first: Sebastien

middle: A

last: Smallwood

first: Gavin

last: Kelsey

first: Oliver

last: Stegle

first: Benjamin

middle: D

last: Simons

first: Wolf

last: Reik


title: Endogenous transcripts control miRNA levels and activity in mammalian cells by a target-dependent miRNA degradation mechanism

city: Milano

first: Francesco

last: Nicassio

address: Via Adamello 16

department: Center for genomic science

country: Italy

institute: Istituto Italiano di Tecnologia

type: Non-coding RNA profiling by high throughput sequencing


title: Expression profile of hematopoietic stem and progenitor cells (HSPCs) after conditional deletion of the histone 3 lysine 4 (H3K4) methyltransferase Setd1b in mice

city: Dresden

first: Konstantinos

last: Anastassiadis

address: Tatzberg 47

department: Biotechnology Center

institute: Technische Universitaet Dresden

summary: Loss of Setd1b in adult mice using both ubiquitous and hematopoietic-specific deletion strategies caused lethality arising from severe hematopoietic defects. To identify genes regulated by Setd1b in HSPCs we performed RNA-seq analysis of the LSK (Lin-, Sca+, Kit+) compartment in the bone marrow. Mice carrying heterozygous and homozygous conditional Setd1b alleles and Rosa26-Cre-ERT2 were treated with Tamoxifen for 6 days (4 days gavage - 3 days pause - 2 days gavage). Mice were sacrificed 3 days after the last treatment and RNA was isolated and processed for next generation sequencing. We found that Setd1b regulates key lineage specification components including Cebpa, Gata-1 and Klf-1.

overall_design: HSPC (LSK) mRNA profiles of Setd1b control (heterozygous) and knockout (homozygous) littermates were generated 3 days after the last tamoxifen administration, in triplicates.

first: Kerstin

last: Schmidt


title: p21 misexpression in the left growth plates

first: Alberto

last: Rosello-Diez

address: Physiology building. Downing St.

department: Physiology, Development and Neuroscience

summary: To study mechanisms of long bone growth regulation, p21 misexpression was induced in the left hindlimb of mouse embryos using an intersectional approach that requires both Cre and (r)tTA activity. Doxycycline was provided to the pregnant female at embryonic day (E)12.5 to activate the transgene, and embryos were collected at E17.5. Distal femur and proximal tibia growth plates were dissected out, keeping left and right separated, deprived of perichondrium and flash frozen. After RNA extraction, mRNA libraries were prepared and all samples were deep sequenced in parallel

overall_design: 6 samples (left and right growth plates from embryos #386, #387, #388) were sequenced in parallel

first: Alexandra

middle: L

last: Joyner

last: Roselló-Díez


title: HNF4alpha P1 and P2 isoforms regulate different gene networks in the colorectal cancer cell line Caco2/15.

city: Sherbrooke

first: Francois

last: Boudreau

state: Quebec

address: 3201 rue Jean-Mignault

department: Anatomie et biologie cellulaire

country: Canada

institute: Université de Sherbrooke

summary: The nuclear receptor HNF4alpha is a master regulator of gene transcription in intestinal epithelial cells where P1 and P2 isoforms are expressed. HNF4alpha has been associated with colorectal cancer  where its role remains controverisal since both tumor suppressor and  oncogene characteristics have been observed. Evidences gathered from mouse models suggested however that this controversy on HNF4alpha function in colorectal cancer could be explained by the different functions of its two isoforms classes. In order to clarify this hypothesis in a human context, we downregulated the expression of HNF4alpha P1 or P2 isoforms using isoforms specific shRNA in the colorectal cancer cell line Caco2/15. Transcriptomic data were obtained by RNA-seq from these shRNA cell populations and compared to a non-target shRNA control.

overall_design: Colorectal cancer cell line Caco2/15 (clone of Caco-2) was infected for 48 hours with lentiviral shRNAs targeting HNF4alpha P1 or P2 isoforms classes or a control shRNA targeting no human genes. Lentiviruses were removed and cells incubated for another 24 hours before extracting RNA. Cells reached one day post-confluence at the time RNA was extracted. RNA librairies were prepared using TrueSeq mRNA kit (Illumina) and paired reads generated using Illumina Hiseq 2000/2500 sequencer. Base calls were made using Illumina CASAVA pipeline. Three biological independant replicates were analyzed in each condition.

first: Jean-Philippe

last: Babeu

first: François


title: Genome-wide analysis reveals dual roles for Ikaros in regulation of macrophage chromatin state and inflammatory gene expression

city: Bethesda

first: Songjoon

last: Baek

state: MD

address: 41 Library Drive

department: CCR

institute: NCI / NIH

type: Genome binding/occupancy profiling by high throughput sequencing


title: RNA-seq of cytosolic and chromatin-associated transcripts following TNFα and Spt5 KD

city: Rehovot

first: Rivka

last: Dikstein

address: 2 Hertzl st.

country: Israel

institute: The Weizmann Institute of Science

summary: We examined the effects of TNFα and Spt5, the major DSIF subunit, on nascent and mature transcripts using RNA-Seq of chromatin-associated and cytoplasmic transcripts.

overall_design: RNA was extracted from the cytosolic and chromatin fractions of control and Spt5 KD cells that were treated with TNFα for 1 hour

first: Gil

last: Diamant


title: zUMIs: a fast and flexible pipeline for RNA sequencing data with UMIs

status: Public on Jun 22 2018

city: Martinsried

first: Christoph

last: Ziegenhain

state: Bavaria

address: Großhaderner Str. 2

department: Department Biology II

institute: Ludwig-Maximilians University Munich

summary:
Background
Single-cell RNA-sequencing (scRNA-seq) experiments typically analyze hundreds or thousands of cells after amplification of the cDNA. The high throughput is made possible by the early introduction of sample-specific bar codes (BCs), and the amplification bias is alleviated by unique molecular identifiers (UMIs). Thus, the ideal analysis pipeline for scRNA-seq data needs to efficiently tabulate reads according to both BC and UMI.

Findings
zUMIs is a pipeline that can handle both known and random BCs and also efficiently collapse UMIs, either just for exon mapping reads or for both exon and intron mapping reads. If BC annotation is missing, zUMIs can accurately detect intact cells from the distribution of sequencing reads. Another unique feature of zUMIs is the adaptive downsampling function that facilitates dealing with hugely varying library sizes but also allows the user to evaluate whether the library has been sequenced to saturation. To illustrate the utility of zUMIs, we analyzed a single-nucleus RNA-seq dataset and show that more than 35% of all reads map to introns. Also, we show that these intronic reads are informative about expression levels, significantly increasing the number of detected genes and improving the cluster resolution.

Conclusions
zUMIs flexibility makes if possible to accommodate data generated with any of the major scRNA-seq protocols that use BCs and UMIs and is the most feature-rich, fast, and user-friendly pipeline to process such scRNA-seq data.

overall_design: HEK293T cells were sequenced using the mcSCRB-seq protocol (Bagnoli et al., 2017)

first: Swati

last: Parekh


title: Prep1 and Pbx2/4 dependent genes in zebraffish embryos

city: Worcester

first: Franck

last: Ladam

state: Massachusetts

address: 364 plantation street , LRB 870D

department: Biochemistry and Molecular Pharmacology

institute: University of Massachusetts Medical School

summary: Identification of Prep1 and Pbx2/4 regulated genes during early zebrafish embryogenesis (6 and 12hpf) by RNA-seq

overall_design: zebrafish embryos were injected with morpholinos targeting Prep1, Pbx2/4 (TALE MO) mixed with water and red phenol dye or injected with water and red phenol dye alone (Control). Then total RNA was extracted from 6 or 12hpf injected embryos and polyA+RNA libraries constructed using the TruSeq stranded RNAseq LT kit from illumina

middle: G

first: William

last: Stanney

first: Ian

last: Donaldson

first: Nicoletta

last: Bobola

first: Charles

last: Sagerstrom


title: DNA methylation oscillations at CpG poor enhancers in primed embryonic stem cells [single cell Bisulfite-seq]

first: Felix

last: Krueger

overall_design: scBS-Seq: A total of 81 single cells from 5 different cell populations

first: Steffen

last: Rulands

first: Heather

last: Lee

first: Stephen

last: Clark

first: Christof

last: Angermueller

first: Sebastien

last: Smallwood

first: Gavin

last: Kelsey

first: Oliver

last: Stegle

first: Benjamin

last: Simons

first: Wolf

last: Reik


title: Genome-wide analysis reveals dual roles for Ikaros in regulation of macrophage chromatin state and inflammatory gene expression [RNA-seq]

first: Songjoon

last: Baek

summary: Macrophage activation by bacterial LPS leads to induction of a complex inflammatory gene program dependent on numerous transcription factor families. Here we describe an unexpected role for the lymphoid lineage-determining factor Ikaros in the macrophage LPS response using comprehensive genomic analysis of Ikaros-dependent transcription, DNA binding, and chromatin accessibility. Ikaros exhibited dual repressor and activator roles in LPS-induced gene expression. Consistent with the described function of Ikaros as transcriptional repressor, Ikzf1-/- cells showed enhanced induction for select responses. In contrast, we observed a dramatic defect in expression of delayed response genes and ChIP-seq analyses support a key role for Ikaros in sustained NF-kB chromatin binding. Furthermore, in the absence of Ikaros, a constitutively open chromatin state was coincident with dysregulation of LPS-induced chromatin dynamics, gene expression, and cytokine responses. Our data suggest a central role for Ikaros in coordinating the complex macrophage transcriptional program in response to pathogen challenge.

overall_design: RNA-seq time course (0h, 2h, 4h, 10h) and DNase-seq time course (0h, 3h, 8h) sample replicates from WT and Ikzf KO macrophages; RelA/Ikaros ChIP-seq time course (0h, 3h, 8h) sample replicates and input control DNA samples

first: Kyu-Seon

last: Oh

first: Rachel

last: Gottschalk

first: Nicolas

middle: W

last: Lounsbury

first: Jing

last: Sun

first: Ze

last: Wang

first: Bhaskar

last: Dutta

first: Gordon

last: Hager

first: Myong-Hee

last: Sung

first: Iain

last: Fraser


title: Transcriptome data involved in Chinese tongue sole (Cynoglossus semilaevis) sexual growth dimorphism

city: Qingdao

state: Shandong

address: 106 Nanjing Road

country: China

institute: Yellow Sea Fisheries Research Institute, Chinese Academy Fishery Science

summary: In the present study, four important somatotropic and reproductive tissues including brain, pituitary, liver, and gonad from 15 females and 15 males were used for transcriptome analysis by RNA-seq. A mean of 37,533,991 high quality clean reads were obtained from each library and 806, 1,482, 818, and 14,695 differentially expressed genes in female and male were identified from brain, pituitary, liver, and gonad, respectively (fold change ≥ 2 and q < 0.05). GO terms and KEGG pathways enrichment analysis revealed nucleic acid binding transcription factor activity, G-protein coupled receptor activity, MAPK signaling pathway, steroid biosynthesis, and neuroactive ligand-receptor interaction may involve in sexual growth differences.

overall_design: Examination of four important somatotropic and reproductive tissues including brain, pituitary, liver and gonad from 15 females and 15 males individuals

first: Renkai

first: Ruoqing

first: Songlin

last: Chen


title: Transcriptomic sequencing after MALAT1 knockdown in A431 cells

status: Public on Jun 23 2018

city: Guangzhou

first: Lin

last: Gao

state: Guangdong province

address: No. 1023, South Shatai Road

department: School of Public Health

institute: Southern Medical University

summary: Cutaneous squamous cell carcinoma (CSCC) is the most common metastatic skin cancer, and its incidence is increasing globally. In this study, metastasis-associated in lung adenocarcinoma transcript 1 (MALAT1), a long non-coding RNA, was identified to be significantly up-regulated in CSCC tumors and cell lines. We conducted RNA sequencing to characterize gene expression changes in MALAT1-silenced cutaneous squamous cell carcinoma cells (A431).

overall_design: RNA-Seq analysis of CSCC cells followed by siRNA-induced gene knockdown of MALAT1. Total RNAs from negative control (NC ASO) and MALAT1 ASO-treated CSCC cells (antisense oligonucleotide) were extracted. The samples were sequenced using Illumina sequencing.

first: Zhenhua

last: Ding

first: Liang

last: Zhou


title: Temporal Changes in Macrophage Phenotype after Peripheral Nerve Injury

status: Public on Jun 21 2018

city: Ithaca

first: Jennifer

middle: K

last: Grenier

state: NY

address: 526 Campus Rd

department: Biomedical Sciences

institute: Cornell University

summary: Identification of temporal changes in gene expression in macrophages isolated from the site of nerve injury.

overall_design: Macrophages were profiled at 3 timepoints (5, 14, and 28 days) after nerve injury with 2-3 independent biological replicates per timepoint.

first: Jonathan

last: Cheetham

first: Joy

middle: E

last: Tomlinson


title: Transcriptomic analysis of senescent cells upon PTBP1 knockdown and EXOC7 knockdown

city: London

first: Gopuraja

last: Dharmalingam

address: Faculty of Medicine, Imperial College, Hammersmith Hospital Campus

department: Epigenetics section

institute: MRC London Institute of Medical Sciences


title: Transcriptomic analysis of senescent cells upon EXOC7 knockdown.

summary: IMR90 ER:RAS cells were transfected with scramble siRNA or 2 deconvoluted siRNAs targeting EXOC7 or an siRNA targeting PTBP1. The next day the cells were treated with 4OHT to induce senescence. 5 days later the cells were collected for total mRNA analysis. Our previous experiments had shown that alternative splicing of EXOC7 depends on PTBP1 and PTBP1 is a target for suppressing pro-inflammatory factors in senescence. By performing RNA-seq we confirmed that the pro-inflammatory factors whose expression depends on PTBP1 equally depends on EXOC7.

overall_design: 5 conditions were examined: proliferating cells (si_Scramble), senescent cells (si_Scramble + 4OHT), senescent cells transfected with an siRNA targeting EXOC7 (si_EXOC7_3 + 4OHT), senescent cells transfected with a different siRNA targeting EXOC7 (si_EXOC7_4 + 4OHT) and senescent cells transfected with an siRNA targeting PTBP1 (si_PTBP1_5 + 4OHT).

first: Athena

last: Georgilis

first: Jesús

last: Gil

first: Thomas

last: Carroll


title: Transcriptomic analysis of senescent cells upon PTBP1 knockdown.

summary: IMR90 ER:RAS cells were stably transduced with either an empty vector or 2 deconvoluted shRNAs targeting PTBP1. Following selection with puromycin, the cells were treated with 4OHT to induce senescence. 6 days later the cells were collected for total mRNA analysis. PTBP1 is a regulator of alternative splicing. Our previous experiments had shown that PTBP1 depletion inhibits the expression of pro-inflammatory genes without affecting other senescence-associated phenotypes. By performing RNA-seq we confirmed those observations at a global level and analysed how PTBP1 knockdown alters alternative splicing as a potential mechanism of action.

overall_design: 4 conditions were examined: proliferating cells (Vector), senescent cells (Vector + 4OHT), senescent cells expresssing an shRNA targeting PTBP1 (shPTBP1_53 + 4OHT) and senescent cells expressing a different shRNA targeting PTBP1 (shPTBP1_86 + 4OHT).


title: Transcriptomic analysis of SM21 resistant and susceptible haploid Candida albicans strains

city: Singapore

first: Thuyen

last: Truong

address: 11 Lower Kent Ridge Road

department: Faculty of Dentistry

country: Singapore

institute: National University of Singapore

summary: A comprehensive RNASeq analysis was performed on the haploid parent GZY803 and strains resistant to SM21 antifungal. Results imply the mechanisms underlying resistance responses.

overall_design: Wild type GZY803 and resistant strains were exposed to 2 µg/ml SM21 for 1h and 4h. RNA from treated and untreated cultures were extracted for RNASeq analysis.

first: Tanujaa

last: Suriyanarayanan

first: Guisheng

last: Zeng

first: Thuc

last: Le

last: Liu

first: Jiuyong

last: Li

first: Cao

last: Tong

first: Yue

first: Chaminda

last: Seneviratne


title: Capturing the onset of PRC2-mediated repressive domain formation (RNA-Seq)

city: New York

first: Danny

last: Reinberg

address: 522 First Avenue

institute: NYU School of Medicine

summary: Our study defines nucleation and spreading regions for Polycomb repressive complex 2 (PRC2), demonstrating the principle of PRC2 domain formation in mammals. We elucidate the role of genome architecture in formation of these domains and identify JARID2 and MTF2 as being crucial for full recruitment of PRC2 to chromatin.

overall_design: In this experiment, we performed RNA-seq in mESCs and embryoid bodies with indicated genotypes.

first: Ozgur

last: Oksuz

first: Varun

last: Narendra

first: Chul-Hwan

last: Lee

first: Nicolas

last: Descostes

first: Gary

last: LeRoy

first: Ramya

last: Raviram

first: Kelly

last: Karch

first: Pedro

middle: R

last: Rocha

first: Benjamin

last: Garcia

first: Jane

last: Skok


title: Gene expression analysis in mouse mature T-cells either upon overexpression of the NICD or knockout of H2A.Z

city: Giessen

first: Marek

last: Bartkuhn

state: Hessen

address: Aulweg 132

department: Biomedical Informatics and Systems Medicine

institute: Justus-Liebig-University Giessen

summary: Genes induced by the Notch-Intracellular domain (NICD) in mouse mature T-cells (MT) are identifed. Among the induced Notch target genes we also identify those genes, which are regulated by the histone variant H2A.Z

overall_design: RNA-Seq

first: Benedetto

last: Giaimo

first: Tobias

last: Zimmermann


title: New BRD4-inhibitor quinoline compound targets a distinct latent HIV-1 reservoir for re-activation from canonical “shock” drugs

city: Barcelona

first: Guillaume

last: Filion

address: Dr. Aiguader 88

department: GRSCC

country: Spain

institute: CRG

summary: Upon HIV-1 infection, a reservoir of latently infected resting T cells prevents the eradication of the virus from patients. To achieve complete depletion, the existing virus-suppressing antiretroviral therapy must be combined with drugs that reactivate the dormant viruses. We previously described a novel chemical scaffold compound, MMQO (8-methoxy-6-methylquinolin-4-ol) that is able to reactivate viral transcription in J-Lat T cell model of latency through an unknown mechanism. Here we demonstrate that MMQO activates HIV-1 in ex vivo latently infected primary CD4 T cells, where it synergizes with the PKC agonist prostratin. Gene expression microarray analysis in Jurkat cells indicated that MMQO treatment results in robust immunosuppression, affects cell proliferation by rapidly diminishing expression of the transcription factor c-Myc, and causes the dysregulation of acetylation sensitive genes. These hallmarks indicated that MMQO mimics acetylated lysines of core histones and might function as a bromodomain and extraterminal domain protein family inhibitor (BETi). MMQO functionally mimics the effects of JQ1, a well-known BETi. We confirmed that MMQO interacts with the BET family bromodomain BRD4. Utilizing MMQO and JQ1, we demonstrate how the inhibition of BRD4 targets a distinct subset of latently integrated barcoded proviruses from those targeted by HDAC inhibitors or PKC pathway agonists. Thus, the scaffold compound MMQO is a new member of the BET family inhibitors, which due to its minimalistic structure holds promise for further optimization for increased affinity and specificity for distinct bromodomain family members and could potentially be of use against a variety of diseases, including HIV.

overall_design: We used the system of Jurkat T cells with latent barcoded HIV infections and measured the fold induction of HIV expression by the drugs including MMQO, JQ1, Prostratin and SAMA through the B-HIVE technology. Each sample preparation and data analysis was done in parallel in 2 replicates. DMSO treatment as a negative control.

first: Heng-Chang

last: Chen

first: Erik

last: Abner

first: Eduard

last: Zorita

first: Albert

last: Jordan


title: Instability of WT AAV2 genome and AAV promoter activities in the Baculovirus/Sf9 cells system

city: Kuopio

first: Minna

middle: U

last: Kaikkonen

department: A.I. Virtanen Institute, Department of Biotechnology and Molecular Medicine

country: Finland

institute: University of Eastern Finland

summary: The human Adeno-Associated Virus serotype 2 (WT AAV2) is a common non-pathological virus and its recombinant form (rAAV) is widely used as gene therapy vector. However, it has been shown that WT AAV2 and recombinant AAV display significantly different characteristics, especially regarding infection rate, with a near perfect infectivity and better encapsidation rate of WT AAV2. Even though rAAVs are routinely produced in the Baculovirus/Sf9 cell system, WT AAV2 has never been produced in this context. To understand the infectivity and encapsidation rate differences between WT AAV2 and rAAV, we tried to produce WT AAV2 in baculovirus/Sf9 cells system hypothesizing that the WT AAV2 may be considered as a normal recombinant AAV transgene. Through our attempts to produce WT AAV2 in Baculovirus/Sf9, we found that WT AAV2 p5 promoter, which controls the expression of large Rep proteins in mammalian cells, was active in this system. p5 promoter activity in the baculovirus/Sf9 cell system led to the expression of Rep78 that finally excises WT AAV2 genome from the baculovirus genome during the earliest phase of baculovirus stock production. The p5 promoter expression kinetics and the specific strand RNA-Seq analysis of the WT AAV2, rAAV Rep2/Cap2 cassettes in the baculovirus context was performed. We demonstrate that the WT AAV2 native promoters, p5, p19 and p40 are all active and lead to the expression of different proteins and peptides. In addition, this study demonstrates that the baculovirus brings at least some of the helper functions needed in the AAV replication/life cycle.

overall_design: RNA-Seq analysis of the expression of WT AAV2, rAAV Rep2/Cap2 cassettes in the baculovirus system

first: Adrien

last: Savy

last: Leger

first: Yohann

last: Dickx

first: Lionel

last: Galibert

first: Otto

last: Merten


title: Zeb2 is Essential for Bergmann Glial Development and Cerebellar Organization

city: Cincinnati

first: Richard

last: Lu

state: OH

address: 3333 Burnet Ave

department: CBDI

institute: Cincinnati Children's Hospital Medical Center

summary: Cerebellar circuitry is critical for balance and motor control among a wide array of functions and largely consists of granule and Purkinje neurons. Bergmann glia in the cerebellum form distinct morphological structures that facilitate granule neuron migration during development and that maintain the cerebellar organization and functional integrity. At present, molecular control of the formation and morphogenesis of Bergmann glia remains obscure. In this study, we found that Zeb2 (a.k.a. Sip1 or Zfhx1b), a Mowat-Wilson syndrome-associated transcriptional regulator, is highly restricted to Bergmann glia and is essential for their development and maturation. The mice with Zeb2 ablation in the cerebellar neural progenitor exhibit dysgenesis of cerebellar cortical lamination and locomotion defects. Deletion of Zeb2 markedly reduced Bergmann glial proliferation, differentiation and the establishment of the normal radial scaffold, disrupting migration of granule cell progenitors from external to internal granular layers. Transcriptome profiling indicated that Zeb2 regulates multiple pathways including FGF and Notch signaling as well as axonal guidance cues including Netrin G2 and Gdf10 to control Bergmann glial development. Our data reveal that Zeb2 acts as a transcriptional integrator of diverse signaling pathways to regulate the formation and morphogenesis of Bergmann glia ensuring maintenance of cerebellar integrity, suggesting that Zeb2 dysfunction in Bergmann glia might contribute to motor deficits in Mowat-Wilson syndrome.

overall_design: 6 RNA-Seq samples from P0 cerebellum of Ctrl and Zeb2 CKO mice (3 Ctrl and 3 CKO).

first: Li

last: He

first: Haibo

last: Wang


title: Genome-scale DNA methylation oscillations in pluripotent cells

city: Cambridge

first: Felix

last: Krueger

address: Granta Park

department: Bioinformatics

institute: Altos Labs

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

overall_design: Refer to individual Series


title: Gene expression profiling in pre-leukemic hematopoietic stem cells carrying both NrasG12D/+ and Tet2+/- mutations

city: Ann Arbor

first: Qing

last: Li

state: Michigan

address: 109 Zina Pitcher Place, BSRB 1520

department: Internal Medicine

institute: University of Michigan

summary: By using a genetically accurate mouse model, we demonstrate that endogenous expression of oncogenic N-RasG12D and Tet2 haploinsufficiency collaborate to accelerate CMML development in mice. Gene expression was compared across all genotypes (WT, Tet2+/-, NrasG12D/+ and double mutants) in bone marrow-derived hematopoietic stem cells (CD150+CD48-Lin-Sca1+cKit+) using RNA-seq. N-RasG12D and Tet2 haploinsufficiency cooperate to induce both unique and overlapping effects on HSC gene expression programs.

overall_design: Gene expression profiling in FACS-sorted SLAM HSCs from 10-12 week old wild type control (n=3), NrasG12D/+ single mutant (n=3), Tet2+/- single mutant (n=3) and NrasG12D/+;Tet2+/- double mutant (n=3) mice.

first: Xi

last: Jin

first: Tingting

last: Qin

first: Maria

last: Figueroa


title: Capturing the onset of PRC2-mediated repressive domain formation (4C-Seq)

first: Danny

last: Reinberg

overall_design: In this experiment, we performed 4C-seq in mESCs using nucleation sites, spreading sites, and two control sites as viewpoints.

first: Ozgur

last: Oksuz

first: Varun

last: Narendra

first: Chul-Hwan

last: Lee

first: Nicolas

last: Descostes

first: Gary

last: LeRoy

first: Ramya

last: Raviram

first: Kelly

last: Karch

first: Pedro

last: Rocha

first: Benjamin

last: Garcia

first: Jane

last: Skok


title: Capturing the onset of PRC2-mediated repressive domain formation (ChIP-Seq)

overall_design: In this experiment, we performed ChIP-seq with the indicated antibodies and genotpyes of mESCs.


title: Regulation of alternative splicing induced by paclitaxel in human non-small cell lung cancer

status: Public on Jun 20 2018

city: Dalian

first: Yang

last: Wang

address: No.9 West section, South Lv Road

institute: Dalian Medical University

summary: RNA-seq was performed on A549 cells treated with paclitaxel or control, in order to profile the alternative splicing events that were regulated upon paclitaxel treatment.

overall_design: Examination of different alternative splicing events in 2 types of cells

first: Wang

last: Yang

first: Zhu

last: Ziran


title: Transcriptome analysis of the Golgi stress response

city: West Haven

first: Francesc

last: Lopez

state: CT

department: Department of Genetics

institute: Yale University

summary: We explore the transcriptional response of mammalian cells undergoing various insults to Golgi homeostasis. HEK293 cells (Flp-In T-REx 293 cells) stably containing a doxycycline-inducible Golgi-localized HaloTag2 construct (GA-HT2) were treated with the ionophore nigericin, the glycosylation inhibitor xyloside, or were induced by doxycycline and treated with the hydrophobic tag HyT36 to induce destabilization of GA-HT2. We found that while nigericin and xyloside induce global transcriptional changes, destabilization of GA-HT2 induces a Golgi-specific response.

overall_design: HEK293/GA-HT2 cells were treated in duplicate with nigericin, xyloside, HyT36, or the vehicle DMSO in the absence of doxycycline. In addition, cells were treated with DMSO or HyT36 in the presence of doxycyline.

first: Yevgeniy

middle: V

last: Serebrenik

first: Doris

last: Hellerschmied

last: López-Giráldez

first: Craig

middle: M

last: Crews


title: Metabolite and transcript profile of grapevine clusters at 6th,7th,8th WAA and the effect of 1-MCP on ripening

status: Public on Jun 17 2018

city: San Michele all'Adige

first: Carmen

last: Leida

address: E. Mach, 1

institute: Fondazione E. Mach

summary: We report the trasncriptomic profile of Pinot Noir at ethylene peak. The main aim is to understand the effect of 1-MCP on gene transcription and during this ripening window.

overall_design: Total RNA profile of CONTROL and TREATED from berry tissue were generated by deep sequencing in three/five biologial replicates with Illumina Hiseq2000. From the same plant (considered a biological replicate) treated and control were obtained, in order to minimize the variability due to individual.   Bioproject accession (PRJNA323698)

first: Joaquin

last: Cañizares

first: Claudio

last: Moser


title: ChIP-Seq analysis of NRF2 binding sites in human BEAS-2B cell line treated with sulforaphane or vehicle

status: Public on Jun 19 2018

city: Research Triangle Park

first: Xuting

state: NC

address: 111 TW Alexander Dr

institute: NIEHS/NIH

summary: Cellular oxidative and electrophilic stress triggers a protective response in mammals regulated by NRF2 (nuclear factor (erythroid-derived) 2-like; NFE2L2) binding to DNA-regulatory sequences near stress responsive genes. Studies using Nrf2-deficient mice suggest that hundreds of genes may be regulated by NRF2. To identify human NRF2-regulated genes, we conducted ChIP-sequencing experiments in human BEAS-2B cell line treated with the dietary isothiocyanate, sulforaphane (SFN) and carried out follow-up biological experiments on candidates.

first: Michelle

last: Campbell

first: Brian

middle: N

last: Chorley

first: Douglas

last: Bell


title: Constrained pioneering and partner factor redirection by PU.1 shape early T-cell gene regulation

city: Pasadena

first: Ellen

last: Rothenberg

state: CA

address: 1200 E. California Blvd

department: Division of Biology and Biological Engineering

institute: California Institute of Technology

summary: A major problem with linking transcription factor binding to function is that many factors bind to a large number of, at least in any given cell type, seemingly irrelevant regions. This makes it hard to filter out which binding sites are responsible for the regulation of a given gene. PU.1 (Spi1, Sfpi1) is an excellent example of a transcription factor that works both to mediate developmental choices and to serve the alternative developmental fates that emerge from these choices. Its role in T-cell development is confined to the early stages where high PU.1 expression persists through multiple cell divisions and is sharply downregulated over the DN2a-to-DN2b T-cell commitment stage. Even though it is known that PU.1 is necessary for the survival of the earliest T-cell progenitors, where it is needed for optimal proliferation, control of alternative lineage genes, and correct timing of the access to T-lineage genes, it has not been well-studied how PU.1 finds it targets and exerts its functions within this context.    Here, we show in detail how PU.1 selects its binding sites and subsequently regulate gene expression. We show that PU.1 has two effective affinity thresholds for occupancy depending on current chromatin openness as measured by ATAC-sequencing, but that it is easily capable of binding sites in closed chromatin. Unexpectedly, although its binding to promoters is least constrained, promoters are the only major class of sites where it exerts a predominantly negative effect; otherwise it works locally as an activator, mainly mediated through binding to sites in closed or dynamically closing distal enhancer elements, where it can rapidly open chromatin and induce histone acetylation. However, its ability to open the chromatin depends not only on its own affinity but also on the presence of collaborating factors, because we show that PU.1 introduction into PU.1-negative cells triggers a massive reorganization of occupancy patterns of at least three other factors: Runx1, Satb1, and Gata3. Most strikingly, PU.1’s “theft” of Runx1 and Satb1 from many sites where they were binding in the absence of PU.1 enables PU.1 to exert a novel form of repression even on genes where it has no binding sites itself. We show here that PU.1 requires domains outside of its DNA binding domain to properly open chromatin, and this structural requirement is directly connected with its ability to bind to Runx1 and to Satb1, and moreover, Runx1, in particular, is an important collaborator to activate many of its target genes in the context of early T-cell development. Thus, PU.1 regulates gene expression via two distinct mechanisms. First, PU.1 steals Satb1 and Runx1 from many genomic sites, thereby repressing T cell gene expression indirectly. Second, PU.1, opens chromatin, recruits Satb1 and Runx1 to new sites, and directly activates its target gene expression.    In summary, we here present a model where a transcription factor can work through redeployment of other factors and not only through sites that it binds itself.

overall_design:
ChIP-Seq:  Primary cell experiments:  Briefly, lin- FLPs were expanded for 7 days on OP9-DL1, CD25+ enriched and retrovirally infected with empty vector control or HA-tagged versions of Lzr-PU.1WT, -PU.1ENG, -PU.1ETS. Cells were co-cultured on OP9-DL1 for ~40h and then sorted as LiveGFP+CD25+ or LiveGFP+CD44+CD25- on BD FACSAriaTM.    Scid.adh.2c2:    HA-tagged-ChIP: Scid.adh.2c2 cells were infected with empty vector control or HA-tagged versions of Lzr-PU.1WT, -PU.1ENG, -PU.1ETS. Cells were expanded for ~40h and then sorted as LiveGFP+on BD FACSAriaTM.  4-OHT experiment: Scid.adh.2c2 cells were transduced with Lzr-PU.1ert2 (Scid.adh.2c2-PU.1ert2) or Lzr-ert2 (Scid.adh.2c2-ert2) control, cells were expanded for 48h and infected (GFP+) cells followed by sorting of GFP+ cells on BD FACSAriaTM and further expanded in vitro prior to 2, 8 or 24h activation with 0.1 μM 4-OHT (4-hydroxytamoxifen).  Satb1, Runx1, Fog1 and GATA3 ChIP: Scid.adh.2c2 cells were infected with retroviral supernatant of pMxs-Myc-Flag-PU.1-IRES-hNGFR and cultured for 48h.    Approximately 3-5 million primary sorted CD25+ -or 10 million Scid.adh.2c2 cells for ChIP were fixed at RT in 1 mg/ml DSG (Thermo Scientific) in PBS for 30 min followed by additional 10 min after addition of formaldehyde (FA) up to 1% (Satb1, Runx1, Fog1 and Ets1) or 10 min of 1% FA only (PU.1, HA,-tag, GATA3, H3-modifications). The reaction was quenched by addition of 1/10 volume of 0.125M glycine and the cells were washed in ice-cold 1xHBSS (Gibco). Cell pellets were snap frozen on dry ice and stored in -80°C. For DSG+FA crosslinked cells, nuclei were isolated by 10 min incubation in Nuclei Isolation buffer (50 mM Tris-pH 8.0, 60 mM KCl, 0.5% NP40) + protease inhibitor cocktail (PIC) (Roche) on ice. Pelleted nuclei were dissolved in 0.5% Lysis buffer (0.5% SDS, 10 mM EDTA, 0.5 mM EGTA, 50 mM Tris-HCl (pH 8)) + PIC. FA crosslinked cells only were resuspended 1% Lysis buffer (1% SDS, 10 mM EDTA, 50 mM Tris-HCl (pH 8). Lysates were sonicated on a Bioruptor (Diagenode) for 18 cycles, max power for 30s followed by 30 s rest. Sonication was followed by pelleting of debris and the supernatant was transferred to a new tube and chromatin was diluted 3X in 1xHBSS+PIC followed by 2xRIPA (2% Triton, 2mM EDTA, 200 mM NaCl, 20 mM Tris-HCl (pH 7.5), 1% sodium deoxycholate, 0.1%SDS + PIC). Approximately 10 μg per 107 cells of antibody (Rabbit anti-PU.1 polyclonal IgG (sc-352x, Santa Cruz Biotechnology), Rabbit HA-probe polyclonal IgG (sc-805x, Santa Cruz Biotechnology), Rabbit anti-Ets1 polyclonal IgG (sc-350x, Santa Cruz Biotechnology), Mouse anti-Gata3 monoclonal IgG (a mixture of sc-268, Santa Cruz Biotechnology and MAB26051, R&D Systems), Rabbit anti-Satb1 monoclonal IgG (ab109122, Abcam), Rabbit anti-Runx1 polyclonal IgG (ab23980, Abcam), Goat anti-Fog1 polyclonal IgG (sc-9361, Santa Cruz Biotechnolog), Rabbit polyclonal anti-H3K27Ac (ab4729, Abcam), Rabbit polyclonal anti-H3K4me2 (07-030, Millipore) or Rabbit polyclonal anti-H3K27me3 (07-449, Millipore) were hybridized to Dynabeads M-280 Sheep anti- Rabbit or Dynabeads M-280 Sheep anti- Mouse or Dynabeads Protein A/G (Invitrogen), respectively in 1ml 1xRIPA+PIC for 4h, washed twice with 1xRIPA, resuspended in 100 μl 1xRIPA+PIC and added to the diluted chromatin. For polyclonal and monoclonal antibodies, 20 and 50 μl of Dynabeads were used per μg antibody, respectively. ChIP was performed over night at 4°C, and subsequently washed (1 time with 1 ml Low Salt Immune Complex Wash Buffer (0.1% SDS, 1 % Triton-X, 2 mM EDTA, 50 mM Tris-Hcl pH8, 150 mM NaCl), 1 time with 1 ml High Salt Immune Complex Wash Buffer (0.1% SDS, 1 % Triton-X, 2 mM EDTA, 50 mM Tris-Hcl pH8, 500 mM NaCl), 1 time with 1 ml LiCl Immune Complex Wash Buffer (0.25 M LiCl, 1% Igepal-CA630, 1% natrium deoxycholate, 1 mM EDTA, 10 mM Tris-Hcl pH8), 2 times with 1 ml TE buffer (10 mM Tris–HCl, pH 8.0, 10 mM EDTA)) and eluted for 6 h to O/N at 65°C in ChIP elution buffer (20 mM Tris-HCl, pH 7.5, 5 mM EDTA 50 mM NaCl, 1% SDS, and 50 μg proteinase K) treated and finally cleaned up using Zymo ChIP DNA Clean & Concentrator. ChIP-seq libraries were constructed using NEBNext ChIP-Seq Library Preparation Kit (NEB #E6240) following manufacturer’s instructions. Libraries were sequenced on Illumina HiSeq2500 in single read mode with the read length of 50 nt following manufacturer's instructions. Base calls were performed with RTA 1.13.48.0 followed by conversion to FASTQ with bcl2fastq 1.8.4 and produced approximately 30 million reads per sample.

RNA-seq: CD25+ cell experiments:  Briefly, lin- FLPs were expanded for 7 days on OP9-DL1, CD25+ enriched and retrovirally infected with empty vector control or HA-tagged versions of Lzr-PU.1WT, -PU.1ENG, -PU.1ETS. Cells were co-cultured on OP9-DL1 for ~40h and then sorted as LiveGFP+CD25+ or LiveGFP+CD44+CD25- on BD FACSAriaTM.    Scid.adh.2c2: Scid.adh.2c2 cells were transduced with Lzr-PU.1ert2 (Scid.adh.2c2-PU.1ert2) or Lzr-ert2 (Scid.adh.2c2-ert2) control, cells were expanded for 48h and infected (GFP+) cells followed by sorting of GFP+ cells on BD FACSAriaTM and further expanded in vitro prior to 2, 8 or 24h activation with 0.1 μM 4-OHT (4-hydroxytamoxifen).    Total RNA was isolated from 200.000-500.000 primary or Scid.adh.2c2 cells using RNAeasy MicroKit (Qiagen) according to manufacturer’s recommendations. Libraries were constructed using NEBNext Ultra RNA Library Prep Kit for Illumina (NEB #E7530) from ~1 μg of total RNA following manufacturer’s instructions. Libraries were sequenced on Illumina HiSeq2500 in single read mode with the read length of 50 nt following manufacturer's instructions. Base calls were performed with RTA 1.13.48.0 followed by conversion to FASTQ with bcl2fastq 1.8.4 and produced approximately 30 million reads per sample.

ATAC-seq: Eighty thousand Scid.adh.2c2 or Scid.adh.2c2 transduced Lzr-PU.1-ert2 or Lzr-er2 control vector followed by 0.1 uM 4-hydroxytamoxifen stimulation, 18.4 thousand primary sorted thymic ETP or 50,000 thymic DN3 cells were washed in ice cold HBSS-HEPES prior to the assay. For the primary cells, the final amplified libraries were purified and size-selected (x1.2 bead-to-DNA ratio) using SPRIselect-beads (Beckman-Coultier). Libraries were single-end sequenced on a HiSeq2500 (Illumina) (primary cells) or a NextSeq500 (Scid.adh.2c2) and produced approximately 30-50 million reads per sample.

Please note that processed data was not provided for some ChIP-seq samples since the provided HOMER peak files are merged in various ways throughout the manuscript and then read counts from  various combinations of the bed-files are used to generate the plots. Instead, information of the peak files (used to get the data) was provided in the corresponding sample description field. The listed chip-seq peak files (that have been used to drive reads) are not processed versions of those per se, but explain which files and how they were used in the data analysis using  the HOMER software (http://homer.salk.edu/).

first: Jonas

last: Ungerbäck

first: Hiroyuki

last: Hosokawa

first: Xun

first: Tomoaki

last: Tanaka

first: Tobias

last: Strid

last: Williams

first: Masaki

last: Matsumoto

first: Mikael

last: Sigvardsson

first: Keiichi

middle: I

last: Nakayama


title: Bisulfite-Seq and RNA-seq profiling of Arabidopsis aneuploids

city: Taipei

first: Pao-Yang

last: Chen

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

department: Institute of Plant and Microbial Biology

country: Taiwan

institute: Academia Sinica

summary: Aneuploidy refers to gains and/or losses of individual chromosomes from the normal chromosome set. The resulting gene dosage imbalance usually has a deleterious effect on the phenotype, as illustrated in humans by Down Syndrome (trisomy 21) and solid tumor cells, which are typically highly aneuploid. Aneuploidy has been studied for many years in plants and has been a factor contributing to plant chromosome evolution. Nevertheless, there is still relatively little information about how chromosome numerical imbalances affect the molecular constitution of cells and confer altered phenotypes. To investigate these questions we have performed comparative transcriptome (RNA-seq) and methylome (bisulfite-seq) analyses on trisomics (2n+1) of all five chromosomes of Arabidopsis thaliana as well as matched diploids, triploid and tetraploids.

overall_design: Trisomics of each of the five Arabidopsis thaliana chromosomes were screened out microscopically at the seedling stage from a population of selfed progeny of a triploid parent using fluorescent-tags on specific chromosomes as markers. The triploid parents are obtained as progeny from a cross between a diploid and a tetraploid. The trisomics were grown in soil and the number of chromosomes was confirmed by counting metaphase chromosomes prepared from flower pistils and by comparative genome hybridization. At least two biological replicates representing each confirmed karyotype (trisomies 1 through 5, diploid, triploid, tetraploid) were used for DNA and RNA isolation. DNA and RNA preparations as well as RNA-seq and bisulfite-seq were carried out according to standard procedures.

first: Marjori

last: Matzke

first: Wen-Ching

last: Chan

first: Fei-Man

last: Hsu

first: Bruno

last: Huettel

first: Ming-Ren

last: Yen

first: Yih-Shien

last: Chiang

first: Antonius


title: Next Generation Sequencing Analysis of Wild Type Mouse Embryonic Fibroblast (MEFs) Transcriptomes Stimulated by Viral Mimics

status: Public on Jun 15 2018

city: Abu Dhabi

first: Piergiorgio

last: Percipalle

state: Abu Dhabi

address: Saadiyat Campus, Abu Dhabi

department: Biology Department, Science Division

country: United Arab Emirates

institute: New York University Abu Dhabi

summary:
Purpose: The goals of this study are to use NGS to perform transcriptome profiling (RNA-seq) to find the changes of the global gene expression programs after viral mimic stimulation in Mouse Embryonic Fibroblasts (MEFs)
Methods: MEFs were staimulated by 10 ug/ml Poly(dA:dT) or Poly(I:C) using Lipofectamine 2000 (Invitrogen) as transfection regaent in Opti-MEM medium for 6 hours. Mock groups were treated only with transfection regaent. Total RNA was extracted using RNeasy Mini Kit (Qiagen) according to the manufacturer protocol. Total RNA was used for library construction at the Center for Genomics and Systems Biology, New York University Abu Dhabi, using TruSeq RNA Library Prep Kit v2 (Illumina). Deep-sequencing was performed using Illumina HiSEq 2500 sequencing platform (New York University Abu Dhabi Sequencing Center). Data was processed through the standard RNAseq analysis pipeline at NYUAD. Read aligmnet was performed using tophat2 v2.1.0 against Mus musculus GRCm38.p4 genome version. Raw counts of mapped reads of each genes were derived using HTseq count. DESeq2 was used for differential expression analysis.
Results: Using an optimized data analysis workflow, we identified genes up-regulated or down-regulated after anti-viral immunity activation. Gene ontology analysis further identified biological processes, cellular components and molecular functions that were overrepresented in the differentially expressed genes.
Conclusions: Our study provides a good reourses for profiling genes regulations after viral immunity activation

overall_design: mRNA profiles of Wild Type MEFs stimulated by viral mimics to study how global transcriptome changes after innate vrial immunity activation in non-immune cells

first: Xin

last: Xie

first: Nizar

last: Drou


title: The conserved regulatory RNA RsaE down-regulates the arginine degradation pathway in Staphylococcus aureus

status: Public on Jun 16 2018

city: Jouy-en-Josas

first: Tatiana

last: Rochat

address: Domaine de Vilvert

country: France

institute: INRA


title: Mutations in the SWI/SNF chromatin remodeling complex induce metabolic rewiring and dependence on oxidative phosphorylation

city: Houston

first: Jennifer

last: Wang

state: TX

address: 1515 Holcombe Blvd

institute: University of Texas MD Anderson Cancer Center

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus


title: The conserved regulatory RNA RsaE down-regulates the arginine degradation pathway in Staphylococcus aureus [Hybrid-Trap-seq]

summary: Bacterial regulatory RNAs (sRNA) generally act by base-pairing with target mRNAs. While identification of sRNA targets is the essential step in sRNA characterization, it remains a stumbling block in most studies. To study sRNA-regulated networks in the major human pathogen Staphylococcus aureus, we used a RNA-RNA interactome screening method for identifying sRNA targets based on synthetic sRNAs that are used in vitro as bait to trap their corresponding targets. The key to target discovery lies in the differential analysis of RNA-seq data from captures with different sRNAs and from ΔrsaE and RsaE over-expressing strains. This strategy was applied to study RsaE, a regulatory RNA highly conserved amongst Firmicutes and revealed that RsaE targets the arginase rocF mRNA via direct interactions involving G-rich motifs. Two duplicated C-rich motifs of RsaE can independently downregulate rocF expression. However, the relative activity of these motifs is substrate dependent. Metabolite quantifications in wild-type and ΔrsaE cultures indicate that the absence of RsaE affects amino acid metabolism. Collectively, the data support the model that RsaE acts as a global regulator downregulating functions associated to metabolic adaptation.

overall_design: The protocol called Hybrid-Trap-seq used in this study allows identifying RNA targets interacting in vitro with a sRNA. In brief, a synthetic sRNA was produced, biotinylated, and fixed to streptavidin-associated magnetic beads. The resulting complex was incubated with total RNA extracts, washed, and RNAs bound to sRNAs were eluted. Recovered RNAs were quantified by RNA-seq using Illumina Genome Analyzer IIx generating single-end 40-nt reads. Hypothesizing that unspecific RNAs recovered via this protocol would be the same with different sRNAs used as baits; therefore, background noise could be subtracted by a differential analysis of Hybrid-Trap-seq data obtained with several different baits. In this study, we run Hybrid-Trap-seq with Staphylococcus aureus RsaE regulatory RNA and we used 3 other sRNAs (RsaA, RsaH and RNAIII) dataset as control. The processed data file (putative_RsaE-targets_revised_20180425.csv) compiles read counts for S. aureus CDSs and bona-fide sRNAs (described by Liu et al 2018, doi: 10.3389/fmicb.2018.00228) for the 4 RNA-seq samples as well as DEseq output (RsaE sample versus the 3 other samples used as control).

first: Philippe

last: Bouloc


title: DC-SCRIPT binds GA-rich enhancers to control MAPK-signaling and IL-10 production in human dendritic cells

city: Nijmegen

first: Simon

last: van Heeringen

address: Geert Grooteplein 28

department: Molecular Developmental Biology

country: Netherlands

institute: Radboud University

summary: Dendritic cells (DCs) uniquely direct the adaptive immune response towards activation or inhibition, yet little is known how these opposite programs are regulated at the transcriptional level. Here we applied genome-wide approaches to delineate the molecular function of DC-Specific transCRIPT (DC-SCRIPT/ZNF366), an 11 Zn finger-containing transcription factor potentiating DC-function by limiting IL-10 production. Transcriptome analysis identified DC-SCRIPT to affect expression of genes involved in MAPK signaling, and ChIP-Seq analysis showed binding of DC-SCRIPT to GA-rich enhancers nearby genes encoding MAPK Dual-Specificity Phosphatases (DUSPs). Functional studies demonstrated that DC-SCRIPT-knockdown DCs express much less DUSP4 and exhibit increased phosphorylation of all the three major MAPKs (ERK, JNK and p38). Enhanced ERK signaling in DC-SCRIPT-knockdown DCs led to higher production of the immune-inhibitory cytokine IL-10, which could be reverted by DUSP4 overexpression. These results delineate the molecular mechanism DC-SCRIPT employs to limit IL-10 production in DCs, thereby fine-tuning these professional antigen-presenting cells towards immune activation.

overall_design: ChIP-seq of DC-SCRIPT (ZNF366) in monocyte-derived DCs from three different donors, stimulated with the TLR ligand R848 for 0, 1 and 24 hours, as well as ChIP-seq for the histone modifications H3K4me3 and H3K27ac in monocyte-derived DCs, stimulated with R848 for 0, 1 and 24 hours.

first: Jonas

last: Søndergaard

first: Maaike

last: Looman

first: Chunling

last: Tang

first: Vassilis

last: Triantis

first: Pauline

last: Louche

first: Eva

last: Janssen-Megens

first: Anieta

last: Sieuwerts

first: John

last: Martens

first: Colin

last: Logie

first: Hendrik

last: Stunnenberg

first: Marleen

last: Ansems

first: Gosse

last: Adema


title: Transcriptomic analysis of mouse embryonic stem cells during early differentiation and the effect of mTORC1 regulators knockout

city: Hinxton

first: Meng

last: Li

address: Genome Campus

institute: Sanger Institute

summary: This study analysed the transcriptome of mouse Rex1GFPd2 cells before and during early differentiation and further investigated the transcriptomic changes of Nprl2 and Tsc2 knockout.

overall_design: RNA samples were collected before differentiation, and on day 1, 2, 3 of differentiation; RNA samples of Rex1GFP positive population were collected for Nprl2, Tsc2 knockout and compared to wild type cells.

first: Kosuke

last: Yusa


title: The conserved regulatory RNA RsaE down-regulates the arginine degradation pathway in Staphylococcus aureus [ssRNA-Seq]

first: Tatiana

last: Rochat

summary: Bacterial regulatory RNAs (sRNA) generally act by base-pairing with target mRNAs. While identification of sRNA targets is the essential step in sRNA characterization, it remains a stumbling block in most studies. To study sRNA-regulated networks in the major human pathogen Staphylococcus aureus, we used a RNA-RNA interactome screening method for identifying sRNA targets based on synthetic sRNAs that are used in vitro as bait to trap their corresponding targets. In addition, the transcriptomic effects of sRNA deletion and sRNA accumulation were analyzed by differential expression analyzes of RNA-seq data. This strategy was applied to study RsaE, a regulatory RNA highly conserved amongst Firmicutes and revealed that RsaE targets the arginase rocF mRNA via direct interactions involving G-rich motifs. Two duplicated C-rich motifs of RsaE can independently downregulate rocF expression. However, the relative activity of these motifs is substrate dependent. Metabolite quantifications in wild-type and ΔrsaE cultures indicate that the absence of RsaE affects amino acid metabolism. Collectively, the data support the model that RsaE acts as a global regulator downregulating functions associated to metabolic adaptation.

overall_design: Two transcriptome comparisons were analyzed: i) Staphylococcus aureus ΔrsaE mutant compared to HG003 wild-type strain, and ii) ΔrsaE containing rsaE controlled by an inducible promoter (PtetO-rsaE) under induced relative to non-induced conditions. Total RNA were extracted, mRNA enriched using the MICROBExpress kit (Ambion), then RNA-seq libraries were generated with the TruSeq SBS Kit v3 (Illumina). The sequencing was performed with a HiSeq using 100 x 2 (Paired-End) bases run. Transcriptomic changes were determined by running a differential expression analysis of RNA-seq data using DESeq2.

first: Daniel

last: Gautheret

first: Claire

last: Toffano-Nioche


title: Expression of TGFβ-inducible myosin-X predicts survival and chemotherapy resistance in squamous cell lung cancer

city: Lübeck

first: Hauke

last: Busch

state: Schleswig-Holstein

address: Ratzeburger Allee 160

department: Lübeck Institute of Experimental Dermatology

institute: University of Lübeck

summary: Squamous cell lung carcinoma (SCC) corresponds to about 25% of all lung cancers. Therapeutic approaches are very limited and platinum-based chemotherapy remains the main treatment option. Despite multiple studies, there are no generally accepted predictive biomarkers for SCC. Transforming growth factor-β (TGFβ) signaling was shown to be implicated in numerous pro-tumorigenic processes, including immune evasion, inflammation and cancer metastasis. In the context of SCC epithelial-to-mesenchymal transition phenotype that is commonly mediated by TGFβ was widely observed in surgically resected specimens. However, the relation between TGFβ-induced changes and SCC progression remains to be elucidated. In the presented work, we combined phenotypic and transcriptome-wide approaches to identify novel predictive biomarkers for SCC. We show that TGFβ treatment activated Smad-mediated signal transduction and resulted in increase of migratory and invasive properties of SK-MES1 cells. Multiple actin cytoskeleton-related proteins, including myosin motor proteins such as Myosin-X, were up-regulated upon TGFβ stimulation. siRNA-mediated knockdown of Myosin-X completely abrogated TGFβ-induced collagen gel invasion. Finally, analysis of mRNA expression in paired surgically resected tissues of 151 SCC patients with corresponding 80-month clinical follow-up, showed that the mRNA expression ratio of Myosin-X in tumor and adjacent non-tumor tissue is predictive for overall survival and chemotherapy resistance independently of tumor stage. Given Myosin-X role in cellular motility and invasion, it can represent a new biomarker for aggressive disease and serve as a potential molecular target for therapeutic intervention in patients with SCC.

overall_design: SK-MES-1 cells were treated with TGFb or left untreated and mRNA isolated at t=[0h, 8h, 24h, 48h] in biological triplicates.

first: Melanie

last: Börries

first: Sebastian

last: Ohse

first: Dmytro

last: Dvornikov

first: Ursula

last: Klingmüller


title: Differentially expressed genes after siRNA transfection in human cancer cell lines

status: Public on Jun 18 2018

city: Chiba

first: Naohiko

last: Seki

state: Chiba

address: 1-8-1, Inohana

department: Functional Genomics

country: Japan

institute: Chiba University

summary: To identify differentially expressed genes by anti cancer treatments (siRNAs) in human cancer, several cell lines (lung cancer, breast cancer and pancreatic cancer) were subjected to Agilent whole genome microarrays.

overall_design: Human cell lines (A549, SK-MES, SW1990 and MDA-MB-231) were treated with siRNAs (si-TNS4, si-NCS1, si-RACGAP and si-AP1S3).

first: Yasutaka

last: Yamada


title: Mutation of S-adenosyl-methionine synthetase 3 (SAMS3) releases transcriptional silencing by reducing DNA and histone methylation in Arabidopsis

city: Beijing

first: Jinkui

last: Cheng

address: No.2 Yuanmingyuan West Road

institute: China Agricultural University

summary: Arabidopsis SAMS3 regulates epigenetic status through controlling DNA and histone methylation.The mutation in SAMS3 decreased the CHG and CHH methylation and reactivation of certain transposable elements.

overall_design: Examination of whole genomic DNA methylation and gene expression in L119 (a transgenic lines used as WT) and sams3.

first: Jingjing

last: Meng

first: Lishuan

last: Wang

first: Xiaowen

last: Zhao

first: Dan

last: Jin

first: Qing

first: Zhizhong

last: Gong


title: The C-terminal multimerization domain is essential for leukemia development by CBFβ-SMMHC

city: Bethesda

first: Pu

middle: Paul

last: Liu

address: 49 Convent Dr.

department: TFGB

institute: NHGRI/NIH

summary: The recurrent chromosome 16 inversion in acute myeloid leukemia generates a fusion gene between CBFB and MYH11, which in turn encodes a chimeric protein CBFβ-SMMHC (core binding factor β-smooth muscle myosin heavy chain).   We previously demonstrated that CBFβ-SMMHC needs its C terminal domains for leukemogenesis.  In this study, we generated a new Cbfb-MYH11 knock-in mouse model to dissect the role of the multimerization domain at the C terminus of CBFβ-SMMHC.  Specifically, we mutated six amino acids in the helices D and E (mDE) of the assembly competent domain, which is important for SMMHC multimerization.  We found that the embryos with the mDE mutation (Cbfb+/mDE) did not develop hematopoietic defects seen in embryos with full-length CBFβ-SMMHC (Cbfb+/MYH11).  More importantly leukemia development was abolished in the adult Cbfb+/mDE mice even after mutagenesis treatment.  In addition, gene expression profile of the hematopoietic cells from the Cbfb+/mDE mice was more similar to that of Cbfb+/+ mice than the Cbfb+/MYH11 mice. Our data suggest that the C terminal multimerization domain is required for the defects in primitive and definitive hematopoiesis caused by CBFβ-SMMHC, and it is also essential for leukemogenesis caused by CBFβ-SMMHC.

overall_design: RNA-seq of Bone marrow c-KIT+ cells from WT, Cbfb +/mDE, Cbfb +/MYH11 mice

first: Ling

first: P

middle: P


title: Expression charcaterization of an internal protocol developed to differentiate RPE cells

city: Basel

first: Roland

last: Schmucki

address: Grenzacherstrase

country: Switzerland

institute: F. Hoffmann - La Roche AG

summary: We performed RNA-seq and miRNA-seq  in fetal RPE cells differentated during 5 weeks in a transwell set up

overall_design: Samples from days 7, 14, 21, 28 and 35 were characterized. Cells were grown in a proliferation medium during the first week (EpiCM) and then in a maturation medium (MAM medium) that enahnces differentiation towards the desired phenotype.

first: Roberto

last: Iacone

first: Esther

last: Melo

first: Zeynep

last: Eren Ghiani

first: Laura

last: Badi

first: Tony

last: Kam-Thong

first: Nicholas

last: Flint

first: Jean-Christophe

last: Hoflack


title: Identification and characterization of long noncoding RNAs and mRNAs expression profiles related to postnatal liver maturation of breeder roosters using Ribo-zero RNA Sequencing

city: Yangling

first: Shengru

last: Wu

address: 22 St.XiNong

department: College of Animal Science and Technology

institute: Northwest A&F University

summary: The liver, which is mainly hematopoietic in the embryo but converts into a major metabolic tissue in the adult, is extensively remodeled after birth to adapt and perform adult functions. Long non-coding RNAs (lncRNAs) are involved in organ development and cell differentiation, which indicate that they have potential roles in regulating postnatal liver development.

overall_design: In order to understand the roles of lncRNAs in postnatal liver maturation, we analyzed the lncRNAs and mRNAs expression profiles in immature and mature livers from one-day-old and adult (40 weeks of age) breeder roosters by Ribo-Zero RNA-Sequencing. Around 27,353 mRNAs and 9,063 predicted lncRNAs were expressed in breeder rooster livers, and the identified chicken lncRNAs shared fewer exons, shorter transcript length, and significantly lower expression levels compared to protein-coding genes. Notably, in comparison between the livers of newborn and adult breeder roosters, a total of 1480 mRNAs and 622 lncRNAs were differentially expressed with the criteria of log2fold change > 1 or < -1 and P values < 0.05, which were validated by qPCR using randomly selected five mRNAs and five lncRNAs. Further GO and KEGG analyses have revealed that the differentially expressed mRNAs were involved in the hepatic metabolic and immune change, as well as some biological processes and pathways including cell division, proliferation, apoptotic and cell cycle that are implicated in the development of liver. We also investigated the cis- and trans- regulatory effects of lncRNAs on its target genes. Further GO and KEGG analyses indicated that these differentially expressed lncRNAs have their neighbor protein coding genes and co-expressed genes associated with adapting of adult hepatic functions, as well as some pathways involved in liver development, such as cell cycle pathway, Notch signaling pathway, Hedgehog signaling pathway, adherens junction and Wnt signaling pathway.

first: Xiaojun

last: Yang


title: RNA-seq data analysis of Smad5 knockout and wild type individual mouse amnion samples collected during early embryonic development

city: Leuven

first: Stein

last: Aerts

address: Herestraat 49

department: Center for Human Genetics

country: Belgium

institute: KULeuven

summary: Mice deficient in the BMP-effector, Smad5 (Smad5 KO), develop severe defects in embryonic morphogenesis as well as a delay in amnion-chorion separation, important extraembryonic tissues. After closure of the proamniotic canal, a remarkable ectopic primitive streak-like aggregate develops in the amnion of these mutants. We investigated the earliest steps of mutant amnion misdifferentiation by RNAseq of single Control (Ctrl) and Smad5 KO amnion samples collected before the appearance of the aggregate. The transcriptome analysis revealed two separate sets of non-squamous amnion defects. One set of mutants (KO-SetA) robustly overexpressed streak mesoderm-related genes conform former analyses (Pereira et al., 2012). The other set overexpressed extraembryonic ectoderm markers suggestive of chorionic inclusion in amnion (KO-SetB). Tetraploid chimera analyses confirmed that SMAD5 deficiency in the epiblast can result in two distinct sets of amnion defects: one with impaired anterior amnion expansion and differentiation, and another with inclusion of chorionic extraembryonic ectoderm in the space normally occupied by amnion.

overall_design: To investigate the molecular differences between WT and Smad5 mutant amnion, we assessed global gene expression by RNA-Seq before morphological differences were apparent between individual control and knockout amnion samples. Amnion was microdissected of control (n=6), and stage-matched littermate Smad5 knockout (n=6)(Chang et al., 1999 Development, 126, 1631–1642) embryos ranging between early pre-headfold (EPHF) and 3-somites (3S) stages (Table S1).  cDNA was synthesized and amplified from individual amnion samples. cDNA for RNA-seq was prepared and amplified following adaptation of a procedure originally designed for RNA-seq of single-cells (Tang et al., 2009) on the SOLiD platform (“Double stranded cDNA synthesis from a single cell”, Applied Biosystems demonstrated protocol). Briefly, reverse transcription and second-strand cDNA synthesis was performed in the sample lysate without RNA isolation using SuperScript III Reverse Transcriptase (Invitrogen) and poly-dT universal primers. This was followed by two rounds of PCR amplification (18 and 12 cycles, respectively) and column purifications (QIAquick, QIAGEN) generating up to 3 kb coverage at the 3’ end of all poly-adenylated transcripts. The resulting cDNA was used for RT-qPCR and RNA-seq. The cDNA yield for the 12 RNA-seq samples is shown in Table S1.

first: Mariya

last: Dobreva

first: Marina

last: Naval-Sanchez

first: Danny

last: Huylebroeck

first: An

last: Zwijsen

first: Rekin's

last: Janky


title: Mice Exposed to Bisphenol A Exhibit Depressive-like Behavior with Neurotransmitter and Neuroactive Steroid Dysfunction

city: Philadelphia

first: Yemin

last: Lan

state: PA

address: 3400 Civic Center Blvd

institute: University of Pennsylvania

summary: We report here hippocampal RNA-seq data from adult (20 weeks) male offspring exposed to 0 mg (control) or 50 mg BPA/kg diet during gestation and lactation.

overall_design: RNA-seq in adult hippocampus from control and BPA-exposed males, four independent biological replicates per treatment group, sequenced on Illumina NextSeq 500 platform.

first: Frances

last: Xin

first: Erin

last: Fischer

first: Christopher

last: Krapp

first: Elizabeth

last: Krizman

first: Clementina

last: Mesaros

first: Nathaniel

last: Snyder

first: Amita

last: Bansal

first: Michael

middle: B

last: Robinson

first: Rebecca

last: Simmons

first: Marisa

middle: S

last: Bartolomei


title: Comparison of DEG between neonatal male and female mice age of P25 by CHD8 Asn2373LysfsX2 heterozygote mutation.

city: Daejeon

first: Eunjoon

last: Kim

address: Kuseong-dong

department: Center for Synaptic Brain Dysfunctions/Department of Biological Sciences

country: South Korea

institute: IBS/KAIST

summary: CHD8, encoding a chromatin remodeling protein, is one of the most frequently mutated genes in autism spectrum disorders. However, how such mutations cause autistic behaviors remain unclear. In mice carrying a heterozygous frame-shift mutation in the Chd8 gene (Asn2373LysfsX2) identified in autistic human individuals, we observed autistic-like behaviors that are much stronger in males than in females, similar to human cases. These behaviors included enhanced mother-seeking ultrasonic vocalizations in pups, mother-attachment behaviors in juveniles, and isolation-induced self-grooming in adults. These behaviors were associated with opposite changes in synaptic excitation/inhibition and neuronal firing in male and female mice, but with strong changes in gene expression in female mice. Therefore, this CHD8 mutation may cause male-preponderant autistic-like behaviors in mice through differential synaptic/neuronal changes and gene expression

overall_design: Whole brain transcriptome of 3 neonatal WT and CHD8 Asn2373LysfsX2 heterozygote mutant mice age of P25 in both sex.

first: Hwajin

last: Jung

first: Hyojin

last: Kang


title: RNA sequencing of ESC/iPSC-derived purified PAX6-GFP neural progenitors form control and Phelan-Mcdermid patients

city: Cambridge

first: Reinhard

last: Roessler

address: 455 Main street

country: USA

institute: Whitehead Institute

summary: The goal of this study was to generated and transcriptionally analyze purified populations of Phelan-McDermid and control pluripotent stem cells-derived neural progenitors. This type of analysis allowed us to identify prevoisly unknown transcriptional phenotypic changes in patient neural progenitors which appear to be independent of functional synapses.

overall_design: Patient and control pluripotent stem cells were targeted by CRISPR/Cas9 and a PAX6-GFP reporter plasmid. Correctly modified clones were selected and subsequently differentiated into PAX6 expressing neural progenitors.

first: Rudolf

last: Jaenisch


title: Comparison of DEG between neonatal male and female mice age of P0 by CHD8 Asn2373LysfsX2 heterozygote mutation

summary: CHD8, encoding a chromatin remodeling protein, is one of the most frequently mutated genes in autism spectrum disorders (ASDs) 1-8. However, whether and how such mutations cause autistic behaviors remain unclear. Here we show that a human CHD8 mutation causes autistic-like behaviors and enhanced excitatory drive specifically in male mice. We found that knockin mice carrying a heterozygous frame-shift mutation in the Chd8 gene (Asn2373LysfsX2) identified in human individuals with ASDs (Chd8+/N2373K mice) display male-specific autistic-like behaviors. Gene transcript analysis shows that male and female Chd8+/N2373K neurons exhibit largely opposite changes in the levels of 29 mRNAs, nine of which correspond to known ASD-risk genes. These results suggest that a human CHD8 mutation causes male-specific autistic-like behaviors in mice in association with gender-specific differential changes in gene transcripts and excitatory drive in the brain.

overall_design: Whole brain transcriptome of 3 neonatal WT and CHD8 Asn2373LysfsX2 heterozygote mutant mice in both sex.


title: RNAseq of genetically engineered mouse model (GEMM)  tumors

first: Jennifer

last: Wang

summary: We report the expression profiling of lung adenocarcinoma tumors arising from genetically engineered mice with activation of KRAS(K) and loss of either SMARCA4 (S), p53(P) or both.

overall_design: Examination of RNA expression of tumors with different genotypes

first: Chia-Chin

last: Wu

first: Yonathan

last: Deribe


title: Crispr-Cas9-mediated Aire gene editing in medullary thymic epithelial (mTEC) cells shows its role as a gene expression modulator during thymocyte adhesion

city: Ribeirao Preto

first: Geraldo

middle: A

last: Passos

state: SP

address: 3900, Bandeirantes Avenue

department: Genetics

country: Brazil

institute: University of Sao Paulo

summary: The aim of this study is to evaluate the effect of Autoimmune regulator (Aire) gene disruption in a murine medullary thymic epithelial cells (mTEC 3.10 cell line) on the transcriptome of these cells during its adhesion with thymocytes. The mTEC-thymocyte adhesion is a crucial step for the negative selection of autoreactive thymocytes and prevention of autoimmune diseases. To generate Aire mutant cell clones, a total of 5x10^5 mTEC 3.10 cells were electro-transfected (Lonza Nucleofector) with CRISPR-Cas9 plasmid targeting the Aire Exon 3 (plasmid "all in one"  encoding Aire Exon 3 gRNA + Cas9 + GFP, from Sigma-Aldrich). The GFP positive mTEC single cells were sorted by using a FACS Aria III cytometer and cells were cloned by expansion in culture. Sanger sequencing of PCR products from the Aire Exon 3 of these clones was used in order to evaluate the occurrence of indel mutations within the targeted Exon 3. The mTEC 3.10 clone E6 was identified and validated as a compound heterozygous Aire KO (Aire +/-). This clone features the Aire allele 1 that encodes a mutant Aire protein carring a neutral aminoacid substitution (A118P) and allele 2 encoding a truncated Aire protein. Wild type (WT) mTEC 3.10 cells or mTEC 3.10 clone E6 were cultured in the presence (or not) of thymocytes in order to establish cell adhesion. The total RNA preparations from WT or clone E6 mTEC cells (before or after mTEC- thymocyte co-cultures) were then sequenced  through RNA-sequencing using a Illumina HiSeq 2500 instrument and the TruSeq Stranded mRNA Library Preparation kit resulting in about 50 million paired-end stranded specific 100 bp reads per sample. Sequencing reads were mapped to Mus musculus reference genome (mm10) using STAR v.2.5.0a. Read counts over transcripts were calculated using HTSeq v.0.6.1p2  based on a current UCSC annotation file for GRCm38/mm10 (Dec. 2011).

overall_design: The mRNA profiles of mTEC 3.10 cells carring WT Aire (before or after co-culture with thymocytes) or heterozygous KO mTEC 3.10 cells (clone E6, Aire +/-) (before or after co-culture with thymocytes) were generated by sequencing, in duplicates, using a Illumina HiSeq 2500 instrument.

first: Cesar

last: Speck-Hernandez

first: Amanda

middle: F

last: Assis

first: Rafaela

last: Felício

first: Larissa

last: Cotrim-Sousa

first: Silvana

last: Giuliatti


title: RNA sequencing reveals levamisole target genes PTPRZ1 and MDK and their links to interferon pathway in human podocytes

city: Bristol

first: Lulu

last: Jiang

state: State...

address: Academic Renal Unit,Dorothy Hodgkin Building,Whitson Street

department: School of Clinical Sciences

country: United Kingdom

institute: University of Bristol

summary: To investigate the mechanisms underlying levamisole's efficacy in disease, we performed RNA sequencing using 3 human podocyte cell lines aiming to generate transcriptome profiles of podocyte in response to levamisole.

overall_design: Transcriptome profiles of human podocytes treated with levamisole were generated by RNA-sequencing using Illumina HiSeq 2500

first: Charlie

last: Hindmarch

first: Mark

last: Rogers

first: Christy

last: Waterfall

first: Jane

last: Coghill

first: Peter

last: Mathieson

first: Gavin

last: Welsh


title: Isolation and differential transcriptome of vascular smooth muscle cells and mid-capillary pericytes from the rat brain

city: paris

first: bruno

last: saubamea

address: 4 avenue de l'observatoire

department: Faculté de Pharmacie de Paris

institute: Université Paris Descartes

summary: VSMC and mcPC were freshly isolated from the rat brain by magnetic-activated cell sorting and their transcriptome was analyzed by RNA-Seq

first: Bruno

last: Saubaméa

first: Stéphanie

last: Chasseigneaux


title: Hippocampus transcriptome analysis of CHD8 Asn2373LysfsX2 heterozygote mutation mice age of P25

first: Eunjoon

last: Kim

overall_design: Hippocampus transcriptome of WT and CHD8 Asn2373LysfsX2 heterozygote mutant mice age of P25 in both sex.

first: Hwajin

last: Jung

first: Hyojin

last: Kang


title: A tetraspanin family member functionally resolves and facilitates the purification of adult pluripotent stem cells used for whole-body regeneration (Bulk)

status: Public on Jun 14 2018

city: Kansas City

first: Hua

last: Li

state: MO

address: 1000 E 50th Street

institute: Stowers Institute for Medical Research

summary: Proliferating cells known as neoblasts include pluripotent stem cells (PSCs) capable of sustaining tissue homeostasis and regeneration of lost body parts in planarians. However, the lack of markers to prospectively identify and isolate these adult PSCs has significantly hampered their characterization. We used single-cell RNA sequencing (scRNA-seq) and single cell transplantation to address this long-standing issue. Large-scale scRNA-seq of fluorescently sorted neoblasts unveiled a novel subtype of neoblast (Nb2) characterized by high levels of PIWI-1 mRNA and protein, and marked by a conserved cell-surface protein coding gene, tetraspanin 1 (tspan1). tspan1-positive cells survived sub-lethal irradiation, underwent clonal expansion to repopulate whole animals, and when purified with an anti-TSPAN-1 antibody rescued with high efficiency the viability of lethally irradiated animals after single-cell transplantation. Our work demonstrates the first prospective isolation of an adult PSC, bridges a conceptual dichotomy between operationally and molecularly defined neoblasts, and sheds light on the mechanisms governing in vivo pluripotency and the source of regeneration in planarians.

overall_design: 1) Examine gene expression profiles of FACS sorted X1, X2 and Xins cell populations from adult flatworms. The experiment was performed in triplicate, resulting in 9 samples; 2) Examine gene expression profiles of wildtype untreated and lethal (10K) irradiated animals. The experiment was performed in triplicate, resulting in 6 samples; 3) Examine gene expression profiles of Flow Cytometry sorted PIWI-1_high,PIWI-1_low and PIWI-1_negative cell populations from adult flatworms. The experiment was performed in quadruplicate, resulting in 12 samples; 4) To examine gene expression profiles of regenerating fragment over the regeneration time-course, we performed RNA-seq on 17 time-points of regeneration, starting from amputation of small-sized fragments to fully functional regenerated animals. The experiment was performed in quadruplicate, resulting in 68 samples.

first: An

last: Zeng

first: Alejandro

last: Sánchez Alvarado


title: Comparison of DEG between neonatal male and female mice by CHD8 Asn2373LysfsX2 heterozygote mutation

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

overall_design: Refer to individual Series


title: Tetraspanin family member functionally resolves and facilitates the purification of adult pluripotent stem cells used for whole-body regeneration (Single)

overall_design: 1) Examine gene expression profiles of sorted 80 X1 and 80 Xins cell at single cell level using the plate based single cell RNA-seq platform. Two batch of (40 X1+40 Xins) cells were sorted into individual 96-well plates, and libraries were made by ClonTech v4 full reactions, and quarter-size reactions, respectively.  2) Examine gene expression profiles of sorted X1 cells, and X1+X2 cells survived in 7-day irradiation by 10X scRNA-Seq platform.


title: A tetraspanin family member functionally resolves and facilitates the purification of adult pluripotent stem cells used for whole-body regeneration


title: SOX9 has distinct regulatory roles in alternative splicing and transcription

city: Montpellier

first: Peggy

last: Raynaud

address: 1919 Route de Mende

institute: CRBM

summary: SOX9 is known as a crucial transcription factor for various developmental processes and for tissue homeostasis. We examined here its potential role in alternative splicing by analyzing global splicing changes, using RNA-seq of colon tumor cells. We show that SOX9 knockdown alters the splicing of hundreds of genes without affecting their expression levels, revealing that SOX9 controls distinct splicing and transcriptional programs. SOX9 does not affect splicing patterns through the control of splicing factors expression. We identify mutants that uncouple SOX9 splicing function from its transcriptional activity. We demonstrate that SOX9 binds to RNA and associates with several RNA-binding proteins, including the core Exon Junction Complex component Y14. Half of SOX9 splicing targets are also modulated by Y14 and are no longer regulated by SOX9 upon Y14 depletion. Altogether, our work reveals that SOX9 is a moonlighting protein which modulates either transcription or splicing of distinct sets of targets.

overall_design: DLD-1 colon tumor cells RNA-seq analysis of transcriptomic and splicing profiles

first: Michael

last: Girardot


title: Age-specific biological and molecular profiling distinguishes paediatric from adult acute myeloid leukaemias

city: St. Lucia

first: Othmar

last: Korn

state: Queensland

address: University of Queensland

country: Australia

institute: Australian Institute for Bioengineering and Nanotechnology

summary: Acute myeloid leukaemia (AML) affects children and adults of all ages. AML remains one of the major causes of death in children with cancer and for children with AML relapse is the most common cause of death. Here, by modelling AML in vivo we demonstrate that AML is discriminated by the age of the cell of origin. Young cells give rise to myeloid, lymphoid or mixed phenotype acute leukaemia, whereas adult cells give rise exclusively to AML, with a shorter latency. Unlike adult, young AML cells do not remodel the bone marrow stroma. Transcriptional analysis distinguishes young AML by the upregulation of immune pathways. Analysis of human paediatric AML samples recapitulates a paediatric immune cell interaction gene signature, highlighting two genes, RGS10 and FAM26F as prognostically significant. This work advances our understanding of paediatric AML biology, and provides murine models that offer the potential for developing paediatric specific therapeutic strategies.

overall_design: 4 samples, 3 replicates: AML 10 weeks, AML 3 weeks, AML more than 52 weeks, AML foetal liver

first: Shahzya

last: Chaudhury

first: Aine

last: Prendergast

first: Graham

last: Hamilton

first: Karen

last: Keeshan


title: A Dynamic Interplay of Klf4  Enhancer Elements Drives Naïve-pluripotency

city: Berkeley

first: Robert

last: Tjian

address: LKS

institute: UC Berkeley/HHMI

summary: We characterized a long-range enhancer cluster driving Klf4 expression in naïve pluripotent stem cells. CRISPR/Cas9 mediated screening and genome editing identified DNA motifs corresponding to key pluripotency TFs on these enhancer regions. We employed ChIP-exo genome-wide sequencing to Esrrb and Stat3 in mESCs and validated their functional engagement on the Klf4 enhancer cluster. Moreover, we performed ATAC-seq on wild type and Oct4/Sox2 composite motif biallelic deletion ESCs on Klf4 enhancer E2 to examine the accessible chromatin structure upon loss of lead TFs Oct4 and Sox2 in a single enhancer region.

overall_design: Examine high resolution genome wide TF binding in naïve pluripotent stem cells and the accessible chromatin upon loss of key TF binding.

first: Liangqi

last: Xie

first: Li


title: RNA-seq of naked mole-rat liver from breeders and non-breeders [livers]

city: Jena

first: Marco

last: Groth

address: Beutenbergstraße 11

department: Core Facility - Next Generation Sequencing

country: Germany

institute: Leibniz Institute on Aging - FLI

summary: Comparison of gene expression level by Illumina sequencing of naked mole-rat liver from breeders and non-breeders.

overall_design: 23 liver samples, 5-6 animals per group, 4 groups: 1) female non-breeder, 2) female breeder, 3) male non-breeder, 4) male breeder

first: Martin

last: Bens

first: Karol

last: Szafranski

first: Susanne

last: Holtze

first: Arne

last: Sahm

first: Thomas

last: Hildebrandt

first: Matthias

last: Platzer


title: The fate of CD8+ T cells during infection is linked to their developmental origin

city: Ithaca

first: Jennifer

last: Grenier

address: 526 Campus Rd

department: Biomedical Sciences

institute: Cornell University


title: Defining the Rio1 transcriptome in the yeast Saccharomyces cerevisiae

city: Milan

first: Peter

last: De Wulf

address: via Adamello 16

department: Department of Experimental Oncology

country: Italy

institute: European Institute of Oncology

summary: The conserved Saccharomyces cerevisiae kinase/ATPase  Rio1 downregulates rDNA transcription to promote rDNA stability and segregation. To uncover additional roles in transcriptional regulation beyond the rDNA locus we defined the global Rio1 transcriptiome. By NGS we identify 818 differentially expressed genes that are under the transcriptional control of Rio1.

overall_design: By depleting Rio1, C-terminally labeled with  an auxin-inducible degron cassette, we identified the genes that are differentially expresse din yeast lacking Rio1 activity, as compared to those expressed in the isogenic wild-type strain.

first: Maria

last: Iacovella

first: Luciano

last: Giacó


title: Genome-wide chromatin localization maps of the endogenously expressed AID-labelled kinase/ATPase Rio1 in Saccharomyces cerevisiae W303, as determined under unperturbed culture conditions (YPD medium, exponential growth, 23°C)

summary: We report the the first  single-molecule-based NGS analysis of chromatin regions that co-immunoprecipitate with Rio1 in the yeast Saccharomyces cerevisiae.

overall_design: We examined the association of Rio1-AID with chromatin by anti-AID chromatin co-immunoprecipitation followed by MiSeq deep sequencing. As the negative control experiment, we probedthe isogenic S. cerevisiae parent strain endogenously expressing untagged Rio1.


title: RNA sequencing of Drosophila melanogaster optic lobe cell types

city: Paris

first: NIKOLAOS

last: KONSTANTINIDES

address: 15 rue Helene Brion

institute: Institut Jacques Monod

summary: Transcription factors regulate the molecular, morphological, and physiological characters of neurons and generate their impressive cell type diversity. To gain insight into general principles that govern how transcription factors regulate cell type diversity, we used large-scale single-cell mRNA sequencing to characterize the extensive cellular diversity in the Drosophila optic lobes. We sequenced 57,000 single optic lobe neurons and glia and assigned them to 52 clusters of transcriptionally distinct single cells. We validated the clustering and annotated many of the clusters using RNA sequencing of characterized FACS-sorted single cell types, as well as marker genes specific to given clusters. To identify transcription factors responsible for inducing specific terminal differentiation features, we used machine-learning to generate a ‘random forest’ model. The predictive power of the model was confirmed by showing that two transcription factors expressed specifically in cholinergic (apterous) and glutamatergic (traffic-jam) neurons are necessary for the expression of ChAT and VGlut in many, but not all, cholinergic or glutamatergic neurons, respectively. Therefore, the same terminal characters can be regulated by different transcription factors in different cell types, arguing for extensive phenotypic convergence. Our data provide a deep understanding of the developmental and functional specification of a complex brain structure.

overall_design: We sequenced 17 FACS-sorted single cell types from the Drosophila optic lobe, as well as FACS-sorted neurons as a control. We also sequenced a line that had GFP expression in two cell types - Tm1 and T1.

first: Katarina

last: Kapuralin

first: Nikos

last: Konstantinides

first: Claude

last: Desplan


title: Changes in DNA methylation pattern following intervention in PTSD patients

city: Bethesda

first: Sijung

last: Yun

state: Maryland

address: 8908 Ewing Drive

institute: NIH/JHU

summary:
Objective: The purpose of this study was to observe  changes in DNA methylation in service members who experienced PTSD.
Methods: We compared DNA Methylation profiles in the blood of those with or without PTSD and also compared those with PTSD at both baseline and follow-up with those service members that only developed PTSD at follow-up (3-months after baseline). Blood was collected and analyzed at baseline and follow up.
Results: In participants with PTSD, 10 genes were significantly hyper-methylated and 18 were significantly hypo-methylated. In the second analysis, participants that had PTSD at both time points did not present any differentially methylated gene regions. Interestingly, for individuals’ whose PTSD scores increased from baseline to follow-up, there were 8 hypo-methylated genes detected at the baseline.

overall_design:
Methyl-CpG-Binding Domain (MBD) seq
All the sample names starting with 'c' is from control, meaning "no PTSD"
'i' means 'increased' PTSD, and 'p' means 'persistent' PTSD when we compare
at time point 2 when we have a reference from at time point 1 (3-months after baseline).

first: J

last: Gill


title: Immunomodulatory effects of dietary seaweeds in LPS challenged Atlantic salmon Salmo salar as determined by deep RNA sequencing of the head kidney transcriptome

city: Leiden

first: Ron

last: Dirks

address: Sylviusweg 74

institute: Future Genomics Technologies BV

summary: Seaweeds may represent immunostimulants that could be used as health-promoting fish feed components thereby offering an alternative for the use of antibiotics. This study was performed to gain insights into the immunomodulatory effects of dietary seaweeds in Atlantic salmon. Specifically tested were 10% inclusion levels of Laminaria digitata (SW1) and a commercial blend of seaweeds (Oceanfeed®) (SW2) against a fishmeal based control diet (FMC). Differences between groups were assessed in growth, feed conversion ratio (FCR) and blood parameters hematocrit (Hct) and hemoglobin (Hb). After a LPS challenge of fish representing each of the three groups, RNAseq was performed on head kidneys to determine transcriptomic differences in response to the immune activation, to our knowledge for the first time in fish in this context. Atlantic salmon fed with dietary seaweeds showed slightly higher FCRs and more homogenous growth but in general no major differences in performance in comparison with fishmeal fed fish. RNAseq resulted in ~154 million reads which were mapped against a NCBI Salmo salar reference and against a de novo assembled Salmo salar reference for analyses of expression of immune genes and ontology of immune processes among the 87,600 cDNA contigs. The dietary seaweeds provoked a more efficient immune response which involved more efficient identification of the infection site, and processing and presentation of antigens. More specifically, chemotaxis and the chemokine-mediated signaling pathway with involvement of genes such as C-C motif chemokine 19 were improved and the defense response to Gram-positive bacterium reduced. The predicted integrin alpha-2-like gene had by far the highest up-regulated expression and may therefore represent a key marker gene of the LPS immune response in salmonids. Specific Laminaria digita effects included reduction of the cytokine-mediated signalling pathway as indicated by the cytokine macrophage migration inhibitory factor, and interferon-gamma-mediated signalling as indicated by STAT1 and the gamma-interferon-inducible lysosomal thiol reductase precursor. Highly upregulated and specific for this diet was the expression of Major histocompatibility complex class I-related gene protein. The commercial blend of seaweeds caused more differential expression than Laminaria digita and improved immune processes such as receptor-mediated endocytosis, inflammatory response, cell adhesion and response to lipopolysaccharide. Particularly expression of many important immune receptors was up-regulated illustrating increased responsiveness. NF-kappa-B inhibitor alpha  is an important gene that marked the difference between both seaweed diets as Laminaria digita inhibits the production of this cytokine while the blend of seaweeds stimulates it. It can be concluded that replacing fishmeal partly with seaweeds such as Laminaria digita can have important modulatory effects on the immune capacity of Atlantic salmon resulting in a more efficient immune response.

overall_design: Deep-sequencing transcriptome analysis of head kidney samples derived from LPS-challenged Salmo salar specimens after a 42-day seaweed or control diet: Laminaria digitata (Seaweed 1 (SW1); n=3), Oceanfeed (Seaweed 2 (SW2); n=3), fishmeal-based control diet (FMC; n=3).

first: Arjan

last: Palstra


title: RNA-seq of breeding and non-breeding naked mole-rats and guinea pigs

first: Marco

last: Groth

sample_organism:
Cavia porcellus
Heterocephalus glaber

platform_organism:
Cavia porcellus
Heterocephalus glaber


title: Triple Inhibition of TGFb/BMP/GSK3 Signaling in Human Pluripotent Stem Cells Generates PAX6- Neural Progenitors with robust Self-renewal

city: Guangzhou

first: qianyu

last: chen

address: 190 Kai Yuan Avenue, Science Park

country: China

institute: Guangzhou Institutes of Biomedicine and Health,Chinese Academy of Sciences

summary: In this report, we show that in addition to dual SMAD inhibition, blocking Glycogen synthase kinase 3(GSK3) signaling by adding CHIR99021 generated novel NPCs with significantly enhanced proliferation. By RNA-seq, we show that S/D/C NPCs and S/D NPCs display distinct expression profiles, as S/D/C triggered NPCs are Pax6- and express hindbrain genes while . S/D triggered NPCs are Pax6+ and highly express forebrain genes. Our data reveal that Pax6 labels distinct regionally specified hNPCs and manipulating GSK3 signaling could pattern hNPCs with positionally specified fate and different self-renewal capability.

overall_design: We compared the transcriptome of through triple Inhibition of TGFb/BMP/GSK3 Signaling  and dual SMAD inhibition derived  NPCs  by RNA-Seq.

first: Zhenghui

last: Su

first: Qianyu

last: Chen

first: Xi

last: Shi

first: Guangjin

last: Pan

first: Juan

last: Du