title: Repression of the soma-specific transcriptome by Polycomb-repressive complex 2 promotes male germ cell development

city: Chapel Hill

first: Joshua

last: Starmer

state: NORTH CAROLINA

address: 120 Mason Farm Road

department: Genetics

institute: University of North Carolina at Chapel Hill

summary: Polycomb-repressive complex 2 (PRC2) catalyzes the methylation of histone H3 Lys27 (H3K27) and functions as a critical epigenetic regulator of both stem cell pluripotency and somatic differentiation, but its role in male germ cell development is unknown. Using conditional mutagenesis to remove the core PRC2 subunits EED and SUZ12 during male germ cell development, we identified a requirement for PRC2 in both mitotic and meiotic germ cells. We observed a paucity of mutant spermatogonial stem cells (SSCs), which appears independent of repression of the known cell cycle inhibitors Ink4a/Ink4b/Arf. Moreover, mutant spermatocytes exhibited ectopic expression of somatic lamins and an abnormal distribution of SUN1 proteins on the nuclear envelope. These defects were coincident with abnormal chromosome dynamics, affecting homologous chromosome pairing and synapsis. We observed acquisition of H3K27me3 on stage-specific genes during meiotic progression, indicating a requirement for PRC2 in regulating the meiotic transcriptional program. Together, these data demonstrate that transcriptional repression of soma-specific genes by PRC2 facilitates homeostasis and differentiation during mammalian spermatogenesis.

overall_design: Examination of two different types of histone modifications (H3K27me3 and H3K4me3) in spermatocytes at two different stages of development (P12 and P17) and the DNA binding protein, EED, in spermatocytes at one stage of development, P17. At P12, there was a single replicate of H3K27me3, H3K4me3 and input.  At P17, there were two replicates of H3K27me3, H3K4me3 and input and a single replicate for EED.

first: Weipeng

last: Mu

first: Andrew

last: Fedoriw

first: Della

last: Yee

first: Terry

last: Magnuson


title: Transcriptional changes in the aging mouse hippocampus

status: Public on Oct 01 2014

city: Cork

first: Roman

middle: M

last: Stilling

address: Western Road

department: Anatomy and Neuroscience

country: Ireland

institute: University College Cork

type: Expression profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Adenylation of maternally inherited microRNAs by Wispy

city: Seoul

first: Yeon

last: Choi

address: Building 504 Room 501, School of Biological Sciences, Seoul National University, 1 Gwanangno, Gwanak-gu

department: School of Biological Sciences

country: South Korea

institute: Seoul National University

type: Non-coding RNA profiling by high throughput sequencing

summary: Early development depends heavily on accurate control of maternally inherited mRNAs, and yet it remains unknown how maternal microRNAs (miRNAs) are regulated during maternal to zygotic transition (MZT). We here find that maternal miRNAs are highly adenylated at their 3' ends in mature oocytes and early embryos. Pervasive adenylation is observed in oocytes of fly, sea urchin and mouse, indicating that maternal miRNA adenylation may be widely conserved in animals. We identify Wispy as the enzyme responsible for miRNA adenylation in flies. Wispy is known to be expressed specifically in oocytes and early embryos and function as a noncanonical poly(A) polymerase. Knockout of wispy abrogates miRNA adenylation and induces miRNA accumulation in fly eggs whereas overexpression of Wispy increases adenylation and reduces miRNA levels in S2 cells. Adenylation occurs on both the 5p and 3p miRNAs, indicating that Wispy acts on miRNAs after Dicer processing. We further find that Wispy interacts with Ago1 through protein-protein interaction, which may allow the effective and selective adenylation of miRNAs. Thus, adenylation may contribute to the clearance of maternally deposited miRNAs during MZT. Our work provides the first mechanistic insights into the regulation of maternal miRNAs and illustrates the importance of RNA tailing in development.

sample_organism:
Drosophila melanogaster
Danio rerio

platform_organism:
Drosophila melanogaster
Danio rerio

overall_design: MiRNA expression and modification profile during early embryo development of fruit fly and zebra fish using high throughput sequencing

first: Mihye

last: Lee

first: Kijun

last: Kim

first: Hua

last: Jin

first: Jaechul

last: Lim

first: Minsun

last: Jeong

first: Antonio

middle: J

last: Giraldez

first: V

middle: N


title: Genome accessibility is widely preserved and locally modulated during mitosis [Dnase-seq]

city: University Park

first: Ross

last: Hardison

state: PA

address: 303 Wartik Lab

country: USA

institute: Pennsylvania State University

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Mitosis entails global alterations to chromosome structure and nuclear architecture, concomitant with transient silencing of transcription. How cells transmit transcriptional states through mitosis remains incompletely understood. While many nuclear factors dissociate from mitotic chromosomes, the observation that certain nuclear factors and chromatin features remain associated with individual loci during mitosis originated the hypothesis that they could provide transcriptional memory through mitosis. To obtain the first genome-wide view of the dynamics of chromatin structure during mitosis, we compared the DNase sensitivity of interphase and mitotic chromatin at two stages of cellular maturation in a rapidly dividingmurine erythroblastmodel. Despite global chromosome condensation visible during mitosis at the microscopic level, the chromatin accessibility landscape is largely unaltered. However, mitotic chromatin accessibility is locally dynamic, with individual loci maintaining none, some, or all of their interphase accessibility. Mitotic reduction in accessibility occurs primarily within narrow, highly hypersensitive sites that frequently coincide with transcription factor binding sites, whereas broader domains of moderate accessibility tend to be more stable. In mitosis, proximal promoters generally maintain their accessibility, whereas distal regulatory elements preferentially lose accessibility. Promoters with the highest degree of accessibility preservation in mitosis tend to also be accessible across many murine tissues in interphase. Transcription factor GATA1 exerts site-specific changes in interphase accessibility that are most pronounced at distal regulatory elements, but does not visibly influence mitotic accessibility. We conclude that features of open chromatin are remarkably stable through mitosis and are modulated at the level of individual genes and regulatory elements.

overall_design:
Dnase-Seq data is integrated with Chip-seq [GSE36589, GSE30142] and RNA-seq to examine epigentic changes in mitosis.
We performed DNase-seq on  two cell lines, G1E and G1E-ER4, both on an asynchronus population, and on a sample of cells  in mitosis; each of the 4 experiments in triplicate.

first: Christapher

middle: S

last: Morrissey

first: Chris

last: Hsiung

first: Maheshi

last: Udugama

first: Christopher

middle: L

last: Frank

first: Cheryl

middle: A

last: Keller

first: Songjoon

last: Baek

first: Belinda

last: Giardine

first: Gregory

middle: E

last: Crawford

first: Myong-Hee

last: Sung

middle: C

first: Gerd

last: Blobel


title: ASCL1 is a lineage oncogene providing therapeutic targets for high-grade neuroendocrine lung cancers

city: Dallas

first: Tao

last: Wang

state: TX

address: 5323 Harry Hines Boulevard

institute: UT Southwestern Medical Center

summary: ChIP-Seq analysis performed on 5 ASCL1(+) cell lines and 2 ASCL1 (-) cell lines in order to understand the transcriptome of ASCL1 as it pertains to high-grade neuroendocrine lung cancers

overall_design: 5 ASCL1(+) lung cancer cell lines and 2 ASCL1(-) lung cancer cell lines were compared using ChIP-Seq analysis

first: Alexander

last: Augustyn

first: Mark

middle: D

last: Borromeo

first: Yang

last: Xie

first: Melanie

middle: H

last: Cobb

first: Jane

last: Johnson

first: John

last: Minna


title: Two distinct promoter architectures centered on dynamic nucleosomes control ribosomal protein gene transcription

city: Lausanne

first: Jacques

last: Rougemont

address: Station 15

department: School of Life Sciences

country: Switzerland

institute: EPFL

summary: In yeast, ribosome production is controlled transcriptionally by tight coregulation of the 138 ribosomal protein genes (RPGs). RPG promoters display limited sequence homology, and the molecular basis for their coregulation remains largely unknown. Here we identify two prevalent RPG promoter types, both characterized by upstream binding of the general transcription factor (TF) Rap1 followed by the RPG-specific Fhl1/Ifh1 pair, with one type also binding the HMG-B protein Hmo1. We show that the regulatory properties of the two promoter types are remarkably similar, suggesting that they are determined to a large extent by Rap1 and the Fhl1/Ifh1 pair. Rapid depletion experiments allowed us to define a hierarchy of TF binding in which Rap1 acts as a pioneer factor required for binding of all other TFs. We also uncovered unexpected features underlying recruitment of Fhl1, whose forkhead DNA-binding domain is not required for binding at most promoters, and Hmo1, whose binding is supported by repeated motifs. Finally, we describe unusually micrococcal nuclease (MNase)-sensitive nucleosomes at all RPG promoters, located between the canonical +1 and –1 nucleosomes, which coincide with sites of Fhl1/Ifh1 and Hmo1 binding. We speculate that these ‘‘fragile’’ nucleosomes play an important role in regulating RPG transcriptional output.

overall_design: Genome-wide examination of binding sites for transcription factors controlling ribosomal protein genes expression and nucleosome positions in budding yeast cells

first: Britta

last: Knight

first: Slawomir

last: Kubik

first: Philippe

last: Jacquet

first: David

last: Shore


title: Genome-wide DNA methylation map of Zebrafish liver

city: Dunedin

first: Aniruddha

last: Chatterjee

state: Otago

address: 270 Great king Street

department: Pathology

country: New Zealand

institute: University of Otago

type: Methylation profiling by high throughput sequencing

summary: Using a pooled (n=10) zebrafish liver DNA, we generated base-resolution DNA methylation maps to document epigenetic landscape in zebrafish genome.

overall_design: Here we generated single-nucleotide resoultion DNA methylation map of zebrafish pooled liver sample using Reduced Representation Bisulfite Sequencing (RRBS)

first: Peter

last: Stockwell

first: Julia

last: Horsfield

first: Ian

last: Morison

first: Shinichi

last: Nakagawa


title: Dopamine Signaling leads to loss of Polycomb Repression and Aberrant Gene [ChIP-Seq]

city: Copenhagen N

first: Mads

last: Lerdrup

address: Blegdamsvej 3C

department: Department of Cellular and Molecular Medicine

country: Denmark

institute: University of Copenhagen

summary: Polycomb group (PcG) proteins bind to and repress genes in embryonic stem cells and through lineage commitment to the terminal differentiated state. PcG repressed genes are commonly characterized by the presence of the epigenetic histone mark, H3K27me3, catalyzed by the Polycomb repressive complex 2. Here, we present in vivo evidence for a previously unrecognized plasticity of PcG-repressed genes in terminal differentiated brain neurons of parkisonian mice. We show that acute administration of the dopamine precursor, L-DOPA, induces a remarkable increase in H3K27me3S28 phosphorylation. The induction of the H3K27me3S28p histone mark specifically occurs in medium spiny neurons expressing the dopamine D1 receptors and is dependent on Msk1 kinase activity and DARPP-32-mediated inhibition of protein phosphatase-1. Chromatin immunoprecipitation (ChIP) experiments showed that increased H3K27me3S28p was accompanied by reduced PcG binding to regulatory regions of genes. An analysis of the genome wide distribution of L-DOPA induced H3K27me3S28 phosphorylation by ChIP sequencing (ChIP-seq) in combination with expression analysis by RNA-sequencing (RNA-seq) showed that the induction of H3K27me3S28p correlated with increased expression of a subset of PcG repressed genes. We found that induction of H3K27me3S28p persisted during chronic L-DOPA administration to parkisonian mice and correlated with aberrant gene expression. We propose that dopaminergic transmission can activate PcG repressed genes in the adult brain and thereby contribute to long-term maladaptive responses including the motor complications, or dyskinesia, caused by prolonged administration of L-DOPA in Parkinsons disease.

overall_design: 8 ChIP-seq libraries were generated from a pool of chromatin generated from 45 mice.

first: Erik

last: Södersten

first: Klaus

last: Hansen

first: Gilberto

last: Fisone

first: Michael

last: Feyder


title: H3K9me3 ChIP-Seq of baboon SVZ primary cells

city: San Antonio

first: Chin-Hsing Annie

last: Lin

address: One UTSA Circle

department: Biology

institute: University of Texas at San Antonio

summary: Histone 3 Lysine 9 (H3K9) methylation is known to be associated with pericentric heterochromatin and important in genomic stability. In this study, we show that trimethylation at H3K9 (H3K9me3) is enriched in an adult neural stem cell niche- the subventricular zone (SVZ) on the walls of the lateral ventricle in both rodent and non-human primate baboon brain.  Previous studies have shown that there is significant correlation between baboon and human regarding genomic similarity and brain structure, suggesting that findings in baboon are relevant to human. To understand the function of H3K9me3 in this adult neurogenic niche, we performed genome-wide analyses using ChIP-Seq (chromatin immunoprecipitation and deep-sequencing) and RNA-Seq for in vivo SVZ cells purified from baboon brain. Through integrated analyses of ChIP-Seq and RNA-Seq, we found that H3K9me3-enriched genes associated with cellular maintenance, post-transcriptional and translational modifications, signaling pathways, and DNA replication are expressed, while genes involved in axon/neuron, hepatic stellate cell, or immune-response activation are not expressed. As neurogenesis progresses in the adult SVZ, cell fate restriction is essential to direct proper lineage commitment. Our findings highlight that H3K9me3 repression in undifferentiated SVZ cells is engaged in the maintenance of cell type integrity, implicating a role for H3K9me3 as an epigenetic mechanism to control cell fate transition within this adult germinal niche.

overall_design: SVZ H3K9me3 ChIP-seq profile of an adult baboon subventricular zone was generated by deep sequencing with Illumina HiSeq2000

first: Richard

last: Sandstrom

first: Christopher

middle: T

last: Rhodes

first: Chin-Hsing


title: Molecular mechanisms of two-component system RhpRS regulating type III secretion system in Pseudomonas syringae

city: Hong Kong

first: Xin

last: Deng

address: To Yuen Building

department: Biomedical Sciences

country: Hong Kong

institute: City University of Hong Kong

summary: Pseudomonas syringae uses two-component system RhpRS to regulate the expression of type III secretion system (T3SS) genes and bacterial virulence. However, the molecular mechanism and the regulons of RhpRS have yet to be fully elucidated. Here we show that RhpS functions as an autokinase, an RhpR kinase, and a P-RhpR phosphatase. RhpR can also be phosphorylated by small phosphodonor acetyl phosphate. A specific RhpR-binding site containing an inverted repeat (IR) motif GTATC-N6-GATAC, was mapped to its own promoter by DNase I footprint analysis. Electrophoretic mobility shift assay (EMSA) indicated that P-RhpR has higher binding activity than RhpR to the IR motif. To identify additional targets of RhpR in P. syringae, we performed chromatin immunoprecipitation followed by high-throughput DNA sequencing (ChIP-seq), which detected 167 enriched loci including the hrpR promoter, indicating the direct regulation of T3SS cascade genes by RhpR. Genome-wide microarray analysis showed that, besides the T3SS cascade genes, RhpR differentially regulates a large set of genes of various functions in response to different growth conditions. Together, these results suggested that RhpRS is a global regulator that allows P. syringae to sense and respond to environmental changes to coordinate the T3SS and many other biological processes.

overall_design: ChIP-seq analysis of RhpR


title: Profiling microRNAs in Eucalyptus grandis reveals no mutual relationship between alterations in miR156 and miR172 expression and adventitious root induction during development

city: Bet Dagan

first: Tzahi

last: Arazi

state: ISRAEL

address: Institute of Plant Sciences, ARO the Volcani Center

department: Institute of Plant Sciences

country: Israel

institute: ARO - Volcani Center

summary:
Background: The change from juvenile to mature phase in woody plants is often accompanied by a gradual loss of rooting ability, as well as by reduced microRNA (miR) 156 and increased miR172 expression.
Results: We characterized the population of miRNAs of Eucalyptus grandis and compared by Northern blot the gradual reduction in miR156 and increase in miR172 expression during development to the loss of rooting ability.  Forty known and eight novel miRNAs were discovered and their predicted targets are listed. The expression pattern of nine miRNAs was determined during adventitious root formation in juvenile and mature cuttings. While the expression levels of miR156 and miR172 were inverse in juvenile and mature tissues, no mutual relationship was found between high miR156 expression and rooting ability, or high miR172 expression and loss of rooting ability.  This is shown both in E. grandis and also in E. brachyphylla, in which explants that underwent rejuvenation in tissue culture conditions were also examined.
Conclusions: It is suggested that in these Eucalyptus species, there is no correlation between the switch of miR156 with miR172 expression in the stems and the loss of rooting ability.

overall_design: Examination of microRNA in seedlings of Eucalyptus grandis

first: Aviv

last: Levy

last: Szwerdszarf

first: Mohamad

last: Abu-Abied

first: Inna

last: Mordehaev

first: Yossi

last: Yaniv

first: Joseph

last: Riov

first: Einat

last: Sadot


title: Dopamine Signaling leads to loss of Polycomb Repression and Aberrant Gene [RNA-Seq]

first: Mads

last: Lerdrup

overall_design: 12 mice were used for RNAseq, 4 conditions, 3 mice per condition.


title: Dopamine Signaling leads to loss of Polycomb Repression and Aberrant Gene


title: Nova1 is a master regulator of alternative splicing in pancreatic beta cells

city: Bruxelles

first: Olatz

last: Villate

address: Route de Lennik 808 CP618

department: Faculty of Medecine

country: Belgium

institute: Universite Libre de Bruxelles

summary: Alternative splicing (AS) is a fundamental mechanism for the regulation of gene expression. It affects more than 90% of human genes but its role in the regulation of pancreatic beta cells, the producers of insulin, remains unknown. Our recently published data indicated that the “neuron specific” Nova1 splicing factor is expressed in pancreatic beta cells. We have presently coupled specific knockdown (KD) of Nova1 with RNA-sequencing to determine all splice variants and downstream pathways regulated by this protein in beta cells. Nova1 KD altered the splicing of nearly 5000 transcripts. Pathway analysis indicated that these genes are involved in exocytosis, apoptosis, insulin receptor signalling, splicing and transcription. In line with these findings, Nova1 silencing inhibited insulin secretion and induced apoptosis basally and after cytokine treatment in rodent and human beta cells. These observations identify a novel layer of regulation of beta cell function, namely AS controlled by key splicing regulators such as Nova1.

overall_design: 3 batch of primary rat pancreatic beta cells were examined under 2 conditions: control and with Nova1 splicing factor knock-down

first: Jean-Valery

last: Turatsinze

last: Sammeth

first: Decio

last: Eizirik


title: Genome-wide characterizaion of H3K27me3 levels upon inhibition of JMJD3 histone demethylase using a small molecule inhibitor in T cell leukamia

city: NEW YORK

first: Panagiotis

last: Ntziachristos

address: 522 FIRST AVENUE

department: PATHOLOGY

institute: NYU MEDICAL SCHOOL

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus


title: Characterizing the contrasting roles of JMJD3 and UTX histone demethylases in T cell acute lymphoblastic leukemia [JMJD3_CUTLL1_ChIP-seq]

summary: T-cell acute lymphoblastic leukemia (T-ALL) is an immature hematopoietic malignancy driven mainly by oncogenic activation of NOTCH1 signaling. In this study we assayed for genome-wide localization of JMJD3 enrichment. This piece of data was further integrated to expression changes using RNA sequencing as well as ChIP-Sequencing analysis of H3K27me3 upon genomic/genetic knock-down or chemical inhibition of JMJD3 and UTX. These results, coupled to genomic analysis of primary samples for the genomic status of the UTX gene in T-ALL, helped us to identify a hitherto unknown role of JMJD3 as an oncogenice facilitator in leukemia whereas UTX plays a tumor suppressor role.

overall_design: JMJD3 ChIP: 10 million cells were used for the ChIP and precipitated using 10micrograms of antibody (abgent, AP1022a) against human JMJD3.

first: Aristotelis

last: Tsirigos


title: Characterizing the contrasting roles of JMJD3 and UTX histone demethylases in T cell acute lymphoblastic leukemia [shJMJD3_ChIP-seq]

summary: T-cell acute lymphoblastic leukemia (T-ALL) is an immature hematopoietic malignancy driven mainly by oncogenic activation of NOTCH1 signaling. In this study we abrogated the expression of JMJD3 (KDM6B) and UTX (KDM6A) H3K27me3 demethylases in human T-ALL lines and assayed for genome-wide changes in H3K27me3 levels. This piece of data was further integrated to expression changes using RNA sequencing in the same cells as well as ChIP-Sequencing analysis of H3K27me3 and JMJD3 genome-wide analysis from treatment of T-ALL lines with the GSKJ4 inhibitor. These results, coupled to genomic analysis of primary samples for the genomic status of the UTX gene in T-ALL, helped us to identify a hitherto unknown role of JMJD3 as an oncogenice facilitator in leukemia whereas UTX seems to play a tumor suppressor role.

overall_design: Half to one million cells were treated with micrococcal nuclease (MNASE) to generate mononucleosomal particles and an adaptation of the Upstate ChIP protocol was used.


title: Histone H3.3 and its proteolytically processed form drive a cellular senescence program

city: New York

first: Zichen

last: Wang

state: NY

address: 1468 Madison Ave

department: Pharmacological Sciences

institute: Icahn School of Medicine at Mount Sinai

summary: In this study, we have uncovered novel proteolytic processing of the histone H3 tail in senescence models in primary fibroblasts and melanocytes. Cleavage of H3 tail occurs at two distinct residues and is mediated by Cathepsin L. We show that variant H3.3 is preferentially cleaved, and that cleaved histones are associated with chromatin and incorporated into nucleosomes.  We also found that the histone chaperone ASF1a is required for chromatin incorporation of the cleaved histone species. Further, we show that overexpression of cleaved H3.3 induces a senescence program in fibroblasts in the absensence of oncogenic signaling.

overall_design: For the RNA-seq studies, growing IMR90 fibroblasts were compared to cells induced to senesce via oncogene activation or cleaved H3.3 overexpression. Growing controls consist of IMR90 cells infected with empty retroviral construct pBabe and grown under normal conditions for 13 days prior to RNA isolation. For oncogene-induced senescence samples, IMR90s carrying a tamoxifen-inducible H-RasV12 retroviral construct were induced to senesce by addition of 10nM tamoxifen to the media for 8 days. Finally, IMR90s were infected with a retroviral construct expressing the cleaved form of H3.3 with a C-terminal Flag tag. RNA samples form this group were isolated at days 3 (early) and 13 (late) post-infection. In all cases, total RNA samples were isolated using RNeasy kit (Qiagen) and prepared at the Icahn School of Medicine at Mount Sinai Genomics Core Facility for poly A library construction and sequencing on IlluminaHiSeq 2500.

first: Emily

last: Bernstein

first: Luis

middle: F

last: Duarte


title: Genome-wide DNA methylation map of Zebrafish male and female brain

first: Aniruddha

last: Chatterjee

summary: Using 2 male and 2 female zebrafish (pool of 6) brain samples, we generated base-resolution DNA methylation maps to document sex-specific epigenetic differences.

overall_design: Here we generated single-nucleotide resoultion DNA methylation map of 4 zebrafish brain samples using Reduced Representation Bisulfite Sequencing (RRBS)

first: Peter

last: Stockwell

first: Julia

last: Horsfield

first: Ian

last: Morison

first: Shinichi

last: Nakagawa


title: ADAR2 reproducibly changes abundance and sequence of mature microRNAs in the mouse brain

status: Public on Oct 02 2014

city: Vienna

first: Fritz

last: Sedlazeck

state: Vienna

address: Dr. Bohr-Gasse 9

department: CIBIV

country: Austria

institute: MFPL


title: Serpinb3a contributes to early skin inflammation following allergen exposure

city: Cincinnati

first: Mario

last: Medvedovic

state: OH

address: 3223 Eden Av. ML 56

department: Department of Environmental Health

institute: University of Cincinnati

summary: Serpinb3a knockout mice show attenuated Transepidermal Water Loss (TEWL) and S100A8 expression (an early inflammatory marker) following treatment with Aspergillus fumigatus extract. The aim of the RNAseq experiments was to provided mechanistic insight for the role of Serpinb3a by identifying early differences in gene expression following allergen exposure.

overall_design: Wild type Balb/c and Serpinb3a null mouse backs were shaved and 24 hours later a patch containing 200ug of Aspergillus fumigatus extract was applied to the skin. After 3 days the patch was removed and the skin was harvested after an additional 24 hours.

first: Umasundari

last: Sivaprasad

first: Kayla

middle: G

last: Kinker

first: Gurjit

middle: K

last: Hershey


title: Micrococcal nuclease sequencing analysis for nucleosome localization of mouse fetal liver erythroblasts

city: Chicago

first: Peng

last: Ji

state: IL

address: 303 E Chicago Ave

institute: Northwestern University

summary: Nucleosomal DNA was prepared using Simple ChIP Enzymatic Chromatin IP Kit according to manufacturer’s instruction. Briefly, Nuclei were isolated from purified Ter119 negative or in vitro cultured erythroblasts. Cross-linked native chromatin was then digested with MNase into mononucleosomal DNA. Sequencing libraries were generated from nucleosomal DNA, and sequencing was carried out using the Illumina system according to the manufacturer’s specification.

overall_design: In this study, we purified chromatin from in vitro cultured mouse fetal liver erythroblasts on day 0, day 1, and day 2. The chromatins were digested by micrococcal nuclease to make mononucleosomal products, which were further analyzed by next generation sequencing analysis. We aim to determine the dynamic changes of nucleosome during terminal erythropoiesis.

first: Baobing

last: Zhao


title: Characterizing the contrasting roles of JMJD3 and UTX histone demethylases in T cell acute lymphoblastic leukemia [GSKJ4_RNA-seq]

first: Panagiotis

last: Ntziachristos

summary: T-cell acute lymphoblastic leukemia (T-ALL) is an immature hematopoietic malignancy driven mainly by oncogenic activation of NOTCH1 signaling. In this study we chemically inhibited the H3K27me3 demethylase JMJD3 using the GSKJ4 inhibitor and assayed for genome-wide changes in H3K27me3 and JMJD3 enrichment. This piece of data was further integrated to expression changes using RNA sequencing as well as ChIP-Sequencing analysis of H3K27me3 upon genomic knock-down of JMJD3 and UTX. These results, coupled to genomic analysis of primary samples for the genomic status of the UTX gene in T-ALL, helped us to identify a hitherto unknown role of JMJD3 as an oncogenice facilitator in leukemia whereas UTX seems to play a tumor suppressor role.

overall_design: Whole RNA was extracted from 1-5 million primary cells from CUTLL1 human T cell leukemia cells untreated or treated with 2micromolar GSKJ4 using the RNAeasy kit (Qiagen) according to the manufacturer’s protocol. Poly-A+ (magnetic oligodT-containing beads (Invitrogen)) or Ribominus RNA was used for library preparation. cDNA preparation and strand-specific library construction was performed using the dUTP method. Libraries were sequenced on the Illumina HiSeq 2000 using 50bp single-read method. Differential gene expression analysis was performed between knockout vs wild-type background samples. Analysis was performed using DEGseq package leading to very similar conclusions.


title: Characterizing the contrasting roles of JMJD3 and UTX histone demethylases in T cell acute lymphoblastic leukemia [short_hairpins_RNA-seq]

summary: T-cell acute lymphoblastic leukemia (T-ALL) is an immature hematopoietic malignancy driven mainly by oncogenic activation of NOTCH1 signaling. In this study we abrogated the expression of JMJD3 (KDM6B) and UTX (KDM6A) H3K27me3 demethylases in human T-ALL lines and assayed for genome-wide expression changes using RNA sequencing. This piece of data was further integrated to ChIP-Sequencing analysis of H3K27me3 from the same treatment as well as H3K27me3 and JMJD3 genome-wide analysis from treatment of T-ALL lines with the GSKJ4 inhibitor. These results, coupled to genomic analysis of primary samples for the genomic status of the UTX gene in T-ALL, helped us to identify a hitherto unknown role of JMJD3as an oncogenice facilitator in leukemia whereas UTX seems to play a tumor suppressor role.

overall_design: Whole RNA was extracted from 1-5 million T-ALL (lines) cells or primary cells using the RNAeasy kit (Qiagen) according to the manufacturer’s protocol. Poly-A+ (magnetic oligodT-containing beads (Invitrogen)) or Ribominus RNA was used for library preparation. cDNA preparation and strand-specific library construction was performed using the dUTP method. Libraries were sequenced on the Illumina HiSeq 2000 using 50bp single-read method. Differential gene expression analysis was performed for each matched knockdown vs control pairs, separately in each biological or technical replicate in each of two cell lines (CUTLL1, CEM). Three types of comparisons were tested: (a) JMJD3 knockdown vs Renilla, (b) JMJD3 knockdown vs UTX knockdown, and (c) UTX knockdown vs Renilla. Analysis was performed using both DEGseq and Cufflinks packages leading to very similar conclusions.

first: Aristotelis

last: Tsirigos


title: Genome-wide binding of two new insulator-like proteins Pita and ZIPIC

city: Giessen

first: Marek

last: Bartkuhn

state: Hessen

address: Aulweg 132

department: Biomedical Informatics and Systems Medicine

country: Germany

institute: Justus-Liebig-University Giessen

summary: Insulators are multiprotein–DNA complexes that regulate nuclear architecture. The Drosophila CP190 protein is a common cofactor of two main insulator complexes formed by the DNA-binding proteins Su(Hw) and dCTCF. Here, we have identified two new DNA-binding proteins, Pita and ZIPIC, that interact with CP190. In vitro, CP190 interacts with Pita through the BTB domain and with ZIPIC through the centrosomal targeting domain. The artificial binding sites for Pita or ZIPIC partially block enhancers and protect gene expression from PRE-mediated silencing in transgenic lines. Pita binds adjacent to dCTCF in the well-studied Mcp insulator and is required for its activity, which indicates that two insulator proteins can cooperate in organizing a functional insulator. Pita and ZIPIC preferentially bind to the promoter regions and extensively colocalize with dCTCF and BEAF. These results suggest that insulator proteins can fulfill multiple functions in chromosome architecture and gene expression.

overall_design: ChIP-seq

first: Pavel

last: Georgiev


title: High resolution specificity of DNA sequencing highlights alternative mode of lac repressor binding

city: Palo Alto

first: Zheng

last: Zuo

state: CA

address: Stanford

institute: Stanford university

summary: In this work we introduce a new high-throughput method, Spec-seq, that directly measures specificity by sequencing. It has several advantages over existing methods to quantify large collections (thousands) of binding site energies in one experiment. Using lac repressor as an example, we show that this method has excellent reproducibility giving energy measurements generally consistent within about 0.1kT. While not measuring in parallel as many different sequences as some of the higher throughput methods, we obtain high accuracies because we measure exactly what is necessary, the relative affinity to a large collection of sequences, without any mathematical fitting procedures or approximations required. It is similar to MITOMI in the number of different sites that can be analyzed in parallel, but it is much simpler to perform, requiring only a means to separate bound and unbound fractions which are then sequenced using standard high throughput, short read sequencing machines that are now readily available. When applied to the lac repressor we obtain, in a single experiment, data covering a large fraction of all the previous studies, plus thousands of additional variants, allowing us to compile a much more comprehensive profile of its specificity. We learn that the lac repressor can bind to sites of different lengths but that the preferred sequence, and the mode of binding, depends on the length. We also apply the method to two other members of the LacI/GalR protein family, PurR and YcjW, to obtain extensive models of their specificity and test the generality of the lac repressor's ability to bind to operators of variable length. We find that the lac repressor is apparently unique in its ability to bind with high affinity to sites of different lengths and with different modes of binding.

overall_design: 4 independent experiments.

first: Gary

last: Stormo


title: Genome-wide discovery of human splicing branchpoints [RNAse]

city: Sydney

first: Tim

middle: R.

last: Mercer

state: New South Wales

address: 384 Victoria Street Darlinghurst NSW 2010

country: Australia

institute: Garvan

summary: Gene splicing requires three basal genetic elements; the 3’ and 5’ splice sites and the branchpoint to which the 5’ intron termini is ligated to form a closed lariat during the splicing reaction. The 5’ and 3’ splice sites that define exon boundaries have been widely identified, revealing pervasive transcription and splicing of human genes. However, the locations of the third requisite element, the branchpoint, are still largely unknown. Here we employ two complementary approaches, targeted RNA sequencing and exoribonuclease digestion, to distil sequenced reads that traverse the lariat junction and, via non-conventional alignment, locate human branchpoint nucleotides. Alignments identify 88,748 branchpoints that correspond to 20% of known introns, with 76% supported by diagnostic sequence mismatch errors. This affords a first genome-wide analysis of branchpoints, describing their distribution, selection, and the existence of a diverse array of overlapping sequence motifs with distinct usage, evolutionary histories, and co-variation with distal splicing elements. The overlap of branchpoints with noncoding human genetic variation also indicates a notable contribution to disease. This annotation and analysis incorporates branchpoints into transcriptomic research and reflects a core role for this element in the regulatory code that governs gene splicing and expression.

overall_design: RNaseR validation of branchpoint nucleotides

middle: R

first: John

last: Mattick


title: Genome-wide discovery of human splicing branchpoints

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

overall_design: Refer to individual Series


title: Cassava responses to Pathogenic and non-pathogenic Xanthomonas axonopodis pv manihotis

city: Montpellier

first: Alvaro

middle: Luis

last: Pérez-Quintero

state: Montpellier

address: 911 Avenue Agropolis

department: RBP

country: France

institute: IRD

summary: Xanthomonas axonopodis pv. manihotis (Xam) is a gram negative bacterium causing Cassava Bacterial Blight (CBB), an important limitation for cassava production. The genetic bases underlying cassava resistance and susceptibility to different Xam strains are currently unknown. To identify genes and pathways important for the interaction, we used RNA-seq data to study transcriptomic changes in cassava plants inoculated with the non-pathogenic Xam strain, (ORST4) and a pathogenic strain, ORST4 transformed with the TAL effector TALE1Xam (ORST4+TALE1Xam). This analysis revealed that transcriptomic responses to both strains were very similar and were dominated by the induction of genes related to photosynthesis and phenylpropanoid biosynthesis and the down-regulation of genes related to jasmonic acid signaling, features possibly related to defense responses. Among the genes induced exclusively in cassava plants inoculated with ORST4 + TALE1Xam we found one gene containing a predicted binding site for TALE1Xam in its promoter region. This gene encodes for a Heat Shock Transcription Factor B3 (HsfB3) and likely acts a transcriptional repressor. HsfB3 may constitute a new type of susceptibility gene activated by a TAL effector that manages to be sufficient for symptom development without suppressing defense responses in the plant.

overall_design: mRNA of Cassava stems inoculated with a non-pathogenic (ORST4) and pathogenic (+TALE1Xam) strain of Xanthomonas axonopodis pv. Manihotis, tissues collected at 0, 5 and7 days post-inoculation, 2 technical replicates used

first: Camilo

last: Lopez

first: Alejandra

last: Muñoz-Bodnar


title: Comparisons of Phlebiopsis gigantea transcript profiles when cultured different substrates

city: Madison

first: Dan

last: Cullen

state: WI

address: One Gifford Pinchot Dr

institute: UW/FPL

summary: Collectively classified as white-rot fungi, certain basidiomycetes efficiently degrade the major structural polymers of wood cell walls. A small subset of these Agaricomycetes, exemplified by Phlebiopsis gigantea, is capable of colonizing freshly exposed conifer sapwood despite its high pitch content, which retards the establishment of other fungal species. The mechanism(s) by which P. gigantea tolerates and metabolizes resinous compounds have not been explored. Here, we report the annotated P. gigantea genome and compare profiles of its transcriptome and secretome when cultured on fresh-cut versus solvent-extracted loblolly pine. The P. gigantea genome contains a conventional repertoire of hydrolase genes involved in cellulose/hemicellulose degradation, whose patterns of expression were relatively unperturbed by the absence of extractives. The expression of genes typically ascribed to lignin degradation was also largely unaffected. In contrast, genes likely involved in the transformation and detoxification of pitch were highly induced in its presence. Their products included an ABC transporter, lipases, cytochrome P450s, glutathione S-transferase and aldehyde dehydrogenase. Other regulated genes of unknown function and several constitutively expressed genes are also likely involved in P. gigantea’s pitch metabolism. These results contribute to our fundamental understanding of conifer colonization and carbon cycling processes.

overall_design: Phlebiopsis gigantea was cultivated in media containing one of three carbon sources:  freshly harvested loblolly pine (3 replicates), acetone extracted lobollly pine (3 replicates), or glucose (2 replicates). RNA was extracted and processed for Illumina sequencing as described below.


title: Characterizing the contrasting roles of JMJD3 and UTX histone demethylases in T cell acute lymphoblastic leukemia [GSKJ4_ChIP-seq]

overall_design: Histone ChIP: Half to one million cells were treated with micrococcal nuclease (MNASE)  to generate mononucleosomal particles and an adaptation of the Upstate ChIP protocol was used.


title: Transcriptome response to salinity stress in Crassostrea hongkongensis

city: Qingdao

first: Xuelin

address: Yushan Road 5

department: Fisheries College

country: China

institute: Ocean University of China

summary: Low salinity is one of the main factors limiting the distribution and survival of marine species. As estuarine species, Crassostrea hongkongensis can live in relative low salinity. Through Illumina sequencing, we generated two transcriptomes with samples taken from gills of oysters exposed to the low salinity seawater versus the optimal seawater. By RNAseq technology, we found 13550 up-regulation genes and 9914 down-regulation genes that may regulate osmotic stress in C. hongkongensis. As blasted by GO annotation and KEGG pathway mapping, functional annotation of the genes recovered diverse biological functions and processes. The genes regulated significantly were dominated in structural molecule activity, intracellular,cytoplasm protein metabolism, biosynthesis,cell and transcription regulator activity according to GO annotation. The study aimed to compare the expression data of the two transcriptomes to provide some useful insights into signal transduction pathways in oysters and offer a number of candidate genes as potential markers of tolerance to hypoosmotic stress for oysters. In addition, the characterization of C. hongkongensis transcriptome will facilitate research into biological processes underlying physiological adaptations to hypoosmotic shock for marine invertebrates.

overall_design: Twelve oysters were exposed in low salinity (8‰) seawater and in optimal salinity (25‰) seawater,respectively. Gills from six oysters in each condition were balanced mixed respectively. The transcriptomes of two samples were generated by deep sequencing, using Illumina HiSeq2000

first: Hong

last: Yu

first: Lingfeng

last: Kong

first: Qi

last: Li


title: Genome-wide discovery of human splicing branchpoints [BPCapture]

overall_design: CaptureSeq identification of branchpoint nucleotides


title: Functional Annotation of Colon Cancer Risk SNPs

city: Los Angeles

first: Peggy

last: Farnham

address: 1450 Biggy Street NRT 6503

department: GMCB

institute: University of Southern California Graduate Student

summary: To understand the funtion of Colorectal cancer GWAS results, we perform a comprehensive analysis using biofeatures of HCT116 colon cancer cell line and got a list of risk-asscociated SNP. Risk-associated SNP are likely exerting their effects through promoters or enhancer. In order to understand the importance of the genes with risk-associated SNP in their promoters and enhancers' putatively targeted genes,  we did a comparison of these genes between HCT116 colon cancer cell and normal colon and try to understand their function

overall_design: Two biological replicates of HCT116 were compared to the data of two normal colon samples already deposited in GEO (GSM1010974 and GSM1010942).

first: Adam

last: Blattler

first: Fides

last: Lay

first: Peter

last: Jones


title: The role of small RNAs in wide hybridisation and allopolyploidisation between Brassica rapa and Brassica nigra

status: Public on Sep 30 2014

city: Hangzhou

first: Muhammad Awais

last: Ghani

state: Zhejiang

address: Room# 517, Buliding A, College of agriculture and biotechnology, Zhejiang university

department: Horticulture

institute: Zhejiang University

summary: An allopolyploid formation consists of the two processes of hybridisation and chromosome doubling. Hybridisation makes a different genome combined in the same cell, and genome “shock” and instability occur during this process, whereas chromosome doubling results in doubling and reconstructing the genome dosage. Recent studies have demonstrated that small RNAs, mainly siRNAs and miRNAs, play an important role in maintaining the genome reconstruction and stability. However, to date, little is known regarding the role of small RNAs during the process of wide hybridisation and chromosome doubling, which is essential to elucidate the mechanism of polyploidisation. Therefore, the genetic and DNA methylation alterations and changes in the siRNA and miRNA were assessed during the formation of an allodiploid (genome: AB) and its allotetraploid (genome: AABB) between Brassica rapa (♀) and Brassica nigra (♂) in the present study.The phenotypic analysis exhibited that the allotetraploid had high heterosis compared with their parents and the allodiploid. The methylation-sensitive amplification polymorphism (MSAP) analysis indicated that the proportion of changes in the methylation pattern of the allodiploid was significantly higher than that found in the allotetraploid, while the DNA methylation ratio was higher in the parents than the allodiploid and allotetraploid. The high-throughput sequencing results obtained for the small RNAs showed that the expression levels of miRNAs increased in the allodiploid and allotetraploid compared with the parents, and the expression levels of siRNAs increased and decreased compared with the parents B. rapa and B. nigra, respectively. Moreover, the percentages of miRNAs increased with an increase in the polyploidy levels, but the percentages of siRNAs and DNA methylation alterations decreased with an increase in the polyploidy levels. Furthermore, 320 known and 52 novel miRNAs were obtained from the parents in both the allodiploid and allotetraploid. However, quantitative real-time polymerase chain reaction (qRT PCR) analysis showed that the expression levels of the targets genes were negatively corrected with the expressed miRNAs.The study showed that siRNAs and DNA methylation play an important role in maintaining the genome stability in the formation of an allotetraploid. The miRNAs regulate gene expression and induce the phenotype variation, which may play an important role in the occurrence of heterosis in the allotetraploid. The findings of this study may provide new information for elucidating that the allotetraploids have a growth advantage over the parents and the allodiploids.

sample_organism:
Brassica nigra
Brassica rapa
Brassica rapa x Brassica nigra

overall_design: High throughput sequence of the parents (Brassica rapa and Brassica nigra) and their hybrids (allodiploid and allotetraploid)

first: Muhammad

first: Junxing

first: Linli

last: Rao

last: Raza

first: Liwen

last: Cao

first: Ningning

first: Xiaoxia

last: Zou

first: Liping

last: Chen


title: Astrocyte elevated gene-1 (AEG-1) and c-Myc cooperate to promote hepatocarcinogenesis

city: San Antonio

first: Yidong

state: Texas

address: 8403 Floyd Curl Drive, MSC 7784

department: Population Health Sciences

institute: UT Health Science Center at San Antonio

summary: Double transgenic mice with hepatocyte-specific expression of AEG-1 and c-Myc show aggressive HCC compared to single transgenics. Gene expression was analyzed to understand the molecular mechanism by which AEG-1 and c-Myc cooperate to promote hepatocarcinogenesis.

overall_design: Livers were collected from naïve adult mice (3 mice/group). Total RNA was extrancted and subjected to RNA-Seq.

first: Jyoti

last: Srivastava

first: Ayesha

last: Siddiq

first: Paul

middle: B

last: Fisher

first: Devanand

last: Sarkar


title: Characterizing the contrasting roles of JMJD3 and UTX histone demethylases in T cell acute lymphoblastic leukemia [UTXKO_RNA-seq]

first: Panagiotis

last: Ntziachristos

summary: T-cell acute lymphoblastic leukemia (T-ALL) is an immature hematopoietic malignancy driven mainly by oncogenic activation of NOTCH1 signaling. In this study we conduct expression analysis in NOTCH1-IC-induced tumors in Utx wild-type (Utx+/+ or Utx+/Y) and knockout (Utx-/Y) background. These results, coupled to genomic analysis of primary samples for the genomic status of the UTX gene in T-ALL, helped us to characterize the hitherto understudied role of Utx as an oncogenic facilitator in leukemia and the contrasting expression signatures between JMJD3 and UTX in this disease.

overall_design: Whole RNA was extracted from 1-5 million primary cells from Notch1-IC-expressing (sorted populations of) mouse T-ALL tumors using the RNAeasy kit (Qiagen) according to the manufacturer’s protocol. Poly-A+ (magnetic oligodT-containing beads (Invitrogen)) or Ribominus RNA was used for library preparation. cDNA preparation and strand-specific library construction was performed using the dUTP method. Libraries were sequenced on the Illumina HiSeq 2000 using 50bp single-read method. Differential gene expression analysis was performed between knockout vs wild-type background samples. Analysis was performed using DEGseq package leading to very similar conclusions.

first: Aristotelis

last: Tsirigos


title: PARALLEL IN VIVO AND IN VITRO MELANOMA RNAi DROPOUT SCREENS

city: Amsterdam

first: Oscar

last: Krijgsman

state: Outside the US or Canada

address: Plesmanlaan 121

department: Molecular Oncology and Immunology

country: Netherlands

institute: Netherlands Cancer Institute

summary: To identify factors preferentially necessary to drive tumor expansion we performed parallel in vitro and in vivo negative selection shRNA screens. Melanoma cells harboring shRNAs targeting several DNA Damage Response (DDR) kinases had a greater selective disadvantage in vivo than in vitro, indicating an essential contribution of these factors during tumor expansion. In growing tumors, DDR kinases were activated following hypoxia. Correspondingly, depletion or pharmacologic inhibition of DDR kinases was toxic to melanoma cells, including those that were resistant to BRAF inhibitor, and this could be enhanced by angiogenesis blockade. These results reveal that hypoxia sensitizes melanomas to targeted inhibition of the DDR and illustrate the utility of in vivo shRNA dropout screens for identification of pharmacologically tractable targets.

overall_design: A lentivirus-based kinome shRNA library (four pools) was used to transduce 888mel cells (MOI<0.2). After puromycin selection (1μg/ml), two reference samples were collected as controls. Next, tumor cells (5x105 per injection) were either injected s.c. into 6 NSG mice or plated into 6 independent plates (5x105) for in vitro culture. Tumors were removed from the mice and genomic DNA used to recover shRNAs by PCR amplification followed by deep sequencing. Three analyses were performed independently in parallel: (1) Tumors versus cultured cells (Log2 Fold Change < -1); (2) Tumors versus references (Log2 Fold Change < -2.5) and (3) Cultured cells versus references (Log2 Fold Change < -2.5). Genes targeted with at least 2 shRNAs in each of the analysis were considered hits with an enhanced in vivo effect.

first: Patricia

last: Possik

first: Daniel

last: Peeper


title: Large-scale identification and comparative analysis of miRNA expression profile in the respiratory tree of the sea cucumber Apostichopus japonicus during aestivation

city: QINGDAO

first: Muyan

address: 5 YUSHAN ROAD


title: Genome-wide profiling of mouse RNA secondary structures reveals key features of the mammalian transcriptome

city: Torino

first: Francesco

last: Neri

state: Italy

address: Via Nizza 52

country: Italy

institute: University of Torino

summary: By using a novel high-throughput method, named chemical inference of RNA structures (CIRS-seq), that uses dimethyl sulfate (DMS), and N-cyclohexyl-N'-(2-morpholinoethyl)carbodiimide metho-p-toluenesulfonate (CMCT) to modify RNA residues in single-stranded conformation within native RNA secondary structures, we investigated the structural features of mouse embryonic stem cell (ESC) transcripts.  Our analysis revealed an unexpected higher structuring of the 5' and 3' untranslated regions (UTRs) compared to the coding regions, a reduced structuring at the Kozak sequence and stop codon, and a three-nucleotide periodicity across the coding region of messenger RNAs. We also observed that ncRNAs exhibit a higher degree of structuring with respect to protein coding transcripts. Moreover, we found that the Lin28a binding protein binds selectively to RNA motifs with a strong preference toward a single stranded conformation.

overall_design: Transcritome-wide mapping of RNA secondary structures in mouse embryonic stem cells (mESC)

first: Danny

last: Incarnato

first: Francesca

last: Anselmi

first: Salvatore

last: Oliviero


title: Digital gene expression (DGE) of PaWB phytoplasma-infected Paulownia witches

city: Beijing

first: Jie

last: Lu

address: Shuangqiaozhonglu

institute: CAIQ

summary: Here we performed a transcriptomic study on PaWB phytoplasma-infected Paulownia sp. using Solexa/Illumina’s high-throughput digital gene expression (DGE) system. 4 DGE libraries (from 2 virus-infected samples and 2 healthy samples) were constructed, and the gene expression variations between the PaWB phytoplasma-infected (diseased) sample and the corresponding healthy sample were compared. Thousands of differentially expressed genes were obtained by the comparison, and KEGG pathway analysis of these genes suggested that many biological processes were responded to PaWB infection.

overall_design: To investigate the response of Paulownia sp. to PaWB infection, we collected four samples in two groups, namely the tissue cultured group (containing healthy sample TH and diseased sample TD) and field-grown group (containing healthy sample FH and diseased sample FD). Four individual tag libraries from these samples were constructed in parallel. For the gene expression analysis, the digital gene expression (DGE) data of diseased sample were compared to that of healthy sample in each group to obtain the gene expression variations.

first: Wenjun

last: Zhao

first: Haiqing

last: Mou


title: BMP Signaling and Its pSMAD1/5 Target Genes Differentially Regulate Hair Follicle Stem Cell Lineages

city: New York

first: Maria

last: Genander

address: 1130 York Avenue

department: Laboratory of Mammalian Cell Biology and Development

institute: Rockefeller University


title: Single-cell analysis reveals prominent classes within the planarian stem cell compartment

city: Cambridge

first: Josien

last: van Wolfswinkel

state: MA

address: 9 Cambridge Center

department: Planarian regeneration

institute: Whitehead Institute

summary: Single cell qPCR analysis identified two prominent expression signatures within the planarian stem cells, the neoblasts. RNAi against zinc finger protein zfp-1 results in elimination of one of the two classes.

overall_design: Worms were treated with RNAi by feeding. X1 or X2 cells were isolated by FACS, or blastemas were dissected. mRNA libraries were sequenced.


title: RNA-Seq in hair follicle stem cell lineage

summary: We performed RNA_Seq on purified hair follicle stem cells (HFSCs)and their direct progenty, transit amplifying cells (TACs) using temorally and spatially regulated Cre lines.

overall_design: Consequences of loss of Bmpr1a in either HFSC (K15-CrePGR;Bmpr1a fl/fl), or TACs (1. Shh-CreER:Bmpr1a fl/fl or 2. K15-CrePGR;Bmpr1a fl/fl derived)

first: Elaine

last: Fuchs


title: Polycomb Repressor Complex2 and REST-associated histone deacetylases are independent pathways toward a mature neuronal phenotype

city: Portland

first: Gail

last: Mandel

state: OR

address: 3181 SW Sam Jackson Park Rd.

department: Vollum Institute

institute: Oregon Health & Science Universtiy

summary: These experiments were designed to test the hypothesis that REST and Polycomb Repressor Complex 2 function cooperatively in undifferentiated ESCs. Our results show that H3K27me3-enriched genes show no de-repression in REST-/- ESCs, while REST target genes show significant de-repression.

overall_design: RNA-seq was performed and analyzed using two biological replicates for each genotype, WT (N6) and REST-/- (N8) mouse embryonic stem cells.

first: James

last: McGann


title: RNA-seq analysis of alternative splicing events in duplicated genes of Arabidopsis thaliana indicates considerable qualitative and quantitative divergence

city: Vancouver

first: David

middle: Christopher

last: Tack

state: B. C.

address: 3529-6720 University Boulevard

department: Botany

country: Canada

institute: Universtiy of British Columbia

summary: Divergence of alternative splicing in duplicated genes is investigated

overall_design: Three replicates of RNA-seq for Arabidopsis first true leaf tissue

first: Keith

last: Adams


title: A circadian gene expression atlas in mammals: Implications for biology and medicine

city: Philadelphia

first: Nicholas

last: Lahens

state: PA

address: Smilow Center for Translational Research 10-110 3400 Civic Center Blvd, Bldg 421

department: ITMAT

institute: University of Pennsylvania

overall_design: Refer to individual Series


title: Expression profile of trace metal element responsive genes in Brassica rapa

first: Jian

last: Wu

address: Zhongguancun Nan Da Jie 12

institute: Institute of Vegetables and Flowers, CAAS

summary:
Purpose: Zinc deficiency (ZnD) and iron deficiency (FeD), excess Zn (ZnE) and cadmium exposure (CdE) are major environmental problems for crop cultivation.
Methods: Applying Tag-Seq technology to leaves of Brassica rapa grown under FeD, ZnD, ZnE or CdE conditions, with normal conditions as a control, we examined global gene expression changes and compared the expression patterns of multiple paralogs.
Results: We identified 812, 543, 331 and 447 differentially expressed genes under FeD, ZnD, ZnE and CdE conditions, respectively, in B. rapa leaves.Further analysis revealed that genes associated with Zn, Fe and Cd responses tended to be over-retained in the B. rapa genome. Most of these multiple-copy genes showed the same direction of expression change under stress conditions.
Conclusion: We conclude that the duplicated genes involved in trace element responses in B. rapa are functionally redundant, making the regulatory network more complex in B. rapa than in Arabidopsis thaliana.

overall_design: In total, there were 15 Digital gene expression libraries, one for each of the three replicates under the four trace metal element treatments and normal nutrient supply conditions as a control.

first: Jimeng

last: Li

first: Bo

last: Liu

first: Feng

last: Cheng

first: Xiaowu

last: Wang

first: Mark

last: MAarts


title: Small RNA analysis for fetal testes of Hsp90-alpha knockout mice

city: Nagoya

first: Kenji

last: Ichiyanagi

address: Furo-cho, Chikusa-ku

department: Graduate School of Bioagricultural Sciences

country: Japan

institute: Nagoya University

summary: HSP90, found in all kingdoms of life, is a major chaperone protein regulating many client proteins. We demonstrated that HSP90α, one of two paralogs duplicated in vertebrates, plays an important role in the biogenesis of fetal PIWI-interacting RNAs (piRNA), which act against the transposon activities, in mouse male germ cells. The knockout mutation of Hsp90α resulted in a large reduction in the expression of primary and secondary piRNAs and mislocalization of MIWI2, a PIWI homolog. Whereas the mutation in Fkbp6 encoding a co-chaperone reduced piRNAs of 28-32 nucleotides in length, the Hsp90α mutation reduced piRNAs of 24-32 nucleotides, suggesting the presence of both FKBP6-dependent and -independent actions of HSP90α. Although DNA methylation and mRNA levels of L1 retrotransposon were largely unchanged in the Hsp90α mutant testes, the L1-encoded protein was increased, suggesting the presence of post-transcriptional regulation. This study revealed the specialized function of the HSP90α isofom in the piRNA biogenesis and repression of retrotransposons during the development of male germ cells in mammals.

overall_design: Total RNA was extracted from WT and Hsp90α KO testes at E16.5 (20 testes for each), and used for making a small RNA library with TruSeq Small RNA Library Preparation Kit (Illumina).  The libraries were sequenced on MiSeq (Illumina) by 50-bp single-end sequencing.

first: Tomoko

first: Ayako

last: Ogawa

first: Satomi

last: Kuramochi-Miyagawa

first: Toru

last: Nakano

first: Shinichiro

last: Chuma

first: Hiroyuki

last: Sasaki

first: Heiichiro

last: Udono


title: pSMAD1/5/8 ChIP-Seq in hair follicle stem cell lineage

first: Maria

last: Genander

state: NY

summary: We performed ChIP_Seq on purified hair follicle stem cells (HFSCs)and their direct progenty, transit amplifying cells (TACs) using and antibody against pSMAD1/5/8. We report novel common and unique in vivo pSMAD targets for each population.

overall_design: Examination of pSMAD1/5/8 targets in a stem cell lineage

first: Elaine

last: Fuchs


title: Whole transcriptomic sequencing of zebrafish rx3 mutants during optic vesicle morphogenesis

city: Dublin

first: Breandan

last: Kennedy

address: Belfield

department: UCD School of Biomolecular and Biomedical Science

country: Ireland

institute: UCD Conway Institute, University College Dublin

summary: To identify genes regulated by Rx3 during optic vesicle morphogenesis, adult zebrafish carriers of a null rx3 mutation were mated. Before 13 hours post fertilization (hpf), the earliest time point at which optic vesicle evagination phenotypes could be reliably detected, offspring were phenotypically separated into pools comprising of mutants with an absence of optic vesicles or siblings exhibiting a wild-type phenotype. Three replicates of pooled RNA samples from 13 hpf eyeless mutants (rx3-/-) or phenotypically wild-type siblings (rx3+/+ or rx3+/-), and one replicate of 13 hpf wild-type zebrafish larva were collected for whole transcriptome sequencing.

overall_design: Whole transcriptome sequencing (RNA-seq) was performed on zebrafish rx3-/- mutants, wild-type siblings and wild-type AB strains at 13 hpf

first: Jun

last: Yin

last: Morrissey

first: Lisa

last: Shine

first: Breandán

middle: N


title: A chromatin code for alternative splicing involving an association between CTCF and HP1α proteins

city: Stockholm

first: Eneritz

last: Agirre

address: Solnavägen 9

department: MBB

country: Sweden

institute: Karolinska Institutet

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


title: Neurons gradually attain a post-mitotic state

status: Public on Sep 29 2014

first: Li-Huei

last: Tsai

state: Massachusetts

department: Picower Institute for Learning and Memory

country: USA

institute: MIT

summary: Once generated, neurons are thought to permanently exit the cell cycle and become irreversibly differentiated. However, neither the precise point at which this post-mitotic state is attained nor the extent of its irreversibility is clearly defined. Here we report that newly born neurons from the upper layers of the mouse cortex, despite initiating axon and dendrite elongation, continue to drive gene expression from the neurogenic intermediate neural progenitor tubulin ?1 promoter (T?1p). These observations suggest an ambiguous post-mitotic neuronal state. Whole transcriptome analysis of sorted upper cortical neurons further revealed that neurons continue to express genes related to cell cycle progression long after mitotic exit until at least post-natal day 3 (P3). These genes are however down regulated thereafter and are associated with a concomitant up regulation of tumor suppressors at P5. Interestingly, newly born neurons located in the cortical plate (CP) at embryonic day 18-19 (E18-E19) and P3 challenged with calcium influx are found in S/G2/M phases of the cell cycle, and still able to undergo division at E18-E19 but not at P3. At P5 however, calcium influx becomes neurotoxic and leads instead to neuronal loss. Our data delineate for the first time the temporal window during which developing neurons acquire a post-mitotic state.

overall_design: 3 biological replicates per group were analyzed. The material analyzed was upper layer cortical neurons labeled with GFP and isolated by FACS sorting. Total RNA was extracted from those cells.

first: Froylan

last: Calderon de Anda

first: Jia

last: Meng


title: A circadian gene expression atlas in mammals assayed by RNA-seq

first: Nicholas

last: Lahens

summary: To characterize the role of the circadian clock in mouse physiology and behavior, we used RNA-seq and DNA arrays to quantify the transcriptomes of 12 mouse organs over time. We found 43% of all protein coding genes showed circadian rhythms in transcription somewhere in the body, largely in an organ-specific manner. In most organs, we noticed the expression of many oscillating genes peaked during transcriptional “rush hours” preceding dawn and dusk. Looking at the genomic landscape of rhythmic genes, we saw that they clustered together, were longer, and had more spliceforms than nonoscillating genes. Systems-level analysis revealed intricate rhythmic orchestration of gene pathways throughout the body. We also found oscillations in the expression of more than 1,000 known and novel noncoding RNAs (ncRNAs). Supporting their potential role in mediating clock function, ncRNAs conserved between mouse and human showed rhythmic expression in similar proportions as protein coding genes. Importantly, we also found that the majority of best-selling drugs and World Health Organization essential medicines directly target the products of rhythmic genes. Many of these drugs have short half-lives and may benefit from timed dosage. In sum, this study highlights critical, systemic, and surprising roles of the mammalian circadian clock and provides a blueprint for advancement in chronotherapy.

overall_design: 96 samples total covering 12 different tissues, with no replicates. Each tissue sampled every 6 hours for 2 days (8 samples per tissue).

first: Ray

last: Zhang

first: Heather

middle: I

last: Ballance

first: Michael

last: Hughes

first: John

last: Hogenesch


title: Identification of building principles of methylation states at CG rich regions by high-throughput editing of a mammalian genome

city: Basel

first: Dirk

last: Schuebeler

address: Fabrikstrasse 24

country: Switzerland

institute: Friedrich Miescher Institute for Biomedical Research

summary: Methylation is a repressive modification of DNA prevalent throughout mammalian genomes yet mostly absent at CG rich stretches referred to as CGI. Here we identify their building principles by parallel genomic targeting of sequence libraries.  Iterative insertions generated over 3,000 variants of genome-derived and artificial sequences at the same genomic site.  Single molecule profiling of the methylation status of this collection allowed modeling the contribution of CG content and DNA binding factors towards the unmethylated state. It made the surprising prediction that the majority of CGs within endogenous islands are susceptible to methylation changes modulated by the presence of transcription factors, which is indeed confirmed by genome-wide methylation dynamics during multiple cellular differentiations.   Our model further predicts blocks of constitutively unmethylated CGs independent from TF binding, which have a median size of ~300bp but are only present in half of all islands. Their constitutively unmethylated state is a hallmark of untransformed cells but their increased methylation is a specific and predictive feature of cancer.   This study quantifies the two principal mechanisms governing methylation patterns in mammalian genomes. It provides a framework to interpret methylation data across normal and cancer samples and refines the concept of CpG islands. Methylation is a repressive modification of DNA prevalent throughout mammalian genomes yet mostly absent at CG rich stretches referred to as CGI. Here we identify their building principles by parallel genomic targeting of sequence libraries.  Iterative insertions generated over 3,000 variants of genome-derived and artificial sequences at the same genomic site.  Single molecule profiling of the methylation status of this collection allowed modeling the contribution of CG content and DNA binding factors towards the unmethylated state. It made the surprising prediction that the majority of CGs within endogenous islands are susceptible to methylation changes modulated by the presence of transcription factors, which is indeed confirmed by genome-wide methylation dynamics during multiple cellular differentiations.   Our model further predicts blocks of constitutively unmethylated CGs independent from TF binding, which have a median size of ~300bp but are only present in half of all islands. Their constitutively unmethylated state is a hallmark of untransformed cells but their increased methylation is a specific and predictive feature of cancer.   This study quantifies the two principal mechanisms governing methylation patterns in mammalian genomes. It provides a framework to interpret methylation data across normal and cancer samples and refines the concept of CpG islands.

sample_organism:
Escherichia coli
Mus musculus
synthetic construct

overall_design: Libraries of DNA sequences were constructed either by mouse genome (129S6) or E.coli genome (NC_010473.1) subrepresentation or custom synthesis. DNA fragments were inserted into the genome of mouse embryonic stem cells by recombination mediated casette exchange (RMCE) at the B-globin locus. Methylation status of the inserted DNA sequences was profiled by bisulfite sequencing using a pair of universal primers flanking the fragments.

first: Arnaud

last: Krebs

last: Schübeler


title: Large-scale identification and comparative analysis of miRNA expression profile in the respiratory tree of the sea cucumber Apostichopus japonicus during aestivation (part 2)

first: Muyan

last: Chen

institute: Ocean University of China

summary: The sea cucumber Apostichopus japonicus withstands high water temperatures in the summer by suppressing metabolic rate and entering a state of aestivation. We hypothesized that changes in the expression of miRNAs could provide important post-transcriptional regulation of gene expression during hypometabolism via control over mRNA translation. The present study analyzed profiles of miRNA expression in the sea cucumber respiratory tree using Solexa deep sequencing technology. We identified 279 sea cucumber miRNAs, including 15 novel miRNAs specific to sea cucumber. Animals sampled during deep aestivation (DA; after at least 15 days of continuous torpor) were compared with animals from a non-aestivation (NA) state (animals that had passed through aestivation and returned to an active state). We identified 30 differentially expressed miRNAs ([RPM (reads per million) >10, |FC| (|fold change|) ≥1, FDR (false discovery rate) <0.01]) during aestivation, which were validated by two other miRNA profiling methods: miRNA microarray and real-time PCR. Among the most prominent miRNA species, miR-124, miR-124-3p, miR-79, miR-9 and miR-2010 were significantly over-expressed during deep aestivation compared with non-aestivation animals, suggesting that these miRNAs may play important roles in metabolic rate suppression during aestivation.

overall_design: analyze the global profile of small RNAs in non-aestivation (NA) and deep aestivation (DA) sea cucumbers' respiratory tree using Solexa sequencing technology

first: Kenneth

last: Storey


title: Rapid neurogenesis through transcriptional activation in human stem cell (RNA-Seq)

city: La Jolla

first: Nathan

last: Lewis

department: Pediatrics

institute: University of California, San Diego

summary: Advances in cellular reprogramming and stem cell differentiation now enable ex vivo studies of human neuronal differentiation. However, it remains challenging to elucidate the underlying regulatory programs because differentiation protocols are laborious and often result in low neuron yields. Here, we overexpressed two murine Neurogenin transcription factors in human induced pluripotent stem cells, and obtained neurons with bipolar morphology in four days at greater than 90% purity. The high purity enabled mRNA and microRNA expression profiling during neurogenesis, thus revealing the genetic programs involved in the transition from stem cell to neuron. These profiles were then analyzed to identify the regulatory networks underlying the differentiation of the neurons.

overall_design: Paired end RNA sequencing of iPS cells (PGP1) at 0, 1, 3, and 4 days post- doxycycline induction of murine NGN1 and NGN2. This was done using an Illumina HiSeq, and reads were aligned to hg19

first: Volker

last: Buskamp


title: Polycomb Repressor Complex 2 and REST-associated histone deacetylases are independent pathways toward a mature neuronal phenotype.

first: Gail

last: Mandel

summary: These experiments were designed to test the hypothesis that REST and Polycomb Repressor Complex 2 function cooperatively in undifferentiated ESCs. Our results showed that a majority of REST-bound genomic regions were not associated with H3K27me3 enrichment and loss of H3K27me3 enrichment was not a general observation in REST -/- ESCs. These results support the conclusion that REST and PRC2 function independently in ESCs and similarly contribute to maintaing a transcriptionally poised state through antagonism of H3K4me3.

overall_design: Examination of REST-bound regions in undifferentiated mouse embryonic stem cells (ESC), and a comparison of H3K27me3 distribution between WT and REST-/- ESCs.

first: Jon

last: Oyer

first: James

last: McGann

first: Xin

last: Feng


title: Human stem cell and mouse transchromosomic (hsa11) embryonic stem cell (with and without ectopic ZNF91 expression) ChIP-seq

city: Santa Cruz

first: Sofie

last: Salama

address: MS: CBSE, 1156 High St

department: Center for Biomolecular Science and Engineering

institute: Univerity of California, Santa Cruz

summary: We compared the binding patterns in embryonic stem cells of KAP1 and KRAB Zinc Finger (KZNF) proteins as well as H3K4me3 DNA under several conditions. Native human stem cells. Mouse stem cells containing a transchromosomic copy of human chromosome 11, with and without the introduction of plasmids containing KRAB Zinc Finger sequences. We show that certain KZNFs are responsible for the repression of certain retrotransposons in embryonic stem cells, preventing their spread across the genome.

overall_design: ChIP-seq of HESCs and mouse TC11 ESCs with ZNF91 plasmids and with empty vector plasmids. ChIP of KAP1, ZNF486, H3K4me3. At least 2 replicates of each condition.

first: Frank

middle: M

last: Jacobs

first: David

last: Greenberg

first: Ngan

last: Nguyen

first: Maximilian

last: Haeussler

first: Adam

last: Ewing

first: Sol

last: Katzman

first: Benedict

last: Paten

last: Haussler


title: An evolutionary arms race between KRAB zinc finger genes 91/93 and SVA/L1 retrotransposons

sample_organism:
Macaca mulatta
Homo sapiens
Mus musculus


title: Human embryonic stem cell, rhesus embryonic stem cell and mouse transchromosomic (hsa11) embryonic stem cell (with and without ectopic ZNF91 expression) RNA-seq

summary: We compared the expression in embryonic stem cells of KRAB ZNF proteins and retrotransposon elements in the human genome under several conditions. Native human stem cells. Mouse stem cells containing a transchromosomic copy of human chromosome 11, with and without the introduction of plasmids containing KRAB Zinc Finger sequences. We show that certain KZNFs are responsible for the repression of certain retrotransposons in embryonic stem cells, preventing their spread across the genome.

platform_organism:
Macaca mulatta
Homo sapiens
Mus musculus

overall_design: RNA-seq of hESCs, rhESCs and mouse TC11 ESCs with ZNF91 plasmids (2 replicates), with empty vector plasmids (2 replicates), or no plasmids.


title: CK2 regulates transcriptional elongation

first: Yuliang

last: Tan

state: California

address: Room 345, CMM West, 9500 Gilman Drive, Mail Code 0648

department: School of Medicine

summary: CK2 binds to actively transcribed regions in the genome and regulates transcriptional elongation.

overall_design: Identification of genome-wide binding sites of CK2α, and examination of the effect of Pol II binding upon CK2 inhibition in LNCaP cells

first: Michael

last: Rosenfeld

first: Harihar

last: Basnet


title: NF-kB coordinates rapid, BRD4-dependent remodeling of proinflammatory super-enhancers [Chem-Seq]

city: Boston

last: Bradner

address: 450 Brookline

department: Medical Oncology

institute: Dana-Farber Cancer Institute

summary: Proinflammatory stimuli rapidly and globally remodel chromatin landscape, thereby enabling transcriptional responses. Yet, the mechanisms coupling chromatin regulators to the master regulatory inflammatory transcription factor NF-kB remain poorly understood.  We report in human endothelial cells (ECs) that activated NF-kB binds to enhancers, provoking a rapid, global redistribution of BRD4 preferentially at super-enhancers, large enhancer domains highly bound by chromatin regulators.  Newly established NF-kB super-enhancers drive nearby canonical inflammatory response genes. In both ECs and macrophages BET bromodomain inhibition prevents super-enhancer formation downstream of NF-kB activation, abrogating proinflammatory transcription. In TNFa-activated endothelium this culminates in functional suppression of leukocyte rolling, adhesion and transmigration.  Sustained BET bromodomain inhibitor treatment of LDLr -/- animals suppresses atherogenesis, a disease process rooted in pathological vascular inflammation involving endothelium and macrophages. These data establish BET-bromodomains as key effectors of inflammatory response through their role in the dynamic, global reorganization of super-enhancers during NF-kB activation.

overall_design: Chem-Seq for the biotinylated small molecule JQ1 in untreated or TNFalpha treated human endothelial cells

first: Jonathan

last: Brown

first: Charles

middle: Y

last: Lin


title: mRNAs that co-purify with OMA-1 in the C. elegans germline (sequencing)

city: Minneapolis

first: Caroline

last: Spike

state: MN

address: 420 Washington Avenue, SE

department: Genetics, Cell Biology, and Development

institute: University of Minnesota

summary: The oocytes of most animals arrest at diplotene or diakinesis, but resume meiosis (meiotic maturation) in response to hormones. In C. elegans, maturation of the –1 oocyte requires the presence of sperm, Gas-adenylate cyclase-PKA signaling in the gonadal sheath cells, and germline function of two Tis11-like CCCH zinc-finger proteins, OMA-1 and OMA-2 (OMA proteins).  Prior studies indicate that the OMA proteins redundantly repress the translation of specific mRNAs in oocytes (zif-1, mom-2, nos-2, glp-1) and early embryos (mei-1).  We purified OMA-1-containing ribonucleoprotein particles (RNPs) and identified mRNAs that associate with OMA-1 in oocytes using microarrays. We examined the relative abundances of mRNAs in OMA-1 RNPs using high-throughput RNA sequencing.  Previously identified targets of OMA-dependent translational repression in oocytes were found to be both enriched (>2-fold relative to input RNA) and abundant in purified OMA-1 RNPs.  Furthermore, we verified that some of the newly identified mRNAs that share these characteristics are translationally repressed by OMA-1/2 in oocytes through sequences in their 3’UTRs.  Although meiotic maturation is stimulated by sperm, we found that the mRNAs copurifying with OMA-1 are not significantly different in the presence and absence of sperm, suggesting that sperm-dependent signaling does not modify the suite of mRNAs stably associated with OMA-1.  Further, several tested OMA-1-associated mRNAs were shown to be translationally repressed in both the presence and absence of sperm.

overall_design: C. elegans mRNAs that co-purify with OMA-1 were identified by deep-sequencing using the Illumina HiSeq 2000

middle: A

last: Greenstein


title: Argonaute-1 binds transcriptional enhancers and controls constitutive and alternative splicing in human cells

first: Eneritz

last: Agirre

summary: The roles of Argonaute proteins in cytoplasmic miRNA and RNAi pathways are well established. However, their implication in small RNA-mediated transcriptional gene silencing in the mammalian cell nucleus is less understood. We have recently shown that intronic siRNAs cause chromatin modifications that inhibit RNA polymerase II elongation and modulate alternative splicing in an Argonaute-1 (AGO1) dependent manner. Here we used chromatin immunoprecipitation followed by deep sequencing (ChIP-seq) to investigate the genome-wide distribution of AGO1 nuclear targets. Unexpectedly we found that about 80% of AGO1 clusters are associated with cell-type specific transcriptional enhancers, most of them (73%) overlapping active enhancers. This association seems to be mediated by long, rather than short, enhancer RNAs; and to be more prominent in intragenic, rather than intergenic, enhancers. Paradoxically, crossing ChIP-seq with RNA-seq data upon AGO1 depletion revealed that enhancer bound AGO1 is not linked to the global regulation of gene transcription but to the control of constitutive and alternative splicing, which was confirmed by an individual gene analysis explaining how AGO1 controls inclusion levels of the cassette exon 107 in the SYNE2 gene.

overall_design: Genome-wide analysis of AGO-1 and different histone modifications in 2 cell types

first: Mariano

last: Alló

first: Sergey

last: Bessonov

first: Paola

last: Bertucci

first: Luciana

last: Gómez Acuña

first: Valeria

last: Buggiamo

first: Nicolás

last: Bellora

first: Babita

last: Singh

first: Ezequiel

last: Petrillo

first: Matías

last: Blaustein

first: Belén

last: Miñana Gómez

first: Gwendal

last: Dujardin

first: Berta

last: Pozzi

first: Federico

last: Pelisch

first: Elias

last: Bechara

first: Dmitry

last: Agafonov

first: Anabella

last: Srebrow

first: Reinhard

last: Lührmann

first: Juan

last: Valcárcel

first: Eduardo

last: Eyras

first: Alberto

last: Kornblihtt


title: mRNAs that co-purify with OMA-1 in the C. elegans germline

first: Caroline

last: Spike

overall_design: Refer to individual Series


title: CREBBP (CBP) ChIP-seq in wild type and CBP null MEFs +/- dipyridyl

city: Memphis

first: Suzanne

middle: J

last: Baker

state: TN

address: 262 Danny Thomas Place

department: Developmental Neurobiology

institute: St Jude Children's Research Hospital

summary: Genome-wide distribution of histone H3K18 and H3K27 acetyltransferases, Crebbp (CBP) and Ep300 (p300), is used to map enhancers and promoters, but whether these elements functionally require CBP/p300 remains largely uncertain.  We investigated this relationship by comparing genomic CBP recruitment with gene expression in wild type and CBP/p300 double-knockout fibroblasts.  ChIP-seq revealed nearby CBP recruitment for 20 percent of constitutively expressed genes, but surprisingly, three-quarters of these were unaffected or slightly activated by CBP/p300 deletion. Computationally defined enhancer-promoter-units (EPUs) having a CBP peak within two kilobases of the enhancer-like element provided better predictive value, with CBP/p300 deletion attenuating expression of 40 percent of such EPU assigned constitutively expressed genes. We next examined signaling-responsive (Hypoxia Inducible Factor) gene expression and CBP recruitment, and found that 97 percent of inducible genes were within 50 kilobases of an inducible CBP peak, and 70 percent of these required CBP/p300 for full inducible expression. Unexpectedly however, most inducible CBP peaks occurred near signal-nonresponsive genes.

overall_design: eight samples total; Two wild type and two CBP null primary mouse embryonic fibroblast (MEF) lines, each treated with 100uM 2,2-dipyridyl or ethanol vehicle for 2 hours

first: Paul

last: Brindle

first: Lawryn

last: Kasper


title: RNA-seq of laser-capture microdissected 50x50x50 um tissue cubes from the medial ganglionic eminence of wild-type and GFRa1 mutant mice

first: Sten

last: Linnarsson

address: Scheeles väg 1

department: Medical Biochemistry and Biophysics

summary: Cortical interneurons originating from the medial ganglionic eminence (MGE) are among the most diverse cells within the CNS. Different pools of proliferating progenitor cells are thought to exist in the ventricular zone of the MGE, but whether the underlying subventricular and mantle regions of the MGE are spatially patterned has not yet been addressed. Here, we combined laser-capture microdissection and multiplex RNA-sequencing to map the transcriptome of MGE cells at a spatial resolution of 50 µm. Distinct groups of progenitor cells showing different stages of interneuron maturation were identified and topographically mapped based on their genome-wide transcriptional pattern.

overall_design: One 50  µm coronal section from the MGE was taken from each of two wildtype and one GFRa1 mutant E12.5 C57bl6/J mouse. Each section was laser microdissected into approximately 100 cubes, covering the whole MGE, and each cube was further processed for RNA-seq analysis.

first: Sabrina

last: Zechel

first: Pawel

last: Zajac

first: Peter

last: Lönnerberg

first: Carlos

last: Ibáñez


title: NF-kB coordinates rapid, BRD4-dependent remodeling of proinflammatory super-enhancers

first: James

last: Bradner


title: Genome-wide and single-cell analyses reveal target gene context-dependent relationship between Crebbp recruitment and both constitutive and signal-responsive gene expression.


title: Small RNA sequencing of plasma RNA

city: Belfast

first: David

middle: Arthur

last: Simpson

state: N. Ireland

address: QUB Institue of clinical Science, Block A, RVH

department: Centre for Experimental Medicine

country: United Kingdom

institute: Queen's University Belfast

summary: We report the sequencing of small RNAs present in the plasma of three normal subjects. In addition to microRNAs we identified abundant non-human small RNA sequences. The organisms from which these were derived were identified by BLAST searches with contigs assembled from the short sequences. The taxonomic profiles were very consistent between individuals, including plants and microbes reported previously in the microbiome, but in proportions distinct from other sites. The majority of bacterial reads were from the phylum Proteobacteria, whilst for 5 of 6 individuals over 90% of the more abundant fungal reads were from the phylum Ascomycota; of these over 90% were from the order Hypocreales. Most reads mapped to rRNA sequences and the presence of specific common sequences was confirmed by RT-PCR. In addition, extremely abundant small RNAs derived from human Y RNAs were detected. We conclude that a characteristic profile of a subset of the human microbiome can be obtained by sequencing small RNAs present in the blood. The origin and potential function of these molecules remains to be determined, but the specific profile is likely to reflect health status. This facile test has immense potential to provide biomarkers for the diagnosis and prognosis of human disease.

overall_design: The profile of small RNAs present in the plasma of three normal subjects was determined


title: NF-kB coordinates rapid, BRD4-dependent remodeling of proinflammatory super-enhancers [ChIP-Seq]

overall_design: ChIP-Seq for various transcription factors, RNA Polymerase II, and histone modifications in human endothelial cells

first: Jonathan

middle: D

last: Brown

first: Charles

last: Lin

middle: E


title: Characterisation of p-hydroxycinnamate catabolism in a soil Actinobacterum

status: Public on Sep 28 2014

city: Berkeley

first: Hiroshi

last: Otani

state: CA

address: 1 Cyclotron Road

department: DOE Joint Genome Institute

institute: Lawerence Berkeley National Laboratory

summary: p-Hydroxycinnamates, such as p-coumarate and ferulate, are components of plant cell walls and have a number of commercial applications. Previously, we had shown that the soil Actinobacterium Rhodococcus jostii RHA1 (RHA1) grows on ferulate, catabolizing it via vanillate and the β-ketoadipate pathway. We used transcriptomics to identify genes in RHA1 that were specifically up-regulated during growth on ferulate. These include three operons predicted to encode the uptake and β-oxidative deacetylation of ferulate and p-coumarate: couHLT, couMNO and couR. A couL mutant did not grow on p-coumarate, ferulate or their dihydro derivatives, but grew on vanillate. Purified CouL catalyzed the thioesterification of several p-hydroxycinnamates. Among the tested substrates, the best were p-coumarate and caffeate (kcat/KM ~400 mM-1s-1), and sinapate was not transformed. Of these, p-coumarate was also RHA1’s preferred growth substrate. Although the data indicate that p-hydroxycinnamates are catabolized via β-oxidation, the pathway lacks a typical β-ketothiolase. The data further suggest the involvement of two formaldehyde detoxification pathways in vanillate catabolism. This study augments our understanding of the bacterial catabolism of biomass and facilitates the production of aromatics from renewable feedstocks.

overall_design: Transcriptomes of R. jostii RHA1 from ferulate and benzoate cultures were analysed using Ion PGMTM system.

first: Lindsay

last: Eltis


title: Metabolic Respiration Induces AMPK- and Ire1p-Dependent Activation of the p38-Type HOG MAPK Pathway

status: Public on Sep 26 2014

city: Buffalo

last: Cullen

address: 341 Cooke Hall

department: Biology

institute: SUNY-Buffalo

summary: In fungal species, differentiation to the filamentous/hyphal cell type is critical for entry into host cells and virulence. Comparative RNA sequencing was used to explore the pathways that regulate differentiation to the filamentous cell type in yeast. This approach uncovered a role for the stress-response MAPK pathway, HOG, during the increased metabolic respiration that induces filamentous growth. In this context, the AMPK Snf1p and ER stress kinase Ire1p regulated the HOG pathway. Cross-modulation between the HOG and filamentous growth (ERK-type) MAPK pathways optimized the differentiation response. The regulatory circuit described here may extend to behaviors in metazoans.

overall_design: Comparison of expression patterns of wild-type and mutant yeast cells grown in salt, tunicamycin or galactose by comparative RNA sequencing analysis.

first: Hema

last: Adhikari


title: PPARα mouse liver cistrome

status: Public on Sep 27 2014

city: Houston

last: Moore

address: One Baylor Plaza

institute: Baylor College of Medicine

summary: Autophagy is an evolutionally conserved catabolic process that recycles nutrients upon starvation and maintains cellular energy homeostasis1-3. Its acute regulation by nutrient sensing signaling pathways is well described, but its longer-term transcriptional regulation is not. The nuclear receptors PPARα and FXR are activated in the fasted or fed liver, respectively4,5. Here we show that both regulate hepatic autophagy. Pharmacologic activation of PPARα reverses the normal suppression of autophagy in the fed state, inducing autophagic lipid degradation, or lipophagy. This response is lost in PPARα knockout (PPARα-/-) mice, which are partially defective in the induction of autophagy by fasting. Pharmacologic activation of the bile acid receptor FXR strongly suppresses the induction of autophagy in the fasting state, and this response is absent in FXR knockout (FXR-/-) mice, which show a partial defect in suppression of hepatic autophagy in the fed state. PPARα and FXR compete for binding to shared sites in autophagic gene promoters, with opposite transcriptional outputs. These results reveal complementary, interlocking mechanisms for regulation of autophagy by nutrient status.

overall_design: Mouse liver PPARα cistromes in fed 8-week-old male WT or PPARα KO mice treated with or without its synthetic agonist ligand GW7647twice a day were generated by deep sequencing in quadruplicate using illumina

first: Mitchell

last: Lazar


title: microRNA sequencing of HEK293 and H9c2 cells

city: Indianapolis

first: Scot

last: Matkovich

state: IN

address: Lilly Corporate Center

institute: Eli Lilly and Company

summary: The human HEK293 / 293T and rat cardiomyoblast H9c2 cell lines are commonly employed for microRNA-mRNA interaction studies. Here, I provide microRNA sequencing data obtained from each of these lines to better document which microRNAs are endogenously expressed at high or low levels.

sample_organism:
Homo sapiens
Rattus norvegicus

platform_organism:
Homo sapiens
Rattus norvegicus

overall_design: Small RNA sequencing profiles were generated from cultured HEK293 and H9c2 cells on Illumina HiSeq 2000 instruments.


title: The miR-155/PU.1 axis acts on Pax5 to enable efficient terminal B cell differentiation [RNA-Seq]

city: Cambridge

first: Simon

middle: Richard

last: Andrews

state: Cambs

address: Babraham Research Campus

department: Science Services

institute: The Babraham Institute

summary: miR-155 inhibits PU.1 expression, leading to the initiation of the initiation of plasma cell differentiation pathway. Additional PU.1 targets include a network of genes whose products are involved in adhesion, with direct links to B:T interactions.

overall_design: Sequencing analysis of B cell transcriptome using Illumina TruSeq mRNA sample prep kit and Illumina platform. RNA was isolated from ex-vivo LPS and IL-4 activated (96h) splenic B cells from wild type, PU.1[155-/155-] and miR-155[-/-] mice. 4-5 biological replicates per genotype and condition.

first: Dong

last: Lu

first: Rinako

last: Nakagawa

first: Sandra

last: Lazzaro

first: Philipp

last: Staudacher

first: Cei

last: Abreu-Goodger

first: Tom

last: Henley

first: Sara

last: Boiani

first: Rebecca

last: Leyland

first: Alison

last: Gallowway

first: Geoffrey

last: Butcher

first: Stephen

middle: L

last: Nutt

first: Martin

last: Turner

first: Elena

last: Vigorito


title: H3K27me3 ChIP-seq profiling in MCF7 cell lines under hypoxia

city: Amsterdam

first: Michiel

middle: Emanuel

last: Adriaens

address: Meibergdreef 15

department: Experimental Cardiology

institute: AMC

summary:
Purpose: Study hypoxia induced changes in genome-wide H3K27me3 occupancy
Methods: Using the MCF7 breast epithelial adenocarcinoma cell line as a model, we studied epigenomic reprogramming as a function of fluctuating oxygen tension. To this end, we combined chromatin-immunoprecipitation and deep-sequencing analysis to identify H3K27me3-marks in MCF7 cells subjected to changes in oxygenation (i.e. acute hypoxia, chronic hypoxia).
Results: H3K27me3-marks showed a rapid global increase at specific sites throughout the genome under hypoxia, both genic and inter-genic.
Conclusions: Our data show that oxygen availability dynamically regulates the epigenetic state of the genome.

overall_design: Genome-wide H3K27me3-mark profiles were generated by combining ChIP analysis with deep sequencing using Illumina GAIIx.

first: Peggy

last: Prickaerts

first: Willem

last: Voncken

first: Brad

last: Wouters

first: Twan

last: Van Den Beucken


title: Myc mild overexpression in the heart in a mosaic fashion induces matabolic changes and triggers hipertrophic protection pathways without a pathological phenotype

city: Madrid

first: Fatima

last: Sanchez-Cabo

address: Melchor Fernandez Almagro

country: Spain

institute: CNIC

summary: Recently, it was described that mammalian cells are able to eliminate those with relative lower Myc levels in the epiblast through cell competition. We have described that cardiomyocytes during heart development are also able to complete eliminating cells with lower Myc levels. We have also shown that adult cardiomyocytes respond in the same way over long periods of time when cell competition is induced by overexpressing Myc in a mosaic fashion. We therefore have developed an RNASeq assay to further understand the mechanism of elimination of WT cells and the effect of mild Myc overexpression in cardiomyocytes.

overall_design: Myc overexpression in a mosaic fashion in adult cardiomyocytes, 2 hearts were analyzed and two wild type littermates were used as controls

first: Miguel

last: Torres

first: Cristina

last: Villa del Campo


title: Control of embryonic stem cell identity by BRD4-dependent transcriptional elongation of super-enhancer associated pluripotency genes

city: New york

first: RAFFAELLA

last: Di MICCO

state: New York

address: 550 1st avenue,  Smilow 305

department: Pathology

institute: NYU School of Medicine

summary:
BET-regulated transcriptome and BRD4, BRD2, BRD3 and Pol II ChIP-seq datasets in human ESCs before and after BET inhibition.
Transcription factors and chromatin remodeling complexes are key determinants of embryonic stem cell (ESC) identity. In this study, we investigate the role of BRD4, a member of the bromodomain and extra-terminal domain (BET) family of epigenetic reader proteins, in control of ESC identity. We performed RNA-seq analyiss in the presense of small molecule inhibitors of BET proteins to show that BRD4 positively regulates the ESC transcriptome. We also integrated RNA-seq analysis with ChIP-sequencing datasets s for BRD4 (and for other BRD2 and BRD3) to demonstrate that BRD4 binds SEs and regulates the expression of SE-associated pluripotency genes. We have also conducted ChIP-seq analysis for Pol II binding to demonstrate that SE-associated genes depend on BRD4-dependent Pol II binding at TSS and gene body for their productive transcriptional elongation.

overall_design: Total RNA was extracted from samples using the RNeasy Qiagen kit according to the manufacturer’s instructions. Deep sequencing of RNA (1ug) from hESCs FGF- or MS436-treated at day 1 and day 5 was performed as described in (Higgin et al., 2010c). Samples were subjected to PolyA  selection using magnetic oligo-dT beads. The resulting RNA samples were then used as input for library construction  as described by the manufacturer (Illumina, CA, USA). RNA libraries were then sequenced on the GAIIx system using 50bp single reads. Chromatin for ChIP-sequencing was obtained from FGF-maintained hESCs, vehicle or MS417-treated (at 250nM concentration for 6h) (10 to 20x106 cells/IP). ChIP-Seq libraries were generated using standard Illumina kit and protocol as described in (Ntziachristos et al., 2012). We performed cluster amplification and single read 50 sequencing-method using the Illumina HiSeq 2000, following manufacturer’s protocols.

first: Raffaella

last: Di Micco