title: Mouse CD4+/CD8+ thymocytes from miR-181a1/b1 KO vs WT

status: Public on Apr 09 2014

city: Baltimore

first: Loyal

last: Goff

state: MD

address: 733 N. Broadway Avenue

department: Institute of Genetic Medicine

institute: Johns Hopkins University

summary: miR-181a1/b1, miR-181a2/b2, and miR-181c/d belong to a highly conserved family of microRNA clusters, yet their role in vivo is poorly understood. Here we show that the miR-181a1/b1 cluster is absolutely essential for NKT development and is a critical determinant of thymocyte proliferation, survival and T-cell receptor α locus rearrangement. Furthermore, while individual ablation of miR-181a2/b2 and miR-181c/d revealed no overt phenotypes, compound mutant mice lacking both miR-181a1/b1 and miR-181a2/b2 display decreased survival, reduced body weight, and abnormal B cell development. Mechanistically, we reveal that miR-181 regulates PTEN, a key tumor suppressor whose abundance determines key metabolic adaptations required to meet the biosynthetic demands of highly proliferative tissues. These results provide important insights into the physiological function of this family of microRNAs in vivo; moreover, it places miR-181 as a central regulator of the PI3K signaling pathway and cellular metabolism.

overall_design: Three sorted populations of double positive (DP) thymocytes from each of two conditions (WT and miR181a1/b1 -/- mice)

first: Jorge

last: Henao-Mejia

first: Adam

last: Williams

first: Matthew

last: Staron

first: Paula

last: Licona-Limon

first: Susan

last: Kaech

first: Maki

last: Nakayama

first: John

last: Rinn

first: Richard

last: Flavel


title: Holocentromeres are dispersed point centromeres localized at transcription factor hotspots

status: Public on Apr 08 2014

city: Seattle

first: Jorja

last: Henikoff

state: WA

address: 1100 Fairview AV N, A1-162

department: Basic Sciences

country: USA

institute: Fred Hutchinson Cancer Research Center

type: Genome binding/occupancy profiling by high throughput sequencing

summary: The centromere is a defining feature of eukaryotic chromosomes and is essential for the segregation of chromosomes during cell division. Centromeres are universally marked by the histone variant cenH3 and are restricted to specialized chromatin that most commonly localized to a single position along the chromosome. However, the DNA on which centromeric nucleosomes assemble is not conserved and varies greatly in size and composition. It ranges from genetically defined point centromeres that assemble a single cenH3-containing nucleosome to epigenetically defined regional centromeres embedded in megabases of tandemly repeated DNA to holocentromeres that extend along the length of the entire chromosomes. The organization of regional and holocentric centromeres has so far been elusive, as the precise locations of cenH3-containing sequences could not be determined. Our results show that the point centromere is the basic unit of holocentromeres and provide a basis for understanding how centromeric chromatin is maintained.

overall_design: We use high-resolution mapping of cenH3-associated DNA to show that Caenorhabditids elegans holocentromeres are organized as dispersed but discretely localized point centromeres.

first: Florian

middle: A

last: Steiner

first: Steven


title: Quantitative sequencing of 5-formylcytosine in DNA at single-base resolution

city: Cambridge

first: Giovanni

last: Marsico

address: Robinson Way

country: United Kingdom

institute: CRUK Cambridge Institute

type: Methylation profiling by high throughput sequencing

summary: Cytosine base modifications 5-methylcytosine (5mC), 5-hydroxymethylcytosine (5hmC) and 5-formylcytosine (5fC) are present in mammalian DNA. Here, reduced bisulfite sequencing is developed for quantitatively sequencing 5fC at single-base resolution. This method is then applied with oxidative bisulfite sequencing to gain a map of 5mC, 5hmC and 5fC in mouse embryonic stem cells.

overall_design: 12 samples, reduced representation bisulphite treatment: 4 replicates each for bisulphite (BS), oxidative BS (oxBS) and reduced BS (redBS) for the detection of 5mC, 5hmC and 5fC. Mouse (strain B6C) embryonic stem cells.

first: Michael

middle: J

last: Booth

first: Martin

last: Bachman

first: Dario

last: Beraldi

first: Shankar

last: Balasubramanian


title: Genome-wide measuement of methylated cytosin in regulatory T cells (Treg) and conventional T cells (Tconv)  depleted with Uhrf1

status: Public on Apr 07 2014

city: Yokohama

first: Takaho

middle: A.

last: Endo

state: Kanagawa

address: 1-7-22 Suehiro, Tsurumi

department: IMS

country: Japan

institute: RIKEN

summary: To investigate the molecular mechanism how Uhrf1 regulate gene expression and cell cycle progression in Treg, we precipitated methylated DNA using recombinant MBD1 followed by  genome-wide analysis of methylated gene loci by next generation sequencer

overall_design: Examination of methylated DNA in Treg and Tconv isolated from Cd4cre-Uhrf1+/+ and Cd4cre-Uhrf1flox/flox (Cd4cre-Uhrf1fl/fl)

first: T

first: Y

last: Furusawa

last: Obata

first: K

last: Hase


title: The epigenetic regulator Uhrf1 facilitates functional expansion of colonic regulatory T cells

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

overall_design: Refer to individual Series


title: RNA-seq and Proteogenomics Reveal the Importance of Leaderless mRNAs in the Radiation-tolerant Bacterium Deinococcus deserti

city: Saint Paul lez Durance

first: Arjan

last: de Groot

address: CEA Cadarache

department: BIAM-SBVME

country: France

institute: CEA

type: Expression profiling by high throughput sequencing

summary: Deinococcus deserti is a desiccation- and radiation-tolerant desert bacterium. Differential RNA sequencing was performed to explore the specificities of its transcriptome. Strikingly, for 1174 (60%) mRNAs the transcription start site was found exactly at (916 cases, 47%) or very close to the translation initiation codon AUG or GUG. Such proportion of leaderless mRNAs, which may resemble ancestral mRNAs, is unprecedented for a bacterial species. Proteomics showed that leaderless mRNAs are efficiently translated in D. deserti. Interestingly, we also found 173 additional transcripts with a 5’-AUG or 5’-GUG that would make them competent for ribosome binding and translation into novel small polypeptides. Fourteen of these are predicted to be leader peptides involved in transcription attenuation. Another 30 correlated with new gene predictions and/or showed conservation with annotated and non-annotated genes in other Deinococcus species, and five of these novel polypeptides were indeed detected by mass spectrometry. The data also allowed re-annotation of the start codon position of 257 genes, including several DNA repair genes. Moreover, several novel highly radiation-induced genes were found and their potential roles are discussed. Based on our RNA sequencing and proteogenomics data, we propose that translation of many of the novel leaderless transcripts, which may have resulted from single nucleotide changes and maintained by selective pressure, provides a new explanation for the generation of a cellular pool of small peptides important for protection of proteins against oxidation and thus for radiation/desiccation tolerance and adaptation to harsh environmental conditions.

overall_design: Using Illumina HiSeq 2000, RNA-Seq was performed to explore the transcriptome, including transcription start sites identification, in non-irradiated and irradiated Deinococcus deserti.

first: Laurence

last: Blanchard


title: Phosphate Starvation in Chlamydomonas reinhardtii

status: Public on Apr 05 2014

city: Stanford

first: Arthur

last: Grossman

state: CA

address: 260 Panama Street

department: Plant Biology

institute: Carnegie Institute for Science

summary: Phosphorus (P) is an essential nutrient that is limiting in many environments. When P is scarce organisms employ strategies for conservation of internal stores, and to efficiently scavenge P from their external surroundings. In this study we investigated the acclimation response of Chlamydomonas reinhardtii to P deficiency, comparing the transcriptional profiles of P starved wild-type cells to the P replete condition. RNA was prepared from P-containing or P-deprived logarithmic growth phase cells and subjected to RNA-Seq analysis. During the 24 hours after the imposition of P starvation we observed that from the 407 significantly changing genes (> 2 fold change, corrected p-value < 0.05) in the wild-type 317 genes were up-regulated, in average 8.36-fold, and 90 genes were down-regulated by 3.43-fold, in average. Many of the upregulated genes encoded enzymes involved in specific responses to P starvation, including PHOX, encoding the major secreted alkaline phosphatase, and multiple putative, high-efficiency phosphate transporter genes. More general responses included the up-regulation of genes involved in photoprotective processes (LHCSR3, LHCSR1, LHCBM9, PTOX1) and genes involved in protein modification and degradation. Down-regulated mRNAs indicated an early stage of the reduction of chloroplast ribosomal proteins, which are considered to be a reservoir for P in the cell.

overall_design: Chlamydomonas reinhardtii strain 21 gr (CC1690, wild-type) grown in TAP medium (Harris 1989) in a rotary incubator (200 rpm) at 25 °C in continuous light (70 µmol m-2 s-1). For 24 hours, either 1.1 mM phosphate or 0 mM were provided with the growth media. P deprivation was achieved by washing cells twice in midlogarithmic growth phase with liquid TAP medium without P (TAP-P) and cells were resuspended at a density of 2.5 mg/ml Chlorophyll in TAP or TAP-P. Cell aliquots were collected for mRNA isolation 24 h after being transferred either to TAP or TAP-P medium.

first: Jeffrey

last: Moseley


title: ChIPseq examination of EAP-1 binding across generations of spr-5(by101) mutant worms

city: San Diego

first: Yun

last: Zhu

address: 9300 Gilman Dr

department: Chemistry Departmant

institute: UCSD

summary: How epigenetic information is transmitted from generation to generation remains largely unknown. Deletion of the C. elegans Histone H3 lysine 4 dimethyl (H3K4me2) demethylase spr-5 leads to inherited accumulation of the euchromatic H3K4me2 mark and progressive decline in fertility. Here we identified a genetic network of chromatin-modifying factors, including the H3K4me1/me2 methyltransferases SET-17 and SET-30, the H3K9me1/me2 methyltransferase MET-2, an H3K9me3 methyltransferase, SET-26, the H3K9me3 demethylase JMJD-2, and an H3K9me reader EAP-1, which regulate the trans-generational flow of epigenetic information. Importantly, genetic ablation of set-17, set-30, jmjd-2, or eap-1 suppresses the progressive transgenerational phenotypes, while loss of SET-26 or MET-2 accelerates the infertility of spr-5 mutant worms. We further show that loss of spr-5 also causes a trans-generational increase in lifespan, which is dependent on these chromatin regulators as well as DAF-36 and DAF-12, which control a germline to soma longevity signaling pathway. These findings suggest that the balance between the euchromatic H3K4 and the heterochromatic H3K9 methylation regulates trans-generational effects on longevity and fertility.

overall_design: EAP-1 binding ChIPseq libraries were prepared from 50 ul of packed young adult worms which were maintained at 16 degrees until the appropriate generation and then shifted to 25 degrees after birth. Two biological repeats were generated for wildtype and generation 20 spr-5(by101) mutant worms and a single repeat for generation 10. There were four input samples and eap-1 null mutant worms were also analyzed.

first: Eric

middle: L

last: Greer

first: Emily

last: Brookes


title: A Smooth Muscle-Like Origin for Beige Adipocytes

status: Public on Apr 04 2014

city: Boston

first: Jonathan

middle: Z

last: Long

state: MA

address: 3 Blackfan Circle, Rm 11115

institute: Dana-Farber Cancer Institute

summary: A comparison of the translating RNAs in UCP1-positive cells from the classical BAT, the inguinal WAT, and the perigonadal WAT. We find genes common to all UCP1-positive cells independent of anatomical location, as well as genes specifically enriched in one group versus the others. Results provide a portrait of gene expression in UCP1-positive cells in vivo.

overall_design: Translating RNAs in UCP1-positive cells were purified from 6 week old female UCP1-TRAP mice after 2 weeks at 4 degC. N=3 for iWAT, N=2 for pgWAT, and N=3 for BAT.

first: Bruce

middle: M

last: Spiegelman


title: Identification of small ORFs in vertebrates using ribosome footprinting and evolutionary conservation

city: New Haven

first: Antonio

middle: J.

last: Giraldez

state: CT

address: 333 Cedar Street

department: Genetics

institute: Yale University

type: Non-coding RNA profiling by high throughput sequencing

summary: Identification of the coding elements in the genome is a fundamental step to understanding the building blocks of living systems. Short peptides (< 100 aa) have emerged as important regulators of development and physiology, but their identification has been limited by their size. We have leveraged the periodicity of ribosome movement on the mRNA to define actively translated ORFs by ribosome footprinting. This approach identifies several hundred translated small ORFs in zebrafish and human. Computational prediction of small ORFs from codon conservation patterns corroborates and extends these findings and identifies conserved sequences in zebrafish and human, suggesting functional peptide products (micropeptides). These results identify micropeptide‐encoding genes in vertebrates, providing an entry point to define their function in vivo.

overall_design: Ribosome profiling experiments at five timepoints across zebrafish development in WT embryos

first: Ariel

last: Bazzini

first: Timothy

middle: G

last: Johnstone

first: Romain

last: Christiano

first: Sebastian

middle: D

last: Mackowiak

first: Benedikt

last: Obermayer

first: Elizabeth

middle: S

last: Fleming

first: Charles

middle: E

last: Vejnar

first: Miler

middle: T

last: Lee

first: Nikolaus

last: Rajewsky

first: Tobias

middle: C

last: Walther


title: MOV10 Is a 5' to 3' RNA Helicase Contributing to UPF1 mRNA Target Degradation by Translocation along 3'UTRs

status: Public on Apr 11 2014

city: Berlin

first: Markus

last: Landthaler

address: Robert-Rössle-Straße 10

department: Berlin Institute for Medical Systems Biology

country: Germany

institute: Max-Delbrück-Center for Molecular Medicine


title: Transcriptome analysis reveals an activation of MHC-I and MHC-II pathways in chicken trachea immunized with infectious laryngotracheitis virus vaccine

city: College Park

first: Jose

middle: Adrian

last: Carrillo

address: 1413 Animal Sciences Center

department: Department of Animal and Avian Sciences

institute: University of Maryland

summary: Infectious laryngotracheitis (ILT) is an acute, contagious, upper respiratory disease, which is caused by gallid herpesvirus 1 (GaHV-1). Due to the mortality rates up to 70% depending on the virulence of the virus, it is of economic importance of the disease to explore the etiology of the ILT in the poultry industry. In this study, 15-day-old SPF white leghorn chickens were used to transcriptome analysis in chicken trachea immunized with infectious laryngotracheitis virus vaccine. In conclusion, chicken embryo origin (CEO) vaccine activation of the MHC-I and MHC-II pathways provides insight into the molecular mechanism of immune response in chickens, and holds potential for evaluation and design of new ILT vaccines in a manner adapted to the host immune response to the virus.

overall_design: Ten vaccine inoculated birds were randomly divided in two groups. Each group represents one replication of five pooled tissues, for inoculated birds. Control group consists of five birds that received sterile vaccine diluent.

first: José

first: Juan

last: Luo

first: Kimberly

middle: R

last: Menendez

first: Nathaniel

last: Tablante

first: Keji

last: Zhao

first: Jiuzhou

last: Song


title: Domains of genomewide gene expression dysregulation in Down syndrome [Dnase HS mapping]

city: Geneva

first: Audrey

last: Letourneau

address: Rue Michel-Servet 1

department: Genetic Medicine and Development

country: Switzerland

institute: University of Geneva Medical School

summary: Trisomy 21 (T21) is the most frequent genetic cause of cognitive impairment. To assess the perturbations of gene expression in T21, and to eliminate the noise of the genomic variability, we studied the transcriptome of fetal fibroblasts from a pair of monozygotic twins discordant for T21. Here we show that the differential expression between the twins is organized in domains along all chromosomes that are either up- or downregulated. These gene expression dysregulation domains (GEDDs) can be defined by the expression level of their gene content, and are well conserved in induced pluripotent stem cells derived from the twins’ fibroblasts. Comparison of the transcriptome of the Ts65Dn mouse model of DS and wild-type, also showed GEDDs along the mouse chromosomes that were syntenic in human. The GEDDs correlate with the lamina-associated (LADs) and replication domains of mammalian cells. The overall LADs position was not altered in trisomic cells. However, the H3K4me3 profile of the trisomic fibroblasts was modified and accurately followed the GEDD pattern. These results suggest that the nuclear compartments of trisomic cells undergo modifications of the chromatin environment influencing the overall transcriptome and that GEDDs may therefore contribute to some T21 phenotypes.

overall_design: DNaseI HS mapping in monozygotic twins discordant for trisomy 21 (2 replicates of each).

first: Federico

last: Santoni

first: Ximena

last: Bonilla

last: Sailani

first: David

last: Gonzalez

first: Jop

last: Kind

first: Claire

last: Chevalier

first: Robert

last: Thurman

first: Richard

last: Sandstrom

first: Youssef

last: Hibaoui

first: Konstantin

last: Popadin

first: Emilie

last: Falconnet

first: Maryline

last: Gagnebin

first: Corinne

last: Gehrig

first: Anne

last: Vannier

first: Michel

last: Guipponi

first: Laurent

last: Farinelli

first: Daniel

last: Robyr

first: Marco

last: Garieri

first: Eugenia

last: Migliavacca

first: Christelle

last: Borel

first: Samuel

last: Deutsch

first: Anis

last: Feki

first: John

last: Stamatoyannopoulos

first: Yann

last: Herault

first: Bas

last: van Steensel

first: Roderic

last: Guigo

first: Stylianos

last: Antonarakis


title: RNA sequencing reveals novel gene clusters in bovine conceptuses associated with maternal recognition of pregnancy and implantation

city: Belfield

first: Paul

last: McGettigan

state: Co Dublin

address: Veterinary Sciences Centre, UCD

department: School of Agriculture, Food Science and Veterinary Medicine

country: Ireland

institute: University College Dublin

summary: Successful establishment and maintenance of pregnancy can be attained only through optimum conceptus-maternal cross talk. Despite significant progress in our understanding of the temporal changes in the transcriptome of the uterine endometrium, we have only a rudimentary knowledge of the genes and pathways governing growth and development of the bovine conceptus. In particular, very little information exists for the posthatchingembryo and elongating conceptus. This period of development is arguably the most important, as approximately 40% of all embryonic loss occurs between Days 8 and 17 of pregnancy in cattle. Here, we describe the global transcriptome profile of the bovine conceptus at five key stages of its pre- and peri-implantation growth (Days 7, 10, 13, 16, and 19) using state-of-the-art RNA sequencing techniques. More than 287 million reads were generated at the five stages, and more than 22 700 unique transcripts were detected. Analysis of variance followed by self-organizing maps identified differentially regulated (P < 0.05) genes organized in nine gene clusters forming a sequential transcript dynamics across these developmental stages. Of particular interest, genes in clusters 3 (n = 236) and 6 (n = 1409) were significantly up-regulated on Days 16 and 19, suggesting a role in maternal recognition and initiation of implantation. This transcriptome analysis of the bovine conceptus will provide a blueprint of the dynamic changes in gene expression occurring during maternal recognition and implantation and will complement existing knowledge of the temporal changes in the endometrial transcriptome, thus facilitating a better understanding of conceptus-maternal cross talk during the peri-implantation period of pregnancy.

overall_design: mRNA-seq study of bovine conceptuses at 5 stages of development  (days 7, 10, 13, 16 and 19) post fertilization.

first: Solomon

last: Mamo

first: Jai

middle: P

last: Mehta

first: Trudee

last: Fair

first: Tom

last: Spencer

first: Fuller

middle: W

last: Bazer

first: Patrick

last: Lonergan


title: Domains of genomewide gene expression dysregulation in Down syndrome [RRBS]

first: Audrey

last: Letourneau

overall_design: Reduced representation bisulfite sequencing in monozygotic twins discordant for trisomy 21

first: Federico

middle: A

last: Santoni

first: Ximena

last: Bonilla

last: Sailani

first: David

last: Gonzalez

first: Jop

last: Kind

first: Claire

last: Chevalier

first: Robert

last: Thurman

first: Richard

last: Sandstrom

first: Youssef

last: Hibaoui

first: Konstantin

last: Popadin

first: Emilie

last: Falconnet

first: Maryline

last: Gagnebin

first: Corinne

last: Gehrig

first: Anne

last: Vannier

first: Michel

last: Guipponi

first: Laurent

last: Farinelli

first: Daniel

last: Robyr

first: Marco

last: Garieri


title: Whole transcriptome analysis of postmortem human hippocampus dentate gyrus granlule cells

first: Ruth

last: Kohen

address: 1959 Pacific Ave NE

department: Psychiatry and Behavioral Sciences

institute: University of Washington

summary: We investigated the transcriptome of dentate gyrus (DG) granule cells in postmortem hippocampus from 79 subjects with mental illness (schizophrenia, bipolar disorder, major depression) or non-psychiatric controls.

overall_design: Material for RNA-seq analysis was harvested from tissue slides using laser capture microdissection (LCM) and aRNA amplification. Equimolar amounts of triplicate aRNA samples for each of the 79 subjects were then pooled for the preparation of sequencing libraries. Sequencing libraries were prepared using Applied Biosystem's Total RNA Sequencing Kit, following the directions for Whole Transcriptome Libraries, and analyzed with an Applied Biosystems SOLiD 4 high-throughput sequencer. We compared the performance of different normalization methods (length normalization vs. noise reductin scaling), and assembled evidence for dysruption of signaling by the micro RNA miR-182 in subjects with major depression and schizophrenia.

first: Adrian

last: Dobra

first: Julia

middle: H

last: Tracy

first: Eric

last: Haugen


title: RNAseq to investigate transcriptional changes in human MM cell lines due to panobinostat, 5-Azacytidine, panobinostat+5-Azacytidine or n-methyl-2-pyrroldine (NMP) treatments.

city: Melbourne

first: Geoffrey

middle: Mark

last: Matthews

state: Victoria

address: St Andrews Place

department: Cancer Therapeutics

country: Australia

institute: Peter MacCallum Cancer Centre

summary:
Purpose: We applied RNA sequencing technology for high-throughput analysis of transcriptional changes within human MM cell lines JJN3 and U266 due to individual and combination drug treatment.
Methods: JJN3 and U266 cells were treated with pan-HDACi panbobinostat, DNMTi 5-Azacytidine, panobinostat+5-Azacytidine or NMP for 4h or 24h in triplicate and transcriptional changes assessed by RNAseq using Illumina HiSeq platform. Specifically, JJN3 cells were treated with 10nM panobinostat, 2.5µM 5-Azacytidine, panobinostat+5-Azacytidine (at given doses), or 10mM NMP. U266 cells were treated with 10nM panobinostat, 10µM 5-Azacytidine, panobinostat+5-Azacytidine (at given doses), or 10mM NMP.
Results: We report unique and overlapping transcriptional signatures that lead to the induction of apoptosis in human MM cell lines in a cell-specific manner due to individual or combination treatments.
Conclusions: A detailed analysis of differential transcriptional events in human MM cell lines due to HDACi, DNMTi, HDACi+DNMTi and NMP appear to define the molecular events leading to apoptosis and drug mechanism of action.

overall_design: We tested triplicate experiments at 4h and 24hr time points in JJN3 and U266 cell lines against vehicle control treated cells.

first: Jake

last: Shortt


title: High throughput quantitative whole transcriptome analysis of distal mouse lung epithelial cells from various developmental stages (E14.5, E16.5, E18.5 and adult)

city: Stanford CA

first: Barbara

last: Treutlein

address: 318 Campus Drive

department: Bioengineering

institute: Stanford University

summary: We used microfluidic single cell RNA-seq on 198 individual mouse lung epithelial cells at 4 different stages throughout development to measure the transcriptional states which define the developmental and cellular hierarchy of the distal mouse lung epithelium. We classified 80 cells comprising the distal lung epithelium at E18.5 into distinct populations using an unbiased genome-wide approach that did not require a priori knowledge of the underlying cell types or prior purification of cell types.  This “reverse tissue engineering” approach confirmed the basic outlines of the conventional model of cell type diversity in the distal lung and led to the discovery of a large number of novel transcriptional regulators and cell type markers that discriminate between the different populations. Moreover, we reconstructed the steps during maturation of bipotential progenitors into both alveolar lineages based on the presence of undifferentiated, differentiated as well as differentiation intermediate cells at the single time point E18.5. Finally, we followed Sftpc-positive cells throughout their lifecycle (E14.5, E16.5, E18.5, adult) and identified 7 gene sets that differentiate between the multipotential, bipotential, mature, as well as intermediate states of the AT2 lineage.

overall_design: 198 single-cell transcriptomes from mouse lung epithelium were analyzed in total, two 200-cell bulk control samples as well as one no-cell control; All single cell and control samples contain 92 external RNA spike-ins; For time point E18.5, three individual experiments were performed using 3 different pregnant mice (3 biological replicates): Replicate 1 (pooled sibling lungs) yielded 20 single cell transcriptomes, replicate 2 (one single embryonic lung) yielded 34 single cell transcriptomes and replicate 3 (pooled siibling lungs) yielded 26 single cell transcriptomes; In addition, a 200-cell bulk control sample was prepared for E18.5 replicate 1 and E18.5 replicate 3 experiments; A no-cell control sample was generated for the E18.5 replicate 1 experiment; For time point E14.5, one experiment (one pregnant mouse, pooled sibling lungs) was performed yielding 45 single cell transcriptomes; For time point E16.5, one experiment (one pregnant mouse, pooled sibling lungs) was performed yielding 27 single cell transcriptomes; For the adult time point, one 107 day old mouse was used and transcriptomes of 46 single cells were obtained; All single cell samples were processed on the microfluidic platform, 200-cell-bulk and no-cell control samples were processed in microliter volumes in PCR tubes.

first: Stephen

last: Quake


title: Nuclear pore complex evolution: A trypanosome Mlp analog functions in chromosomal segregation but lacks transcriptional barrier activity

status: Public on Apr 03 2014

first: Alexander

last: Ratushny

address: 307 Westlake Avenue North, Suite 500

institute: Seattle BioMed

summary: The nuclear pore complex (NPC) has dual roles in nucleocytoplasmic transport and chromatin organisation. In many eukaryotes the coiled coil Mlp/Tpr proteins of the NPC nuclear basket have specific roles in interactions with chromatin and defining specialised regions of active transcription, while Mlp2 associates with the mitotic spindle in a cell-cycle dependent manner. We previously identified two putative Mlp-related proteins in African trypanosomes, TbNup110 and TbNup92, the latter of which associates with the spindle. We now provide evidence for independent ancestry for TbNup92/TbNup110 and Mlp/Tpr proteins. However, TbNup92 is required for correct chromosome segregation, with knockout cells exhibiting microaneuploidy and low fidelity telomere segregation. Further, TbNup92 is intimately associated with the mitotic spindle and spindle anchor site, but apparently has minimal roles in the control of gene transcription, indicating that TbNup92 lacks major barrier activity. TbNup92 therefore acts as a functional analog of Mlp/Tpr proteins, and together with the lamina analog NUP-1, represents a cohort of novel proteins operating at the nuclear periphery of trypanosomes, uncovering complex evolutionary trajectories for the NPC and nuclear lamina.

overall_design: Whole transcriptome comparison between parental and TbNup92Δ cells

first: Jennifer

last: Holden

first: Ludek

last: Koreny

first: Samson

last: Obado

middle: V

first: Wei-Ming

last: Chen

first: Steven

last: Kelly

first: Brian

last: Chait

first: John

last: Aitchison

first: Michael

last: Rout

first: Mark

last: Field


title: Integrated genomic analyses of adrenocortical tumors (SNP array, DNA methylation, mRNA and miRNA expression).

city: Paris

first: Nabila

last: Elarouci

address: 14 rue Corvisart

department: Recherche

institute: Ligue Nationale contre le Cancer

type: Methylation profiling by genome tiling array

type: Genome binding/occupancy profiling by array


title: Different miRNA expression profiles between human breast cancer turmors and serum

status: Public on Apr 10 2014

city: Wenzhou

first: ZhaoYun

last: Li

state: Zhejiang province

address: Chashan Higher Education Park

department: School of Laboratory Medicine and Life Science

country: China

institute: Wenzhou Medical University

summary: We identified total 174 significantly differentially expressed miRNAs between tumors and the normal tissues, and 109 miRNAs between serum from patients and serum from healthy individuals. There are only 10 common miRNAs. This suggests that only a small portion of tumor miRNAs are released into serum selectively. Interestingly, the expression change pattern of 28 miRNAs is opposite between breast cancer tumors and serum. Functional analysis shows that the differentially expressed miRNAs and their target genes form a complex interaction network affecting many biological processes and involving in cancer-related pathways such as prostate, basal cell carcinoma, acute myeloid leukymia, and more.

overall_design: A bunch of miRNAs have been demonstrated to be aberrantly expressed in cancer tumor tissue and serum. The miRNA signatures identified from the serum samples could serve as potential noninvasive diagnostic markers for breast cancer. The roles of the miRNAs in cancerigenesis is unclear. In this study, we generated the expression profiles of miRNAs from the paired breast cancer tumors, normal, tissue, and serum samples from eight patients using small RNA-sequencing. Serum samples from eight healthy individuals were used as normal controls.

first: Jie

last: Zhu

first: ZhiBao

last: Zheng

first: Jia

last: Wang

first: JinHua

last: Sun

first: Pang

first: XianYing

last: Cheng

first: Lun

last: Fu

first: LiMing

last: Zhang

first: ZuoJun


title: Effect of Aid deletion on the global DNA methylation status of iPS cells

city: Kyoto

first: Ren

last: Shimamoto

address: 53 Shogoin Kawahara-cho, Sakyo-ku

department: Dept. of Reprogramming Science

institute: Center for iPS Cell Research and Application (CiRA)

summary: It has been shown that DNA demethylation has a pivotal role in the generation of induced pluripotent stem (iPS) cells. However, the underlying mechanism is still unclear. Previous reports indicated that activation-induced cytidine deaminase (Aid) is involved in DNA demethylation in several developmental processes and cell fusion-mediated reprogramming. Based on the reports, we hypothesized that Aid may be involved in DNA demethylation during the iPS cell generation. In this study, we examined the function of Aid in iPS cell generation using Aid knockout (Aid-/-) mice expressing a GFP reporter under the control of a pluripotent stem cell marker, Nanog. By the introduction of Oct3/4, Sox2, Klf4 and c-Myc, Nanog-GFP positive iPS cells could be generated from the fibroblasts and primary B cells of Aid-/- mice. The Aid-/- iPS cells showed normal proliferation and gave rise to chimeras, indicating their capacity for self-renewal and pluripotency. The comprehensive DNA methylation analysis by MBD-sequening demonstrated that there were only a few differences between Aid+/+ and Aid-/- iPS cells.

overall_design: Aid+/+ and Aid-/- iPS colonies were generated from Aid+/+ and Aid-/- MEFs and picked up mechanically. The clones were passaged four times on feeder cells and two times on gelatin-coated dishes to exclude the contamination of feeder cells. Subsequently, the genome was isolated. Four Aid+/+ iPS cell clones and four Aid-/- iPS cell clones were compared. To confirm the validity of MBD-sequencing, four Aid+/+ iPS cell clones were compared with three ES cell clones or three Aid+/+ MEFs.


title: The cohesin acetyltransferase Eco1 coordinates rDNA replication and transcription

city: Kansas City

first: Chris

last: Seidel

state: MO

department: Genomics

institute: Stowers Institute

type:
Other
Expression profiling by array

platform_organism:
Schizosaccharomyces pombe
Saccharomyces cerevisiae


title: ERa and PolII ChIP seq from KIKO mouse uterus

city: Research Triangle Park

first: Sylvia

last: Hewitt

state: NC

address: 111 Alexander Dr

department: RDBL

institute: NIEHS

summary:
ChIP-seq from mice with DNA binding mutations in Esr1 (KIKO mouse).
Estrogen Receptor α (ERα) interacts with DNA, directly, or indirectly via other transcription factors, referred to as “tethering”. Evidence for tethering is based on in vitro studies and a widely used “KIKO” mouse model containing mutations that prevent direct estrogen response element (ERE) DNA-binding. KIKO mice are infertile, due in part to the inability of estrogen (E2) to induce uterine epithelial proliferation. To elucidate the molecular events that prevent KIKO uterine growth, regulation of the pro-proliferative E2 target gene Klf4, and of Klf15, a progesterone (P4) target gene that opposes KLF4’s pro-proliferative activity, were evaluated. Klf4 induction was impaired in KIKO uteri; however, Klf15 was induced by E2 rather than by P4. Whole uterine ChIP-seq revealed enrichment of KIKO ERα binding to hormone response elements (HRE), motifs. KIKO binding to HRE motifs was verified using reporter gene and DNA-binding assays. Because the KIKO ERα has HRE DNA-binding activity, we evaluated the “EAAE” ERα, which has more severe DBD mutations, and demonstrated lack of ERE or HRE reporter gene induction or DNA binding. The EAAE mouse has an ERα-null like phenotype, with impaired uterine growth and transcriptional activity. Our findings demonstrate that the KIKO mouse model, which has been used by numerous investigators, cannot be used to establish biological functions for ERα tethering, as KIKO ERα effectively stimulates transcription using HRE motifs. The EAAE-ERα DBD mutant mouse demonstrates that ERα DNA-binding is crucial for biological and transcriptional processes in reproductive tissues, and that ERα-tethering may not contribute to estrogen-responsiveness in vivo.

overall_design: one sample each, vehicle ER-alpha ChIP seq,1 hour estradiol ER-alpha ChIP seq, vehicle RNA polymerase II ChIP seq,1 hour estradiol RNA polymerase II ChIP seq, input DNA

first: Leping

first: Sara

last: Grimm

first: David

last: Fargo

first: Kenneth

last: Korach


title: Genome-wide replication analysis of a yeast eco1 mutant by deep sequencing.

summary: Eco1 is the acetyltransferase that establishes sister-chromatid cohesion during DNA replication. Budding yeast with an eco1 mutation that genocopies Roberts syndrome displaysreduced ribosomal DNA (rDNA) transcription and a transcriptional signature of starvation. Weshow that deleting FOB1, a gene encoding a specific replication fork blocking protein for therDNA region, rescues rRNA production and partially rescues transcription genome-wide. This experiment examines the effect of eco1 mutation on replication genome-wide. Furtherstudies show that deletion of FOB1 corrects the genome-wide replication defects, nucleolarstructure, and rDNA segregation in an eco1 mutant. Our study highlights cohesin's central role atthe rDNA for global control of DNA replication and gene expression.

overall_design: DNA content in eco1-W216G mutant and wt yeast is measured in duplicate by sequencing at 0, 20, and 40 minutes following release from G1 arrest.

first: Shuai

last: Lu

first: Jennifer

middle: L

last: Gerton


title: Incorporation of transposable element small RNAs into the Arabidopsis AGO1 protein

city: Columbus

first: Richard

middle: Keith

last: Slotkin

state: OH

address: 318 West 12th Ave.

department: Molecular Genetics

institute: The Ohio State University

summary:
Background: Transposable element 24 nucleotide small RNAs are not efficiently incorporated into the AGO1 protein, which is involved in endogenous RNAi and gene regulation through the microRNA and tasiRNA pathways.

Results: The AGO1 protein incorporates large quantities of transposable element siRNAs when transposable elements are epigenetically activated and transcribed. The incorporation of transposable element siRNAs is at the expense of the most abundant microRNAs. These transposable element siRNAs can act as tasiRNAs, regulating genes that they have partial complementarity to.

Conclusion: Transposable element small RNAs are more dynamic than previously thought. They can be incorporated into AGO1 and regulate genes.

overall_design: Three biological replicates of small RNA sequencing from two genotypes

first: R

middle: K

first: Kaushik

last: Panda


title: The 8q24 cleft lip susceptibility locus is a remote Myc enhancer required for normal face morphology

city: Heidelberg

first: Massimo

last: Petretich

address: Meyerhofstraße 1

department: Developmental biology

institute: EMBL

summary: A 640kb non-coding interval at 8q24 has been associated with an increased risk of non-syndromic cleft lip and palate (CLP) in humans, but the genes and pathways involved in this genetic susceptibility have remained elusive. With a large series of rearrangements engineered over the syntenic mouse region, we showed that this interval contains very remote cis-acting enhancers that control c-myc expression in the developing face. Deletion of this interval led to mild alteration of facial morphologies in mice and, sporadically, to CLP. At a molecular level, we identified mis-expression of several downstream genes, highlighting a combined impact on cranio-facial developmental network and general metabolic capacity. This dual molecular etiology may account for the prominent role to the 8q24 region in human facial dysmorphologies.

overall_design: ChIP-seq and transcriptomics analysis in wt or/and mutant mice

first: Veli

last: Uslu

first: François

last: Spitz


title: ER  and RNA PolII ChIP-seq in WT and ER  mutant mouse uterus

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

overall_design: Refer to individual Series


title: Quantifying ChIP-seq data: a spiking method providing an internal reference for sample-to-sample normalization.

status: Public on Apr 02 2014

city: Lausanne

first: Gergana

last: Bounova

state: VD

address: Quartier Sorge, bâtiment Génopode

department: Center for Integrative Genomics

institute: University of Lausanne

summary:
Purpose:  Description of a spike-adjusting-method (SAM) to normalize ChIP-seq data .
Methods: We performed ChIP-seq of POLR3D and POLR2B with mouse liver supplemented with 2.5% of human DNA. Human DNA will be used as an internal control for ChIP-seq quantification.
Results: We show that using the SAM for ChIP-seq quantification improve similarity of POLR3D and POLR2B ChIP-seq replicates samples and improve difference between samples originate from different conditions.
Conclusions: The SAM improves comparison of ChIP-seq samples, either by increasing similarity between replicates or by emphasise differences between conditions.

overall_design: Chromatin Immuno-precipitations were performed with antibodies directed against POLR3D (Pol III) and POLR2B (Pol II) using mouse liver material supplemented with human DNA. Immuno-precipitated DNA was next sequenced using Illumina HiSeq. Three different concentrations of human spiked DNA were tested for the Pol III ChIP (2.5%, 5% and 10%). We also sequenced the corresponding inputs (crosslinked DNA from mouse liver). Two concentrations of human spiked DNA (5% and 10%) were tested for the Pol 2 ChIP. We also sequenced the corresponding inputs (crosslinked DNA from mouse liver).

first: Nicolas

last: Bonhoure

last: Bernasconi

first: Viviane

last: Praz

first: Fabienne

last: Lammers

first: Donatella

last: Canella

first: Ian

middle: M

last: Willis

first: Winship

last: Herr

first: Nouria

last: Hernandez

first: Mauro

last: Delorenzi


title: Domains of genomewide gene expression dysregulation in Down syndrome [ChIP-seq]

first: Audrey

last: Letourneau

overall_design: H3K4me3 ChIP-Seq profiling of monozygotic twins discordant for trisomy 21.

first: Federico

last: Santoni

first: Ximena

last: Bonilla

last: Sailani

first: David

last: Gonzalez

first: Jop

last: Kind

first: Claire

last: Chevalier

first: Robert

last: Thurman

last: Sandstrom

first: Youssef

last: Hibaoui

first: Konstantin

last: Popadin

first: Emilie

last: Falconnet

first: Maryline

last: Gagnebin

first: Corinne

last: Gehrig

first: Anne

last: Vannier

first: Michel

last: Guipponi

first: Laurent

last: Farinelli

first: Daniel

last: Robyr

first: Marco

last: Garieri

first: Eugenia

last: Migliavacca

first: Christelle

last: Borel

first: Samuel

last: Deutsch

first: Anis

last: Feki

first: John

last: Stamatoyannopoulos

first: Yann

last: Herault

first: Bas

last: van Steensel

first: Roderic

last: Guigo

first: Stylianos

last: Antonarakis


title: Dissection of thousands of cell type-specific enhancers identifies dinucleotide repeat motifs as general enhancer features

city: Vienna

first: J. Omar

last: Yanez-Cuna

address: Dr. Bohr-Gasse 7

country: Austria

institute: Research Institute of Molecular Pathology

summary:
Gene expression is determined by genomic elements called enhancers, which contain short motifs bound by different transcription factors (TFs). However, how enhancer sequences and TF motifs relate to enhancer activity is unknown and general sequence requirements for enhancers or comprehensive sets of important enhancer sequence elements have remained elusive.

Here, we computationally dissect thousands of functional enhancer sequences from three different Drosophila cell lines. We find that the enhancers display distinct cis-regulatory sequence signatures, which are predictive of the enhancers’ cell type-specific or broad activities. These signatures contain transcription factor motifs and a novel class of enhancer sequence elements, dinucleotide repeat motifs (DRMs). DRMs are highly enriched in enhancers, particularly in enhancers that are broadly active across different cell types. We experimentally validate the importance of the identified TF motifs and DRMs for enhancer function and show that they can be sufficient to create an active enhancer de novo from non-functional sequence. The function of DRMs as a novel class of general enhancer features that are also enriched in human regulatory regions might explain their implication in several diseases and provides important insights into gene regulation.

overall_design: STARR-seq was performed in BG3 cells with paired-end sequencing in two replicates and respective inputs.

first: J

middle: O

last: Yáñez-Cuna

first: Cosmas

middle: D

last: Arnold

first: Gerald

last: Stampfel

first: Lukasz

last: Boryn

last: Gerlach

first: Martina

last: Rath

first: Alexander

last: Stark


title: Identification of new regulators of the three dimensional Polycomb organization by a microscopy-based genome-wide RNAi screen

city: Montpellier

first: Giacomo

last: Cavalli

address: IGH - CNRS, 141, rue de la Cardonille

institute: CNRS

summary: Polycomb group (PcG) proteins dynamically define cellular identities through epigenetic repression of key developmental genes. PcG target gene repression can be stabilized through the interaction in the nucleus at PcG foci. Here, we report the results of a high-resolution microscopy genome-wide RNAi screen that identifies 129 genes that regulate the nuclear organization of Pc foci. Candidate genes include PcG components and chromatin factors, as well as many novel protein-modifying enzymes, including components of the SUMOylation pathway. In the absence of SUMO Pc foci coagulate into larger aggregates. Conversely, loss of function of the SUMO peptidase velo disperses Pc foci. Moreover, SUMO and velo colocalize with PcG proteins at PREs and Pc SUMOylation affects its chromatin targeting, suggesting that the dynamic regulation of Pc SUMOylation regulates PcG-mediated silencing by modulating the kinetics of Pc binding to chromatin as well as its ability to form Polycomb foci.

overall_design: ChIP-Seq mapping of Polycomb (PC), SUMO and Velo  on Drosophila Melanogaster

first: Inma

last: Gonzalez

first: Julio

last: Mateos-Langerak

first: Aubin

last: Thomas

first: Thierry

last: Cheutin


title: Transcriptional landscape of Rag2 -/- thymocytes [Mnase-Seq]

status: Public on Apr 01 2014

city: Freiburg

first: Pierre

middle: Daniel

last: Cauchy

state: Baden-Württemberg

address: Breisacher Str. 115

department: Zentrum für Translationale Zellforschung

institute: University Medical Center Freiburg

summary: We performed ChIP-Seq for hallmark TFs (Ets1, Runx1), histone modification marks (H3K4me1, H3K4me2, H3K4me3, H3K27me3, H3K36me3), total RNA Pol II, short RNA-Seq as well as nucleosome mapping mainly in murine Rag2 -/- thymocytes. We also performed ChIP-Seq for E47 as well as nucleosome mapping, gene expression microarray analysis in CD4+ CD8+ DP thymocytes. Overall, we find a key role for the transcription factor Ets1, contributing towards alpha beta T cell lineage commitment via differential transactivation of stage-specific genes orchestrated by dynamic, co-association -mediated chromatin remodeling, as well as transcription dependent generation of a specialized chromatin structure at the TCR beta locus.

overall_design:
Genome-wide analysis via ChIP-Seq for Ets1, Runx1, total RNA Pol II binding,  H3K4me1, H3K4me2, H3K4me3, H3K27me3, H3K36me3, short RNA-Seq, Mnase-Seq in murine Rag2 -/- thymocytes, ChIP-Seq for E47, Mnase-Seq and gene expression microarray analysis in DP thymocytes

This Series represents Mnase-Seq data.

first: Joaquin

last: Zacarias-Cabeza

first: Laurent

last: Vanhille

first: Frederic

last: Koch

first: Romain

last: Fenouil

first: Angelica

last: Santana

first: Aleksandra

last: Pekowska

first: Ahmad

last: Maqbool

first: Hélène

last: Holota

first: Aurélien

last: Griffon

first: Jean

last: Imbert

last: Ferrier

first: Salvatore

last: Spicuglia

first: Jean-Christophe

last: Andrau


title: Genome-wide expression analysis of young, senescent and p38MAPK-inhibitited senescent human fibroblasts.

city: St. Louis

first: Maureen

middle: J

last: Donlin

address: 1100 South Grand

department: Biochemistry and Molecular Biology

institute: Saint Louis University School of Medicine

summary: We utilized whole genome sequencing of mRNA (RNA-seq) to understand the extent to which the senescence-associated secretory phenotype is regulated by p38MAPK

overall_design: Examination of replicates of young, senescent or p38MAPK-inhibited senescent BJ human foreskin fibroblasts.

first: Sheila

last: Stewart

first: Elise

last: Alspach


title: Evidence for an early endometrial response to pregnancy in cattle: both dependent upon and independent of interferon tau

first: Paul

last: McGettigan

institute: University College Dublin

summary: The aims of this study were to 1) identify the earliest transcriptional response of the bovine endometrium to the presence of the conceptus (using RNAseq), 2) investigate if these genes are regulated by interferon tau (IFNT) in vivo, and 3) determine if they are predictive of the pregnancy status of postpartum dairy cows. RNAseq identified 459 differentially expressed genes (DEGs) between pregnant and cyclic endometria on day 16. Quantitative real-time PCR analysis of selected genes revealed PARP12, ZNFX1, HERC6, IFI16, RNF213, and DDX58 expression increased in pregnant compared with cyclic endo- metria on day 16 and were directly upregulated by intrauterine infusion of IFNT in vivo for 2 h (P < 0.05). On day 13 following estrous endometrial expression of nine genes increased [ARHGAP1, MGC127874, LIMS2, TBC1D1, FBXL7, C25H16orf71, LOC507810, ZSWIM4, and one novel gene (ENSBTAT00000050193)] and seven genes decreased (SERBP1, SRGAP2, AL7A1, TBK1, F2RL2, MGC128929, and WBSCR17; P < 0.05) in pregnant compared with cyclic heifers. Of these DEGs, significant differences in expression between pregnant and cyclic endometria were maintained on day 16 for F2RL2, LIMS2, LOC507810, MGC127874, TBC1D1, WBSCR17, and ZSWIM4 (P < 0.05) both their expression was not directly regulated by IFNT in vivo. Analysis of the expression of selected interferon-stimulated genes in blood samples from postpartum dairy cows revealed a significant increase (P < 0.05) in expression of ZXFX1, PARP12, SAMD9, and HERC6 on day 18 following artificial insemination in cows subsequently confirmed pregnant compared with cyclic controls. In conclusion, RNAseq identified a number of novel pregnancy-associated genes in the endometrium of cattle during early pregnancy that are not regulated by IFNT in vivo. In addition, a number of genes that are directly regulated by short term exposure to IFNT in vivo are differentially expressed on day 18 following estrus detection in the blood of postpartum dairy cows depending on their pregnancy status.

overall_design: mRNA profiles of pregnant vs not pregnant cross-bred beef heifers at days 13 and 16 (n=5 per group)

first: Niamh

last: Forde

first: Gillian

middle: B

last: Duffy

first: John

last: Browne

first: Jai

last: Mehta

first: A

last: Kelly

first: N

last: Mansouri-Attia

first: O

last: Sandra

first: Brendan

last: Loftus

first: Mark

last: Crowe

first: Trudee

last: Fair

first: James

middle: F

last: Roche

first: Patrick

last: Lonergan

first: Alexander

middle: C

last: Evans


title: HDAC-regulated myomiRs control BAF60 variant exchange and direct the functional phenotype of fibroadipogenic progenitors in dystrophic muscles

first: Iros

last: Barozzi

address: Borschkegasse 8a

institute: Medical University Vienna

summary: Fibro-adipogenic progenitors (FAPs) are emerging cellular components of the skeletal muscle regenerative environment. The alternative functional phenotype of FAPs - either supportive of muscle regeneration or promoting fibro-adipogenic degeneration – is a key determinant in the pathogenesis of muscular diseases, including Duchenne Muscular Dystrophy (DMD). However, the molecular regulation of FAPs is still unknown. We show here that an “HDAC-myomiR-BAF60 variant network” regulates the functional phenotype of FAPs in dystrophic muscles of mdx mice. Combinatorial analysis of gene expression microarray and genome-wide chromatin remodeling by Nuclease accessibility (NA)-seq revealed that HDAC inhibitors de-repress a “latent” myogenic program in FAPs from dystrophic muscles at early stages of disease progression. In these cells HDAC inhibition promoted the expression of two core components of the myogenic transcriptional machinery, MyoD and BAF60C, and upregulated the myomiRs (miRs) 1.2, 133 and 206, which target two alternative BAF60 variants (BAF60A and B) ultimately leading to the activation of a pro-myogenic program at the expense of the fibro-adipogenic phenotype. By contrast, FAPs from dystrophic muscles at late stages of disease progression displayed resistance to HDACi-induced chromatin remodeling at myogenic loci and fail to activate the pro-myogenic phenotype. These results reveal a previously unappreciated disease stage-specific bipotency of mesenchimal cells within the regenerative environment of dystrophic muscles. Resolution of such bi-potency by epigenetic interventions, such as HDACi, provides the molecular rationale for the in situ reprogramming of target cells to promote therapeutic regeneration of dystrophic muscles.

overall_design: Genome-wide chromatin accessibility patterns in FAPs cells derived from mdx mice at different ages treated either with HDAC inhibitor Trichostatin A (TSA) or with vehicle (saline solution) were assessed using NA-seq (PMID: 19289091)

first: V

last: Saccone

first: S

last: Consalvi

first: L

last: Giordani

first: C

last: Mozzetta

first: T

last: Ryan

last: Madaro

first: Munoz

last: Rojas

first: B

last: Bruneau

first: P

last: Fasanaro

first: I

last: Termanini

first: M

last: Mercola

last: Minucci

last: Puri


title: Quality assessment and control of tissue specific RNA-seq libraries of Drosophila transgenic RNAi models

city: Lisbon

first: Andreia

middle: Jesus

last: Amaral

state: Estremadura

address: Camp Grande

department: Gene Expression and Bioinformatics Unit

country: Portugal

institute: Center for Biosystems, Integrative and Functional Genomics

summary: Next-generation RNA-sequencing (RNA-seq) is rapidly emerging as the technology of choice for whole-transcriptome studies. However, RNA-seq is not a bias free technique. It requires large amounts of RNA and library preparation can introduce multiple artifacts, compounded by problems from later stages in the process. Nevertheless, RNA-seq is increasingly used in multiple studies, including the characterization of tissue-specific transcriptomes from invertebrate models of human disease. The generation of samples in this context is complex, involving the establishment of mutant strains and the delicate contamination prone process of dissecting the target tissue. Moreover, in order achieve the required amount of RNA, multiple samples need to be pooled. Such datasets pose extra challenges due to the large variability that may occur between similar pools, mostly due to the presence of cells from surrounding tissues. Therefore, in addition to standard quality control of RNA-seq data, analytical procedures for control of 'biological quality’ are critical for successful comparison of gene expression profiles. In this study, the transcriptome of the central nervous system of a Drosophila transgenic strain with neuronal-specific RNAi of a ubiquitous gene was profiled using RNA-seq. While conducting our investigation, we observed the existence of an unusual variance in the libraries and showed that the expression profile of a small panel of genes was sufficient to identify libraries with low levels of contamination from neighboring tissues, enabling the selection of a robust dataset for differential expression analysis. We additionally analyzed the potential of profiling a complex tissue to identify cell-type specific changes in response to target gene down-regulation. Finally, we demonstrated that trimming 5’ ends of reads decreases nucleotide frequency biases, increasing the coverage of protein coding genes with a potential positive impact in the incurrence of sampling errors.

overall_design: Central Nervous System profiles of third instar larvae WT (N=6), C24 (N=7), and X7/C24 D. melanogaster were generated by sequencing using the Illumina Hiseq2000.

first: Francisco

last: Brito

first: Tamar

last: Chobanyan

first: Seiko

last: Yoshikawa

first: Takakazu

last: Yokokura

first: David

last: Van Vactor

first: Margarida

last: Gama-Carvalho


title: DNA methylation in adult queend and worker bees

city: Canberra

first: Sylvain

last: Foret

state: ACT

address: Gould Building #116

department: Research School of Biology

institute: The Australian National University

summary: In honey bees (Apis mellifera) the behaviorally and reproductively distinct queen and worker female castes derive from the same genome as a result of differential intake of royal jelly and are implemented in concert with DNA methylation. To determine if these very different diet-controlled phenotypes correlate with unique brain methylomes, we conducted a study to determine the methyl cytosine (mC) distribution in the brains of queens and workers at single-base-pair resolution using shotgun bisulfite sequencing technology. The whole-genome sequencing was validated by deep 454 sequencing of selected amplicons representing eight methylated genes. We found that nearly all mCs are located in CpG dinucleotides in the exons of 5,854 genes showing greater sequence conservation than non-methylated genes. Over 550 genes show significant methylation differences between queens and workers, revealing the intricate dynamics of methylation patterns. The distinctiveness of the differentially methylated genes is underscored by their intermediate CpG densities relative to drastically CpG-depleted methylated genes and to CpG-richer non-methylated genes. We find a strong correlation between methylation patterns and splicing sites including those that have the potential to generate alternative exons. We validate our genome-wide analyses by a detailed examination of two transcript variants encoded by one of the differentially methylated genes. The link between methylation and splicing is further supported by the differential methylation of genes belonging to the histone gene family. We propose that modulation of alternative splicing is one mechanism by which DNA methylation could be linked to gene regulation in the honey bee. Our study describes a level of molecular diversity previously unknown in honey bees that might be important for generating phenotypic flexibility not only during development but also in the adult post-mitotic brain.

overall_design: This study looked at the DNA methylation levels of queen and worker honeybees

first: Frank

last: Lyko

first: Robert

last: Kucharski

first: Stephan

last: Wolf

first: Cassandra

last: Falckenhayn

first: Ryszard

last: Maleszka


title: Mapping of Notch1 binding sites in a mouse immature T cell line

city: Illkirch

first: Susan

last: Chan

address: 1, rue Laurent Fries

institute: IGBMC

summary: We used an immature mouse T cell line engineered to express a biotinylated form of the cleaved form of Notch1 (ICN1). ICN1-bound sites were precipitated with streptavidin-coated beads and subjected to ChIP-sequencing.

overall_design: Beko cells correspond to a spontaneous T lymphoma immature cell line derived from a TCRb deficient mouse. These cells were engineered to express a biotin-tagged-ICN1 and the bacterial biotin ligase BirA (Bio-ICN1) or just BirA as a control (Bio). Chromatin from both cell lines was subjected to strepatavidin-mediated precipitation and subjected to sequencing with the Illumina GAII sequencer as single end 36 base pair reads.

first: Philippe

last: Kastner


title: Breast tumor specific mutation in GATA3 impacts protein stability and genomic location

first: Paul

last: Wade

address: 111 TW Alexander Drive

department: Laboratory of Molecular Carcinogenesis

summary: The transcription factor GATA3 is a favorable prognostic indicator in estrogen receptor-α (ERα)-positive breast tumors in which it participates with ERa and FOXA1 in a complex transcriptional regulatory program driving tumor growth. Paradoxically, GATA3 mutations are frequent in breast cancer and have been classified as drivers. To elucidate the contribution(s) of GATA3 alterations to oncogenesis, we studied two breast cancer cell lines, MCF7, which carries a heterozygous frameshift mutation in the second zinc finger of GATA3, and T47D, wild-type at this locus. Heterozygosity for the truncating mutation conferred protection from regulated turnover of GATA3, ERa and FOXA1 following estrogen stimulation.  Thus, mutant GATA3 uncouples protein-level regulation of master regulatory transcription factors from hormone action. Consistent with increased protein stability, ChIP-seq profiling identified stronger accumulation of GATA3 in cells bearing the mutation, albeit with a similar distribution across the genome. We propose that this specific, cancer-derived mutation in GATA3 deregulates physiologic protein turnover, stabilizes GATA3 binding across the genome and modulates the response of mammary epithelial cells to hormone signaling, thus conferring a selective growth advantage.

overall_design: Genome-wide mapping of GATA3 in two cell lines. There were two biological replicates and unchipped (input) DNA was used as reference.

first: Aleksandra

last: Adomas

first: Sara

last: Grimm

first: Christine

last: Malone

first: Motoki

last: Takaku

first: Jennifer

last: Sims


title: Regulation of the mouse heart transcriptome by Celf1

city: Gainesville

first: Eric

middle: T

last: Wang

state: FL

address: 2033 Mowry Rd

institute: University of Florida

summary: The CUG-BP and ETR-3-like factor 1 (Celf1) RNA binding protein plays an important role in heart and muscle development, and is over-expressed in the disease myotonic dystrophy.  Celf1 has known roles in regulation of RNA splicing, RNA stability, and protein translation.  To identify transcriptome-wide targets of the Celf1 protein in heart, we performed RNA-Seq of polyA+ RNA from mice inducibly expressing Celf1 in the heart.

overall_design: Mice were engineered to express the reverse tetracycline trans-activator (rtTA) from a heart-specific alpha myosin heavy chain promoter, and an N-terminal Flag-tagged version of the LYLQ isoform of human Celf1 from a tet-inducible promoter.  Mice were fed doxycycline to induce Celf1 expression in heart, and hearts were harvested from 3 mice each at 12 hour, 24 hour, 72 hour, and 7 day time points.  To account for potential doxycycline-dependent effects, control mice were fed doxycycline for 72 hours but these mice did not contain the tet-inducible Celf1 cassette.  In total, 15 hearts were analyzed by RNA-Seq.

first: Amanda

last: Ward

first: Daniel

last: Treacy

first: Thomas

last: Housman

first: Tom

last: Cooper

first: Christopher

last: Burge


title: A signature predicting poor prognosis in gastric and ovarian cancer represents a coordinated macrophage and stromal-response.

city: East Melbourne

first: Rita

middle: Ann

last: Busuttil

state: VIC

address: St Andrews Pl

department: Research

institute: Peter MacCallum Cancer Centre

summary: Genome wide mRNA expression profiling of 94 gastric tumours derived from Australian based cohort was performed. . From this data we identified a cluster of co-expressed genes termed the stromal response cluster which almost perfectly differentiates tumor from its non-malignant gastric tissue and hence can be regarded as a highly tumor-specific gene expression signature.  We show that these genes are consistently co-expressed across a range of independent gastric datasets as well as other cancer types suggesting a conserved functional role in cancer.

overall_design: Profiling of 94 primary gastric tumors on Affymetrix GeneChip Human Genome U133 Plus 2.0 Arrays. All tumors were collected with approvals from Peter MacCallum Cancer Center, Australia; the Research Ethics Review Committee; and signed informed patient consent.

first: Alex

last: Boussioutas

middle: A


title: Digital gene expression analysis of ACMV-infected cassava

city: Shanghai

first: Jiao

last: Liu

address: 300 Fenglin Road

institute: Shanghai Institute of Plant Physiology & Ecology

summary: Cassava mosaic disease caused by cassava begomoviruses is the most serious disease of cassava in Africa. However, the molecular mechanisms leading to symptom development of infected cassava plants are poorly understood. Here a high throughput digital gene expression profiling (DGE) based on Illumina Solexa sequencing technology was used to investigate the global transcriptional response of cassava to the African cassava mosaic virus infection. Results showed that 3,210 genes were differentially expressed in virus-infected cassava leaves. Gene Ontology (GO) term and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis indicated that photosynthesis related genes were most affected, which was consistent with the chlorotic symptom on the infected leaves. The upregulation of chlorophyll degradation genes, e.g. the genes encoding chlorophyllase and pheophorbide a oxygenase, as well as the downregulation of the major apoproteins genes in light harvesting complex II (LHCII) identified by the DGE analysis were confirmed by qRT-PCR. Together with the reduction of chlorophyll b content and fewer grana stacks in the infected leaf cells, this study reveals that the degradation of chlorophyll plays an important role during ACMV symptom development for the first time. Meanwhile, we believe that the non-lethal effect on photosystem is a trick for virus to avoid fierce host immune response and a result of the long-term co-evolution. This study will provide a road map for future investigations into virus symptom development.

overall_design: ACMV-infected cassava leaves mixture from three independent replicates were collected for RNA extractions at 20 dpi. Control samples were harvested from empty agrobacteria treated leaves incubated under the same conditions.

first: Jun

last: Yang

first: Peng

last: Zhang


title: Comparison of Transcriptome Profiles of Human Ehrlichiosis Agents

city: Baltimore

first: Sean

last: Daugherty

state: Maryland

address: 801 W. Baltimore St.

department: Institute for Genome Sciences

institute: University of Maryland School of Medicine

summary: This study is to determine and compare the transcriptomes in two eukaryotic hosts (mammalian host - DH82 canine cell line and tick vector - ISE6 Ixodes scapularis cell line) of eight Ehrlichia chaffeensis strains in three divergent genetic groups, including Arkansas, Heartland, Jax, Liberty, Osceola, St. Vincent, Wakulla, and West Paces, and one Ehrlichia sp. HF strain.

sample_organism: Ehrlichia chaffeensis str. Saint Vincent

overall_design:
RNA samples for the Arkansas (reference), Heartland, Jax, Liberty, Osceola, St. Vincent, Wakulla, and West Paces strains in both tick and canine cell lines, including two uninfected controls, will be isolated in triplicate. As such 108 libraries will be constructed and sequenced corresponding to (8 strains + 1 control) x 2 conditions (tick cells + dog cells) x 3 biological triplicates x 2 molecules sequenced (eukaryotic RNA + prokaryotic RNA).
The transcriptomes are obtained with Illumina HiSeq reads obtained from paired end libraries. The bacterial reads will be mapped against the reference genome and compared through a differential expression analysis. The eukaryotic reads from the same cell types (i.e. tick or canine) will be assembled with Trinity. The reads will then be mapped back to this assembly for the differential expression analysis.

first: Mingqun

last: Lin

first: Sushma

last: Parankush

first: Nikhil

last: Kumar

first: Zhihui

last: Cheng

first: Qingming

last: Xiong

first: Julie

last: Dunning Hotopp

first: Yasuko

last: Rikihisa


title: A Genome-wide RNAi screen identifies factors required for distinct stages of C. elegans piRNA biogenesis

city: Shenzhen

first: Wee Siong Sho

last: Goh

state: Guangdong

address: Shenzhen Bay Laboratory, No. 5, Kelian Road, Goh Lab, A1819, Guangming District

department: Institute of Molecular Physiology

institute: Shenzhen Bay Laboratory

summary: In animals, piRNAs, and their associated Piwi proteins, guard germ cell genomes against mobile genetic elements via an RNAi-like mechanism. In C. elegans, 21U-RNAs comprise the piRNA class and these collaborate with 22G RNAs, via unclear mechanisms, to discriminate self from non-self and selectively and heritably silence the latter. Recent work indicates that 21U-RNAs are post-transcriptional processing products of individual transcription units that produce ~26 nucleotide capped precursors. Yet, nothing is known of how the expression of precursors is controlled or of how primary transcripts give rise to mature small RNAs. We conducted a genome-wide RNAi screen to identify components of the 21U biogenesis machinery. Screening by direct, qPCR-based measurements of mature 21U-RNA levels, we identified 22 genes important for 21U-RNA production, termed TOFUs (Twenty-One-u Fouled Up). We also identified 7 genes that normally repress 21U production. By measuring mature 21U-RNA and precursor levels for the 7 strongest hits from the screen, we have assigned factors to discrete stages of 21U-RNA production. Our work has identified factors separately required for the transcription of 21U precursors, and the processing of these precursors into mature 21U-RNAs, thereby providing an essential resource for studying the biogenesis of this important small RNA class.

overall_design: Small RNA and capped small RNA sequencing from total RNA of C. elegans subjected to different RNAi and different C. elegans mutants

first: Wee

middle: S

first: Gregory

last: Hannon


title: Small RNA sequencing study of C elegans piRNA biogenesis factors

city: London

first: Peter

last: Sarkies

address: Du Cane Road

department: MRC Clinical Sciences Centre

institute: Imperial College London

summary: Identification of defects in piRNAs and piRNA precursors

overall_design:
Sequencing of small RNAs (18-30nt).
Sample treatments:
Secondary siRNAs in C. elegans carry a 5' triphosphate and this has to be removed prior to cloning, hence the use of 5' polyphosphatase treatment to reveal this population.
TAP (tobacco acid pyrophosphatase) is another enzyme that removes 5' triphosphate but it also removes 5' CAP structures- we used this to look for piRNA precursor sequences in C. elegans that have a 5' cap.

first: Eric

last: Miska


title: Abscisic acid regulates plant growth and flowering through SINGLE FLOWER TRUSS pathway in tomato.

first: Han

last: Xiao

address: 300 Fenglin Rd

department: National Key Laboratory of Plant Molecular Genetics

institute: Shanghai Institutes for Biological Sciences

summary: Abscisic acid (ABA) regulates seed and bud dormancy. We show by forward and reverse genetic analysis that the tomato transcription factor SlZFP2 is required for release of bud and seed dormancy through negative regulation of ABA biosynthesis. We also demonstrated that ABA promotes growth and represses flowering in tomato both through transcriptional control on the florigen-encoding gene SINGLE FLOWER TRUSS (SFT) in tomato. To gain further insight on transcriptome changes by overexpresion of HA-SlZFP2, we sequenced two lines of p35S:HA-SlZFP2 in LA1589 background and their nontransgenic siblings on Illumina Hiseq2000 platform.

overall_design: Two homozygous transgenic lines 103 and 104 showing very similar phenotypes in flowering and branching were chosen for profiling gene expression via RNA sequencing. Their respective nontransgenic siblings were served as controls (103N and 104N).

first: Lin

last: Weng


title: SND1 Transcription Factor-Directed Quantitative Functional Hierarchical Genetic Regulatory Network in Wood Formation in Populus trichocarpa

city: Raleigh

first: Vincent

last: Chiang

address: 840 Main Campus Drive

department: Forestry and Environmental Resources

institute: North Carolina State University

summary: We focused on RNA-seq-based full transcriptome responses to PtrSND1-B1 overexpression at 7, 12, and 25 h in stem defferentiating xylem (SDX) protoplasts

overall_design: We transfected PtrSND1-B1 and sGFP into stem differentiating xylem protoplasts and performed RNA-seq to reveal the whole transcriptome.


title: Mapping of Ikaros and RBPJ binding sites in the mouse T cell leukemia cell line T29

first: Susan

last: Chan

summary: The transcription factor Ikaros represses Notch signaling. Since Ikaros and Notch treanscriptional mediator RBPJ both recognize sequences that contain the same core TGGGAA motif, it was hypothesized that Ikaros represses Notch signaling by targeting Notch response elements and competing with RBPJ for their binding. Here we used the mouse T-cell leukemia cell line T29 to compare the genomic binding profiles of Ikaros and RBPJ by ChIP-seq.

overall_design: The T29 cell line is derived from a Ikaros-deficient T-cell leukemia (Dumortier et al, MCB 26, 209-220, 2006) and exhibits strong Notch activation. We performed two chip-seq experiments with an anti-RBPJ antibody to map RBPJ binding sites. To map Ikaros binding sites, we engineered a T29-derived cell line that expresses a fusion protein between Ikaros and the ligand binding domain of the estrogen receptor (Ik1-ER) which is activated by addition of 4-hydroxy-tamoxifen (4OHT). We used an anti-Ikaros antibody to map the sites bound by Ik1-ER after treatment of the cells with 4OHT. Sequencing were performed with the Illumina GAII sequencer as as single end 36 base pair reads.

first: Philippe

last: Kastner


title: Whole-genome landscape of histone H2AX and γ-H2AX along with related factors in activated cells

city: Daejeon

first: Jungmin

last: Seo

address: #211, BVC, KRIBB, Eoung-dong, Yuseong-gu

department: Bioinformatics Research Institute

country: South Korea

summary: Phosphorylation of the histone variant H2AX forms γ-H2AX, which serves as a marker of DNA repair response. Here we provide ChIP-seq-based maps of histone H2AX, γ-H2AX, H2AZ, INO80, SRCAP, and RNA polymerase II in activated T cells. Matched data for H2AX and γ-H2AX in resting T cells and Jurkat cancer T cells are available in GSE25577.

overall_design: CD4+ T cells were stimulated in two different ways (IL-2 alone or IL-2 plus anti-CD3 and anti-CD28), and H2AX, γ-H2AX, H2AZ, INO80, and SRCAP profiles were examined by ChIP-seq

first: Jung

last: Choi


title: The enhancer and promoter landscape of human regulatory and conventional T cell subpopulations

city: Regensburg

first: Michael

last: Rehli

address: F.-J.-Strauss-Allee 11

department: Internal Med III

institute: University Hospital Regensburg

summary: CD4+CD25+FOXP3+ human regulatory T cells (Treg) are essential for self-tolerance and immune homeostasis. Here, we generated genome-wide maps of poised and active enhancer elements marked by histone H3 lysine 4 monomethylation and histone H3 lysine 27 acetylation for CD4+CD25highCD45RA+ naive and CD4+CD25highCD45RA- memory Treg and their CD25- conventional T cell (Tconv) counterparts after in vitro expansion . In addition we generated genome-wide maps of the transcription factors STAT5, FOXP3, RUNX1 and ETS1 in expanded  CD4+CD25highCD45RA+ Treg- and CD4+CD25- Tconv to elucidate their role in cell type-specific gene regulation.

overall_design: ChIP-seq of 2 histone marks and transcription factors ETS1, STAT5, FOXP3 and RUNX1 in expanded T cell subpopulations

first: Christian

last: Schmidl


title: BCL11A cistrome in mouse erythroleukemia (MEL) cells

city: Memphis

first: Jian

last: Xu

state: Tennessee

address: 262 Danny Thomas Place, MS 345

department: Pathology

institute: St. Jude Children's Research Hospital

summary: ChIP-seq analyses were performed in MEL cells expressing BirA alone or BirA and FLAG-Biotin tagged BCL11A (XL isoform).

overall_design: BCL11A chromatin occupancy in MEL cell line.

first: Stuart

middle: H

last: Orkin


title: Site-Specific Association with Host and Viral Chromatin by KSHV LANA and its Reversal during Lytic Reactivation

city: EMERYVILLE

first: ALEXANDRE

last: MERCIER

state: CA

address: 5300 CHIRON WAY

department: INFECTIOUS DISEASES

institute: NOVARTIS INSTITUTES FOR BIOMEDICAL RESEARCH

summary: Latency-associated nuclear antigen (LANA), a multifunctional protein expressed by the Kaposi sarcoma-associated herpesvirus (KSHV) in latently-infected cells, is required for stable maintenance of the viral episome. This is mediated by two interactions: LANA binds to specific sequences (LBS1 and 2) on viral DNA, and also engages host histones, tethering the viral genome to host chromosomes in mitosis. LANA has also been suggested to affect host gene expression, but both the mechanism(s) and role of this dysregulation in KSHV biology remain unclear. Here we have examined LANA interactions with host chromatin on a genome-wide scale using ChIP-seq, and show that LANA predominantly targets human genes near their transcriptional start sites (TSSs). These host LANA-binding sites are generally found within transcriptionally active promoters and display striking overrepresentation of a consensus DNA sequence virtually identical to the LBS1 motif in KSHV DNA. Comparison of the ChIP-seq profile with whole transcriptome (RNA-seq) data reveals that few of the genes that are differentially regulated in latent infection are occupied by LANA at their promoters. This suggests that direct LANA binding to promoters is not the prime determinant of altered host transcription in KSHV-infected cells. Most surprisingly, the association of LANA to both host and viral DNA is strongly disrupted during the lytic cycle of KSHV. This disruption can be prevented by the inhibition of viral DNA synthesis, suggesting the existence of novel and potent regulatory mechanisms linked to either viral DNA replication or late gene expression.

overall_design: Profiling of KSHV LANA positioning on the host genome and examination of gene expression from promoters bound by KSHV LANA.

first: Alexandre

last: Mercier

first: Carolina

last: Arias

first: Alexis

last: Madrid

first: Meghan

last: Holdorf

first: Don

last: Ganem


title: Meiotic recombination cold spots in chromosomal cohesion sites [ChIP-Seq]

status: Public on Mar 31 2014

city: Suita

first: Masaru

last: Ito

state: Osaka

address: Yamadaoka 3-2

institute: Osaka University

summary: Meiotic chromosome architecture called “axis-loop structures” and histone modifications have been demonstrated to regulate the Spo11-dependent formation of DNA double-strand breaks (DSBs) that trigger meiotic recombination. Using genome-wide chromatin immunoprecipitation (ChIP) analyses followed by deep sequencing, we compared the genome-wide distribution of the axis protein Rec8 (the kleisin subunit of meiotic cohesin) with that of oligomeric DNA covalently bound to Spo11, indicative of DSB sites. The frequency of DSB sites is overall constant between Rec8 binding sites. However, DSB cold spots are observed in regions spanning ±0.8 kb around Rec8 binding sites. The axis-associated cold spots are not due to exclusion of Spo11 localization from the axis, since ChIP experiments revealed that substantial Spo11 persists at Rec8 binding sites during DSB formation. Spo11 fused with Gal4 DNA binding domain (Gal4BD-Spo11) tethered in close proximity (≤0.8 kb) to Rec8 binding sites hardly forms meiotic DSBs, in contrast with other regions. In addition, H3K4 tri-methylation (H3K4me3) remarkably decreases at Rec8 binding sites. These results suggest that reduced histone H3K4me3 in combination with inactivation of Spo11 activity on the axis discourages DSB hot spot formation.

overall_design:
ChIP-seq analyses of Rec8, Spo11, and Gal4BD-Spo11 on budding yeast meiotic chromosomes
• Distribution of Rec8 in wt and Gal4BD-Spo11-expressing cells at 4h after meiotic induction
• Distribution of Spo11 at 3h, 4h, and 5h after meiotic induction
• Distribution of Gal4BD-Spo11 at 0h after meiotic induction

first: Kazuto

last: Kugou

first: Jeffrey

last: Fawcett

first: Sachiko

last: Mura

first: Sho

last: Ikeda

first: Hideki

last: Innan

first: Kunihiro

last: Ohta


title: Generation and characterization of a tomato DCL3 silencing mutant

city: Bet Dagan

first: Tzahi

last: Arazi

state: ISRAEL

address: Institute of Plant Sciences, ARO the Volcani Center

department: Institute of Plant Sciences

country: Israel

institute: ARO - Volcani Center

summary: DICER-like 3 (DCL3) is a major player in heterochromatic 24-nucleotide siRNA and long miRNA biogenesis and mutants have been characterized from Arabidopsis and rice. Here, a tomato DCL3 mutant was generated through the use of trans-activated artificial microRNA and characterized. Global tomato DCL3 (SlDCL3) silencing was induced by crossing the generated responder line (OP:amiRDCL3) with constitutive driver line. Constitutive trans-activation knocked down SlDCL3 levels by ~77%, resulting in dramatically decreased 24-nucleotide small RNA levels, but a significant increase in 21- and 22- nucleotide small RNAs, which was correlated with specific upregulation of SlDCL4 and SlDCL2b. Moreover, in the majority of small RNA-generating loci an almost complete overlap between the control and 35S>>amiRSlDCL3 siRNA signatures, was observed, strongly suggesting that the reduction in 24-nucleotide siRNAs was compensated by increased biogenesis of 21-22 nt siRNAs from the same genomic sequences. Collectively, these results suggest the requirement of SlDCL3 for the biogenesis of 23-24 nt sRNAs and its substitution by SlDCL4 and SlDCL2b, which function in the biogenesis of 21- and 22-nt small RNAs. In addition, differential expression between control and SlDCL3-silenced small RNAs indicated a significant reduction in the abundance of 24-nt putative long miRNAs, which was validated by northern blots, implicating SlDCL3 in their biogenesis.

overall_design: Examination of small RNA populations in control (35S:LhG4) and SlDCL3 silenced seedlings (35S>>amiRSlDCL3) 2 biological replicates each

first: T

first: M

last: Kravchik

first: S

last: Damodharan


title: The regulation of cambial activity in Cunninghamia lanceolata involves extensive transcriptome remodeling

city: Beijing

first: Jinxing

last: Lin

address: No.20 Nanxincun, Xiangshan

institute: Institute of botany

summary: We sequenced mRNA from dormant, reactivating, and actively growing C. lanceolata vascular cambium using tangential cryosections  to generate the first transcriptome dynamics that may serve as a gene expression profile blueprint for cambium development and wood formation in the conifer.

overall_design: Examination of mRNA levels in dormant, reactivating, and actively growing C. lanceolata vascular cambium, respectively

first: zongbo

last: qiu

first: lichuan

last: wan

first: jinxing

last: lin


title: Increased Neanderthal ancestry in genomic regions associated with lipid catabolism in contemporary Europeans

first: Liu

last: He

address: 320 Yue Yang Road

institute: PICB

summary: While Neanderthals are extinct, fragments of their genome still persist in the genomes of contemporary humans. Here, we show that such Neanderthal-like sequences are not distributed randomly in contemporary human genomes. Specifically, while genome-wide frequency of Neanderthal-like sites is close to 6% in all out-of-Africa populations, genes involved in lipid catabolism contain large excess Neanderthal-like sequences in Europeans (24.3%), but not in Asians (12.4%). While lipid catabolism cannot be assayed in Neanderthals, we took advantage of genetic divergence between human populations, chimpanzees and Neanderthals to predict metabolic divergence expected from the observed excess of Neanderthal gene flow into Europeans. We confirmed predicted changes in lipid catabolism using hydrophobic metabolome measurements in the brain tissue and further linked these metabolic changes to gene expression divergence.

sample_organism:
Pan troglodytes
Homo sapiens

platform_organism:
Pan troglodytes
Homo sapiens

overall_design: 14 human and 6 chimpanzee samples were sequenced.

first: Philipp

last: Khaitovich


title: Isolation and Molecular Characterization of Circulating Melanoma Cells

city: Boston

first: Ben

middle: S.

last: Wittner

state: MA

address: 149 13th Street

department: Center for Cancer Research

institute: Massachusetts General Hospital

summary: Melanoma is an invasive malignancy with a high frequency of blood-borne metastases, but circulating tumor cells (CTCs) have not been readily isolated. We adapted microfluidic CTC capture to a tamoxifen-driven B-RAF/PTEN mouse melanoma model. CTCs were detected in all tumor-bearing mice, rapidly declining after B-RAF inhibitor treatment. CTCs were shed early from localized tumors and a short course of B-RAF inhibition following surgical resection was sufficient to dramatically suppress distant metastases. The large number of CTCs in melanoma-bearing mice enabled comparison of RNA sequencing profiles with the matched primary tumor. A mouse melanoma CTC-derived signature correlated with invasiveness and cellular motility in human melanoma. In patients with metastatic melanoma, CTCs were detected in smaller numbers in patients with metastatic melanoma and declined with successful B-RAF targeted therapy. Together, the capture of CTCs and their molecular characterization provide insight into the hematogenous spread of melanoma.

overall_design: We adapted a microfluidic platform, the HbCTC-Chip (Stott et al., 2010, Pubmed ID: 20930119), to capture melanoma CTCs derived from mouse tumors, using panels of antibodies against melanoma-specific cell surface markers, followed by staining for melanoma antigens and optimized on-chip fluorescent imaging. We used a tamoxifen inducible BRAF(CA/+)/PTEN(flox/flox) melanoma mouse model (Dankort et al., 2009, Pubmed ID: 19282848) derived from a C57BL/6 background. Such mice received focal subcutaneous injection of tamoxifen (Sigma) (50ul at 5mg/ml in 50% EtOH suspension) at the left flank at 6-7 weeks after birth. Blood samples were collected from five mice with high tumor burden following tamoxifen injection. Blood from each mouse was split and processed through the CTC-chips functionalized with anti-CSPG4/MCAM antibody and control IgGs, respectively. Matched primary (from the tamoxifen injection site) and metastatic (from upper or lower back) tumors were harvested from the same mouse and immediately flash-frozen in liquid nitrogen. RNA extraction, single molecular sequencing and determination of Digital Gene Expression was as in Yu et al., 2012 (Pubmed ID: 22763454). One of the five mice yielded no CTC or IgG data. In addition, skin was taken from a sixth mouse, which was a complete wild type C57BL/6 mouse (without the BRAF/PTEN transgenes).The skin was taken after euthanasia of the animal and was processed as the tissue from the matched primary and metastatic tumors.

first: Xi

last: Luo

first: Devarati

last: Mitra

first: Ryan

last: Sullivan

first: Anya

last: Kimura

first: Shiwei

last: Pan

first: Mai

middle: P

last: Hoang

first: Brian

middle: W

last: Brannigan

first: Donald

last: Lawrence

first: Keith

last: Flaherty

first: Lecia

middle: V

last: Sequist

first: Martin

last: McMahon

first: Marcus

last: Bosenberg

first: Shannon

middle: L

last: Stott

first: David

last: Ting

first: Sridhar

last: Ramaswamy

first: Mehmet

last: Toner

middle: E

last: Fisher

first: Shyamala

last: Maheswaran

first: Daniel

last: Haber


title: RNA expression of both host and virus during 24 hour time course

status: Public on Mar 30 2014

first: Amol

middle: Carl

last: Shetty

address: 670 W. Baltimore Street, Floor 3

institute: University of Maryland Baltimore

summary: human lung epithelial cell line infected with hRSV to measure gene expression over time

sample_organism:
Homo sapiens
human respiratory syncytial virus

overall_design: 8 samples, no duplicates

first: T

last: McCarty

first: P

last: Collins


title: Characterization of transcriptome dynamics in response to contact with host cells

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

sample_organism:
Candida albicans
Homo sapiens
Mus musculus

overall_design: Refer to individual Series


title: MNase-seq analysis of Col-0 and chr11-1 chr17-1

city: shanghai

first: yijing

last: zhang

address: 2005 Songhu Road

department: Biochemistry

institute: Fudan University

summary: Chromatin has a highly organized structure with the repeating nucleosome subunits. The position of nucleosomes on the chromatin is dynamically regulated by ATP-dependent chromatin remodeling factors (remodelers), therefore providing specific epigenetic information. However, the in vivo nucleosome distribution pattern in plants and how plant remodelers control the pattern formation are not yet clear. Here we use the Micrococcal Nuclease digestion followed by deep sequencing (MNase-seq) assay to show the genome-wide nucleosome pattern in plants and the role of the Arabidopsis thaliana Imitation Switch (AtISWI) subfamily remodelers in the pattern formation. Our data revealed three patterns in wild-type plants: 1) Promoters have a relative low nucleosome density compared with gene bodies; 2) Nucleosome density is negatively associated with gene expression levels; 3) The evenly and periodically spaced nucleosomes in gene bodies are positively associated with gene expression levels. Double mutations in the AtISWI genes CHROMATIN REMODELING 11 (CHR11) and CHR17 resulted in loss of the evenly-spaced-nucleosome pattern in gene bodies, but did not affect nucleosome density, suggesting that the primary role of AtISWI is to slide nucleosomes in gene bodies for promotion of transcription.

overall_design: The overall design of the experiment: (1) Nuclear extraction from Col-0 and chr11-1 chr17-1. (2) MNase treatment. (3) DNA purification. (4) Library construction. (5) Deep sequencing. (6) Data analysis.

first: Lin

last: Xu

first: Yijing

last: Zhang

first: Guang

last: Li


title: Characterization of transcriptome dynamics in response to contact with host cells [Phase II In Vivo]

summary:
Candida albicans is the major invasive fungal pathogen of humans, causing diseases ranging from superficial mucosal infections to disseminated, systemic infections which are often life threatening. Hematogenously disseminated candidiasis (HDC) has a 47% mortality rate despite current antifungal therapy. An increase in prevalence, as well as an increasing resistance to most of the clinically important antifungal therapies, provides a strong impetus to understand the molecular mechanisms of pathogenesis and the acquisition of drug resistance. This information holds promise to identify novel therapeutic targets. A complete and accurate characterization of how the transcriptome of C. albicans responds to its interaction with cells from the host is an absolute necessity to accomplish this goal.
RNA-seq (deep-sequencing of cDNA) provides an unbiased method to define comprehensively and systematically the transcriptome of an organism. We propose a comprehensive characterization of the C. albicans transcriptome in two different human tissue culture models, using RNA-seq. Such a characterization will shed unprecedented light on how fungal pathogens sense and respond to the host environment and how the host tissue responds to fungal invasion.
We performed a comprehensive characterization of the C. albicans transcriptome in two different murine models of candidiasis (HDC and oropharyngeal candidiasis (OPC)), and two different human tissue culture models, using RNA-seq. Because the two murine models accurately recapitulate the pathology that is observed in clinical cases of candidiasis, we are confident that gene expression levels will provide an accurate representation of what occurs during the course of an infection in humans.  Such a characterization of the in vivo transcriptome will shed unprecedented light on how fungal pathogens sense and respond to the host environment and how the host tissue responds to fungal invasion.
PRJNA211732

sample_organism:
Candida albicans
Mus musculus

platform_organism:
Candida albicans
Mus musculus

overall_design: C. albicans strain SC5314 was used to infect mice in a murine model of systemic candidiasis as well as a murine model of oropharyngeal candidiasis. For the disseminated model, kidneys were harvested at 6 hours, 12 hours, 24 hours and 48 hours after infection. For the oropharyngeal model, tongues were harvest at 1 day, 2 days, 3 days, and 5 days after infection. Total RNA was extracted from all harvested tissues and subject to RNA-seq. We also tested 3 C. albicans mutants in the disseminated model. For the infections with the mutant strains, only 1 timepoint (24 hours post-infection) was analyzed.

first: Scott

last: Filler

first: Yaoping

last: Liu

first: Aaron

last: Mitchell


title: PHF6 OCCUPANCYIN HUMAN T-CELL ACUTE LYMPHOBLASTIC LEUKEMIA

city: Cambridge

first: Richard

last: Young

address: 9 Cambridge Center

institute: Whitehead Institute for Biomedical Research

summary: Here we identify the genome-wide occupancy of PHF6 in T-ALL cells.

overall_design: Human T-ALL cells were cross-linked with formaldehyde for 20 min. DNA was enriched by chromatin immunoprecipitation (ChIP) and analyzed by Solexa sequencing. A sample of whole cell extract (WCE) was sequenced and used as the background to determine enrichment. ChIP was performed using an antibody against total PHF6 (Bethyl A301-451A). This represents the ChIP-seq portion of this dataset.

first: Lee

middle: N

last: Lawton


title: Rate of elongation by RNA polymerase II is influenced by specific gene features and histone modifications

city: Ann Arbor

first: Mats

last: Ljungman

state: Michigan

address: NCRC, B520 Room 1346 2800 Plymouth Rd.

institute: University of Michigan

summary: The rate of transcription elongation plays important roles in the timing of expression of full-length transcripts as well as for the regulation of alternative splicing. In this study we coupled Bru-Seq technology with 5,6-dichlorobenzimidazole 1-β-D-ribofuranoside (DRB) to estimate the elongation rates of over 2,000 individual genes in human cells. This technique, BruDRB-Seq, revealed gene-specific differences in elongation rates with a median rate of around 1.5 kb/min. We found that genes with fast elongation rates showed higher densities of H3K79m2 and H4K20me1 marks compared to slower elongating genes. Furthermore, fast elongation rates had a positive correlation with gene length, low complexity DNA sequence and distance from nearest active transcription unit. Features that negatively correlated with elongation rate included exon density and the number of LINE sequences in the gene. The BruDRB-Seq technique offers new opportunities to interrogate mechanisms of regulation of transcription elongation.

overall_design:
Measurement of RNA Pol II elogation rate. Normal fibroblasts (HF1 and TM), Cockayne syndrome group B fibroblasts, K562 and MCF-7 cells were exposed to DRB for 60 minutes, after which a washout was performed. Nascent RNA was labeled using bromouridine for 10 minutes immediately after the washout. The genomic region extending from actice Trancription Start Sites was used to determine the gene's elongation rate.
Please note that the nf_0h_3* samples are duplicated sample records of GSM1062445 and GSM1062446, for the convenient retrieval of the complete raw data from SRA.

first: Artur

last: Veloso

first: Killeen

last: Kirkconnell

first: Brian

last: Magnuson

first: Benjamin

last: Biewen

first: Michelle

last: Paulsen

first: Thomas

last: Wilson


title: Next-generation sequencing facilitates quantitative analysis of AcKLF5 and unAcKLF5 transcriptomes in xenografts of DU-145 cell line

status: Public on Mar 29 2014

city: Tianjin

first: jintang

last: dong

address: Weijin road

institute: Nankai university

summary: KLF5 possesses both tumor-suppressing and tumor-promoting activities, though the mechanism controlling these opposing functions is unknown. In cultured non-cancerous epithelial cells, KLF5 converts from pro-proliferative to anti-proliferative activity upon TGFβ-induced acetylation, which sequentially alters the KLF5 transcriptional complex and the expression of genes such as p15 and MYC. In nude mice, KLF5 also suppressed tumor growth in an acetylation-dependent manner. Furthermore, deacetylation switched KLF5 to tumor-promoting activity, and blocking TGFβ signaling attenuated the tumor suppressor activity of KLF5. The results from RNA-Seq indicated the different downstream genes of AcKLF5 and unAcKLF5 and the mechanisms that determine the opposing functions of AcKLF5 and unAcKLF5. These results provide novel insights into the mechanism by which KLF5 switches from anti-tumorigenic to pro-tumorigenic function, and also suggest the roles of AcKLF5 and unAcKLF5, respectively, in the tumor-suppressing and tumor-promoting functions of TGFβ.

overall_design: Xenografts of DU-145 cell lines expressing KLF5, KLF5 mutant K369R or the empty vector PLHCX were surgically isolated from mice at day 38 after injection. RNA-Seq was performed using these three groups of xenografts.

first: Jin

last: Dong

first: Xin

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