title: Two distinct transcription termination modes dictated by promoters

city: Basel

first: Sarah

last: Carl

address: Maulbeerstrasse 66

department: Epigenetics

country: Switzerland

institute: Friedrich Miescher Institute

summary: Transcription termination determines the ends of transcriptional units and thereby ensures integrity of gene regulation. Here we perform genome-wide investigation of RNA polymerase II (Pol II) transcription termination in Caenorhabditis elegans and observe two distinct modes of termination. Whereas a subset of genes requires the exoribonuclease XRN2, a known termination factor, termination of other genes appears independent of, and refractory to, XRN2. Unexpectedly, promoters instruct the choice of termination mode, but XRN2-independent termination additionally requires a compatible region downstream of the 3’ end cleavage site. Hence, different termination mechanisms may affect different configurations of Pol II complexes dictated by promoters.

overall_design: RNAi-treated, polyA-selected RNA-seq data: 3 replicates of mock and xrn-2 RNAi were analyzed, 2 of which can be found under series GSE79994 (1 included here). ChIP-seq: 2 replicates of ChIP and input samples for each factor/condition were analyzed, including untagged (control) strains for XRN2 ChIP. RNAi-treated, ribosomal RNA-depleted RNA-seq: 2 replicates of xrn-2 RNAi and one replicate of mock were analyzed. TS-mutant RNA-seq: one replicate of WT and xrn-2(xe31) RNA per timepoint (hours after temperature shift) were analyzed. Timepoints 06h and 10h were re-sequenced at approximately 4-fold deeper coverage. Catalytic-dead mutant RNA-seq: one replicate of WT and xrn-2cd;xrn-2(xe31) RNA per timepoint (06h and 10h after temperature shift) were analyzed.

first: Takashi

last: Miki

first: Helge

last: Grosshans


title: Tacrolimus-induced nephrotoxicity in mice is associated with microRNA deregulation [transcriptome analysis]

status: Public on Feb 06 2018

city: Valbonne - Sophia-Antipolis

first: Kevin

last: Lebrigand

address: 660 route des lucioles

country: France

institute: IPMC/CNRS

summary: Although Tacrolimus is an immunosuppressive drug widely used in renal transplantation, its chronic use paradoxically induces nephrotoxic effects, in particular renal fibrosis, which is responsible for chronic allograft dysfunction and represents a major prognostic factor of allograft survival. As molecular pathways and mechanisms involved in Tacrolimus-induced fibrogenic response are poorly elucidated, we assessed whether miRNAs are involved in the nephrotoxic effects mediated by Tacrolimus. Treatment of CD-1 mice with Tacrolimus (1mg/kg/d for 28 days) resulted in kidney injury and was associated with alteration of a gene expression signature associated with cellular stress, fibrosis and inflammation. Tacrolimus also affected renal miRNA expression, including miRNAs previously involved in fibrotic and inflammatory processes including “fibromirs” such as miR-21-5p. Specifically, Renal Proximal Tubular Epithelial cells exposed to tacrolimus showed up-regulation of miR-21-5p and the concomitant induction of epithelial phenotypic changes, inflammation and oxidative stress. In conclusion, this study suggests for the first time that miRNAs, especially fibromiRs, mediate Tacrolimus-induced nephrotoxic effects.

overall_design: CD-1 mice were treated with Tacrolimus (1mg/kg/d for 28 days) or physiological serum as control (n=4) and total RNA was prepared from kidneys. Experiment performed in a one color design, corresponding to 2 conditions (Control: CTL and Tacrolimus: TAC) for a total of 8 samples.

first: C

last: Cauffiez

first: G

last: Savary

first: N

last: Nottet

last: Pottier

first: B

last: Mari


title: Analysis of the impact of CDK activity regulation on DNA replication

status: Public on Feb 04 2018

city: Bordeaux

first: Pei-Yun Jenny

last: Wu

address: 1 Rue Camille Saint Saens

institute: Institute of Biochemistry and Cellular Genetics

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

overall_design: Refer to individual Series


title: Functional analysis of evening complex component LUX in Arabidopsis

city: Beijing

first: Lei

last: Wang

address: Nanxincun 20, Xiangshan, Haidian District

department: Key Laboratory of Plant Molecular Physiology

country: China

institute: insitute of botany, chinese academy of sciences

type: Expression profiling by high throughput sequencing

summary: Plants trigger leaf senescence to relocate energy and nutrients from aging leaves to developing tissues or storage organs to optimize the growth and reproduction under limited nutrients and energy conditions. Jasmonate signaling is one of the major endogenous hormone signals to induced leaf senescence in Arabidopsis. However, whether circadian clock will gate Jasmonate signaling to induce leaf senescence and the underlying precise mechanism is unclear. Here we find that the Evening Complex (EC) of core oscillator closely regulates leaf senescence. To identify the underlying mechanism of EC regulating leaf senescence, we conducted RNA-sequencing. Transcriptomic data reveals Evening complex extensively involves into JA signal transduction and responses. Moreover, the mutants of ELF3, ELF4 and LUX universly display the accelerated JA-induced leaf senescence phenotype, while their overexpression lines act reversely. In accordance with the transcript levels of JA immediate early induced JA-responsive gene MYC2 are up-regulated in lux mutants. Futhermore we demonstrated LUX can bind to to the promoter of MYC2 in vivo to represses its transcription. In addition, the accelerated JA-induced leaf senescence in mutants of evening complex can be overturned by myc2, myc3 and myc4 mutants redundantly. Collectively, our findings demonstrated the underlying molecular basis for circadian clock gating jasmonate signaling to induce leaf senescence through the module of evening complex to directly repressing MYC2 transcription. This novel established molecular module also refines complicated nodes between circadian clock and jasmonate signal in Arabidopsis.

overall_design: We analyzed the transcriptomic profile of Col-0 and lux-6 mutant using 10-day-old seedlings, as evening complex negatively regulates leaf senescence through its transcription repressive activity. Two biological replicates were performed with the tissues harvesting at ZT12, the peak expression time of evening complex components.

first: Yuanyuan

last: Zhang


title: Transcription of bovine milk and meat factors (BMMFs) in HEK293TT cells

status: Public on Feb 05 2018

city: Heidelberg

first: Sebastian

last: Eilebrecht

address: Im Neuenheimer Feld 280

department: Episomal-persistent DNA in Cancer and Chronic Diseases

country: Germany

institute: German Cancer Research Center

summary: We analyze the transcription of these bovine milk and meat factors (BMMFs) and MS isolates in human HEK293TT cells. While all analyzed DNA agents are readily transcribed, one of the MS brain isolates (MSBI1.176), sharing homology with a transmissible spongiform encephalopathy (TSE)-associated DNA molecule, is transcribed at highest levels. Our results support the transcription of an open reading frame (ORF) encoding a replication-associated protein, which is highly conserved among all isolated BMMFs. In the case of the cow milk isolate CMI1.252, transcription also occurs in a second large ORF. Transcriptome profiling upon BMMF expression identified host cellular gene expression changes related to cell cycle progression and cell viability control, indicating potential pathways for a pathogenic involvement of BMMFs.

sample_organism:
Homo sapiens
Sphinx1.76-related DNA

platform_organism:
Homo sapiens
Sphinx1.76-related DNA

overall_design: The BMMFs CMI1.252, CMI3.168, MSBI1.176 and MSBI2.176 were transfected into HEK293TT cells. RNA-Seq was performed in triplicates per isolate at day 3 post transfection

first: Agnes

last: Hotz-Wagenblatt

first: Victor

last: Sarachaga

first: Corinna

last: Whitley

first: Timo

last: Bund


title: Single-cell full-length total RNA sequencing uncovers dynamics of recursive splicing and enhancer RNAs

status: Public on Feb 12 2018

city: Tsukuba

first: Hiroaki

last: Tateno

state: Ibaraki

address: Tsukuba Central 2, 1-1-1 Umezono

country: Japan

institute: National Institute of Advanced Industrial Science and Technology (AIST)

summary: We report a novel single-cell total RNA-seq method, RamDA-seq, by combining a novel reverse transcription (RT) technology, RT with random displacement amplification (RT-RamDA), and not-so-random (NSR) primers. RT-RamDA provides global cDNA amplification directly from RNA during RT without any universal adopters, which benefits RT efficiency, simplification of the procedure, avoiding the step of PCR amplification, and decontamination of genomic DNA. NSR enables random priming while preventing cDNA synthesis from rRNAs. RamDA-seq showed high sensitivity to non-poly(A) RNAs and full-length coverage even for extremely long transcripts (>10 kb). Moreover, RamDA-seq revealed recursive splicing, a multi-step, splicing, within > 300 kbp-long introns. Finally, RamDA-seq enabled the first genome-wide analysis of enhancer RNAs (eRNAs) in single cells.

overall_design:
Total 1,172 single cells (mouse ES, PrE and differentiating cells from ES cells) were sequenced by using RamDA-seq protocol:
(1) Cell cycle samples (277 RamDA-seq samples with mouse ES cells from G1, S, and G2M phases) were sequenced.
(2) Cell differentiation time series RamDA-seq samples (456 RamDA-seq samples with mouse ES cells collected 0, 12, 24, 48, and 72 h after the induction of cell differentiation into PrE cells ) were sequenced.
(3) 55 C1-RamDA-seq samples with mouse ES and PrE cells were sequenced.
(4) To evaluate plate-to-plate variability (batch effect) of RamDA-seq, two experimenters performed RamDA-seq with two plates (384 cells) of single-cell samples (consisting of mouse ES and PrE cells) on two separate days.

For assesment of RamDA-seq, 9 averaged single-cell lysate samples and 16 purified total RNA samples diluted to 10 pg were sequenced by using RamDA-seq protocol. We also sequenced 4 SMART-Seq v4 samples using 10 pg of total RNA. Moreover, 6 poly(A) RNA-selected and 6 rRNA-depleted samples derived from 1 ug of total RNA were sequenced by using a commercially available library prepararion kit. Furthermore, to evaluate reproducibility, we sequenced ~96 diluted RNA samples (corresponding to the entire plate) with RamDA-seq, C1-RamDA-seq, and C1-SMART-seq v4.

To compare RamDA-seq and rRNA-depleted RNA-seq prepared with equal amounts of input RNA, we sequenced RamDA-seq samples using 1 ng of diluted total RNA and rRNA-depleted RNA-seq samples using using 1 and 10 ng of diluted total RNA.

first: Tetsutaro

last: Hayashi

first: Haruka

last: Ozaki

first: Yohei

last: Sasagawa

first: Itoshi

last: Nikaido


title: Characterizing steroid hormone receptor chromatin binding landscapes in male and female breast cancer

status: Public on Feb 02 2018

city: Amsterdam

first: Wilbert

last: Zwart

address: Plesmanlaan 121

country: Netherlands

institute: The Netherlands Cancer Institute

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Male breast cancer (MBC) is rare and poorly characterized. Like the female counterpart, most MBCs are hormonally driven, but resistance to hormonal treatment is common. The pan-hormonal action of steroid hormonal receptors including Estrogen Receptor alpha (ERα), Androgen Receptor (AR), Progesterone Receptor (PR) and Glucocorticoid Receptor (GR) in this understudied tumor type remains wholly unexamined. This pioneering study reveals genomic cross-talk of steroid hormone receptor action and interplay in human tumors, here in the context of MBC, in relation to patient outcome.  Using chromatin immunoprecipitation coupled with massively-parallel sequencing (ChIP-seq), we characterized human MBCs for epigenetic make-up of hormonal regulation in human tumors, revealing genome-wide chromatin binding landscapes of ERα, AR, PR and GR, along with pioneer factor FOXA1 and enhancer-enriched histone mark H3K4me1. These data were integrated with transcriptomics analyses to reveal gender-selective and genomic location-specific hormone receptor action, that are associated with survival in MBC patients.

overall_design: Binding of steroid hormone receptors (ER, AR, PR, GR) together with pioneer factors (FOXA1, GATA3) and a histone mark (H3K4me1) profiled in subsets of 30 males and 8 females breast tumors, and MCF-7 by ChIP-seq (Chromatin Immunoprecipitation followed by deep sequencing).

first: Karianne

last: Schuurman

first: Stacey

last: Joosten

first: Yongsoo

last: Kim

first: Tesa

middle: M

last: Severson

first: Petra

last: van der Groep

first: Paul

last: van Diest


title: Replication origin mapping of cdc25-22 and Cdc13-Cdc2 cells after G2 block

first: Pei-Yun Jenny

last: Wu

summary:
Identification of sites of replication initiation by copy number analysis, comparing samples before and after entry into S phase.
Experiments were performed in the following strains and conditions: cdc25-22 (temperature-sensitive mutation arrests cells at G2/M for synchrony, otherwise wild type), G2 arrest; Cdc13-Cdc2, G2 arrest.

overall_design: Comparison of samples before and after S phase entry; dye-swap experiments were performed and averaged.

first: Anthony

last: Perrot

first: Pei-Yun


title: Dynamic Quantitative Analytic Response of Human Natural Killer Cells at Single-Cell Resolution in B-cell Non-Hodgkin Lymphoma

status: Public on Feb 01 2018

city: Mountain View

first: Afshin

last: Beheshti

state: CA

address: Moffett Blvd

country: USA

institute: NASA Ames Research Center

summary: Natural Killer (NK) cells are phenotypically and functionally diverse lymphocytes that recognize and kill cancer cells. The susceptibility of target cancer cells to NK cell-mediated cytotoxicity depends on the strength and balance of regulatory (activating/inhibitory) ligands expressed on target cell surface. We performed gene expression arrays to determine patterns of NK cell ligands associated with B-cell non-Hodgkin lymphoma (b-NHL). Microarray analyses revealed differential upregulation of a multitude of NK-activating and costimulatory ligands across varied lymphoma cell lines and b-NHL cells including ULBP1, CD72, CD48 and SLAMF6. To correlate genetic signatures with functional anti-lymphoma activity, we developed a dynamic and quantitative cytotoxicity assay in an integrated microfluidic droplet array. This assay facilitated the examination of transient interactions between NK cells and target lymphoma cells and also the functional heterogeneity in NK cell effector outcomes at single-cell level. Individual NK cells and target lymphoma cells were co-encapsulated in picoliter-volume droplets and imaged over time to quantify heterotypic cell-cell interaction. We identified significant variability in NK-lymphoma cell contact duration, frequency and subsequent cytolysis. The death of lymphoma cells undergoing single contact with NK cells occurred faster than cells that made multiple short contacts. NK cells also killed target cells in droplets via contact-independent mechanisms that partially relied on calcium-dependent processes and perforin secretion, but not on cytokines (IFN-γ or TNF-α). We extended this technique to characterize cytolysis of primary cells from b-NHL patients. Diffuse large B-cell lymphoma cells from two patients showed similar contact durations with NK cells; primary Burkitt lymphoma cells made longer contacts and were lysed at later times. We further tested the cytotoxic efficacy of NK-92, a continuously growing NK cell line being investigated as an anti-tumor therapy, using our droplet-based bioassay. NK-92 cells were found to be more efficient in killing b-NHL cells compared with primary/fresh NK cells, requiring shorter contacts for faster killing activity. Taken together, our combined genetic and microfluidic analysis demonstrate significant heterogeneity in the dynamic interaction and associated cytotoxicity between NK cellular therapy and b-NHL cells.

overall_design:
For genome-wide expression profiling of SUDHL4, SUHDL10, EL-2 (also called DLBCL.1), EL-5 (also called DLBCL.2), and Raji cells were grown under normal conditions and Human HT-12 BeadArray chips (Illumina, San Diego, CA) were used. RNA was isolated using RNeasy Minikit (Qiagen), following instructions recommended supplied by the manufacturer. Purity and the yield of isolated RNA was determined using Bioanalyzer. These experiments were performed in biological triplicates. All replicate samples were run individually. Total RNA of 500ng per sample was amplified using AmbionTotalPrep, and 1.5ug of the product was loaded onto the chips. Following hybridization at 55C, the chips were washed and then scanned using the Illumina iScan System. The data was checked with GenomeStudio (Illumina) for quality control. Data were corrected through normalization with the housekeeping genes, quantile normalized, collapsed to genes from probes, then imported into MultiExperiment Viewer, MeV for analysis. Statistically significant genes were determined by applying a one-way ANOVA with an adjusted Bonferroni correction and false discovery rate (FDR) < 0.05 that resulted in a list of significant genes.
------------------------------------
This dataset does not include experimental samples

first: Ravi

last: Dashnamoorthy

first: Saheli

last: Sarkar

first: Andrew

last: Evens

first: Tania

last: Konry


title: Role of BmoR in Bacteroides fragilis response to oxidative stress

city: Rio de Janeiro

first: Felipe

middle: Lopes

last: Teixeira

state: Rio de Janeiro

address: Av. Carlos Chagas Filho 373

department: Microbiologia Médica

country: Brazil

institute: Universidade Federal do Rio de Janeiro

summary: Comparison of transcriptional profile of wild-type and bmoR mutant strains of Bacteroides fragilis grown under anaerobic conditions and after exposure to atmospheric oxygen for 1 hour.

overall_design: This is an eight chip study using two biological replicates of each strain under anaerobic or oxygen exposure conditions. Anaerobic grown wild-type strain was used as reference for alterations in gene expression.

middle: L

first: Edson

middle: R

last: Rocha

first: Charles

middle: J

last: Smith


title: Genome-wide DNA-binding analysis for E(spl)m8 in intestinal stem cells by DamID

first: huanwei

last: huang

address: 7 Science Park Rd., ZGC life science park

institute: NIBS

summary:
Purpose:  Genome-wide DNA-binding analysis for E(spl)m8 in intestinal stem cells in Drosophila midgut by DNA adenine methyltransferase identification(DamID)
Methods: DNA adenine methyltransferase identification (DamID) on E(spl)m8 driven by Dl-Gal4
Results:

overall_design: Dam(Ctrl) or E(spl)m8-Dam transgenes were expressed in intestinal stem cells by Dl-Gal4.The guts within 2 days was used for DamID-seq, followed by deep sequencing analysis.

first: Jun

last: Chen

first: Rongwen

last: Xi


title: A SRp55-regulated alternative splicing network controls pancreatic beta cell survival and function

city: Brussels

first: Jonas

last: Juan Mateu

address: Route de Lennik 808 CP 618

department: Faculty of Medecine

country: Belgium

institute: Universite Libre de Bruxelles

summary: Progressive failure of insulin-producing beta cells is the central event leading to diabetes, yet the signalling networks controlling beta cell fate remain poorly understood. Here we show that SRp55, a splicing factor regulated by the diabetes susceptibility gene GLIS3, has a major role in maintaining function and survival of human beta cells. RNA-seq analysis revealed that SRp55 regulates the splicing of genes involved in cell survival and death, insulin secretion and JNK signalling. Specifically, SRp55-mediated splicing changes modulate the function of the pro-apoptotic proteins BIM and BAX, JNK signalling and endoplasmic reticulum stress, explaining why SRp55 depletion triggers beta cell apoptosis. Furthermore, SRp55 depletion inhibits beta cell mitochondrial function, explaining the observed decrease in insulin release. These data unveil a novel layer of regulation of human beta cell function and survival, namely alternative splicing modulated by key splicing regulators such as SRp55 that may crosstalk with candidate genes for diabetes.

overall_design: Five independent preparations of EndoC-βH1 cells exposed to control (siCTL) or SRp55 (siSR#2) siRNAs

last: Mateu

first: Maria

middle: I

last: Alvelos

first: Jean-Valery

last: Turatsinze

first: Decio

last: Eizirik


title: Lung Epithelial Response to Cigarette Smoke and Modulation by the Nicotinic Alpha 7 Receptor

city: Salt Lake City

first: Robert

middle: B

last: Weiss

state: UT

department: Human Genetics

institute: University of Utah

summary: We report the impact of side-stream cigarette smoking on baseline tracriptional status of enriched epithelium from the distal lung of both male and female control mice or mice harboring a mutation in the nicotinc alpha7 recptor that selectivley diminshes the calcium current (E260A).

overall_design: Mice (male or female) of each nicotinic recptor alpha7 genotype (Control (c) or mutant (E260A)) were exposed to side-stream cigarette smoke 5 days per week for four months.  The distal lung epithelium was enriched and poly-adenylated strand-specific RNA-Seq libraries using Illumina TruSeq stranded mRNA were preared for analysis.

first: Scott

last: Rogers


title: TGF-beta signature in cholangiocarcinoma cell lines

city: Rennes

first: Cedric

last: Coulouarn

address: CLCC Eugène Marquis, Rue de la Bataille Flandres Dunkerque, Bat D, 1er étage

institute: INSERM

summary: The aim of the study was to characterize the transcriptional profiles of two cholangiocarcinoma cell lines (HuCCT1 and Huh28) after a treatment with Transforming Growth Factor beta (TGF-beta).

overall_design: HuCCT1 (RCB-1960) and Huh28 (RCB-1943) cholangiocarcinoma cell lines after treatment with Transforming Growth Factor beta (TGF-beta).


title: Temporal control of metabolic amplitude by Nocturnin

city: Dallas

first: Gokhul

middle: Krishna

last: Kilaru

state: TEXAS

address: 6000 Harry Hines Boulevard

department: Department of Neuroscience

institute: University of Texas Southwestern Medical Center

summary: The timing of food intake and nutrient utilization is critical to health and regulated in part by the circadian clock.  Recently, increased amplitude of circadian oscillations and metabolic output was found to improve health in diabetic and obesity mouse models.  Here, we report a novel function for the circadian deadenylase Nocturnin as a regulator of metabolic amplitude across the day/night cycle and in response to nutrient challenge.  We show that mice lacking Nocturnin (Noct-/-) display significantly increased amplitudes of mRNA expression of hepatic genes encoding key metabolic enzymes regulating lipid and cholesterol synthesis, both over the daily circadian cycle and in response to fasting and refeeding.  Noct-/- mice have increased plasma triglyceride throughout the night and increased amplitude of hepatic cholesterol levels.  Therefore, posttranscriptional control by Nocturnin regulates the amplitude of these critical metabolic pathways and loss of this activity results in increased metabolic flux and reduced obesity.

overall_design: Total RNA was isolated from liver samples of Wildtype (WT) mice and mice lacking Nocturnin (Noct-/-) and the hepatic transcriptome analyzed by mRNA-seq.  WT and Noct-/- mice were on a light:dark LD 12:12 cycle and fed standard chow Ad Libitum.

first: Jeremy

last: Stubblefield

first: Peng

last: Gao

middle: K

first: Bilal

last: Mukadam

last: Terrien

first: Carla

last: Green


title: HNRNPH1 is required for rhabdomyosarcoma cell growth and survival

city: Memphis

first: David

last: Finkelstein

state: TN

address: 332 N. Lauderdale St.

department: Computational Biology

institute: St Jude Children's Research Hospital

summary: Rhabdomyosarcoma (RMS) is an aggressive and difficult to treat cancer characterized by a muscle-like phenotype. Although the average 5-year survival rate is 65% for newly diagnosed RMS, the treatment options for metastatic disease are limited in efficacy, with the 5-year survival rate plummeting to 30%. Heterogenous nuclear ribonucleoprotein H1 (HNRNPH1) is an RNA-binding protein that is highly expressed in many cancers, including RMS. To determine the role HNRNPH1 plays in RMS tumorigenesis, we investigated its expression and effect on growth in 3 cellular models of RMS: RD, RH30, and RH41 cells. Upon knockdown of HNRNPH1, growth of all cell lines was dramatically reduced, most likely through a combination of apoptosis and cell cycle arrest. We then recapitulated this finding by performing in vivo xenograft studies, in which knockdown of HNRNPH1 resulted in a striking reduction in tumor formation and growth. We used RNA sequencing to identify changes in gene expression after HNRNPH1 knockdown and found altered splicing of some oncogenes. Our data show that HNRNPH1 may be an important molecular target for the development of future RMS therapy.

overall_design: Two individual siRNAs targeting HNRNPH1 were used to knockdown HNRNPH1 in RD, RH30, and RH41 cells. Both siRNAs were done in triplicate in all cell lines. Briefly, 20nM siRNA was transfected with RNAiMax in 6-well dishes (2 × 105 RD and RH30 or 2.4 × 105 RH41) for 12 hours, after which media was changed and cells collected 48 hours post transfection.

first: Yangfeng

last: Li

first: Jesse

last: Bakke

first: Hu

last: Zeng

first: Jing

last: Wu

first: Taosheng


title: Spatially resolved RNA-sequencing of the embryonic zebrafish heart

city: Utrecht

first: Silja

middle: Barbara

last: Burkhard

address: Uppsalalaan 8

department: Cardiac Development and Genetics

institute: Hubrecht Institute

summary: Development of specialized cell types and structures in the vertebrate heart is regulated by spatially-restricted molecular pathways. Disruptions in these pathways can cause severe congenital cardiac malformations or functional defects. To better understand these pathways and how they regulate cardiac development and function we used tomo-seq, combining high-throughput RNA sequencing with tissue sectioning, to establish a genome-wide expression dataset with high spatial resolution for the developing zebrafish heart. Analysis of the dataset revealed over 1100 genes differentially expressed in sub-compartments. Pacemaker cells in the sinoatrial region induce heart contractions, but little is known about the mechanisms underlying their development and function. Using our transcriptome map, we identified spatially restricted Wnt/β-catenin signaling activity in pacemaker cells, which was controlled by Islet-1 activity. Moreover, Wnt/β-catenin signaling at a specific developmental stage in the myocardium controls heart rate by regulating pacemaker cellular response to parasympathetic stimuli. Thus, this high-resolution transcriptome map incorporating all cell types in the embryonic heart can expose spatially-restricted molecular pathways critical for specific cardiac functions.

overall_design: To generate spatially-resolved RNA-seq data for the developing zebrafish hearts (2 days post fertilization), we cryosectioned 3 hearts, extracted RNA from the individual sections, amplified and barcoded mRNA using the CEL-seq protocol (Hashimshony et al., Cell Reports, 2012) with a few modifications. Libraries were sequenced on Illumina NextSeq using 75bp paired end sequencing. Sample Heart #1 is the primary sample. Heart #2 and #3 are biological replicates used for comparison.

first: Jeroen

last: Bakkers


title: PARP14 controls the nuclear accumulation of a subset of type I Interferon-inducible proteins

status: Public on Feb 07 2018

city: Milano

first: Viviana

last: Piccolo

state: Milano

address: Via Adamello 16

department: Department of Experimental Oncology

country: Italy

institute: European Institute of Oncology (IEO)


title: Functions of Arabidopsis REI1-LIKE cytosolic ribosome biogenesis factor homologs

city: Potsdam-Golm

first: Dirk

last: Walther

address: Am Mühlenberg 1

department: Bioinformatics

institute: Max Planck Institute for Molecular Plant Physiology

summary:
Functions of Arabidopsis REI1-LIKE (REIL) proteins, two homologs of a yeast ribosome biogenesis protein (RBP) that takes part in the late cytoplasmic steps of 60S ribosomal subunit maturation, were characterized by systems analyses of the 10°C cold acclimating reil1-1 reil2-1 double mutant compared to Col-0 wildtype. The mutant lacked both REIL proteins, was strongly growth inhibited in the cold and complemented by constitutive expression of N-terminal FLUORESCENT PROTEIN (FP)-REIL1 and FP-REIL2 fusion proteins under control of the UBIQUITIN 10 promoter. Wildtype acclimation to 10°C causes relative accumulation of cytosolic ribosome subunits and rRNA. Expression of cytosolic ribosomal genes, known cytosolic RBPs, translation initiation- and elongation-factors was activated. Conserved function of Arabidopsis REIL proteins was indicated by delay of these processes in reil1-1 reil2-1, cytosolic localization of FP-REIL proteins and native REIL protein interactions with 60S containing ribosome fractions. Non-acclimated reil1-1 reil2-1 triggered plant specific metabolic and transcriptomic cold acclimation responses that included activation of the DREB/CBF-regulon with a preference for the cold acclimation factors, CBF1/DREB1B, CBF2/DREB1C, and CBF3/DREB1A. Cold-acclimating reil1-1 reil2-1 maintained cellular integrity and acquired freezing tolerance but did not activate FLOWERING LOCUS T expression in mature leaves. This block was independent of FLOWERING LOCUS C and AGAMOUS-LIKE 19 mediated vernalization. We conclude that Arabidopsis REIL proteins enhance accumulation of cytosolic ribosome subunits after cold shift and either directly or indirectly feedback on temperature perception by suppression of premature cold acclimation at optimized temperature and by triggering growth and the vegetative to generative phase transition in the cold.
Transcriptomic profiling demonstrated a hidden acclimation phenotype of the morphologically inconspicuous reil1-1reil2-1 mutant under optimized temperature conditions. Premature triggering of cold acclimation, a severe growth defect at 10°C and compensation responses indicate that REIL function may extend beyond cytosolic ribosome biogenesis towards translation initiation.

overall_design: Cold temperature induced gene expression in Arabidopsis reil1-1reil2-1 mutant in comparison to the wild type Col-0 was measured at 20°C, 24 hours, 1 week and 3 weeks after shift from 20°C to the 10°C temperatures.  3 to 6 replicates were harvested for each line and at each time point before or after cold treatment

first: Olga

last: Beine-Golovchuk

first: Joachim

last: Kopka


title: SHED: control vs Hepatocyte-like cells

city: Fukuoka

first: Haruyoshi

last: yamaza

address: 3-1-1 Maidashi, Higashi-ku

department: Division of Oral Health, Growth & Development

institute: Faculty of Dental Science, Kyushu University

type: Genome variation profiling by genome tiling array

summary: Profiling of chromosomal gains and losses in a genomic complement of stem cells from human exfoliated deciduous teeth comparing control undifferentieated SHED with hepatocyte differentiated SHED.

overall_design: Two-condition experiment, control vs. Hepatocyte differentiated cells. Biological replicates: 6 control replicates, 6 differentiated replicates.

last: Yamaza


title: iCLIP-based modeling uncovers 3’ splice site definition: how U2AF65 specificity relies on regulation by co-factors [in vitro iCLIP co-factors]

city: Mainz

first: Stefanie

last: Ebersberger

address: Ackermannweg 4

institute: Institute of Molecular Biology

summary: Alternative splicing generates distinct mRNA isoforms and is crucial for proteome diversity in eukaryotes. The RNA-binding protein (RBP) U2AF65 is central to splicing decisions, as it recognizes 3' splice sites and recruits the spliceosome. We established 'in vitro iCLIP' experiments, in which recombinant RBPs are incubated with long transcripts, to study how U2AF65 recognizes RNA sequences and how this is modulated by trans-acting RBPs. We quantitatively measure U2AF65 affinities at hundreds of binding sites, and compare in vitro and in vivo binding landscapes by mathematical modelling. We find that trans-acting RBPs extensively regulate U2AF65 binding in vivo, including enhanced recruitment to 3' splice sites and clearance of intronic regions. Using machine learning, we identify novel trans-acting RBPs (including FUBP1, BRUNOL6 and PCBP1) that modulate U2AF65 binding and affect splicing outcomes. Our study offers a blueprint for the high-throughput characterization of in vitro mRNP assembly and in vivo splicing regulation.

overall_design: Comparison of in vitro and in vivo iCLIP data of U2AF65 and other regulatory RBPs

first: Reymond

last: Sutandy


title: iCLIP-based modeling uncovers 3’ splice site definition: how U2AF65 specificity relies on regulation by co-factors [in vivo iCLIP]


title: Lipid addiction via Fatty Acid Synthase activates PI3K signaling in B cell non-Hodgkin’s Lymphoma (bNHL) [SUDHL2, SUHDL10, and Raji]

first: Afshin

last: Beheshti

summary: Dysregulated lipid metabolism is an oncogenic mechanism that leads to increased de novo fatty acid biosynthesis via elevated expression of fatty acid synthase (FASN) in cancer. FASN catalyzes biosynthesis of (C16) palmitic acid by using Acetyl CoA, malonyl CoA and NADPH as substrates. Palmitic acid and its derivatives participate as signaling lipids andinvolved in the regulation of receptor functions associated with many biological processes. and furtherThis includes contribute formation of cell structures, and all these processes which are essential for both normal and malignant cell growth and proliferation. In this study, we investigated the biological impact of FASN inhibition in B cell Non Hodgkin Lymphoma (bNHL) cells using FASN inhibitors Cerulenin and other novel agents (TVB3166, TVB3567).  Inhibition of FASN resulted in a dose dependent increase in cell death and apoptosis in the bNHL and primary tumor cells. Transcriptomic and comprehensive systems biology analysis of cerulenin, TVB3166 or TVB3567 treated bNHL cells revealed conserved activation of (TNF) immune and PI3K signaling, and inhibition of cell cycle pathways as prominent responses to FASN inhibition. Cell fractionation and western blot analysis of cCerulenin treated cells showed translocation of TNF receptor adaptor proteins (TRAF2 and RIP), docking of AKT and PI3K/p110 subunits to cell membrane and activation of PI3K/AKT pathways by phosphorylation in SUDHL2 and SUDHL4 bNHL cells, while these mechanisms were constitutively active in cerulenin treated SUDHL10 bNHL cells. Disruption of lipid raft formation by methyl β cyclodextrin inhibited PI3K signaling and downregulated FASN expression in SUDHL10 cells.  Subsequently, we observed that combination of FASN (cerulenin) and PI3K (Buparlisib) inhibitors resulted in synergistic cell death in bNHL cell lines and increased apoptosis accompanied with down-regulation of FASN expression in bNHL cell lines and primary cells, implicating the importance of PI3K in the regulation of lipid metabolism. Further mass spectrometric evaluation of cerulenin and Buparlisib treated DLBCL cells fed with C14C glucose, synergistically decreased the biosynthesis of palmitic acid and its derivatives compared to single agent treatments. Our conclusions from these investigations suggest that simultaneous blocking of PI3K and FASN is likely to exert stronger inhibition of lipid metabolism and enhance cell death in lymphoma.

overall_design: For genome-wide expression profiling of SUDHL2, SUHDL10, and Raji cells treated with 12.5uM Cerulenin and IC50 drug concentrations for TVB3567 and TVB3166 for 48 hours, Human HT-12 BeadArray chips (Illumina, San Diego, CA) were used. RNA was isolated using RNeasy Minikit (Qiagen), following instructions recommended supplied by the manufacturer. Purity and the yield of isolated RNA was determined using Bioanalyzer. These experiments were performed in biological triplicates. All replicate samples were run individually. Total RNA of 500ng per sample was amplified using AmbionTotalPrep, and 1.5ug of the product was loaded onto the chips. Following hybridization at 55C, the chips were washed and then scanned using the Illumina iScan System. The data was checked with GenomeStudio (Illumina) for quality control. Data were corrected through COMBAT correction, quantile normalized, collapsed to genes from probes, then imported into MultiExperiment Viewer, MeV for analysis. Statistically significant genes were determined by applying a one-way ANOVA with an adjusted Bonferroni correction and false discovery rate (FDR) < 0.05 that resulted in a list of significant genes.

first: Ravi

last: Dashnamoorthy

first: Nasséra

last: Abermil

first: Andrew

last: Evens


title: LSD1 inhibitor T-3775440 inhibits SCLC cell proliferation by disrupting LSD1 interactions with SNAG domain proteins INSM1 and GFI1B

city: Fujisawa

first: SHINJI

last: TAKAGI

address: 26-1 Muraoka Higashi 2-chome

department: Oncology Drug Discovery Unit

institute: Takeda Pharmaceutical Company

summary: T-3775440 is an irreversible inhibitor of the chromatin demethylase LSD1. Here we describe the anti-cancer effects and mechanism of action of T-3775440 in small cell lung cancer (SCLC). T-3775440 inhibited proliferation of SCLC cells in vitro and retarded SCLC tumor growth in vivo. Our results argue that LSD1 plays an important role in neuroendocrine-associated transcription and cell proliferation of SCLC via interactions with the SNAG domain proteins INSM1 and GFI1B. Targeting these critical interactions with LSD1 inhibitors offers a novel rational strategy to therapeutically manage SCLC.

overall_design: Total RNA was obtained from cells treated with DMSO or 500 nmol/L T-3775440 for 72 h in vitro (n=3/group). Gene expressions in T-3775440-treated SCLC cells were compared to those in DMSO-treated cells.

first: Shinji

last: Takagi

first: Koji

last: Yamamoto


title: Transcriptional Pause Sites Delineate Stable Nucleosome-Associated Premature Polyadenylation Suppressed by U1 snRNP [2P-seq]

city: Nagoya

first: Hiroshi

middle: I.

last: Suzuki

address: 65 Tsurumai-cho, Showa-ku

department: DIvision of Molecular Oncology, Graduate School of Medicine

institute: Nagoya University

summary: Regulation of RNA polymerase II (Pol II) elongation is a critical step in gene regulation. Here, we report that U1 snRNP recognition and transcription pausing at stable nucleosomes are linked through premature polyadenylation signal (PAS) termination. By generating RNA exosome conditional deletion mouse embryonic stem cells, we identified a large class of polyadenylated short transcripts in the sense direction destabilized by the RNA exosome. These PAS termination events are enriched at the first few stable nucleosomes flanking CpG islands and suppressed by U1 snRNP. Thus, promoter-proximal Pol II pausing consists of two processes: TSS-proximal and +1 stable nucleosome pausing, with PAS termination coinciding with the latter. While pausing factors NELF/DSIF only function in the former step, flavopiridol-sensitive mechanism(s) and Myc modulate both steps. We propose that premature PAS termination near the nucleosome-associated pause site represents a common transcriptional elongation checkpoint regulated by U1 snRNP recognition, nucleosome stability, and Myc activity.

overall_design: 3'-End sequencing (2P-seq) in Exosc3 conditional KO mESCs

first: Anthony

middle: C

last: Chiu

first: Phillip

middle: A

last: Sharp


title: Transcriptional Pause Sites Delineate Stable Nucleosome-Associated Premature Polyadenylation Suppressed by U1 snRNP [RNA-seq]

overall_design: RNA sequencing (RNA-seq) in Exosc3 conditional KO mESCs


title: Transcriptional Pause Sites Delineate Stable Nucleosome-Associated Premature Polyadenylation Suppressed by U1 snRNP

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

overall_design: Refer to individual Series


title: Unexpected conformational variations of the human centromeric chromatin complex

status: Public on Jan 31 2018

city: Seattle

first: Jorja

last: Henikoff

state: WA

address: 1100 Fairview AV N, A1-162

department: Basic Sciences

institute: Fred Hutchinson Cancer Research Center

summary: Centromeres of most eukaryotes are multi-megabase arrays of satellite DNA that assemble proteinaceous kinetochores to facilitate faithful chromosome segregation. However, the nature of the chromatin landscape at centromeres remains unclear, in part due to the difficulty of isolating intact centromeric chromatin rendered insoluble by megadalton-size kinetochore protein complexes. To address this challenge we combined classical salt fractionation with chromatin immunoprecipitation to recover human centromeric chromatin under native conditions. We found that >85% of the total centromeric chromatin is insoluble under conditions typically used for native chromatin extraction. To map both soluble and insoluble chromatin in situ, we combined CUT&RUN, a targeted nuclease method, with salt fractionation. Using this approach, we found that the strength of Centromere Protein B (CENP-B) binding and the density of CENP-B motifs corresponds to the occupancy of Constitutive Centromere-Assocated Network (CCAN) complexes bound to α-satellite arrays. We also observed unexpected structural variations of CENP-A-containing complexes on different α-satellite dimeric units within highly homogenous arrays. Our results suggest that slight α-satellite sequence differences controls the structure and occupancy of the associated centromeric chromatin complex.

overall_design: We used Cleavage under targets and Release using nuclease (Cut-and-Run), a chromatin profiling strategy in which antibody-targeted controlled cleavage by micrococcal nuclease releases specific protein-DNA complexes into the supernatant for paired-end DNA sequencing.

first: Jitendra

last: Thakur

first: Steven


title: iCLIP-based modeling uncovers 3’ splice site definition: how U2AF65 specificity relies on regulation by co-factors [in vitro iCLIP]


title: iCLIP-based modeling uncovers 3’ splice site definition: how U2AF65 specificity relies on regulation by co-factors

sample_organism:
Homo sapiens
synthetic gene

platform_organism:
Homo sapiens
synthetic gene


title: An Mll4/COMPASS-Lsd1 epigenetic axis governs enhancer function and pluripotency transition in embryonic stem cells

city: Chicago

first: Ali

last: Shilatifard

state: IL

address: 320 E Superior St

department: Department of Biochemistry and Molecular Genetics

institute: Northwestern University Feinberg School of Medicine

summary: Chromatin regulators control cellular differentiation by orchestrating dynamic developmental gene expression programs, and hence, malfunctions in the regulation of chromatin state contribute to both developmental disorders and disease state. Mll4 (Kmt2d), a member of the COMPASS (COMplex of Proteins ASsociated with Set1) protein family that implements histone H3 lysine 4 monomethylation (H3K4me1) at enhancers, is essential for embryonic development and functions as a pancancer tumor suppressor. We define the roles of Mll4/COMPASS and its catalytic activity in the maintenance and exit of ground-state pluripotency in murine embryonic stem cells (ESCs). Mll4 is required for ESC to exit the naive pluripotent state; however, its intrinsic catalytic activity is dispensable for this process. The depletion of the H3K4 demethylase Lsd1 (Kdm1a) restores the ability of Mll4 null ESCs to transition from naive to primed pluripotency. Thus, we define an opposing regulatory axis, wherein Lsd1 and associated co-repressors directly repress Mll4-activated gene targets. This finding has broad reaching implications for human developmental syndromes and the treatment of tumors carrying Mll4 mutations.

overall_design: Studying the functions of Mll4/COMPASS in ESC self-renewal and pluripotency transition by examining epigenetic modifications and gene expression.

first: Kaixiang

last: Cao

first: Clayton

last: Collings


title: Surveillance of rRNA synthesis by an RNA helicase mediates tissue-specific developmental disorders

city: Cambridge

first: Eliezer

last: Calo

state: MASSACHUSETTS

department: Biology

institute: Massachusetts Institute of Technology

summary: Myriad of craniofacial disorders are caused by heterozygous mutations in general regulators of housekeeping cellular functions such as ribosome biogenesis. While it is understood that many of these highly tissue-specific malformations are a consequence of defects in cranial neural crest cells (cNCCs), an embryonic cell group that gives rise to most of the facial structures during embryogenesis, the mechanism underlying cell type-selectivity of these effects remains largely unknown. Here we show that DDX21, a DEAD-box RNA helicase involved in control of both RNA polymerase (Pol) I and Pol II dependent transcriptional arms of ribosome biogenesis, is a key mediator of the nucleolar stress response. Using an in vitro model of Treacher Collins Syndrome (TCS), a craniofacial disorder caused by heterozygous mutations in components of the Pol I transcriptional machinery, we demonstrate that perturbations in ribosomal RNA (rRNA) transcription induce DDX21-mediated surveillance mechanism in cNCCs, leading to DDX21 relocalization from the nucleolus to the nucleoplasm, its eviction from Pol I and Pol II chromatin targets and inhibition of rRNA processing. Importantly, these effects are cell type-selective, cell-autonomous and involve activation of the p53 pathway. We further show that cNCCs are characterized by the inherently high reservoir of p53 mRNA and sensitivity to p53-mediated apoptosis. Remarkably, preventing the loss of DDX21 from nucleolus and chromatin can rescue both the sensitivity to apoptosis and TCS-associated craniofacial phenotypes in vivo. This mechanism is not restricted to TCS, as gene function perturbations linked to other ribosomopathies with craniofacial manifestations also lead to the activation of DDX21 surveillance. Lastly, we provide evidence that inhibition of rRNA synthesis leads to rDNA damage, and furthermore, that rDNA damage is sufficient to activate DDX21 surveillance and elicits tissue-selective and dosage-dependent effects on craniofacial development in vivo.

overall_design:
Examination of DDX21 and TCOF1 binding to chromatin

Examination of DDX21 irCLIP from HeLa with DMSO or ActD treatments

first: Joanna

last: Wysocka


title: Viral like infection induces Human β cell dedifferentiation

city: Paris

first: Masaya

last: OSHIMA

address: 123 boulevard de Port Royal

institute: INSERM U1016


title: Enhancer of Polycomb maintains germline activity and genome integrity in Drosophila testis

city: baltimore

first: Xin

last: Chen

state: MD

address: 3400 N Charles St

institute: The Joihns Hopkins University

summary: Tissue homeostasis depends on the activities of tissue-specific adult stem cells to maintain a balance between proliferation and differentiation and ensure DNA damage repair. Here, we use the Drosophila male germline stem cell lineage to study how a chromatin factor, Enhancer of Polycomb [E(Pc)], regulates the proliferation-to-differentiation (mitosis-to-meiosis) transition and DNA damage repair. We identified two critical target genes of E(Pc). First, E(Pc) directly represses CycB transcription through modulating H4 acetylation. Second, E(Pc) is required for accumulation of an important germline differentiation factor, Bag-of-marbles (Bam) protein, through post-transcriptional regulation. When E(Pc) is downregulated, increased CycB transcription and decreased Bam protein are both responsible for defective mitosis-to-meiosis transition in germ cells. Moreover, E(Pc) is required for the DNA double-strand break repair, the failure of which leads to germ cell death. Finally, compromising the activity of Tip60, a histone acetyltransferase, leads to germline defects similar to E(Pc) loss-of-function, suggesting that E(Pc) acts cooperatively with Tip60 . Together, our data demonstrate that E(Pc) has pleiotropic roles in maintaining male germline activity and genome integrity. E(Pc) is highly conserved with implications in cancers; consequently, our findings will help elucidate the in vivo mechanisms of E(Pc), in turn making this chromatin factor a promising target for cancer treatment.

overall_design: Examination of direct binding site of E(Pc) in Drosophila S2 cells

first: Lijuan

last: Feng

first: Zhen

last: Shi


title: MicroRNA-130a and miR-212 disrupt the intestinal epithelial barrier through modulation of peroxisome proliferator-activated receptor gamma and occludin expression in chronic SIV-infected rhesus macaques

city: San Antonio

first: Mahesh

last: Mohan

state: Texas

address: 8715 West Military Road

department: Host Pathogen Interaction Program

institute: Southwest National Primate Research Center

summary: We profiled and characterized mRNA expression exclusively in colonic epithelium (CE) of 4 chronically SIV-infected and 4 control rhesus macaques (RMs). Further analysis identified significant dowregulation of genes Genes associated with ion transport, epithelial barrier integrity/function, protection against oxidative injury, double stranded DNA damage repair and autophagy. Similarly, mRNA expression of genes associated with interferon response, anti-microbial defense, apoptosis, oxidative DNA damage, endoplasmic reticulum stress and inflammasome signaling were markedly upregulated.

overall_design: Colon tissue was harvested from 4 chronically SIV-infected and 4 control rhesus macaques. Colonic epithelial cells were isolated from intact colon tissue and then lysed in Qiazol solution (Qiagen Inc) for total RNA extraction. Comparisons in gene expression were made between 4 chronic SIV and 4 uninfeced control animals.


title: Estradiol and ICI 182,780 infusions induce similar transcriptomic response in late gestation fetal hypothalamus

first: Maria

middle: Belen

last: Rabaglino

address: Yalelaan 7

department: Population Health Science

institute: Utrecht University

summary: Estradiol plays a critical role stimulating the fetal hypothalamus-pituitary-adrenal axis at the end of gestation. Estradiol action is mediated through nuclear and membrane receptors that can be modulated by ICI 182,780, a pure anti-estrogen compound. The objective of this study was to evaluate the transcriptomics of estradiol and ICI 182,780, testing the hypothesis that ICI 182,780 blocks the action of estradiol in the fetal hypothalamus. However, we found that a short term (48 hrs) infusion with ICI 182,780 induces a similar transcriptomic response than estradiol infusion in the late gestation ovine fetal hypothalamus, being more evident with higher doses of ICI 182,780. These results suggest that ICI 182,780 is primarily an agonist of estradiol in the developing brain.

overall_design: For direct TCS exposure, chronically catheterized late gestation fetal sheep were infused with vehicle (n=4) or TCS (250 μg/day; n=4) iv. For indirect TCS exposure, TCS (100 μg/kg/day; n=3) or vehicle (n=3) was infused into the maternal circulation. Fetal hypothalami were collected after 2 days of infusion, and gene expression was measured using Agilent 15k ovine microarrays.

first: Maureen

last: Keller-Wood

first: Charles

middle: E

last: Wood


title: PacBio sequencing of Cp/Wp-driven transcripts from B95-8 LCLs

city: Durham

first: Bryan

last: Cullen

state: NC

address: 213 Research Dr., 0045 CARL Bldg.

department: MGM

institute: Duke University

summary: PacBio sequencing was performed to identify alternative splice patterns of Cp/Wp-driven transcripts. We were particularly interested in splicing from Cp/Wp exons to the HF exon. The HF exon contains pri-miR-BHRF1-2 and 1-3, so splicing to the HF exon would indicate that these pri-miRNAs are capable of regulating the 5' EBNA-LP ORF in cis through excision by Drosha.

overall_design: Cp/Wp driven transcripts were isolated from B95-8 infected LCLs using a biotinylated oligo complimentary to the W2 exon. PacBio sequencing was then performed for these transcripts. Two SMRT cells were used looking at two different size selected samples (800-2kb and 2kb+)

first: Brigid

last: Poling


title: Gender differences of gene expression profiles in brain cardiovascular centers of SHRs

status: Public on Jan 30 2018

city: Tokyo

first: Sabine

last: Gouraud

address: 3-10-2  Osawa, Mitaka

institute: International Christian University


title: Expression data from SOX9 overexpressing EndoC-ßH1 cells

summary: Human ß cell dedifferentiation as a potent mechanism of diabetes is gaining prominence. Several data suggest an upregulation of the transcription factor SOX9, a progenitor and duct cell marker during ß cell dedifferentiation. However, its targets in such cells need more understanding. Here, we overexpressed SOX9 and a constitutively active mutant (VP16-SOX9∆TAD) in Human pancreatic beta EndoC-ßH1 cells in order to understand its targets.

overall_design: EndoC-ßH1 cells were transfected with either MCS-ires-GFP, VP16-ires-GFP, SOX9WT-ires-GFP or VP16-SOX9∆TAD-ires-GFP and FAC sorted 48h post transfection for RNA extraction and hybridization on Affymetrix microarrays

last: Oshima

first: Klaus

middle: P

last: Knoch

first: Marc

last: Diedisheim

first: Pierra

last: Cattan

first: Piero

last: Marchetti

first: Marco

last: Bugliani

first: Michele

last: Solimena

first: Olivier

last: Albagli

first: Raphaël

last: Scharfmann


title: Expression data from PolyIC treated EndoC-ßH1 cells

first: Masaya

last: OSHIMA

summary: Type 1 diabetes (T1D) is a chronic disease characterized by an autoimmune-mediated destruction of insulin-producing pancreatic β cells. Environmental factors such as viruses play an important role in the onset of T1D and interact with predisposing genes. Recent data suggest that viral infection of human islets leads to a decrease in insulin production rather than β cell death, suggesting loss of β cell identity. We undertook this study to examine whether viral infection could induce human ß cell dedifferentiation. Using the functional human β cell line EndoC-βH1, we demonstrate that polyinosinic-polycitidilic acid (PolyI:C), a synthetic double-stranded RNA that mimics a by-product of viral replication induces a decrease in β cell-specific gene expression. In parallel to this loss, the expression of progenitor-like genes such as SOX9 was activated following PolyI:C treatment or enteroviral infection. SOX9 was induced by the NF-kB pathway and also in a paracrine non-cell autonomous fashion through the secretion of IFNA. Finally, we identified new SOX9 targets in human β cells as new markers of dedifferentiation in T1D. These findings reveal that inflammatory signaling has clear implications in human β cell dedifferentiation.

overall_design: EndoC-ßH1 cells were treated with PolyIC for RNA extraction and hybridization on Affymetrix microarrays


title: Estrogen regulated genes in BCK4 tumors

city: Aurora

first: Britta

last: Jacobsen

state: CO

department: Pathology

institute: University of Colorado Anschutz Medical Campus

summary: Genes regulated in different tumor regions, cells isolated by LCM

overall_design: BCK4 cells grown in nod scid mice supplemented with either cellulose or beta-estradiol for 5 months

first: J

last: Harrell


title: Transcription factors drive Tet2-mediated enhancer demethylation to reprogram cell fate

city: Barcelona

first: Thomas

last: Graf

address: c/Dr. Aiguader, 88. PRBB Building

department: Gene Regulation, Stem Cells and Cancer

country: Spain

institute: CRG (Center for Genomic Regulation)

type: Methylation profiling by high throughput sequencing


title: SALP, a new NGS library construction method especially useful for the high-throughput characterization of chromatin openness states

city: NanJing

first: Jian

last: Wu

state: Jiangsu

address: Sipailou 2#, Sipailou 2#

institute: Southeast University

summary:
Background: Next-generation sequencing (NGS) is fundamental to the current biological and biomedical research. Construction of sequencing library is a key step of NGS. Therefore, various library construction methods have been explored. However, the current methods are still limited by some shortcomings.
Results: This study developed a new NGS library construction method, Single strand Adaptor Library Preparation (SALP), by using a novel single strand adaptor (SSA). SSA is a double-stranded oligonucleotide with a 3ʹ overhang of 3 random nucleotides, which can be efficiently ligated to the 3′ end of single strand DNA by T4 DNA ligase. SALP can be started with any denatured DNA fragments such as those sheared by Tn5 tagmentation, enzyme digestion and sonication. When started with Tn5-tagmented chromatin, SALP can overcome a key limitation of ATAC-seq and become a high-throughput NGS library construction method, SALP-seq, which can be used to comparatively characterize the chromatin openness state of multiple cells unbiasly. In this way, this study successfully characterized the comparative chromatin openness states of four different cell lines, including GM12878, HepG2, HeLa and 293T, with SALP-seq. Similarly, this study also successfully characterized the chromatin openness states of HepG2 cells with SALP-seq by using 105 to 500 cells.
Conclusions: This study developed a new NGS library construction method, SALP, by using a novel kind of single strand adaptor (SSA), which should has wide applications in the future due to its unique performance.

overall_design: The chromatin openness state of 9 samples (GM12878, HepG2, HeLa, 293T and 5 cell level of HepG2 cell ) were analyzed with SALP-seq. And two HepG2 genome DNA NGS libraries,  fragmented by Hind III digestion and sonication were constructed with SALP and sequenced.

first: Wei

last: Dai

first: Lin

first: Jinke

last: Wang


title: Enhancer invasion shapes MYCN dependent transcriptional amplification in neuroblastoma [ChIP-seq Th-MYCN]

city: Boston

first: James

last: Bradner

state: MA

address: 450 Brookline

department: Medical Oncology

institute: Dana-Farber Cancer Institute

summary: In neuroblastoma, amplification of the oncogenic basic helix-loop-helix (bHLH) transcription factor (TF) MYCN is the defining prognosticator of high-risk disease, occurs in one-third of neuroblastoma, and drastically reduces overall survival rates. As a proto-oncogene, targeted MYCN overexpression in peripheral neural crest is sufficient to initiate disease in mouse models. In MYCN amplified neuroblastoma, elevated expression of the factor is crucial to maintain tumor stemness and is associated with increased proliferation and aberrant cell cycle progression, as these tumors lack the ability to arrest in G1 in response to irradiation. MYCN down-regulation broadly reverses these oncogenic phenotypes in a variety of neuroblastoma models and recent thereapeutic strategies to indirectly target MYCN production or protein stability have reduced tumor growth in vivo. These observations motivate an investigation of MYCN binding in MYCN amplified tumors as it remains fundamentally unclear how elevated levels of the factor occupy the genome and alter transcriptional programs in neuroblastoma. Here we present the first dynamic chromatin and transcriptional landscape of direct MYCN perturbation in neuroblastoma. We find that at oncogenic levels, MYCN associates with E-box (CANNTG) binding motifs in an affinity dependent manner across most active cis-regulatory promoters and enhancers. MYCN shutdown globally reduces histone acetylation and transcription, consistent with prior descriptions of MYC proteins as non-linear amplifiers of gene expression. We establish that MYCN load at the promoter and proximal enhancers predicts transcriptional responsiveness to MYCN shutdown and that MYCN enhancer binding occurs prominently at the most strongly occupied and down-regulated genes, suggesting a role for these tissue specific elements in predicating MYCN responsive “target” genes. At these invaded enhancers, we identify the lineage specific bHLH TWIST1 as a key collaborator and dependency of oncogenic MYCN. These data suggest that MYCN enhancer invasion helps shape transcriptional amplification of the neuroblastoma gene expression program to promote tumorigenesis.

overall_design: ChIP-Seq in Th-MYCN genetically engineered mouse model of neuroblastoma of tumors, celiac ganglia (CG), and superior cervical ganglia (SCG). ChIPs for H3K27ac and MYCN

first: Rhamy

last: Zeid

first: Charles

middle: Y

last: Lin

first: Louis

last: Chesler

first: Evon

last: Poon


title: Gender differences of gene expression profiles in brain cardiovascular centers of SHRs [hypothalamus]

first: Sabine

last: Gouraud

summary: Arterial pressure (AP) is lower in pre-menopausal women than in men of similar age. Pre-menopausal women exhibit a lower sympathetic outflow and a greater baroreceptor reflex, however molecular mechanisms for the gender differences of AP regulation are still not well understood. Since the hypothalmus is strongly functionnaly connected to the nucleus tractus solitarius (NTS), a pivotal region of the medulla oblongata for regulating the set-point of AP, we hypothesized that a different neuronal functions at the level of hypothalamus between men and women could contribute to the gender difference in cardiovascular homeostasis.  Since females Spontaneous Hypertensive Rats (SHRs) clearly exhibit lower AP levels than their male counterparts at similar age, we investigated whether the hypothalamus of SHRs exhibit gender differences in gene expression by using microarray technique.

overall_design: 2 groups of adult SHR rats (9 weeks old) were formed : males (n=5) and females(n=5). Hypothalamus was dissected from each rat and total RNAs were extracted. For each group, 100ng of total RNA from each rats were pooled together before being processed in the microarray experiment.

first: Makiko

last: Onishi

first: Ko

last: Yamanaka

first: Yasunori

last: Miyamoto

first: Hidefumi

last: Waki


title: Mechanisms of the innate defense regulator peptide 1002 anti-inflammatory activity in a sterile inflammation mouse model

city: Vancouver

first: Erin

last: Gill

state: British Columbia

address: 2259 Lower Mall

country: Canada

institute: University of British Columbia

summary: Innate defense regulator (IDR) peptide-1002 is a synthetic host defense peptide derivative with strong anti-inflammatory properties.To investigate the anti-inflammatory mechanisms of IDR-1002 in vivo, the phorbol 12-myristate 13-acetate (PMA)-induced mouse ear inflammation model was used. Topical IDR-1002 treatment successfully dampened PMA-induced ear edema, proinflammatory cytokine production, reactive oxygen and nitrogen species release, and neutrophil recruitment in the ears of CD1 mice. Advanced RNA-Seq analysis on the mouse ear transcriptome revealed that IDR-1002 reduced sterile inflammation by suppressing the expression of transmembrane G-protein coupled receptors (Class A/1 rhodopsin-like), including receptors for chemokines, prostaglandins, histamine, platelet activating factor and anaphylatoxin. IDR-1002 also dampened IFNγ response and repressed the Interferon regulatory factor (Irf) 8-regulated network that controls central inflammatory pathways.

overall_design: RNA-Seq profiling of gene expression in mouse ear tissue exposed to three treatment conditions

first: Bing

first: Amy

middle: H

last: Lee

first: Robert

last: Hancock


title: Temporal response of Mycobacterium tuberculosis H37Rv to vitamin C exposure.

city: Bangalore

first: Genotypic

last: technology

state: Karnataka

address: 259, Apoorva 4th cross,80 feet Road,RMV 2ND STAGE

country: India

institute: Genotypic Technology

summary: Transcriptional profiling of M.tuberculosis to 10 mM vitamin C at 0.25, 0.5, 1, 2 , 4, 8 and 24 h compared to gene expression profile of untreated M. tuberculosis culture (0 h).

overall_design: Organism :Mycobacterium tuberculosis, Agilent's M. tb Gene Expression  8x15k Array (AMADID: 026323) designed by Genotypic Technology Private Limited

first: Kriti

last: Sikri

first: Sakshi

middle: D

last: Batra

first: Jaya

middle: S

last: Tyagi


title: Evolution of Barrett’s Esophagus through space and time at single-crypt and whole-biopsy levels [Batch 7 of 7]

city: Lyon cedex 08

first: Pierre

last: Martinez

address: 28 rue Laennec

institute: Cancer Research Centre of Lyon

summary:
Barrett’s esophagus is a condition in which the squamous epithelium of the esophagus is replaced with a metaplastic crypt-structured epithelium and progresses to esophageal adenocarcinoma in a minority of cases. It is an ideal human system in which to study the evolutionary dynamics of progression. Previous results suggested that there is a low mutation rate in Barrett’s, based on measures of whole biopsies. However, it was unclear if this was due to a low mutation rate in crypts or a high mutation rate in crypts with a low clonal expansion rate. We used phylogenetic analyses to study copy-number alterations in the epithelium of 67 whole biopsies and 291 single crypts from those biopsies in 8 individuals with Barrett’s esophagus, including 4 cancer progressors.  While crypts contained some private mutations, the mutation load and inferred mutation rate in crypts were similar to those in whole biopsies; the latter thus being a reasonable substrate for studies of the mutational process. Diversity between biopsies and within biopsies were strongly correlated. Mutations were more frequent near the gastro-esophageal junction and genomic instability appeared to precede genome doubling, leading to clonal expansions in two cases. This suggests that progression is driven by rare, carcinogenic alterations of large effect, rather than the gradual accumulation of many alterations of small effect, and explains why most patients with Barrett’s esophagus do not progress to esophageal adenocarcinoma. These results shed light on the evolutionary dynamics underlying neoplastic progression in Barrett’s esophagus.
Batch 7 of 7

overall_design: 96 samples per batch (note: 63 samples in batch 7 of 7), taken either from a crypt or a whole biopsy. Paired normal samples are included for each patient in every batch. Normal samples with 5ng of DNA were used to normalise crypt samples, those with 200ng of DNA to normalise whole-biopsy samples.

first: Thomas

middle: G

last: Paulson

first: Carissa

last: Sanchez


title: Gender differences of gene expression profiles in brain cardiovascular centers of SHRs [nucleus of solitarii tract]

summary: Arterial pressure (AP) is lower in pre-menopausal women than in men of similar age. Pre-menopausal women exhibit a lower sympathetic outflow and a greater baroreceptor reflex, however molecular mechanisms for the gender differences of AP regulation are still not well understood. Since the nucleus tractus solitarius (NTS), a pivotal region of the medulla oblongata for regulating the set-point of AP, is strongly associated with the AP level, we hypothesized that a different neuronal functions at the level of NTS between men and women could contribute to the gender difference in cardiovascular homeostasis.  Since females Spontaneous Hypertensive Rats (SHRs) clearly exhibit lower AP levels than their male counterparts at similar age, we investigated whether the NTS of SHRs exhibit gender differences in gene expression by using microarray technique.

overall_design: 2 groups of adult SHR rats (9 weeks old) were formed : males (n=5) and females(n=5). NTS was dissected and total RNAs were extracted from each rat. For each group, 100ng of total RNAs from each rats were pooled together before being processed in the microarray experiment.


title: Evolution of Barrett’s Esophagus through space and time at single-crypt and whole-biopsy levels [Batch 6 of 7]

summary:
Barrett’s esophagus is a condition in which the squamous epithelium of the esophagus is replaced with a metaplastic crypt-structured epithelium and progresses to esophageal adenocarcinoma in a minority of cases. It is an ideal human system in which to study the evolutionary dynamics of progression. Previous results suggested that there is a low mutation rate in Barrett’s, based on measures of whole biopsies. However, it was unclear if this was due to a low mutation rate in crypts or a high mutation rate in crypts with a low clonal expansion rate. We used phylogenetic analyses to study copy-number alterations in the epithelium of 67 whole biopsies and 291 single crypts from those biopsies in 8 individuals with Barrett’s esophagus, including 4 cancer progressors.  While crypts contained some private mutations, the mutation load and inferred mutation rate in crypts were similar to those in whole biopsies; the latter thus being a reasonable substrate for studies of the mutational process. Diversity between biopsies and within biopsies were strongly correlated. Mutations were more frequent near the gastro-esophageal junction and genomic instability appeared to precede genome doubling, leading to clonal expansions in two cases. This suggests that progression is driven by rare, carcinogenic alterations of large effect, rather than the gradual accumulation of many alterations of small effect, and explains why most patients with Barrett’s esophagus do not progress to esophageal adenocarcinoma. These results shed light on the evolutionary dynamics underlying neoplastic progression in Barrett’s esophagus.
Batch 6 of 7

overall_design: 96 samples per batch, taken either from a crypt or a whole biopsy. Paired normal samples are included for each patient in every batch. Normal samples with 5ng of DNA were used to normalise crypt samples, those with 200ng of DNA to normalise whole-biopsy samples.


title: Interplay between genome topology and gene regulation during somatic cell reprogramming [ATAC-seq]

last: Graf

summary: Studies of genome topology, chromatin state and transcriptome dynamics during highly-efficient somatic cell reprogramming

overall_design: Chromatin accessibility profiling of B cells reprogramming to pluripotent cells and (induced) pluripotent stem cells using ATAC-Seq

first: Ralph

last: Stadhouders

first: Enrique

last: Vidal

first: Antonio

last: Gomez


title: Interplay between genome topology and gene regulation during somatic cell reprogramming

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

overall_design: Refer to individual Series


title: MYC favors the onset of tumorigenesis by inducing epigenetic reprogramming of mammary epithelial cells towards a stem cell-like state [microarray]

city: Trento

first: CIBIO

last: Zippo

state: Not Applicable

address: via sommarive 9

department: Cellular, Computation and Integrative Biology (CIBIO)

institute: University of Trento

summary: We address the molecular mechanisms through which MYC promotes loss of cell identity and acquisition of stem cell-like traits, favouring the onset of tumorigenesis, by performing gene expression profile analyses in a transition from WT IMEC, IMEC over-expressing MYC and mammospeheres formed from IMEC-MYC (named M2). We then investigated the global gene expression profile of the fraction of cells hyper-activating the WNT pathway in M2 spheres, compared to the ones with low activation

overall_design: Total RNAs were extracted from log-phase WT IMEC, IMEC-MYC, M2 spheres and GFP+ and GFP- cells sorted from M2 spheres

first: Luca

last: Fagnocchi

first: Vittoria

last: Poli

first: Alessio


title: Evolution of Barrett’s Esophagus through space and time at single-crypt and whole-biopsy levels [Batch 5 of 7]

summary:
Barrett’s esophagus is a condition in which the squamous epithelium of the esophagus is replaced with a metaplastic crypt-structured epithelium and progresses to esophageal adenocarcinoma in a minority of cases. It is an ideal human system in which to study the evolutionary dynamics of progression. Previous results suggested that there is a low mutation rate in Barrett’s, based on measures of whole biopsies. However, it was unclear if this was due to a low mutation rate in crypts or a high mutation rate in crypts with a low clonal expansion rate. We used phylogenetic analyses to study copy-number alterations in the epithelium of 67 whole biopsies and 291 single crypts from those biopsies in 8 individuals with Barrett’s esophagus, including 4 cancer progressors.  While crypts contained some private mutations, the mutation load and inferred mutation rate in crypts were similar to those in whole biopsies; the latter thus being a reasonable substrate for studies of the mutational process. Diversity between biopsies and within biopsies were strongly correlated. Mutations were more frequent near the gastro-esophageal junction and genomic instability appeared to precede genome doubling, leading to clonal expansions in two cases. This suggests that progression is driven by rare, carcinogenic alterations of large effect, rather than the gradual accumulation of many alterations of small effect, and explains why most patients with Barrett’s esophagus do not progress to esophageal adenocarcinoma. These results shed light on the evolutionary dynamics underlying neoplastic progression in Barrett’s esophagus.
Batch 5 of 7


title: Evolution of Barrett’s Esophagus through space and time at single-crypt and whole-biopsy levels [Batch 3 of 7]

summary:
Barrett’s esophagus is a condition in which the squamous epithelium of the esophagus is replaced with a metaplastic crypt-structured epithelium and progresses to esophageal adenocarcinoma in a minority of cases. It is an ideal human system in which to study the evolutionary dynamics of progression. Previous results suggested that there is a low mutation rate in Barrett’s, based on measures of whole biopsies. However, it was unclear if this was due to a low mutation rate in crypts or a high mutation rate in crypts with a low clonal expansion rate. We used phylogenetic analyses to study copy-number alterations in the epithelium of 67 whole biopsies and 291 single crypts from those biopsies in 8 individuals with Barrett’s esophagus, including 4 cancer progressors.  While crypts contained some private mutations, the mutation load and inferred mutation rate in crypts were similar to those in whole biopsies; the latter thus being a reasonable substrate for studies of the mutational process. Diversity between biopsies and within biopsies were strongly correlated. Mutations were more frequent near the gastro-esophageal junction and genomic instability appeared to precede genome doubling, leading to clonal expansions in two cases. This suggests that progression is driven by rare, carcinogenic alterations of large effect, rather than the gradual accumulation of many alterations of small effect, and explains why most patients with Barrett’s esophagus do not progress to esophageal adenocarcinoma. These results shed light on the evolutionary dynamics underlying neoplastic progression in Barrett’s esophagus.
Batch 3 of 7


title: MYC favors the onset of tumorigenesis by inducing epigenetic reprogramming of mammary epithelial cells towards a stem cell-like state


title: Evolution of Barrett’s Esophagus through space and time at single-crypt and whole-biopsy levels [Batch 4 of 7]

summary:
Barrett’s esophagus is a condition in which the squamous epithelium of the esophagus is replaced with a metaplastic crypt-structured epithelium and progresses to esophageal adenocarcinoma in a minority of cases. It is an ideal human system in which to study the evolutionary dynamics of progression. Previous results suggested that there is a low mutation rate in Barrett’s, based on measures of whole biopsies. However, it was unclear if this was due to a low mutation rate in crypts or a high mutation rate in crypts with a low clonal expansion rate. We used phylogenetic analyses to study copy-number alterations in the epithelium of 67 whole biopsies and 291 single crypts from those biopsies in 8 individuals with Barrett’s esophagus, including 4 cancer progressors.  While crypts contained some private mutations, the mutation load and inferred mutation rate in crypts were similar to those in whole biopsies; the latter thus being a reasonable substrate for studies of the mutational process. Diversity between biopsies and within biopsies were strongly correlated. Mutations were more frequent near the gastro-esophageal junction and genomic instability appeared to precede genome doubling, leading to clonal expansions in two cases. This suggests that progression is driven by rare, carcinogenic alterations of large effect, rather than the gradual accumulation of many alterations of small effect, and explains why most patients with Barrett’s esophagus do not progress to esophageal adenocarcinoma. These results shed light on the evolutionary dynamics underlying neoplastic progression in Barrett’s esophagus.
Batch 4 of 7


title: Evolution of Barrett’s Esophagus through space and time at single-crypt and whole-biopsy levels [Batch 1 of 7]

summary:
Barrett’s esophagus is a condition in which the squamous epithelium of the esophagus is replaced with a metaplastic crypt-structured epithelium and progresses to esophageal adenocarcinoma in a minority of cases. It is an ideal human system in which to study the evolutionary dynamics of progression. Previous results suggested that there is a low mutation rate in Barrett’s, based on measures of whole biopsies. However, it was unclear if this was due to a low mutation rate in crypts or a high mutation rate in crypts with a low clonal expansion rate. We used phylogenetic analyses to study copy-number alterations in the epithelium of 67 whole biopsies and 291 single crypts from those biopsies in 8 individuals with Barrett’s esophagus, including 4 cancer progressors.  While crypts contained some private mutations, the mutation load and inferred mutation rate in crypts were similar to those in whole biopsies; the latter thus being a reasonable substrate for studies of the mutational process. Diversity between biopsies and within biopsies were strongly correlated. Mutations were more frequent near the gastro-esophageal junction and genomic instability appeared to precede genome doubling, leading to clonal expansions in two cases. This suggests that progression is driven by rare, carcinogenic alterations of large effect, rather than the gradual accumulation of many alterations of small effect, and explains why most patients with Barrett’s esophagus do not progress to esophageal adenocarcinoma. These results shed light on the evolutionary dynamics underlying neoplastic progression in Barrett’s esophagus.
Batch 1 of 7

overall_design: 96 samples per batch, taken either from a crypt or a whole biopsy. Paired normal samples are included for each patient in every batch. Normal samples with 5ng of DNA were used to normalize crypt samples, those with 200ng of DNA to normalize whole-biopsy samples.


title: Evolution of Barrett’s Esophagus through space and time at single-crypt and whole-biopsy levels [Batch 2 of 7]

summary:
Barrett’s esophagus is a condition in which the squamous epithelium of the esophagus is replaced with a metaplastic crypt-structured epithelium and progresses to esophageal adenocarcinoma in a minority of cases. It is an ideal human system in which to study the evolutionary dynamics of progression. Previous results suggested that there is a low mutation rate in Barrett’s, based on measures of whole biopsies. However, it was unclear if this was due to a low mutation rate in crypts or a high mutation rate in crypts with a low clonal expansion rate. We used phylogenetic analyses to study copy-number alterations in the epithelium of 67 whole biopsies and 291 single crypts from those biopsies in 8 individuals with Barrett’s esophagus, including 4 cancer progressors.  While crypts contained some private mutations, the mutation load and inferred mutation rate in crypts were similar to those in whole biopsies; the latter thus being a reasonable substrate for studies of the mutational process. Diversity between biopsies and within biopsies were strongly correlated. Mutations were more frequent near the gastro-esophageal junction and genomic instability appeared to precede genome doubling, leading to clonal expansions in two cases. This suggests that progression is driven by rare, carcinogenic alterations of large effect, rather than the gradual accumulation of many alterations of small effect, and explains why most patients with Barrett’s esophagus do not progress to esophageal adenocarcinoma. These results shed light on the evolutionary dynamics underlying neoplastic progression in Barrett’s esophagus.
Batch 2 of 7


title: Expression profiling of circular RNAs in human islet samples

status: Public on Jan 29 2018

city: Lausanne

first: Romano

last: Regazzi

address: Rue du Bugnon 9

department: Department of fundamental neurosciences

institute: University of Lausanne

summary:
There is already strong evidence indicating that different types of non-coding RNAs, including microRNAs (miRNAs) and long non-coding RNAs, are key players in the regulation of β-cell functions and in the development of diabetes.
However, the role of the newly discovered class of circular RNAs remains to be elucidated. We therefore analysed circular RNA expression in human islet samples.

overall_design: Human islet were incubated for 48 h in culture medium (CMRL medium supplemented with 10 % FCS, 100 U/ml Penicillin, 100 µg/ml Streptovidin, 2 mM L-Glutamine and 250 µM Hepes)

first: Lisa

last: Stoll


title: Effects of circHIPK3 silencing on mRNA expression

summary:
circHIPK3 silencing impairs ß-cell functions leading to a decrease in insulin secretion, proliferation, and survival.
Therefore, we wanted to identify the mode of action of circHIPK3 by measuring gene expression changes upon circHIPK3 silencing in MIN6B1 cells.

overall_design: MIN6B1 cells were incubated in culture medium (DMEM Glutamax medium supplemented with supplemented with 15 % fetal calf serum, 50 U/ml penicillin, 50 µg/ml streptomycin, and 70 µM β-mercaptoethanol) and cells were transfected with siGFP (Control) or circHIPK3 siRNA for 48 h with Lipofecatime RNAiMax.


title: Enhancer invasion shapes MYCN dependent transcriptional amplification in neuroblastoma [ChIP-seq]

first: James

last: Bradner

summary: In neuroblastoma, amplification of the oncogenic basic helix-loop-helix (bHLH) transcription factor (TF) MYCN is the defining prognosticator of high-risk disease, occurs in one-third of neuroblastoma, and drastically reduces overall survival rates. As a proto-oncogene, targeted MYCN overexpression in peripheral neural crest is sufficient to initiate disease in mouse models. In MYCN amplified neuroblastoma, elevated expression of the factor is crucial to maintain tumor stemness and is associated with increased proliferation and aberrant cell cycle progression, as these tumors lack the ability to arrest in G1 in response to irradiation. MYCN down-regulation broadly reverses these oncogenic phenotypes in a variety of neuroblastoma models and recent thereapeutic strategies to indirectly target MYCN production or protein stability have reduced tumor growth in vivo. These observations motivate an investigation of MYCN binding in MYCN amplified tumors as it remains fundamentally unclear how elevated levels of the factor occupy the genome and alter transcriptional programs in neuroblastoma. Here we present the first dynamic chromatin and transcriptional landscape of direct MYCN perturbation in neuroblastoma. We find that at oncogenic levels, MYCN associates with E-box (CANNTG) binding motifs in an affinity dependent manner across most active cis-regulatory promoters and enhancers. MYCN shutdown globally reduces histone acetylation and transcription, consistent with prior descriptions of MYC proteins as non-linear amplifiers of gene expression. We establish that MYCN load at the promoter and proximal enhancers predicts transcriptional responsiveness to MYCN shutdown and that MYCN enhancer binding occurs prominently at the most strongly occupied and down-regulated genes, suggesting a role for these tissue specific elements in predicating MYCN responsive “target” genes. At these invaded enhancers, we identify the lineage specific bHLH TWIST1 as a key collaborator and dependency of oncogenic MYCN. These data suggest that MYCN enhancer invasion helps shape transcriptional amplification of the neuroblastoma gene expression program to promote tumorigenesis.

overall_design: ChIP-Seq in SHEP21, BE2C, KELLY, and NGP neuroblastoma cell lines for H3K27ac, H3K27me3, H3K4me3, RNA PolII, MYCN, BRD4, or TWIST1

first: Rhamy

last: Zeid

first: Charles

last: Lin


title: Evolution of Barrett's Esophagus through space and time at single-crypt and whole-biopsy levels


title: Breaching self-tolerance to Alu duplex RNA underlies MDA5-mediated inflammation [WT HEK293T and mutant MDA5]

first: Sun

last: Hur

address: 3 Blackfan Circle, Boston, MA, 02115

department: Program in Cellular and Molecular Medicine (PCMM)

institute: Boston Children's Hospital

summary: Aberrant activation of innate immune receptors can cause a spectrum of immune disorders, such as Aicardi-Goutières syndrome (AGS). One such receptor is MDA5, a viral double-stranded RNA (dsRNA) sensor that induces antiviral immune response. We here demonstrate that constitutive activation of MDA5 in AGS results from the loss of tolerance to cellular dsRNAs formed by Alu retroelements. While wild-type MDA5 cannot efficiently recognize Alu-dsRNA because its filament formation on dsRNA is impaired by the imperfect duplex structure, AGS-variants of MDA5 display reduced sensitivity to duplex structural irregularities, assembling signaling-competent filaments on Alu-dsRNA. Moreover, we identified an unexpected role of RNA-rich cellular environment in suppressing aberrant MDA5 oligomerization, highlighting context-dependence of self vs. non-self discrimination. Overall, our work demonstrates that the increased efficiency of MDA5 to recognize dsRNA comes at a cost of self-recognition, and implicates a unique role of Alu RNAs as virus-like elements that shape the primate immune system.

overall_design: cytoplasmic RNA profiles of a footprinting assay were generated in duplicate using Illumina Miseq V2.

first: Sadeem

last: Ahmad

first: Xin

last: Mu

first: Fei

last: Yang

first: Emily

last: Greenwald

first: Ji

middle: W

last: Park

first: Etai

last: Jacob

first: Cheng-Zhong

last: Zhang


title: Breaching self-tolerance to Alu duplex RNA underlies MDA5-mediated inflammation


title: Natural regulatory mutations elevate fetal globin via disruption of BCL11A or ZBTB7A binding

city: Sydney

first: Merlin

last: Crossley

state: NSW

address: UNSW

department: BABS

country: Australia

institute: University of New South Wales

summary: B-hemoglobinopathies such as Sickle Cell Disease (SCD) and b-thalassemia result from mutations in the adult b-globin gene. Reactivating the developmentally silenced fetal g-globin gene is a therapeutic goal for treating SCD and b-thalassemia1. Some forms of Hereditary Persistence of Fetal Hemoglobin (HPFH), a rare benign condition in which individuals express g‑globin throughout adulthood, are caused by point mutations in the g‑globin gene promoter at regions residing ~115 and 200 base pairs upstream of the transcription start site. Here we show that the major fetal globin repressors BCL11A and ZBTB7A/LRF directly bind to the -115 and ‑200 sites, respectively. Furthermore, introduction of naturally occurring HPFH mutations into erythroid cells by CRISPR/Cas9 disrupts repressor binding and raises g‑globin expression. These findings resolve the mystery surrounding how these HPFH mutations operate and demonstrate that BCL11A and ZBTB7A/LRF are major direct repressors of the fetal globin gene.

overall_design: ChIP-seq experiments were performed in two different cell lines, HUDEP-2 and K562 against BCL11A and ZBTB7A. Input samples were used as control.

first: Gabriella

last: Martyn

first: Beeke

last: Wienert

first: Lu

first: Manan

last: Shah

first: Laura

middle: J

last: Norton

first: Jon

last: Burdach

first: Ryo

last: Kurita

first: Yukio

last: Nakamura

first: Richard

last: Pearson

first: Alister

last: Funnell

first: Kate

last: Quinlan


title: Bacterial artificial chromosomes establish replication timing and sub-nuclear compartment de novo as extra-chromosomal vectors

city: Tallahassee

first: David

middle: M

last: Gilbert

state: FL

address: 319 Stadium Drive

institute: Florida State University


title: Breaching self-tolerance to Alu duplex RNA underlies MDA5-mediated inflammation [ADAR1KO HEK293T and WT MDA5]

first: Sun

last: Hur

first: Sadeem

last: Ahmad

first: Xin

last: Mu

first: Fei

last: Yang

first: Emily


title: ZBTB2 as a novel reader for unmethylated DNA

city: Zürich

first: Ino

last: Karemaker

address: Winterthurerstrasse 190

institute: University of Zurich


title: ZBTB2 as a novel reader for unmethylated DNA [ChIP-seq]

summary: DNA methylation is a dynamic epigenetic modification that plays a key role in various cellular processes. Proteins that bind to DNA depending on its methylation status are thought to play an important role in DNA methylation-mediated gene expression. Using a variety of genomics and proteomics approaches, we identified ZBTB2 as a novel reader of unmethylated DNA. ZBTB2, which forms a complex with ZBTB25 and ZNF639, preferentially binds at CpG island promoters in mouse embryonic stem cells, from where it regulates genes that are involved in the exit from pluripotency. Binding of ZBTB2 to target genes is mostly associated with gene activation. Furthermore, ZBTB2 is intricately interwoven with DNA methylation, as we found not only that its binding to DNA is methylation-sensitive, but also that ZBTB2 regulates the turnover of methylated DNA. Summarising, we propose that ZBTB2 is a DNA methylation-sensitive transcription factor that is involved in cellular differentiation.

overall_design: ChIP-seq samples of ZBTB2 in ESCs (in two different culture conditions) and NPCs


title: Suckling Piglets; Fructooligosaccharides

city: Wageningen

first: Dirkjan

last: Schokker

address: Droevendaalsesteeg 1

department: Wageningen Livestock Research

institute: Wageningen UR

summary: Emerging knowledge shows the importance of early life events in programming the intestinal mucosal immune system and development of the intestinal barrier function. These processes depend heavily on close interactions between gut microbiota and host cells in the intestinal mucosa. In turn, development of the intestinal microbiota is largely dependent on available nutrients and substrates required for the specific microbial community structures to expand. It is currently not known what the specificities are of intestinal microbial community structures in relation to the programming of the intestinal mucosal immune system and development of the intestinal barrier function. The objective of the present study was to investigate the effect of a nutritional intervention on intestinal development of suckling piglets by daily oral administration of fructooligosaccharides (FOS) over a period of 12 days. At the microbiota community level a clear “bifidogenic” effect of the FOS administration was observed in colon digesta at day 14. The former, however, did not translate into significant changes of local gene expression in the colonic mucosa. In the jejunum, significant changes were observed for microbiota composition at day 14, and microbiota diversity at day 25. In addition, significant differentially expressed gene sets in mucosal tissues of jejunum were identified at both days 14 and 25 of age. At the age of 14 days, lower activity of cell cycle-related processes and a higher activity of extracellular matrix processes were observed in jejunal scrapings of piglets supplemented with FOS compared to control piglets. At day 25, lower activity of immune-related processes in jejunal tissue were seen in piglets supplemented with FOS. Histological parameters, villi height and crypt depth, were significantly different at day 25 between the experimental and control group, where piglets supplemented with FOS had higher villi and deeper crypts. We conclude that oral FOS administration during the suckling period of piglets has significant bifidogenic effects on the microbiota in the colon and on gene expression in jejunal mucosa scrapings. We hypothesize that FOS supplementation of suckling piglets results in a higher butyrate production in the colon due to the increase in bifidobacteria and lactobacilli in the hindgut. We further speculate that a higher butyrate production in colonic digesta relates to changes in gene expression in the jejunum by thus far unknown mechanisms.

overall_design: At day 2, 8 control piglets were sacrificed  to obtain the jejunum scrapings. Whereas at days 14 and 25, 16 piglets were sacrificed (8 controls and 8 piglets that received FOS) to obtain the jejunum scrapings. At day 14, we also extracted colon tissue from 8 controls and 8 piglets that received FOS. After quality control in total 51 samples were left (37 jejunum and 14 colon samples).

first: Stephanie

last: Vastenhouw


title: DNA microarray analysis in hippocampus of Wild-type (WT), AtrxΔE2, and TMPyP4-treated AtrxΔE2 mice.

status: Public on Jan 28 2018

city: Kamakura

first: Satoshi

last: Kondo

state: Kanagawa

address: Tebiro 6-10-1

department: New Projects Development Division

institute: Toray Industries,Inc.

summary: TMPyP4 treatment antagonizes cognitive deficits seen in ATR-X model (AtrxΔE2) mice. Our findings suggest a potential therapeutic strategy to target G-quadruplexes and decrease cognitive impairment associated with ATR-X syndrome.

overall_design: We administered TMPyP4 (30mg/kg, i.p.) to AtrxΔE2 mice twice weekly between post-natal day 30 to 90. (n = 5 mice per group per pool)

first: Norifumi

last: Shioda

first: Kohji

last: Fukunaga


title: RNA-directed DNA methylation involves co-transcriptional small RNA-guided slicing of Pol V transcripts in Arabidopsis

city: Haining

first: Wanlu

last: Liu

state: Zhejiang

address: 718 East Haizhou Rd.,

department: Zhejiang University - University of Edinburgh Institute

institute: Zhejiang University

summary: Small RNAs regulate chromatin modifications such as DNA methylation and gene silencing across eukaryotic genomes. In plants, RNA-directed DNA methylation (RdDM) requires 24-nucleotide (nt) small RNAs (siRNAs) that bind ARGONAUTE4 (AGO4) and target genomic regions for silencing. It also requires non-coding RNAs transcribed by RNA POLYMERASE V (Pol V), although their function is largely unknown. We utilized a modified global nuclear run-on (GRO) protocol followed by deep sequencing to capture Pol V nascent transcripts genome-wide. We uncovered unique characteristics of Pol V RNAs, including a uracil (U) common at position 10. This uracil was complementary to the 5’ adenine found in many AGO4-bound 24-nt siRNAs and was eliminated in an siRNA-deficient mutant. This indicates co-transcriptional slicing of Pol V transcripts by AGO4-bound siRNAs. Pol V transcript slicing also required the elongation factor SPT5L. These results highlight a novel step in RNA-mediated transcriptional gene silencing, which is a conserved process in many eukaryotes.

overall_design: 24 GROseq and 4 ChIPseq samples.


title: Non-canonical DNA structures at eukaryotic centromeres can resolve the CENP-B paradox

city: Seattle

first: Jorja

last: Henikoff

address: 1100 Fairview AV N, A1-162

institute: Fred Hutchinson Cancer Research Center

summary: Epigenetic mechanisms are thought to specify the identity of centromeres, yet the degree to which DNA sequence contributes to centromere determination remains unresolved. For example, the conserved CENP-B protein binds the 17-bp âCENP-B boxâ motif found in Î±-satellite arrays at most human centromeres. Although CENP-B is required for artificial centromere function, it is non-essential and some species lack CENP-B boxes entirely. To address this âCENP-B paradox,â we determined the distribution of CENP-B boxes in primates and directly demonstrated the absence of CENP-B at Old World Monkey centromeres. We found extensive interspecific variation in Î±-satellite arrays including abundance of <10-bp dyad symmetries and of non-B-form DNA structures. We confirmed the presence of non-canonical DNA at human centromeres and neocentromeres and detected similar structures at mouse, chicken and yeast centromeres. To resolve the CENP-B paradox, we propose that CENP-B enhances non-canonical DNA formation, thus providing a general basis for centromere specification.

sample_organism:
Chlorocebus aethiops
Homo sapiens

platform_organism:
Chlorocebus aethiops
Homo sapiens

overall_design: We used Cleavage under targets and Release using nuclease (Cut-and-Run), a chromatin profiling strategy in which antibody-targeted controlled cleavage by micrococcal nuclease releases specific protein-DNA complexes into the supernatant for paired-end DNA sequencing.

first: Sivakanthan

last: Kasinathan

first: Steven