title: Longitudinal Transcriptional Response of Endothelial Cells to Shear Stress

city: La Jolla

first: Shankar

last: Subramaniam

address: 9500 Gilman Dr MC 0412

department: Bioengineering

institute: University of California, San Diego

summary: The goal of this study was to find longitudinal transcriptional response of Human Umbilical Vein Endothelial Cells (HUVECs) to pulsatile shear (PS) and oscillatory shear (OS). PS is associated with an atheroprotective endothelial phenotype, while OS is associated with an atheroprone endothelial phenotype. Using RNASeq method (single-ended 50-bp sequencing on Illumina Hi-seq 2000 instrument), we measured the transcriptional response at 10 time-points (1, 2, 3, 4, 6, 9, 12, 16, 20, 24 hr) under PS and OS conditions. Low flow scenario was used as static condition. Two replicates were used for each condition/time-point. Results: Through combining the temporal data on differentially expressed transcription factors and their targets with existing knowledge on relevant functional pathways, we infer the causal relationships between disparate endothelial functions through common transcriptional regulation mechanisms. Our study presents the first comprehensive temporally longitudinal experimental study and mechanistic model of shear stress response. By comparing the relative endothelial expressions of genes between OS and PS, we provide novel insights and an integrated perspective into endothelial cell function in response to differential shear.

overall_design: HUVECs were cultured in medium M199 (Gibco) supplemented with 15% FBS (Omega), 3 ng/mL β-EC growth factor (Sigma), 4 U/mL heparin (Sigma), and 100 U/mL penicillin-streptomycin. Pulsatile shear ﬂow (PS) or oscillatory shear ﬂow (OS) was applied to ECs with a shear stress of 12 ± 5 dyn/cm2 or 0.5 ± 5 dyn/cm2, respectively. Samples for RNA sequencing analysis were collected at 1, 2, 3, 4, 6, 9, 12, 16, 20, and 24 hours after exposure to shear. Endothelial cells (ECs) were also exposed to a “low flow” condition, i.e. 0.5 dyn/cm2 laminar flow, and designated as a “static” flow condition (ST). Samples for RNA sequencing analysis were collected at hour 1 for the ST condition. Two biological replicates were collected for each time point and for each shear condition. Total RNA was extracted from ECs, processed and sequenced on the Illumina HiSeq 2000 50-bp single-end reads platform. More details are provided in Ajami et al. (Systems Biology Analysis of Longitudinal Functional Response of Endothelial Cells to Shear Stress, Proc Natl Acad Sci U S A., 2017).

first: Zhen

last: Chen

first: John

middle: Y

last: Shyy

first: Shu

last: Chien

first: Mano

middle: R

last: Maurya

first: Nassim

last: Ajami

first: Shakti

last: Gupta

first: Phu

last: Nguyen

first: Julie

last: Li


title: Bone marrow derived macrophage vs peritoneal macrophage

status: Public on Oct 13 2017

city: Frederick

first: Luke

middle: Cynlais

last: Davies

state: MARYLAND

address: 1050 Boyles Street

department: Cancer and Inflammation Program

country: USA

institute: National Cancer Institute

type: Expression profiling by high throughput sequencing

summary: Immunometabolism is a rapidly growing field, which has led to greater understanding of innate immune cell functions. Macrophages are at the core of this research: polarized subsets of in vitro-derived cells reportedly utilize select metabolic pathways to maintain their phenotype. However, relevance of these in vitro studies to the in vivo setting is not known, and metabolic requirements are likely dependent on unique physiological and cellular metabolic environments. Here we define the metabolic requirements of peritoneal tissue-resident macrophages, the accessibility of these metabolites to cells in the peritoneum, and we dissect the role of this unique environment in maintaining a crucial macrophage function. We find that the peritoneal cavity is enriched in amino acids, most notably glutamate. Peritoneal tissue-resident macrophages have an extraordinarily large mitochondrial capacity compared with other phagocytes; this is primarily fueled by glutaminolysis, which is additionally required to maintain an extensive respiratory burst. Glutaminolysis fuels the electron transport chain, which is enhanced during tissue-resident macrophage respiratory burst via a switch to dependence of mitochondrial complex-II. This is not dependent on the level of NADPH, but requires p47 maintained NADPH-oxidase activity. Therefore, we propose that tissue-resident macrophages exploit their unique metabolic niche by implementing their glutamine-fueled mitochondrial-rich phenotype to sustain respiratory burst to assault pathogens, showing that cell-specific metabolic underpinning is important for function. Importantly, we also find that glutamine is required for the respiratory burst in human monocytes, which highlights that metabolites are not species-specific and can be the link between cellular mechanism in mouse and man.

overall_design: M-CSF bone marrow derived macrophage (BMDM) (7 days) vs peritoneal tissue-resident macrophage, 2 vs 2 independent samples

middle: C

first: Daniel

middle: W

last: McVicar


title: Regulation of m6A transcripts by the 3ʹ→5ʹ RNA helicase YTHDC2 is essential for a successful meiotic program in the mammalian germline

status: Public on Oct 14 2017

city: Gneveva

first: Ramesh

last: Pillai

address: 30, Quai Ernest-Ansermet

department: Department of Molecular Biology

country: Switzerland

institute: University of Geneva

summary: N6-methyladenosine (m6A) is an essential internal RNA modification that is critical for  gene expression control in most organisms. Proteins with a YTH domain recognize  m6A marks and are mediators of molecular functions like RNA splicing, mRNA decay  and translation control. Here we demonstrate that YTH domain-containing 2 (YTHDC2)  is an m6A reader that is essential for male and female fertility in mice. In mutant males,  mitotic spermatogonia fail to differentiate into meiotic spermatocytes. Analysis identifies  transcripts upregulated in the Yhtdc2 mutant testes to be those that are expressed in  purified spermatogonia, while high-throughput mapping of the m6A transcriptome in  the mouse male germline demonstrates the upregulated transcripts to be those that  are enriched in m6A marks. Our biochemical studies indicate that YTHDC2 is an RNAinduced  ATPase that fuels its 3ʹ→5ʹ RNA helicase activity. Interestingly, we find an  RNA-independent interaction between the 5ʹ→3ʹ exoribonuclease XRN1 and the  Ankyrin repeat of YTHDC2 that is wedged in between the two RecA helicase domains.  Our studies reveal a role for YTHDC2 in modulating the levels of m6A-modified  germline transcripts that is essential for successful meiosis.

sample_organism:
Mus musculus
synthetic construct

platform_organism:
Mus musculus
synthetic construct

overall_design: RNA sequencing was used to identify transcript abundance differences between Ythdc2 mutant and wt mice. The distribution of m6A in testicular transcripts was analyzed by m6A RNA-Seq.  RNA Bind-N-Seq analysis was performed to determine the YTH domain bound sequences.

first: Magdalena

middle: N

last: Wojtas

first: Radha

last: Pandey

first: Mateusz

last: Mendel

first: David

last: Homolka

first: Ravi

last: Sachidanandam

middle: S


title: RNA Seq of C2C12 cells stimulated with Control, Jag1-expressing or Jag1Ndr-expressing cells

city: Stockholm

first: Urban

last: Lendahl

address: Von Eulers vag 3

country: Sweden

institute: Karolinska Insttitutet

summary: RNA sequencing of control or Notch1-expressing mouse cells co-cultured with control, Jag1WT, or Jag1Ndr-expressing human cells. Deep sequencing and bioinformatical separation of mouse and human reads reveals transcripts specifically regulated in mouse receptor-expressing cells.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Mouse C2C12 control and C2C12-FLNotch1, and human HEK-293-Flp-In cells (Hansson et al., 2010): HEK293-Flp control (Flp Ctrl), HEK293-Flp-Jag1WT (Flp Jag1+), HEK293-Flp-Jag1Ndr (Flp Jag1Ndr) were used in this experiment. In one 12-well plate, we seeded 3 wells of mouse C2C12 control cells and 3 wells of C2C12-FLN1 cells, with 3.6x105 cells in 1 mL antibiotic-free medium per well. Cells were allowed to settle for 8 hours. C2C12 control and C2C12-FLN1 cells were transfected with pcDNA5 (1.6 ug/well). All transfections were done using Lipofectamine® 2000 (InvitrogenTM, cat. no. 11668-019) with Opti-MEM® I Reduced Serum Medium (Gibco®, cat. no. 31985-062), according to manufacturer’s instructions. The following day (18 hours post transfection), 3.6x105 cells in 0.5 mL antibiotic-free medium of Flp Ctrl, Flp Jag1+, or Flp Jag1Ndr cells were added. Cells were co-cultured for 6 hours, then lysed in 350 uL per well Buffer RLT (QIAGEN, cat. no. 79216) with 1% 2-Mercaptoethanol (Sigma-Aldrich®, cat. no. M3148) and stored at -80°C until RNA extraction.

first: Emma

last: Andersson

first: Simona

last: Hankeova

first: Indira

middle: V

last: Chivukula

first: Helena

last: Storvall

last: Ramsköld

first: Yat

middle: L

last: Tsoi


title: RNA sequencing of the urethral epithelial progenitor cells

city: Edinburgh

first: GUDMAP

last: Developers

address: Crewe Road

country: United Kingdom

institute: IGMM MRC Human Genetics Unit

summary: We will use micro-dissection to isolate cells from the embryonic mouse cloaca, and perform RNA-Seq analysis to identify molecular markers of the region of the cloaca that will give rise to the urethral epithelium.

overall_design: Total RNA was obtained from LCM dissected tissue originating from the dorsal and ventral regions of the embryonic day 10.5 and 13.5 cloaca. The long terms goal is to develop a transcriptional atlas for the developing genital tubercle and associated urethra

first: Brad

last: Barbazuk

first: Martin

middle: J

last: Cohn

first: Ruth

last: Davenport

first: Brooke

middle: A

last: Armfield

first: Emily

last: Merton


title: RNA Seq of Alagille liver biopsies

summary: Needle biopsies were performed to obtain liver samples from patients for clinical purposes from patients with Alagille syndrome. A small portion was snap frozen and later used for RNA sequencing analysis. Needle biospies from 5 patients with other liver disorders were included as controls.

overall_design: Examination of RNA expression in Alagille patients' liver samples, compared to other control liver samples (with other chronic liver diseases).

first: Daniel


title: Alagille_Nodder

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

overall_design: Refer to individual Series


title: Characterization of changes in enhancer activity and transcription factor binding following pro-inflammatory cytokine treatments in primary mouse hepatocytes

status: Public on Oct 10 2017

city: Bethesda

first: Songjoon

last: Baek

state: MD

address: 41 Library Drive

department: CCR

institute: NCI / NIH

type: Genome binding/occupancy profiling by high throughput sequencing

summary: The cytokines interleukin 1β and 6 (IL-1β, IL-6) mediate the systemic acute and chronic inflammatory response. In liver, they both regulate the secretion of acute-phase proteins which curb infection and tissue damage. Using RNA-seq, we show that these cytokines regulate gene transcription in a multifaceted manner to modulate gene expression, leading both to synergistic and antagonistic expression patterns. By mapping changes in enhancer landscape and transcription factor occupancy following cytokine treatment, we show that synergistic gene expression was achieved by cooperation between cytokine-induced transcription factors on the chromatin template. At a subset of enhancers proximal to synergistically-induced acute-phase genes, IL-1β led to enhancer priming and an increase in STAT3 binding. These effects were mediated by NF-κB and were enhancer-specific rather that influencing the entirety of STAT3 binding. Our findings reveal that STAT3-NF-κB crosstalk at the chromatin template plays a major role in transcriptional regulation during inflammation

overall_design: the enhancer mark H3K27ac and various transcription factors were profiled by ChIP-seq following cytokine treatemnts (treatment regimen and replicates specified in sample title, NT is the control samples)

first: Ido

last: Goldstein

first: Gordon

last: Hager


title: mRNA expression levels in splenic human CD4-positive T cells of mock-infected, HIV-1-infected and pegIFN-treated humanized mice

city: Minato-ku

first: Kei

last: Sato

state: Tokyo

address: 4-6-1 Shiroganedai

department: Department of Microbiology and Immunology

country: Japan

institute: The Institute of Medical Science, The University of Tokyo

summary: Through RNA sequencing and reactome analyses, we report that type I interferon response is elicited in the spleen of HIV-1-infected humanized mice when compared to mock-infected humanized mice.

overall_design: mRNA expression profiles in the splenic human CD4-positive T cells of HIV-1-infected humanized mice at 6 weeks post-infection (n=2), mock-infected humanized mice (n=2) and pegIFN-treated humanized mice (n=2) were generated by RNA sequencing. RNA was extracted using QIAamp RNA Blood Mini kit (Qiagen). RNA sequencing was conducted in Medical & Biological Laboratories, co (Nagoya, Japan).

first: Eri

last: Yamada

first: Shinji

last: Nakaoka

first: Yoshio

last: Koyanagi


title: Pro-inflammatory cytokine treatments in primary mouse hepatocytes


title: RNAseq and ChIPseq on Mouse Embryonic Stem Cells and Sorted Mesendoderm Progenitor Cells [ChIP-seq]

city: Lausanne

first: Frederic

last: Burdet

address: Quartier Sorge - Batiment Amphipole

institute: Swiss Institute of Bioinformatics

summary: We mapped the enhancer and long non-coding transcriptional landscape during mesendoderm specification. Mesendodermal progenitors were sorted from differentiating ESCs according to Eomes expression. Enhancer usage was coordinated with mesendoderm-specific expression of key lineage-determining transcription factors. We demonstrated that many of these enhancers are associated with the expression of lncRNAs.

overall_design: ChIPseq (on H3K4me3 and H3K27Ac) on mESC and sorted mesendoderm progenitors.

first: Michael

last: Alexanian

last: Maric

first: Stephen

middle: P

last: Jenkinson

first: Marco

last: Mina

first: Clayton

middle: E

last: Friedman

first: Ching

last: Ting

first: Rudi

last: Micheletti

first: Isabelle

last: Plaisance

first: Mohamed

last: Nemir

first: Damien

last: Maison

first: Jasmin

last: Kernen

first: Iole

last: Pezzuto

first: Dominic

last: Villeneuve

first: Frédéric

first: Mark

last: Ibberson

last: Leib

first: Nathan

last: Palpant

first: Nouria

last: Hernandez

first: Samir

last: Ounzain

first: Thierry

last: Pedrazzini


title: Characterization of gene expression changes following pro-inflammatory cytokine treatments in primary mouse hepatocytes

first: Songjoon

last: Baek

overall_design: transcriptome evaluated in 6 groups (2 replicates each). NT is the control group

first: Ido

last: Goldstein

first: Gordon

last: Hager


title: BEX1 is an RNA-dependent mediator of cardiomyopathy [RNA-Seq]

city: Cincinnati

first: Tobias

last: Schips

address: 3333 Burnet Ave

institute: Cincinnati Children's Hospital Medical Center

summary: Regulation of mRNA splicing, processing and stability is increasingly recognized as a critical control point in dynamically altering gene expression during stress or disease states. Very little is understood of this process in the heart despite the wide ranging changes in gene expression that occurs during heart failure; a disease of epidemic proportions in the western world. Here we identified BEX1 as a heart-failure-induced gene and identified its function as an mRNA binding protein responsible for enhancing expression of a subset of cardiac disease promoting genes. Modeling this increase in BEX1 that occurs in disease, BEX1 cardiac-specific overexpressing transgenic mice were generated and shown to have worse cardiac disease with stress stimulation while Bex1 gene-deleted mice were protected from insults that would otherwise promote heart failure. Since BEX1 was of unknown molecular function we performed a series of proteomic and interactive screening assays, which identified Bex1 as part of a large ribonucleoprotein processing complex involved in regulating pro-inflammatory mRNA expression in the heart. Specifically, induction of BEX1 augmented the stability and expression of AU-rich element containing mRNAs typically found within pro-inflammatory genes. Thus, BEX1 functions as an mRNA modulatory effector that augments pathology promoting gene expression during heart failure.

overall_design: Analysis of neonatale neonatale rat cardiomyocytes overexpressing Bex1 or beta galactosidase

first: Federica

last: Accornero

first: Jeff

last: Molkentin


title: Genome-wide DNA methylation and expression profiling of MEFS and SKOM induced reprogramming cells after modulation of Cell cycle and Dnmt1 expression

city: Guangzhou

first: Hui

last: Zheng

state: Guangdong

address: 190 Kaiyuan Avenue

country: China

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

type: Methylation profiling by high throughput sequencing


title: Role of Microglial C5aR1 in the Arctic Alzheimer’s Disease Mouse Model

city: Irvine

first: Andrea

middle: J.

last: Tenner

state: California

address: 3205 McGaugh Hall

institute: University of California, Irvine

summary: C5aR1, a receptor for the complement activation proinflammatory fragment, C5a, is primarily expressed on cells of the myeloid lineage, and to a lesser extent on endothelial cells and neurons in brain. Previous work demonstrated C5aR1 antagonist, PMX205, decreased amyloid pathology and suppressed cognitive deficits in Alzheimer Disease (AD) mouse models. In the Arctic AD mouse model, genetic deletion of C5aR1 prevented behavior deficits at 10 months. However, the molecular mechanisms of this protection has not been definitively demonstrated.  To understand the role of microglial C5aR1 in the Arctic AD mouse model, we have taken advantage of the CX3CR1GFP and CCR2RFP reporter mice to distinguish microglia as GFP-positive and infiltrating monocytes as GFP and RFP positive, for subsequent transcriptome analysis on specifically sorted myeloid populations from wild type and AD mouse models. Immunohistochemical analysis of mice aged to 2, 5, 7 and 10 months showed no change in amyloid beta (Ab) deposition in the Arctic C5aR1 knockout (KO) mice relative to that seen in the Arctic mice. Of importance, no CCR2+ monocytes/macrophages were found near the plaques in the Arctic brain with or without C5aR1. RNA-seq analysis on microglia from these mice identified inflammation related genes as differentially expressed, with increased expression in the Arctic mice relative to wildtype and decreased expression in the Arctic/C5aR1KO relative to Arctic. In addition, phagosomal-lysosomal proteins and protein degradation pathways that were increased in the Arctic mice were further increased in the Arctic/C5aR1KO mice. These data are consistent with a microglial polarization state with restricted induction of inflammatory genes and enhancement of clearance pathways.

overall_design: Microglia mRNA profiles of wildtype (WT), C5aR1 knockout (C5aR1KO), Arctic (ARC) and Arctic C5aR1 knockout (ARCKO) mice at 2, 5, 7 and 10-11 month. Duplicates were sequenced for each genotype on Illumina HiSeq 2500 platform.

first: Michael

middle: X

first: Shan

last: Jiang

first: Tracy

last: Cole

first: Shu-Hui

last: Chu

first: Maria

middle: I

last: Fonseca

first: Rick

last: Wetsel

first: Ali

last: Mortazavi


title: Molecular profiling of prelimbic projections to NAc, BLA, and VTA using vTRAP

city: New York

first: Alexander

last: Nectow

state: New York

address: 630 W 168th St., Physicians and Surgeons

department: Medicine

institute: Columbia University

summary: Translational profiling of prelimbic cortical projections to nucleus accumbens, basolateral amygdala, and ventral tegmental area was performed. Studies were performed using AAV5-IV-GFPL10 (in prelimbic cortex) in tandem with CAV2-Cre (in the projection target), which enables rapid, projection-specific translational profiling.

overall_design: Polysome-bound mRNA in each of the projections was immunoprecipitated (IP) and compared to the total (Input) RNA in the surrounding prelimbic cortex.

first: Malavika

last: Murugan

first: Ilana

last: Witten


title: Differentially expressed LncRNAs and mRNA identified by SEQ analysis in colorectal cancer patients

status: Public on Oct 12 2017

city: Shijiazhuang

first: Li

last: Meng

address: 12# Jiankang road

institute: colon tissues

type: Non-coding RNA profiling by high throughput sequencing

summary: LncRNA plays an important role in gene regulation, but its impact on the pathogenesis of colorectal cancer and the biological function of cancer cells is unclear. In this study, we will use the next generation sequencing technique to study the differences of the expression profiles of lncRNA and mRNA in colorectal cancer tissues, analyzing the differentially expressed genes by GO/KEGG enrichment, and predicting the new lncRNAs’ function. Our results revealed that Comparing with the nontumor colorectal tissues, 1019 lncRNAs (512 upregulated, 507 downregulated) and  3221 mRNAs (1606 upregulated, 1615 downregulated) were differentially expressed in tumor colorectal tissues (fold change>2 and P<0.05).

overall_design: lncRNA and mRNA profiles of 10 paired tissues (nontumor and colon cancer tissues)  were generated by deep sequencing, in triplicate, using Illumina.

first: Guiying

last: Wang

first: Meng

last: Li


title: YAP drives proliferation and tumorigenesis by recruiting Mediator to super- enhancer elements (H69)

city: Torino

first: Raffaele

last: Calogero

state: To

address: Via Nizza 52

department: Molecular Biotechnology Center

country: Italy

institute: University of Torino

summary:
The Hippo/YAP signaling pathway is a crucial regulator of tissue growth and stem cell activity. YAP is also a powerful driver of tumor growth and its nuclear accumulation is frequently observed in human cancer.
However, the molecular mechanism employed by YAP to orchestrate transcriptional outputs are undefined. Here, we utilize genomic technologies to demonstrate that YAP occupancy is restricted to a small number of distal regulatory elements highly enriched for superenhancers.
Target genes associated with these elements are selectively sensitive to the loss of YAP in cancer cells. YAP directs recruitment of the Mediator complex to activate transcriptional regulation from these enhancers without affecting chromatin organization
We also provide in vivo genetic evidence that loss of Mediator rescues YAP-driven cancer cell proliferation and organ overgrowth. Our data provide a molecular mechanism behind YAP-driven tumorigenesis, and highlights transcriptional control as a potential therapeutic strategy for YAP-driven cancers.

overall_design: A total of four ChIP-Seq samples are available. Two directed against YAP using two different antibodies (Santa Cruz and Avruch), one directed against TEAD1, an unrelated IgG for background noise definition

first: Giorgio

middle: G

last: Galli

first: Matteo

last: Carrara

first: Christian

last: Valdes-Quezada

first: Basanta

last: Gurung

first: Brian

last: Pepe-Mooney

first: Geert

last: Geeven

first: Wei-Chien

last: Yuan

first: Wouter

last: de Laat

first: Fernando

middle: D

last: Camargo


title: Sympathetic neuron-associated macrophages contribute to obesity by importing and metabolizing norepinephrine

status: Public on Oct 09 2017

first: Christopher

last: Glass

state: CA

address: 9500 Gilman Dr.

department: CMM

summary: The cellular mechanism(s) linking macrophages to norepinephrine (NE)-mediated regulation of thermogenesis has been a topic of debate. Here, we identify sympathetic neuron-associated macrophages (SAMs) as a population of macrophages that mediate clearance of NE via expression of Slc6a2, an NE transporter, and monoamine oxidase A (MAOa), a degradation enzyme. Optogenetic activation of the SNS upregulates NE uptake by SAMs and shifts the SAM profile to a more pro-inflammatory state. NE uptake by SAMs is prevented by genetic depletion of Slc6a2 or inhibition of the transporter. We also found that obesity increases SAM content in the SNS. In two mouse models of obesity, genetic ablation of Slc6a2 in SAMs increases brown adipose tissue (BAT) content, causes browning of white fat, increases thermogenesis and leads to significant and sustained weight loss. We further show that this pathway is conserved as human sympathetic ganglia also contain SAMs and the analogous molecular machinery for NE clearance, thus constituting a potential target for obesity treatment.

overall_design: The authors sorted and profiled gene expression by low input RNAseq of F4/80+CD45+ double positive cells from sympathetic ganglia (SAM ganglia), sympathetic nerve fibers from inguinal fat (SAM fibers), neighboring subcutaneous fat (sATM), visceral fat (vATM), spleen (SpM) and brain (microglia)

first: Ana

last: Domingos


title: The juvenile rhesus monkey (Macaca mulatta) testicular transcriptome and differential gene expression associated with the decision of undifferentiated spermatogonia to commit to the differentiation pathway

city: Pittsburgh

first: Uma

last: Chandran

state: PA

address: 5150 Center Ave

department: BioMedical Informatics

institute: University of Pittsburgh

summary:
Goal:  To identify the genes associated with the decision of undifferentiated spermatogonia to commit to a pathway of differentiation.
Methods: testis mRNA profiles of 10 juvenile wild-type rhesus monkeys (3 vehicle-treated; 6 gonadotropin treated) were generated by deep sequencing, in triplicate and quadruplicate, using Illumina NextSeq500. The sequence reads that passed filters were analyzed at the transcript isoform level with TopHat followed by Cufflinks. qRT–PCR validation was performed using TaqMan and SYBR Green assays
Results: We mapped about 45-55 million sequence reads per sample to the rhesus monkey genome (rheMac 8.0.1) and identified 26,813 transcripts in the testes of wild type monkeys with TopHat workflow.  Approximately a combined 7% of unique transcripts showed differential expression between vehicle and gonadotropin treatment for 48 and 96h, with a fold change ≥1.5 and p value <0.05. Altered expression of 12 genes was confirmed with qRT–PCR, substantiating the RNA-seq findings.
Conclusions: The testis transcriptome of the juvenile monkey contained 15,475 genes of which 15,174 were protein encoding genes or pseudogenes.  Gonadotropin stimulation for 48 h resulted in the commitment of spermatogonia to differentiate and this was associated with the emergence of 1587 differentially expressed genes.
Funding support: NIH R01 HD072189 to Tony Plant

overall_design: Whole testis mRNA profiles of juvenile wild type rhesus monkeys were generated by deep sequencing, in triplicate or quadruplicate, using Illumina NextSeq500.

first: Rahil

last: Sethi

first: William

last: Walker


title: RNA Seq of Jag1Ndr/ndr and Jag1+/+ liver

first: Urban

last: Lendahl

summary: RNA sequencing of livers from postnatal day 10 (P10) control mice (Jag1+/+) and an Alagille mouse model (Jag1Ndr/Ndr).

overall_design: Transcriptome profiling of postnatal day 10 livers from Jag1Ndr/Ndr and Jag1+/+ in triplicates using Illumina TruSeqTM

first: Emma

middle: R

last: Andersson

first: Simona

last: Hankeova

first: Indira

last: Chivukula

first: Helena

last: Storvall

first: Daniel

last: Ramsköld

first: Yat

last: Tsoi


title: Clustering gene expression time series data using an infinite Gaussian process mixture model

status: Public on Oct 08 2017

city: Durham

first: Timothy

last: Reddy

state: NC

address: 101 Science Drive

institute: Duke University

summary: In order to identify and characterize novel human gene expression responses to glucocorticoids, we exposed the human lung adenocarcinoma cell line, A549, to the synthetic glucocorticoid dexamethasone for 1, 3, 5, 7, 9, and 11 hrs in duration as well as to a paired vehicle control, ethanol. We assayed gene expression with RNA-seq and clustered gene expression profiles using an infinite Gaussian process mixture model.

overall_design: Time series treatment of human A549 cells with dexamethasone or paired vehicle control.

middle: M

last: Vockley

first: Ian

last: McDowell

first: Dinesh

last: Manandhar

first: Amy

middle: K

last: Schmid

first: Barbara

last: Engelhardt


title: DNA Methylation Profiling of Bone Sarcomas

city: New York City

first: Shengyang

last: Wu

state: NY

address: 560 First Avenue

department: Radiation Oncology

institute: New York University School of Medicine

type: Methylation profiling by genome tiling array

summary: DNA Methylation-Based Classifier for Accurate Molecular Diagnosis of Bone Sarcomas: DNA methylation status of 482,421 CpG sites in 10 Ewing’s sarcoma, 11 synovial sarcoma, and 15 osteosarcoma samples were determined using the Illumina HumanMethylation450 array.

overall_design: DNA methylation status of 482,421 CpG sites in 10 Ewing’s sarcoma, 11 synovial sarcoma, and 15 osteosarcoma samples were determined using the Illumina HumanMethylation450  array.  We developed a random forest classifier trained from the 400 most differentially methylated CpG sites within the training set of 36 sarcoma samples. This classifier was validated on data drawn from The Cancer Genome Atlas (TCGA) synovial sarcoma, TARGET Osteosarcoma, and a recently published series of Ewing’s sarcoma tumors.

first: S

first: Benjamin

middle: T

last: Cooper


title: Single cell RNA Seq data of BMDM from mouse infected with Salmonella [stat log]

status: Public on Oct 07 2017

first: Itai

last: Yanai

address: 430 East 29th St., Office 510

department: Biochemistry and Molecular Pharmacology

institute: New York University, School of Medicine

summary: The interaction between a pathogen and a host is a highly dynamic process in which both agents activate complex programs. Here, we introduce a single-cell RNA-Seq method (scDual-Seq) that simultaneously captures both host and pathogen transcriptomes and use it to study the process of infection of individual mouse macrophages with the intracellular pathogen Salmonella typhimurium. Among the infected macrophages, we found three subpopulations and we show evidence for a linear progression through these subpopulations, supporting a model in which these three states correspond to consecutive stages of infection.

sample_organism: Salmonella enterica subsp. enterica serovar Typhimurium str. SL1344

overall_design: salmonella RNA extracted from liquid stationary culture

first: Gal

last: Avital

first: Roi

last: Avrahanm

last: Fan

first: Tamar

last: Hashimshony

first: Deborah

last: Hung


title: The transcriptome of estrogen-independent mammary growth induced by dietary conjugated linoleic acid

city: Davis

first: Grace

last: Berryhill

address: One Shields Avenue

department: Animal Science

institute: University of California, Davis

summary: RNA sequencing confirmed that both E and CLA increased the expression of genes involved in proliferation and cell cycle progression. The transcriptome of the mammary glands from E-treated mice was characterized by canonical E-induced genes including Pgr and Areg, whereas CLA differentially regulated the expression of genes implicated in inflammation – a finding in-line with perturbation of the mammary stroma stimulated by CLA.

overall_design: A total of 8 groups (Control_60h, Estrogen_60h, Control_84h, Estrogen_84h, Control 108h, CLA_108h, Control_132h, CLA_132h) were analyzed with 5 biological replicates per group for a total of 40 samples.

first: Russell

middle: C

last: Hovey


title: Single cell RNA Seq data of BMDM from mouse infected with Salmonella

platform_organism: Salmonella enterica subsp. enterica serovar Typhimurium str. SL1344

overall_design: Refer to individual Series


title: Single-cell RNA sequencing of EPLM subpopulations (152 Ly6D+ and 213 TN)

status: Public on Oct 11 2017

city: Basel

first: Llucia

last: Alberti-Servera

state: Basel stat

address: Mattenstrasse 28

department: Department of Biomedicine

institute: Uni Basel

summary: Single-cell RNA-sequencing has emerged as a powerful technology to assess heterogeneity within defined cell populations. Here, we comprehensively study the heterogeneity of a previously described B220+CD117intCD19-NK1.1- uncommitted hematopoietic progenitor with combined lymphoid and myeloid potential (EPLM). Using staining for surface markers together with functional assays, we describe four subpopulations of this progenitor with distinct lineage developmental potentials. Amongst them, the Ly6D+SiglecH-CD11c- fraction was lymphoid restricted exhibiting strong B-cell potential, whereas the Ly6D-SiglecH-CD11c- fraction showed mixed lympho-myeloid potential. Single-cell RNA-sequencing of these subsets revealed that the latter population is composed of a mixture of cells with distinct lymphoid and myeloid genetic signatures and identified a subgroup as the potential precursor of Ly6D+SiglecH-CD11c-. We observed a B-cell priming gradient within the Ly6D+SiglecH-CD11c- subset and propose a herein newly identified subgroup as the direct precursor of the first B-cell committed stage. Our study demonstrates that the multipotency of B220+CD117intCD19-NK1.1- progenitors is a result of underlying heterogeneity at the single-cell level. Moreover, it highlights the validity of single-cell transcriptomics to resolve cellular heterogeneity and identify developmental relationships in hematopoietic progenitors.

overall_design: Single cells were captured from ex-vivo sorted hematopoietic progenitors (from Flt3Ltg mice) on a small-sized (5-10μm) C1 chip using the Fluidigm C1 system. A total of 3 chips per population were used yielding to 178 Ly6D+ and 232 TN single cells captured. During the quality control, cells having less than 60% of mapped reads, less than 2x10e5 counts, or less than 800 detected genes were filtered out from further analysis. In total, 152 Ly6D+ and 213 TN passed these criteria, which are the samples provided in the repository.

first: Robert

last: Ivanek

first: Antonius

last: Rolink


title: Single-cell RNA sequencing reveals developmental heterogeneity among early lymphoid progenitors

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


title: N6-methyladenosine-Mediated Nuclear Export of Messenger RNA

city: Chicago

first: Ian

last: Roundtree

state: IL

address: 929 E 57th Street E307

department: Department of Chemistry

institute: University of Chicago

summary: N6-methyladenosine (m6A) is the most common internal modification in eukaryotic messenger RNA (mRNA). The effects of this reversible cheimcal modification are mediated in part by methyl-specific RNA binding protein 'readers' of the YTH family. In this study we present the function of YTHDC1 (YT521) in promoting the export of mRNA from the nucleus to the cytoplasm. Additionally, we identify a role for YTHDC1 in exon retention in mouse embryonic stem cells (mESCs).

overall_design: Two complementary methods were used to determine YTHDC1 target transcripts within HeLa cells. Subcellular RNA-sequencing following knockdown of YTHDC1 was used to analyze transcript distribution. In mESCs, knockout of Ythdc1 showed minor splicing differences. We analyzed the differences in RNA binding by NXF1 and SRSF3 by RIP-seq in the presence and absence of YTHDC1.

first: Guan-Zheng

last: Luo

first: Chuan

last: He


title: Liver Macrophage Infiltration and Inflammation are associated with Antiviral Responses During SIV Infection in Macaques that is Only Partly Reduced during cART

city: Minneapolis

first: Michael

last: Gale

state: Minnesota

address: 689 23rd Ave SE

department: Microbiology and Immunology

institute: University of Minnesota

summary: Non-AIDS- related conditions have become increasingly more prevalent in those infected with HIV, with liver disease being one of the most predominant complications. Using the macaque simian immunodeficiency virus (SIV) model, we identified upregulation of numerous inflammatory pathways in the liver, which only partially resolved in cART (combination antiretroviral therapy) treated macaques.

overall_design: 11 adults (4 uninfected, 5 SIV-infected, 2 SIV-infected + cART treatment) and 20 infants (7 uninfected, 9 SIV-infected, 4 SIV-infected + cART treatment). We performed microarray analysis on liver samples collected from each animal during necropsy.

first: Bridget

middle: S

last: Fisher

first: Richard

last: Green

first: Rachel

last: Brown

first: Matthew

last: Wood

first: Andrew

last: Miller

first: Jeffrey

last: Lifson

last: Gale Jr

first: Ann

last: Chahroudi

first: Guido

last: Silvestri

first: Nichole

last: Klatt

first: Donald

last: Sodora


title: Transcriptional response to lactic acid stress in the hybrid yeast Zygosaccharomyces parabailii

status: Public on Oct 06 2017

city: Delft

first: Raúl

middle: Antonio

last: Ortiz-Merino

address: van der Maasweg 9

department: Biotechnology

country: Netherlands

institute: Delft University of Technology

summary: In this study we used stringent mapping methods and a high-quality genome sequence to study the transcriptional response to lactic acid stress in Zygosaccharomyces parabailii ATCC60483, a natural interspecies hybrid yeast

overall_design: The RNA seq was performed on bioreactor grown cells either with addition of lactic acid or without. The samples were taken at two timepoints corresponding to exponential growth phase and post diaxulic shift

first: Nurzhan

last: Kuanyshev

first: Kenneth

middle: H

last: Wolfe

first: Paola

last: Branduardi


title: Single nuclei RNA-seq using droplet microfluidics (DroNc-seq)

city: Cambridge

first: Naomi

last: Habib

state: MA

address: 75 Ames st

department: Zhang

institute: Broad Institute

summary: We report the application of DroNc-seq on archived tissue from mouse adult hippocampus and cortex barin regions

overall_design: DroNc-seq application in 4 mouse brain tissue on two different brain regions


title: RNA sequencing analysis of triple cytokine-captured human CD4 T cells

city: Oxford

first: Mohammad Hussein

last: Al-Mossawi

address: Botnar Research Centre

department: Nuffield Department of Orthopaedics Rheumatology and Musculoskeletal sciences

institute: University of Oxford

summary: GM-CSF positve CD4 cells are found at sites of inflammation. The purpose of this study was to understand their transcriptional profile relative to known Th1 and Th17 subsets.

overall_design: Human CD4 T cells were isolated by magnetic negative selection and activated with PMA and ionomycin. A cytokine capture assay was used to isolate CD45RA-positive, cytokine negative, IFN-gamma-single-positive, IL-17A-single-positive, GM-CSF-single positive and IL-17A-GM-CSF-double positive cells.

first: Mohammad

first: Hai

last: Fang

first: Paul

last: Bowness


title: Single cell RNA Seq data of BMDM from mouse infected with Salmonella [hundred_ten_singles]

first: Itai

last: Yanai

overall_design: 40 single cells, 6 ten cells bulk, 2 hundred cells bulk,  in two time point of infection (0,4 hours after infection)

first: Gal

last: Avital

first: Roi

last: Avrahanm

first: Amy

last: Fan

first: Tamar

last: Hashimshony

first: Deborah

last: Hung


title: Quantitative analysis of primary human and pluripotent stem cell-derived brain microvascular endothelial cells transcriptomes via next generation sequencing

city: Madison

first: Tongcheng

last: Qian

state: WI

address: 1415 Engineering Drive, Room 3745

institute: UW-Madison

summary: Next-generation sequencing (NGS) has significantly facilitated the analysis of the gene profile and elucidated the molecular mechanisms important for specific cell lineage differentiated from human pluripotent stem cells (hPSCs). Here we report the application of RNA-sequencing technology for transcriptome profile of primary human brain microvascular endothelial cells (BMECs) and hPSC-derived BMECs, and comparison of transcriptomes among cells, including hPSCs, mesodermal progenitors, ectodermal progenitors, endodermal progenitors, hBMECs and hPSC-derived BMECs from hPSCs. Four different BMEC samples differentiated from hPSCs by two different methods and hBMECs were performed in IIIumina HiSeq2500. The resulting sequence reads (about 20 million reads per sample) were mapped to human genome (hg19) using HISAT, and the RefSeq transcript levels (FPKMs) were quantified using the python script rpkmforgenes.py. We next analyzed the expression of a subset of genes that regulate key BBB attributes, including tight junctions and molecular transporters. The gene set comprises 20 tight junction-related genes and an unbiased list of all 25 CLDN genes, all 407 solute carrier (SLC) transporters, and all 53 ATP-binding cassette (ABC) transporters regardless of prior knowledge of BBB association. Primary human BMECs expressed 234 of these genes. BMECs differentiated from hPSCs via the defined method expressed many of these same genes (206 of 234 (88%)) as did BMECs differentiated via the UM method (208 of 234 (89%), indicating a close similarity between hPSC-derived BMECs and primary human BMECs with respect to transcripts having likely relevance to BBB function. RNA sequencing was used to compare global gene expression profiles in the hPSC-derived BMECs with BMECs generated from our previously reported co-differentiation system and primary human BMECs. As expected, hPSC-derived BMECs from three independent differentiations clustered closely and were similar to those generated from the undefined UM platform. Moreover, the hPSC-derived BMECs clustered with primary human BMECs and were distinct from undifferentiated hPSCs and hPSC-derived ectoderm, endoderm and mesodermThis study provides detailed transcriptome comparison between hBMECs and hPSC-derived BMECs and unveils important genes related BMEC phenotypes.

overall_design: Transcriptome profiles of human primary brain microvascular endothelial cells and hPSC-derived brain microvascular endothelial cells were generated by RNA-seq technology using IIIumina HiSeq2500

first: Eric

middle: V

last: Shusta

first: Sean

last: Palecek


title: BEX1 is an RNA-dependent mediator of cardiomyopathy

city: Cincinnati

first: Tobias

last: Schips

address: 3333 Burnet Ave

institute: Cincinnati Children's Hospital Medical Center


title: BEX1 is an RNA-dependent mediator of cardiomyopathy [RIP-Seq]

summary: Regulation of mRNA splicing, processing and stability is increasingly recognized as a critical control point in dynamically altering gene expression during stress or disease states. Very little is understood of this process in the heart despite the wide ranging changes in gene expression that occurs during heart failure; a disease of epidemic proportions in the western world. Here we identified BEX1 as a heart-failure-induced gene and identified its function as an mRNA binding protein responsible for enhancing expression of a subset of cardiac disease promoting genes. Modeling this increase in BEX1 that occurs in disease, BEX1 cardiac-specific overexpressing transgenic mice were generated and shown to have worse cardiac disease with stress stimulation while Bex1 gene-deleted mice were protected from insults that would otherwise promote heart failure. Since BEX1 was of unknown molecular function we performed a series of proteomic and interactive screening assays, which identified Bex1 as part of a large ribonucleoprotein processing complex involved in regulating pro-inflammatory mRNA expression in the heart. Specifically, induction of BEX1 augmented the stability and expression of AU-rich element containing mRNAs typically found within pro-inflammatory genes. Thus, BEX1 functions as an mRNA modulatory effector that augments pathology promoting gene expression during heart failure.

overall_design: Analysis of neonatale neonatale rat cardiomyocytes overexpressing Bex1 or beta galactosidase

first: Federica

last: Accornero

first: Jeff

last: Molkentin


title: RNAseq and ChIPseq on Mouse Embryonic Stem Cells and Sorted Mesendoderm Progenitor Cells [RNA-seq]

city: Lausanne

first: Frederic

last: Burdet

address: Quartier Sorge - Batiment Amphipole

institute: Swiss Institute of Bioinformatics

summary: We mapped the enhancer and long non-coding transcriptional landscape during mesendoderm specification. Mesendodermal progenitors were sorted from differentiating ESCs according to Eomes expression. Enhancer usage was coordinated with mesendoderm-specific expression of key lineage-determining transcription factors. We demonstrated that many of these enhancers are associated with the expression of lncRNAs.

overall_design: RNA profile of mESC and sorted mesendoderm progenitors.

first: Michael

last: Alexanian

first: Daniel

last: Maric

first: Stephen

last: Jenkinson

first: Marco

last: Mina

first: Clayton

last: Friedman

first: Ching

last: Ting

first: Rudi

last: Micheletti

first: Isabelle

last: Plaisance

first: Mohamed

last: Nemir

first: Damien

last: Maison

first: Jasmin

last: Kernen

first: Iole

last: Pezzuto

first: Dominic

last: Villeneuve

first: Frédéric

first: Mark

last: Ibberson

middle: L

last: Leib

first: Nathan

middle: J

last: Palpant

first: Nouria

last: Hernandez

first: Samir

last: Ounzain

first: Thierry

last: Pedrazzini


title: RNAs enriched at protrusions of control or APC knockdown fibroblasts

city: Bethesda

first: Margaret

last: Cam

institute: NIH

summary: We used a fractionation scheme to isolate protrusions and cell bodies from control or APC knockdown fibroblasts, which were induced to extend protrusions in response to addition of lysophosphatidic acid (LPA). Four replicate protrusion and cell body samples for each cell type were analyzed by RNA-Seq. To identify RNAs enriched in protrusions, a protrusion/cell body ratio was calculated. We find that ca. 7% of detected RNAs were enriched in protrusions of control cells. Knockdown of APC reduced the protrusion enrichement of a subset of these RNAs, indicating that they are dependent on APC for their localization. Localization of the remaining RNAs was not significantly affected upon APC knockdown suggesting that their localization is mediated by an APC-independent mechanism.

overall_design: RNA-seq profiles of protrusions and cell bodies from cells transfected with control siRNAs or with siRNAs against APC. Four replicates were sequenced on Illumina HiSeq2500.

first: Stavroula

last: Mili

first: Maggie


title: Effect of overexpressed UTR constructs on RNA enrichment at protrusions

summary: We generated fibroblast lines overexpressing beta-globin constructs under a doxycycline-inducible promoter. These constructs contain the beta-globin coding sequence followed by either a control 3'UTR (HBB) or the Pkp4 3'UTR. The Pkp4 3'UTR carries a protrusion-targeting element and is expected to compete with localization of endogenous protrusion-localized mRNAs.  We used a fractionation scheme to isolate protrusions and cell bodies from HBB and Pkp4 UTR-expressing fibroblasts, which were induced to extend protrusions in response to addition of lysophosphatidic acid (LPA). 4 replicate protrusion and cell body samples for each cell type were analyzed by RNA-Seq. To identify RNAs enriched in protrusions, a protrusion/cell body ratio was calculated. We find that ca. 4.5% of detected RNAs were enriched in protrusions of control HBB cells. Overexpression of the Pkp4 UTR reduced the protrusion enrichement of a subset of these RNAs, while the degree of localization of the remaining RNAs was not significantly affected.

overall_design: RNA-seq profiles of protrusions and cell bodies from cells stably overexpresing beta-globin constructs carrying either a control 3' UTR (HBB) or the Pkp4 3'UTR. 4 replicates were sequenced on Illumina HiSeq2500.


title: Transcriptomic profiling of CD8+ T cells from steroid-sensitive and steroid-resistant asthmatics and healthy controls

city: Denver

first: Brian

middle: Patrick

last: OConnor

state: Colorado

address: 1400 Jackson St

department: Center for Genes, Environment and Health

institute: National Jewish Health

summary: The heterogeneity of asthma has prompted attempts at classifications based on clinical phenotypes and cluster designations. Although helpful, reliance on clinical parameters fails to address fundamental issues resulting in specific phenotypes, in particular, steroid-resistant (SR) asthma. Limited data are available defining the mechanisms or pathways (endotypes) causing the failure of inhaled corticosteroids (ICS) in SR asthmatics. Unlike CD4+ T cells, both human and mouse CD8+ T cells fail to undergo apoptosis in the presence of corticosteroids (CS) positioning them as unique effector cells. Whole transcriptome analyses of peripheral CD8+ T cells from steroid-sensitive or steroid-resistant asthmatics or healthy controls on days 0 and 8 were performed.

overall_design: RNA-sequencing of activated (anti-CD3/CD2) CD8+ T cells purified from the peripheral blood from steroid-sensitive and steroid-resistant asthmatics and healthy controls on day 0 and cultured in the presence of IL-2 for 8 days.

middle: P

first: Laura

last: Harmacek

first: Thomas

last: Danhorn

first: Sonia

last: Leach

first: Erwin

middle: W

last: Gelfand

first: Michaela

last: Schedel

first: Sven

last: Michel


title: Mitochondrially-imported RNA in Drug Discovery

city: Tuscaloosa

first: Carl

middle: A.

last: Pinkert

state: AL

address: 201 Rose Administration Bldg

institute: University of Alabama

summary: The import of nuclear transcribed RNAs into mitochondria is an emerging area that presents tremendous opportunity to develop human metabolic therapeutics. However, our knowledge base is quite limited. Much remains to be discovered regarding specific RNA localization and mechanisms of import. In order to identify novel RNAs imported into mitochondria, all RNAs within the mitochondria were characterized using next generation sequencing technology. Several nuclear transcribed RNAs were found within mitochondrial RNA samples, including nuclear ribosomal RNAs, gamma satellite RNA and VL30 retroelement RNA. The presence of these RNAs within mitochondria coupled with RNA sequencing data (RNAseq) from other laboratories investigating mitochondrial RNA processing, lead us to hypothesize that nuclease treatment of mitoplasts is insufficient for removing contaminating cytoplasmic RNAs. In contrast to traditional methodology, mitochondrial import was evaluated by qRT-PCR after stepwise removal of the outer mitochondrial membrane and subsequent lysis of mitochondria. This allowed identification of RNAs lost from the mitochondria with the same kinetics as mtDNA-transcribed RNAs. This approach provided an improved evaluation of nuclear RNA enrichment within mitochondrial membranes in order to characterize nuclease protection and mitochondrial import and identify false-positive detection errors. qRT-PCR results confirmed the presence of VL30 retroelement RNA within mitochondria and question the hypothesis that the RNA component of RNase P is imported. These results illustrate a reliable approach for evaluating the presence of RNAs within mitochondria and open new avenues of investigation relating to mitochondrial RNA biology and in targeting mitochondrial based therapeutics.

overall_design: RNA isolated from purified mitoplasts was sequenced on an Illumina Genome Analyzer IIx

first: Matthew

last: Cannon

first: Michael

last: Irwin

middle: A


title: Genomic profiling of human spermatogonial stem cells [ATAC-Seq]

status: Public on Oct 05 2017

city: Salt Lake City

first: Jingtao

last: Guo

state: UT

address: 2000 Circle of Hope

department: Department of Surgery Division of Urology

institute: University of Utah School of Medicine

summary: To better understand human spermatogonial stem cells (SSCs), we profiled their transciptome and epigenome, which revealed the mechanism how human SSCs regulates their self-renewal versus differentiation dermination, as well as how latent pluripotency is established in human SSCs. Remarkly, we discovered signaling pathways (e.g. LIF, BMP, WNT) that differentially regulated self-renewal vesus differentiation in SSCs. We also discovered that SSCs repress core pluripotent factors (Sox2, Pou5f1 and Nanog) yet activate ancillary factors (e.g. Klf4, Mbd3, Tcf3, Sall4) transcriptionally and epigenetically.

overall_design: Using SSEA4 as self-renewal marker, we isolated self-renewal SSCs by magnetic antibody cell sorting (MACS). We also collected hESCs for comparison. Standard ATAC-seq protocol was then performed on collected cells.

first: Edward

last: Grow

first: Chongil

last: Yi

first: James

last: Hotaling

first: Bradley

last: Cairns


title: HNF1 regulates critical functions of the human epididymis epithelium. [ChIP-Seq]

first: Ann

last: Harris

state: Illinois

address: 2430 N Halsted

institute: Northwestern University

summary: HNF1α and HNF1β recognize the same DNA consensus sequence in the genome, to which they bind as homodimers or heterodimers. Both factors share a high degree of homology their DNA binding and dimerization (N-terminus) regions but have a more divergent C-terminal transactivation domain. HNF1β is essential for the generation of a functional male reproductive tract in mice and genital tract abnormalities are evident in humans with recessive mutations in HNF1β. The functions of HNF1α and HNF1β have been studied in epithelia from other several tissues (liver, kidney, intestine, and pancreas) but their role in the adult human epididymis epithelium (HEE) remains unexplored. We established that HNF1α/β are expressed in caput HEE cells and are predicted to occupy cis-regulatory elements in these cells. We next used Chromatin immunoprecipitation followed by deep sequencing (ChIP-Seq) to examine the genome-wide binding of HNF1 in caput HEE cells from two donors. The majority of the HNF1 peaks localized within intergenic regions and intronic regions rather than at promoter regions. This implies an important role for HNF1 in regulating gene expression though distal cis-regulatory elements. HNF1 occupies many genomic sites that are close to genes with a role in epididymis epithelial transport.

overall_design: Examination of HNF1 binding in Caput HEE cells. There are two replicates of the HNF1 ChIP-seq plus input samples (control).

last: Browne

first: Rui

last: Yang


title: Single cell RNA Seq data of BMDM from mouse infected with Salmonella [dilution; CEL-Seq2]

city: New York

first: Itai

last: Yanai

summary: The interaction between a pathogen and a host is a highly dynamic process in which both agents activate complex programs. Here, we introduce a single-cell RNA-Seq method (scDual-Seq) that simultaneously captures both host and pathogen transcriptomes and use it to study the process of infection of individual mouse macrophages with the intracellular pathogen Salmonella typhimurium. Among the infected macrophages, we found three subpopulations and we show evidence for a linear progression through these subpopulations, supporting a model in which these three states correspond to consecutive stages of infection.

overall_design:
a dilution series of mouse and salmonella RNA

Please note that the samples are identical to GSM2729237-GSM2729241 and the RNA was extracted simultaneously. The only difference between them is the different protocol by which the libraries were made for sequencing (i.e. CEL-Seq2 or scDual-Seq).

first: Gal

last: Avital

first: Roi

last: Avrahanm

first: Amy

last: Fan

first: Tamar

last: Hashimshony

first: Deborah

last: Hung


title: Profiling and regulation of protrusion-enriched RNAs from mouse fibroblasts

first: Margaret

last: Cam

state: MD

overall_design: Refer to individual Series


title: Genomic profiling of human spermatogonial stem cells [scRNA-Seq]

overall_design: Using SSEA4 as self-renewal marker and Kit as differentiating marker, we isolated self-renewal and differentiation SSCs by magnetic antibody cell sorting (MACS). SSEA4+ or Kit+ cells were loaded into 5-10 μm integrated fluidic circuits (IFCs) using Fluidigm C1 instrument. Single cells in IFCs were lysed and total RNA was harvested for polyadenylation selection, reverse transcription and PCR amplification. Library constructions were performed according to Fluidigm Library preparation with Nextera XT protocol and sequenced on a 50-cycle single end run.

first: Ed


title: Single cell RNA Seq data of BMDM from mouse infected with Salmonella [full time course]

overall_design: 96 single cells in 4 time point of infection (0,2.5,4,8 hours after infection)


title: Temporal tracking of microglia activation in neurodegeneration at single-cell resolution

first: Li-Huei

last: Tsai

state: Massachusetts

department: Picower Institute for Learning and Memory

institute: MIT

summary: In this study, we used single-cell RNA-sequencing to gain unprecedented insight into the phenotypic heterogeneity and the transcriptional dynamics of microglia cells during the progression of neurodegeneration. Briefly, by using a severe neurodegeneration mouse model with Alzheimer’s-like pathology and phenotypes (CK-p25 model), we surveyed microglia activation by RNA sequencing longitudinally at fine temporal- and single-cell resolution. In summary, our work identified previously unobserved heterogeneity in the response of microglia to neurodegeneration, discovered novel microglia cell states, revealed the trajectory of cellular reprogramming of microglia in response to neurodegeneration, and uncovered the underlying transcriptional programs. These insights into the molecular programs underlying microglia activation provided by our study may pave the way for designing new rational and efficient strategies to treat Alzheimer’s and other neurodegenerative diseases.

overall_design: To determine the phenotypic heterogeneity and the transcriptional dynamics of microglia cells during the progression of neurodegeneration, we used the CK-p25 mouse model of severe neurodegeneration which develops AD-like pathology in an inducible and temporally predictable manner. We isolated cells expressing the microglia markers CD11b and CD45 from the hippocampus of three to four CK-p25 animals and three CK control littermates at each of the following four time points during the progression of neurodegeneration: before p25 induction (0 weeks), 1 week, 2 weeks, and 6 weeks after p25 induction. Single microglial cells (expressing the cell surface markers CD11b and CD45) were sorted directly into RNA lysis buffer in 96-well plates using Fluorescence-Activated Cell Sorting (FACS). We prepared a total of 2183 single-cell RNA-sequencing libraries using a modified version of the Smart-Seq2 protocol and sequenced the libraries to a depth of 251353 (median) mapped reads per cell.

first: Hansruedi

last: Mathys

first: Fan

last: Gao


title: Transcriptome Data of Peritoneal Macrophages Isolated from ApoE-/-/Ch25h-/- Mice

city: La Jolla

first: John

last: Shyy

state: California

address: 9500 Gilman Dr., BRF2 Rm 1217 Bay LL

institute: University of California, San Diego

summary: Cholesterol 25-hydroxylase (Ch25h) has been previously demonstrated to be required for anti-inflammatory effect and anti-viral activity, while also promoting foam cell formation.  In order to delineate the role of Ch25h in atherosclerosis, we isolated peritoneal macrophages (PMs) from mice lacking ApoE and Ch25h (i.e., ApoE-/-/Ch25h-/-) compared to their wildtype littermates (i.e., ApoE-/-/Ch25h+/+) for transcriptome analysis, in biological duplicates (n=12 pooled mice per group).  Deep-sequencing analysis revealed that PMs ablated with Ch25h were prone to the pro-inflammatory M1 phenotype as well as increased cholesterol biosynthesis.  Overall, we found that Ch25h is integral to atheroprotection through the preservation of M2 polarization and suppression of intracelllular cholesterol content.

overall_design: Transcriptome data of PMs isolated from ApoE-/-/Ch25h-/- mice (n=12 per group) followed by deep sequencing, in biological duplicate, using Illumina NextSeq 500.

first: Zhao

last: Li

first: Marcy

last: Martin

middle: Y


title: Single cell RNA Seq data of BMDM from mouse infected with Salmonella [dilution]

institute: New York University, School of Medicine

overall_design: a dilution series of mouse and salmonella RNA


title: Identification of Super-enhancer-associated Cancer Genes Provides Novel Therapeutic Targets in Adult T-cell Leukemia/Lymphoma

city: Lund

first: Phuong

last: Cao Thi Ngoc

address: BMC, B1222b, Klinikgatan 26

institute: Lund University

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


title: RNA-seq data of the Streptomyces venezualae WT and mtrB::apr mutant strains at time point 14 and 20 h (biological duplicates).

city: Norwich

middle: Tyson

last: Munnoch

address: Norwich Research Park

department: Schol of Biological Sciences

institute: University of East Anglia

summary: We report here the sequence results of S. venezualae WT and the mutant mtrB::apr strains at 14h and 20h and the differential expression profile of WT vs. Mutant.

overall_design: WT vs mtrB::apr mutant at 14h and 20h.  Total RNA was isolated and processed by vertis Biotechnologie AG.  Ribodepletion was carried out to remove ribosomal RNA, the sample Tex treated, split in two, 1 half TAP treated the other TAP minus, adaptors were then ligated with the TAP plus sample containing sequence/data enriched for TSS.  EAch was then converted to cDNA and sequenced.  WT vs Mutant was compared at 16h while the others were used for the purposes of TSS identification.


title: HNF1 regulates critical functions of the human epididymis epithelium.

first: Ann

last: Harris


title: RNA Pol II ChIP-seq analysis after THZ1 treatment in TL-Om1 cells

summary: ChIP-seq analysis was performed in an adult T-cell leukemia/lymphoma cell line (TL-Om1) to analyze DNA bindings of RNA polymerase II (Pol II) after treatment with the THZ1 CDK7 inhibitor.

overall_design: TL-Om1 cells were treated with control DMSO or THZ1 for 3 hours.  The cells were crosslinked with formaldehyde for 10 min. DNA was enriched by chromatin-immunoprecipitation (ChIP) and analyzed by Illumina Hiseq 4000.  A sample of whole cell extract (WCE) was sequenced and used as the background to determine enrichment.  ChIP was performed using an antibody against RNA polymerase II (Abcam, ab817: 8WG16).

first: Regina

last: Wong

last: Ngoc

first: Wei

middle: Z

last: Leong

first: Alice

last: Yam

first: Takaomi

last: Sanda


title: HNF1 regulates critical functions of the human epididymis epithelium. [RNA-Seq]

summary: HNF1α and HNF1β recognize the same DNA consensus sequence in the genome, to which they bind as homodimers or heterodimers. Both factors share a high degree of homology their DNA binding and dimerization (N-terminus) regions but have a more divergent C-terminal transactivation domain. HNF1β is essential for the generation of a functional male reproductive tract in mice and genital tract abnormalities are evident in humans with recessive mutations in HNF1β. The functions of HNF1α and HNF1β have been studied in epithelia from other several tissues (liver, kidney, intestine, and pancreas) but their role in the adult human epididymis epithelium (HEE) remains unexplored. We established that HNF1α/β are expressed in caput HEE cells and are predicted to occupy cis-regulatory elements in these cells. To investigate the contribution of HNF1 in controlling gene expression in caput cells we performed siRNA-mediated depletion of HNF1α and HNF1β together, followed by RNA-seq analysis. Three replicas of caput cells were transfected with the specific siRNAs or with a non-targeting control siRNA. RNA-seq after HNF1 depletion showed significant alterations in the expression of genes encoding ion channels and exchangers that are involved in controlling the luminal environment in the caput epididymis.

overall_design: mRNA profiles from Caput HEE cells transfected with negative control (NC) or HNF1alpha and HNF1beta siRNA, in triplicate.

first: James

last: Browne

first: Rui

last: Yang


title: Transcriptome analysis for identification of novel biomarker for disease progression in Dengue patients

city: Faridabad

first: ABHAY

middle: DEEP

last: PANDEY

state: HARYANA

address: 3RD MILE STONE, FARIDABAD GURGAON EXPRESSWAY, NCR BIOTECH CLUSTER

department: VIROLOGY

country: India

institute: Translational Health Science and Technology Institute

summary: To study the early transcriptional signature in the peripheral blood mononuclear cells (PBMCs) in a large number of clinically and virologically well characterized patients with mild and severs dengue infection and establishes their correlation with disease progression

overall_design: mRNA profiling of Dengue fever(DF), Dengue hemorrhagic fever [DHF], Dengue shock syndrome (DSS) and Disease control.

first: Arup

last: Banerjee

first: Abhay

middle: D

last: Pandey

first: Vikas

last: Sood

first: Sudhanshu

last: Vrati

first: Arjun

last: Malhotra

first: Priynka

first: Shweta

last: Shukla

first: Amitesh

last: Agarwal

first: Shukla

last: Das

first: V

middle: G

last: Ramachandran

first: Saptamita

last: Goswami

first: Bhaswati

last: Bandyopadhyay

first: Nimai

last: Bhattacharya


title: Exploring the gene expression profile upon FXR1 knockdown in H358 cells using RNA-seq

status: Public on Oct 04 2017

city: Shanghai

first: Bin

last: Xiang

address: Building 3, 4218 Jinke Road, Pudong

institute: novartis institutes for biomedical research

summary: We performed the RNA-seq in control samples and FXR1 knockdown samples, and compared the gene expression profiles to explore the effect of FXR1 knockdown on gene expression. The study was performed in H358 cells. Doxycycline inducible shRNA3 (sh3) was used to knockdown FXR1. Control shRNA (ctrl) samples were used to get rid of the effect of Doxycycline treatment. Both the Doxycycline treament for 3 days (D3) and 5 days (D5) samples were collected. Each sample has three repeats (rep 1, rep 2, and rep 3). The mRNA profiles were generated by deep sequencing using Illumina.Sequenced reads were trimmed for adaptor sequence, then mapped to hg19 whole genome using STAR v2.5.3 with parameters --bamRemoveDuplicatesType UniqueIdentical --outSAMmultNmax 1. Raw reads and Reads Per Kilobase per Megabase of library size (RPKM) were calculated using HOMER (PMID: 20513432). Differential gene expression was analyzed using R package DESeq2 using the raw reads.

overall_design: The mRNA profiles of control sample and FXR1 knockdown samples of H358 cells were generated by deep sequencing, in triplicate, using Illumina.

first: Jiao

last: Yue

first: Chun

last: Ma

first: Yao

last: Wang


title: Ancestry as a potential modifier of gene expression in breast tumors from Colombian women

city: New Orleans

first: Jovanny

last: Zabaleta

state: LA

address: 1700 Tulane Ave

department: Interdisciplinary Oncology

institute: LSUHSC

summary:
Background: Differences in breast cancer outcomes according to race/ethnicity have been reported. Hispanic/Latino (H/L) populations are a genetically admixed and heterogeneous group, with variable fractions of European, Indigenous American and African ancestries. Some studies suggest that breast cancer-specific mortality is higher in U.S. Hispanic/Latinas compared to non-Hispanic Whites (NHW) even after adjustment for socioeconomic status and education. The molecular profile of breast cancer has been widely described in NHWs but equivalent knowledge is lacking in Hispanic/Latinas. We have previously reported that the most prevalent breast cancer intrinsic subtype in Colombian H/L women was Luminal B as defined by surrogate St. Gallen 2013 criteria. In this study we explored ancestry-associated differences in molecular profiles of Luminal B tumors among these highly admixed women.
Methods: We performed whole-transcriptome RNA-seq analysis in 42 Luminal tumors (21 Luminal A and 21 Luminal B) from Colombian women. Genetic ancestry was estimated from a panel of 80 ancestry-informative markers (AIM). We categorized patients according to Luminal subtype and to the proportion of European and Indigenous American ancestry and performed differential expression analysis comparing Luminal B against Luminal A tumors according to the assigned ancestry groups.
Results: We found 5 genes potentially modulated by genetic ancestry: ERBB2 (Fold Change = 2.367, padj < 0.01), GRB7 (Fold Change = 2.327, padj < 0.01), GSDMB (Fold Change = 1.723, padj < 0.01, MIEN1 (Fold Change = 2.195, padj < 0.01 and ONECUT2 (Fold Change = 2.204, padj < 0.01). In the replication set we found a statistical significant association between European ancestry fraction and the expression levels of ERBB2 (p = 0.02, B = 2.49) and ONECUT2 (p = 0.04, B = -4.87). We also observed statistical significant associations for ERBB2 expression with Indigenous American ancestry (p < 0.001, B = 3.82). This association was not biased by the distribution of HER2+ tumors among the groups analyzed.
Conclusions: Our results suggest that genetic ancestry in Hispanic/Latina women might modify ERBB2 gene expression in Luminal tumors. Further analyses are needed to confirm these findings and explore their prognostic value.

overall_design: RNA profile of 42 luminal breast cancer tumors (21 luminal A and 21 luminal B) from Colombian patients

first: Silvia

last: Serrano-Gomes

first: María

middle: C

last: Sanabria-Salas

first: Jone

last: Garay

first: Melody

last: Baddoo

first: Gustavo

last: Hernández-Suarez

first: Juan

last: Mejía

first: Oscar

last: García

first: Lucio

last: Miele

first: Laura

last: Fejerman


title: Profiling of 5-methylcytosine (m5C) methylation in Arabidopsis thaliana wild-type

city: Singapore

first: Hao

last: Yu

address: Science Drive 4

country: Singapore

institute: National University of Singapore

summary: Methylation of carbon 5 in cytosine (5-methylcytosine; m5C) is a well-characterized DNA modification, and is also predominantly reported in highly abundant noncoding RNAs, such as rRNA and tRNA, in both prokaryotes and eukaryotes. However, the distribution and biological functions of m5C in plant mRNAs remain largely unknown. Here we develop an m5C RNA immunoprecipitation followed by deep sequencing approach (m5C-RIP-seq) to achieve transcriptome-wide profiling of RNA m5C in Arabidopsis thaliana. Liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) and dot blot analyses reveal a dynamic pattern of m5C mRNA modification in various tissues and at different developmental stages. m5C-RIP-seq analysis identifies 6,045 putative m5C peaks in 4,465 expressed genes in young seedlings. m5C is enriched in coding sequences with two peaks located immediately after start codons and before stop codons, and is associated with mRNAs with low translation activity. We further show that a RNA (cytosine-5)-methyltransferase, tRNA specific methyltransferase 4B (TRM4B), exhibits the m5C mRNA methyltransferase activity. Mutations in TRM4B display defects in root development and decreased m5C levels in root mRNA. Furthermore, TRM4B affects transcript levels of the genes involved in root development, which is positively correlated with their mRNA stability and m5C levels. Our results suggest that m5C in mRNA is a new epitranscriptome marker widely distributed in plant genes, and that regulation of this modification is an integral part of gene regulatory networks underlying plant development.

overall_design: m5C-seq in Arabidopsis thaliana (Col-0) wild-type, two replicates for each sample

first: Xuean

last: Cui

first: Zhe

last: Liang

first: Lisha

last: Shen

first: Qian

last: Zhang

first: Shengjie

last: Bao

first: Yuke

last: Geng

first: Vonny

last: Leo

first: Leah

last: Vardy

first: Tiegang

last: Lu

first: Xiaofeng

last: Gu


title: 5-Methylcytosine RNA methylation in Arabidopsis thaliana


title: Influence of PepFect14 transfection on cellular response

city: Stockholm

first: Moataz

last: Dowaidar

state: Stockholm

address: S.Arrheniusv. 16B

department: Neurochemistry

institute: Dpt. Neurochemistry Stockholm University

summary: Cell-penetrating peptides (CPP) uptake mechanism is still to be clarified to have a better understanding of their action in the mediation of oligonucleotide transfection. In this study, the effect on early events (1 h treatment) in transfection by Pepfect 14, with or without oligonucleotide cargo on gene expression, on HeLa cells, have been investigated. The RNA expression was characterized by RNA sequencing.

overall_design: The quality of purified total RNA was estimated by Agilent 2200 TapeStation analysis (Agilent Technologies, Santa Clara, USA). One µg of total RNA was used as an input to prepare next-generation sequencing libraries according to the Illumina TruSeq Stranded mRNA sample preparation protocol (Illumina, San Diego, USA). Final library mixtures were quantified by Qubit 2.0 Fluorometer (Life Technologies, Grand Island, USA) and validated with Agilent 2200 TapeStation analysis. Libraries were quantified by qPCR with Kapa Library Quantification Kit (Kapa Biosystems, Woburn, USA) to optimize cluster generation and sequenced on HiSeq2500 platform (Illumina, San Diego, USA) with 2 x 50 bp paired-end reads. Over 93.9% of the bases sequenced were above the quality of Q30. Demultiplexing was done with CASAVA 1.8.2. (Illumina, San Diego, USA) Allowing one mismatch in 6 bp index read. Initial data analysis was conducted by the RNA-Seq pipeline of Estonian Genome Centre, University of Tartu. Shortly, fastQ files were trimmed (removal of adapter sequences and bases below the quality Q20) with FASTX-Toolkit version 0.013 (http://hannonlab.cshl.edu/fastx_toolkit) and then aligned to the human reference genome (hg19/GRCh37) with Bowtie version 2.1.019 in combination with TopHat version 2.0.1320. Transcript quantification (measured as FPKM) was conducted with Cuffdiff program from Cufflinks version 2.2.121 with reference annotation Homo_sapiens.GRCh37.72.gtf (http://ftp.ensembl.org/pub/release-72/gtf/homo_sapiens) Cuffdiff analysis, which summarizes expression changes for all annotated gene variations, was filtered by lowest q-values (corrected p-values for multiple testing) from output file gene_exp.diff and the top list of differentially expressed genes were analyzed through the use of QIAGEN’s Ingenuity® Pathway Analysis (IPA®, QIAGEN Redwood City, www.qiagen.com/ingenuity).

first: Maxime

last: Gestin

first: Carmine

last: Cerrato

first: Helerin

last: Margus

first: Paula

last: Kivistik

first: Margus

last: Pooga

first: Mattias

last: Hällbrink

first: Ülo

last: Langel


title: Integration of the tricarboxylic acid (TCA) cycle with cAMP signaling and Sfl2 pathways in the regulation of CO2 sensing, filamentation, and virulence in Candida albicans

city: Beijing

first: guanghua

last: huang

address: NO.1 West Beichen Road, Chaoyang District

institute: Institute of Microbiology

summary: Candida albicans is the most common opportunistic fungal pathogen of humans and is also a benign member of the gastrointestinal (GI) tract microbiota. Morphological transitions and metabolic regulation are critical for C. albicans to adapt to the changing host environment. We generated a library of central metabolic pathway mutants in the tricarboxylic acid (TCA) cycle, and investigated the functional consequences of these gene deletions on C. albicans biology. Inactivation of the TCA cycle impairs the ability of C. ablicans to utilize non-fermentable carbon sources and dramatically attenuates cell growth rates under several culture conditions. Through integrations with the Ras1-cAMP  signaling pathway and the heat shock factor-type transcription regulator Sfl2, we found that the TCA cycle plays fundamental roles in the regulation of CO2 sensing and filamentation. The TCA cycle and cAMP signaling pathways form a regulatory feedback loop, in which ATP and CO2 may function as molecular linkers. We further demonstrate that inactivation of the TCA cycle leads to lowered intracellular ATP and cAMP levels and thus affects the activation of the Ras1-regulated cAMP signaling pathway. In turn, the Ras1-cAMP signaling pathway controls the TCA cycle through both Efg1- and Sfl2-mediated transcriptional regulation in response to elevated CO2 levels. The protein kinase A (PKA) catalytic subunit Tpk1, but not Tpk2, may play a major role in this regulation. Sfl2 specifically binds to several TCA cycle and filamentation-associated genes under high CO2 conditions. Global transcriptional profiling experiments indicate that Sfl2 is indeed required for the gene expression changes occurring in response to these elevated CO2 levels. Finally, we also demonstrate that several key genes of the TCA cycle are essential for virulence and successful colonization of the GI tract in two mouse infection models.

overall_design: total RNA profiles of 22h old WT or sfl2/sfl2 strains grown on YPD plates in air or in 5% CO2.

first: Guanghua

last: Huang

first: Li

last: Tao

first: Yulong


title: Role of N-a-acetyltransferase 10 Protein in DNA Methylation and Genomic Imprinting

city: Taipei

first: Li-Jung

last: Juan

address: 128 Academia Road, Section 2, Nankang

department: Genomic Research Center

country: Taiwan

institute: Academia Sinica

summary: We report that Naa10-null mice display partial embryonic lethality, growth retardation, brain disorder and maternal-effect lethality, phenotypes commonly observed in defective genomic imprinting. Genome-wide analyses further reveal enriched dysregulation of imprinted genes in Naa10p-knockout ES cells.

overall_design: mRNA profiles of 2 biological ESC replicates of each WT and Naa10-KO group

first: Chen-Cheng

last: Lee

first: Shih-Huan

last: Peng


title: Global analysis of AGO2-bound RNAs in teratoma-derived fibroblasts

city: Seoul

first: Young-Soo

last: Kwon

address: Nungdongro 209, Gwangjingu

department: Department of Bioscience and Biotechnology

country: South Korea

institute: Sejong University

summary: Among the four Argonaute family members in mammals, only AGO2 protein retains endonuclease activity, and facilitates cleavage of target RNAs base-pairing with highly complementary guide RNAs. Despite the deeply conserved catalytic activity, only a small number of targets have been reported to extensively base pair with cognate miRNAs to be cleaved by AGO2. Here, we analyzed AGO2-bound RNAs by CrossLinking ImmunoPrecipitation (CLIP) of genetically modified cells that express epitope-tagged AGO2 from the native genomic locus. We found that HMGA2 mRNA is cleaved by AGO2 loaded with let-7 and miR-21. In contrast to the generally accepted notion, the base-pairing from the seed region to the cleavage site, rather than perfect or near perfect complementarity, was required for cleavage of the target mRNA in cells. Non-templated addition of nucleotides at the 3’ end of the cleaved RNA was observed, further supporting the AGO2-mediated cleavage. Based on the observation that the limited complementarity is the minimum requirement for cleavage, we found that AGO2-mediated cleavage of targets is more common than previously thought. Our result may explain the vital role of endonuclease activity in controlling miRNA-mediated gene regulation.

overall_design: CLIP-seq of FLAG-AGO2 by using an anti-FLAG-antibody (Wako) and IgG controls.

first: Young

middle: S


title: Genomic profiling of human spermatogonial stem cells

first: Jingtao

last: Guo

overall_design: Refer to individual Series


title: Genomic profiling of human spermatogonial stem cells [WGBS]

summary: To better understand human spermatogonial stem cells (SSCs), we profiled their transciptome and epigenome, which revealed the mechanism how human SSCs regulates their self-renewal versus differentiation dermination, as well as how latent pluripotency is established in human SSCs. Remarkly, we discovered signaling pathways (e.g. LIF, BMP, WNT) that differentially regulated self-renewal vesus differentiation in SSCs. We also discovered that SSCs repress core pluripotent factors (Sox2, Pou5f1 and Nanog) yet activate ancillary factors (e.g. Klf4, Mbd3, Tcf3, Sall4) transcriptionally and epigenetically.

overall_design: Using SSEA4+ as self-renewal marker, we isolated self-renewal SSCs by magnetic antibody cell sorting (MACS). Genomic DNA were extracted from isolated cells, and bisulfite converted. Sequencing libraries were then prepared and sequenced on a 125-cycle paired-end run on an Illumina HiSeq 2500 instrument.

first: Ed

last: Grow

first: Chongil

last: Yi

first: James

middle: M

last: Hotaling

first: Bradley

middle: R

last: Cairns


title: Genomic profiling of human spermatogonial stem cells [BulkRNA-Seq]

overall_design: Using SSEA4 as self-renewal marker and Kit as differentiating marker, we isolated self-renewal and differentiation SSCs by magnetic antibody cell sorting (MACS). Total RNA were extracted from those populations, and standard RNA sequencing libraries were prepared for sequnecing on a 50-cycle single end run.


title: High-resolution gene expression datasets of ontogenetic zones in the root apical meristem

status: Public on Oct 03 2017

city: Gent

first: Jos

last: Wendrich

address: Technologiepark 71

department: PSB

country: Belgium

institute: VIB-UGent

summary: In plants, apical meristems allow continuous growth along the body axis. Within the root apical meristem (RAM), a group of slowly dividing quiescent center (QC) cells is thought to limit stem cell activity to directly neighboring cells (Cowels, 1956; van den Berg et al., 1997), thus endowing them with unique properties, distinct from displaced daughters. This binary identity of the stem cells stands in apparent contradiction with the more gradual changes in cell division potential (Bennett and Scheres, 2010) and differentiation (Yamaguchi et al., 2008; 2010; Furuta et al, 2014; Geldner, 2013; Masucci et al., 1996; Dolan and Costa, 2001) that occur as cells move further away from the QC. To address this paradox and to infer molecular organization of the root meristem, we used a whole-genome approach to determine dominant transcriptional patterns along root ontogeny zones. We found that the prevalent patterns are expressed in two opposing gradients. One is characterized by genes associated with development, the other enriched in differentiation genes. We confirmed these transcript gradients, and demonstrate that these translate to gradients in protein accumulation and gradual changes in cellular properties. We also show that gradients are genetically controlled through multiple pathways. Based on these findings, we propose that cells in the Arabidopsis root meristem gradually transition from ‘stemness’ towards differentiation.

overall_design: This study contains high-resolution datasets from cell populations from the enitre root meristem and xylem-specific cell populations. Using fluorescence activated cell sorting, three cell populations were isolated based on their GFP expression intensity. Two-Three replicates were used per sample

first: Dolf

last: Weijers


title: Transcriptome Comparison of phoU Homologies: Genes Deletion strain and the Parent Strain in stationary phase (10 h)

first: Xiaofei

last: Wang

address: Room 1103, Zhidao building, Dong'an Road, Xuhui district, Shanghai

department: Shanghai Medical College

institute: Fudan University

summary:
Purpose: To compare the transcriptome changes in phoU1(serp0956) or phoU2(serp0316)deletion strain with the parent strain SE1457 in stationary phase (10 h).

Methods: Total RNA was isolated and sequenced from the phoU1 deletion strain, phoU2 deletion strain and the parent strain SE1457 in stationary phase (10 h) in triplicate using an illumina high-seq 2500. Raw data was analyzed using TopHat. Genes were considered changed  which performed fold-change>=1.5 and FDR<=0.05. Changed genes were then filtered to reveal the downstream targets of phoU1 or phoU2.

Results: The expression of 2 genes were significantly decreased and no genes was significantly increased in expression level due to the loss of phoU1 expression.The expression of 279 genes were significantly decreased and 716 genes were significantly increased in expression level due to the loss of phoU2 expression.

overall_design: Total three biological samples are analyzied, each of them had triplication.The three samples are parent strain SE1457,the phoU1 deletion strain and phoU2 deletion strain.

first: Di

last: Qu


title: Transcriptome Comparison of phoU Homologies Genes Deletion strain  and the Parent Strain in log-phase (6 h).

summary:
Purpose: To compare the transcriptome changes in phoU1(serp0956) or phoU2(serp0316)deletion strain with the parent strain SE1457  in log-phase (6 h)
Methods: Total RNA was isolated and sequenced from the phoU1 deletion strain, phoU2 deletion strain and the parent strain SE1457  in log-phase (6 h) in triplicate using an illumina high-seq 2500. Raw data was analyzed using TopHat. Genes were considered changed  which performed fold-change>=1.5 and FDR<=0.05. Changed genes were then filtered to reveal the downstream targets of phoU1 or phoU2 in log-phase (6 h).
Results: The expression of 23 genes were significantly decreased and 69 genes was significantly increased in expression level due to the loss of phoU1 expression.The expression of 474 genes were significantly decreased and 471 genes were significantly increased in expression level due to the loss of phoU2 expression.


title: The zinc transporter ZIP10 regulates epidermal morphogenesis

city: Pohang

first: Bhin

last: Jin-hyuk

address: Ji-gok research center, hyoja-dong, Nam-gu

department: Department of Chemical engineering

institute: POSTECH

summary: We performed an mRNA-sequencing experiment using ZIP10 positive and negative cells isolated from the ventral skins of WT mice and analyzed gene expression profiles of those cells to identify the functional differences between the two cell types.

overall_design: ZIP10 positive and negative cells from the ventral skins of one-week-old WT mice were sorted by flow cytometry. From those cells, we isolated mRNAs and analyzed gene expression profiles by mRNA sequencing.

first: Bum-Ho

last: Bin

first: Toshiyuki

last: Fukada

first: Jinhyuk

last: Bhin


title: Spontaneous generation of a novel fetal human retinal pigment epithelium (RPE) cell line available for investigation on phagocytosis and morphogenesis

first: Zhihua

last: Shao

address: No.1239, Siping Road

institute: Tongji University

summary:
Purpose: Primary retinal pigment epithelium (RPE) cells have a limited capacity to re-establish epithelial morphology and to remain native RPE function in vitro, and all commercially available RPE cell lines have drawbacks of morphology or function; therefore, the establishment of new RPE cell lines with typical characteristics of RPE would be helpful in further understanding of their physiological and pathological mechanism. Here, we firstly report a new spontaneously generated RPE line, fhRPE-13A, from a 13-week aborted foetus. We aimed to investigate its availability as a RPE model.

Methods:RNA-seq data were mapped to the human genome assembly hg19. Global transcriptional data were analyzed by Weighted Gene Co-expression Network Analysis (WGCNA) and differentially expressed genes(DEGs) to identify gene expression profiles. The morphology and molecular characteristics were examined by immunofluorescence, transmission electron micrographs, PCR and western blot. Photoreceptor outer segments (POS) phagocytosis assay and transepithelial resistance measurement (TER) were performed to assess phagocytic activity and barrier function, respectively.

Results: The fhRPE-13A cells showed typical polygonal morphology and the normal biological processes of RPE, had ability of POS phagocytosis in vitro, and expressed TYR and TYRP1 significantly higher than that in ARPE-19 cells.

Conclusions: The fetal human RPE line fhRPE-13A is a valuable system for researching phagocytosis function and morphogenesis of RPE in vitro.

overall_design: mRNA profiles of RPE cell lines and passaged primary RPE at different stage were generated by deep sequencing using Illumina HiSeq2000.

first: Zhixue

last: Liu

first: Siguang

last: Li


title: Transcriptome Data of KLF4 Overexpression in Human Umbilical Vein Endothelial Cells

first: John

last: Shyy

department: Medicine

summary: Atheroprotective flow (e.g., pulsatile shear stress) and statins drastically induce the expression of krüppel-like factor 4 (KLF4) in vascular endothelial cells (ECs).  We therefore investigated the role of KLF4 in EC function through comparing the transciptional profiling of human umbilical vein endothelial cells (HUVECs) that were infected with adenovirus overexpression KLF4 (Ad-KLF4) or with empty vector (Ad-null) control.  Gene ontology analysis revealed that KLF4 not only regulates EC homeostasis through the upregulation of nitric oxide synthesis and vascular development, but also mediates response to lipid.  Among the lipid responsive genes, KLF4 exhibited induction of cholesterol efflux and oxidation [i.e., liver X receptor (LXR) and cholesterol 25-hydroxylase (Ch25h)], while suppressing cholesterol biosynthesis [i.e., sterol regulatory element-binding protein 2 (SREBP2)].

overall_design: Transcriptome data of HUVECs overexpressing KLF4 using adenovirus followed by deep sequencing, in biological duplicate, using Illumina NextSeq 500.

first: Zhao

first: Marcy

last: Martin


title: Genome-wide transcriptional profiling of YAP target genes in human endothelial cells

city: Rochester

first: Suowen

last: Xu

address: 603 Elmwood Avenue

institute: University of Rochester

summary: Endothelial cell is the major cell type that senses and transduces mechanosignal generated by shear stress. We have recently shown that Hippo/YAP pathway is a mechanosensitive pathway that is critical for maintaining endothelial cell homeostasis. However, the transcritpional targets and biological functions of YAP in endothelial cells remain largely unknown. To evaluate YAP-dependent gene expression in endothelial cells, we performed RNA-sequencing in YAP depleted (by transfection with by YAP siRNA) and overexpressed (by infection with YAP-S127A catalytically active adenovirus) human endothelial cells. We observed that YAP critically regulates endothelial function by modulating multiple atherosclerosis-related genes. Our study provides mechanistic insights into the question how YAP regulates endothelial function and atherosclerosis by modulating endothelial transcriptional profile.

overall_design: Human Umbilical Vein Endothelial Cells (HUVEC, passage 3 to passage 5) were seeded in 0.2% gelatin-coated dishes the day before experiment. The next day, 80% confluent HUVECs were transfected with non-targetting siRNA pool (siNC, GE Dharmacon, #D-001810-10)  or YAP1 siRNA pool (siYAP1, GE Dharmacon, #M-012200) for 48 hours. In the overexpression experiment, 80% confluent HUVECs were infected with control adenovirus (Ad-NC) or YAP-S127A catalytically active adenovirus (Ad-YAP-S127A, gifted by Prof. Junichi Sadoshima, New Jersey Medical School, NJ) for 48 hours. After treatment, total RNA was isolated using the RNA-Easy Mini Plus kit (QIAGEN). High quality RNA samples (pre-assessed by Nanodrop measurements) were further processed in the Genome Research Center of the University of Rochester Medical Center. The TruSeq RNA Sample Preparation Kit V2 (Illumina, San Diego, CA) was used for next generation sequencing library construction per manufacturer’s protocols. Briefly, mRNA was purified from 100ng total RNA with oligo-dT magnetic beads and fragmented. First-strand cDNA synthesis was performed with random hexamer priming followed by second-strand cDNA synthesis. End repair and 3’ adenylation was then performed on the double stranded cDNA. Illumina adaptors were ligated to both ends of the cDNA, purified by gel electrophoresis and amplified with PCR primers specific to the adaptor sequences to generate amplicons of approximately 200-500bp in size. The amplified libraries were hybridized to the Illumina single end flow cell and amplified using the cBot (Illumina, San Diego, CA) at a concentration of 8pM per lane. Single end reads of 100nt are generated for each sample and aligned to the organism specific reference genome. Sequenced reads were cleaned using Trimmomatic-0.32 before mapping some of to the human reference genome (GRCh38.p2) with STAR-2.4.2a. Raw read counts were obtained using HTSeq and gencode 23 human gene annotations. DESeq2-1.10.1 was used to perform data normalization and differential expression analysis with an adjusted p-value threshold of 0.05. Then, Cufflinks-2.0.2 Software was used with the gencode 23 human gene annotations to perform differential expression analysis with an FDR cutoff of 0.05.

first: Zheng

last: Jin

last: Ashton

first: Jason

last: Myers


title: RSPO1 downregulation and a CFA9 Insertion are Associated with Inherited XX DSD in a Canine Model

status: Public on Oct 02 2017

city: Ithaca

first: Charles

last: Danko

address: Hungerford Hill Rd

department: Baker Institute

institute: Cornell University

summary: Remarkable progress has been achieved in understanding the mechanisms controlling sex determination, yet the cause for many Disorders of Sexual Development remains unknown. Of particular interest is a rare XX DSD subtype in which individuals are negative for SRY, the testis determining factor on the Y chromosome, yet develop testes or ovotestes, and both phenotypes occur in the same family. This is a naturally occurring disorder in humans and dogs (Canis lupus familiaris), and phenotypes in the canine XX DSD model are strikingly similar to those in this type of human XX DSD. The purposes of this study were to identify 1) a variant associated with XX DSD in the canine model and 2) gene expression alterations in canine embryonic gonads that could be informative to causation. Using a genome wide association study (GWAS) and whole genome sequencing (WGS), we identified a variant on Canis familiaris autosome 9 (CFA9) that is significantly associated with XX DSD in the canine model and in affected purebred dogs. This is the first marker and candidate mutation identified for inherited canine XX DSD, and it lies within the canine ortholog for the human disorder (OMIM 278850). Inheritance of the variant indicates that XX DSD is a complex trait in which breed genetic background affects penetrance. Furthermore, the homozygous variant genotype is associated with embryonic lethality in at least one breed. Gene expression studies (RNA-seq and PRO-seq) in embryonic gonads at risk of XX DSD from the canine model identified significant RSPO1 downregulation without significant upregulation of SOX9 or other known testis pathway genes. A novel mechanism is proposed in which the canine XX DSD mutation acts upstream of RSPO1 to induce epigenomic gonadal mosaicism.

overall_design: PRO-seq was used to analyze eight canine gonad samples isolated from day 39 embryos.

first: Vicki

last: Meyers Wallen


title: Impact of control interventions on lung gene expression in post-natal lung development

city: Bad Nauheim

first: Rory

last: Morty

address: Parkstr. 1

department: W.G. Kerckhoff-Institut

country: Germany

institute: MPI für Herz- und Lungenforschung

summary: During the studies of post-natal lung development in the laboratory C57BL/6 mouse, broad spectrums of interventions are applied by intraperitoneal (i.p.) injections. We explored whether injection of saline, tamoxifen, “scrambled” antagomiRs (to control studies that target microRNA with antagomiRs), migloyl or water, in well-established protocols, had any impact on the post‑natal gene expression in mouse lungs.

overall_design: Pups were randomized on the day of birth and litters were balanced. Saline (daily treatment) group was treated via i.p. injection (25µl/g) daily from the postnatal day (P)1 to P13. Scramble antagomiR controls: Scrambled 1 (Custom miRCURY™  LNA Inhibitor probe, batch number 184196, Exiqon) and Scrambled 2 (Custom miRCURY™ LNA Inhibitor probe, New scramble control design, Exiqon) were applied at P1 and P3, by i.p. injection (10 mg/kg) dissolved in nuclease-Free Water (Ambion). Nuclease-Free Water (Ambion) was applied via i.p injection at P1 and P3 at (10µl/g). At P14 mice from Saline (daily treatment), Scrambled 1, Scrambled 2, Water and Untreated groups were sacrificed. Tamoxifen (Sigma-Aldrich) was applied at P1 and P2, by i.p. injection (0.4 mg; 25 µl) dissolved in Miglyol. Migloyl was applied at P1 and P2 at 25 µl via i.p. injection. Saline (treatment day 1+2) was applied at P1 and P2 at 25 µl via i.p. route. At P6 mice from Tamoxifen, Migloyl and Saline (treatment day 1+2) treatment groups were sacrificed.

first: Agnieszka

last: Pozarska

first: Joshua

last: Fehl

middle: E


title: Characterization of small RNAs expressed in roots and galls from Arabidopsis thaliana induced by the plant parasitic nematodes Meloidogyne incognita

city: Valbonne - Sophia-Antipolis

first: Kevin

last: Lebrigand

address: 660 route des lucioles

country: France

institute: IPMC/CNRS

summary: By sequencing small RNAs from uninfected Arabidopsis roots and from galls seven and 14 days post infection with Meloidogyne incognita, we sequenced by SOLiD technology the RNA fraction below 50nt. We identified 24 miRNAs differentially expressed in gall as putative regulators of gall development.

sample_organism:
Arabidopsis thaliana
Meloidogyne incognita

overall_design: Uninfected Arabidopsis roots and galls seven and 14 days post infection with Meloidogyne incognita (3 replicates)

first: C

last: Medina

first: K

first: M

last: Da Rocha

first: B

last: Favery

first: S

last: Jaubert-Possamai


title: Identification of differential gene expression in WT and DGKzeta deficient CD8 T cells by RNA-seq

city: Milwaukee

first: Sridhar

last: Rao

address: 8727 Watertown Plank Road

department: Blood Research Institute

institute: BloodCenter of Wisconsin

summary:
Purpose: The goal of this study is to identify changes in gene expression between wild type and DGKzeta deficient splenic CD8+ T cells upon sitmulation with anti-CD3 and anti-CD28.
Methods: mRNA profiles from WT and DGKzeta deficient CD8 T cells were generated.

overall_design: Comparison of stimulated splenic CD8 T cells from WT and DGKzeta deficient mice.

first: Matthew

last: Riese

first: Kirthi

last: Pulakanti