title: Epigenetics markers ChIPSeq  in mouse Alkbh1 KO and WT ESC

first: Andrew

last: Xiao

summary:
It has widely accepted that 5-methylcytosine is the only form of DNA methylation in mammalian genomes, whereas the other forms, such as N6-methyladenine, primarily exist in prokaryotes and only a few eukaryotes.  Herein, we demonstrated the surprising presence of N6-methyladenine in mammalian genomes, especially, mouse embryonic stem cells. This modification is enriched at histone variant H2A.X-deposited genomic regions in wild type embryonic stem cells.  Our work also showed that a previously unknown DNA demethylase, Alkbh1, is the major demethylase for N6-methyladenine in embryonic stem cells.  Increase of N6-methyladenine levels in Alkbh1 deficient cells leads to silencing of genes that regulate embryonic development. Surprisingly, genes located on the X-chromosome, but not the Y-chromosome or autosomes are preferentially silenced by N6-methyladenine. Strikingly, N6-methyladenine in Alkbh1 deficient cells are specifically deposition at young, full-length subfamilies of LINE1 transposons that are strongly enriched on the X chromosome. Furthermore, N6-methyladenine deposition on LINE1s pattern is inversely correlated with their evolutionary age.   The deposition of N6-methyladenine results in epigenetic silencing of such L1s, which are otherwise actively transcribed in wild type embryonic stem cells, and the neighboring enhancers and genes.  Furthermore, N6-methyladenine induced-silencing resists gene activation signals during embryonic stem cell differentiation. Thus, N6-methyladenine adopts a new function in epigenetic silencing in evolution, distinct from its role in gene activation in other organisms. In summary, our results demonstrate that N6-methyladenine unexpectedly constitutes a crucial component of the epigenetic regulation repertoire in mammalian genomes.
First, we used different histone antibodies to enrich for DNA molecules with histone modification or specific variant in mouse ESCs as previously described Native-ChIP methods. Then, co-purified DNA molecules from WT or KO ESCs were subject to HiSeq2000 sequencing and data analysis for histone modification or variant peaks.

overall_design: Native ChIP coupling with HiSeq sequencing

first: Tao

last: Wu

first: Kaixuan

last: Lin


title: RNASeq in mouse Alkbh1 KO and WT ESC

status: Public on Mar 30 2016

city: New Haven

middle: Zhuo

state: CT

address: 10 Amistad St

department: Genetics

country: USA

institute: Yale Stem Cell Center

type: Expression profiling by high throughput sequencing

summary:
It has widely accepted that 5-methylcytosine is the only form of DNA methylation in mammalian genomes, whereas the other forms, such as N6-methyladenine, primarily exist in prokaryotes and only a few eukaryotes.  Herein, we demonstrated the surprising presence of N6-methyladenine in mammalian genomes, especially, mouse embryonic stem cells. This modification is enriched at histone variant H2A.X-deposited genomic regions in wild type embryonic stem cells.  Our work also showed that a previously unknown DNA demethylase, Alkbh1, is the major demethylase for N6-methyladenine in embryonic stem cells.  Increase of N6-methyladenine levels in Alkbh1 deficient cells leads to silencing of genes that regulate embryonic development. Surprisingly, genes located on the X-chromosome, but not the Y-chromosome or autosomes are preferentially silenced by N6-methyladenine. Strikingly, N6-methyladenine in Alkbh1 deficient cells are specifically deposition at young, full-length subfamilies of LINE1 transposons that are strongly enriched on the X chromosome. Furthermore, N6-methyladenine deposition on LINE1s pattern is inversely correlated with their evolutionary age.   The deposition of N6-methyladenine results in epigenetic silencing of such L1s, which are otherwise actively transcribed in wild type embryonic stem cells, and the neighboring enhancers and genes.  Furthermore, N6-methyladenine induced-silencing resists gene activation signals during embryonic stem cell differentiation. Thus, N6-methyladenine adopts a new function in epigenetic silencing in evolution, distinct from its role in gene activation in other organisms. In summary, our results demonstrate that N6-methyladenine unexpectedly constitutes a crucial component of the epigenetic regulation repertoire in mammalian genomes.
RNASeq compare the differential expressed genes in Alkbh1 KO and WT ES cell

overall_design: total RNA with mRNA HiSeq sequencing


title: Novel DNA modification in mouse ESC

type: Genome binding/occupancy profiling by high throughput sequencing

type: Methylation profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Hormones, Genes, and Athleticism: Effect of Androgens on the Avian Muscular Transcriptome

city: Seoul

first: Jae-Hyung

last: Lee

address: 26 Kyungheedae-ro, Dongdaemun-gu

country: South Korea

institute: Kyung Hee University

summary: Male vertebrate social displays vary from physically simple to complex, with the latter involving exquisite motor command of the body and appendages. Studies of these displays have, in turn, provided substantial insight into neuromotor mechanisms. The neotropical golden-collared manakin (Manacus vitellinus) has been used previously as a model to investigate intricate motor skills because adult males of this species perform an acrobatic and androgen-dependent courtship display. To support this behavior, these birds express elevated levels of androgen receptors (AR) in their skeletal muscles. Here we use RNA sequencing to explore how testosterone (T) modulates the muscular transcriptome to support male manakin courtship displays. In addition, we explore how androgens influence gene expression in the muscles of the zebra finch (Taenopygia guttata), a model passerine bird with a limited courtship display and minimal muscle AR. We identify androgen-dependent, muscle-specific gene regulation in both species. In addition, we identify manakin-specific effects that are linked to muscle use during the manakin display, including androgenic regulation of genes associated with muscle fiber contractility, cellular homeostasis, and energetic efficiency. Overall, our results point to numerous genes and gene networks impacted by androgens in male birds, including some that underlie optimal muscle function necessary for performing acrobatic display routines. Manakins are excellent models to explore gene regulation promoting athletic ability.

sample_organism:
Taeniopygia guttata
Manacus vitellinus

platform_organism:
Taeniopygia guttata
Manacus vitellinus

overall_design: Compare gene expression profiles between two species in the absence of testosterone or in the presence of testosterone using RNA-Seq (Illumina platform: Hi-Seq 2000)

first: Matthew

middle: J

last: Fuxjager

first: Jae

middle: H

last: Bahn

first: Tak-Ming

last: Chan

first: Jenifer

middle: G

last: Chew

first: Xinshu

first: Barney

middle: A

last: Schlinger


title: Defining the Roles and Regulation of the GATA-factor ELT-2 in the C. elegans Endoderm

city: Fort Collins

first: Erin

last: Osborne Nishimura

state: CO

address: 1870 Campus Delivery, 200 W. Lake St.

department: Biochemistry and Molecular Biology

institute: Colorado State University

summary: We identified genome-wide sites of occupancy for the intestine-specific transcription factor ELT-2 in L3-staged N2 worm by performing ELT-2 ChIP-seq on whole worms; we performed RNA-seq on L3-staged N2 whole worms

overall_design: To identify DNA regions of direct ELT-2 occupancy, we performed the following ChIP-seq assays 1) ELT-2 ChIP-seq on L3 staged N2 C. elegans worms; 2) H3K4me3 ChIP-seq on matched L3 staged N2 worms; 3) Mock IgG-only ChIP-seq negative control on L3 stage N2 worms. Input DNA was also sequenced for each replicate. In addition, we performed RNA-seq on two replicates of L3 N2 C. elegans worms.

first: Jason

middle: D

last: Lieb

first: Tobias

last: Wiesenfahrt

first: Adam

last: Robinson

first: James

last: McGhee


title: H2A.X SMRT-ChIP

summary:
It has widely accepted that 5-methylcytosine is the only form of DNA methylation in mammalian genomes, whereas the other forms, such as N6-methyladenine, primarily exist in prokaryotes and only a few eukaryotes.  Herein, we demonstrated the surprising presence of N6-methyladenine in mammalian genomes, especially, mouse embryonic stem cells. This modification is enriched at histone variant H2A.X-deposited genomic regions in wild type embryonic stem cells.  Our work also showed that a previously unknown DNA demethylase, Alkbh1, is the major demethylase for N6-methyladenine in embryonic stem cells.  Increase of N6-methyladenine levels in Alkbh1 deficient cells leads to silencing of genes that regulate embryonic development. Surprisingly, genes located on the X-chromosome, but not the Y-chromosome or autosomes are preferentially silenced by N6-methyladenine. Strikingly, N6-methyladenine in Alkbh1 deficient cells are specifically deposition at young, full-length subfamilies of LINE1 transposons that are strongly enriched on the X chromosome. Furthermore, N6-methyladenine deposition on LINE1s pattern is inversely correlated with their evolutionary age.   The deposition of N6-methyladenine results in epigenetic silencing of such L1s, which are otherwise actively transcribed in wild type embryonic stem cells, and the neighboring enhancers and genes.  Furthermore, N6-methyladenine induced-silencing resists gene activation signals during embryonic stem cell differentiation. Thus, N6-methyladenine adopts a new function in epigenetic silencing in evolution, distinct from its role in gene activation in other organisms. In summary, our results demonstrate that N6-methyladenine unexpectedly constitutes a crucial component of the epigenetic regulation repertoire in mammalian genomes.
First, we used a native-ChIP approach to enrich for DNA molecules residing in H2A.X-deposition regions in mouse ESCs as previously described. Then, co-purified DNA molecules from WT or KO ESCs were subject to SMRT sequencing and data analysis for DNA modifications (Pacific Biosciences).

overall_design: H2A.X Native ChIP coupling with SMRT sequencing (separate the short "S" and long "L" part to sequencing)


title: Functional analysis of KAI1

status: Public on Mar 31 2016

first: Tae-Young

last: Roh

state: --- Select One ---

address: 52 Ewhayeodae-gil, Seodaemun-gu

department: Life Sciences

institute: Ewha Womans University

summary: We sequenced mRNA from mouse EML cell line under Kai1 knockdown or Kai1 overexpressing condition. KAI1 belongs to tetraspanin superfamily and is known as suppressor of tumor metastasis.

overall_design: Examination of mRNA levels in wildtype (WT) and KAI1 knockdown EML cells were generated by deep sequencing, in duplicate, using Illumina GAIIx. Examination of mRNA levels in wildtype (WT) and KAI1 overexpressing EML cells were also generated in single set.

first: Jin

last: Hur

first: Seokjin

last: Ham

first: Hyeongrin

last: Jeon

first: Hyo-Soo

last: Kim


title: Romidepsin and azacitidine synergize in their epigenetic modulatory effects to induce apoptosis in CTCL

city: Geneva

first: Phil

middle: Fang

last: Cheng

address: Rue Gabrielle-Perret-Gentil 4

department: Oncology

country: Switzerland

institute: Hopitaux universitaires de Geneve

summary: We demonstrate that the combination of the HDAC inhibitor, romidepsin, and the demethylating agent, azacitidine, exerts synergistic anti-proliferative effects and induction of apoptosis involving activation of the caspase cascade in CTCL cell lines as well as CD4+ T cells derived from patients with Sézary Syndrome (SS) with high tumor burden. We identified genes that were selectively induced by the combination treatment, such the tumor suppressor gene RhoB.

overall_design: We performed global CpG methylation-sequencing in a CTCL cell line and in patient cells. Results showed a subset of genes with a unique change in methylation profile in the combination treatment as compared to single drugs.

first: Sima

last: Rozati

middle: F

first: Daniel

middle: S

last: Widmer

first: Kazuyasu

last: Fujii

first: Mitchell

middle: P

last: Levesque

first: Reinhard

last: Dummer


title: Overproduction of indole-3-acetic acid in free-living rhizobia mimics physiological conditions required for nitrogen fixation inside root nodules

city: Naples

first: Carmen

last: Bianco

address: Via P. Castellino 111

department: IBBR

country: Italy

institute: CNR

summary:
Background: The overproduction of the main auxin indole-3-acetic acid (IAA) in Sinorhizobium meliloti RD64 leads to beneficial effects both in free-living rhizobium and in host plant during symbiosis. These effects could be related to alteration in gene expression induced by IAA. Preliminary studies of these alterations induced in free-living cells have already started. In this work the transcriptional profiles of free-living S. meliloti wild type 1021 and RD64 strains obtained by RNA-seq analysis were used to better investigate this issue.
Results: Transcriptome analysis by next-generation sequencing (RNA-seq) allowed the identification of the genes differentially expressed in RD64 cells, grown in a rich medium under aerobic conditions. Most of the up-regulated genes can be classified into two main categories. The first class comprised the main regulator fixJ and the two intermediate regulators fixK and nifA, which control the expression of genes involved in nitrogen-fixation. For these two latter genes the highest expression levels were observed. Several other genes, known to be FixJ targets, such as those involved in denitrification process and proline metabolism, were also identified.  The second class of up-regulated genes comprised the gene coding for the sigma factor RpoH1 and other genes regulated in RpoH1-dependent manner in S. meliloti and involved in stress response. The main category of genes negatively affected can be classified in transport of sugars and other small molecules, chemotaxis and cell envelope functions. Under microaerobic condition, quantitative real-time PCR analysis of genes coding for the two-component regulatory system FixJL and for the regulator NifA showed that for all three genes the induction levels measured in RD64 cells were even higher when compared to aerobic conditions. In particular, for the fixJ gene the expression level increased more than tenfold. The up-regulation of nifA gene was also observed under aerobic conditions both when nitrogen level in the medium was limiting and when it was high.
Conclusion: This work provided evidence that the overexpression of the auxin IAA in S. meliloti free-living cells triggers the induction of many genes involved in bacterial response to different abiotic stresses and allows the expression of key genes involved in nitrogen fixation.

overall_design: mRNA profiles of S. meliloti wild type 1021 and its derivative IAA-overproducing RD64 strain were generated in triplicate by using HiSeq2000 sequencer

first: Roberto

last: Defez

first: Roberta

last: Esposito

first: Claudia

last: Angelini


title: ChIP-seq analysis of Arid1a and C/ebpα in the mouse livers

city: shanghai

first: Xuxu

last: Sun

address: 227 south chongqing road

department: school of medicine

country: China

institute: shanghai jiao tong university

summary: Arid1a is the subunit of SWI/SNF complex, which was reported to guide SWI/SNF to DNA. Here, we found that loss of Arid1a in the liver and other adult tissues results in improved organ regeneration. Within SWI/SNF complexes, Arid1a physically interacts with C/ebpα, a hepatocyte transcription factor that drives maturation and limits proliferation. Genome-wide analysis showed that loss of Arid1a reduces the recruitment and activity of C/ebpα on target promoters, resulting in expression programs that favor regeneration and cellular fitness during injury. Arid1a binding is enriched in promoters near transcriptional start sites (TSSs), and C/ebpα binds at precisely the same positions, indicating that Arid1a facilitates C/ebpα binding across the genome.

overall_design: Perfuse and isolate primary hepatocytes from mice livers, analysis of genomic occupancy of C/ebpα and H3K4me2 in hepatocytes from Arid1a WT and Arid1a liver specific KO mice by ChIP-seq. Analysis of genomic occupancy of Arid1a in the hepatocytes from V5-Arid1a transgenic mouse by ChIP-seq.

first: Chao

last: Xing

first: Hao

last: Zhu


title: Profiling gene expression in M. tuberculosis under different in vitro conditions.

city: College Station

first: Thomas

middle: R

last: Ioerger

state: TX

address: 3112 TAMUS

department: Department of Computer Science

institute: Texas A&M University

summary: This RNA-Seq study of gene expression in M. tuberculosis was done as part of the FLUTE project (NIH grant U19-AI107774, Eric Rubin, program director).

sample_organism:
Mycobacterium tuberculosis H37Rv
Mycobacterium tuberculosis HN878

overall_design: There are 3 replicates each for Mtb H37Rv grown under the following conditions: 0.1% butyrate, 0.4% glucose, butyrate+glucose, low iron, acidic conditions, and several RIF-resistant mutants of Beijing strain HN878 with different mutations in RpoB.

first: Sarah

last: Fortune

first: Jonathan

last: Livny


title: A mechanism of resistance to gefitinib mediated by cellular reprogramming and the acquisition of an FGF2-FGFR1 autocrine growth loop

status: Public on Mar 29 2016

city: Cleveland

first: Jihye

last: Kim

state: Ohio

address: 9500 Euclid Ave.

department: Quantitative Health Sciences

institute: Cleveland Clinic

summary: Despite initial and often dramatic responses of epidermal growth factor receptor (EGFR)-addicted lung tumors to the EGFR-specific tyrosine kinase inhibitors (TKIs), gefitinib and erlotinib, nearly all develop resistance and relapse. To explore novel mechanisms mediating acquired resistance, we employed non-small-cell lung cancer (NSCLC) cell lines bearing activating mutations in EGFR and rendered them resistant to EGFR-specific TKIs through chronic adaptation in tissue culture. In addition to previously observed resistance mechanisms including EGFR-T790M 'gate-keeper' mutations and MET amplification, a subset of the seven chronically adapted NSCLC cell lines including HCC4006, HCC2279 and H1650 cells exhibited marked induction of fibroblast growth factor (FGF) 2 and FGF receptor 1 (FGFR1) mRNA and protein. Also, adaptation to EGFR-specific TKIs was accompanied by an epithelial to mesenchymal transition (EMT) as assessed by changes in CDH1, VIM, ZEB1 and ZEB2 expression and altered growth properties in Matrigel. In adapted cell lines exhibiting increased FGF2 and FGFR1 expression, measures of growth and signaling, but not EMT, were blocked by FGFR-specific TKIs, an FGF-ligand trap and FGFR1 silencing with RNAi. In parental HCC4006 cells, cell growth was strongly inhibited by gefitinib, although drug-resistant clones progress within 10 days. Combined treatment with gefitinib and AZD4547, an FGFR-specific TKI, prevented the outgrowth of drug-resistant clones. Thus, induction of FGF2 and FGFR1 following chronic adaptation to EGFR-specific TKIs provides a novel autocrine receptor tyrosine kinase-driven bypass pathway in a subset of lung cancer cell lines that are initially sensitive to EGFR-specific TKIs. The findings support FGFR-specific TKIs as potentially valuable additions to existing targeted therapeutic strategies with EGFR-specific TKIs to prevent or delay acquired resistance in EGFR-driven NSCLC.

overall_design: Examination of mRNA levels in DMSO and gefitinib-resistant cultures of HCC4006 and HCC827. Each group has two replicates.

first: K

middle: E

last: Ware

first: T

middle: K

last: Hinz

first: E

last: Kleczko

last: Singleton

first: L

last: Marek

first: B

last: Helfrich

first: C

middle: T

last: Cummings

first: D

last: Graham

last: Astling

first: A-C

last: Tan

last: Heasley


title: RISC-mediated control of selected chromatin regulators stabilizes ground state pluripotency of mouse embryonic stem cells (TRAP)

city: Genova

first: Luca

last: Pandolfini

state: Genova

address: Via Enrico Melen 83, Building B

department: Center for Human Technologies

institute: Istituto Italiano di Tecnologia - I

summary: We performed an integrated analysis of RNA and proteins at the transition between naïve ES cells and cells primed to differentiate. During this transition, mRNAs coding for chromatin regulators were specifically released from translational inhibition mediated by RNA-Induced Silencing Complex (RISC). This suggests that, prior to differentiation, the propensity of ES cells to change their epigenetic status is hampered by RNA interference. The expression of these chromatin regulators was reinstated following acute inactivation of RISC, and it correlated with loss of stemness markers and activation of early cell differentiation markers in treated ES cells.

overall_design: We evaluated ribosome occupancy of mRNAs in embryonic stem cells and Epiblast-Like Aggregates by means of polysome profiling with a modified Translating Ribosome Affinity Purification protocol (TRAP).

first: Ettore

last: Luzi

first: Dario

last: Bressan

first: Nadia

last: Ucciferri

first: Michele

last: Bertacchi

first: Rossella

last: Brandi

first: Silvia

last: Rocchiccioli

first: Mara

last: D'Onofrio

first: Federico

last: Cremisi


title: Lung microbiome enriched with anaerobes associated with Th17 immune phenotype

city: New York

first: Leopoldo

middle: Nicolas

last: Segal

state: NY

address: 462 First Avenue NBV 6N5-6N8

department: Medicine

institute: NYU School of Medicine

summary: In a prior report, we observed two distinct lung microbiomes in healthy subjects that we termed “pneumotypes”: pneumotypeSPT, characterized by high bacterial load and supraglottic predominant taxa (SPT) such as the anaerobes  Prevotella and Veillonella; and pneumotypeBPT, with low bacterial burden and background predominant taxa (BPT) found in the saline lavage and bronchoscope. Here, we determined the prevalence of these two contrasting lung microbiome types, in a multi-center study of healthy subjects. We confirmed that a lower airway microbiome enriched with upper airway microbes (pneumotypeSPT) was present in ~45% of healthy individuals.

overall_design:
Cross-sectional Multicenter cohort. BAL of 49 healthy subjects from three cohort had their lower airway microbiome assessed by 16S rDNA sequencing and microbial gene content (metagenome) was computationally inferred from taxonomic assignments.

The amplicons from total 100 samples are barcoded; the barcode and other clinical characteristics (e.g. inflammatory biomarkers and metabolome data) for each sample are provided in the 'Pneumotype.sep.Map.A1.txt' file.

first: Michael

last: Weiden


title: Ribosome hibernation factor promotes Staphyloccocal survival and differentially represses translation

city: Chicago

first: Mee-Ngan

middle: F.

last: Yap

state: IL

address: 320 E Superior St

department: Microbiology-Immunology

institute: Northwestern University Feinberg School of Medicine

summary: We sequenced a total of 16 cDNA libararies derived from fragmented total mRNA and ribosome protected mRNAs from S. aureus hpf mutant (NE838) and its parental strain JE2 grown in tryptic soy broth (TSB) or minimal medium (MM). The data represented 2 independent biological replicates.

overall_design: Examination of the impact of hpf on global S.aureus translatome, mRNA abundance and the ribosome density along the transcripts.

first: A

last: Basu


title: RISC-mediated control of selected chromatin regulators stabilizes ground state pluripotency of mouse embryonic stem cells

type: Non-coding RNA profiling by high throughput sequencing


title: STAT5 paralog dose governs T cell effector and regulatory function

status: Public on Mar 28 2016

city: Bethesda

first: Yuka

last: Kanno

state: MD

address: 10 Center Drive, Rm13C120

department: NIAMS

institute: NIH

summary: The transcription factor STAT5 is fundamental to the mammalian immune system. However, the relationship between its two paralogs, STAT5A and STAT5B, and the extent to which they are functionally distinct, remains controversial. We addressed this longstanding question in primary CD4+ 'helper' T cells, the principal orchestrators of adaptive immunity. Using a combination of genetic and genomic approaches, we demonstrate that, although both influence regulatory (Treg) and effector T cell responses, and control many of the same genes, they are not functionally equivalent and, in fact, only the latter is required for immunological tolerance. Differences in genomic distribution and transcriptomic output support the conclusion that STAT5B is dominant and, surprisingly, point towards relative abundance (i.e. paralog dose), rather than unique functional capabilities, as the principal distinguishing feature. Collectively, our data provide a unifying model for the discrete and redundant activities of STAT5A and STAT5B, establishing that asymmetrical expression underlies paralog specificity (or dominance) in the face of widespread structural homology.

overall_design: This dataset includes 55 individual samples of transcriptome or STAT5 distibution data from cytokine treated CD4+T cells. Each culture condition includes at least 2 biological replicates per genotype.

middle: V

last: Villarino

first: H

middle: W

last: Sun

first: G

last: Gutierrez-Cruz

first: Y

first: J

last: O'Shea


title: Integrated epigenomic analyses of enhancer as well as promoter regions in gastric cancer

city: Daejeon

first: sujin

last: Baek

address: Gwahank-ro 125

department: MGRC

institute: KRIBB


title: Enhanced CLIP (eCLIP) enables robust and scalable transcriptome-wide discovery and characterization of RNA binding protein binding sites [iCLIP]

city: La Jolla

first: Gene

last: Yeo

state: CA

address: 2880 Torrey Pines Scenic Dr. Room 3805/Yeo Lab

institute: UCSD

summary: RNA binding proteins (RBPs) play essential roles in cellular physiology by interacting with target RNAs. As defects in protein-RNA recognition lead to human disease, UV-crosslinking and immunoprecipitation (CLIP) of ribonuclear complexes followed by deep sequencing (-seq) is critical in constructing protein-RNA maps to expand our understanding of RBP function. However, current CLIP protocols are technically demanding and involve low complexity libraries that yield squandered sequencing of PCR duplicates and high experimental failure rates. To enable truly large-scale implementation of CLIP-seq, we have developed an enhanced CLIP methodology (eCLIP) that features a decrease of ~10 cycles of requisite amplification with a concomitant >60% decrease in discarded PCR duplicate reads, while maintaining the ability to identify RNA binding with single-nucleotide resolution. By simplifying the generation of paired IgG and size-matched input controls, eCLIP also dramatically improves specificity in discovery of authentic binding sites. To demonstrate that eCLIP enables large-scale and robust profiling of RBPs, 102 eCLIP experiments in biological duplicate for a diverse collection of 74 RBPs in HepG2 and K562 cells were completed (available at https://www.encodeproject.org). We establish that eCLIP is comparable in amplification and sample requirements to ChIP-seq, and enables integrative analysis of diverse RBPs to reveal factor-specific profiles, common artifacts for CLIP experiments and RNA-centric perspectives of RBP activity.

overall_design: iCLIP-seq against RBFOX2 in 293T Cells

first: Eric

last: Van Nostrand

first: Gabriel

last: Pratt

first: Alexander

last: Shishkin

first: Chelsea

last: Gelboin-Burkhart

first: Mark

last: Fang

first: Balaj

last: Sundararaman

first: Steven

last: Blue

first: Thai

last: Nguyen

first: Christine

last: Surka

first: Keri

last: Elkins

first: Rebecca

last: Stanton

first: Frank

last: Rigo

first: Mitchell

last: Guttman


title: Enhanced CLIP (eCLIP) enables robust and scalable transcriptome-wide discovery and characterization of RNA binding protein binding sites [eCLIP - 293T cells]

overall_design: eCLIP-seq against RBFOX2 and SLBP in 293T Cells


title: Classical dendritic cells are required for dietary antigen-mediated peripheral regulatory T cell and tolerance induction I

city: NEW YORK

first: Thiago

middle: Y

last: Oliveira

state: New York

address: 1230 YORK AVE

department: Immunology, Virology and Microbiology

institute: The Rockefeller University

summary: Oral tolerance prevents pathological inflammatory responses towards innocuous foreign antigens via peripheral regulatory T cells (pTreg cells). However, whether a particular subset of antigen-presenting cells (APCs) is required during dietary antigen exposure to instruct naïve CD4+ T cells to differentiate into pTreg cells has not been defined. Using myeloid lineage-specific APC depletion in mice, we found that monocyte-derived APCs are dispensable, while classical dendritic cells (cDCs) are critical for pTreg cell induction and oral tolerance. CD11b¬– cDCs from the gut-draining lymph nodes efficiently induced pTreg cells, and conversely, loss of IRF8-dependent CD11b– cDCs impaired their polarization, although oral tolerance remained intact. These data reveal the hierarchy of cDC subsets in pTreg cell induction and their redundancy during oral tolerance development.

overall_design: Four dendritic cell subpopulations from mouse mesenteric lymphnodes were sorted and compared in their gene expression profile

first: Daria

last: Esterházy

first: Jakob

last: Loschko

first: Mariya

last: London

first: Veronica

last: Jove

first: Daniel

last: Mucida


title: Identification of CLF-regulated transcripts in embryos at the mature-green stage by strand-specific RNA-sequencing in Arabidopsis

city: No.12 Zhongguancun South Street, Haidian

first: Jun

last: Liu

state: Beijing

address: 1230 York Avenue

department: Chinese Academy of Agricultural Sciences

institute: Institute of Crop Science

summary: CURLY LEAF (CLF), the major histone methyltransferase of Polycomb Repressive Complex 2 (PRC2), modifies trimethylation of histone H3 lysine 27 (H3K27me3) and mediates dynamical chromatin repression in Arabidopsis. Our previous results suggested that CLF may play regulatory roles on seed size and lipid biosynthesis in embryos at the mature-green stages. Here we used strand specific RNA-sequencing to profile Arabidopsis transcriptomes obtained from isolated embryos at the mature-green stage of Col-0 and clf-28 plants. Our analysis identified a large number of CLF-regulatedd transcripts in developmenting embryos in Arabidopsis.

overall_design: Issolated embryos of WT and clf-28 with 2 biological replicates

first: Hui-Wen

last: Wu

first: Shulin

last: Deng

first: Huan

last: Wang

first: Nam-Hai

last: Chua


first: sujin

last: Baek

summary: We performed H3K27ac ChIP-seq data in seven gastric cancer cell lines to investigate the functional significance of epigenetic changes at distal regulatory regions

overall_design: H3K27ac ChIP-seq profiling of gastric cancer cell lines were generated by deep sequencing, in each seven samples, using Illumina GAIIx and Hiseq-2000

first: Seon-Young

last: Kim

first: Su-Jin


title: Identification of CLF-regulated transcripts using strand-specific RNA-sequencing in Arabidopsis

summary: CURLY LEAF (CLF), the major histone methyltransferase of Polycomb Repressive Complex 2 (PRC2), modifies trimethylation of histone H3 lysine 27 (H3K27me3) and mediates dynamical chromatin repression in Arabidopsis. Here we used strand specific RNA-sequencing to profile Arabidopsis transcriptomes obtained from roots, shoots, flowers and siliques of Col-0 and clf-28 plants. Our analysis identified a large number of CLF-regulatedd transcripts in Arabidopsis.

overall_design: Transcriptome profiling in roots, shoots, inflorescences and siliques of WT and clf-28 plants with 3 biological replicates.

first: Jian

last: Ye


title: Transcriptome profiling of siliques at 4 developmental stages in Arabidopsis

summary: To search for the differential expressed genes during postferlization and embryonic development in Arabidopsis, We profiled transcriptome of siliques at 4 developmental stages using RNA-seq based on poly(A) selection. Each RNA library yielded 100 million 101-bp paired-end reads.Using Tophat and Cufflinks with the Arabidopsis TAIR10 annotation as reference, we identifed 33,451 assembled transcripts in 4 samples. Of them, 5,608 were up-regulated genes in at least one sample. A group of them are preferentially expressed at mature green-stage of seeds.

overall_design: Transcriptome profiling in 0-5 day, 6-10 day, 11-15 day and 16-20 day post-anthesis siliques


title: Enhanced CLIP (eCLIP) enables robust and scalable transcriptome-wide discovery and characterization of RNA binding protein binding sites

first: Gene

last: Yeo

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

overall_design: Refer to individual Series


summary: To understand epigenetic changes in the distal regulatory as well as proximal regions, we performed RNA-seq, MBD-seq, and H3K27ac ChIP-seq on gastric tissues and cell lines

overall_design: MBD sequencing of normal tissue (n), purified gastric cancer (sc), and cultured gastric cancer cell (dc) were generated by deep sequencing, in five samples from three patients (csc1, csc2, csc3) and two replicates (csc1_sc2, csc1_sc3), using Illumina GAIIx.

first: Sae-Ock

last: Oh


city: Stanford

first: Yang

middle: I

last: Li

state: CALIFORNIA

address: Department of Genetics

department: genetics

institute: Stanford University

summary: Noncoding variants play a central role in the genetics of complex traits, but we still lack a full description of the main molecular pathways through which they act. Here we used molecular data to quantify the contribution of cis-acting genetic effects at each major stage of gene regulation from chromatin to proteins, within a population sample of Yoruba lymphoblastoid cell lines (LCLs). We performed 4sU metabolic labeled transcripts in 65 YRI LCLs to identify genetic variants that affect transcription rates. As expected, we found an important contribution of genetic variation via chromatin, contributing ∼65% of eQTLs (expression Quantitative Trait Loci). The remaining eQTLs, which are not asso- ciated with chromatin-level variation, are highly enriched in transcribed regions, and hence may affect expression through co- or post-transcriptional processes.

overall_design: International HapMap lymphoblastoid cell lines (LCLs) derived from YRI (Yoruba in Ibadan, Nigeria); We adapted the 4sU labelling method from (PMID 21516085). Briefly, cell cultures were grown to log phase in volumes sufficient to yield about 300 ng of 4sU-labeled RNA. Cells were incubated with 4sU for the required length of time (0, 30, or 60 minutes), then washed, pelleted, and frozen. Total RNA was extracted, and 4sU-labeled RNA was separated from total RNA using a bead-based biotin-streptavidin purification protocol. We sequenced metabolic labeled transcripts in 65 YRI LCLs 30 minutes and 60 minutes after incubation.

first: Jonathan

last: Pritchard

first: Allegra

last: Petti

first: Yoav

last: Gilad


title: XX-650-23, an inhibitor of CREB-CBP interaction, effect on acute myeloid leukemia cell line

status: Public on Mar 26 2016

first: Kathleen

middle: M

last: Sakamoto

state: California

address: CCSR 1215C, 235 Campus Drive

department: Pediatrics

summary: Analysis of acute myeloid leukemia KG1 cells treated with XX-650-23 (5 mM) for 12hrs. XX-650-23 inhibits proliferation of KG1 cells. Results provides insight into the molecular mechanisms underlying by the inhibitory activity of XX-650-23 on acute leukemia.

overall_design:
mRNA profiles of KG1 cells treated with XX-650-23 were generated by deep sequencing.
CREB genome binding profiles of KG1 cells treated with XX-650-23 were generated by deep sequencing.

first: Bryan

last: Mitton

first: Hee-Don

last: Chae

first: Roberto

last: Ferrari


title: Tristetraprolin (TTP) regulated splenic transcriptome in "Triple KO" mice

status: Public on Mar 27 2016

city: Research Triangle Park

first: Perry

middle: J

last: Blackshear

state: NC

address: 111 T.W. alexander drive

department: LST

institute: NIEHS

summary: Tristetraprolin (TTP)  binds to specific AU-rich elements in the 3'UTR of certain transcripts and regulates post-transcriptional gene expression by increasing the rate of mRNA turnover.  In this study, we evaluated the effects of TTP deficiency on the overall gene expression of spleen tissue, in order to discover tissue specific targets of TTP under normal physiologic conditions.  We utilized "Triple KO" (Zfp36-/-/TNFR1-/-/TNFR2-/-) mice that are deficient in TTP and two TNF receptors and compared the transcriptomic changes to "Double KO" (TNFR1-/-/TNFR2-/-) and WT mice.  Spleen mRNA from four WT, four "Double KO", and four "Triple mice" was subjected to RNA-Seq in two phases.  All the animals used in this study were males between the ages of 12-14 weeks and were on a mixed (75% C57BL/6NTac, 25% 129/SVEV) background.

overall_design: Examination of splenic gene expression difference between "Triple KO"-WT and "double KO"-WT data sets

first: Sonika

last: Patial

first: Deborah

last: Stumpo

first: W

last: Young III

first: James

last: Ward

first: Gordon

last: Flake


first: sujin

last: Baek

summary: To understand epigenetic changes in the distal regulatory as well as proximal regions, we performed RNA-seq, MBD-seq, and H3K27ac ChIP-seq on gastric tissues and cell lines.

overall_design: mRNA sequencing profiles of normal tissue (n), purified gastric cancer (sc), and cultured gastric cancer cell (dc) were generated by deep sequencing, in five samples from three patients (csc1, csc2, csc3) and two replicates (csc1_sc2, csc1_sc3), using Illumina GAIIx and HiSeq2000.

first: Seon-Young

last: Kim

first: Su-Jin


title: Pho and Ph, H3K27me3 and Cg ChIP-seq data in Drosophila third instar larval brains and discs

city: North Bethesda

first: sandip

last: de

address: 6 center drive

department: rm 3B-331, bldg 6B

institute: NICHD/NIH

summary: Genome-wide binding profile of PcG proteins- Ph, H3K27me3 and Cg in Drosophila third instar larval brains and disks, in duplicate, using Illumina HiSeq 2500. The sequence reads that passed quality filters were analyzed.

overall_design: Drosophila third instar larval brains and discs were dissected and fixed with 2% HCHO before sonication. After sonication, chromatin immunoprecipitation was carried out.

first: Sandip

last: De

first: Judith

last: Kassis

first: Harold

last: Smith


title: Transcriptomic and Epigenetic analysis of cell line derivatives from H2087 and HCC1954 cell lines

first: Danilo

middle: G

last: Macalinao

address: 1275 York Avenue

institute: MSKCC

summary: We profiled the transcriptomes of latency-competent cells derived from the human cancer cell lines H2087 (lung adenocarcinoma) and HCC1954 (breast adenocarcinoma) in mitogen-rich and mitogen-low media (MRM and MLM, respectively). In addition, we analyzed the epigenetic landscape of these cell lines under MLM conditions.

overall_design: H2087 and HCC1954 parental and latency-competent cell derivatives (LCCs) were grown for 48hr in mitogen-rich or mitogen-low conditions in vitro, and whole RNA was extracted for RNA-seq profiling. Cell lines were also grown in MLM conditions and DNA extracted for ChIP-Seq experiments.

first: Joan

last: Massague

first: Srinivas

last: Malladi


title: Dnmt3a haploinsufficiency transforms Flt3-ITD myeloproliferative disease into a rapid, spontaneous, and fully-penetrant acute myeloid leukemia (Single Cell RNA-Seq)

city: Cincinnati

first: Nathan

last: Salomonis

state: OH

address: 3333 Burnet Avenue

department: Biomedical Informatics

institute: Cincinnati Children's Hospital

summary: Cytogenetically normal acute myeloid leukemia (CN-AML) represents nearly 50% of human acute myeloid leukemia (AML) cases with a 5-year overall survival of approximately 30%. In CN-AML with poorer prognosis, mutations in the de novo DNA methyltransferase (DNMT3A) and the FMS-like tyrosine kinase 3 (Flt3) commonly co-occur (1-3). We demonstrate that mice with Flt3-internal-tandem duplication (Flt3ITD) and inducible deletion of Dnmt3a spontaneously develop a rapidly-lethal, completely-penetrant, and transplantable AML of normal karyotype. These murine AML retain a single Dnmt3a floxed allele, revealing the oncogenic potential of Dnmt3a haploinsufficiency. FLT3-ITD/DNMT3A-mutant primary human and murine AML demonstrate a similar pattern of global DNA methylation. In the murine model, rescuing DNMT3A expression was accompanied by DNA re-methylation and loss of clonogenic potential, suggesting that Dnmt3a-mutant oncogenic effects are reversible.  Differentially methylated genomic regions were associated with changes in the expression of nearby genes. Moreover, dissection of the cellular architecture of the AML model using single-cell RNA-Seq, flow cytometry and colony assays identified clonogenic subpopulations that differentially express genes that are sensitive to the methylation of nearby genomic loci and varied in response to Dnmt3a levels. Thus, Dnmt3a haploinsufficiency transforms Flt3ITD myeloproliferative disease by modulating methylation-sensitive gene expression within a clonogenic AML subpopulation.

overall_design: To identify the gene expression changes associated with Dnmt3a loss of function in human and murine Flt3-ITD and Dnmt3a-mutant AML (Single Cell RNA-Seq).

first: Sara

last: Meyer

first: Tingting

last: Qin

first: David

last: Muench

first: Kohei

last: Masuda

first: Meenakshi

last: Venkatasubramanian

first: Emily

last: Orr

first: Elisabeth

last: Paietta

first: Martin

last: Tallman

first: Hugo

last: Fernandez

first: Ari

last: Melnick

first: Michelle

last: Beau

first: Scott

last: Kogan

first: Maria

last: Figueroa

first: H

middle: L

last: Grimes


title: Single cell analysis of long non-coding RNAs in the developing human neocortex

city: SAN FRANCISCO

first: John

last: Liu

address: 35 Medical Center Way

institute: UCSF

summary: Long non-coding RNAs (lncRNAs) comprise a diverse class of transcripts that can regulate molecular and cellular processes in brain development and diseasee.  LncRNAs exhibit cell type- and tissue-specific expression, but little is known about the expression and function of lncRNAs in the developing human brain.  Here, we deeply profiled lncRNAs from polyadenylated and total RNA obtained from human neocortex at different stages of development and integrated this resource to analyze the transcriptomes of single cells.  While lncRNAs were generally detected at low levels in whole tissues, single cell transcriptomics revealed that many lncRNAs are abundantly expressed in individual cells and are cell type-specific.  Furthermore, we used CRISRPi to show that LOC646329, a lncRNA enriched in radial glia but detected at low abundance in tissues, regulates cell proliferation. The discrete and abundant expression of lncRNAs among individual cells has important implications for both their biological function and utility for distinguishing neural cell types.

overall_design:
16 Bulk Tissue Samples from GW13-23; 226 Single Cells from GW19.5-23.5
------------------
bulk_tpm.polya.txt: bulk RNA-seq expression; using polyA full reference
scell_ncounts.genes.thresh.txt: single cell RNA-seq expression; using polyA stringent reference; includes 50 GW16, GW21, GW21p3 cells previously analyzed (Pollen et. al. 2014)
polya_RNA_stringent_ref.gtf: bulk RNA-seq polyA stringent transcriptome reference
polya_RNA_full_ref.gtf: bulk RNA-seq polyA full transcriptome reference
total_RNA_stringent_ref.gtf: bulk RNA-seq total stringent transcriptome reference
total_RNA_full_ref.gtf: bulk RNA-seq total full transcriptome reference
GW13_1_polya_minus.bw: strand-specific bulk RNA-seq alignment signal
GW13_1_polya_plus.bw: strand-specific bulk RNA-seq alignment signal
GW13_1_total_minus.bw: strand-specific bulk RNA-seq alignment signal
GW13_1_total_plus.bw: strand-specific bulk RNA-seq alignment signal
GW14.5_1_polya_minus.bw: strand-specific bulk RNA-seq alignment signal
GW14.5_1_polya_plus.bw: strand-specific bulk RNA-seq alignment signal
GW14.5_1_total_minus.bw: strand-specific bulk RNA-seq alignment signal
GW14.5_1_total_plus.bw: strand-specific bulk RNA-seq alignment signal
GW16_1_polya_minus.bw: strand-specific bulk RNA-seq alignment signal
GW16_1_polya_plus.bw: strand-specific bulk RNA-seq alignment signal
GW16_1_total_minus.bw: strand-specific bulk RNA-seq alignment signal
GW16_1_total_plus.bw: strand-specific bulk RNA-seq alignment signal
GW16_2_polya_minus.bw: strand-specific bulk RNA-seq alignment signal
GW16_2_polya_plus.bw: strand-specific bulk RNA-seq alignment signal
GW16_2_total_minus.bw: strand-specific bulk RNA-seq alignment signal
GW16_2_total_plus.bw: strand-specific bulk RNA-seq alignment signal
GW21_1_polya_minus.bw: strand-specific bulk RNA-seq alignment signal
GW21_1_polya_plus.bw: strand-specific bulk RNA-seq alignment signal
GW21_1_total_minus.bw: strand-specific bulk RNA-seq alignment signal
GW21_1_total_plus.bw: strand-specific bulk RNA-seq alignment signal
GW21_2_polya_minus.bw: strand-specific bulk RNA-seq alignment signal
GW21_2_polya_plus.bw: strand-specific bulk RNA-seq alignment signal
GW21_2_total_minus.bw: strand-specific bulk RNA-seq alignment signal
GW21_2_total_plus.bw: strand-specific bulk RNA-seq alignment signal
GW23_1_polya_minus.bw: strand-specific bulk RNA-seq alignment signal
GW23_1_polya_plus.bw: strand-specific bulk RNA-seq alignment signal
GW23_1_total_minus.bw: strand-specific bulk RNA-seq alignment signal
GW23_1_total_plus.bw: strand-specific bulk RNA-seq alignment signal
GW23_2_polya_minus.bw: strand-specific bulk RNA-seq alignment signal
GW23_2_polya_plus.bw: strand-specific bulk RNA-seq alignment signal
GW23_2_total_minus.bw: strand-specific bulk RNA-seq alignment signal
GW23_2_total_plus.bw: strand-specific bulk RNA-seq alignment signal

first: J

first: T

last: Nowakowski

first: A

last: Pollen

last: Lui

last: Kriegstein

first: Diaz

last: Aaron

first: Lim

last: Daniel


title: Next Generation Sequencing Facilitates Quantitative Analysis of Wild Type and dek35 mutant Transcriptomes

status: Public on Mar 25 2016

city: Shanghai

first: Xinze

last: Chen

address: 333 Nanchen Road

institute: Shanghai University

summary: Analysis of  gene expression level. The hypothesis tested in the present study was that dek35 mutant influence the expression of mitochondrial oxidative phosphorylation associated genes. Results provide important information of the gene expression level regulation of mitochondrial complexes assembly proteins, translational elongation factors and genes associated with other biological processes.

overall_design: Endosperm mRNA profiles of 18DAP wild type (WT) and dek35 mutant were generated by deep sequencing, in triplicate, using Illumina Hiseq 2500 (Zea mays).

first: Weiwei

last: Qi


title: Cis-regulatory architecture of a brain-signaling center predates the origin of chordates

city: Old Westbury

first: Ying-Tao

last: Zhao

address: Riland building, Room 024 Northern Boulevard, P.O. Box 8000

department: Biomedical Sciences

institute: New York Institute of Technology

summary: Genomic approaches have predicted hundreds of thousands of tissue specific cis-regulatory sequences, but the determinants critical to their function and evolutionary history are mostly unknown1-4. Here, we systematically decode a set of brain enhancers active in the zona limitans intrathalamica (zli), a signaling center essential for vertebrate forebrain development via the secreted morphogen, Sonic hedgehog (Shh)5,6. We apply a de novo motif analysis tool to identify six position-independent sequence motifs together with their cognate transcription factors that are essential for zli enhancer activity and Shh expression in the mouse embryo. Using knowledge of this regulatory lexicon, we discover novel Shh zli enhancers in mice, and a functionally equivalent element in hemichordates, indicating an ancient origin of the Shh zli regulatory network that predates the chordate phylum. These findings establish a paradigm for delineating functionally conserved enhancers in the absence of overt sequence homologies, and over extensive evolutionary distances.

overall_design: Gene expression profiles from the mouse zona limitans intrathalamica (ZLI) region at E10.5

first: Yao

last: Yao

first: Paul

last: Minor

first: Yongsu

last: Jeong

first: Ariel

last: Pani

first: Anna

middle: N

last: King

first: Orsolya

last: Symmons

first: Lin

last: Gan

first: Wellington

last: Cardoso

first: François

last: Spitz

first: Christopher

last: Lowe

first: Douglas

last: Epstein


title: RISC-mediated control of selected chromatin regulators stabilizes ground state pluripotency of mouse embryonic stem cells (miRNA-Seq)

first: Luca

last: Pandolfini

address: Via Enrico Melen 83, Building B

summary: We performed an integrated analysis of RNA and proteins at the transition between naïve ES cells and cells primed to differentiate. During this transition, mRNAs coding for chromatin regulators were specifically released from translational inhibition mediated by RNA-Induced Silencing Complex (RISC). This suggests that, prior to differentiation, the propensity of ES cells to change their epigenetic status is hampered by RNA interference. The expression of these chromatin regulators was reinstated following acute inactivation of RISC, and it correlated with loss of stemness markers and activation of early cell differentiation markers in treated ES cells.

overall_design: We evaluated global miRNA profiles of embryonic stem cells cultured in either 10%FCS+LIF or 2i+LIF and of Epiblast-Like Aggregates derived from ES cells in 10%FCS+LIF or 2iL+LIF.

first: Ettore

last: Luzi

first: Dario

last: Bressan

first: Nadia

last: Ucciferri

first: Michele

last: Bertacchi

first: Rossella

last: Brandi

first: Silvia

last: Rocchiccioli

first: Mara

last: D'Onofrio

first: Federico

last: Cremisi


title: Organic Nitrogen Source is Critical for Welan Production in Sphingomonas sp. ATCC 31555

city: Wuxi

first: xiaopeng

last: Xu

address: Jiangnan University, Wuxi 214122, Jiangsu, China

department: school of biotechnology

institute: Jiangnan University

summary: Welan gum is mainly produced by Sphingomonas sp. ATCC 31555 and has broad applications in industry such as that in cement production. Both carbon and nitrogen sources are essential for welan production. However, how nitrogen sources affect the metabolism and gene transcription of welan remains elusive. Here, we used next-generation sequencing RNA-seq to analyze the transcriptome of Sphingomonas sp. ATCC 31555 in the presence of inorganic or organic nitrogen sources. Enriched gene expression and pathway analysis suggest that organic nitrogen sources significantly enhanced the expression of genes in central metabolic pathways of Sphingomonas sp. ATCC 31555 and those critical for welan synthesis compared to that observed using inorganic nitrogen sources. The present study improves our understanding of the molecular mechanism underlying the use of nitrogen in welan synthesis in Sphingomonas sp., as well as provides an important transcriptome resource for Sphingomonas sp. in relation to  nitrogen sources.

overall_design: Sphingomonas sp. ATCC 31555 strain (stored in our laboratory) was first seeded in an inoculum medium (20 g/L glucose, 3 g/L yeast extract, 3 g/L malt extract, and 5 g/L fish meal protein peptone, pH 7.0), and then cultured in a fermentation medium containing 40 g/L sucrose, 4.0 g/L nitrogen source, 0.6 g/L KH2PO4, and 0.2 g/L MgSO4.7H2O at 37°C. The nitrogen sources used in the present study were as follows: NaNO3 (4.0 g/L) as inorganic nitrogen (IN), beef extract (4.0 g/L) as organic nitrogen (ON), and NaNO3 (1.5 g/L) + beef extract (2.5 g/L) as complex nitrogen (CN). All cultivations were conducted in flasks with constant rotary shaking at 400–1,000 rpm and 37°C.

first: Xiaopeng

first: Zuoming

last: Nie

first: Zhiyong

last: Zheng

first: Li

last: Zhu

first: Hongtao

last: Zhang

first: Xiaobei

last: Zhan


title: Anchoring of Heterochromatin to the Nuclear Lamina Reinforces Dosage Compensation-Mediated Gene Repression

city: Ann Arbor

first: Gyorgyi

last: Csankovszki

state: MI

address: 830 N. University Ave.

department: MCDB

institute: University of Michigan

summary: Higher order chromosome structure and nuclear architecture can have profound effects on gene regulation. We analyzed how compartmentalizing the genome by tethering heterochromatic regions to the nuclear lamina affects dosage compensation in the nematode C. elegans. In this organism, the dosage compensation complex (DCC) binds both X chromosomes of hermaphrodites to repress transcription two-fold, thus balancing gene expression between XX hermaphrodites and XO males. X chromosome structure is disrupted by mutations in DCC subunits. Using X chromosome paint fluorescence microscopy, we found that X chromosome structure and subnuclear localization are also disrupted when the mechanisms that anchor heterochromatin to the nuclear lamina are defective. Strikingly, the heterochromatic left end of the X chromosome is less affected than the gene-rich middle region, which lacks heterochromatic anchors. These changes in X chromosome structure and subnuclear localization are accompanied by small, but significant levels of derepression of X-linked genes as measured by RNA-seq, without any observable defects in DCC localization and DCC-mediated changes in histone modifications. We propose a model in which heterochromatic tethers on the left arm of the X cooperate with the DCC to compact and peripherally relocate the X chromosomes, contributing to gene repression.

overall_design:
RNA-seq profiles of C. elegans L1 wild type hermaphrodites, cec-4, met-2 set-25, and DPY-27 RNAi.
RNA-seq profiles or C. elegans. Strains are N2 Bristol strain (wild type), RB2301 cec-4(ok3124) IV, and EKM99 met-2(n4256) set-25(n5021) III. Biological replicates for each strain/stage listed separately.

first: Martha

last: Snyder

first: Alyssa

middle: C

last: Lau

first: Elizabeth

middle: A

last: Brouhard

first: Michael

middle: B

last: Davis

first: Jianhao

last: Jiang

first: Margarita

middle: H

last: Sifuentes


title: Hypothalamic transcriptome of male mice on high fat diet, from 99 strains

status: Public on Mar 24 2016

city: Los Angeles

first: Yehudit

last: Hasin-Brumshtein

address: 695 Charles E. Young Drive South

department: David Geffen School of Medicine, Cardiology

institute: UCLA

summary: Previous studies had shown that integration of genome wide expression profiles, in metabolic tissues, with genetic and phenotypic variance, provided valuable insight into the underlying molecular mechanisms.  We used RNA-Seq to characterize hypothalamic transcriptome in 99 inbred strains of mice from the Hybrid Mouse Diversity Panel (HMDP), a reference resource population for cardiovascular and metabolic traits. We report numerous novel transcripts supported by proteomic analyses, as well as novel non coding RNAs. High resolution genetic mapping of transcript levels in HMDP, reveals both local and trans expression Quantitative Trait Loci (eQTLs) demonstrating 2 trans eQTL "hotspots" associated with expression of hundreds of genes. We also report thousands of alternative splicing events regulated by genetic variants. Finally, comparison with about 150 metabolic and cardiovascular traits revealed many highly significant associations. Our data provides a rich resource for understanding the many physiologic functions mediated by the hypothalamus and their genetic regulation.

overall_design: 282 samples, 3 biological replicates per strain

first: Aldons

last: Lusis

first: Desmond

last: Smith


title: cis regulated role of lncRNA in transcriptional or developmental process (ChIRP-Seq)

city: Rochester

first: jinlong

last: lu

address: 213 Hutchinson Hall

institute: University of Rochester

summary: We report that long noncoding RNAs contribute to transcription and developmental process. Thousands of lncRNAs have been identified in the whole genome, and tend to located closely to protein-coding genes. To study position relationship between lncRNA and protein-coding genes, we classified all of lncRNA to several subgroups based on the genome position with their coding neighbors. XH, the head to head subgroup is associated with transcription and development in GO analysis. Here, we knockdown serveral XH lncRNA by shRNA in embryonic stem cells and induce nondirectional differnetiation by removing LIF or neural differnetiation by RA. Knockdown of XH lncRNAs led to uniform downregulation of nearby coding genes, and form regulatory circuits with its nearby coding genes to fine-tune embryonic lineage development. In addition, we also knockout one lncRNA-Evx1as and its nearby protein-coding gene-EVX1 by CRISPR, and get similar results as knockdown.We propose that XH lncRNA may function primarily as 'cis-regulators' of the expression of nearby protein-coding genes, and tend to participate in transcriptional or development regulations as their coding neighbors.

overall_design: All RNA-seq(s) were designed to reveal the differentially expressed genes between wild-type and XH lncRNA knockdown/knockout ESCs during differentiation.

first: Sai

last: Luo

first: Yuyang

last: Lu

first: Xiaohua

last: Shen


title: Identification of plant vacuolar transporters mediating phosphate storage

city: Taipei

first: Tzyy-Jen

last: Chiou

address: No. 128, Sec. 2, Academia Road, Nankang

department: Agricultural Biotechnology Research Center

country: Taiwan

institute: Academia Sinica

summary: Plant vacuoles serve as the primary intracellular compartments for inorganic phosphate (Pi) storage. Passage of Pi across vacuolar membranes plays a critical role in buffering the cytoplasmic Pi level against fluctuations of external Pi and metabolic activities. Here we demonstrate that the SPX-MFS proteins, designated as Phosphate Transporter 5 family (PHT5), also named Vacuolar Phosphate Transporter (VPT), function as vacuolar Pi transporters. Based on 31P-magnetic resonance spectroscopy analysis, Arabidopsis pht5;1 loss-of-function mutants accumulate less Pi and exhibit a lower vacuolar-to-cytoplasmic Pi ratio than controls. Conversely, overexpression of PHT5 leads to massive Pi sequestration into vacuoles and altered regulation of Pi starvation-responsive genes. Furthermore, we show that heterologous expression of OsSPX-MFS1, the rice PHT5 homolog, mediates Pi influx to yeast vacuoles. Our findings uncover a group of Pi transporters in vacuolar membranes that regulate the cytoplasmic Pi homeostasis required for the fitness of plant growth.

overall_design: 10-day seedlings grown on Pi-sufficient medium or followed by 1-day or 3-day Pi starvation, shoot RNA and root RNA were extracted separately

first: Tzu

last: Liu

first: Teng

middle: K

last: Huang

first: Yu

last: Hong

first: Su

middle: F

last: Chiang

first: Tzyy


title: Next Generation Sequencing Facilitates Quantitative Analysis of Transcriptomes for Wild Type and Mutants including opaque2, PbfRNAi and PbfRNAi;o2 in Maize (Zea mays)

city: Piscataway

first: Joachim

last: Messing

state: NJ

address: 190 Frelinghuysen Rd

institute: Waksman Institute of Microbiology, Rutgers University

summary: The goals of this study are to study the regulatory network of the two maize endosperm-specific transcription factors O2 and PBF by 16-DAP endosperm transcriptome profiling (RNA-seq) of their mutants and wild type. The results utilize the expression pattern of global genes regulated by PBF and O2 to elucidate their control for storage compounds synthesis in maize kernels.

overall_design: The 16-DAP endosperm transcriptome of  wild type (WT) and mutants including opaque2, PbfRNAi and PbfRNAi;o2 were generated by RNA-seq with three biological replicates per genotype on Illumina HiSeqTM2500.

first: Zhiyong

last: Zhang

first: Yongrui

last: Wu


title: Bivalency - Histone H3 Lysine4 and Histone H3 Lysine27 Methylation Dynamics

city: Burlington

first: Jonathan

middle: AR

last: Gordon

state: VT

address: 89 Beaumont Ave Given E209

department: Biochemistry

institute: University of Vermont

summary: The molecular signature at histone H3K4me3 and H3K27me3 involved in epigenetic regulation of normal (MCF10A) and transformed (MCF7, MDA-MB-231) breast cells using ChIP-Seq technology.  This study examines the dynamic distribution of H3K4me3, associated with active chromatin, and H3K27me3, associated with repressed chromatin, histone modifications to provide an understanding of the changes in epigenetic regulation associated with the unique breast cancer subtypes.

overall_design:
histone H3K4me3 and H3K27me3 ChIP-seq normal (MCF10A) and transformed (MCF7, MDA-MB-231) breast cells

Please note that the 'H3K4me3' and 'input' data are duplicated records of the samples represented in GSE69377 for the convenient retrieval of the complete raw data from SRA.

first: Terri

last: Messier

first: Joseph

middle: R

last: Boyd

first: Gary

last: Stein


title: Fine-mapping of DNA modifications in the lactase gene region of humans and mice

city: Toronto

first: Arturas

last: Petronis

address: 250 College St Rm606

country: Canada

institute: Centre for Addiction and Mental Health

summary: Bisulfite padlock probe technique was used to examine DNA modifications at the lactase gene region for human and mouse tissues

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: DNA modifications were investigated in human sperm, blood, jejunal enterocytes and jejunum lacking enterocytes, as well as mouse jejunal enterocytes and jejunum lacking enterocytes. For WGA samples, a genome devoid of DNA modifications was used to verify the efficiency of the bisulfite conversion reactions (the tissue sources were human or mouse intestine).

first: Art

first: Viviane

last: Labrie

first: Orion

last: Buske


title: Epigenetics of lactose intolerance and lactase persistence

type: Methylation profiling by genome tiling array

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

overall_design: Refer to individual Series


title: Steady state growth under inorganic carbon limitation increases energy consumption for maintenance and enhances nitrous oxide production in Nitrosomonas europaea

city: Corvallis

first: Luis

last: Sayavedra-Soto

state: OR

address: Cordley Hall 2082

department: Botany and Plant Pathology

institute: Oregon State University

summary: Nitrosomonas europaea is a chemolithoautotrophic bacterium that oxidizes ammonia (NH3) to obtain energy for growth on carbon dioxide (CO2), and can also produce nitrous oxide (N2O), a greenhouse gas. We interrogated the growth, physiological, and transcriptome responses of N. europaea to replete (> 5.2 mM) and limited inorganic carbon (IC) provided by either 1.0 mM or 0.2 mM sodium carbonate (Na2CO3) supplemented with atmospheric CO2. IC-limited cultures oxidized 25 to 58% of available NH3 to nitrite, depending on dilution rate and Na2CO3 concentration. IC limitation resulted in a 1.5-fold increase in cellular maintenance energy requirements compared to NH3-limited cultures. Rates of N2O production increased 2- and 6.3 fold under the two IC-limited conditions increasing the percentage of oxidized NH3-N being transformed to N2O-N from 0.5% (replete) to 4.4% (0.25 mM Na2CO3). Transcriptome analysis showed differential expression (p ≤ 0.05) of 488 genes (20% of inventory) between replete and IC-limited conditions, but few differences were detected between the two IC-limiting treatments. IC-limited conditions resulted in decreased expression of ammonium/ammonia transporter and ammonia monooxygenase subunits, and increased expression of genes involved in C1 metabolism including RuBisCO (cbb gene cluster), carbonic anhydrase, folate-linked metabolism of C1 moieties, and putative C salvage due to oxygenase activity of RuBisCO. Increased expression of nitrite reductase (gene cluster NE0924-0927) correlated with increased production of N2O. Together, these data suggest that N. europaea adapts physiologically during IC-limited steady state growth, which leads to uncoupling of NH3 oxidation from growth and increased N2O production.

overall_design: Transcriptome responses of N. europaea under three inorganic carbon (IC) conditions: IC replete (2 mM Na2CO3, 100 mL min-1 0.2-µm-filtered atmospheric air), 1.0 mM Na2CO3, and 0.2 mM Na2CO3 (both IC-limiting conditions with 50 mL min-1 0.2-µm-filtered atmospheric air) were measured. There were three experimental replicates of each treatment and fold change in mRNA transcripts was calculated in comparison to IC replete.

first: Brett

last: Mellbye

first: Andrew

last: Giguere

first: Frank

last: Chaplen

first: Peter

last: Bottomley


title: FOXE3 Contributes to Peters Anomaly through Transcriptional Regulation of an Autophagy Associated Protein termed DNAJB1

city: Baltimore

first: S. Amer

last: Riazuddin

state: Maryland

address: 600 N. Wolfe St

institute: Johns Hopkins University School of Medicine

summary: FOXE3 is a lens specific transcription factor that has been associated with anterior segment ocular dysgenesis. To determine the transcriptional target(s) of FOXE3 that are indispensable for the anterior segment development, we examined the transcriptome and the proteome of cells expressing truncated FOXE3 responsible for Peters anomaly identified through linkage-coupled next-generation whole exome sequencing. We found that DNAJB1, an autophagy-associated protein, was the only candidate exhibiting differential expression in both screens. We confirmed the candidacy of DNAJB1 through chromatin immunoprecipitation and luciferase assays while knockdown of DNAJB1 in human lens epithelial cells resulted in mitotic arrest. Subsequently, we targeted dnajb1a in zebrafish through injection of a splice-blocking morpholino. The dnajb1a morphants exhibited underdeveloped cataractous lenses with persistent apoptotic nuclei. In conclusion, we have identified DNAJB1 as a transcriptional target of FOXE3 in a novel pathway that is crucial for development of the anterior segment of the eye.

overall_design: Human Embryonic Kidney (HEK293FT) cells were transfected with the expression vector (pT-RexTM-DEST30) harboring either the wild type or the mutant (C240*) FOXE3 ORF (open reading frame). The experimental design included a total of eight biological replicates of cells expressing the wild type and eight replicates of mutant FOXE3 along with eight non-transfected controls. Cells were harvested 24-hour post-transfection and subjected to total RNA isolation for the preparation of whole transcriptome next-generation sequencing libraries. Initially, we examined the quality of transcriptome libraries on a MiSeq genome analyzer. Subsequent to confirmation of the quality, all libraries were paired-end sequenced (2 x 100 bp) using Illumina TruSeq Cluster V3 flow cell at a concentration of 13.0 pM in two separate lanes (12 bar-coded mRNA pooled libraries in each lane) on a HiSeq 2000 genome analyzer.

first: Shahid

last: Khan

first: Shivakumar

last: Vasanth

first: Firoz

last: Kabir

first: John

middle: D

last: Gottsch

first: Arif

middle: O

first: Raghothama

last: Chaerkady

first: Mei-Chong

last: Lee

first: Carmen

last: Leitch

first: Zhiwei

last: Ma

first: Julie

last: Laux

first: Rafael

last: Villasmil

first: Shaheen

first: Sheikh

first: Javed

last: Akram

first: Robert

last: Cole

first: C

last: Talbot Jr

first: Nader

last: Pourmand

first: Norann

last: Zaghloul

first: J

last: Hejtmancik

first: S


title: A Small RNA-Catalytic Argonaute Pathway Tunes Germline Transcript Levels to Ensure Embryonic Divisions

city: Bethesda

first: Shaohe

last: Wang

address: 9000 Rockville Pike, Bldg 30, Rm 403

department: DIR

institute: NIDCR, NIH

summary: Multiple division cycles without growth are a characteristic feature of early embryogenesis. The female germline deposits proteins and RNAs into oocytes to support these divisions, which lack many of the quality control mechanisms operating in somatic cells undergoing growth. How the composition of the oocyte maternal load is regulated to ensure its ability to support early embryogenesis is not known. Here we describe a small RNA-Argonaute pathway, operating in the C. elegans germline, that ensures early embryonic divisions by employing catalytic slicing activity to broadly tune, instead of silence, germline gene expression. Misregulation of one target, a kinesin-13 microtubule depolymerase, underlies a major embryonic phenotype associated with pathway loss. Tuning of target expression is guided by small RNA density, which must ultimately be related to target sequence. Thus, C. elegans employs a single catalytic Argonaute for small RNA-mediated tuning of the mRNA levels of germline-expressed genes that support early embryogenesis.

overall_design: mRNA profiling of 2 replicates each for 3 genotypes of adult-stage C. elegans worms

first: Adina

last: Gerson-Gurwitz

first: Shashank

last: Sathe

first: Rebecca

last: Green

first: Gene

middle: W

last: Yeo

first: Karen

last: Oegema

first: Arshad

last: Desai


title: Identification of MOL1-dependent genes II

city: Heidelberg

first: Thomas

last: Greb

address: Im Neuenheimer Feld 360

department: Centre for Organismal Studies

country: Germany

institute: Heidelberg University

summary: We used RNA-seq to generate the transcriptional profile of WT and mol1-1 mutants in the second internode of the infloresence stem of 15 to 20 cm tall Arabidopsis. We then used stastical analysis to identify differentially expressed genes that potentially contribute to the mol1-1 mutant phenotype.

overall_design: RNA from three replicates each of WT and mol1-1 mutants were sequenced and differentially expressed genes were identified.

first: Nial

last: Gursanscky

first: Virginie

last: Jouannet


title: Next-generation sequencing analyses of the differently expressed genes of wild type and atcep1 mutant Arabidopsis thaliana

city: Beijing

first: dan

middle: dan

last: Zhang

address: No.35 Qinghua East Road, Haidian Distract

department: College of Biological Sciences and Biotechnology

institute: Beijing Forestry University

summary:
Purpose: The goal of this study is to compare the differently expressed genes in the wild type and the KDEL-tailed cysteine protease AtCEP1 knockout (atcep1) Arabidopsis using RNA-sequencing (RNA-seq).

Methods: Arabidopsis buds mRNA profiles of anther development stages 5-6, 7-9, and 10-11 of the wild type (WT) and atcep1 mutant were generated by deep sequencing via Illumina HiSeqTM 2000. The sequence reads that passed quality filters were analyzed at the transcript isoform level with the following steps: Remove reads with adaptor sequences; Remove reads in which the percentage of unknown bases (N) is greater than 10%; Remove low quality reads, in which the percentage of the low quality base (base with quality value ≤ 5) is greater than 50%. The clean reads were mapped to the Arabidopsis reference genome and reference genes using SOAP aligner/SOAP2. No more than 2 mismatches were allowed in the alignment. The gene expression level was calculated using RPKM (Reads Per Kb per Million reads). Differential expression analysis between the wild type and the atcep1 mutant was performed using the DEGseq R package (1.12.0) based on normalized read counts. A corrected P value of < 0.005 and |log2Ratio| > 1 were set as the threshold for significantly differential expression.

Results: We identified 872 genes showing significant differential expression, and in the atcep1 mutant, the upregulated genes significantly outnumbered the downregulated genes at the three time points. The GO analysis of the differently expressed genes showed that the expression of genes participating in anther tapetal secretory structure formation, pollen wall development, and tapetal cell wall generation, clearly changed.

overall_design: Arabidopsis buds mRNA profiles of development stages 5-6, 7-9, and 10-11 of the wild type (WT) and atcep1 muant were generated by deep sequencing via Illumina HiSeqTM 2000.

first: Hai

last: Lu

first: Dandan


title: C9orf72 is required for proper macrophage and microglial function in mice

first: Jacqueline

middle: Gire

last: O'Rourke

address: 8700 Beverly Blvd

institute: Cedars Sinai Medical Center

summary: Expansions of a hexanucleotide repeat (GGGGCC) in the noncoding region of the C9orf72 gene are the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia. Decreased expression of C9orf72 is seen in expansion carriers, suggesting loss of function may play a role in disease. We find that two independent mouse lines lacking the C9orf72 ortholog (3110043O21Rik) in all tissues developed normally and aged without motor neuron disease. Instead, C9orf72 null mice developed progressive splenomegaly and lymphadenopathy with accumulation of engorged macrophage-like cells. C9orf72 expression was highest in myeloid cells, and loss of C9orf72 led to lysosomal accumulation and altered immune responses in macrophages and microglia, with age-related neuroinflammation similar to C9orf72 ALS but not sporadic ALS patient tissue. Thus, C9orf72 is required for normal function of myeloid cells, and altered microglial function may contribute to neurodegeneration in C9orf72 expansion carriers.

overall_design: To compare the RNA Seq profiles from the lumbar region of spinal cords from mice  lacking one copy or both copies of the C9orf72 ortholog (3110043O21Rik) compared to wild type control with two copies at 3 months (n=3) and 17 months (n=4).

first: Robert

last: Baloh


title: SON and its alternatively spliced isoforms control MLL complex-mediated H3K4me3 and transcription of leukemia-associated genes (ChIP-seq data)

city: BALTIMORE

first: Linda

last: Resar

address: 720 Rutland Avenue

department: Department of Medicine / Division of Hematology

institute: Johns Hopkins University

summary: Dysregulation of MLL complex-mediated histone methylation plays a pivotal role in gene expression associated with diseases, but little is known about cellular factors modulating MLL complex activity.  Here, we report that SON, previously known as an RNA splicing factor, controls MLL complex-mediated transcriptional initiation.  SON binds to DNA near transcription start sites, interacts with menin, and inhibits MLL complex assembly, resulting in decreased H3K4me3 and transcriptional repression. Importantly, alternatively spliced short isoforms of SON are markedly upregulated in acute myeloid leukemia. The short isoforms compete with full-length SON for chromatin occupancy, but lack the menin-binding ability, thereby antagonizing full-length SON function in transcriptional repression while not impairing full-length SON-mediated RNA splicing. Furthermore, overexpression of a short isoform of SON enhances replating potential of hematopoietic progenitors. Our findings define SON as a fine-tuner of the MLL-menin interaction and reveal short SON overexpression as a marker indicating aberrant transcriptional initiation in leukemia.

overall_design: We performed chromatin immunoprecipitation and sequencing (ChIP-seq) in K562 leukemia cells with two different SON antibodies (SON-N and SON-C Abs)

first: Eun-Young Erin

last: Ahn


title: cis regulated role of lncRNA in transcriptional or developmental process

first: jinlong

last: lu


title: Splicing of nascent RNA coincides with intron exit from RNA polymerase II

city: Berlin

first: Lydia

last: Herzel

address: Takustr. 6

department: Biology, Chemistry and Pharmacy

institute: Freie Universität

sample_organism:
Schizosaccharomyces pombe
Saccharomyces cerevisiae


title: Targeted paired-end sequencing of nascent RNA from S. cerevisiae

summary:
Targeted paired-end sequencing of cDNA from unfragmented nascent RNA from exponentially growing S. cerevisiae cells was employed to obtain Pol II transcription elongation and splicing information from single transcripts.
Nascent RNA was prepared from the yeast chromatin fraction (Carrillo Oesterreich, Preibisch, Neugebauer, Mol Cell 2010) or enriched from total RNA with polyadenylated RNA depletion. The nascent 3’ end was labeled with a 3’ DNA adaptor through ligation.
A PCR with a forward primer in the first exon of select intron-containing genes amplifies nascent transcripts of specific genes and ensures sequencing adaptor attachment for paired-end sequencing.
With this approach co-transcriptional splicing progression with distance from the intron end could be analyzed for 87 genes. Note that the unmapped and mapped data also include genes that did not pass the read coverage requirements in SMIT analysis.

overall_design: Nascent RNA profiles for mainly intron-containing genes were generated with paired-end sequencing with Illumina HiSeq technology.

first: Fernando

last: Carrillo Oesterreich

first: Korinna

last: Straube

first: Karla

middle: M

last: Neugebauer


title: Primary cilium-autophagy-Nrf2 (PAN) axis links the cell cycle to neuroectoderm fate in human embryonic stem cells

city: Santa Barbara

first: Kenneth

middle: S.

last: Kosik

address: UC Santa Barbara

department: NRI

institute: UCSB

summary: Under defined differentiation conditions human embryonic stem cells (hESCs) can be directed toward a mesendodermal (ME) or neuroectoderm (NE) fate, the first decision during hESC differentiation. Coupled with G1 lengthening a divergent ciliation pattern emerged within the first 24 hours of induced lineage specification and these changes heralded a neuroectoderm decision before any neural precursor markers were expressed. By day 2, increased ciliation in NE precursors induced autophagy that resulted in the inactivation of Nrf2. Nrf2 binds directly to upstream regions of the OCT4 and NANOG genes to promote their expression and represses NE derivation. Nrf2 suppression was sufficient to rescue poorly neurogenic iPSC lines. Only after these events have been initiated do neural precursor markers get expressed at day 4. Thus we have identified a primary cilium-autophagy-Nrf2 (PAN) axis coupled to cell cycle progression that directs hESCs toward NE.

overall_design: Transcriptome analysis of hESC-derived neuroectoderm and mesendoderm cells

first: Jiwon

last: Jang

first: Yidi

last: Wang

first: Matthew

last: Lalli

first: Elmer

last: Guzman

first: Sirie

middle: E

last: Godshalk

first: Hongjun

last: Zhou

middle: S


title: A Tunable Mechanism Determines the Transgenerational Duration of Small RNA Inheritance in C.elegans

city: Tel-Aviv

first: Leah

last: Houri-Ze'evi

address: Haim Levanon

department: Neurobiology

country: Israel

institute: Tel-Aviv University

summary: In C.elegans nematodes small RNAs enable transmission of epigenetic responses across multiple generations. While RNA interference (RNAi) inheritance mechanisms that enable “memorization” of ancestral responses are being elucidated, it is not known why or how, after a few generations, epigenetic effects are “forgotten”. We show that exposure to dsRNA activates a feedback loop that determines the duration of inherited silencing. We find that gene-specific RNAi responses dictate the transgenerational duration of RNAi responses mounted against unrelated genes, elicited separately in previous generations. RNA-seq analysis reveals that aside from silencing of genes with complementary sequences, dsRNA-induced RNAi affects the production of heritable endogenous small RNAs, which regulate the expression of RNAi factors. Manipulating genes in this feedback pathway changes the duration of heritable silencing. Active control of transgenerational effects could be adaptive, since ancestral responses would be detrimental if the environments of the progeny and the ancestors would differ.

overall_design:
Examination of dsRNA induced transgenerational inheritance in lineages with and without second trigger enhancment
Small-RNA sequencing of P0-F3 generations of [1] worms that were exposed to anti-gfp dsRNA at the P0 generation and transferred to control bacteria ("empty vector") at the next generations (F1-F3) [2] worms that were exposed to anti-gfp dsRNA at the P0 generation, transferred to an additional anti-mCherry RNAi plates at the F1 generation ("2nd trigger") and moved to control bacteria at the F2 generations for two generations (F2-F3) and [3] control group of worms that were grown on "empty-vector" plates and were not exposed to RNAi at any stage.

first: Oded

last: Rechavi


title: Long read sequencing of nascent RNA from S. cerevisiae and S. pombe

summary:
Long read SMRT cDNA sequencing of nascent RNA from exponentially growing S. cerevisiae and S. pombe cells was employed to obtain transcription elongation and splicing information from single transcripts.
Nascent RNA was prepared from the yeast chromatin fraction (Carrillo Oesterreich, Preibisch, Neugebauer, Mol Cell 2010).
The nascent 3’ end was labeled with a 3’ DNA adaptor through ligation.
The adaptor sequence served as template for full-length reverse transcription and double-stranded cDNA was obtained in a PCR (gene-specific or transcriptome-wide). SMRT DNA sequencing libraries were prepared subsequently.

platform_organism:
Schizosaccharomyces pombe
Saccharomyces cerevisiae

overall_design: Nascent RNA profiles for mainly intron-containing genes were generated with long-read SMRT cDNA sequencing.


title: Atad2 is a generalist facilitator of chromatin dynamics in embryonic stem cells [Atad2_ES_ChIPseq2_MG2]

city: La Tronche

first: Sophie

last: Rousseaux

address: Bd de la Chantourne

department: IAB

country: France

institute: INSERM-UGA

summary: Although the conserved AAA ATPase – bromodomain factor, ATAD2, has been described as a transcriptional co-activator upregulated in many cancers, its function remains poorly understood. Here, using a combination of ChIP-seq, ChIP-proteomics and RNA-seq experiments in embryonic stem cells, we found that Atad2 is an abundant nucleosome-bound protein present on active genes, associated with chromatin remodelling, DNA replication and DNA repair factors. A structural analysis of its bromodomain and subsequent investigations demonstrate that histone acetylation guides ATAD2 to chromatin, resulting in an overall increase of chromatin accessibility and histone dynamics, which is required for the proper activity of the highly expressed gene fraction of the genome. While in exponentially growing cells Atad2 appears dispensable for cell growth, in differentiating ES cells, Atad2 becomes critical in sustaining specific gene expression programs, controlling proliferation and differentiation. Altogether, this work defines Atad2’s function as a facilitator of general chromatin-templated activities such as transcription.

overall_design: Chromatin immunoprecipitation of Atad2-TAP tag following tandem affinity purification (TAP) protocol in 46C ES wild type cells (control) and in Atad2-TAP tag expressing ES cells. Cross-linked sheared chromatin fragments underwent two rounds of ChIP. After the first ChIP, using an anti-Flag antibody, Atad2-bound fragments were released with an excess of Flag peptides and were then used for a second round of ChIP using an anti-Ha antibody.

first: Alexandra

last: Debernardi

first: Yuichi

last: Morozumi

first: Faycal

last: Boussouar

first: Philippe

last: Guardiola

first: Matthieu

last: Gerard

first: Davide

last: Gianni

first: Norbert

last: Schweifer


title: Atad2 is a generalist facilitator of chromatin dynamics in embryonic stem cells [Atad2_ES_RNAseq_BI]

overall_design: We used a siRNA approach to knock-down Atad2 and measure the resulting variations in gene expression by RNA-seq


title: Transitive silencing of mobile small RNAs in Arabidopsis Pollen

city: St. Louis

first: R. Keith

last: Slotkin

state: Missouri

address: 975 North Warson Road

institute: Donald Danforth Plant Science Center

summary:
To assay siRNA movement in the pollen grain, we took advantage of the known behavior of 22nt siRNAs to target transcripts for cleavage and initiate secondary siRNA biogenesis. Previous research has demonstrated that when targeted by a 22nt microRNA, a transcript containing a truncated non-fluorescent ~400 bp version of GFP (trGFP) will produce GFP secondary siRNAs that can target a second copy of a functional full-length GFP mRNA in trans. Therefore, we created a ~400 bp non-fluorescent version of trGFP with a 5’ 22nt microRNA target site driven by the vegetative cell-specific KRP6 promoter and transformed this construct into a line homozygous for the sperm-specific functional full-length GFP driven by the MGH3 promoter. If cleaved by a 22nt microRNA, the vegetative cell-driven trGFP will be cleaved into secondary siRNAs, and if the RNA components of this system are mobile and transferred into the sperm cells, the sperm-specific fluorescence of GFP will be reduced.
We used two versions of the microRNA target site, which included a mock target site that is not targeted by small RNAs, the target site of microRNA173 (which is a 22nt known to induce the production of secondary siRNAs). We demonstrate that when the 22nt microRNA173 target site is used in the vegetative cell transcript, a corresponding decrease in sperm cell fluorescence is observed. In the mock small RNA target site this reduction in fluorescence was not observed.
To ensure the transitive silencing sperm cell transcripts is occurring via secondary small RNAs, we transferred this trGFP experiment into a dcl2/dcl4 mutant background that is defective in TE 21-22nt siRNA silencing. In dcl2/dcl4 double mutants with the trGFP system, we find that the GFP fluorescence of the mock target site transgene does not change, while the repression found in wild-type pollen with the 22nt microRNA173 target site is alleviated. The data for microRNA173 is particularly important, as DCL2 and DCL4 are not necessary for microRNA173 production, and therefore this demonstrates that specifically secondary siRNAs acting downstream of microRNA action are required to silence the sperm cell GFP. We further analyzed the production of sRNAs derived from GFP in pollen grains of transgenic lines expressing the 22nt miR173 or 22nt mock target sites inserted on the 5’ region of the trGFP transgene in both wt and dcl2/dcl4 backgrounds.

overall_design: Four small RNA sequencing libraries. There are two genotypes: wt Col and a dcl2/dcl4 double mutant. Each genotype has one of two truncated-GFP transgenes: one with a 22nt microRNA173 target site and one with 22nt 'mock' target site not targeted by small RNAs.

first: R

first: Germán

last: Martínez-Arias

first: Kaushik

last: Panda


title: Atad2 is a generalist facilitator of chromatin dynamics in embryonic stem cells


title: Identification of MOL1-dependent genes

first: Thomas

last: Greb

summary: We used RNA-seq to generate the transcriptional profile of WT and mol1-1 mutants in a 1cm fragment above the base of the infloresence stem of 15 to 20 cm tall Arabidopsis. We then used stastical analysis to identify differentially expressed genes that potentially contribute to the mol1-1 mutant phenotype.

overall_design: RNA from two replicates each of WT and mol1-1 mutants were sequenced and differentially expressed genes were identified.

first: Nial

last: Gursanscky

first: Virginie

last: Jouannet


title: Evx1as and EVX1 knockouts by CRISPR

first: jinlong

last: lu

address: 213 Hutchinson Hall

institute: University of Rochester

summary: We report that long noncoding RNAs contribute to transcription and developmental process. Thousands of lncRNAs have been identified in the whole genome, and tend to located closely to protein-coding genes. To study position relationship between lncRNA and protein-coding genes, we classified all of lncRNA to several subgroups based on the genome position with their coding neighbors. XH, the head to head subgroup is associated with transcription and development in GO analysis. Here, we knockdown serveral XH lncRNA by shRNA in embryonic stem cells and induce nondirectional differnetiation by removing LIF or neural differnetiation by RA. Knockdown of XH lncRNAs led to uniform downregulation of nearby coding genes, and form regulatory circuits with its nearby coding genes to fine-tune embryonic lineage development. In addition, we also knockout one lncRNA-Evx1as and its nearby protein-coding gene-EVX1 by CRISPR, and get similar results as knockdown.We propose that XH lncRNA may function primarily as 'cis-regulators' of the expression of nearby protein-coding genes, and tend to participate in transcriptional or development regulations as their coding neighbors.

overall_design: All RNA-seq(s) were designed to reveal the differentially expressed genes between wild-type and XH lncRNA knockdown/knockout ESCs during differentiation.

first: Sai

last: Luo

first: Jinlong

last: Lu

first: Xiaohua

last: Shen


title: Sequence Features Accurately Predict Genome-wide MeCP2 Binding in vivo

city: Urbana

first: Hans

middle: Tomas

last: Rube

state: Illinois

address: 1110 West Green Street

department: Physics

institute: University of Illinois at Urbana Champaign

summary: MECP2 is critical for proper brain development and expressed at near-histone levels in neurons, but the mechanism of its genomic localization remains poorly understood. Using high-resolution MeCP2 binding data, we show that genetic features alone can predict binding with 88% accuracy. Integrating MeCP2 binding and DNA methylation in a probabilistic graphical model, we demonstrate that previously reported methylation preferences may be due to MeCP2’s affinity to GC-rich chromatin, a result replicated using published data. Furthermore, MeCP2 co-localized with nucleosomes, and Mecp2 deletion led to nucleosome repositioning. Finally, MeCP2 binding downstream of promoters correlated with increased expression in Mecp2 deficient neurons.

overall_design: Study of genetic and epigenetic determinants of MeCP2 binding using MeCP2 ChIP-seq, MNase-seq, Bisulfite-seq and  RNA-seq. Please see individual  sample record  for details on experimental design.

first: H

middle: T


title: Atad2 is a generalist facilitator of chromatin dynamics in embryonic stem cells [Atad2_ES_Chipseq1_BI]

first: Sophie

last: Rousseaux

overall_design: Native ChIP protocol with a single anti-Ha immunoprecipitation in 46C ES wild type cells (control) and in Atad2-TAP tag expressing ES cells

first: Alexandra

last: Debernardi

first: Yuichi

last: Morozumi

first: Faycal

first: Philippe


title: Pretreatment microRNA expression impacting on epithelial to mesenchymal transition predicts intrinsic radiosensitivity in head and neck cancer cell lines and patients [miRNA-seq]

status: Public on Mar 23 2016

city: Amsterdam

first: Monique

middle: C.

last: de Jong

address: Plesmanlaan 121

department: Dept of Biological Stress Response and Dept. of Radiation Oncology

country: Netherlands

institute: Netherlands Cancer Institute

summary:
Purpose: Predominant causes of head and neck cancer recurrence after radiotherapy are rapid repopulation, hypoxia, fraction of cancer stem cells and intrinsic radioresistance. Currently, intrinsic radioresistance can only be assessed by ex-vivo colony assays. Besides being time-consuming, colony assays do not identify causes of intrinsic resistance. We aimed to identify a biomarker for intrinsic radioresistance to be used before start of treatment and to reveal biological processes that could be targeted to overcome intrinsic resistance.
Experimental design: We analyzed both micro- and messenger RNA expression in a large panel of HNSCC cell lines. Expression was measured on both irradiated and unirradiated samples. Results were validated using modified cell lines and a series of laryngeal cancer patients.
Results: MiRs, mRNAs and gene sets that correlated with resistance could be identified from expression data of unirradiated cells. The presence of epithelial to mesenchymal transition (EMT) and low expression of miRs involved in the inhibition of EMT were important radioresistance determinants. This finding was validated in two independent cell line pairs, in which the induction of EMT reduced radiosensitivity. Moreover, low expression of the most important miR (miR-203) was shown to correlate with local disease recurrence after radiotherapy in a series of laryngeal cancer patients.
Conclusions: These findings indicate that EMT and low expression of EMT-inhibiting miRs, especially miR-203, measured in pre-treatment material, causes intrinsic radioresistance of HNSCC, which could enable identification and treatment modification of radioresistant tumors.

overall_design: Matched pairs of pre-treatment biopsies of 34 patients treated at The Netherlands Cancer Institute between 2002 and 2010. Patients with T2-3 laryngeal cancers, all treated with radiotherapy alone with a curative intent. The series was designed to be a matched cohort of 17 patients with local recurrences matched with 17 local cures. There were no significant differences between groups with and without local recurrence in age, gender, subsite, T-stage or treatment year.

middle: C

first: Jelle

middle: J

last: ten Hoeve

first: Reidar

last: Grénman

first: Lodewyk

last: Wessels

first: Ron

last: Kerkhoven

first: Hein

last: te Riele

first: Michiel

last: van den Brekel

first: Marcel

last: Verheij

first: Adrian

last: Begg


title: ToxR Antagonizes H-NS Regulation of Horizontally Acquired Genes to Drive Host Colonization.

city: Austin

first: Bryan

last: Davies

state: TX

address: 2506 Speedway

institute: University of Texas at Austin

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

overall_design: Refer to individual Series


title: Single cell global gene profiling reveals molecular and functional platelet bias of aged hematopoietic stem cells

city: Oxford

first: Claus

last: Nerlov

address: John Radcliffe Hospital, University of Oxford

department: MRC Molecular Haematology Unit

country: United Kingdom

institute: Weatheral Institute of Molecular Medicine

summary: Single cell whole transcriptome analysis of  young (2-3 months) and old (20-25 months) mouse HSCs, defined as Lin–Sca-1+c-Kit+150+CD48– .

overall_design: Differential gene expression analysis of young and old mouse HSCs (Lin–Sca-1+c-Kit+150+CD48– )

first: A

last: Grover

first: S

last: Thongjuea

last: Giustacchini

first: Sten

last: Jacobsen


title: Genomic deletion by CRISPR/Cas9 reveals a critical role for Evx1as/Evx1 locus in regulating embryonic stem cell differentiation

city: Rochester

first: jinlong

last: lu

state: NY

summary: We report that head to head long noncoding RNAs contribute to transcription and developmental process. Thousands of lncRNAs have been identified in the whole genome, and classified to several subgroups based on the genome position with their coding neighbors. XH, the head to head subgroup is associated with transcription and development in GO analysis. Here, we deleted the promoter and exon1 region which shared by Evx1as/Evx1 XH pair in embryonic stem cells by two sgRNAs-mediated CRISPR. Two homozygous mutants came out from 46 single clones with high efficiency, while two control clones were picked from cells which were only Cas9 transfected. These two mutants failed to activate Evx1 in -LIF differentiation compared to control cells, while Evx1as still have 30% mRNA residual. Meanwhile, we characterized gene expression level among different stages of ESC differentiation and found many differentiated related makers exhibited aberrant regulation, especially endodermal genes. We propose that XH lncRNA and its coding neighbor could regulate each other, and function as a unit to promote ESC differentiation and biological development.

overall_design: All RNA-seq(s) were designed to reveal the differentially expressed genes between wild-type or Evx1as/Evx1 promoter and exon1 KO cells during -LIF differentiation.

first: Sai

last: Luo

first: Jinlong

last: Lu

first: Xiaohua

last: Shen


title: Characterization of the TcpP Regulon Vibrio cholerae El Tor

summary: The transcriptional factor ToxR initiates a virulence regulatory cascade required for V. cholerae to express critical host colonization factors and cause disease. Genome-wide expression studies suggest that ToxR regulates many genes important for V. cholerae pathogenesis, yet our knowledge of the direct regulon controlled by ToxR is limited to just four genes. Here, we determine ToxR’s genome-wide DNA-binding profile and show that ToxR is a global regulator of both progenitor genome-encoded genes and horizontally acquired islands encoding the majority of V. cholerae’s major virulence factors. Our results suggest that ToxR has gained regulatory control over important acquired elements that not only drive V. cholerae pathogenesis but that also define the major transitions of V. cholerae pandemic lineages. We demonstrate that ToxR shares nearly half its regulon with the histone-like nucleoid structuring protein H-NS, and antagonizes H-NS for control of critical colonization functions. This regulatory interaction is the major role of ToxR in V. cholerae colonization since deletion of H-NS abrogates the need of ToxR in V. cholerae host colonization. By comparing the genome-wide binding profiles of ToxR and other critical virulence regulators, we show that despite similar predicted DNA binding requirements, ToxR is unique in its global control of progenitor-encoded and acquired genes. Our results suggest that, like H-NS, factors in addition to linear DNA sequence drive selection of ToxR binding sites.

overall_design: We used ChIP-seq to identify TcpP binding sites across the genome to determine the direct regulon of TcpP

first: Misha

middle: I

last: Kazi


title: Characterization of the HNS Regulon Vibrio cholerae El Tor

overall_design: We used ChIP-seq to identify HNS binding sites across the genome to determine the direct regulon of HNS