title: Stellate cells, hepatocytes and endothelial cells imprint the Kupffer cell identity on monocytes colonizing the liver macrophage niche (RNA-Seq)

status: Public on Aug 14 2019

city: Zwijnaarde

first: Liesbet

last: Martens

address: Technologiepark 927

department: VIB Inflammation Research Center

country: Belgium

institute: VIB-University of Ghent

summary: Macrophages are strongly adapted to their tissue of residence. Yet, we know little about the cell-cell interactions that imprint the tissue-specific identities of macrophages in their respective niches. Using conditional depletion of liver Kupffer cells, we traced the developmental stages of monocytes differentiating into Kupffer cells and mapped the cellular interactions imprinting the Kupffer cell identity. Kupffer cell loss induced the tumor necrosis factor (TNF) and interleukin-1 (IL-1) receptor-dependent activation of stellate cells and endothelial cells, resulting in the transient production of chemokines and adhesion molecules orchestrating monocyte engraftment. Engrafted circulating monocytes transmigrated into the perisinusoidal space, and acquired the liver-associated transcription factors ID3 and LXRα. Coordinated interactions with hepatocytes induced ID3 expression, while endothelial cells and stellate cells induced LXRα via a synergistic NOTCH-BMP pathway. This study shows that the Kupffer cell niche is composed of stellate cells, hepatocytes and endothelial cells that together imprint the liver-specific macrophage identity.

overall_design:
Microarray = different time points during development of Kupffer cells during embryogenesis and after depletion. This data was also compared to other tissue resident macrophages from the Immgen Consortium Database (GSE15907 and GSE37448).
Bulk RNA Seq = Hepatic Stellate Cells (HSCs), Liver Sinusoidal Endothelial Cells (LSECs) and hepatocytes. For each cell type there are 4 replicates from Pbs mice, 12h after DT-injection mice and 36h after DT-injection mice.

first: Johnny

last: Bonnardel

first: Wouter

last: T’Jonck

first: Djoere

last: Gaublomme

first: Robin

last: Browaeys

first: Charlotte

last: Scott

first: Bavo

last: Vanneste

first: Sofie

last: De Prijck

first: Sergei

last: Nedospasov

first: Anna

last: Kremer

first: Evelien

last: Van Hamme

first: Peter

last: Borghgraef

first: Wendy

last: Toussaint

first: Pieter

last: De Bleser

first: Inge

last: Mannaerts

first: Alain

last: Beschin

first: Leo

last: van Grunsven

first: Bart

middle: N

last: Lambrecht

first: Tom

last: Taghon

first: Saskia

last: Lippens

first: Dirk

last: Elewaut

first: Yvan

last: Saeys

first: Martin

last: Guilliams


title: Stellate cells, hepatocytes and endothelial cells imprint the Kupffer cell identity on monocytes colonizing the liver macrophage niche

first: Liesbet

last: Martens

type: Expression profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: The conserved 3’ UTR-derived small RNA NarS counteracts the risk of toxic nitric oxide accumulation during nitrate respiration

status: Public on Aug 13 2019

city: Shanghai

first: Chuan

last: Wang

address: 131 # Dong An Road

country: China

institute: Fudan University

summary: In pursuit of a biological role of Salmonella 3' UTR derived sRNA NarS, we sought to determine potential target mRNAs of NarS under anaerobic conditions. To this end, we compared gene expression in NarS-deficient and NarS overexpression (from plasmid pPL-NarS) strains following a 30-minute anaerobic shock by RNA-seq.

sample_organism: Salmonella enterica subsp. enterica serovar Typhimurium str. SL1344

overall_design: Total RNA of Salmonella NarS-deficient strain (ΔnarK ΔNarS) carrying either empty plasmid pJV300 or NarS-overexpressed plasmid pPL-NarS were compared.


title: RNA sequencing human monocytes

city: Leuven

first: Paul

last: HOLVOET

address: Herestraat 49

department: Cardiovascular Sciences

institute: KULeuven

summary:
Aim:
To improve risk stratification in patients with stable coronary artery disease (CAD), we aimed to identify genes in monocytes predictive of new ischemic events in patients with CAD and determine to what extent expression of these transcripts resembles expression in acute myocardial infarction (AMI).
Results:
COX10 and ZNF484 distinguished between AMI and the whole group of stable CAD patients with an accuracy of 90%. COX10 and ZNF484 together with MT-COI and WNK1 distinguished AMI patients from stable CAD patients with and without a new event with a sensitivity of 89% and a specificity of 98%. MT-COI and COX10 increased the accuracy for separating stable CAD patients with and without a new coronary event from 68 to 80% in addition to age, gender, BMI, diabetes, lipids, blood pressure and hs-CRP. Interestingly, expression of MT-COI, COX10 and WNK1 (but not ZNF484) in PBMCs paired with that in monocytes; COX10 in whole blood was similar to that in monocytes.
Conclusions:
This work showed that COX10 and ZNF484, eventually combined with MT-COI and WNK1 have the potential to accurately discriminate between AMI and stable CAD patients, and may improve the risk assessment of stable CAD patients.

overall_design: We included 19 AMI and 66 CAD patients which were stable at time of blood sampling; 24 of the latter developed a new coronary event: sudden death, new AMI, stroke or urgent coronary revascularization. Circulating CD14+ monocytes were isolated with magnetic beads. Exploratory RNA sequence analysis (36 individual samples; 11 in group 1 i.e CAD pateints without new event; 13 patients in group 2 i.e. CAD patients with new event; 12 patients in group 3 i.e. AMI patients. Expressions were validated with qPCR in monocytes (85 patients).


title: Transcriptional profiling of microglia; current state of the art and future perspectives

status: Public on Aug 10 2019

city: Groningen

first: Emma

last: Gerrits

email: e.gerrits@rug.nl, e.gerrits@umcg.nl

state: Groningen

address: Antonius Deusinglaan 1

department: BSCS

country: Netherlands

institute: UMCG

summary: Microglia are the tissue macrophages of the central nervous system (CNS) and the first to respond to CNS dysfunction and disease. Gene expression profiling of microglia during development, under homeostatic conditions and in the diseased CNS provided insight in microglia functions and changes thereof. Single cell sequencing studies further contributed to our understanding of microglia heterogeneity in relation to age, sex and CNS disease. Recently, single nucleus gene expression profiling was performed on (frozen) CNS tissue. Transcriptomic profiling of CNS tissues by (single) nucleus RNA-sequencing has the advantage that it can be applied to archived and well stratified frozen specimens. Here, we give an overview of the significant advances recently made in microglia transcriptional profiling. In addition, we present matched cellular and nuclear microglia RNA-seq datasets we generated from mouse and human CNS tissue to compare cellular versus nuclear transcriptomes from fresh and frozen samples. We demonstrate that microglia can be similarly profiled with cell and nucleus profiling, and importantly also with nuclei isolated from frozen tissue. Nuclear microglia transcriptomes are a reliable proxy for cellular transcriptomes. Interestingly, lipopolysaccharide- (LPS)-induced changes in gene expression were even more pronounced in the nuclear transcriptome. In addition, heterogeneity in microglia observed in fresh samples is similarly detected in frozen nuclei of the same donor. Together, these results show that microglia nuclear RNAs obtained from frozen CNS tissue are a reliable proxy for microglia gene expression and cellular heterogeneity and may prove an effective strategy to study of the role of microglia in neuropathology.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: 12 mouse samples for bulk RNAseq, 4 mouse samples from 3 replicates per group for single-cell RNA seq and 6 samples from 2 human donors for single-cell RNA seq

first: Yang

last: Heng

first: Erik

last: Boddeke

last: Eggen


title: Comprehensive epigenome characterization reveals the diverse transcriptional regulation across human vascular endothelial cells (ChIP-seq)

status: Public on Aug 08 2019

city: Bunkyo-ku

first: Ryuichiro

last: Nakato

state: Tokyo

address: 1-1-1 Yayoi

department: Institute for Quantitative Biosciences

country: Japan

institute: The University of Tokyo

type: Genome binding/occupancy profiling by high throughput sequencing

summary:
Endothelial cells (ECs) make up the innermost layer throughout the entire vasculature. Their phenotypes and physiological functions are initially regulated by developmental signals and extracellular stimuli. The underlying molecular mechanisms responsible for the diverse phenotypes of ECs from different organs are not well understood.
To characterize the transcriptomic and epigenomic landscape in the vascular system, we cataloged gene expression and active histone marks in nine types of human ECs (generating 148 genome-wide datasets) and carried out a comprehensive analysis with chromatin interaction data. We developed a robust procedure for comparative epigenome analysis that circumvents variations at the level of the individual and technical noise derived from sample preparation under various conditions. Through this approach, we identified 3765 EC-specific enhancers, some of which were associated with disease-associated genetic variations. We also identified various candidate marker genes for each EC type. We found that the nine EC types can be divided into two subgroups, corresponding to those with upper-body origins and lower-body origins, based on their epigenomic landscape. Epigenomic variations were highly correlated with gene expression patterns, but also provided unique information. Most of the deferentially expressed genes and enhancers were cooperatively enriched in more than one EC type, suggesting that the distinct combinations of multiple genes play key roles in the diverse phenotypes across EC types. This comprehensive analysis of epigenome characterization of EC types reveals diverse transcriptional regulation across human vascular systems. These datasets provide a valuable resource for understanding the vascular system and associated diseases.

overall_design: ChIP-Seq experiments were conducted in human vascular endothelial cells. Fastq files of all samples except for IMR90 have been deposited in the SRA database under accession numbers SRP192924 (BioProject accession: PRJNA532996).

first: Youichiro

last: Wada

first: Toutai

last: Mitsuyama

first: Hiroyuki

last: Aburatani

first: Hiroshi

last: Kimura

first: Katsuhiko

last: Shirahige


title: Identification of proteins and miRNAs that specifically bind an mRNA in vivo

city: Berlin

first: Kathrin

last: Theil

address: Hannoversche Str. 28

country: Germany

institute: BIMSB/MDC

type: Non-coding RNA profiling by high throughput sequencing


title: Identification of proteins and miRNAs that specifically bind an mRNA in vivo [small RNA]

summary: Understanding regulation of an mRNA requires knowledge of its regulators. However, methods for reliable de-novo identification of proteins binding to a particular RNA are scarce and were so far only successfully applied to abundant noncoding RNAs in cell culture. Here, we present vIPR, an RNA-protein crosslink, RNA pulldown, and shotgun proteomics approach to identify proteins bound to a selected mRNA in C. elegans. Applying vIPR to the germline-specific transcript gld-1 led to enrichment of known and novel interactors. By comparing enrichment upon gld-1 and lin-41 pulldown, we demonstrate that vIPR recovers both common and specific RNA-binding proteins, and we validate DAZ-1 as a novel and specific gld-1 regulator. Finally, combining vIPR with small RNA sequencing, we recover known and biologically important transcript-specific miRNA interactions, and we identify miR-84 as a specific interactor of the gld-1 transcript. We envision that vIPR provides a platform for investigating RNA in vivo regulation in diverse biological systems.

overall_design: small RNA sequencing of 16 vIPR RNA pulldown samples (C. elegans), with 3 replicates of the gfp control pulldown, 2 replicates each of pulldowns of the transcripts gld-1::gfp, gld-1, alg-1, lin-41, and 1 replicate of gfp::lin-41

first: Koshi

last: Imami

first: Nikolaus

last: Rajewsky


title: Genome-wide Assessment of Antimicrobial Tolerance in Yersinia pseudotuberculosis Under Ciprofloxacin Stress

status: Public on Aug 02 2019

city: London

first: Sam

last: Willcocks

address: Keppel St

country: United Kingdom

institute: LSHTM

summary: Yersinia pseudotuberculosis is a Gram-negative bacterium capable of causing gastrointestinal infection and is closely related to the highly virulent plague bacillus Yersinia pestis. Infection by both species are currently treatable with antibiotics such as ciprofloxacin, a quinolone-class drug of major clinical importance in the treatment of many other infections. Our current understanding of the mechanism of action of ciprofloxacin is that it inhibits DNA replication by targeting DNA gyrase, and that resistance is primarily due to mutation at this target site, along with generic efflux and detoxification strategies. We utilised transposon directed insertion site sequencing (TraDIS or TnSeq) to identify the non-essential chromosomal genes in Y. pseudotuberculosis that are required to tolerate sub-lethal concentrations of ciprofloxacin in vitro. As well as highlighting recognised antibiotic resistance genes, we provide evidence that a multitude of genes involved in regulating DNA replication and repair are central in enabling Y. pseudotuberculosis to tolerate the antibiotic, including dksA (yptb0734), a regulator of RNA polymerase and hda (yptb2792), an inhibitor of DNA replication initiation. We furthermore demonstrate that even at sub-lethal concentrations, ciprofloxacin causes severe cell-wall stress, requiring lipopolysaccharide lipid A, O-antigen and core biosynthesis genes to resist the sub-lethal effects of the antibiotic. It is evident that coping with the consequence(s) of antibiotic-induced stress requires the contribution of scores of genes that are not exclusively engaged in drug-resistance.

overall_design: TnSeq was used to identify the non-essential chromosomal genes in Y. pseudotuberculosis that are required to tolerate sub-lethal concentrations of ciprofloxacin in vitro.

first: S

middle: J

first: K

middle: K

last: Huse

first: R

last: Stabler

first: P

middle: C

last: Oyston

first: A

last: Scott

first: H

middle: S

last: Atkins

first: B

middle: W

last: Wren


title: Identification of proteins and miRNAs that specifically bind an mRNA in vivo [total RNA]

overall_design: RNA sequencing of 4 vIPR pulldown input samples, 7 vIPR pulldown elution samples, and 1 total RNA sample from wild-type and transgenic C. elegans strains


title: Auxin response in tomato

status: Public on Aug 01 2019

city: Rehovot

first: Idan

last: Efroni

address: POB 12

department: Plant Sciences

country: Israel

institute: Hebrew University

summary: The dataset described the short term transcriptional response to auxin in M82 tomatoes

overall_design: Root and shoot samples were treated with mock or auxin. Root experiments were performed in duplicates. Shoot experiments were performed in triplicates.

first: Alon

last: Israeli

first: Chen

last: Yahav


title: Tumor Heterogeneity in Metastatic Potential is Indicated by Adhesion Strength

city: La Jolla

first: Adam

middle: Jeffrey

last: Engler

state: CA

address: 9500 Gilman Drive, MC 0412

department: Bioengineering

country: USA

institute: UC San Diego

summary: Tumors are heterogeneous and comprised of cells with varying function and ability to disseminate. Despite significant effort, no universal biological marker currently serves as a metric for metastatic potential of solid tumors. Common to disseminating cells from such tumors, however, is the need to modulate their adhesion as they detach from the tumor and migrate through stroma to intravasate. Adhesion strength is heterogeneous even amongst cancer cells within a given population, and using a parallel plate flow chamber, we separated and sorted these populations into weakly and strongly adherent groups; when cultured under stromal conditions, this adhesion phenotype is stable over multiple days, sorting cycles, and common across all epithelial tumor lines investigated. Subpopulations do not show differences in expression of proteins involved in the focal adhesion complex but do exhibit intrinsic focal adhesion assembly as well as contractile differences that result from differential expression of genes involved in microtubules, cytoskeleton linkages, and motor activity. Collectively, these differences drive migration in both 2D and 3D migration assays for weakly adherent cells; these differences are maintained over days in culture, suggesting that adhesion strength can serve as a stable marker for migration and metastatic potential within a given tumor population. Moreover, these data suggest that the fraction of weakly adherent cells present within a tumor could act as a novel physical marker for metastatic potential.

overall_design: RNA-sequencing profiles of weakly adherent (n = 3) and strongly adherent (n = 2) MDA-MB231 cells isolated via application of shear stress to heterogeneous populations.

last: Beri


title: RNA-seq analyses of two intestinal dendritic cell subsets, CD103+CD11b- and CD103-CD11b+, from wild type and CbfbF/F:CD11c-Cre mice.

city: Tsurumi-ku, Yokohama

first: Hideyuki

last: Yoshida

state: Kanagawa

address: 1-7-22 Suehiro-cho

institute: RIKEN IMS

summary: We report phenotypes in gut dendritic cell development by loss of Runx/Cbfb transcription factor complexes. To further examine function of residual CD103+CD11b- and CD103-CD11b+ DCs, we performed RNA-seq analyses and compared gene expression signatures between control and Cbfb-deficient cells.

overall_design: Examination of transcriptome of CD103+CD11b- and CD103-CD11b+ intestinal DCs.

first: Mari

last: Tenno

first: Alicia

middle: Y

last: Wong

first: Mika

last: Ikegaya

first: Bradley

last: Smith

first: Ichiro

last: Taniuchi


title: Pro-survival Lipid Sphingosine-1-Phosphate Metabolically Reprograms T cell to Limit its Anti-Tumor Activity

status: Public on Jul 31 2019

city: Maywood

first: Michael

last: Zilliox

state: IL

address: 2160 S. First Ave

department: Ophthalmology

institute: Loyola University Chicago

summary: Sphingosine 1-phosphate (S1P), a pro-tumor survival bioactive lysophospholipid, generated by sphingosine kinase 1 (SphK1), regulates lymphocyte egress into circulation via transducing S1P receptor 1 (S1PR1) signalling, and also plays a role in the differentiation of regulatory T cells and T helper-17 cells. However, the mechanisms by which receptor-independent SphK1/S1P signallng regulates anti-tumor T cell activity remain unknown. Using melanoma antigen gp100 reactive T cells deficient in Sphk1, we found that Pmel-Sphk1-/- T cells maintained central memory phenotype, showed higher mitochondrial respiration, and exhibited resistance to TGF-β mediated suppression. Mechanistically, we discovered a direct correlation between SphK1 generated intracellular S1P and PPARγ (peroxisome proliferator-activated receptor gamma) activity, which in turn regulate lipolysis in T cells. Genetic, molecular or pharmacologic inhibition of SphK1 or PPARγ improved metabolic fitness and anti-tumor activity of T cells. Further, inhibition of SphK1 and PD1 together led to long-term control of melanoma. Overall, these data highlight the clinical potential of targeting SphK1/S1P for improved anti-cancer adoptive T cell immunotherapy.

overall_design: Duplicate samples were analyzed from the WT and Sphingosine Kinase deficient mouse. The splenic T cells were activated wih cognate antigen (gp100) for three days before the RNA was prepared and RNA seq. performed.

first: Shikhar

last: Mehrotra


title: A tumor specific super-enhancer drives immune evasion by guiding synchronous expression of PD-L1 and PD-L2

status: Public on Jul 30 2019

city: Nantong

first: Yingcheng

last: Wu

state: Jiangsu

address: No. 19 Qixiu Road, Chongchuan District, Nantong, Jiangsu 226001, China

department: Basic Medical Research Center

institute: Nantong University

overall_design: RNA-seq of SgVec and PD-L1L2-SE deficient SUM-159 cells.

first: Yihui

last: Fan


title: Genome-wide identification of transcripts associated with KIAA1429 by RIP-seq

status: Public on Jul 27 2019

city: Chengdu

first: Tian

last: Lan

state: Sichuan

address: No. 37 of Guoxue Lane

department: Department of Liver Surgery & Liver Transplantation

institute: West China Hospital

summary: N6-methyladenosine (m6A) modification, as the most abundant internal methylation of eukaryotic RNA transcripts, is critically implicated in RNA processing, decay, transport and translation. Here we showed that KIAA1429, the largest known component in the m6A methyltransferase complex, was considerably upregulated in hepatocellular carcinoma (HCC) tissues. High expression of KIAA1429 was significantly associated with the malignant clinical features and the poor prognosis of HCC patients. Silencing KIAA1429 suppressed the cell proliferation and metastasis in vitro and in vivo. Integrated MeRIP-seq, RIP-seq and RNA-seq data identified GATA3 as the direct downstream target of KIAA1429-mediated m6A modification. KIAA1429 knockdown markedly impaired the m6A levels of GATA3 mRNA and increased the expression of GATA3. Mechanistically, KIAA1429 induced the m6A methylation on 3’ UTR of GATA3 pre-mRNA, leading to the separation of RNA-binding protein HuR and the degradation of GATA3 pre-mRNA, which was followed by the downregulation of GATA3. Strikingly, a long noncoding RNA (lncRNA) GATA3-AS, transcribed from the antisense strand of GATA3 gene, functioned as a cis-acting element for the preferential interaction of KIAA1429 with GATA3 pre-mRNA. Accordingly, we found that the tumor growth and metastasis driven by KIAA1429 or GATA3-AS were mediated by GATA3. Patients with low KIAA1429 and high GATA3 expressions showed greatly better poor overall survival and disease-free survival. In conclusion, our study proposed a complex KIAA1429-GATA3 regulatory model based on m6A modification and provided insights into the epi-transcriptomic dysregulation in hepatocarcinogenesis and metastasis.

overall_design: To identify mRNA species bound by KIAA1429, we performed the RIP-seq to immunoprecipitate endogenous mRNAs associated with the KIAA1429 and sequenced the retrieved RNA.

first: T

last: Li

first: D

last: Zhang

last: Liu


title: Next Generation Sequencing Facilitates Quantitative Analysis of Streptococcus agalactiae and Nitrite Exposure Leukocyte Transcriptomes

city: guangzhou

first: zhao

last: zaoya

address: Waihuan East Road

institute: SUN YAT-SEN University

summary:
Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare NGS-derived head kidney leukocytes transcriptome profiling (RNA-seq) to microarray and quantitative reverse transcription polymerase chain reaction (qRT–PCR) methods and to evaluate protocols for optimal high-throughput data analysis
Methods: The leukocytes of tilapia were treated with PBS(A), heat-killed S.agalactiae(B), sodium nitrite(C) or heat-killed S.agalactiae and sodium nitrite co-treatment(D),respectively. Then the mRNA profiles were generated by deep sequencing, in triplicate, using Illumina GAIIx. The sequence reads that passed quality filters were analyzed at the transcript isoform level. qRT–PCR validation was performed using SYBR Green assays
Results: Using an optimized data analysis workflow, we mapped about 40 million sequence reads per sample to the tilapia genome in the head kidney leukocytes of tilapia.10 candidate genes were randomly selected to quantify their mRNA expression levels of treatment and control groups by q-PCR. Results showed that the Pearson’s correlation coefficient between q-PCR and RNA-seq was 0.939. Total of 6173 transcripts were differently expressed, with a fold change ≥2 and p value <0.05.
Conclusions: Our study represents the first detailed analysis of tilapia head kidney leukocytes transcriptomes, with biologic replicates, generated by RNA-seq technology. The optimized data analysis workflows reported here should provide a framework for comparative investigations of expression profiles. Our results show that NGS offers a comprehensive and more accurate quantitative and qualitative evaluation of mRNA content within a fish cell. We conclude that RNA-seq based transcriptome characterization would expedite genetic network analyses and permit the dissection of complex biologic functions.

overall_design: The mRNA profiles of leukocytes treated by A) PBS, B) Streptococcus agalactiae, C) nitrite or D) Streptococcus agalactiae and nitrite co-treatment were generated by deep sequencing, in triplicate, using Illumina HiSeq 2000.

first: Zhao

last: Zaoya

first: Li

last: Wensheng

first: Sun

last: Caiyun


title: Profiling of N6-Adenosine (m6A) methylation in HCCLM3 cells with shCtrl or shKIAA1429

status: Public on Jul 23 2019

overall_design: To map the altered m6A modification caused by KIAA1429, MeRIP-seq was performed in HCCLM3 cells with shCtrl or shKIAA1429.

first: H


title: Promoter activity profiling reveals on- and off-target transcriptional responses to drug treatment

status: Public on Jul 25 2019

city: Yokohama

first: Erik

last: Arner

address: Suehiro-cho 1-7-22

institute: RIKEN

summary: Owing to safety concerns or insufficient potential for efficacy, only 0.01% to 0.02% of new drug candidates are approved for marketing. Drugs already on the market may be withdrawn or restricted to certain uses due to adverse effects (AEs) discovered after market introduction. Comprehensively investigating the on-/off-target effects of drugs can help expose AEs during the drug development process. In this study, we developed an integrative framework for systematic identification of on-/off-target pathways and elucidation of the underlying mechanisms, by combining expression profiling after drug treatment with gene perturbation of the primary drug target. Expression profiles from statin-treated cells and HMG-CoA reductase knockdowns were analyzed using the framework, allowing for identification of not only reported adverse effects but also novel candidates of off-target effects from statin treatment. Our findings may provide new insights for finding new usages or potential side effects of drug treatment.

overall_design: CAGE profiling in three different cell types after treatment with four different statins and two siRNAs targeting HMG-CoA reductase (HMGCR), including replicates and control samples.


title: Gene expression profiling for HCCLM3 cells with shCtrl or shKIAA1429

overall_design: To identify candidate downstream genes of KIAA1429, RNA-seq analysis was implemented in HCCLM3 cells with shCtrl or shKIAA1429.


title: Obesity synergizes with HBx, Src and p53 mutation accelerate hepatocarcinogenesis [Microarray]

status: Public on Jul 19 2019

city: Hsinchu

first: Yong Chun

last: Luo

address: Boai Street

country: Taiwan

institute: National Chiao Tung University

summary: Previous studies have identified liver cancer associated to NASH, diabetes, obesity, and some genetic risk factors. We want to investigate the synergism between diet and genetic risk factors in hepatocarcinogenesis using omics data of four background zebrafish in three types of diet. We found the fishes have more genetic risk factors at the same time, and have higher probability to accelerate cancer formation. Overfed and high fat diet will increase the chance. Moreover, the results showed metabolism and genetic information processing, including the pathways of fatty acid metabolism, steroid biosynthesis and ribosome biogenesis are highly affected in hepatocellular carcinoma.

overall_design: Four background zebrafish in three types of diet: Wild type, HBx(p53-), Src(p53-), and HBx,Src(p53-) zebrafish in normal, overfed, and high fat diet. RNA was prepared from liver and analyzed by microarray.

first: Wan

last: Yang

first: Pei

last: Rao

first: Yong

first: Hua

last: Lin

first: Sing

middle: H

last: Huang

first: Jinn

middle: M

first: Chiou

last: Yuh


title: Obesity synergizes with HBx, Src and p53 mutation accelerate hepatocarcinogenesis [RNA-seq]

overall_design: Four background zebrafish in three types of diet: Wild type, HBx(p53-), Src(p53-), and HBx,Src(p53-) zebrafish in normal, overfed, and high fat diet. RNA was prepared from liver and analyzed by RNAseq.


title: Obesity synergizes with HBx, Src and p53 mutation accelerate hepatocarcinogenesis

overall_design: Refer to individual Series


title: Ribosome profiling of GC7/vehicle-treated mouse primary B cells

status: Public on Jul 09 2019

city: Oxford

first: Jack

middle: Edward

last: Feltham

state: Oxfordshire

address: South Parks Road

department: Biochemistry

institute: University of Oxford

summary: Mouse primary B cells were treated with 10 uM GC7 or vehicle control and processed for ribosome profiling. During harvesting, samples were spiked with S. cerevisiae lysate containing stable polysomes.

overall_design: Examination of ribosome-protected fragment abundances and distributions in drug-treated and control cells. N = 1.

first: Hanlin

first: Katja

last: Simon

first: Jane

last: Mellor


title: FLASH: ultra-fast protocols to identify RNA-protein interactions in cells

city: Freiburg

first: Asifa

last: Akhtar

address: Stuebeweg 51

department: Chromatin Regulation

institute: Max Planck Institute of Immunobiology and Epigenetics

summary: Determination of the in vivo binding sites of RNA-binding proteins (RBPs) is paramount to understanding their function and how they affect different aspects of gene regulation. With hundreds of RNA-binding proteins identified in human cells, a flexible, high-resolution, high-throughput, highly multiplexible and radioactivity-free method to determine their binding site has not been described to date. Here we report FLASH (Fast Ligation of RNA after some sort of Affinity Purification for High-throughput Sequencing), which uses a special adapter design and an optimized protocol to determine protein-RNA interactions in living cells. We demonstrate the flexibility, speed and versatility of FLASH by using it to determine binding sites of both tagged and endogenously expressed proteins under diverse conditions. The entire FLASH protocol, starting from cells-on-plates to a sequencing library, takes 1.5 days.

overall_design: FLASH (Fast cloning of RNA After some Sort of affinity purification for High-throughput sequencing)

first: Ibrahim

last: Ilik

first: Daniel

last: Maticzka


title: A compendium of promoter-centered long-range chromatin interactions in diverse human tissues and cell types

city: Daejeon

first: Inkyung

last: Jung

address: KAIST, 291 Daehak-ro, Yuseong-gu

department: Biological Sciences

country: South Korea

institute: KAIST

summary: A large number of genetic variants associated with human diseases are found in non-coding DNA and may contribute to illness by affecting gene regulation, but mechanistic study of these variants has been hindered by a lack of information of their potential regulatory targets. In order to overcome this limitation, we generate maps of long-range chromatin interactions centered on 19,539 promoters in 27 human cell/tissue types, and use this information to infer putative target genes of candidate cis-regulatory sequences throughout the human genome. In additional to known enhancer-promoter interactions, we also confirm widespread promoter-promoter interactions and obtain evidence suggesting enhancer-like function for many promoter regions. These promoter-centered chromatin interaction maps corroborate target genes of genetic variants defined in previous genome-wide association studies, predict new target genes of many disease-associated genetic variants, and contribute novel insights into a broad spectrum of human traits and diseases.

overall_design:
The study contains promoter capture Hi-C results for diverse human cell/tissue types
.bz2, text files for promoter capture Hi-C chromatin interaction between DNA fragments for promoter-promoter interactions (PP) or promoter-other interactions (PO)
pp: P-P interaction frequency
po: P-O interaction frequency

first: Anthony

last: Schmitt

first: Andrew

last: Lee

first: Bing

last: Ren


title: MicroRNA profile of pediatric pilocytic astrocytomas identifies two tumor-specific signatures when compared to non-neoplastic white matter

city: Ribeirão Preto

first: Carlos

middle: Alberto

last: Scrideli

state: São Paulo

address: Avenida Bandeirantes 3900

department: Pediatrics

country: Brazil

institute: Ribeirao Preto Medicine School - University of Sao Paulo

summary: Pilocytic astrocytoma (PA) is a low-grade neoplasm frequently found in childhood. PA is characterized by slow growth and a relatively good prognosis. Genetic mechanisms such as activation of MAPK, BRAF gene deregulation and neurofibromatosis type 1 (NF1) syndrome have been associated with PA development. Epigenetic signature and miRNA expression profile are providing new insights about different types of tumor, including PAs. In the present study we evaluated global miRNA expression in 16 microdissected pediatric PA specimens, three NF1-associated PAs and 11 cerebral white matter (WM) samples by the microarray method. An additional cohort of 20 PAs was used to validate by qRT-PCR the expression of six miRNAs differentially expressed in the microarray data. Unsupervised hierarchical clustering analysis distinguished one cluster with nine PAs, including all NF1 cases and a second group consisting of the WM samples and seven PAs. Among 88 differentially expressed miRNAs between PAs and WM samples, the most underexpressed ones regulate classical pathways of tumorigenesis, while the most overexpressed miRNAs are related to pathways such as focal adhesion, P53 signaling pathway and gliomagenesis. The PAs/NF1 presented a subset of underexpressed miRNAs, which was also associated with known deregulated pathways in cancer such as cell cycle and hippo pathway. In summary, our data demonstrate that PA harbors at least two distinct miRNA signatures, including a subgroup of patients with NF1/PA lesions.

overall_design: In order to identify miRNA signatures associated with pilocytic astrocytoma, we evaluated the miRNA expression profile of a group of PAs (including a cohort of three cases of PAs occurring in NF-1 patients) compared to white matter (WM) samples by the miRNA microarray technique.

first: Luiz

middle: D

last: Júnior

first: Regia

middle: P

last: Lira

first: Paola

middle: F

last: Fedatto

first: David

last: Antonio

first: Elvis

middle: T

last: Valera

first: Simone

last: Aguiar

first: José

middle: A

last: Yunes

first: Silvia

middle: R

last: Brandalise

first: Luciana

last: Neder

first: Fabiano

last: Saggioro

first: Aline

last: Becker

first: Ricardo

last: Oliveira

first: Hélio

last: Machado

first: Rodrigo

last: Panepucci

middle: G

last: Tone


title: Real-time quantitative PCR analysis of human Vastus Lateralis muscle biopsies

status: Public on Jul 03 2019

city: Chieti

first: Rosa

last: Mancinelli

address: via dei Vestini

country: Italy

institute: university of Chieti

summary: Vastus Lateralis muscle biopsies were obtained before and after the training protocol from 6 healthy elderly male people. We chose custom TaqMan low density array cards (Life Technologies) preloaded with 48 selected genes related to the following pathways: myogenesis, apoptosis, protein anabolism/catabolism and antioxidant enzymes to quantitate their gene expression in human skeletal muscle.

overall_design: RT-PCR gene expression profiling. Vastus Lateralis muscle biopsies were obtained before and after the training protocol from 6 healthy elderly male people. From each biopsies, total RNA was purified and then cDNA was synthesized from it. Equal amount of cDNA from each donor was analysed for its gene expression. For each donor, post-training gene expression was compared to pre-training. In the end mean and standard error was calculated.


title: Transcriptome analysis of purified microglial cells from striatum and midbrain

status: Public on Jul 02 2019

city: Pamplona

first: Marisol

last: Aymerich

state: Navarra

address: CIMA Universidad de Navarra, Avda. Pío XII, 55

country: Spain

institute: University of Navarra

summary: Microglia constitutes a diverse population of cells that present a broad spectrum of responses when they become activated. Here, microglial status was studied under steady-state conditions from different brain regions involved in neurodegenerative diseases. Under basal conditions, midbrain microglia showed an immune-alert state not observed in striatum. Unique subpopulations of microglia expressing TLR4 and MHC-II with antigen presenting properties, and a higher proportion of infiltrating CD4+ T cells were identified in the midbrain.  These results highlight that the inflammatory tone is context-dependent and reveal the unique properties of the midbrain related to the interaction with the immune system.

overall_design: Analysis of two cohorts of control animals


title: Analysis of transcriptome, selected intracellular signaling pathways, proliferation and apoptosis of LNCaP cells exposed to high leptin concentration

city: Poznan

first: Marcin

last: Rucinski

address: Swiecicki 6

department: Histology and Embriology

country: Poland

institute: Poznan University of Medical Science

summary:
The androgen-sensitive LNCaP line of prostate cancer cells is one of the most commonly used cells in oncological research. These include, but are not limited to, research into the influence of leptin (LEP) on prostate cancer. However, the results of studies on the effects of this cytokine on the proliferation and apoptosis of LNCaP cells are controversial. Therefore, we performed studies on the effects of high LEP concentration (1x10-6 M) on gene expression profile, change of selected signaling pathways, proliferation and apoptosis of LNCaP cells. RTCA (real-time cell analyzer) revealed inhibitory effect of LEP on cell proliferation, but lower LEP concentrations (10-8 and 10-10 M) did not affect cell division. Within 24 h LEP (10-6 M) increases expression of 297 genes and decreases expression of 119 genes. Differentially expressed genes (DEGs) were subjected to functional annotation and clusterization using the DAVID bioinformatics tools
Most ontological groups are associated with proliferation and apoptosis (7 groups), immune response (6) and extracellular matrix (2). These results were confirmed by the Gene Set Enrichment Analysis (GSEA). The leptin's effect on apoptosis stimulation was also confirmed using Pathview library. These results were also confirmed by qPCR method. The results of Western Blot analysis (exposure to LEP 10 min, 1, 2, 4 and 24h) suggest (after 24 h) decrease of p38 MAPK, p44-42 mitogen-activated protein kinase and Bcl-2 phosphorylated at threonine 56. Moreover, exposure of LNCaP cells to LEP significantly stimulates the secretion of matrix metallopeptidase 7 (MMP7). Obtained results suggest activation of apoptotic processes in LNCaP cells cultured at high LEP concentration. At the same time, this activation is accompanied by inhibition of proliferation of the tested cells.

overall_design: LNCaP - the human prostate carcinoma cell line [LNCaP clone FGC (ATCC® CRL-1740DTM)] was purchased from ATCC (American Type Culture Collection, Manassas, Rockville, MD, USA). LNCaP cells were cultivated in the RPMI Medium 1640 (1X), supplemented with GlutaMAX, HEPES (all from Gibco, Life technologies, Carlsbad, California, USA) and antibiotics (Antibiotic/antimycotic Sigma Aldrich, Saint Louis, MO 63103, USA). Cells were grown in the 75 cm2 flask (NUNC EasyFlask with Nunclon surface, Thermo Fisher Scientific) at 37°C in a humidified atmosphere of 5% CO2. The culture medium was changed every 2 days. When cells reached approximately 80% confluence (about 7-8 days of cultivation), they were subcultured into 6-well plates (Nunc, Thermo Fisher Scientific, approximately 343 000 cells per one well – 9.6 cm2), to determine the effect of LEP on LNCaP transcriptome modulation. The applied experimental protocol was as follows: during the first 48 h of cultivation, the cells grew in a standard medium mentioned above. For the next 24 h, the cells were grown in starvation medium (FBS free). Afterwards, the cells were cultivated for 48 h, in a starvation medium supplemented with LEP (Recombinant Human Leptin, PeproTech, Germany) at the following concentrations: 0 (control), 1x10-6 M. After the mentioned period cell supernatants were collected and stored at - 80°C for further analyses.

first: Marta

last: Szyszka

first: Marianna

last: Tyczewska

first: Karol

last: Jopek

first: Ludwik

middle: K

last: Malendowicz


title: Alterations of miRNAs and mRNAs in MDBK cells expressing  BPV-13 E5

city: Haikou

first: Feng

last: Pang

address: Renmin Road 58

institute: Hainan University

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


title: Sphingobium sp. SYK-6 treated with several carbon sources versus SEMP

city: Vancouver

first: Hirofumi

last: Hara

state: BC

address: 2350 Health Science Mall

department: Microbilogy and Immunology

country: Canada

institute: University of British Columbia

summary: Transcriptional profile of RNA extracted from Sphingobium sp. SYK-6 treated with 10 mM pinoresinol (PR), 10 mM acetovanillone (AV), 10 mM phenylcoumarane (PC), 10 mM vanillate (VA), 10 mM vanillin (VN), 10 mM protocatechuate (PCA), 10 mM fellulate (FA),  10 mM 5,5'- dehydrodivanillate (DDVA), 10 mM 1,2‐bis(4‐hydroxy‐3‐methoxyphehyl)‐1,3‐propanediol (B1), 10 mM syringate (SA), 10 mM guaiacylglycerol‐β‐guaiacyl ether (GGE) comparing RNA extracted from SYK-6 treated with SEMP (10 mM sucrose, 10 mM glutamate, 0.34 mM methionine, and 10 mM proline).

overall_design: Dual channel experiment, RNA from cells treated with 11 substrates vs. RNA from cells treated with SEMP, independently grown and harvested.

first: Masaya

last: Fujita

first: Kosuke

last: Mori

first: Shoji

last: Hishiyama

first: Naofumi

last: Kamimura

first: Eiji

last: Masai


title: Alterations of miRNAs in MDBK cells expressing  BPV-13 E5

summary: To find out differentially expressed miRNAs in MDBK cells expressing BPV-13 E5

overall_design: A MDBK sample expressiong BPV-13 E5 and a negative control

first: Fengyang

last: Wang


title: Cap analysis gene expression (CAGE) of Aspergillus luchuensis mut. kawachii control and sirtuin D disruptant strains

status: Public on Jun 15 2019

city: Kagoshima

first: Taiki

last: Futagami

address: Korimoto 1-21-24

department: Department of Agriculture

institute: Kagoshima University

summary: Sirtuin is considered to play a significant role in the growth phase-dependent gene expression. In this study, we characterized sirtuin in the white koji fungus, Aspergillus kawachii, to examine their role on regulation of amylolytic enzymes and citric acid productions during the solid-state culture (koji). Characterization of rice-koji made using five putative sirtuin gene disruptants indicated that they are involved in amylolytic activity and acidity of rice koji; especially the sirD disruptant showed lower levels of α-amylase activity and citric acid production per mycelial weight in the koji compared the control strain. In addition, the sirD disruptant also showed a change in mycelial pigmentation, higher sensitivity to cell wall biogenesis inhibitor such as calcofluor white and Congo red, and reduced conidia formation, indicating that SirD is required for secondary metabolism, cell wall integrity, and conidial development. The cap analysis gene expression (CAGE) indicated that the transcriptional changes related to the characteristic phenotype of the sirD disruptant (e.g., a reduced transcript level of acid-stable α-amylase gene and a citric acid exporter) in rice koji. These results indicated that SirD has a significant role on the global transcriptional regulation including productions of α-amylase and citric acid in A. kawachii during the solid-state fermentation process.

overall_design: CAGE profiles of Aspergillus luchuensis mut. kawachii control and sirtuin D disruptant strains were investigated by Illumina NextSeq 500.

first: Kazuki


title: m6A in mRNA coding regions promotes translation via the RNA helicase-containing YTHDC2

status: Public on Jul 01 2019

city: Hangzhou

first: Yuanhui

last: Mao

state: Zhejiang

address: 1369 West Wenyi Road

institute: Zhejiang University

summary: Dynamic mRNA modification in the form of N6-methyladenosine (m6A) adds considerable richness and sophistication to gene regulation. The m6A mark is asymmetrically distributed along mature mRNAs, with a strong preference around the stop codon and 3’UTR. Nevertheless, approximately 35% of m6A residues are located within the coding region (CDS). It has been suggested that methylation in CDS slows down translation elongation by interfering the decoding process. However, neither the decoding feature of endogenous mRNAs nor the physiological significance of CDS m6A has been clearly defined. By integrating Ribo-seq and m6A-seq data sets, we found that CDS m6A methylation leads to ribosome pausing in a codon-specific manner. Unexpectedly, removing CDS m6A modification from these transcripts results in a further decrease of translation. A systemic analysis of RNA structural datasets revealed that CDS m6A methylation positively regulates translation by resolving mRNA secondary structures. We further demonstrate that the elongation-promoting effect of CDS methylation requires the RNA helicase-containing m6A reader YTHDC2. Our findings established the physiological significance of CDS methylation and uncovered non-overlapping function of m6A reader proteins.

overall_design: Using Ribo-seq to investigate the effect of m6A/m6A reader on translation efficiency

first: Shu-Bing

last: Qian


title: Transcriptome of FliC-stimulated mouse bladder and dependency on TLR5

status: Public on Jun 25 2019

city: Gold Coast

first: Glen

middle: C

last: Ulett

state: QLD

address: Parklands Drive

department: School of Medical Science

country: Australia

institute: Griffith University

summary: We report a transcriptional response in the mouse bladder following exposure to the major flagella filament protein, FliC. The response is shown to encompasses multiple innate immune networks not previously associated with FliC. Additionally, we provide analysis of components of this response that depend on the presence of the known flagellin receptor, TLR5.

overall_design: We used RNA-based sequencing technology for high-throughput profiling of innate immune responses in mouse bladder exposed to FliC, and the role of TLR5 in these responses


title: CSF-1 and IL-34 drive distinct microglial programs in the brain tissue

status: Public on Jun 27 2019

city: New York

first: Christie

last: Chang

state: New York

address: 1470 Madison Avenue, 5-301

institute: Icahn School of Medicine at Mount Sinai

summary: Bulk RNA sequencing of FACS sorted primary murine microglia from WT mixed glial cultures isolated from P2-3 forebrain and cerebellum, stimulated with either IL-34 or CSF-1

overall_design: Microglia isolated murine forebrain or cerebllar mixed glial cultures were treated with either IL-34 or CSF-1 for 24hrs before sequencing. N=3 replicates of microglia from independent mixed glial culutres, per brain region, per ligand treatment

first: Veronika

last: Kana

first: Fiona

last: Desland

first: Miriam

last: Merad


title: Regional transcriptional differences between human cerebellar and forebrain (superior temporal gyrus) microglia

summary: Bulk RNA sequencing of purified primary human microglia from superior temporal gyrus and cerebellum

overall_design: Primary microglia were obtained from the Netherlands Brain Bank and isolated from patient matched superior temporal gyrus and cerebellum

first: Lotje

last: de Witte


title: Alterations of mRNAs in MDBK cells expressing  BPV-13 E5

first: Feng

last: Pang

summary: To find out differentially expressed mRNAs in MDBK cells expressing BPV-13 E5


title: RNAseq of prostate cancer cells (PC3 and DU145)  treated with 2.5 uM pentamidine or vehicle

status: Public on Jun 14 2019

city: Shanghai

first: Lin

last: Liu

state: Shanghai

address: Shanghai

department: Renji‐Med X Clinical Stem Cell Research Center, Ren Ji Hospital

institute: Shanghai Jiao Tong University

summary: To uncover the mechanism underlying the effect of pentamidine on prostate cancer cells, we performed RNA sequencing (RNA-seq) to compare the transcriptional difference between pentamidine- and vehicle-treated prostate cancer cells (PC3 and DU145 cells).

overall_design: mRNA profiles of PC3 and DU145 cells treated with 2.5 uM pentamidine or vehicle were generated by deep sequencing, in triplicate, using Illumina Novaseq 6000 PE150.


title: Disruption of CSF-1R signaling affects cerebellar and forebrain homeostasis

summary: Ultra low input sequencing of FACS sorted primary murine microglia from CSF-1 or IL-34 deficient forebrain and cerebella, at P8 and 9 weeks

overall_design: Csf1fl/fl vs NesCreCsf1fl/fl: 3-4 biological replicates per timepoint per group; Il34wt/wt vs Il34Lacz/Lacz: 2-3 biological replicates per timepoint per group. P8, 9weeks


title: Temporal analysis of in vivo Toxoplasma gondii cysts shows novel and late stage specific transcripts.

status: Public on Jul 11 2019

city: Madison

first: Laura

last: Knoll

state: WI

address: 1550 Linden Drive

department: Medical Microbiology and Immunology

institute: UW-Madison

summary: The protozoan pathogen Toxoplasma gondii has the unique ability to develop a chronic infection in the brain of its host. The T. gondii develops cysts within the neurons that are resilient against the host immune response and current therapeutics. The bradyzoite parasites within the cyst have a sugar-protein-rich wall and a slow-replication cycle, allowing them to remain hidden from the host. This intracellular encysted lifestyle of T. gondii has made them recalcitrant to molecular analysis in vivo. Here, we have enriched for T. gondii brain cysts 21 to 150 days post-infection (DPI). RNA and protein from bradyzoites was isolated from each timepoint. Deep sequencing of transcripts expressed during these three timepoints showed many of the identified proteins are transcriptionally expressed at a high level. Approximately one-third are more enriched during bradyzoite conditions compared to tachyzoites, and approximately half are expressed at similar levels during each phase. We have expanded the transcriptional profile of in vivo bradyzoites to 120 DPI. The RNA sequencing depth of in vivo bradyzoite T. gondii was over 250-fold greater than was have been previously, which allowed us to identify low level transcripts and novel bradyzoite-specific isoforms.

overall_design: Illumina sequecing of bradyzoites isolated from mouse brain cysts, 28, 90, and 120 days postinfection in duplicate. Sequencing ran on two lanes and collated together.

first: Andrew

middle: L

last: Garfoot

middle: J


title: Bioavailability of clay-adsorbed dioxin to Sphingomonas wittichii RW1 and its associated genome-wide shifts in gene expression

status: Public on Jun 30 2019

city: East Lansing

first: Benli

last: Chai

state: Michigan

address: 567 Wilson Road Rm 2225 A

department: Center for Microbial Ecology

institute: Michigan State University

summary: Polychlorinated dibenzo-p-dioxins and dibenzofurans are a group of chemcially-related pollutants categorically known as dioxins. We used Sphingomonas wittichii strain RW1 (RW1), one of the few strains able to grow on dioxin, to characterize its ability to respond to and degrade clay-bound dioxin. Strain RW1 grew on and completely degraded dioxin intercalated in smectite clay. To characterize the effects of sorption and bioavailability of dioxin on RW1, transcriptomes of RW1 either grown with dioxin intercalated to clay (DDSAP) or with free crystalline dioxin (DD) were sequenced using RNA-Seq. While either condition caused RW1 large-scale shifts in gene expression compared to succinate control (SUC), differences in gene expression between these two conditions were marked by a small number (86) of differentially expressed genes. The differences in gene expression may reflect the underlying adaptive mechanisms by which RW1 cells sense and deploy pathways to access dioxin intercalated in the clay.

overall_design: Transcriptomes were sampled in triplicate (3 biological replicates) at early to mid log phase from cultures grown under three different conditions: succinate (SUC), dibenzo-p-dioxin (DD), and clay-sorbed dibenzo-p-dioxin (DDSAP).

first: Tamara

last: Tsoi

first: J

middle: B

last: Sallach

first: Cun

first: Jeff

last: Landgraf

first: Mark

last: Bezdek

first: Gerben

last: Zylstra

first: Hui

last: Li

first: Cliff

last: Johnston

first: Brian

last: Teppen

first: James

last: Cole

first: Steve

last: Boyd

last: Tiedje


title: Elongation Factor TFIIS Prevents Transcription Stress and R-Loop Accumulation to Maintain Genome Stability [ RNA-seq]

status: Public on Jun 13 2019

city: London

first: Richard

middle: James

last: Mitter

state: LONDON

address: 1 Midland Road

department: Bioinformatics & Biostatistics

institute: The Francis Crick Institute

summary: Connections between RNA polymerase II (RNAPII) transcription stress, R-loops, and genome instability have been established however, the underlying mechanisms remain poorly understood. Here we used a mutant version of elongation factor TFIIS (TFIISmut) to specifically induce increased levels of RNAPII pausing, arrest, and/or backtracking in human cells. TFIISmut expression results in slower elongation rates, relative depletion of polymerases from the end of genes, and increased levels of stopped RNAPII. It affects mRNA splicing and termination as well. Remarkably, however, TFIISmut expression also dramatically increases R-loops, which may form at the anterior end of backtracked RNAPII and trigger genome instability, including DNA strand breaks. These results shed new light on the relationship between transcription stress and R-loops, and suggest that different classes of R-loops exist, potentially with distinct consequences for genome instability.

overall_design: To study RNAPII backtracking and its effects in human cells, we used HEK293 TREX cells in which we overexpressed, under the control of a dox-promoter, a dominant negative form of TFIIS (TFIIS mut), an elongation factor necessary for stimulating RNAPII intrinsic cleavage activity. TFIISmut cells were maintained in the presence of Dox to ensure over-expression for 48 hours prior to harvest..

first: Diana

last: Zatreanu

first: Zhong

last: Han

first: Emanuela

last: Tumini

first: Hannah

last: Williams

first: Lea

last: Gregersen

first: A

last: Dirac-Svejstrup

first: Andres

last: Aguilera

first: Jesper

middle: Q

last: Svejstrup


title: Elongation Factor TFIIS Prevents Transcription Stress and R-Loop Accumulation to Maintain Genome Stability [ChIP-seq]


title: Effects of a low carbohydrate ketogenic diet on the expression of genes in mouse tissues

status: Public on Jun 10 2019

city: Tsukuba

first: Tetsuya

last: Okuda

state: Ibaraki

address: Central6, 1-1-1, Higashi

department: Bioproduction Research Institute

institute: National Institute of Advanced Industrial Science and Technology (AIST)

summary: Low carbohydrate ketogenic diet (LCKD) has been used for treatment obesity and epilepsy, however, the molecular mechanism of effect of a LCKD (F3666) on tissues has not been fully investigated. In this study, novel molecular targets of LCKD were explored by gene expression profiling in the liver and cerebral cortex of an LCKD-fed mouse model.

overall_design: Total RNA prepared from mouse tissues after 7 weeks LCKD feeding was analyzed by Agilent Gene Expression Microarray. By comparing the results with the data of regular chow (CE-2)-fed mice, significantly altered genes were extracted.


title: Stellate cells, hepatocytes and endothelial cells imprint the Kupffer cell identity on monocytes colonizing the liver macrophage niche (microarray)

status: Public on Aug 14 2019

city: Zwijnaarde

first: Liesbet

last: Martens

address: Technologiepark 927

department: VIB Inflammation Research Center

institute: VIB-University of Ghent

type:
Expression profiling by array
Third-party reanalysis

summary: Macrophages are strongly adapted to their tissue of residence. Yet, we know little about the cell-cell interactions that imprint the tissue-specific identities of macrophages in their respective niches. Using conditional depletion of liver Kupffer cells, we traced the developmental stages of monocytes differentiating into Kupffer cells and mapped the cellular interactions imprinting the Kupffer cell identity. Kupffer cell loss induced the tumor necrosis factor (TNF) and interleukin-1 (IL-1) receptor-dependent activation of stellate cells and endothelial cells, resulting in the transient production of chemokines and adhesion molecules orchestrating monocyte engraftment. Engrafted circulating monocytes transmigrated into the perisinusoidal space, and acquired the liver-associated transcription factors ID3 and LXRα. Coordinated interactions with hepatocytes induced ID3 expression, while endothelial cells and stellate cells induced LXRα via a synergistic NOTCH-BMP pathway. This study shows that the Kupffer cell niche is composed of stellate cells, hepatocytes and endothelial cells that together imprint the liver-specific macrophage identity.

overall_design:
Microarray = different time points during development of Kupffer cells during embryogenesis and after depletion. This data was compared to previously published data (GSE76999, GSE75225  and GSE117080) and to other tissue resident macrophages from the Immgen Consortium Database (GSE15907).
Bulk RNA Seq = Hepatic Stellate Cells (HSCs), Liver Sinusoidal Endothelial Cells (LSECs) and hepatocytes. For each cell type there are 4 replicates from Pbs mice, 12h after DT-injection mice and 36h after DT-injection mice.

first: Johnny

last: Bonnardel

first: Wouter

last: T’Jonck

first: Djoere

last: Gaublomme

first: Robin

last: Browaeys

first: Charlotte

last: Scott

first: Bavo

last: Vanneste

first: Sofie

last: De Prijck

first: Sergei

last: Nedospasov

first: Anna

last: Kremer

first: Evelien

last: Van Hamme

first: Peter

last: Borghgraef

first: Wendy

last: Toussaint

first: Pieter

last: De Bleser

first: Inge

last: Mannaerts

first: Alain

last: Beschin

first: Leo

last: van Grunsven

first: Bart

middle: N

last: Lambrecht

first: Tom

last: Taghon

first: Saskia

last: Lippens

first: Dirk

last: Elewaut

first: Yvan

last: Saeys

first: Martin

last: Guilliams


title: Elongation Factor TFIIS Prevents Transcription Stress and R-Loop Accumulation to Maintain Genome Stability

first: Richard

last: Mitter

overall_design: Refer to individual Series


title: Clone-specific transcript profiling of ‘Nebbiolo’ grape berries unveils environmental responses mediated by sugar and secondary metabolite signalling

status: Public on Jun 03 2019

city: Verona

first: Massimo

last: Delledonne

address: Strada le Grazie 15

institute: Universita' degli Studi di Verona

summary:
Goal: In the last years, many research efforts were addressed to uncover how grapevine cultivars respond to environment, nevertheless little is known on the molecular processes underlying the interplay between clones (vegetatively propagated lines of selected mother plants) of the same cultivar and environment. The goal of this study was to explore the whole complexity of the clone x environment (C x E) interaction through the identification of molecular changes controlled by clone, vineyard, phenological phase or a combination of them.

Methods: We analyzed the transcriptome of berries collected over the ripening period from three Vitis vinifera cv Nebbiolo clones (CVT 71, CVT 423, CVT 185) grown in different vineyards, in two vegetative seasons. A multidisciplinary approach was adopted by implementing RNA-seq data with: i) expression analysis of candidate genes; ii) quantification of abscisic acid, flavonoids and stilbenoids; iii) agronomical parameters; and iv) climatic data.

Results: Transcripts involved in sugar signalling, anthocyanin biosynthesis and transport were differently modulated among diverse clones, accordingly to berry agronomical features. Conversely, genes linked to the production of secondary metabolites enhancing defence responses, such as stilbene synthase genes, were significantly affected by vineyard location, consistently with stilbenoid accumulation patterns.

Conclusion: The collected results attest that clone-specific metabolic responses do play a key role in shaping agronomic performances of a grape variety in different environments, thus providing valuable indications for orienting viticultural practices.

overall_design: mRNA profiles of berries collected from 3 clones (CVT 71, CVT 423, CVT 185) of Vitis vinifera cv Nebbiolo in 3 vineyards (V1, V2, V3) at 3 developmental stages (E-L31, E-L35, E-L38) were generated by Illumina deep sequencing. Three biological replicates were processed for each thesis.

first: Chiara

last: Pagliarani

first: Paolo

last: Boccacci

first: Walter

last: Chitarra

first: Emanuela

last: Cosentino

first: Marco

last: Sandri

first: Irene

last: Perrone

first: Alessia

last: Mori

first: Danila

last: Cuozzo

first: Luca

last: Nerva

first: Marzia

last: Rossato

first: Paola

last: Zuccolotto

first: Mario

last: Pezzotti

first: Franco

last: Mannini

first: Ivana

last: Gribaudo

first: Giorgio

last: Gambino


title: NASC-seq monitors RNA synthesis in single cells

status: Public on Jul 18 2019

city: Solna

first: Gert-Jan

last: Hendriks

address: Solnavägen 9

department: Cell and Molecular Biology

country: Sweden

institute: Karolinska Institutet

summary: Sequencing of newly synthesized RNA can monitor transcriptional dynamics with great sensitivity and high temporal resolution, but is currently restricted to populations of cells. Here, we develop new transcriptome alkylation-dependent single-cell RNA sequencing (NASC-seq), to monitor newly synthesized and pre-existing RNA in single cells. We validate the method on pre-alkylated RNA, and by demonstrating that more newly synthesized RNA was detected for genes with known high mRNA turnover. NASC-seq reveals rapidly up- and down-regulated genes during T-cell activation, and RNA arising from induced genes is essentially only newly synthesized. The newly synthesized and pre-existing transcriptomes after T-cell activation are distinct, confirming that we simultaneously measure gene expression at two time points in single cells. Altogether, NASC-seq is an accessible and powerful tool to investigate transcriptional dynamics that enables the precise monitoring of RNA synthesis at flexible time periods during homeostasis, perturbation responses and cellular differentiation.

overall_design: For each single-cell RNA-seq experiment, 188 single cells were sequenced according to the NASC-seq protocol. The following experiments are included:  exp1: K562 cells were labelled for 60 minutes with 50uM 4sU. Negative controls and unlabelled cells are included in the experiment  exp2: Jurkat cells were stimulated with PMA and ionomycin and simultaneously labelled with 50uM 4sU for 15 minutes. Negative controls, unlabelled cells and unstimulated cells are included in the experiment.  exp3: Jurkat cells were stimulated with PMA and ionomycin and simultaneously labelled with 50uM 4sU for 30 minutes. Negative controls, unlabelled cells and unstimulated cells are included in the experiment.  exp4: Jurkat cells were stimulated with PMA and ionomycin and simultaneously labelled with 50uM 4sU for 60 minutes. Negative controls, unlabelled cells and unstimulated cells are included in the experiment.  TT-seq: Jurkat cells were stimulated with PMA and ionomycin for 15 or 30 minutes and labelled with 500uM 4sU during the last 5 minutes of stimulation. Biological duplicates of TT-seq samples were sequenced.

first: Lisa

last: Jung

first: Anton

last: Larsson

first: Michael

last: Lidschreiber

first: Oscar

last: Forsman

first: Katja

first: Patrick

last: Cramer

first: Rickard

last: Sandberg


title: Elongation Factor TFIIS Prevents Transcription Stress and R-Loop Accumulation to Maintain Genome Stability [TT-seq]

first: Richard

last: Mitter

first: Diana

last: Zatreanu

first: Zhong

last: Han

first: Emanuela

last: Tumini

first: Hannah

last: Williams

first: Lea

last: Gregersen

first: A

last: Dirac-Svejstrup

first: Andres

last: Aguilera

first: Jesper

last: Svejstrup


title: Integrative analyses of rhabdoid tumors across all anatomical sites reveal subgroups with possible immune modulation through epigenetic dysregulation

status: Public on Jun 01 2019

city: Heidelberg

first: Pascal

middle: David

last: Johann

address: Im Neuenheimer Feld 580

department: Pediatric Neurooncology

institute: German Cancer Research Center

type: Methylation profiling by genome tiling array

summary: Extra-cranial malignant rhabdoid tumors (MRTs) and cranial atypical teratoid RTs (ATRTs) are pediatric cancers driven by SMARCB1 loss. To understand biological relationships between MRTs and ATRTs, we performed integrative data analyses of 140 MRTs and 161 ATRTs, and detected features conserved between MRTs and the MYC subgroup of ATRTs, including global DNA hypomethylation and over-expression of HOX and mesenchymal development genes, thereby distinguishing them from other ATRT subgroups that exhibit neural-like features. Our analyses revealed five DNA-methylation subgroups associated with distinct anatomical sites, SMARCB1 alteration patterns, and distinct gene-expression and enhancer profiles. We also report on RT subgroups with expression patterns indicative of both increased immune infiltration and expression of immune checkpoint genes, which were confirmed using immunohistochemical techniques.

overall_design: Comparative analysis of the methyolme of eMRT and ATRT by methylation arrays

first: Elizabeth

last: Chun


title: Gene expression of Porphyromonas gingivalis ATCC 33277 when growing in an in vitro multispecies biofilm

status: Public on Jun 05 2019

city: Madrid

first: Patricia

last: Romero Lastra

address: Plaza de Ramon y Cajal s/n

institute: Universidad Complutense de Madrid

summary: The aim of the present study to compare the transcriptomic profile of  P.gingivalis when growing within an in vitro multispecies biofilm or in a planktonic state, using microarray technology.

overall_design: P. gingivalis ATCC 33277 was grown in anaerobiosis within multi-well culture plates at 37ºC for 96h under two conditions: (a) planktonic samples (no hydroxyapatite discs) or (b) within a multispecies-biofilm containing Streptococcus  oralis, Actinomyces naeslundii, Veillonella parvula, Fusobacterium nucleatum and  Aggregatibacter actinomycetemcomitans deposited on hydroxyapatite discs. Scanning Electron Microscopy (SEM) and Confocal Laser Scanning Microscopy (CLSM) combined with Fluorescence In Situ Hybridization (FISH) were used to verify the formation of the biofilm and the presence of P. gingivalis in it. Total RNA was extracted from both the multispecies biofilm and planktonic samples, then purified and, with the use of a microarray, its differential gene expression was analyzed. A linear model was used for determining the differentially expressed genes using a filtering criterion of two fold change (up or down) and a significance p-value of <0.05. Differential expression was confirmed by reverse transcription quantitative polymerase chain reaction (RTqPCR).

last: Romero-Lastra

first: María

last: Sánchez

first: Arancha

last: Llama-Palacios

first: Elena

last: Figuero

first: David

last: Herrera

first: Mariano

last: Sanz


title: Evaluation of immunostimulating effects of mineral heat ware in pig

status: Public on May 29 2019

city: Seoul

first: Suji

last: Kim

state: Gwanak-gu

address: 1, Gwanak-ro

department: Veterinary Medicine

institute: Seoul National University

summary: Thermal condition is an important envrionment factor to produce healthy pig. A mineral heat ware, RAVI is a regenerative far-infrared heating system. The purpose of this sutyd was to evaluate immunostimulating effects in pigs when RAVI was applied to maintain thermal condition compared with gasoline heater.

overall_design: Twenty of six-week-old waened pigs were raised in two rooms, each equipped with a gasoline heater and a mineral heat ware for 8 weeks. Two weeks after the stocking, the porcine atropic thinitis vaccine was administered and microarray analysis was performed using the whole blood RNA collected form pigs at 2 weeks, 4 weeks, and 8 weeks post vaccination.

first: Han

middle: S

last: Yoo


title: Dynamic chromatin structure during early ILC progenitor differentiation

status: Public on May 30 2019

city: Bethesda

first: Keji

last: Zhao

state: MD

address: 9000 Rockville Pike

department: Systems Biology Center

institute: NHLBI,NIH

summary: To characterize the epigenomes during early ILC progenitor differentiation, we did DNase-seq, CUT&RUN and scMNase-seq assays in all-lymphoid progenitor (ALP), early innate lymphoid progenitor (EILP), and innate lymphoid cell progenitor (ILCP) cells.

overall_design: We did the CUT&RUN for identifing genome-wide TCF1 binding in EILP cells and DNase-seq in ALP, EILP and ILCP cells.  Samples that include "_tech1" or "_tech2" indicate that those samples with same GB identifier were technical repeats with different libraries made independently from the same RNA extraction.

first: Gang

last: Ren

first: Binbin

last: Lai


title: Comprehensive epigenome characterization reveals the diverse transcriptional regulation across human vascular endothelial cells (RNA-seq)

status: Public on May 24 2019

city: Bunkyo-ku

first: Ryuichiro

last: Nakato

state: Tokyo

address: 1-1-1 Yayoi

department: Institute for Quantitative Biosciences

institute: The University of Tokyo

summary:
Endothelial cells (ECs) make up the innermost layer throughout the entire vasculature. Their phenotypes and physiological functions are initially regulated by developmental signals and extracellular stimuli. The underlying molecular mechanisms responsible for the diverse phenotypes of ECs from different organs are not well understood.
To characterize the transcriptomic and epigenomic landscape in the vascular system, we cataloged gene expression and active histone marks in nine types of human ECs (generating 148 genome-wide datasets) and carried out a comprehensive analysis with chromatin interaction data. We developed a robust procedure for comparative epigenome analysis that circumvents variations at the level of the individual and technical noise derived from sample preparation under various conditions. Through this approach, we identified 3765 EC-specific enhancers, some of which were associated with disease-associated genetic variations. We also identified various candidate marker genes for each EC type. We found that the nine EC types can be divided into two subgroups, corresponding to those with upper-body origins and lower-body origins, based on their epigenomic landscape. Epigenomic variations were highly correlated with gene expression patterns, but also provided unique information. Most of the deferentially expressed genes and enhancers were cooperatively enriched in more than one EC type, suggesting that the distinct combinations of multiple genes play key roles in the diverse phenotypes across EC types. This comprehensive analysis of epigenome characterization of EC types reveals diverse transcriptional regulation across human vascular systems. These datasets provide a valuable resource for understanding the vascular system and associated diseases.

overall_design: RNA-Seq experiments were conducted in human vascular endothelial cells. Fastq files have been deposited in the SRA database under accession numbers SRP188296 (BioProject accession: PRJNA526601).

first: Youichiro

last: Wada

first: Toutai

last: Mitsuyama

first: Hiroyuki

last: Aburatani

first: Hiroshi

last: Kimura

first: Katsuhiko

last: Shirahige


title: Transcriptional analysis of tendons from mice with a tenocyte-specific deletion of IGF1R

status: Public on May 28 2019

first: Christopher

last: Mendias

state: NY

address: 535 E 70th St

institute: Hospital for Special Surgery

summary: We report RNA sequencing data from the plantaris tendons of adult male mice in the C57Bl/6 background that either have the IGF1 receptor (IGF1R) present in their tendons (Scx:IGF1R+) or mice in which IGF1R has been deleted in tenocytes expressing scleraxis (Scx:IGF1R-). Mice were created by crossing ScxCreERT2 mice with IGF1R flox/flox mice. Mice were treated with tamoxifen for 5 days to induce recombination at the IGF1R allele, and then subjected to a synergist ablation procedure in which the Achilles tendon is removed, resulting in compensatory growth of the plantaris tendon. Mice were analyzed either 7 or 14 days after synergist ablation. Control mice that did not undergo tamoxifen treatment or synergist ablation were also studied.

overall_design: Differential expression of mRNAs were evaluated from the plantaris tendons of Scx:IGF1R+ and Scx:IGF1R- mice subjected to a growth stimulus for either 7 or 14 days, along with control mice that did not undergo a growth stimulus.


title: Regulation of tendon growth by IGF1 signaling

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


title: RNA sequencing results of EMT and CSC phenotypes induced by modulated expression of wild-type and mutated PSPC1 and PTK6

status: Public on May 22 2019

city: Taipei, Taiwan

first: Yaw-dong

last: lang

state: Taiwan

address: 28 Sec. 2, Academia Rd. Nankang, Taipei 115

department: Institute of biomedical science

institute: Academia Sinica

summary: We found that PSPC1 is the master activator for transcription factors of EMT and stemness and accompanies c-Myc activation to facilitate tumour growth.  To elucidate mechanisms that the underlying post-translational modification of PSPC1, we performed the RNA-seq analysis of overexpression of PSPC1, alone and with PTK6, and PSPC1 tyrosine residue mutation in SK-Hep1 cells. We performed c-terminal of PSPC1 and its mutated nuclear localization site (NLS) in Mahlavu cells to evaluate its inhibitory effects on gene expression. Moreover, we further treated with HGF in Huh-7 cells to examineThe role of downstream signaling pathways of the PSPC1 and PTK6 regulation in microenviroment.

overall_design: Gene expression profiles of PSPC1 with modulated overexpression in SK-Hep1 cells and Mahlavu cells were generated by Illumna RNA sequencing and compared to profiles derived from mock control cells. Gene expression profiles of HGF treatment in Huh-7 cells were generated by Illumna RNA sequencing and compared to profiles derived from mock control cells.

first: Y

middle: D

last: Lang

last: Jou


title: Transcriptional analysis of IGF1 treatment of mouse tenocytes over a 24 hour time course

summary: We report RNA sequencing data from tenocytes treated with IGF1. Tenocytes were obtained from the tail tendons of adult C57Bl/6 mice via collagenase digestion. Tenocytes were grown to 60% confluence, and then treated with 100ng/mL of recombinant IGF1 for a period of 0, 1, 2, 6, or 24 hours. Experiments were conducted in quadruplicate. RNA was isolated and prepared for RNA sequencing.

overall_design: Differential expression of mRNAs were evaluated from tenocytes isolated from tail tendons of adult wild type C57Bl/6 mice that were treated with recombinant IGF1 for 0, 1, 2, 6, and 24 hours.


title: Expression analysis of ethanol treatment on SK-N-BE(2)

status: Public on May 21 2019

city: Indianapolis

first: Andy

last: Chen

state: IN

address: 410 W 10th Street, Suite 5000

department: MMGE

institute: IUSM

summary: To evaluate the impact of ethanol treatment on neuronal cells, we examined the expression of SK-N-BE(2) neuronal cell line 24 h and 42 h after treating with 10 mM and 20 mM ethanol by RNA sequencing.

overall_design: RNA-seq results from 4 replicates of SK-N-BE(2) cells with 0, 10 mM, and 20 mM ethanol harvest after 24 h and 42 h

first: Kriti

last: Thapa

first: Hongyu

last: Gao

first: Xiaoling

last: Xuei

first: Howard

last: Edenberg

first: Yunlong

last: Liu


title: Plasmacytoid dendritic cells develop from Ly6D+ lymphoid progenitors distinct from the myeloid lineage

status: Public on May 10 2019

city: Singapore

first: Jinmiao

address: 8A Biomedical Grove, #04-06 Immunos

country: Singapore

institute: Singapore Immunology Network

summary: We provide a comprehensive single cell mRNA-seq analyses of mouse bone marrow progenitors revealing early lineage commitment in the bone marrow.

overall_design: Gene profiles of ex vivo sorted single cell mouse bone marrow progenitor populations were obtained by C1 single cell mRNA sequencing.

first: Regine

last: Dress

first: Andreas

last: Schlitzer

first: Florent

last: Ginhoux


title: Identification of SERPINE1 as a Regulator of Glioblastoma Cell Dispersal via Analyzing Dynamic Transcriptome of Dispersing Cells

status: Public on May 09 2019

city: Istanbul

first: Tugba

last: Bagci-Onder

state: Sariyer

address: Koc University School of Medicine

department: School of Medicine

country: Turkey

institute: Koc University

summary: With a model mimicking GBM tumor cell dispersal, transcriptome changes between core (immotile) and dispersive (motile) cells were analyzed. Many genes are differentially expressed between these populations. This study focused on the genes that are significantly upregulated in dispersive cells. Besides gene sets related with the cell cycle and cell survival, epithelial to mesenchymal transition gene set is upregulated in dispersive cells. In this gene set, this study identified SERPINE1 gene as an important regulator of GBM cell dispersal.

overall_design: Examination of core and dispersive populations' transcriptome during U373 cell spheroid dispersal. 2 sets of samples were prepared each for core and dispersive cells.

first: Tuğba

last: Bağcı-Önder

first: Fidan

last: Şeker

first: Fırat

last: Uyulur


title: Single-cell RNA-Seq Investigation of Foveal and Peripheral Expression in the Human Retina

status: Public on May 17 2019

city: Iowa City

first: Todd

last: Scheetz

state: IA

address: 3181B MERF

institute: UNIVERSITY OF IOWA

summary:
Purpose: Single-cell RNA sequencing has revolutionized cell-type specific gene expression analysis. The goals of this study are to compare cell specific gene expression patterns between retinal cell types originating from the fovea and the periphery of human eyes.
Methods: Independent libraries were prepared for foveal and peripheral samples of neural retina from three donors using the 10x Chromium system. Libraries were sequenced on a HiSeq4000. Sequenced reads were mapped to the human genome build hg19 will CellRanger(v3.0.1) and filters removed cells likely to be doublets or cells with a high proportion of mitochondrial reads. Clustering of cells with similar expression profiles was performed with Seurat (v2.3.4).
Results: Independent libraries were prepared for foveal and peripheral samples of neural retina from three donors using the 10x Chromium system. Libraries were sequenced on a HiSeq4000. Sequenced reads were mapped to the human genome build hg19 will CellRanger(v3.0.1) and filters removed cells likely to be doublets or cells with a high proportion of mitochondrial reads. Clustering of cells with similar expression profiles was performed with Seurat (v2.3.4).
Conclusions: Our study generates a large atlas of human retinal transcriptomes at the single cell level. We identified the majority of expected neural and supportive cell types, and describe regional differences in gene expression between the fovea and the periphery. Our results show that that single-cell RNA sequencing can be performed on human retina after cryopreservation, and that cone photoreceptors and Muller cells demonstrate region-specific patterns of gene expression.

overall_design: mRNA profiles for thousands of cells from foveal and peripheral retinal isolates were generated from three human donor eyes using 10X Genomics Chromium single-cell system followed by sequencing on an Illumina HiSeq 4000.

middle: E

first: Robert

last: Mullins


title: Transcriptome sequencing of a keystone aquatic herbivore yields insights on the temperature-dependent metabolism of essential lipids

status: Public on May 04 2019

city: Düsseldorf

first: Heidrun

middle: Sigrid

last: Windisch

state: NRW

address: Universitätsstrasse 1

department: Cell Biology and Zoology

institute: Heinrich-Heine-University


title: Transcriptomic analysis of gastric cancer cells in response to stable over-expression of circular RNA derived from AGO2 gene (circAGO2)

city: Wuhan

first: Qiangsong

last: Tong

state: Hubei

address: 1277 Jiefang Avenue

department: Department of Surgery

country: China

institute: Union Hospital of Tongji Medical College

summary: Argonaute 2 (AGO2) is the core component of microRNA (miRNA)-induced silencing complex, and plays a compelling role in tumorigenesis and aggressiveness. Recent studies have implicated the emerging roles of circular RNAs (circRNAs) in cancer progression. However, the roles of circRNAs in regulating the functions of AGO2 in cancer still remain elusive. We indentify one intronic circRNA derived from AGO2 gene (circAGO2) as a novel regulator of AGO2-miRNA complexes and cancer progression. To investigate the mechanisms underlying the oncogenic functions of circAGO2, we employed the Illumina HiSeq X Ten as a discovery platform to analyze the transcriptome profiling changes of human gastric cancer MKN-45 cells in response to stable over-expression of circAGO2. The results showed that stable over-expression of circAGO2 led to altered expression of 624 human mRNAs, including 257 up-regulated genes and 367 down-regulated genes. Furthermore, we validated the RNA-seq results by real-time RT-PCR with high identity. Overall, our results provided fundamental information about the transcriptomic changes in response to circAGO2 over-expression in human cancer cells, and these findings will help us understand the pathogenesis of cancer progression.

overall_design: Total RNA of cells stably transfected with empty vector or circAGO2 was extracted using the TRIzol® reagent according to the manufacturer's instructions. RNA concentration was measured using a Qubit® RNA Assay Kit with a Qubit® 2.0 Fluorometer (Life Technologies, Inc.), and integrity was assessed using the RNA Nano 6000 Assay Kit with a Bioanalyzer 2100 system (Agilent Technologies, CA). Library preparation and transcriptome sequencing on an Illumina HiSeq X Ten platform were performed by Novogene Bioinformatics Technology Co., Ltd. (Beijing, China), and 100 bp paired-end reads were generated. HTSeq v0.6.0 was used to count the reads numbers mapped to each gene.

first: Tong

last: Qiangsong

first: Zheng

last: Liduan


title: A mouse model of acute post-surgical pain

status: Public on May 01 2019

city: St Lucia

first: Alexander

last: Mueller

state: Queensland

address: 306 Carmody Rd

department: Institute for Molecular Bioscience

institute: University of Queensland

summary: Pain is the leading cause of disability in the developed world but remains a poorly treated condition. Specifically, post-surgical pain continues to be a frequent and undermanaged condition. Here, we investigate the analgesic potential of pharmacological NaV1.7 inhibition in a mouse model of acute post-surgical pain, based on incision of the plantar skin and underlying muscle of the hind paw. We demonstrate that local and systemic treatment with the selective NaV1.7 inhibitor μ-theraphotoxin-Pn3a is effectively anti-allodynic in this model and completely reverses mechanical hypersensitivity in the absence of motor adverse effects. In addition, the selective NaV1.7 inhibitors ProTx-II and PF-04856264 as well as the clinical candidate CNV1014802 also reduced mechanical allodynia. Interestingly, co-administration of the opioid receptor antagonist naloxone completely reversed analgesic effects of Pn3a, indicating an involvement of endogenous opioids in the analgesic activity of Pn3a. Additionally, we found super-additive antinociceptive effects of sub-therapeutic Pn3a doses not only with the opioid oxycodone but also with the GABAB receptor agonist baclofen. Transcriptomic analysis of gene expression changes in dorsal root ganglia of mice post-surgery revealed decreased expression of several pro-nociceptive genes including N- and P/Q-type voltage-gated calcium channels important for neurotransmitter release, which suggest a reactive compensatory mechanism to reduce excessive pain similar to the endogenous opioid system. In summary, these findings suggest that pain after surgery can be successfully treated with NaV1.7 inhibitors alone or in combination with baclofen or opioids, which may present a novel and safe treatment strategy for this frequent and poorly managed condition.

overall_design: RNA was extracted from Dorsal Root Ganglia L3, L4 and L5 tissue from wild type C57BL/6 mice, either naïve or from mice that underwent surgery, in triplicates, and sequenced using Illumina NextSeq 500 as 75-nucleotide single-end runs

first: Hana

last: Starobova

first: Irina

last: Vetter


title: RNA sequencing results of wild type and overexpresssion or knockdown PTTG3P in LUAD cell lines based on IlluminaNovaSeq 6000 platform

city: Taipei

first: JOU-HO

last: SHIH

state: Please Select

address: 128 Sec. 2, Academia Rd. Nankang, Taipei

institute: Institute of Biomedical Sciences, Academia Sinica

summary: We found that PTTG3P expression perturbed cancer cell proliferation in lung adenomacarcinoma.

overall_design: Gene expression profiles of PTTG3P with modulated overexpression in A549 cells or knockdown in H1299 cells were generated by Illumna RNA sequencing and compared to profiles derived from control cells.

first: J

middle: H

last: Shih

first: Y

last: Jou


title: FRY Impacts Breast Cancer Malignancy by Inducing an Enrichment of Genes with Tumor-Suppressive Functions and Affecting EMT Potential of Breast Cancer Cells

city: Buffalo

first: Jie

last: Wang

address: Elm and Carlton Streets

department: Department of Biostatistics and Bioinformatics

institute: Roswell Park Comprehensive Cancer Center

summary: Breast cancer is a disease with diverse phenotypes, and while the progression of breast cancer from a localized lesion to metastatic disease is well understood clinically. Our previous study genetically linked the FRY gene with differential susceptibility to mammary carcinogenesis; however, little is known about the mechanistic insights of FRY in mammalian cells and breast cancer progression. Through RNA-Seq analysis of series of constructed cell lines based on highly malignant triple-negative human breast cancer cells MDA-MB-231, we show that enhanced FRY induces massive changes in gene expression, which favor signalings and molecules with tumor-suppressive functions and engage in promoting cell differentiation, maintaining the normal and histopathological characteristics of epithelial cells, and in regulating EMT potential of breast cancer cells. As a result, the proliferation, migration and invasion potentials of breast cancer cells are significantly inhibited both in vitro and in xenograft tumor models. Importantly, consistent with FRY’s anti-tumor progression role, decreased FRY protein levels are correlated with poorly differentiated and aggressive tumor phenotypes in the analysis of two clinically annotated breast cancer cohorts. Our results identify FRY’s potent tumor-suppressive role at the gene transcription and molecular function levels, providing a novel biomarker for breast cancer prognosis and a target for therapeutic intervention.

overall_design: Examination of two repeats for parental, vector-ctrl and FRY expressing MDA-MB-231 cells.

first: Xusheng

last: Chen

first: Xuefeng

last: Ren


title: IL-1β driven amyloid plaque clearance is associated with an expansion of transcriptionally reprogrammed microglia

city: Rochester

first: M. Kerry

last: O'Banion

address: 601 Elmwood Avenue

department: Neuroscience

institute: University of Rochester

summary: Neuroinflammation is thought to contribute to the pathogenesis of Alzheimer’s disease (AD), yet numerous studies have demonstrated a beneficial role for neuroinflammation in amyloid plaque clearance. We have previously shown that sustained expression of IL-1β in the hippocampus of APP/PS1 mice decreases amyloid plaque burden independent of recruited CCR2+ myeloid cells, suggesting resident microglia as the main phagocytic effectors of IL-1β-induced plaque clearance. To date, however, the mechanisms of IL-1β-induced plaque clearance remain poorly understood. To determine whether IL-1β-induced plaque clearance is due to enhanced microglial phagocytosis of Aβ, APP/PS1 mice induced to express mature human IL-1β in the hippocampus via adenoviral transduction were treated with the Aβ fluorescent probe methoxy-X04 (MX04) and microglial internalization of Aβ was analyzed by flow cytometry and immunohistochemistry. We found that resident microglia (CD45loCD11b+) constituted >70% of the MX04+ cells in both control and IL-1β-treated conditions, and that <10% of MX04+ cells were recruited myeloid cells (CD45hiCD11b+). However, we found that IL-1β treatment did not augment the percentage of MX04+ microglia nor the quantity of Aβ internalized by individual microglia. Instead, we found that IL-1β treatment resulted in a significant increase in the total number of MX04+ microglia in the hippocampus due to IL-1β-induced proliferation. Consistent with these results, transcriptomic analyses revealed very similar gene expression profiles between MX04+ and MX04- microglia, indicating IL-1β does not drive enhanced expression of phagocytosis-related genes. By contrast, IL-1β treatment was associated with large-scale changes in the expression of genes related to proliferation, immune function and inflammation. Together, these studies demonstrate that IL-1β induces microglial proliferation and the expression of genes involved in inflammatory immune functions that may be related to Aβ clearance.

overall_design: Hippocampal microglia mRNA profiles of 8-month-old APP/PS1-rAAV2-Phe and APP/PS1-rAAV2-IL-1β further sorted on MX04+ (Aβ+) and MX04- (Aβ-). Two biological replicates of pooled cells from 6-8 APP/PS1 mouse hippocampi for each experimental group were sequenced using the Illumina HiSeq 2500.

first: Fátima

last: Rivera-Escalera

first: M


title: mRNA expression profiling in MDA-MB-231 (LM1) cells with a tet-incible MBD2 or p66α knock down, or treated with MBD2-targeting small molecule ABA or APC

status: Public on May 03 2019

first: Chul Geun

last: Kim

address: wangsimni-ro 222

department: Life Science

institute: Hanyang University

summary: The goal of this study is to compare mRNA expression profiles among wild type, MBD2 knock down, p66α knock down, and ABA- and APC-treated cells.. All cell lines were maintained in DMEM supplemented with 10% fetal bovine serum and 1% penicillin/streptomycin. For the doxycycline inducible gene expression regulation, each cell lines were incubated in the presence of 1 μg/ml doxycycline for 2 days. For the drug treatment, MDA-MB-231 (LM1) cells were incubated in the presence of 10 μM ABA or APC for 2 days. Total cellular RNAs were extracted using Qiazol reagent. The RNA quality was examined by spectrophotometry, agarose gel electrophoresis (calculating the 18S and 28S rRNA ratio) and an Agilent Technologies 2100 Bioanalyzer (ensuring a RIN value greater than 7). Library was prepared with QuantSeq 3’ mRNA-Seq Library Prep Kit FWD. The constructed libraries were subjected to 75-bp single-end sequencing using an Illumina NextSeq 500 sequencer. Using an optimized data analysis workflow, we mapped to the human genome (build hg19) and identified 25,737 transcripts in these cell lines.

overall_design: Duplicated mRNA profiles of WT, tet-inducible MBD2 or p66α knock down, and ABA- or APC-treated MDA-MB-231 (LM1) cells are generated by quant-sequencing using Illumina Casava1.8.and then examine the expression patterns among these groups of cells.

first: C


title: WaGa single cell RNA sequencing

status: Public on Apr 26 2019

city: Essen

first: Jan

last: Gravemeyer

address: Universitätstraße 1

institute: DKZF

summary: The Merkel cell carcinoma cell line (MCC) WaGa was sequenced using the 10x Genomics 3' Chromium v2.0 platform in order to analyse transcriptional heterogeneity on a single cell level

overall_design: scRNAseq of a single sample of the MCC cell line WaGa

first: Kaiji

last: Fan

first: Jürgen

last: Becker


title: Analysis of MLLT3 genomic distribution and related chromatin features in fetal liver hematopoietic stem-progenitor cells [ChIP-seq_cultured_FL-HSPC]

status: Public on Apr 29 2019

city: Los Angeles

first: Vincenzo

last: Calvanese

address: 615 Charles E Young Dr S

institute: UCLA

summary: MLLT3 binds to TSS of genes active in FL-HSPC

overall_design: ChIP analysis of sorted FL-HSPC, uncultured or cultured for 4, and K562 cells. This is an addition to the other experiments in the superseries.  Sonicated chromatin (250,000-100,000 cells per IP) DNA was immunoprecipitated with antibodies against the indicated mark. DNA was amplified with Nugen Ovation V2 kit and sequenced in single end 50bp in multiplexed run on an Illumina HiSeq4000 instrument.

first: Andrew

middle: T

last: Nguyen

first: Timothy

last: Bolan

first: Yanling

first: Fides

last: Lay

first: Mattias

last: Magnusson

first: Siavash

last: Kurdistani

first: Hanna

last: Mikkola


title: Charting in vitro beta cell differentiation by single cell RNA sequencing

city: Cambridge

first: Adrian

last: Veres

state: Massachusetts

address: 7 Divinity Ave

institute: Harvard University

summary: In vitro differentiation of human stem cells can produce pancreatic beta cells, the insulin-secreting cell type whose loss underlies Type 1 Diabetes. As a step towards mastery of this process, we report on transcriptional profiling of >100,000 individual cells sampled during in vitro beta cell differentiation and describe the cells that emerge. We resolve populations corresponding to beta cells, alpha-like poly-hormonal cells, non-endocrine cells that resemble pancreatic exocrine cells and a previously unreported population resembling enterochromaffin cells. We show that the beta and alpha-like cells are stable for weeks in culture without exogenous growth factors and that gene expression changes associated with in vivo beta cell maturation are recapitulated in vitro. We demonstrate that stem-cell derived enterochromaffin cells can synthesize and secrete serotonin in vitro. To remove exocrine cells, we characterize a scalable re-aggregation technique that efficiently selects endocrine cells. Finally, we use a high-resolution sequencing time course to characterize gene expression dynamics during human pancreatic endocrine induction from which we develop a lineage model of in vitro beta cell differentiation. This study provides a deeper perspective on the current state of human stem cell differentiation and is a jumping-off point for future endeavors in in vitro differentiation of pancreatic islet cells and their application in regenerative medicine.

overall_design:
Single-cell mRNA sequencing of pluripotent stem cells differentiating in vitro towards pancreatic beta cells.

The data & metadata match the initial submission of the manuscript, not the final version.

first: Aubrey

last: Faust

first: Henry

last: Bushnell

first: Elise

last: Engquist

first: Jennifer

last: Hyoje-Ryu Kenty

first: George

last: Harb

first: Yeh-Chuin

last: Poh

first: Elad

last: Sintov

first: Mads

last: Gürtler

first: Felicia

middle: W

last: Pagliuca

first: Quinn

middle: P

last: Peterson

first: Douglas

middle: A

last: Melton


title: Transcriptomically-guided mesendoderm induction of human pluripotent stem cells using a systematically defined culture scheme

status: Public on Apr 11 2019

city: Ottawa

first: William

last: Stanford

state: Ontario

address: 501 Smyth Road

institute: Ottawa Hospital Research Institute

summary: Human pluripotent stem cells (hPSCs) are an essential cell source in tissue engineering, studies of development, and disease modeling.  Efficient and broadly amenable protocols for rapid lineage induction of hPSCs are of great interest in the stem cell biology field.  We describe a simple yet robust method for differentiation of hPSCs into mesendoderm in defined conditions utilizing single-cell seeding and BMP4 and Activin A (BA) treatment.  Gene sets and gene ontology terms related to mesoderm and endoderm differentiation were enriched after 48 hours of BA treatment.  After 14 days of differentiation, BA-treated cells expressed genes indicative of mesoderm and endoderm-derived tissue types, including lung, kidney and adipose tissue.  BA treatment was readily incorporated into existing protocols for chondrogenic and endothelial progenitor cell differentiation.  Teratomas formed from BA-treated cells contained higher ratios of mesoderm and endoderm to ectoderm tissue compared to PSC-derived teratomas.  Collectively, these data demonstrate that single-cell seeding with BA treatment is a powerful method for induction of mesendoderm that can be integrated into protocols for mesoderm and endoderm differentiation.

overall_design: 35 samples total. BA (mesendoderm) samples: 6hr, 12hr, 18hr, 24hr x 3, 30hr, 36hr, 42hr, 48hr x 3.  E8 (pluripotency control) samples (6hr, 12hr, 18hr, 24hr, 30hr, 36hr, 42hr, 48hr x 3). E6 (spontaneous differentiation control) samples: 6hr, 12hr, 18hr, 24hr, 30hr, 36hr, 42hr, 48hr x 3. time_zero control

first: Richard

last: Carpenedo

first: R

middle: M

last: Tanner

first: Theodore

last: Perkins