title: Identifying the genome-wide genetic variation between precocious trifoliate orange and its wild type and developing new markers for genetics research

city: WuHan

first: jin

middle: zhi

last: zhang

address: College of Horticulture and Forestry Science, Huazhong Agricultural University, Wuhan 430070, China

institute: Huazhong Agricultural University

summary: To increase our understanding of the genes involved in flowering in citrus, we performed genome resequencing of an early flowering trifoliate orange mutant (Poncirus trifoliata L. Raf.) and its wild type. At the genome level, 3,932,628 single nucleotide polymorphisms (SNPs), 1,293,383 insertion/deletion polymorphisms (InDels), and 52,135 structural variations (SVs) were identified between the mutant and its wild type based on the citrus reference genome. Based on integrative analysis of resequencing and transcriptome analysis, 233,998 SNPs and 75,836 InDels were also identified between the mutant and its wild type at the transcriptional level. Also, 272 citrus homologous flowering-time transcripts containing genetic variation were also identified. GO and KEGG annotation revealed that the transcripts containing the mutant and the wild-type-specific InDel were involved in diverse biological processes and molecular function. Among these transcripts, there were 131 transcripts that were expressed differently in the two genotypes. When 268 selected InDels were tested on 32 genotypes of the three generas of Rutaceae for the genetic diversity assessment, these InDel-based markers showed high transferability. This work provides important information that will allow a better understanding of the citrus genome and that will be helpful for dissecting the genetic basis of important traits in citrus.

overall_design: Two samples, the terminal bud and the five following buds (the major node position for flower formation) from spring flushes of the MT and the WT 2-year-old trees were collected at flower initiation (after self-pruning).  And then subjected to RNA-seq

first: Chun-Gen

last: Hu

first: Jin-Zhi

last: Zhang


title: Comparative gene expression analysis of chloroquine sensitive and resistant strains of Plasmodium falciparum using RNA-Seq

status: Public on May 06 2016

city: Pondicherry

first: Hiasindh Ashmi

last: Antony

state: Pondicherry

address: Dhanvantri Nagar

department: Microbiology

country: India

institute: Jawaharlal Institute of Postgraduate Medical Education and Research (JIPMER)

type: Expression profiling by high throughput sequencing

summary: Drug resistance in Plasmodium falciparum is an important public health burden since it reverses the malaria control achieved so far. Understanding the mechanism of drug resistance will help us to develop novel drug/vaccine targets for malaria treatment. In the present study, we have used the whole transcriptome sequencing to characterize the transcriptional difference between chloroquine sensitive and resistant P. falciparum strains (3D7 and Dd2). The differential gene expression between these strains was analyzed to understand their phenotypic properties like drug sensitivity, immune evasion and pathways involved.

overall_design: Gene expression analysis of chloroquine sensitive and resistant strains of P. falciparum, grown in the presence and absence of CQ, using Illumina Hiseq 2500 platform.

first: Hiasindh

middle: A

first: Vrushali

last: Pathak

first: Subhash

middle: C

last: Parija

first: Kanjaksha

last: Ghosh

first: Amrita

last: Bhattacherjee


title: Cell freezing protocol suitable for ATAC-Seq on motor neurons derived from human induced pluripotent stem cells

status: Public on May 05 2016

city: Cambridge

first: Pamela

last: Milani

address: 32 Vassar Street

department: Biological Engineering

country: USA

institute: Massachusetts Institute of Technology

type: Genome binding/occupancy profiling by high throughput sequencing

summary: We describe a protocol for freezing neuronal cells that is compatible with ATAC-Seq, producing results that compare well with those generated from fresh cells. We developed our protocol on a disease-relevant cell type, namely motor neurons differentiated from induced pluripotent stem cells from a patient affected by spinal muscular atrophy. We found that while flash-frozen motor neurons are not suitable for ATAC-Seq, the assay is successful with slow-cooled cryopreserved samples. Using this method, we were able to isolate high quality, intact nuclei, and we verified that epigenetic results from fresh and cryopreserved motor neurons agree quantitatively.

overall_design:
We quantitatively compare the results from fresh and cryopreserved motor neurons. We generated sequencing data on three technical replicates from both conditions.

Submitter declares that reads for the transposed naked DNA used as a control will be made available through dbGaP.

first: Renan

last: Escalante-Chong

first: Ernest

last: Fraenkel


title: Genome-wide distribution of proteins Pita, ZIPIC, Zw5 in Drosophila embryos

city: Moscow

first: Pavel

last: Georgiev

address: Vavilova 34/5

country: Russia

institute: Institute of Gene Biology

summary: According to recent models, as yet poorly studied architectural proteins appear to be required for local regulation of enhancer–promoter interactions, as well as for global chromosome organization. Transcription factors ZIPIC, Pita, and Zw5 belong to the class of chromatin insulator proteins and preferentially bind to promoters near the TSS. They are structurally similar in containing the N-terminal zinc finger-associated domain (ZAD) and different numbers of C2H2-type zinc fingers at the C-terminus. We have shown that the ZAD domains of ZIPIC, Pita, and Zw5 usually form homodimers. In Drosophila transgenic lines, these proteins can support long-distance interaction between GAL4 activator and the reporter gene promoter. However, no functional interaction between binding sites for different proteins has been revealed, suggesting that such interactions are highly specific. ZIPIC facilitates long-distance stimulation of reporter gene by GAL4 activator in yeast. Thus, ZAD-containing zinc-finger proteins probably belong to the class of architectural proteins.

overall_design: ChIP-Seq for Pita, ZIPIC, Zw5 proteins from Drosophila embryos

first: Oksana

last: Maksimenko

first: Ivan

last: Kulakovskiy


title: TAp73 is a master regulator of airway multiciliogenesis [sRNA-Seq]

city: Göttingen

first: Orr

last: Shomroni

address: Grisebachstraße 5

country: Germany

institute: DZNE

type: Non-coding RNA profiling by high throughput sequencing

summary: Comparative small RNA seq analysis of WT and global p73KO Mouse Tracheal Epithelial Cell  (MTECs) during the course of their differentiation (Air-Liquid Interface ALI D0, D4, D7, D14) aimed to determine the role of p73 in motile multiciliogenesis.

overall_design: Two independent biological replicates of murine primary airway epithelial cell cultures (MTECs) from wild type and global p73KO mice were differentiated under air-liquid interface (ALI) conditions and harvested  at Day 0, Day 4 , Day 7 and Day 14  post ALI.

first: A

last: Nemajerova

first: U

middle: M

last: Moll

first: S

last: Bonn

first: O

first: T

last: Pena

first: R

last: Vidal


title: Simultaneous pathway activity inference and global gene expression analysis using RNA-sequencing

city: Boston

first: Raivo

last: Kolde

state: MASSACHUSETTS

address: 185 Cambridge St.

department: Center for Computational and Integrative Biology

institute: Massachusetts General Hospital

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

overall_design: Refer to individual Series


title: TAp73 is a master regulator of airway multiciliogenesis [RNA-Seq]

summary: Comparative RNA seq analysis of WT and global p73KO Mouse Tracheal Epithelial Cell  (MTECs) during the course of their differentiation (Air-Liquid Interface ALI D0, D4, D7, D14) aimed to determine the role of p73 in motile multiciliogenesis.

overall_design: Three independent biological replicates of murine primary airway epithelial cell cultures (MTECs) from wild type and global p73KO mice were differentiated under air-liquid interface (ALI) conditions and harvested  at Day 0, Day 4 , Day 7 and Day 14  post ALI.


title: TAp73 is a master regulator of airway multiciliogenesis


title: Transcriptome Analysis of Drosophila Mushroom Body Neurons by Cell Type Reveals Memory-Related Changes in Gene Expression

city: Middlebury College

first: Amanda

last: Crocker

state: Vermont

address: 276 Bicentennial Way

department: Neuroscience

institute: Middlebury College

summary: We report the application of low cell number sequencing of identifiable Drosophila melanogaster neurons following behavior. We demonstate the feasibility of identifying the transcriptome of 5 Mushroom Body output Neurons and 2 classes of Kenyon Cells.  We find these neurons display a diverse repertoire of receptors and signaling transcripts.  This information alone seems to be enough to identify each class of neurons in the study.  In additional we show that aversive long-term memory induces changes in gene transcript levels in a subset of these neurons. This study provides a framework for identifying neuronal classes in Drosophila melanogaster and gaining insight into the interplay between behavior and gene regulation.

overall_design: 5 Mushroom Body output neurons and 2 classes of kenyon cells are used to look at general gene expression and changes following aversive long term memory.   Paired control and trained animals were used and a minimum of 4 pairs up to 6 pairs. Animals were of the same background (w1118).  Animals were aged and parental matched.  Cells were harvested at the same chronological time for the animals across all experiments.  All animals were exposed to 1 minute of each odor and 1 minute of a series of 12 5second 60V shocks. This was considered one block and then the animals had spaced training of each block so there was a 10 minute break between 8 blocks of training.  Trained animals had an odor paired with a shock, control animals received the shock then the odor stimulus.  All cells were harvested usign a patch pipet from living animals on an electrophysiology rig within a half hour of the end of training.  Cells were amplified using the Clontech SMARTer Ultra Low Input RNA version 2 High Volume kit. 2 Brain samples were also collected and 3-4 whole fly samples for each genotype were collected to account for background differences across flies.

first: Mala

last: Murthy


title: Setd1a and NURF mediate chromatin dynamics and gene regulation during erythroid cell differentiation

city: New Haven

first: Vince

last: Schulz

state: CT

address: 333 Cedar St. LMP 4085

department: Department of Pediatrics

institute: Yale University

summary: The modulation of chromatin structure is a key step in transcription regulation in eukaryotic cells. Mammalian erythropoiesis is accompanied by dynamic alterations in chromatin structure and gene expression, but the epigenetic regulators that modulate and coordinate these changes are largely unknown. USF, Setd1a and NURF complexes interact to regulate chromatin architecture in erythropoiesis, but the basis for this regulation is unknown. We studied this process using both murine and human models of erythropoiesis. In human primary erythroid cells, H3K4me3, USF1/2, and BPTF, a component of the NURF complex, were significantly co-enriched at more than 50% of transcription start sites (TSSs) of erythroid genes, associated with repositioning of promoter- or enhancer-associated nucleosomes. Depletion of Setd1a, an H3K4 trimethylase, led to a reduction of promoter-associated H3K4 methylation, inhibition of gene transcription and blockade of erythroid differentiation. This was associated with alterations in NURF complex-mediated nucleosome repositioning at erythroid gene promoters and chromatin accessibility. Setd1a deficiency caused decreased associations between the LCR and the -major globin promoter as well as reduced expression of the adult β-globin gene. In vivo, mice deficient for Setd1a in the erythroid compartment resulted in reduced Ter119/CD71 positive bone marrow erythroblasts, peripheral blood RBCs, and hemoglobin levels. These data indicate that Setd1a and NURF complexes coordinately regulate erythroid promoter chromatin dynamics during erythroid differentiation.

overall_design: CD34+-selected stem and progenitor cells were expanded for three days in the absence of EPO.  The cells were further cultured in the presence of EPO, and formaldehyde crosslinked chromatin was isolated after cells differentiated into R3/R4 nucleated erythroid cells.  Chromatin Immunoprecipitation followed by sequencing (chIP-seq) was performed using antibodies against BPTF, SET1, USF1 and USF2.

first: Ying

last: Li

first: Vincent

middle: P

first: Changwang

last: Deng

first: Guangyao

first: Yong

last: Shen

first: Betsabeh

middle: K

last: Tusi

first: Jared

last: Stees

first: Yi

last: Qiu

first: Lei

last: Zhou

first: Laurie

last: Steiner

first: Jorg

last: Bungert

first: Patrick

middle: G

last: Gallagher

first: Suming

last: Huang


title: Cryptic transcription is the primary driving force for nucleosome instability in Spt16 mutant cells

city: New York

first: zhiguo

last: zhang

state: NY

address: 1130 St. Nicholas Avenue

department: Pediatric and Genetics and Development

institute: Columbia University

summary: FACT consists of two essential subunits Spt16 and Pob3 and is a histone chaperone. Depletion of Spt16 using a an spt16 mutant results in a global alteration of nucleosome positions as well as aberrant transcription. Here we show that the majority of nucleosomal changes at gene body upon Spt16 depletion are independent of gene activity, but correlates with cryptic gene transcription and are suppressed by inhibition of RNA Polymerase II activities. In addition, A small fraction of nucleosomal changes is resistant to Pol II inhibition, and Spt16 is enriched at this subgroup of nucleosomes. Moreover, nucleosomes surrounding the initiation sites of cryptic transcription in the spt16 mutant cells are more dynamic than other regions. These results support a model that Spt16 maintains nucleosome stability locally to inhibit the initiation from cryptic transcription, which that once initiated drives additional nucleosome loss upon Spt16 depletion.

overall_design: To gain further insight into how Spt16 inactivation/depletion alters chromatin structure, we mapped the location of nucleosomes of wild type, spt16(G132D), spt16(G132D) rpb1-1, rpb1-1 cells by high-throughput next generation sequencing of MNase digested fragments.  To resolve the apparent discrepancy, we first analyzed transcription including mRNA and non-coding RNA transcription in wild type, spt16(G132D), spt16(G132D) rpb1-1, rpb1-1 cells using strand-specific RNA-seq.

first: Qing

first: Jianxun

last: Feng

first: Haiyun

last: Gan

first: Zhiguo

last: Zhang


title: The GATA transcription factor gene gtaG is required for terminal differentiation in Dictyostelium.

city: Houston

first: Gad

last: Shaulsky

state: TX

address: One Baylor Plaza

department: Molecular and Human Genetics

institute: Baylor College of Medicine

summary: The GATA transcription factor GtaG is conserved in Dictyostelids and essential for terminal differentiation in Dictyostelium discoideum, but its function is not well understood. Here we show that gtaG is expressed in prestalk cells at the anterior region of fingers and in the extending stalk during culmination. The gtaG- phenotype is cell-autonomous in prestalk cells and non-cell-autonomous in prespore cells. Transcriptome analyses reveal that GtaG regulates prestalk gene expression during cell differentiation before culmination and is required for progression into culmination. GtaG-dependent genes include genetic suppressors of the Dd-STATa-defective phenotype as well as Dd-STATa target-genes, including extra cellular matrix genes. We show that GtaG may be involved in the production of two culmination-signaling molecules, cyclic di-GMP and the spore differentiation factor SDF-1 and that addition of c-di-GMP rescues the gtaG- culmination and spore formation deficiencies. We propose that GtaG is a regulator of terminal differentiation that functions in concert with Dd-STATa and controls culmination through regulating c-di-GMP and SDF-1 production in prestalk cells.

overall_design: Total 28 samples were used for this analysis. 7 developmental timepoints of wild-type and gtaG- strains were analyzed with 2 biological replication.

first: Mariko

last: Katoh-Kurasawa

first: Balaji

last: Santhanam


title: Gene expression differences in yolk sac tissue between wild-type and Zfp36l3 knockout mice

city: Durham

first: Perry

middle: J

last: Blackshear

address: 101 Alexander Dr.

institute: NIEHS

summary: The ZFP36L3 protein is a rodent-specific, placenta- and yolk sac-specific member of the tristetraprolin (TTP) family of CCCH tandem zinc finger proteins.  These proteins bind to AU-rich elements in target mRNAs, and promote their deadenylation and decay.  Mice deficient in ZFP36L3 exhibited decreased neonatal survival rates, but no apparent morphological changes in the placenta or surviving offspring.  Zfp36l3 is paternally imprinted, with profound parent-of-origin effects on gene expression.  RNASeq of KO placental mRNA revealed many significantly affected transcripts, some of which exhibited decreased decay rates in differentiated trophoblast stem cells derived from KO blastocysts.  The type 1 transferrin receptor mRNA was unexpectedly decreased in KO placentas, despite an increase in its stability.  This receptor is critical for placental iron uptake from the maternal circulation, and its decrease was accompanied by decreased iron stores in the KO fetus, suggesting that this intrauterine deficiency might have deleterious consequences in later life.

overall_design: Examination of gene expression differences in yolk sac tissue between wild-type and knockout mice groups with 4 biological replicates in each group

first: Deborah

last: Stumpo

first: Weichun

first: Carol

middle: S

last: Trempus

first: Charles

last: Tucker

first: Leping

last: Li

first: Kimberly

last: Kluckman

first: Donna

last: Bortner


title: Deficiency of the placenta- and yolk sac-specific tristetraprolin family member ZFP36L3 identifies likely mRNA targets and an unexpected link to placental iron metabolism


title: Gene expression differences in mouse placenta tissue bewteen wild-type and Zfp36l3 knockout mice

overall_design: Examination of gene expression differences in mouse placenta tissue bewteen wild-type and Zfp36l3 knockout groups, each with 5 biological replicates


title: 5-hydroxymethylcytosine-mediated alteration of transposon activity associated with Preeclampsia

status: Public on May 09 2016

city: Atlanta

first: Peng

last: Jin

state: Georgia

address: 615 Michael St., Rm 325

department: Human Genetics

institute: Emory University

type: Methylation profiling by high throughput sequencing

summary: Preeclampsia and gestational diabetes mellitus (GDM) are two of the most common clinical conditions during pregnancy that could result in adverse utero environments of the fetus. Fetal exposure to poor environments in uterus also raises the risk of future adulthood disorders known as fetal origins of adult disease (FOAD). Epigenetic modifications like cytosine methylation and histone modification have been proposed to be involved in FOAD. Recent research has implicated 5-hydroxymethylcytosine (5hmC), a DNA base derived from 5-methylcytosine (5mC) via oxidation by ten-eleven translocation (Tet) enzymes, in DNA methylation-related plasticity. Here we show that the expression of Tet2 and 5hmC abundance significantly altered in the umbilical vessels of preeclampsia. Genome-wide profiling of 5hmC revealed differentially hydroxymethylated regions (DhMRs) associated with preeclampsia and GDM, and DhMRs were enriched among the genes involved in unique biological pathways for each condition. In particular, 5hmC significantly changed in selective transposons and led to the alteration of transposon activity in preeclampsia. The 5hmC-mediated alteration of transposon activity was further confirmed using established hypoxia cell culture model and could be rescued by Vitamin C, a known activator of Tet proteins. Together our results suggest that adverse utero environments induced by preeclampsia could influence 5hmC-mediated epigenetic profile and contribute to FOAD.

overall_design: To determine the genome-wide 5hmC distribution in the the umbilical vessels of control, preeclampsia and GDM, we employed a previously established chemical labeling and affinity purification method, coupled with high-throughput sequencing (Song et al, Nature Biotechnology, 2011).

first: Miao

last: Sun


title: TET2 Regulates Mast Cell Differentiation and Proliferation through Catalytic and Non-catalytic Activities.

status: Public on May 04 2016

city: Milano

first: Viviana

last: Piccolo

state: Milano

address: Via Adamello 16

department: Department of Experimental Oncology

country: Italy

institute: European Institute of Oncology (IEO)

summary: Dioxygenases of the TET family impact genome functions by converting 5-methylcytosine (5mC) in DNA to 5-hydroxymethylcytosine (5hmC). Here, we identified TET2 as a crucial regulator of mast cell differentiation and proliferation. In the absence of TET2, mast cells showed disrupted gene expression and altered genome-wide 5hmC deposition, especially at enhancers and in proximity of down-regulated genes. Impaired differentiation of Tet2-ablated cells could be compensated or further exacerbated by modulating the activity of other TET family members, and mechanistically it could be linked to the dysregulated expression of C/EBP family transcription factors. Conversely, the marked increase in proliferation induced by the loss of TET2 could be rescued exclusively by re-expression of wild-type or catalytically inactive TET2. Our data indicate that in the absence of TET2, mast cell differentiation is under the control of compensatory mechanisms mediated by other TET family members, while proliferation is strictly dependent on TET2 expression.

overall_design: Chromatin-IP was performed as described (Ostuni et al. PMID: 23332752).  5–15 × 10^6 fixed primary bone marrow-derived mast cells were lysed with RIPA buffer and, after chromatin shearing by sonication, incubated overnight at 4°C with protein G Dynabeads coupled with 3–10 μg of anti-H3K4me1 antibody.

first: S

last: Montagner

first: C

last: Leoni

last: Emming

first: G

middle: D

last: Chiara

last: Balestrieri

first: I

last: Barozzi

first: V

last: Togher

first: M

last: Ko

first: A

last: Rao

last: Natoli

last: Monticelli


title: Next Generation Sequencing to determine MneHV7 Transcriptome in pigtailed macaque salivary gland tissues

city: seattle

first: Serge

last: Barcy

state: WA

address: 1900 Ninth Avenue

department: SCRI

institute: Seattle Children's

summary:
Purpose: The goal of this study is to determine the expression profile of a new macaque HHV-7 homolog in salivary gland tissues from naturally infected pigtailed macaques
Methods: Viral mRNA profiles of 9 different pigtailed macaques naturally infected with the HHV7 homolog provisionnally named MneHV7 were generated by deep sequencing using Illumina Highseq. The sequence reads that passed quality filters were analyzed at the transcript isoform level with two methods: Burrows–Wheeler Aligner (BWA) followed by ANOVA (ANOVA) and TopHat followed by Cufflinks. qRT–PCR validation was performed using TaqMan and SYBR Green assays
Results: . Hierarchical clustering of differentially expressed genes uncovered several as yet uncharacterized genes that may contribute to retinal function. Data analysis with BWA and TopHat workflows revealed a significant overlap yet provided complementary insights in transcriptome profiling.
Conclusions: Our study represents the first detailed analysis of viral transcriptomes associated with the macaque HHV-7 homolog generated by RNA-seq technology. Our results show that NGS offers a comprehensive and more accurate quantitative and qualitative evaluation of mRNA content within a cell or tissue.

overall_design: Examine MneHV7 viral transcriptome in macaque salivary gland with different viral loads determined by qPCR

first: Jeannette

last: Staheli

first: Michael

middle: R

last: Dyen

first: Gail

middle: H

last: Deutsch

first: Ryan

last: Basom

first: Matthew

last: Fitzgibbon

first: Patrick

last: Lewis


title: Prolyl hydroxylation regulates protein degradation, synthesis, and splicing in human induced pluripotent stem cell-derived cardiomyocytes

city: Bethesda

first: Yanqin

last: Yang

state: Maryland

address: 10 Center Dr.

institute: NIH

summary: In this study we report the gene expression profile and MISO analysis for alternative splicing events such as exon skipping in iPSC-derived cardiomyocytes which were treated with a drug  inhibiting α-ketoglutarate-dependent hydroxylases (dimethyloxalylglycine) and compared to vehicle control. α-ketoglutarate-dependent hydroxylase inhibition plays a central role in cardiac hypoxia and the goal of this study was to identify new pathways in hypoxia beyond HIF-1α.

overall_design: Biological replicates of RNA-seq data from iPSC-derived cardiomyocytes treated with dimethyloxalylglycine or vehicle control

first: Andrea

last: Stoehr

first: Elizabeth

last: Murphy


first: Viviana

last: Piccolo

summary: Dioxygenases of the TET family impact genome functions by converting 5-methylcytosine in DNA to 5-hydroxymethylcytosine (5hmC), but their contribution to differentiation and function of immune cells is still incompletely understood. Here, we identified TET2 as a crucial regulator of mast cell differentiation and proliferation. Specifically, mast cell differentiation was impaired in absence of TET2, a defect that could be however compensated or further exacerbated modulating the activity of other TET family members. Mechanistically, delayed mast cell differentiation was related to the dysregulated expression of C/EBP transcription factors. Vice versa, the marked increase in proliferation induced by the absence of TET2 could be rescued exclusively by the specific re-expression of TET2 itself, regardless of its enzymatic activity. Our data indicate that in absence of TET2, mast cell differentiation is largely under the control of compensatory mechanisms mediated by other TET family members, while proliferation is strictly dependent on TET2 expression.

overall_design: Genomic DNA of FACS-sorted Kit+ FcεRIα+ populations of primary bone marrow-derived mast cells (BMMCs) from Tet2-/- and Tet2+/+ animals was extracted. Changes in 5hmC deposition were assessed genome-wide by biotin-based enrichment of 5hmC (Hydroxymethylation Collector-seq Kit, Active Motif) and  multiparallel sequencing.

first: S

last: Montagner

last: Leoni

last: Emming

last: Della Chiara

last: Balestrieri


title: The Common Stress Responsive Transcription Factor ATF3 Binds Active Enhancers and Bookmarks Genomic Sites for Transcriptional Regulation

first: Jindan

last: Yu

state: GA

address: E330, 1760 Haygood Dr NE

department: Urology


title: Disruption of Na+/H+ exchanger regulatory factor 2 scaffold suppresses colon cancer proliferation

first: Viren

last: Patel

address: 101 Woodruff Circle

department: EICC

institute: Emory University School of Medicine

summary: A key function of Na+/H+ exchanger regulatory factor 2 (NHERF2) is spatial organization of signaling proteins to facilitate signal transduction. The role of NHERF2 in cancer progress is not well understood. This study determines how loss of NHERF2 alter colon cancer progress.

overall_design:
We show that loss of NHERF2 decreases colon cancer cell proliferation. To compare the effects of NHERF2 and LPA2 at the molecular level, HCT116 colon cancer xenograft with knockdown of NHERF2 or LPA2 was analyzed by RNAseq.

Please note that standard cufflinks/cuffdiff output files are provided in the compressed tar files as processed data and Cufflinks/Cuffdiff output file content/formats are described at:
https://cole-trapnell-lab.github.io/cufflinks/cuffdiff/#cuffdiff-output-files
https://cole-trapnell-lab.github.io/cufflinks/file_formats/
(also included in the 'readme.txt' file)

first: Chris

last: Yun


title: The Common Stress Responsive Transcription Factor ATF3 Binds Active Enhancers and Bookmarks Genomic Sites for Transcriptional Regulation [ChIP-seq]

summary: Comprehensive analysis of global ATF3-binding profiles in the human genome under quiescent and stressed (DNA damage) conditions. Although expressed at a low level, quiescent ATF3 was found to bind a large number of genomic sites that are often associated with genes required for cellular stress responses. While these ATF3-binding sites often contained motifs for other transcription factors (TFs), we found that ATF3 bound a large portion of active enhancers characterized by p300 binding, and as a consequence, ATF3 often regulates expression of the genes proximal to these enhancers. While DNA damage elicited by camptothecin dramatically altered the ATF3 binding profile, most of the genes regulated by ATF3 upon DNA damage were pre-bound by ATF3 before the stress. Moreover, we demonstrated that ATF3 was co-localized with the major stress responder p53 at genomic sites, thereby contributing to induction of p53 target gene expression upon DNA damage. Therefore, our results suggest that ATF3 bookmarks genomic sites and interacts with other TFs for transcriptional regulation under quiescent and stressed conditions.

overall_design: ATF3-knockout HCT116 cells were generated by AAV-mediated homologous recombination. ATF3-wildtype (ATF3-WT), -knockout (ATF3-KO), p53-knockout (p53-KO) cells were treated with DMSO (Ctrl) or 1.5 uM of camptothecin (CPT) for 4 h, and then crosslinked by 1 mM of di(N-succinimidyl) glutarate (DSG) for 45 min and 1% of formaldehyde for 10 min. Chromatin was immunoprecipitated with 2 ug of ATF3 antibody (Santa Cruz, cat.# sc-188, lot# F2614) or p53 antibody (Santa Cruz, cat.# sc-126, lot# L3013) and subjected to ChIP-seq analyses.

first: Chunhong

last: Yan

first: Jonathan

last: Zhao


title: Transcriptome profiling of Caki2 cells re-expressing Polybromo-1 (PBRM1)

city: West Lafayette

first: Emily

last: Dykhuizen

state: IN

address: 201 S University St

department: Medicinal Chemistry and Molecular Pharmacology

institute: Purdue University

summary: PBRM1 is a component of the PBAF chromatin remodelling complex and has been observed to be deregulated in a significant proportion of patients with clear-cell Renal Cell Carcinoma (ccRCC). This study employs RNA-Seq to identify differentially expressed genes in cellular models of ccRCC by expressing PBRM1 in PBRM1-deficient Caki2 cells.

overall_design: Using lentiviral mediated mechanism, stable Caki-2 cell line expressing PBRM1 was developed (Caki2-PBRM1). Empty vector inserted in Caki2 cells served as control (Caki2-Ø)

first: Basudev

last: Chowdhury


summary: Dioxygenases of the TET family impact genome functions by converting 5-methylcytosine in DNA to 5-hydroxymethylcytosine, but the individual contribution of the three family members to differentiation and function of myeloid cells is still incompletely understood. Using cells with a deletion in the Tet2 gene, we show that TET2 contributes to the regulation of mast cell differentiation, proliferation and effector functions. The differentiation defect observed in absence of TET2 could be however completely rescued or further exacerbated by modulating TET3 activity, and it was primarily linked to dysregulated expression of the C/EBP family of transcription factors. In contrast, hyper-proliferation induced by the lack of TET2 could not be modified by TET3. Together, our data indicate the existence of both overlapping and unique roles of individual TET proteins in regulating myeloid cell gene expression, proliferation and function.

overall_design: Total mRNA of FACS-sorted Kit+ FcεRIα+ populations of primary bone marrow-derived mast cells (BMMCs) from Tet2-/- and Tet2+/+ animals was extracted and subjected to multiparallel sequencing.

last: Monticelli

first: G

last: Natoli

first: V

first: I

last: Barozzi

first: Chiara

last: Della


last: Piccolo

overall_design: Refer to individual Series


title: Comparison of gene expression profiles between FRA1 knockdown and vector control Bel-7402 cells by RNA-sequencing

city: Hong Kong

first: Chi Ho

last: Lin

address: 6th Floor, The Hong Kong Jockey Club Building for Interdisciplinary Research, 5 Sassoon Road, Pokfulam

department: Centre for PanorOmic Sciences

country: Hong Kong

institute: The University of Hong Kong

summary: The purpose of this experiment is to search for the transcriptional target of FRA1.

overall_design: We subjected FRA1 knockdown and vector control Bel-7402 cells to RNA-sequencing, and analyzed the genes that were shown to be differentially expressed between the two groups.

first: Yuen

middle: T

last: Lau

first: Kin

middle: W

last: Lee


title: Comparative proteomics and transcriptomics study of signaling network proteins across multiple normal and cancer cell lines

status: Public on May 03 2016

first: Stephen

last: Lindemann

address: 745 Agriculture Mall Drive

department: Food Sciences

summary: It is not known whether cancer cells generally show quantitative differences in the expression of signaling pathway proteins that could dysregulate signal transduction. To explore this issue, we first defined the primary components of the EGFR-MAPK pathway in normal human mammary epithelial cells, identifying 16 core proteins and 10 feedback regulators. We then quantified their absolute abundance at both the protein and mRNA level across a panel of normal and breast cancer cell lines. We found that core pathway proteins were present at very similar levels across all cell types. In contrast, the EGFR and transcriptionally controlled feedback regulators were present at highly variable levels. The absolute abundance of most core proteins was between 50,000-70,000 copies per cell, but the adaptors SOS1, SOS2, and GAB1 were found at far lower levels (2,000-5,000 per cell). MAPK signaling showed saturation in all cells between 3,000-10,000 occupied EGFR, consistent with the idea that adaptors limit signaling. Our results suggest that the relative stoichiometry of core MAPK pathway proteins is very similar across different cell types, with cell-specific differences mostly restricted to variable levels of feedback regulators. The low abundance of adaptors relative to the EGFR could be responsible for previous observation of saturable signaling, endocytosis, and high affinity EGFR.

overall_design: Cells grown to ~80% confluence were serum starved and processed for both RNA-Seq and proteomics anaysis, using parallel samples. A total of seven samples. RNA was extracted from duplicate 15cm plates and poled before library preparation.

first: Tujin

last: Shi

first: Mario

last: Niepel

first: William

middle: B

last: Chrisler

first: Lye

last: Markillie

first: Wei-Jun

last: Qian

first: Ronald

last: Taylor

first: H

last: Wiley


title: Functional organization of the human 4D Nucleome

city: Ann Arbor

first: Indika

last: Rajapakse

state: MI

address: 1600 Huron Parkway, NCRC Building 520, Rm. 3392

department: Department of Computational Medicine & Bioinformatics

institute: University of Michigan

summary: The 4D organization of the interphase nucleus, or the 4D Nucleome (4DN), reflects a dynamical interaction between 3D genome structure and function and its relationship to phenotype. We present initial analyses of the human 4DN, capturing genome-wide structure using chromosome conformation capture and 3D imaging, and function using RNA-sequencing. We introduce a quantitative index that measures underlying topological stability of a genomic region. Our results show that structural features of genomic regions correlate with function with surprising persistence over time. Furthermore, constructing genome-wide gene-level contact maps aided in identifying gene pairs with high potential for coregulation and colocalization in a manner consistent with expression via transcription factories. We additionally use 2D phase planes to visualize patterns in 4DN data. Finally, we evaluated gene pairs within a circadian gene module using 3D imaging, and found periodicity in the movement of clock circadian regulator and period circadian clock 2 relative to each other that followed a circadian rhythm and entrained with their expression.

overall_design: Genome wide chromosome conformation capture (Hi-C) data from human fibroblasts, every 8 hours for 56 hours.

first: Laura

last: Seaman

first: Haiming

last: Chen


title: Differential expression of RNAs in E9 tuft rostrums with NTDs

city: Honolulu

first: Keith

last: Fong

state: HI

address: 651 Ilalo Street; BSB 110

department: Anatomy, Biochemistry and Physiology

institute: University of Hawai'i John A. Burns School of Medicine

summary:
Purpose: Identify genes and pathways affected in tuft embryos with NTDs
Results: Expression of genes associated with neural tube closure and components of non-canonical WNT signaling/PCP pathways were affected
Conclusions: TET1 regulates genes associated with neural tube closure

overall_design: RNA pooled from the rostrums of E9 (18-22 somites) tuft/tuft embryos with NTD compared with respective wildtype background strain

first: Vedbar

last: Khadka


title: Shh and ZRS enhancer co-localization is specific to the zone of polarizing activity

city: Edinburgh

first: Iain

last: Williamson

address: Crewe Road

department: Chromosomes and Gene Expression

country: United Kingdom

institute: IGMM MRC Human Genetics Unit

summary: Limb-specific expression of Shh is regulated by the long-range (~one megabasepair distant) ZRS enhancer. In the mouse, murine limb bud restricted spatiotemporal expression of Shh occurs from ~E10 until E11.5 at the distal posterior margin is essential for the correct formation of the autopod. Here, we have analyzed the higher-order chromatin conformation of Shh in expressing and non-expressing tissues, both by fluorescence in situ hybridization (FISH) and by chromosome conformation capture (5C). Conventional and super-resolution light microscopy identified significantly elevated frequences of Shh/ZRS co-localization only in the Shh expressing regions of the limb bud consistent with the formation of an enhancer-promoter loop. However, Shh-ZRS spatial distances were consistently shorter than intervening distances to a neural enhancer in all tissues and developmental stages analyzed – regardless of Shh expression. 5C also identified a topologically associating domain (TAD) over the Shh-ZRS genomic region and enriched interactions between Shh and ZRS, but in the head, body and limb buds of E11.5 embryos, so also not linked to Shh expression. We show that gene-enhancer (Shh/ZRS) co-localization correlates with the spatiotemporal domain of limb bud-specific Shh expression, but that close Shh/ZRS proximity in the nucleus occurs regardless of whether the gene or enhancer is active. We suggest that this constrained chromatin configuration optimises the opportunity for the active enhancer to locate and instigate Shh expression.

overall_design:
Here, we have analyzed the higher-order chromatin conformation of Shh in expressing and non-expressing tissues by chromosome conformation capture (5C).
5C oligonucleotides were designed around HindIII restriction sites following an alternative scheme

last: Lettice

first: Robert

middle: E

last: Hill

first: Wendy

last: Bickmore


title: Identification of in vivo binding sites for YY2.

city: Zaragoza

first: Jon

last: Schoorlemmer

address: c/Juan Bosco 13, planta A

department: Retroviral elements in development and disease

country: Spain

institute: ARAID

summary: We report the application of next generation sequencing for high-throughput profiling of Yin Yang 2 (YY2) binding sites in mammalian Trophectoderm Stem cells. Sequencing of chromatin obtained in chromatin-immunoprecipitation (ChIP) assays carried out with αYY2 serum yielded over ten million sequence reads. Peak finding allowed us allowed us to identify a limited number of chromatin targets for YY2. Putative binding sites were validated in regular ChIP assays and gene expression of a few genes nearby was altered in the absence of Yy2. This study provides a framework to understand the role of YY2 in diverse mammalian cell populations.

overall_design: Examination of YY2 binding sites in TSB7 cells.

first: Pedro

last: Muniesa


title: Arabidopsis SAURs Are Critical for Differential Light Regulation of the Development of Various Organs

city: Beijing

first: Xu

middle: Zhao

last: Gao

state: Beijing

address: The Summer Palace Road No. 5

department: College of Biological Sciences

country: China

institute: Peking University

summary: During de-etiolation of Arabidopsis seedlings, light promotes the expansion of cotyledons but inhibits the elongation of hypocotyls. The mechanism of this differential regulation of cell enlargement is unclear. Our organ-specific transcriptomic analysis identified 32 Small Auxin Up RNA (SAUR) genes whose transcripts were light-induced in cotyledons and/or repressed in hypocotyls. We therefore named these SAURs as lirSAURs. Both overexpression and mutation analyses demonstrated that lirSAURs could promote cotyledon expansion and opening and enhance hypocotyl elongation, possibly by inhibiting phosphatase activity of PP2C-Ds. Light reduced auxin levels to down-regulate the expression of lirSAURs in hypocotyls. Further, phytochrome-interacting factors (PIFs) were shown

overall_design: Totally eighteen wild type samples, three treatments and two organs, and each has three replicates. Totally six pifq samples, two organs, and each has three replicates.

first: Ning

last: Sun

first: Jiajun

last: Wang

first: Zhaoxu

first: Jie

last: Dong

first: Hang

last: He

last: Terzaghi

last: Wei

first: Xing

last: Deng

first: Haodong


title: Mitochondrial stress induces chromatin reorganization to promote longevity and UPRmt

city: Berkeley

first: Andrew

last: Dillin

state: CA

address: 188 Li Ka Shing

institute: University of California, Berkeley

summary:
Organisms respond to mitochondrial stress through
the upregulation of an array of protective genes,
often perpetuating an early response to metabolic
dysfunction across a lifetime. We find that mitochondrial
stress causes widespread changes in chromatin
structure through histone H3K9 di-methylation
marks traditionally associated with gene silencing.
Mitochondrial stress response activation requires
the di-methylation of histone H3K9 through the activity
of the histone methyltransferase met-2 and the
nuclear co-factor lin-65. While globally the chromatin
becomes silenced by these marks, remaining portions
of the chromatin open up, at which point the
binding of canonical stress responsive factors such
as DVE-1 occurs. Thus, a metabolic stress response
is established and propagated into adulthood of
animals through specific epigenetic modifications
that allow for selective gene expression and lifespan
extension.

overall_design: comparison of gene expression changes in response to cco-1 RNAi treatment in N2, lin-65(n3441) and met-2(ok2307) populations of C. elegans L4 animals

first: Kristan

last: Steffen

first: Ye

last: Tian


title: Two Conserved Histone Demethylases Regulate Mitochondrial Stress-Induced Longevity

summary: Across eukaryotic species, mild mitochondrial stress can have beneficial effects on the lifespan of organisms. Mitochondrial dysfunction activates an unfolded protein response (UPRmt), a stress signaling mechanism designed to ensure mitochondrial homeostasis. Perturbation of mitochondria during larval development in C. elegans not only delays aging but also maintains UPRmt signaling, suggesting an epigenetic mechanism that modulates both longevity and mitochondrial proteostasis throughout life. Here we identify the conserved histone lysine demethylases jmjd-1.2/PHF8 and jmjd-3.1/JMJD3 as positive regulators of lifespan in response to mitochondrial dysfunction across species. Reduction-of-function of the demethylases potently suppresses longevity and UPRmt induction while gain-of-function is sufficient to extend lifespan in an UPRmt-dependent manner. A systems genetics approach in the BXD mouse reference population further indicated conserved roles of the mammalian orthologs in longevity and UPRmt signaling. These findings illustrate an evolutionary conserved epigenetic mechanism that determines the rate of aging downstream of mitochondrial perturbations.

overall_design: Comparison of gene expression changes in response to cco-1 RNAi treatment, overexpression of jmjd-1.2 and overexpression of jmjd-3.1 in populations of C. elegans L4 animals

first: Carsten

last: Merkwirth


title: RNA-seq of 50 paired hepatocellular carcinoma

city: Shanghai

first: Gang

last: Liu

address: 320 Yueyang Rd, Shanghai

institute: Shanghai Institutes for Biological Sciences

summary: RNA-seq of Illumina sequencing of 50 paired normal and tumor samples

first: Guojun

last: Hou

first: Lei

first: Weiping

last: Zhou


title: Chronic Activation of γ2 AMPK Induces Obesity and Reduces Beta Cell Function

city: Cambridge

first: Brian

middle: Yee Hong

last: Lam

address: Box 289 Addenbookes Hospital, Hills Road

department: Metabolic Research Laboratories

institute: University of Cambridge

summary: Despite significant advances in our understanding of the biology determining systemic energy homeostasis, the treatment of obesity remains a medical challenge. Activation of AMP-activated protein kinase (AMPK) has been proposed as an attractive strategy for the treatment of obesity and its complications. AMPK is a conserved, ubiquitously expressed, heterotrimeric serine/threonine kinase whose short-term activation has multiple beneficial metabolic effects. Whether these translate into long-term benefits for obesity and its complications is unknown. Here, we observe that mice with chronic AMPK activation, resulting from mutation of the AMPK γ2 subunit, exhibit ghrelin signalling-dependent hyperphagia, obesity and impaired pancreatic islet insulin secretion. Humans bearing the homologous mutation manifest a congruent phenotype. Our studies highlight that long-term AMPK activation can have adverse metabolic consequences with implications for pharmacological strategies seeking to chronically activate AMPK systemically to treat metabolic disease.

overall_design: Transcriptomic profiling of the hypothalamic arcuate nucleus from AMPK γ2 R299Q knock-in mice

middle: Y

first: Arash

last: Yavari

first: Kasparas

last: Petkevicius

first: Marcella

last: Ma

first: Giles

last: Yeo

first: Houman

last: Ashrafian


title: CHIP-seq KAT2B in mouse isolated islet

city: Lille

first: Jean Sébastien

last: Annicotte

address: Faculté de Médecine - Pôle recherche 2ème étage 1 place de Verdun

country: France

institute: CNRS UMR8199

summary: The Endoplasmic Reticulum (ER) unfolded protein response (UPRer) pathway plays an important role for pancreatic β cells to adapt their cellular responses to environmental cues and metabolic stress. Although altered UPRer gene expression appears in rodent and human type 2 diabetic (T2D) islets, the underlying molecular mechanisms remain unknown. We show here that germ-line and β-cell specific disruption of the lysine acetyltransferase 2B (Kat2b) gene in mice leads to impaired insulin secretion and glucose intolerance. Genome wide analysis of Kat2b-regulated genes and functional assays revealed a critical role for KAT2B in maintaining UPRer gene expression and subsequent β-cell function. Importantly, Kat2b expression was decreased in db/db and in human T2D islets and correlated with UPRer genes in normal human islets. In conclusion, KAT2B is a crucial transcriptional regulator for adaptive β-cell function during metabolic stress by controlling UPRer and represents a promising target for T2D prevention and treatment

overall_design: CHIP-seq KAT2B in mouse isolated islet

first: Nabil

last: Rabhi

first: Pierre-Damien

last: Denechaud

first: Xavier

last: Gromada

first: Sarah

middle: A

last: Hannou

first: Hongbo

last: Zhang

first: Talha

last: Rashid

first: Elisabet

last: Salas

first: Emmanuelle

last: Durand

first: Olivier

last: Sand

first: Amélie

last: Bonnefond

first: Loic

last: Yengo

first: Carine

last: Chavey

first: Caroline

last: Bonner

first: Julie

last: Kerr-Conte

first: Amar

last: Abderrahmani

first: Johan

last: Auwerx

first: Lluis

last: Fajas

first: Philippe

last: Froguel

first: Jean-Sébastien


title: RNA-Seq Quantification of Xenobiotic Processing Genes in Various Sections of the Intestine in Comparison to the Liver of Male Mice

city: Seattle

first: Julia

last: Cui

state: Washington

address: 4225 Roosevelt Way NE Suite # 100

institute: University of Washington

summary: This study aimed to quantify and compare the mRNA abundance of all major XPGs in liver and intestine using RNA-Seq.  The mRNA profiles of 304 XPGs, including phase-I, phase-II enzymes, phase-II cosubstrate synthetic enzymes, xenobiotic transporters, as well as xenobiotic-related transcription factors, were systematically examined in liver and various sections of the intestine in adult male C57BL/6J mice.  By two-way hierarchical clustering, over 80% of the XPGs had tissue-divergent expression, which partitioned into liver-, small intestine-, and large intestine-predominant patterns.  Among the genes, 54% were highest expressed in liver, 21% in duodenum, 4% in jejunum, 6% in ileum, and 15% in large intestine.  The highest expressed XPG in liver was Mgst1, in duodenum Cyp3a11, in jejunum and ileum Ces2e, and in large intestine Cyp2c55.  Interestingly, XPGs in the same family usually exhibited highly different tissue distribution patterns, and many XPGs were almost exclusively expressed in one tissue and minimally expressed in others.  In conclusion, the present study is among the first and the most comprehensive investigation of the real mRNA abundance and tissue-divergent expression of all major XPGs in mouse liver and intestine, which aids in understanding the tissue-specific biotransformation and toxicity of drugs and other xenobiotics.

overall_design: mRNA profiles of liver and four sections of intestine of wild-type (WT) mice were generated by RNA-Seq, in triplicate, by illumina HiSeq 2000.


title: Comparative Systems Pharmacology of HIF Stabilization in the Prevention of Retinopathy of Prematurity

city: Cleveland

first: Ernest

last: Chan

state: OH

address: 10900 Euclid Ave, Robbins E211

department: PQHS

institute: CWRU

summary: Retinopathy of prematurity (ROP) is the most common cause of childhood blindness worldwide and is caused by oxygen therapy necessary to prevent mortality after premature birth. We have previously demonstrated the efficacy of systemic hypoxia inducible factor (HIF) stabilization through HIF prolyl hydroxylase inhibition (HIF PHi) in protecting retinal vasculature from oxygen toxicity in a mouse model of ROP or oxygen induced retinopathy (OIR). We definitively demonstrated that hepatic HIF-1 can be activated to confer this protection using systemic dimethyloxalylglycine (DMOG) to prevent HIF-1α degradation. In this study we compare Roxadustat, a small molecule stabilizer of HIF-1 currently in phase 3 clinical trials for increasing erythropoiesis in adult patients with chronic kidney disease, to DMOG. We demonstrate that Roxadustat induces vascular protection during hyperoxia to induce the coordinated sequential growth of retinal vasculature with a 3-fold reduction in oxygen induced capillary loss (p-=0.001). In order to define the molecular mechanism of protection, we further compared the transcriptome of both liver and retina after systemic treatment with Roxadustat or DMOG. Similar gene expression profiles were identified in liver but very different effects on transcription were found in retinal tissues because Roxadustat, in contrast to DMOG, directly targets retina, confirmed by western blot and by rescue of the hepatic HIF-1 KO, two criteria that DMOG treatment is unable to fulfill. Systems pharmacologic analysis demonstrates that Roxadustat induces typical HIF regulated genes critical to aerobic glycolysis in liver and retinal tissues whereas DMOG, acting through either secreted hepatokines or by influence of systemic DMOG, downregulates cell adhesion/extracellular matrix interaction pathways while increasing expression of histone cluster genes. Stratification of liver transcriptomes to secreted gene products again shows close consensus of hepatic genes induced by both small molecules, and includes upregulation of a plethora of angiogenic proteins such as plasminogen activator inhibitor (PAI-1), erythropoietin (EPO), and orosomucosoid 2 (ORM2). Secondary validation of these transcripts by serum ELISA confirms secretion of EPO and PAI-1 into blood from liver. These findings definitively demonstrate that HIF stabilization can prevent OIR by two pathways: direct retinal HIF stabilization and induction of aerobic glycolysis or indirect, hepatic HIF-1 stabilization and increased serum angiokines. Systems pharmacology analysis therefore explains why intermittent, low dosage of small molecule HIF stabilizers creates a profound protective phenotype, because both pathways can take advantage of cytoprotection induced by the liver and by retina synergistically. These data provide a rationale for considering low dose, intermittent systemic administration of Roxadustat, currently in phase 3 trials in adults with chronic kidney disease, to eradicate ROP in children.

overall_design: RNA-Seq of mice treated with PBS (control), DMOG, or Roxadustat from liver or retina.

first: Jonathan

last: Sears

first: George

last: Hoppe


title: Nuclear RNA in the nervous system

city: St Louis

first: Joseph

last: Dougherty

state: Missouri

address: 4566 Scott Ave, box 8232

department: Genetics and Psychiatry

institute: Washington University

summary:
Background: Recent characterization of the transcriptional landscape of cell lines and whole tissues has suggested widespread transcription of the genome, including loci that produce regulatory non-coding RNAs that function within the nucleus.

Methods: Here, we have defined the nuclear transcriptional landscape of the three major cellular divisions of the nervous system using flow sorting of genetically labeled nuclei from bacTRAP mouse lines followed by characterization the unique expression of coding, non-coding and intergenic RNAs in the mature mouse brain with RNAseq, and validation with independent methods.

Results: Our findings reveal diverse expression across the cell-types of all classes of RNAs, including long non-coding RNAs - several of which were confirmed as highly enriched in the nuclei of specific cell-types using anatomical methods. Finally, we also discovered several examples of cell-type specific expression of tandem gene fusions, and report the first cell-type specific expression of circular RNAs, notably a neuron specific and nuclear enriched RNA arising from the gene Hnrnpu.

Conclusion: These non-coding RNA expression data should provide an important context for studies evaluating the function of a variety of ncRNA in the nervous system.

overall_design: Three to four independent replicate samples (each from one mouse) were collected for each of three sample types: Neuronal nuclear RNA, Astrocyte nuclear RNA, Oligodendrocyte nuclear RNA.  Controls include low-coverage presorted nuclear RNA from each mouse.

first: A

last: Reddy

first: D

last: O'Brien

first: N

last: Pisat

first: C

last: Weichselbaum

first: M

last: Lisci

first: K

last: Sakers

first: J

last: Dalal


title: Effect of Cited2 knockdown on global transcript expression in Rcho-1 cell differentiation

city: Kansas City

first: Pramod

last: Dhakal

state: Kansas

address: 3901 Rainbow Boulevard

department: Pathology and Laboratory Medicine

institute: University of Kansas Medical Center

summary: We had previously discovered that the transcription factor Cited2 was highly induced during trophoblast differentiation. In this study, we used an lentiviral shRNA strategy to decrease Cited2 expression in Rcho-1 trophoblast cells. A RNA-seq approach was used to determine global transcript differences inRcho-1 knockdown cells compared to control cells.

overall_design: Rcho-1 cells transduced with control shRNAs were used as controls. Cells transduced with shRNAs targetingCited2 were used as treatment.Cells were differentiated for 8 days and the analyses were done.

first: Kazuhiko

last: Imakawa

first: Michael

last: Soares


title: Cooperation of Nutlin-3a and a Wip1 inhibitor to induce p53 activity

status: Public on May 02 2016

city: Göttingen

first: Anusha

last: Sriraman

address: Justus-von-liebig Weg 11

department: Department of Molecular Oncology

institute: Georg-August University of Göttingen

summary: Targeting the Mdm2 oncoprotein by drugs has the potential of re-establishing p53 function and tumor suppression. However, Mdm2-antagonizing drug candidates, e. g. Nutlin-3a, often fail to abolish cancer cell growth sustainably. To overcome these limitations, we inhibited Mdm2 and simultaneously a second negative regulator of p53, the phosphatase Wip1/PPM1D. When combining Nutlin-3a with the Wip1 inhibitor GSK2830371 in the treatment of p53-proficient but not p53-deficient cells, we observed enhanced phosphorylation (Ser 15) and acetylation (Lys 382) of p53, increased expression of p53 target gene products, and synergistic inhibition of cell proliferation. Surprisingly, when testing the two compounds individually, largely distinct sets of genes were induced, as revealed by deep sequencing analysis of RNA. In contrast, the combination of both drugs led to an expression signature that largely comprised that of Nutlin-3a alone. Moreover, the combination of drugs, or the combination of Nutlin-3a with Wip1-depletion by siRNA, activated p53-responsive genes to a greater extent than either of the compounds alone. Simultaneous inhibition of Mdm2 and Wip1 enhanced cell senescence and G2/M accumulation. Taken together, the inhibition of Wip1 might fortify p53-mediated tumor suppression by Mdm2 antagonists.

overall_design: Expression profiling by high throughput sequencing

first: Sriraman

last: Anusha

first: Radovanovic

last: Marija

first: Matthias

last: Dobbelstein


title: Determination of Transcriptional start sites in S. aureus USA300 (JE2) during exponential phase

city: Cologne

first: Konrad

middle: U.

last: Förstner

state: North Rhine-Westphalia

address: Gleueler Str. 60

department: Information Services

institute: ZB MED - Information Centre for Life Sciences

summary: Transcriptional start sites determination of USA300 (JE2)  by using dRNA-Seq

overall_design: In order to globally identify transcriptional start sites in Staphylococcus aureus we depleted processed transcripts from RNA extractions with Terminator 5'-phosphate-dependent exonuclease.

first: Sudip

last: Das

middle: U

first: Martin

last: Fraunholz


title: Transcriptionally inactive ATF2 variant drives melanomagenesis

city: San Diego

first: brian

last: james

address: 10901 North Torrey Pines Road

institute: Sanford Burnham

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

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Refer to individual Series


title: Next Generation Sequencing Facilitates Quantitative Analysis of Wild Type and dek10 mutant Transcriptomes

status: Public on May 01 2016

first: Weiwei

last: Qi

address: 333 Nanchen Road

institute: Shanghai University

summary: Analysis of  gene expression level. The hypothesis tested in the present study was that dek10 mutant influences the expression of mitochondrial ETC associated genes. Results provide important information of the gene expression level regulation of mitochondrial envelop, proton transport,translation and genes associated with other biological processes.

overall_design: Endosperm mRNA profiles of 15DAP wild type (WT) and dek10 mutant were generated by deep sequencing, in triplicate, using Illumina Hiseq 2500 (Zea mays).

first: Zhongrui

last: Tian


title: LHP1, CLF binding sites and their role in H3K27me3 spreading in Arabidopsis thaliana

city: Thuwal

first: Alaguraj

last: Veluchamy

address: Thuwal

department: BESE

country: Saudi Arabia

institute: King Abdullah University of Science and Technology

summary: LHP1 regulates H3K27me3 spreading

overall_design: Examination of LHP1 binding sites by ChIP-seq

first: Teddy

last: Jégu

first: Federico

last: Ariel

first: David

last: Latrasse

first: Kiruthiga

last: Mariappan

first: Soonkap

last: Kim

first: Abdelhafid

last: Bendahmane

last: Crespi

first: Heribert

last: Hirt

first: Catherine

last: Bergounioux

first: Cécile

last: Raynaud

first: Moussa

last: Benhamed


title: mRNA expression of breast cancer cell lines across different densities [SCRB-Seq]

city: Boston

first: Marc

last: Hafner

state: MASSACHUSETTS

address: WAB 438, 200 Longwood Av.

department: Systems Biology

institute: Harvard Medical School

summary: mRNA expression profiles for 3 breast cancer cell lines seeded at different density and grown for different duration

overall_design: This experiment is part of a study fo the effect of cell density on drug sensitivity [1]. Cells plated at different densities in 384-well plates were harvested at the indicated times and RNA was extracted using the RNeasy mini kit (Qiagen). To ensure sufficient RNA amounts wells with low cell numbers were pooled. Some conditions have been tested in biollogical replicates grown at the same time. Libraries were prepared by the Broad Technology Labs (BTL) following the protocol for SCRB-Seq described in [2]. Transcripts were quantified by the BTL computational pipeline using Cuffquant version 2.2.1 [3].   [1] Hafner, M., Niepel, M., Chung, M., Sorger, P.K., Growth rate inhibition metrics correct for confounders in measuring sensitivity to cancer drugs. DOI:10.1038/NMETH.3853  [2] Soumillon, M., Cacchiarelli, D., Semrau, S., van Oudenaarden, A. & Mikkelsen, T.S. Characterization of directed differentiation by high-throughput single-cell RNA-Seq http://biorxiv.org/content/early/2014/03/05/003236   [3] Trapnell, C., Roberts, A., Goff, L., Pertea, G., Kim, D., Kelley, D.R., Pimentel, H., Salzberg, S.L., Rinn, J.L. & Pachter, L. Differential gene and transcript expression analysis of RNA-seq experiments with TopHat and Cufflinks, Nat. Protoc. 7, 562-578 (2012).

first: Mario

last: Niepel

first: Mirra

last: Chung

first: Magali

last: Soumillon

first: Peter

last: Sorger


title: Polycomb Ezh2 controls the fate of GABAergic neurons in the embryonic cerebellum

city: Bethesda

first: Hossein

last: Zare

address: 5o South Dr #1347

institute: NIH

summary: Genetic ablation of Ezh2 in Pax7-Cre driven lineages resulted in developmental defects in E13.5 cerebellum. The transcriptional change and H3K27me3 mark change were investigated by RNA-seq and ChIP-seq on H3K27me3.

overall_design: The Ezh2 gene was genetically ablated by crossing Ezh2 flox allele line and Pax7-Cre line mice, which resulted in Ezh2 gene deletion in embryonic cerebellum. To aid the precise dissection of embryonic cerebellum. A Rosa-YFP allele was also breeded along with the Ezh2 flox allele. E13.5 cerebella were dissected, then immediatedly dropped into RNAlater for mRNA abstraction and RNA-seq, or fixed with 1%PFA for H3K27me3 ChIP-seq assay.

first: Xuesong

last: Feng

first: Aster

last: Juan

first: Hongjun

last: Wang

first: Kyung

last: Ko

first: Gustavo

last: Gutierrez-Cruz

first: Vittorio

last: Sartorelli


title: A master virulence regulator of S. aureus inactivated during carriage in man

city: Oxford

first: Julius

last: Muller

state: Oxfordshire

address: ORCRB, Roosevelt Drive

department: Nuffield Department of Medicine

institute: The Jenner Institute, University of Oxford

summary: It remains unclear how the bacterial populations which colonise many healthy humans occasionally give rise to severe disease. Staphylococcus aureus represents one such population. Here we describe mutations in invasive S. aureus which arose during human colonisation and inactivated rsp, a transcription factor. RNA-Seq was used to generate mRNA profiles of genes controlled by rsp.

overall_design: mRNA samples were sequenced in duplicates from Staphylococcus aureus strains isolated from two patients with concomitant nasal carriage and bloodstream infection as well as from USA300 wild type and USA300 NE1304 NARSA (Network on Antimicrobial Resistance in Staphylococcus aureus) strain.

first: Claudia

last: Lindemann

first: Bernadette

last: Young

first: Daniel

last: Wilson

first: David

last: Wyllie


title: Single-cell 5hmC sequencing reveals chromosome-wide cell-to-cell variability and enables lineage reconstruction

city: Utrecht

first: Siddharth

middle: Subhas

last: Dey

address: Uppsalalaan 8

country: Netherlands

institute: Hubrecht Institute

summary: The epigenetic DNA modification 5-hydroxymethylcytosine (5hmC) has crucial roles in development and gene regulation. Quantifying the abundance of this epigenetic mark at the single-cell level could enable us to understand its roles. We present a single-cell, genome-wide and strand-specific 5hmC sequencing technology based on 5hmC glucosylation and glucosylation-dependent digestion of DNA, that reveals pronounced cell-to-cell variability in the abundance of 5hmC on the two DNA strands of a given chromosome. We develop a mathematical model that reproduces the strand bias and use this model to make two main predictions. First, the variation in strand bias should decrease when 5hmC turnover increases. Second, the strand bias of two sister cells should be strongly anti-correlated. We validate these predictions experimentally, and use our model to reconstruct lineages of 2- and 4-cell mouse embryos, showing that single-cell 5hmC sequencing can be used as a lineage reconstruction tool.

sample_organism:
Mus musculus
synthetic construct

platform_organism:
Mus musculus
synthetic construct

overall_design: In this study, 5-hydroxymethylcytosine (5hmC) was detected on a genome-wide level in single cells from E14tg2a (E14) mouse embryonic stem cells, Vitamin C treated E14 cells, B6/CBA mouse oocytes, 2-cell mouse embryos and 4-cell mouse embryos using a new method called as scAba-seq.

first: Dylan

last: Mooijman

first: Jean-Charles

last: Boisset

first: Nicola

last: Crosetto

first: Alexander

last: van Oudenaarden


title: Vascular smooth muscle cell contractile protein expression is increased through protein kinase G-dependent and –independent pathways by glucose-6-phosphate dehydrogenase inhibition and deficiency

city: Rochester

first: Joseph

last: Miano

state: NY

address: 601 Elmwood Avenue, Box CVRI

department: Aab Cardiovascular Research Institute

institute: University of Rochster

summary:
Purpose: Homeostatic control of vascular smooth muscle cell (VSMC) differentiation is critical for contractile activity and regulation of blood flow. Recently, we reported that pre-contracted blood vessels are relaxed and the phenotype of VSMC is regulated from a synthetic to contractile state by glucose-6-phosphate dehydrogenase (G6PD) inhibition. In the current study, we investigated whether the increase in the expression of VSMC contractile proteins by inhibition and knockdown of G6PD is mediated through a protein kinase G (PKG)-dependent pathway and whether it regulates blood pressure
Methods: Coronary arteries (LAD) isolated from bovine heart mRNA profiles of 12-16 week old wild type (WT) and G6PD-deficient mice were generated by deep sequencing, in triplicate, using Illumina HiSeq 2500. The sequence reads that passed quality filters (Trimmomatic-0.32) were analyzed at the transcript isoform level using STAR_2.4.2a for mapping to reference GRCm38.p4 + Gencode-M6 Annotation and processed with Cufflinks-2.0.2. miR analysis was performed by quantitative RT-PCR (qRT-PCR) for validation using miR-specific TaqMan miR assays (Applied Biosystems, Foster City, CA). Quantitative PCR was performed in triplicate using TaqMan Universal PCR Master mix. Standard curves were made for each miR using synthetic miR oligonucleotides (IDT, Coralville, IA) with the following sequence:  Rno-miR-145: GUCCAGUUUUCCCAGGAAUCCCU,  Rno-miR-1: UGGAAUGUAAAGAAGUGUGUAU,  Rno-miR-143: UGAGAUGAAGCACUGUAGCUC,   Rno-miR-133a: UUUGGUCCCCUUCAACCAGCUG
Results: We found that the expression of VSMC-restricted contractile proteins, myocardin (MYOCD), and miR-1 and miR-143 are increased by G6PD inhibition or knockdown. Importantly, RNA-sequence analysis of aortic tissue from G6PD-deficient mice revealed uniform increases in VSMC-restricted genes, particularly those regulated by the MYOCD-serum response factor (SRF) switch.  Conversely, expression of Krüppel-like factor 4 (KLF4) is decreased by G6PD inhibition. Interestingly, the G6PD inhibition-induced expression of miR-1 and contractile proteins was blocked by Rp-β-phenyl-1,N2-etheno-8-bromo-guanosine-3’,5’-cyclic monophosphorothioate, a PKG inhibitor. On the other hand, MYOCD and miR-143 levels are increased by G6PD inhibition through a PKG-independent manner. Furthermore, blood pressure was lower in the G6PD-deficient as compared to wild-type mice
Conclusions: Therefore, our results suggest that the expression of VSMC contractile proteins induced by G6PD inhibition occurs via PKG1-dependent and –independent pathways

overall_design:
Coronary arteries (LAD) isolated from bovine heart mRNA profiles of 12-16 week old wild type (WT) and G6PD-deficient mice were generated by deep sequencing, in triplicate, using Illumina HiSeq 2500
genotype/variation: CYPKO: Sample 1,Sample 2,Sample 3
genotype/variation: G6PD: Sample 4,Sample 5,Sample 6
biological replicate: Sample 1,Sample 2,Sample 3
biological replicate: Sample 4,Sample 5,Sample 6

first: Rakee

last: Gupte

first: Sukrutha

last: Chettimada

first: Sachindra

last: Joshi

first: Anand

last: Lahkar

first: Vidhi

last: Dhagia

first: Alessandro

last: Aiezza II

first: Thomas

middle: M

last: Lincoln

first: Sachin


title: Epigenetic Profile in RBPJ Maintains Brain Tumor Initiating Cells through CDK9-mediated Transcriptional Elongation

first: Tyler

last: Miller

state: Ohio

department: Stem Cell Biology and Regenerative Medicine

institute: Cleveland Clinic Lerner Research Institute

summary: Glioblastomas display hierarchies with self-renewing cancer stem-like cells (CSCs). RNA sequencing and enhancer mapping revealed regulatory programs unique to CSCs causing upregulation of the iron transporter transferrin, the top differentially expressed gene compared to tissue-specific progenitors. Direct interrogation of iron uptake demonstrated CSCs potently extract iron from the microenvironment more effectively than other tumor cells. Systematic interrogation of iron flux determined that CSCs preferentially require transferrin receptor and ferritin - two core iron regulators - to propagate and form tumors in vivo.  Depleting ferritin disrupted CSC mitotic progression, through the STAT3-FoxM1 regulatory axis, revealing an iron-regulated CSC pathway. Iron is a unique, primordial metal fundamental for earliest life forms, and on which CSCs have an epigenetically programmed, targetable dependence.

overall_design: H3K27Ac histone modifications and RBPJ transcription factor binding were mapped by ChIP-seq in Glioblastoma initiating cells

first: Qi

last: Xie

first: Brian

last: Liau

first: Bradley

last: Bernstein

first: Jeremy

middle: N

last: Rich


title: RNA-seq Profiles in RBPJ Maintains Brain Tumor Initiating Cells through CDK9-mediated Transcriptional Elongation

summary: Glioblastomas coopt stem cell regulatory pathways to maintain brain tumor initiating cells (BTICs), also known as cancer stem cells.  Notch signaling has been a molecular target in BTICs, but Notch antagonists have demonstrated limited efficacy in clinical trials.  RBPJ is considered a central transcriptional mediator of Notch activity. Here, we report that pharmacologic Notch inhibitors were less effective than targeting RBPJ in suppressing tumor growth. While Notch inhibitors decreased canonical Notch gene expression, RBPJ regulated a distinct profile of genes critical to BTIC stemness and cell cycle progression. RBPJ was preferentially expressed by BTICs and required for BTIC self-renewal and tumor growth. MYC, a key BTIC regulator, bound the RBPJ promoter and treatment with a BET family bromodomain inhibitor decreased MYC and RBPJ expression.  Proteomic studies demonstrated that RBPJ binds CDK9, a component of P-TEFb (positive transcription elongation factor), to target gene promoters, enhancing transcriptional elongation.  Collectively, RBPJ links MYC and transcriptional control through CDK9, providing potential nodes of fragility for therapeutic intervention, potentially distinct from Notch.

overall_design: RNA-seq of primary patient-derived GBM brain tumor initiating cells with DAPT treatment or shRBPJ treatment


title: Simultaneous pathway activity inference and global gene expression analysis using RNA-sequencing [myd88]

first: Raivo

last: Kolde

address: 185 Cambridge St.

department: Center for Computational and Integrative Biology

institute: Massachusetts General Hospital

summary: Two 96-well plates per genotype wild type and Myd88 knockout,  4 hour time series in 0.5 hr increments

overall_design: Myd88 BMDM transcriptional profiling to complement TF-seq data

first: Daniel

last: O'Connell


title: RBPJ Maintains Brain Tumor Initiating Cells through CDK9-mediated Transcriptional Elongation


title: Sensitivity and engineered resistance of myeloid leukemia cells to BRD9 inhibition (ChIP-seq)

city: Cold Spring Harbor

first: Anja

middle: Fides

last: Hohmann

address: 1 Bungtown Road

institute: Cold Spring Harbor Laboratory

summary: Our study shows that acute myeloid leukemia (AML) cells require the BRD9 subunit of the SWI/SNF chromatin remodeling complex to sustain MYC transcription, rapid cell proliferation, and a block in differentiation. Based on these observations, we derived small-molecule inhibitors of the BRD9 bromodomain, which selectively suppressed the proliferation of mouse and human AML cell lines. To establish these effects as on-target, we engineered a bromodomain-swap allele of BRD9, which retains functionality despite a radically altered bromodomain pocket. Expression of this allele in AML cells conferred resistance to the anti-proliferative effects of our compound series, thus establishing BRD9 as the relevant cellular target. Furthermore, we used an analogous domain-swap strategy to generate an inhibitor-resistant allele of EZH2. Our study provides the first evidence for a role of BRD9 in cancer and reveals a simple genetic strategy for constructing resistance alleles to demonstrate on-target activity of chemical probes in cells.

overall_design: ChIP-seq of Brd9, Brg1, and FLAG-BRD9 with or without BI-7273 treatment in MLL-AF9 transformed acute myeloid leukemia cells (murine: RN2, human: NOMO-1)

first: A

middle: F

first: C

last: Vakoc


title: Sensitivity and engineered resistance of myeloid leukemia cells to BRD9 inhibition


title: Sensitivity and engineered resistance of myeloid leukemia cells to BRD9 inhibition (RNA-seq)

overall_design: PolyA selected RNA-seq for shRNA-expressing or inhibitor-treated mouse and human leukemia cells and fibroblasts

middle: R


title: m6A and YTHDF1, YTHDF2, YTHDF3 mapping of the HIV-1 RNA pool with PAR-CLIP and pA-m6A-seq

city: Durham

first: Edward

middle: Matthew

last: Kennedy

state: NC

address: Department of Molecular Genetics and Microbiology, Box 3025

department: Molecular Genetics and Microbiology

institute: Duke University

summary: To replicate in the host, HIV-1 must produce RNA and protein, and this depends cellular factors that also control gene expression. Recently, it has been shown that novel components of this machinery post-transcriptionally modify mRNAs with direct consequences for their structure and stability. The most common among these modifications is the addition of a methyl group to the N6 position of adenosine, termed m6A methylation. Here we are interested the interface of HIV-1 with this posttranscriptional mRNA modification pathway. We show that viral transcripts harbor a number of discrete methylation sites and are bound at those sites by these YTHDF proteins. These sites are conserved among distinct HIV-1 isolates and are observed during viral replication in numerous cell types. With reporter assays we also show that these sites stabilize mRNAs when compared to similar sequences mutated to be incapable of methylation. We further show that overexpression of these reader proteins boosts viral protein expression.

overall_design: In this GEO submission we include the PAR-CLIP data sets for numerous HIV-1 isolates in 293T cells, and pA-m6A-seq in CEM-SS cells

first: Bryan

last: Cullen


title: Large-scale rewiring of innate immunity circuitry and microRNA regulation during initial rice blast infection

city: New Haven

first: Haifan

last: Lin

state: Connecticut

address: 10 Amistad St.

department: Cell Biology

institute: Yale Univeristy


title: Large-scale rewiring of innate immunity circuitry and microRNA regulation during initial rice blast infection [smallRNA-Seq]

summary: Rice blast is a recurrent fungal disease, and resistance to fungal infection is a complex trait. Therefore, a comprehensive examination of rice transcriptome and its variation during fungal infection is necessary to understand the complex gene regulatory networks. In this study, adopting Next-Generation Sequencing we profiled the transcriptomes and microRNAomes of rice varieties, one susceptible and the other resistant to M. oryzae, at multiple time points during the fungal infection.

overall_design: In order to investigate variations of rice transcriptome and microRNAome in response to blast infection, we sequenced, profiled, compared and analyzed the gene expressions of two rice cultivars, one susceptible (CO39) and the other resistant (C101LAC) to M. oryzae, at different time points after M. oryzae infection (Fig. 1).

first: Ze-Yuan

last: Li

first: Cong-Ying

last: Wang

first: Weixiong

last: Zhang

first: Yue-Qin

last: Chen


title: Genome-wide identification of snoRNA-guided 2’-O-methylation sites [CLIP-seq]

city: Basel

first: Rafal

last: Gumienny

state: CH

address: Im Ettingerhof 8

department: Biozentrum

country: Switzerland

institute: University of Basel

summary: We combine two experimental high-throughput sequencing methods to identify new 2'-O-methylation sites in human and assign snoRNA guides to sites with previously unknown guides.

overall_design: CLIP-Seq profiling for snoRNP core protein Fibrillarin in HEK293 cells.

first: Dominik

middle: J

last: Jedlinski

first: Mihaela

last: Zavolan


title: Genome-wide identification of snoRNA-guided 2?-O-methylation sites with CLIP and Ribometh-seq


title: Targetting super enhancer associated oncogenes in esophageal squamous cell carcinoma

city: Singapore

first: Phillip

middle: H

last: Koeffler

address: 14 Medical Drive

country: Singapore

institute: Cancer Science Institute


title: Genome-wide identification of snoRNA-guided 2’-O-methylation sites [Ribometh-seq]

overall_design: We applied RiboMeth-seq to map 2’-O-Me sites in rRNA and poly(A)-enriched RNA obtained from either the nucleus or cytoplasmic fraction of HEK293 cells. We carried out the alkaline hydrolysis for different time intervals of 8, 14 or 20 minutes. The sample titles report  protocol, cell type, extraction and digestion time.


title: The transcription factor Zeb2 regulates development of conventional and plasmacytoid DCs by repressing Id2

city: Zwijnaarde

first: Liesbet

last: Martens

address: Technologiepark 927

department: VIB Inflammation Research Center

country: Belgium

institute: VIB-University of Ghent

summary: Plasmacytoid dendritic cells (pDCs) develop from pre-pDCs, while two lineages of conventional DCs (cDC1s and cDC2s) develop from lineage-committed pre-cDCs. A number of transcription factors (TFs) have been implicated in regulating the development of pDCs (E2-2, Id2) and cDC1s (IRF8, Id2 and Batf3) however, those required for the early commitment of pre-cDCs towards the cDC2 lineage are unknown. Here we identified the TF Zinc finger E box binding homeobox 2 (Zeb2), to play a crucial role in regulating DC development. Zeb2 was expressed from the pre-pDC and pre-cDC stage onwards, and highly expressed in mature pDCs and cDC2s.  Mice conditionally lacking Zeb2 in CD11c+ cells had a cell intrinsic reduction in pDCs and cDC2s, coupled with an increase in cDC1s. Conversely, mice in which CD11c+ cells overexpressed Zeb2 displayed a reduction in cDC1s. This was accompanied by altered expression of Id2, which was upregulated in cDC2s and pDCs from conditional knock-out mice. Zeb2 ChIP analysis revealed Id2 to be a direct target of Zeb2. Thus, we conclude that Zeb2 regulates commitment to both the cDC2 and pDC lineages through repression of Id2.

overall_design: cDC1s and cDC2s were isolated and sorted from spleens of CD11cCRExZeb2fl/fl, CD11cCRExR26-Zeb2Tg/Tg or CRE- littermate controls. 24 samples (RNA-Seq) in total. RNA was isolated, amplified by SMARTer amplification, converted to cDNA and then sequenced using Illumina NextSeq.

first: Charlotte

middle: L

last: Scott

first: Bieke

last: Soen

first: Nicolas

last: Skrypek

first: Wouter

last: Saelens

first: Joachim

last: Taminau

first: Gillian

last: Blancke

first: Gert

last: Van Isterdael

first: Danny

last: Huylebroeck

first: Jody

last: Haigh

first: Yvan

last: Saeys

first: Martin

last: Guilliams

first: Bart

last: Lambrecht

first: Geert

last: Berx


title: Targetting super enhancer associated oncogenes in esophageal squamous cell carcinoma [ChIP-seq]

summary: Esophageal squamous cell carcinoma (ESCC) is an aggressive malignancy and the major histological subtype of esophageal cancer. Although recent large-scale genomic analysis has improved the description of the genetic abnormalities of ESCC, few targetable genomic lesions were identified and no molecular-based therapy is available. To identify druggable candidates in this tumor, we performed high-throughput small-molecule inhibitor screening. The unbiased results led us to discover a highly potent anti-ESCC compound, THZ1, a newly developed covalent CDK7 inhibitor. Further in vitro and in vivo experiments showed that THZ1 has powerful anti-neoplastic properties against ESCC cells with minimal toxic effect on healthy tissues. Importantly, whole-transcriptome sequencing (RNA-Seq) revealed that low-dose THZ1 treatment led to selective inhibition of a number of oncogenic transcripts. Notably, further characterization of the genomic features of these THZ1-sensitive transcripts demonstrated that they were frequently associated with Super-Enhancer (SE). Moreover, SE analysis alone uncovered many lineage-specific master regulators in ESCC. Finally, integrative analysis of both THZ1-sensitive and SE-associated transcripts identified a number of novel oncogenes in the context of ESCC, such as PAK4, RUNX1, DNAJB1, SREBF2 and YAP1, with PAK4 being a potential druggable kinase. Together, our integrative molecular approaches identified a catalog of SE-associated lineage-specific master regulators and oncogenic transcripts in ESCC, which will significantly promote the understanding of ESCC biology. The preclinical results of THZ1 may help establish the therapeutic merit of targeting transcriptional regulation program for the treatment of this deadly malignancy.

overall_design: ChIP-seq for H3K27Ac, H3K4Me1 in two esophageal cell lines TE7 and KYSE510.

first: Yan-Yi

last: Jiang

first: De-Chen

last: Lin

first: Anand

last: Mayakonda

first: Masaharu

last: Hazawa

first: Ling-Wen

last: Ding

first: Wen-Wen

last: Chien

first: Xu

last: Liang

first: Chen

last: Ye

first: Jin-Fen

last: Xiao

first: William

last: Senapedis

first: Erkan

last: Baloglu

first: Deepika

last: Kanojia

first: Li

last: Shang

first: Xin

last: Xu

first: Ming-Rong

last: Wang

first: Henry

last: Yang

first: Jeffrey

middle: W

last: Tyner


title: Targetting super enhancer associated oncogenes in esophageal squamous cell carcinoma [RNA-seq]

first: Phillip

last: Koeffler

overall_design: RNA-seq of esophageal cell lines treated with either DMSO or THZ1 at various time points of treatment.

first: Yan-Yi

last: Jiang

last: Mayakoda


title: Transcriptomic and behavioural characterisation of a mouse model of burns pain

city: St Lucia

first: Kathleen

last: Yin

state: Queensland

address: 306 Carmody Rd

department: Institute of Molecular Biosciences

country: Australia

institute: University of Queensland

summary:
Aim: We performed a transcriptomic analysis of affected dorsal root ganglions in mice following a partial thickness locl burn injury on the left hind feet to gain a better understanding of the molecular changes that accompany burns pain
Methods: Male C57BL/6 mice aged 6-8 weeks were sacrificed three days after the burn injury and lumbar dorsal root ganglions L3, L4, and L5 were extracted. Four animals were pooled together to form one biological replicate, and samples were gathered in triplicates. Sequencing was conducted on Illumina NextSeq 500 as 75-nucleotide single-end runs and libraries wre prepared using TruSeq stranded total RNA library preparation. Reads were mapped using STAR aligner and samtools, mapped to mm10 (Ensembl), and count tables generated using HTSeq_count
Results: We discovered 30 genes differentially expressed following the burns injury in dorsal root ganglions, among which are Atf3, Npy, Lipn, and others. We identified Cckbr and Grin2b as potential pharmaceutical targets for pain.
Conclusion: Neurons undergo mainly an inflammatory and regenerative process following a local burn and enriched neuropeptide signalling processes. Cckbr and Grin2b in particular were identified as potential targets in the treatment of burns pain together with other genes identified in our study

overall_design:
Dorsal root ganglion (DRG) neurons RNA were extracted and sequenced in triplicates using Illumina NextSeq 500 as 75-nucleotide single-end runs
Please note that the IP (in the BURN_IP sample title) stands for 'ipsilateral' (not immunoprecipitation).

first: Jennifer

last: Deuis

first: Irina

last: Vetter


title: Neuronal cell lines as model dorsal root ganglion neurons: a transcriptomic comparison

summary:
Aim: Dorsal root ganglion neuron-derived immortal cell lines including ND7/23 and F-11 cells have been used extensively as in vitro model systems of native peripheral sensory neurons. However, while it is clear that some sensory neuron-specific receptors and ion channels are present in these cell lines, a systematic comparison of the molecular targets expressed by these cell lines with intact peripheral neurons is lacking.
Methods: we examined the expression of RNA transcripts in the human neuroblastoma-derived cell line, SH-SY5Y, and two dorsal root ganglion hybridoma cell lines, F-11 and ND7/23, using Illumina next-generation sequencing.
Results: The expression profile of these three cell lines did not resemble any specific dorsal root ganglion neuron subclass. The cell lines lacked many markers for nociceptive sensory neurons, such as the transient receptor potential V1 gene, but expressed markers for both myelinated and unmyelinated neurons.
Conclusion: This paper provides insights into the receptor repertoire expressed in common dorsal root ganglion neuron-derived cell lines, and illustrates the limits and potentials of these cell lines as tools for neuropharmacological exploration.

sample_organism:
Homo sapiens
Mus musculus
Rattus rattus

platform_organism:
Homo sapiens
Mus musculus
Rattus rattus

overall_design: RNA from mouse dorsal root ganglions, SH-SY5Y cells, F-11 cells, and ND7/23 cells were extracted and sequenced using Illumina HiSeq-2000 as 100-nucleotide single-end runs

first: Gregory

last: Baillie


title: Methylation profile generated by reduced representation bisulfite sequencing (RRBS) of H2O2-treated and untreated HCT116 cells

city: Chong Qing

first: Xianfeng

last: Li

address: Da ping

department: Department of Gastroenterology, Daping Hospital

institute: Army Medical University (Third Military Medical University)

summary: DNA methylation is an important regulatory mechanism that contributes to transcriptional repression. To examine whether DNA methylation affects the transcriptional changes of DNA repair genes under H2O2-induced oxidative stress, we performed reduced representation bisulfite sequencing (RRBS) in H2O2-treated and untreated HCT116 cells.

overall_design: Reduced representation bisulfite sequencing (MspI,~40-220bp size fraction) data for HCT116 cell line with or without H2O2 treatement were generated by deep sequencing, using Illumina Illumina HiSeq 2000.

first: Jianing

last: Zhong

first: Zhongsheng

last: Sun


title: RNA-seq Generated Transcriptional Profile of H2O2 Treated HCT116 vs. Untreated HCT116

summary: Oxidative stress, is considered to be a key risk state for a variety of human diseases, which can upregulate transcriptional expression of DNA repair genes. However, it remains elusive whether all or just a portion of DNA repair genes can be upregulated in response to oxidative stress. Therefore, the changes in expression pattern of DNA repair genes and how they are regulated in response to oxidative stress need to be elucidated. In this study, we performed RNA-seq to generate expression profile and identified subsets of induced and repressed DNA repair genes in colorectal cancer cell line exposure to hydrogen peroxide.

overall_design: Total RNAs were extracted from H2O2-treated HCT116 and untreated HCT116 cells with RNeasy Mini Kit (QIAGEN, Germany). Quality control was performed using Bioanalyser 2100 (RNA nano kits, Agilent). mRNA-Seq libraries were generated from total RNA with polyA+ selection of mRNA using the TruSeq RNA Sample Prep Kit v2 (Illumina, San Diego, CA), and then subjected to transcriptome sequencing on the Illumina Hiseq 2000


title: Genes regulated by dimethyl-2-ketoglutarate (DKG) in breast cancer cells

city: Duarte

first: Ching-Ying

last: Kuo

state: California

address: 1500 Duarte Rd.

institute: City of Hope/Beckman Research Institute

summary: We report the gene expression profile of DKG-treated MDA-MB-231 cells.

overall_design: 2 replicates of parental MDA-MB-231 cells and MDA-MB-231 cells treated with DKG (10 mM, 4-days) were subjected to RNA-seq

first: David

middle: K

last: Ann


title: H2O2-treated and untreated HCT116 cells

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

overall_design: Refer to individual Series


title: U1 snRNP regulates alternative polyadenylation in DNA damage response

city: Seoul

first: Jiyeon

last: Park

address: 222, Banpo-daero

department: Catholic Center for Precision Medicine Research

country: South Korea

institute: The Catholic University of Korea

summary: The DNA damage response (DDR) involves coordinated control of gene expression and DNA repair. Using deep sequencing we found widespread changes of  alternative cleavage and polyadenylation (APA) site usage upon UV-treatment in mammalian cells. APA regulation in the 3’ untranslated region (3’UTR) is substantial, leading to both shortening and lengthening of 3’UTRs. Interestingly, a strong activation of intronic APA sites is detected, resulting in widespread expression of truncated transcripts. Intronic APA events are biased to the 5’ end of genes and affect gene groups with important functions in DDR. Moreover, intronic APA site activation during DDR correlates with a decrease in U1 snRNA levels, and this is reversed by U1 snRNA overexpression. Importantly, U1 snRNA overexpression decreases UV-induced apoptosis. Together, these studies describe a significant gene regulatory scheme in DDR where U1 snRNP impacts gene expression via APA.

overall_design: APA profiles were defined after UV-treatment (0h, 0.5h, 2h) in RKO colon cancer cells.

first: Emral

last: Devany

first: Ji-Yeon

first: Michael

last: Murph

first: Jorge

last: Baquero

first: George

last: Zakusilo

first: Xiaokan

last: Zhang

first: Mainul

last: Hoque

first: Bin

last: Tian

first: Frida

middle: E

last: Kleiman


title: A Mutation in LTBP2 Causes Congenital Glaucoma in Domestic Cats (Felis catus)

city: Iowa City

first: S. Scott

last: Whitmore

state: IA

address: 375 Newton Road

department: The University of Iowa Institute for Vision Research

institute: The University of Iowa

summary: The glaucomas are a group of diseases characterized by optic nerve damage that together represent a leading cause of blindness in the human population and in domestic animals. Here we report a mutation in LTBP2 that causes primary congenital glaucoma (PCG) in domestic cats. We identified a spontaneous form of PCG in cats and established a breeding colony segregating for PCG consistent with fully penetrant, autosomal recessive inheritance of the trait. Elevated intraocular pressure, globe enlargement and elongated ciliary processes were consistently observed in all affected cats by 8 weeks of age. Varying degrees of optic nerve damage resulted by 6 months of age. Although subtle lens zonular instability was a common feature in this cohort, pronounced ectopia lentis was identified in less than 10% of cats examined. Thus, glaucoma in this pedigree is attributed to histologically confirmed arrest in the early post-natal development of the aqueous humor outflow pathways in the anterior segment of the eyes of affected animals. Using a candidate gene approach, significant linage was established on cat chromosome B3 (LOD 18.38, q = 0.00) using tightly linked short tandem repeat (STR) loci to the candidate gene, LTBP2. A 4 base-pair insertion was identified in exon 8 of LTBP2 in affected individuals that generates a frame shift that completely alters the downstream open reading frame and eliminates functional domains. Thus, we describe the first spontaneous and highly penetrant non-rodent model of PCG, identifying a valuable animal model for primary glaucoma that closely resembles the human disease providing valuable insights into mechanisms underlying the disease and a valuable animal model for testing therapies.

overall_design: Polyadenylated RNA from the iridiocorneal angles of one PCG-affected cat and one normal cat was sequenced to confirm expression of LTBP2.

first: Markus

last: Kuehn

first: S

middle: S

first: Todd

last: Scheetz

first: Gillian

last: McLellan


title: Transcriptional and proteomic analyses of different herbivore-induced responses in a wild soybean line ED059 and a cultivar Tianlong 2

city: Wuhan

first: xiaoyi

last: wang

address: Xudong 2nd Road,Hubei,P.R.ChinaWuha

institute: Oil crops research institute, Chinese academy of agriculture sciences

summary: Soybean is one of the main sources of oil worldwide, and the cotton bollworm [Helicoverpa armigera (Hübner)] is one of the common herbivores seriously affecting soybean production. After being attacked by herbivores, multiple signaling patheways will be triggered in plants, which induces the production of toxic, repellent, or anti-digestive compounds. Genetic engineering and breeding of resistant plant lines are the most effective and environmentally friendly approaches to control herbivores and improve the performance of plants. This study focuses on founding changes genes and proteins in soybean defense response induced by H. armigera.  Illumina Solexa sequencing platform and iTRAQ were used for the comparative analysis of transcriptome and proteome profiles in the leaves of Tianlong 2 (a cultivated soybean susceptible to herbivores) and ED059 (a wild soybean resistant to herbivores) in response to infestation by H. armigera. Transcriptome data show that 2073 differentially expressed genes (DEGs) were uniquely expressed in ED059 compared with those in Tianlong 2 after 24 h of H. armigera treatment. Based on gene ontology classification of these 2073 DEGs, we identified 233 DEGs corresponded to various signaling pathways, including protein kinases, transcriptional factors, hormone biosynthesis and signal transduction pathways. In addition, a large number of genes related to antinutritive or toxic proteins were special induced in ED059. By combining the gene expression at the transcriptomic and proteomic levels, results reveal that no significantly high correlation was detected between the expression of mRNAs and proteins. Functional analysis of three candidate genes was accomplished by Agrobacterium-mediated transformation in Arabidopsis thaliana, and a MYB transcription factor 109 (MYB109) was identified with herbivore defense function in ED059.  This study provides insights into the identification of the potential genes and proteins involved in herbivore defense, and a possible understanding of the herbivore defense mechanism in a wild soybean by combining analysis of transcriptome and proteome data. Moreover, a novel gene associated with herbivore resistance function was identified and validated.

sample_organism:
Glycine max
Glycine soja

platform_organism:
Glycine max
Glycine soja

overall_design: Totally four samples, two treatments and two genotypes.

first: X

last: Wang

last: Zhou