title: Gene expression profiling of neural crest progenitor cultures derived from human embryonic stem cells carrying nonsense mutations in the Polycomb gene ASXL1 [HET]

summary: We observed impaired neural crest cell differentiation from human embryonic stem cells that harbor nonsense mutations in the Polycomb gene ASXL1. To investigate the underlying molecular mechanisms, we harvested neural crest differentiation cultures from wildtype and heterozygous ASXL1 mutant lines at day 7 of a neural crest differentiation protocol and performed total RNA sequencing using the Illumina HiSeq2500 system. Analyses of global transcriptomes revealed profound changes between wildtype and mutant cultures, and we identified downregulation of the neural crest transcription factor ZIC1.

overall_design: Global mRNA profiles of day 7 neural crest differentiation cultures derived from different human embryonic stem cell lines (three replicates of wildtype controls, and three replicates of a line harbouring a 500 bp deletion in the central part of ASXL1) were generated on the Illumina HiSeq 2500.

first: Miha

last: Modic

first: Micha

last: Drukker


title: A Brain-Melanocortin-Vagus axis mediates adipose tissue expansion independently of energy intake.

status: Public on Apr 18 2019

city: Cincinnati

first: Mario

last: Medvedovic

state: OH

address: 3223 Eden Av. ML 56

department: Department of Environmental Health

country: USA

institute: University of Cincinnati

type: Expression profiling by high throughput sequencing

summary: The melanocortin system is a brain circuit that influences energy balance by regulating energy intake and expenditure. In addition, the brain-melanocortin system controls adipose tissue metabolism to optimize fuel mobilization and storage. Specifically, increased brain-melanocortin signaling or negative energy balance promotes lipid mobilization by increasing Sympathetic Nervous input to adipose tissue. In contrast, calorie-independent mechanisms favoring energy storage are less understood. Here we demonstrate that obesogenic signals, including reduction of brain-melanocortin signaling or high-fat feeding, actively promote fat mass gain independently of caloric intake via efferent nerve fibers conveyed by the common hepatic branch of the vagus nerve. These signals promote adipose tissue expansion by activating lipogenic program and adipocyte and endothelial cell proliferation independently of insulin action or the sympathetic tone to adipose tissue.  These data reveal a novel physiological mechanism whereby the brain controls energy stores that may contribute to increased susceptibility to obesity.

overall_design: This experiment involves a 2 x 2 design (Subdiaphragmatic Vagotomy x icv infusion of SHU9119) with 4 replicates per group (16 samples total).

first: Diego

last: Perez-Tilve


title: Selective binding of the PHD6 finger of MLL4 to histone H4K16ac links MLL4 and MOF

status: Public on Apr 20 2019

city: Bethesda

first: Kai

last: Ge

state: MD

address: 50 South Dr Rm 4154

department: NIDDK

institute: NIH

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Histone methyltransferase MLL4 is centrally involved in transcriptional regulation and is often mutated in human diseases, including cancer and developmental disorders. MLL4 contains a catalytic SET domain that mono-methylates histone H3K4 and seven PHD fingers of unclear function. Here, we identify the PHD6 finger of MLL4 (MLL4-PHD6) as the first selective reader of the epigenetic modification H4K16ac. The solution NMR structure of MLL4-PHD6 in complex with a H4K16ac peptide along with binding and mutational analyses reveal unique mechanistic features underlying recognition of H4K16ac. Genomic studies show that one third of MLL4 chromatin binding sites overlap with H4K16ac-enriched regions in vivo and that MLL4 occupancy in a set of genomic targets depends on the acetyltransferase activity of MOF, a H4K16ac-specific acetyltransferase. The recognition of H4K16ac is conserved in the PHD7 finger of paralogous MLL3. Together, our findings highlight a novel acetyllysine reader and suggest that selective targeting of H4K16ac by MLL4 provides a direct functional link between MLL4, MOF and H4K16 acetylation.

overall_design: Histone H3K4 mono-methyltransferase MLL4 and histone modifications (histone H4K16ac) by ChIP-seq in brown preadipocytes in culture.

first: Jang

last: Younghoon

first: Lee

last: Ji-Eun

first: Ge

last: Kai


title: Gene expression profiling of neural crest progenitor cultures derived from human embryonic stem cells carrying nonsense mutations in the Polycomb gene ASXL1 [HOM]

status: Public on Apr 23 2019

city: München

first: Friederike

last: Matheus

address: Ingolstädter Landstrasse 1

department: Institute for Stem Cell Research

country: Germany

institute: Helmholtz Center Munich

summary: We observed impaired neural crest cell differentiation from human embryonic stem cells that harbor nonsense mutations in the Polycomb gene ASXL1. To investigate the underlying molecular mechanisms, we harvested neural crest differentiation cultures from wildtype and homozygous ASXL1 mutant lines at day 7 of a neural crest differentiation protocol and performed total RNA sequencing using the Illumina NextSeq system. Analyses of global transcriptomes revealed profound changes between wildtype and mutant cultures, and we identified downregulation of several genes of the neural crest regulatory network, most prominently of the transcription factor ZIC1.

overall_design: Global mRNA profiles of day 7 neural crest differentiation cultures derived from different human embryonic stem cell lines (three replicates of wildtype controls,  and 7 replicates from 3 different clones harbouring 500 bp deletions/inversions in the central part of ASXL1) were generated on the Illumina NextSeq system.

first: Lukas

last: Simon


title: Expression changes in early postnatal mouse sciatic nerve after Schwann cell-specific deletion of the Ep400 chromatin remodeler

city: Erlangen

first: Michael

last: Wegner

state: Bavaria

address: Fahrstr. 17

department: Institut of Biochemistry

summary: To address the role of INO80/SWR-type remodeling complexes, we deleted Ep400 in developing Schwann cells. Ep400 becomes important for proper Schwann cell development at differentiation. Its absence in Schwann cells leads to a peripheral neuropathy. RNA-Seq studies were performed on sciatic nerve from control mice and mice with a Schwann cell-specific deletion of Ep400  to analyze changes in gene expression. These revealed substantial changes in the expression of genes associated with immune and inflammatory response, myelination-associated lipid metabolism and developmental regulatory genes whose occurrence and action is normally limited to earlier stages of development.

overall_design: Schwann cell mRNA profiles of 9-day old control (ctrl) and Ep400 cko mice were generated using Illumina HiSeq 2500 platform.

first: Franziska

last: Fröb


title: Chronic Liver Disease in Humans Causes Expansion and Differentiation of Liver Lymphatic Endothelial Cells

city: Aurora

first: Matthew

middle: Alan

last: Burchill

state: Colorado

department: Department of Medicine

institute: University of Colorado - Anschutz Medical Campus

summary: Liver lymphatic vessels support liver function by draining interstitial fluid, cholesterol, fat, and immune cells for surveillance in the liver draining lymph node. Chronic liver disease is associated with increased inflammation and immune cell infiltrate. However, it is currently unknown if or how lymphatic vessels respond to increased inflammation and immune cell infiltrate in the liver during chronic disease. Here we demonstrate that lymphatic vessel abundance increases in patients with chronic liver disease and is associated with areas of fibrosis and immune cell infiltration. Using single-cell mRNA sequencing and multi-spectral immunofluorescence analysis we identified liver lymphatic endothelial cells and found that chronic liver disease results in lymphatic endothelial cells (LECs) that are in active cell cycle with increased expression of CCL21. Additionally, we found that LECs from patients with NASH adopt a transcriptional program associated with increased IL13 signaling. Moreover, we found that oxidized low density lipoprotein, associated with NASH pathogenesis, induced the transcription and protein production of IL13 in LECs both in vitro and in a mouse model. Finally, we show that oxidized low density lipoprotein reduced the transcription of PROX1 and decreased lymphatic stability. Together these data indicate that LECs are active participants in the liver, expanding in an attempt to maintain tissue homeostasis. However, when inflammatory signals, such as oxidized low density lipoprotein are increased, as in NASH, lymphatic function declines and liver homeostasis is impeded.

overall_design: Single-cell RNA-seq analysis of human liver lymphatic endothelial cells.

first: Beth

middle: A

last: Tamburini

first: Jeffrey

middle: M

last: Finlon

first: Austin

middle: E

last: Gillen

middle: S

last: Kriss

first: Kent

last: Riemondy

first: Rui

last: Fu

first: Ronald

last: Schuyler

first: Jay

middle: R

last: Hesselberth

first: Hugo

last: Rosen


title: The NuRD complex suppresses transcriptional noise during lineage commitment [RNA-Seq]

status: Public on Apr 17 2019

city: St Louis

first: Sabine

last: Dietmann

state: MO

address: 4565 McKinley Avenue

institute: Washington University School of Medicine

summary: Here we set out to define the biochemical and functional diversity encoded by one such group of proteins within the mammalian Nucleosome Remodelling and Deacetylation (NuRD) complex: Mta1, Mta2 and Mta3. We find that, in contrast to what has been described in somatic cells, MTA proteins are not mutually exclusive within ES cell NuRD and, despite subtle differences in chromatin binding and biochemical interactions, serve largely redundant functions. ES cells lacking all three MTA proteins represent a complete NuRD null and are viable, allowing us to identify a highly previously unreported function for NuRD in reducing transcriptional noise, which is essential for maintaining a proper differentiation trajectory during early stages of lineage commitment.

overall_design: comparison of gene expresison patterns in wild type and mutant cells in self-renewing and differentiating stem cells

first: Brian

last: Hendrich

last: Barber


title: The NuRD complex suppresses transcriptional noise during lineage commitment [ChIP-Seq]

overall_design: comparison of gene expression patterns in wild type and mutant cells in self-renewing and differentiating stem cells


title: The NuRD complex suppresses transcriptional noise during lineage commitment

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

overall_design: Refer to individual Series


title: Paternal tobacco smoke correlated to offspring asthma and prenatal epigenetic programming

status: Public on Apr 13 2019

city: Kaohsiung

first: Jen-Chieh

last: Chang

address: 123 Ta-Pei Road, Niaosong

department: Department of Medical Research

country: Taiwan

institute: Kaohsiung Chang Gung Memorial Hospital

summary: To investigate whether paternal tobacco smoke (PTS) was associated with prenatal epigenetic programming of CG site methylation, 20 cord blood DNA samples from newborns with and without PTS were subjected to microarray assay of 1,505 CG loci in 807 genes by Illumina GoldenGate® technology bead system to obtain high-throughput DNA Methylation profiles. Samples included 14 newborns without PTS exposure, and 6 newborns with PTS exposure.

overall_design: 20 umbilical cord blood samples were subjected to DNA extraction, following bisulfite conversion and hybridized to the Illumina GoldenGate Methylation Cancer Panel I array.

first: Chih-Chiang

last: Wu

first: Kuender

middle: D

last: Yang


title: A bioactive mammalian disaccharide associated with autoimmunity activates a STING-TBK1-dependent immune response

status: Public on Apr 16 2019

city: Dallas

last: Yan

state: TX

address: 6000 Harry Hines Blvd

institute: UT Southwestern Medical Center

summary: DNase III, also known as TREX1, is a 314 amino acid endoplasmic reticulum (ER) tail-anchored 3’ exonuclease where the N-terminal region contains the DNase domain and the C-terminal end controls TREX1 localization to the surface of the ER. Disease mutations in the C-terminal region alters TREX1 ability to interact with the oligosaccharyltransferase (OST) subunits Ribophorin 1 (RPN1) and DDOST leading to rapid hydrolysis of lipid-linked oligosaccharides (LLOs) into bioactive free oligosaccharides (fOS) in a switch-like manner. Bioassay-guided fractionation of the fOS pool revealed that the structure responsible for the bioactivity is a mannose (Man) b1-4 N-acetylglucosamine (GlcNAc) disaccharide. The bioactive disaccharide is produced from OST’s hydrolyzed LLOs in the cytoplasm and activates a STING-TBK1 dependent immune signal that leads to the upregulation of interferon-stimulated genes (ISGs) and chemokine genes.

overall_design: Examine the trancriptome of RAW 264.7 cells when treated with 10uM of Trex1-/- fOS or the ManGn disaccharide for 24 hours.

first: Charles

last: Fermaintt


title: Atypical function of a centrosomal module in WNT signalling drives contextual cancer cell motility

status: Public on Apr 26 2019

city: Toronto

first: Jeff

last: Wrana

address: 600 University Avenue

country: Canada

institute: Lunenfeld-Tanenbaum Research Institute

summary: Centrosomes control cell motility, polarity and migration that are thought to be mediated by their microtubule-organizing capacity. In this study we demonstrate that WNT signalling drives a distinct form of non-directional cell motility that requires a key centrosome module, but not microtubules or centrosomes.  Upon exosome mobilization of Planar Cell Polarity proteins, we show that DVL2 orchestrates recruitment of a CEP192-PLK4/AURKB complex to the cell cortex where PLK4/AURKB act redundantly to drive protrusive activity and cell motility. This is mediated by coordination of formin-dependent actin remodelling through displacement of cortically localized DAAM1 for DAAM2.  Furthermore, abnormal expression of PLK4, AURKB and DAAM1 is associated with poor outcomes in breast and bladder cancers.  Thus, a centrosomal module plays an atypical function in WNT signalling and actin nucleation that is critical for cancer cell motility and is associated with more aggressive cancers.  These studies have broad implications in how contextual signalling controls distinct modes of cell migration.

overall_design: Total RNA profilling of 158 archival bladder cancer samples and comparison between 107 high grade and 51 low grade samples. This study involves 120 Samples. Thirty eight Samples were reanalyzed from previously submitted GSE59483.

first: Jess

last: Shen

first: Kin

last: Chan

first: Ying

middle: Y

last: Zhang

first: Yu

last: Liu

middle: L


title: In utero lead (Pb) exposure and neuron-specific DNA methylation in mice

status: Public on Apr 12 2019

city: Ann Arbor

first: Kelly

last: Bakulski

state: MI

address: 1415 Washington Heights

department: Department of Epidemiology

institute: University of Michigan

type: Methylation profiling by genome tiling array

summary: Lead (Pb) exposure is associated with a wide range of neurological deficits. Epigenetic changes, such as DNA methylation, may be impacted by environmental exposures and can affect neurodevelopmental outcomes over the life-course.  Mating mice were obtained from a C57BL/6J background agouti Avy strain. Virgin dams (a/a) were randomly assigned exposure to 2.1 ppm (low) or 32 ppm (high) Pb-acetate through the drinking water, started two weeks prior to mating with viable yellow agouti male mice (Avy/a) and continued throughout gestation and three weeks after birth. At 10 months of age, we separated brain NeuN+ (a marker for neuronal nuclei) neuronal nuclei from NeuN  non-neuronal nuclei in mice. We investigated neuron specific genome-wide promoter DNA methylation associated with early-life Pb exposures using the Roche NimbleGen Mouse DNA methylation 3x720K CpG island Promoter Array.

overall_design: Sample pooling was necessary to reach the amount of genomic DNA necessary to perform hybridization. Three pooled samples were used for each control, low, and high Pb exposure group, resulting in nine total pooled samples. Each pool consisted of DNA from at least two isogenic mice.

first: John

middle: F

last: Dou

first: Zishaan

last: Farooqui

first: Christopher

last: Faulk

first: Amanda

middle: K

last: Barks

first: Tamara

last: Jones

first: Dana

middle: C

last: Dolinoy


title: Acute Effects of Single Doses of Bonito Fish Peptides and Vitamin D on Whole Blood Gene Expression Levels

status: Public on Apr 11 2019

city: Quebec

first: Frederic

last: Guenard

address: 2440, Hochelage blvd

institute: Laval University

summary: Fish contains high quality proteins and essential nutrients including vitamin D (VitD3). Fish peptide consumption can lower cardiovascular disease (CVD) risk factors and studies showed an association between VitD3 deficiency, CVD and CVD risk factors such as diabetes. This study investigated acute effects of a single dose of VitD3, bonito fish peptide hydrolysate (BPH), or a combination of both on CVD risk factors and whole blood gene expression levels. A randomized, crossover, placebo controlled trial was conducted in 22 adults. They ingested, in random order and at 7-day intervals, 1000 IU of VitD3, 3g of BPH, a combination of both, or a placebo. A 180 min oral glucose tolerance test was done. Differences in whole-genome expression levels after vs. before each supplementation were computed for 18 subjects. We observed that 16, 1 and 5 transcripts were differentially expressed post- vs. pre-ingestion for VitD3, BPH or VitD3+BPH treatments, respectively. VitD3-containing treatments affected the SLC25A20 gene involved in fatty acid oxidation, various transcription factors and genes related to glucose metabolism. These results suggest that VitD3 rapidly modulates genes related to CVD risk factors in blood while BPH seems to moderately modulate gene expression levels.

overall_design: A randomized, crossover, placebo controlled trial was conducted on gene expression levels in 18 adults. They ingested, in random order and at 7-day intervals, 1000 IU of VitD3, 3g of bonito fish peptide hydrolysate, a combination of both, or a placebo.

first: Frédéric

last: Guénard

first: Hélène

last: Jacques

first: Claudia

last: Gagnon

first: André

last: Marette

first: Marie-Claude

last: Vohl


title: Common regulatory pathways mediate activity of microRNAs inducing cardiomyocyte proliferation

city: Trieste

first: Danilo

last: Licastro

department: Genomica Funzionale

country: Italy

institute: CBM

summary: Loss of functional cardiomyocytes is a major determinant of heart failure after myocardial infarction. Previous high throughput screening studies have identified a series of microRNAs that induce cardiomyocyte proliferation and stimulate cardiac regeneration after myocardial infarction. Here we investigate the mechanism of action of the top ten most effective of these miRNAs. Analysis of the transcriptional profile of miRNA-treated cardiomyocytes revealed an involvement of the Hippo-YAP pathway in their action. All the investigated miRNAs activated YAP-mediated transcription, nuclear localization of active YAP and increased expression of YAP responsive genes. In particular, miR-199a-3p, one of the most effective miRNAs, directly downregulated two mRNA targets impinging on the Hippo pathway, the upstream inhibitory kinase TAOK1and the E3 ubiquitin ligase destroying YAP, b-TrCP. Most of the miRNAs that promoted proliferation also modulated the dynamics of the cardiomyocyte actin cytoskeleton. In particular, four miRNAs targeted Cofilin2 and increased the ratio between polymerized F-actin and monomeric G-actin. Cells treated with the most effective miRNAs (including miR-199a-3p) were round-shaped, with formation of gross bundles of actin fibers at the cytoplasm periphery. During mitosis, these miRNA-treated cardiomyocytes displayed disruption of the sarcomeric architecture. Downregulation of Cofilin2 itself was sufficient to promote cardiomyocyte proliferation, activate nuclear translocation of YAP and stimulate transcription of TEAD-responsive genes. Inhibition of F-actin polymerization decreased YAP activation. Collectively, these results indicate that activation of YAP and modulation of the acting cytoskeleton are major components of the pro-proliferative effect of miR-199a-3p and other miRNAs inducing cardiomyocyte proliferation

overall_design: Cardiomyocyte transcriptomic  profiles of 0-day old wild type (WT) rats were generated by deep sequencing using Illumina Hiseq 2000

first: Consuelo

last: Torrini

first: Mauro

last: Giacca


title: Altered genomic occupancy of H2A.Z in early postnatal mouse sciatic nerve after Schwann cell-specific deletion of the Ep400 chromatin remodeler

first: Michael

last: Wegner

summary: To address the role of INO80/SWR-type remodeling complexes, we deleted Ep400 in developing Schwann cells. Ep400 is involved in exchanging H2A-containing standard with H2A.Z-containing nucleosomes. Ep400 becomes important for proper Schwann cell development at differentiation. Its absence in Schwann cells leads to a peripheral neuropathy. ChIP-Seq studies were performed with H2A.Z-specific antibodies on sciatic nerve from control mice and mice with a Schwann cell-specific deletion of Ep400  to determine the genomic occupancy of this histone variant. One group of genes with strongly altered H2A.Z occupancy around the transcriptional start site consisted of developmental regulatory genes whose occurrence and action is normally limited to earlier stages of development.

overall_design: Examination of histone isoform H2A.Z in sciatic nerves of control and Ep400 cko mice.

first: Franziska

last: Fröb


title: ChiCMaxima: a robust and simple pipeline for detection and visualization of chromatin looping in Capture Hi-C (4C validation)

city: Illkirch

first: Tom

last: Sexton

address: 1 rue Laurent Fries

country: France

institute: IGBMC (Institute of Genetics and Molecular and Cellular Biology)

summary: Capture Hi-C (CHi-C) is a new technique for assessing genome organization based on chromosome conformation capture coupled to oligonucleotide capture of regions of interest, such as gene promoters. Chromatin loop detection is challenging because existing Hi-C/4C-like tools, which make different assumptions about the technical biases presented, are often unsuitable. We describe a new approach, ChiCMaxima, which uses local maxima combined with limited filtering to detect DNA looping interactions, integrating information from biological replicates. ChiCMaxima shows more stringency and robustness compared to previously developed tools. The tool includes a GUI browser for flexible visualization of CHi-C profiles alongside epigenomic tracks. The results of the analysis on existing mouse ES Capture Hi-C data (ArrayExpress E-MTAB-2414) were validated by two 4C-seq experiments, submitted here.

overall_design: Single 4C-seq experiment on mouse ES cells with two baits


title: High-throughput single-cell ChIP-seq identifies heterogeneity of chromatin states in breast cancer

city: Paris

first: Kevin

last: Grosselin

address: 10 rue Vauquelin

department: CBI

institute: ESPCI Paris

summary: The dynamic nature of chromatin and transcription plays a critical role in normal differentiation and is expected to contribute to tumor evolution. Studying chromatin heterogeneity with single-cell resolution is mandatory to understand the role of epigenetic plasticity in cancer. Here, we describe a droplet microfluidics system that enables the profiling of chromatin marks of individual cells with an average coverage of up to 10,000 loci per cell. In patient-derived xenograft (PDX) models of breast cancer with acquired drug resistance, single-cell chromatin immunoprecipitation followed by sequencing (scChIP-seq) identified rare populations of cells in untreated, drug-sensitive tumors with chromatin features that match those of resistant cells. Loci depleted for transcriptional repressive mark H3K27me3 included genes know to promote resistance to chemotherapy, highlighting the potential to discover new drug targets.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Single-cell ChIP-seq and single-cell RNA-seq in patient-derived xenograft models of breast cancer with acquired drug resistance

first: Annabelle

last: Gerard

first: Andrew

last: Griffiths

first: Céline

last: Vallot


title: Transcriptomic analysis of cultured isogenic myotonic dystrophy type 1 myoblasts with and without the DMPK CTG repeat

city: Nijmegen

first: Remco

last: van Cruchten

address: Geert Grooteplein Zuid 26-28

department: Cell Biology

country: Netherlands

institute: Radboud Institute for Molecular Life Sciences

summary: RNA-seq on proliferating myoblasts of the DM11 line carrying 13 and 2600 CTG repeats in the DMPK gene, compared to their genome-edited counterparts without both the 13 and 2600 CTG repeat or just lacking the 2600 CTG repeat.

overall_design: DM1 patient-derived and immortalized myoblasts (DM11) were genome-edited using CRISPR-Cas9 to excise the CTG repeat from the DMPK gene, thereby creating isogenic healthy controls for the DM1 cell line. After the CRISPR procedure, cells were clonally expanded and genetically characterized. We performed RNAseq on polyA selected RNA of one myoblast clone of which exclusively the expanded CTG repeat was excised (1E6), three clones of which the CTG repeat of both alleles was excised (3E3, 3B11 and 4A3) and three clones of which the repeat was still present after this procedure (4F9, EA11 and EA7), along with the DM11 source cell line (DM11 parental). This procedure and initial characterisation is described in Van Agtmaal et al. (2017) Molecular Therapy vol. 25 issue 1 p24-43

first: L

last: André

first: R

middle: T

first: D

middle: G

last: Wansink


title: MULTI-seq: Universal sample multiplexing for single-cell RNA sequencing using lipid-tagged indices

city: San Francisco

first: Christopher

last: McGinnis

state: CA

address: 600 16th Street

department: Pharmaceutical Chemistry

institute: University of California, San Francisco

summary: We report the development of MULTI-seq: a scRNA-seq and snRNA-seq sample multiplexing approach using lipid- or cholesterol-modified oligonucleotides. We demonstrate MULTI-seq utility in the following scRNA-seq and snRNA-seq contexts: (1) Live HEKs and HMECs with and without stimulation with TGF-β, (2) Purified nuclei from HEKs, MEFs, and Jurkats stimulated for 0-24 hrs with ionomycin and PMA, (3) 96 unique HMEC cultures (with one technical replicate), and (4) human metastases and mouse immune cells isolated from cryopreserved primary tissue samples dissected from a patient-derived xenograft mouse model of triple-negative breast cancer at progressive stages of metastasis to the lung.

overall_design: Single-cell or single-nucleus gene expression profiles for (1) HEKs and HMECs, (2) HEKs, MEFs, and Jurkats, (3) HMECs in distinct culture conditions, and (4) Human metastases and mouse immune cells from PDX mice.

first: Zev

middle: J

last: Gartner


title: Role of microRNA in CB1 antagonist-mediated regulation of adipose tissue macrophages during diet-induced obesity.

status: Public on Apr 04 2019

city: Columbia

first: Prakash

last: Nagarkatti

state: SC

address: 6439 Garners Ferry Road

institute: University of South Carolina School of Medicine

summary: We used microRNA microarrays to identify dysregulated microRNAs in macrophages isolated from epididymal adipose tissue.

overall_design: F4/80+ adipose tissue macrophages (ATMs) were isolated from epididymal fat of mice following diet-induced obesity intervention with CB1 receptor antagonists.

first: Pegah

last: Mehrpouya-Bahrami

first: Kathryn

last: Miranda

first: Mitzi


title: The HRP3 PWWP domain recognizes the minor groove of double-stranded DNA and recruits HRP3 to chromatin

city: Marburg

first: Robert

last: Liefke

state: Hessen

address: Hans-Meerwein-Str. 2

department: Institute of Molecular Biology and Tumor Research (IMT)

institute: Philipps University of Marburg

summary:
HDGF-related protein 3 (HRP3, also known as HDGFL3) belongs to the family
of HDGF-related proteins (HRPs) and plays an essential role in hepatocellular
carcinoma pathogenesis. All HRPs have a PWWP domain at the N-terminus
that binds both histone and DNA substrates. Despite previous advances in
PWWP domains, the molecular basis by which HRP3 interacts with chromatin
is unclear. In this study, we solved the crystal structures of the HRP3 PWWP
domain in complex with various double-stranded DNAs with/without bound
histone peptides. We found that HRP3 PWWP bound to the phosphate
backbone of the DNA minor groove and showed a preference for DNA
molecules bearing a narrow minor groove width. In addition, HRP3 PWWP
preferentially bound to histone peptides bearing the H3K36me3/2 modification.
HRP3 PWWP uses two adjacent surfaces to bind both DNA and histone
substrates simultaneously, enabling us to generate a model illustrating the
recruitment of PWWP to H3K36me3-containing nucleosomes. Cell-based
analysis indicated that both DNA and histone binding by the HRP3 PWWP
domain are important for HRP3 recruitment to chromatin in vivo. Our work
establishes that HRP3 PWWP is a new family of minor groove-specific DNA
binding proteins, which improves our understanding of HRP3 and other
PWWP domain-containing proteins.

overall_design: HRP3 (HDGFL3) ChIP-Seq after ectopic expression of GFP-tagged HRP3 proteins. Two replicates were performed.

first: Wei

last: Tian

first: Peiqiang

last: Yan

first: Ning

last: Xu

first: Arghya

last: Chakravorty

first: Qiaoran

last: Xi

first: Zhanxin

last: Wang


title: Global donor and acceptor splicing site kinetics in human cells

city: Göttingen

first: Livia

last: Caizzi

address: Am Fassberg 11

department: Molecular Biology

institute: Max Planck Institute For Biophysical Chemistry

summary: Transient transcriptome sequencing and kinetic modeling suggest how the kinetics and yield of RNA splicing are encoded in the human genome.

overall_design: In vivo kinetics (TT-seq) to quantify donor and acceptor splice-sites metabolic rates

first: Leonhard

last: Wachutka

first: Julien

last: Gagneur

first: Patrick

last: Cramer


title: Myc and Dnmt1 impede the pluripotent to totipotent state transition in embryonic stem cell

city: Doha

first: Mohamed Nadhir

last: Djekidel

state: Doha

address: Gharafa

department: Translational Medicine Department

country: Qatar

institute: Sidra Medicine

type: Methylation profiling by high throughput sequencing


title: Genome wide VDR binding sites in RWPE1 human prostate epithelial cells

status: Public on Apr 10 2019

city: Austin

first: James

last: Fleet

address: 1400 Barbara Jordan Blvd

department: Nutritional Sciences

institute: University of Texas at Austin

summary: Signaling through the vitamin D receptor (VDR) has been proposed to suppress the development of epithelial cancers, including prostate cancer.   We conducted ChIP-seq to identify the VDR binding sites in the genome of the prostate epithelial cell line, RWPE1.  This analysis reveals a large number of VDR binding sites in both control cells and in cells treated with 10 nM 1,25 dihydroxyvitamin D3 for 3 hours.   These peaks are associated with genes controlling a wide variety of cellular functions.

overall_design: 80% cultures of RWPE1 cells were treated with 10 nM 1,25 dihydroxyvitamin D3 (1,25(OH)2D) or ethanol vehicle for 3 h. Chromatin immunoprecipitation (ChIP) was conducted using either normal rabbit IgG polyclonal antibody as a control (12-370, Millipore Sigma, Burlington, MA) or anti-VDR polyclonal antibody (C-20, sc-1008, Santa Cruz Biotechnology, Inc., Dallas, TX).  Prior to sending samples for next generation sequencing, we conducted ChIP-PCR to detect ligand-inducible VDR binding to the -252 to -51 bp region of the CYP24A1 gene promoter and lack of VDR binding to the GAPDH gene promoter (42).   Three biological replicates that met this criterion were pooled by treatment to generate three samples (control treated-VDR immunoprecipitation, 1,25(OH)2D treated-VDR immunoprecipitation, pooled control and 1,25(OH)2D treated-IgG immunoprecipitation).  The ChIP procedure was repeated on these pooled samples and the resultant immunoprecipitations were sent to the Michael Smith Genome Science Centre (Vancouver, Canada) where 100-300 bp fragments were isolated from 12% polyacrylamide gels for library construction and 36 bp single-end reads were generated by Illumina sequencing. Each sample was sequenced in two lanes and the data for the two lanes were pooled for final analysis.


title: Pathologically distinct fibroblast subsets drive inflammation and tissue damage in arthritis

status: Public on Apr 09 2019

city: Birmingham

first: Adam

last: Croft

address: Edgbaston

country: United Kingdom

institute: University of Birmingham

summary: The identification of lymphocyte subsets with non-overlapping effector functions has been pivotal to the development of targeted therapies in immune mediated inflammatory diseases (IMIDs). Yet, despite their key role in disease, it remains unclear whether fibroblast subclasses with non-overlapping functions also exist and are responsible for the wide variety of tissue driven pathologies observed in IMIDs such as inflammation and damage . Here we identify and describe the biology of distinct subsets of fibroblasts responsible for mediating either inflammation or tissue damage in arthritis. We show that deletion of FAPa+ synovial cells suppressed both inflammation and bone erosions in murine models of resolving and persistent arthritis. Single cell transcriptional analysis identified two distinct fibroblast subsets: FAPa+ THY1+ immune effector fibroblasts located in the synovial sub-lining, and FAPa+ THY1- destructive fibroblasts restricted to the synovial lining. When adoptively transferred into the joint, FAPa+ THY1- fibroblasts selectively mediate bone and cartilage damage with little effect on inflammation whereas transfer of FAPa+ THY1+ fibroblasts  resulted in a more severe and persistent inflammatory arthritis, with minimal effect on bone and cartilage. Our findings describing anatomically discrete, functionally distinct fibroblast subsets with non-overlapping functions have important implications for cell based therapies aimed at modulating  inflammation and  tissue damage.

overall_design: Serum transfer inflammatory arthritis (STIA) was induced by intravenous injection of 100 µl of arthritogenic KRN serum into naive C57BL/6 mice. From these mice, CD45-ve live Podoplanin (PDPN)+ synovial cells from hind limb joints were sort purified at day 9 (n=3 biological replicates, each comprised of cells from the joints of three animals). Individuals subsets of CD45- PDPN+ cells were further sort puified in the following populations FAP⍺+ THY1- (n=10 mice); FAP⍺+ THY+ (n=13 mice); FAP⍺- THY1+ (n=7 mice) and FAP⍺- THY1- (n=5 mice). Small bulk RNA sequencing was performed on each of these cell populations with each sample representing a biological replicate comprising of cells isolated from the synovial joints of both hind limbs from a single mouse).

middle: P

first: Chris

last: Buckley


title: ChIP-Seq analysis of SMARCA4 binding sites in human NSC

city: Dresden

first: Maciej

last: Paszkowski-Rogacz

address: Tatzberg 47/49

department: UCC / Medical Faculty and University Hospital Carl Gustav Carus

institute: Technische Universität Dresden

summary:
We performed large-scale, pooled shRNA screens targeting 538 epigenetic regulators in genetically identical human stem cells, where each gene was targeted on avarage by 12 shRNAs. CD34+ HSPCs, used as a starting point of the study, were reprogrammed to iPSCs, which were differentiated to NSC. These cells were transduced with the pooled shRNA library and samples were collected 2 days post transduction (dpt), 12 dpt, and 22dpt - allowing 5 cell doublings. PCR amplified gDNA carrying the shRNA barcodes was sequenced on a HiSeq2500 Illumina sequencer. Analysis of NGS results, based on the change of shRNA representation (read count) and represented by z-scores, identified SMARCA4 as a differential regulator (SMARCA4 shRNAs were depleted in HSPC vs enriched in NSC sreens).
We performed RNA-seq experiments as part of the characterization dataset for the isogenic HSPC, HSPC-derived iPSC, and iPSC-derived NSC. RNA samples from CD34+ HSPC were harvested in parallel to gDNA isolation on 12- and 22-days post transduction, during the RNAi screen. In addition, untreated iPSC and NSC RNA samples were sequenced as part of their characterization. RNA samples were sequenced with poly-dT enrichment using single-end sequencing. Each sample was sequenced at a depth of yielding at least 20 million reads.
We performed a ChIP-seq analysis of the SMARCA4 binding sites in hNSC, which we identified in the large-scale RNAi screens as a candidate, involved in regulation of self-renewal and differentiation. The pull-down experiment was performed on the untreated NSCs. We show that SMARCA4 has a prominent binding tendency for the TSS throughout the genome, suggesting a role for SMARCA4 in transcriptional regulation.

overall_design: Pooled shRNA screen on genetically identical CD34+ HSPC and NSC, RNA-Seq transcriptome profiling of isogenic CD34+ HSPC, iPSC, and NSC and examination of SMARCA4 binding sites in an iPSC-derived NSC line by ChIP-Seq.

first: Ceren

last: Güneş


title: Gene-Centric Functional Dissection of Human Genetic Variation Uncovers Regulators of Hematopoiesis

city: Boston

first: Vijay

last: Sankaran

state: MA

address: 300 Longwood Ave

institute: Boston Children's Hospital

summary: Follow-up work was performed for SF3A2, a gene among the hits identified in a red blood cell trait GWAS-informed shRNA screen. Differential splicing effects were assayed to investigate resulting effects on the differentiating erythroid cell spliceome and explore potential modifier relationships with other known splicing defects associated with human disease.

overall_design: Examination of differential splicing events resulting from knockdown of splicing factor 3a subunit 2 (SF3A2) in three unique donor CD34+ cells populations undergoing erythroid differentiation. Two shRNA targeting SF3A2 were tested, along with a negative control shRNA targeting luciferase (which should not be expressed) using paired-end sequencing.

first: Sean

last: McFarland

first: Satish

last: Nandakumar

first: Laura

last: Mateyka


title: A Molecular Roadmap of the Phased Progression of Pluripotency

city: RTP

first: Raja

last: Jothi

state: NC

address: 111 TW Alexander Drive; A314

department: National Institute of Environmental Health Sciences

institute: National Institutes of Health

summary: Pluripotency is highly dynamic and progresses through a continuum of pluripotent stem cell states. The two states that bookend the pluripotency continuum, naïve and primed, are well characterized, but our understanding of the intermediate states and the transitions between them remain incomplete. To address this gap in knowledge, using a previously validated system to induce naive mouse embryonic stem cells (ESCs) to post-implantation pre-grastrulating epiblast-like cells (EpiLCs), we generated comprehensive high-temporal-resolution maps of the proteome, phosphoproteome, transcriptome, and epigenome of ESCs transitioning from naïve to primed pluripotency. Our data provide a comprehensive molecular description of the phased progression of pluripotency and a foundation for investigating mechanisms that regulate pluripotent state transitions.

overall_design: RNA-Seq and ChIP-Seq experiments at various time-points during the 72-hour ESC to EpiLC transition

first: P

last: Yang

first: S

last: Humphrey

last: Cinghu

first: R

last: Pathania

first: A

last: Oldfield

first: D

last: Kumar

first: K

last: Perera

first: J

last: James

first: M

last: Mann


title: ATAC-seq on human pancreatic islets

status: Public on Apr 06 2019

city: Malmö

first: Charlotte

last: Ling

address: Jan Waldenströms gata, 35

department: Department of Clinical Sciences

country: Sweden

institute: Lund University

summary: We have studied the impact of T2D on open chromatin in human pancreatic islets. We used assay for transposase-accessible chromatin using sequencing (ATAC-seq) to profile open chromatin in islets from T2D and non-diabetic  donors. We identified ATAC-seq peaks representing open chromatin regions  in islets of non-diabetic and diabetic donors. The majority of ATAC-seq peaks mapped near transcription start sites. Additionally, peaks were enriched in enhancer regions and in regions where islet-specific TFs bind. Islet ATAC-seq peaks overlap with SNPs associated with T2D and with additional SNPs in LD with known T2D SNPs. There was enrichment of open chromatin regions near highly expressed genes in human islets.

overall_design: Maps of open chromatin in human pancreatic islets from non-diabetic donors and donors with type 2 diabetes were generated by using ATAC-seq

first: Madhusudhan

last: Bysani

first: Rasmus

last: Agren

first: Cajsa

last: Davegårdh

first: Petr

last: Volkov

first: Tina

last: Rönn

first: Per

last: Unneberg

first: Karl

last: Bacos


title: Fgf10-Fgfr2b signaling generates basal cells and drives alveolar epithelial regeneration by bronchial epithelial stem cells after lung injury

status: Public on Mar 30 2019

city: Rochester

first: Stijn

last: De Langhe

state: Minnesota

address: 200 First street SW

department: Medicine

institute: Mayo Clinic Rochester Minnesota

summary: Idiopathic pulmonary fibrosis (IPF) is a common form of interstitial lung disease (ILD) resulting in alveolar remodeling and progressive loss of pulmonary function due to chronic alveolar injury and failure to regenerate the respiratory epithelium. Histologically, fibrotic lesions and honeycomb structures expressing atypical proximal airway epithelial markers replace alveolar structures, the latter normally lined by alveolar type 1 (AT1) and AT2 cells. Bronchial epithelial stem cells (BESCs) can give rise to AT2 and AT1 cells or honeycomb cysts following bleomycin-mediated lung injury. However, little is known about what controls this binary decision or whether this decision can be reversed. Here we report that inactivation of Fgfr2b in BESCs impairs their contribution to both alveolar epithelial regeneration and honeycomb cysts after bleomycin injury. By contrast overexpression of Fgf10 in BESCs enhances fibrosis resolution by favoring the more desirable outcome of alveolar epithelial regeneration over the development of pathologic honeycomb cysts.

overall_design: Sox2CreERT2;mTmG mice were placed on tamoxifen food for 3 weeks starting at 2 months of age, followed by a 2 month washout period prior to saline (3 mice) or bleomycin treatment (3 mice). Whole lung tissues were collected, from lineage traced mice 6 weeks after saline or bleomycin treatment.


title: Charting DENR-dependent translation reinitiation uncovers predictive uORF features and links to circadian timekeeping via Clock

status: Public on Apr 01 2019

city: Lausanne

first: David

last: Gatfield

address: Genopode

department: Center for Integrative Genomics

country: Switzerland

institute: University of Lausanne

summary: The non-canonical initiation factor DENR promotes translation reinitiation on uORF-containing mRNAs. Moreover, DENR depletion shortens circadian period in mouse fibroblasts, suggesting that uORF usage and reinitiation regulate clock function. To identify DENR-regulated translation events transcriptome-wide and, in particular, specific core clock transcripts affected by this mechanism, we have used ribosome profiling in DENR-deficient NIH3T3 cells. We found 240 transcripts with altered translation rate, and used linear regression analysis to extract uORF features predictive of DENR dependance. Among core clock genes, we identified Clock as a DENR target. Using Clock 5'UTR mutants, we mapped the specific uORF through which DENR acts to regulate CLOCK protein biosynthesis. Notably, these experiments identified an alternative downstream start codon, which likely represents the true CLOCK N-terminus. Our findings provide insights into uORF-mediated translational regulation that can regulate the mammalian circadian clock and gene expression at large.

overall_design: iCLIP experiments: Denr-overexpressing (OE) or non-transfected (NT) NIH3t3 cells were grown to subconfluency in 15 cm plates/replicate. Cells were crosslinked with UV-C (254 nm) at 150 mJ/cm2,prior to lysate preparation. RNA was partially digested, and DENR-RNA complexes were pulled down with anti-DENR antibody (pre-immobilised in protein G dynabeads). An infrarred adaptor was ligated in the 3' end of RNAs and complexes were visualised after PAGE and nitrocellulose transfer in a Licor machine. RNA-protein complexes of 60-130 Kda were excised and used for library preparation. 24 PCR cycles were used for final library amplification. An IgG immunoprecipitation was used as IP control.

first: Violeta

last: Castelo-Szekely

first: Mara

last: De Matos

first: Marina

last: Tusup

first: Steve

last: Pascolo

first: Jernej

last: Ule


title: Integrated epigenomic and transcriptomic profiling of terminal human erythropoiesis

city: New York

first: Caleb

last: Lareau

state: New York

address: 417 E 68th St, Zuckerman - ZRC 1132

institute: Memorial Sloan Kettering

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

overall_design: Refer to individual Series


title: Identification of DDX5 target genes in spermatogonia

status: Public on Apr 02 2019

city: CLAYTON

first: Robin

middle: Mark

last: Hobbs

state: VIC

address: 27-31 Wright Street

department: Centre for Reproductive Health

country: Australia

institute: Hudson Institute of Medical Research

summary: Sustained spermatogenesis in adult males and recovery of fertility following germ cell depletion are dependent on undifferentiated spermatogonia with self-renewal potential. Self-renewal of undifferentiated spermatogonia is dependent on a complex network of transcriptional and post-transcriptional regulatory factors. DDX5 is known to act as a transcriptional co-regulator in other systems through interactions with key transcription factors. Here, we performed ChIP-Seq to identify target genes of DDX5 in cultured undifferentiated spermatogonia in order to elucidate its role in maintenance of the male germline.

overall_design: Chromatin immunoprecipitations using anti-DDX5 were performed using digested chromatin from two independently derived cultured undifferentiated spermatogonia from CBA mice.

first: Julien

last: Legrand

first: Ai-Leen

last: Chan

first: Hue

last: La

first: Frances

middle: V

last: Fuller-Pace


title: Sensory lesioning induces microglial synapse elimination via ADAM10 and fractalkine signaling

city: Worcester

first: Georgia

last: Gunner

state: Massachusetts

address: 364 Plantation Street, LRB 770 Y-Z

department: Neurobiology

institute: UMass Medical School

summary: Single-cell (inDrops) analysis of primary somatosensory cortex after early postnatal unilateral whisker lesioning

overall_design: CX3CR1-GFP (JAX stock # 005582) heterozygous (HET) and homozygous (KO) littermates underwent unilateral whisker lesioning at postnatal day 4 (P4). 24hours after whisker removal the primary somatosensory cortex was dissected from either hemisphere within each animal (Control- Undeprived/spared whiskers; Experimental- Deprived/whiskers removed) and single cell RNASeq performed.

first: Lucas

last: Cheadle

first: Kasey

last: Johnson

first: Pinar

last: Ayata

first: Ana

last: Badimon

first: Erica

last: Mondo

first: M

middle: A

last: Nagy

first: Lliwang

last: Liu

last: Bemiller

first: Ki-Wook

last: Kim

first: Sergio

last: Lira

first: Bruce

last: Lamb

first: Andrew

last: Tapper

first: Richard

last: Ransohoff

first: Michael

last: Greenberg

first: Anne

last: Schaefer

first: Dorothy

last: Schafer


first: David

last: Gatfield

overall_design: Denr knock-down or control NIH3t3 cells were grown to subconfluency in 15 cm plates/replicate. Cells were pre-treated with cycloheximide for 2 min, prior to lysate preparation. Two different sh-RNAs were used for knock-down experiments, and  Ribosome profiling (RPF-seq) and parallel RNA-seq libraries were prepared in triplicate for each. For control, non-targeting scramble sh-RNA in triplicate,  and Gfp-shRNA in duplicate was used.

first: Violeta

last: Castelo-Szekely

first: Mara

last: De Matos


title: DENR-regulated reinitiation events uncover predictive uORF features and links to circadian timekeeping via Clock regulation


city: Koblenz

first: Andreas

last: Schüttler

address: Am Mainzer Tor 1

department: Department Biochemistry and Ecotoxicology

institute: German Federal Institute of Hydrology

summary:
Time and concentration dependent transcriptome signatures in the ZFE of diruon, diclofenac and naproxen

Keywords: Expression profiling by array

overall_design: Wild-type adult zebrafish, originally received from OBI petshop (Leipzig, Germany) were kept in 120L fish tanks containing carbon-filtered tap water at 26°C under a 12h:12h dark-light cycle. Eggs were collected approximately 1 hour after light onset and inspected using a light microscope. Fertilized eggs were incubated at 26°C, while unfertilized eggs were discarded. For lethality experiments, which were conducted to find adequate exposure concentrations for transcriptome experiments, nine embryos per replicate were distributed into three 7.5ml glas vials, each containing 6ml exposure or control medium. Vials were sealed and incubated in a climate chamber shaking (Edmund Bühler SM-30 Control, 26°Celsius, 75 rpm, 12h:12h light:dark) till 96 hours post fertilization (hpf). At 48 hpf, 72 hpf and 96 hpf sublethal and lethal effects were recorded. For transcriptome experiments 20 embryos per replicate were transfered to two 20ml GC-glas vials at 24 hpf. A volume of 18ml of exposure medium was added to each vial and vials were sealed and incubated in a climate chamber (same settings as above) until sampling. Embryos were exposed to a range of 5 different concentrations of toxicant and RNA was extracted at 6 different timepoints. Exposure medium for all experiments was prepared by dissolving the substance in freshly prepared ISO-water (pH 7.4, oxygenized) (Diclofenac, Naproxen) or in 0.1 % methanol as solubilising agent (Diuron). Time points of RNA-extraction during the exposure were 3, 6, 12, 24, 48, and 72 hour post exposure start. The exposure concentrations were phenotypically anchored to the LC at 96 hpf/72 hpe. The LC25, modeled from experimental observations served as highest and the LC0.5 as lowest exposure concentration with 6 equal dilution steps in between, with dilution steps 1,2,4 and 6 chosen for exposure.

first: Janet

last: Krüger

first: Rolf

last: Altenburger

first: Madeleine

last: Ammar

first: Marcella

last: Bader-Blukott

first: Gianina

last: Jakobs

first: Johanna

last: Knapp

first: Wibke

last: Busch


title: Gene expression differences between activated B cells expressing GL7 and not in Peyer´s patches

city: Gothenburg

first: Mats

last: Bemark

address: PO Box 435

department: Departement of microbiology and immunity

institute: University of Gothenburg

summary: We report two populations of activated germinal centre B cells in the Peyer´s patches that are distinguished by their expression of GL7 or not. Using an antigen-specific system, we sort GL7+ and GL7- activated cells and performe RNASeq. We also included GL7 expressing B cells activated against the gut microbiota, and resting B cells from Peyer´s patches and spleen.

overall_design: GFP expressing NP-specific B cells from spleen were transfered into recipient wild type C57Bl/6 mice and the mice were orally immunized with NP conjugated to cholera toxin (NP-CT). Antigen-specific cells were isolated based on GFP and GL7 expression and compared to endogenous GL7+ and GL7- cells.

first: Nils

last: Lycke


title: Integrated epigenomic and transcriptomic profiling of terminal human erythropoiesis [ATAC-seq]

status: Public on Mar 31 2019

first: Caleb

last: Lareau

summary: In vitro cultured CD34+ derived erythroblasts were sorted using surface markers and processed using ATAC-seq

overall_design: Biological replicates (3 or 4 per population) were processed across 2-3 biological donors for 8 sorted populations for ATAC-seq

first: Leif

last: Ludwig


title: Integrated epigenomic and transcriptomic profiling of terminal human erythropoiesis [TMCC2]

summary: HUDEP-2 cells were lentivirally infected with CRISPRi constructs using a nontargeting guide or guides targeting an enhancer in the TMCC2 locus

overall_design: Whole transcriptome libraries were sequenced for three replicates of non-targeting gRNA and two replicates each for two different gRNA targeting a regulatory region upstream of the TMCC2 erythroid-specific isoform


title: Regulation of gene expression in undifferentiated spermatogonia by DDX5

first: Robin

last: Hobbs

summary: The maintenance of spermatogenesis in adult males is dependent on a population of mitotic germ cells with self-renewal potential known as undifferentiated spermatogonia. Regulation of undifferentiated spermatogonia function is dependent on transcriptional and post-transcriptional mechanisms. We have identified an essential role for the RNA helicase DDX5 in undifferentiated spermatogonia through generation of a UBC-CreERT2;Ddx5flox/flox mouse model that allows tamoxifen-dependent Ddx5 ablation. To identify genes regulated by DDX5, we generated lines of cultured undifferentiated spermatogonia from these mice and treated cells with tamoxifen (TMX) to induce Ddx5 knockout or vehicle (VEH) as a control, and performed RNA-sequencing analysis to compare these conditions.

overall_design: 4 independently generated lines of cultured undifferenatiated spermatogonia from UBC-CreERT2;Ddx5flox/flox mice treated with vehicle (VEH) as a control or tamoxifen (TMX) to induce Ddx5 knockout.

middle: M

first: Julien

last: Legrand

first: Ai-Leen

last: Chan

first: Hue

last: La

first: Fernando

last: Rossello

first: Minna-Liisa

last: Anko

first: Frances

last: Fuller-Pace


title: Integrated epigenomic and transcriptomic profiling of terminal human erythropoiesis [RNA-seq]

summary: In vitro cultured CD34+ derived erythroblasts were sorted using surface markers and processed using RNA-seq

overall_design: Biological replicates (3 or 4 per population) were processed across 2-3 biological donors for 8 sorted populations for RNA-seq


title: A c-type lectin triggers immune surveillance by innate lymphoid  cells for eradication of attaching-and-effacing bacteria.

city: Saint-Malo

first: Phuong

middle: Hang

last: PHAM

address: 7 allée Métis

department: Data science

institute: Parean Biotechnologies

summary: Although gut protection to infection is ensured by innate lymphoid cells (ILCs), how ILCs are regulated remains poorly understood. Here, we show that secretion of the c-type lectin Reg3b downstream of Ripk2 signaling pathway was crucial for immune surveillance by ILCs against Citrobacter rodentium that mimics human infection with attaching-and-effacing bacteria. Unexpectedly, phosphorylation of signal transducer and activator of transcription 3 (STAT3) and secretion of both tumor necrosis factor alpha (Tnf-a) and interleukin-17A (IL-17A) were blunted in mutant mice contributing to their susceptibility to a primary infection. We further determined that influx of neutrophils was triggered by Reg3b through an intact IL-17A signaling. Loss of either Reg3b or Ripk2 weakened Tnf-a secretion by intestinal phagocytes lowering their propensity to promote IL17A secretion by ILCs early in infection. These results provide a previously unrecognized mechanism by which intestinal epithelial cells reciprocally regulate early production of IL-17A by ILCs preventing from overcolonization by opportunistic enteropathogenic bacterium.

overall_design: Microarray analysis was performed by using total tissue from caecum tip of mice on days 0, 4 and 12 post-infection (n = 5). Changes (log2-fold) in gene expression profiles of Ripk2-/-, Nod2-/- and WT mice on day 4 p.i. and on day 12 p.i. were calculated relative to non-infected WT controls and compared by correlation analysis.

first: Nadine

last: Waldschmitt

first: Myriam

last: Delacre

first: Hang

last: Pham

first: Adrien

last: Six

first: Harry

last: Sokol

first: Mathias

last: Chamaillard


title: Identification of gene expression pattern in Brassica rapa floral tissues including Flowerbuds (≤ 2mm), Flowerbuds (≤ 4mm), Stamen and Carpel

status: Public on Mar 29 2019

city: Jeonju-si

first: Soo In

last: Lee

state: Jeollabuk-do

address: Nongsaengmyeong-ro, Wansan-gu

department: Agricultural Biotechnology

country: South Korea

institute: National Institute of Agricultural Sciences (NAS)

summary:
Investigation of differentially expressed gene in floral tissues of of Brassica rapa in comparison with leaves as control
To unravel the transcriptomic changes associated with small early floral buds (<2 mm; FB2), large early floral buds (2-4 mm; FB4), stamen (ST) and carpel (CP) tissues, transcriptome profiling was carried out with Br300K oligo microarray.

overall_design: A twelve chip study using total RNA recovered from small early floral buds (<2 mm; FB2), large early floral buds (2-4 mm; FB4), stamen (ST) and carpel (CP) tissues of vernalization sensitive rassica rapa plants plants and non-vernalised leaves (vegetative tissue) as control

first: Soo-In

first: Myung-Ho

last: Lim


title: DNA microarray expression analysis of mice spleen following cadmium exposure

status: Public on Mar 28 2019

city: Zhenjiang

first: Haifeng

last: Shi

state: Jiangsu

address: 301 Xuefu Rd

department: Institute of life sciences

country: China

institute: Jiangsu University

summary: This study use DNA microarray to examine gene expression in the spleen of mice exposed to cadmium. Mice were exposed via a single subcutaneous injection CdCl2, with spleen harvested at 24h after exposure for microarray analysis. This work provides insight into the mechanistic pathways involved in acute cadmium intoxication, providing a theoretical basis for identifying candidate biomarkers.

overall_design: Total RNA was isolated from spleen tissue of ICR mice which subcutaneous injected with 5μmol/kg or 10μmol/kg CdCl2 compared to  subcutaneous injected Saline

first: Jie

last: Gu


title: Slx5/Slx8-dependent ubiquitin hotspots on chromatin contribute to stress tolerance [Gene expression]

city: Martinsried/Munich

first: Markus

last: Höpfler

address: Am Klopferspitz 18

department: Molecular Cell Biology

institute: Max-Planck Institute of Biochemistry

summary: Chromatin is a highly regulated environment, and protein association with chromatin is often controlled by post-translational modifications and the corresponding enzymatic machinery. Specifically, SUMO-targeted ubiquitin ligases (STUbLs) have emerged as key players in nuclear quality control, genome maintenance and transcription. However, how STUbLs select specific substrates amongst myriads of SUMOylated proteins on chromatin remains unclear. Here, we reveal a remarkable co-localization of the budding yeast STUbL Slx5/Slx8 and ubiquitin at seven genomic loci that we term ‘ubiquitin hotspots’. Ubiquitylation at these sites depends on Slx5/Slx8 and protein turnover on the Cdc48 segregase. We identify the transcription factor-like Ymr111c/Euc1 to associate with these sites and be a critical determinant of ubiquitylation. Euc1 specifically targets Slx5/Slx8 to ubiquitin hotspots via bipartite binding of Slx5 that involves the Slx5 SUMO-interacting motifs and an additional, novel substrate recognition domain. Interestingly, the Euc1-ubiquitin hotspot pathway acts redundantly with chromatin modifiers of the H2A.Z and Rpd3L pathways in specific stress responses. Thus, our data suggest that STUbL-dependent ubiquitin hotspots shape chromatin during stress adaptation.

platform_organism:
Schizosaccharomyces pombe
Saccharomyces cerevisiae

overall_design: Gene expression profiling was performed in biological triplicates to compare transcriptomes of ymr111c/euc1∆ or ymr111c/euc1-K231R mutant strains with a wild-type strain as reference. RMA-normalized expression intensities were obtained for each sample and pair-wise comparisons between sample groups were performed to obtain log2-fold-change values.

first: Maximilian

last: Kern

first: Tobias

last: Straub

first: Roman

last: Prytuliak

first: Bianca

middle: H

last: Habermann

first: Boris

last: Pfander

first: Stefan

last: Jentsch


title: Slx5/Slx8-dependent ubiquitin hotspots on chromatin contribute to stress tolerance [ChIP-chip]

type: Genome binding/occupancy profiling by genome tiling array

overall_design: ChIP-chip profiling of ubiquitylated proteins (FK2-antibody and K48-specific Apu2-clone), Slx8-9myc and Ymr111c/Euc1 in S. cerevisiae. Custom-made Nimblegen arrays (100304_Scer2_MS_Chip_Stefan Jentsch MPI Biochemistry S.cerevisiae 135K Tiling Array Version 2) were used to compare ChIP versus input DNA. Typically, replicate experiments were performed including a dye-swap for ChIP/input DNA (except for rad6-delta-and one of the IgG-control experiments in Fig. 1A).


title: Slx5/Slx8-dependent ubiquitin hotspots on chromatin contribute to stress tolerance [RNA-Seq]

overall_design: Gene expression profiling was performed in biological triplicates to compare transcriptomes of ymr111c/euc1∆ to WT or YMR111C/EUC1-overexpression (pGAL-EUC1) to a control strain in Saccharomyces cerevisiae. Total RNA was isolated and sequenced on an Illumina NextSeq 500 instrument.


title: Transcriptome of Dp1Tyb and wild-type mouse embryonic fibroblasts and hippocampus

city: London

first: Victor

last: Tybulewicz

address: 1 Midland Road

institute: Francis Crick Institute


title: Neuron-astrocyte metabolic coupling protects against activity-induced fatty acid toxicity

city: Ashburn

first: Zhe

last: Liu

state: Virginia

address: 19700 Helix Dr.

department: Janelia Farm Research Campus

institute: Howard Hughes Medical Institute

summary: Gene expression profiling of cultured glia with low or high levels of lipid droplets

overall_design: We stained lipid droplets (LDs) in cultured rat glia with nile red stain. We seperated glia containing high numbers of LDs using fluorescence activated cell sorting (FACS). We performed mRNA-seq experiments on glia with low or high levels of LDs.

first: Maria

last: Ioannou


title: Slx5/Slx8-dependent ubiquitin hotspots on chromatin contribute to stress tolerance

overall_design: Refer to individual Series


title: Transcriptome of Dp1Tyb and wild-type mouse embryonic fibroblasts [ERCC spike-ins]

summary: Purpose: to identify the effects of the Dp1Tyb mutation on the transcriptome of mouse embryonic fibroblasts

overall_design:
RNAseq libraries were prepared from RNA isolated from mouse embryonic fibroblasts. Libraries were prepared from total RNA using the TruSeq Stranded mRNA Sample Prep Kit (Illumina) by the Advanced Sequencing Facility, The Francis Crick Institute. Libraries were sequenced (100 bases paired end) on the Illumina Hiseq 4000
Please note that this dataset contains ERCC spike ins to normalise the data

first: Nneka

last: Anyanwu

first: Helena

last: Ahlfors


title: Identification of two distinct pathways of human myelopoiesis.

status: Public on Mar 27 2019

city: Oxford

first: Claus

last: Nerlov

address: John Radcliffe Hospital, University of Oxford

department: MRC Molecular Haematology Unit

institute: Weatheral Institute of Molecular Medicine

summary: The myeloid innate immune system plays key roles in the defence against parasites and bacteria, and carries out essential functions during tissue homeostasis and repair. It consist of neutrophils, macrophages, eosinophils, basophils and mast cells. Myelopoietic dysfunction or deregulation underlies numerous pathologies, ranging from immune deficiencies and allergies to myeloproliferative disorders and myeloid leukemia. To study malignant myelopoiesis, it is essential to understand the ontology of the different celltypes. In a generally accepted model for hematopoietic differentiation, all myeloid cells are preceded by a common granulocyte and macrophage progenitor that has the potential for all myeloid celltypes at the single cell level. However, we recently proposed a model for mouse hematopoiesis in which myeloid cells are formed via two independent pathways, one leading towards macrophages and neutrophils and one leading towards eosinophils, basophils and mast cells. The first pathway is affiliated with lymphoid potentials whereas the latter with megakaryocyte/erythroid development. Here, we demonstrate that those findings translate to human hematopoiesis. We demonstrate prospective isolation of myeloid progenitor cells that have restricted multipotent myeloid potential and are affiliated with lineages similar to those found in the mouse hierarchy.

overall_design: Gene expression of human single common myeloid progenitor cells (Lin- CD34+ CD38+ CD45RA- CD123+), and defined bulk populations (100 cells) within the CD34+ compartment from bone marrow of healthy adult donors.

first: Roy

last: Drissen

first: Supat

last: Thongjuea


title: Effect of NOX2 (CYBB) knock out on the transcriptome of leukemia stem cells isolated from NUP98/HOXA9+BCR/ABL murine syngeneic leukemia model

status: Public on Mar 26 2019

city: Aurora

first: Shanshan

last: Pei

state: CO

institute: University of Colorado

summary: To investigate the function of NOX2 (CYBB) in leukemia stem cells, we created leukemia by tranforming primitive hematopoetic cells isolated from wild type and NOX2-KO mice with NUP98/HOXA9 and BCR/ABL oncogenes, isolated mRNA, and performed RNA-seq experiments to profile transcriptional changes upon NOX2 loss.

overall_design: For each condition (wt or KO), we purified LSCs from 4 leukemic recipient mice and pooled them into one technical replicate. Two technical replicates per condition were used.

first: Biniam

last: Adane

first: Craig

last: Jordan


title: Yb body assembly on the flamenco piRNA precursor transcripts reduces genic piRNA production

city: Stockholm

first: Artem

last: Ilin

address: Svante Arrhenius väg 20C

department: MBW

institute: Stockholm University

type: Non-coding RNA profiling by high throughput sequencing

summary: In Drosophila ovarian somatic cells, piRNAs against transposable elements are mainly produced from the ~180-kb flamenco (flam) locus. flam transcripts are gathered into one-two foci, located close to nuclear envelope, and processed into piRNAs in the cytoplasmic Yb bodies. The mechanism of Yb body formation remains unknown. Using RNA FISH, we found that in the follicle cells of ovaries the 5’-ends of flam transcripts are usually located in close proximity to the nuclear envelope and outside of Yb bodies, while their extended downstream regions mostly overlap with Yb bodies. In flamKG mutant ovaries, flam transcripts containing the 1st and, partially, 2nd exons but lacking downstream regions are gathered into foci at the nuclear envelope, but Yb bodies are not assembled. Strikingly, piRNA production from the protein-coding gene transcripts is enhanced in flamKG ovaries. We propose that normally in ovarian follicle cells flam downstream transcript regions function not only as a substrate for generation of piRNAs but also as a scaffold for Yb body assembly, which is necessary to minimize piRNA production from protein-coding gene transcripts. By contrast, in ovarian somatic cap and escort cells Yb body assembly does not require flam transcription.

overall_design: The fractions of small RNAs (19-29 nt) from ovaries of flam heterozygous flies (flamKG/FM7 control) and flamKG/Df mutants were sequenced using Illumina NovaSeq 6000

last: Ilyin

first: Olesya

last: Sokolova


title: Sequence and Expression Levels of Circular-RNAs in Progenitor Cell Types During Mouse Corticogenesis

city: Modena

first: Martina

last: Dori

address: Via G. Campi 287

department: Department of Life Sciences

institute: Center for Genome Research

summary: Circular (circ) RNAs have recently emerged as a novel class of non coding transcripts whose identification and function remain elusive. Among many tissues and species, the mammalian brain is the organ in which circRNAs are more abundant and in which the first evidence of functional significance started to emerge. Yet, even within this well studied organ, annotation of circRNAs remains fragmentary, their sequence is unknown and their expression patterns in specific cell types was never investigated. Overcoming these limitations, here we provide the first comprehensive identification of circRNAs and their expression in proliferating neural stem cells, neurogenic progenitors and newborn neurons of the developing mouse cortex. Extending the current knowledge about the diversity of this class of transcripts by the identification of nearly 4,000 new circRNAs, our study is the first to provide the full sequence information and expression patterns of circRNAs in specific cell types and representing the lineage toward neurogenic commitment. We further exploited our data by evaluating the coding potential, evolutionary conservation and biogenesis of circRNAs that we found to arise from a specific sub-class of linear mRNAs. Our study provides the arising field of circRNA biology with a new resource essential to address the complexity and potential biological significance of this new class of transcripts

overall_design: Circular RNA profiles in different cell population (proliferating and differentiating progenitors and newborn neurons) of E14.5 developing mouse cortex. Profiles were obtained by deep-sequencing of RNaseR treated total RNA, in triplicate, using Illumina HiSeq 2500

first: Leila

last: Haj Abdullah Alieh

first: Daniel

last: Cavalli

first: Simone

last: Massalini

first: Mathias

last: Lesche

first: Andreas

last: Dahl

first: Federico

last: Calegari


title: Stem cell proliferation is kept in check by the chromatin regulators Kismet/CHD7/CHD8 and Trr/MLL3/4

city: Paris cedex 05

first: Marius

last: van den Beek

state: Ile-de-France

address: 11-13 rue Pierre et Marie Curie, Equipe Bardin/UMR3215/Batiment BDD

department: GENETICS AND DEVELOPMENTAL BIOLOGY

institute: Institut Curie

summary: Chromatin remodeling accompanies differentiation, however, its role in self-renewal is less-well understood. We report that in Drosophila the chromatin remodeler Kismet/CHD7/CHD8 limits intestinal stem cells (ISC) number and proliferation without affecting differentiation. Stem cell-specific whole-genome profiling of Kismet revealed its enrichment at transcriptionally active regions bound by RNA Polymerase II and Brahma, its recruitment to transcription start site of activated genes and developmental enhancers and its depletion from regions bound by Polycomb, Histone H1 and heterochromatin Protein 1. We demonstrate that the Trithorax-related/MLL3/4 chromatin modifier regulates ISC proliferation, colocalizes extensively with Kismet throughout ISC genome, and co-regulates genes in ISCs, including Cbl a negative regulator of Epidermal Growth Factor Receptor (EGFR). Loss of kismet or trr leads to elevated levels of EGFR protein and signaling, thereby promoting ISC self-renewal. We propose that Kismet with Trr establishes a chromatin state that limits EGFR proliferative signaling, preventing tumor-like stem cell overgrowths.

overall_design: genome wild profiling of different chromatin factors in Drosophila intestinal stem cells by DamID and RNAseq on FACS sorted ISC expressing Kis RNAi Trr RNAi or white RNAi

first: Louis

last: Gervais

first: Manon

last: Josserand

first: Jérémy

last: Sallé

first: Marine

last: Stefanutti

first: Carolina

middle: N

last: Perdigoto

first: Patricia

last: Skorski

first: Khallil

last: Mazouni

first: Owen

last: Marshall

first: Andrea

last: Brand

first: François

last: Schweisguth

first: Allison

last: Bardin


title: Probabilistic modeling of personalized drug combinations from integrated chemical screen and molecular data in sarcoma

status: Public on Mar 25 2019

city: Beaverton

first: Noah

last: Berlow

address: 12655 SW Beaverdam Rd

institute: CC-TDI

summary: DNA and RNA were isolated from snap frozen primary tumor tissue (human and mouse), and DNA was isolated from matched normal tissue (region matched non-tumor tissue for human, tail section for mouse). A mouse leg was also injected with cardiotoxin to induce tissue damage was taken following sacrifice and was snap frozen, then processed for RNA isolation to serve as a mouse tissue-normal for RNA sequencing. All sequencing was performed at the OHSU MPSSR core.

overall_design: Tumor and normal tissues were unpurturbed from procurement to snap freezing, so there are no experimental conditions for any of the samples submitted. A mouse limb was treated with cardiotoxin to induce physical damage and stimulate the wound response to create a similar environment to tissue bearing alveolar rhabdomyosarcoma.

middle: E

first: Charles

last: Keller


title: Transcriptome of Dp1Tyb and wild-type mouse embryonic fibroblasts

first: Victor

last: Tybulewicz

overall_design: RNAseq libraries were prepared from RNA isolated from mouse embryonic fibroblasts. Libraries were prepared from total RNA using the TruSeq Stranded mRNA Sample Prep Kit (Illumina) by the Advanced Sequencing Facility, The Francis Crick Institute. Libraries were sequenced (100bp paired end) on the Illumina Hiseq 2500.

first: Nneka

last: Anyanwu

first: Natalia

last: Dinischiotu

first: Helena

last: Ahlfors


title: Gamma entrainment binds higher order brain regions and alleviates neurodegeneration

city: Cambridge

first: Li-Huei

last: Tsai

department: Picower Institute for Learning and Memory

institute: MIT

summary: We report the RNA-Seq data of microglia from CK-p25 mice visual cortex

overall_design:
Summary of mice and treatments included in this series:

CK-p25 mice - in which the expression of the Cdk5 activator p25 is driven by the excitatory neuron-specific CaMKIIα promoter in an inducible manner (CaMKIIα promoter- tTA x TetO- p25+GFP) (Cruz et al., 2003). Following withdrawal of doxycycline from the diet, CK-p25 exhibit progressive neuronal and synaptic loss with cognitive impairment, which is severe by 6 weeks of p25 induction (Cruz et al., 2003).

Tau P301S mice, which express high levels of humanized mutant microtubule-associated protein tau and have tau aggregates that are associated with frontotemporal dementia as early as 5 months of age (Yoshiyama et al., 2007). 8 months old P301S mice, at which age they have synaptic and neuronal loss and cognitive deficits.

CK = wild type control mice; CK-p25 + No Stim = CK-p25 mice that did not under go any stimulation; CK-p25 +GENUS = Ck-p25 mice that was stimulated with 40 Hz visual stimulation

WT = wild type control mice; P301S + No Stim = P301S mice that did not under go any stimulation; P301S +GENUS = P301S mice that was stimulated with 40 Hz visual stimulation

first: Chinnakkaruppan

last: Adaikkan

first: Fan

last: Gao


title: Transcriptome of Dp1Tyb and wild-type mouse hippocampus

summary: Purpose: to identify the effects of the Dp1Tyb mutation on the transcriptome of mouse hippocampus

overall_design: RNAseq libraries were prepared from RNA isolated from whole hippocampi. Libraries were prepared from total RNA using the TruSeq Stranded mRNA Sample Prep Kit (Illumina) by the Advanced Sequencing Facility, The Francis Crick Institute. Libraries were sequenced (75bp paired end) on the Illumina Hiseq 4000.

first: Justin

last: Tosh

first: Elizabeth

last: Fisher


title: Transcriptome of Trophoblast Stem Cells

city: Shanghai

first: Shaorong

address: Siping Road NO.1239

institute: Tongji University

summary: Three kinds of TSCs with different genotype of mouse Tet2 including Tet2+/-, Tet2-/-and wild-type TSCs  were harvested . Total RNA was isolated from cell pellets by Trizol reagent and RNA-Seq library were generated using KAPA Strandard mRNA-Seq Kits according to the manufacturer’s recommendations.

overall_design: mRNA profiles of 3 cell types, generated by deep sequencing in diplicate, using HiSeq 2500. Paired-end 125-bp sequencing was applied.

first: Linfeng

last: Zhang

first: Wenchao

last: Xiu


title: Identification of microRNA signatures in umbilical cord blood associated with maternal characteristics

city: Brno

first: Jaroslav

last: Juracek

address: Kamenice 5

country: Czech Republic

institute: CEITEC MU

summary: Umbilical cord blood could serve as useful source of blood markers enabling more efficient and reliable prenatal and neonatal diagnostics. MicroRNAs (miRNAs) are ubiquitous in body fluids where they were used for detecting and monitoring various physiological and pathological conditions. In this descriptive study, we aimed to identify changes in miRNA expression profiles associated with basic maternal somatic and epidemiological characteristics.

overall_design: Cord blood from 24 mothers was collected at time of delivery and global miRNA profiling was performed. Expression profiles were statistically evaluated in relation to maternal age, BMI, pregnancy weight gain, blood type, Rh factor status, allergies during pregnancy, addictive substance abuse and smoking status.

first: Ondrej

last: Slaby


title: Nutrient sensing by the intestinal epithelium orchestrates mucosal antimicrobial defense via translational control of Hes1

city: Beijing

first: Shaonan

last: Liang

state: Beijing

address: Room D311, Medical Science Building, Tsinghua University

department: School of Medicine

institute: Institute for Immunology

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


title: Bloom syndrome protein restrains innate immune sensing of DNA damage by cGAS.

city: Paris

first: Nicolas

last: Manel

address: 12 rue Lhomond

department: INSERM U932

summary: Cellular innate immune sensors of DNA are essential for host defense against invading pathogens. However, the presence of self-derived DNA inside cells poses a biological risk of triggering an inappropriate immune response. The mechanisms limiting induction of inflammation by self nucleic acids are poorly understood. BLM RecQ like helicase is essential for the maintenance of genome integrity and deficient in Bloom syndrome, a rare genetic disease characterized by genome instability, susceptibility to cancer and immunodeficiency. Here, we show that BLM-deficient cells exhibit a constitutively upregulated inflammatory interferon-stimulated gene (ISG) signature. Restoring BLM expression reverts ISG expression to baseline. ISG expression in BLM-deficient fibroblasts is mediated by the cGAS-STING-IRF3 cytosolic DNA sensing pathway. Increased DNA damage or downregulation of the cytoplasmic exonuclease TREX1 enhances ISG expression in BS fibroblasts, and cytoplasmic micronuclei positive for cGAS are increased in BLM-deficient fibroblasts. Finally, BS patients demonstrate elevated ISG expression in peripheral blood. We conclude that BLM is essential to limit the expression of pro-inflammatory genes resulting from the sensing of DNA damage products by the cGAS-STING-IRF3 pathway. These results reveal an unexpected role for BLM in limiting ISG induction, thus connecting DNA damage to innate immunity, which may contribute to human pathogenesis.

overall_design: Human SV40-immortalized fibroblasts depleted for BLM or not by CRISPR-Cas9 (3 replicates each) were seed and allow to grow. After 48h, cells were harvest and total RNA was extract for gene expression analysis.

first: Matthieu

last: Gratia


title: Transcriptional characterization of  PD-1 positive and negative Foxp3 negative CD4 positive T cells from liver of 5- and 10-day-old Aire-deficient C57BL/6 mice

city: Boston

first: CBDM

last: Lab

address: 77 Avenue Louis Pasteur

department: Microbiology and Immunobiology

institute: Harvard Medical School

summary: The first T cells to arrive in the liver were mostly T regulatory (Treg) cells and metabolically active, highly proliferative T conventional (Tconv) cells. The Tconv cells had unusually high expression of PD-1 and the IL-33 receptor, ST2. As these PD-1+ Tconv cells accumulated in the tissue, they gradually lost their expression of ST2, ceased to proliferate and acquired an anergic phenotype.

overall_design: Gene expression profiles of flow cytometry sorted DAPI negative CD45 positive TCRb positive CD4 positive Foxp3 negative cells from liver of 5- and 10-day-old B6.Aire-KO mice

first: Jonatan

last: Tuncel

first: Diane

last: Mathis

first: Christophe

last: Benoist


title: Light can synchronise peripheral clocks autonomously from each other [darkness experiment (DD)]

city: Barcelona

first: Aikaterini

last: Symeonidi

state: Barcelona

address: Baldiri i Reixac, 10

department: Oncology

country: Spain

institute: Institut for Research in Biomedicine (IRB-Barcelona)

summary: Organisms have adapted to the changing environmental conditions within the 24h cycle of the day by temporally segregating tissue physiology to the optimal time of the day. On the cellular level temporal segregation of physiological processes is established by the circadian clock, a Bmal1 dependent transcriptional oscillator network. The circadian clocks within individual cells of a tissue are synchronised by environmental signals, mainly light, in order to reach temporally segregated physiology on the tissue level. However, how light mediated synchronisation of peripheral tissue clocks is achieved mechanistically and whether circadian clocks in different organs are autonomous or interact with each other to achieve rhythmicity is unknown. Here we report that light can synchronise core circadian clocks in two peripheral tissues, the epidermis and liver hepatocytes, even in the complete absence of functional clocks in any other tissue within the whole organism. On the other hand, tissue extrinsic circadian clock rhythmicity is necessary to retain rhythmicity of the epidermal clock in the absence of light, proving for the first time that the circadian clockwork acts as a memory of time for the synchronisation of peripheral clocks in the absence of external entrainment signals. Furthermore, we find that tissue intrinsic Bmal1 is an important regulator of the epidermal differentiation process whose deregulation leads to a premature aging like phenotype of the epidermis. Thus, our results establish a new model for the segregation of peripheral tissue physiology whereby the synchronisation of peripheral clocks is acquired by the interaction of a light dependent but circadian clock independent pathway with circadian clockwork dependent cues.

overall_design: Determining the epidermal circadian transcriptome in the presence or absence of non-epidermal clocks after 6-7 days in complete darkness (DD).

first: Patrick-Simon

last: Welz

first: Valentina

last: Zinna

first: Kevin

last: Koronowski

first: Kenichiro

last: Kinouchi

first: Inés

last: Marín Guillén

first: Andrés

last: Castellanos

first: Paolo

last: Sassone-Corsi

first: Salvador

last: Aznar Benitah


summary: To investigate whether intestinal Hes1 expression was translationally regulated during nutrient deprivation, we performed ribosome profiling of ileum from fed and fasted mice.

overall_design: Examination of mRNA translation in ileum of mice fed or fasted for 48 hrs

first: Rongyao

last: Huang


first: Shaonan

last: Liang

summary: Metabolic programs and host defense are highly integrated to ensure proper immune responses during stress. Central to these responses, mTOR regulates immune functions by sensing and integrating environmental cues, yet how these systems are coordinated at the intestinal surface remains undefined. We show that the antimicrobial peptide α-defensin is functionally sustained during nutrient deprivation due to regulation of defensin-processing enzyme MMP7 by microbiota- and host-derived factors. Unlike other antimicrobial peptides, the MMP7-α-defensin axis remains active during nutrient fluctuations, providing essential protection against enteric pathogens. Sustained Mmp7 expression requires the microbiota and is mediated by de-repression of the transcription activator Atoh1 upon attenuation of the transcriptional repressor Hes1 in intestinal epithelial cells. Hes1 levels are regulated via mTOR and controlled translationally, constituting a metabolism-translation-transcription loop. Disrupting this loop by supplying nutrients paradoxically compromises anti-bacterial defense. Together, these results uncover a regulatory circuit that couples host nutrient status to epithelial anti-microbial immunity.

overall_design:
Mice were fasted for 0 hr (also called fed), 48 hrs or 72 hrs. Ileum mRNA profiles of fed and fasted mice were generated by deep sequencing.
NOTE:  the 'Fed RNA' sample, prepared and sequenced together with 'Fasted_48 hrs RNA', is the control sample of 'Fasted_48 hrs RNA'. The 'Fed' sample, prepared and sequenced together with 'Fasted_72 hrs', is the control sample for the 'Fasted_72 hrs' sample.

first: Xuekun

last: Guo

first: Bin

last: Zhang


title: Expression profile of Lo19S state cells in the presence and absence of bortezomib treatment

first: Peter

last: Tsvetkov

address: 455 Main St

institute: Whitehead Institute

summary: We transiently induce the Lo19S state with a dox inducible shRNa targeting PSMD2 and explore the gene expression in the presence and absence of bortezomib

overall_design: one cell type (T47D), two states (Control and Lo19S) with and without treatment with 20nM bortezomib , all in triplicates


title: Expression profile of Lo19S state cells and MM.1S tumors

overall_design: Refer to individual Series


title: Exosomes from CD99-deprived Ewing sarcoma cells reverse tumor malignancy by inhibiting cell migration and promoting neural differentiation

status: Public on Mar 22 2019

city: Bologna

first: Annalisa

last: Astolfi

address: via massarenti 11

department: "Giorgio Prodi" Cancer Research center

institute: University of Bologna

summary: Ewing sarcoma (EWS) is an aggressive mesenchymal tumor with unmet clinical need and significant social impacts on children, adolescents and young adults. CD99, a hallmark surface molecule of EWS, participates in crucial biological processes including cell migration, differentiation and death. EWS cells can release CD99 through exosomes, specialized extracellular vesicles with major cell communication roles. Here we show that, as a consequence of CD99 silencing, EWS cells deliver exosomes with oncosuppressive functions which significantly reduce tumor aggressiveness. These CD99-lacking microvesicles modulate gene expression of the EWS recipient cells, reduce proliferation and migration, in turn inducing a more differentiated less malignant phenotype. The most relevant effects were detected on the AP-1 signaling pathway whose regulation was found to be dependent on the specific cargo loaded in vesicles after CD99 shutdown. Investigation of the miRNA content of CD99-deprived exosomes identified miR-199a-3p as a key driver to reverse EWS malignancy in experimental models as well as in clinical specimens.  All together our data provide evidence that the abrogation of CD99 in EWS tumor cells leads to produce and release exosomes capable to transfer their antineoplastic effects into the nearby tumor cells, suggesting a novel atypical role for these microvesicles in reversion of malignancy rather than in priming the soil for progression and metastatic seeding.  This conceptually innovative approach might offer a new therapeutic opportunity to treat a tumor still refractory to most treatments.

overall_design: The analysis was performed comparing the parental EWS cell line TC-71 and TC-71 after treatment with exosomes isolated by CD99-silenced cells. The aim of the study was to identify specific players responsible for the biological effects observed after exosome exposure.

first: Katia

last: Scotlandi


title: Detecting cytosine-5 methylation in RNA

first: Susanne

last: Bornelöv

address: Tennis Court Road

department: WT-MRC Cambridge Stem Cell Institute

institute: University of Cambridge

summary: We generated a map of NSUN6-dependent RNA 5 methyl-cytosine in the human transcriptome by applying RNA bisulfite conversion and sequencing to two sets of cell lines (HUES9 and HEK293) where the RNA cytosine 5 methyl-transfearse NSUN6 has been either knocked out by CRISPR Cas9 or overexpressed using a PiggyBac vector. We apply this and other molecular methods to validate NSUN6 RNA methylation targets with single nucelotide resolution in the human transcriptome.

overall_design: Whole transcriptome bisulfite sequencing of wildtype, Nsun6 -/-, Nsun6 over-expresson and Nsun6 rescue human H9 and/or HEK cells; four replicates per sample.

first: Tommaso

last: Selmi

first: Sabine

last: Dietmann

first: Michaela

last: Frye


title: Human keratinocytes RNA bisulfite sequencing

summary: Human keratinocytes were cultured either as undifferentiated or differentiated cells. Total RNA extracted from both conditions and depleted from rRNA then treated with three cycles of Sodium bisulfite. DNA libraries were then generated from converted RNA for sequecing.

overall_design: Whole transcriptome bisulfite sequencing of undifferentiated and differentiated human keratinocytes; 5 replicates per condition.

first: Abdulrahim

last: Sajini


title: Light can synchronise peripheral clocks autonomously from each other

first: Aikaterini

last: Symeonidi

overall_design: Determining the epidermal circadian transcriptome in the presence or absence of non-epidermal clocks under light entrainment (LD).

first: Patrick-Simon

last: Welz

first: Valentina

last: Zinna

first: Kevin

last: Koronowski

first: Kenichiro

last: Kinouchi


title: RNA sequencing on LNCaP cells

city: Oslo

first: Adnan

last: Hashim

state: Oslo

address: NCMM - Forskningsparken, House 1, 3rd Floor - Gaustadalléen 21

department: NCMM (Centre for Molecular Medicine Norway)

country: Norway

institute: UiO (University of Oslo)

summary: RNA sequencing on LNCaP cells was carried out to study how tunicamycin-induced gene expression is affected by knockdown of EIF2AK3 and ATF4.

overall_design: Samples from the below setup (treatments protocol) were harvested from four independent experiments. RNA integrity of total RNA samples was assessed by Bioanalyzer. All samples had RIN ≥ 9.7.

first: Morten

last: Luhr

first: Nikolai

last: Engedal


title: The lung environment controls alveolar macrophage metabolism and responsiveness during type-2 inflammation.

city: Edinburgh

first: Alasdair

last: Ivens

address: Kings Buildings

institute: Centre for Immunity, Infection and Evolution

summary: Fine control of macrophage activation is required to prevent inflammatory disease, particularly at barrier sites such as the lung. However, the dominant mechanisms that regulate pulmonary MΦs during inflammation are currently poorly understood. Here we show that airway MΦs are substantially less able to respond to the canonical type-2 cytokine IL-4, which underpins allergic disease and parasite worm infections, than lung tissue or peritoneal cavity MΦs. We reveal that MΦ hypo-responsiveness to IL-4 is dictated by the lung environment, though independent of the host microbiota or the prominent lung extracellular matrix components surfactant protein D and mucin 5b. Rather, compared to cavity MΦs,  airway MΦs display severely dysregulated metabolism. Strikingly, upon removal from the lung, alveolar MΦs regain IL-4 responsiveness in a process dependent upon glycolysis. Thus, we propose that impaired glycolysis within the pulmonary niche is a central determinant for regulation of MΦ responsiveness during type-2 inflammation.

overall_design: The 13 analysed samples belong to 6 different groups, each group consisted of 2 or 3 samples. The groups consist of 3 separate macrophage populations, from either control or IL-4 complex treated mice. Each individual sample was generated from 3-5 pooled biological replicate mice.

first: Freya

last: Svedberg

last: Cook

first: Andrew

last: Macdonald


title: Expression profile of MM.1S tumors folloiwing treatment with bortezomib

first: Peter

last: Tsvetkov

country: USA

summary: MM.1S orthotopic tumors were analyzed fro their gene expression upon tumor outgrowth. In contorl/bortezomib/elesclmol and combo treatments.

overall_design: examination of three tumors for each condition.


title: Targeting IDH1 as a pro-senescent therapy in high-grade serous ovarian cancer

city: Hershey

first: Yuka

last: Imamura Kawasawa

state: PA

address: 500 University Dr.

department: College of Medicine, Pharmacology

institute: Penn State University

summary: We found that targeting IDH1 in HGSC induces senescence by altering the repressive histone landscape of E2F target genes

overall_design: Ovcar3 cells were infected with control or shIDH1. Cells were selected in 3ug/mL puromycin for 7 days. RNA was isolated and RNA-seq was performed comparing control and knockdown RNA.

first: Erika

middle: S

last: Dahl

first: Raquel

last: Buj

first: Kelly

last: Leon

first: Jordan

last: Newell

last: Imamura

first: Benjamin

last: Bitler

first: Nathaniel

middle: W

last: Snyder

first: Katherine

last: Aird


title: BET bromodomain inhibitor iBET151 impedes human ILC2 activation and prevents experimental allergic lung inflammation

status: Public on Mar 23 2019

first: Mayuri

last: Gogoi

state: Cambridgeshire

address: Francis Crick Avenue

department: PNAC

institute: Laboratory of Molecular Biology

summary: Group 2 innate lymphoid cells (ILC2) increase in frequency in eczema and allergic asthma patients, and thus represent a new therapeutic target cell for type-2 immune-mediated disease. The bromodomain and extra-terminal (BET) protein family of epigenetic regulators are known to support the expression of cell cycle and pro-inflammatory genes during type-1 inflammation, but have not been evaluated in type-2 immune responses.  We isolated human ILC2 and examined the capacity of the BET protein inhibitor, iBET151, to modulate human ILC2 activation following IL-33 stimulation. iBET151 profoundly blocked expression of genes critical for type-2 immunity, including type-2 cytokines, cell surface receptors and transcriptional regulators of ILC2 differentiation and activation.  Furthermore, in vivo administration of iBET151 during experimental mouse models of allergic lung inflammation potently inhibited lung inflammation and airways resistance in response to cytokine or allergen exposure.  Thus, iBET151 effectively prevents human ILC2 activation and dampens type-2 immune responses.

overall_design: Lymphocytes were enriched from 150 ml human peripheral blood using lymphoprep (Axis-Shield) according to the manufacturer’s instructions. The collected PBMC fraction was washed with PBS/2% FCS followed by red blood cell lysis with ACK solution for 5 min at room temperature. To pre-enrich ILCs, cells were stained with biotinylated anti-CD3, CD14 and B220 in the presence of TruStain FcX (anti-CD16/CD32) followed by depletion with M280 streptavidin beads (Invitrogen). Subsequently, the enriched fraction was stained with CD45-AF488, CD127-PE-Cy7, CD117-PE, CRTH2-BV421, a lineage cocktail (APC-streptavidin, CD8a, CD11b, CD11c, FceRIa, CD123, CD20, CD56, CD71) and Draq7-viability dye, followed by sorting of ILCs as indicated on an Influx FACS. ILC2s designated as Draq7–(live cells)CD45+Lin–(streptavidin-APC, CD8a, CD11b, CD11c, FceRIa, CD123, CD20, CD56, CD71)CD127+CRTH2+ cells, were purified by flow cytometry from the peripheral blood of 3 individual donors and cultured separately. Purified human ILC2s were grown in media containing IL-2 and IL-7 (both at 10 ng/ml) with IL-33 (10 ng/ml) added to the cultures every alternate week.  ILC2s were then rested for 3 days in IL-2 and IL-7-containing media (no IL-33) before being divided and added to the following culture conditions: IL-2/IL-7, +/-IL33 (all at 10 ng/ml), +/- 125 nM iBET151 for 24 hours. Cells were pelleted and RNA prepared for RNAseq analysis.

first: Bernhard

last: Kerscher

first: Jillian

middle: L

last: Barlow

first: Batika

last: Rana

first: Helen

last: Jolin

first: Michelle

last: Bartholomew

first: Deepali

last: Jhamb

first: Ashutosh

last: Pandey

first: David

middle: F

last: Tough

first: Antoon

last: van Oosterhout

last: McKenzie


title: G1E cells infected with control (HMD empty vector), human GATA1, or human GATA1 mutant cDNA

status: Public on Mar 20 2019

first: Vijay

last: Sankaran

address: 1 Blackfan Cr

institute: Boston Children's Hospital

overall_design: 3 Biological replicates per condition for RNA-seq

first: Jeffrey

last: Verboon

first: Claudia

last: Fiorini

first: Leif

last: Ludwig