title: Genome-wide maps of the histone H3K4 demethylase Jhd2 occupany in Saccharomyces cerevisiae

city: Taipei

first: Wan-Sheng

last: Lo

state: Taiwan

address: 128 Academia Road, Section 2, Nankang

department: IPMB

country: Taiwan

institute: Academia Sinica

summary: By using of paired-end sequencing technology, we report the high-throughput profiling of Jhd2 targeting in S. cerevisiae. We obtained more than 1.8E+7 Illumina reads and generated 1.1E+7 mapped reads from Jhd2-ChIP DNA .  Sequence reads for Jhd2-ChIP and Input DNA were mapped to yeast genome by using the BWA (Burrows Wheeler Aligner), SAMtools (Sequence Alignment Map) programs and followed by SICER (Spatial clustering for Identification of ChIP-Enriched Regions) program to obtain the Significant peaks. We found that the overall Jhd2 distribution across the gene body of targets is fairly correlated to the levels of H3K4 di- and tri-methylation, which are enriched at 5’ through 3’ of ORF regions. These observations demonstrate that the Jhd2, a histone H3K4 demethylase, is linked to gene transcription by locating mainly at the coding regions to balance the H3K4me of its target genes at steady state. Furthermore, we observed that Jhd2 are significantly enriched at several specific chromosomal loci including telomeres, centromeres, silent HM loci, LTR-tRNAs, and rDNA arrays.  Our study demonstrated that Jhd2, H3K4-demethylase, plays a dynamic role as a chromatin insulator to define boundary regions between euchromatin and heterochromatin by association with specific chromatin loci in the yeast genome.

overall_design: Input DNA and ChIP DNA from immunopurified Jhd2-Flag were subjected to paired-end sequencing

first: Chia-Yi

last: Chien


title: Dissection of functionally relevant Wnt/beta-catenin target genes in an organ culture model of Wilms tumor

status: Public on Feb 20 2017

city: New York

first: Emily

last: Hsu

state: NY

address: 1130 St. Nicholas Ave.

department: ICG

country: USA

institute: Columbia University

type: Expression profiling by high throughput sequencing

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

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Refer to individual Series


title: RNA sequencing data from beta-catenin activated, E2-2 wildtype and beta-catenin activated, E2-2 deleted cultured fetal mouse kidneys.

summary: Oncogenic stabilizing mutations in the beta-catenin gene CTNNB1 are common in one class of human Wilms tumors. To model these tumors, we crossed mice carrying an inducible Cre recombinase (Rosa26-CreERT2) with a conditional gain-of-function beta-catenin allele (Ctnnb1Ex3flox). We obtained kidneys at embryonic day 13.5 and placed them in organ culture with tamoxifen. Multiple small tumors formed in the Ctnnb1∆Ex3 kidneys while the control kidneys developed normally. Immunostaining for Sall1, a marker of condensing metanephric mesenchyme, indicated that the tumors arise from this tissue compartment, analogous to the origin of human Wilms tumors. Expression profiling identified a set of induced genes overlapping with those up-regulated in human Wilms tumors with CTNNB1 mutations, including genes encoding feedback inhibitors of Wnt/beta-catenin signaling, the epithelial-mesenchymal transition (EMT) inducers, and the transcription factor E2-2 (ITF2/TCF4). A conditional deletion of E2-2 showed that this gene is important for growth of the nephroblastomas, in part via promoting an epithelial to mesenchymal transition downstream of beta-catenin. Given these cross-species validations, and genetic proof of the oncogenic function of a key downstream target gene, we expect that this rapid and efficient organ culture model will be useful for further studies to dissect and inhibit oncogenic Wnt/beta-catenin signaling.

overall_design: The expression of 4 beta-catenin activated, E2-2 wildtype cultured fetal mouse kidneys was compared with 3 beta-catenin activated, E2-2 deleted cultured fetal mouse kidneys.

first: Iveta

last: Yotova

first: Benjamin

last: Tycko


title: Early transcriptional and epigenetic regulation of CD8+ T cell differentiation revealed by single-cell RNA-seq

city: La Jolla

first: Gene

last: Yeo

state: CA

address: 2880 Torrey Pines Scenic Dr. Room 3805/Yeo Lab

institute: UCSD

type: Genome binding/occupancy profiling by high throughput sequencing

summary: During microbial infection, responding CD8+ T lymphocytes differentiate into heterogeneous subsets that together provide immediate and durable protection. To elucidate the dynamic transcriptional changes that underlie this process, we applied a single-cell RNA sequencing approach and analyzed individual CD8+ T lymphocytes sequentially throughout the course of a viral infection in vivo. Our analyses revealed a striking transcriptional divergence among cells that had undergone their first division and identified previously unknown molecular determinants controlling CD8+ T lymphocyte fate specification, including Ezh2, the catalytic component of the Polycomb Repressive Complex 2. Our data provide a revised model of terminal effector cell differentiation initiated by an early burst of transcriptional activity and subsequently refined by epigenetic silencing of transcripts associated with memory lymphocytes. These findings provide unexpected insights into tightly coupled transcriptional and epigenetic mechanisms underlying CD8+ T lymphocyte fate specification and highlight the power and necessity of single-cell approaches.

overall_design: Gene expression profiles for key stages of T cell differentiation were derived from 256 single-cell RNAseq libraries prepared on Fluidigm C1 and sequenced on Illumina HiSeq2500 (with 224 unique sequencing samples and 64 duplicates).  Epigenetic profiles of Ezh2 binding, its associated H3K27me3 mark, and PolII occupancy in invitro-activated T cells were derived from ChIP libraries prepared with NEB Next ChIP (4 Ezh2 fl/fl and 4 WT samples, each with a biological duplicate).  These ChIPseq libraries and their matched IGG input DNA controls were sequenced on Illumina HiSeq 4000 together with bulk RNAseq libraries from T cells invivo-activated in either C57BL/6 wild-type or Ezh2-knockout mice (each with a biological duplicate).

first: Boyko

last: Kakaradov

first: Janilyn

last: Arsenio

first: Christella

middle: E

last: Widjaja

first: Zhaoren

last: He

first: Stefan

last: Aigner

first: Patrick

middle: J

last: Metz

first: Bingfei

last: Yu

first: Ellen

last: Wehrens

first: Justine

last: Lopez

first: Stephanie

middle: H

last: Kim

first: Elina

middle: I

last: Zuniga

first: Ananda

middle: W

last: Goldrath

first: John

middle: T

last: Chang


title: Polycomb-mediated deactivation of AR-feedback loop leads to castration-resistant prostate cancer [ChIP-seq]

status: Public on Feb 19 2017

city: Atlanta

first: Jindan

state: GA

address: E330, 1760 Haygood Dr NE

department: Urology

institute: Emory University

summary: Castration-resistant prostate cancer (CRPC) is a lethal disease, often characterized by aberrant  androgen receptor (AR) activation independently of androgen. We have previously reported that  AR can directly repress the expression of many target genes, one of which is NOV/CCN3. Here  we show that NOV, primarily localized in the cytoplasm, physically interacts with AR at its Nterminus  and sequesters AR and AR variants in the cytoplasm, thereby reducing AR chromatintargeting.  This negative feedback loop, however, is disrupted in CRPC due to epigenetic silencing of NOV by the   Polycomb group protein EZH2, rendering AR transcriptional activities and drug-resistant prostate cancer   progression. Our findings thus suggest a working model wherein AR-repressed genes critically prevent   CRPC through negative feedback loops inhibiting AR signaling.

overall_design: ChIP_Seq examination of AR and NOV binding sites in LNCaP cells

first: Ka-wing

last: Fong

first: Jonathan

last: Zhao


title: Polycomb-mediated deactivation of AR-feedback loop leads to castration-resistant prostate cancer


title: The complex transcriptional response of Acaryochloris marina to different oxygen levels

city: Sydney

first: Miguel

middle: A

last: Hernández-Prieto

state: NSW

address: Macleay Building A12

department: School of life and environmental sciences

country: Australia

institute: University of Sydney

summary: Ancient oxygenic photosynthetic prokaryotes produced oxygen as a waste product, but existed for a long time under an oxygen-free (anoxic) atmosphere, before an oxic atmosphere emerged. The change in oxygen levels in the atmosphere influenced the chemistry and structure of many enzymes that contained prosthetic groups that were inactivated by oxygen.   In the genome of Acaryochloris marina, multiple gene copies exist for proteins that are normally encoded by a single gene copy in other cyanobacteria. Using high throughput RNA sequencing to profile transcriptome responses from cells grown under microoxic and hyperoxic conditions, we detected 8446 transcripts out of the 8462 annotated genes in the Cyanobase database. Two thirds of the 50 most abundant transcripts are key proteins in photosynthesis. Microoxic conditions negatively affected the levels of expression of genes encoding photosynthetic complexes, with the exception of some subunits. In addition to the known regulation of the multiple copies of psbA, we detected a similar transcriptional pattern for psbJ and psbU, which might play a key role in the altered components of photosystem II. Furthermore, regulation of genes encoding proteins important for reactive oxygen species-scavenging is discussed at genome level, including, for the first time, specific small RNAs having possible regulatory roles under varying oxygen levels.

overall_design: Acaryochloris marina MBIC11017 was routinely kept in a culture room at 27°C under 15–30 µmol photons•m˗2•s˗1 of cool white light. Sterilized K+ES (artificial seawater), buffered with 25 mM TES at pH 8.0 was used as the culture medium for all three treatment groups. To make sure photosynthesis was not limited by CO2, NaHCO3 was dissolved in a small volume of autoclaved media, and injected into the enclosed culture flasks every two days (yielding an initial concentration of 0.375 mM). The initial cell density of all culture groups was adjusted to an optical density at 750 nm of 0.2. The cultures were shaken on an orbital flat-bed shaker at ~90 rpm.  Cultures under normal O2 levels were inoculated in 1-l Erlenmeyer glass flask containing 500 ml of medium, capped with a cotton stopper that permitted gas exchange. Thus, the O2 concentration of the gas phase inside the bottle was similar to atmospheric levels ~21% (v/v).  Microoxic conditions were achieved by using a 500-ml two-necked round-bottom flask sealed tightly by a rubber stopper. Cultures were vacuumed, and refilled with pure nitrogen gas (99.95% purity) to ensure normal atmospheric pressure. We repeated this process several times, yielding a final O2 concentration of <0.2% inside the sealed culture flask. To maintain a microoxic condition, a positive pressure was created by bubbling nitrogen through the culture.  A similar set-up was used for hyperoxic conditions, with the exception that pure O2 gas was used to refill the flask after vacuuming. Because the cells generate O2 under illumination, ongoing input of O2 gas to maintain the high-oxygen concentration was not required. However, to avoid pressure build-up, the flask was re-vacuumed and refilled with O2 gas every 48 h, as described above. The O2 concentration inside the flask remained within the range of 65–75% (v/v) during the experiment.  Acaryochloris cultures were harvested after five days from all three different treatments. The experiment was triplicated under ambient conditions, and duplicated under microoxis and hyperoxis conditions. The harvested cell pellets were mixed with TRIzol (TRIzol® Reagent, Life Technologies, Australia), and frozen immediately using liquid nitrogen. They were stored at ˗80°C for at least 60 min. The frozen samples were thawed in a water bath of 37°C, and spun down at 16,000×g for 5 min to eliminate cellular debris. This supernatant was mixed at a volume ratio of 4:1, with chloroform and spun down at 16,000×g for 10 min at 4°C. The upper layer, containing RNA and DNA, was carefully transferred to a new tube, without disturbing the white middle layer of solid components. The RNA was precipitated by addition of an equal volume of isopropanol and incubated at ˗20°C for at least 45 min. RNA pellets were washed with 70% (v/v) ethanol and the DNA removed using the Baseline-ZERO™ DNase kit (Epicentre, WI, USA), following the manufacturer's instructions. Prior to RNA quality assessment, the absence of DNA was confirmed by polymerase chain reaction (data not shown).  The quality of RNA was assessed on a 2100 Bioanalyzer (Agilent Technologies, CA, USA), using a RNA 6000 Nano RNA Kit (Agilent Technologies) to obtain an RNA integrity number of >8.0. Transcripts corresponding to rRNAs (5S, 16S and 23S) were reduced from the samples with a Ribo-Zero Kit (Epicentre), following the manufacturer's instructions. Further processing, including quality assessment, was undertaken prior to RNAseq analysis by Beijing Genomics Institution, China.

first: Min

last: Chen


title: Genome-wide identification of glucocorticoid receptor binding sites in the whole hippocampus of adrenalectomised male rats given corticosterone infusion and stressed or non-stressed.

city: Bristol

middle: Robert

last: Pooley

address: Dorothy Hodgkin Building, Whitson Street

department: Henry Wellcome Laboratories for Integrated Neuroscience and Endocrinology

country: United Kingdom

institute: University of Bristol

summary: We have applied next-generation sequencing technology to obtain high through-put profiling of glucocorticoid receptor (GR) DNA binding in whole hippocampus from male rat brain. By obtaining sequence from immunoprecipitated, sonicated chromatin, we have generated genome-wide binding maps for GR in two contexts, testing whether chromatin is primed for GR binding according to the animal's acute experience. In one context (stressed) adrenalectomised rats are infused with corticosterone while undergoing restraint stress. In the second context (non-stressed control) rats are infused with the same corticosterone profile but restraint stress is omitted. We find that GR binding was highly similar suggesting factors other than GR binding may account for any context-dependent alterations in GR function associated with acute stress. We show that GR is positioned near to genes associated with structural and functional organisation of the hippocampus at largely intergenic and intronic sites. Glucocorticoid response element sequences support a majority of GR binding in this tissue but may be reinforced by NF-1 and/or basic-helix loop helix transcription factors binding their nearby recognition motifs. We found little evidence for tethering of GR to other factors, or for the usage of negative glucocorticoid response elements.

overall_design: Examination of glucocorticoid receptor binding in two different physiological contexts (stressed and non-stressed controls).

middle: R

first: Becky

middle: L

last: Conway-Campbell


title: Identification of methylation haplotype blocks aids in deconvolution of heterogeneous tissue samples and tissue-of-origin mapping from plasma DNA

first: Kun

last: Zhang

address: 9500 Gilman Dr Mailcode: 0412

department: Bioengineering

institute: University of California, San Diego

type: Methylation profiling by high throughput sequencing


title: Identification of methylation haplotype blocks aids in deconvolution of heterogeneous tissue samples and tissue-of-origin mapping from plasma DNA [Plasma RRBS]

summary: Adjacent CpG sites in mammalian genomes tend to be co-methylated due to the processivity of enzymes responsible for adding or removing the methyl group. Yet discordant methylation patterns have also be observed, and found to be related to stochastic or uncoordinated molecular processes. Here we focused on a systematic search and investigation of regions in the human genome that exhibit highly coordinated methylation.   By examining the co-methylation patterns of multiple adjacent CpG sites, termed methylation haplotypes, in single bisulfite sequencing reads, we applied a greedy-searching strategy to defined blocks of tightly coupled CpG sites, called Methylation Haplotype Blocks (MHBs), based on 53 sets of whole genome bisulfite sequencing (WGBS) data, including 43 published sets from human adult tissues, ESC and in vitro differentiated cell lines, as well as 10 sets from human adult tissues generated in this study. The MHBs were then further validated with 101 sets of RRBS ENCODE data, and 637 sets of Illumina 450k methylation array data from TCGA tumor and normal samples. Globally, MHBs are enriched in but only partially overlap with several well-known genomic features, including CpG islands, promoters, enhancers and VMRs.  To perform quantitative analysis of the MHBs, we defined a metric called Methylation Haplotype Load (MHL), which is covered both average methylation level and methylation complexity and therefore more informative than average methylation level or Shannon entropy. Using a feature selection strategy, we identified a set of tissue-specific MHBs that cluster by developmental germ-layers. Interestingly, examination of these MHBs revealed two distinct mechanisms for fate commitment during development: epigenetic silencing of pluripotent genes, such as NANOG, for mesoderm induction; and epigenetic induction (or de-suppression) of lineage-specific factors for ectoderm commitment.

overall_design: Examination of co-methylation in human cfDNA

first: Shicheng

last: Guo

first: Dinh

last: Diep


title: Identification the target genes of microRNA in Ammopiptanthus mongolicus

city: Beijing

first: Fei

last: Gao

address: 27 Zhongguancun South Street

country: China

institute: Minzu University of China

summary: Ammopiptanthus mongolicus is an ecologically important desert plant. miRNA are endogenous 20-22 nt small RNAs that play important roles in regulating gene expression in plants. To identify to the target genes of miRNA in Ammopiptanthus mongolicus, degradome sequencing was performed and a batch of transcripts were found to be targeted by miRNA. The result is helpful in understanding the roles on miRNA in plant development and environmental stress response.

overall_design: A degradome sequencing library was constructed using pooled mRNA isolated from leaves and roots of A. mongolicus seedlings. The library was sequenced using Illumina Hiseq 2500 platform.

first: Yijun

last: Zhou


title: Identification of methylation haplotype blocks aids in deconvolution of heterogeneous tissue samples and tissue-of-origin mapping from plasma DNA [Tumor RRBS]

overall_design: Examination of co-methylation in primary tumor tissues


title: Immunological Imbalance in Fibrodysplasia Ossificans Progressiva Revealed by PBMC Transcriptome Analysis

status: Public on Feb 18 2017

city: Shanghai

first: Kunshan

address: 389 Xincun Road

department: Stem cell Translational Research center

institute: Tongji Hospital, Tongji University

summary: In order to study the inflammation mechanism of FOP patients’ peripheral blood system, we profiled the transcriptome of peripheral blood mononuclear cells (PBMC) from 3 FOP patients as well as 3 healthy people. After comparing the PBMC transcriptomes between FOP patients and healthy control, we identified 335 differentially expressed genes from FOP patients, including 135 significantly up-regulated and 200 significantly down-regulated genes. Using functional predictive analysis, we discovered these genes are enriched in GO terms such as proliferation, differentiation of T cells and anti-apoptotic process. By cross comparison with previous published data, we found 21 differentially expressed genes overlapped with SMAD signaling target genes.Our observations indicate the role of adaptive immune system activation during the pathology of abnormal muscular ossification. Our study further highlights the significance of anti-inflammatory therapy in the treatment of FOP and provide novel molecular markers for the clinical diagnosis of FOP disease.

overall_design: PBMC RNA-seq

first: Jing

last: Yu

first: Keqin


title: CAN1 Arginine Permease Deficiency Extends Yeast Replicative Lifespan via Translational Activation of Stress Response Genes

city: Boston

first: Vyacheslav

last: Labunskyy

state: Massachusetts

address: 609 Albany Street

department: Department of Dermatology

institute: Boston University School of Medicine

summary: Transcriptional regulation plays an important role in the control of gene expression during aging. However, translation efficiency likely plays an equally important role in determining protein abundance, but has been relatively under studied in this context. Here we used RNA-seq and ribosome profiling to investigate the role of translational regulation in lifespan extension by CAN1 gene deletion in yeast. Through comparison of the transcriptional and translational changes in cells lacking CAN1 with other long-lived mutants, we were able to identify critical regulatory factors, including transcription factors and mRNA-binding proteins, that coordinate transcriptional and translational responses. Together, our data support a model in which deletion of CAN1 extends replicative lifespan through increased translation of proteins that facilitate cellular response to stress. This study extends our understanding of the importance of translational control in regulating stress resistance and longevity.

overall_design: RNA-seq and ribosome profiling analyses of wild-type, can1, rpl22a, and tor1 knockout strains

first: Carine

last: Beaupere

first: Brian

middle: M

last: Wasko

first: Jared

last: Lorusso

middle: K

last: Kennedy

first: Matt

last: Kaeberlein


title: Deletion of the thyroid hormone-activating type 2 deiodinase rescues cone photoreceptor degeneration but not deafness in mice lacking type 3 deiodinase

city: Bethesda

first: Hong

last: Liu

state: MD

address: 9000 Rockville Pike

department: NIDDK

institute: NIH

summary: Using Next-generation sequencing (NGS) to get the retinal transcriptome profiles (RNA-seq) for understanding gene regulations during retina development

overall_design: Retinal mRNA profiles from embryo day 16.5 to postnatal day 28 wild type (WT) mice were generated by NGS sequencing

first: Douglas

last: Forrest

first: Lily

last: Ng


title: Ascorbate concentration in Arabidopsis is controlled at the gene expression level mainly by GDP galactose phosphorylase

status: Public on Feb 17 2017

city: Palmerston North

first: Helge

last: Dzierzon

address: Bachelar Road

department: Bioinformatics

country: New Zealand

institute: Plant and Food Research Ltd.

summary: We explore where transcriptional regulation of ascorbate concentration lies in plants. Is it in biosynthesis, recycling, regulation or consumption? Arabidopsis thaliana plants were grown under controlled environment at four photon flux density levels (PFD). Rosettes from plants were harvested at the four PFD levels and over a diurnal cycle and after a step change in PFD and analysed for ascorbate concentration and transcript levels measured by RNAseq. Ascorbate concentrations and expression of genes in the L-galactose ascorbate biosynthesis, recycling, consumption pathways and regulation are presented to provide a full analysis of the control of ascorbate by environmentally modulated gene expression. Ascorbate concentration responded to PFD levels but not to time of day and showed only a small response to change of PFD after 2 days. Of the L-galactose pathway genes, only GDP galactose phosphorylase (GGP) showed a significant response in to different PFDs, time of day and to change in PFD.  Other genes also showed limited responses. This study compares gene expression of a range of ascorbate related genes to changes in environment in a unified way and supports the concept that GGP is the key regulatory gene in ascorbate biosynthesis and that post transcriptional regulation is also important.

overall_design: Arabidopsis were grown in controlled envrionment and ascorbate and gene expression measured at different light intensities, time of day and change in light intensity. 24 samples are analysed with two replicates per treatment.

first: William

last: Laing


title: RNAseq analysis of B cells inducible for murine gammaherpesvirus 68 reactivation

city: Stony Brook

first: Laurie

last: Krug

address: 130 Life Sciences Bldg

department: Molecular Genetics and Microbiology

institute: Stony Brook University

summary: RTA, the viral Replication and Transcription Activator, is essential for rhadinovirus lytic gene expression upon de novo infection and reactivation from latency. LPS/TLR4 engagement enhances rhadinovirus reactivation. We developed the HE-RIT cell line, a latent murine A20 B cell inducible for Flag-RTA expression and murine gammaherpesvirus 68 reactivation.  LPS acted as a second stimulus to drive virus reactivation from latency in the context of induced expression of FLAG-RTA. We applied RNAseq to examine for genome-wide changes in viral gene expression in response to doxycycline-induced RTA-FLAG, alone or in combination with LPS.

overall_design: 6 samples total: Replicates of Mock (untreated controls), Dox (induced for RTA expression), and Dox + LPS (dual treatment)

first: Laraib

last: Malik


title: Next-Generation Sequencing Facilitates Quantitative Analysis of Wild Type and csrA Mutant Helicobacter pylori J99 Transcriptomes

first: Cheng-Yen

last: Kao

address: Department of Biotechnology and Laboratory Science in Medicine, School of Biomedical Science and Engineering, National Yang Ming University, No.155, Sec. 2, Linong St., Beitou Dist

institute: National Yang Ming University

summary:
Purpose: The goals of this study are to clarify the CsrA-regulatory system in H. pylori by NGS-derived transcriptome profiling (RNA-seq).

Method: CsrA regulatory system was determined by comparative transcriptomic analysis carried out with RNA-seq on strain J99 and csrA mutant.

Results: Using an optimized data analysis workflow, fifty-three genes in the csrA mutant were found to be differentially expressed compared with the wild-type.

Conclusions: Our study represents the first detailed analysis of CsrA-regulatory system in H. pylori J99.

overall_design: mRNA profiles of wild type (WT) and csrA mutant J99 were generated by deep sequencing, in triplicate, using 5500XL SOLiD™ Sequencer System.

first: Cheng

middle: Y

first: Jiunn

last: Wu


title: Generation of mature T cells from human hematopoietic stem/progenitor cells in artificial thymic organoids

city: Pasadena

first: Michael

last: Bethune

address: 1200 E California Ave, MC147-75

department: BBE

institute: California Institute of Technology

summary: Mechanistic studies of human T cell development require robust model systems that recapitulate the full span of thymopoiesis, from hematopoietic stem and progenitor cells (HSPC) through to mature naïve T cells. We developed a serum free, artificial thymic organoid (ATO) system that supports highly efficient and reproducible in vitro differentiation and positive selection to generate conventional human T cells from all sources of HSPCs. Deep-sequencing of the TCR variable genes was performed to compare the diversity of the TCR repertoire between ATO-derived CD8SP T cells and naive CD8SP T cells from thymus and blood.

overall_design: TCR repertoire analysis of CD3+TCRab+CD8SP T cells derived from cord blood CD34+CD3- HSPCs in ATOs compared with human naïve CD8SP T cells from the thymus and blood

first: Christopher

middle: S

last: Seet


title: RNA-seq analysis of bone marrow peri-vascular stromal cells

city: Bronx

first: Paul

last: Frenette

address: 1301 Morris Park Avenue , Room 101

department: Ruth L. and David S. Gottesman Institute for Stem Cell and Regenerative Medicine Research

institute: Albert Einstein College of Medicine

summary: Fate decisions of haematopoietic stem cells (HSCs) to self-renew or differentiate in response to various demands are finely tuned by specialized microenvironments called “niches” in the bone marrow. Recent studies suggest that arterioles and sinusoids accompanied with distinct stromal cells marked by nerve/glial antigen 2 (NG2) and leptin receptor (LepR), compose distinct niches regulating quiescence and proliferation of HSCs, respectively. However, it remains unknown how the distinct niche cells differentially regulate the HSC functions. Here we show that effects of cytokines regulating HSC functions are dependent on the producing cell sources. Deletion of chemokine C-X-C motif ligand 12 (CXCL12) in NG2-cre targeted cells, which exclusively overlap with Nestin-GFP (Nes-GFP)+ stromal cells associated with arterioles and sinusoids, resulted in a robust reductions of HSCs in the bone marrow and massive mobilization. Deletion of CXCL12 from arteriolar NG2+ vascular smooth muscle cells caused a significant decrease of HSCs and altered HSC location in the marrow, while CXCL12 depletion from sinusoidal LepR+ cells did not reduce HSC numbers in the bone marrow. By contrast, deletion of stem cell factor (SCF) in LepR+ cells led to significant reductions in HSC numbers whereas SCF deletion in arteriolar NG2+ cells showed no effect on HSC numbers in the marrow. These results uncover the distinct contributions of cytokines derived from perivascular cells in separate vascular niches for HSC maintenance and mobilization.  We sought to obtain comprehensive understanding of differences between peri-arteriolar and peri-sinusoidal niche cells by the present RNA-seq analysis.

overall_design: mRNA profiles of sorted LepR-cre/Tomato+, NG2-cre/Tomato+, Myh11-creER/Tomato+, and CD31+ bone marrow stromal cells were generated by illumina HiSeq2500.

first: Noboru

last: Asada


title: Overcoming AXL Activation and Epithelial–Mesenchymal Transition is Critical in Conquering ALK-positive Non–Small Cell Lung Cancer

city: Tokyo

first: Shinji

last: Nakamichi

address: 1-1-5, Sendagi, Bunkyo-ku

department: Department of Pulmonary Medicine and Oncology

country: Japan

institute: Graduate School of Medicine, Nippon Medical School

summary: Anaplastic lymphoma kinase tyrosine kinase inhibitors (ALK-TKIs) induce a dramatic response in non–small cell lung cancer (NSCLC) patients with the ALK fusion gene. However, acquired resistance to ALK-TKIs in lung cancer cells remains an inevitable problem: ALK secondary mutations and bypass pathways have been reported as major resistance mechanisms. In this study, we aimed to discover a novel mechanism of acquired resistance to ALK-TKIs and a strategy to conquer ALK-positive lung cancer. We established three types of ALK-TKI (crizotinib, alectinib and ceritinib)–resistant H2228 non-small cell lung cancer cell lines by high exposure and stepwise methods. We found these cells showed a loss of ALK signaling, overexpressed AXL with epithelial–mesenchymal transition (EMT), and had cancer stem cell–like properties. Similarly, we demonstrated that TGF-β1 treated H2228 cells also showed AXL overexpression with EMT features and ALK-TKI–resistance. The AXL inhibitor, R428, or HSP90 inhibitor, ganetespib, were effective in reversing ALK-TKI–resistance and EMT changes in both ALK-TKI–resistant and TGF-β1–exposed H2228 cells. Progression-free survival of ALK-positive NSCLC patients with AXL overexpression was shorter than that of patients who underwent crizotinib therapy and showed low AXL expression. Thus, we found ALK signaling-independent AXL overexpression and EMT features were commonly involved in intrinsic and acquired resistance to first and second generation ALK-TKIs. This suggests AXL and HSP90 inhibitors may be promising therapeutic drugs to overcome tumor cells in ALK-positive NSCLC patients.

overall_design: We established ALK-TKI-resistant H2228 cells to discover a novel mechanism of acquired resistance to ALK-TKIs and a strategy to conquer ALK-positive lung cancer.

first: Seike

last: Masahiro

first: Akihiko

last: Miyanaga

first: Mika

last: Chiba

first: Fenfei

last: Zou

first: Akiko

last: Takahashi

first: Arimi

last: Ishikawa

first: Shinobu

last: Kunugi

first: Rintaro

last: Noro

first: Kaoru

last: Kubota

last: Gemma


title: Stage-specific human induced pluripotent stem cells map the progression of myeloid transformation to transplantable leukemia.

first: Samir

last: Parekh

address: 1470 Madison Ave, 5th FL, Room 202

institute: Icahn School of Medicine at Mount Sinai, New York

summary: We used Illumina NGS for high-throughput profiling of the DNA methylome (ERRBS) for myelodysplastic syndrome (MDS) iPSC cell lines. We performed DNA methylation analysis of the low-risk MDS1.12 cells (iPSC cell line)  treated with 5-AzaC- and show that profound hypomethylation changes can be detected following treatment with 5-AzaC and support the use of our stage-specific induced pluripotent stem cells system to study mechanisms of drug action.

overall_design: For ERRBS experiments, MDS-1.12 cells used to assess  colony formation, were collected on day 14 of hematopoietic  differentiation and treated with 0, 100 nM and 500 nM 5-AzaC every day for 3 days. Genomic DNA was extracted and library fragment lengths of 150-250 bp  and 250-400 bp were prepared, gel isolated and sequenced on an Illumina Hi-seq 2500. All samples passed quality control with methylation assayed at >2  million CpG dinucleotides per sample on average 80x coverage per cytosine.  Methylation data analysis was performed using an established computational pipeline (Akalin et al., PLoS Genetics 2012). Briefly, reads were aligned to the genome using the bismark alignment software with a maximum of 2 mismatches in a directional manner and only uniquely aligning reads were retained. Read bases aligning to positions with at least 20 phred quality score were retained for subsequent analysis. Percentage of bisulfite converted Cs (representing unmethylated Cs) and non-converted Cs (representing methylated Cs) were recorded for each C position in a CpG context. Differential methylation analysis was performed using R package methylKit (Akalin et al., Genome Bio 2012). Differentially methylated loci between the 5-AzaC-treated and untreated groups were identified (25% methylation difference, q<0.01). The  differentially methylated loci were annotated with hg19 to infer gene associations.

first: Andriana

last: Kotini

first: Deepak

last: Perumal

first: Eirni

last: Papapetrou


title: Expression analysis of primary human monocytes stimulated with an anti-TLR10 mAb (5C2C5)

city: Urbana

first: Nicholas

middle: J.

last: Hess

state: IL

address: B103 CLSL MC110, 601 S. Goodwin Ave

department: Microbiology

institute: University of Illinois at Urbana-Champaign

summary: We report that engagment of TLR10 via the 5C2C5 anti-TLR10 mAb causes differential expression is a subset of inflammatory, miRNA and uncharacterized genes compared to an isotype control antibody

overall_design: Examination of gene expression on primary human monocytes under 4 different conditions

first: Richard

last: Tapping


title: Differential responses of human fetal brain neural stem cells to Zika virus infection

city: Galveston

first: Ping

last: Wu

state: Texas

address: 301 University Blvd

institute: University of Texas Medical Branch

summary:
Purpose: The goal of this study is to investigate the effect of Zika virus infection on neural stem cells
Methods: RNA-Seq transcriptome analysis of neural stem cells infected with Zika virus compared to mock infected

overall_design: RNA-seq transcriptome profiling of Zika infected human fetal neural stem cells

first: Erica

last: McGrath

first: Shannan

last: Rossi

first: Junling

last: Gao

first: Steven

middle: G

last: Widen

first: Auston

middle: C

last: Grant

first: Tiffany

last: Dunn

first: Sasha

last: Azar

first: Christopher

last: Roundy

first: Ying

last: Xiong

first: Deborah

last: Prusak

first: Bradford

middle: D

last: Loucas

first: Thomas

last: Wood

first: Yongjia

last: Yu

first: Ildefonso

last: Fernández-Salas

first: Scott

last: Weaver

first: Nikos

last: Vasilakis


title: Seq-Well: A portable, low-cost platform for single-cell RNA-seq of low-input samples

city: Cambridge

first: Alex

last: Shalek

state: MA

address: 77 Masachusetts Ave

department: Chemistry

institute: Massachusetts Institute of Technology

summary: Single-cell RNA -seq can precisely resolve cellular states, but applying this method to low-input samples is challenging. Here, we present Seq-Well, a portable, low-cost platform for massively parallel single-cell RNA -seq. Barcoded mRNA capture beads and single cells are sealed in an array of subnanoliter wells using a semipermeable membrane, enabling efficient cell lysis and transcript capture. We characterize Seq-Well extensively and use it to profile thousands of primary human macrophages exposed to tuberculosis.

overall_design: Single-cell RNA sequencing experiments of HEK/3T3 mixing, PBMCs, and TB-exposed Macrophages.

first: Travis

last: Hughes


title: Specific and redundant roles of TEAD transcription factors in C2C12 cell and primary myoblast differentiation (ChIP-Seq)

city: Illkirch

first: Shilpy

last: Joshi

address: 1 Rue laurent Fries

country: France

institute: IGBMC

summary: Defining the function of TEAD transcription factors in myogenic differentiation has proved elusive due to overlapping expression and functional redundancy. Here, we show that siRNA silencing of either Tead1, Tead2 or Tead4 did not effect differentiation of primary myoblasts (PMs) while their simultaneous knockdown strongly impaired differentiation. In contrast in C2C12 cells, silencing of Tead1 or Tead4 impaired differentiation showing a differential requirement for these factors in PMs and C2C12 cells that involved both differential regulation of their expression and intracellular localisation. Through integration of Tead1 and Tead4 ChIP-seq with chromatin modifications, we identify active enhancers associated with genes activated during C2C12 cell differentiation that are bound by combinations of Tead4, Myod1 or Myog and show a signature of frequently co-occuring motifs. We show that distinct but overlapping sets of genes are deregulated by Tead silencing in C2C12 cells and PMs therefore describing for the first time in a comprehensive manner the specific and redundant regulatory roles of Tead factors in myogenic differentiation. We also performed ChIP-seq from mouse muscle in vivo identifying a set of highly transcribed muscle cell-identity genes and revealing that Tead4 binds a distinct repertoire of sites in C2C12 cells and muscle.

overall_design: Examination of Tead1 and Tead4 occupancy during muscle differentiation and in adult muscle by ChIP-seq

first: Irwin

last: Davidson

first: Stéphanie

last: Le Gras

first: Guillaume


title: Specific and redundant roles of TEAD transcription factors in C2C12 cell and primary myoblast differentiation

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

overall_design: Refer to individual Series


title: Identification of methylation haplotype blocks aids in deconvolution of heterogeneous tissue samples and tissue-of-origin mapping from plasma DNA [WGBS]

city: La Jolla

first: Kun

last: Zhang

summary: Adjacent CpG sites in mammalian genomes tend to be co-methylated due to the processivity of enzymes responsible for adding or removing the methyl group. Yet discordant methylation patterns have also be observed, and found to be related to stochastic or uncoordinated molecular processes. Here we focused on a systematic search and investigation of regions in the human genome that exhibit highly coordinated methylation.   By examining the co-methylation patterns of multiple adjacent CpG sites, termed methylation haplotypes, in single bisulfite sequencing reads, we applied a greedy-searching strategy to defined blocks of tightly coupled CpG sites, called Methylation Haplotype Blocks (MHBs), based on 53 sets of whole genome bisulfite sequencing (WGBS) data, including 43 published sets from human adult tissues, ESC and in vitro differentiated cell lines, as well as 10 sets from human adult tissues generated in this study. The MHBs were then further validated with 101 sets of RRBS ENCODE data, and 637 sets of Illumina 450k methylation array data from TCGA tumor and normal samples. Globally, MHBs are enriched in but only partially overlap with several well-known genomic features, including CpG islands, promoters, enhancers and VMRs.  To perform quantitative analysis of the MHBs, we defined a metric called Methylation Haplotype Load (MHL), which is covered both average methylation level and methylation complexity and therefore more informative than average methylation level or Shannon entropy. Using a feature selection strategy, we identified a set of tissue-specific MHBs that cluster by developmental germ-layers. Interestingly, examination of these MHBs revealed two distinct mechanisms for fate commitment during development: epigenetic silencing of pluripotent genes, such as NANOG, for mesoderm induction; and epigenetic induction (or de-suppression) of lineage-specific factors for ectoderm commitment.

overall_design: Examination of co-methylation in primary human tissues

first: Shicheng

last: Guo

first: Dinh

last: Diep


title: SIRT7 is an RNA-activated protein lysine deacylase [RIP ribosomal RNA]

city: Ithaca

first: Jennifer

last: Grenier

address: 526 Campus Rd

department: Biomedical Sciences

institute: Cornell University

summary: We report that RNA can increase the catalytic efficiency of SIRT7 and that SIRT7 can remove long chain fatty acyl groups more efficiently than removing acetyl groups. Truncation and mutagenesis studies revealed residues at both the amino and carboxyl termini of SIRT7 that are involved in RNA-binding and important for activity. RNA immunoprecipitation-sequencing (RIP-seq) identified ribosomal RNA (rRNA) as the predominant RNA binding partners of SIRT7. The associated RNA was able to effectively activate the deacetylase and defatty-acylase activities of SIRT7. These findings provide important insights into the biological functions of SIRT7, as well as an improved platform to develop SIRT7 modulators.

overall_design: SIRT7 with a Flag tag was expressed in HEK293T cells. Anti-Flag antibody (Sigma #A2220) was used to pull down SIRT7 along with the RNA that were bound to SIRT7. The RNA associated with SIRT7 was examined using next-generation sequencing.

first: Zhen

last: Tong

first: Hening

last: Lin


title: SIRT7 is an RNA-activated protein lysine deacylase [RIP-seq ribo-zero]


title: Composition and Function of Mutant Swi/Snf Complexes

city: Kansas City

first: Madelaine

last: Gogol

state: MO

address: 1000 E. 50th Street

department: Computational Biology Core

institute: Stowers Institute

summary: Loss of Swi/Snf subunits regulate transcription.

overall_design: RNA-seq and ChIP-seq for Snf2 was done in Swi/Snf subunit deletion mutant yeast strains

first: Arnob

last: Dutta

first: Jerry

last: Workman


title: SIRT7 is an RNA-activated protein lysine deacylase


title: MicroRNA-independent functions of DGCR8 are critically required for neocortical development and TBR1 expression

city: Milano

first: Francesco

last: Nicassio

address: Via Adamello 16

department: Center for genomic science

country: Italy

institute: Istituto Italiano di Tecnologia

type: Non-coding RNA profiling by high throughput sequencing

summary: Recent evidence indicates that DROSHA, DGCR8 and DICER exert non-overlapping functions, including miRNA-independent regulatory mechanisms. It is currently unknown whether miRNA-independent functions of DGCR8 play any role in corticogenesis. Here, by phenotypic comparison of conditional knockout cortices for Dgcr8 and Dicer, we uncover that Dgcr8 deletion, in contrast to Dicer, leads to premature differentiation of neural progenitor cells and overproduction of Tbr1 positive neurons. Remarkably, miRNA depletion does not account for these phenotypic differences, indicating that miRNA-independent functions of DGCR8 are critical for corticogenesis. We propose Tbr1 transcript, bearing evolutionary conserved hairpins that resemble miRNA precursors, as one of DGCR8 targets. Our study uncovers, for the first time in the nervous system, an earlier and stronger phenotype occurring in Dgcr8 compared to Dicer mutant and proposes that this effect is mediated by miRNA-independent functions of DGCR8.

overall_design: miRNA expression by deep-sequencing in the cortex of Wild-Type, DICER and DGCR8 conditional knock-out mice. Two mice for each genotype.

first: Federica

last: Marinaro

first: Matteo

middle: J

last: Marzi

first: Nadin

last: Hoffmann

first: Hayder

last: Amin

first: Roberta

last: Pelizzoli

last: Niola

first: Davide

last: De Pietri Tonelli


title: Differentiation of ncRNAs from small mRNAs in Escherichia coli O157:H7 EDL933 (EHEC) by combined RNAseq and RIBOseq – ryhB encodes the regulatory RNA RyhB and a peptide, RyhP

city: Freising

first: Klaus

last: Neuhaus

address: Weihenstephaner Berg 3

department: Core Facility Microbiome/NGS

country: Germany

institute: Technische Universität München

summary: While NGS allows rapid global detection of transcripts, it remains difficult to distinguish ncRNAs from short mRNAs. To detect potentially translated RNAs, we developed an improved protocol for bacterial ribosomal footprinting (RIBOseq). This allowed distinguishing ncRNA from mRNA in EHEC. A high ratio of ribosomal footprints per transcript (ribosomal coverage value, RCV) is expected to indicate a translated RNA, while a low RCV should point to a non-translated RNA. Based on their low RCV, 150 novel non-translated EHEC transcripts were identified as putative ncRNAs, representing both antisense and intergenic transcripts, 74 of which had expressed homologs in E. coli MG1655. Bioinformatics analysis predicted statistically significant target regulons for 15 of the intergenic transcripts; experimental analysis revealed 4-fold or higher differential expression of 46 novel ncRNA in different growth media. Out of 329 annotated EHEC ncRNAs, 52 showed an RCV similar to protein-coding genes, of those, 16 had RIBOseq patterns matching annotated genes in other enterobacteriaceae, and 11 seem to possess a Shine-Dalgarno sequence, suggesting that such ncRNAs may encode small proteins instead of being solely non-coding. To support that the RIBOseq signals are reflecting translation, we tested the ribosomal-footprint covered ORF of ryhB and found a phenotype for the encoded peptide in iron-limiting condition. Determination of the RCV is a useful approach for a rapid first-step differentiation between bacterial ncRNAs and small mRNAs. Further, many known ncRNAs may encode proteins as well.

overall_design: Comparison of RNAseq with RIBOseq data from EHEC grown in LB medium

first: Richard

last: Landstorfer

first: Svenja

last: Simon

first: Steffen

last: Schober

first: Patrick

last: Wright

first: Cameron

last: Smith

first: Rolf

last: Backofen

first: Romy

last: Wecko

first: Daniel

last: Keim

first: Siegfried

last: Scherer


title: Mechanisms of variable ventilation in experimental lung injury

city: Dresden

first: Peter

last: Spieth

address: Fetscherstraße 74

department: Klinik und Poliklinik für Anästhesiologie und Intensivtherapie

institute: Universitätsklinikum Carl Gustav Carus an der Technischen Universität Dresden

summary: In the present study we seek to identify changes in lung gene expression under mechanical ventilation in uninjured as well as acutely and chronically injured lungs. A standard volume-controlled lung-protective ventilatory protocol is compared to a concept of mechanical ventilation using variable tidal volumes.

overall_design: Comparison of mRNA profiles in whole-lung samples after mechanical ventilation with constant or variable tidal volumes

first: Hannes

last: Krause

first: Thomas

last: Bluth

first: Maria

last: Lippmann

first: Nadine

last: Oppermann

first: Robert

last: Huhle

first: Ines

last: Rentzsch

first: Andreas

last: Dahl

first: Dimitra

last: Alexopoulou

first: Micheal

last: Kasper

first: Claudia

last: dos Santos

last: Hegeman

first: Thea

last: Koch

first: Marcelo

last: Gama de Abreu


title: FOXR2 Interacts with MYC to Promote Its Transcriptional Activities and Tumorigenesis

city: Houston

first: Yuanxin

last: Xi

state: TX

address: 1400 Pressler St

department: Bioinformatics and Computational Biology

institute: The University of Texas MD Anderson Cancer Center

summary: Combining the results of a large scale proteomic analysis of human transcription factor interaction network with knowledge databases, we identified FOXR2 as one of the top-ranked candidate proto-oncogenes. Here, we show that FOXR2 forms a stable complex with MYC and MAX and subsequently regulates cell proliferation by promoting MYC’s transcriptional activities. We demonstrated that FOXR2 is highly expressed in several breast, lung, and liver cancer cell lines and related patient tumor samples, while reduction of FOXR2 expression in a xenograft model inhibits tumor growth. These results indicate that FOXR2 acts with MYC to promote cancer cell proliferation, which is a potential tumor-specific target for therapeutic intervention against MYC-driven cancers.

overall_design: ChIP-seq analysis of FOXR2, MAX and MYC in MB468 cells

first: Xu

last: Li


title: Systematic investigation of transcription factor activity in the context of chromatin using massively parallel DNA binding and expression assays

status: Public on Feb 16 2017

city: Rehovot,

first: Michal

last: Levo

address: Herzl Street 234

country: Israel

institute: Weizmann Institute of Science

summary: Precise gene expression patterns are established by timely and specific binding of transcription factors (TFs) to regulatory sequences. While these events occur in the context of chromatin, our understanding of how TF-nucleosome interplay affects gene expression is highly limited. Here we present a novel assay for high-resolution measurements of both DNA occupancy and gene expression on large-scale libraries of systematically designed regulatory sequences. Our assay is capable of revealing occupancy patterns at the single–cell level. It provides an accurate quantification of the fraction of the population bound by a nucleosome and captures distinct, even adjacent, TF binding events. By applying this assay to over 1500 promoter variants in yeast, we reveal pronounced differences in the dependency of TF activity on chromatin. Our approach provides means to classify TFs by their differential capacity to alter chromatin and promote expression. We further demonstrate how different regulatory sequences give rise to nucleosome-mediated TF collaborations that quantitatively account for the resulting expression. We show the utility of this approach in dissecting the logic of native regulatory sequences, highlighting the importance of TF-nucleosome interactions in the quantitative readout of regulatory information

overall_design:
A library of ~1600 promoter variants of length 210bp was designed and synthesized on Agilent programmable microarrays. Each sequence includes 19bp 5’ primer sequence, a 10bp unique barcode sequence, 163bp variable region and 18 bp 3’ primer. Sequences were integrated into the yeast genome upstream of a YFP reporter. Yeast population carrying this library was subjected to expression measurements as described in Sharon et al, Nature Biotechnology, 2012. In brief, cells were grown to mid-log and sorted in FACSAria to 16 bins based on YFP fluorescence. Sorted cells were grown to stationary phase, DNA was extracted and PCR amplification of the region of interest was performed. The 3′ primer was common to all bins (5′-NNNNNTTATGTGATAATGCCTAGGATCGC-3′, where the Ns represent random nucleotides). The 5′ primer had a common sequence and a unique upstream 5-bp barcode sequence (underlined) that was specific to each bin (5′-XXXXXGGGGACCAGGTGCCGTAAC-3′, where the Xs represent the bin's unique sequence). Three 5’ primers were used for the amplification of each bin, reads were later joined after we found them to be highly consistent. The list of primers is shown below. Amplified product was gel-extracted and 10 nanograms were used in library preparation for sequencing (protocol adopted from Blecher-Gonen et al. 2013). The DNA was amplified using 14 amplification cycles and library was analyzed using the 2200 TapeStation system and sequenced with 50-bp, single read flowcell on an Illumina HiSeq 2000 sequencer. This sample is referred as the “Expression measurements”. Yeast population carrying the library was subjected to a novel binding assay, aimed to reveal the occupancy pattern on the designed promoter variants. Cells were crosslinked and spheroplasts were then obtained. Spheroplasts were then subjected to methylation by the CpG Methyltranferase M.SssI. DNA was extracted and Bisulfite conversion was performed. We then PCR amplify the region of interest (520bp) with the following primers (that include 5bp tails underlined): 5’ primer – 5’-CATTCTAAAAGAATAGACCTAGATAGGGTT-3’, 3’ primer – 5’-NNNNNCATCACCATCTAATTCAACCAAAAT-3’. Amplified product was gel-extracted and 10 nanograms were used in library preparation for sequencing (protocol adopted from Blecher-Gonen et al. 2013). The DNA was amplified using 14 amplification cycles and library was analyzed using the 2200 TapeStation system. Libraries were sequenced on a 300 paired-end flow cells (MiSeq Reagent Kit v3, MS-102-3001) on MiSeq desktop sequencer (Illumina). This sample is referred as the “Binding measurements”. As the samples have low complexity they were mixed with other material prior to sequencing, the relevant reads can be identified by the sequence of the primers.      Expression bins 5’ primers:
Bin1 AACGAGGGACCAGGTGCCGTAAC  Bin2 GCATAGGGACCAGGTGCCGTAAC
Bin3 AGATAGGGACCAGGTGCCGTAAC  Bin4 GAGTAGGGACCAGGTGCCGTAAC
Bin5 TCGTAGGGACCAGGTGCCGTAAC  Bin6 CTGTAGGGACCAGGTGCCGTAAC
Bin7 TATTAGGGACCAGGTGCCGTAAC  Bin8 CGTTAGGGACCAGGTGCCGTAAC
Bin9 ACGAAGGGACCAGGTGCCGTAAC  Bin10 TAAGCGGGACCAGGTGCCGTAAC
Bin11 TTATCGGGACCAGGTGCCGTAAC  Bin12 TACTCGGGACCAGGTGCCGTAAC
Bin13 CATTCGGGACCAGGTGCCGTAAC  Bin14 CCTAAGGGACCAGGTGCCGTAAC
Bin15 GCAATGGGACCAGGTGCCGTAAC  Bin16 CTACAGGGACCAGGTGCCGTAAC
Bin1 ACTATGGGACCAGGTGCCGTAAC  Bin2 TACCAGGGACCAGGTGCCGTAAC
Bin3 CAAGTGGGACCAGGTGCCGTAAC  Bin4 TGAGTGGGACCAGGTGCCGTAAC
Bin5 ATAGTGGGACCAGGTGCCGTAAC  Bin6 AAGGTGGGACCAGGTGCCGTAAC
Bin7 ATCCAGGGACCAGGTGCCGTAAC  Bin8 GATGTGGGACCAGGTGCCGTAAC
Bin9 TCTGTGGGACCAGGTGCCGTAAC  Bin10 AGTGTGGGACCAGGTGCCGTAAC
Bin11 CTTGTGGGACCAGGTGCCGTAAC  Bin12 TCATTGGGACCAGGTGCCGTAAC
Bin13 AAGCAGGGACCAGGTGCCGTAAC  Bin14 CGATTGGGACCAGGTGCCGTAAC
Bin15 GTATTGGGACCAGGTGCCGTAAC  Bin16 CACTTGGGACCAGGTGCCGTAAC
Bin1 ATGTTGGGACCAGGTGCCGTAAC  Bin2 CATCAGGGACCAGGTGCCGTAAC
Bin3 TCTCAGGGACCAGGTGCCGTAAC  Bin4 AGTCAGGGACCAGGTGCCGTAAC
Bin5 GTTCAGGGACCAGGTGCCGTAAC  Bin6 ACAGAGGGACCAGGTGCCGTAAC
Bin7 GTAGAGGGACCAGGTGCCGTAAC  Bin8 TTGGAGGGACCAGGTGCCGTAAC
Bin9 TGTGAGGGACCAGGTGCCGTAAC  Bin10 ATTGAGGGACCAGGTGCCGTAAC
Bin11 GACAAGGGACCAGGTGCCGTAAC  Bin12 CAATAGGGACCAGGTGCCGTAAC
Bin13 TCCAAGGGACCAGGTGCCGTAAC  Bin14 AGCAAGGGACCAGGTGCCGTAAC
Bin15 CTCAAGGGACCAGGTGCCGTAAC  Bin16 CAGAAGGGACCAGGTGCCGTAAC

first: Tali

last: Avnit-Sagi

first: Maya

last: Lotan-Pompan

first: Yael

last: Kalma

first: Adina

last: Weinberger

first: Zohar

last: Yakhini

first: Eran

last: Segal


title: Stage-specific human induced pluripotent stem cells map the progression of myeloid transformation to transplantable leukemia

city: South San Francisco

first: Han

last: Yuan

address: 1170 Veterans Blvd

institute: Calico Life Sciences

summary: Comparison of disease stage transitions between a panel of iPSC clones derived  from different MDS patients

overall_design: RNA from CD34+ cells derived from in vitro hematopoietic differentiation of iPSCs from MDS patients

first: Eirini

last: Papapetroui

first: Andriana

last: Kotini


title: HEPATIC DEFICIENCY OF THE PIONEER TRANSCRIPTION FACTOR FOXA RESTRICTS HEPATITIS B VIRUS BIOSYNTHESIS BY THE DEVELOPMENTAL REGULATION OF VIRAL DNA METHYLATION

city: Chicago

first: Mark

last: Maienschein-Cline

address: 900 S Ashland Ave, Rm 1058, M/C 937

department: Research Resources Center

institute: University of Illinois at Chicago

summary: The FoxA family of pioneer transcription factors regulate hepatitis B virus (HBV) transcription, and hence viral replication. Hepatocyte-specific FoxA-deficiency in the HBV transgenic mouse model of chronic infection prevents the transcription of the viral DNA genome as a result of the failure of the developmentally controlled conversion of 5-methylcytosine residues to cytosine during postnatal hepatic maturation.  These observations suggest that pioneer transcription factors such as FoxA, which mark genes for expression at subsequent developmental steps in the cellular differentiation program, mediate their effects by reversing the DNA methylation status of their target genes to permit their ensuing expression when the appropriate tissue-specific transcription factor combinations arise during development.  Furthermore, as the FoxA-deficient HBV transgenic mice are viable, the specific developmental timing, abundance and isoform type of pioneer factor expression must permit all essential liver gene expression to occur at a level sufficient to support adequate liver function.  This implies that pioneer transcription factors can recognize and mark their target genes in distinct developmental manners dependent upon, at least in part, the concentration and affinity of FoxA for its binding sites within enhancer and promoter regulatory sequence elements.  This selective marking of cellular genes for expression by the FoxA pioneer factor compared to HBV may offer the opportunity for the specific silencing of HBV gene expression and hence the resolution of chronic HBV infections which annually are responsible for approximately one million deaths worldwide due to liver cirrhosis and hepatocellular carcinoma.

overall_design: 31 bisulfite sequencing samples, each with 11 regions targeted by amplicon sequencing

first: Vanessa

last: Reese

first: Rasha

middle: E

last: Shalaby

first: Saira

last: Iram

last: Oropeza

first: Jennifer

last: Landolfi

first: Alexander

middle: V

last: Lyubimov

first: Stefan

last: Green

first: Klaus

middle: H

last: Kaestner

first: Alan

last: McLachlan


title: TFAP2A ChIP-seq in mouse immortalized melanocytes

city: Bethesda

first: Stacie

last: Loftus

address: 49 Convent Dr

institute: National Human Genome Research Institute

summary: Damage to the gene regulatory network governing terminal differentiation of melanocytes leads to pigmentation phenotypes and increases the risk for melanoma. Microphthalmia-associated transcription factor (MITF) directly activates expression of melanocyte differentiation effectors, and levels of MITF have been proposed to govern the melanoma phenotype. Mutations in the gene encoding Transcription Factor Activator Protein 2 alpha (TFAP2A) cause reduced pigmentation in model organisms and premature hair graying in humans, and TFAP2A expression tends to be lower in advanced melanoma tumors than in benign nevi. However, the transcriptional targets of TFAP2A in melanocytes, and the epistatic relationship of TFAP2A and MITF, have been unclear. Using microarray-based analysis of zebrafish tfap2a mutant embryos, we generated a profile of genes whose expression is Tfap2a-dependent. We conducted anti-TFAP2A chromatin immunoprecipitation followed by next-generation sequencing (ChIP-seq) in immortalized mouse melanocytes and human primary melanocytes, and discovered that TFAP2A peaks are present near the promoters of Tfap2a-dependent genes expressed in melanocytes, and also at the majority of enhancers active in melanocytes. Comparison of TFAP2A ChIP-seq data to published MITF ChIP-seq data showed that the set of genes with promoters bound by both MITF and TFAP2A is enriched for the gene ontology term “pigment cell differentiation.” Deletion analysis of one such co-bound promoter, for Transient Receptor Potential Melastatin-like 1(TRPM1), confirmed that its expression depends on the presence of MITF binding sites as previously shown, but also depends on the presence of TFAP2A binding sites. Finally, we find that mitfa and tfap2a interact genetically in zebrafish. Collectively, these results show that TFAP2A, operating in parallel with MITF, directly regulates effectors of terminal differentiation in melanocytes and melanoma.

overall_design: TFAP2A ChIP-seq was conducted in a mouse immortalized melanocyte cell line (melan-a cells).  TFAP2A enriched chromatin was compared to an input control.

first: William

last: Pavan

first: Robert

last: Cornell


title: Diffuse Large B Cell Lymphoma cell line with Acquired Resistance to PI3Kδ Inhibitor Idelalisib

city: Foster City

first: Li

last: Li

address: 333 Lakeside Drive

institute: Gilead Sciences

summary: RNAseq profile of TMD8 cell lines resistant to Idelalisib treatment.  Idelalisib resistant TMD8 cells were generated by continuous passage in the presence of 1 μM idelalisib for 8 weeks until stable resistance to idelalisib was established. Parallel cultures were grown in the presence of 0.1% DMSO as passage-matched, drug-sensitive control lines. Sensitive and resistant TMD8 cells were clonally isolated through two rounds of single cell limiting dilution

overall_design: 8 Idela-resistant TMD8 clones were evaluated and compared to 3 passage-matched idela-sensitive TMD8 clones

first: Rick

last: Sorensen

first: Stacey

last: Tannheimer

first: Anella

last: Yahiaoui

first: Sarah

last: Meadows


title: RNA-seq analysis of hepatocellular carcinoma with portal vein tumor thrombosis

city: Beijing

first: Jin

last: Gu

address: FIT Building 1-112C

department: Automation

institute: Tsinghua University

summary: Portal vein tumor thrombosis is a strong poor indication of HCC. Characterizing the molecular alterations of these metastatic HCCs is important for understanding the molecular mechanisms during HCC progression and metastasis. Previous genomic studies mainly focus on single molecular layer, such as gene expressions or exonic mutations. In this study, we systematically examined the copy number variation (CNV), DNA methylation, miRNA and transcriptome of matched adjacent normal tissues, primary tumors and PVTTs from 19 HCC patients. Based on the integrative multi-omics profiles, we established the molecular landscape of the metastatic HCCs and identified a set of the recurrent genomic alterations and candidate driver genes.

overall_design: RNA-seq of matched adjacent normal, primary tumor and PVTT samples from HCC patients

first: Gu

last: Jin

first: Chen

last: Lei


title: Genes regulated by TAZ in a lung fibroblast cell line

city: Tokyo

first: Satoshi

last: Noguchi

address: 7-3-1 Hongo, Bunkyo-ku

department: Department of Respiratory Medicine

institute: Graduate School of Medicine, The University of Tokyo

summary: To investigate the roles of TAZ in lung fibroblasts, we compared the expression profiles of a lung fibroblast cell line, HFL-1, transfected with control siRNA and siTAZ.

overall_design: We collected RNA from HFL-1 cells transfected with control siRNA and siTAZ. Two kinds of TAZ siRNAs (siTAZ #1 and siTAZ #2) were used. Two biological replicates (rep1 and rep2) were used for each condition.

first: Akira

last: Saito

first: Masafumi

last: Horie

first: Takahide

last: Nagase


title: miRNA-seq analysis of hepatocellular carcinoma with portal vein tumor thrombosis

overall_design: miRNA-seq of matched adjacent normal, primary tumor and PVTT samples from HCC patients

first: Lei

last: Chen


title: Diverse interventions that extend mouse lifespan suppress shared age-associated epigenetic changes at critical gene regulatory regions (WGBS 2)

city: Glasgow

first: Neil

last: Robertson

state: Scotland

address: Switchback Road

department: Beatson Institute for Cancer Research

institute: University of Glasgow

summary: Investigation into changes in the methylome in rapamycin and caloric restriction in aged mice.

overall_design: Examination of methylation states via high-throughput, whole-genome bisulfite sequencing in mouse hepatocytes of young (2 month) and aged (22 month) mice that display increased longevity through a variety of interventions

first: John

last: Cole

first: Mohammed

middle: I

last: Rather

first: Peter

last: Adams


title: The molecular landscape of hepatocellular carcinoma with portal vein tumor thrombosis

type: Methylation profiling by genome tiling array

type: Genome variation profiling by high throughput sequencing

overall_design: Refer to individual Series


title: Diverse interventions that extend mouse lifespan suppress shared age-associated epigenetic changes at critical gene regulatory regions

middle: A


title: Diverse interventions that extend mouse lifespan suppress shared age-associated epigenetic changes at critical gene regulatory regions (WGBS 1)

summary: Investigation into changes in the methylome in young (2 months) and aged (22 months) Ames dwarf (Prop1 Hom) and WT (Prop1 Het) mice


title: Specific and redundant roles of TEAD transcription factors in C2C12 cell and primary myoblast differentiation (RNA-Seq)

first: Shilpy

last: Joshi

summary: Defining the function of TEAD transcription factors in myogenic differentiation has proved elusive due to overlapping expression and functional redundancy. Here, we show that siRNA silencing of either Tead1, Tead2 or Tead4 did not effect differentiation of primary myoblasts (PMs) while their simultaneous knockdown strongly impaired differentiation. In contrast in C2C12 cells, silencing of Tead1 or Tead4 impaired differentiation showing a differential requirement for these factors in PMs and C2C12 cells that involved both differential regulation of their expression and intracellular localisation. Through integration of Tead1 and Tead4 ChIP-seq with chromatin modifications, we identify active enhancers associated with genes activated during C2C12 cell differentiation that are bound by combinations of Tead4, Myod1 or Myog and show a signature of frequently co-occuring motifs. We show that distinct but overlapping sets of genes are deregulated by Tead silencing in C2C12 cells and PMs therefore describing for the first time in a comprehensive manner the specific and redundant regulatory roles of Tead factors in myogenic differentiation. We also performed ChIP-seq from mouse muscle in vivo identifying a set of highly transcribed muscle cell-identity genes and revealing that Tead4 binds a distinct repertoire of sites in C2C12 cells and muscle.

overall_design: Examination of genome-wide gene expression changes upon simultaneous knockdown of Tead1 and Tead4 by siRNA

first: Irwin

last: Davidson

first: Stéphanie

last: Le Gras

first: Guillaume


title: Defining Age- and Lactocrine-Sensitive Elements of the Neonatal Porcine Uterine microRNA-mRNA Interactome

city: New Brunswick

first: Caroll

last: Bagnell

state: New Jersey

address: 59 Dudley Road

department: Animal Sciences

institute: Rutgers University

summary: Factors delivered to offspring in colostrum within two days of birth support neonatal porcine uterine development. The uterine mRNA transcriptome is affected by age and nursing during this period. Whether uterine microRNA (miRNA) expression is affected similarly is unknown. Objectives were to: 1) determine effects of age and nursing on porcine uterine miRNA expression between birth and postnatal day (PND) 2 using small RNA sequencing (smRNA-Seq) and; 2) define affected miRNA-mRNA interactions and associated biological processes using integrated target prediction analysis.

overall_design: Pig uterine tissues were obtained from neonatal gilts (n = 4/group) within 1 h of birth [postnatal day (PND) 0], or 48 h after nursing ad libitum (PND 2N), or feeding a commercial milk replacer (PND 2R).

first: Carol

first: Frank

middle: F

last: Bartol


title: Diverse interventions that extend mouse lifespan suppress shared age-associated epigenetic changes at critical gene regulatory regions (RNA-Seq)

overall_design: We quantify transcription via high throughput RNA sequencing in young (2 months) and old (22 months) mice that show increased longevity through a variety of interventions


title: ZSCAN5B and its primate-specific paralogs bind RNA polymerase III genes and extra-TFIIIC (ETC) sites to modulate mitotic progression

city: Urbana

first: Lisa

last: Stubbs

address: 1206 West Gregory Drive | MC-195

department: IGB

institute: University of Illinois Urbana-Champaign

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


title: ZSCAN5B and its primate-specific paralogs bind RNA polymerase III genes and extra-TFIIIC (ETC) sites to modulate mitotic progression [Mouse Placenta ChIP-Seq data set]

summary: Particularly in the context of differentiation and development, the importance of three-dimensional chromatin architecture to gene regulatory mechanisms is becoming increasingly clear.  The most ancient known mechanism of chromatin organization involves TFIIIC, a transcription factor (TF) that recruits RNA polymerase III (Pol III) for transcription of tRNA and other types of non-coding RNA genes. From yeast to mammals, TFIIIC binds to tRNA genes (tDNAs) and scattered “extra-TFIIIC” (ETC) loci and serves to tether these loci together as anchors of chromatin loops. TFIIIC activities are modulated by MAF, MYC, and other TF proteins that are still unidentified.  Here we identify the ZSCAN5 TF family - including mammalian ZSCAN5B and its primate-specific paralogs - as proteins that occupy mammalian Pol III promoters and ETC sites.  We show that ZSCAN5B binds with high specificity to a conserved subset of tDNA loci and other Pol III genes in human and mouse and that primate-specific ZSCAN5A and ZSCAN5D also bind Pol III genes, although ZSCAN5D preferentially localizes to MIR SINE- and LINE2-associated ETC sites. ZSCAN5 genes are expressed in proliferating cell populations and are cell-cycle regulated, and gene expression data suggested that they might cooperate to regulate basic cellular functions including mitotic progression. Consistent with this predicted role, ZSCAN5A knockdown led to increasing numbers of cells in mitotic cells and aneuploidy in cultured cells. Together these data implicate ZSCAN5 genes in regulation of Pol III gene transcription and nearby Pol II genes, ultimately influencing cell cycle progression and differentiation in a variety of tissues.

overall_design: We used a custom made polyclonal antibody to the mouse Zscan5b protein for Chromatin immunoprecipitation (ChIP) experiments in chromatin prepared from the fetal portion of embryonic placenta at 17.5 days.  ChIP was done in duplicate samples and compared to genomic input control.

first: Younguk

last: Sun


title: Transcriptome of iPSC-derived Cerebral Organoids with Heterozygous Knockout in CHD8

city: Bronx

first: Deyou

last: Zheng

address: 1301 Morris Park Ave

institute: Albert Einstein College of Medicine

summary: CHD8 (chromodomain helicase DNA binding protein 8), which codes for a member of the CHD family of ATP-dependent chromatin-remodeling factors, is the most commonly mutated gene in autism spectrum disorders (ASD) identified in exome-sequencing studies. Loss of function mutations in the gene have also been found in schizophrenia (SZ) and intellectual disabilities, and affects cancer cell proliferation. To better understanding the molecular links between CHD8 functions and ASD, we have applied the CRISPR/Cas9 technology to knockout (KO) one copy of CHD8 in induced pluripotent stem cells (iPSCs) and build cerebral organoids, a model for the developing telencephalon. RNA-seq was carried out on KO organoids (CHD8+/-) and isogenic controls (CHD8+/+). Differentially expressed genes (DEGs) revealed an enrichment of genes involved in neurogenesis, forebrain development, Wnt/β-catenin signaling and axonal guidance. The SZ and bipolar disorder (BD) candidate gene TCF4 was significantly upregulated. Our CHD8 KO DEGs were significantly overlapped with those found in a transcriptome analysis using cerebral organoids derived from a family with idiopathic ASD and another transcriptome study using iPS cell-derived neurons from patients with BD, a condition characterized in a subgroup of patients by dysregulated WNT/β-catenin signaling. Overall, the findings show that distinct ASD, SZ and BD candidate genes converge on common molecular targets - an important consideration for developing novel therapeutics in genetically heterogeneous complex traits.

overall_design: iPSCs derived from a healthy subject were transduced with CRISPR/Cas9 vectors with  single guide RNA sequences to target the N-terminal of CHD8 protein to generate truncated mutations. The CHD8+/- iPSC lines were used to generate cerebral organoids for RNA-seq analysis, together with samples prepared from the parental clones, for a total of 6 samples (two biological replicates of wild-type (WT) and 4 biological replicates of CHD8+/-).

first: Ping

last: Wang

first: Ryan

last: Mokhtari

first: Erika

last: Pedrosa

first: Michael

last: Kirschenbaum

first: Can

last: Bayrak

first: Herbert

last: Lachman


title: Comparison of whole-genome bisulfite sequencing library preparation strategies identifies sources of biases affecting DNA methylation data

city: Cambridge

first: Gavin

last: Kelsey

address: Babraham Research Campus

institute: The Babraham Institute

summary: We performed a parallel analysis of commonly used pre- and post-bisulfite WGBS library preparation protocols for their performance and quality of sequencing outputs. Our results show that bisulfite conversion per se generates pronounced sequencing biases, and subsequent fragmentation and amplification steps lead to several-fold overrepresentation of these artefacts. Standard pre-bisulfite library preparation methods lead to a significantly biased genomic sequence representation and a marked overestimation of methylation levels. We have integrated a bias diagnostic tool in the Bismark package and propose that amplification-free and post-bisulfite procedures should become the gold standard for WGBS library preparation.

overall_design: WGBS (BS-seq) was performed on unmethylated or in vitro methylated triple-Dnmt-KO (TKO) mouse embryonic stem cells with two bisulfite conversion protocols - with either alkaline or heat denaturation. MeDIP input was used as a non-bisulfute converted genomic DNA control.

first: Nelly

last: Olova

first: Felix

last: Krueger

first: Simon

middle: R

last: Andrews

first: Miguel

last: Branco

first: Wolf

last: Reik


title: Germline- and soma-specific heritable epigenetic silencing at an endogenous locus

first: Olga

last: Minkina

address: 16 Divinity Ave.

department: Molecular and Cellular Biology

institute: Harvard University


title: Germline- and soma-specific heritable epigenetic silencing at an endogenous locus (RNA-Seq)

summary: The emergence and transmission of epigenetic signals across generations can quickly and efficiently alter gene expression in a population. We describe an epigenetic silencing signal whose initiation, transmission properties, genetic requirements and site of action are distinct from previously described epigenetic inheritance in C. elegans. A multi-copy transgene containing the region upstream of sid-1 silences sid-1 and upstream genes. Once  established, silencing is stable in the absence of the array and can be maintained without selection for 13 generations. We show that the silenced state can be transmitted to progeny in the absence of the silenced locus, but that inherited silencing is dependent on the nuclear RNAi Argonaute HRDE-1, which stabilizes silencing siRNAs that target sid-1 exons. Notably, at each generation, the RNAi-dependent germline silenced sid-1 locus transitions to a chromatin-dependent silenced state in somatic cells, indicating that the mechanisms of transgenerational silencing in the soma and germline are distinct.

overall_design: RNA-seq in wild-type (4 samples, 2 replicates per experiment) and Psid-1::gfp 4 samples, 2 replicates per experiment) worms

first: Craig

middle: P

last: Hunter


title: A single miR390 targeting event is sufficient for triggering TAS3-tasiRNA biogenesis in Arabidopsis.

city: Zürich

first: Alexis

last: Sarazin

address: Universitätstrasse 2

country: Switzerland

institute: ETH

summary: In plants, tasiRNAs form a class of endogenous secondary siRNAs produced through the action of RNA-DEPENDENT-RNA-POLYMERASE-6 (RDR6) upon microRNA-mediated cleavage of non-coding TAS RNAs. In Arabidopsis thaliana, TAS1, TAS2 and TAS4 tasiRNA production proceeds via a single cleavage event mediated by 22nt-long or/and asymmetric miRNAs in an ARGONAUTE-1 (AGO1)-dependent manner. By contrast, tasiRNA production from TAS3 seems to follow the so-called ‘two-hit’ process, where dual targeting of TAS3, specifically mediated by the 21nt-long, symmetric miR390, initiates AGO7-dependent tasiRNA production. Interestingly, features for TAS3 tasiRNA production differ in other plant species and we show here that such features also enable TAS3 tasiRNA biogenesis in Arabidopsis, and that a single miR390 targeting event is, in fact, sufficient for this process, suggesting that the ‘one-hit’ model underpins all the necessary rudiments of secondary siRNA biogenesis from plant TAS transcripts. Further results suggest that the two-hit configuration likely enhances the fidelity of tasiRNA production and, hence, the accuracy of downstream gene regulation. Finally, we show that a ‘non-cleavable one-hit’ process allows tasiRNA production from both TAS1 and TAS3 transcripts, indicating that RDR6 recruitment does not require miRNA cleavage, nor does the recruitment, as we further show, of SUPRRESSOR-OF-GENE-SILENCING-3, indispensable for tasiRNA generation.

overall_design: We have analysed the sRNA populations from 5 different reporter lines designed to test the biogenesis of tasiRNAs from TAS3. These lines were generated using artificial tasiRNA constructs carrying mutations in either one or both of the miR390 target sites.

first: Felipe

last: de Felippes

first: Antonin

last: Marchais

first: Stefan

last: Oberlin

first: Olivier

last: Voinnet


title: ZSCAN5B and its primate-specific paralogs bind RNA polymerase III genes and extra-TFIIIC (ETC) sites to modulate mitotic progression [Human cell line ChIP-Seq data set]

first: Lisa

last: Stubbs

overall_design: Antibodies specific to ZCSAN5A, ZCSAN5B and ZCSAN5D were used in Chromatin immunoprecipitation (ChIP) experiments in chromatin from the BeWo choriocarcinoma cells; ZSCAN5B was also used in HEK 293 human embryonic kidney cell lines.  Experiments were done in duplicate and compared to genomic input controls.

first: Younguk

last: Sun


title: RNA substrate length as an indicator of exosome interactions in vivo

status: Public on Feb 15 2017

city: Edinburgh

first: Clementine

last: Delan-Forino

address: Michael Swann Building 5.1, King's Buildings, Mayfield Road

institute: University of Edinburgh - WTCCB

summary: The exosome complex plays key roles in RNA processing and degradation in Eukaryotes and Archaea. Outstanding structural studies have identified multiple pathways for RNA substrates into the exosome complex in vitro, but determining which pathways are preferentially followed by individual RNA species in vivo remains challenging. Here we attempted to address this important question by an RNase protection approach. We applied an in vivo RNA-protein crosslinking technique to the exosome component Rrp44/Dis2, which has both endonuclease and exonuclease activity. The resulting high-throughput sequence reads were stratified by length of the cDNA. This should reflect the length of the RNA fragments, and therefore the RNA region that was protected by exosome binding. Striking effects were seen for short fragments, which presumably arise from RNA species that follow the “direct access” route to the Rrp44 exonuclease active site. These were strongly elevated in strains lacking exonuclease activity and, surprisingly, suppressed by point mutations in the endonuclease active site. This indicates that the short fragments are degraded by the exonuclease activity of Rrp44, but also suggest that they are generated by the endonuclease activity. Confident mapping of the short reads is challenging but it seems clear that they are derived from only a subset of exosome targets. In particular, pre-rRNA species, which are major exosome targets, are strongly disfavored for the generation of short reads.

overall_design: 1 sample was analysed from strains carrying HTP tagged protein.

first: David

last: Tollervey


title: SC3-consensus clustering of single cell RNA-Seq data

first: Anthony

last: Green

state: Cambridgeshire

address: Hills Road

department: Department of heamatology

institute: University of Cambridge

summary: We report a new unsupervised clustering tool for single cell RNA-seq data called SC3. We show that biologically relevant information can be obtained from preneoplastic cells of patients with myeloprolifertive disease.

overall_design: examination of three different patients with myeloproloferative disease

first: Kristina

last: Kirschner


title: Germline- and soma-specific heritable epigenetic silencing at an endogenous locus (small RNA-Seq)

overall_design: Small RNA-seq in wild-type (2 replicates), Psid-1::gfp (4 replicated), hrde-1;Psid-1::gfp (2 replicates), non-array silenced F6 (2 replicates)  and non-array de-silenced F21 (2 replicates) worms


title: ZSCAN5B and its primate-specific paralogs bind RNA polymerase III genes and extra-TFIIIC (ETC) sites to modulate mitotic progression [SiRNA RNA-Seq data set]

overall_design: Human HEK 293 cells were treated with siRNA designed to knock down RNA from ZSCAN5A or ZSCAN5B genes respectively, or a scrambled negative control siRNA. Gene expression was compared between knocked down samples and control samples to determine differentially expressed genes.


title: Analysis of C. elegans muscle transcriptome using trans-splicing-based RNA tagging (SRT)

first: Xiaopeng

last: Ma

state: Outside US and Canada

address: HaiDian district

department: Biology

institute: Tsinghua Univeristy

summary: Current approaches to profiling tissue-specific gene expression in C. elegans require delicate manipulation and are difficult under certain conditions, e.g. from dauer or aging worms. We have developed an easy and robust method for tissue-specific RNA-seq by taking advantage of the endogenous trans-splicing process. In this method, transgenic worms are generated in which a spliced leader (SL) RNA gene is fused with a sequence tag and driven by a tissue-specific promoter. Only in the tissue of interest, the tagged SL RNA gene is transcribed and then trans-spliced onto mRNAs. The tag allows enrichment and sequencing of mRNAs from that tissue only. As a proof of principle, we profiled the muscle transcriptome, which showed high coverage and efficient enrichment of muscle specific genes, with low background noise. To demonstrate the robustness of our method, we profiled muscle gene expression in dauer larvae and aging worms, revealing gene expression changes consistent with the physiology of these stages. The resulting muscle transcriptome also revealed 461 novel RNA transcripts, likely muscle-expressed long non-coding RNAs. In summary, the splicing-based RNA tagging (SRT) method provides a convenient and robust tool to profile trans-spliced genes and identify novel transcripts in a tissue-specific manner, with a low false positive rate.

overall_design: the tissue-specific transcriptome was analysed by SRT method using U2-3 promoter for multiple-tissue profiling, myo-3 promoter for muscle-specific profiling, rgef-1 promoter for pan-neuron specific profiling with 2 biological replicates for most cases.

first: Ge

last: Zhan

first: Monica

last: Sleumer

first: Siyu

last: Chen

first: Weihong

last: Liu

first: Michael

middle: Q

last: Zhang

first: Xiao


title: Measuring signaling and RNA-Seq in the same cell links gene expression to dynamic patterns of NF-κB activation

city: Stanford

middle: Ashley

last: Van Valen

address: 443 Via Ortega, Shriram Center Rm 62

department: Bioengineering

institute: Stanford

summary: We measure translocation dynamics of NF-κB and perform single-cell RNA-sequencing in the same individual cell. We use this data to identify genes that are responsive to the dynamics of NF-κB activation

overall_design: Single-cell RNA sequencing libraries (Smart-Seq2) for ~800 RAW 264.7 cells across 10 libraries are given. Cells were stimulated with LPS on a fluidigm C1 chip - data for both stimulated and unstimulated (control) cells are provided.

first: Keara

last: Lane

first: Markus

last: Covert


title: TP53 modulates oxidative stress in Gata1+ erythroid cells

city: Minneapolis

first: Ying

state: MN

address: 117 Pleasant St S.E.

department: Minnesota Supercomputing Institute

institute: University of Minnesota

summary: In this report, we have found that gata1 expressing erythroid cells contribute to a significant proportion of total body oxidative stress when animals were exposed to a strong pro-oxidant. RNA-seq of zebrafish under oxidative stress revealed the induction of tp53. Zebrafish carrying tp53 with mutation in its DNA binding domain were acutely sensitive to pro-oxidant exposure and displayed significant reactive oxygen species (ROS) and tp53-independent erythroid cell death resulting in an edematous phenotype. We found that a major contributing factor to ROS was increased basal mitochondrial respiratory rate without reserve. These data add to the concept that tp53, while classically a tumor suppressor and cell cycle regulator, has additional roles in controlling cellular oxidative stress.

overall_design:
We performed RNA-seq in two experiments. (1) Wild-type zebrafish embryos were exposed to 1-naphthol (vs no exposure) from 24 - 72 hpf (n = 5/group).
(2) tp53 mutant zebrafish embryos were exposed to 1-naphthol (vs no exposure) from 24 - 72 hpf  (n = 5/group).

first: Troy

last: Lund


title: Sorting zebrafish thrombocyte lineage cells with a Cd41 monoclonal antibody enriches hematopoietic stem cell activity

city: Boston

first: Leonard

last: Zon

address: 1 Blackfan Circle

department: Oncology/Hematology

institute: Boston Children's Hospital

summary: The goal of the experiment was to determine the transcriptional expression profile of zebrafish thrombocytes in order to enable comparison with mouse and human platelets.

overall_design: Thrombocyte isolation from Tg(cd41:EGFP) zebrafish peripheral blood was performed using a novel monoclonal antibody (3H9) to Cd41

first: John

last: Gansner

first: Song

last: Yang


title: A short splicing isoform of HBS1L links the cytoplasmic exosome and SKI complexes in humans

city: Warszawa

first: Tomasz

middle: M.

last: Kuliński

state: Mazowieckie

address: ul. Ks. Trojdena 4

country: Poland

institute: International Institute of Molecular and Cell Biology in Warsaw

summary: A multi-subunit exosome complex is a major eukaryotic exoribonuclease that in the cytoplasm requires the SKI complex for activity. In yeast, SKI forms a heterotetramer and delivers RNA substrates directly into the exosome channel. Such cooperation requires Ski7 protein, which links the exosome and SKI complexes. However, since the human genome does not encode an orthologue of the yeast Ski7, the factor mediating SKI and exosome linkage in human cells is unknown. Proteomic analysis revealed that the human cytoplasmic exosome interacts with HBS1LV3, a protein encoded by a newly discovered short splicing isoform of HBS1L. HBS1LV3 recruits the SKI complex to the exosome. In contrast, the canonical HBS1L variant, HBS1LV1, acting as a ribosome dissociation factor, does not associate with the exosome and instead interacts with the mRNA surveillance factor PELOTA. HBS1LV3 contains a new domain of unknown structure with the short linear motif RxxxFxxxL, which is responsible for exosome binding, and may interact with the exosome core subunit RRP43 in way that resembles the association between Rrp6 RNase and Rrp43 in yeast. Depletion of HBS1LV3 and the SKI complex helicase SKI2W similarly affected the transcriptome by strongly upregulating a large number of genes. Moreover, following HBS1LV3 or SKI2W depletion the half-lives of representative upregulated mRNAs were increased, thus supporting the involvement of HBS1LV3 and SKI2W in the same mRNA degradation pathway. In contrast, HBS1LV1 depletion had little effect on transcriptome homeostasis. Our data indicate that human HBS1LV3 is the long-sought factor that links the exosome and SKI complexes to regulate cytoplasmic mRNA decay.

overall_design: Examination of siRNA-mediated silencing in HEK293 cell lines. To identify transcripts that are degraded by cytoplasmic SKI/HBS1LV3/exosome supercomplexes, we used specific siRNAs to knock down HBS1LV1, HBS1LV3 or SKIV2L gene expression in (i) WT HEK293 cells and (ii) HEK293 cells rescued with siRNA insensitive protein.  Analyses were performed in triplicate.

first: Katarzyna

last: Kalisiak

middle: M

last: Kulinski

first: Andrzej

last: Dziembowski


title: Impact of NOD2 Genotype on the Innate Monocyte Transcriptome in Crohn’s Disease

city: Hørsholm

first: Yun

address: Bøge Alle 10

department: Health & Nutrition

country: Denmark

institute: Chr Hansen A/S

summary:
Short title: Innate Gene/miRNA MDP-response in CD

RNA-seq and small RNA-seq were used to identify the expression of gene and microRNA (miRNA) in monocytes from healthy controls and patients with Crohn's Disease (CD), with and without NOD2 (nucleotide-binding oligomerization domain-containing protein 2) single nucleotide polymorphisms (SNPs) after stimulation with muramyl dipeptide (MDP).

overall_design: Identification of the gene expression of mRNA and miRNA in monocytes from healthy controls and Crohn's patients with and without NOD2 SNPs before and after stimulation with the NOD2-agonist MDP.

first: Ole

last: Nielsen

first: Jakob

middle: B

last: Seidelin

first: Albin

last: Sandelin


title: RNA sequencing of Esophageal adenocarcinomas

city: Baltimore

first: Zhixiong

last: Wang

state: Maryland

address: 1503 E. Jefferson St., Rm. 112.

department: gastroenterology

institute: Johns Hopkins University

summary: Identify EAC-relative fusion genes,differential genes  and mutations

overall_design: 40 EACs and matched non-malignant tissues

first: Stephen

last: Meltzer


title: Ettlia oleoabundans under varying nitrate, NaCl, light and cell density conditions

city: Singapore

first: Neil

last: Clarke

address: 16 College Avenue West

country: Singapore

institute: Yale-NUS College

summary: Ettlia oleoabundans is a triacylglyceride-producing chlorophyte.  We investigated gene expression and lipid changes after 6 days under varying concentrations of nitrate and NaCl.

overall_design: E. oleoabundans was grown under phototrophic growth conditions, under varying conditions.

first: Falicia

last: Goh

first: Justin

last: Jeyakani

first: Zhenxuan

last: Yeo


title: Seb1 is recruited co-transcriptionally to non-coding regions to drive transcriptional interference

status: Public on Feb 14 2017

city: Oxford

first: Beth

last: Watts

address: South Parks Road

department: Biochemistry

institute: University of Oxford

overall_design: Refer to individual Series


title: Radiation Induced Pulmonary Fibrosis as a Model of Progressive Fibrosis: Contributions of DNA Damage, Inflammatory Response and Cellular Senescence Genes

city: Rochester

first: Tyler

last: Beach

state: NY

address: 601 Elmwood Ave., Box 850

institute: University of Rochester

summary: Utilizing an established model of Radiation Induced Pulmonary Fibrosis, low input RNA sequencing was performed on whole lung cell suspensions obtained from 12.5 Gy thorax only radiation treated C57BL/6J mice and compareed to 0 Gy (Sham irradiation) age matched controls

overall_design: Differential expression of gene transcripts identified in pooled samples of RNA obtained from 3 mice per time point and un-irradiated controls were examined at 24 hrs, 1, 4, 12, and 32 weeks post radition treatment  during the onset and progression of pulmonary fibrosis which were then compared to genes identified in literature as bieng associated with DNA damage, inflammatory response, cellular senescence and pulmonary fibrosis


title: Genome-wide maps of TBP-associated factor (TAF, Taf1) in S.cerevisiae

first: Yoo Jin

last: Joo

address: 240 Longwwod Ave.

department: BCMP

institute: HMS/Harvard University

summary: The goals of this study are to compare Next-Generation-Sequecing (NGS)-derived genome-wide occupancy of TAF (Taf1) comparing with other basal initiation components (TBP and TFIIB), histones (H3, H4, Htz1 and H4 acetylation) and histone regulator complexes (Swr1, Bdf1) in S. cerevisiae.

overall_design:
Examination of H3, H4ac, TBP, Bdf1 (SRP010134), Taf1 (SRP010134), Swr1 (SRP028548/PRJNA214015), Sua7 (SRP036647/PRJNA237486) and Htz1 (SRP035413/PRJNA235191) in normal growing cells. Examination of Taf1 and TBP (Spt15) in Rpb1- and Bdf1- nuclear depleted cells.

The following bedGraph files were generated using the following third-party SRA data:
Bdf1.bedGraph: SRP010134/SRX114994/SRR397550
Htz1.bedGraph: SRP035413/SRX433134/SRR1118262 (GEO GSE54105/GSM1308195)
Swr1.bedGraph: SRP028548/SRX332103/SRR948403
Taf1.bedGraph: SRP010134/SRX114691/SRR396796
Sua7.bedGraph: SRP036647/SRX472138/SRR1197807

first: Yoo

first: Luis

last: Soares

first: Steve

last: Buratowski


title: Histone deacetylase inhibition in triple-negative breast cancer

city: Santa Monica

first: Dave

last: Hoon

state: CA

address: 2200 Santa Monica Boulevard

department: Translational Molecular Medicine

institute: Saint John's Cancer Institute

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


title: Retrotransposon Activation Contributes to Neurodegeneration in a Drosophila TDP-43 Model of ALS

city: Cold Spring Harbor

first: Wen-Wei

last: Liao

state: New York

address: 1 Bungtown Road

institute: Cold Spring Harbor Laboratory

summary: Functional abnormality of TAR DNA-binding protein 43 (TDP-43), an aggregation-prone RNA and DNA binding protein, is observed in the vast majority of both familial and sporadic ALS cases and in ~40% of FTLD cases, but the cascade of events leading to cell death are not understood. We have expressed human TDP-43 (hTDP-43) in Drosophila neurons and glia, a model that recapitulates many of the characteristics of TDP-43-linked human disease including protein aggregation pathology, locomotor impairment, and premature death. We report that such expression of hTDP-43 impairs small interfering RNA (siRNA) silencing, which is the major post-transcriptional mechanism of retrotransposable element (RTE) control in somatic tissue. This is accompanied by de-repression of a panel of both LINE and LTR families of RTEs, with somewhat different elements being active in glia vs neurons. hTDP-43 expression in glia causes an early and severe loss of control of a specific RTE, the endogenous retrovirus (ERV) gypsy. We demonstrate that gypsy causes the degenerative phenotypes in these flies because we are able to rescue the toxicity of glial hTDP-43 either by genetically blocking expression of this RTE or by pharmacologically inhibiting RTE reverse transcriptase activity. Moreover, we provide evidence that activation of DNA damage-mediated programmed cell death underlies both neuronal and glial hTDP-43 toxicity, consistent with RTE-mediated effects in both cell types. Our findings suggest a novel mechanism in which RTE activity contributes to neurodegeneration in TDP-43-mediated diseases such as ALS and FTLD.

overall_design: Human TDP-43 expressed in glia (repo) or neurons (elav).

first: Lisa

last: Krug

first: Nabanita

last: Chatterjee

first: Rebeca

last: Borges-Monroy

last: Hearn

first: Kathleen

last: Morrill

last: Prazak

first: Nikolay

last: Rozhkov

first: Delphine

last: Theodorou

first: Molly

last: Hammell

first: Joshua

last: Dubnau


title: Genome-wide maps of chromatin accessibility of histone deacetylase inhibition in triple-negative breast cancer

summary: Analysis of chromatin accessibility changes after prolonged exposure of triple-negative breast cancer cell lines to low doses of Panobinostat (LBH589), a pan-histone deacetylase inhibitor.

overall_design: Examination of accessible chromatin after histone deacetylase inhibition in 2 triple-negative cell lines cell types.

first: Diego

last: Marzese

first: Matthew

last: Salomon

middle: S

first: Matias

last: Bustos


title: Effect of Varroa infestation, at the pupal stage, on the transcriptome of newly emerged bees, maintained under artificial conditions

city: University Park

first: David

middle: Asher

last: Galbraith

state: Pennsylvania

address: 19a Chemical Ecology Lab

institute: Penn State University

summary: The parasitic mite Varroa destructor causes both direct and indirect damage to infested honeybees but precise information about the identity of the metabolic pathways that are affected is still missing. In order to gain a deeper knwoledge about the effects of parasitization at various levels, while reducing to the miminum the impact of possible confounding factors, a standardized experiment was carried out under artificial condition.

overall_design: 15 Apis mellifera larvae (late fifth instar) were infested with none, one or three V. destructor mites (5 bees per treatment) and maintained in gealtin capsules, at 35 °C, 75% R.H., for 12 days; at eclosion, bees were individually extracted and the extract processed.

first: Francesco

last: Nazzi

first: Desiderato

last: Annoscia