title: Genome-wide profilings of transcriptome and translatome in mouse hippocampi after contextual fear conditioning

city: Seoul

first: Jun

last: Cho

address: Building 504 Room 509, School of Biological Sciences, Seoul National University, 1 Gwanangno, Gwanak-gu

department: School of Biological Sciences

country: South Korea

institute: Seoul National University

summary: Memory stabilization after learning requires transcriptional and translational regulations in the brain, yet the temporal molecular changes following learning have not been explored at the genomic scale. We here employed ribosome profiling and RNA sequencing to quantify the translational status and transcript levels in mouse hippocampus following contextual fear conditioning. We identified 104 genes that are dynamically regulated. Intriguingly, our analysis revealed novel repressive regulations in the hippocampus: translational suppression of ribosomal protein-coding genes at basal state; learning-induced early translational repression of specific genes; and late persistent suppression of a subset of genes via inhibition of ESR1/ERα signaling. Further behavioral analyses revealed that Nrsn1, one of the newly identified genes undergoing rapid translational repression, can act as a memory suppressor gene. This study unveils the yet unappreciated importance of gene repression mechanisms in memory formation.

overall_design: The application of ribosome profiling and RNA-seq techniques to mouse hippocampi tissues after contextual fear conditioning and to mouse hippocampal primary cultures.  Mouse ESCs were also examined.

first: V

middle: N

last: Kim


title: HDAC inhibition impedes epithelial–mesenchymal plasticity and suppresses metastatic, castration-resistant prostate cancer

status: Public on Oct 07 2015

city: Los Angeles

first: Yu-Ting

last: Tseng

state: California

address: 650 Charles E. Young Drive South

department: Department of Microbiology, Immunology, & Molecular Genetics

country: USA

institute: University of California, Los Angeles

type: Expression profiling by high throughput sequencing

summary: PI3K (phosphoinositide 3-kinase)/AKT and RAS/MAPK (mitogen-activated protein kinase) pathway coactivation in the prostate epithelium promotes both epithelial–mesenchymal transition (EMT) and metastatic castration-resistant prostate cancer (mCRPC), which is currently incurable. To study the dynamic regulation of the EMT process, we developed novel genetically defined cellular and in vivo model systems from which epithelial, EMT and mesenchymal-like tumor cells with Pten deletion and Kras activation can be isolated. When cultured individually, each population has the capacity to regenerate all three tumor cell populations, indicative of epithelial–mesenchymal plasticity. Despite harboring the same genetic alterations, mesenchymal-like tumor cells are resistant to PI3K and MAPK pathway inhibitors, suggesting that epigenetic mechanisms may regulate the EMT process, as well as dictate the heterogeneous responses of cancer cells to therapy. Among differentially expressed epigenetic regulators, the chromatin remodeling protein HMGA2 is significantly upregulated in EMT and mesenchymal-like tumors cells, as well as in human mCRPC. Knockdown of HMGA2, or suppressing HMGA2 expression with the histone deacetylase inhibitor LBH589, inhibits epithelial–mesenchymal plasticity and stemness activities in vitro and markedly reduces tumor growth and metastasis in vivo through successful targeting of EMT and mesenchymal-like tumor cells. Importantly, LBH589 treatment in combination with castration prevents mCRPC development and significantly prolongs survival following castration by enhancing p53 and androgen receptor acetylation and in turn sensitizing castration-resistant mesenchymal-like tumor cells to androgen deprivation therapy. Taken together, these findings demonstrate that cellular plasticity is regulated epigenetically, and that mesenchymal-like tumor cell populations in mCRPC that are resistant to conventional and targeted therapies can be effectively treated with the epigenetic inhibitor LBH589.

overall_design: RNA was extracted from pooled Epithelial, EMT and Mesenchymal-like tumor cells isolated by FACS sorting CD45-CD31-Ter119-EpCAM+GFP-, CD45-CD31-Ter119-EpCAM+GFP+, and CD45-CD31-Ter119-EpCAM-GFP+ cells, respectively, from the prostates of 10-12 week old Pb-Cre+/-;PtenL/L;KrasG12D/+;Vim-GFP (CPKV) mice (n=17) and separated into two technical replicates. Paired-end sequencing data with read lengths of 100 bp were generated using the Illumina HiSeq2000 system.

first: Marcus

last: Ruscetti

first: Eman

middle: L

last: Dadashian

first: Weilong

last: Guo

first: Bill

last: Quach

first: David

middle: J

last: Mulholland

first: Juw

middle: W

last: Park

first: Linh

middle: M

last: Tran

first: Naoko

last: Kobayashi

first: Daniella

last: Bianchi-Frias

first: Yi

last: Xing

first: Pete

middle: S

last: Nelson

first: Hong

last: Wu


title: Transcriptome profiling of the ovine pars tuberalis in long and short photoperiod conditions

status: Public on Oct 09 2015

city: Manchester

first: Shona

middle: Hiedi

last: Wood

address: Oxford road

country: United Kingdom

institute: University of Manchester

summary: Persistent free-running circannual (circa 1-year) rhythms have  evolved in animals to regulate hormone cycles and time annual reproduction.  In mammals, these are synchronized by environmental photoperiod and the  melatonin signal. Long summer-like photoperiods (LP) activates thyrotrophs  and thyroid-stimulating hormone (TSH) expression in the melatonin target  tissue, the pars tuberalis (PT) region of the pituitary gland, driven by the  transcriptional co-activator EYA3. TSH in turn stimulates thyroid hormone  (TH) conversion in hypothalamic ependymal tanycytes, thus timing breeding  seasons. In contrast to our new knowledge defining photoperiodic input, it is  still not known which cells, tissues and pathways generate the underlying  circannual cycle. We used seasonal sheep to characterise the circannual cycle, transferring animals from short winter-like photoperiods (SP)  to prolonged LP conditions, and assaying prolactin to track circannual phase  in individual animals. Using RNA-seq, we profiled PT cells at different phases of  the circannual cycle, and defined extensive changes of cellular-remodeling  pathways and genes encoding synaptic guidance proteins in the PT.

overall_design: Short photoperiod (SP) - 5 samples, Long photoperiod (LP) day 1 - 6 samples, LP day 7 - 6 samples, LP day 28 - 6 samples

middle: H

first: Helen

last: Christian

first: Katarzyna

last: Miedzinska

first: Ben

middle: R

last: Saer

first: Mark

last: Johnson

first: Bob

last: Paton

first: Le

last: Yu

first: Judith

last: McNeilly

first: Julian

last: Davis

first: Alan

middle: D

last: Burt

first: Andrew

last: Loudon


title: Reprogramming of primary metabolism facilitates metabolic engineering of bioactives in tomato fruit (ChIP-Seq)

city: Norwich

first: Ghanasyam

last: Rallapalli

address: Norwich Research Park

institute: The Sainsbury Laboratory

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Phenylpropanoids are derived from phenylalanine and comprise an important class of plant secondary metabolites that include specialized bioactives with medicinal properties, important phytonutrients, a broad range of natural colours, phytoanticipins, phytoalexins and phytoestrogens. A number of transcription factors have been used to upregulate specific branches of phenylpropanoid metabolism, but by far the most effective has been the fruit-specific expression of AtMYB12 in tomato, which resulted in an astonishing 10% of fruit dry weight accumulating as flavonoids and hydroxycinnamates. We show that AtMYB12 not only increases the demand of flavonoid biosynthesis, but also increases the supply of carbon from primary metabolism, and the supply of energy and reducing power, by upregulating glycolysis, the TCA cycle, the oxidative pentose phosphate pathway, fuelling the shikimate and phenylalanine biosynthetic pathways to supply more aromatic amino acids for secondary metabolism. AtMYB12 directly activates at least some genes encoding enzymes of primary metabolism. The enhanced supply of precursors, energy and reducing power achieved by AtMYB12 expression can be harnessed to engineer high levels of novel phenylpropanoids in tomato fruit, offering an effective production system for bioactives and other high value ingredients.

overall_design: ChIP-Seq experiments were done using AtMYB12 tomato fruit at 3-4 days after breaker (when the expression of AtMYB12 is strongly induced).

first: Yang

last: Zhang

first: Eugenio

last: Butelli

first: Saleh

last: Alseekh

first: Takayuki

last: Tohge

first: Jie

last: Luo

first: Prashant

last: Kawar

first: Lionel

last: Hill

first: Angelo

last: Santino

first: Alisdair

last: Fernie

first: Cathie

last: Martin


title: Genome wide map of heterochromatin state in fission yeast Schizosaccharomcyces pombe (S. pombe)

city: Basel

first: Tim

last: Roloff

address: Maulbeerstrasse 66

department: Functional Genomics

country: Switzerland

institute: FMI

summary: Proteins of the conserved HP1 family are elementary components of heterochromatin and are generally assumed to play a central role in creating a rigid heterochromatic network that is densely packed and inaccessible to the transcription machinery. In this study we demonstrate that the fission yeast HP1 protein Swi6 exists as a single highly dynamic population and rapidly exchanges in cis and in trans between different heterochromatic regions. Binding to methylated H3K9 or to heterochromatic RNA decelerates Swi6 mobility. We further show that Swi6 is largely dispensable to maintain heterochromatin domains. In contrast, our results disclose an unexpected role of Swi6 in demarcating constitutive heterochromatin from neighboring euchromatin. Our results are consistent with a stochastic model of heterochromatin and imply that heterochromatin is permissive for transcription throughout the cell cycle. Rather than promoting maintenance and spreading of heterochromatin, Swi6 appears to limit these processes to ensure that heterochromatin is appropriately confined.

overall_design: Examination of a single histone modification in a Swi6 deletion fission yeast strain

first: Raghavendran

last: Kulasegaran-Shylini

first: Dimosthenis

last: Gaidatzis

first: Sophie

last: Dessus-Babus

first: Marc

last: Buehler


title: Human beta cell proliferation induced by inhibition of Dyrk1a and GSK3b

status: Public on Oct 22 2015

city: San Diego

first: John

last: Walker

state: CA

address: 10675 John Jay Hopkins

institute: Genomics Institute of the Novartis Research Foundation

summary:
Purpose: Single-cell whole transcriptome sequencing was used to better understand the mechanism of action of our Dyrk1a inhibitor's proliferation of pancreatic islets.
Methods: primary pancreatic islets were isolated, cultured, and stimulated with either 0.1% DMSO or 3 µM GNF4877.  Single cells were captured and cDNA isolated on a Fluidigm C1 instrument.  Sequencing libraries were made with Nextera XT reagents (Illumina) and single-end 50 bp reads were generated on an Illumina HiSeq 1000.  Reads were mapped to the rat transcriptome.
Results:  Consistent with GNF4877 eliciting beta cell proliferation, we observed an increase in the number of beta cells co-expressing insulin 1 and genes involved in cell cycle including the M phase marker Cyclin B1. Comparison of Cyclin B1 expressing cells from GNF4877-treated islets to beta cells from DMSO-treated islets further revealed a significant increase expression of genes associated with full cell cycle progression and enrichment of Gene Ontology (GO) categories for proliferation.
Conclusions:  Since only a small subset of islet cells proliferate when stimulated with GNF4877, single-cell transcriptome sequencing allowed us to examine expression of genes co-regulated with known proliferation markers and will hopefully allow us to characterize beta cell subsets which are responsive to proliferation-associated therapies.

overall_design: 84 GNF4877-treated and 86 DMSO-treated rat islet cells containing greater than 100,000 mapped sequencing reads per cell and having a single verified cell per port were compared

first: Weijun

last: Shen

first: Andrew

middle: F

last: Powers

first: Ann

middle: E

last: Herman

first: Anwesh

last: Kamireddy

first: Austin

last: Swafford

first: Badry

last: Bursalaya

first: Bernhard

last: Hering

first: Brandon

last: Taylor

first: Carolina

last: Turk

first: Christophe

last: Filippi

first: George

last: Harb

first: Jing

last: Li

last: Lamb

first: Lisa

last: Deaton

first: Martin

last: Seidel

first: Minhua

last: Qiu

first: Peter

last: McNamara

first: Qihui

last: Jin

first: Richard

last: Glynne

first: Bryan

last: Laffitte

first: Shanshan

last: Yan

first: Shelly

last: Meeusen

first: Tingting

last: Mo

first: Tom

last: Wu

first: Van

last: Nguyen-Tran

first: Wilma

last: Suarez-Pinzon

first: Xueshi

last: Hao

first: Yahu

last: Liu

first: You-Qing


title: Reprogramming of primary metabolism facilitates metabolic engineering of bioactives in tomato fruit

first: Ghanasyam

last: Rallapalli

overall_design: WT (MicroTom), AtMYB12, Del/Ros and Indigo trasgenic fruits were harvested after 1 week and 4 weeks after breaker stage. Pericarp from harvested fruits was used for total RNA extraction. mRNA profiles were generated by deep sequencing on Illumina GAIIx using EXPRSS tag-seq protocol.

first: Yang

last: Zhang

first: Eugenio

last: Butelli

first: Saleh

last: Alseekh

first: Takayuki

last: Tohge

first: Jie

last: Luo

first: Prashant

last: Kawar

first: Lionel

last: Hill

first: Angelo

last: Santino

first: Alisdair

last: Fernie

first: Cathie

last: Martin


title: Identification and characterization of circannual calendar cells within the pituitary driving seasonal cycles in mammals

first: Shona

last: Wood

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

overall_design: Refer to individual Series


title: Genome-wide profiling of the  DNA methylation landscape at base-pair resolution in human acute myeloid leukemia cell lines.

city: L'Hospitalet de Llobregat

first: Manel

last: Esteller

state: Barcelona

address: Hospital Duran i Reynals Av. Gran Via s/n km, 2.7

department: PEBC

country: Spain

institute: IDIBELL

type: Methylation profiling by high throughput sequencing

summary: Whole genome bisulphite sequencing of 2 human cancer cell line samples. One cell line harbours a heterozygous mutation in the DNA methyl-transferase DNMT3a, a common alteration observed in AML patients. The main goal is to find the differentially methylated regions (DMR) at genome-wide level between the mutant and a wild-type cell lines to elucidate the effect of the mutation and the implication in leukomogenesis.

overall_design: Sequencing of bisulfite converted DNA of a wild-type  and a DNMT3a mutant AML cancer cell line.

first: Holger

last: Heyn


title: DNA methylation landscape in AML


title: Transcriptome analysis in MDCK cells on anti-influenza A activity of bakuchiol

status: Public on Oct 06 2015

city: Tokushima

first: Takashi

last: Kuzuhara

address: Nishihama, Yamashiro-cho

country: Japan

institute: Tokushima Bunri University

summary: Bakuchiol is a phenolic isoprenoid compound present in Babchi (Psoralea corylifolia Linn.) seeds. We examined the anti-influenza activity of synthetic bakuchiol using Madin-Darby canine kidney (MDCK) cells. We found that the naturally occurring form, (+)-(S)-bakuchiol, and its enantiomer, (-)-(R)-bakuchiol, inhibited influenza A infection and growth, and reduced the expression of viral mRNAs and proteins in MDCK cells. However, the molecular mechanism in anti-influenza activity of bakuchiol remains unclear. Thus, to evaluate its molecular mechanism in host cells, we performed the next generation sequencing analysis of the MDCK transcriptome in cells treated with bakuchiol and influenza A/PR/8/34. These results provide keys to elucidate the molecular mechnism of anti-influenza activity of bakuchiol for us, and these data will contribute to the development of novel approaches to anti-influenza treatments.

overall_design: We used next generation sequence (NGS) to conduct a comprehensive transcriptome analysis in MDCK cells treated with bakuchiol and influenza virus (A/PR/8/34). (+)-(S)-Bakuchiol or (–)-(R)-bakuchiol (25 uM) were mixed with A/PR/8/34 at a MOI of 0.1. DMSO (0.25%) was used as a negative control. Each mixture was added to the MDCK cells and incubated for 24 h before extracting total RNA from the cell lysates. mRNA sequencing libraries were constructed from each total RNA extract using SureSelect Strand Specific RNA Library Preparation Kit. Thirty-six-base-pair, single-end-read RNA sequencing tags were generated using an Illumina Hiseq2500 sequencer. RNA sequencing tags that mapped to the dog reference genome sequences (CanFam3 genome) were analyzed. The reads per kilobase per million mapped reads (RPKM) was calculated for the mRNA transcripts in Ensemble gene. Genes that showed a > 1.5 fold-change in RPKM value in (+)-(S)-bakuchiol-treated MDCK cells are indicated. The complete NGS transcriptome analysis has been deposited in the DNA Data Bank of Japan database (accession number: DRA003499).

first: Masaki

last: Shoji


title: RNA-sequencing analysis of human fibrolamellar hepatocellular carcinoma and 4 maturational stages of the human hepatobiliary system

city: Ithaca

first: Praveen

last: Sethupathy

state: NY

address: T7 006 Vet Research Tower

institute: Cornell University

summary: The etiology of fibrolamellar hepatocellular carcinoma (FL-HCC), a liver cancer occurring increasingly in children to young adults, is poorly understood. We performed high-throughput sequencing on mRNA isolated from a newly developed model of FL-HCC and 4 different maturational lineages of the hepatobiliary system. Our results indicate that the transcriptome of FL-HCC is most similar to that of biliary tree stem cells.

overall_design: mRNA profiles of a newly developed model of FL-HCC and 4 maturational stages of the human hepatobiliary system (biliary tree stem cells, hepatic stem cells, hepatoblasts, and adult hepatocytes) were generated by high throughput sequencing using the Illumina HiSeq 2500.

first: Lola

last: Reid

first: Timothy

middle: A

last: Dinh

first: Sara

last: Selitsky


title: A mechanism for expansion of the regulatory T cell repertoire and its role in enforcing self-tolerance

status: Public on Oct 05 2015

city: New York

first: Alexander

middle: Y

last: Rudensky

department: Immunology

institute: Memorial Sloan Kettering Cancer Center

summary: To investigate the influence of CNS3, a cis-regulatory element in the Foxp3 locus, on the selection of T cell antigen receptor (TCR) repertoire of regulator CD4+ T cells (Treg), we crossed Foxp3ΔCNS3-gfp or control Foxp3gfp mice to DO11.10 TCRβ transgene and Tcra-/+ background. We isolated Treg and conventional CD4+T cells from thymus, spleen and lymph nodes of Foxp3ΔCNS3-gfp DO11.10 TCRβ Tcra-/+ or Foxp3gfp DO11.10 TCRβ Tcra-/+ male littermates, and sequenced the TCRα chains. Analysis of the diversity of Complementary Determining Region 3 (CDR3) of TCRα showed a distinct clustering of CNS3-deficient Treg cells from the CNS3-sufficient ones.

overall_design: DO11.10 TCRβ transgene inhibits the recombination of endogenous Tcrb loci thus restricting TCR repertoire to TCRα chains expressed by T cells. Further limitation of the TCR repertoire was achieved by the presence of one functional Tcra gene. With restricted TCR repertoire, mRNA of TCRα was extracted from Treg and conventional CD4+ T cells for library preparation and high throughput sequencing.

first: Yongqiang

last: Feng


title: Genetic and epigenetic variation in the lineage specification of regulatory T cells

first: Aaron

last: Arvey

institute: Memorial Sloan-Kettering Cancer Center

summary: ChIP-seq of H3K27ac was performed in regulatory T cells (resting and activated) and conventional T cells (naïve, effector, memory) in mouse and human.  A small number of regulatory elements were lineage specific in both mouse and human and represented the 'core' lineage specification program.  Regulatory element acetylation levels were associated with genetic variation in humans and lineage-specific loci were enriched for autoimmune risk-alleles (especially type 1 diabetes) identified in classic and fine-resolution genome-wide association studies.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: H3K27ac ChIP-seq of human and mouse

first: Joris

last: van der Veeken

first: George

last: Plitas


title: Network Modelling using Transcription Sequence Data Reveals Novel MAPK Interactions Important for Drug Resistance

city: Berlin

first: Robert

last: Altwasser

address: Robert-Rössle-Str. 10

department: AG Di Virgilio

country: Germany

institute: Max-Delbrück-centrum

summary: Caspofungin study on A. fumigatus. Applied to the Wild type as well as akuB, mpkA and sakA mutant. RNASeq-data

overall_design: Samples were taken before treatment (0h) as well as 0.5h, 1h, 4h, 8h after treatment for the wild type and the akuB mutant. For the sakA and mpkA mutant, samples were taken at 0h, 1h and 4h after treatment.

first: Vito

last: Valiante

first: Clara

last: Baldin

first: Anne

last: Beauvais

first: Karine

last: Guilloux

first: Reinhard

last: Guthke

first: Olaf

last: Kniemayer

first: Jean

middle: P

last: Latgé

first: Jörg

last: Linde

first: Axel

last: Brakhage


title: JunB control of keratinocyte-mediated inflammation [RNA-seq]

city: Durham

first: Jennifer

middle: Yunyan

last: Zhang

state: NC

address: 3 Genome Ct, rm4130

department: Dermatology

institute: Duke University

summary:
To determne JunB target gene in human keratincoytes
Mice with epidermal deletion of JunB transcription factor displayed a psoriasis-like inflammation. The relevance of these findings to humans and the mechanisms mediating JunB function are not fully understood. Here, we demonstrate that impaired JunB function via gene silencing or overexpression of a dominant negative mutant increased human keratinocyte cell proliferation but decreased cell barrier function. RNA-seq revealed over 500 genes affected by JunB loss-of-function which included an upregulation of an array of proinflammatory molecules relevant to psoriasis. Among these were TNFa, CCL2, CXCL10, IL6R and SQSTM1, an adaptor protein involved in NF-kB activation. ChIP-Seq and gene reporter analyses showed that JunB directly suppressed SQSTM1 through binding to a consensus AP-1 cis-element located around 2 Kb upstream of SQSTM1-trasncription start site. Similar to JunB loss-of-function, SQSTM1-overexpression induced TNFa, CCL2 and CXCL10. Conversely, NF-kB-inhibition genetically with a mutant IkBa or pharmacologically with PDTC prevented cytokine, but not IL6R, induction by JunB-deficiency.  Taken together, our findings indicate that JunB controls epidermal growth, barrier formation and proinflammatory responses through direct and indirect mechanisms, pinpointing SQSTM1 as a key mediator of JunB-suppression of NF-kB-dependent inflammation.

overall_design: 3 indepdent set of priamry human keratinocytes isolated from foreskin skin samples were transfected with nonsilencing control or siRNA oligonucleotides targeting JunB. mRNA was then isolated and used for cDNA library construction followed by RNA-sequencing.

first: Xiaoling

first: Jane

last: Jin

first: Joseph

last: Wu

first: Xiaoxia

last: Qin

last: Streilein

first: Russell

last: Hall


title: Bap1 loss results in EZH2 dependent transformation

first: Barbara

last: Spitzer

address: 411 E 67th St

summary: BAP1 and ASXL1 interact to form a polycomb deubiquitinase complex that removes monoubiquitin from histone H2A lysine 119 (H2AK119Ub). However, BAP1 and ASXL1 are mutated in distinct cancer types, consistent with independent roles in regulating epigenetic state and malignant transformation. Here we demonstrate that Bap1 loss results in increased trimethylated histone H3 lysine 27 (H3K27me3), elevated Ezh2 expression, and enhanced repression of Polycomb Repressive Complex 2 (PRC2) targets. These findings contrast with the reduction in H3K27me3 seen with Asxl1 loss. Conditional deletion of Bap1 and Ezh2 in vivo abrogates the myeloid progenitor expansion induced by Bap1 loss alone.

overall_design: Whole bone marrow cells from mice treated with polyI:polyC at three weeks of age were extracted from Bap1 f/f  and Mx1-Cre negative Bap1f/f littermates. Bone marrow cells were red cell lysed and cells were sorted for granulocyte macrophage progenitors (GMP). Genomic DNA was extracted for ChIP-Seq studies to assess transcriptional changes with Bap1 loss in GMPs.

first: Lindsay

last: LaFave

first: Richard

last: Koche

first: Ross

last: Levine


title: RNA-Seq and ChIP-Seq reveals SQSTM1/p62 as a key mediator of JunB suppression of NF-kB-dependent inflammation


title: Pyronema confluens circadian rhythm analysis

city: Bochum

first: Minou

last: Nowrousian

address: Universitaetsstr. 150

department: Molecular and Cellular Botany

institute: Ruhr-University Bochum

summary: The aim of this analysis was the identification of candidate genes that might be regulated in a circadian manner in the filamentous ascomycete Pyronema confluens. The fungus was grown in submerged culture to promote vegetative growth, and after a period in constant light, the samples were shifted to darkness (DD). Cultures were sampled after 24 and 36 hours in DD. The samples were used for RNA-seq analysis, and candidate genes with significantly higher expression after 24 vs. 36 hours or the reverse were further analyzed with RT-qPCR.

overall_design: For each growth condition (DD24 and DD36), three independent biological replicates were performed and sequenced.

first: Stefanie

last: Traeger


title: In beta-actin knockout mouse embryonic fibroblasts epigenetic reprogramming and chromatin modifications lead to rDNA transcription inactivation and defects in cell growth and proliferation

status: Public on Oct 04 2015

city: Abu Dhabi

first: Piergiorgio

last: Percipalle

state: Abu Dhabi

address: Saadiyat Campus, Abu Dhabi

department: Biology Department, Science Division

country: United Arab Emirates

institute: New York University Abu Dhabi

summary: Actin and nuclear myosin 1 (NM1) are emerging as regulators of transcription and chromatin organization. Using a genome-wide approach we report here that β-actin binds both intergenic and genic regions across the entire mammalian genome, associated with both protein-coding and rRNA genes. Across the rDNA transcription unit the distribution of β-actin correlated with NM1 and the other subunits of the B-WICH complex, WSTF and SNF2h. In β-actin-/- mouse embryonic fibroblasts (MEFs), we found that rRNA synthesis levels are down-regulated concomitantly with drops in Pol I and NM1 occupancies across the rRNA gene. In knock-in experiments, reintroduction of wild-type β-actin but not mutated forms with polymerization defects, rescued rRNA synthesis underscoring the direct role for β-actin in Pol I transcription and the importance of polymerization during the transcription process. The rRNA defects in the β-actin-/-MEFs are accompanied by epigenetic reprogramming that leads to up-regulation of the repressive mark H3K4me1. In addition, a high resolution chromatin accessibility assay showed that the absence of β-actin leads to a more compact chromatin at specific locations, including promoter-proximal enhancer (T0 sequence) and Sal boxes (T1-T10), disturbing binding of the transcription termination factor 1 (TTF1). We propose a novel mechanism where the polymerase-associated β-actin synergizes with NM1 to coordinate the establishment of permissive chromatin with the correct rDNA topology for Pol I transcription enhancement.

overall_design: Association of beta-actin with the genome in mouse embryonic fibroblasts.

first: Bader

last: Almuzzaini

first: Ann

middle: K

last: Östlund Farrants


title: Chromatin dynamics and the RNA exosome function in concert to regulate transcriptional homeostasis

status: Public on Oct 02 2015

city: Berkeley

first: Xavier

last: Darzacq

address: 475D Li Ka Shing Center

department: Molecular and Cell Biology

institute: University of California, Berkeley

summary: The histone variant H2A.Z is a hallmark of nucleosomes flanking the promoters of protein coding genes, and is often found in nucleosomes that also carry lysine 56- acetylated histone H3 (H3-K56Ac), a mark which promotes rapid replication- independent turnover of nucleosomes. Although H2A.Z and H3-K56Ac have been generally implicated in transcriptional activation, their exact contributions have remained elusive. Here we find that H3-K56Ac promotes RNA polymerase II occupancy at a large number of protein coding and noncoding loci, yet neither H3- K56Ac nor H2A.Z has a significant impact on steady state mRNA levels in yeast. Instead, broad effects of H3-K56Ac or H2A.Z on levels of both coding and noncoding RNAs (ncRNAs) are only revealed in the absence of the nuclear RNA exosome. H2A.Z is also necessary for expression of divergent, promoter-proximal ncRNAs in mouse embryonic stem cells, suggesting a conserved role for H2A.Z across eukaryotes. Finally, we show that H2A.Z functions with H3-K56Ac in chromosome folding, facilitating formation of chromosome interaction domains (CIDs). Our study suggests that H2A.Z and H3-K56Ac work in concert with the RNA exosome to control mRNA and ncRNA expression, perhaps in part by regulating higher order chromatin structures.

overall_design: Two replicates of swr1d

first: Oliver

last: Rando

first: Craig

last: Peterson


title: Engineered mammalian RNAi can elicit antiviral protection that negates the requirement for the interferon response

status: Public on Oct 03 2015

city: Tarrytown

first: Asiel

middle: Arturo

last: Benitez

state: New York

address: 777 Old Saw Mill River Rd, 92-516A

department: Immunology & Inflammation

institute: Regeneron Pharmaceuticals, Inc

summary: While the intrinsic antiviral cell defenses of many kingdoms utilize pathogen-specific small RNAs, the antiviral response of chordates is primarily protein-based and not uniquely tailored to the incoming microbe. In an effort to explain this evolutionary bifurcation, we determined whether antiviral RNA interference (RNAi) was sufficient to replace the protein-based type I interferon (IFN-I) system of mammals. To this end, we recreated an RNAi-like response in mammals and determined its effectiveness to combat influenza A virus in vivo in the presence and absence of the canonical IFN-I system. Mammalian antiviral RNAi, elicited by either host- or virus-derived small RNAs, effectively attenuated virus and prevented disease independently of the innate immune response.  These data find that chordates could have utilized RNAi as their primary antiviral cell defense and suggest that the IFN-I system emerged as a result of natural selection imposed by ancient pathogens.

overall_design: WT C57BL/6 and Ifnar1-/- C57BL/6 were infected with WT Influenza A virus (IAV-WT) or a miRNA-targeted virus (IAV-NPt).  RNA from lungs was harvested 2 and 9 days post infection.

first: Bejamin

last: tenOever


summary: Analysis of sorted granulocyte macrophage progenitors (GMPs) in control and Bap1-deficient bone marrow cells. Loss of Bap1 in the hematopoietic compartments results in an MDS-like disease. These data allow for the examination of the genetic underpinnings of Bap1 loss in disease.

overall_design: Whole bone marrow cells from mice treated with polyI:polyC at three weeks of age were extracted from Mx1-Cre positive and Mx1-Cre negative Bap1f/f littermates. Bone marrow cells were red cell lysed, stained and sorted for GMP cells (Lineage-c-Kit+Sca1-CD34+Fc?). RNA was extracted for RNA-Seq studies to assess transcriptional changes with Bap1 loss in GMPs.

first: Nick

last: Socci


title: Next Generation Sequencing Facilitates Quantitative Analysis of Endometrium Transcriptomes During Porcine Embryo Implantation.

city: Jinan

first: Haichao

last: Lin

state: Shandong

address: Sangyuan Road 8.

country: China

institute: Institute of animal science and veterinary medicine Shandong academy of agricultural sciences

summary: In this study, deep sequencing of endometrial RNA samples during early pregnancy were performed using RNA sequencing (RNA-Seq). The changes of genes expressed among three stages (day 12, 18 and 25) of pregnancy were investigated by the multiple comparisons. Analysis of reads, between 1557 and 8951 differentially expressed genes (DEGs) were revealed among different periods of implantation. Expression of selected genes was validated by the use of quantitative real-time RT-PCR. Bioinformatics analysis of differentially expressed genes in the endometrium revealed a number of biological processes and pathways potentially involved in embryo implantation in the pig, most noticeably cell proliferation, regulation of immune response, interaction of cytokine-cytokine receptor, and cell adhesion. This revealed specific gene expression patterns reflecting the different functions of the endometrium on three stages (maternal recognition, conceptus attachment and embryo implantation). This study identified comprehensive transcriptomic profile in the porcine endometrium, it could be a foundation for targeted studies of genes and pathways potentially involved in abnormal endometrial receptivity and embryo loss in early pregnancy.

overall_design: Porcine endometrium mRNA profiles during early pregnancy were generated by deep sequencing using Illumina Hiseq2000.

first: Jianfeng

last: Guo


title: Planarian yorkie/YAP functions to integrate adult stem cell proliferation, organ homeostasis, and maintenance of axial patterning

city: Portland

first: Bret

last: Pearson

state: OR

address: 3181 SW Sam Jackson Park Rd., BRB Room 305

department: Pediatrics

institute: Oregon Health and Science University

summary: During adult homeostasis and regeneration, the freshwater planarian must accomplish a constant balance between cell proliferation and cell death, while also maintaining proper tissue and organ size and patterning. Yet how these ordered processes are precisely modulated, remains relatively unknown.  Here we show that planarians use the downstream effector of the Hippo signalling cascade, yorkie (yki; YAP in vertebrates) to control a diverse set of pleiotropic processes in organ homeostasis, stem cell regulation, regeneration, and axial patterning.  We show that yki functions to maintain the homeostasis of the planarian excretory (protonephridial) system and to limit stem cell proliferation, while not affecting the differentiation process or cell death.  Finally, we show that yki synergizes with WNT/β-catenin signalling to repress head determination by limiting the expression domains of posterior WNT genes and the WNT-inhibitor notum.  Together, our data show that yki is a key factor in planarians that integrates stem cell proliferation control, organ homeostasis, and the spatial patterning of tissues.

overall_design: RNA deep sequencing (RNAseq) was performed on whole uninjured animals 6 days following 3 RNAi feedings (3fd6) for control(RNAi) and yki(RNAi).  Experiments were performed in biological-triplicate, sequenced to a depth of 50 million reads per sample, and multiplexed on an Illumina HiSeq2500 with 50 basepair, single-end reads.

first: Alexander


title: Hypoxia-induced TET-TNFa-p38-MAPK signaling axis drives tumor malignancy of human breast cancer

city: La Jolla

first: Christopher

last: Benner

address: 9500 Gilman Dr. MC 0640

department: Medicine

institute: University of California, San Diego (UCSD)

summary: Hypoxia, a hallmark of most solid tumors, leads to aberrations in epigenetic modifications promoting malignant tumor phenotypes, including metastatic features and stem cell-like characteristics. Aberrant DNA methylation has been considered to play an essential role during tumor progression and tightly associate with tumor malignancy. However, the mechanism by which hypoxia alters DNA methylation to promote tumor malignancy remains poorly understood. Ten-eleven translocation 1-3 (TET1-3) proteins, which catalyze the conversion of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), play a critical role in the DNA demethylation that controls different biological processes. Here we demonstrate that the expression of TET1 and TET3 is closely associated with tumor hypoxia, tumor malignancy and poor prognosis of patients with breast cancer. Hypoxia results in deregulation of TET1 and TET3, leading to breast tumor initiating cell (BTIC) properties. Mechanically, hypoxia regulates expression of TET1 and TET3 via hypoxia-inducible factor-1a (HIF-1a), thereby resulting in 5hmC genome-wide changes, which in turn leads to the upregulation of TNFa expression and activation of its downstream p38-MAPK pathway. Importantly, signal transduction through the TET-TNFa-p38-MAPK signaling axis is required for the acquisition of BTIC characteristics and chemotherapy resistance, leading to more malignant tumor phenotypes. Inhibition of the hypoxia-TET-TNFα-p38-MAPK signaling pathway results in compromised BTIC properties and tumorigenicity in vitro and in vivo, suggesting a possible therapeutic strategy.

overall_design: Examination of 5hmc profile in MCF7 breast cancer cell line with or without hypoxia treatment using hMeDIP-Seq

first: Min-Zu

last: Wu

first: Juan

last: Carlos

first: Izpisua

last: Belmonte


title: RNA-SEQ Reveals Changes in the Staphylococcus aureus Transcriptome following blue light illumination

city: Waco

first: Tamarah

middle: L

last: Adair

state: Tx

address: One Bear Place 97388

department: Biology

institute: Baylor University

type: Non-coding RNA profiling by high throughput sequencing

summary:
Purpose: In an effort to better understand the mechanism of blue light inhibition in Staphylococcus aureus, a whole transcriptome analysis of S. aureus isolate BUSA2288 was performed using RNA-seq to analyze the differential gene expression in response to blue light exposure.
Methods: RNA was extracted from S. aureus cultures pooled from 24 one ml well samples illuminated with a dose of 250 J/cm2 of 465 nm blue light, and from   control cultures grown in the dark.  Complementary DNA (cDNA) were generated from the RNA samples and then sequenced using the Illumina MiSeq Next Generation Sequencer.  The combined results of 2 independent experiments were analyzed using the Pairwise Analysis tools in GeneSifter®. The genes were normalized by Mapped Reads using EdgeR statistics including a Benjamini and Hochberg false discovery rate correction.  Quality was set at a minimum number of 10 reads and a p value of 0.05. The lower threshold for change was 5 fold.
Results: Transcriptomic comparisons using a cutoff of 5 fold identified a total of 28 down-regulated genes and 6 up-regulated genes in the samples that were exposed to blue light.  All but 6 of the differentially regulated genes fall into 8 functional categories: amino acid    biosynthesis, cell envelope components, cellular processes, central intermediary metabolism, energy metabolism, protein synthesis, regulatory function, and transport and binding proteins. Five genes encode conserved proteins of unknown function.
Conclusions: Blue light has been shown to be bactericidal against S. aureus and is a potential alternative therapy for antibiotic resistant organisms.  The mechanism for the inactivation of bacteria is hypothesized to involve ROS. We have found evidence that supports this hypothesis that involves multiple pathways including essential metabolic pathways and known virulence pathways.

overall_design: S. aureus mRNA profiles were produced from 24 pooled samples of S. aureus cultures exposed to blue light (BL) and compared to parallel samples grown in the dark (NL). Two independent experiments were performed.

first: Bayless

last: Drum


title: Fixed single-cell transcriptomic characterization of human radial glial diversity

city: Seattle

first: Zizhen

last: yao

state: WA

department: MAT

institute: Allen Institute for Brain Science

summary: The human neocortex is created from diverse progenitors that are intermixed with multiple cell types in the prenatal germinal zones. These progenitors have been difficult to profile with unbiased transcriptomics since progenitors-particularly radial glia (RG)-are rare cell types, defined by a combination of intracellular markers, position and morphology. To circumvent these problems, we developed a method called FRSCR for transcriptome profiling of individual fixed, stained, and sorted cells. After validation of FRSCR with human embryonic stem cells, we profiled primary human RG that constitute only 1% of the mid-gestation cortex. These data showed that RG could be classified into ventricle zone-enriched RG (vRG) that expressed ANXA1 and CRYAB, and outer subventricular zone-localized RG (oRG) that expressed HOPX. Our study identified the first markers and molecular profiles of vRG and oRG cells, and provides an essential step for understanding molecular networks that control the development and lineage of human neocortical progenitors. Furthermore, FRSCR allows targeted single-cell transcriptomic profiling of many tissues that currently lack live-cell markers.

overall_design: 26 Llive and 19 Fixed cultured hESCs were prepared and sequenced using both FRISCR and TritonX-100 Lysis as proof of principal for FRSCR.

first: Boaz

last: Levi

first: John

last: Mich

last: Yao

first: Elliot

last: Thomsen

first: Rebecca

last: Hodge

first: Adele

last: Doyle

first: Sumin

last: Jang

first: Soraya

last: Shehata

first: Angelique

last: Nelson

first: Nadiya

middle: V

last: Shapovalova

first: Sharad

last: Ramanathan


title: MiRNA analysis of rice seed germination

city: Wuhan

first: Dongli

last: He

state: HuBei

address: Wuchang

department: plant breeding

institute: Wuhan Botanic Garden

summary: In this study, we sequenced the sRNAs population from embryos of 0, 12, and 24 HAI rice seeds using next-generation deep sequencing technology. A series of miRNAs were identified, including both known and novel miRNAs. We also predicted the potential targets for the miRNAs. RT-PCR and 5’ RACE assay were performed to confirm some deep sequencing and target prediction results. This study provides the unique composition and expressional profiles of miRNAs and their potential regulations in the embryo at the early stages of rice seed germination.

overall_design: Examination of  miRNA expressional profiles in rice embryo during early stages of seed germination

first: Qiong

last: Wang

first: Kun

first: Pingfang

last: Yang


title: CLIP-seq analysis of cytoplasmic poly(A) binding protein-1 reveals binding to genomically encoded sequences within mammalian mRNAs

city: Philadelphia

first: Ian

last: Silverman

state: Pennsylvania

address: 433 South University Ave, Room 104

institute: University of Pennsylvania

summary: Pabpc1 is the major cytoplasmic poly(A) binding protein in mammalian cells. Pabpc1 functions have been characterized predominantly in context of its binding to 3’ poly(A) tails of mRNAs. Here we performed CLIP-seq to identify additional Pabpc1 binding sites within mammalian mRNAs that may impact on gene regulation. Our analysis revealed that Pabpc1 binds directly to the canonical polyadenylation signal on thousands of mRNAs in the mouse transcriptome. Pabpc1 binding was also observed at sites coincident with the translational initiation and termination sites bracketing open reading frames, exemplified by non-polyadenylated replication-dependent histone mRNAs. A less abundant set of Pabpc1 interactions were mapped to A-rich sites within 5’ UTRs in a restricted subset of mRNAs including Pabpc1 itself. Mechanistic analyses of the subset of 5’UTR-Pabpc1 interactions revealed evidence for auto-regulatory and trans-regulatory translational control mediated by defined A/U-rich elements. These data, in their entirety, demonstrate that the repertoire of Pabpc1 binding and actions is substantially broader than previously recognized and has the potential to impact and coordinate post-transcriptional controls over a critical to an array of cellular functions

overall_design:
Pabpc1 CLIP-seq (3 replicates)
mRNA-seq (2 replicates)

first: Hemant

last: Kini

first: Brian

last: Gregory

first: Stephen

last: Liebhaber


title: The miRNome of Fasciola hepatica juveniles endorse the existence of a reduced set of highly divergent miRNAs in parasitic flatworms.

city: Montevideo

first: José

middle: Francisco

last: Tort

state: Montevideo

address: Avenida Gral. Flores 2125

department: Department of Genetics

country: Uruguay

institute: Faculty of Medicine, UdelaR

summary: The liver fluke Fasciola hepatica is a foodborne zoonotic parasite affecting livestock worldwide with increasing relevance in human health. The first developmental stage that the host meets after ingestion of the parasite is the newly excysted juvenile (NEJ), that actively transverse the gut wall and migrates to their final location in the liver. The regulation of the early developmental events in NEJs is still poorly understood and a relevant target for control strategies. Here we investigated the putative involvement of small regulatory RNAs in the invasion process. The small RNA population of the NEJ fall into two classes, one represented by miRNAs and a secondary group of larger (32- 33 nucleotides) tRNA derived sequences. We identified more than 30 different miRNAs most of them belonging to ancient miRNAs conserved in protostomes and metazoans, notably with an miR-125b variant as highly predominant. Remarkably, several protostomian and metazoan conserved families were not detected in consonance with previous reports of drastic miRNome reduction in parasitic flatworms. Additionally, a set of 11 novel miRNAs was identified, probably associated with specific gene regulation expression needs in F. hepatica. While sequence conservation in mature miRNA is high across the metazoan tree, we observed that flatworm miRNAs are more divergent suggesting that mutation rates in parasitic flatworms could be high. Finally, the distinctive presence of tRNA derived sequences, mostly 5' tRNA halves of selected tRNAs in the small RNA population of NEJs could indicate that this parasite uses both miRNA and tRNA fragments for the regulation of gene expression.

overall_design: SmallRNA profile of Fasciola hepatica juvenile stage generated by deep sequencing using illumina GAIIx

first: Santiago

last: Fontenla

first: Nicolas

last: Dell'Oca

first: Pablo

last: Smircich

first: Mar

last: Siles-Lucas


title: RNAseq in the whole whole freshwater planarian, Schmidtea mediterranea, following RNAi of lhx1/5-1, pitx or yorkie

first: Bret

middle: J

last: Pearson

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

overall_design: Refer to individual Series


title: Comparative analysis of Rtf1- and PAF1C-regulated genes

status: Public on Oct 01 2015

city: Yokohama

first: Yuki

last: Yamaguchi

address: 4259 Nagatsuta

department: Life Science and Technology

institute: Institute of Science Tokyo

summary: Paf1 and Ski8 were selected as representative subunits of the Paf1 complex (PAF1C), and RNA-seq analysis was performed in triplicate to compare the genes affected by Paf1, Ski8, and Rtf1 knockdown in HeLa cells.

overall_design: Total RNA was harvested from control HeLa and Ski8 knockdown cells at day 4 and from Rtf1 or Paf1 knockdown cells at day 7 and was subjected to RNA-seq in triplicates.

first: Qing-Fu

last: Cao

first: Junichi

last: Yamamoto

first: Tomoyasu

last: Isobe

first: Shumpei

last: Tateno

last: Murase

first: Yexi

last: Chen

first: Hiroshi

last: Handa


title: Synergistic gene expression signature observed in TK6 cells upon co-exposure to UVC-irradiation and protein kinase C-activating tumor promoters

city: Wilmington

first: Zhongqiang

state: DE

institute: E.I. du Pont de Nemours and Company


title: Method for accurate epigenome profiling using a few hundred cells and its applications

city: Baltimore

first: Yixian

last: Zheng

state: MD

address: 3520 San Martin Drive

department: Embryology

institute: Carnegie Institution for Science

summary: Difficulties to accurately map epigenomes in a few cells sorted or dissected from tissues have hampered our understanding of how chromatin modification regulates development and diseases. Despite recent progress, all reported chromatin-immunoprecipitation-based deep sequencing (ChIP-seq) methods have not achieved high quality mapping of rare cell populations. We report Recovery via Protection (RP)-ChIP-seq and favored amplification RP-ChIP-seq (FARP-ChIP-seq) for as few as 500 cells with superior quality compared to all reported techniques to date. FARP-ChIP-seq accurately mapped histone H3 lysine 4 trimethylation (H3K4me3) and H3K27me3 in long-term hematopoietic stem cells (LT-HSCs), short-term HSCs (ST-HSCs), and multi-potent progenitors (MPPs) sorted from one mouse. These high quality datasets not only implicate genes involved in HSC differentiation but also demonstrate a general lack of H3K4me3/H3K27me3 bivalency on hematopoietic genes in HSCs. Thus the method offers accurate mapping for fewest cells.

overall_design: two H3K4me3 replications for mESC, two to three replications of H3K4me3 and H3K27me3 for LT-HSC, ST-HSC and MPP, 2 replicates of H3K4me3 for young and old lens

first: Xiaobin

first: Sibiao

last: Yue


title: Canonical poly(A) polymerase activity promotes the decay of a wide variety of mammalian nuclear RNAs

city: Edinburgh

first: Stefan

last: Bresson

state: Scotland

address: Max Born Crescent, Swann 5.1

department: Wellcome Trust Centre for Cell Biology

institute: University of Edinburgh

summary: The human nuclear poly(A)-binding protein PABPN1 has been implicated in the decay of nuclear noncoding RNAs (ncRNAs). In addition, PABPN1 stimulates hyperadenylation by poly(A) polymerase, and this activity is thought to be required for decay. Here, we inactivated hyperadenylation by two distinct mechanisms and examined changes in gene expression in HEK293 cells by RNAseq. We observed the upregulation of various ncRNAs, including snoRNA host genes, primary miRNA transcripts, and upstream antisense RNAs, confirming that hyperadenylation is broadly required for the degradation of PABPN1-targets. In addition, we found that mRNAs with retained introns are susceptible to PABPN1 and PAPα/γ-mediated decay (PPD). Transcripts are targeted for degradation due to inefficient export, which is a consequence of reduced intron number or incomplete splicing. We conclude that PPD is an important mammalian nuclear RNA decay pathway for the removal of poorly spliced and nuclear-retained transcripts.

overall_design: Poly(A)+ RNA from HEK293 cells was analyzed by next generation sequencing following depletion of PAPα and PAPγ or expression of a dominant negative allele of PABPN1 (LALA) designed to inhibit polyadenylation. For each condition, we collected both total RNA and a nuclear-enriched sample. Each sample was collected in duplicate.

middle: M

first: Olga

last: Hunter

first: Allyson

middle: C

first: Nicholas

last: Conrad


title: JunB ChIP of human keratinocytes [ChIP-seq]

city: Durham

first: Jennifer

last: Zhang

institute: Duke University

summary:
This study is designed to determine the direct target genes of JunB transcription factor in human keratinocytes
Mice with epidermal deletion of JunB transcription factor displayed a psoriasis-like inflammation. The relevance of these findings to humans and the mechanisms mediating JunB function are not fully understood. Here, we demonstrate that impaired JunB function via gene silencing or overexpression of a dominant negative mutant increased human keratinocyte cell proliferation but decreased cell barrier function. RNA-seq revealed over 500 genes affected by JunB loss-of-function which included an upregulation of an array of proinflammatory molecules relevant to psoriasis. Among these were TNFa, CCL2, CXCL10, IL6R and SQSTM1, an adaptor protein involved in NF-kB activation. ChIP-Seq and gene reporter analyses showed that JunB directly suppressed SQSTM1 through binding to a consensus AP-1 cis-element located around 2 Kb upstream of SQSTM1-trasncription start site. Similar to JunB loss-of-function, SQSTM1-overexpression induced TNFa, CCL2 and CXCL10. Conversely, NF-kB-inhibition genetically with a mutant IkBa or pharmacologically with PDTC prevented cytokine, but not IL6R, induction by JunB-deficiency.  Taken together, our findings indicate that JunB controls epidermal growth, barrier formation and proinflammatory responses through direct and indirect mechanisms, pinpointing SQSTM1 as a key mediator of JunB-suppression of NF-kB-dependent inflammation.

overall_design: Genomic DNA isolated from human keratinocytes were used for immunoprecipitation with an antibody against JunB.

first: Xiaoling

first: Jane

last: Jin

first: Joseph

last: Wu

first: Xiaoxia

last: Qin

first: Robert

last: Streilein

first: Russell

last: Hall


title: Perturbations of PIP3 signaling trigger a global remodeling of mRNA landscape and reveal a transcriptional feedback loop

city: Cambridge

first: Felix

last: Krueger

address: Granta Park

department: Bioinformatics

institute: Altos Labs

summary: PIP3 is synthesized by PI3Ks and regulates complex cell responses, such as growth and migration. Signals that drive long-term reshaping of cell phenotypes are difficult to resolve because of complex feedback networks that operate over extended times. It is clear PIP3-dependent modulation of mRNA accumulation is important in this process but is poorly understood. We have quantified the genome-wide mRNA-landscape of non-transformed, breast epithelium-derived MCF10a cells and its response to transient (EGF or PI3Ka-selective inhibitor) or chronic (isogenic cells expressing an oncomutant PI3Ka allele or lacking the PIP3-phosphatase /tumour-suppressor, PTEN) perturbations of PIP3.These results show that whilst many mRNAs are changed by long-term  genetic perturbation of PIP3 signaling (“butterfly effect”), a much smaller number change with a directional logic that aligns with different PIP3 perturbations, allowing discrimination of more directly regulated mRNAs. Our results also indicate that mRNAs can be differentially sensitive to specific features of PIP3 signals, that PIP3-sensitive mRNAs encode PI3K pathway components and identify the transcription factor binding motifs SRF and PRDM1 as important regulators of PIP3-sensitive mRNAs involved in cell movement.

overall_design: RNA-seq on WT MCF10a, treated or not with A66 (Pi3Kalpha inhibitor), PIK3CA H1047R MCF10a and PTEN KO MCF10a. EGF time course stimulation applied (0, 15, 40, 90, 180, 300 min). A66 no EGF when A66 was applied for 300min w/o EGF simulation. All samples made in triplicate. Total of 75 samples.

first: Vladimir

last: Kiselev

first: Veronique

last: Juvin

first: Mouhannad

last: Malek

last: Luscombe

first: Phillip

last: Hawkins

first: Nicolas

last: Le Novère

first: Len

last: Stephens


title: Synergistic gene expression signature observed in TK6 cells upon co-exposure to UVC-irradiation and protein kinase C-activating tumor promoters, a dosage study (study 2/2)

first: Zhongqiang

last: Chen

summary: The purpose of this study, including this dosage series and another  time course series, was to establish gene expression signatures representing the interaction of pathways deregulated by tumor promoting agents and pathways induced by DNA damage.  Human lymphoblastoid TK6 cells were pretreated with the protein kinase C activating tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) and exposed to UVC-irradiation.  The time and dose-responsive effects of the co-treatment were captured with RNA-sequencing (RNA-seq) in two separate experiments.  TK6 cells exposed to both TPA and UVC had significantly more genes differentially regulated than the theoretical sum of genes induced by either stress alone, thus indicating a synergistic effect on global gene expression patterns.  Further analysis revealed that TPA+UVC co-exposure caused synergistic perturbation of specific genes associated with p53, AP-1 and inflammatory pathways important in carcinogenesis.  The 17 gene signature derived from this model was confirmed with other PKC-activating tumor promoters including phorbol-12,13-dibutyrate, sapintoxin D, mezerein, (-)-Indolactam V and resiniferonol 9,13,14-ortho-phenylacetate (ROPA) with quantitative real-time PCR (QPCR).  Here we show a novel gene signature that may represent a synergistic interaction in the tumor microenvironment that is relevant to the mechanisms of chemical induced tumor promotion.

overall_design: mRNA profiles of TK6 cells treated with tumor promoting agent at different dose, co-exposed to UVC

first: Kyle

last: Glover

first: Lauren

last: Markell

first: Xing

last: Han


title: Atrx facilitate normal neuronal differentiation

city: Houston

first: Haoqiang

last: Ying

state: Texas

address: 1515 Holcombe Blvd

institute: M. D. Anderson Cancer Center

summary: Purpose: the chromatin-remodeler ATRX is frequently mutated in a range of human malignancies including pediatric and adult gliomas, and its loss underlies induction of alternative lengthening of telomeres (ALT) pathway for chromosome-end maintenance. But how ATRX exters its glioma suppresor role remains unclear. Here, we combine RNAi and somatic deletion approaches to demonstrate ATRX as a glioma suppressor controlling neural differentiation. Loss of ATRX promotes gliomagenesis by blocking neuronal progenitors from undergoing terminal maturation. Our findings provide the molecular and cellular mechanism underline ATRX's function in gliomagenesis.

overall_design: Examined gene expression profiles of three Atrx-deleted neural stem cells and two control ones cultured in two differentiation conditions.

first: Fei

last: Li

first: Jin

first: Jun

last: Yao

first: Hongwu

last: Zheng


title: Conservation and modification of genetic and physiological toolkits underpinning diapause in bumble bee queens

city: State college

first: Etya

last: Amsalem

address: Orchard Road  University Park

department: Entomology

institute: Pennsylvania State University

summary: Here, we examined the transcriptomic changes associated with diapause and CO2 treatment in B. terrestris queens before, during and post diapause and in the short and long term after CO2 treatment

overall_design: Examination of transcriptomic changes in 25 queens assinged to 5 different groups: 5-10 days old mated, 5-10 days old mated right after CO2 treatment, 2.5 months old diapausing queens that were kept under 4 C for 9-10 weeks and two different egg-layer founders: 4 months old, post diapause and 1.5 months old post CO2 treatment

first: David

last: Galbraith

first: Jonathan

last: Cnaani

first: Peter

last: Teal

first: Christina

last: Grozinger


title: Rice gene expression in irrigated and rainfed fields during two seasons

city: New York

first: Christoph

last: Hafemeister

address: 101 Avenue of the Americas

institute: New York Genome Center

summary: To identify the concurrent effects of multiple environmental factors on gene expression under natural climatic fluctuations in rice, we examined global gene expression patterns in two fields typical of the main modes of rice cultivation, over two seasons, each comprising 15 time-points in three different genotypes

overall_design: Two time-series of 15 time-points, one for each season (dry and wet) over two different fields

first: Anne

last: Plessis

first: Richard

last: Bonneau

first: Michael

last: Purugganan


title: Dose-dependent role of the cohesin complex in normal and malignant hematopoiesis [RNA-Seq]

city: Boston

first: James

last: Bradner

state: MA

address: 450 Brookline

department: Medical Oncology

institute: Dana-Farber Cancer Institute

summary: Cohesin complex members have recently been identified as putative tumor suppressors in hematologic and epithelial malignancies. The cohesin complex guides chromosome segregation, however cohesin-mutant leukemias do not show genomic instability. We hypothesized reduced cohesin function alters chromatin structure and disrupts cis-regulatory architecture of hematopoietic progenitors.  We investigated the consequences of Smc3 deletion in normal and malignant hematopoiesis. Bi-allelic Smc3 loss induced bone marrow aplasia with premature sister chromatid separation, and revealed an absolute requirement for cohesin in hematopoietic stem cell function. In contrast, Smc3 haploinsufficiency increased self-renewal in vitro and in vivo including competitive transplantation. Smc3 haploinsufficiency reduced coordinated transcriptional output, including reduced expression of transcription factors and other genes associated with lineage commitment.  Smc3 haploinsufficiency cooperated with Flt3-ITD to induce acute leukemia in vivo, with potentiated Stat5 signaling and altered nucleolar topology.  These data establish a dose-dependency for cohesin in regulating chromatin structure and hematopoietic stem cell function.

overall_design: mRNA-seq in murine c-kit+ cells for the following genotypes: Smc3 fl/+, Smc3 del/+, Flt3-ITD, Smc3 fl/del Flt3-ITD

first: Christopher

last: Ott

first: Aaron

last: Viny


title: Global microRNA expression profiling of mouse livers following ischemia-reperfusion injury at different stages

city: Shanghai

first: Weisheng

address: Siping Road, No.1239

department: Department of Bioinformatics

institute: Tongji University


title: Synergistic gene expression signature observed in TK6 cells upon co-exposure to UVC-irradiation and protein kinase C-activating tumor promoters, a time course study (study 1/2)

first: Zhongqiang

last: Chen

summary: The purpose of this study, including this time course series and another dosage series, was to establish gene expression signatures representing the interaction of pathways deregulated by tumor promoting agents and pathways induced by DNA damage.  Human lymphoblastoid TK6 cells were pretreated with the protein kinase C activating tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) and exposed to UVC-irradiation.  The time and dose-responsive effects of the co-treatment were captured with RNA-sequencing (RNA-seq) in two separate experiments.  TK6 cells exposed to both TPA and UVC had significantly more genes differentially regulated than the theoretical sum of genes induced by either stress alone, thus indicating a synergistic effect on global gene expression patterns.  Further analysis revealed that TPA+UVC co-exposure caused synergistic perturbation of specific genes associated with p53, AP-1 and inflammatory pathways important in carcinogenesis.  The 17 gene signature derived from this model was confirmed with other PKC-activating tumor promoters including phorbol-12,13-dibutyrate, sapintoxin D, mezerein, (-)-Indolactam V and resiniferonol 9,13,14-ortho-phenylacetate (ROPA) with quantitative real-time PCR (QPCR).  Here we show a novel gene signature that may represent a synergistic interaction in the tumor microenvironment that is relevant to the mechanisms of chemical induced tumor promotion.

overall_design: mRNA profiles of TK6 cells in 0,4,8,24 hours, with individual or combined treatment between UV irradiation and tumor promoting agent;

first: Kyle

last: Glover

first: Lauren

last: Markell

first: Xing

last: Han


title: Global microRNA expression profiling of mouse livers following ischemia-reperfusion injury at different stages. [miRNA]

last: Zheng

summary: Hepatic ischemia reperfusion injury is a dynamic process consisting of two stages: ischemia and reperfusion, and triggers a cascade of physiological and biochemical events. Given the important role of microRNAs in regulating gene expression, we analyzed gene expression changes in mouse livers at sham control, ischemia stage, and reperfusion stage. We generated global expression profiles of microRNA and mRNA genes in mouse livers subjected to ischemia reperfusion injury at the three stages, respectively. Comparison analysis showed that reperfusion injury had a distinct expression profile whereas the ischemia sample and the sham control were clustered together. Consistently, there are 69 differentially expressed microRNAs between the reperfusion sample and the sham control whereas 28 differentially expressed microRNAs between the ischemia sample and the sham control. We further identified two modes of microRNA expression changes in ischemia reperfusion injury. Functional analysis of both the differentially expressed microRNAs in the two modes and their target mRNAs revealed that ischemia injury impaired mitochondria function, nutrient consumption, and metabolism process. In contrast, reperfusion injury led to severe tissue inflammation that is predominantly an innate-immune response in the ischemia reperfusion process. Our staged analysis of gene expression profiles provides new insights into regulatory mechanisms of microRNAs in mouse hepatic ischemia reperfusion injury.

overall_design: Liver microRNA profiles of male mouse liver under Sham, warm ischemia (WI) and Ischemia-reperfusion (IR) conditions were generated by deep-sequencing using Solexa/Illumina platform.

first: Jinzhen

last: Cai


title: Impact of two RNaseIII-like protein on the small RNA repertoire

city: Castanet

first: Matthias

last: Zytnicki

address: 24, Chemin de Borde-Rouge

department: Applied Mathematics and Informatics

country: France

institute: INRA

summary: RNaseIII enzymes catalyze the cleavage of double stranded RNA (dsRNA) and have diverse functions in RNA maturation. Arabidopsis RNASE THREE LIKE2 (RTL2) carries one RNaseIII and two dsRNA-binding (DRB) domains, and is the unique Arabidopsis RNAse III enzyme resembling the budding yeast siRNA-producing Dcr1 enzyme. Here we show that RTL2 modulates the production of siRNAs, and that this activity depends on both RNaseIII and DRB domains. However, RTL2 mode of action differs from that of Dcr1. Indeed, whereas Dcr1 directly cleaves dsRNAs into 23-nt siRNAs, RTL2 likely cleaves dsRNAs into molecules longer than siRNAs, which are subsequently processed by the DCL enzymes. Depending on the dsRNA considered, RTL2-mediated maturation either improves (RTL2- dependent loci) or reduces the efficiency of DCL processing (RTL2-sensitive loci). Because the vast majority of plant small RNAs are 24-nt siRNAs that guide DNA methylation at transposons and intergenic regions, RTL2 depletion mostly modifies DNA methylation in these regions. Nevertheless, 13% of the siRNA-producing loci affected by RTL2 depletion correspond to protein coding genes. We show that changes in 24-nt siRNA levels also affect DNA methylation levels at such loci, and inversely correlate with mRNA steady-state levels, thus implicating RTL2 in the control of protein-coding gene expression.

overall_design: Small RNA sequencing in different A. thaliana RnaseIII-like mutants

first: Hervé

last: Vaucheret


title: Global microRNA expression profiling of mouse livers following ischemia-reperfusion injury at different stages. [mRNA]

overall_design: Liver mRNA profiles of male mouse liver under Sham, warm ischemia (WI) and Ischemia-reperfusion (IR) conditions were generated by deep-sequencing using Solexa/Illumina platform.


title: Cell type-specific chromatin states differentially prime squamous cell carcinoma tumor-initiating cells for epithelial to mesenchymal transition

city: Brussels

first: Cédric

last: Blanpain

address: Route de Lennik 808

country: Belgium

institute: ULB

summary: Epithelial to mesenchymal transition (EMT) in cancer cells has been associated with metastasis, stemness and resistance to therapy. The reason why some tumors undergo EMT and other not might reflect intrinsic properties of their cell of origin, although this possibility is largely unexplored. By targeting the same oncogenic mutations to discrete skin compartments, we show cell type-specific chromatin and transcriptional states differentially prime tumors to EMT.  Squamous cell carcinomas (SCCs) derived from intrafollicular epidermis (IFE) are generally well-differentiated, while hair follicle (HF) stem cell-derived SCCs frequently exhibit EMT, efficiently form secondary tumors, and possess increased metastatic potential.  Transcriptional and epigenomic profiling revealed IFE and HF tumor-initiating cells possess distinct chromatin landscapes and gene regulatory networks associated with tumorigenesis and EMT that correlate with accessibility of key epithelial and EMT transcription factor binding sites.  These findings highlight the importance of chromatin states and transcriptional priming in dictating tumor phenotypes and EMT.

first: M

last: Latil

first: D

last: Nassar

first: B

last: Beck

first: S

last: Boumhadi

first: L

last: Wang

first: A

last: Brisebarre

first: R

last: Yi

first: C


title: Nucleosome dynamics and gene expression in glucocorticoid-treated acute lymphoblastic leukemia, RS4;11

first: Jennifer

middle: Nan-Wah

last: Wu

address: 450 Brookline Ave.

department: Pediatric Oncology


title: Hoxa5 undergoes dynamic DNA methylation and transcriptional repression in the adipose tissue of mice exposed to high-fat diet

city: Naples

first: Valerio

last: Costa

state: Italy

address: Via P. Castellino, 111

department: Institute of Genetics and Biophysics

country: Italy

institute: National Research Council

summary: Here we report the genome-wide DNA methylation analyses by MeDIP-seq, performed on epididymal fat of mice fed either high-fat or regular chow diet.

overall_design: Mice have been fed in high-fat (HFD) or regular chow diet (STD) for 5 months. Epididymal fat of both mice (HFD and STD) was extracetd and used for genome-wide DNA methylation analyses by methylated DNA immunoprecipitation sequencing (MeDIP-seq).

last: Parrillo

first: V

first: G

last: Raciti

last: Longo

last: Spinelli

last: Esposito

last: Nigro

last: Vastolo

last: Desiderio

first: F

last: Zatterale

last: Ciccodicola

first: P

last: Formisano

last: Miele

last: Beguinot


title: Evasion of affinity-based selection in germinal centers by Epstein-Barr virus LMP2A

first: Hufeng

last: Zhou

address: 181 Longwood Ave,

department: Department of Microbiology and Molecular Genetics

institute: Harvard Medical School

summary:
Abstract: Epstein-Barr virus (EBV) infects germinal center (GC) B cells and establishes persistent infection in memory B cells. EBV-infected B cells sometimes cause B cell malignancies in humans with T- or NK-cell deficiency. We now find EBV-encoded Latent Membrane Protein 2A (LMP2A) to mimic B cell antigen receptor (BCR) signaling in murine GC B cells, causing altered humoral immune responses and autoimmune diseases. Investigation of the impact of LMP2A on B cell differentiation in mice that conditionally express LMP2A in GC B cells, or all B-lineage cells, found LMP2A expression enhanced not only BCR signals but also plasma-cell differentiation, in vitro and in vivo. Conditional LMP2A expression in GC B cells resulted in preferential selection of low-affinity antibody–producing B-cells despite apparently normal GC formation. GC B cell-specific LMP2A expression led to systemic lupus erythematosus-like autoimmune phenotypes in an age-dependent manner. Epigenetic profiling of LMP2A B cells found increased H3K27ac and H3K4me1 signals at the Zbtb20 locus. We conclude that LMP2A reduces the stringency of GC B cell selection and may contribute to persistent EBV infection and pathogenesis by providing GC B cells with pro-survival signals.
Significance Statement: Epstein-Barr virus (EBV) is a human herpesvirus that establishes persistent infection of the B-cell compartment. EBV is associated with autoimmune diseases, including systemic lupus erythematosus (SLE). However, the molecular mechanisms by which EBV contributes to autoimmunity remain unclear. We used transgenic mouse models to study the role of EBV-encoded Latent membrane protein 2A (LMP2A), which mimics B-cell receptor signaling.　Interestingly, LMP2A not only enhanced B-cell survival, but also upregulated the transcription factor Zbtb20 and promoted plasma cell differentiation. When expressed late in B-cell development, LMP2A also caused prominent features of SLE, including autoantibody production with kidney immune complex deposition. Our findings suggest that LMP2A has important roles in B cell activation, differentiation and the development of EBV-associated autoimmune diseases.

overall_design: GFP transgenic mouse B cells and EBV LMP2a transgenic mouse B cells are subjected to ChIP-seq analysis for histone mark profiling. 2 mice of each genotype serving as replicates were analyzed. After the quality and reproducibility of the sequencing results were validated, results from replicates were combined

first: Takeharu

last: Minamitani

first: Teruhito

last: Yasui

first: Yijie

last: Ma

first: Daisuke

last: Okuzaki

first: Chiau-Yuang

last: Tsai

first: Shuhei

last: Sakakibara

first: Benjamin

last: Gewurz

first: Elliott

last: Kieff

first: Hitoshi

last: Kikutani


title: mRNA export through an additional cap-binding complex consisting of NCBP1 and NCBP3

city: Martinsried

first: Andreas

last: Pichlmair

state: Bayern

address: Am Klopferspitz 18

department: Mann

institute: Max-Planck Institut für Biochemie

summary: The flow of genetic information from DNA to protein requires polymerase-II transcribed RNA characterized by the presence of a 5’cap. The cap-binding complex (CBC), consisting of the nuclear cap-binding protein (NCBP) 2 and its adaptor NCBP1, is believed to bind all capped RNA and to be necessary for its processing and intracellular localization. Here, we show that NCBP1 but not NCBP2 is required for cell viability and poly(A) RNA export. We identify C17orf85 (here named NCBP3) as cap-binding protein that together with NCBP1 forms an alternative CBC in higher eukaryotes. NCBP3 binds mRNA, associates with components of the mRNA processing machinery and contributes to poly(A) RNA export. Loss of NCBP3 can be compensated by NCBP2 under steady state conditions. However, NCBP3 becomes pivotal under stress conditions such as virus infection. We propose the existence of an alternative CBC involving NCBP1 and -3 that plays a key role in mRNA biogenesis.

overall_design: GFP-tagged NCBP3 (C17orf85), NCBP2 or control protein RAB5C stably expressed in HeLa cells were precipitated and associated RNAs were analyzed by deep sequencing on the Illumina HiSeq platform

first: Arno

last: Meiler

first: Matthias

last: Habjan

first: Anna

last: Gebhardt


title: Genome-wide nucleosome dynamics in glucocorticoid-treated acute lymphoblastic leukemia cell line, RS4;11

first: Jennifer

last: Wu

summary: Precise nucleosome positioning is an increasingly recognized feature of promoters and enhancers, reflecting complex contributions of DNA sequence, nucleosome positioning, histone modification and transcription factor binding to enhancer activity and regulation of gene expression. Changes in nucleosome position and occupancy, histone variants and modifications, and chromatin remodeling are also critical elements of dynamic transcriptional regulation, but poorly understood at enhancers. We investigated glucocorticoid receptor-associated (GR) nucleosome dynamics at enhancers in acute lymphoblastic leukemia. For the first time, we demonstrate functionally distinct modes of nucleosome remodeling upon chromatin binding by GR, which we term central, non-central, phased, and minimal. Central and non-central remodeling reflect nucleosome eviction by GR and cofactors, respectively. Phased remodeling involves nucleosome repositioning and is associated with rapidly activated enhancers and induction of gene expression. Minimal remodeling sites initially have low levels of enhancerassociated histone modification, but the majority of these regions gain H3K4me2 or H3K27Ac to become de novo enhancers. Minimal remodeling regions are associated with gene ontologies specific to decreased B cell number and mTOR inhibition and may make unique contributions to glucocorticoid-induced leukemia cell death. Our findings form a novel framework for understanding the dynamic interplay between transcription factor binding, nucleosome remodeling, enhancer function, and gene expression in the leukemia response to glucocorticoids.

overall_design: We performed ChIP-Seq for histone modifications (H3K4me1, H3K4me2, H3K4me3, H3K27Ac), ChIP-Seq for transcription factors (GR and PU.1), and DNase-Seq from RS4;11 cells for two conditions:  1) no stimulation (steroid starved for 2 days), and 2) 10 nM dexamethasone for 1 hour.

middle: N

first: Luca

last: Pinello

first: Elinor

last: Yissachar

first: Jonathan

middle: W

last: Wischhusen

first: Guo-Cheng

last: Yuan

first: Charles

last: Roberts


title: LncRNA HOTAIR enhances ER signaling and confers tamoxifen resistance in breast cancer

city: Atlanta

first: Jindan

last: Yu

state: GA

address: E330, 1760 Haygood Dr NE

department: Urology

institute: Emory University

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

overall_design: Refer to individual Series


title: ABCC5 functions as a transporter of glutamate conjugates and analogs

city: Amsterdam

first: Koen

last: van de Wetering

address: Plesmanlaan 121

department: Molecular Oncology

country: Netherlands

institute: Netherlands Cancer Institute

summary: The ubiquitous efflux transporter ATP-binding cassette sub-family C member 5 (ABCC5) is present at high levels in the blood-brain barrier, neurons and glia, but its in vivo substrates and function are not known. Untargeted metabolomic screens revealed that Abcc5-/- mice accumulate endogenous glutamate conjugates and analogs in several tissues, but brain in particular. The abundant neurotransmitter N-acetylaspartylglutamate (NAAG), for example, was over 2-fold higher in Abcc5-/- brain. In line with ABCC5-mediated transport, the metabolites that accumulated in Abcc5-/- tissues were depleted in cultured cells that overexpressed human ABCC5. Using membrane vesicles, we show that ABCC5 not only transports the metabolites detected in our screen, but also a wide range of peptides containing a C-terminal glutamate. Glutamate conjugates are of physiological relevance because they can affect the function of glutamate, the principal excitatory neurotransmitter in the brain. We found that ABCC5 also transports exogenous glutamate analogs, like the classic excitotoxic neurotoxins kainic acid, domoic acid and N-methyl-D-aspartate (NMDA) and the therapeutic glutamate analog ZJ43. Taken together, we have identified ABCC5 as a general glutamate conjugate and analog transporter that affects the disposition of endogenous metabolites, toxins and drugs.

overall_design: A set of 5 wildtype brains was compared to a set of 5 Abcc5-knockout mouse brains

first: Robert

middle: S

last: Jansen

first: Sunny

last: Mahakena

first: Marcel

last: de Haas

first: Piet

last: Borst


title: MicroRNA 19a replacement partially rescues fin and cardiac defects in zebrafish model of Holt Oram syndrome [miRNA-seq]

city: Pisa

first: Romina

last: D'Aurizio

state: PI

address: via Moruzzi 1

department: IIT

institute: CNR

summary: The regulative role of miRNAs in Holt-Oram Syndrome is investigated in a zebrafish model.

overall_design: The zebrafish gene Tbx5a was silenced with antisense morpholino oligonucleotide MO-Tbx5a against the TSS of the gene. A MO-CT was used in CT sample. 1.5ng morpholinos were injected into the yolk of 1-cell stage embryos and total RNA extracted from 50 embryos collected at 48hpf.The Zebrafish  wild-type AB strain line was used.

first: Mario

last: Baumgart

first: Marco

last: Groth

first: Elena

last: Chiavacci

first: Letizia

last: Pitto


title: Akt1/Protein Kinase B Enhances Transcriptional Reprogramming of Fibroblasts to Functional Cardiomyocytes

city: Dallas

first: Min Soo

last: Kim

address: 5323 Harry Hines Blvd.

department: Department of Clinical Sciences

institute: University of Texas Southwestern

summary: Conversion of fibroblasts to functional cardiomyocytes represents a potential approach for restoring cardiac function following myocardial injury, but the technique thus far has been slow and inefficient. To improve the efficiency of reprogramming fibroblasts to cardiac-like myocytes (iCMs) by cardiac transcription factors (Gata4, Hand2, Mef2c, and Tbx5=GHMT), we screened 192 protein kinases and discovered that Akt/protein kinase B dramatically accelerates and amplifies this process. Approximately 50% of reprogrammed fibroblasts displayed spontaneous beating after three weeks of induction by Akt plus GHMT. Furthermore, addition of Akt1 to GHMT evoked a more mature cardiac phenotype for iCMs, as seen by enhanced polynucleation, cellular hypertrophy, gene expression, and metabolic reprogramming. Igf1 and Pi3 kinase acted upstream of Akt, whereas mTORC1 and Foxo3a acted downstream of Akt to influence fibroblast-to-cardiomyocyte reprogramming. These findings provide new insights into the molecular basis of cardiac reprogramming and represent an important step toward further application of this technique.

overall_design: We performed RNA-Seq using either isolated adult mouse ventricular cardiomyocytes (CMs) or MEFs treated for three weeks with empty vector, GHMT (iCMs cell sorted using αMHC-GFP before RNA-Seq), or AGHMT (iCMs cell sorted using αMHC-GFP before RNA-Seq).

first: Huanyu

last: Zhou

first: Matthew

middle: E

last: Dickson

first: Rhonda

last: Bassel-Duby

first: Eric

last: Olson


title: Promoter targeted bisulfite sequencing reveals DNA methylation profiles associated with low sperm motility in asthenozoospermia

city: Shenzhen

first: Desheng

last: Gong

address: No.7 PengFei road

institute: Agricultural Genomes Institute at Shenzhen

summary: Aberrant DNA methylation profiles have been associated with male infertility and some semen parameters. However, few studies systematically surveyed DNA methylation profiles associated with sperm motility in normozoospermia and asthenozoospermia. In this study, based on promoter targeted bisulfite sequencing technology, we provided a quantitative description on global DNA methylation profiles. The average global methylation values were 24.7% and the inter-individual variance was about 14.4%, while the intra-individual variance was about 3.9%. The difference between different motile sperm population or different participant groups was subtle and not significant. Furthermore, we identified 134 differentially methylated CpGs and 134 differentially variable CpGs in low motile sperm from asthenozoospermic patient (P < 0.05). Based on the literature, we further found 16 differentially methylated or variable genes which were required for spermatogenesis or sperm motility or dominantly expressed in testis. This study will provide potential markers for clinical diagnosis and a promising basis for understanding the effect of DNA methylation on asthenozoospermia.

overall_design: This is case-control study investigating DNA methylation of CpGs in functionally important regions of high and low motile sperm population from normozoospermic controls (n=8) and asthenozoospermic cases (n=7).

first: Ye

last: Du

first: Meiyan

last: Li


title: Conservation of m5C in rRNA in the kingdom Plantae

city: Adelaide

first: Alice

middle: Louise

last: Burgess

state: SA

address: Frome road

department: Genetics and Evolution

country: Australia

institute: University of Adelaide

summary: Here we use bisulfite conversion of RNA combined with high-throughput IIlumina sequencing (RBS-seq) to identify single-nucleotide resolution of m5C sites in non-coding ribosomal RNAs of all three sub-cellular transcriptomes across six diverse species that included, the single-celled algae Nannochloropsis oculata, the macro algae Caulerpa taxifolia and multi-cellular higher plants Arabidopsis thaliana, Brassica rapa, Triticum durum and Ginkgo biloba. RNA-seq was performed on total RNA for all species except for Arabidopsis in order to generate rRNA reference sequences using the Arabidopsis rRNA sequences (TAIR10) as a guide.

sample_organism:
Ginkgo biloba
Arabidopsis thaliana
Brassica rapa
Triticum turgidum subsp. durum
Nannochloropsis oculata
Caulerpa taxifolia

overall_design: RNA Bisulfite sequencing was used to identify candidate m5C sites in tRNAs and rRNAs of diverse plant species. One biological replicate was performed for each RNA-seq library. One biological replicate was performed for each RNA Bisulfite-seq library except for Arabidopsis thaliana, which was performed in biological triplicate. Illumina sequencing was performed on a MiSeq platform.

first: Rakesh

last: David

first: Iain

middle: R

last: Searle


title: tRNA 5-methylcytosine in Arabidopsis thaliana

summary: Here we use bisulfite conversion of RNA combined with high-throughput IIlumina sequencing (RBS-seq) to identify single-nucleotide resolution of m5C sites in  transfer RNAs of all three sub-cellular transcriptomes of Arabidopsis thaliana. 5-methylcytosine sites in tRNAs were also determined in Arabidopsis T-DNA knockouts for the RNA methyltransferases TRM4A, TRM4B, TRDMT1, NSUN5 and NOP2A.

overall_design: RNA Bisulfite sequencing was used to identify candidate m5C sites in Arabidopsis tRNAs. Illumina sequencing was performed on a MiSeq platform. Two experiments were performed. In experiment 1, all samples were performed with one biological replicate, except for Arabidopsis thaliana wild type, which was performed in biological triplicate. In experiment 2, only one biological replicate was performed for each sample.


title: rRNA 5-methylcytosine in Arabidopsis thaliana

summary: Here we use bisulfite conversion of RNA combined with high-throughput IIlumina sequencing (RBS-seq) to identify single-nucleotide resolution of m5C sites in ribosomal RNAs of all three sub-cellular transcriptomes in Arabidopsis thaliana. m5C sites in rRNAs were also anlyzed in Arabidopsis T-DNA knockouts for the RNA methyltransferases TRM4A, TRM4B, TRDMT1, NSUN5, NOP2A, NOP2B and NOP2C.

overall_design: RNA Bisulfite sequencing was used to identify candidate m5C sites in rRNAs of Arabidopsis thaliana. Illumina sequencing was performed on a MiSeq platform. Two experiments were performed; experiment 1 and experiment 2. In experiment 1, one to three biological replicates were performed for the genotypes wild type, nop2a-2, nop2b-1, nop2b-2, nop2c-1, nop2c-2, nsun5-1 and nsun5-2. In experiment 2, one biological replicate was performed for each of the genotypes wild type, trm4a, trm4b-1, trdmt1 and trdmt1 trm4b.


title: MicroRNA 19a replacement partially rescues fin and cardiac defects in zebrafish model of Holt Oram syndrome


title: RNA-Seq analysis facilitates quantitative analysis to identify genes in hepatic tissue of partially hepatectomized rats regulated by ASCs-miR27b introduction in comparison with ASCs-siRNC or saline controls

city: Kaohsiung

first: Kuang-Den

last: Chen

address: 123, Ta-Pei Rd., Niao-Sung Dist.

department: Translational Research Center in Biomedical Sicences

country: Taiwan

institute: Kaohsiung Chang Gung Memorial Hospital

summary:
Methods: The cDNA libraries from 3 pooled samples of liver tissues for each group were sequenced to generate RNA profiles using Illumina Miseq platform
Results: The sequencing runs yielded a total of 22.67 M reads with an average length of 100 bp. The high-throughput sequencing performed for liver samples with different treatments showed similar numbers of yielded reads ranged from 5.57 to 5.74 M and the same average length. The Strand NGS software (version 2.1) was used using default parameters for pre-alignment and post-alignment quality control analysis and 100% of the raw reads remained in the dataset. Of these, 19.02 M reads (84%) were mapped into contigs of the rat genome (rn5) and identified 31457 transcripts in liver samples.

overall_design: Hepatic mRNA profiles of partially hepatectomized rats intravenously treated by ASCs-miR27b, ASCs-siRNC, saline control and naïve control were generated by sequencing, using Illumina Miseq platform

first: Wei-Teng

last: Weng


title: Genome-wide identification of differentially expressed microRNAs in leaves and the developing head of four durum genotypes during water deficit stress

city: Urrbrae

first: Jason

middle: A

last: Able

address: GN10b, Waite Main Buiding, Waite Road

department: School of Agriculture, Food and Wine

summary:
Background: MicroRNAs are endogenous small noncoding RNAs that play critical roles in plant abiotic stress responses. The interaction between miRNA-mRNA targets and their regulatory pathways in response to water deficit stress has been investigated in many plant species. However, the miRNA transcriptome of durum wheat (Triticum turgidum L. ssp. durum) is poorly characterised, with little known about miRNA functions related to water deficit stress. Yield loss in durum wheat can be exacerbated due to minimal rainfall in the early reproductive stages of development during Spring in Australia. This study describes genotypic differences in the miRNAome between water deficit tolerant/sensitive durum, using flag leaf and developing head tissue, and more specifically identifies miRNAs associated with water deficit stress.
Results: Small RNA libraries (96 in total) were constructed from flag leaf and developing head tissues of four durum genotypes (Tamaroi, Yawa, EGA Bellaroi, Tjilkuri), with or without water deficit stress. Illumina sequencing and subsequent analysis detected 110 conserved miRNAs and 159 novel candidate miRNA hairpins. Statistical analysis of the abundance of sequencing reads revealed 66 conserved miRNAs and five novel miRNA hairpins showing differential expression under water deficit stress. During stress, several conserved and novel miRNAs showed unambiguous inverted regulatory profiles between the durum genotypes studied. Several miRNAs were also identified to have different abundance in the flag leaf compared to the developing head regardless of treatment. Predicted mRNA targets from four novel durum miRNAs were characterised using Gene Ontology (GO) which revealed functions common to stress responses and plant development.
Conclusion: For the first time, we present a comprehensive study of the miRNA transcriptome of flag leaf and developing head tissues in different durum genotypes under water deficit stress. The identification of differentially expressed miRNAs provides molecular evidence that miRNAs are potential determinants of water stress tolerance in durum wheat. GO analysis of predicted targets contributes to the understanding of genotype-specific physiological responses leading to stress tolerance capacity. Further functional analysis of specific stress responsive miRNAs identified, and their interaction with mRNA targets is ongoing and will assist in developing future durum wheat varieties with enhanced water deficit stress tolerance.

overall_design: A total of 96 small RNA libraries were analysed [4 durum genotypes * 2 tissue types * 2 treatment groups (control and water stress) * 6 biological replicates] in this study. Please see Methods, Liu et al.(2015)

first: Haipei

last: Liu

first: Nathan

last: Watson-Haigh

first: Ute

last: Baumann

first: Diane

last: Mather

first: Amanda


title: A mechanism for expansion of the regulatory T cell repertoire and its role in enforcing self-tolerance.

city: Pittsburgh

first: Hatice

middle: Ulku

last: Osmanbeyoglu

state: PA

address: 5117 Centre Ave

institute: University of Pittsburgh

summary: Thymic Treg cells, mature non-Treg CD4+ single positive thymocytes, peripheral (spleen) resting and activated Treg cells were sorted from Foxp3-gfp reporter (wid type, WT) mice or Foxp3 enhancer CNS3 knockout (KO, carrying the same GFP reporter) mice. Total RNA was extracted and used for RNA sequencing to assess gene expression profiles.

overall_design: Two 6-8 week old littermates of male Foxp3-gfp and Foxp3ΔCNS3-gfp mice were used to sort Treg cells and conventional CD4+ T cells. Lymphocyte preparation and electronic sorting were performed at the same time. RNA extraction, SMART amplification, library preparation were conducted in parallel.

first: Yongqiang

last: Feng

middle: U

first: Alexander

middle: Y

last: Rudensky


title: LncRNA HOTAIR enhances ER signaling and confers tamoxifen resistance in breast cancer [ChIP-Seq]

first: Jindan

last: Yu

summary: HOTAIR is up-regulated in tamoxifen-resistant breast cancer tissues compared to their primary counterparts.  Mechanistically, HOTAIR is a direct target of ER-mediated transcriptional repression and is thus restored upon the blockade of ER signaling, either by hormone deprivation or tamoxifen treatment.

overall_design: ChIP-Seq examination of ER binding sites in MCF7 cells.

first: Xingyang

last: Xue


title: Conservation of m5C in tRNAs in the kingdom Plantae

summary: Here we use bisulfite conversion of RNA combined with high-throughput IIlumina sequencing (RBS-seq) to identify single-nucleotide resolution of m5C sites in transfer RNAs of all three sub-cellular transcriptomes across six diverse species that include, the single-celled algae Nannochloropsis oculata, the macro algae Caulerpa taxifolia and multi-cellular higher plants Arabidopsis thaliana, Brassica rapa, Triticum durum and Ginkgo biloba.

platform_organism:
Ginkgo biloba
Arabidopsis thaliana
Brassica rapa
Triticum turgidum subsp. durum
Nannochloropsis oculata
Caulerpa taxifolia

overall_design: RNA Bisulfite sequencing was used to identify candidate m5C sites in tRNAs of diverse plant species. RNA-seq was used to examine abundance of tRNAs in Arabidopsis thaliana wild type. Arabidopsis thaliana RNA Bisulfite-Seq libraries were performed in biological triplicate. All other samples had only one biological replicate. Illumina sequencing was performed on a MiSeq platform.

middle: L


title: Expression divergence of chemosensory genes between Drosophila sechellia and its sibling species and its implications for host shift [Dsec JP]

city: Taipei

first: Jia-Ming

last: Chang

address: NO.64, Sec. 2, ZhiNan Rd., Wenshan District

department: Department of Computer Science

institute: National Chengchi University

summary: Drosophila sechellia relies exclusively on the fruits of Morinda citrifolia, which are toxic to most insects, including its sibling species D. melanogaster and D. simulans.  Although several odorant binding protein (Obp) genes and olfactory receptor (Or) genes were suggested to be associated with the D. sechellia host shift, a broad view of how chemosensory genes have contributed to this shift is still lacking.  We therefore studied the antennal transcriptomes, the main organ responsible for detecting food resource and oviposition, of D. sechellia and its two sibling species. We wanted to know whether gene expression, particularly chemosensory genes, has diverged between D. sechellia and its two sibling species.  Using a very stringent definition of differential gene expression, we found 147 genes (including 11 chemosensory genes) were up-regulated while only 81 genes (including 5 chemosensory genes) were down-regulated in D. sechellia.  Interestingly, Obp50a exhibited the highest up-regulation, a ~100 fold increase, and Or85c – previously reported to be a larva-specific gene– showed ~20 fold up-regulation in D. sechellia.  Furthermore, Ir84a, proposed to be associated with male courtship behavior, is significantly up-regulated in D. sechellia.  We also found expression divergence in most of the receptor gene families between D. sechellia and the two sibling species.  Our observations suggest that the host shift of D. sechellia is associated with expression profile divergence in all chemosensory gene families and is achieved mostly by up-regulation of chemosensory genes.

overall_design: RNAseq experiments in wild type drosophila antennaes. The strain is D. sechellia (k-s10; #14021-0248.25).

first: Meng-Shin

last: Shiao

first: Wen-Lang

last: Fan

first: Mei-Yeh

last: Lu

first: Cedric

last: Notredame

first: Shu

last: Fang

first: Rumi

last: Kondo

first: Wen-Hsiung


title: Expression divergence of chemosensory genes between Drosophila sechellia and its sibling species and its implications for host shift [Dsim JP]

summary: Drosophila simulans relies exclusively on the fruits of Morinda citrifolia, which are toxic to most insects, including its sibling species D. melanogaster and D. simulans.  Although several odorant binding protein (Obp) genes and olfactory receptor (Or) genes were suggested to be associated with the D. simulans host shift, a broad view of how chemosensory genes have contributed to this shift is still lacking.  We therefore studied the antennal transcriptomes, the main organ responsible for detecting food resource and oviposition, of D. simulans and its two sibling species. We wanted to know whether gene expression, particularly chemosensory genes, has diverged between D. simulans and its two sibling species.  Using a very stringent definition of differential gene expression, we found 147 genes (including 11 chemosensory genes) were up-regulated while only 81 genes (including 5 chemosensory genes) were down-regulated in D. simulans.  Interestingly, Obp50a exhibited the highest up-regulation, a ~100 fold increase, and Or85c – previously reported to be a larva-specific gene– showed ~20 fold up-regulation in D. simulans.  Furthermore, Ir84a, proposed to be associated with male courtship behavior, is significantly up-regulated in D. simulans.  We also found expression divergence in most of the receptor gene families between D. simulans and the two sibling species.  Our observations suggest that the host shift of D. simulans is associated with expression profile divergence in all chemosensory gene families and is achieved mostly by up-regulation of chemosensory genes.

overall_design: RNAseq experiments in wild type drosophila antennaes. The strain is D. simulans (k-s05; #14021-025.194).

last: Li