title: Role of the LytSR two-component regulatory system in Staphylococcus lugdunensis biofilm formation and pathogenesis

city: Lyon

first: jean-michel

last: flaman

address: 28 rue Laennec

department: CRCL

institute: Inserm 1052

summary: Staphylococcus lugdunensis is a coagulase negative Staphylococcus, recognized as a virulent pathogen. It is responsible for a wide variety of infections, some of which being associated with biofilm production, such as implanted medical devices infections or endocarditis. However, little is known about S. lugdunensis virulence regulation. Two-component regulatory systems (TCS) play critical roles in bacterial adaptation, survival and virulence. Among them, LytSR is widely conserved, but has variable roles in different organisms, all being connected to metabolism or cell death and lysis into biofilms. Therefore, we investigated here the functions of LytSR in S. lugdunensis pathogenesis.    Deletion of lytSR in S. lugdunensis DSM 4804 strain did not alter neither the susceptibility to Triton X-100 induced autolysis nor the death induced by antibiotics targeting cell wall synthesis. Interestingly, ΔlytSR biofilm was characterized by a lower biomass, a lack of tower structures and a higher rate of dead cells compared to the wild-type strain. Virulence towards Caenorhabditis elegans using slow-killing assay was significantly reduced for the mutant compared to the wild-type strain. In contrast, the deletion of lytSR had no effect on S. lugdunensis cytotoxicity towards the human keratinocyte cell line HaCaT. Transcriptional analyses conducted at mid- and late-exponential phases showed that lytSR deletion affected the expression of 286 genes. Most of them were involved in basic functions such as metabolism of amino acid, carbohydrates and nucleotides. Furthermore, LytSR appeared to be involved in the regulation of genes coding known or putative virulence and colonization factors, including the fibrinogen-binding protein Fbl, the major autolysin AtlL and the type VII secretion system.    Overall, our data suggest that LytSR TCS is implicated in S. lugdunensis pathogenesis, through its involvement in biofilm formation and potentially by the control of genes encoding putative virulence factors.

overall_design: Transcriptome of S. lugdunensis ΔlytSR and WT strains was compared by DNA microarray at 6 h and 8 h of growth. Three replicates per strain were included.

first: Sandrine

last: Dahyot

first: Virginie

last: Oxaran

first: Maïté

last: Niepceron

first: Eddy

last: Dupart

first: Stéphanie

last: Legris

first: Jennifer

last: Didi

first: Thomas

last: Clamens

first: Olivier

last: Lesouhaitier

first: Yasmine

last: Zerdoumi

first: Jean

last: Flaman

first: Martine

last: Pestel-Caron


title: Expression analysis of Caulobacter crescentus NA1000 bearing a CCNA_00690 (cdnL) deletion [dcdnL] vs Caulobacter crescentus NA1000 wild type [wt]

status: Public on Nov 05 2019

city: Lausanne

first: Diego

last: Gonzalez

address: Quartier UNIL-Sorge, Bâtiment Biophore

department: Department of Fundamental Microbiology

country: Switzerland

institute: University of Lausanne

summary:
Investigation of whole genome gene expression level changes in a Caulobacter crescentus NA1000 dcdnL mutant, compared to the wild-type strain.
In bacteria, transcription of housekeeping genes required for metabolic homeostasis and cell proliferation is guided by the sigma factor σ70. The conserved CarD-like transcriptional regulator, CdnL, associates with promoter regions where σ70 localizes and stabilizes the open promoter complex. Caulobacter crescentus cells lacking CdnL have severe morphological and growth defects. Our microarray experiment demonstrates how cdnL deletion affects the transcriptome of Caulobacter crescentus.

overall_design: A six chip study using total RNA recovered from three separate wild-type cultures of Caulobacter crescentus NA1000 and three separate cultures of a mutant strain, Caulobacter crescentus NA1000 dcdnL (deletion of CCNA_00690), in which the cdnL gene coding for an RNA polymerase associated factor is deleted. Each chip measures the expression level of 3933 genes from Caulobacter crescentus NA1000 with 3 probes per gene and with three-fold technical redundancy.

first: Justine

last: Collier-Close


title: Connect-seq to superimpose molecular on anatomical neural circuit maps

status: Public on Nov 06 2019

city: Seattle

first: Linda

last: Buck

state: Washington

address: 1100 fairview ave North

department: Basic Sciences

country: USA

institute: Fred Hutchinson Cancer Research Centre

type: Expression profiling by high throughput sequencing

summary: We report RNA sequencing of single virus-infected neurons upstream of neurons producing corticotropin-releasing hormone (CRH). Analyses of PRV-infected transcriptomes reveal neurons upstream of CRH neurons express diverse and a large variety of signaling molecules. The molecular identities of upstream neurons can be used to superimpose a molecular on neuroanatomical circuit maps.

overall_design: Identification of neuronal signalling molecules in individual neuronal components in neural circuits


title: LSD1 mediates AKT activity in PIK3CA mutant colorectal cancer [ChIP-Seq]

city: Boston

first: Robert

last: Policastro

state: Massachusetts

address: 451 D St Unit #600

institute: Ensoma Inc.

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Activation of the epithelial-mesenchymal transition (EMT) program is a critical mechanism for initiating cancer progression and migration. Colorectal cancers (CRCs) contain many genetic and epigenetic alterations that can contribute to EMT. Mutations activating the PI3K/AKT signaling pathway are observed in >40% of patients with CRC contributing to increased invasion and metastasis. Little is known about how oncogenic signaling pathways such as PI3K/AKT synergize with chromatin modifiers to activate the EMT program. Lysine Specific Demethylase 1 (LSD1) is a chromatin-modifying enzyme that is overexpressed in colorectal cancer (CRC) and enhances cell migration. In this study we determine that LSD1 expression is significantly elevated in CRC patients with mutation of the catalytic subunit of PI3K, PIK3CA, compared to CRC patients with WT PIK3CA. LSD1 enhances activation of the AKT kinase in CRC cells through a non-catalytic mechanism, acting as a scaffolding protein for the transcription-repressing CoREST complex. Additionally, growth of PIK3CA mutant CRC cells is uniquely dependent on LSD1.  Knockdown or CRISPR knockout of LSD1 blocks AKT-mediated stabilization of the EMT-promoting transcription factor Snail and effectively blocks AKT-mediated EMT and migration. Overall we uniquely demonstrate that LSD1 mediates AKT activation in response to growth factors and oxidative stress, and LSD1-regulated AKT activity promotes EMT-like characteristics in a subset of PIK3CA mutant cells.

overall_design: Examination of histone modification and transcription factor binding in SW480 cells.

first: Heather

middle: M

last: O'Hagan

first: Samuel

middle: A

last: Miller


title: Rac -dependent signaling from keratinocytes promotes differentiation of intradermal white adipocytes

city: Kobe

first: Takehiko

last: Ueyama

state: Hyogo

address: Rokkodai-cho 1-1, Nada-ku

department: Biosignal Research Center

country: Japan

institute: Kobe university

summary: Rac signaling affects numerous downstream targets; however, few studies have established in vivo levels using model animals. In  the present study, wWe generated mice with a single knockout (KO) of Rac1 (Keratin5 (K5)-Cre;Rac1flox/flox, hereafter Rac1-KO) and double KO of Rac1 and Rac3 (K5-Cre;Rac1flox/flox;Rac3−/−, hereafter Rac1/Rac3-DKO) in keratinocytes. Hairless phenotype in Rac1-KO mice was markedly exacerbated in Rac1/Rac3-DKO mice. Notably, Rac1-KO mice also exhibited thinner dermal white adipose tissue, which was considerably further reduced in Rac1/Rac3-DKO mice. DNA microarray using primary keratinocytes from Rac1/Rac3-DKO mice exhibited decreased mRNA levels of Bmp2, Bmp5, Fgf20, Fgf21, Fgfbp1, and Pdgf-alpha. Combinational treatment with BMP2 and FGF21, BMP2 and FGF20, or PDGF and FGF21 in culture medium, but not individual purified recombinant proteins, could differentiate 3T3-L1 fibroblasts into adipocytes, as could culture media obtained from primary keratinocytes (both human and mouse). Conversely, addition of anti-BMP2 or anti-FGF21 antibodies into the culture medium inhibited fibroblast differentiation. Furthermore, combinational treatment with BMP2 and FGF21 promoted adipocyte differentiation only of rat primary white adipocyte precursors but not rat primary brown adipocyte precursors. Notably, brown adipogenesis by FGF21 was inhibited by BMP2. Thus, we here proposed a novel paracrine pathway from keratinocytes to intradermal pre-adipocytes using Rac1/Rac3-DKO mice, which is Rac-dependent and functions as an inducer of white adipocytes.

overall_design: The expression levels of mRNA in mouse primary keratinocytes: from Rac3-KO mice and Rac1/Rac3-DKO mice.


title: A lysosome-plasma membrane-sphingolipid axis linking lysosomal storage to cell growth arrest

city: Pieve Emanuele-Milan

first: Giulia

last: Solda'

address: via Rita Levi Montalcini 4

country: Italy

institute: Humanitas University

summary: We used human fibroblasts loaded with sucrose as a simple model of lysosomal accumulation, and we verified by high-throughput RNA sequencing which processes were altered by the treatment at two different time points. Moreover, we hypothesized that ceramide production due to the ectopic catabolism of plasma-membrane glycosphingolipids is involved in  cell cycle arrest upon sucrose treatment.  To test this hypothesis, we treated sucrose-loaded fibroblasts with two specific inhibitors for the β-glucosidases GBA1 and GBA2, and analyzed the cell responses using high-throughput RNA sequence.

overall_design: Poly(A) RNA capture followed by multiparallel sequencing performed in human skin fibroblasts cultured in presence or absence of sucrose. Three series of experiments have been performed. Series nr 1 includes samples cultured in presence or absence of sucrose for 14 days, and was performed on 3 different fibroblast cell lines (F1, L37 and L40, biological replicates), each in duplicate or triplicate, N=14. Series nr 2 includes samples cultured in presence or absence of sucrose for 2 days, and was performed on one fibroblast cell lines (L40) in triplicate, N=6. Series nr 3 includes samples cultured in presence of sucrose for 14 days, and treated for the last 48 hours with CBE and AMP-DNM, which specifically inhibits β-glucosidases GBA1 and GBA2 (N=3).

last: Soldà

first: Letizia

last: Straniero

first: Maura

last: Samarani

first: Massimo

last: Aureli


title: Transcriptional response of Tetrahymena thermophila to exogenous cholesterol

city: Barcelona

first: Sebastián

middle: Rodrigo

last: Najle

state: Catalonia

address: Passeig Maritim de la Barceloneta, 37-49

country: Spain

institute: Institut de Biologia Evolutiva (CSIC-Universitat Pompeu Fabra)

summary: The model ciliate Tetrahymena thermophila satisfies its growth using triterpenoid alcohols, mainly tetrahymanol, as a surrogate of sterols.  When sterols are present in the environment, T. thermophila efficiently incorporates and modifies them. T. thermophila can modify exogenous sterols by desaturation at positions C5(6), C7(8) and C22(23), and also by de-ethylation of C24. Three out of four of the enzymes involved in the sterol modification pathway were previously described by our group. However, identification of the sterol C22 desaturase remained elusive, as well as other basic aspects of this metabolism. To get more insights into this peculiar metabolism we here perform a whole transcriptome analysis of T. thermophila in response to exogenous cholesterol. We found 356 T. thermophila genes to be differentially expressed after supplementation with cholesterol for two hours. Among those genes that were upregulated under our experimental conditions, we identified two genes belonging to the long-spaced family of desaturases that we propose as putative sterol C22 desaturases. Moreover, we determined that the inhibition of tetrahymanol synthesis after sensing exogenous cholesterol occurs by a transcriptional downregulation of genes involved in squalene synthesis and cyclization. Finally, we also identified several uncharacterized genes that can be putatively involved in sterols transport and signaling.

overall_design: mRNA profiles of T. thermophila strain CU428 (wild type) cultivated with (TREATED) and without (CONTROL) the addition of cholesterol to the growth media, by triplicate.

middle: R

first: Antonio

middle: D

last: Uttaro


title: Transcriptomics analysis of cells transfected with miR-183 cluster mimics and immunostimulated with poly(I:C)

status: Public on Nov 04 2019

city: Ottawa

first: John

middle: Paul

last: Pezacki

state: ON

address: 150 Louis-Pasteur Pvt

department: Chemistry and Biomolecular Sciences

country: Canada

institute: University of Ottawa

summary:
Our work demonstrated that miR-183 cluster regulates IFN production and signaling
We used microarrays to detail the effect of miR-183 cluster on hepatic gene expression during immunostimulation with poly(I:C). Our finding suggests enrichment of pathways involved in IFN production and signaling. Amoung the miR-183 repressed genes, we identifies PPP2CA and TRIM27, which are negative regulators of the innate immune response.

overall_design: Human liver heptoma cells (HepG2) were were transfected with control mimic (con-miR) or miR-183 cluster mimics (miR-96,miR-182 and mIR-183) individually for 48h and then immunostimulated with poly(I:C) for 24h. RNA was extracted and analysed using a Affymetrix Human Gene ST.2.0 array.

first: Nadine

last: Ahmed

first: Ragunath

last: Singaravelu


title: Pol5 is required for recycling of small subunit biogenesis factors and for formation of the polypeptide exit tunnel of the large ribosomal subunit

city: Goettingen

first: Markus

middle: T.

last: Bohnsack

address: Humboldtallee 23

department: Institute for Molecular Biology

country: Germany

institute: Goettingen University

summary: More than 200 assembly factors (AFs) are required for the production of ribosomes in yeast. The stepwise association and dissociation of these AFs with the pre-ribosomal subunits occurs in a strictly hierarchical manner to ensure correct maturation of the pre-rRNAs and assembly of the ribosomal proteins. Although decades of research have provided a wealth of insights into the functions of many AFs, others remain poorly characterized. Pol5 was initially classified with B-type DNA polymerases, however, several lines of evidence indicate the involvement of this protein in ribosome assembly. Here, we show that depletion of Pol5 affects the processing of pre-rRNAs destined for the both the large and small subunits. Furthermore, we identify binding sites for Pol5 in the 5’ external transcribed spacer and within domain III of the 25S rRNA sequence. Consistent with this, we reveal that Pol5 is required for recruitment of ribosomal proteins that form the polypeptide exit tunnel in the LSU and that depletion of Pol5 impairs the release of 5’ ETS fragments from early pre-40S particles. The dual functions of Pol5 in 60S assembly and recycling of pre-40S AFs suggest that this factor could contribute to ensuring the stoichiometric production of ribosomal subunits.

overall_design: Wild-type BY4741 yeast cells or cells expressing genomically C-terminally His-Tev-ProtA-tagged Pol5 were analysed by CRAC (Bohnsack et al., 2012).

first: Christina

last: Braun

first: Philipp

last: Hackert

first: Catharina

last: Schmid

middle: T

first: Katherine

middle: E

first: Jorge

last: Perez-Fernandez


title: Extracellular matrix hydrogel derived from decellularized tissues enables endoderm organoids culture

city: Würzburg

first: Kai

last: Kretzschmar

address: Josef-Schneider-Str. 2

institute: University Hospital Würzburg

summary: Organoids have extensive therapeutic potential and are increasingly opening up new avenues within regenerative medicine. However, the clinical application is greatly limited by the lack of effective GMP-compliant systems for organoid expansion in culture. Here, we envisage that the use of extracellular matrix hydrogels derived from decellularized tissues (DT) can provide an environment capable of directing cell growth. These gels possess the biochemical signature of tissue-specific ECM, and have the potential for clinical translation. Gels from decellularized porcine small intestinal mucosa/submucosa enable formation and growth of endoderm-derived human organoids, such as gastric, liver, pancreas, and small intestine. ECM gels could be used for direct organoids derivation from human biopsies, multiple passages with transcriptomic signature comparable to gold standard culture system, and in vivo organoid delivery. The development of these ECM-derived hydrogels opens up the potential for human organoids to be used clinically.

overall_design: We generated transcriptome data from human hepatocyte organoids (HepOrg) and human liver ductal organoids (DuctOrg) grown in standard BME hydrogel or newly developed ECM hydrogel purified from piglet liver ECM.

first: Giovanni

middle: G

last: Giobbe

first: Claire

last: Crowley

first: Camilla

last: Luni

first: Sara

last: Campinoti

first: Moustafa

last: Khedr

first: Martina

last: De Santis

first: Elisa

last: Zambaiti

first: Qianjiang

last: Hu

first: Federica

last: Michielin

first: Laween

last: Meran

first: Luca

last: Urbani

first: Anna

last: Manfredi

first: Gijs

last: van Son

first: Simon

last: Eaton

first: Davide

last: Cacchiarelli

first: Vivian

middle: S

last: Li

first: Hans

last: Clevers

first: Paola

last: Bonfanti

first: Nicola

last: Elvassore

first: Paolo

last: De Coppi


city: Derio

first: Ana

middle: Maria

last: Aransay

state: BIZKAIA

address: Parque tecnologico de Bizkaia, Building 801-A

department: Genome Analysis Platform

institute: CIC bioGUNE

summary: Glucocorticoids (GCs) are important hormones involved in the regulation of multiple physiologic functions. GCs are also widely used in anti-inflammatory/immunosuppressant drugs. GCs are synthesized by the adrenal cortex as part of the hypothalamus-pituitary-adrenal axis and also by intestinal epithelial cells, among other peripheral sites. GCs are one of the main therapy choices for the exacerbations of inflammatory bowel disease, but they are not useful to prolong remission, and development of tolerance with secondary treatment failure is frequent. Thus, GC actions at the intestinal epithelial level are of great importance, both physiologically and pharmacologically. We generated a tamoxifen-inducible nuclear receptor subfamily 3 group C member 1 (NR3C1)ΔIEC mouse model to study the effects of GCs on epithelial cells in vivo. Nr3c1 deletion in epithelial cells of the small intestine and colon was associated with limited colonic inflammation at 1 wk postdeletion, involving augmented epithelial proliferation and mucus production, plus local and systemic immune/inflammatory changes. This phenotype regressed substantially, but not completely, after 2 wk. The mechanism may involve augmented inflammatory signaling by epithelial cells or defective barrier function. We conclude that the epithelial GC receptor plays a significant role in colonic homeostasis in basal conditions, but its deficiency can be compensated in the short term. Future studies are required to assess the impact of Nr3c1 deletion in other conditions such as experimental colitis.-Aranda, C. J., Arredondo-Amador, M., Ocón, B., Lavín, J. L., Aransay, A. M., Martínez-Augustin, O., Sánchez de Medina, F. Intestinal epithelial deletion of the glucocorticoid receptor NR3C1 alters expression of inflammatory mediators and barrier function.

overall_design: mRNA profile from Colonic Epithelial Cells of Wild Type and Nr3c1IEC-/- mice were generated by deep sequencing in triplicate.

first: Carlos

middle: J

last: Aranda

first: María

last: Arredondo-Amador

first: Borja

last: Ocón

first: José

middle: L

last: Lavín

first: Olga

last: Martínez-Augustin

first: Fermín

last: de Medina


title: Neutralization of Oxidized Phospholipids Restrains Nonalcoholic Steatohepatitis

city: La Jolla

first: Jason

middle: Samuel

last: Seidman

state: CA

address: 9500 Gilman Drive, Mail code 0651

department: Cellular and Molecular Medicine

institute: University of California San Diego

summary: Oxidized phospholipids (OxPL) are pro-inflammatory and pro-atherogenic, but their roles in non-alcoholic steatohepatitis (NASH) are unknown. Here, we show that OxPL accumulate in human and murine NASH. Using a transgenic mouse that expresses a functional single chain variable fragment of E06, a natural antibody that neutralizes OxPL, we demonstrate the casual role of OxPL in NASH. Targeting OxPL in hyperlipidemic Ldlr-/- mice decreased multiple aspects of NASH, including steatosis, inflammation, fibrosis, hepatocyte death and progression to hepatocellular carcinoma. Mechanistically, we found that OxPL promote ROS accumulation to induce mitochondrial dysfunction in hepatocytes. Neutralizing OxPL in AMLN diet-fed Ldlr-/- mice reduced oxidative stress, improved hepatic and adipose tissue mitochondrial function and fatty acid oxidation. Since neutralizing OxPL also protects against atherogenesis, targeting OxPL may be an effective therapeutic strategy for both NASH and atherosclerosis.

overall_design: Bulk RNA-Seq of RNA isolated from Ldlr-/- or E06-scFvLdlr-/- mice fed AMLN diet.

first: Sun

last: Xiaoli


city: Pozzuoli

first: Lucio

last: Di Filippo

address: Via Campi Flegrei, 34

institute: TIGEM

summary: Organoids have huge therapeutic potential and are increasingly opening up new avenues within regenerative medicine. However, the clinical application is greatly limited by the lack of effective GMP-complaint systems for organoid expansion in culture. Here, we envisage that the use of extracellular matrix hydrogels derived from decellularized tissues (DT) can provide the instructive native environment necessary to direct cell growth. These gels possess the biochemical signature of tissue-specific ECM, and have the potential for clinical translation due to their natural, non-carcinogenic origin.  We show that gels developed from decellularized porcine small intestinal mucosa have similar physiological range and mechanical properties to commercially available gels. ECM-derived gels enable formation and growth of endoderm-derived organoids and, moreover, supports in vivo organoid growth. The development of these ECM-derived hydrogels opens up the potential for human organoids to be used clinically.

overall_design: RNA-seq profiles of organoids expanded on a decellularized porcine small intestinal mucosa-derived gel respect to control on Matrigel, with 4 replicates from the same pediatric small intestine cell line.

first: Giovanni

last: Giobbe

first: Camilla

last: Luni

first: Claire

last: Crowley

first: Sara

last: Campinoti

first: Moustafa

last: Khedr

first: Martina

last: De Santis

first: Elisa

last: Zambaiti

first: Qianjiang

last: Hu

first: Kai

last: Kretzschmar

first: Luca

last: Urbani

first: Laween

last: Meran

first: Federica

last: Michielin

first: Anna

last: Manfredi

last: Clever


title: Impact of UCP1 and FGF21 for the development of diet-induced obesity at standard housing temperatures

city: Neuherberg

first: Dominik

last: Lutter

address: Ingolstädter Landstraße 1

department: Institute for Diabetes and Obesity

institute: Helmholtz Zentrum München

summary: Global RNA sequencing analysis of brown fat (BAT), inguinal white fat (iWAT), liver and muscle (quadriceps) of high-fat diet fed WT, FGF21 KO, UCP1 KO and UCP1/FGF21 double KO mice.

overall_design: The experiment was performed in homozygous, male WT, FGF21 KO, UCP1 KO and UCP1/FGF12 double KO (dKO) mice. All mice were bred, born and weaned at 30°C. At the age of 10 to 12 weeks, mice were transferred from 30°C to room temperature and chow diet was changed to high-fat diet for 3 weeks. At the end of the experiment, mice were euthanized after 2-3 hours food withdrawal, followed by serum and tissue sample collection.

first: S

last: Keipert

first: M

last: Jastroch


title: Developmental Cytoplasmic-to-Nuclear Translocation of RNA-Binding Protein HuR Is Required for Adult Neurogenesis

city: Beijing

first: Jing

last: Sun

address: No. 1 West Beichen Road

department: IGDB

country: China

institute: Chinese Academy of Sciences

summary: Although neurogenesis in the adult brain recapitulates processes that occur during embryonic development, adult neurogenesis exhibits distinct characteristics from its embryonic counterpart. However, the intrinsic mechanism underlying the differential regulation of neurogenesis between these two stages remains unclear. Herein, we show that the ablation of RNA-binding protein HuR in neural stem cells (NSCs) impairs adult, but not embryonic, neurogenesis. HuR is predominantly expressed in the cytoplasm of embryonic NSCs but translocates into the nucleus of adult NSCs. Transcriptomic analysis of HuR-deficient adult NSCs revealed that nuclear HuR primarily regulates alternative splicing of numerous premRNA transcripts, including focal adhesion kinase (FAK). HuR-deficient adult NSCs generate increased FAK mRNA isoforms with shorter 5’ UTRs, leading to enhanced FAK mRNA translation and hyperactivated FAK signaling, and inhibition of FAK ameliorates defective adult neurogenesis and impaired hippocampus-dependent learning in HuR-deficient mice. Taken together, these findings reveal novel mechanistic insights into the differential regulation of embryonic and adult neurogenesis through developmental cytoplasmic-to-nuclear translocation of HuR in NSCs.

overall_design: Total RNA was extracted from HuRf/f aNSCs infected with lenti-dCre or lenti-Cre using TRIzol (Invitrogen). The integrity of the extracted total RNA was analyzed using Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA, USA). The samples with RNA Integrity Number (RIN) ≥ 7 were subjected to the subsequent analysis. The libraries were constructed using TruSeq Stranded mRNA LTSample Prep Kit (Illumina, San Diego, CA, USA) according to the manufacturer’s instructions. The standard Illumina protocol was used to prepare the libraries for RNA-Seq. RNA-seq was performed using Illumina HiSeq2500  > 45 million 2 × 100 reads per sample was produced and alignment was performed using mouse genome database GRCm38 version 67. Only uniquely and properly mapped read pairs were used for further analysis. FPKM value of each gene was calculated using cufflinks, and the read counts of each gene were obtained by htseq-count. Differentially expressed genes (DEGs) were identified using the DESeq (2012) functions estimateSizeFactors and nbinomTest. P value < 0.05 and foldChange >2 or foldChange < 0.5 was set as the threshold for significantly differential expression. Hierarchical cluster analysis of DEGs was performed to explore genes expression pattern. GO enrichment and KEGG pathway enrichment analysis of DEGs were respectively performed using R based on the hypergeometric distribution. The alternatively splicing analysis of differentially regulated transcripts isoforms or exons was performed using multivariate analysis of transcript splicing (MATS).

first: Yuhan

last: Wang

first: Weixiang

last: Guo


title: Mevalonate metabolism-dependent protein geranylgeranylation regulates thymocyte egress

city: Memphis

first: Hongbo

last: Chi

state: TN

address: 262 Danny Thomas Place

department: Immunology

institute: St Jude Children's Research Hospital

summary:
Loss of Pggt1b leads to marked defects in thymocyte egress and T cell lymphopenia in peripheral lymphoid organs in vivo
CD4SP thymocytes (n = 4 per genotype) were isolated from the thymus of WT and Pggt1b KO mice

overall_design: In total 8 samples were analyzed: (4 WT and 4 Pggt1b KO).

first: Xingrong

last: Du

first: Yogesh

last: Dhungana


title: LSD1 mediates AKT activity in PIK3CA mutant colorectal cancer [RNA-Seq]

first: Robert

last: Policastro

overall_design: RNA sequencing profiles were generated for LSD1 knockdown and GSK-LSD1 treated SW480 cells as well as LSD1 knockdown in HT29 cells compared to controls.

first: Heather

last: O'Hagan

first: Samuel

last: Miller


title: Transcriptome profiling of subcutaneous and visceral adipose tissue from obese individuals

city: Kansas City

first: Elin

last: Grundberg

state: Missouri

address: 2401 Gillham Road

institute: CHILDREN'S MERCY HOSP (KANSAS CITY, MO)

summary: Distinct characteristics of adipose tissue at different localization of human body has shown greater significance in development of metabolic disorders. Visceral adipose tissue in particular is known to be associated with obesity related metabolic complications that include type II diabetes. In this experiment, we attempt to profile transcriptome signatures of adipocyte, stromal vascular fraction (SVF) and adipose tissue from subcutaneous and visceral adipose tissue from obese individuals.

overall_design: Comparitive analysis of mRNA from subcutaneous and visceral adipose tissue using RNASeq

first: Jinchu

last: Vijay


title: Single Cell RNASeq profiling of stromal vascular fraction from visceral adipose tissue of young individual

summary: Visceral adipose tissue (VAT) is studied extensively to understand its involvement in development of metabolic disorders. In this experiment, we attempt to profile different cell types in VAT of a healthy young individual.

overall_design: Profiling of different cell types found in adipose derived stromal vascular fraction (SVF) of young individual


title: Transcriptome analysis in individual mouse blastocyst with Xist deletion

city: Isehara-shi

first: Atsushi

last: Fukuda

institute: Tokai University

summary: We reprot transcriptome states in mouse blastocyst cultured with various conditons as well as Xist deletion. By comparing the gene expression states of blastocyst grown in different culture conditions with Xist deletion, we identified the differentially expressed genes, which are dependent culture conditions and Xist RNA.

overall_design: Examination of transcriptome states in mouse 18 blastocysts


title: Single-cell sequencing of enteroendocrine cells (EEs) in Drosophila midgut.

first: xi

last: rongwen

address: No. 7 Science Park Road, zhongguancun Life Science Park, Changping District,

institute: National Institute of Biological Sciences, Beijing

summary: Enteroendocrine cells (EEs) in the intestinal epithelium have important endocrine functions, yet this cell lineage exhibits great local and regional variations that have hampered detailed characterization of EE subtypes. Through single cell RNA-sequencing analysis, combined with a collection of neuropeptide/receptor knock-in strains, here we provide a comprehensive analysis of cellular diversity, spatial distribution and transcription factor (TF) code of EEs in adult Drosophila midgut. We identify 10 major EE subtypes that totally produced 14 different classes of hormone peptides. Each EE on average co-produces 2-5 different classes of hormone peptides. Functional screen with subtype-enriched TFs suggests a combinatorial TF code that controls EE cell diversity, with class-specific TFs Mirr and Ptx1that respectively define two major classes of EEs, and regional TFs such as Esg, Drm, Exex and Fer1that further define regional EE identity. Our single-cell data should greatly facilitate Drosophila modeling of EE differentiation and function.

overall_design: FACS sort EE cells using CG32547-KI-GAL4> uas-GFP line and perform 10X genomics single cell sequencing. Raw data was further processed by Seurat algorithm and EE subtypes were visualized by generating a t-distributed stochastic neighborhood embedding (t-SNE) plot.

first: Xingting

first: Chang

last: Yin

first: Fu

last: Yang

first: Yongchao

last: Zhang

first: Rongwen

last: Xi


title: MYCi975 regulates MYC target genes

city: Chicago

first: Zachary

middle: Rockow

last: Chalmers

state: IL

address: 303 E. Superior Ave

institute: Northwestern University

summary: We report MYCi975 regulates MYC taraget genes in PC3 and P493-6 cells by deep sequencing (RNA-seq).

overall_design: We used PC3 cells with high endougenous c-MYC level and P493-6 with induceble c-MYC expressing system to demonstrate MYCi975 sepcifically regulates MYC target genes. Transcriptome profiling of PC3 and P493-6 treated with MYCi975, and P493-6 treated with tetracycline to turn off MYC expression were generated.

first: Sarki

last: Abdulkadir

first: Huiying

last: Han


title: ZFP281 recruits MYC to active promoters in regulating transcriptional initiation and elongation

city: Nanjing

first: Xiaoxu

last: Liu

state: Jiangsu

address: 2 Sipailou

department: Institute of Life Sciences

institute: Southeast University

summary: In this study, we analyzed the function of ZFP281 in transcription control by genome wide ChIP-Seq analyses in mouse embryonic stem cells.

overall_design: SRP175199 : PRJNA512974_ChIP-seq of whole cell extract in mES cells

first: Chengq

last: i Lin


title: Condensin II is recruited to promoters by pRb and regulates expression of divergently paired genes [expression array]

city: London

first: Fred

last: Dick

address: Cancer Research Labs 790 Commissioners Road East

institute: Western University

summary: In this study, use of a gene-targeted mouse model that is defective for pRb-condensin II interactions, Rb1L, has revealed instances where condensin II localization in interphase nuclei is dependent on pRb. Condensin II binding overlaps significantly with marks of active chromatin, and is enriched at the promoters of genes, including closely spaced divergent promoters. Interestingly, there are some bidirectional promoters where condensin II binds in an pRb-dependent manner and transcription of only one of the two genes is upregulated in Rb1L/L MEFs, suggesting the pRb-condensin II complex may be acting as a repressor or an insulator at distinct genomic locations to influence transcription. Chromatin conformations at these locations demonstrated that the pRb-condensin II complex may be responsible for maintaining more static, long-range interactions. In addition, pRb also recruits TFIIIC, another condensin II interactor, to many genomic loci. These recently discovered non-canonical transcriptional functions of the pRb-condensin II complex may represent an additional mechanism of pRb-mediated tumor suppression.

overall_design: Total RNA from proliferating MEFs was isolated. Three Rb1+/+ and Rb1L/L MEF pairs were used for this analysis and expression between genotypes was compared.

first: Frederick

first: Aren

last: Marshall


title: BEN-solo factors partition active chromatin to ensure proper gene activation in Drosophila

city: CANBERRA

first: Jiayu

last: Wen

state: ACT

address: 133 Garran Rd

department: Department of Genome Science

country: Australia

institute: John Curtin School of Medical Research, The Australian National University

summary: We generated ChIP-seq datasets for four transcriptional regulators, Elba1, Elba2, Elba3 and Insv from wild-type, cognate mutants and non-cognate mutants. We generated ChIP-nexus data for these four factors. We produced RNA-seq and PRO-seq data from wild-type and mutant embryos.

overall_design: The ChIP-seq datasets for four transcriptional regulators, Elba1, Elba2, Elba3 and Insv from wild-type, cognate mutants and non-cognate mutants allow identification of factor binding in these five genetic conditions. The ChIP-nexus data is used to increase the resolution of TF binding sites and determine symmetric versus asymmetric binding patterns. The RNA-seq data was used to determine gene expression changes upon factor deletion. The PRO-seq data was used to determine real-time transscription changes in mutant embryos for each factor.

first: Qi

last: Dai


title: Single Cell RNASeq profiling of stromal vascular fraction from Subcutaneous and visceral adipose tissue

first: Elin

last: Grundberg

summary: The complex relationship between metabolic disease risk and body fat distribution in humans involves cellular characteristics which are specific to each body fat compartment.  We applied single-cell RNA sequencing (scRNA-Seq) to identify these depot-specific differences in the stromal vascular fraction of visceral (VAT) and subcutaneous (SAT) adipose tissue of obese individuals. We characterized multiple immune cells, endothelial cells, fibroblasts, adipose and hematopoietic stem cell progenitors. Subpopulations of adipose-resident immune cells were found to be metabolically active and associated with metabolic disease status including a population of potential dysfunctional CD8+ T cells expressing metallothioneins. Adipocyte progenitors were identified at different stages of adipogenesis where a subset was pronounced in type 2 diabetics. Depot-specific analysis revealed a class of adipocyte progenitors unique to VAT possessing features similar to ‘inducible brown preadipocytes’. Our first generation human scRNA-Seq transcriptome atlas across fat depots provides a new resource to dissect functional genomics of metabolic disease.

overall_design: Profiling of different cell types found in adipose derived stromal vascular fraction (SVF) of obese individuals with or without type 2 diabetes

first: Jinchu


title: Condensin II is recruited to promoters by pRb and regulates expression of divergently paired genes [ChIP-seq]

overall_design: Chromatin was isolated from passage 4 MEFs cross-linked with either ethylene glycol bissuccinimidylsuccinate (EGS) and formaldehyde or formaldehypde alone, and then sonicated so most chromatin was ≤ 400 bp. DNA fragments were immunoprecipitated using different antibodies, washed, then purified. DNA was prepared for sequencing according to the NEBNext Ultra II DNA library prep kit. Each protein was immoprecpitated in both wild type and Rb1L/L to compare locatlization between genotypes.


title: Microglial microRNAs mediate sex-specific responses to tau pathology

city: San Francisco

first: Lay

last: Kodama

address: 1650 Owens St.

department: Gladstone Institute of Neurological Disease

institute: University of California San Francisco

summary: Sex is a key modifier of neurological disease outcomes. Microglia are implicated in neurological diseases and modulated by miRNAs, but it is unknown whether microglial miRNAs have sex-specific influences on disease. We show that microglial miRNA expression differs in males and females, and loss of miRNAs leads to sex-specific changes in the microglial transcriptome and to tau pathology. These findings suggest microglial miRNAs influence tau pathogenesis in a sex-specific manner.

overall_design: Examination of mRNA expression in single cell microglia isolated from female and male Dicer KO P301S tauopathy mouse samples sorted by FACS into 96-well plates. Libraries were prepared using the Smart-Seq2 protocol.

first: Elmer

last: Guzman

first: Jon

middle: I

last: Etchegaray

first: Yaqiao

last: Li

first: Faten

last: Sayed

first: Lu

last: Zhou

first: Yungui

first: Lihong

last: Zhan

first: David

last: Le

first: Joe

middle: C

last: Udeochu

first: Claire

last: Clelland

first: Zuolin

last: Cheng

first: Guoqiang

last: Yu

first: Qingyun

first: Kenneth

last: Kosik

first: Li

last: Gan


title: Condensin II is recruited to promoters by pRb and regulates expression of divergently paired genes [4C-seq]

first: Fred

last: Dick

overall_design: Four Rb1+/+ and Rb1L/L MEF pairs were used for 4C-Seq. Interactions at two different viewpoints, the bidrectional promoter between Cdca3 and Usp5 and the bidirectional promoter between Pole and Pxmp2, were examined.

first: Frederick

middle: A

first: Aren

last: Marshall


title: Differentially expressed genes following ARID1A depletion

first: Qingyu

last: Luo

address: Panjiayuannanli No.17#, Chaoyang District

department: State Key Laboratory of Molecular Oncology

institute: Chinese Academy of Medical Sciences and Peking Union Medical College

summary: We konckdown ARID1A and then detected differentially expressed genes

overall_design: ARID1A was knockdown using shRNA then total RNA was collected


title: Affymetrix Mouse Diversity Genotyping Array for 129S2/SvPasCrl and 129S4/SvJae

city: Cambridge

first: Peter

last: Nicholls

state: MA

address: 9 Cambridge Center

institute: Whitehead Institute

summary: SNP comparison between two closely related 129S mouse strains

overall_design: Affymetrix SNP arrays were performed on singlicate samples according to the manufacturer's directions on DNA extracted from mouse ear skin samples

middle: K

last: Page


title: Brainstem control of transcription after spinal cord injury (SCI)

status: Public on Nov 03 2019

city: Miami

first: Ian

last: Hentall

state: FL

address: 1095 NW 14th Terrace

department: Neurological Surgery

institute: University of Miami Miller School of Medicine

summary: The spinal cord after injury shows altered transcription in numerous genes. We tested in a pilot study whether the nucleus raphé magnus, a descending serotonergic brainstem region whose stimulation improves recovery after incomplete spinal cord injury, can influence these transcriptional changes. Rats received 2 hours of low-frequency electrical stimulation in the raphé magnus three days after an impact contusion at segment T8. Comparison groups lacked injuries or activated stimulators or both. Immediately following stimulation, spinal cords were extracted, their RNA transcriptome sequenced, and differential gene expression quantified. Confirming many previous studies, injury primarily increased inflammatory and immune transcripts and decreased those related to lipid and cholesterol synthesis and neuronal signaling. Stimulation plus injury, contrasted with injury alone, caused significant changes in 43 transcripts (39 increases, 4 decreases), all protein-coding. Injury itself decreased only four of these 43 transcripts, all reversed by stimulation, and increased none of them. The non-specific 5-HT7 receptor antagonist pimozide reversed 25 of the 43 changes. Stimulation in intact rats principally caused decreases in transcripts related to oxidative phosphorylation, none of which were altered by stimulation in injury. Gene ontology (biological process) annotations comparing stimulation with either no stimulation or pimozide treatment in injured rats highlighted defense responses to lipopolysaccharides and microorganisms, and also erythrocyte development and oxygen transport (possibly yielding cellular oxidant detoxification). Connectivity maps of human orthologous genes generated in the CLUE database of perturbagen-response transcriptional signatures showed that drug classes whose effects in injured rats most closely resembled stimulation without pimozide include peroxisome proliferator-activated receptor agonists and angiotensin receptor blockers, which are reportedly beneficial in spinal cord injury. Thus, the initial transcriptional response of injured spinal cord to raphé magnus stimulation is upregulation of genes that in various ways are mostly protective, some probably located in recently arrived myeloid cells.

overall_design:
Group A: no SCI + no stim, n=4
Group B: SCI + no stim, n=3
Group C: SCI + stim, n=3
Group D: SCI + stim +pimozide, n=2
Group E: no SCI + stim, n=5

first: Walter

last: Jermakowicz


title: Chromera velia de novo transcriptome assembly, annotation and comparative transcriptomics

status: Public on Nov 02 2019

city: Brisbane

first: Amin

middle: Roushdy

last: Mohamed

state: Queensland

address: 306 Carmody Rd, University of Queensland

department: Agriculture and Food

institute: CSIRO

summary: Reef-building corals live in a mutualistic relationship with photosynthetic algae (family Symbiodiniaceae) that usually provide the bulk of the energy required by the coral host. This relationship is very sensitive to temperature stress, with as little as 1°C increase above mean in sea surface temperatures (SSTs) often leading to the collapse of the association. The meta-stability  of these associations has led to interest in the potential of more stress tolerant algae to supplement or substitute for the normal Symbiodiniaceae mutualists. In this respect, the apicomplexan-like microalga Chromera is of particular interest as it is considerably more temperature tolerant than are most members of the Symbiodiniaceae. Here we generated a de novo transcriptome for a Chromera strain isolated from a GBR coral (“GBR Chromera”) and compared to those of the reference strain of Chromera (“Sydney Chromera”), and to those of Symbiodiniaceae algae (Fugacium, Cladocopium and Breviolum), as well as the apicomplexan parasite, Plasmodium falciparum. By contrast with the Symbiodiniaceae, the two Chromera strains had a high level of sequence similarity evident by very low levels of divergence in orthologous genes. Although surveys of specific KEGG categories provided few general criteria by which true coral mutualists might be identified, they provide a molecular rationalization for the near ubiquitous association of Cladocopium strains with Indo-Pacific reef corals in general and with Acropora spp. in particular.  In addition, HSP20 genes may underlie the higher thermal tolerance shown by Chromera compared to Symbiodiniaceae

overall_design: We assembled a de novo transcriptome for a new strain of Chromera velia isolated from Montipora digitata at Magnetic Island on the Great Barrier Reef (GBR) "GBR Chromera". This transcripome was compared to the reference Chromera strain "Sydney Chromera", members of the Symbiodiniaceae and the apicomplexan parasite Plasmodium Falciparum. We sought to address two specific objectives here, in both cases use of a denovo-assembled transcriptome generated for a Chromear strain isolated from a GBR coral to estimate the degree of divergence between this strain and the Sydney strain. Second is to investigate the repretoires of genes that play roles in symbioses and stress tolerance between the two Chromera and three members of the Symbiodiniaceae including mtualists.

middle: R

first: Cheong

middle: X

last: Chan

first: Mark

last: Ragan

first: Jia

last: Zhang

first: Ira

last: Cooke

first: Eldo

last: Ball

last: Miller


title: Structure of a zinc-finger antiviral protein in complex with RNA reveals mechanism for selective targeting of CG-rich viral sequences

status: Public on Nov 01 2019

city: New York

first: Matthew

last: Takata

state: NY

address: 1230 York Ave

institute: The Rockefeller Univeristy

summary: Infection of animal cells by many viruses is detected and countered by a variety of means, including recognition of non-self nucleic acids. The zinc-finger antiviral protein (ZAP) depletes cytoplasmic RNA that is recognized as foreign in mammalian cells by virtue of its elevated CG dinucleotide content compared to endogenous mRNAs. Here, we determined a crystal structure of a protein-RNA complex containing the N-terminal, four-zinc finger human (h) ZAP RNA binding domain (RBD), and a CG dinucleotide-containing RNA target. The structure reveals in molecular detail how hZAP is able to bind selectively to CG-rich RNA. Specifically, the four zinc fingers create a basic patch on the hZAP RBD surface. The highly basic second zinc finger contains a pocket that selectively accommodates CG dinucleotide bases. Structure guided mutagenesis, crosslinking-immunoprecipitation-sequencing assays, and RNA affinity assays show that the structurally defined CG-binding pocket is not required for RNA binding per se in human cells. However, the pocket is a crucial determinant of high affinity specific binding to CG-dinucleotide-containing RNA. Moreover, variations in the RNA binding specificity a panel of CG-binding pocket mutants quantitatively predict their selective antiviral activity against a CG-enriched HIV-1 strain. Overall, the hZAP RBD:RNA structure provides an atomic-level explanation for how ZAP selectively targets foreign, CG-rich RNA.

overall_design: CLIP-seq data sets from 293T cells

first: Paul

middle: D

last: Bieniasz


title: Chromatin dynamics reveal circadian control of human in vitro islet maturation [RNA-seq]

first: Juan

last: Alvarez-Dominguez

address: 7 Divinity Ave

department: Stem Cell and Regenerative Biology

institute: Harvard University

summary: Stem cell-derived tissues could transform disease research and therapy, yet most methods generate functionally immature products. We investigate how human stem cells differentiate into pancreatic islets in vitro by profiling DNA methylation, chromatin accessibility, and histone modification changes. We find that enhancer potential is reset upon lineage commitment, and show how pervasive epigenetic priming steers endocrine cell fates. Modeling islet differentiation and maturation regulatory circuits reveals genes critical for generating endocrine cells and identifies circadian control as limiting for in vitro islet function. Entrainment to circadian feeding/fasting cycles triggers islet metabolic maturation by inducing cyclic synthesis of energy metabolism and insulin secretion effectors, including antiphasic insulin and glucagon pulses. Following entrainment, stem cell-derived islets gain persistent chromatin changes and rhythmic insulin responses with a raised glucose threshold, a hallmark of functional maturity, and function within days of transplantation. Thus, stem cell-derived tissues are amenable to functional improvement by circadian modulation.

overall_design:
RNA-seq of purified live cells at defined developmental stages from stem cells to mature pancreatic islet α/β cells.
RNA-seq of human islets / islets differentiated from human pluripotent stem cells entrained to circadian metabolic rhythms.


title: Effects of SLS or urushiol on the whole-genome scale transcription of human epidermal keratinocytes.

city: Seoul

first: Minsoo

last: Noh

address: 1 Gwanak-ro, Gwanak-gu

department: Pharmacy

country: South Korea

institute: Seoul National University

summary: To understand molecular pathogenesis of chemical induced contact dermatitis, cultured normal human epidermal keratinocytes (NHKCs) were treated with sub-cytotoxic concentration of sodium lauryl sulfate (SLS, 10 microM) and urushiol (1 microM). In analysis, 259 genes were selected as up-regulated DEGs in the SLS-treated NHKCs and 193 genes as down-regulated DEGs. In urushiol-treated NHKCs, 173 up-regulated and 124 down-regulated DEGs were identified. Gene Ontology (GO) biological process (BP) in the SLS-induced upregulated DEGS in NHKCs were inflammation-associated biological processes, such as inflammatory response (GO:0006954), cellular response to calcium ion (GO:0071277), immune response (GO:0006955) and cellular response to lipopolysaccharide (GO:0071222) and in urushiol-induced upregulated DEGs were inflammation-associated biological processes such as cytokine-mediated signaling pathway (GO:0019221) and inflammatory response (GO:0006954). SLS downregulated proteins in chemotaxis (GO:0006935), oxidation-reduction process (GO:0055114), cornification (GO:0070268), G1/S transition of mitotic cell cycle (GO:0000082), keratinization (GO:0031424) and cellular protein metabolic process (GO:0044267) and urushiol downregulated proteins in nervous system development (GO:0007399), positive regulation of cell proliferation (GO:0008284) and cellular protein metabolic process (GO:0044267) significantly.

overall_design: Normal Human Epidermal Keratinocytes (NHKCs) were treated with 10 microM of SLS or 1 microM of urushiol for 24 hours and harvested for total RNA extraction and hybridization on Affymetrix Human GeneChip U133 2.0 microarrays.


title: Single cell sequencing of mouse muscles

status: Public on Oct 31 2019

city: Dallas

first: Adwait Amod

last: Sathe

state: Texas

address: 5323 Harry Hines Blvd.

department: McDermott Center of Human Growth and Development

institute: UT Southwestern Medical Center

summary:
Purpose: The goals of this study are to compare the Quadriceps and diaphragm muscles
Methods: Library prepared followed by 10X Genomics standard protocol. Transcriptome were generated by high throughput sequencing

overall_design: Single cell RNA sequencing was conducted on samples of cells isolated from limb muscle (Quadriceps) and respiratory muscle (Diaphragm) in mouse.

first: Lan

last: Zhou

first: Xingyu

last: Wang

first: Gregory

last: Smith

first: Adwait


title: A Spatiomolecular Map of the Striatum

city: Stockholm

first: Konstantinos

last: Meletis

address: Solnavagen 9

department: Neuroscience

country: Sweden

institute: Karolinska Institutet

summary: The striatum is organized into two major outputs formed by striatal projection neuron (SPN) subtypes with distinct molecular identities. In addition, the histochemical division into patch and matrix compartments represents an additional spatial organization, proposed to mirror a functional specialization in a motor-motivation dimension. To map the molecular diversity of SPNs in the context of the patch and matrix division, we genetically labeled mu-opioid receptor (Oprm1) expressing striatal neurons and performed single-nucleus RNA sequencing (snRNA-seq). This allowed us to establish new molecular definitions of the patch-matrix compartments, resulting in a molecular code for mapping patch SPNs at the cellular level. In addition, Oprm1 expression labeled exopatch SPNs, which we found to be molecularly distinct from both patch as well as neighboring matrix SPNs, thereby forming a separate molecular entity. At the cell-type level, we found an unexpected SPN diversity, leading to the identification of a new Col11a1+ striatonigral SPN type. At the tissue level, we found that mapping the spatial expression of a number of markers revealed new definitions of spatial domains in the striatum, which were conserved in the non-human primate brain. Interestingly, the spatial markers were cell-type independent and instead represented a spatial code that was found across all SPNs within a spatially restricted domain. This spatiomolecular map establishes a formal system for targeting and studying the striatal subregions and SPNs subtypes, beyond the classical striatonigral and striatopallidal division.

overall_design: Single-nucleus RNA sequencing of mouse striatal neurons using two different genotypes to include patch neurons (Oprm1-Cre genotype) and GABAergic neurons (Slc32a1-Cre genotype)

first: Antje

last: Märtin

first: Daniela

last: Calvigioni

first: Ourania

last: Tzortzi

first: Janos

last: Fuzik

first: Emil

last: Wärnberg


title: Comparative genome-scale analysis of Pichia pastoris variants   informs selection of an optimal base strain [non-expressing]

first: Charles

middle: Arthur

last: Whittaker

institute: Koch Institute

summary: Komagataella phaffii, also known as Pichia pastoris, is a common host for the production of biologics and enzymes, due to fast growth, high productivity, and advancements in host engineering. Several K. phaffii variants are commonly used as interchangeable base strains, which confounds efforts to improve this host. In this study, genomic and transcriptomic analyses of Y-11430 (CBS7435), X-33, GS115, and eight other variants enabled a comparative assessment of the relative fitness of these hosts for recombinant protein expression. Cell wall integrity explained the majority of the variation among strains, impacting transformation efficiency, growth, methanol metabolism, and secretion of heterologous proteins. Y-11430 exhibited the highest activity of genes involved in methanol utilization, up to two-fold higher transcription of heterologous genes, and robust growth. With a more permeable cell wall, X-33 displayed a six-fold higher transformation efficiency and up to 1.2-fold higher titers than Y-11430. X-33 also shared nearly all mutations, and a defective variant of HIS4, with GS115, precluding robust growth. Transferring two beneficial mutations identified in X-33 into Y-11430 resulted in an optimized base strain that provided up to four-fold higher transformation efficiency and three-fold higher protein titers, while retaining robust growth. The approach employed here to assess unique banked variants in a species and then transfer key beneficial variants into a base strain should also facilitate rational assessment of a broad set of other recombinant hosts.

overall_design: Characterization of 11 different Komagataella phaffii strains under 3 growth conditions with replicate total mRNA samples collected for each strain and condition

first: Joseph

last: Brady

first: M

last: Tan

first: Duan

last: Ma

first: L

last: Kristensen

first: Neil

last: Dalvie

first: Kerry

last: Love

first: J


title: ERα over-expression does not accelerate development of p53-deficient mammary tumors in mice [ChIP-Seq]

status: Public on Nov 13 2019

city: Amsterdam

first: Lisette

last: Cornelissen

address: Plesmanlaan 121

country: Netherlands

institute: The Netherlands Cancer Institute

summary: About 75% of all breast cancers express the nuclear hormone receptor oestrogen receptor α (ERα). However, the majority of mammary tumors from genetically engineered mouse models are ERα-negative. To model ERα-positive breast cancer in mice, we exogenously introduced expression of mouse and human ERα in an existing p53-deficiency driven breast cancer mouse model. After initial ERα expression during development of the mammary gland, expression was reduced or lost in adult glands and p53-deficient mammary tumors. ChIP-sequencing analysis of primary mouse mammary epithelial cells (MMECs) derived from these models, in which expression of the ERα constructs was induced in vitro, confirmed interaction of ERα with the DNA. In human breast and endometrial cancer, the pioneer factor FOXA1 is known to be essential to facilitate ERα/DNA binding. Surprisingly, the ERα binding sites identified in primary MMECs, but also in mouse mammary gland and uterus, showed a high enrichment of ERE motifs, but were devoid of Forkhead motifs. Furthermore, exogenous introduction of FOXA1 and GATA3 in ERα-expressing MMECs was not sufficient to promote ERα-responsiveness of these cells. Together, this suggests that species-specific differences in ERα-cistromes between mouse and human are dictated by the DNA sequence, resulting in ERα-dependencies in mice that are not FOXA1 driven and potentially not tumorigenic. These species-specific differences in ERα-biology can limit the use of mouse models in ERα-positive breast cancer research.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Binding of ERa in MCF7 cells and HA-tagged human ERa or HA-tagged mouse ERa in primary MMECs profiled by ChIP-seq (Chromatin Immunoprecipitation followed by deep sequencing).

middle: M

first: Linda

last: Henneman

first: Anne

last: Paulien Drenth

first: Eva

last: Schut

first: Roebi

last: de Bruijn

first: Wilbert

last: Zwart

first: Jos

last: Jonkers


title: gabrg2-KO whole zebrafish larval brain differential expression analysis

city: MONTREAL

first: ERIC

last: SAMARUT

state: QC

address: 900 St DENIS

department: NEUROSCIENCES

institute: CRCHUM

summary: We conducted a whole transcripome sequencing analysis (RNAseq) from whole larval brains extracted from 5 dpf gabrg2+/+ and gabrg2-/- larvae

overall_design: Triplicates of gabrg2+/+ (WT) and gabrg2-/- (HM) samples that consist of a pool of 5 extracted whole brains from 5 dpf larvae.

first: Éric

last: Samarut


title: RNA-seq of lipid droplet-low and lipid droplet-rich microglia isolated from the hippocampi of aged mice

status: Public on Oct 30 2019

city: Stanford

first: Julia

last: Marschallinger

address: 1201 Welch Road

department: Neurology

institute: Stanford University

summary: Sequencing data related to the manuscript entitled, "Lipid droplet accumulating microglia represent a dysfunctional and pro-inflammatory state in the aging brain".

overall_design: 18-month old mice were perfused and hippocampi were dissected (Hippocampi of 3 mice were pooled per sample). Primary microglia were isolated by gentle dounce homogenization of the brain, Percoll-based myelin removal, and FACS-purification of ~10,000 live CD11b+CD45lo cells. BODIPYlo (=lipid droplet-low) and BODIPYhi (=lipid droplet-high) microglia were sorted into RLT Plus buffer (Qiagen) containing beta-mercaptoethanol. RNA was extracted using a RNeasy Micro Plus kit (Qiagen) according to the manufacturer’s protocol. RNA integrity was assessed on a Bioanalyzer (Agilent), and high quality samples (RIN >9) were used for library preparation. cDNA synthesis and amplification was performed using the SmartSeq v4 Ultra-low input kit (Takara), and libraries were tagmented, adaptor ligated, and indexed using the Nextera XT kit (Illumina). Sequencing of isolated microglia was carried out with Illumina HiSeq 2000/2500, paired end, 2x 100 bp depth sequencer. The quality of fastq files was assessed using FASTQC (v 0.11.4). Reads were mapped to mouse mm9 reference genome using STAR (v 2.5.1b). Raw read counts were generated with STAR using the GeneCounts function. Differential expression analysis was performed using DESeq2 with standard settings.

first: Tal

last: Iram

first: Macy

last: Zardeneta


title: TFAP2C signalling in human fibroblasts

first: Erik

last: Sahai

address: 1 Midland Road

country: United Kingdom

institute: Francis Crick Institute

summary: Our work has shown that the transcription factor TFAP2C is upregulated in fibroblasts that generate an anisotrophic extracellular matrix. To determine what is acting downstream of TFAP2C we compared fibroblasts that had been mock transfected, or treated with a non-targeting siRNA control or siRNA against TFAP2C.

overall_design: Vulval carcinoma associated fibroblasts (VCAF8) underwent a mock transfection (Ctrl_Tfn) or were treated with a non-targeting control siRNA (Ctrl_AllS), or siRNA against TFAP2C (siTFAP2C1) in biological replicates.

first: Danielle

last: Park

first: Stefan

last: Boeing


title: Control of RNA pol II speed by PNUTS-PP1 and Spt5 dephosphorylation facilitates termination by a “sitting duck torpedo” mechanism

city: Aurora

first: David

last: Bentley

state: Colorado

department: BMG

institute: UCD AMC

summary: Control of transcription speed, which influences many co-transcriptional processes, is poorly understood. We report that PNUTS-PP1 phosphatase is a negative regulator of RNA pol II elongation rate. The PNUTS W401A mutation, which disrupts PP1 binding, causes genome-wide acceleration of transcription associated with hyper-phosphorylation of the Spt5 elongation factor. Immediately downstream of poly(A) sites, pol II decelerates from >2kb/min to <1 kb/min, which correlates with Spt5 dephosphorylation. Pol II deceleration and Spt5 dephosphorylation require poly(A) site recognition and the PNUTS-PP1 complex, which is in turn necessary for transcription termination. These results lead to a new model for termination, the “sitting duck torpedo” mechanism, where poly(A) site-dependent deceleration caused by PNUTS-PP1 and Spt5 dephosphorylation is required to convert pol II into a viable target for the Xrn2 terminator exonuclease. Spt5 and its bacterial homologue NusG therefore have related functions controlling kinetic competition between RNA polymerases and the termination factors that pursue them.

overall_design: Transcription elongation rates were determined within gene bodies and dwonstream of termination sites by mapping pol II positions after release of a DRB block or after inhibiting initiation with triptolide. Elongation rates were estimated under conditions where termination was inhibited by a dominant negative mutants of the exonuclease Xrn2 or the PP! nuclear targeting subunit, PNUTS. In addition Spt5 phosphorylation was measured by ChIP and found to correlate closely with chages in elongation rate.

first: Benjamin

last: Erickson


title: Silencing p300 in MCF7 cells to study expression and alternative splicing

city: Jerusalem

first: Maayan

last: Salton

address: Ein-Kerem Canpus

country: Israel

institute: Hebrew University

summary: Splicing of precursor mRNA (pre-mRNA) is an important regulatory step in gene expression. Recent evidence points to a regulatory role of chromatin-related proteins in alternative splicing (AS) regulation. Using an unbiased approach, we have identified the acetyltransferase p300 as a key chromatin-related regulator of AS. P300 promotes genome-wide exon inclusion in both a transcription-dependent and ‑independent manner. Using CD44 as a paradigm, we found that p300 regulates AS by modulating the binding of splicing factors to pre-mRNA. Employing a tethering strategy, we found that binding of p300 to the CD44 promoter region promotes CD44v exon inclusion independently of RNAPII transcriptional elongation rate. Promoter-bound p300 regulates AS by acetylating splicing factors, leading to exclusion of hnRNP M from CD44 pre-mRNA and activation of Sam68. P300-mediated CD44 AS reduces cell motility and promotes epithelial features. Our findings reveal a mechanism through which chromatin-related proteins regulate AS and show the impact of this mechanism on cell function.

overall_design: MCF7 cells were transfected with non-targeting siRNA (siNT) and sip300 for 72 h, each in triplicates. RNA was extracted, libraries prepared and RNA-seq was conducted.


title: Condensin II is recruited to promoters by pRb and regulates expression of divergently paired genes [RNA-seq]

first: Fred

last: Dick

address: Cancer Research Labs 790 Commissioners Road East

institute: Western University

summary: In this study, use of a gene-targeted mouse model that is defective for pRb-condensin II interactions, Rb1L, has revealed instances where condensin II localization in interphase nuclei is dependent on pRb. Condensin II binding overlaps significantly with marks of active chromatin, and is enriched at the promoters of genes, including closely spaced divergent promoters. Interestingly, there are some bidirectional promoters where condensin II binds in an pRb-dependent manner and transcription of only one of the two genes is upregulated in Rb1L/L MEFs, suggesting the pRb-condensin II complex may be acting as a repressor or an insulator at distinct genomic locations to influence transcription. Chromatin conformations at these locations demonstrated that the pRb-condensin II complex may be responsible for maintaining more static, long-range interactions. In addition, pRb also recruits TFIIIC, another condensin II interactor, to many genomic loci. These recently discovered non-canonical transcriptional functions of the pRb-condensin II complex may represent an additional mechanism of pRb-mediated tumor suppression.

overall_design: Total RNA from proliferating MEFs was isolated. Three Rb1+/+ and Rb1L/L MEF pairs were used for this analysis and expression between genotypes was compared.

first: Frederick

first: Aren

middle: E

last: Marshall


title: Analysis of gene expression in CNOT2-depleted A549 lung cancer cells

first: Miyong

last: Yun

address: 209, Neungdong-ro, Gwangjin-gu

department: Bioindustry and bioresource engineering

institute: Sejong University

summary: CNOT2 depletion partially decreased number of colony in colony forming assay and cell number in proliferation assay for NSCLC cells. Co-treatment with TRAIL and depletion of CNOT2 dramatically declined survival rate and increased apoptosis compared with TRAIL only in TRAIL resistant NSCLC cells. So, to find major signaling molecules in CNOT2 depleted condition of A549 cells, gene expression analysis was designed.

overall_design: NSCLC cells were plated at 60% confluency in 60mm dish 24 hours before transfection. shRNAs were designed against specific target sequence of human CNOT2 mRNAs.

first: Sung-Hoon

last: Kim


title: Transcriptome profiling reveals transcriptional and alternative splicing regulation in the early embryonic development of hair follicles in the cashmere goat

city: Wuhan

first: Dong

last: Chen

state: Hubei

address: 388 GaoXin 2nd Road, East Lake Hi-Tech Development Zone

department: Center for Genome Analysis

institute: ABLife, Inc.

type: Non-coding RNA profiling by high throughput sequencing

summary: The undercoat fiber of the cashmere goat, from the secondary hair follicle (HF), possesses commercial value. However, very few studies have focused on the molecular details of primary and secondary HF initiation and development in goat embryos. In this study, skin samples at embryonic day 45, 55, and 65 (E45, E55, and E65) were collected and prepared for RNA sequencing (RNA-seq). We found that the HF probably initiated from E55 to E65 by analyzing the functional pathways of differentially expressed genes (DEGs). Most key genes in canonical signaling pathways, including WNT, TGF-β, FGF, Hedgehog, NOTCH, and other factors showed clear expression changes from E55 to E65. We, for the first time, explored alternative splicing (AS) alterations, which showed distinct patterns among these three stages. Functional pathways of AS-regulated genes showed connections to HF development. By comparing the published RNA-seq samples from the E60, E120, and newborn (NB) stages, we found the majority of WNT/β-catenin signaling genes were important in the initiation of HF development, while other factors including FOXN1, GATA3, and DLX3 may have a consistent influence on HF development. Our investigation supported the time points of embryonic HF initiation and identified genes that have potential functions of embryonic HF initiation and development. We further explored the potential regulatory roles of AS in HF initiation, which extended our knowledge about the molecular mechanisms of HF development.

overall_design: Nine samples for RNA sequencing: three skin samples from cashmere goat embryo at embryonic day 45, three skin samples from cashmere goat embryo at embryonic day 55, and three skin samples from cashmere goat embryo at embryonic day 65. Three samples for small RNA sequencing: three skin samples from cashmere goat embryo at embryonic day 45 were mixed together as one sample, three skin samples from cashmere goat embryo at embryonic day 55 were mixed together as one sample, and three skin samples from cashmere goat embryo at embryonic day 65 were mixed together as one sample.

first: Yanjun

last: Zhang

first: Lele

last: Wang

first: Zhen

last: Li

first: Wenjing

last: Han

first: Rui

last: Su

first: Ruijun

first: Zhixin

first: Yi

first: Jinquan


title: ARID1A and the BAF complex are determinants of breast cancer treatment response [ATAC-Seq]

first: Jason

last: Carroll

address: Li Ka Shing Centre, Robinson Way

institute: Cancer Research UK, Cambridge Institute

summary: Global CRISPR screens provide an unparalleled, longer-term experimental approach for the identification of essential genes in drug resistance. We used an ~18,000 gene deletion screen to discover ARID1A and other BAF complex components as the most critical factors required for response to two classes of Estrogen Receptor (ER) antagonists, namely ER degraders and Selective Estrogen Receptor Modulators (SERMs). Unexpectedly, ARID1A was also the top candidate for response to the BET inhibitor JQ1, but in the opposite direction, where loss of ARID1A sensitised breast cancer cells to BET inhibition. We show that ARIDA binds chromatin at ER cis-regulatory elements and can physically associate with ER in model systems and primary tumour samples. ARID1A binding to ER enhancer elements, can occur in the absence of ER, suggesting that its repressive activity occurs in an enhancer-specific, but ER-independent manner. Specific targeting of ARID1A validated the CRISPR screen and shows that depletion of BAF activity, does not result in redundancy from P-BAF, the other ATP-dependent chromatin remodelling complex, but instead results in loss of HDAC1 binding, increased Histone 4 lysine acetylation and subsequent BRD4-driven growth. ARID1A and the BAF complex therefore function as a critical mechanism of antiestrogen activity and mutation or depletion in BAF activity drives a BRD4-mediated proliferative program that is refractory to ER targeted agents. Since ARID1A is mutated in a subset of treatment-resistant disease, these findings provide mechanistic insight and treatment strategies for patients, based on BAF complex fidelity status.

overall_design: ATAC-sequencing on MCF7 wild type and ARID1A knockout clones Clone 11 and 14 with Vehicle and Tamoxifen treatment

first: Sankari

last: Nagarajan

middle: S

first: Igor

last: Chernukhin


title: ARID1A and the BAF complex are determinants of breast cancer treatment response [ChIP-seq]

overall_design:
Examination of ARID1A, BRG1 and SNF5 binding sites in asynchronous MCF7 cells treated with vehicle (DMSO) or 100 nM 4-Hydroxytamoxifen, 250 nM JQ1 and 10 nM Fulvestrant for 6 hours or double times charcoal stripped serum (DCDT)- treated cells for 72 hours and treated with Vehicle or 10 nM b-estradiol for 6 hours. Examination of ER, H3K27ac, ARID2, BRG1, HDAC1 and BRD4 in MCF7 parental cell lines and ARID1A knockout clones 11 and 14 treated with vehicle (ethanol) or 100 nM 4-Hydroxytamoxifen.
Please note that the processed data was generated from merged replicates and is linked to the corresponding rep 1 sample records.

Examination of ARID1A and BRG1  binding in hormone-deprived synchronous MCF7  and ZR-75-1 cells with FOXA1 siRNA. Examination of histone H4 acetylation marks on MCF7 ARID1A knockout cells treated with vehicle (DMSO) or 100 nM 4-Hydroxytamoxifen.


title: ARID1A and the BAF complex are determinants of breast cancer treatment response [RNA-seq]

overall_design: RNAseq in the MCF7 cells with ARID1A Wild type and knockout clones with treatment of Fulvestrant, 4-HydroxyTamoxifen and JQ1 for 6 hours

first: Nagarajan

last: Sankari

first: Chernukhin

last: Igor


title: Analysis of transcriptomes in intestinal stem cells in response to Interferon gamma

first: Bic

last: MSKCC

address: 1275 York Ave.

institute: Memorial SLoan-Kettering Cancer Center

summary: There is limited understanding of how immune-mediated damage impacts intestinal stem cells (ISCs). We found that stem cell compartment injury is a shared feature of both alloreactive and autoreactive intestinal immunopathology, reducing ISCs and impairing their recovery in T-cell-mediated injury models. Modeling with ex vivo epithelial cultures indicated ISC depletion and impaired human as well as murine organoid survival upon co-culture with activated T cells, and screening of effector pathways identified Interferon-γ (IFNγ) as a principal mediator of ISC compartment damage. Flow cytometry analysis confirmed expression of IFNγ receptor on ISCs. To investigate if the receptor was functional, we performed transcriptome analysis of ISCs treated with IFNγ. In brief, mRNA profiles of sorted Lgr5-high cells treated with/without IFNγ were generated by deep sequencing, in triplicate, using the TruSeq SBS Kit V4 (Illumina). RNA profiles confirmed a robust IFNγ transcriptional signature in the stem cells, as several IFNγ-related genes were upregulated shortly after exposure, indicating direct activity of IFNγ in ISCs. Epithelial cultures with IFNγ-receptor-deficient ISCs and purified stem cell colonies also indicated direct targeting of the ISCs. This study showed that dysregulated T cell activation and Interferon-γ production are potent mediators of ISC injury.

overall_design: mRNA profiles of isolated Lgr5-high intestinal stem cells treated w/wo interferon gamma were generated by deep sequencing, in triplicate, using the TruSeq SBS Kit V4(Illumina).

first: Alan

last: Hanash

first: Shuichiro

last: Takashima


title: Overexpression of the adenylate cyclase gene cyaC facilitates current generation by Shewanella oneidensis in bioelectrochemical systems

city: Hachioji

first: Atsushi

last: Kouzuma

state: Tokyo

address: 1432-1 Horinouchi

institute: Tokyo University of Pharmacy and Life Sciences

summary: Electrochemically active bacteria (EAB) are capable of electrochemical interactions with electrodes via extracellular electron transfer (EET) pathways and serve as essential components in bioelectrochemical systems. Previous studies have suggested that EAB, such as Shewanella oneidensis MR-1, use cyclic AMP (cAMP) receptor proteins for coordinately regulating the expression of catabolic and EET-related genes, allowing us to hypothesize that the intracellular cAMP concentration is an important factor determining electrochemical activities of EAB. The present study constructed an MR-1 mutant, cyaC-OE that overexpressed cyaC, a gene encoding a membrane-bound class III adenylate cyclase, and examined its electrochemical and transcriptomic characteristics. We show that intracellular cAMP concentration in cyaC-OE is more than double that in wild-type MR-1, and cya-OE generates approximately two-fold higher current in BES than the wild type. In addition, the expression of genes involved in EET and anaerobic carbon catabolism is up-regulated in cya-OE as compared to that in the wild type. These results suggest that enhancement of the intracellular cAMP level is a promising approach for constructing an EAB with high catabolic and electrochemical activities.

overall_design: Total 8 samples, 4 biological replicates for each sample.

first: Takuya

last: Kasai

first: Kazuya

last: Watanabe


title: DUX4-induced transcripts from pericentric human satellite II repeats form double-stranded RNA [stranded-RNAseq]

city: Seattle

first: Stephen

last: Tapscott

state: WASHINGTON

address: 1100 Fairview N. Ave

department: Human Biology

institute: Fred Hutch Cancer Research Center

summary: The DUX4 transcription factor is normally expressed in the cleavage stage embryo and regulates genes involved in zygotic genome activation. Mis-expression of DUX4 in skeletal muscle, however, is toxic and causes facioscapulohumeral muscular dystrophy (FSHD). We recently showed DUX4-induced toxicity is due, in part, to the accumulation of double-stranded RNAs (dsRNAs) with concomitant activation of the PKR viral response pathway. Here, using dsRNA immunoprecipitation coupled with next-generation sequencing, we determined that DUX4-induced dsRNAs originate from intergenic regions enriched for Alu and LINE-1 elements, endogenous retroviruses, and pericentric human satellite II (HSATII) repeats. DUX4-induced HSATII dsRNAs are formed via temporally controlled bidirectional expression with predominant transcription of one strand preceding the other. Whereas DUX4 activation of dsRNAs derived from intergenic regions such as HSATII contributes to toxicity in FSHD, in the early embryo these dsRNAs might facilitate heterochromatin formation, as has been proposed for cleavage stage expression of mouse major satellites.

overall_design: MB135-DUX4i myoblast cells were induced for approximately 18 hours in duplicate. Stranded RNA-seq libraries were made from TRIzol extracted RNA.

first: Sean

last: Shadle

first: Chao-Jen

last: Wong

last: Bennett

first: Nancy

last: Karreman

first: Amy

last: Campbell

first: Silvère

last: van der Maarel

first: Brenda

last: Bass


title: DUX4-induced transcripts from pericentric human satellite II repeats form double-stranded RNA

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

overall_design: Refer to individual Series


title: Chromatin dynamics reveal circadian control of human in vitro islet maturation

first: Juan

last: Alvarez-Dominguez

institute: Harvard University

type: Methylation profiling by high throughput sequencing


title: Identification of circulating microRNA signatures as potential biomarkers in the serum of elk infected with chronic wasting disease

status: Public on Oct 29 2019

city: WInnipeg

first: Sarah

last: Medina

state: Manitoba

address: 1015 Arlington Street

department: One Health

institute: Public Health Agency of Canada

summary:
Purpose: Idenfication of microRNA biomarkers of Chronic wasting disease in serum from infected elk
Methods: Illumina next generation was used to profile abundance of serum miRNA in elk naturally infected with chronic wasting disease and Hamsters experimentally infected with the 263K scrapie prion strains
Results: A signature of 21 miRNAs with diagnostic potential was found to be altered in abundance in serum from CWD infected elk. Of these, 6 were similarily altered in the serum from the 263K infected hamsters
Conclusion: These altered miRNA signatures may serve as the basis for non-invasive diagnostic assays for chronic wasting disease and may shed light on the pathogenesis of prion infection

sample_organism:
Mesocricetus auratus
Cervus canadensis

platform_organism:
Mesocricetus auratus
Cervus canadensis

overall_design: Serum miRNA was profiled in prion infected elk and hamsters using illumina next generation sequencing

first: Jessy

last: Slota

first: Megan

last: Klassen

first: Damian

last: Gorski

first: Christine

last: Mesa

first: Catherine

last: Robertson

first: Michael

middle: B

last: Coulthart

first: Sandra

last: Pritzkow


title: Expression data from Arabidopsis thaliana Col-0 transformed with constructs misexpressing NAC052 transcription factor involving leaf morphology and anatomy

city: Duesseldorf

first: Roxanne

last: van Rooijen

address: Universitaetsstrasse 1

institute: Heinrich Heine University

summary: In an effort to identify genetic regulators for the cell ontogeny around the veins in Arabidopsis thaliana leaves, an activation-tagged mutant line with altered leaf morphology and altered bundle sheath anatomy was characterized. This mutant had a bundle sheath-specific promoter from the gene GLYCINE DECARBOXYLASE SUBUNIT-T from the C4 species Flaveria trinervia (GLDTFt promoter) inserted in the coding region of the transcriptional repressor NAC052. Reconstruction of the mutation event of our activation-tagged line by creating a line expressing an amino-terminally truncated sequence of NAC052 under control of the GLDTFt promoter as well as creating a line expressing the full sequence of NAC052 under control of the GLDTFT promoter, as well as creating a CRISPR/Cas mutant line of NAC052, confirmed the involvement of NAC052 in leaf development.

overall_design: Complete Arabidopsis rosettes of 5 lines (reference line, activation tagged line, GLDTFT::5'truncatedNAC052 line, GLDTFT::NAC052 line, and CRISPR/Cas nac052) were harvested on day 28 after sowing.

first: Stefanie

last: Schulze

first: Patrick

last: Petzsch

first: Peter

last: Westhoff


title: Mouse spleen expression profile after systemic Candida glabrata infection

city: Vienna

last: Tscherner

address: Dr. Bohr Gasse 9/2

department: Max F. Perutz Laboratories, Department for Medical Biochemistry

country: Austria

institute: Medical University of Vienna

summary: Pleiotropic type I interferons (IFNs-I) fulfil multiple protective functions during infectious diseases, but can also exert detrimental effects for the host leading to immunopathology. Here, we report that IFNs-I promote the dysregulation of Fe homeostasis in macrophage populations and in the spleen during systemic infections with the intracellular pathogen Candida glabrata, leading to fungal survival and persistence. By using microarray expression profiling with RNA isolated from mouse spleens upon systemic Candida glabrata infection and subsequent in vitro interaction experiments we show that IFN-I signaling induces global transcriptional downregulation of key players in Fe homeostasis and profoundly alters Fe transport mechanisms within macrophages.

overall_design: Wild-type (C57BL/6J) and Ifnar1-/- mice (8-12 weeks old) were infected intravenously (i.v.) with 5 x 10^7 colony-forming units (CFUs) of C. glabrata (ATCC2001) (in 100 μl) per 21.5 g mouse weight. On day 1, 3, 7 and 14 post-infection, spleens were collected and stored in RNAlaterTM Stabilization Solution (Thermo Fisher Scientific). Three biological replicates were done for each time point and mouse strain, respectively. Four biological replicates of PBS infected mice for each mouse strain were used as control samples and spleens were harvested 14 days post PBS injection. Organs were homogenized in 1.5 ml RNA lysis buffer (Promega) with occasional cooling on ice by using an Ika T10 basic Ultra-Turrax homogenizer (IKA). For RNA isolation, the SV Total RNA Isolation System (Promega) was used according to the manufacturer’s instructions. RNA quality was checked on RNA 6000 Nano chips (Agilent) using a Bioanalyzer 2100 (Agilent).  The Low Input Quick Amp Labeling Kit (one-color) (Agilent) was used to generate fluorescent cRNA. The amplified cyanine 3-labeled cRNA samples were then purified using SV Total RNA Isolation System (Promega) and hybridized to the SurePrint G3 Mouse GE 8x60K microarray (Agilent). Microarray slides were washed and scanned with a DNA Microarray Scanner (Agilent), according to the standard protocol of the manufacturer. Information from probe features was extracted from microarray scan images using the Feature Extraction software v10.7.3 (Agilent).

last: Riedelberger

first: Philipp

last: Penninger

first: Sabrina

last: Jenull

first: Andriy

last: Petryshyn

first: Christelle

last: Bourgouis

first: Walter

last: Glaser

first: Karl

last: Kuchler


title: ARID1A and the BAF complex are determinants of breast cancer treatment response

city: Cambridge

first: Jason

last: Carroll


title: Epstein-Barr Virus (EBV) Latent Membrane Protein 1 (LMP1) Regulates Host B Cell miR-155 and its Target FOXO3a via PI3K p110a Activation

status: Public on Oct 28 2019

city: Colorado Springs

first: Olivia

last: Hatton

state: CO

address: 14 E Cache La Poudre St

department: Molecular Biology

institute: Colorado College

summary: The purpose of this study was to investigate the host cell miRs regulated by LMP1 from the B95-8 strain of EBV. In this study, we utilized an established chimeric, inducible NGFRLMP1 siganling molecule stably expressed in EBV- BL41 Burkitt’s Lymphoma cell lines (BL41 NGFRLMP1 cells). A qPCR array was used to measure the expression level of 377 previously identified miRs from RNA isolated from two sources: BL41 NGFRLMP1 cells in which LMP1 signaling was not activated (n=2, control) and BL41 NGFRLMP1 cells in which LMP1 signaling was activated for 12 hours (n=2).

overall_design: miRNA expression was assayed in EBV negative BL41 Burkitt's lymphoma cells stably expressing chimeric, inducible NGFRLMP1 molecules where LMP1 signaling was not activated (n=2) and where LMP1 signaling was activated for 12 hours (n=2). The expression of miRNA in the samples without LMP1 activation was used as a control.

first: Aleishia

last: Harris-Arnold

first: Sheri

last: Krams

last: Martinez


title: DUX4-induced transcripts from pericentric human satellite II repeats form double-stranded RNA [dsRIPseq]

first: Stephen

last: Tapscott

overall_design: Double-stranded RNA immunoprecipitations were conducted on MB135-DUX4i cells using K1 or J2 mouse monoclonal antibodies with three replicates per each -/+doxycycline condition which induces DUX4 expression. Mouse nonspecific IgG was used as comparator for analysis.

first: Sean

last: Shadle

first: Chao-Jen

last: Wong

last: Bennett

first: Nancy

last: Karreman

first: Amy

last: Campbell

first: Silvère

last: van der Maarel

first: Brenda

last: Bass

middle: J


title: RNA Seq Analysis of HeLa cell lines Transcriptomes at different condition

status: Public on Oct 24 2019

city: Beijing

first: Miao

last: Liu

address: Xitoutiao No.10, Youanmen Waidajie

institute: Capital Medical University

summary: The goal of this study are to compare the transcriptome profiling (RNA-seq) of human cervical cells (HeLa) transcriptome profiling (RNA-seq) under the condition of inhibition of respiratory chain by antimycin A or hypoxia.

overall_design: Examination of mRNA profiling of HeLa cells under the normal condition or treatment with antimycin A or hypoxia.

first: Binghui

last: Li


title: The hippo pathway effector protein YAP modulated resistance to trametinib neuroblastomas with hyperactivated RAS pathway signalling

status: Public on Oct 26 2019

city: Los Angeles

first: Miller

last: Huang

state: CA

address: 4650 Sunset Blvd, #57

department: Pediatrics

institute: Children's Hospital Los Angeles

summary: We sequenced seven neuroblastoma cell lines, including the parental NLF neuroblastoma cell line and three modified NLF cell lines.

overall_design: The parental NLF line (N=3) was used as a comparison to the NLF sgCon cell line. NLF sgCon was transduced with a scrambled sgRNA-targeted CRISPR-Cas9 lentivirus, and NLF_YAPKO1 and NLF_YAPKO2 represent two NLF cell lines with YAP1 genetic knockout. NLF sgCon, NLF YAPKO1, and NLF YAPKO2 were treated with either 0.1% DMSO  (N=3) or 20 nM trametinib (N=3) for 72 hours prior to RNA isolation. NLF sgCon serves as a control for the NLF YAP1 knockout cell lines.

first: Grace

last: Coggins

first: Alvin

last: Farrel

first: Komal

last: Rathi

first: JoLynne

last: Rokita

first: John

last: Maris


title: Gene expression analysis of a panel of human fibroblasts that generate an aligned (anisotropic) vs non-aligned extracellular matrix

city: London

first: Erik

last: Sahai

institute: Francis Crick Institute

summary: A panel of human fibroblasts derived from patient samples (vulval, oral, cervical and breast carcinoma and metastatic melanoma) were seperated according to their ability to generate an aligned (anisotropic) or non-aligned extracellular matrix.

overall_design: Human fibroblasts were isolated from a patient tissues of vulval (VCAF8, VCAF2B) cervical (CerCAF), oral (OCAF), breast (1901T) carcinoma and metastatic melanoma (MAF2) and immortalised with lentiviral HTERT as described in Gaggioli, C., et al. Fibroblast derived matrix assays were performed as described in Franco-Barraza, J., et al and RNA extracted after 6 days in culture.

first: Danielle

last: Park

first: Stefan

last: Boeing


title: ARID1A and the BAF complex are determinants of breast cancer treatment response [CRISPR gRNA-seq]

first: Jason

last: Carroll

overall_design: CRISPR gRNA sequencing of Cas9 expressing MCF7 cells infected with pooled gRNA libraries with different days of infection and with drug treatments (JQ1, 4-hydroxytamoxifen and Fulvestrant).

first: Sankari

last: Nagarajan

first: Igor

last: Chernukhin


title: Sphingosine 1-phosphate-regulated transcriptomes in heterogenous arterial and lymphatic endothelium of the aorta

status: Public on Oct 18 2019

city: Boston

first: Eric

last: Engelbrecht

address: 1 Blackfan Street

department: Vascular Biology Program

institute: Boston Children's Hospital

summary: Despite the medical importance of G protein-coupled receptors (GPCRs), in vivo cellular heterogeneity of GPCR signaling and downstream transcriptional responses are not understood. While endothelial S1pr1 expression is ubiquitous (Kaur et al., 2017), receptor activation in vivo, as reported by GFP in S1PR1-GS mice, revealed heterogeneous signaling in multiple organs (Kono et al., 2014), including in the aorta (Galvani et al., 2015).    To gain insights into the molecular mechanisms of S1PR1 regulation of endothelial transcription and the heterogenous nature of S1PR1 signaling in vivo, we performed transcriptome (bulk and single-cell RNA-seq) and open chromatin profiling (ATAC-seq) of FACS-sorted adult mouse aortic endothelial cells (MAECs) with high (GFPhigh) or low (GFPlow) reporter expression. We also performed bulk transcriptome and open chromatin profiling of MAECs from tamoxifen-treated Cdh5-CreERT2 S1pr1f/f (S1pr1-ECKO) and S1pr1f/f (S1pr1-WT) littermates. MAECs were defined as CD45-Ter119-DAPI-CD31+. The CD31 antibody used was PE-conjugated rat anti-mouse clone MEC13.3 from BD Biosciences.

overall_design: Cells from 2-4 aortae of age and sex-matched adult mice were pooled for each individual experiment (ATAC-seq, RNA-seq and scRNA-seq). N=3 for each MAEC population for all bulk comparisons.

first: Michel

middle: V

last: Levesque

first: Liqun

last: He

first: Michael

last: Vanlandewijck

first: Christer

last: Betsholtz

first: Timothy

last: Hla


title: Single Cell RNA sequencing of mouse CD11b+, Gr1+ cells from the spleen from FVB/n and PYMT mice

status: Public on Oct 21 2019

city: Irvine

first: Kai

last: Kessenbrock

address: 839 Health Sciences Road

department: Biological Chemistry

institute: UC Irvine

summary: Myeloid-derived suppressor cells (MDSCs) are innate immune cells that acquire the capacity to suppress adaptive immune responses during cancer. It remains elusive how MDSCs differ from their normal myeloid counterparts, which limits our ability to specifically detect and therapeutically target MDSCs during cancer. Here, we used single-cell RNAseq to compare MDSC-containing splenic myeloid cells from breast tumor-bearing mice to wildtype controls. Our computational analysis of 14,646 single-cell transcriptomes reveals that MDSCs emerge through a previously unrealized aberrant neutrophil maturation trajectory in the spleen giving rise to a unique chemokine-responsive, immunosuppressive cell state that strongly differs from normal myeloid cells. We establish the first MDSC-specific gene signature and identify novel surface markers for improved detection and enrichment of MDSCs in murine and human samples. Our study provides a single-cell transcriptional map defining the development of MDSCs, which will ultimately enable us to specifically target these cells in cancer patients.

overall_design: Droplet Based Single Cell RNA sequencing libraries were generated for 5 pooled FVB/n mouse spleens and 3 pooled PYMT mouse spleens  using the 10x Genomics Chromium Platform and sequenced on the Illumina HighSeq 4000

first: Hamad

last: Alshetaiwi

first: Nicholas

last: Pervolarakis


title: Transcriptome analysis of mouse tumors driven by PIK3CA mutation variants

status: Public on Oct 09 2019

city: Houston

first: Chad

last: Creighton

state: TX

address: One Baylor Plaza, Mail Stop: BCM305

department: Biostatistics, Ducan Cancer Center

institute: Baylor College of Medicine

summary: The RTK/RAS/PI3K pathway is a key driver of tumorigenesis across all cancers, with 90% of Glioblastoma (GBM) tumors exhibiting alterations in this pathway. Among the specific genes in this pathway mutations in PI3K’s catalytic subunit, PIK3CA, are found in 11% of GBM tumors. Sequencing of GBM samples has revealed several known hotspot mutations that drive tumorigenesis in several cancers (E545K, H1047R), as well as a series of 63 in frame mutations (as indexed in COSMIC) that remain largely unclassified. To decode which of these PIK3CA variants function as drivers of GBM, we established an in vivo complementation screening platform for GBM. Our mouse GBM model relies on in utero electroporation (IUE) and CRISPR-Cas9 mediated knockout of NF1, p53, and PTEN (termed 3xCR). Because PTEN catalyzes the reverse reaction of PIK3CA, phosphorylation of PIP2 to PIP3, we complemented loss of PTEN with overexpression of E545K or H1047R, two bona fide hot spot driver mutations of PIK3CA. Both these variants accelerated tumor associated death, demonstrating that known PIK3CA drivers can complement PTEN loss in our system.

overall_design: We examined the global transcriptional effects of PIK3CA variants in our model system via RNA-Sequencing on end-stage tumors driven by the variants.

first: Fengju

last: Chen

first: Benjamin

last: Deneen

first: Yiqun

last: Zhang


title: Genome-wide mapping of oxidative DNA damage via engineering of 8-oxoguanine DNA glycosylase

first: Yuxin

last: Fang

state: Beijing

address: Yiheyuan Road No.5

institute: Peking University

summary: 7,8-dihydro-8-oxoguanine (OG) is one of the major DNA oxidative damages, which has been concerned by researchers for its potential of mutagenesis and epigenetic regulation. Genome-wide mapping of oxidative DNA damage is of great value for understanding the biological impact of these modifications. Here we present a novel appoach to profile guanine oxidation in the mouse genome. Through structure-guided mutagenesis, we've successufully engineered 8-oxoguanine DNA glycosylase into a useful tool for OG detection. By enrichment of glycosylase-DNA complex, thousands of OG peaks were identified in the mouse genome.

overall_design: Enrichment procedure was perform on mouse embryonic fibroblasts (MEFs) with two biological replicates. Genomic samples without any treatment were denoted as input; samples enriched by engineered glycosylase were denoted as enrich; samples pretreated with Fpg to remove oxidative damages before enrichment were denoted as control.

first: Peng

last: Zou


title: The miR-199-CDK2 axis modulates neutrophil migration and systemic inflammation

status: Public on Oct 08 2019

city: West Lafayette

first: Qing

last: Deng

state: Indiana

address: 915 W. State Street

department: Department of Biological Sciences

institute: Purdue University

summary: We performed RNAseq to identify miRNAs that were specifically down regulated in neutrophils, and RNAseq for identification of mRNA targets of  miRNA-199 in neutrophils

overall_design: for miRNA profiles, we compared the miRNA present in zebrafish neutrophils and keratinocytes to the whole embryo. for miR-199 target identification we sequenced messenger RNAs in zebrafish neutrophils over expressing miR199 or vector control

first: Alan

middle: Y

last: Hsu

first: Sheng

last: Liu

first: Jun

last: Wan