title: Genomic targets sites of the Candida glabrata transcription factor Pdr1

status: Public on Sep 12 2014

city: Iowa City

first: Thomas

last: Bair

state: IA

address: 335 EMRB

institute: University of Iowa

summary: Pdr1 is the major regulator of azole resistance in the fungal pathogen Candida glabrata.  Earlier experiments demonstrated that expression of Pdr1 itself is increased when cells lose their mitochondrial genome (rho0).  Here we use chromatin immunoprecipitation coupled with highthroughput sequencing (ChIP-seq) to map the genomic binding sites for Pdr1 in both normal and rho0 cells.  These data provide the first look at genes that are likely to represent the direct targets of Pdr1 in this important pathogen.

overall_design: Examination of Pdr1 binding sites using two different antibodies in five different strains. Strains labeled TAP carry the TAP-tagged allele and capture was done with anti-TAP. The control was performed by carrying out a ChIP reaction on rho0 TAP-PDR1 cells but with no primary anti-TAP antibody.  Since Pdr1 expression is the highest in this background, this was selected as the best control for antibody specificity.  We also performed ChIP reactions on rho+, rho0 and cells lacking the PDR1 gene (pdr1D) cells using the anti-Pdr1 antibody.  The control for these reactions was a ChIP experiment using anti-Pdr1 antibody on cells lacking the endogenous Pdr1 protein.

first: Sanjoy

last: Paul

first: W

middle: S

last: Moye-Rowley


title: Calorie Restricted and Non-Restricted Yeast

status: Public on Sep 15 2014

city: Philadelphia

first: Shelley

middle: L

last: Berger

state: PA

address: 3400 Civic Center Blvd, Bldg 421

department: Departments of Cell & Developmental Biology; Biology; Genetics

country: USA

institute: University of Pennsylvania

type: Genome binding/occupancy profiling by high throughput sequencing

type: Expression profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: MNase-seq Experiments from Saccharomyces cerevisiae

summary: MNase-seq Experiments from Calorie Restricted and Non-Restricted Yeast from WT, ISW2DEL and ISW2K215R strains

overall_design: We used MNase-seq to study genome-wide nucleosome positions under Calorie Restricted and Non-restricted Saccharomyces cerevisiae

first: Weiwei

last: Dang

first: Gabriel

last: Otte

first: Jean

middle: A

last: Dorsey


title: Transcriptome and transposable element dynamics  during PIWI regulation in Drosophila ovarian cell cultures

city: BOSTON

first: NELSON

middle: C

last: LAU

state: MA

address: 72 E Concord St K219, Dept of Biochemistry

department: Biochemistry and Cell Biology

institute: Boston University Chobanian & Avedisian School of Medicine

type: Non-coding RNA profiling by high throughput sequencing

summary: This study examines the regulatory capacity of the Piwi protein in the Drosophila OSS cell. Piwi CLIP-Seq, mRNA-Seq and nascent RNAseq datasets were integrated to determine how Piwi proteins where using piRNAs and binding interactions to regulate the expression of transcripts.  We also sequenced the genomes of various OSS cell lines.

overall_design:
We first performed several replicates of a Piw CLIP-Seq experiment to isolate RNA fragments as CLIP-tags to discover which transcripts are preferentially bound by the Piwi protein.  Then we performed several types of mRNA expression profiling experiments using several forms of mRNA-Seq library construction formats.  Finally, we sequenced the genomes from various OSS cell lines.

The genomic sequencing component of the study is represented by BioProject PRJNA240323. The genomic sequencing raw data have been deposited at SRA (SRP039565).

first: Yuliya

last: Sytnikova

first: Gung-wei

last: Chirn

first: Nelson

last: Lau

first: Reazur

last: Rahman


title: Enhancer chromatin signatures predict Smad2/3 binding in Xenopus

status: Public on Sep 17 2014

city: Stanford

first: Julie

last: Baker

state: CA

address: 300 Pastuer Drive

department: Genetics

institute: Stanford University

summary: In this study we have examine the deposition of H3K4me1,H3K4Me3 and H3K27Ac and the Nodal transcription factor, Smad2/3, immediately following zygotic transcription and continuing through gastrulation.

overall_design: We profiled 4 histone modifications (H3K4Me3, H3K27Me3, H3K27AC, H3K4Me1) and one transcription factor smad2/3 (+ chromatin input) using ChIP-Seq, and expression profiles (3' RNA-Seq) for Xenopus tropicalis embryos stage8, stage9 and stage10.5. Furthermore, we have profile two histone modifications (H3K4Me1 and H3K27Ac) in absance of nodal signaling in stage9 Xenopus tropicalis embryos using ChIP-seq and 3-seq

first: Rakhi

last: Gupta


title: Heritable variation of mRNA decay rates in yeast

status: Public on Sep 16 2014

city: Seattle

first: Joshua

last: Akey

state: WA

address: 3720 15th Ave NE

department: Genome Sciences

institute: University of Washington

summary: Gene expression levels are determined by the balance between rates of mRNA transcription and decay, and genetic variation in either of these processes can result in heritable differences in transcript abundance. Although the genetics of gene expression has been the subject of intense interest, the contribution of heritable variation in mRNA decay rates to gene expression variation has received far less attention. To this end, we developed a novel statistical framework and measured allele-specific differences in mRNA decay rates in a diploid yeast hybrid created by mating two genetically diverse parental strains. In total, we estimate that 31% of genes exhibit allelic differences in mRNA decay rate, of which 350 can be identified at a false discovery rate of 10%. Genes with significant allele-specific differences in mRNA decay rate have higher levels of polymorphism compared to other genes, with all gene regions contributing to allelic differences in mRNA decay rate. Strikingly, we find widespread evidence for compensatory evolution, such that variants influencing transcriptional initiation and decay having opposite effects, suggesting steady-state gene expression levels are subject to pervasive stabilizing selection. Our results demonstrate that heritable differences in mRNA decay rates are widespread, and are an important target for natural selection to maintain or fine-tune steady-state gene expression levels.

overall_design: We measured rates of allele-specific mRNA decay (ASD) in a diploid yeast produced by mating two genetically diverse haploid Saccharomyces cerevisiae strains: the laboratory strain BY4716 (BY), which is isogenic to the reference sequence strain S288C, and the wild Californian vineyard strain RM11-1a (RM). Briefly, we introduced rpb1-1, a temperature sensitive mutation in an RNA polymerase II subunit, to each of the haploid yeast strains, mated the strains, and grew the resulting hybrid diploid to mid-log phase at 24 °C, before rapidly shifting the culture to 37 °C to inhibit transcription. RNA-seq was performed on culture samples taken at 0, 6, 12, 18, 24, and 42 minutes subsequent to the temperature shift. To identify ASD, we used transcribed polymorphisms to distinguish between parental transcripts, and compared the relative levels of transcript abundance over the time course. Note, this experimental design internally controls for trans-acting regulatory variation as well as environmental factors. Under the null hypothesis of no ASD, the proportion of reads from the BY transcript (p_BY = N_BY / (N_BY + N_RM)) observed over the time course remains unchanged. However, genes with ASD will exhibit an increasing or decreasing proportion of BY reads as a function of time. In total, we measured ASD from three independent biological replicates.

first: Jennifer

middle: M

last: Andrie

first: Jon

last: Wakefield


title: Environmental enrichment modulates 5-hydroxymethylcytosine dynamics in hippocampus

status: Public on Sep 11 2014

city: Atlanta

first: Bing

last: Yao

state: GA

address: 615 Michael St. Rm 323

department: Human Genetics

institute: Emory University

type: Methylation profiling by high throughput sequencing

summary: Gene-environment interactions mediated at the epigenetic level may provide an initial step in delivering an appropriate response to environmental changes. 5-hydroxymethylcytosine (5hmC), a DNA base derived from 5-methylcytosine (5mC), accounts for ~40% of modified cytosine in brain and has been implicated in DNA methylation-related plasticity. To identify the role of 5hmC in gene-environment interactions, we exposed both young (6-week-old) and aged (18-month-old) mice to both an enriched environment and a standard environment. Exposure to EE significantly improves learning and memory in aged mice and reduces 5hmC abundance in mouse hippocampus. Furthermore, we mapped the genome-wide distribution of 5hmC and found that the alteration of 5hmC modification occurred mainly at gene bodies. In particular, genes involved in axon guidance are enriched among the genes with altered 5hmC modification. These results together suggest that environmental enrichment could modulate the dynamics of 5hmC in hippocampus, which could potentially contribute to improved learning and memory in aged animals.

overall_design:
To identify the role of 5hmC in gene-environment interactions, we exposed both young (6-week-old) and aged (18-month-old) C57B/6 mice to both an enriched environment (EE) and a standard environment. Exposure of mice to EE was achieved by keeping a group of mice in the EE chamber, which is larger than the standard cage, includes novel objects, such as toys of varied color and texture, tunnels, an exercise wheel for voluntary exercise, and extra bedding material, along with free access to food and water. Daily exposure to EE was kept at 5 hours during the daylight cycle for 4 consecutive weeks. Control animals were kept in their small cages that are used as standard housing cages (CE) containing bedding, food and access to water. YC, young mice exposing to control environment, YE, young mice exposing to enriched environment, AC, aged mice exposing to control environment, AE, aged mice exposed to enriched environment.

Please note that 5hmC-containing DNA enrichment method was inspired by a unique character of beta-glucotransferase that can specifically add glucose to 5hmC modification. With a modified glucose conjugated with biotin, we are able to purify the 5hmC-containing DNA by biotin-streptavidin-based immunoprecipitation.

first: Peng

last: Jin


title: Generation of a Panel of Induced Pluripotent Stem Cells From Chimpanzees: a Resource for Comparative Functional Genomics

city: Chicago

first: Irene

last: Gallego Romero

state: IL

address: 920 E 58th St, CLSC 317

institute: University of Chicago

sample_organism:
Pan troglodytes
Homo sapiens

platform_organism:
Pan troglodytes
Homo sapiens


title: Transcriptome comparison of mouse pancreatic islets cultured at low vs high ambient glucose

city: Davis

first: Mark

middle: O.

last: Huising

address: One Shields Avenue

department: Neurobiology, Physiology and Behavior

institute: University of California, Davis

summary: Islets are known to respond to changes in ambient glucose. To quantify the transcriptome-wide changes in ambient glucose, we compared transcriptome of islets exposed to low and high glucose.

overall_design: Isolated islets from wild type male mice. Islets from adult males were pooled, cultured overnight in RPMI containing 11 mM glucose. The next day, all islets were  starved in RPMI containing 2.8 mM glucose for 2 hours before stimulation with 2.8 mM glucose or 16.8 mM glucose for 12 hours. Islets were lysed in Trizol for RNA isolation and library construction.

middle: O


title: Methylated DNA is over-represented in whole-genome bisulfite sequencing data

city: Athens

first: Robert

middle: J

last: Schmitz

address: B416 Davison Life Sciences

institute: University of Georgia

summary: The development of whole-genome bisulfite sequencing (WGBS) has led to a number of exciting discoveries about how genomes utilize DNA methylation and has led to a plethora of novel testable hypotheses. Methods for constructing sodium bisulfite-converted and amplified libraries have recently excelled to the point that the bottleneck for experiments that use WGBS has shifted to data analysis and interpretation. Here we present empirical evidence for an over-representation of methylated DNA from WGBS. This enrichment for methylated DNA is exacerbated by higher cycles of PCR and is influenced by the type of uracil-insensitive DNA polymerase used for amplifying the sequencing library. Future efforts to computationally correct for this enrichment bias will be essential to increasing the accuracy of determining methylation levels for individual cytosines.

overall_design: MethylC-Seq of Arabidopsis thaliana


title: Ebf2 is a selective marker of brown and beige adipogenic precursor cells (RNA-seq)

city: Cincinnati

first: Hee-Woong

last: Lim

state: Ohio

address: 3333 Burnet Ave. MLC 7024

department: Division of Biomedical Informatics

institute: Cincinnati Children's Hospital Medical Center

summary: Brown adipocytes, muscle and dorsal dermis descend from precursor cells in the dermomyotome, but the factors that regulate commitment to the brown adipose lineage are unknown. Here, we prospectively isolated and determined the molecular profile of embryonic brown preadipose cells. Brown adipogenic precursor activity in embryos was confined to Pdgfrα+, Myf5Cre-lineage-marked cells. RNAseq analysis identified Early B Cell Factor-2 (Ebf2) as one of the most selectively expressed genes in this cell fraction. Importantly, Ebf2-expressing cells purified from Ebf2-GFP embryos or brown fat tissue did not express myoblast or dermal cell markers and uniformly differentiated into brown adipocytes. Interestingly, Ebf2-expressing cells from white fat tissue in adult animals differentiated into brown-like (or beige) adipocytes. Loss of Ebf2 in brown preadipose cells reduced the expression levels of brown preadipose-signature genes, whereas ectopic Ebf2-expression in myoblasts activated brown preadipose-specific genes. Altogether, these results indicate that Ebf2 specifically marks and regulates the molecular profile of brown preadipose cells.

overall_design: Embryonic fibroblasts, isolated from dorsal body wall of E14.5 Myf5-CrE;mTmG embryos, were further fractionated based on the expression of PDGFRα, Itga7 and Myf5-cre (GFP).  Total mRNA profiles from Myf5-cre(GFP)+PDGFRα+ and  Myf5-cre(GFP)+PDGFRα-Itga7+ cells were generated by deep sequenceing

first: Wenshan

last: Wang

first: Megan

last: Kissig

first: Sona

last: Rajakumari

first: Li

last: Huang

first: Kyoung-Jae

last: Won

first: Patrick

last: Seale


title: Transcriptome-wide mapping reveals widespread dynamic regulated pseudouridylation of mRNA

city: Rehovot

first: Schraga

last: Schwartz

state: Choose a State or Province

address: Herzl 234, Department of Molecular Genetics

country: Israel

institute: WEIZMANN INSTITUTE OF SCIENCE

summary: Pseudouridine is the most abundant modification occurring on RNA, yet with the exception of a few well-studied RNA molecules little is known about the modified positions and their function(s). Here, we develop Ψ-seq, a method for transcriptome-wide quantitative mapping of pseudouridine. We validate Ψ-seq with synthetic spike-ins and de novo identification of the vast majority of previously reported pseudouridylated positions. Ψ-seq permits discovery of hundreds of novel pseudouridine modifications in human and yeast mRNAs and snoRNAs. Knockdown and knockout of pseudouridine synthases uncovers the cognate PUSs mediating pseudouridine catalysis at these individual novel sites and their target sequence features. In both human and yeast pseudouridine formation on mRNA depends on both site-specific PUSs – often guided by a specific sequence motif - and snoRNA-guided PUSs. Importantly, upon heat shock in yeast, Pus7-mediated pseudouridylation is induced at >200 sites in diverse mRNAs. Pus7 deletion in yeast leads to decreased recovery from heat shock and decreased RNA levels at otherwise pseudouridylated messages, suggesting a role for pseudouridine in enhancing transcript stability. Pseudouridine stoichiometries in rRNA are highly conserved from yeast to mammals, but are reduced in cells derived from dyskeratosis congenita patients, where the pseudouridine synthase DKC1 is mutated, compared to age matched controls. Our results establish pseudouridine as a ubiquitous and dynamic modification in mRNA, and provide a sensitive, quantitative and transcriptome-wide methodology to address its underlying mechanisms and function.

sample_organism:
Saccharomyces cerevisiae
Candida albicans
Homo sapiens
Mus musculus

overall_design: Examination of m6A methylation in human Hek293 and A549 cell lines, in human embryonic stem cells (ESCs) undergoing differentiation to neural progenitor cells (NPCs),  in OKMS inducible fibroblasts reprogrammed into iPSC, and upon knockdown of factors using siRNAs or shRNAs.

first: Douglas

last: Bernstein

first: Maxwell

last: Mumbach

first: Eric

last: Lander

first: Gerald

last: Fink

first: Aviv

last: Regev


title: Extensive translation of small Open Reading Frames revealed by Poly-Ribo-Seq

city: Brighton

first: Juan Pablo

last: Couso

address: Falmer

country: United Kingdom

institute: University of Sussex

summary: We present a genome-wide assessment of small open reading frames (smORF) translation by ribosomal profiling of polysomal fractions in Drosophila S2 cell. In this way, mRNAs bound by multiple ribosomes and hence actively translated can be isolated and distinguished from mRNAs bound by sporadic, putatively non-productive single ribosomes or ribosomal subunits.

overall_design: Ribosomal profiling of large and small polysomal fractions in Drosophila S2 cells to assess translation of smORFs

first: Julie

last: Aspden

first: Ying

last: Eyre-Walker

first: Juan

middle: P


title: Tissue-Specific RNA Expression Marks Distant Acting Developmental Enhancers

city: Berkeley

first: Han

last: Wu

state: California

address: 1 Cyclotron Road, Mail Stop 84-171

department: Genomics Division

institute: Lawrence Berkeley National Lab

summary: In this study, we explored the in vivo expression dynamics of tissue-specific non-coding RNAs in embryonic mouse tissues via in-depth transcriptome profiling. Overall, approximately 80% of validated in vivo enhancers show tissue-specific RNA expression that correlates with tissue-specific enhancer activity. Globally, we identified thousands of tissue-specifically transcribed non-coding regions (TSTRs) displaying various genomic hallmarks of bona fide enhancers. Together, our results demonstrate that tissue-specific eRNA expression is a common feature of in vivo enhancers, as well as a major source of extragenic transcription, and that eRNA expression signatures can be used to predict tissue-specific enhancers independent of known epigenomic enhancer marks.

overall_design: Examimation of total RNA expression in mouse heart and limb tissues collected at mouse embryonic day 11.5.

first: H

first: A

last: Nord


title: Genome wide transcriptome analysis of skin harboring tissue-resident memory CD8+ T cells

city: Amsterdam

first: silvia

last: ariotti

address: Plesmanlaan 121

country: Netherlands

institute: NKI

summary: Comparison of the transcriptional profiles of full-thickness murine skin harboring tissue resident memory T cells exposed to specific or control trigger

overall_design: Expression profiling by high throughput sequencing

first: Silvia

last: Ariotti

first: Marc

last: Hogenbirk

first: Heinz

last: Jacobs

first: Ton

middle: N

last: Schumacher


title: Active and Passive Demethylation of Male and Female Pronuclear DNA in the Mammalian Zygote

city: Beijing

first: Xianlong

last: Li

address: 5, Yiheyuan Rd., Haidian District

department: Biodynamic Optical Imaging Center, College of Life Sciences

country: China

institute: Peking University

summary: The epigenomes of mammalian sperm and oocytes, characterized by gamete-specific 5-methylcytosine (5mC) patterns, are reprogrammed during early embryogenesis to establish full developmental potential. Previous studies have suggested that the paternal genome is actively demethylated in the zygote while the maternal genome undergoes subsequent passive demethylation via DNA replication during cleavage. Active demethylation is known to depend on 5mC oxidation by Tet dioxygenases and excision of oxidized bases by thymine DNA glycosylase (TDG). Here we show that both maternal and paternal genomes undergo widespread active and passive demethylation in zygotes before the first mitotic division. Passive demethylation was blocked by the replication inhibitor aphidicolin, and active demethylation was abrogated by deletion of Tet3 in both pronuclei. At actively demethylated loci, 5mCs were processed to unmodified cytosines. Surprisingly, the demethylation process was unaffected by the deletion of TDG from the zygote, suggesting the existence of other demethylation mechanisms downstream of Tet3-mediated oxidation.

overall_design: The dataset includes RRBS anlysis of 2 MII oocyte samples, 3 WT female pronuclei samples PN3-4 stage, 2 Tet3 KO female pronuclei samples and 2 Aphidicolin treated female pronuclei samples. Also as male counterpart, a Sperm sample, 2 WT male pronuclei samples PN3-4 stage, 2 Tet3 KO male pronuclei samples and 2 Aphidicolin treated male pronuclei samples were included.

first: Fan

last: Guo

first: Dan

last: Liang

first: Tong

first: Ping

last: Zhu

first: Hongshan

first: Xinglong

first: Lu

last: Wen

first: Tian-Peng

last: Gu

first: Boqiang

last: Hu

first: Colum

last: Walsh

first: Jin-Song

first: Fuchou

last: Tang

first: Guo-Liang

last: Xu


title: A global regulatory mechanism for activating an exon network required for neurogenesis

first: Bushra

last: Raj

address: 421 Curie Boulevard

summary: The vertebrate and neural-specific SR-related protein nSR100/SRRM4 regulates an extensive program of alternative splicing with critical roles in nervous system development. However, the mechanism by which nSR100 controls its target exons is poorly understood. We demonstrate that nSR100-dependent neural exons are associated with a unique configuration of intronic cis-elements that promote rapid switch-like regulation during neurogenesis. A key feature of this configuration is the insertion of specialized intronic enhancers between polypyrimidine tracts and acceptor sites that bind nSR100 to potently activate exon inclusion in neural cells, while weakening 3' splice site recognition and contributing to exon skipping in non-neural cells. nSR100 further operates by forming multiple interactions with early spliceosome components bound proximal to 3' splice sites. These multifaceted interactions achieve dominance over neural exon silencing mediated by the splicing regulator PTBP1. The results thus illuminate a widespread mechanism by which a critical neural exon network is activated during neurogenesis.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: RNA-Seq was used to obtain mRNA profiles of various N2A and 293T cell lines from human and mouse, respectively, to investigate the roles of nSR100, Ptbp1 and U2af65 in alternative splicing regulation. PAR-iCLIP and iCLIP experiments followed by high throughput sequencing were conducted to obtain RNA binding profiles of nSR100, PTBP1 and U2af65.

first: Manuel

last: Irimia

first: Ulrich

last: Braunschweig

first: Timothy

last: Sterne-Weiler

first: Dave

last: O'Hanlon

first: Zhen-Yuan

last: Lin

first: Ginny

middle: I

last: Chen

first: Laura

last: Easton

first: Jernej

last: Ule

first: Anne-Claude

last: Gingras

first: Eduardo

last: Eyras

first: Benjamin

last: Blencowe


title: RNA-sequencing analysis of NGN3-GFP positive and negative cells purified from differentiating human embryonic stem cells

city: Shenzhen

first: ilmen

last: zhang

state: Guangdong Province

address: Beishan Industrial Zone,Yantian District

institute: BGI

summary: NGN3-eGFP could be used to mark endocrine progenitor cells and their progeny in differentiating hESCs, suggesting that profiling analysis of NGN3-eGFP+ cells could lead to the discovery of novel gene participation. Six batches of NGN3-eGFP+ and NGN3-eGFP -cells purified by FACS from a reporter hES cell line cell cultures at stage 4 were pooled together, respectively. RNA-seq-based analysis of the two cell fractions identified a total of 3976 differentially expressed genes (P-value<0.05; Fold >2), of which 72% are enriched in NGN3-GFP+ cells. As expected, all the genes expressed in pancreatic progenitor cells are enriched in NGN3-eGFP-cells; while all endocrine-associated genes are enriched in NGN3-eGFP+ cells. Among the 3976 genes, 195 have potential transcriptional factor activity and 453 are potential non-coding RNA genes, most of which have not been well investigated in pancreatic endocrine cells. The gene ontology term ‘integral to membrane’ GO: 0016021 identified 1003 differentially expressed genes that encoded ion channels, adhesion molecules and transporters.

overall_design: NGN3-eGFP Postive and negtive cells were purified at stage 4 and subjected to RNA-sequncing

first: Haisong

last: Liu

first: Long

first: Xi

last: Zhang

first: Jian

last: Wang

first: Hongkui

last: Deng


title: RNase ZS1-mediated UbL40 mRNA decay controls thermo-sensitive genic male sterility in rice.

first: xiaofeng

last: cao

address: West Lincui Road, Chaoyang District

department: State Key Laboratory of Plant Genomics

institute: Institute of Genetics and Developmental Biology

summary: Male sterility is an important trait in hybrid crop breeding. Thermo-sensitive genic male sterility (TGMS) lines, which are male-sterile at restrictive (high) temperatures but convert to male-fertile at permissive (low) temperatures, have been widely utilized in two-line hybrid rice breeding3. However, the molecular mechanism underlying TGMS remains unclear. Here we show that the rice (Oryza sativa L.) thermo-sensitive genic male sterile gene 5 (tms5) locus, which in 2010 was present in cultivars occupying more than 70% (2.4 million hectares) of two-line hybrid rice-growing land in China, confers the TGMS trait through a loss-of-function mutation of RNase ZS1, resulting in failure to mediate mRNA decay of three temperature-responsive ubiquitin fusion ribosomal protein L40 genes (UbL40). TMS5 encodes an evolutionarily conserved endonuclease, RNase ZS1. RNase ZS1 can process tRNAs in vitro, but does not do so in vivo due to its localization in the cytoplasm. Defective RNase ZS1 in tms5 plants leads to over-accumulation of UbL401, UbL402 and UbL404 mRNAs at restrictive but not permissive temperatures. Furthermore, over-expression of UbL401 and UbL404 in wild-type plants causes male sterility, while knockdown of UbL401 and UbL404 in tms5 partially restores its fertility. Our results uncover a novel mechanism of RNase ZS1-mediated UbL40 mRNA decay which controls TGMS in rice and has potential applications in hybrid breeding not only of rice but also of other crops.

overall_design: Examination of differences in mRNA accumulation between 93-11, NIL5 and NIL8.

first: Xiaofeng

last: Cao

first: Ming

last: Zhou

first: Lianfeng

last: Gu

first: Yuanzhu

last: Yang

first: Mingji

last: Feng


title: DNA methylation profiles of isolated embryonic and adult cardiomyocytes, cardiac fibroblasts, tissue and stem cells.

status: Public on Sep 10 2014

city: Tokyo

first: Mayumi

last: Oda

address: 35 Shinanomachi, Shinjuku

department: School of Medicine

country: Japan

institute: Keio University

summary: To investigate the DNA methylation profiles of matured differentiated cardiomyocytes, DNA methylation profiles were obtained by HELP tagging assay using the isolated cardiomyocytes and cardiac fibroblasts from different developmental stages, tissues and cultured stem cells.

overall_design: DNA methylation in mouse tissues and cells

first: Shinji

last: Makino


title: Global loss of DNA methylation uncovers intronic enhancers in genes

city: Los Angeles

first: Peggy

last: Farnham

address: 1450 Biggy Street NRT 6503

department: GMCB

institute: University of Southern California Graduate Student

summary: We used HCT116 colorectal cancer cells with and without mutations in DNA methyltransferases (resulting in a 95% reduction in global DNA methylation levels) to study the relationship between DNA methylation, histone modifications, and gene expression. (The double knockout cell line is called DKO1)

overall_design:
Examination of DNA methylation, two histone modifications, RNA polymerase II ChIP-seq and RNA expression in two cell line (HCT116 and DKO1).
One of relipates of HCT116 RNA P II ChIP-seq is from GSM970210. Histone modification data(H3K27ac, H3K4me3) of HCT116 are from GSM945304, GSE31755. Two replicates of RNA-seq data in HCT116 are from GSM1266733 and GSM1266734

first: Adam

last: Blattler

first: Heather

last: Witt

first: Yu

last: Guo

first: Charles

last: Nicolet

first: Fides

last: Lay

first: Peter

last: Jones


title: Resetting Transcription Factor Control Circuitry Towards Ground State Pluripotency In Human

city: Cambridge

first: Felix

last: Krueger

address: Granta Park

department: Bioinformatics

institute: Altos Labs

summary: Current human pluripotent stem cells lack the transcription factor circuitry that governs the ground state of mouse embryonic stem cells (ESC). Here we report that short-term expression of two components, NANOG and KLF2, is sufficient to ignite other elements of the network and reset the human pluripotent state. Inhibition of ERK and protein kinase C signalling sustains a transgene-independent rewired state. Reset cells self-renew continuously without ERK signalling, are phenotypically stable and karyotypically intact. They differentiate in vitro and form teratomas in vivo. Metabolism is reprogrammed in reset cells with activation of mitochondrial respiration as in ESC. DNA methylation is dramatically reduced and transcriptome state is globally realigned across multiple cell lines. Depletion of ground state transcription factors, TFCP2L1 or KLF4 has marginal impact on conventional human pluripotent stem cells, but collapses the reset state. These findings demonstrate feasibility of installing and propagating functional control circuitry for ground state pluripotency in human cells.

overall_design: DNA methylation analysis in Conventional and Reset human embryonic stem cells by whole genome bisulfite sequencing, in triplicate, using the Illumina platform

first: Yasuhiro

last: Takashima

first: Gabriella

last: Ficz

first: Wolf

last: Reik

first: Austin

last: Smith


title: Tranposable elements modulate human mRNAs and lncRNAs via specific RNA-protein interactions.

first: David

middle: R

last: Kelley

address: 7 Divinity Ave.

institute: Harvard University

summary: Transposable elements (TEs) have significantly influenced the evolution of transcriptional regulatory networks in the human genome. Post-transcriptional regulation of human genes by TE-derived sequences has been observed in specific contexts, but has yet to be systematically and comprehensively investigated. Here, studied a collection of CLIP-Seq (CrossLinked ImmunoPrecipitation) experiments mapping the RNA binding sites for a diverse set of 46 human proteins across 68 experiments to explore the role of TEs in post-transcriptional regulation genome-wide via RNA-protein interactions. We detected widespread interactions between RNA binding proteins (RBPs) and various families of TE-derived sequence in the CLIP-Seq data. Alignment coverage clustered on specific positions of the TE consensus sequences, illuminating a diversity of TE-specific motifs for many RBPs. Evidence of binding and conservation of these motifs in the nonrepetitive transcriptome suggest that TEs have appropriated existing sequence preferences of the RBP. Upon depletion of the RBPs, transcripts possessing TE-derived binding sites were similarly regulated as those bound in nonrepetitive sequence.  However, in a few cases the effect of RBP binding depended on the specific TE family bound—e.g., the ubiquitously expressed RBP HuR conferred opposite effects on stability to transcripts when bound to Alu elements versus other families. Our meta-analysis suggests a widespread role for TEs in shaping RNA-protein regulatory networks in the human genome.

overall_design: HuR formaldehyde RIP-Seq in K562 cells, with RIP and input sequenced in triplicate.

first: John

last: Rinn


title: Generation of a Panel of Induced Pluripotent Stem Cells From Chimpanzees: a Resource for Comparative Functional Genomics (RNA-Seq)

first: Irene

last: Gallego Romero

summary: Comparative studies in primates are extremely restricted because we only have access to a few types of cell lines from non-human apes and to a limited collection of frozen tissues. In order to gain better insight into regulatory processes that underlie variation in complex phenotypes, we must have access to faithful model systems for a wide range of tissues and cell types. To facilitate this, we have generated a panel of 7 fully characterized chimpanzee (Pan troglodytes) induced pluripotent stem cell (iPSC) lines derived from fibroblasts of healthy donors. All lines are free of integration from exogenous reprogramming vectors, can be maintained using standard iPSC culture techniques, and have proliferative and differentiation potential similar to human and mouse lines. To begin demonstrating the utility of comparative iPSC panels, we collected RNA-seq data and methylation profiles from the chimpanzee iPSCs and their corresponding fibroblast precursors, as well as from 7 human iPSCs and their precursors, which were of multiple cell type and population origins. Overall, we observed much less regulatory variation within species in the iPSCs than in the somatic precursors, indicating that the reprogramming process has erased many of the differences observed between somatic cells of different origins. We identified 4,918 differentially expressed genes and 1,986 differentially methylated regions between iPSCs of the two species, many of which are novel inter-species differences and not observed between the somatic cells of the two species. Our panel will help realize the potential of iPSCs, and in combination with genomic technologies, transform studies of comparative evolution in primates.

overall_design: We obtained RNA sequencing and methylation profiles from 7 chimpanzee iPSCs and the fibroblasts used to generate them, as well as 7 human iPSCs and the LCLs and fibroblasts used to generate them.

first: Bryan

last: Pavlovic

last: Hernando-Herraez

first: Nicholas

middle: E

last: Banovich

first: Courtney

last: Kagan

first: Jonathan

last: Burnett

first: Constance

middle: H

last: Huang

first: Amy

last: Mitrano

first: Claudia

last: Chavarria

first: Inbar

middle: F

last: Ben-Nun

first: Yingchun

last: Li

first: Karen

last: Sabatini

first: Trevor

last: Leonardo

first: Mana

last: Parast

first: Tomas

last: Marques-Bonet

first: Louise

last: Laurent

first: Jeanne

last: Loring

first: Yoav

last: Gilad


title: Argonaute HITS-CLIP to identify miRNA target sites in the bovine retina

city: Cleveland

first: Thomas

last: Sundermeier

state: OHIO

address: 2109 Adelbert Road/W317

department: Pharmacology

institute: Case Western Reserve University

summary: We report identification of 348 high confidence miRNA target sites in 261 genes in the bovine retina

overall_design: Employ Argonaute HITS-CLIP analysis to identify miRNA target sites in the bovine retina

first: Krzysztof

last: Palczewski


title: The cistrome and transcriptome landscape of androgen receptor splice variants in castration-resistant prostate cancer cells

city: Rochester

first: LIGUO

last: WANG

state: MN

address: 200 1st St SW

department: Division of Computational Biology

institute: Mayo Clinic

summary: The spliced variant forms of androgen receptor (AR-Vs) have been identified recently in castration-resistant prostate cancer (CRPC) cell lines and clinical samples. Here we identified the cistrome and transcriptome landscape of AR-Vs in CRPC cell lines and determine the clinical significance of AR variants regulated gene.The AR variants binding sites can be identified in 22Rv1 cell line in the absence of androgen. Knocking down full-length AR (AR-FL) doesn't affect AR-Vs binding sites in genome-wide. A set of genes were identified to be regulated uniquely by AR-Vs, but not by AR-FL in androgen-depleted condition. Integrated analysis showed that some genes may be modulated by AR-Vs directly. Unsupervised clustering analysis demonstrated that AR variants gene signature can separate not only the benign and malignant prostate tissue, but also the localized prostate cancer and metastatic CRPC specimens. Some genes modulated uniquely by AR variants were also identified to correlate with the Gleason Pattern of prostate cancer and PSA failure. We conclude that AR spliced variants bind to DNA independent of full-length AR, and can modulate a unique set of genes which is not regulated by full-length AR in the absence of androgen. AR variants gene signature correlate with CRPC and prostate cnacer disease progress.

overall_design: Androgen receptor (AR) binding sites in human prostate cancer 22Rv1 cell lines were studied using ChIP-seq. ChIP enriched and input DNA were sequenced using Illumina HiSeq 2000.

first: Donald

last: Tindall

first: Ji

last: Lu

first: Liguo

last: Wang


title: RNAseq analysis of the global bovine retinal transcriptome

summary: We report RNAseq analysis of the transcriptome of 3 biological replicates of bovine retina

overall_design: Examine retinal transcriptome of 3 biological replicates with tissue collected between 7:00 - 10:00AM


title: HITS-CLIP and RNA-Seq of bovine retina

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

overall_design: Refer to individual Series


title: Transforming growth factor β/activin signaling functions as a sugar-sensing feedback loop to regulate digestive enzyme expression.

city: Lausanne

first: Maroun

last: Bou Sleiman

state: Vaud

address: Rue Cantonale

department: IBI

country: Switzerland

institute: EPFL

summary: Organisms need to assess their nutritional state and adapt their digestive capacity to the demands for various nutrients. Modulation of digestive enzyme production represents a rational step to regulate nutriment uptake. However, the role of digestion in nutrient homeostasis has been largely neglected. In this study, we analyzed the mechanism underlying glucose repression of digestive enzymes in the adult Drosophila midgut. We demonstrate that glucose represses the expression of many carbohydrases and lipases. Our data reveal that the consumption of nutritious sugars stimulates the secretion of the transforming growth factor β (TGF-β) ligand, Dawdle, from the fat body. Dawdle then acts via circulation to activate TGF-β/Activin signaling in the midgut, culminating in the repression of digestive enzymes that are highly expressed during starvation. Thus, our study not only identifies a mechanism that couples sugar sensing with digestive enzyme expression but points to an important role of TGF-β/Activin signaling in sugar metabolism.

overall_design: RNA-sequencing of whole guts from Drosophila melannogaster OregonR adult females was performed under three feeding conditions: Standard medium, glucose, and agar. Three biological repeats were performed for each condition.

first: Wen-bin

last: Chng

first: Fanny

last: Schüpfer

first: Bruno

last: Lemaitre


title: Transcriptome profiling of Arabidopsis smg7 mutant in immune signaling blocked background

city: Vienna

first: Jiradet

last: Gloggnitzer

address: Dr.-Bohr-Gasse 3

country: Austria

institute: Gregor Mendel Institute of Molecular Plant Biology

summary: Goal of the study is the identification of transcriptome deregulation of smg7 pad4 mutants, which are deficient in nonsense-mediated mRNA decay and are blocked in immune signaling, which should avoid secondary responses from immune signaling

overall_design: Total RNA (ribosomal RNA depleted) illumina sequencing of three single end libraries respectively of adult pad4 leaves (control) vs adult smg7 pad4 mutant leaves (mutant)

first: J

first: A

last: Srinivasan

last: Jimenez-Gomez

first: K

last: Riha


title: Strand-specific RNA sequencing profile of dbp2Δ  vs. wild type strains

city: West Lafayette

first: Elizabeth

last: Tran

state: IN

address: 175 S. University Street

department: Biochemistry

institute: Purdue University

summary: We report widespread changes in ribosome biogenesis factor, aerobic respiration and hexose tranporter gene expression. We also identify antisense transcripts to hexose transporter genes in dbp2∆ cells.

overall_design: Strand-specific RNA profiles of a dbp2Δ strain and a wild type (reference) strain were generated using two replicates of each strain.


title: Differential miRNAs between Sensitive and Resistant Pig to Escherichia coli F18 by deep sequencing

status: Public on Sep 21 2014

city: Yangzhou

first: Lan

last: Ye

state: Jiangsu

address: Daxue Road  NO 88

department: animal science and technology

institute: Yangzhou university

summary: An M307 G/A point mutation of FUT1 gene has been considered as a usful marker to select the piglets that are sensitive(GG/AG genotype) or resistant(AA) to Escherichia coli F18 in foreign pig breeds. However,it is not suitable to Chinese native breeds. Duodenal tissues were collected from 3 full-sib pairs of Sutai pigs (a new hybrid between the Duroc and Taihu breeds) at the age of 28-day differing in adhesion phenotype to find the differential miRNAs that can  provide the basis for analyzing the different mechanisms of E.coli F18 resistance between foreign and Chinese native breeds,as well as for breeding for disease resistance in Chinese native breeds in the future.

overall_design:
piglets at the age of 28-day sensitive(GG genotype) to Escherichia coli F18  vs resistant(AA) ones.Biological replicates:3 full-sib pairs of Sutai pigs(3 pairs for GG/AA genotype of FUT1 gene).One GG sample (512 piglet) was sequenced twice to verify the result.
2385 piglet and 2383 piglet were a pair of full-sib with AA and GG genotype at M307 positionof FUT1 gene, respectively. Unfortunately, the sequencing of 2385 was a failure and was excluded.
2385 piglet has an AA genotype of the FUT1 gene

first: Wenbin

last: Bao


title: Global MEF2 target gene analysis in cardiac and skeletal muscle reveals novel regulation of DUSP6 by p38MAPKMEF2 signaling [RNA-seq]

status: Public on Sep 09 2014

city: Toronto

first: John

last: McDermott

state: ON

address: 4700 Keele Street

country: Canada

institute: York University

summary: Identfification of MEF2A target genes using ChIP-exo and RNA-seq in skeletal muscle and primary cardiomyocytes. MEF2 plays a profound role in the regulation of transcription in cardiac and skeletal muscle lineages. To define the overlapping and unique MEF2A genomic targets, we utilized ChIP-exo analysis of cardiomyocytes and skeletal myoblasts. Of the 2783 and 1648 MEF2A binding peaks in skeletal myoblasts and cardiomyocytes, respectively, 294 common binding sites were identified. Genomic targets were compared to differentially expressed genes in RNA-seq analysis of MEF2A depleted myogenic cells.

overall_design: The effect of MEF2A gene silencing on gene expression in myoblasts was assessed at 48 hr DM. Up and downregulated genes were then compared to MEF2A target genes identified in ChIP-exo analysis of  48 hr DM C2C12 myoblasts cells and primary cardiomyocytes.

first: S

last: Wales


title: Global MEF2 target gene analysis in cardiac and skeletal muscle reveals novel regulation of DUSP6 by p38MAPKMEF2 signaling

sample_organism:
Mus musculus
Rattus norvegicus

platform_organism:
Mus musculus
Rattus norvegicus


title: Transcriptional profiles of bovine in vivo pre-implantation development

city: Gainesville

first: Zongliang

last: Jiang

state: FL

address: 2033 Mowry Rd

department: Animal Sciences

institute: University of Florida

summary: During mammalian pre-implantation embryonic development dramatic and orchestrated changes occur in gene transcription.  The identification of the complete changes has not been possible until the development of the Next Generation Sequencing Technology. Here we report the first transcriptome dynamics of single matured bovine oocytes and all stages of pre-implantation embryos developed in vivo.  Surprisingly, nearly half of the bovine genome, 11,488 to 12,729 genes involved in more than 100 pathways, is expressed in oocytes and early embryos.  Despite the similarity in the total numbers of genes expressed across stages, the nature of the expressed genes is dramatically different.  A total of 2,845 genes were differentially expressed among different stages, of which the largest change was observed between the 4- and 8-cell stages, demonstrating that the bovine embryonic genome activation occurs at this transition.  Additionally, 774 genes were identified as only expressed/highly enriched in particular stages of development.  Using weighted gene co-expression network analysis, we found 12 stage-specific modules of co-expressed genes that can be used to represent the corresponding stage of development.  Furthermore, we identified conserved key members (or hub genes) of the bovine expressed gene networks.  Their vast association with other embryonic genes suggests that they may have important regulatory roles in embryogenesis; yet, the majority of the hub genes are relatively unknown/under-studied in embryos.  We also conducted the first embryonic expression profile comparison across three mammalian species, human, mouse and bovine, for which RNA-seq data are available.  We found that the three species share more maternally deposited genes than embryonic genome activated genes.  More importantly, there are more similarities in embryonic transcriptomes between bovine and humans than between humans and mice, demonstrating that bovine embryos are better models for human embryonic development. This study provides the first comprehensive examination for gene activities in bovine embryos and identified little-known potential master regulators of pre-implantation development.

overall_design: RNA-Seq profiles from single in vivo matured oocytes and in vivo developed embryos from the 2-cell to the blastocyst stages generated using Solid RNA-seq platform.

first: Jiangwen

last: Sun

first: Hong

last: Dong

first: Oscar

last: Luo

first: Xinbao

last: Zheng

first: Craig

last: Obergfell

first: Yong

last: Tang

first: Jinbo

last: Bi

first: Rachel

last: O'Neill

first: Yijun

last: Ruan

first: Jingbo

last: Chen

first: Xiuchun

last: Tian


title: Competence for reprogramming sex

city: Madison

first: Elena

last: Sorokin

state: WI

address: 433 Babcock Drive

institute: UW-Madison

summary: We used RNA-Seq to compare transcriptomes of chemical reprogramming competent worms versus worms not competent for chemical reprogramming. We also performed RNA-seq during a time course of chemical reprogramming.

overall_design: Three replicates of each of two reprogramming non-competent strains and three replicates of each of two reprogramming competent strains were collected. For the time course, five time points were analyzed (1, 2, 4, 6, and 18 hours) in either DMSO or DMSO + U0126 in three genotypes (non-reprogramming competent worms, reprogramming competent, and wildtype worms).

first: Judith

last: Kimble


title: Transcriptome sequencing of Fukomys damarensis

city: shenzhen

first: zhiyong

last: huang

state: Guangdong

address: F5, Main Building, Beishan Industrial Zone, Beishan Road, Yantian District

institute: BGI-Shenzhen, Shenzhen 518083, China

summary: Deep sequencing of mRNA from Fukomys damarensis to uncover the reproduce regulation

overall_design: Analysis of ploy(A)+ RNA of Fukomys damarensis

first: Xiaodong

last: Fang

first: Vadim

last: Gladyshev

first: Zhiyong

last: Huang


title: Global MEF2 target gene analysis in cardiac and skeletal muscle reveals novel regulation of DUSP6 by p38MAPKMEF2 signaling [ChIP-seq]

first: John

last: McDermott

summary: Identfification of MEF2A target genes using ChIP-exo in skeletla muscle and primary cardiomyocytes. Identfification of MEF2A target genes using ChIP-exo and RNA-seq in skeletal muscle and primary cardiomyocytes. MEF2 plays a profound role in the regulation of transcription in cardiac and skeletal muscle lineages. To define the overlapping and unique MEF2A genomic targets, we utilized ChIP-exo analysis of cardiomyocytes and skeletal myoblasts. Of the 2783 and 1648 MEF2A binding peaks in skeletal myoblasts and cardiomyocytes, respectively, 294 common binding sites were identified. Genomic targets were compared to differentially expressed genes in RNA-seq analysis of MEF2A depleted myogenic cells.

overall_design: MEF2A target genes were identified in 48 hr DM C2C12 myoblasts cells and primary cardiomyocytes using ChIP-exo. Binding profiles on MEF2A in each cell type were compared. Cross sectional-analysis between ChIP-exo identified targets and RNA-seq analysis of MEF2A deplted myoblasts was also done.

first: S

last: Wales

first: J


title: Deciphering the Fur transcriptional regulatory network highlights its complex role beyond iron metabolism in Escherichia coli [RNA-seq]

city: Ulsan

first: Donghyuk

last: Kim

address: 50 UNIST-gil, Eonyang-eup, Ulju-gun

department: Department of Chemical Engineering

country: South Korea

institute: UNIST

summary: The ferric uptake regulator (Fur) plays a critical role in the transcriptional regulation of iron metabolism in many bacteria. However, the full regulatory potential of Fur beyond iron metabolism remains undefined. Here, we comprehensively reconstructed the Fur transcriptional regulatory network in Escherichia coli K-12 MG1655 in response to iron availability using genome-wide measurements (ChIP-exo and RNA-seq). Polyomic data analysis revealed that a total of 81 genes in 42 transcription units (TUs) are directly regulated by three different modes of Fur regulation, including apo- and holo-Fur activation as well as holo-Fur repression. We showed that Fur connects iron transport and utilization enzymes with negative-feedback loop pairs for iron homeostasis. In addition, direct involvement of Fur in the regulation of DNA synthesis, energy metabolism, and biofilm development was found. These results indicate that Fur exhibits a comprehensive regulatory role affecting many fundamental cellular processes linked to iron metabolism in order to coordinate E. coli responses to the availability of iron.

sample_organism: Escherichia coli str. K-12 substr. MG1655

overall_design: [RNA-seq]:  A total of eight samples were analyzed. WT and fur mutant cells were cultured in the presense and absence of iron with biological duplicates. DPD = iron chelator.

first: Sang Woo

last: Seo


title: Different nucleosomal landscapes at early and late replicating origins in Saccharomyces cerevisiae

city: Madrid

first: Enrique

last: Vazquez de Luis

state: Madrid

address: Melchor Fernandez Almagro, 3

country: Spain

institute: CNIC

summary: Sequencing of mononucleosomal DNA during G1 and S phases in Saccharomyces cerevisiae

overall_design: Samples from mononucleosomal DNA from WT and rpd3 mutant strains (W303-1a background) in G1 or in S phase in the presence of 0.2 M HU were sequenced (Illumina Genome Analyzer IIx) using the single-end read protocol

first: Ignacio

last: Soriano

first: Esther

last: Cabañas

last: Vázquez

first: Francisco

last: Antequera

first: Monica

last: Segurado


title: Deciphering the Fur transcriptional regulatory network highlights its complex role beyond iron metabolism in Escherichia coli

platform_organism: Escherichia coli str. K-12 substr. MG1655


title: The exon junction complex controls transposable element activity by ensuring the faithful splicing of the piwi transcript

status: Public on Sep 08 2014

first: Jean-Yves

last: Roignant

address: genopode building

institute: University of Lausanne

summary: The exon junction complex (EJC) is a highly conserved ribonucleoprotein complex which binds RNAs at a late stage of the splicing reaction and remains associated following export to the cytoplasm. This complex is involved in several cellular post-transcriptional processes including mRNA localization, translation and degradation. The EJC plays an additional role in the splicing of a subset of genes in Drosophila and in human cells but the underlying mechanism remains to be elucidated. Here, we have found a novel function for the EJC and its splicing subunit RnpS1 in preventing transposon accumulation in both Drosophila germline and surrounding follicular cells. This function is mediated specifically through the control of the splicing of the piwi transcript. In absence of RnpS1 one of the piwi intron is retained. This intron contains a weak 5’ splice site as well as degenerate transposon fragments, reminiscent of heterochromatic introns. In addition, we identified a small A/T rich region, which alters its polypyrimidine tract (PPT) and confers the RnpS1’s dependency. Finally, we showed that the removal of this intron by RnpS1 requires the initial splicing of the flanking introns, suggesting a model in which the EJC facilitates the splicing of challenging introns following its initial deposition to adjacent exon junctions.

overall_design: In total there are 4 different conditions. Comparisons were made between piwi mutant vs control piwi and rnps1 KD vs controls RnpS1

first: Junaid

last: Akhtar

first: Colin

last: Malone

first: Nastasja

last: Kreim


title: Identification of differentially expressed genes during development of the zebrafish pineal complex using RNA sequencing

city: Nashville

first: Sataree

last: Khuansuwan

state: TN

address: 465 21st Avenue South

department: Biological Sciences

institute: Vanderbilt University

summary: We describe a method for isolating RNA suitable for high-throughput RNA sequencing (RNA-seq) from small numbers of fluorescently labeled cells isolated from live zebrafish (Danio rerio) embryos without using costly, commercially available columns. This method ensures high cell viability after dissociation and suspension of cells and gives a very high yield of intact RNA. We demonstrate the utility of our new protocol by isolating RNA from fluorescence activated cell sorted (FAC sorted) pineal complex neurons in wild-type and tbx2b knockdown embryos at 24 hours post fertilization. Tbx2b is a transcription factor required for pineal complex formation. We describe a bioinformatics pipeline used to analyze differential expression following high-throughput sequencing and demonstrate the validity of our results using in situ hybridization of differentially expressed transcripts. This protocol brings modern transcriptome analysis to the study of small cell populations in zebrafish.

overall_design: Differential expression analysis of mRNA levels in a single time-point (24 hpf) between wild-type and Tbx2b deficient FAC sorted pineal complex cells

first: Joshua

middle: T

last: Gamse


title: H2A.Z knockdown RNAseq

city: Birmingham

first: Jeremy

middle: Jason

last: Day

state: AL

address: 1825 University Blvd

department: Neurobiology

institute: UAB

summary: This experiment seeks to ascertain the transcriptional changes in the adult mouse hippocampus (CA1 subregion) that occur following viral knockdown of the histone variant H2A.Z. We are especially interested in understanding the role of this histone variant in memory formation and memory maintenance in the adult central nervous system.

overall_design: This experiment includes 3 groups, each with 3 biological replicates. Samples S108, S109, and S110 are from controls infected with an AAV expressing a scrambled shRNA control. Samples A100, A101, A102, A104, A106, and A107 were infected with an AAV expressing an shRNA against H2A.Z. Samples A100, A101, and A102 were naive animals, whereas samples A104, A106, and A107 were trained in contextual fear conditioning.

first: Iva

middle: B

last: Zovkic

first: Brynna

last: Paulukaitis

first: Deepa

last: Etikala

middle: D

last: Sweatt


title: Differential expression of miRNAs and their target genes in senescing leaves and siliques: insights from massive sequencing of small RNAs and cleaved target RNAs

city: Newark

first: Shawn

last: Thatcher

state: DE

address: 15 Innovation Way

institute: University of Delaware

summary: To identify senescence-regulation of miRNAs in Arabidopsis thaliana, eight small RNA libraries were constructed and sequenced at four different stages of development and senescence from both leaves and siliques. Parallel Analysis of RNA Ends (PARE) libraries were also constructed and sequenced to enable the large-scale examination of miRNA-guided cleavage products.

overall_design: Genome-wide small RNA profiling was done by Illumina TruSeq sample preparation followed by high-throughput sequencing with Illumina HiSeq 2000 platform.

first: Pamela

last: Green


title: An activating NLRC4 inflammasome mutation causes autoinflammation with recurrent macrophage activation syndrome

city: BETHESDA

first: Scott

last: Canna

state: MD

department: MIIB

institute: NIAMS

summary: Inflammasomes are intracellular innate immune sensors that respond to pathogen and damage-associated signals with the proteolytic cleavage of caspase-1, resulting in IL-1_ and IL-18 secretion and macrophage pyroptosis. The discovery that heterozygous gain-of-function mutations in NLRP3 lead to oversecretion of IL-1_ and cause the autoinflammatory disease spectrum Cryopyrin Associated Periodic Syndrome (CAPS), led to the successful use of IL-1 blocking therapies1. We found that a de novo missense mutation in the regulatory domain of the NLRC4 (IPAF, CARD12) inflammasome causes early-onset recurrent fever flares and Macrophage Activation Syndrome (MAS). Functional analyses demonstrated spontaneous production of the inflammasome-dependent cytokines IL-1² and IL-18 exceeding levels in CAPS patients. The NLRC4 mutation led to constitutive caspase-1 cleavage in transduced cells and enhanced spontaneous production of IL-18 by both patient and NLRC4 mutant macrophages. Thus, we describe a novel monoallelic inflammasome defect that expands the autoinflammatory paradigm to include MAS and suggests novel targets for therapy.

overall_design:
Whole blood RNA-seq from seven timepoints of one patient with NLRC4-MAS as compared to five healthy pediatric controls, 7 NOMID patients with active disease prior to anakinra treatment and the same 7 NOMID patients with inactive disease after anakinra treatment.

Please note that seven time points are chronologic time point. They are ordinal, in that "1" was drawn before "2", but the distance in time between points is not constant. Thus, time points 4 through 7 correspond to samples drawn while the patient was well AND on treatment. However there may be differences between 4 and 7 pertaining to the length of treatment, and for that reason any of these samples were not considered replicates.

middle: W

first: Adriana

middle: A

last: de Jesus

first: Stephen

last: Brooks

first: Raphaela

last: Goldbach-Mansky


title: Genome wide mapping of histone 3 lysine 79 mono, di and trimethylation and histone 3 lysine 27 trimethylation in LSKs, GMPs and retroviral MLL-AF9 driven murine leukemias

city: New York

last: Armstrong

address: 1275 York Avenue

institute: Memorial Sloan-Kettering Cancer Center

summary: We report the genome wide distribution of the three states of H3K79 methylation (H3K79me1/me2/me3) and H3K27me3 in mouse lineage negative Sca-1 positive Kit positive cells (LSKs), granulocyte macrophage progenitors (GMPs) and LSK derived MLL-AF9 leukemias in the presence or absence of the Af10 OM-LZ domain. Legend- MIT:MSCV-IRES-tdTomato (Empty vector control) and CRE (MIT vector with the Cre recombinase).

overall_design: We examined the H3K79 me1,me2,me3 and H3K27me3 profiles by ChIP-seq in  lineage negative Sca-1 positive, Kit positive  (LSK) cells, granulocyte macrophage progenitors (GMPs) and bone marrow cells from sacrificed terminally ill secondary MLL-AF9 positive leukemic mice. In case of the MLL-AF9 leukemias, the ChIP-seq experiments were performed in 2 conditions in the presence or absence of the Dot1l interacing octapeptide-motif leucine zipper (OM-LZ) domain of Af10.  For the leukemia experiments, leukemias derived from the Af10 OM-LZ homozygous floxed background were transduced with MSCV-IRES-tdTomato control vector (MIT) or its Cre-recombinase expressing counterparts (CRE). Subsequently, we sorted tdTomato positive cells and injected them into sub-lethally irradiated syngenic secondary recipient mice. Seconday leukemias obtained from these MIT or CRE expressing cells were used for ChIP -seq studies.

first: Aniruddha

last: Deshpande


title: Chickpea Small RNA Atlas - Development

city: New Delhi

first: Mukesh

last: Jain

state: New Delhi

address: New Mehrauli Road

country: India

institute: Jawaharlal Nehru University

summary: In this study, we sequenced small RNA content from seven major tissues/organs employing Illumina technology. More than 154 million reads were generated using Illumina high-throughput sequencing GAII platform, which represented more than 20 million distinct small RNA sequences. After pre-processing, several conserved and novel miRNAs were identified in chickpea. Further, the putative targets of chickpea miRNAs were identified and their functional categorization was analyzed. In addition, we identified miRNAs exhibitng differential and specific expression in various tissues/organs.

overall_design: We collected different tissue samples used in this study and total RNA isolated was subjected to Illumina sequencing. The sequenced data was further filtered using NGS QC Toolkit to obtain high-quality reads. The filtered reads were pre-processed using modified perl script provided in the miRTools software. After quality control, the identical reads were collapsed into a unique read and read count for each sequence was recorded. All the filtered unique reads from each sample were screened stepwise against annotated non-coding RNA sequences, including plant snoRNA, tRNA and rRNA. The remaining  reads were screened against repeat sequences from RepBase and chickpea chloroplast sequence. Conserved miRNAs were identified based on similarity with miRBase database and novel miRNAs were identified using miRDeep-P pipeline. For differential expression analysis, the read count for each miRNA was normalized using DESeq software. The genes preferentially and specifically expressed in various tissues/organs were identified.

first: Narayana

middle: V

last: Chevala

first: Rohini

last: Garg


title: The developmental potential of iPSCs is greatly influenced by the selection of the reprogramming factors (SNEL iPSC and H2A.X deposition pattern)

status: Public on Sep 07 2014

city: New Haven

first: Andrew

middle: Zhuo

last: Xiao

state: CT

address: 10 Amistad St

department: Genetics

institute: Yale Stem Cell Center

summary: Induced pluripotent stem cells (iPSCs) are commonly generated by transduction of Oct4, Sox2, Klf4 and Myc (OSKM) into somatic cells. Though iPSCs are pluripotent, they frequently exhibit high variation in their quality as measured by chimera contribution and tetraploid (4n) complementation. Thus, improving the quality of iPSCs is an indispensable prerequisite for future iPSC-based therapy. Here we show that one major determinant for iPSCs quality is the selection of the reprogramming factors combination. Ectopic expression of Sall4, Nanog, Esrrb and Lin28 (SNEL) in MEFs efficiently generated high quality iPSCs as compared to other combinations of factors. SNEL-iPSCs produced approximately 5 times more efficiently “all-iPSC” mice compared to OSKM-iPSCs. While differentially methylated regions, transcript number of master regulators, establishment of ESC-specific super enhancers, and global aneuploidy were comparable between the lines,  aberrant expression of 1,765 genes, trisomy of chromosome 8 and abnormal H2A.X deposition were frequently observed  in poor quality OSKM-iPSCs. For high-quality iPSCs, H2A.X pattern of SNEL is most similar to that of ESC, OSK and OSKM have more devoid regions than SNEL iPSCs.

overall_design: Compare H2A.X deposition pattern of the OSKM 4-factor iPS cell lines (4N-), SNEL 4-factor iPS cell lines (4N+) with ChIP-Seq. The same background ES cell line as the control line.

first: Yosef

last: Buganim

first: Rudolf

last: Jaenisch

first: Tao

last: Wu


title: Role of H3K79 methylation states in HOX gene expression and leukemogenesis

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

overall_design: Refer to individual Series


title: Histone Variant H2A.X Plays Novel Roles in Cell Lineage Commitment and Genomic Stability in ESC and iPSC

summary: Herein, we demonstrated that the cell lineage commitment is unexpectedly regulated by the novel functions of H2A.X, a histone variant which was only well-known for its role in genome integrity maintenance previously. Surprisingly, only in ESCs but not differentiated cells, H2A.X is specifically targeted to genomic regions encoding early embryonic and extra-embryonic lineage genes to repress their expression. In addition, H2A.X is also enriched at genomic regions sensitive to replication stress and maintains genomic stability thereat. Most interestingly, faithful H2A.X deposition plays critical roles in maintaining both cell lineage commitment and genome integrity in iPSC. In iPSC lines which support the development of "all-iPS" animals, H2A.X deposition faithfully recapitulates the ESC pattern and therefore, the genome stability and cell lineage commitment are maintained. In iPSC lines that fail to support embryonic development, defective H2A.X depositions result in aberrant upregulation of early embryonic and extra-embryonic lineage genes and H2A.X-dependent genome instability.

overall_design: mRNA-Seq of WT ESC and H2A.X KO ESC; and 4N+, 4N- iPSC.


title: Histone H3.3 regulates dynamic chromatin states during spermatogenesis

city: Sacramento

first: Paul

last: Knoepfler

state: CA

address: 2425 Stockton Blvd

department: Cell Biology and Human Anatomy

institute: UC Davis

summary: We used H3f3b-/- testes to analyze the impact of H3f3b derived H3.3 on spermatogenesis.

overall_design: Analysis of Histone H3 (Lys 4) tri-methylation in H3f3b WT/WT and H3f3b -/- testes tissue.

first: Ben

last: Yuen

first: Rebecca

last: Cotterman

first: Kelly

last: Bush


title: H2A.X native ChIP-Seq in TSC; Histone Variant H2A.X Mediated Epigenetic Mechanisms are Critical for Pluripotency in ES and iPS Cells

summary: With the advent of the induced pluripotent stem cell (iPSC) technology, how to distinguish the developmental potentials of the iPSC clones with molecular approaches becomes an imperative issue.  Herein, we demonstrated that histone variant H2A.X plays an unexpected role in distinguishing the developmental potentials of iPSC.  We showed that H2A.X is specifically targeted to and negatively regulates extra-embryonic lineage gene expression in embryonic stem cell (ESCs) and therefore, it prevents trophectoderm (TE) lineage differentiation under inductive conditions.  ESC-specific H2A.X deposition and functions are faithfully recapitulated in the iPSC lines that support the development of “all-iPS” animals.   In iPSC lines that fail to support embryonic development, aberrant H2A.X depositions result in upregulation of extra-embryonic lineage genes and predisposition to extra-embryonic tissue differentiation.   In summary, our work has revealed novel epigenetic mechanisms for maintaining cell lineage commitment, which can be used to distinguish the quality of the iPSC lines.

overall_design: Detect and compare different H2A.X deposition patterns in ES cells [GSE42306] and TS cells, with Illumina HiSeq 2000


title: MicroRNA and mRNA expression analyses of postnatal and adult rat pancreatic islets

status: Public on Sep 06 2014

city: Indianapolis

first: Scot

last: Matkovich

address: Lilly Corporate Center

institute: Eli Lilly and Company


title: Islet-specific microRNA changes driven by nutrient shift trigger postnatal beta-cell maturation - dispersed islet cell mRNAs after miR-17-5p transfection

summary: We found that in rodents, postnatal beta-cell maturation is associated with changes in the expression of several islet microRNAs and discovered that these modifications are driven by changes in the nutrient supply. Mimicking the microRNA changes observed during β-cell maturation in newborn rat islet cells was sufficient to promote glucose-induced insulin release and to achieve a mature β-cell secretory phenotype. Moreover, the modifications in the level of some of these microRNAs reduced the proliferation of newborn β-cells, suggesting that they contribute to the limited proliferative capacity of adult β-cells. These findings demonstrated that miRNAs contribute to postnatal beta-cell maturation and development. Their role is likely to promote beta-cell adaptation to fule supply and to maintain glucose homeostasis by regulating insulin release and proliferation.

overall_design: Islets from 10-day-old rats (P10) were taken, dispersed and transfected with control miRNA mimic or miR-17-5p. Total RNA was extracted and mRNA profiling via Illumina single-end sequencing of mRNA-seq libraries was performed.  In parallel, Ago2 immunoprecipitation with RNA recovery and mRNA-seq was performed (RISC-seq).

first: Cecile

last: Jacovetti

first: Romano

last: Regazzi


title: H2A.X native ChIP-Seq in ESC and iPSC: Histone Variant H2A.X Mediated Epigenetic Mechanisms are Critical for Maintaining Genome Stability and Pluripotency in ES and iPS Cells

summary: It is well-known that embryonic stem cells (ESC) are much more sensitive to replication-induced stress than differentiated cells but the underpinning mechanisms are largely unknown. H2A.X, a minor variant of H2A, constitutes only 1-10% of the mammalian genome.  H2A.X plays a well-known for role in the DNA damage response and maintaining stability in the genome, including the regions frequently experiencing replication stress, such as the fragile sites. Intriguingly, several recent studies have reported that H2A.X function is elevated in ESC; and others reported that H2A.X function is provoked during cellular reprogramming (in induced pluripotent stem cells, iPSC), indicating that increased proliferation during iPS may trigger replication stress and the H2A.X DNA damage response.  However, several studies of genomic instability in iPSC led to different conclusions on this important issue. For example, frequent copy number variants (CNV) were reported at the genomic regions sensitive to replication stress, such as the fragile sites. On the other hand, another study reported the lack of genomic instability in mouse iPS clones that are able to generate “all-iPS” animals in tetraploid complementation assays (4N+ iPSC), indicative of a potential link between pluripotency and genome integrity. However, whether if high level genomic instability occurs in the 4N- iPSC iPSC clones at replication stress sensitive regions is unknown.  Moreover, due to the lack of mechanistic insights on genome integrity maintenance, how pluripotency and genome integrity are connected remains elusive. Here we show that H2A.X plays unexpected roles in maintaining pluripotency and genome integrity in ESC and iPSC. In ESC, it is specially enriched at genomic regions sensitive to replication stress so that it protects genome integrity thereat. Faithful H2A.X deposition is critical for genome integrity and pluripotency in iPSC. H2A.X depositions in 4N+ iPSC clones faithfully recapitulate the ESC pattern and therefore, prevent genome instability. On the other hand, insufficient H2A.X depositions in 4N- iPSC clones at such regions lead to genome instability and defects in replication stress response and DNA repair, reminiscent of the H2A.X deficient ESC.

overall_design: Detect and compare different H2A.X deposition patterns in ES cells and iPS cells, with Illumina HiSeq 2000 and Illumina Genome Analyzer IIx


title: Islet-specific microRNA changes driven by nutrient shift trigger postnatal beta-cell maturation - intact islet mRNAs

overall_design: Islets from 10-day-old rats (P10) (n=3) or 3-month-old male rat (n=3) were taken. Total RNA was extracted and mRNA profiling via Illumina single-end sequencing of mRNA-seq libraries was performed.


title: Deciphering the Fur transcriptional regulatory network highlights its complex role beyond iron metabolism in Escherichia coli [ChIP-Seq]

first: Donghyuk

last: Kim

overall_design: [ChIP-exo]:  A total of twelve samples were analyzed. WT and Fur-8-myc tagged cells were cultured in the presense and absence of iron with biological duplicates. To analyze static RNAP binding, rifampicin was also added to the media with biological duplicates. DPD = iron chelator.

first: Sang Woo

last: Seo


title: Islet-specific microRNA changes driven by nutrient shift trigger postnatal beta-cell maturation - intact islet microRNAs

overall_design: Islets from 10-day-old rats (P10) (n=5) or 3-month-old male rat (n=6) were taken. Total RNA was extracted and microRNA profiling was performed using the Illumina TruSeq small RNA kit and single-end sequencing.


title: Upf2 in NMD pathway

city: reno

first: Wei

last: Yan

state: NV

address: 1664 N. Virginia Street

department: Physiology

institute: University of Nevada

summary:
Purpose: Probe the transcriptome-wide changes in the expression pattern between WT and Sertoli-specific Upf2 KO testes
Methods: Total RNA were extracted from WT and Sertoli-specific Upf2 KO testes in triplicates and subject to deep-sequencing in Ion Torrent seq platform.
Results: Using an optimized data analysis workflow, we mapped about 30 million sequence reads per sample to the mouse genome (build mm9) and identified 16,014 transcripts in the retinas of WT and Nrl−/− mice with BWA workflow and 34,115 transcripts with TopHat workflow. RNA-seq data confirmed stable expression of 25 known housekeeping genes, and 12 of these were validated with qRT–PCR. RNA-seq data had a linear relationship with qRT–PCR for more than four orders of magnitude and a goodness of fit (R2) of 0.8798. Approximately 10% of the transcripts showed differential expression between the WT and Nrl−/− retina, with a fold change ≥1.5 and p value <0.05. Altered expression of 25 genes was confirmed with qRT–PCR, demonstrating the high degree of sensitivity of the RNA-seq method. Hierarchical clustering of differentially expressed genes uncovered several as yet uncharacterized genes that may contribute to retinal function. Data analysis with BWA and TopHat workflows revealed a significant overlap yet provided complementary insights in transcriptome profiling.
Conclusions: Our study represents the first detailed analysis of Upf2-mediated NMD pathway in Sertoli cell development

overall_design: Testis mRNA profiling was generated from postnatal day 4 WT and Amh-cKO (Sertoli specific Upf2 KO) testes, in triplicates.

first: Jianqiang

last: Bao

first: Chong

last: Tang

first: Tian

last: Yu


title: H2A.X function in mouse ESC and iPSC (ChIPSeq and CNVs)

first: Andrew

last: Xiao

type: Genome variation profiling by genome tiling array


title: Twi1p-loaded scnRNAs in GIW1 KO Tetrahymena cells

status: Public on Sep 23 2014

city: Montpellier

first: Kazufumi

last: Mochizuki

address: 141 rue de la Cardonille

country: France

institute: Institute of Human Genetics, CNRS

summary: To understand the importance of Giw1p in the loading of small RNAs to the Argonaute protein Twi1p in Tetrahymena, we compared Twi1p-bound small RNAs from wild-type cells and GIW1 KO cells. We found that the Twi1p-bound small RNA populations in wild-type cells and GIW1 KO cells are indistinguishable in their size distributions and base profiles.

overall_design: We immunopurified Twi1p-containing complex from wild-type and GIW1 KO cells and co-purified small RNAs were compared.

first: Tomoko

last: Noto


title: Temporal RNA-seq expression profiling of Sox17-medaited XEN conversion of ESCs

status: Public on Sep 05 2014

city: Toronto

first: Angela

middle: C

last: McDonald

state: Ontario

address: 686 Bay Street

institute: The Hospital for Sick Children

summary: The transcription factor Sox17 is expressed in early primitive endoderm-fated cells of the mouse embryo and in embryo-derived extraembryonic endoderm (ExEn) stem (XEN) cells. We have shown that overexpression of Sox17 in mouse embryonic stem cells (ESCs) drives cell fate to a committed XEN-like cell state (Sox17-XEN cells). When placed back into the embryo, Sox17-XEN cells contribute exclusively to the ExEn. Transient Sox17 expression is sufficient to drive this fate change during which time cells transit through distinct intermediate states prior to the generation of functional XEN-like cells. We identified dynamic regulatory networks driving Sox17-mediated XEN conversion by analyzing a dynamic regulatory map of gene expression bifurcation points throughout conversion, created using RNA-seq time series data. We found that Sox17 orchestrates this conversion process by acting in autoregulatory and feed-forward network motifs, regulating dynamic gene regulatory networks (GRNs) directing cell fate. We have shown that Sox17-mediated XEN conversion provides a powerful tool for understanding the regulation of cell fate changes and for the elucidation of GRNs regulating lineage decisions in the mouse embryo.

overall_design: Total RNA was extracted during a time course of Sox17 overexpression in mouse ESCs at 7 time points as well as from wild-type ESCs and wild-type XEN cells.

first: Steffen

last: Biechele

first: Janet

last: Rossant

first: William

middle: L

last: Stanford


title: Subclone of resistant CT26 murine tumor shows high sensitivity to VA7 virotherapy: A novel system to study innate immunity and antigen dominance in oncolytic immuno-virotherapy

city: Kuopio

first: Minna

middle: U

last: Kaikkonen

department: A.I. Virtanen Institute, Department of Biotechnology and Molecular Medicine

country: Finland

institute: University of Eastern Finland

summary: Recently, attenuated Semliki Forest virus vector VA7 completely eliminated type I interferon (IFN) unresponsive human U87 glioma xenografts while IFN responsive mouse GL261 and CT-2A gliomas proved refractory to the oncolytic virotherapy. Here we describe in two clones of a well established Balb/c mouse tumor cell line, CT26 murine colon carcinoma, diametrically opposed IFN responsiveness and sensitivity to oncolytic virus. Both CT26WT and CT26LacZ clones secreted biologically active type I IFN in vitro upon infection but virus replication was self-limiting only in CT26WT cells. Total transcriptome sequencing (RNA-Seq) and western blotting experiments revealed that in sharp contrast to CT26LacZ cells, CT26WT cells had strong constitutive expression of 56 different genes associated with pattern recognition and type I interferon signaling pathways, spanning two reported anti-RNA virus gene signatures and22 genes that have been reported to have direct anti-Alphaviral activity. Correspondingly, only CT26LacZ tumors were infectable in vivo, resulting in rapid central necrosis of the  tumors by 96 hours post infection and complete tumor eradication both in immunocompetent and in SCID mice. CT26LacZ tumor eradication by oncolysis induced 100% protective immunity against homologous CT26LacZ challenge but only 50% protection against heterologous CT26WT challenge, indicating LacZ immune dominance over shared antigens. We believe the two clone CT26 system  described herein constitutes a challenging yet realistic model for clonally and immunologically heterogeneous cancer where a strong therapy efficacy bias toward sensitive tumor subpopulations might falsely predict therapeutic success on a broad patient scale highlighting the necessity of successful pre-screening for responsive tumors.

overall_design: RNA-Seq in CT26 tumor cell line

first: Janne

last: Ruosalainen


title: Integrative study of cooperative lineage-survival oncogenes in gastric cancer

city: Singapore

first: Niantao

last: Deng

state: Singapore

address: Center for Computational Biology, level 6, 8 College Road

country: Singapore

institute: Duke NUS Graduate Medical School


title: IsomiRage: from functional classification to differential expression of miRNAs and their isoforms

city: Milan

first: Heiko

last: Muller

address: Via Adamello 16

country: Italy

institute: Italian Institute of Technology

summary: Characterization and analysis of miRNAs and their variants (called "IsomiRs") in next-generation sequencing datasets in breast epithelium. Our pipeline, called IsomiRage, allows distinguishing canonical miRNAs from templated and non-templated isomiRs by alignment to a custom database, which comprises all possible 3’-, 5’- and trimmed variants. Functionally equivalent isomiRs can be grouped together according to the type of modification (e.g. uridylation, adenylation, trimming...) to assess which miRNAs are more intensively modified in a given biological context. When applied to the analysis of primary epithelial breast cancer cells, our methodology provided a 40% increase in the number of detected miRNA species and allowed to easily identify and classify more than 1000 variants. Most modifications were compatible with templated IsomiRs, as a consequence of  imprecise Drosha or Dicer cleavage. However, some non-templated variants were consistently found either in the normal or in the cancer cells, with the 3’-end adenylation and uridylation as the most frequent events, suggesting that miRNA post- transcriptional modification frequently occurs. Our analytical tool permits the deconvolution of miRNA heterogeneity and could be used to explore the functional role of miRNA isoforms.

overall_design: A matched normal/tumor primary culture from breast epithelium

first: Matteo

middle: J

last: Marzi

first: Francesco

last: Nicassio


title: Whole Transcriptomic Sequencing of Metastatic Castration Resistant Prostate Cancer Samples

city: Bethesda

first: Adam

middle: G.

last: Sowalsky

address: 37 Convent Drive Building 37 Room 1062B

department: Genitourinary Malignancies Branch

institute: National Cancer Institute

summary: We report the gene expression profile of 8 metastatic castration resisistant prostate cancer samples analyzed by paired-end RNA-seq. We found evidence of extensive abnormal splicing as well as several novel fusion genes. Finally, we also observed several recurrent high-confidence somatic mutations.

overall_design: Paired-end RNA-seq by rRNA depletion

middle: G

first: Zheng

last: Xia

first: Liguo

last: Wang

first: Hao

last: Zhao

first: Shaoyong

last: Chen

first: Glenn

last: Bubley

first: Steven

middle: P

last: Balk

first: Wei

last: Li


title: Study of Huntington disease by analyzing RNA-Seq data using fly and mouse models

city: Houston

first: hao

last: zhao

address: 1 Baylor Plaza

institute: BAYLOR COLLEGE OF MEDICINE

summary: In this study, we perform deep mRNA sequencing to study the effect of a few gene modifiers in Huntington disease using fly models.

sample_organism:
Drosophila melanogaster
Mus musculus

platform_organism:
Drosophila melanogaster
Mus musculus

overall_design: Analysis of 3 fly RNA-Seq samples and 4 mouse RNA-Seq samples

first: Ismael

last: Al-Ramahi

first: Juan

last: Botas


title: Transcriptome profiling of severe spinal muscular atrophy mouse embryonic stem cell-derived motor neurons

city: Philadelphia

first: Matthew

middle: E R

last: Butchbach

state: PA

address: 1020 Locust St

institute: Thomas Jefferson University

summary: Proximal spinal muscular atrophy (SMA) is an early onset, autosomal recessive motor neuron disease caused by loss of or mutation in SMN1 (survival motor neuron 1). Despite understanding the genetic basis underlying this disease, it is still not known why motor neurons (MNs) are selectively affected by the loss of the ubiquitously expressed SMN protein. Using a mouse embryonic stem cell (mESC) model for severe SMA, the RNA transcript profiles (transcriptomes) between control and severe SMA (SMN2+/+;mSmn-/-) mESC-derived MNs were compared in this study using massively parallel RNA sequencing (RNA-Seq). The MN differentiation efficiencies between control and severe SMA mESCs were similar. RNA-Seq analysis identified 3094 upregulated and 6964 downregulated transcripts in SMA mESC-derived MNs when compared against control cells. Pathway and network analysis of the differentially expressed RNA transcripts showed that pluripotency and cell proliferation transcripts were significantly increased in SMA MNs while transcripts related to neuronal development and activity were reduced. The differential expression of selected transcripts such as Crabp1, Crabp2 and Nkx2.2 was validated in a second mESC model for SMA as well as in the spinal cords of low copy SMN2 severe SMA mice. Furthermore, the levels of these selected transcripts were restored in high copy SMN2 rescue mouse spinal cords when compared against low copy SMN2 severe SMA mice. These findings suggest that SMN deficiency affects processes critical for normal development and maintenance of MNs.

overall_design: RNA profiles were generated from FACS-purified control and SMA mESC-derived motor neurons (n=3/genotype) by deep sequencing using Illumina HighSeq 2500

middle: E


title: Loss of MBNL Leads to Disruption of Developmentally Regulated Alternative Polyadenylation in RNA-Mediated Disease

city: La Jolla

first: Ranjan

last: Batra

state: California

address: 2880 Torrey Pines Scenic Drive

department: Cellular and Molecular Medicine

institute: University of California San Diego

summary: Mapping MBNL-regulated genome-wide alternative polyadenylation: We report that depletion of Mbnl proteins in mouse embryo fibroblasts (MEFs), DM mouse model quadriceps muscle, and DM-autopsy muscle tissue leads to mis-regulation of alternative polyadenylation

overall_design:
We compared WT, Mbnl1/2KO, Mbnl1/2KO/3siRNA, and Mbnl1/2KO/scrambled siRNA MEFs (n=2 for each group) to evaluate alternative polyadenylation shifts that occur due to progressive loss of Mbnl proteins. We also compared WT (1 day old, and 4 months old, n=2 each) and HSALR mouse model (4 months old, n=2) of myotonic dystrophy for developmental alternative polyadenylation defects in myotonic dystrophy. Finally, we compared control and DM1 autopsy muscle tissues (n=3) for changes in alternative polyadenylation.
We performed HITS-CLIP analysis of binding sites of Mbnl1, Mbnl2 and Mbnl3 in MEFs (n=3 each). We also performed HITS-CLIP analysis for major skeletal muscle Mbnl protein, Mbnl1 in FVB WT adult muscle (4 months, n=3). Finally we performed HITS-CLIP analysis for CPSF6 in WT and Mbnl1/2 KO MEFs (n=3 each)

Please note that the 'readme_Table.txt' describes the contents of 'Table S*.xlsx' files, and the readme_method.txt include additional details about experiemenal procedures.

first: Konstantinos

last: Charizanis

first: Chaolin

last: Zhang


title: Pseudouridine profiling reveals widespread regulated mRNA pseudouridylation in yeast and human cells

city: Cambridge

first: Thomas

middle: M

last: Carlile

state: Massachusetts

address: 77 Massachusetts Ave

department: Biology

institute: MIT

summary: We report the development of Pseudo-Seq, a sequencing based technique for identification of pseuoduridine (Ψ) modifications within RNA. We performed Pseudo-Seq on S. cerevisiae and HeLa cells and find the first evidence for pseusouridylation of endogenous mRNAs.

sample_organism:
Saccharomyces cerevisiae
Homo sapiens

platform_organism:
Saccharomyces cerevisiae
Homo sapiens

overall_design: RNA was treated with CMC to modify Ψ residues, which cause RT stops. The sites of CMC-dependent RT stops were used to identify sites of pseudouridyation.

first: Wendy

last: Gilbert


title: Explicit DNase sequence bias modeling enables high resolution transcription factor footprint detection

status: Public on Sep 04 2014

city: Durham

first: Christopher

last: Frank

state: NC

address: 101 Science Dr.

institute: Duke University

summary: DNase I footprinting is an established assay for identifying transcription factor (TF)-DNA interactions with single base pair resolution. High throughput DNase-seq assays have recently been used to detect in vivo DNase footprints across the genome. A number of computational approaches have been developed to accurately identify DNase-seq footprints and these methods have been used as a predictor of TF-DNA interactions by itself or in combination with other epigenetic features.  However, recent studies have pointed to a substantial cleavage bias of DNase and its impact on footprinting, casting doubts on its predictive performance. To assess the potential for using DNaseI to identify individual binding sites, we performed DNase-seq experiments on deproteinized naked genomic DNA isolated from two different cell types and determined sequence cleavage bias associated with the DNase-seq protocol. This allowed us to build cleavage bias corrected footprint models specific to individual transcription factors. The predictive performance of these DNase-seq-based binding site models demonstrated that predicted footprints corresponded to high confidence TF-DNA interactions.

overall_design: To quantify the DNase I sequence-dependent cleavage bias, we performed DNase-seq experiments using deproteinized DNA from K562 and MCF7 cell lines.

first: Galip

last: Yardimci

first: Gregory

last: Crawford

first: Uwe

last: Ohler


title: Comparative transcriptomics of Mycobacterium bovis and Mycobacterium marinum

city: Edinburgh

first: Robin

last: Allshire

state: Midlothian

address: Max Born Cresent

institute: The University of Edinburgh

summary: Related surrogate species are often used to study the molecular basis of pathogenicity of a pathogen on the basis of a shared set of biological features generally attributable to a shared core genome consisting of orthologous genes. An important and understudied aspect, however, is the extent to which regulatory features affecting the expression of such shared genes are present in both species. Here we report on an analysis of whole transcriptome maps for an important member of the TB complex Mycobacterium bovis and a closely related model organism for studying mycobacterial pathogenicity Mycobacterium marinum.

sample_organism: Mycobacterium tuberculosis variant bovis

overall_design: Predict transcription start site

first: Pin

last: Tong

first: Brendan

last: Loftus


title: ChIP-Seq experiments of KLF5, GATA4 and GATA6 in YCC3/AGS/KATOIII cell lines

first: Niantao

last: Deng

summary: Three transcription factors KLF5, GATA4 and GATA6 are recurrently amplified in multiple gastric cancer cohorts, representing one type of lineage-survival oncogenes in gastric cancer. ChIP-Seq analysis of these three factors in multiple cell lines revealed that significant number of genomic sites are co-occupied by KLF5 and GATA4 and/or GATA6. Integrative analysis of ChIP-Seq and gene expression identified several targets of the three transcription factors in both cell lines and primary tumors, including HNF4A. These results suggest that KLF5, GATA4 and GATA6 interact and co-operate to regulate HNF4A and other genes to promote tumorigenesis in gastric cancer.

overall_design: ChIP-Seq experiments of KLF5, GATA4 and GATA6 were performed in three gastric cancer cell lines YCC3, AGS and KATOIII

first: Na-Yu

last: Chia

first: Patrick

last: Tan


title: Genome-wide identification and characterization of functional neuronal activity-dependent enhancers

city: Boston

first: Athar

middle: N.

last: Malik

state: MA

address: 200 Longwood Avenue, Goldenson 405

institute: Harvard Medical School

summary: Experience-dependent gene transcription is required for nervous system development and function.  However, the DNA regulatory elements that control this program of gene expression are not well defined.  Here we characterize the enhancers that function across the genome to mediate activity-dependent transcription in neurons. While ~12,000 putative activity-regulated enhancer sequences have previously been identified that are enriched for H3K4me1 and the histone acetyltransferase CBP, we find that this chromatin signature is not sufficient to distinguish which of these regulatory sequences are actively engaged in promoting activity-dependent transcription. We show here that a subset of H3K4me1/CBP positive enhancers that is enriched for H3K27 acetylation (H3K27ac) in vivo, and shows increased H3K27ac upon membrane depolarization of cortical neurons, function to regulate activity-dependent transcription.  The function of many of these activity-regulated enhancers appears to be dependent on the binding of FOS, a protein that had previously been thought to interact primarily with the promoters of activity-regulated genes.  Furthermore, many of these target genes in cortical neurons encode neuron specific proteins that regulate synaptic development and function.  These findings suggest that FOS functions at enhancers to control activity-dependent gene programs that are critical for nervous system function, and provide a resource of activity-dependent enhancers that may give insight into genetic variation that contributes to brain development and disease.

overall_design: Genome-wide maps of H3K27ac and AP1 transcription factors (CFOS, FOSB, JUNB) before and after neuronal activity in mouse cortical neurons.

middle: N


title: Gastric cancer subtyping (Singapore Patient Cohort, batch B)

first: Zhengdeng

last: Lei

address: 8 College Road

institute: Duke-NUS graduate medical school

summary:
Genome-wide mRNA expression profiles of 56 primary gastric tumors from the Singapore patient cohort, batch B. Like many cancers, gastric adenocarcinomas (gastric cancers) show considerable heterogeneity between patients. Thus, there is intense interest in using gene expression profiles to discover subtypes of gastric cancers with particular biological properties or therapeutic vulnerabilities.
Identification of such subtypes could generate insights into the mechanisms of cancer progression or lay the foundation for personalized treatments. Here we report a robust gene-expression-based clustering of a large collection of gastric adenocarcinomas (with GSE15459) from Singaporean patients.

overall_design: Profiling of 56 primary gastric tumors on Affymetrix GeneChip Human Genome U133 Plus 2.0 Array. All tumors were collected with approvals from the National Cancer Centre, Singapore; the Research Ethics Review Committee; and signed patient informed consent.

first: Steve

last: Rozen


title: Dormancy of green-cotyledon Col seeds matured at 20°C vs 15°C

city: Harpenden

first: Dana

middle: Reid

last: MacGregor

state: Hertfordshire

address: Rothamsted Research

department: Biointeractions and Crop Protection

institute: Rothamsted Research

summary: To determine genes involved in induction of dormancy in response to reduced maturation temperatures, RNA sequencing was performed on developing seeds matured at 20°C, where low dormancy is observed, and compared to seeds matured at 15°C where high dormancy is observed.

overall_design: Col green cotyledon seeds matured at 20°C and 15°C were dissected from siliques between five to seven hours after dawn and frozen in liquid nitrogen.  RNA was extracted as described previously (Penfield et al., 2005).

middle: R

first: Steven

last: Penfield


title: Direct roles of SPEECHLESS in the specification of stomatal self-renewing cells [RNA-seq]

city: Stanford

first: On Sun

last: Lau

address: 371 Serra Mall, Rm 210

institute: Stanford University

summary: Lineage-specific stem cells are critical for the production and maintenance of specific cell types and tissues in multicellular organisms. In higher plants, the initiation and proliferation of stomatal stem cells is controlled by the basic helix-loop-helix transcription factor SPEECHLESS (SPCH). The stomatal stem cells and SPCH, which represent an innovation in seed plants, allow flexibility in the production of stomata, but how SPCH generates these stem cells is unclear. Here, we developed a highly sensitive chromatin immunoprecipitation (ChIP) assay and profiled the cell-type specific genome-wide targets of Arabidopsis SPCH in vivo. We found that SPCH directly controls key and novel regulators that drive cell fate and asymmetric cell divisions and enhances responsiveness to cell-cell communication. Our results provide molecular insights on how a master transcription factor generates an adult stem cell lineage that contributes to the success of land plants.

overall_design: RNA-Seq profiles of inducible SPCH and wild-type upon induction

first: Kelli

middle: A

last: Davies

first: Jessica

last: Chang

first: On

middle: S

first: Dominique

last: Bergmann


title: Direct roles of SPEECHLESS in the specification of stomatal self-renewing cells [ChIP-Seq]

overall_design: Genome-wide identification of SPCH binidng sites


title: Insights into the epigenomic landscape of the human malaria vector Anopheles gambiae

city: Armilla

first: Elena

last: Gómez-Díaz

state: Granada

address: Avda. del Conocimiento, 17

department: Institute of Parasitology and Biomedicine Lopez-Neyra (IPBLN)

institute: Spanish National Research Council (CSIC)

summary: We characterize the epigenome of the human malaria vector Anopheles gambiae in midgut cells by mapping the distribution and levels of two post-translational histone modifications, H3K27ac and H3K27me3. These histone profiles were then correlated with levels of gene expression obtained by RNA-seq.

overall_design: ChIP-seq and RNA-seq were performed on adult female A. gambiae midguts. RNA-seq was performed on adult female A. gambiae salivary glands.

first: Ana

last: Rivero

first: Fabrice

last: Chandre

first: Victor

last: Corces


title: Direct roles of SPEECHLESS in the specification of stomatal self-renewing cells

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

overall_design: Refer to individual Series