title: Reduced representation bisulfite sequencing from G0 & G4/G5 CMP

city: Houston

first: Ronald

last: Depinho

address: 1400 Pressler St

institute: University of Texas MD Anderson Cancer Center

summary:
Differential hyper- and hypo-methylation regions in G0 versus G4/G5 CMP
The goal of this study is to evaluate changes in CpG methylation profilings of telomere dysfunctional common myeloid progenitor cells (CMP) as compared to their wild type controls

overall_design: Genomic DNA was extracted from sorted CMP populations isolated from 3 pools of G0 or 2 pools of G5 mice using UltraPure Phenol:Chloroform:Isoamyl Alcohol according to manufacturer’s instructions (Life Technologies). 14,000 to 30,000 cells were available for each sample, resulting in a minimum of 45ng of DNA. Genome-wide DNA methylation profiling was performed by RRBS. Library preparation and sequencing were performed at the UT MD Anderson Cancer Center’s DNA Methylation Analysis Core and Sequencing and Microarray Facility, according to published protocols. RRBS sequencing data were aligned and methylation was called using Bismark v0.7.119. In brief, bisulphite-treated DNA was aligned to UCSC Genome Browser mm10 reference genome using Bowtie. In total 29-38 million reads were generated per sample with alignment rates around 63%. Next, MethylKit10 implemented with Fisher’s exact test was used to compare the cytosine methylation profiles of G0 and G5 CMP. Gene promoter regions were calculated based on RefSeq gene annotations with regions starting 1 kb upstream of the annotated transcription start site (TSS) and extending 500 base pairs downstream of TSS. Exons, introns, and CpG islands coordinates were collected from the UCSC Genome Browser mm10 version.

first: Simona

last: Colla

first: Derrick

last: Ong

first: Ting

last: Gong


title: Telomere dysfunction drives aberrant hematopoietic differentiation and Myelodysplastic Syndrome.

status: Public on Apr 09 2015

country: USA

type: Methylation profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: A comprehensive catalog of the C. elegans dsRNAome

city: Salt Lake City

first: Joseph

middle: M.

last: Whipple

state: UTAH

address: 15 N Medical Drive East, Rm 4800

department: Department of Biochemistry

institute: University of Utah

type: Expression profiling by high throughput sequencing

summary: Recent studies hint that endogenous dsRNA plays an unexpected role in cellular signaling. However, a complete understanding of the mechanisms underlying endogenous dsRNA signaling is hindered by an incomplete annotation of dsRNA-producing genes. In order to identify all dsRNAs expressed in C. elegans, we developed a bioinformatics pipeline that identifies dsRNA by detecting clustered RNA editing sites, which are strictly limited to long dsRNA substrates of Adenosine Deaminases that act on RNA (ADAR). Using a combination of alignment algorithms to map both unique and repetitive reads, we detected as many as 664 editing-enriched regions (EERs). Most EERs were associated with protein-coding genes, with ~1.7% of all C. elegans mRNAs containing an EER, located primarily in introns and 3’UTRs. Introns having an embedded EER were up to 9-fold longer than the average C. elegans intron. A significant fraction of EERs were located less than 1000 base pairs downstream of protein-coding genes and likely represent a previously undetected population of edited 3’UTRs. Many EERs are predicted to possess strong internal secondary structures as well as sequence complementarity with other EERs, indicative of both intramolecular and intermolecular dsRNAs. In addition to numerous EERs associated with coding genes, we identified a population of prospective noncoding EERs, distant from protein coding genes with no overlapping genome annotation and little to no coding potential. By combining next-generation RNA sequencing with freely available bioinformatics tools, our workflow provides an easily accessible approach for the identification of dsRNAs, and more importantly, a comprehensive catalogue of the C. elegans dsRNAome.

overall_design:
RNA-seq of dsRNA isolated from N2 and dcr-1(mg375) embryos and mixed stage worms using immunoprecipitation with various antibodies.
Please note that the data from 6 biological samples were combined to generate a single set of alignments used to make all the processed data files  (*bed and *bw files linked as Series supplementary files).

middle: M

first: Osama

middle: A

last: Youssef

first: P

middle: J

last: Aruscavage

first: David

last: Nix

first: Changjin

last: Hong

first: W

middle: E

last: Johnson

first: Brenda

middle: L

last: Bass


title: Transcriptome profile form endometrial biopsy samples of pregnant and non-pregnant cows

city: Pirassununga

first: Mario

last: Binelli

state: SP

address: Rua Duque de Caxias Norte, 255

department: department of animal reproduction

country: Brazil

institute: University of Sao Paulo (USP)

summary:
Purpose: characterize the uterine transcriptome profiles of pregnant (P) versus non-pregnant (NP) cows during early pregnancy and attempted to define a potential set of marker genes that can be valuable for predicting pregnancy outcome.
Methods: beef cows were synchronized and artificially inseminated at detected estrus. Six days after AI, jugular blood samples and a biopsy from the uterine horn contralateral to the ovary containing the corpus luteum were collected. Based on pregnancy outcome on day 30, samples were retrospectively allocated to the following groups: Pregnant and Non-Pregnant. Both groups had similar plasma progesterone concentrations on D6. Uterine biopsies were submitted to RNA-Seq analysis in a Illumina HiScanSQ platform.
Results: The 272,685,768 million filtered reads were mapped to the Bos taurus reference genome and 14,654 genes were analyzed for differential expression between groups. Transcriptome data showed that 216 genes are differently expressed when comparing NP versus P uterine tissue (Padj≤0.1). More specifically, 36 genes were up-regulated in P cows and 180 are up-regulated in NP cows.
Conclusions: this study characterized a unique set of genes, expressed in the uterus on 6 days after insemination, that indicate a receptive state leading to pregnancy success. Furthermore, expression of such genes can be used as potential markers to efficiently predict pregnancy success.

overall_design:
endometrial mRNA profiles of pregnant and non-pregnant cows were generated by deep sequencing using Illumina HiScanSQ platform
BioProject PRJNA268916
SRA Study SRP05036

first: Saara

last: Scolari

first: Angela

last: Gonella-Diaza

first: Sonia

last: Andrade

first: Gustavo

last: Gasparin

first: Luiz

last: Lehmann Coutinho


title: CapStarr-seq: a high-throughput method for quantitative assessment of enhancer activity in mammals

city: Marseille - cedex 09

first: Aurélien

last: Griffon

address: 163, Avenue de Luminy - case 928

country: France

institute: TAGC - Inserm U1090

summary: Here we developed CapStarr-Seq, a novel high-throughput strategy to quantitatively assess enhancer activity in mammals. This approach couples capture of regions of interest to previously developed Starr-seq technique. Extensive assessment of CapStarr-seq demonstrated accurate quantification of enhancer activity. Furthermore, we found that enhancer strength correlates with binding complexity of tissue-specific transcription factors and super-enhancers, while additive enhancer activity isolates key genes involved in cell identity and function.

overall_design: CapStarr-seq analysis in P5424 cell line (2 replicates), 3T3 cell line and in the plasmid library before (Input) and after transfection

first: Laurent

last: Vanhille

first: Ahmad

last: Maqbool

first: Joaquin

last: Zacarias

first: Nicolas

last: Fernandez

first: Benoit

last: Ballester

first: Jean-Christophe

last: Andrau

first: Salvatore

last: Spicuglia


title: Neurotrophic Tyrosine Kinase Receptor 1 is Epigenetically Regulated by IL-13 and Contributes to Allergic Inflammation

city: Cincinnati

first: Mark

last: Rochman

state: OH

address: 3333 Burnett Ave

department: Allergy and Immunology

institute: CCHMC

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Despite the well-recognized role of IL-13–induced transcriptional responses in allergic inflammation, the epigenetic mechanisms driven by IL-13 have not been well defined. We interrogated the transcriptional and epigenetic signatures of IL-13-induced epithelial responses focusing on the chromatin activation marks H3K4me3, H3K9Ac, and H3K27Ac. ChIP-sequencing analysis revealed that IL-13–inducible genes were epigenetically poised for induction and continued to accumulate epigenetic changes in response to IL-13. By intersecting the transcriptome and the epigenome of the IL-13 response, we identified neurotrophic tyrosine kinase receptor 1 (NTRK1) as a major target of IL-13 in epithelial cells. Using eosinophilic esophagitis as a model system for human allergic inflammation, we found that NTRK1 was dramatically induced in inflamed esophageal biopsies, and downstream mediators of NTRK1 signaling were elevated in diseased tissue. The NTRK1 ligand nerve growth factor (NGF) was constitutively expressed in control and disease states, indicating that induction of the receptor by IL-13 limited pathway activation. In epithelial cells, NGF and IL-13 synergistically induced transcription and secretion of the key eosinophil chemoattractant CCL26 (eotaxin-3). In summary, we demonstrate that IL-13–mediated allergic responses are epigenetically driven and identify NTRK1 as a novel epigenetic and transcriptional target of IL-13 that uniquely contributes to allergic inflammation.

overall_design: Human esophageal epithelial cell line TE-7 was stimulated with IL-13 at 100 ng/ml for 2 hr, 6 hr and 24 hr and subjected to RNA-sequencing. In parallel, TE-7 cells were induced with IL-13 for 6 hr and subjected to ChIP-sequencing analysis for H3K4me3, H3K9Ac and H3K27Ac activating chromatin marks.

first: Marc

last: Rothenberg


title: Profile of gene expression in U87-MG xenografts expressing control vector (V0), the ubiquitin ligase KPC1 or the p50 subunit of the NF-kB transcription factor, using RNASeq analysis of transcripts mapped independently to the human and murine genomes

city: Haifa

first: Aaron

last: Ciechanover

state: Haifa

address: Efron Street, Bat Galim, POB 9649

department: Faculty of Medicine, Cancer and Vascular Biology Research Center

country: Israel

institute: Technion-Israel Institute of Technology

summary:
Purpose: We identified KPC1 as the ubiquitin ligase that binds to the p105 precursor of NF-kB, ubiquitinates it and mediates its proteasomal processing to generate the p50 active subunit of the transcription factor. Using U87-MG human glioblastoma xenografts, we observed that overexpression of KPC1 results in strong inhibition of tumor growth mediated via excessive generation of p50.The goal of this RNASeq study was to analyze the profile of gene expression in xenografts overexpressing control (V0), KPC1 or p50 vectors, and to further understand how the altered gene expression patterns can explain the tumor suppressive effect we observed.
Results:Transcript analysis of U87-MG xenografts overexpressing control (V0), KPC1 or p50 vector mapped  to the human genome  revealed:
• A strong similarity between overexpression of p50 and KPC1 (correlation of 0.51, p-value<10-300 )
• A specific signature of NF-kB targets [21 of the consistently changed genes are known to be regulated by NF-kB (p-value<3.4×10-9 )]
• A significant (p-value<1.4×10-18) increase in the expression of 40 tumor suppressor genes, with no significant change in other classes.
• A significant down regulation of a cluster of genes including LIN28B, IL-6, HMAGA2 and VEGFA. This finding links well to an established regulatory axis involving LIN28B, Let-7 microRNA, and IL-6 in inflammation and cell transformation that is regulated by NF-kB.

overall_design:
Exponentially growing U87-MG cells were stably transfected with an empty vector (V0) or vectors coding for Myc-KPC1 or Flag-p50. Cells were dissociated with trypsin, washed with PBS, and brought to a concentration of 50×10^6 cells/ml. Cell suspension (5×10^6/0.1 ml) was inoculated subcutaneously at the right flank of 7-weeks old male Balb/C nude mice (n=7). Following 21 days, mRNA from U87-MG xenografts was isolated using an RNA purification kit, and analyzed using the Illumina HiSeq 2500 sequencer. The RNASeq analysis experiment was repeated twice independently.
Run1 included a total of 7 samples. Samples 1-3 were isolated from V0 – control tumors (3 different tumors), samples 4-5 were isolated from KPC1-expressing  tumors (2 different pools of 3 tumors each due to small tumor size), and samples 6-7 were isolated from p50-expressing tumors for (2 different pools of 2-3 tumors each, due to very small tumor size).
Run2 included a total of 5 samples. Samples 8-10 were isolated from V0 (control) tumors (3 different tumors), samples 11-12 were isolated from KPC1 tumors (2 different pools of 3 tumors each due to small tumor size). Several repeated attempts to extract RNA from the p50-expressing tumors did not yield any results, as the tumors were miniscule.

first: Yelena

last: Kravtsova-Ivantsiv

first: Inna

last: Shomer

first: Victoria

last: Cohen-Kaplan

first: Berend

last: Snijder


title: Genome-wide analysis of Smad2 and FoxH1 binding sites in zebrafish blastula

status: Public on Apr 08 2015

city: Cambridge

first: Julien

last: Dubrulle

state: MA

address: 16 Divinity Ave

department: MCB

institute: Harvard University

summary: Morphogen gradients expose cells to different signal concentrations and induce target genes with different ranges of expression. To determine how the Nodal morphogen gradient induces distinct gene expression patterns during zebrafish embryogenesis we characterized the binding sites of the Nodal-specific transcription co-activators Smad2 and FoxH1 in wild-type, Nodal-overexpressing or Nodal signaling inhibited embryos. We found that FoxH1 binds to Nodal targets loci even in the absence of the signal, while Smad2 only binds and colocalizes to FoxH1 upon induction of the Nodal pathway.

overall_design: 6 samples total.

first: Laila

last: Akhmetova

first: Alexander

middle: F

last: Schier


title: Mutational analysis of measles virus suggests constraints on antigenic variation of the glycoproteins

city: New York

first: Nicholas

middle: S

last: Heaton

state: NY

address: One Gustave L Levy Place

department: Microbiology

institute: Icahn School of Medicine at Mount Sinai

summary: RNAseq of mutant measles virus libraries

overall_design:
Insertional mutagenesis of measles to identify tolerated locations
Mutant measles virus libraries were constructed as described in:    Heaton, N.S., Sachs, D., Chen,C.J., Hai, R., and Palese, P. (2013).    Genome-wide mutagenesis of influenza virus reveals unique plasticity of    the hemagglutinin and NS1 proteins. PNAS. 110, 20248-20253.

first: Peter

last: Palese


title: MicroRNA-dependent transposable element transcriptional silencing in Drosophila somatic ovarian cells [RNA-Seq]

city: MONTPELLIER

first: SEVERINE

last: CHAMBEYRON

address: 141, rue de la Cardonille

department: Institute of Human Genetics

institute: CNRS

type: Non-coding RNA profiling by high throughput sequencing

summary: RNAi-related silencing mechanisms concern as diverse biological processes as gene regulation, mostly via the miRNA pathway, and defense against molecular parasites, mainly controlled by the siRNA and the piRNA pathways. In Drosophila somatic ovarian cells, transposable elements (TEs) are repressed by chromatin changes induced by Piwi-interacting RNAs (piRNAs). We show here that a functional miRNA pathway is required for this piRNA-mediated TE transcriptional silencing to operate in this tissue. A general miRNA depletion, caused by tissue- and stage-specific knock-down of either drosha (involved in miRNA biogenesis) or AG01 and gawky (both responsible for miRNA activity) resulted in chromatin-mediated TE transcriptional desilencing and piRNA loss. For unknown reasons, the amount of piRNA produced by the traffic jam 3' UTR was apparently unaffected in miRNA-defective somatic ovarian cells. Although weaker, similar phenotypes could also be observed upon individual titration (by expression of the complementary miR-sponge) of at least three miRNAs, miR-14, miR-34 and miR-989. This work adds the maintenance of genome stability, via the piRNA-mediated TE repression, to the list of the already reported miRNA-controlled biological functions.

overall_design: Study of small regulatory RNA populations present in ovaries expressing either wild-type Drosha protein or the trans-dominant negative Drosha protein coupled or not with the Tub-GAL80[ts]GAL4 inhibitor.

first: Bruno

last: Mugat

first: Abdou

last: Akkouche

first: Vincent

last: Serrano

first: Claudia

last: Armenise

first: Blaise

last: Li

first: Christine

last: Brun

first: Tudor

last: Fulga

first: David

last: Van Vacto

first: Alain

last: Pélisson

first: Séverine

last: Chambeyron


title: MicroRNA-dependent transposable element transcriptional silencing in Drosophila somatic ovarian cells [RIP-Seq]

overall_design: Analyses of RNA present in 3 different genetic backgounds with or without Drosha-IP: a control, a second one expressing the wild-type Drosha protein and a third one expressing the trans-dominant negative Drosha protein in the somatic cells.


title: MIWI and piRNA-mediated cleavage of messenger RNAs in mouse testes

city: Beijing

first: Jiao

last: Yuan

address: 15 Datun Road, Chaoyang District

country: China

institute: Institute of Biophysics, Chinese Academy of Sciences

summary: The piRNA machinery is known for its role in mediating epigenetic silencing of transposons. Recent studies suggest that this function also involves piRNA-guided cleavage of transposon-derived transcripts. As many piRNAs also appear to have the capacity to target diverse mRNAs, this raises the intriguing possibility that piRNAs may act extensively as siRNAs to degrade specific mRNAs. To directly test this hypothesis, we compared mouse PIWI (MIWI)-associated piRNAs with experimentally identified cleaved mRNA fragments from mouse testes, and observed cleavage sites that predominantly occur at position 10 from the 5' end of putative targeting piRNAs. We also noted strong biases for U and A residues at nucleotide positions 1 and 10, respectively, in both piRNAs and mRNA fragments, features that resemble the pattern of piRNA amplification by the 'ping-pong' cycle. Through mapping of MIWI-RNA interactions by CLIP-seq and gene expression profiling, we found that many potential piRNA-targeted mRNAs directly interact with MIWI and show elevated expression levels in the testes of Miwi catalytic mutant mice. Reporter-based assays further revealed the importance of base pairing between piRNAs and mRNA targets and the requirement for both the slicer activity and piRNA-loading ability of MIWI in piRNA-mediated target repression. Importantly, we demonstrated that proper turnover of certain key piRNA targets is essential for sperm formation. Together, these findings reveal the siRNA-like function of the piRNA machinery in mouse testes and its central requirement for male germ cell development and maturation.

overall_design: CLIP-Seq (Crosslinking Immunoprecipitation coupled with high-throughput sequencing) experiments targeting Miwi in isolated round spermatids from mouse testis.

first: Peng

last: Zhang

first: Junyan

last: Kang

first: Lantao

last: Gou

first: Jiajia

last: Wang

first: Yuanchao

last: Xue

first: Geir

last: Skogerboe

last: Dai

first: Dawei

last: Huang

first: Runsheng

last: Chen

first: Xiangdong

last: Fu

first: Mofang

last: Liu

first: Shunmin

last: He


title: The hub protein Loqs connects the miRNA and endo-siRNA pathways in mosquitos

city: College Station

first: Zach

last: Adelman

state: TX

address: 370 Olsen Blvd

department: Entomology

institute: Texas A&M University

summary: Aedes aegypti mosquitoes vector several arboviruses of global health significance, including dengue viruses and chikungunya virus. RNA interference (RNAi) plays an important role in antiviral immunity, gene regulation and protection from transposable elements. Double-stranded RNA binding proteins (dsRBPs) are important for efficient RNAi; in Drosophila functional specialization of the miRNA, endo-siRNA and exo-siRNA pathway is aided by the dsRBPs Loqs-PB, Loqs-PD and R2D2, respectively. However, this functional specialization has not been investigated in other dipterans. Characterization of the gene structure, expression pattern and interactions with other RNAi/miRNA components revealed that mosquito Loqs/R3D1 isoform -PA, but not -PB interacted readily with both AeAGO1 and AeAGO2. This interaction was mapped to a 32 a.a. region predicted to be structurally similar to the unique 22 a.a. tail of Drosophila Loqs-PD. No -PD isoforms could be detected in Ae. aegypti; analysis of other dipteran genomes demonstrated that this isoform is not conserved outside of Drosophila. Overexpression experiments indicated that Loqs/R3D1-PA participates in endo-siRNA, but not exo-siRNA based silencing. We conclude that the functional specialization of Loqs-PD in Drosophila is a recently derived trait, and that in other dipterans, including the medically important mosquitoes, Loqs/R3D1-A participates in both the miRNA and endo-siRNA based pathways.

overall_design: 12 samples, 3 replicates of each type of sample.  The GFP sample is the reference sample

first: Mary

last: Haac

first: Michelle

last: Anderson

first: Heaher

last: Eggleston

first: Kevin

last: Myles

middle: N


title: MicroRNA-dependent transposable element transcriptional silencing in Drosophila somatic ovarian cells

first: SEVERINE

last: CHAMBEYRON


title: PU.1 regulates T-lineage gene expression and progression via indirect repression during early T-cell development

city: Los Angeles

first: Ameya

last: Champhekar

state: California

address: 700 Tiverton Ave, 11-272 Factor Bldg

department: Medicine, HemOnc Division

institute: University of California, Los Angeles

summary: The Ets family transcription factor PU.1 is essential for the development and maintenance of several hematopoietic lineages. In the thymus, PU.1 is expressed only in the early ETP/DN1, DN2a and DN2b stages of development. While PU.1 deletion in multipotent precursors leads to a complete block in T-cell development its function in the intrathymic stages in which it is expressed remains undetermined. The goal of this expression profiling study was to determine if PU.1 regulates the expression of T-lineage genes during the early stages of development. To do this, we generated the PU.1-Eng construct which expresses a fusion protein containing the DNA binding ETS domain of PU.1 (aas 159-260) fused to the obligate repressor domain (aas 1-298) of the Drosophila engrailed protein. The PU.1-ETS construct only expresses the ETS domain of PU.1 (aas 159-260) and serves as a control. Fetal liver precursors were isolated from e14.5 embryos and co-cultured with OP9-DL1 cells in the presence of IL-7 and Flt3L (5 ng/ml each) for 4 days to obtain FLDN1, DN2a and DN2b cells. These were infected with vector only, PU.1-ETS and the PU.1-Eng constructs and DN2 cells were sorted after 20 hours of infection. Total RNA was isolated from these cells and polyA+ fraction was used to prepare libraries for high throughput sequencing. Libraries prepared from 2 independent sets of samples were subjected to non-strand specific single-end sequencing.

overall_design: Two sets of samples generated from fetal liver precursor derived DN2 cells expressing PU.1-ETS and PU.1-Eng constructs were used for expression profiling. The LZRS retroviral vector, without any insert, was used to generate the vector control dataset.

first: A

first: S

last: Damle

first: E

middle: V

last: Rothenberg


title: A species-specific nucleosomal signature defines a periodic distribution of amino acids in proteins

status: Public on Apr 10 2015

city: Salamanca

first: Luis

last: Quintales

address: Calle Zacarías González, 2

department: Universidad de Salamanca / CSIC

country: Spain

institute: Instituto de Biología Funcional y Genómica (IBFG)

summary: Sequencing of mononucleosomal DNA during asynchronous mitosis in Schizosaccharomyces pombe, Schizosaccharomyces octosporus, Schizosaccharomyces japonicus and Saccharomyces cerevisiae

sample_organism:
Schizosaccharomyces pombe
Schizosaccharomyces japonicus
Schizosaccharomyces octosporus
Saccharomyces cerevisiae W303

overall_design: Samples from mononucleosomal DNA from asynchronous mitosis of four species of budding (Saccharomyces cerevisiae W303-1a) and fission yeasts (S. pombe wild type 972h-, S. octosporus CBS1804, S. japonicus var. japonicus ade12- FY53) were sequenced (Illumina Genome Analyzer IIx and HiSeq 2500) using the single read and paired end protocol.

first: Ignacio

last: Soriano

first: Enrique

last: Vázquez

first: Francisco

last: Antequera

first: Mónica

last: Segurado


title: Chromatin immunoprecipitation with next generation sequencing (ChIP-seq) of H3K27ac in primary human macrophages and foam cells

city: Oxford

first: Christopher

last: O'Callaghan

state: Oxfordshire

address: Roosevelt Drive

department: Nuffield Department of Medicine

country: United Kingdom

institute: University of Oxford

summary: We report the enhancer landscape in primary human macrophages and foam cells using ChIP-seq for the H3K27ac histone mark

overall_design: CD14+ monocytes were isolated from the blood of 2 healthy male volunteers. Monocytes were differentiated into macrophages by culture for 7 days with 50ng/ml macrophage colony stimulating factor and then treated for 48 hours with either oxidized low density lipoprotein (oxLDL) to induce foam cell formation or with a control buffer that lacked oxLDL. The resulting 4 samples were then subjected to ChIP-seq for H3K27ac.

first: Michael

last: Reschen


title: Identification of novel, highly expressed retroviral microRNAs in cells infected by bovine foamy virus

status: Public on Apr 07 2015

city: Durham

first: Adam

middle: Wesley

last: Whisnant

state: NC

address: 213 Research Drive 427 CARL Bldg

department: Molecular Genetics and Microbiology

institute: Duke University

summary: While numerous viral microRNAs (miRNAs) expressed by DNA viruses, especially herpesvirus family members, have been reported, there have been very few reports of miRNAs derived from RNA viruses. Here we describe three miRNAs expressed by bovine foamy virus (BFV), a member of the spumavirus subfamily of retroviruses, in both BFV-infected cultured cells and BFV-infected cattle. All three viral miRNAs are initially expressed in the form of an ∼ 122-nucleotide (nt) pri-miRNA, encoded within the BFV long terminal repeat U3 region, that is subsequently cleaved to generate two pre-miRNAs that are then processed to yield three distinct, biologically active miRNAs. The BFV pri-miRNA is transcribed by RNA polymerase III, and the three resultant mature miRNAs were found to contribute a remarkable ∼ 70% of all miRNAs expressed in BFV-infected cells. These data document the second example of a retrovirus that is able to express viral miRNAs by using embedded proviral RNA polymerase III promoters.

overall_design: Small RNA profiling of Bovine Foamy Virus (BFV)-infected cells through total small RNA deep sequencing

middle: W

first: Timo

last: Kehl

first: Qiuying

last: Bao

first: Magdalena

last: Materniak

first: Jacek

last: Kuzmak

first: Martin

last: Löchelt

first: Bryan

middle: R

last: Cullen


title: A study of the effect of OxLDL on C/EBP beta binding in human macrophages using chromatin immunoprecipitation for C/EBP beta with high throughput sequencing (ChIP-seq)

summary: We report the C/EBP beta transcription factor binding landscape in human macrophages and oxLDL-induced foam cells

overall_design: ChIP-seq for C/EBP beta binding was performed in human macrophages and oxLDL induced-foam cells with quadruple biological replicates


title: Lipid-induced epigenomic changes in human macrophages identify a coronary artery disease associated variant that regulates PPAP2B expression through altered C/EBP-beta binding

overall_design: Refer to individual Series


title: A function for the hnRNP A1/A2 proteins in transcription elongation

city: Kansas City

first: Marco

last: Blanchette

state: MO

institute: Stowers Institute for Medical Research

summary: The hnRNP A1 and A2 proteins regulate processes such as alternative pre-mRNA splicing and mRNA stability. Here, we report that a reduction in the levels of hnRNP A1 and A2 by RNA interference or their cytoplasmic retention by osmotic stress drastically increases the transcription of a reporter gene. Based on previous work, we propose that this effect may be linked to a decrease in the activity of the transcription elongation factor P-TEFb. Consistent with this hypothesis, the transcription of the reporter gene was stimulated when the catalytic component of P-TEFb, CDK9, was inhibited with DRB. While low levels of A1/A2 stimulated the association of RNA polymerase II with the reporter gene, they also increased the association of CDK9 with the repressor 7SK RNA, and compromised the recovery of promoter-distal transcription on the Kitlg gene after the release of pausing. Transcriptome analysis revealed that more than 50% of the genes whose expression was affected by the siRNA-mediated depletion of A1/A2 were also affected by DRB. RNA polymerase II-chromatin immunoprecipitation assays on DRB-treated and A1/A2-depleted cells identified a common set of repressed genes displaying increased occupancy of polymerases at promoter-proximal locations, consistent with pausing. Overall, our results suggest that lowering the levels of hnRNP A1/A2 elicits defective transcription elongation on a fraction of P-TEFb-dependent genes, hence favoring the transcription of P-TEFb-independent genes.

overall_design: two treatements and one control

first: Bruno

last: Lemieux

first: Anne

last: Monette

first: Andrew

last: Mouland

first: Raymund

last: Wellinger

first: Benoit

last: Chabot


title: Formaldehyde-assisted isolation of regulatory elements with next generation sequencing (FAIRE-seq) in primary human macrophages and foam cells

summary: We report the open chromatin landscape in primary human macrophages and foam cells using FAIRE-seq

overall_design: CD14+ monocytes were isolated from the blood of 3 healthy volunteers. Monocytes were differentiated into macrophages by culture for 7 days with 50ng/ml macrophage colony stimulating factor and then treated for 48 hours with either oxidized low density lipoprotein (oxLDL) to induce foam cell formation or with a control buffer that lacked oxLDL. The resulting six samples were then subjected to FAIRE-seq using an established protocol (Simon JM, Giresi PG, Davis IJ, Lieb JD. Using formaldehyde-assisted isolation of regulatory elements (FAIRE) to isolate active regulatory DNA. Nature protocols 2012;7:256-67).


title: WT and Mk2/3 KO MEFs response to etoposide

city: Koln

first: Jorge

last: Boucas

address: Joseph-Stelzmann-Str. 9b

department: Bioinformatics

country: Germany

institute: Max Planck Institute for Biology of Ageing

summary: Primary mouse embryonic fibroblasts (MEFs) from wt and MK2/3 KO mice were treated with 20µM etoposide.

overall_design: Primary MEFs from wt and MK2/3 KO mice were treated with 20µM etoposide and RNA extracted after 1h and 6h.

first: Christoph

last: Dieterich

first: H

middle: C

last: Reinhardt


title: RNA-sequencing of healthy human skeletal myocytes

city: Gothenburg

first: Leif

last: Väremo

address: Kemivägen 10

department: Biology and Biological Engineering

country: Sweden

institute: Chalmers University of Technology

summary: Skeletal myocytes are metabolically active and susceptible to insulin resistance, thus implicated in type 2 diabetes (T2D). This complex disease involves systemic metabolic changes and their elucidation at the systems level requires genome-wide data and biological networks. Genome-scale metabolic models (GEMs) provide a network-context to integrate high-throughput data. We generated myocyte-specific RNA-seq data and investigated their correlation with proteome data. These data were then used to reconstruct a comprehensive myocyte GEM. Next, we performed a meta-analysis of six studies comparing muscle transcription in T2D versus healthy subjects. Transcriptional changes were mapped on the myocyte GEM, revealing extensive transcriptional regulation in T2D, particularly around pyruvate oxidation, branched-chain amino acid catabolism, and tetrahydrofolate metabolism, connected through the down-regulated dihydrolipoamide dehydrogenase. Strikingly, the gene signature underlying this metabolic regulation successfully classifies the disease state of individual samples, suggesting that regulation of these pathways is a ubiquitous feature of myocytes in response to T2D.

overall_design: Isolated skeletal muscle precursor cells from six normal glucose tolerant and non-obese males and females were differentiated in vitro. RNA from fully differentiated myotubes was sequenced using RNA-seq.

first: Camilla

last: Scheele

first: Christa

last: Broholm

first: Adil

last: Mardinoglu

first: Caroline

last: Kampf

first: Anna

last: Asplund

first: Intawat

last: Nookaew

first: Mathias

last: Uhlén

first: Bente

last: Klarlund Pedersen

first: Jens

last: Nielsen


title: mRNA profiling of GGT-HIF1aM3 (γ-HIF1αM3, TRACK) kidney cortex

first: Leiping

last: Fu

address: 1300 York Avenue

department: Pharmacology

institute: Weill Cornell Medical College

summary: The complete transcriptomes of kidney cortex from 3 TRACK/γ-HIF1αM3 ~15 month old TG+ male mice and 3 age matched TG- C57BL/6 male mice were sequenced on an Illumina HiSeq2000 Sequencer.

overall_design: Examination of complete transcriptome of kidney cortex between TRACK/γ-HIF1αM3 TG+ male mice and TG- C57BL/6 male mice

first: David

last: Nanus

first: Lorraine

last: Gudas


title: Non-viral generation of marmoset monkey iPS cells by a six-factor-in-one-vector approach

city: Goettingen

first: Gabriela

last: Salinas

state: Lower-Saxony

address: Kreuzbergring 57

department: Department of Pathology

institute: Universitaetsmedizin Goettingen

summary: The analysis compares primary fibroblasts initially used for reprogramming, established marmoset ES cells and a marmoset  iPS cell line which was generated witha non-viral approach using a six-factor-in-one-vector approach

overall_design: Cells were grown under standard conditions for marmoset pluripotent stem cells and primary fibroblasts without additional treatment. For each cell type at least four independent cell preparations were used.

first: Katharina

last: Debowski

first: Rita

last: Warthemann

first: Jana

last: Lentes

last: Salinas-Riester

first: Ralf

last: Dressel

first: Daniel

last: Langenstroth

first: Jörg

last: Gromoll

first: Erika

last: Sasaki

first: Rüdiger

last: Behr


title: RNA-Seq of retina and RPE/choroid tissues from wild type, Abca4 knockout and Abca4 L541P;A1038V knockin mice

city: Cleveland

first: Ning

last: Zhang

address: 10900 Euclid Ave.

institute: Case Western Reserve University

summary: We report RNA-Seq analysis of the transcriptome of retinas and RPE/choroids from Abca4 knockout, Abca4 L541P;A1038V knockin and control wild type mice in order to better understand changes in gene regulation that could lead to retinal pathology in mice with ABCA4 deficiency/defect.

overall_design: Retinal and RPE/choroidal mRNA profiles of 30-day-old wild type (WT), Abca4-/- and Abca4L541P;A1038V/L541P;A1038V mice were generated by RNA-Seq, using Illumina Hiseq 2500

first: Krzysztof

last: Palczewski


title: Effect of Bicaudal C1 deletion on E13.5 mouse pancreatic mRNA profile

city: Copenhagen

first: Laurence

middle: Anne

last: Lemaire

address: Blegdamsvej, 3b building6 floor4

department: DanStem

country: Denmark

institute: University of Copenhagen

summary: Bicaudal C1 KO embryonic pancreas develops cysts and has less endocrine progenitors after E14.5 (14.5 days after fertilization), while no defect is observed at E13.5. Bicaudal C1 is an RNA-binding protein. To understand the molecular mechanisms leading to both phenotypes, the mRNA expression profile of E13.5 WT vs. Bicaudal C1 dorsal pancreas was studied by high-throughput sequencing using the Illumina HiSeq 2000 platform. This time point was selected as a time point where no phenotypic modification was detected. 3 replicates for each condition were sequenced and each replicate consisted of 3 E13.5 dorsal pancreas of the same genotype. There were only few differences between both transcriptomes. Pkd2 was 1.9-fold reduced in Bicaudal C1 KO. Pkd2 inactivation causes renal and pancreatic cyst. Some genes having immune/inflammatory functions were up or downregulated highlighting the early immune cell infiltration observed in Bicaudal C1 KO pancreas. No hit could explain the endocrine progenitor decrease. It may be due to the ability of Bicaudal C1 to regulate mRNA translation without affecting the mRNAs themselves.

overall_design: Comparison of mRNA profiles of E13.5 WT vs. Bicaudal C1 KO dorsal pancreas using the Illumina HiSeq 2000 platform (platform ID SN865), 3 replicates per condition.

first: Solenne

last: Carat

first: Jacques

last: Rougemont

first: Anne

last: Grapin-Botton


title: FBXL10 and the Protection of Polycomb-bound Genes from Hypermethylation (RNA-Seq)

status: Public on Apr 06 2015

city: St. Louis

first: John

middle: R.

last: Edwards

address: 660 S. Euclid Ave, Campus Box 8220

department: Center for Pharmacogenomics

institute: Washington University School of Medicine

summary: Nearly all CpG-dense promoters are occupied by the multi-domain chromosomal protein FBXL10. We show here that complete inactivation of the Fbxl10 gene leads to dense de novo methylation only of the promoters that are co-occupied by both FBXL10 and by Polycomb Repressive Complexes; this results in pervasive defects in embryonic development and death of homozygous Fbxl10 mutant embryos at midgestation. Deletion of key components of Polycomb Repressive Complexes 1 and 2 did not lead to ectopic de novo methylation. These results indicate that FBXL10 defends Polycomb-occupied promoters against ectopic de novo methylation. FBXL10 is the first reported factor whose loss leads to a gain in genomic DNA methylation.

overall_design: DNA methylation analysis using RRBS and expression analysis using RNA-seq was performed on WT and Fbxl10T/T ES cells.

first: Mathieu

last: Boulard

first: Timothy

middle: H

last: Bestor


title: The transcriptional cofactor TRIM33 prevents apoptosis in B lymphoblastic leukemia by deactivating a single enhancer

city: Seoul

first: Jae Seok

last: Roe

address: Yonsei-ro, 50

country: South Korea

institute: Yonsei University

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


title: FBXL10 Protects Polycomb-bound Genes from Hypermethylation


title: The contribution of cohesin-SA1 to chromatin architecture and gene expression in two murine tissues

city: Madrid

first: Ana

last: Losada

state: Madrid

address: C/ Melchor Fernández Almagro, 3.

department: Molecular Oncology Programme

institute: Centro Nacional de Investigaciones Oncológicas (CNIO)

summary: Cohesin, which consists of SMC1, SMC3, Rad21 and either SA1 or SA2, topologically embraces the chromatin fibers to hold sister chromatids together and to stabilize chromatin loops. Increasing evidence indicates that these loops are the organizing principle of higher-order chromatin architecture, which in turn is critical for gene expression. To determine how cohesin contributes to the establishment of tissue-specific transcriptional programs, we compared genome-wide cohesin distribution, gene expression and chromatin architecture in cerebral cortex and pancreas from adult mice. More than one third of cohesin binding sites differ between the two tissues and these are enriched at the regulatory regions of tissue-specific genes. Cohesin colocalizes extensively with the CCCTC-binding factor (CTCF). Cohesin/CTCF sites at active enhancers and promoters contain, at least, cohesin-SA1 whereas either cohesin-SA1 or cohesin-SA2 are present at active promoters independently of CTCF. Analyses of chromatin contacts at the Protocadherin gene cluster and the Regenerating islet-derived (Reg) gene cluster, mostly expressed in brain and pancreas respectively, revealed remarkable differences in the architecture of these loci in the two tissues that correlate with the presence of cohesin. Moreover, we found decreased binding of cohesin and reduced transcription of the Reg genes in the pancreas of SA1 heterozygous mice. Given that Reg proteins are involved in the control of inflammation in pancreas, such reduction may contribute to the increased incidence of pancreatic cancer reported in these animals.

overall_design: Examination of the relationship between gene expression, genome wide cohesin distribution and chromatin structure

last: Cuadrado

first: Silvia

last: Remeseiro

first: Osvaldo

last: Graña

first: David

middle: G

last: Pisano


title: Cell type-specific expression profile and signaling requirements in early hematopoietic reprogramming

city: Dalian

first: Lan

last: Kang

address: 9 West Section, Lvshun South Road

department: Institute of Cancer Stem Cell

institute: Dalian Medical University

summary: Hematopoietic cells represent an attractive starting cell type for induced pluripotent stem (iPS) cells induction, yet the molecular mechanisms in hematopoietic reprogramming are poorly defined. In this study, we showed that long-term hematopoietic stem cells are more amenable for iPS cells induction among several hematopoietic stem and progenitor cell (HSPC) populations, and that this is accompanied by an earlier induction of the transcriptional program that is involved in the promotion of macromolecule metabolism and cell proliferation. Notably, we identified multiple signaling pathways that exhibited distinct expression patterns in HSPCs compared to that of fibroblasts, which is the most commonly used model for probing the somatic reprogramming process. We further experimentally confirmed the differential requirements of the Wnt/β-catenin and transforming growth factor-beta (TGF-β) signaling pathways in these two cell types. These data demonstrate that hematopoietic cells have a cell-type specific transcriptional program and possess unique signaling requirements in the early phase of reprogramming.

overall_design: Examine the differential global gene expression among different hematopietic cells in reprogramming. LT-HSC, ST-HSC, MP and fibroblasts were used in this study. Doxcycline addition could induce expression of oct4, sox2, klf4 and c-myc, consequently reprogramming these cells into iPS cells. Hematopietic cells cultured with doxcycline for 0, 2, 4 days were sampled, and samples at the same timepoint without doxcycline were also taken to exclude vast gene expression change in hematopoietic cell in-vitro culture. Fibroblast samples induced by doxcycline for 0, 2, 4 days were also introduced to provide comparism between lineages.

first: Jun

last: Wu

first: Yu

last: Tao

first: Xiao-lei

last: Liu

first: Yan-ting

last: Ma

first: Shao-rong

last: Gao

first: Qian-fei

last: Wang


title: Pontin functions as an essential coactivator for Oct4 target genes and lincRNAs in embryonic stem cells.

city: Pohang

first: Bhin

last: Jin-hyuk

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

department: Department of Chemical engineering

institute: POSTECH

summary: The actions of transcription factors, chromatin modifiers, and noncoding RNAs are crucial for the programming of cellular states.  Although chromatin remodeling factors regulate the functional status of cells including pluripotency and differentiation, how they cross-talk with embryonic stem (ES) cell-specific transcription factors and noncoding RNAs to coordinate networks controlling of ES cell identity remain unknown.  Here, we find that Pontin chromatin remodeling factor plays an essential role as a coactivator for Oct4 target genes and large intergenic noncoding RNAs (lincRNAs) in ES cells. mRNA- and ChIP-sequencing analyses reveal that Pontin and Oct4 share a substantial set of target genes involved in maintenance of ES cells.  Intriguingly, Oct4-dependent coactivator function of Pontin extends to transcription of lincRNAs that are mainly involved in repression of differentiation in ES cells.  Together, our findings demonstrate newly identified Oct4-Pontin-lincRNA module plays critical roles in the ES cell circuitry to orchestrate cell fate determination program.

overall_design: For mRNA-sequencing, we obtained mRNAs from 1) Pontinf/f; CreER ES cells at 0, 3, or 4 days post-treatment with 4-hydroxy tamoxifen (OHT) for Pontin-depleted ES cells without biological replicates (n=1), 2) ZHBTc4 ES cells at 2 days post-treatment with tetracycline (Tc) for Oct4-depleted ES cells (n=1), and 3) ZHBTc4 ES cells infected by pLKO control or pLKO-shlinc1253 lentivirus at 4 days post infection for knockdown of linc1253 (n=2). For ChIP-sequencing, chromatin extracts containing DNA fragments with an average size of 400bp were immmunoprecipitated by using antibodies against GFP (control) or Pontin. Eluted ChIP DNA (n=1).

first: Kyungjin

last: Boo

first: Jinhyuk

last: Bhin

first: Daehee

last: Hwang

first: Ho

last: Lee

first: Sung

last: Baek


title: eIF3 PAR-CLIP in 293T cells

city: Boston

first: Amy

middle: Si-Ying

address: 450 Brookline Ave SM520

department: Cancer Immunology and Virology

institute: DFCI

summary: Regulation of protein synthesis is fundamental for all aspects of eukaryotic biology by controlling development, homeostasis, and stress responses. The 13-subunit, 800-kDa eukaryotic initiation factor 3 (eIF3) organizes initiation factor and ribosome interactions required for productive translation. However, current understanding of eIF3 function does not explain genetic evidence correlating eIF3 deregulation with tissue-specific cancers and developmental defects. Here we report the genome-wide discovery of human transcripts that interact with eIF3 using photo-activatable crosslinking and immunoprecipitation (PAR-CLIP). eIF3 binds to a highly specific programme of messenger RNAs (mRNAs) involved in cell growth control processes, including cell cycling, differentiation, and apoptosis, via the mRNA 5' untranslated region (5' UTR). Surprisingly, functional analysis of the interaction between eIF3 and two mRNAs encoding cell proliferation regulators, c-Jun and BTG1, reveals that eIF3 employs different modes of RNA stem loop binding to exert either translational activation or repression. Our findings illuminate a new role for eIF3 in governing a specialized repertoire of gene expression and suggest that binding of eIF3 to specific mRNAs could be targeted to control carcinogenesis.

overall_design:
293T cells were treated with 4-thiouridine and protein-RNA complexes were crosslinked, and eIF3-RNA complexes were immunoprecipitated.

Replicate 1 includes use of primer ID on the 5' end ("NNNNNNNNGUAC").

The 3' adapter used for all samples was:
5’ rApp /TGGAATTCTCGGGTGCCAAGG/ 3ddC/


title: Genome-wide analysis reveals dynamic changes in microRNAs expression during vascular cambium development in Chinese fir

city: Beijing

first: Lin

last: Jinxing

address: No.20 Nanxincun, Xiangshan

institute: Institute of Botany

summary: MicroRNAs (miRNAs) are small noncoding regulatory RNAs that play key roles in the process of plant development. To date, extensive studies of miRNAs have been performed in a few model plants, but few efforts have focused on small RNAs in conifers because of the lack of reference sequences for these enormous genomes. In this study, Solexa sequencing of three small RNA libraries obtained from dormant, reactivating, and active vascular cambium in Chinese ﬁr samples identified 76 known miRNAs from 27 miRNA families and 18 new potential miRNAs, of which 15 novel miRNA precursors were validated by RT-PCR and sequencing. More than half of these novel miRNAs displayed stage-specific expression patterns in the vascular cambium. Furthermore, analyzing the 103 miRNAs and their predicted targets indicated that 30% of miRNAs appeared to negatively regulate their targets, of which 4 target genes involved in the regulation of cambial cell division were validated via RNA ligase-mediated rapid amplification of 5’ cDNA ends (RLM 5’-RACE). Interestingly, miRNA156 and miRNA172 may regulate the phase transition in vascular cambium from dormancy to active growth. These results provide new insights into the important regulatory functions of miRNAs in vascular cambium development and wood formation in conifers.

overall_design: The aim of this study is to investigate the small RNA transcriptome in different stages of vascular cambium development in Cunninghamia lanceolata. We herein conducted our comprehensive analysis of a plant sRNAome during development of the vascular cambium in Chinese fir using cryosectioning to isolate cambial meristem, which allowed a higher cellular resolution for defining small RNA profiles and overcomed the limitations resulting from the mixed samples in many previous investigations. Total RNAs of dormant, reactivating, and active cambial meristem, respectively were isolated using the Concert Plant RNA Reagent (Invitrogen, Carlsbad, CA, USA), and were then treated with RNase-free DNase I (Promega, Madison, WI, USA). Twenty micrograms of total RNAs were used and 16 to 30-nt sRNAs were purified using Novex 15% TBE-¬Urea gel (Invitrogen). Two adaptors were sequentially ligated to the 5' and 3' ends of purified sRNAs. The ligation products were further purified from Novex 10% TBE-Urea gel. Reverse transcriptase SuperScript II (Invitrogen) and high-fidelity DNA polymerase Phusion (New England Biolabs, Ipswich, MA, USA) were used in the following RT-PCR reaction. The amplification products were cut from Novex 6% TBE-Urea gel. The purified DNA fragments were used for sequencing on an Illumina 2G Genome Analyzer at the Beijing Genomics Institute, Shenzhen, China.

first: Zongbo

last: Qiu

first: Yuanyuan

last: Zhao

first: Yinglang

last: Wan

first: Dechang

last: Cao

first: Jinxing

last: Lin


title: The transcriptional cofactor TRIM33 prevents apoptosis in B lymphoblastic leukemia by deactivating a single enhancer [ChIP-Seq]

first: Jae Seok

last: Roe

summary: Most mammalian transcription factors and cofactors occupy thousands of genomic sites and modulate the expression of large gene networks to implement their biological functions. In this study, we describe an exception to this paradigm. TRIM33 is identified here as a lineage dependency in B cell neoplasms and is shown to perform this essential function by associating with a single cis element. ChIP-seq analysis of TRIM33 in murine B cell leukemia revealed a preferential association with two lineage-specific enhancers that harbor an exceptional density of motifs recognized by the PU.1 transcription factor. TRIM33 is recruited to these elements by PU.1, yet acts to antagonize PU.1 function. One of the PU.1/TRIM33 co-occupied enhancers is upstream of the pro-apoptotic gene Bim, and deleting this enhancer renders TRIM33 dispensable for leukemia cell survival. These findings reveal an essential role for TRIM33 in preventing apoptosis in B lymphoblastic leukemia by interfering with enhancer-mediated Bim activation.

overall_design: ChIP-Seq for Trim33, Pu.1 and histone modification marks in different types of leukemia cells

first: Jae

first: Christopher

last: Vakoc


title: Glutaraldehyde resistance mechanisms in Pseudomonas fluorescens and Pseudomonas aeruginosa biofilms

city: Pittsburgh

first: Amit

last: Vikram

state: PA

address: 3700 O'Hara Street

department: Civil and Environmental Engineering

institute: University of Pittsburgh

summary: The objective of the current study was to understand the glutaraldehyde resistance mechanisms in P. fluorescens and P. aeruginosa biofilms. Glutaraldehyde is a common biocide used in various industries to control the microbial growth. Recent reports of emergence of glutaraldehyde resistance in several bacterial species motivated this study to understand the genetic factors responsible got glutaraldehyde resistance. Using a combination of phenotypic assays, chemical genetic assays and RNA-seq, we demonstrate that novel efflux pump, polyamine biosynthesis, lipid biosynthesis and phosphonate degradation play significant role in glutaraldehyde resistance and post-glutaraldehyde recovery of Psudomonad biofilms.

overall_design: Examination of P. fluorescens 72 h biofilm transcriptome was elucidated upon exposure to glutaraldehyde. The results were confirmed using qRT--PCR and chemical genetic appraoches in P. fluorescens and P. aeruginosa.

first: Jennifer

last: Bomberger

first: Kyle

last: Bibby


title: Transcriptome Sequencing (RNA-seq) of Normal Human Osteoblasts

city: Minneapolis

first: David

middle: A.

last: Largaespada

state: MN

address: 2231 6th Street SE

department: Dept. of Genetics, Cell Biology and Development, Cancer Center

institute: The University of Minnesota

summary: Three normal human osteoblast samples, acquired from PromoCell, were used as controls to compare to RNA-seq data from prepublished osteosarcoma samples (submitted to the European Bioinformatics Institute; EGAS00001000263) for the purpose of evaluating expression levels of genes identified as common insertions sites in a Sleeping Beauty screen of osteosarcomas in mice.

overall_design: Three normal human osteoblast samples (pellet form in RNAlater) were acquired from PromoCell (Heidelberg, Germany), and RNA was isolated from them immediately upon receipt.

first: Susan

middle: K

last: Rathe

first: Branden

last: Moriarity

middle: A


title: A transcriptional map following the developmental trajectory of the Arabidopsis stomatal lineage

city: Stanford

first: Dominique

last: Bergmann

state: CA

address: 371 Serra Mall

department: Biology Department

institute: Stanford University

summary: Developmental transitions can be described in terms of morphology and individual genes expression patterns, but also in terms of global transcriptional and epigenetic changes. Most of the large-scale studies of such transitions, however, have only been possible in synchronized cell culture systems. Here we generate a cell type specific transcriptome of an adult stem-cell lineage in the Arabidopsis leaf using RNA sequencing and microarrays. RNA profiles of stomatal entry, commitment, and differentiating cells, as well as of mature stomata and the entire aerial epidermis give a comprehensive view of the developmental progression.

overall_design: To obtain pure cell populations corresponding to different stomatal lineage cell types we used Fluorescent Activated Cell Sorting (FACS) in combination with markers of early (SPCHp::SPCH-YFP, SSY), middle (MUTEp::nucGFP, MG), and late (FAMAp::GFP-FAMA, FGF) precursor stages, as well of mature stomata (enhancer trap E1728::GFP, E1728G) and a marker of the entire epidermis (ML1p::YFP-RCI2A, ML1Y). To minimize transcriptional differences due to age, all cell types were sorted from 10-day old aerial rosettes, and the specificity of expression pattern at this common time point confirmed via confocal microscopy. Total RNA was extracted from purified protoplasts (4,000 to 20,000 cells/replicate; 2 replicates/marker line; 3 replicates/SSY marker line) and transcript abundance measured using RNA sequencing.

first: Jessika

last: Adrian

first: Jessica

last: Chang

first: Bastiaan

middle: O

last: Bargmann


title: Comparative transcriptome analyses of citrus root responses to Candidatus liberibacter asiaticus infection

status: Public on Apr 04 2015

city: Guangzhou

first: Chunzhen

last: Cheng

state: Guangdong

address: No.80 Dafeng No.2 street, Tianhe district, Guangzhou, China

department: Guangdong Academy of Agricultural Sciences

institute: Institute of fruit tree research

summary: Root samples of ‘Sanhu’ red tangerine trees infected with and without Candidatus Liberibacter asiaticus (CLas) were collected at 50 days post inoculation and subjected to RNA-sequencingto profile the differentially expressed genes (DEGs) . Results showed that a total of 3956 genes were differentially regulated by HLB-infection. Comparison between our results and those of the previously reported showed that known HLB-modulated biological pathways including cell-wall modification, protease-involved protein degradation, carbohydrate metabolism, hormone synthesis and signaling, transcription activities, and stress responses were similarly regulated by HLB infection but different or root-specific changes did exist. The root unique changes included the down-regulation in genes of ubiquitin-dependent protein degradation pathway, secondary metabolisms, cytochrome P450, UDP-glucosyl transferase and pentatricopeptide repeat containing protein genes. Notably, nutrient absorption was impaired by HLB-infection as the expression of the genes involved in Fe, Zn, N and P adsorption and transportation were significantly changed. HLB-infection induced some cellular defense responses but simultaneously reduced the biosynthesis of the three major classes of secondary metabolites, many of which are known to have anti-pathogen activities. Genes involved in callose deposition were up-regulated whereas those involved in callose degradation were also up-regulated, indicating that the sieve tube elements in roots were hanging on the balance of life and death at this stage. In addition, signs of carbohydrate starvation were already eminent in roots at this stage. Other interesting genes and pathways that were changed by HLB-infection were also discussed based on our findings.

overall_design:
Two-year-old seedlings of ‘Sanhu’ red tangerine were grafted with buds from CLas-infected or CLas-free ‘Gonggan’ mandarin trees. Mature leaves and roots were collected from the CLas-inoculated and the control trees every ten days to detect for CLas by PCR. DNA used for PCR was extracted with the use of the Plant DNA isolation Kit (Trans, Beijing, China) according to manufacturer’s instructions. Primers used for CLas detection were the same A2/J5. The fibrous roots of 3 CLas-positive and 3 control CLas-free trees were individually collected at 50 dpi (days post inoculation) when the HLB-inoculated trees became CLas-positive in both leaves and roots yet showed no visible chlorosis and other HLB symptoms. This should have allowed us not to miss too many early responsive genes but at the same time ensured that the trees were infected as expected. Total RNA was extracted from each sample using RNeasy plant mini kit (Qiagen, Valencia, CA) and further purified using the RQ1 Rnase Free Dnase Kit (Promega, Madison, USA). RNA quality and quantity were assessed by gel-electrophoresis and by absorbance at OD260/OD280, respectively. Aliquot RNA samples were stored at -80 ℃. For RNA-seq analysis, RNA samples from the three trees were mixed in equal amount and used for cDNA library construction following the Illumina mRNA-sequencing sample preparation protocol (Illumina, San Diego, CA). The 90-bp raw paired end reads were generated by Illumina HiSeqTM 2000.

Please note that the features (in the processed data, e.g.,clementine0.9_035093m.g) represent the gene IDs in the Citrus clementina genome at ftp://ftp.jgi-psf.org/pub/JGI_data/phytozome/v9.0/Cclementina/. However, the database has been updated (http://genome.jgi.doe.gov/pages/dynamicOrganismDownload.jsf?organism=PhytozomeV10) with new gene IDs, thus the gene IDs used for this study are not trackable anymore. Therefore, the gene annotation files downloadeded from the previous genome database (v0.9) and used for the data analysis are included in this record.

first: Yun

last: Zhong


title: GAGA Factor maintains promoters in nucleosome-free conformation and allows promoter-proximal pausing [MNase]

city: Ithaca

first: Nicholas

last: Fuda

state: New York

address: 417 Biotechnology Building, Cornell University

department: Molecular Biology and Genetics

institute: Cornell University

summary: Promoter-proximal pausing of RNA polymerase II (Pol II) is a widespread in higher eukaryotes. Previous studies have shown that GAF is enriched at paused genes, but the role of GAF in pausing has not been well characterized on a genome-wide level. To investigate the role of GAF in pausing, we RNAi-depleted GAF from Drosophila S2 cells, and examined the effects on promoter-proximal polymerase. We confirmed the importance of GAF for pausing on the classic pause model gene Hsp70. To determine the dependence of pausing on GAF genome-wide, we assayed the levels of transcriptionally-engaged polymerase genome-wide using GRO-seq in control and GAF-RNAi cells. We found that promoter-proximal polymerase was significantly reduced on a subset of paused genes with GAF-bound promoters. There is a dramatic change in nucleosome distribution at genes with reduction in pausing upon GAF depletion and intergenic GAF binding sites in GAF knock-down, suggesting that GAF allows the establishment of pausing at these genes by directing nucleosome displacement off of the promoter. In addition, the insulator factor BEAF, BEAF-interacting protein Chriz, and transcription M1BP enrichment on unaffected genes suggests that redundant transcription factors or insulators protect other GAF-bound paused genes from GAF knock-down effects.

overall_design: Three biological replicates of MNase digested chromatin from LacZ-RNAi and GAGA factor-RNAi cells.

middle: J

first: John

middle: T

last: Lis


title: GAGA Factor maintains promoters in nucleosome-free conformation and allows promoter-proximal pausing

overall_design: Refer to individual Series


title: Molecular anatomy of palate development

status: Public on Apr 03 2015

city: Cincinnati

first: Steven

last: Potter

address: 3333 Burnet Ave.

department: Developmental Biology

institute: Children's hospital Medical Center

summary:
Background: The FACEBASE consortium was established in part to create a central resource for craniofacial researchers. One purpose is to provide a molecular anatomy of craniofacial development. To this end we have used a combination of laser capture microdissection and RNA-Seq to define the gene expression programs driving development of the murine palate.
Results: We focused on the E14.5 palate, soon after medial fusion of the two palatal shelves. The palate was divided into multiple compartments, including medial and lateral, as well as oral and nasal, for both the anterior and posterior domains. A total of 25 RNA-Seq datasets were generated. The results provide a comprehensive view of the region specific expression of all transcription factors, growth factors and receptors. Paracrine interactions can be inferred from flanking compartment growth factor/receptor expression patterns. The results are validated primarily through very high concordance with extensive previously published gene expression data for the developing palate. In addition selected immunostain validations were carried out.
Conclusions: This report provides an RNA-Seq based atlas of gene expression patterns driving palate development at microanatomic resolution. This FACEBASE resource is designed to fuel discovery by the craniofacial research community.

overall_design: Laser capture microdissection and RNA-seq were used to generate gene expression profiles of different compartments of the mouse E14.5 developing palate

first: Andrew

middle: S

first: S


title: Developmental changes in transcriptomes of two key transport tissues in maize kernels: The Maternal Transfer Zone (MATZ) and Basal Endosperm Transfer Layer (BETL)

city: Gainesville

first: Karen

middle: E

last: Koch

state: Florida

department: Plant Molecular Cellular Biology

institute: University of Florida

summary: Transcriptome profiles of MATZ and BETL tissues are compared across three stages of development. Sugars and other nutrients unloaded from vascular tissues in the MATZ are imported by the BETL for utilization by the developing endosperm.  Pronounced changes in gene expression occur in both tissues during kernel development.

overall_design: RNAseq data were obtained from duplicate tissue samples isolated by cryomicrodissection of developing maize kernels at three developmental time points; 8 days post-pollination (DAP), 14 DAP and 20 DAP.

first: Byung

last: Kang


title: Transcriptional profiling of Eucalyptus grandis challenged with Chrysoporthe austroafricana

city: Pretoria

first: Ronishree

last: Mangwanda

address: Lynwood Road

country: South Africa

institute: University of Pretoria

summary:
Background: Eucalyptus species and interspecific hybrids exhibit valuable growth and wood properties that make them a highly desirable commodity. However, these trees are challenged by a wide array of biotic stresses during their lifetimes. The Eucalyptus grandis reference genome sequence provides a resource to study pest and pathogen defence mechanisms in long-lived woody plants. E. grandis trees are generally susceptible to Chrysoporthe austroafricana, a causal agent of stem cankers on eucalypts. The aim of this study was to characterize the defence response of E. grandis against C. austroafricana.

Results: Hormone profiling of susceptible and moderately resistant clonal E. grandis genotypes indicated a reduction in salicylic acid and gibberellic acid levels at 3 days post inoculation. We hypothesized that these signaling pathways may facilitate resistance. To further investigate other defence mechanisms at this time point, transcriptome profiling was performed. This revealed that cell wall modifications and response to oxidative stress form part of the defence responses common to both genotypes, whilst changes in the hormone signaling pathways may contribute to resistance. Additionally the expression of selected candidate defence response genes was induced earlier in moderately resistant trees than in susceptible trees, supporting the hypothesis that a delayed defence response may occur in the susceptible interaction.

Conclusion: The ability of a host to fine-tune its defence responses is crucial and the responses identified in this study extends our understanding of plant defence, gained from model systems, to woody perennials.

overall_design: E. grandis trees were stem inoculated with C. austroafricana. RNA was extracted from stem material harvested 3 days post inoculation for transcriptome profiling. Three biological replicates of harvested material was sent for RNA-sequencing

first: Sanushka

last: Naidoo


title: Genome-wide analysis of the prmt5 and prmt4a;4b mutant transcriptomes

city: Buenos Aires

first: Marcelo

last: Yanovsky

state: Ciudad Autonoma de Buenos Aires

address: Av. Patricias Argentinas 435

country: Argentina

institute: Fundación Instituto Leloir. IIBBA- CONICET

summary: The goal of this study was to compare the transcriptional profile (RNA-seq) of prmt5 and prmt4a;4b Arabidopsis mutants with their respective wild type plants grown under continous light conditions

overall_design: WT, prmt5 and prmt4a;4b mutant plants were grown together with their respective wild type plants for two weeks under continuous white light at 22 degrees centigrades and the transcriptional profile of these plants was analyzed using RNA-seq.


title: The transcriptional cofactor TRIM33 prevents apoptosis in B lymphoblastic leukemia by deactivating a single enhancer [RNA-Seq]

first: Jae Seok

last: Roe

overall_design: RNA-Seq for shRNAs targeting Trim33 expressing B-ALL

first: Jae

first: Christopher

last: Vakoc


title: FBXL10 and the Protection of Polycomb-bound Genes from Hypermethylation (RRBS)

last: Edwards

summary: Nearly all CpG-dense promoters are occupied by the multi-domain chromosomal protein FBXL10. We show here that complete inactivation of the Fbxl10 gene leads to dense de novo methylation only of the promoters that are co-occupied by both FBXL10 and by Polycomb Repressive Complexes; this results in pervasive defects in embryonic development and death of homozygous Fbxl10 mutant embryos at midgestation. Deletion of key components of Polycomb Repressive Complexes 1 and 2 did not lead to ectopic de novo methylation. These results indicate that FBXL10 defends Polycomb-occupied promoters against ectopic de novo methylation. FBXL10 is the first reported factor whose loss leads to a gain in genomic DNA methylation.

overall_design: DNA methylation analysis using RRBS and expression analysis using RNA-seq was performed on WT and Fbxl10T/T ES cells.

first: Mathieu

last: Boulard

first: Timothy

last: Bestor


title: Pol II chromatin immunoprecipitation in S. cerevisiae

status: Public on Apr 02 2015

city: Oxford

first: Michal

last: Gdula

address: South Parks Road

institute: Oxford University

summary: In Saccharomyces cerevisiae short non-coding RNA (ncRNA) generated by RNA Polymerase II (Pol II) are terminated by the NRD complex consisting of Nrd1, Nab3 and Sen1. We now show that Pcf11, a component of the cleavage and polyadenylation complex (CPAC), is generally required for NRD-dependent transcription termination through the action of its CTD interacting domain (CID). Pcf11 localizes downstream of Nrd1 on NRD terminators, and its recruitment depends on Nrd1. Furthermore mutation of the Pcf11 CID results in Nrd1 retention on chromatin, delayed degradation of ncRNA and restricts Pol II CTD Ser2 phosphorylation and Sen1-Pol II interaction. Finally, the pcf11-13 and sen1-1 mutant phenotypes are very similar as both accumulate RNA:DNA hybrids and display Pol II pausing downstream of NRD terminators. We predict a mechanism whereby Nrd1 and Pcf11 exchange on chromatin facilitates Pol II pausing and CTD Ser2-P phosphorylation. This in turn promotes Sen1 activity that is required for NRD-dependent transcription termination in vivo.

overall_design: ChIP-seq with antibody against pol II in wild type and Pcf11 mutants: Pcf11-2, Pcf11-9 and Pcf11-13 grown at 25C and 37C along with input samples

first: Pawel

last: Grzechnik

first: Nick

last: Proudfoot


title: In vivo targeting of de novo DNA methylation by histone modifications in yeast and mouse (ChIP-Seq mouse)

city: los angeles

first: matteo

last: pellegrini

address: 610 charles young drive east

institute: UCLA

summary: Methylation of cytosines (5meC) is a widespread heritable DNA modification. During mammalian development, two global demethylation events are followed by waves of de novo DNA methylation. In vivo mechanisms of DNA methylation establishment are largely uncharacterized. Here we use Saccharomyces cerevisiae as a system lacking DNA methylation to define the chromatin features influencing the activity of the murine DNMT3B. Our data demonstrate that DNMT3B and H3K4 methylation are mutually exclusive and that DNMT3B is co-localized with H3K36 methylated regions. In support of this observation, DNA methylation analysis in yeast strains without Set1 and Set2 show an increase of relative 5meC levels at the TSS and a decrease in the gene-body, respectively. We extend our observation to the murine male germline, where H3K4me3 is strongly anti-correlated while H3K36me3 correlates with accelerated DNA methylation. These results show the importance of H3K36 methylation for gene-body DNA methylation in vivo.

overall_design: Mouse germ-cells ChIPseq

first: Marco

last: Morselli

first: William

last: Pastor

first: Barbara

last: Montanini

first: Kevin

last: Nee

first: Roberto

last: Ferrari

first: Kai

last: Fu

first: Giancarlo

last: Bonora

first: Liudmilla

last: Rubbi

first: Amander

last: Clark

first: Simone

last: Ottonello

last: Jacobsen

first: Matteo

last: Pellegrini


title: RNA-Seq Samples of siTFE3 in 8988T PDA Cell Line to Investigate Transcriptional Control of the Autophagy-Lysosome System

first: Kenneth

middle: N

last: Ross

address: 450 Brookline Ave., Rm M640

department: Pediatric Oncology

institute: Dana-Farber Cancer Institute

summary: The activation of cellular quality control pathways to maintain metabolic homeostasis and mitigate diverse cellular stresses is emerging as a critical growth and survival mechanism in many cancers. Autophagy, a highly conserved cellular self-degradative process, is a key player in the initiation and maintenance of pancreatic ductal adenocarcinoma (PDA). However, the regulatory circuits that activate autophagy, and how they enable reprogramming of PDA cell metabolism are unknown. We now show that autophagy regulation in PDA occurs as part of a broader program that coordinates activation of lysosome biogenesis, function and nutrient scavenging, through constitutive activation of the MiT/TFE family of bHLH transcription factors. In PDA cells, the MiT/TFE proteins - MITF, TFE3 and TFEB - override a regulatory mechanism that controls their nuclear translocation, resulting in their constitutive activation. By orchestrating the expression of a coherent network of genes that induce high levels of lysosomal catabolic function, the MiT/TFE factors are required for proliferation and tumorigenicity of PDA cells. Importantly, unbiased global metabolite profiling reveals that MiT/TFE-dependent autophagy-lysosomal activation is specifically required to maintain intracellular AA pools in PDA.  This AA flux is part of a program that is essential for metabolic homeostasis and bioenergetics of PDA but not for their non-transformed counterparts. These results identify the MiT/TFE transcription factors as master regulators of the autophagy-lysosomal system in PDA and demonstrate a central role of the autophagosome-lysosome compartment in maintaining tumor cell metabolism through alternative amino acid acquisition and utilization.

overall_design: Examination of mRNA levels in pancreatic ductal adenocarcinoma (PDA) cell line 8988T after treatment with siRNA for control or TFE3

first: Rushika

middle: M

last: Perera

first: Nabeel

last: Bardeesy


title: Epigenome-wide association of liver methylation patterns and complex metabolic traits in mice

city: Los Angeles

first: Luz

last: Orozco

address: 610 Charles Young Drive East

department: Molecular, Cell and Developmental Biology

institute: University of California, Los Angeles

summary: Heritable epigenetic factors can contribute to complex disease etiology. In this study we examine, on a global scale, the contribution of DNA methylation to complex traits that are precursors to heart disease, diabetes and osteoporosis. We profiled DNA methylation patterns in the liver using bisulfite sequencing in 90 mouse inbred strains, genome-wide expression levels, proteomics, metabolomics and sixty-eight clinical traits, and performed epigenome-wide association studies (EWAS). We found associations with numerous clinical traits including bone mineral density, plasma cholesterol, insulin resistance, gene expression, protein and metabolite levels. A large proportion of associations were unique to EWAS and were not identified using GWAS. Methylation levels were regulated by genetics largely in cis, but we also found evidence of trans regulation, and we demonstrate that genetic variation in the methionine synthase reductase gene Mtrr affects methylation of hundreds of CpGs throughout the genome. Our results indicate that natural variation in methylation levels contributes to the etiology of complex clinical traits.

overall_design: Reduced representation bisulfite sequencing in mouse strains using liver genomic DNA

middle: D


title: In vivo targeting of de novo DNA methylation by histone modifications in yeast and mouse (ChIP-Seq yeast)

last: pellegrini

overall_design: Yeast ChIP sequencing

first: Steven


title: In vivo targeting of de novo DNA methylation by histone modifications in yeast and mouse (Mnase-Seq)

first: matteo

overall_design: Nucleosome mapping in yeast

last: Morselli


title: 5-hydroxymethylcytosine dynamics during retinal maturation

city: Munich

first: Arshan

state: Bavaria

department: Pharmacology

institute: LMU Munich

summary: Ten-eleven translocation (Tet) hydroxylases (Tet1-3) oxidize 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC). In neurons increased 5hmC levels within gene bodies correlate positively with gene expression. Here, we studied 5hmC profiles (hMeDIP) during retinal maturation between postnatal week 2 and postnatal week 3 using HiSeq 2000 instrument.

overall_design: Study of retinal 5hmC profile dynamics in 2-week old and 3-week old wild type (WT) mouse.

first: David

last: Eisen

first: Mirko

last: Wagner

first: Silvia

last: Laube

first: Andrea

middle: F

last: Künzel

first: Susanne

last: Koch

first: Jessica

last: Steinbacher

first: Elisabeth

last: Schulze

first: Victoria

last: Splith

first: Nana

last: Mittermeier

first: Markus

last: Müller

first: Martin

last: Biel

first: Thomas

last: Carell

first: Stylianos

last: Michalakis


title: In vivo targeting of de novo DNA methylation by histone modifications in yeast and mouse (Bisulfite-Seq)

overall_design: Collecting Yeast Whole Genome Bisulfite Sequencing Data

first: Marco

first: William

last: Pastor

first: Barbara

last: Montanini

first: Kevin

last: Nee

first: Roberto

last: Ferrari

first: Kai

last: Fu

first: Giancarlo

last: Bonora

first: Liudmilla

last: Rubbi

first: Amander

last: Clark

first: Simone

last: Ottonello

last: Jacobsen

first: Matteo

last: Pellegrini


title: Intestinal microbiota composition modulates choline bioavailability from diet and accumulation of the pro-atherogenic metabolite trimethylamine-N-oxide

city: Madison

first: Federico

last: Rey

state: WI

address: 1550 Linden Dr.

institute: University of Wisconsin-Madison

summary: Choline is a water-soluble nutrient essential for human life. Gut microbial metabolism of choline results in the production of trimethylamine (TMA), which upon absorption by the host is converted in the liver to trimethylamine N-oxide (TMAO). Recent studies revealed that TMAO exacerbates atherosclerosis in mice, and positively correlates with the severity of this disease in human. However, which microbes contribute to TMA production in the human gut; the extent to which host factors, e.g., genotype and diet, affect TMA production and colonization of these microbes; as well as the effects TMA-producing microbes have on bioavailability of dietary choline remain largely unknown. We screened a collection of 78 sequenced human intestinal isolates encompassing the major phyla found in the human gut and identified eight strains capable of producing TMA from choline in vitro. Gnotobiotic mouse studies showed that TMAO accumulates in the serum of animals colonized with TMA-producing species, but not in the serum of animals colonized with intestinal isolates that do not generate TMA from choline in vitro. Remarkably, low levels of colonization of TMA-producing bacteria significantly reduced choline levels available to the host. This effect was more pronounced as the abundance of TMA-producing bacteria increased. Our findings provide a framework for designing strategies aimed at changing the representation or activity of TMA-producing bacteria in the human gut and suggest the TMA producing status of the gut microbiota should be considered when making recommendations about choline intake requirements for humans.

overall_design: Examination of community composition and host (gender and strain)/environmental (diet) factors on TMAO accumulation in gnotobiotic mice.

first: Kymberleigh

last: Romano

first: Eugenio

last: Vivas

first: Daniel

last: Amador-Noguez


title: Immunoglobulin transcript sequence and somatic hypermutation computation from unselected RNA-seq reads in Chronic Lymphocytic Leukemia

city: Columbus

first: James

last: Blachly

state: OH

address: B305 Starling-Loving Hall

institute: The Ohio State University

summary: IGHV mutation status is a well-established prognostic factor in chronic lymphocytic leukemia, and also provides crucial insights into tumor cell biology and function. Currently, determination of IGHV transcript sequence, from which mutation status is calculated, requires a specialized laboratory procedure. RNA sequencing is a method that provides high resolution, high dynamic range transcriptome data that can be used for differential expression, isoform discovery, and variant determination. In this paper, we demonstrate that unselected next-generation RNA sequencing can accurately determine the IGH@ sequence, including the complete sequence of the complementarity-determining region 3 (CDR3), and mutation status of CLL cells, potentially replacing the current method which is a specialized, single-purpose Sanger-sequencing based test.

overall_design: CLL cells were sequenced by mRNA-seq on the Illumina platform then subjected to the costom bioinformatic pipeline Ig-ID which yields IGH data


title: BRG1 recruitment by transcription factors MITF and SOX10 defines a specific configuration of regulatory elements in the melanocyte lineage

city: Illkirch

first: Patrick

last: Laurette

address: 1 Rue Laurent Fries

institute: IGBMC

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

overall_design: Refer to individual Series


title: The DREAM complex promotes gene body deposition of H2A.Z for target repression

city: Cambridge

first: Przemyslaw

middle: Aleksander

last: Stempor

state: United Kingdom

address: Tennis Court Road

department: The Gurdon Institute

institute: University of Cambridge

summary: As part of a study of the function of the C. elegans DREAM complex, here we have used ChIP-seq to analyze the patterns of histone variant HTZ-1/H2A.Z and various histone modifications in lin-35(n745) mutants. In addition, we have used RNA-seq to analyze gene expression in lin-35 and htz-1 mutants as compared with wild-type L3 larvae.

overall_design: EXPERIMENT TYPE: CHIP-seq, RNA-seq. BIOLOGICAL SOURCE: Strain: N2, JA1507, VC2490; Developmental Stage: L3 Larva; Genotype: wild type, lin-35(n745), htz-1(ok3099); Sex: mixed Male and Hermaphrodite population; EXPERIMENTAL FACTORS: Developmental Stage L3 Larva; temp (temperature) 20 degree celsius; Antibodies: HTZ-1 JA00001, H3K4me3 Wako 305-34819, H4ac4 Active Motif 39177, Histone H3 Abcam ab1791

first: Julie

last: Ahringer

first: Michael

last: Chesney

first: Isabel

last: Latorre

first: Alex

last: Appert

first: Paulina

last: Kolasinska-Zwierz


title: BRG1 recruitment by transcription factors MITF and SOX10 defines a specific configuration of regulatory elements in the melanocyte lineage (RNA-seq)

summary: Microphthalmia-associated transcription factor (MITF) is the master regulator of the melanocyte lineage. By tandem affinity purification and mass spectrometry, we present a comprehensive characterisation of the MITF interactome comprising multiple novel cofactors involved in transcription, DNA replication and repair and chromatin organisation, including a BRG1 chromatin remodelling complex comprising CHD7. BRG1 is essential for melanoma cell proliferation in vitro and for normal melanocyte development in vivo. MITF and SOX10 actively recruit BRG1 to a set of MITF-associated regulatory elements (MAREs) at active enhancers. MITF, SOX10 and YY1 bind between two BRG1-occupied nucleosomes thus defining both a combinatorial signature of transcription factors essential for the melanocyte lineage and a specific chromatin organisation of MAREs. Nevertheless, BRG1 silencing enhances MITF occupancy at MAREs showing that BRG1 acts to promote dynamic MITF interactions with chromatin.

overall_design: 19 samples corresponding to mRNA profiles of 501Mel and Hermes3A after MITF, BRG1 or control shRNA-mediated knockdown were generated by deep sequencing in triplicate (in duplicate for 501_shMITF and corresponding control 501_shSCR2), using HiSeq2500.

first: Irwin

last: Davidson


title: Pancreatic Cancer Patient Survival Correlates with DNA Methylation of Pancreas Development Genes

middle: James

last: Thompson

state: California

department: Department of Molecular, Cell and Developmental Biology

summary: DNA methylation is an epigenetic mark associated with regulation of transcription and genome structure.  These markers have been investigated in a variety of cancer settings for their utility in differentiating normal tissue from tumor tissue.  Here, we examine the direct correlation between DNA methylation and patient survival.  We find that changes in the DNA methylation of key pancreatic developmental genes is strongly associated with patient survival.

overall_design: 16 total pancreatic samples including 11 tumors, 2 normal, and 3 pancreatitis were sequenced using Reduced Representation Bisulfite Sequencing in order to measure the methylation status of CpG sites.  These data were then examined for correlation with donor survival time.


title: In vivo targeting of de novo DNA methylation by histone modifications in yeast and mouse

first: matteo

last: pellegrini

sample_organism:
Saccharomyces cerevisiae
Mus musculus

platform_organism:
Saccharomyces cerevisiae
Mus musculus


title: In vivo targeting of de novo DNA methylation by histone modifications in yeast and mouse (RNA-Seq)

overall_design: Yeast mRNA sequencing

first: Marco

last: Morselli

first: William

last: Pastor

first: Barbara

last: Montanini

first: Kevin

last: Nee

first: Roberto

last: Ferrari

first: Kai

last: Fu

first: Giancarlo

last: Bonora

first: Liudmilla

last: Rubbi

first: Amander

last: Clark

last: Ottonello

first: Steven

last: Jacobsen


title: mRNA-Seq profiling of Stra8-deficient and wild-type testes

first: David

middle: C

last: Page

state: MA

address: 9 Cambridge Center

institute: Whitehead Institute

summary: We sequenced mRNA in grossly enlarged testes from 1-year-old Stra8-deficient mice, and in testes from adult male wild-type controls, to verify that Stra8-deficient testes are enriched for genes normally expressed in type A spermatogonia.

overall_design: Examination of mRNA levels in 6 whole-testis samples (3 replicates of each genotype). Specifically, we sequenced mRNA in grossly enlarged testes from three 1-year-old Stra8-deficient male mice, and in normal testes from three adult male wild-type controls

first: Ericka

middle: L

last: Anderson

first: Katherine

last: Romer


title: Jarid2 regulates hematopoietic stem cell function by acting with Polycomb Repressive Complex 2

city: Parkville

first: Marnie

last: Blewitt

state: VIC

address: 1G Royal Parade

department: Molecular Medicine Division

country: Australia

institute: WEHI

summary: Polycomb Repressive Complex 2 (PRC2) has been shown to play a key role in hematopoietic stem and progenitor cell (HSPC) function. Analyses of mouse mutants harboring deletions of core components have implicated PRC2 in fine-tuning multiple pathways that instruct HSPC behavior, yet how PRC2 is targeted to specific genomic loci within HSPCs remains unknown. Here we use shRNA-mediated knockdown to survey the function of known PRC2 accessory factors in HSPCs by testing the competitive reconstitution capacity of transduced murine fetal liver cells. We find that similar to the phenotype observed upon depletion of core subunit Suz12, depleting Jarid2 enhances the competitive transplantation capacity of both fetal and adult, mouse and human HSPCs. Gene expression profiling revealed common Suz12 and Jarid2 target genes that are enriched for the H3K27me3 mark established by PRC2. These data implicate Jarid2 as an important component of PRC2 that has a central role in coordinating HSPC function.

overall_design: RNA-seq of jarid knockdown, suz knockdown and control from HSPC in 16 week old mice.

first: Sarah

last: Kinkel


title: GAGA Factor maintains promoters in nucleosome-free conformation and allows promoter-proximal pausing [GRO-seq]

first: Nicholas

last: Fuda

summary: Promoter-proximal pausing of RNA polymerase II (Pol II) is a widespread in higher eukaryotes. Previous studies have shown that GAF is enriched at paused genes, but the role of GAF in pausing has not been well characterized on a genome-wide level. To investigate the role of GAF in pausing, we RNAi-depleted GAF from Drosophila S2 cells, and examined the effects on promoter-proximal polymerase. We confirmed the importance of GAF for pausing on the classic pause model gene Hsp70. To determine the dependence of pausing on GAF genome-wide, we assayed the levels of transcriptionally-engaged polymerase genome-wide using GRO-seq in control and GAF-RNAi cells. We found that promoter-proximal polymerase was significantly reduced on a subset of paused genes with GAF-bound promoters. There is a dramatic change in nucleosome distribution at genes with reduction in pausing upon GAF depletion and intergenic GAF binding sites in GAF knock-down, suggesting that GAF allows the establishment of pausing at these genes by directing nucleosome displacement off of the promoter. In addition, the insulator factor BEAF, BEAF-interacting protein Chriz, and transcription M1BP enrichment on unaffected genes suggests that redundant transcription factors or insulators protect other GAF-bound paused genes from GAF knock-down effects.

overall_design: Two biological replicates of nascent RNA sequencing using GROseq to map transcriptionally-engaged RNA polymerases in untreated, LacZ-RNAi (control), and GAGA factor-RNAi.

first: John

last: Lis


title: High throughput sequencing of B.henselae infected and non-infected I. ricinus salivary glands

city: Maisons Alfort

middle: Irene

last: Bonnet

address: 23 avenue du general de Gaulle

department: Animal Health

institute: INRA

summary: In order to select genes that are differentially expressed in salivary glands during Ixodes ricinus infection by Bartonella henselae we compare the transcriptome of infected and non-infected salivary glands

overall_design: I. ricinus from a pathogen free colony were infected -or not- with B. henselae at the larval and then the nymphal stage and salivary glands of the resulting females adult were dissected for RNA extraction. 454 sequencing was performed on pooled conditions in order to obtain a reference databank. Infected and non-infected samples were then sequenced separatly with illumina, blaston teh reference datbank and compared.

middle: I

first: Xiang

middle: Y

last: Liu


title: BCLAF1 and its pre-mRNA splicing regulator SRSF10 both regulate tumorigenic capacity of human colon cancer cells

city: Tianjin

first: Xuexia

last: Zhou

address: 154 Anshan Road

department: Tianjin Neurological Institute

institute: Tianjin Medical University General Hospital

summary: Bcl-2-accociated transcription factor 1(BCLAF1) has been shown to be involved in multiple biological processes. Transcript variants encoding different isoforms that are generated by alternative splicing have been found for this gene, but little is known about the mechanisms governing its splicing regulation and whether the misregulation is associated with cancer development. Mechanistic analysis revealed that splicing factor SRSF10 specifically interacts with exon5a and activates its inclusion, as RNAi-mediated knockdown of SRSF10 induced a dramatic skipping of exon5a. To define a comprehensive programm of alternative splicing that is regulated by SRSF10 in RKO cells, we used RNA-seq coupled with a bioinformatic analysis to identify the extensive splicing network regulated by SRSF10 in RKO cells.

overall_design: RNA-seq for control (si-NC) and SRSF10-knockdown (si-SRSF10) human RKO cells

first: Wenwu

last: Wu

first: Ying

last: Feng


title: A genome-wide CRISPR library for high-throughput genetic screening in Drosophila cells

status: Public on Apr 01 2015

first: Ji-Long

department: MRC Functional Genomics Unit, Department of Physiology, Anatomy and Genetics

institute: University of Oxford

summary: Analysis of sgRNA sequences in the raw library and at different time points after transfection into S2R+ cells

sample_organism:
Drosophila melanogaster
synthetic construct

overall_design: Two biological repeats of all samples at three timepoints post transfection, with three dilutions of library transfected.

first: Ji

first: Lesheng

last: Kong

first: Andrew

last: Bassett


title: Transcriptomics profiling of Alzheimer’s disease reveal novel molecular targets

city: Miami

first: Marco

last: Magistri

state: FL

address: 1501 NW 10th Ave

department: Psychiatry

institute: University of Miami

summary: Our data provide a comprehensive list of transcriptomics alterations and warrant holistic approach including both coding and non-coding RNAs in functional studies aimed to understand the pathophysiology of LOAD

overall_design: We performed directional RNA sequencing on high quality RNA samples extracted from hippocampi of 4 late onset Alzheimer's diseas (LOAD) and 4 age-matched controls.

first: Mohammad

middle: A

last: Faghihi

first: Dmitry

last: Velmeshev

first: Madina

last: Makhmutova


title: Genome-wide maps of AR recruitment in primary human ovarian granulosa cells (GCs) transduced with androgen receptor (AR) alternative splice variant (ASV)

city: Hangzhou

first: Fangfang

last: Wang

address: No.1 Xueshi Road.

institute: Zhejiang University

summary: We report the genome-wide AR recruitment patterns in pimary human GCs.

overall_design: Examination of HA binding signal in human GCs transducted with HA tagged AR ASVs.

first: Hefeng

last: Huang


title: Next Generation Sequencing Analysis of Wild Type and SRSF10-/- embryonic day 13.5 heart Transcriptomes

city: Shanghai

first: Ning

last: Wei

address: 320 Yueyang RD.

institute: Institute for Nutritional Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences

summary: We generated the SRSF10 -/- mice, and collected their embryonic heart at 13.5 and controls. Then, we extracted RNAs of embryonic heart and performed next generation sequencing. By comparing sequcing data from WT and SRSF10 -/- samples, we profiled the alternative splicing events and gene expression regulated by SRSF10 during mouse heart development process.

overall_design: E13.5 embryonic heart mRNA profiles of wild type (WT) and SRSF10-/- mice were generated by deep sequencing, using Illumina HiSeq2000.


title: BRG1 recruitment by transcription factors MITF and SOX10 defines a specific configuration of regulatory elements in the melanocyte lineage (ChIP-seq)

first: Patrick

last: Laurette

overall_design: 8 samples corresponding to genomic occupancy profiling of MITF, SOX10, BRG1 after si-control, BRG1 after si-MITF and BRG1 after siSOX10; and their respective controls (MITF Input, SOX10 Input, GFP-siCTRL ChIPseq) in 501Mel cells.

first: Irwin

last: Davidson


title: Analysis of bone marrow derived mast cell transcriptomes in presence or absence of SialostatinL

city: Mainz

first: Tobias

last: Bopp

address: Langenbeckstrasse 1

department: Institute for Immunology

institute: University Medical Center Mainz

summary: Goal of this study was to compare transcriptional changes in stimulated mast cells in the absence or presence of sialostatinL

overall_design: mRNA profiles of 4 weeks old mast cells (BMMC derived from C57BL/6 mice ) stimulated for 24h with ionomycin in absence or presence of tick derived sialostatinL were generated by deep sequencing using Illumina HiSeq2000

first: Matthias

last: Klein


title: Transcriptome-wide analysis of gene expression in 4-day-old WT and athb1-1 mutant seedlings grown under short day conditions

city: Santa Fe

first: Raquel

middle: Lia

last: Chan

state: Santa Fe

address: Centro Científico Tecnológico CONICET Santa Fe, Colectora Ruta Nac. Nº 168 km. 0, Paraje El Pozo

institute: Instituto de Agrobiotecnología del Litoral (CONICET/UNL)

summary: To understand which genes acts downstream AtHB1 affecting hypocotyl growth in Arabidopsis thaliana, we performed transcriptional profiles of 4-day-old seedlings grown in a short-day regime comparing wild-type with athb1-1 mutant plants. These results show that some of the AtHB1-regulated genes modulate cell elongation, particularly cell wall composition and elongation, or encode proteins that serve as a source of carbon, nitrogen, and sulfur for early seedling growth.

overall_design: RNA-Seq data for 4-day-old wild-type (Col-0) and athb1-1 mutant seedlings grown under short-day conditions. Biological triplicates were performed for each genotype analyzed.


title: CHARACTERIZATION AND DYNAMICS OF MOUSE PERICENTROMERE-ASSOCIATED DOMAINS

city: Utrecht

first: Geert

last: Geeven

state: Utrecht

address: Uppsalalaan 8

department: Biomedical Genomics

country: Netherlands

institute: Hubrecht Institute

summary: Despite recent progress in genome topology knowledge, the role of repeats, comprising the majority of mammalian genomes, remains elusive. Satellites are highly abundant sequences that cluster around centromeres, attract pericentromeric heterochromatin and aggregate into nuclear chromocenters that are assumed to form a repressive compartment in the nucleus to which genes are recruited for silencing. Here we designed a strategy for genome-wide identification of pericentromere-associated domains (PADs) in different mouse cell types. The ~1000 PADs and non-PADs have similar chromatin states in embryonic stem cells. During lineage commitment however chromocenters progressively overlap with constitutively inactive genomic regions at the nuclear periphery. This suggests that chromocenters do not actively recruit PADs but are themselves attracted to inactive chromatin compartments. However, we also found that experimentally induced proximity of an active locus to chromocenters was sufficient to cause gene repression. Collectively, our data suggests that rather than being a driver of nuclear organization, pericentromeric satellite repeats mostly co-segregate with inactive genomic regions to nuclear compartments where they can contribute to stably maintaining the repressed status of proximal chromosomal regions.

overall_design: We performed satellite 4C (sat4C) on mouse primary thymus tissue, pluripotent mouse embryonic stem cells (ESC) and neural precursor cells (NPC) and terminally differentiated astrocytes (AC) that were sequentially derived from these ESC in vitro

middle: J

last: Wijchers

first: Wouter

last: de Laat