title: Genome-wide discovery of active regulatory elements and transcription factor footprints in Caenorhabditis elegans using DNase-seq

city: Pasadena

first: Paul

last: Sternberg

address: 1200 E California Blvd

institute: California Institute of Technology

summary: Deep sequencing of size-selected DNaseI-treated chromatin (DNase-seq) allows high resolution measurement of chromatin accessibility to DNaseI cleavage, permitting identification of de novo active cis regulatory modules (CRMs) and individual transcription factor (TF) binding sites. We adapted DNase-seq to nuclei isolated from C. elegans embryos and L1 arrest larvae to generate high-resolution maps of TF binding. Over half of embryonic DNaseI hypersensitive sites (DHS) were annotated in noncoding sequences, with 23% in intergenic, 11% promoter regions and 21% in introns, with similar statistics in data collected from L1 arrest larvae. Noncoding DHS exhibit high evolutionary sequence conservation and are enriched in marks of enhancer activity and transcription. We validated noncoding DHS against a previously investigated set of enhancers from myo-2, myo-3, hlh-1, elt-2 and lin-26/lir-1 gene loci and recapitulated 15 of 17 known enhancers in these loci. We then mined the DNase-seq data to identify putative active CRMs and TF footprints. Our DNase-seq data could also be used to improve predictions of tissue-specific expression compared to motifs alone. In a pilot functional test, 10 of 15 DHS from pha-4, icl-1 and ceh-13 drove reporter gene expression in transgenic C. elegans. Overall, we provide experimental annotation of 26,644 putative CRMs in the embryo containing 55,890 TF footprints, and 15,841 putative CRMs in the L1 arrest larvae containing 32,685 TF footprints.

overall_design: Embryo and L1 arrest larvae nuclei were treated with 0, 20, 40, 80, 120, 160 U/mL DNaseI in 1X DNaseI digestion buffer (containing CaCl2, spermine, spermidine, protease inhibitor) each for 3 minutes at 37°C. DNaseI treatment follows the conditions from the Stamatoyannopoulos lab protocol (Thurman et al. 2012). DNaseI treatment was quenched with STOP buffer containing 20mg/mL Proteinase K and incubated 55°C overnight. After treating with 45ug/mL boiled RNase A for 30 minutes, DNA was purified and concentrated using column purification. The DNA sample was run on 1% agarose, stained with SYBR Gold, and the gel piece containing DNA fragments less than 500bp was isolated and purified. For each nuclei sample, we compared QPCR data using primers designed against positive controls (known enhancers from the lin-39/ceh-13 Hox complex studied in Kuntz et al. 2008) with negative controls (sequences from the same study that did not drive reporter gene expression) in order to measure the regulatory enrichment of CRMs in the DNase-treated nuclei samples. QPCR primers were designed against conserved MUSSA regions of “true positive” N1, N2, N3, N4, N7, N8, N9, N11 lin-39/ceh-13 enhancers and N5 and N6 negative control non-enhancer regions studied by Kuntz et al. (2008). QPCRs were performed with genomic DNA standards and absolute derivative measurement of Cp. Relative fold enrichment was compared within samples by normalizing measured concentration of each region vs. mean of negative controls (N5 and N6) (Figure S4). For each nuclear sample we chose the digested sample from the DNaseI treatment level that yielded the highest regulatory enrichment (typically between 2 and 5 fold) and used it to prepare libraries for sequencing. The libraries were multiplexed sequenced on Illumina HiSeq to yield 50bp single end reads. Raw DNaseI hypersensitive peaks were identified by detecting read enrichment in 150bp consecutive nucleotides using HOTSPOT peak caller specifically designed for DNase-seq (version 3; John et al. 2011). We filtered raw peak calls obtained from HOTSPOT using the irreproducibility discovery rate (IDR) framework developed for ENCODE, which uses a non-parametric copula mixture model to filter peaks into reproducible or irreproducible categories (Li et al. 2011; Landt et al. 2012). Peaks are filtered on the combination of their rank or score as well as their consistency across replicates at an IDR level of 0.1. Peaks overlapping Repeatmasker repeats were omitted. Blacklist regions from ENCODE representing known ce10 genomic regions exhibiting signal artifacts in ChIP-seq experiments were removed (ENCODE Project Consortium 2012). Overlapping peaks were merged to yield 41,825 and 23,670 DHS peaks across embryo and L1 arrest biological replicates, respectively. DHS peak locations were annotated in exons (if 75% of region was located in exon), introns, promoter (<300bp from ATG), and intergenic regions (>300bp from ATG) using custom scripts and WormBase WS241 gene models. Pseudogenes, tRNAs, and ncRNAs were excluded from annotation. Footprints were identified with DNase2TF software package using a FDR threshold of 0.05 (Sung et al. 2014) to identify decreased read coverage within noncoding DHS in regions between 6-40bp with a strand shift in reads in each biological replicate. Replicate data within each stage were merged and used to identify additional TF footprints.

first: Margaret

last: Ho


title: Genome-wide maps of FoxP3 binding in FoxP3WT and FoxP3A384T Treg cells

status: Public on Aug 01 2017

city: Yokohama

first: Shohei

last: Hori

state: Kanagawa

address: 1-7-22 Suehiro-cho, Tsurumi-ku

country: Japan

institute: RIKEN Center for Integrative Medical Sciences

type: Genome binding/occupancy profiling by high throughput sequencing

summary: FoxP3 binding sites in Treg cells expressing WT or A384T mutant FoxP3 were examined.

overall_design: Chromatin was prepared from FoxP3+ Treg cells from FoxP3WT and FoxP3A384T mice, immunoprecipitated with anti-FoxP3 antibody, and the bound DNA were subjected to deep sequencing.

first: Norihito

last: Hayatsu


title: RNA binding protein KSRP mediated miRNAs processing reciprocally regulates granulocytic and monocytic differentiation

city: Beijing

first: Xu

middle: jia

last: yue

address: NO.3 Dongdan street, Beijing

country: China

institute: Peking Union Medical College

type: Non-coding RNA profiling by high throughput sequencing

summary: KSRP is involved in RNA processing at multiple levels including alternative splicing, mRNA stability, mRNA localization and translation efficiency. Dysregulation KSRP can cause to hematological malignancies. However, the direct contribution of KSRP to myeloid lineage differentiation is poorly understood. KSRP, as a KH-type splicing regulatory protein, which interacts with single-stand AU-rich element containing mRNA and mediated mRNA decay, also serves as a component of Drosha and Dicer complexes and regulates the biogenesis of miRNAs. Whether KSRP mediated miRNAs maturation participated in myeloid lineage differentiation is unknown. Here, we analysis the miRNAs processing during over-expression KSRP in THP-1 cells by poly-A(riched) RNA-seq and miRNA-Seq. Our results suggest that KSRP regulate a class of miRNAs maturation and have a critical function in myeloid lineage differentiation.

overall_design: The design was meant to identify differential miRNAs processing which regulated by KSRP in THP-1 cells.

first: Hongmei

last: Zhao

first: Fang

last: Wang


title: Analyses of a mutant FoxP3 allele reveal BATF as a critical transcription factor in the differentiation and accumulation of tissue regulatory T cells

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

overall_design: Refer to individual Series


title: Induction of alternative proinflammatory cytokines accounts for sustained psoriasiform skin inflammation in IL-17C+IL-6KO mice

city: Ann Arbor

first: William

middle: R

last: Swindell

state: MI

address: 1500 E. Medical Center Drive

department: Dermatology

country: USA

institute: University of Michigan

type: Expression profiling by high throughput sequencing


title: Induction of alternative proinflammatory cytokines accounts for sustained psoriasiform skin inflammation in IL-17C+IL-6KO mice [RNA-Seq]

summary: IL-6 inhibition has been unsuccessful in treating psoriasis, despite high levels of tissue and serum IL-6 in patients. Additionally, de novo psoriasis onset has been reported following IL-6 blockade in rheumatoid arthritis patients. To explore mechanisms underlying these clinical observations, we backcrossed an established psoriasiform mouse model (IL-17C+ mice) with IL-6 deficient mice (IL-17C+KO) and examined the cutaneous phenotype.  IL-17C+KO mice initially exhibited decreased skin inflammation, however this decrease was transient and reversed rapidly, concomitant with increases in skin Tnf, Il36α/β/γ, Il24, epigen and S100a8/a9 to levels higher than those found in IL-17C+ mice. Comparison of IL-17C+ and IL-17C+KO mouse skin transcriptomes with that of human psoriasis skin, revealed significant correlation among transcripts of psoriasis patient skin and IL-17C+KO mouse skin, and confirmed an exacerbation of the inflammatory signature in IL-17C+KO mice that aligns closely with human psoriasis. Transcriptional analyses of IL-17C+ and IL-17C+KO primary keratinocytes confirmed increased expression of proinflammatory molecules, suggesting that in the absence of IL-6, keratinocytes increase production of numerous additional proinflammatory cytokines. These preclinical findings may provide insight into why arthritis patients being treated with IL-6 inhibitors develop new onset psoriasis and why IL-6 blockade for the treatment of psoriasis has not been clinically effective.

overall_design: Skin biopsies were obtained from IL-17C+ mice (n = 3) and IL-17C+KO mice (n = 2). Paired lesional and uninvolved skin biopsies were obtained from each mouse (n = 10 biopsies total from 5 mice). Comparisons were made between lesional and uninvolved skin from IL-17C+ and IL-17C+KO mice.

first: Nicole

middle: L

last: Ward

first: Philip

middle: A

last: Klenotic

first: Sarah

middle: G

last: Groft

first: Thomas

middle: S

last: McCormick

first: Patrick

last: Leahy


title: RNA profiling to describe the transcriptional change during white fat browning, brown fat activity and inactivation

city: Singapore

first: Xiaoran

last: Chai

state: Singapore

address: 60 Biopolis Street, Genome, #02-01

country: Singapore

institute: Genome Institute of Singapore

summary: Enhancing brown fat activity and promoting white fat browning are attractive therapeutic strategies for obesity and its associated metabolic disorders. To provide a comprehensive picture of the regulatory network in these processes, we conducted a series of transcriptome studies by RNA-seq and quantified the mRNA and lncRNA changes during white fat browning induced by chronic cold exposure, beta-adrenergic agonist and intensive exercises, brown fat activation by acute cold exposure and inactivation by thermoneutrality.

overall_design: 18 RNA-seqs were performed with 2-3 replicates for each experiment.

first: Lei

last: SUN

first: Sujoy

last: Ghosh


title: Very deep small RNA sequencing of whole rat medial entorhinal cortex

city: Trondheim

first: Lene

middle: Christin

last: Olsen

address: Erling Skjalgsons gt. 1

department: Medical Faculty

country: Norway

institute: Norwegian University of Science and Technology - NTNU

summary: We performed very deep small RNA sequencing on one sample of medial entorhinal cortex of a Long Evans rat aged P23 to see which small RNAs could be detected in this brain area.

overall_design: Small RNA sequencing of medial entorhinal cortex of one rat on one lane of the Illumina HiSeq2000

middle: C


title: Etv2-mediated hemangiogenic fate commitment of mesoderm

city: St. Louis

first: Kyunghee

last: Choi

state: MO

address: 660 S Euclid Ave

institute: Washington University in St. Louis, School of Medicine

summary: A comprehensive understanding of a lineage map and molecular mechanisms underlying lineage specification is fundamental to development. To this end, ETS transcription factor Etv2 functions as the master regulator of hematopoietic and endothelial cell formation. As such, Etv2 regulated hemangiogenesis provides an excellent model for assessing cell lineage specification. Herein, we generated several reporter embryonic stem (ES) cell lines to map developmental route pertaining to hemangiogenesis. We performed transcriptome analysis and high throughput CRISPR screening to further delineate molecular mechanisms regulating hemangiogenic lineage specification. We show a distinct lineage map of hemangiogenesis, in which hemangiogenic fate is specified not simply by the onset of Etv2 expression, but in a threshold-dependent manner.  Importantly, VEGF-FLK1 signaling is necessary for efficiently achieving ETV2-threshold. Moreover, we find forkhead transcription factor Foxh1 to be required for FLK1 mesoderm formation. These studies provide a roadmap in hematopoietic and vascular cell generation and applications in regenerative medicine.

overall_design: We utilized T(Brachyury), Etv2 and Scl expression together with PDGFR-alpha and FLK1 mesodermal markers to track mesoderm differentiation.  (1) We performed transcriptome analysis of different stages in the generation of Etv2-high cells. 7 samples were included. (2) We performed Clustered regularly interspaced short palindromic repeats (CRISPR)-mediated screening for genes implicated in hemangiogenic cell lineage specification, in which 4 samples were included.

first: Haiyong


title: Dynamic of DNA methylation during Arabidopsis reproduction

city: Montpellier

first: Daniel

last: Grimanelli

address: 911 Av Agropolis

country: France

institute: IRD

type: Methylation profiling by high throughput sequencing

summary: Cytosine methylation is a key epigenetic mark in many organisms, important for both transcriptional control and genome integrity. While relatively stable during somatic growth, DNA methylation is reprogrammed genome-wide during mammalian reproduction. Reprogramming is essential for zygotic totipotency, and to prevent transgenerational inheritance of epimutations. The extent of DNA methylation reprogramming in plants however remains unclear. Here, we developed sensors reporting with single-cell resolution CG and non-CG methylation in Arabidopsis. Live imaging during reproduction revealed distinct and sex-specific dynamics for both contexts. We found that CHH methylation in the egg cell depends on DRM2 and Pol V, two main actors of RNA-directed DNA methylation, but does not depend on Pol IV. Our sensors provide insight into global DNA methylation dynamics at the single cell level with high temporal resolution, and offer a powerful tool to track CG and non-CG methylation both during development and in response to environmental cues in all organisms with methylated DNA.

overall_design: ChIP-seq experiments conducted using transgenic lines expressing either CG or non-CG binding proteins. mCG in WT (Col(O)), mCHH and drm1-drm2-cmt2-cmt3, plus inputs.

first: Mathieu

last: Ingouff


title: Time-restricted feeding shifts the skin circadian clock and alters UVB-induced DNA damage

city: Irvine

first: Bogi

last: Andersen

state: CA

address: 250 Sprague Hall

department: Department of Biological Chemistry

institute: University of California, Irvine

summary: The epidermis is as a highly regenerative barrier protecting organisms from environmental insults, including ultraviolet radiation, the main cause of skin cancer and skin aging. Here we show that time-restricted feeding (RF) shifts the phase and alters the amplitude of the skin circadian clock and affects the expression of approximately 10% of the skin transcriptome. Furthermore, a strikingly large number of skin-expressed genes are acutely regulated by food intake. While the circadian clock is required for daily rhythms in DNA synthesis in epidermal stem cells, RF-induced shifts in clock phase do not alter the phase of DNA synthesis. However, both the expression of the key DNA repair gene Xpa, and the diurnal sensitivity to UVB-induced DNA damage, are altered by RF. Together our findings indicate an unexpected regulation of skin function by time of feeding and emphasize the important link between circadian rhythm, food intake, and skin health.

overall_design: Totally 24 samples were sequenced. One sample for each time point and 8 time points for each feeding group. Each sample was pooled from 2-4 biological replicates.

first: Hong

first: Elyse

last: van Spyk

first: Qiang

last: Liu

first: Mikhail

last: Geyfman

first: Michael

last: Salmans

first: Vivek

last: Kumar

first: Alexander

last: Ihler

first: Ning

last: Li

first: Joseph

last: Takahashi


title: Transcriptome profiling from the brain of male and female zebrafish

first: Justin

middle: Jeyakani

last: Joseph Gnanakkan

address: 60, Biopolis street

department: Computational and Systems Biology

summary: Genome-wide analysis  to identify sex specific transcripts in the brain of male and female zebrafish

overall_design: Examination of sex specific gene expression in the brain of male vs female zebrafish.

first: Sinnakaruppan

last: Mathavan


title: Determine the function of lncBATE10 in brown and white adipocyte culture

first: Xiaoran

last: Chai

summary:
Purpose: Determine the function of lncBATE10 in brown and white adipocyte differentiation
Methods: primary brown and white apreadipocytes were isolated by cell culture. We infected brown preadipocytes with retroviral shRNA targeting lncBATE10 or with retrovirus overexpressing lncBATE10. Cells were induced to differetinate for 5 days. Total RNA were harvested for RNA-seq
Conclusions: Our study shows that lncBATE10 is required for BAT-selective program expression. Overexpression of lncBATE10 is not sufficient to promote BAT marker expression.

overall_design: total RNAs from primary brown and white adipocytes cultures (sh-control, shRNA knockdown, overexpression vector, overexpression of lncBATE10)  were generated by deep sequencing using Hi-seq 2000

first: Lei

last: Sun

first: Sujoy


title: Effect of receipt of male ejaculate sex peptide on micro RNA expression in female Drosophila melanogaster, 3 hours after the start of mating, in HeadThorax (HT) and Abdomen (Abd) tissues.

city: Oxford

first: Helio

last: Pais

address: Jonh Radcliffe Hospital

department: Weatherall Institute of Molecular Medicine

country: United Kingdom

institute: University of Oxford

summary: Seminal fluid proteins (Sfps) can have significant effects on female behaviour and physiology. In Drosophila melanogaster a single Sfp, sex peptide (SP), triggers a remarkably diverse array of fundamentally important responses. It increases female fecundity, juvenile hormone (JH) production, antimicrobial peptide production and feeding rates, whilst decreasing sexual receptivity. Although these features are expected to leave a strong genomic signature, previous studies indicated that Sfps affect the expression of relatively modest numbers of genes. A potential explanation is that increased resolution is needed or that SP could also elicit post-transcriptional control at the level of microRNAs (miRs). We hypothesised that responses to SP at both the transcriptional and post-transcriptional levels could provide precise temporal and spatial responsiveness and facilitate the expression of diverse phenotypes from a single ligand. To test this idea, we used microarrays and Solexa deep sequencing to profile mRNA and miR responses to SP in adult female abdomen (Abd) and head+thorax (HT) tissues. SP elicited a rich but subtle signature of temporally and spatially controlled mRNAs related to functions that match the predicted known effects of SP, but that also suggest potentially novel roles for SP such as in phototransduction. mRNA profiles were much more varied in the HT than in the Abd, both within and across time points (3 and 6h after mating). The deep sequencing data revealed post-transcriptional control of SP responses by differential expression of miRs in adults. As for the mRNA profiles, there was evidence for tissue specificity. miRs showing largest expression differences tended to be down regulated in the HT and up regulated in the Abd. Receipt of SP significantly down-regulated the expression of miR279, a micro-RNA with potentially diverse sensory and reproductive roles, across the HT and Abd. We also identified potential differential expression for miRs 31A, 184, 276A and 989. Our study provides the first evidence of changes in both transcriptional and post-transcriptional expression in adults in response to Sfps, and provides a potential mechanism for the extraordinary diversity of responses seen in response to the single SP stimulus.

overall_design:
Examination of microRNA expression level changes in HeadThorax and Abdomen tissues of females 3 hours after receipt of a full ejaculate (SP+) versus an ejaculate minus sex peptide (SP0). 2 biological replicates of each treatment (replicates 3 and 4 for HT and 2 and 4 for Abd).

This submission represents the Solexa deep sequencing component of the study.

first: Anastasia

last: Gioti

first: Stuart

last: Wigby

first: Bregje

last: Wertheim

first: Simon

last: Moxon

first: Tina

last: Rathjen

first: Eugene

last: Schuster

first: Pedro

last: Martinez

first: Caroline

middle: J

last: Pennington

first: Vincent

last: Moulton

first: Tamas

last: Dalmay

first: Linda

last: Partridge

first: Tracey

last: Chapman


title: Transcriptional response of the extremophile red alga Cyanidioschyzon merolae to changes in CO2 concentrations

status: Public on Jul 31 2017

city: Düsseldorf

first: Andreas

middle: PM

last: Weber

address: Universitätsstr. 1

department: Institute for Plant Biochemistry

country: Germany

institute: Heinrich-Heine-University

summary: Cyanidioschyzon merolae (C. merolae) is an acidophilic red alga growing in a naturally low carbon dioxide (CO2) environment. Although it uses a ribulose 1,5-bisphosphate carboxylase/oxygenase with high affinity for CO2, the survival of C. merolae relies on functional photorespiratory metabolism. In this study, we quantified the transcriptomic response of C. merolae to changes in CO2 conditions. We found distinct changes upon shifts between CO2 conditions, such as a concerted up-regulation of photorespiratory genes and responses to carbon starvation. We used the transcriptome data set to explore a hypothetical CO2 concentrating mechanism in C. merolae, based on the assumption that photorespiratory genes and possible candidate genes involved in a CO2 concentrating mechanism are co-expressed. A putative bicarbonate transport protein and two α-carbonic anhydrases were identified, which showed enhanced transcript levels under reduced CO2 conditions. Genes encoding enzymes of a PEP-CK-type C4 pathway were co-regulated with the photorespiratory gene cluster. We propose a model of a hypothetical low CO2 compensation mechanism in C. merolae integrating these low CO2-inducible components.

overall_design: Transcriptomes of C. merolae wild-type cells grown under high CO2 (5% CO2 in air, HC) conditions, shifted to low CO2 (0.04% CO2 in air, LC) for 3 and 24 h, and then shifted back to HC conditions for 24 h.  Samples were sequenced in biological triplicates.

first: Nadine

last: Rademacher

first: Thomas

last: Wrobel

first: Alessandro

middle: W

last: Rossoni

first: Samantha

last: Kurz

first: Andrea

last: Bräutigam

middle: M

first: Marion

last: Eisenhut


title: Hierarchical role for transcription factors and chromatin structure in genome organization along adipogenesis

city: Ramat Gan

first: Ofir

last: Hakim

address: Bar-Ilan University

country: Israel

institute: Bar-Ilan University

summary: The three dimensional folding of mammalian genomes is cell-type specific and hard to convert suggesting that it is an important layer of gene regulation. Analysis of genome-wide chromosomal associations revealed adipogenesis-specific spatial clustering of adipogenic genes in 3T3-L1 cells. High temporal resolution analysis revealed that the adipogenic "hub", sampled by pparg and Lpin1, undergoes orchestrated reorganization along adipogenesis. Coupling the dynamics of genome architecture with multiple chromatin datasets, points that among all transcription factors tested, RXR is central in genome reorganization at the beginning of adipogenesis. Interestingly, at the end of differentiation the adipogenic hub is shifted to H3K27me3 repressive environment in conjunction with attenuation of gene transcription. Together, we propose stage-specific hierarchy among the orchestra of transcription factors contributing to the establishment of the adipogenic genome architecture that brings together the adipogenic genetic program. In addition, the repositioning to H3K27me3 environment in the end of the differentiation may contribute to stabilize gene expression levels and diminish the developmental plasticity of the specialize cell.

overall_design: High resolution 4C-seq on three different baits (pparg, Lpin1 and Ifng) on different time points during adipogenesis (preadipocytes-day0, day1, day2 and mature adipocytes- day6) and on B cells.

first: Avital

last: Sarusi Portuguez

first: Michal

last: Schwartz

first: Rasmus

last: Siersbaek

first: Ronni

last: Nielsen

first: Myong-Hee

last: Sung

first: Susanne

last: Mandrup

first: Tommy

last: Kaplan


title: Ubiquitination of Stalled Ribosome Triggers Ribosome-associated Quality Control [SET2]

city: Wako

first: Shintaro

last: Iwasaki

state: Saitama

address: S205 Bioscience Bldg. 2-1 Hirosawa

department: Cluster for Pioneering Research

institute: RIKEN

summary: Translation elongation stalling has the potential to produce toxic truncated protein fragments. Translation of either non-stop mRNA or transcripts coding for poly-basic sequences induces ribosome stalling, and the arrest product is degraded by the ribosome-mediated quality control (RQC) system. During this process, the stalled ribosome is dissociated into subunits, and the polypeptide is ubiquitinated by the E3 ubiquitin ligase Listerin on the 60S large ribosomal subunit, leading to subsequent proteasomal degradation. However, it is largely unknown how the specific stalled ribosomes are recognized as aberrant to engage the RQC system. Here, we report that ubiquitination of the ribosomal protein uS10 of the 40S small ribosomal subunit, by the E3 ubiquitin ligase Hel2 (or RQC-trigger (Rqt) 1) initiates RQC. We identified a novel RQC-trigger (RQT) complex composed of the RNA helicase-family protein Slh1/Rqt2, the ubiquitin binding protein Cue3/Rqt3, and yKR023W/Rqt4 that is required for RQC. The defects in RQC of the RQT mutants correlated with sensitivity to anisomycin, which stalls ribosome at the rotated form, suggesting that RQT factors rescue ribosomes stalled by this drug. Our un-biased survey by ribosome profiling revealed that ribosomes stalled at the rotated state with specific pairs of codons at P-A sites serve as RQC substrates. Rqt1 specifically ubiquitinates these arrested ribosomes to target them to the RQT complex, allowing subsequent RQC reactions including dissociation of the stalled ribosome into subunits. Our results provide mechanistic insight into the surveillance system for aberrant proteins induced by ribosome stalling and mediated by ribosome ubiquitination.

overall_design: Ribosome profiling samples are obtained from wild type and RQT mutant  (rqt1, rqt2, uS10KR, Rqt1DeltaRingDomain) yeast with 2 replications.

first: Yoshitaka

last: Matsuo

first: Ken

last: Ikeuchi

first: Yasushi

last: Saeki

first: Christian

last: Schmidt

first: Tsuyoshi

last: Udagawa

first: Fumiya

last: Sato

first: Hikaru

last: Tsuchiya

last: Becker

first: Keiji

last: Tanaka

first: Nicholas

middle: T

last: Ingolia

first: Roland

last: Beckmann

first: Toshifumi

last: Inada


title: The impact of Tcf1/Lef1 deficiency and prostagladin E (PGE) treatment on leukemic stem cell (LSC) transcriptome

city: Philadelphia

first: Bing

last: He

state: PA

address: 3501 Civic Center Blvd

institute: Children's Hospital of Philadelphia

summary: Comparison of trasncriptome bewteen control LSCs and Tcf1/Lef1 deficient LSCs, LSCs treated with DMSO, PGE1 or PGE2.

overall_design: Lineage negative BM cells were isolated from control or Tcf1/Lef1-deficient mice, and retrovirally transduced with p210-BCR-ABL retrovirus twice (at 24 hr interval), and 48 hrs after isolation, Lin-c-Kit+ Sca1+GFP+ cells were sorted as LSCs. The LSCs established from control BM cells were treated with DMSO, 20µM PGE1 or 20 µM PGE2 for 6 hrs.

first: Hai-Hui

last: Xue


title: Single glomerular podocyte RNA-seq analysis

city: Nanjing

first: yuqiu

last: lu

state: Jiangsu

address: 305 East Zhongshan Road

institute: National Clinical Research Center for Kidney Disease

summary: Available single-cell RNA-seq analyses have revealed that individual cells of the same type differ substantially in gene expression. We wonder whether glomerular podocytes, the cell type that is part of glomerular filtration, also exhibit big difference in gene expression among individual cells; and what biological information could be obtained from the single-podocyte RNA-seq data. Therefore, we isolated mouse glomeruli by Dynabead/magnetic concentration method, and digested them with enzymes to dissociate the cells. We loaded the single cell suspension to a Fluidigm  C1 Single-Cell Auto Prep System for single cell cDNA preparation. The cDNA samples were amplified and underwent sequecing using Illumina Highseq 2000 system.

overall_design: In total, 20 single podocytes from a C57BL/6 male mouse were analyzed.

first: Yuqiu

last: Lu

first: Yuting

last: Ye

first: Shaolin

last: Shi


title: Molecular signature for anastasis, recovery from the brink of apoptotic cell death

city: Santa Barbara

first: Kenneth

middle: S.

last: Kosik

state: California

address: UC Santa Barbara

department: NRI

institute: UCSB

summary: Cells can survive effector caspase (caspase 3/7) activation in response to transient apoptotic stimuli, a process named anastasis. To characterize the molecular events that occur during anastasis, we performed whole transcriptome RNA sequencing of untreated, apoptotic, and recovering cells. We found that anastasis is an active, two-stage program with unique transcriptional profiles in each stage. We also identified 10 genes that specific to the early stage of anastasis.

overall_design: 3hr ethanol treatment was used to induce apoptosis in Hela cells. Ethanol was washed away after 3hr treatment to allow cells to recover. Total RNA was prepared from mock-treated cells, ethanol-treated cells and cells after 1hr, 2hr, 3hr, 4hr, 8hr, 12hr recovery, followed by ribosomal RNA depletion. 3 biological replicates were included for each group. Sequencing was done using Ion Proton.

first: Gongping

last: Sun

first: Elmer

last: Guzman

first: Hongjun

last: Zhou

first: Denise

last: Montell


title: Silencing SMOC2 protects from kidney fibrosis by inhibiting Fibroblast to Myofibroblast Transformation

city: Boston

first: John

middle: N

last: Hutchinson

state: Massachusetts

address: 201 Wachusett Street

institute: Diamond Age Data Science

summary: Secreted MOdular Calcium-binding protein-2 (SMOC2) belongs to the SPARC (Secreted Protein Acidic and Rich in Cysteines) family of matricellular proteins whose members are known for their secretion into the extracellular space to modulate cell-cell and cel

overall_design: mRNA sequencing of mouse kidney of wildtype and Smoc2 transgenic mice with and without 7 day unilateral uretal obstruction intervention

first: Casimiro

last: Gerarduzzi

first: Vishal

last: Vaidya


title: Epigenetic silencing of the tumor suppressor RASSF4 favors multiple myeloma progression

first: Jerome

last: Moreaux

address: 80 Av. Augustin Fliche

department: Biological Hematology

institute: CHRU Montpellier

summary: RAS is frequently mutated in multiple myeloma (MM) leading to aberrant signaling. Next to their classical oncogenic effects, RAS proteins also mediate anti-tumor effects through the Ras-Association Domain Family (RASSF) proteins. Currently, no data about the role of RASSF proteins in MM disease is available. Here we report that RASSF4 is downregulated during MM progression, correlating with a bad prognosis. Epigenetic modulating agents significantly increased RASSF4 expression, indicating that the RASSF4 downregulation is due to epigenetic silencing. Forced RASSF4 expression induced a G2-phase arrest and caspase-3 mediated apoptosis in human MM cell lines, strongly reduced viability of primary CD138+ MM cells and significantly reduced in vivo tumor growth. Moreover, we demonstrated RASSF4-MST1 binding and the involvement of the downstream signaling pathways JNK/Jun, p38 and p53. RASSF4 overexpression sensitized MM cells to the proteasome inhibitor bortezomib, the specific MEK1/2 inhibitor trametinib and the ROS inducer Prima-1Met. Consequently, combining trametinib with histone deacetylase inhibitors (HDACi) revealed very strong synergistic anti-MM activity. In conclusion, our study identified RASSF4 as a new potent tumor suppressor in MM that is epigenetically silenced during MM progression and provides the basis for novel therapeutic strategies enhancing RASSF4 expression (using HDACi) in combination with MEK/ERK inhibitors and bortezomib.

overall_design: Paired-end RNA sequencing analysis of the RASSF4 transduced cell lines cultured for 5 days in the presence of doxycycline

first: Eva

last: De Smedt

first: Ken

last: Maes

first: Stefaan

last: Verhulst

first: Hui

last: Lui

first: Alboukadel

last: Kassambara

first: Carlo

last: Heirman

first: Anke

first: Dirk

last: Hose

first: Karine

last: Breckpot

first: Leonardus

last: Van Grunsven

first: Elke

last: De Bruyne


title: UTRseq_human_brain

city: New York

first: Shahar

last: Barbash

state: NY

address: 1230 York Ave

institute: The Rockefeller University

summary: Alternative polyadenylation (APA) contributes to post-transcriptional regulation, but its role in Alzheimer’s disease (AD) is largely unknown. Using high-resolution SQUARE multiple 3’ primer-based sequencing, we discovered massive APA differences in temporal gyrus tissues from demented AD patients compared to either healthy controls or non-demented donors with AD neuropathology (NDWP). Advanced statistics, microfluidics RT-PCR and protein measurements validated known and novel APA-modified 3’-intact transcripts. Moreover, APA modifications, more than total transcript counts, distinguished AD patients from both controls and NDWP donors and identified cell type-characteristic, cognition-associated and neuropathology-related changes. AD-enhanced APA variants included known therapeutic targets of brain, vascular and autoimmune disorders, predicting co-involvement of these target genes in AD progression; AD/NDWP increases in 3’-intact proteinopathy-related hnRNP mRNAs were inversely associated with protein decreases, whereas NDWP-potentiated cognition was accompanied by distinct ATP and mitochondrial variants. APA variations thus provide a novel resource of unique value for both basic and translational neuroscience researchers.

overall_design: Examination of polyadenylation sites in brain tissues of Alzheimer`s disease patients, with 3 levels of pathology and 3 levels of cognition

first: Hermona

last: Soreq


title: Hybrid poplar (P. tremula x P. alba ) outer vs inner bark transcriptome

city: Sault Sainte Marie

first: Isabel

last: Molina

state: Ontario

address: 1520 Queen Street East, CC403 Essar Bldg.

department: Biology

country: Canada

institute: Algoma University

summary: The periderm of trees produces cork cells, whose cell walls are modified with suberin. We compared the transcriptome of outer bark (cork) vs inner bark (control containing secondary phloem and vacular meristem) to infer genes related to suberim metabolism.

overall_design: mRNA profiles of periderm cells at two developmental stages of wild type (WT) poplar tree stems were generated by deep sequencing, in triplicate (young) and quadruplicate (older stem), using Illumina HiSeq2000.

first: Meghan

last: Rains


title: Exposure to mixture of Environmental endocrine disruptors induce miRNAs/isomiRs deregulation in mouse testis

city: santiago

first: Julio

middle: Buñay

last: Noboa

state: Santiago

address: Alameda 340

department: Physiology

country: Chile

institute: Pontificia Univerisidad Catolica de Chile

summary:
Purpose: In testis the effects of exposure to mixtures of Endocrine disruptors compounds (EDCs) upon expression of miRNAs were not addressed.

Objective: To identify the expression profiles of the 'miRNome' in mice testis chronic exposed to a defined mixture of five EDCs.

Methods: Pregnant mice from 0.5 post-coital day were exposed in the drinking water to a mixture containing 0.3 mg/Kg-bw/day of each phthalate (DEHP, DBP, BBP), plus 0.05 mg/Kg-bw/day of each alkylphenol (NP, OP) until adulthood of male mouse (60 days old). We characterized the 'miRNome' by next generation sequence (NGS).

Results: In mouse testis exposed to EDCs mixture we detected by NGS 2 up-regulated and 8 down-regulated miRNAs along to 36 isomiRs differentially expressed; these results were validated by RT-qPCR. and functional analysis showed deregulation of testicular hormonal status, spermatogenesis disruption and germ cells apoptosis.

Conclusions: Here we provide the first association between deregulation of miRNAs, isomiRs, with histopathological and hormonal alterations in adult mice testis exposed to mixture of EDCs.

overall_design: Testis snc-RNA profiles of 60-day old mice of control (mice drank water only with the dose of vehicles equivalents diluted) and exposed to EDCs mixture were generated by deep sequency using MiSeq-Illumina sequencer

last: Buñay

first: Eduardo

last: Larriba

first: Ricardo

last: Moreno

first: Jesus

last: del Mazo


title: Ubiquitination of Stalled Ribosome Triggers Ribosome-associated Quality Control

first: Shintaro

last: Iwasaki

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

overall_design: Refer to individual Series


title: Molecular substrates for differences between layers and regions of the entorhinal cortex during postnatal development in rats

first: Lene

last: Olsen

address: Erling Skjalgsons gt. 1

summary: We report the first full transcriptome analysis of layer II and deep layers of the medial and lateral entorhinal cortex during postnatal development. Our analysis showed that postnatal timepoint was the most important element in entorhinal cortex transcriptional dynamics, followed by laminar differences. There were fewer differentially expressed genes between the medial and lateral parts of the entorhinal cortex, and most of these were found in layer II.

overall_design: Ribosome depleted deep sequencing of layer II and deep layers of the medial and lateral entorhinal cortex of Long Evans rats at postnatal ages 2,9,23, and 45.


title: Epigenetic resetting of human pluripotency

city: Cambridge

first: Felix

last: Krueger

address: Granta Park

department: Bioinformatics

institute: Altos Labs

summary: Recent attention has focussed on conversion of human pluripotent stem cells to a more naïve transcriptional and epigenetic status. Here we provide a simple method for resetting without transgenes via transient exposure to histone deacetylase inhibition. This protocol is effective across multiple embryo-derived and induced pluripotent stem cells. Reset cells can retain a normal karyotype over multiple passages. Specific markers and global transcriptome profile, including transposable element signature, diverge markedly from conventional human pluripotent stem cells and are consistent with a naïve-like identity. Global DNA methylation is reduced to a level similar to that reported for the inner cell mass, although gain of methylation is also apparent at certain loci. Imprinted methylation is mostly lost, however. In female cells, X chromosome reactivation is evidenced by bi-allelic transcription detected by RNA-FISH. XIST expression is also regained but is predominantly monoallelic. Reset cells are competent for tri-lineage differentiation, both via embryoid body formation and in adherent culture, and can also undergo germline specification. This reliable conversion system and associated transcriptome and methylome datasets will facilitate routine resetting for side by side evaluation with primed pluripotent stem cells and comprehensive interrogation and refinement of candidate naïve pluripotent stem cells.

overall_design: BS-seq of conventional and reset human pluripotent stem cells

first: Ferdinand

last: von Meyenn

first: Wolf

last: Reik


title: Ubiquitination of Stalled Ribosome Triggers Ribosome-associated Quality Control [SET1]

summary: Translation elongation rates are regulated to ensure proper conformation and biological function of proteins. Translation of either non-stop mRNA or transcripts coding for poly-basic sequences induces ribosome stalling, and the arrest product is degraded by the ribosome-mediated quality control system (RQC). During this process, the stalled ribosome is dissociated into subunits, and the polypeptide is ubiquitinated by the E3 ubiquitin ligase Listerin on the 60S large ribosomal subunit (LSU) leading to subsequent proteasomal degradation. However, it is largely unknown how stalled ribosomes are recognized and dissociated into subunits. Here we report that ubiquitination of the ribosomal protein uS10 by the E3 ubiquitin ligase Hel2 is required for the production of the RQC substrate. RQC-trigger (RQT) factors, a RNA helicase-family protein Slh1/Rqt2, ubiquitin binding protein Cue3/Rqt3 and yKR023W/Rqt4, were also required for the primary steps of RQC, and associated with Hel2-ribosome complexes. Rqt2-4 factors were dispensable for the ubiquitination of uS10 by Hel2/Rqt1 and associated with ribosomes independent of the ubiquitination of uS10. However, the ubiquitin-binding activity of Rqt3 were crucial to trigger RQC. Cryo-electron microscopy (cryo-EM) analysis revealed that Hel2 bound ribosomes are in an rotated state containing hybrid state AP- and PE-tRNAs. Furthermore, ribosome profiling revealed that short footprints, hallmarks of hybrid state ribosomes18, were accumulated at tandem CGA rare codons at the beginning of the poly arginine stalling sequence and long footprints at subsequent codons, respectively. Short footprints at CGA codons were decreased in rqt1 mutant but drastically increased in uS10 mutants defective in the ubiquitination or rqt2 mutant. Collectively, our results demonstrate   that Hel2 stabilizes ratcheted ribosomes leading to ubiquitination of uS10. Subsequently, Rqt2-4 factors target these hybrid state ribosomes specifically, allowing subsequent RQC reactions.

overall_design: Ribosome,profiling

first: Yoshitaka

last: Matsuo

first: Ken

last: Ikeuchi

first: Yasushi

last: Saeki

first: Christian

last: Schmidt

first: Tsuyoshi

last: Udagawa

first: Fumiya

last: Sato

first: Hikaru

last: Tsuchiya

first: Koji

last: Ando

first: Thomas

last: Becker

first: Keiji

last: Tanaka

first: Nicholas

last: Ingolia

first: Roland

last: Beckmann

first: Toshifumi

last: Inada


title: Genomewide Analysis of Clp1 Function in Transcription in Budding Yeast

city: Tehran

first: Ali

last: Sharifi-Zarchi

address: Royan Institute, Bani Hashem Sq.

country: Iran

institute: Royan Institute for Stem Cell Biology and Technology

summary: We report the GRO-Seq analysis in Clp1 mutant and the isogenic wild type strain to determine the the role of Clp1 in termination of transcription on a genomewide scale.

overall_design: Examination of GRO-Seq profile in the temperature-sensitive mutant and the wild type strain at permissive and non-permissive temperature.

first: Athar

last: Ansari

first: Hamidreza

last: Chitsaz

first: Nadra

last: Al Husini

first: Sayed Ahmad

last: Mousavi


title: RNA profiling of sclerotinia-infected leaves of Brassica napus reveals cross talk between redox and hormone pathways

city: Winnipeg

first: Mark

last: Belmonte

state: Manitoba

address: 50 Sifton Road

department: Biological Sciences

institute: University of Manitoba

summary: Global transcriptome profiling of suceptible and tolerant lines of Brassica napus infected with Sclerotinia sclerotiorum using a petal inoculation method that mimics field conditions.

overall_design: A total of 12 samples were sequenced. Three biological replicates of mock- and ascospore-inocluated leaves in both susceptible (Westar) and tolerant (ZY821) genotypes of B. napus. Samples were sequenced with Illumina HiSeq2000 100bp single-end reads at the Genome Québec Innovation Centre in Montréal, Canada

first: Ian

last: Girard

first: Chaobo

last: Tong

first: Xingyu

last: Mao

first: Michael

first: Junyan

last: Huang

first: Teresa

last: de Kievit

first: Dilantha

last: Fernando

first: Shengyi

last: Liu


title: Genome-wide expression analysis of FOXF2-regulated genes in normal and metastatic breast epithelial cells

city: Baltimore

first: Pang-Kuo

last: Lo

state: Maryland

address: 108 N. Greene St.

department: Biochemistry and Molecular Biology

institute: University of Maryland School of Medicine

summary: This genome-wide gene expression studies are aimed at deciphering whether FOXF2 transcriptional activity and specificity are compromised in FOXF2-expressing breast cancer cells (MDA-MB-231) compared with FOXF2-expressing normal breast epithelial cells (MCF10A).

overall_design: Total RNA obtained from MCF10A and MDA-MB-231 (231) cell lines transfected with siRNA for 48 hours.

first: Saraswati

last: Sukumar


title: Gene regulation of candidate cancer gene DIP2C

city: Uppsala

first: L.

last: He

address: Dag Hammarskjölds Väg 20

country: Sweden

institute: Uppsala University

summary:
Purpose: The disco-interacting protein 2 homolog C (DIP2C) gene is an uncharacterized gene found mutated in breast and lung cancers. We want to understand the role of DIP2C in tumor development.
Methods: We engineered human DIP2C knockout cell systems by genome editing, and then use next-generation sequencing to identify the genes affected by the loss of DIP2C.
Results: Inactivation of DIP2C triggers substantial gene expression changes, cellular senescence and epithelial to mesenchymal transition in cancer cells.

overall_design: To investigate if loss of DIP2C affects gene expression we performed RNA sequencing and compared gene expression levels in the DIP2C knockout cells to those in parental RKO cells.

first: Chatarina

last: Larsson

first: Tobias

last: Sjöblom


title: Evolution of Gene Expression in the Uterine Cervix related to Steroid Signaling: Conserved features in the regulation of cervical remodeling.

city: West Haven

first: Gunter

last: Wagner

state: CT

address: 300 Heffernan Drive, ISTC 108

department: Systems Biology Institute, EEB

institute: Yale University

summary: The uterine cervix is the boundary structure between the uterus and the vagina and is a key component of the female reproductive tract for the maintenance of pregnancy and timing of parturition. Here we report on a comparative transcriptomic study of the cervix of four placental mammal species, mouse, guinea pig, rabbit and the basal eutherian mammal, armadillo, Dasypus novemcinctus, and one marsupial species, Monodelphis domestica. Our aim is to investigate the evolution of cervical gene expression as related to putative mechanisms for functional progesterone withdrawal. Our main findings are: 1) The patterns of gene expression found in eutherian species is consistent with the notion that an increase in the ratio of E/P4 signaling strength is critical for cervical remodeling. How the increased E/P4 ratio is achieved, however, is variable between eutherian species. 2) None of the genes related to steroid signaling modulated in eutherian species change expression during opossum gestation, suggesting that the role of P4 signaling in cervical remodeling evolved after the common ancestor of marsupials and placental mammals. 3) A tendency for decreased expression of progesterone receptor co-activators (NCOA1, -2 and -3, and CREBP) towards term is a shared derived feature of eutherians. This finding suggests that parturition is associated with broad scale histone de-acetylation. We tested this hypothesis by performing Western-blotting on mouse cervix and found evidence for a striking degree of histone de-acetylation in labor. This finding may have important implications for the control of premature cervical remodeling and preterm birth in humans.

sample_organism:
Dasypus novemcinctus
Oryctolagus cuniculus
Mus musculus
Monodelphis domestica

overall_design: We sequenced mRNA from cervix samples at different stages of pregnancy of four placental mammal species (one previously uploaded) and one marsupial species.

middle: P

first: Mauris

last: Nnamani

first: Arun

last: Chavan

first: Jamie

last: Maziarz

first: Stella

last: Protopapas

first: Jennifer

last: Condon

first: Roberto

last: Romero


title: Comprehensive Analysis of Nucleocytoplasmic Dynamics of mRNA in Drosophila cells

city: Amsterdam

first: Bas

last: van Steensel

address: Plesmanlaan 121

department: division of Molecular Biology

country: Netherlands

institute: Netherlands Cancer Institute

summary: Eukaryotic mRNAs undergo a cycle of transcription, nuclear export, and degradation. A major challenge is to obtain a global, quantitative view of these processes. Here we measured the genome-wide nucleocytoplasmic dynamics of mRNA in Drosophila cells by metabolic labeling in combination with cellular fractionation. By mathematical modeling of these data we determined rates of transcription, export and cytoplasmic decay for >5,000 genes. We characterized these kinetic rates and investigated links with mRNA features, RNA-binding proteins (RBPs) and chromatin states. We found prominent correlations between mRNA decay rate and transcript size, while nuclear export rates are linked to the size of the 3'UTR.  Transcription, export and decay rates are each associated with distinct spectra of RBPs. Specific classes of genes, such as those encoding cytoplasmic ribosomal proteins, exhibit characteristic combinations of rate constants, suggesting modular control. Overall, transcription and decay rates have a major impact on transcript abundance, while nuclear export is of minor importance. Finally, correlations between rate constants suggest global coordination between the three processes. Our approach should be generally applicable to other cell systems and provides insights into the genome-wide nucleocytoplasmic kinetics of mRNA.

overall_design: 24 RNA-seq experiments comprising 2 biological replicates: pre-exsiting nuclear mRNA time 0h (samples 1&13), pre-exsiting nuclear mRNA time 0.5h (samples 2&14), pre-exsiting nuclear mRNA time 1.5h (samples 3&15) , pre-exsiting nuclear mRNA time 3h (samples 4&16), pre-exsiting nuclear mRNA time 5h (samples 5&17), pre-exsiting nuclear mRNA time 7.5h (samples 6&18), pre-exsiting cytoplasmic mRNA time 0h (samples 7&19), pre-exsiting cytoplasmic mRNA time 0.5h (samples 8&20), pre-exsiting cytoplasmic mRNA time 1.5h (samples 9&21) , pre-exsiting cytoplasmic mRNA time 3h (samples 10&22), pre-exsiting cytoplasmic mRNA time 5h (samples 11&23), pre-exsiting cytoplasmic mRNA time 7.5h (samples 12&24)

first: Tao

last: Chen


title: Hierarchical Analysis of Multi-mapping RNA-Seq Reads Improves the Accuracy of Allele-specific Expression

status: Public on Jul 30 2017

city: Bar Harbor

first: Narayanan

last: Raghupathy

state: ME

address: 600 Main Street

institute: The Jackson Laboratory

summary: RNA-seq experiment on reciprocal F1 hybrid cross between two diverse mouse strains NOD/ShiLtJ and PWK/PhJ to study the extent of Allele-specific expression (ASE) and the cause of ASE.

overall_design: Illumina 68bp single-end RNA-seq data from 48 reciprocal F1 mice. 24 male F1 progeny from each cross direction with two age and diet groups. In total, there are eight groups defined by three experimental factors cross-direction, diet and age with six biological replicates per group.

first: Kwangbom

last: Choi

first: Matthew

last: Vincent

first: Glen

last: Beane

first: Keith

last: Sheppard

first: Steven

last: Munger

first: Ron

last: Korstanje

first: Fernando

last: Pardo-Manual de Villena

first: Gary

last: Churchill


title: The mRNA expression analysis of psoriasis skin lesion mesenchymal stem cell

city: Taiyuan

first: Ruifeng

last: Liu

state: Shanxi

address: No. 1 Dong San Dao Xiang

department: Institute of Dermatology

institute: Taiyuan City Centre Hospital

summary: RNA sequencing for 6 samples from psoriasis skin lesion mesenchymal stem cell, to examine the differentilly expressed mRNAs between two differernt conditions.

overall_design: Examining 2 conditions, each with 3 replicates


title: Activation of the interferon type I response rather than autophagy contributes to myogenesis inhibition in congenital DM1 myoblasts

city: Pisa

first: Romina

last: D'Aurizio

state: PI

address: via Moruzzi 1

department: IIT

country: Italy

institute: CNR

summary: Congenital Myotonic Dystrophy type 1 (CDM1) is characterized by severe symptoms that affect patients from birth, with 40% mortality in the neonatal period and impaired skeletal muscle development. In this paper, we examined the relationship between autophagy and abnormal myogenic differentiation of CDM1 myoblasts. We investigated these pathological features at both ultrastructural and molecular levels, utilizing two CDM1 foetal myoblasts, CDM13 and CDM15, with 1800 and 3200 repeats respectively. The congenital nature of these CDM1 myoblasts was confirmed by the high methylation level at the DMPK locus. Our results indicated that abnormal autophagy was independent of myogenic differentiation as CDM13 myoblasts differentiated as well as control myoblasts but underwent autophagy like CDM15 displaying impaired differentiation. miRNA expression profiles revealed that CDM15 myoblasts failed to upregulate the complex network of myo-miRNAs under MYOD and MEF2A control while this network was upregulated in CDM13 myoblasts. Interestingly, the abnormal differentiation of CDM15 myoblasts was associated with cellular stress accompanied by the induction of the interferon type 1 pathway (innate immune response). Indeed, inhibition the IFN type I pathway restores myogenic differentiation of CDM15 myoblasts suggesting that the inappropriate activation of the innate immune response might contribute to impaired myogenic differentiation and severe muscle symptoms observed in some CDM1 patients. These findings open up the possibility of new therapeutic approaches to treat CDM1.

overall_design: miRNA profiing of two CDM1 fetal myoblasts carryng different numbers of triplets (MCBO and DM15Q with 1800 and 3200 repeats, respectively).  12,000 cells/cm2 were grown in HAM’S/F10 medium with 50 µg/ml Fetuin, 5 µg/ml Insulin, 0.5 mg/ml BSA and 20% FBS to subconflence.  Subconfluent cells were shifted to differentiation medium (MEM with 10 µg/ml, insulin, 10 ng/ml EGF and 0.5 mg/ml BSA) were samples at day 0 and day 3.  Two replicates per sample.

first: Milena

last: Rizzo

first: Marcella

last: Simili


title: Chromatin accessibility dynamics reveal novel functional enhancers in C. elegans

city: Stanford

first: Aaron

last: Daugherty

address: 300 Pasteur Dr

department: Genetics

institute: Stanford University

summary: Chromatin accessibility, a crucial component of genome regulation, has mostly been studied in homogenous and simple systems, such as isolated cell populations or early development models. Whether it can sensitively be assessed in complex, dynamic systems in vivo remains largely unexplored.  In this study, we identify chromatin accessibility changes in a whole organism from embryogenesis to adulthood, using C. elegans as a model. Chromatin accessibility changes between developmental stages are highly reproducible, recapitulate histone modification changes, and reveal key regulatory aspects of the epigenomic landscape throughout organismal development.  Even when derived from the chromatin of a whole organism, dynamic regions of chromatin accessibility are capable of identifying novel cell-type- and temporal-specific enhancers, which we functionally validate in vivo.  Furthermore, by integrating transcription factor binding motifs into a machine-learning framework, we identify EOR-1 as a potential early regulator of chromatin accessibility changes.  Our study provides a unique resource for C. elegans, a system in which the prevalence and importance of enhancers remains poorly characterized, and demonstrates the power of using whole organism chromatin accessibility to identify novel regulatory regions in complex systems.

overall_design: Three independent biological replicates for three key life stages: early embryo, larval stage 3 (L3), and young adults, were tightly temporally synchronized, and nuceli isolated before performing the standard ATAC-seq protocol. In addition, an input control, Tn5 transposition of gDNA purified from mixed stage worms was produced.

first: Anne

last: Brunet


title: RNA-seq of mixed mouse glomerular mesangial and endothelial cells

city: Nanjing

first: yuqiu

last: lu

address: 305 East Zhongshan Road

institute: National Clinical Research Center for Kidney Disease

summary: There are 3 cell types in a glomerulus: podocytes, mesangial cells and endothelial cells. These cell types play distinct roles in the structure and functions of glomeruli. In order to profile the gene expression of single glomerular cells, we isolated mouse glomeruli by Dynabead/magnetic concentration method and digested them with enzymes to dissociate them into single cells. We loaded the single cell suspension to a Fluidigm C1 Single-Cell Auto Prep System for single cell cDNA preparation. We performed qPCR analyses of marker genes of podocytes (Npsh2, Synaptopodin, WT1), mesangial cells (Gata3, IGFbp5) and endothelial cells (CD31, Tie2) to determine the identity of each cDNA sample. To identify podocyte-specific genes, we mixed 15 mesangial cell cDNA samples and 15 endothelial cell cDNA samples and further divided into 3 aliquots as replicates for sequencing using Illumina HiSeq 2000 system. The resulting data are used to compare with that of podocytes in order to identify podocyte-specific genes.

overall_design: A C57BL/6 male mouse was sacrificed for isolation of glomeruli. Glomeruli were dissociated into single cells, which were loaded to a Fluidigm C1 Single-Cell Auto Prep System for single cell cDNA preparation. qPCR analyses of marker genes of podocytes (Npsh2, Synaptopodin, WT1), mesangial cells (Gata3, IGFbp5) and endothelial cells (CD31, Tie2) were conducted to determine sample identities. Fifteen mesangial cells and 15 endothelial cells' cDNA samples were mixed and divided into 3 aliquots as replicates for sequencing.

first: Yuqiu

last: Lu

first: Yuting

last: Ye

first: Shaolin

last: Shi


title: Tek+ cells from the E10 AGM region of Rictorf/f; Tie2-cre and control

city: Beijing

first: Dan

address: NO.1 Beichen West Road, Chaoyang District

institute: Beijing Institute of Genomics

summary: we performed RNAseq analysis on Rictorf/f; Tie2-cre and control tek+ cells, to determine which hematopoietic genes are activated in the endothelium in the AGM by Rictor.

overall_design: Two sample are analyzed.

first: Weili

last: Wang


title: Reduced protein expression in a virus attenuated by codon deoptimization

city: Austin

first: Claus

last: Wilke

state: Texas

address: 2500 Speedway, A4800

department: Integrative Biology

institute: University of Texas at Austin

summary: A general means of viral attenuation involves the extensive recoding of synonymous codons in the viral  genome. The mechanistic underpinnings of this approach remain unclear, however. Using quantitative  proteomics and RNA sequencing, we explore the molecular basis of attenuation in a strain of bacteriophage  T7 whose major capsid gene was engineered to carry 182 suboptimal codons. We do not detect  transcriptional effects from recoding. Proteomic observations reveal that translation is halved for the recoded  major capsid gene, and a more modest reduction applies to several co-expressed downstream genes.  We observe no changes in protein abundances of other co-expressed genes that are encoded upstream.  Viral burst size, like capsid protein abundance, is also decreased by half. Together, these observations  suggest that, in this virus, reduced translation of an essential polycistronic transcript and diminished  virion assembly form the molecular basis of attenuation.

overall_design: mRNA profiles of T7-infected E. coli from three different strains of T7, across three distinct time points post-infection, with three replicates of each strain-time combination.

first: Benjamin

middle: R

last: Jack

first: Daniel

last: Boutz

first: Matthew

last: Paff

first: Bartram

last: Smith

first: James

last: Bull

middle: O


title: Effects of IL2RA non-coding and coding sequence edits on IL2RA expression in primary human T cells.

city: San Francisco

first: Alexander

last: Marson

address: 513 Parnassus Ave, HSW 1033

institute: UCSF

summary: We edited an intronic IL2RA enhancer or exon in primary human naïve T cells. Electroporated cells were activated and sorted on IL2RA expression over three days. Amplicon sequencing was used to assess the effects on sequence edits on IL2RA expression.

overall_design: Cas9 ribonucleoproteins were targeted to the intronic IL2RA enhancer (SNP or CaRE4-Mix) or exon in primary human naïve T cells. Edited cells were activated with anti-CD3 and anti-CD28 antibodies and genomic DNA was extracted from unsorted, IL2RA- or IL2RA+ cells over three days. Amplicon sequencing across the cut sites was performed to assess the sequence edits in the sorted populations.

first: Dimitre

last: Simeonov

last: Gowen


title: The role of IRF6 in salivary gland development

status: Public on Jul 29 2017

city: Houston

first: Walid

middle: D

last: Fakhouri

state: TX

address: 1941 East Rd

department: Diagnostic and Biomedical Sciences

institute: University of Texas School of Dentistry at Houston

summary: Loss of Irf6 leads to disruption of branching morphogenesis and secretory acnii formation in salivary gland. To determine the differentially expressed genes in Irf6 mutant, embryonic salivary gland tissues were extracted at E14.5.

overall_design: Two biological replicates of Irf6 null salivary gland tissues and wild-type littermates at E14.5

first: Goo

last: Jun


title: Igf1r deficiency attenuates acute inflammatory response and oxidative stress following bleomycin-induced lung injury

city: Logroño

first: Jose

middle: Garcia

last: pichel

state: La Rioja

address: Piqueras 98

department: Oncology

country: Spain

institute: FUNDACIÓN RIOJA SALUD - CIBIR

summary: IGF1R (Insulin-like Growth Factor 1 Receptor) is a ubiquitously expressed transmembrane tyrosine kinase receptor with multiple functions including inflammation. IGF activity maintains human lung homeostasis, being involved in relevant pulmonary diseases with an inflammatory component, such as lung cancer, COPD, asthma and pulmonary fibrosis. Here we examined the role of IGF1R in lung inflammation using mice with a postnatal deficiency of Igf1r and a model of bleomycin(BLM)-induced lung injury. Lung transcriptome analysis of Igf1r-deficient mice showed a general inhibition of transcription of genes related to epigenetics, inflammation/immune response and oxidative stress activity with potential pulmonary protective roles. Early upon intratracheal BLM treatment, mutant mice showed improved survival and milder pulmonary injury and inflammation. Their lungs presented down-regulation of macrophage (Marco/Adgre1), neutrophil-related (Cxcl1/Ly6g), pro-inflammatory (Tnf/Il1b/Il6), endothelial adhesion (Icam1/Pecam1) and alveolar damage (Aqp5/Sftpc) markers and up-regulation of resolution phase markers (Csf1/Il13/Cd209a). Changes in mRNA of IGF system genes were also found, in parallel to a hindered response to hypoxia (Hif1a) and increased expression of the anti-oxidative stress marker Gpx8. These findings identify Igf1r as an important player in oxidative stress and inflammation and suggest that targeting Igf1r may block the inflammatory response in lung diseases with this component.

overall_design: Lung mRNA profiles of 2 months-old Igf1rfl/fl normal/control transgenic mice and UBC-CreERT2; Igf1rfl/fl Igf1r-deficient mice were generated by deep sequencing using Illumina GAIIx.

first: Sergio

last: Piñeiro-Hermida

first: Icíar

last: López

first: Raquel

last: Torrens

first: María

last: Iñiguez

first: Lydia

last: Alvarez-Erviti

first: José

middle: G

last: Pichel


title: Tumor-derived exosomes modulate PD-L1 expression in monocytes

city: Heidelberg

first: Martina

last: Seiffert

address: Im Neuenheimer Feld 280

department: Department of Molecular Genetics

institute: German Cancer Research Center

summary: Cancer cell-derived vesicles, so-called exosomes, are important means in cell-cell communication between tumor cells and the tissue microenvironment. Amongst others, exosomes harbor functional RNAs, which are transferred to recipient cells and alter the cellular phenotype of respective cells. Aim of the current study was a detailed characterization of the RNA composition of cancer-cell derived exosomes in CLL. Due to prior results showing an enrichment of small RNAs in exosomes, this was focused on profiling of small RNAs. Further, the impact of high abundant exosomal RNAs in phenotypic alterations of cells in the tumor microenvironment upon exosome uptake was studied.

overall_design: Small RNA sequencing of exosomes (n=5) and respective parental cells (n=3) was conducted. Exosomes were isolated from the CLL cell line MEC-1 and for 3 out of 5 samples, paired cellular samples were harvested.

first: Franziska

last: Haderk

first: Murat

last: Iskar

first: Thomas

last: Worst

first: Marc

last: Zapatka

first: Peter

last: Lichter


title: Gene co-expression networks identify Trem2 and Tyrobp as major hubs in human APOE expressing mice following traumatic brain injury.

city: Pittsburgh

first: Iliya

last: Lefterov

address: 130 De Soto Street

department: Enviromantal and Occupational health

institute: University of Pittsburgh

summary: To determine if there is an APOE isoform-specific response to TBI we performed controlled cortical impact on 3-month-old mice expressing human APOE3 or APOE4 isoforms. Following injury, we used several behavior paradigms to test for anxiety and learning and found that APOE3 and APOE4 targeted replacement mice demonstrate cognitive impairments following moderate TBI. Transcriptional profiling 14 days following injury revealed a significant effect of TBI, which was similar in both genotypes.

overall_design: Examination of the effectes of TBI on APOE expressing mice

first: E

middle: L

last: Castranio

first: A

last: Mounier

first: C

last: Wolfe

first: K

last: Nam

first: N

middle: F

last: Fitz

first: F

last: Letronne

first: J

last: Schug

first: R

last: Koldamova

first: I


title: Genome-wide characterization of the effects of MCMBP deletion in T. brucei

city: New York

first: Eric

last: Fritz

address: 1230 York Ave., Box 39

institute: Rockefeller University

overall_design:
Examination of the effects of MCMBP deletion on transcriptome of T. brucei
4: untreated (wild-type)
4: treated with tet for 1d (d1-ko)
4: treated with tet for 2d (d2-ko)

first: Heesook

last: Kim

last: Papavasiliou


title: PATZ1 is a DNA damage responsive transcription factor that inhibits p53 function [doxorubicin treatment]

city: Istanbul

first: Batu

last: Erman

address: Orhanli, Tuzla

department: Biological Sciences and Bioengineering Program

country: Turkey

institute: Sabanci University

summary: Insults to cellular health cause p53 protein accumulation and loss of p53 function leads to tumorigenesis. Thus, p53 has to be tightly controlled. Here we report that the BTB/POZ domain transcription factor PATZ1 (MAZR), previously known for its transcriptional suppressor functions in T lymphocytes, is a crucial regulator of p53. The novel inhibitory role of PATZ1 on the p53 protein marks it as a proto-oncogene. PATZ1 deficient cells have reduced proliferative capacity which we assess by RNASeq and real time cell growth rate analysis. PATZ1 modifies the expression of p53 target genes associated with cell proliferation gene ontology terms. Moreover, PATZ1 regulates several genes involved in cellular adhesion and morphogenesis. Significantly, treatment with the DNA damage inducing drug doxorubicin results in the loss of the PATZ1 transcription factor, as p53 accumulates. We find that PATZ1 binds to p53 and inhibits p53 dependent transcription activation. We examine the mechanism of this functional inhibitory interaction and demonstrate that PATZ1 excludes p53 from DNA binding. This study documents PATZ1 as a novel player in the p53 pathway.

overall_design: RNA-seq was used to define differentially expressed genes in wild-type and PATZ1-/- MEFs treated with the DNA damage inducing compound Doxorubicin (at a concentration of 1uM for 8 hours). Each sample was represented in triplicate. The doxorubicin untreated samples can be accessed at GSE65864.

first: Emre

last: Deniz

first: Nazli

last: Keskin


title: The prominent transcriptional changes of Mycobacterium bovis BCG str. Tokyo 172 following induction by vitamin B1

status: Public on Jul 28 2017

city: harbin

first: Ningning

last: Song

address: No.678. Haping Road

institute: Harbin Veterinary Research Institute

summary: Vitamin B1 (VB1) is a key dietary nutrient and crucial cofactor, which exhibits a series of regulatory functions on cellular processes and the activation of the immune system. To date, the precise effect of VB1 on Mycobacterium tuberculosis has not been fully described. In the present study, the direct influence of VB1 treatment on M. bovis BCG was determined using RNA-sequencing. The selection of this strain was used due to its common physiological features with M.tuberculosis. The investigation of the M. bovis BCG transcription demonstrated significant changes in certain metabolic and cellular process such as the decrease in fatty acid, cholesterol and glycolipid catabolism, the decrease in DNA replication and protein translation, the reduction in cell division and cell wall formation and the induction of arginine biosynthesis. In addition, growth assays indicated that VB1 inhibited the BCG growth rate in vitro, whereas LC/MS analysis demonstrated that the concentration of arginine was higher following VB1 supplementation. It is suggested that VB1 could be used for the treatment of tuberculosis potentially.

sample_organism: Mycobacterium tuberculosis variant bovis

overall_design: 2 samples

first: Zhaoli

last: Li

first: Ziyin

last: Cui

first: Liping

last: Chen

first: Yingying

first: Guanghui

last: Dang

first: Zhe

first: He

first: Siguo

last: Liu


title: 2-aminoimidazoles potentiate ß-lactam antimicrobial activity against Mycobacterium tuberculosis by reducing ß-lactamase secretion and increasing cell wall permeability

city: East Lansing

first: Robert

middle: B.

last: Abramovitch

state: MI

address: 567 Wilson Road

department: Microbiology and Molecular Genetics

institute: Michigan State University

summary: The purpose of this study was to examine the impact of 2-aminoimidazoles on M. tuberculosis transcriptional responese.

overall_design: M. tuberculosis H37Rv was cultured in 7H9 media to an OD600 of 0.4 then treated with 125 µM 2B8 for 2 and 24 h.  Following treatment, RNA was isolated from the bacterial cultures and used for RNA-seq.

first: Albert

middle: B

last: Jeon

first: Benjamin

middle: K

last: Johnson

first: Randall

last: Basaraba


title: Obstructed defecation – an enteric neuropathy? An exploratory study of patient samples

city: Würzburg

first: Corinna

last: Rosenbaum

address: Röntgenring 11

department: Department of Tissue Engineering and Regenerative Medicine

institute: University Hospital Wuerzburg

summary: The aim of this experimental study is to elucidate alterations of the autonomous enteric nervous system at the molecular level in patients with obstructed defecation, who represent one of the most predominant groups of constipated patients.

overall_design: Full-thickness rectal wall samples of patients with obstructed defecation (OD) were analyzed by RNA-Seq and compared with controls.

first: Mia

first: Marco

last: Metzger


title: Mouse ESC 46C replication analysis by repli-chip

city: Tallahassee

first: David

last: Gilbert

state: FL

address: 319 Stadium Drive

department: Biology

institute: Florida State University

type: Genome variation profiling by genome tiling array

summary: Mouse ESC 46C replication timing has been assessed by repli-chip.

overall_design: Genome-wide replication timing profiles assessed by array in mouse ESC 46C.

first: Juan

middle: C

last: Rivera-Mulia

first: Claudia

last: García


title: Targeting leukemia-initiating cells through pharmacologic inhibition of Pol I transcription to treat MLL-driven acute myeloid leukemia

city: Canberra

first: Nadine

last: Hein

state: ACT

address: Garran Road

department: CBTx

country: Australia

institute: JCSMR

summary: Transcriptome analysis revealed differential responses to Pol I inhibition in p53WT and p53null AML in vivo

overall_design: RNA-seq in p53WT and p53null MLL/ENL Nras AML 10 and 72 hr post treatment

first: Ross

last: Hannan


title: H2A.Z Regulates Astrocyte Proliferation and Differentiation via H3K56ac Modification

first: Jianwei

last: Jiao

state: Beijing

address: 1 Beichen West Road

department: State Key Laboratory of Reproductive Biology

country: China

institute: Institute of Zoology, Chinese Academy of Sciences

summary:
Purpose:To gain a deeper insight into how H2A.Z regulates astrocytogenesis, RNA-sequencing (RNA-seq) was performed to analyze the genome-wide changes by H2AZ deletion at E16.
Methods: Total RNA was extracted from E16 telencephalic tissue of H2A.ZcKO and H2A.Zfl/fl mice. Then total RNA was quality controlled and quantified using an Agilent 2100 Bioanalyzer. After converting to cDNA and building library, high-throughput sequencing was performed using the Illumina HiSeq 2500 platform in Annoroad Genomics.
Results: Approximately approximately one thousand transcripts showed differential expression between the H2A.Zfl/fl and H2A.ZcKO brain, with a fold change ≥1.5 and p value <0.05. Gene ontology (GO) analysis showed that the down-regulated genes were enriched in the terms related to regulation of glial cell differentiation and forebrain development, and cerebellum development. Up-regulated genes showed a significant enrichment of terms involved in negative regulation of cell differentiation. These results reflected the importance of H2A.Z in cortical development.
Conclusions: We conclude that RNA-seq based transcriptome characterization would provide a framework for understanding how H2A.Z gene contribute to cortical development.

overall_design: Total RNA was extracted from E16 telencephalic tissue of H2A.ZcKO and H2A.Zfl/fl mice and generated by deep sequencing, in triplicate, using Illumina HiSeq 2500.

first: Libo

last: Su


title: Repli-seq: genome-wide analysis of replication timing by next-generation sequencing

middle: M

country: USA

summary: Cycling cells duplicate their DNA content during S phase, following a defined program called replication timing (RT). Early and late replicating regions differ in terms of mutation rates, transcriptional activity, chromatin marks and sub-nuclear position. Moreover, RT is regulated during development and is altered in disease . Exploring mechanisms linking RT to other cellular processes in normal and diseased cells will be facilitated by rapid and robust methods with which to measure RT genome wide. Here, we describe a rapid, robust and relatively inexpensive protocol to analyse genome-wide RT by next-generation sequencing (NGS). This protocol yields highly reproducible results across laboratories and platforms. We also provide computational pipelines for analysis, parsing phased genomes using single nucleotide polymorphisms (SNP) for analyzing imprinted RT, and for direct comparison to Repli-chip data obtained by analyzing nascent DNA by microarrays.

overall_design: Repli-seq data of mouse 46C ES cells and ES cells derived from F1 CASTx129 hybrid mouse (clone F121-9).


title: Do natural trypanosome infections induce transcriptomic perturbations in field tsetse flies as do experimental infections in insectary-raised flies?

city: Montpellier

first: Anne

last: Geiger

address: TA A-17 / G Campus international de Baillarguet

institute: IRD-CIRAD

summary:
Purpose: The aim was, in the frame of an anti-vector strategy, to use such genes to control Human and/or animal trypanosomiasis. The present objective was to verify whether field tsetse fly gene expression was modified in response to natural infection with trypanosomes as it was when insectary-raised flies were experimentally infected.
Method: mRNA from 10 samples of Glossina palpalis (5 non-infected and 5 infected by Trypanosoma congolense s.l.) were sequenced on a high-output flow cell (400M clusters) using the NextSeq® 500/550 High Output v2 150 cycles kit (Illumina), in paired-end 75/75nt mode. The sequenced reads that passed quality filter were mapped onto the genome with the local alignment algorithm subread-align (Liao et al., 2013). Differential analysis was done with the SARTools R package (Varet et al., 2016), which runs separately DESeq2 (Love et al., 2014) and egdeR (Robinson et al., 2010). We identified orthologs between Glossina and Drosophila based on the identification of bidirectional best hits (BBH) using blastp (Altschul et al., 1997), and then made functional enrichment and pathway mapping of these DEGs.
Results: Using the RNA-seq approach, differentially expressed genes (DEGs) have been identified in infected versus non-infected tsetse flies, including down-regulated genes and up-regulated genes. Some of the genes, whether down- or up-regulated, were very highly differentially expressed. Down-regulated genes were mainly involved in transcription/translation processes, while up-regulated encoded genes governing amino acid and nucleotide biosynthesis pathways. The data on the molecular cross-talk between the host and the parasite (and the fly microbiome that is always present) recorded when using an experimental biological model have its counterpart in field flies which in turn validates the use of experimental host/parasite couples.
Conclusion: This study is the first evaluation of transcriptomic mechanisms related to infection in field tsetse flies. This opens up prospects for vector-based control strategies, and more precisely the blocking of transmission.

overall_design: mRNA from 10 samples of G. palpalis (5 non-infected and 5 infected by Trypanosoma congolense s.l.) were produced by the TruSeq mRNA-seq Stranded v2 Kit sample preparation (Illumina), according to manufacturer’s instructions.

first: J

last: Tsagmo Ngoune

first: F

last: Njiokou

first: B

last: Loriod

first: G

last: Kame Ngasse

first: N

last: Fernandez-Nunez

first: C

last: Rioualen

last: Van Helden

first: A


title: Cell cycle dynamics and complement expression distinguishes mature hematopoietic subsets arising from hemogenic endothelium

city: SAN FRANCISCO

first: Joan

last: Zape

state: CA

address: 555 Mission Bay Blvd South

summary: We report the transcriptional profiling of intra-aortic clusters (IAHCs) at different time points of the hemogenic window (E10.5 and E11.5) and at different phases of the cell cycle.

overall_design: Transcriptional profiles of intra-aortic clusters (IAHCs) at E10.5 and E11.5 during the G1 and S/G2/M phase of cell cycle were generated by sorting IAHC populations (SSEA-1-CD31+CD117+ ) from FucciRed (mKO2+)and FucciGreen (mAG+)animals. 100 base pair pair-end sequencing was performed in triplicate using the Illumina Hiseq2500.

first: Ann

last: Zovein


title: Transcriptome analysis of lymphoid progenitors in Zbtb1-deficient mice

city: Bethesda

first: Damian

last: Kovalovsky

address: 9000 Rockville Ave

institute: National Cancer Institute

summary: Purpose: Deficiency of Zbtb1 leads to the generation of myeloid cells from lymphoid progenitors when cultured in conditions that do not support myeloid development. We therefore analyzed how was the transcriptional signature altered by Zbtb1 deficiency in lymphoid progenitors ex vivo and after induction of a T-cell developmental program by co-culture with OP9-DL1 stroma cells.

overall_design: Sorted common lymphoid progenitors (CLP) and lymphoid primed multipotent progenitors (LMPP) were subjected to RNAseq (D0) or co-cultured with OP9-DL1 stroma for 24hs (D1) to induce T-cell differentiation in the presence of IL-7 and Flt3l. After co-culture, cells were sorted and subjected to RNAseq.

first: Xin

last: Cao


title: MicroRNA-mediated responses to long-term magnesium-deficiency in Citrus sinensis roots revealed by Illumina sequencing

city: Fuzhou

first: Chen

last: Lisong

address: Shangdian street NO.15

institute: Fujian Agriculture and Forestry University

summary:
Background: Magnesium (Mg)-deficiency occurs most frequently in strongly acidic, sandy soils. Citrus are grown mainly on acidic and strong acidic soils. Mg-deficiency causes poor fruit quality and low fruit yield in some Citrus orchards. For the first time, we investigated Mg-deficiency-responsive miRNAs in ‘Xuegan’ (Citrus sinensis) roots using Illumina sequencing in order to obtain some miRNAs presumably responsible for Citrus Mg-deficiency tolerance.
Results: We obtained 101 (69) miRNAs with increased (decreased) expression from Mg-starved roots. Our results suggested that the adaptation of Citrus roots to Mg-deficiency was related to the several aspects: (a) inhibiting root respiration and related gene expression via inducing miR158 and miR2919; (b) enhancing antioxidant system by down-regulating related miRNAs (miR780, miR6190, miR1044, miR5261 and miR1151) and the adaptation to low-phosphorus (miR6190); (c) activating transport-related genes by altering the expression of miR6190, miR6485, miR1044, miR5029 and miR3437; (d) elevating protein ubiquitination due to decreased expression levels of miR1044, miR5261, miR1151 and miR5029; (e) maintaining root growth by regulating miR5261, miR6485 and miR158 expression; and (f) triggering DNA repair (transcription regulation) by regulating miR5176 and miR6485 (miR6028, miR6190, miR6485, miR5621, miR160 and miR7708) expression. Mg-deficiency-responsive miRNAs involved in root signal transduction also had functions in Citrus Mg-deficiency tolerance.
Conclusions: We obtained several novel Mg-deficiency-responsive miRNAs (i.e., miR5261, miR158, miR6190, miR6485, miR1151 and miR1044) possibly contributing to Mg-deficiency tolerance. These results revealed some novel clues on the miRNA-mediated adaptation to nutrient deficiencies in higher plants.

overall_design: Two libraries were constructed from Citrus sinensis (L.) Osbeck roots subjected to 0(0 mM MgSO4，Mg-deficient，RD) or 1(Mg-sufficient,control, RS) mM MgSO4   for 16 weeks, and sequenced by Illumina sequencing. Size fractionated small RNAs (16-30 bp) from total RNA extracts was ligated to 5' and 3' adapters, and reverse transcribed. After PCR amplification the sample was subjected to Solexa sequencing. The clean tags were used for further analysis.

first: Li-Song

last: Chen

first: Wei-Wei

last: Liang

first: Jiang-Hao

last: Huang


title: Odd skipped-related 1 (Osr1) identifies a population of embryonic fibro-adipogenic progenitors regulating myogenesis during limb development

city: Paris

first: Mickael

last: Orgeur

address: 25-28 rue du Docteur Roux

institute: Institut Pasteur

summary: We sequenced total RNAs that were extracted from Osr1-expressing cells isolated by FACS-sorting from E13.5 limbs of two heterozygous (Osr1 GCE/+) and two homozygous (Osr1 GCE/GCE) mouse embryos.

overall_design: Gene expression profiling of Osr1-expressing cells at E13.5

first: Pedro

last: Vallecillo-Garcia

first: Stefan

middle: T

last: Börno

first: Bernd

last: Timmermann

first: Sigmar

last: Stricker


title: Identification of an LHX target cis-regulatory element through LHX6 ChIP-seq.

first: Juhee

last: Jeong

state: New York

address: 345 E 24th St

department: Basic sciences and Craniofacial development

institute: New York University College of Dentistry

summary: We performed chromatin immunoprecipitation (ChIP) followed by high-throughput sequencing (seq) from mouse E11.5 maxillary arches using anti-LHX6 antibody to identify LHX target cis-regulatory elements.

overall_design: Genome binding/occupancy profiling by high throughput sequencing using only wild-type E11.5 maxillary arch tissue.

first: Jeffry

last: Cesario


title: Continuous suppression of Mek1/2 impairs the developmental potential of mouse embryonic stem cells

status: Public on Jul 27 2017

city: Cambridge

first: Kendell

last: Clement

state: MA

address: 7 Divinity Ave.

institute: Harvard University

summary: Co-suppression of Gsk3 and Mek1/2 signaling (“2i culture”) in mouse embryonic stem cells (ESCs) induces a naïve state that more closely resembles the inner cell mass (ICM) of the pre-implantation embryo, including global DNA hypomethylation. While the ICM exists only transiently in vivo, it remains unclear how continuous suppression of Mek1/2 and Gsk3 signaling and associated hypomethylation affect the stability and functionality of ESCs in vitro. Here we show that, compared to ESCs maintained in conventional serum/LIF media, culturing male ESCs in 2i/LIF for >6 passages results in the erosion of genomic imprints and a concomitant impairment in the ability to generate adult, entirely ESC- derived mice (all-ESC mice). We further demonstrate that female ESCs cultured in serum/LIF phenocopy male ESCs cultured in 2i/LIF, including global hypomethylation, imprint erosion and impaired developmental potential. Unexpectedly, culture of female ESCs in serum/LIF and male ESCs in 2i/LIF for >16 passages causes recurrent chromosomal aberration that have been associated with a selective growth advantage. We provide evidence that inhibition of Mek1/2, but not Gsk3, is responsible for hypomethylation, imprint loss and chromosomal instability in ESCs, and identify alternative conditions that preserve epigenetic integrity. Taken together, our data suggest that while short-term suppression of Mek1/2 in ESCs captures an ICM-like epigenetic state, prolonged suppression results in irreversible changes that compromise developmental potential.

overall_design: Genetically matched mouse EGC and ESC lines were collected for DNA methylation analysis via mRRBS

first: Jiho

last: Choi

first: Alexander

last: Meissner

first: Konrad

last: Hochedlinger

first: Hongcang

last: Gu


title: Next Generation Sequencing Facilitates Quantitative Analysis of Wild Type and FXRalpha-/- testicular Transcriptomes

city: Clermont-Ferrand

first: David

middle: H

last: VOLLE

address: 28 place Henri Dunant

institute: INSERM

summary: The goals of this study are to define the putatitve impacts of FXRalpha deficiency in testicular physiology

overall_design: Testis  mRNA profiles of 10-day old wild type (WT) and FXralpha-/- mice were generated by deep sequencing, on 5 individual mice for each geneotype, sequence on flowcell HS168.

first: Emmanuelle

last: Martinot

last: Volle


title: Extended suppression of Mek1/2 impairs the developmental potential of mouse embryonic stem cells

city: Boston

address: 185 Cambridge St.

department: Molecular Biology

institute: Massachusetts General Hospital

summary: Simultaneous inhibition of Gsk3α/β and Mek1/2 activity in the presence of LIF (2i/L) induces a naïve state in mouse embryonic stem cells (ESCs) that resembles the inner cell mass (ICM) of the pre-implantation embryo. Since the ICM exists only transiently in vivo, it remains unclear how extended propagation of naïve ESCs in vitro affects their stability and functionality. Here we show that extended culture of male ESCs in 2i/L results in the progressive erosion of genomic imprints, loss of H2A.X binding, and accumulation of chromosomal aberrations. Consistent with these observations, we find that the developmental potential of ESCs cultured in 2i/L is impaired. Mechanistically, we demonstrate that Mek1/2 inhibition is predominantly responsible for these effects, in part through downregulation of DNA methyltransferases. Additionally, we demonstrate that female ESCs cultured in conventional serum/LIF media phenocopy male ESCs cultured in 2i/L, including the aforementioned epigenetic and developmental abnormalities. Finally, we show that replacement of the Mek1/2 inhibitor with a Src inhibitor preserves the epigenetic and genomic integrity as well as developmental potential of ESCs. Taken together, our data suggest that, while short-term suppression of Mek1/2 in ESCs maintains an ICM-like epigenetic state, prolonged suppression results in irreversible changes that compromise their developmental potential.

overall_design: Gene expression patterns of mouse ESC lines cultured in multiple culture conditions were compared using RNA-seq.

first: Andrej

last: Savol

first: Ruslan

last: Sadreyev


title: A transcriptionally permissive histone H3 lysine 9 methylation state enables RNAi-mediated heterochromatin assembly

first: Gloria

last: Jih

address: 240 Longwood Ave, LHRRB 517

department: Cell Biology

institute: Harvard Medical School

summary: Heterochromatic DNA domains play important roles in the regulation of gene expression and maintenance of genome stability by silencing repetitive DNA elements and transposons. In organisms ranging from fission yeast to mammals, heterochromatin assembly at DNA repeats involves the activity of small noncoding RNAs (sRNAs) associated with the RNA interference (RNAi) pathway. Typically, sRNAs, originating from long noncoding RNAs transcribed from DNA repeats, guide Argonaute-containing effector complexes to complementary nascent RNAs to initiate histone H3 lysine 9 di- and tri-methylation (H3K9me2 and H3K9me3, respectively) and heterochromatin formation. H3K9me is in turn required for recruitment of RNAi to chromatin and promotes sRNA generation. However, how heterochromatin formation, which silences transcription, can proceed by a co-transcriptional mechanism that also promotes sRNA generation remains paradoxical. Here, using Clr4, the fission yeast homolog of mammalian SUV39H H3K9 methyltransferases, we designed active site mutations, which allow H3K9me2 catalysis but block the transition to H3K9me3. We show that H3K9me2 defines a functionally distinct heterochromatin state that is sufficient for RNAi-dependent co-transcriptional gene silencing (CTGS). Unlike H3K9me3 domains, which are transcriptionally silent, H3K9me2 domains are transcriptionally active, contain modifications associated with euchromatic transcription, and couple RNAi-mediated transcript degradation to the establishment of H3K9me domains. The two H3K9me states recruit reader proteins with different efficiencies, explaining their different downstream silencing functions. Furthermore, transition from H3K9me2 to H3K9me3 is required for RNAi-independent epigenetic inheritance of H3K9me domains. Our findings demonstrate that H3K9me2 and H3K9me3 define functionally distinct heterochromatin states and uncover a mechanism for formation of transcriptionally permissive heterochromatin that is compatible with its broadly conserved role in RNAi-mediated genome defense.

overall_design: ChIP-seq analysis of histone modifications and chromatin associated proteins

first: Danesh

last: Moazed


city: Charlestown

first: Toshihiro

last: Shioda

address: Building 149 - 7th Floor, 13th Street

department: Center for Cancer Research

institute: MGH Cancer Center

summary: Simultaneous inhibition of Gsk3α/β and Mek1/2 activity in the presence of LIF (2i/L) induces a naïve state in mouse embryonic stem cells (ESCs) that resembles the inner cell mass (ICM) of the pre-implantation embryo. Since the ICM exists only transiently in vivo, it remains unclear how continuous propagation of naïve ESCs in vitro affects their stability and functionality. Here we show that extended culture of male ESCs in 2i/L results in the progressive erosion of genomic imprints, loss of H2A.X binding, and accumulation of chromosomal aberrations. Consistent with these observations, we find that the developmental potential of ESCs cultured in 2i/L is impaired. Mechanistically, we demonstrate that Mek1/2 inhibition is predominantly responsible for these effects, in part through downregulation of DNA methyltransferases. Additionally, we demonstrate that female ESCs cultured in conventional serum/LIF media phenocopy male ESCs cultured in 2i/L, including the aforementioned epigenetic and developmental abnormalities. Finally, we show that replacement of the Mek1/2 inhibitor with a Src inhibitor preserves the epigenetic and genomic integrity as well as developmental potential of ESCs. Taken together, our data suggest that, while suppression of Mek1/2 in ESCs maintains an ICM-like epigenetic state, continuous suppression results in irreversible changes that compromise their developmental potential.

overall_design: Profiling allelic expressions of imprinted genes in iPSCs cultured in serum/LIF and 2i/LIF.

first: Aaron

middle: J

last: Huebner

first: Junko

last: Odajima

first: Toshi


title: A novel requirement for Drosha in maintenance of mammalian CG methylation

city: Athens

first: Robert

last: Schmitz

state: GA

address: B416 Davison Life Sciences

institute: University of Georgia

summary: Using a protein interaction screen, we identify the Microprocessor component Drosha as a novel Dnmt1-interactor. Drosha-deficient ES cells display genomic hypomethylation which is not accounted for by changes in the levels of Dnmt proteins. Both genetic and transfection studies show that Drosha stimulates Dnmt1 methyltransferase activity. We identify two transcripts that are specifically upregulated in Drosha but not Dicer-deficient ES cells. Regions within these transcripts predicted to form stem-loop structures are processed by Microprocessor and can inhibit DNMT1-mediated methylation in vitro. Our results highlight Drosha as a novel regulator of mammalian DNA methylation and we propose that Drosha-mediated processing of RNA is necessary to ensure full Dnmt1 activity. This adds to the Drosha repertoire of non-miRNA dependent functions as well as implicating RNA in regulating Dnmt1 activity and correct levels of genomic methylation.

overall_design: Mapping of 5-methylcytosine in wild type and Drosha-deficient ES cells by MethylC-Seq and mapping of RNA in wild type and Drosha-deficient ES cells by RNA-Seq


title: Mapping open chromatin dynamics of cortical neuron differentiation from human induced pluripotent stem cells (iPSCs) by ATAC-Seq

city: Evanston

first: Winton

last: Moy

state: Illinois

address: 1001 University Place

department: Center Psychiatric Genetics

institute: Northshroe Univeristy Health Systems

summary: We aim to profile the dynamic changes of chromatin accessibility (openness) to transcription factors during cortical neuron differentiation from human iPSCs. We used ATAC-seq to map open chromatins in iPSCs, neural stem cells (NSCs) at day 27 and day 33 of neural induction (designated as iN-d30 for simplicity), and neurons at day 41 (iN-d41). We found that there were robust dynamic changes of open chromatins that are corresponding to cell stage-specific gene function both at genome-wide level and at individual loci of interest to neurodevelopment and psychiatric disorders, with NSC (iN-d30) gaining most (89%) of the neuron specific open chromatin peaks. Open chromatin peaks shared by different cell stages were overrepresented in core promoters, while the peaks specific to each cell stage or showing dynamic change of openness were enriched in introns and intergenic sequences. The dynamic change of open chromatins is orchestrated by specific sets of transcription factors (TFs) in each cell stage, providing epigenomic support the central role of NEUROD1 and NEUROG2 in cortical neuron differentiation.

overall_design: We mapped open chromatins by ATAC-seq in iPSCs derived from fibroblasts (GM01835), and iPSC-differentiated cortical neural stem cells (NSCs) at day 27 and day 33 of neural induction (designated as iN-d30 for simplicity), and neurons at day 41 (iN-d41). At each stage (iPSC, iN-d30 or iN-d41), we included two cell culture replicates. We used Hotspot to call open chromatin peaks (FDR<5%) in each cell stage. We used EdgR to quantify the dynamic change of open chromatin during cortical neuron differentiation from iPSCs.  In total, we generated 1 pool of 6 libraries and sequenced in a single lane on HiSeq 2500, and produced >120 million pair-end reads (2x50bp) with average quality scores above Q30.

first: Jubao

last: Duan


title: Ribosome profiling of control and leucine-starved SUM159 cells

city: Amsterdam

first: Koos

last: Rooijers

address: Plesmanlaan 121

department: Gene Regulation

institute: Netherlands Cancer Institute

summary: SUM159 cells were either grown in standard medium or in leucine-depleted medium, or in medium containing 20% of the standard concentration of leucine

overall_design: 1 replicate per condition

first: Fabricio

last: Loayza-Puch

first: Jelle

last: Zijlstra

first: Joachim

last: Oude Vrielink

first: Rui

last: Lopes

first: Alejandro

last: Ugalde

first: Axel

last: Bex

first: Reuven

last: Agami