title: Integrative analysis of reference epigenomes in rice [FAIRE-seq]

city: Wuhan

first: Liang

last: Xie

state: Hubei

address: No.1,Shizishan Street

institute: Huazhong Agricultural University

summary: Approximately 73% of rice genomes were annotated with different epigenomic properties. Refinement of promoter regions using open chromatin and H3K4me3-marked regions provided insight into transcriptional regulation. Active and repressed histone modifications and the predicted enhancers varied largely across tissues, whereas inactive chromatin states were relatively stable. Further, we investigated the impact of genetic variants on epigenomic signals and gene expression. Together, these datasets constitute a resource for functional element annotation in rice and indicate the central role of epigenomic information in understanding transcriptional regulation.

overall_design: Here, we generated genome-wide profiling of five histone modifications and RNA polymerase II occupancy using an enhanced chromatin immunoprecipitation (eChIP) approach. By integrating chromatin accessibility, DNA methylation, and transcriptome datasets, we constructed comprehensive epigenome landscapes across various tissues

first: Lun

last: Zhao

first: Qing

last: Zhang


title: RV infections in asthmatics increase ACE2 expression and stimulate cytokine pathways implicated in COVID19

status: Public on Apr 25 2020

city: Tucson

first: Eugene

middle: H

last: Chang

state: AZ

address: 1501 N. Campbell Avenue

department: Department of Otolaryngology

country: USA

institute: The University of Arizona

type: Expression profiling by high throughput sequencing

summary: We present evidence that (1) viral respiratory infections are potential mechanisms of ACE2 overexpression in patients with asthma and (2) ACE activation regulates multiple cytokine anti-viral responses that could explain a mechanism for cytokine surge and associated tissue damage. These results suggest that the recent finding of severe COVID-19 in asthma patients with recent exacerbations may be attributable to synergistic biomolecular interactions with viral co-infections.

overall_design: In a paired design experiments across discovery and validation cohorts of in asthmatic patients the biological replicates treated with RVA and RVC were compared to non-treated ones.

first: Amanda

middle: L

last: Willis

first: Nima

last: Pouladi

first: Jianrong

last: Li

first: Yves

middle: A

last: Lussier


title: Transcriptome analysis of Arabidopsis mature pollen grains from wild type and heterozygous NOT1 mutants

status: Public on Apr 24 2020

city: Oeiras

first: Jorg

middle: D

last: Becker

address: Rua da Quinta Grande No 6

country: Portugal

institute: Instituto Gulbenkian de Ciencia

summary: In flowering plants, pollen development is under a dynamic and well-orchestrated transcriptional control, characterized by an early phase with high transcript diversity and a late post-mitotic phase skewed to a cell-type specific-transcriptome. Such transcriptional changes require a balance between synthesis and degradation of mRNA transcripts, the latter being initiated by deadenylation. The CCR4-NOT complex is the main evolutionary conserved deadenylase complex in eukaryotes, and its function is essential during germline specification in animals. We hypothesised that the CCR4-NOT complex might play a central role in mRNA turnover during microgametogenesis in Arabidopsis. Disruption of NOT1 gene, which encodes the scaffold protein of the CCR4-NOT complex, showed abnormal seed set. Genetic analysis failed to recover homozygous progeny and reciprocal crosses confirmed reduced transmission through the male and female gametophytes. Concordantly not1 embryo sacs showed delayed development and defects in embryogenesis. not1 pollen grains exhibited abnormal male germ unit configurations and failed to germinate. Transcriptome analysis of pollen from not1/+ mutants revealed that lack of NOT1 leads to an extensive transcriptional deregulation during microgametogenesis. Therefore, our work establishes NOT1 as an important player during gametophyte development in Arabidopsis.

overall_design: Pollen grains were isolated from Arabidopsis flowers and total RNA was isolated for expression profiling on Affymetrix Arabidopsis Gene 1.1 ST Array Strip. The study was performed with biological triplicates.

first: Jörg

first: Patricia

last: Pereira

first: Leonor

middle: C

last: Boavida

first: Mario

middle: R

last: Santos


title: Abundant and equipotent founder cells establish and maintain acute lymphoblastic leukaemia

city: Newcastle Upon Tyne

first: Olaf

last: Heidenreich

address: Brewery Lane

department: Wolfson Childhood Cancer Research Centre

country: United Kingdom

institute: Newcastle University

summary: High frequencies of blasts in primary acute lymphoblastic leukaemia (ALL) samples have the potential to induce leukaemia and to engraft mice. However, it is unclear how individual ALL cells each contribute to drive leukaemic development in a bulk transplant and the extent to which these blasts vary functionally. We used cellular barcoding as a fate mapping tool to track primograft ALL blasts in vivo. Our results show that high numbers of ALL founder cells contribute at similar frequencies to leukaemic propagation over serial transplants, without any clear evidence of clonal succession. These founder cells also exhibit equal capacity to home and engraft to different organs, although stochastic processes may alter the composition in restrictive niches. Our findings enhance the stochastic stem cell model of ALL by demonstrating equal functional abilities of singular ALL blasts and show that successful treatment strategies must eradicate the entire leukaemic cell population.

overall_design: Four different ALL primograft samples were transduced with a lentiviral vector barcode library with an estimated complexity in excess of 100,000 barcodes and transplanted intrafemorally into NSG mice. Samples were collected from spleen, bone marrow and meninges, certain samples were transplanted into secondary and tertiary recipients. Barcode complexity was assessed using Illumina MiSeq.

first: Josef

last: Vormoor

first: Alex

last: Elder


title: Stress sensor Ire1 deploys a divergent transcriptional program in response to lipid bilayer stress

city: Singapore

first: Nurulain

last: Ho

address: 60 Nanyang Drive

department: School of Biological Sciences

country: Singapore

institute: Nanyang Technological University

summary: Membrane integrity at the endoplasmic reticulum (ER) is tightly regulated and its disturbance is implicated in metabolic diseases. Using an engineered sensor that activates the unfolded protein response (UPR) exclusively when normal ER membrane lipid composition is compromised, we identified pathways beyond lipid metabolism that are necessary to maintain ER integrity in yeast and in C. elegans. To systematically validate yeast mutants that disrupt ER membrane homeostasis, we identified a lipid bilayer stress (LBS) sensor in the UPR transducer protein Ire1, located at the interface of the amphipathic and transmembrane helices. Furthermore, transcriptome and chromatin immunoprecipitation (ChIP) analyses pinpoint the UPR as a broad-spectrum compensatory response wherein LBS and proteotoxic stress deploy divergent transcriptional UPR programs. Together, these findings reveal the UPR program as the sum of two independent stress responses, an insight that could be exploited for future therapeutic intervention.

overall_design: Δire1 and Δire1Δopi3 mutants expressing Ire1 and ire1ΔLD were experienced protein stress (DTT treatment, 60 minutes) and lipid stress respectively. The average of the log2 fold change was calculated for the genes.

first: Guillaume

last: Thibault

first: Haoxi

last: Wu


title: LCM-RNAseq on human pyramidal neurons and parvalbumin interneurons from frontal cortex

status: Public on Apr 23 2020

city: Groton

first: Thomas

last: Lanz

state: CT

address: Eastern Point Rd

department: Multi-Omics & Biomarkers

institute: Pfizer

summary: Pyramidal neurons and aggregcan-positive interneurons were collected from healthy control human post-mortem brain tissue (Brodmann Area 9) by laser capture microdissection.  RNA was collected from pooled cells from 4 donors, and compared with full stained sections to determine enriched genes expressed in each cell type by RNAseq.

overall_design: n=4 donors, 3 samples per donor: stained section of BA9, n=200 pyramidal neurons, or n=200 aggrecan-positive interneurons.

first: Hualin

last: Xi


title: Glucocorticoids and the cytokines IL-12, IL-15 and IL-18 present in the tumor microenvironment induce PD-1 expression on human Natural Killer cells

city: Rome

first: Linda

last: Quatrini

address: Via San Paolo 15

department: Immunology

country: Italy

institute: IRCCS Bambino Gesù Pediatric Hospital

summary:
Background: PD-1 immune checkpoint blockade has provided significant clinical efficacy across various types of cancer by unleashing both T- and NK cell-mediated anti-tumor responses. However, resistance to immunotherapy occurs for many patients, rendering the identification of the mechanisms that control PD-1 expression extremely important to increase the response to the therapy
Objective: To identify the stimuli and the molecular mechanisms that induce the de novo PD-1 expression on human NK cells in the tumor setting
Methods: NK cells freshly isolated from peripheral blood of healthy donors were stimulated with different combinations of molecules, and PD-1 expression was studied at the mRNA and protein level. Moreover, ex vivo analysis of tumor microenvironment and NK cell phenotype was performed.
Results: Glucocorticoids (GCs) are indispensable for PD-1 induction on human NK cells, in cooperation with a combination of cytokines that are abundant at the tumor site. Mechanistically, GCs together with IL-12, IL-15 and IL-18 not only upregulate PDCD1 transcription, but also activate a previously unrecognized transcriptional program leading to enhanced mRNA translation and resulting in an increased PD-1 protein amount in NK cells.
Conclusion: Our results provide evidence of a novel immune suppressive mechanism of GCs involving the transcriptional and translational control of an important immune checkpoint

overall_design: CD56bright NK cells were stimulated with cytokines in the presence of Dex or DMSO (ctrl) for 6 days, then RNA was extracted for RNA-seq. Three biological replicates were generated for each condition (Dex and ctrl).


title: A novel Sox9/lncRNA H19 axis contributes to hepatocyte death and liver fibrosis

city: Nanjing

first: Yuhui

last: Zang

address: 163 Xianlin Road

country: China

institute: Nanjing University

summary: We generated the AML12 subline stably expressing H19 (AML12-H19) and performed transcriptome RNA sequence.

overall_design: Total RNA was extracted from AML12 subline cells with TRIzol reagent. The RNA purity and integrity were analyzed with the Agilent Bioanalyzer 2100 (Agilent). The mRNA was purified from total RNA after removal of rRNA and sequenced on an Illumina Hiseq2000 system.

first: Wang

last: Chenqi

first: Deng

last: Jia


title: Patient genetics is linked to chronic wound microbiome composition and healing

city: Lubbock

first: Caleb

last: Phillips

state: Texas

address: 2901 Main Ave.

department: Department of Biological Sciences

institute: Texas Tech University

summary: The clinical importance of microbiomes to the chronicity of wounds is widely appreciated, yet little is understood about patient-specific processes shaping wound microbiome composition. Here, a two-cohort microbiome-genome wide association study is presented through which patient genomic loci associated with chronic wound microbiome diversity were identified. Further investigation revealed that alternative TLN2 and ZNF521 genotypes explained significant inter-patient variation in relative abundance of two key pathogens, Pseudomonas aeruginosa and Staphylococcus epidermidis. Wound diversity was lowest in Pseudomonas aeruginosa infected wounds, and decreasing wound diversity had a significant negative linear relationship with healing rate. In addition to microbiome characteristics, age, diabetic status, and genetic ancestry all significantly influenced healing.  Using structural equation modeling to identify common variance among SNPs, six loci were sufficient to explain 53% of variation in wound microbiome diversity, which was a 10% increase over traditional multiple regression. Focusing on TLN2, genotype at rs8031916 explained expression differences of alternative transcripts that differ in inclusion of important focal adhesion binding domains. Such differences are hypothesized to relate to wound microbiomes and healing through effects on bacterial exploitation of focal adhesions and/or cellular migration. Related, other associated loci were functionally enriched, often with roles in cytoskeletal dynamics. This study, being the first to identify patient genetic determinants for wound microbiomes and healing, implicates genetic variation determining cellular adhesion phenotypes as important drivers of infection type. The identification of predictive biomarkers for chronic wound microbiomes may serve as risk factors and guide treatment by informing patient-specific tendencies of infection.

overall_design: An observational two-cohort microbiome genome wide association study (GWAS) was conducted to first identify candidate SNPs which may influence wound microbiome diversity (n = 79 post QC) [Cohort 1 samples in this series]. An experimental followup GWAS (n = 85) was conducted using only the previously suggested SNPs [Cohort 2 samples in this series]. Patients were considered for study if their wound infections had received 16s-based amplicon microbial profiling at their baseline infection, and baseline microbial measurements were then assessed against patient outcomes for individuals whom we had healing outcome data for (n = 58).

first: Craig

last: Tipton


title: The NSD2 p.E1099K Mutation is Enriched at Relapse and Confers Drug Resistance in a Cell Context Dependent Manner in Pediatric Acute Lymphoblastic Leukemia (ATAC-Seq)

city: Staten Island

first: Joanna

last: Pierro

state: NY

address: 550 1st Avenue

institute: NYU Langone Health

type: Genome binding/occupancy profiling by high throughput sequencing

summary: The NSD2 p.E1099K (EK) mutation increases methyltransferase activity and is observed in 10% of acute lymphoblastic leukemia (ALL) samples with enrichment at relapse indicating a role in clonal evolution and drug resistance. To discover mechanisms that mediate the clonal expansion, we engineered B-ALL cell lines (Reh, 697) to overexpress wildtype (WT) and EK NSD2, but observed no differences in proliferation, clonal growth, or chemosensitivity. To address whether NSD2 EK acts collaboratively with other pathways, we used shRNAs to knockdown expression of NSD2 in B-ALL cell lines heterozygous for NSD2 EK (RS4;11, RCH-ACV, SEM). Knockdown resulted in decreased proliferation in all three lines, decreased clonal growth in RCH-ACV, and increased sensitivity to chemotherapeutic agents routinely used in ALL therapy, although the pattern of drug sensitivity varied among cell lines implying that the oncogenic properties of NSD2 mutations are likely cell context specific and rely on cooperative pathways to exert its effect. Knockdown of both Type II and REIIBP EK isoforms had a greater impact than knockdown of Type II alone, suggesting that both SET containing EK isoforms contribute to phenotypic changes driving relapse. Furthermore, in vivo models using both cell lines and patient samples revealed dramatically enhanced proliferation of NSD2 EK compared to WT and reduced sensitivity to 6-mercaptopurine in the relapse sample relative to diagnosis. Finally, EK-mediated changes in chromatin state and transcriptional output differed dramatically among cell lines further supporting a cell context specific role of NSD2 EK. These results demonstrate a unique role of NSD2 EK in mediating clonal fitness through pleiotropic mechanisms dependent on the genetic and epigenetic landscape.

overall_design: NT (non-targeted control) and sh2 (NSD2 knockdown)

first: Jason

last: Saliba

first: Sonali

last: Narang

first: Gunjan

last: Sethia

first: Shella

middle: S

last: Fleur-Lominy

first: Ashfiyah

last: Chowdhury

first: Anita

last: Qualls

first: Hannah

last: Fay

first: Harrison

last: Kilberg

first: Takaya

last: Moriyama

first: Tori

middle: J

last: Fuller

first: David

middle: T

last: Teachey

first: Kjeld

last: Schmiegelow

first: Jun

last: Yang

first: Mignon

last: Loh

first: Patrick

last: Brown

first: Jinghui

last: Zhang

first: Xiaotu

last: Ma

first: Aristotelis

last: Tsirigos

first: Nikki

last: Evensen

first: William

last: Carroll


title: The NSD2 p.E1099K Mutation is Enriched at Relapse and Confers Drug Resistance in a Cell Context Dependent Manner in Pediatric Acute Lymphoblastic Leukemia (ChIP-Seq)

first: Joanna

last: Pierro

overall_design: Reh B-ALL cell lines engineered to overexpress WT or EK NSD2 were sequenced by ChIPseq; only one replicate was submitted for sequencing. RS4;11 B-ALL cell lines engineered to knockdown NSD2 (NT control and sh2 knockdown) were also sequenced; two replicated were submitted per cell line. The following histone marks were analyzed: H3K9ac, H3K27ac, H3K27me3 and H3K36me2. Input files were used as a reference.

first: Jason

last: Saliba

first: Sonali

last: Narang

first: Gunjan

last: Sethia

first: Shella

last: Fleur-Lominy


title: Transcriptomic comparison of young and aged choroid at the bulk and single-cell levels (bulk)

city: Iowa City

first: Andrew

last: Voigt

state: Iowa

address: 375 Newton Road

institute: University of Iowa

summary:
Purpose: The goals of this study were to identify cell specific expression patterns from infant and aged choroid, with a particular emphasis on how endothelial gene expression changes with age.
Methods: Independent libraries were prepared for infant and adult choroidal cells from seventeen human donors. Trephine punch biopsies were used to acquire central RPE and choroidal tissue (infants: 4 mm punch biopsy; adults: 8 mm hemisected punch biopsy). Poly-A selected RNA transcripts underwent 150 base-pair paired-end sequencing with an Illumina HiSeq 2000. Reads were mapped to the human genome hg19 with STAR (version = 020201) and quantified with the featureCounts function from the Rsubread package (version = 1.28.1). Differential expression analysis was performed with edgeR (version = 3.26.8).
Results: Differential expression analysis identified genes enriched in infant versus adult libraries. Differentially expressed genes (p-value < 0.00001, FDR < 0.001, cpm >2, logFC > 1) were input into WebGestalt for over-representation analysis. Aged choroidal samples were enriched in proinflammatory genes.
Conclusions: This study provides a transcriptomic comparison of infant and aged human donor choroid at the bulk and single-cell levels. The aged choroid, in particular the aged vasculature, is enriched in pro-inflammatory genes.

overall_design: mRNA profiles from infant and adult choroidal isolates were generated from seventeen donor eyes followed by sequencing on a HiSeq 2000.

first: Todd

middle: E

last: Scheetz

middle: P

first: Robert

middle: F

last: Mullins


title: Stress dynamically regulates co-expression networks of glucocorticoid receptor-dependent MDD and SCZ risk genes.

status: Public on Apr 22 2020

city: Munich

first: Janine

last: Arloth

address: Kraepelinstr. 2-10

country: Germany

institute: Max Planck Institute of Psychiatry

summary: Early-life adversity is an important risk factor for major depressive disorder (MDD) and schizophrenia (SCZ) that interacts with genetic factors to confer disease risk through mechanisms that are still insufficiently understood. One downstream effect of early-life adversity is the activation of glucocorticoid receptor (GR)-dependent gene networks that drive acute and long-term adaptive behavioral and cellular responses to stress. We have previously shown that genetic variants that moderate GR-induced gene transcription (GR-response eSNPs) are significantly enriched among risk variants from genome-wide association studies (GWASs) for MDD and SCZ. Here, we show that the 63 transcripts regulated by these disease-associated functional genetic variants form a tight glucocorticoid-responsive co-expression network (termed GCN). We hypothesized that changes in the correlation structure of this GCN may contribute to early-life adversity-associated disease risk. Therefore, we analyzed the effects of different qualities of social support and stress throughout life on GCN formation across distinct brain regions using a translational mouse model. We observed that different qualities of social experience substantially affect GCN structure in a highly brain region-specific manner. GCN changes were predominantly found in two functionally interconnected regions, the ventral hippocampus and the hypothalamus, two brain regions previously shown to be of relevance for the stress response, as well as psychiatric disorders. Overall, our results support the hypothesis that a subset of genetic variants may contribute to risk for MDD and SCZ by altering circuit-level effects of early and adult social experiences on GCN formation and structure.

overall_design: Male Balb/c mice were exposed to either early-life adversity (limited nesting and bedding material; LM) or a caring environment (early handling, EH). LM was performed by placing the animals from postnatal day (P) 2 to P9 in a cage with a metal grid instead of bedding material and reduced nesting material, leading to fragmented maternal care. EH was performed by removing the offspring from the maternal cage to a new cage for 15 min per day on P2 to P9, a procedure that has been shown to increase maternal caregiving behavior. A group with an unmanipulated environment (i.e., animal facility reared mice) was explicitly omitted to test only two opposing rearing environments. At adulthood (12 weeks of age), animals of each group were then either housed with an ovariectomized female (OX; supportive social environment) or underwent chronic social defeat stress (CD; aversive social environment) for three consecutive weeks giving rise to four experimental groups exposed to different qualities of social experience (EHOX, EHCD, LMOX, and LMCD). From each treatment group, eight mouse brains were analyzed. Twelve different brain regions including AMY, bed nucleus of the stria terminalis, cerebellum (CER), dorsal hippocampal Cornu Ammonis (CA) 1 region (dCA1), dorsal hippocampal CA3 region (dCA3), dorsal dentate gyrus (dDG), prefrontal cortex, nucleus accumbens (Nac), paraventricular nucleus (PVN), ventral hippocampal CA1 region (vCA1), ventral hippocampal CA3 region (vCA3), and ventral dentate gyrus were collected.

first: Elisabeth

last: Binder


title: Expression data from undifferentiated and iPS cells differentiated to a myogenic fate [hPAX7]

city: Boston

first: Olivier

last: Pourquie

state: MASSACHUSETTS

address: 60 Fenwood Road

department: Department of Pathology

institute: Brigham and Women’s Hospital

summary:
Here, we report the generation of human induced Pluripotent Stem (iPS) cell reporter line in which a venus fluorescent protein have been introduced into the PAX7 locus. We use microarrays to compare the transcriptome of PAX7-venus+ cells after 3 weeks of myogenic differentiation to that of undifferentiated iPS
Satellite cells (SC) are muscle stem cells which can regenerate adult muscles upon injury. Most SC originate from PAX7-positive myogenic precursors set aside during development. While myogenesis has been studied in mouse and chicken embryos, little is known about human muscle development.  Here, we use microarrays to compare the transcriptome of myogenic cells differentiated in vitro from human and mouse ES and iPS cells reporter cell lines. We generated fluorescent reporter lines in which a fluorescent protein was introduced into the loci coding for Pax7 (mouse ES and human iPS) or MYOG (human iPS). Mouse ES and human iPS cells were differentiated to the myogenic lineage in vitro for three weeks according to the protocols described in Chal et al, Nature Biotech 2015 and to Chal, Al Tanoury et al, Nature Protocols, 2016. Fluorescent Pax7 or MYOG-positive cells were FACS-sorted after 3-weeks of differentiation in vitro and we used Affymetrix microarrays to analyze the transcriptome of the purified mouse and human cell populations and compare it to the transcriptome of undifferentiated mouse ES or human iPS cells.

overall_design: iPS lines were differentiated according to previously described protocols (Chal et al., 2016; Chal et al., 2015). Venus+ cells were purified by FACS from 3-weeks old differentiated cultures and samples were compared to undifferentiated pluripotent iPS cells

first: Ziad

last: Tanoury


title: Temporal analysis of erythropoiesis

city: Ottawa

first: Marjorie

last: Brand

state: Ontario

address: 501 Smyth Road

country: Canada

institute: Ottawa Hospital Research Institute

summary: CD34+ cells were subjected to ex vivo erythropoiesis according to the protocol described in Palii et al. (doi: 10.3791/2813. J Vis Exp. 53, 2011). Samples were harvested at different time points and analyzed by RNAseq.

overall_design: CD34+ cells were subjected to ex vivo erythropoiesis according to the protocol described in Palii et al. (doi: 10.3791/2813. J Vis Exp. 53, 2011). Samples were harvested at different time points (Day0, Day2, Day4, Day6, Day7.5, Day8, Day8.5, Day10, Day10.5, Day11, Day11.5, Day12, Day14 and Day16)  and analyzed by RNAseq.

first: Ted

last: Perkins


title: Transcriptome analysis of pluripotent stem cells and fibroblasts.

city: Cheonan-si, Chungcheongnam-do

first: Man Ryul

last: Lee

address: 25 Bongjeong-ro

country: South Korea

institute: Soonchunhyang Institute of medi-bio science (SIMS)

summary:
Purpose:  The purpose of this data is to compare the transcriptome of embryonic stem cell (ESCs), Induced pluripotent stem cell (iPSCs) and fibroblasts using RNA-seq.
Methods: Total RNA was harvested by Trizol (Invitrogen) and RNA profiles of samples were generated by sequencing. Illumina Casava 1.8 software used for basecalling. Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence (using fastx_trimmer) then mapped to hg19 (genome build) whole genome using Tophat. This study provides a framework for the application of pluripotent stem cells RNA expression dynamics compare to fibroblasts profiling toward characterization.

overall_design: Total RNA extracted from 3 human cell line samples representing hESCs, hiPSCs and fibroblasts.

first: Tae

middle: W

last: Ha

first: Man


title: Expression data from undifferentiated and iPS/ES cells differentiated to a myogenic fate

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

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Refer to individual Series


title: RNA sequencing of the Hordeum vulgare near-isogenic line mnd1.a

city: Düsseldorf

first: Agatha

middle: Alexandra

last: Walla

address: Universitätsstraße 1

department: Institut of Plant Genetics

institute: Heinrich-Heine-University

summary: RNA sequencing of the near-isogenic line mnd1.a in cv. Bowman background, compared to cv. Mesa and cv. Bowman  Purpose: Expression analysis and variant calling.

overall_design: Enriched inflorescence (inflorescence including inflorescence base and small leaves below) tissue of mnd1.a and cv. Bowman plants was isolated at Waddington stage (WD) 2.0, 3.5 and 5.0. Enriched shoot apical meristem tissue at W1.0 was used for cv. Mesa. The samples were harvested at the middle of the day, 6 to 8 h before the end of the light period.

first: Wilma

last: van Esse

first: Maria

last: vonKorff


title: A Chromatin Accessibility Atlas of the Developing Human Telencephalon

city: San Francisco

first: John

last: Rubenstein

state: CA

address: 401 Parnassus Ave

department: Department of Psychiatry

institute: University of California, San Francisco

summary: Gene expression differs between cell types and regions within complex tissues such as the developing brain. To discover regulatory elements underlying this specificity, we generated genome-wide maps of chromatin accessibility in nine anatomically-defined regions of the developing human telencephalon.  Additionally, we defined the histone modification landscape of the prefrontal cortex and generated chromatin accessibility maps of its upper and deep layers. We predicted a subset of open chromatin regions (18%) that are most likely to be active enhancers, many of which are dynamic with 26% differing between early and late mid-gestation and 28% present in only one brain region. These predicted regulatory elements (pREs) are enriched proximal to genes with expression differences across developmental stages, regions, and cortical laminae; they harbor distinct sequence motifs that suggest potential upstream regulators. We leveraged this atlas to predict and validate novel regulatory elements of genes that control cortex laminar identity and genes associated with autism spectrum disorder (ASD).  These include enhancers proximal to FEZF2 and BCL11A that were validated in mouse, and an enhancer of ASD gene SLC6A1 containing two functional de novo mutations in individuals with ASD whose enhancer function we validated via CRISPRa. These applications demonstrate the utility of this atlas for decoding neurodevelopmental gene regulation in health and disease.

overall_design:
We performed ATAC-seq on nuclei isolated from fresh samples of human fetal hemispheres. Nine separate brain regions were dissected.   In prefrontal cortex, for some samples we microdissected upper and deep layers of the cortical plate.  We performed ChIP-seq on native chromatin isolated from nuclei from the prefrontal cortex using antibodies for different histone modifications.
Submitter declares that the human raw data will be deposited in dbGaP due to patient privacy concerns.

first: Eirene

last: Markenscoff-Papadimitriou

first: Sean

last: Whalen

first: Katherine

last: Pollard


title: Clonal Tracking Uncovers Barriers and Validate New Strategies to Enhance Gene Editing in Human Hematopoietic Stem Cells

city: Segrate

first: Ivan

last: Merelli

state: Milano

address: via F.lli Cervi 93

department: Istituto di Tecnologie Biomediche

institute: Consiglio Nazionale delle Ricerche


title: The NSD2 p.E1099K Mutation is Enriched at Relapse and Confers Drug Resistance in a Cell Context Dependent Manner in Pediatric Acute Lymphoblastic Leukemia (RNA-Seq)

first: Joanna

last: Pierro

overall_design: A total of 5 B-ALL cell lines were sequenced by RNAseq, two of which were engineered to overexpress WT or EK NSD2 (Reh, 697) and three others that were engineered to knock down expression of NSD2 using shRNAs (RS4;11, RCH-ACV and SEM). For Reh and 697, there are two and three replicates each, respectively, of each of three cell lines - EV (empty vector control), WT (overexpressed WT) and EK (overexpressed mutant). For the knockdown cell lines, there are three replicates each of each engineered cell line - NT (non-targeted control) and sh2 (NSD2 knockdown).

first: Jason

last: Saliba

first: Sonali

last: Narang

first: Gunjan

last: Sethia

first: Shella

last: Fleur-Lominy

first: Ashfiyah

last: Chowdhury

first: Anita

last: Qualls

first: Hannah

last: Fay

first: Harrison

last: Kilberg

first: Takaya

last: Moriyama

first: Tori

last: Fuller

first: David

last: Teachey

first: Kjeld

last: Schmiegelow

first: Jun

last: Yang

first: Mignon

last: Loh

first: Patrick

last: Brown

first: Jinghui

last: Zhang

first: Xiaotu

last: Ma

first: Aristotelis

last: Tsirigos

first: Nikki

last: Evensen

first: William

last: Carroll


title: Cohesin has a deleterious effect on heterochromatin assembly in fission yeast cells lacking Pds5

city: Bethesda

first: Shiv

last: Grewal

state: MD

address: NCI bldg 37 Rm 6068 9000 Rockville Pike

department: LBMB

institute: NCI

type: Genome binding/occupancy profiling by genome tiling array

summary:
Occupancy profiling of lysine 9 dimethylated histone H3 in fission yeast.
Occupancy profiling of Pds5 proteins in fission yeast.

overall_design: Agilent 60mer array was used to analyze DNA recovered by immunoprecipitation of lysine 9 dimethylated histone H3 from asynchronous culture of fission yeast.

first: H

last: Folco

first: Andrea

last: McCue

first: Vanivilasini

last: Balachandran

middle: I


title: The NSD2 p.E1099K Mutation is Enriched at Relapse and Confers Drug Resistance in a Cell Context Dependent Manner in Pediatric Acute Lymphoblastic Leukemia

first: Joanna

last: Pierro


title: ATACseq measurements during cord blood-derived human erythropoiesis

first: Marjorie

last: Brand

summary: CD34+ cells were subjected to ex vivo erythropoiesis according to the protocol described in Palii et al. (doi: 10.3791/2813. J Vis Exp. 53, 2011). Samples were harvested at different time points and analyzed by ATACseq.

overall_design: ATAC-seq on CD34+ cells subjected to ex vivo erythropoiesis.

first: Jim

last: Hughes


title: Clonal Tracking Uncovers Barriers and Validate New Strategies to Enhance Gene Editing in Human Hematopoietic Stem Cells [RNA-seq]

first: Ivan

last: Merelli

summary: By applying a barcoding strategy to clonal tracking of edited cells (BAR-seq), we show that p53 activation triggered by HDR editing significantly shrink the HSPC clonal repertoire in hematochimeric mice, despite engrafted edited clones preserved multilineage and self-renewing capacity. Transient inhibition of p53 restored polyclonal graft composition. We then overcame HSC constraints to HDR by forcing cell cycle progression and upregulating components of the HDR machinery through transient expression of Adenovirus 5 E4orf6/7 protein, which operates the major cell cycle controller E2F. Combined E4orf6/7 expression and p53 inhibition resulted in high and stable HDR editing efficiencies in the human graft, without perturbing repopulation and self-renewing properties of edited HSCs.

overall_design: Examination of the transcriptional profile in HSPCs treated with 5 different experimental conditions or untreated (UT). The experiment was performed in 3 replicates. Three-days cultured HSPCs were treated as follows: electroporation without adding any further component beside electroporation solution (UT_electro); electroporated with Cas9 RNP targeting the AAVS1 locus and transduced with the cognate AAV6 for HDR (RNP + AAV6); edited as in RNP + AA6 adding to the electroporation solution an mRNA encoding for the p53 inhibitor GSE56 (GSE56); edited as in RNP + AA6 adding to the electroporation solution an mRNA encoding for Ad5-E4orf6/7 (Ad5-E4orf6/7); edited as in RNP + AA6 adding to the electroporation solution an mRNA encoding for GSE56 and Ad5-E4orf6/7 (GSE56/Ad5-E4orf6/7). Transcriptomic analysis was performed 12 hours after treatments.

first: Samuele

last: Ferrari

first: Aurelien

last: Jacob

first: Stefano

last: Beretta

first: Pietro

last: Genovese

first: Luigi

last: Naldini


title: Expression data from undifferentiated and embryonic stem (ES) cells differentiated to a myogenic fate [mPAX7]

status: Public on Apr 21 2020

first: Olivier

last: Pourquie

summary:
Here, we use microarrays to compare the transcriptome of mouse Pax7-GFP ES reporter cell line after 3 weeks of myogenic differentiation in vitro to that of undifferentiated ES
Satellite cells (SC) are muscle stem cells which can regenerate adult muscles upon injury. Most SC originate from PAX7-positive myogenic precursors set aside during development. While myogenesis has been studied in mouse and chicken embryos, little is known about human muscle development.  Here, we use microarrays to compare the transcriptome of myogenic cells differentiated in vitro from human and mouse ES and iPS cells reporter cell lines. We generated fluorescent reporter lines in which a fluorescent protein was introduced into the loci coding for Pax7 (mouse ES and human iPS) or MYOG (human iPS). Mouse ES and human iPS cells were differentiated to the myogenic lineage in vitro for three weeks according to the protocols described in Chal et al, Nature Biotech 2015 and to Chal, Al Tanoury et al, Nature Protocols, 2016. Fluorescent Pax7 or MYOG-positive cells were FACS-sorted after 3-weeks of differentiation in vitro and we used Affymetrix microarrays to analyze the transcriptome of the purified mouse and human cell populations and compare it to the transcriptome of undifferentiated mouse ES or human iPS cells.

overall_design: Mouse ES cell lines were differentiated according to previously described protocols (Chal et al., 2015). GFP+ cells were purified by FACS from 3-weeks old differentiated cultures and samples were compared to undifferentiated Embryonic Stem (ES) cells

first: Ziad

last: Tanoury


title: ChIP-seq analysis of H3K27ac, PPARG and RXRA in human leukemia cell lines

city: Memphis

first: Jian

last: Xu

state: Tennessee

address: 262 Danny Thomas Place, MS 345

department: Pathology

institute: St. Jude Children's Research Hospital

summary: Mutations in protein-coding genes are well established as the basis for human cancer, yet it remains elusive how alterations within non-coding genome, a substantial fraction of which contain cis-regulatory elements (CREs), contribute to cancer pathophysiology. Here we developed an integrative approach to systematically identify and characterize non-coding regulatory variants with functional consequences in human hematopoietic malignancies. Combining targeted resequencing of hematopoietic lineage-specific CREs and mutation discovery, and uncovered 1,837 recurrently mutated CREs containing leukemia-associated non-coding variants. By enhanced CRISPR/dCas9-based CRE perturbation screening and functional analyses, we identified 218 variant-associated oncogenic or tumor suppressive CREs in human leukemia. Non-coding variants at KRAS and PER2 enhancers reside in nuclear receptor (NR) binding regions and modulate transcriptional activities in response to NR signaling in situ in leukemia cells. NR binding sites frequently co-localize with non-coding variants across cancer types. Hence, recurrent non-coding variants connect enhancer dysregulation with nuclear receptor signaling in hematopoietic malignancies.

overall_design: ChIP-seq was performed to determine the chromatin occupancy of H3K27ac, PPARG and RXRA in human leukemia cell lines

first: Kailong

last: Li

first: Yuannyu

last: Zhang

first: Xin

last: Liu

first: Yuxuan


title: Expression data from undifferentiated and iPS cells differentiated to a myogenic fate [MYOG]

summary:
Here, we report the generation of human induced Pluripotent Stem (iPS) cell reporter line in which a venus fluorescent protein have been introduced into the MYOGENIN (MYOG) locus. We use microarrays to compare the transcriptome of MYOG-venus+ cells after 3 weeks of myogenic differentiation to that of undifferentiated iPS
Satellite cells (SC) are muscle stem cells which can regenerate adult muscles upon injury. Most SC originate from PAX7-positive myogenic precursors set aside during development. While myogenesis has been studied in mouse and chicken embryos, little is known about human muscle development.  Here, we use microarrays to compare the transcriptome of myogenic cells differentiated in vitro from human and mouse ES and iPS cells reporter cell lines. We generated fluorescent reporter lines in which a fluorescent protein was introduced into the loci coding for Pax7 (mouse ES and human iPS) or MYOG (human iPS). Mouse ES and human iPS cells were differentiated to the myogenic lineage in vitro for three weeks according to the protocols described in Chal et al, Nature Biotech 2015 and to Chal, Al Tanoury et al, Nature Protocols, 2016. Fluorescent Pax7 or MYOG-positive cells were FACS-sorted after 3-weeks of differentiation in vitro and we used Affymetrix microarrays to analyze the transcriptome of the purified mouse and human cell populations and compare it to the transcriptome of undifferentiated mouse ES or human iPS cells.


title: microRNA profiling of mouse cortical progenitors and neurons reveals miR-486-5p as a regulator of neurogenesis

city: Modena

first: Martina

last: Dori

address: Via G. Campi 287

department: Department of Life Sciences

institute: Center for Genome Research

type: Non-coding RNA profiling by high throughput sequencing

summary: MicroRNAs (miRNAs) are short (~22 nt) single-stranded non-coding RNAs that regulate gene expression at the post-transcriptional level. Over the past years, many studies have extensively characterized the involvement of miRNA-mediated regulation in neurogenesis and brain development. However, a comprehensive catalog of cortical miRNAs cell-specifically expressed in progenitor types of the developing mammalian cortex is still missing. Overcoming this limitation, here we exploited a double reporter mouse line previously validated by our group to allow the identification of the transcriptional signature to neurogenic commitment and provide the field with the complete atlas of miRNAs expression in proliferating neural stem cells, neurogenic progenitors and newborn neurons during corticogenesis. By extending the currently known list of miRNAs expressed in the mouse brain by over two fold, our study highlights the power of cell type-specific analyses for the detection of transcripts that would otherwise be diluted out when studying bulk tissues. We further exploited our data by predicting putative novel miRNAs and validated the power of our approach by providing novel evidence for the involvement of miR-486 as a novel player in brain development.

overall_design: micro RNA profiles in different cell population (proliferating and differentiating progenitors and newborn neurons) of E14.5 developing mouse cortex. Profiles were obtained by deep-sequencing after small RNA enrichment, using Illumina HiSeq 2500

first: Daniel

last: Cavalli

first: Mathias

last: Lesche

first: Simone

last: Massalini

first: Leila

last: Haj Abdullah Alieh

first: Beatriz

last: Cardoso de Toledo

first: Sharof

last: Khudayberdiev

first: Gerhard

last: Schratt

first: Andreas

last: Dahl

first: Federico

last: Calegari


title: RNA-seq of primary human aortic smooth muscle cells treated with AEA

city: bad nauheim

first: Stefan

last: Günther

address: ludwigtr. 43

institute: MPI for heart and lung research

summary: Primary human aortic smooth muscle cells (HAoSMC) were starved over night in 1% FCS. On the next day cells were treated with 100nM Anandamide (AEA) or EtOH as control for 4h in presence of 10µM diclofenac. Subsequently cells were treated with or without 10ng/mL IL-1β for last 1.5h.

overall_design: RNA Seq analysis of human aortic smooth muscle cells after treatment with the endocannabinoid anandamide (AEA) with or without IL-1β.

first: Beatrice

last: Pflüger-Müller

first: Matthias

last: Leisegang

first: Ralf

last: Brandes


title: m6A-Label-seq directly reports m6A methylome at base resolution

city: Hangzhou

first: Jie

last: Cao

address: Zheda Road 38

department: Department of Polymer Science and Engineering

institute: Zhejiang University

type: Methylation profiling by high throughput sequencing

summary: Here we report a metabolic labeling method to map mRNA N6-methyladenosine (m6A) modification transcriptome-wide at base resolution, termed m6A-label-seq. The cells were fed with Se-allyl-L-selenohomocysteine, an analog of methoine, which serves as the precursor of methylation enzyme cofactor, so that cellular RNAs were continuously deposited with N6-allyladenosine (a6A) at supposed m6A sites. We enriched a6A-containing mRNAs and sequenced their a6A sites which are identical to m6A sites, based on iodination-induced misincorporation during reverse transcription.

overall_design:
We profiled m6A sites at base resolution in human HeLa and HEK293T cells and mouse H2.35 cells by metabolically incorporation of Se-derived methionine analog.
We assessed the potential stress response in HeLa cells by an RNA-seq experiment.


title: Clonal Tracking Uncovers Barriers and Validate New Strategies to Enhance Gene Editing in Human Hematopoietic Stem Cells [BAR-seq]

first: Ivan

last: Merelli

overall_design: We performed clonal tracking of HDR-edited HSPCs by a barcoding-based strategy in xenotransplanted NSG mice. We performed deep-sequencing of the on-target barcoded region in bulk and human sorted hematopoietic subpopulations. Transplanted HSPCs were edited in the AAVS1 locus using the barcoded AAV6 vector in presence or absence of GSE56 and/or Ad5-E4orf6/7.

first: Samuele

last: Ferrari

first: Aurelien

last: Jacob

first: Stefano

last: Beretta

first: Pietro

last: Genovese

first: Luigi

last: Naldini


title: Cell-of-Origin Subtyping Diffuse Large B-Cell Lymphoma (DLCBL) by Using a 32-Gene Expression Assay in Formalin-Fixed Paraffin-Embedded Tissues

city: Shanghai

first: Qinghhua

last: Xu

address: Cao'An Road 4800

institute: Tongji University

summary:
Objection: To evaluate a qPCR-based 32-gene expression assay to determine the cell-of-origin (COO) of diffuse large B-cell lymphoma (DLBCL) with formalin-fixed paraffin-embedded (FFPE) tissue.
Methods: The most established subtyping algorithm, the COO model, categorizes DLBCL into activated B-cell (ABC) and germinal center B-cell (GCB)-like subgroups through gene expression profiling. Biopsy of DLBCL patients with paired FFPE and fresh frozen tissue were collected to assign COO based on the immunohistochemistry (IHC) algorithm (Han’s algorithm), qPCR-based 32-gene expression assay (DLBCL-COO assay) and global gene expression profiling with RNA-seq.
Results: The DLBCL-COO assay demonstrated a significantly superior concordance of COO determination with the “gold standard” RNA-seq, comparing with the IHC assignment with Han’s algorithm (91.9% versus 77.5%; P = 0.005). Furthermore, overall survival of GCB patients defined by DLBCL-COO assay was superior significantly towards the ABC patients (Figure 2B, P = 0.023). This effect was not seen when tumors were classified by IHC algorithm.
Conclusions: The DLBCL-COO assay provides flexibility and accuracy in DLBCL subtype characterization. These subtype distinctions should help guiding prognostic and therapeutic options for the patients in our daily practice.

overall_design: RNA-seq results  of 111 DLBCL specimens were generated by paired-end sequencing on Illumina NovaSeq 6000 system

first: Wanhui

last: Yan

first: Xiangnan

last: Jiang

first: Weige

last: Wang

first: Yifeng

last: Sun

first: Yixin

last: Wo

first: Zhengzhi

last: Luo

first: Qinghua

first: Xiaoyan

last: Zhou

first: Junning

first: Xiaonan

last: Hong

first: Xiaoqiu

last: Li


title: Identifying microRNAs that bind to cytoplasmic TDP43

city: Grand Forks

first: Junguk

last: Hur

state: ND

address: 1301 N. Columbia Rd. Stop 9037

department: Biomedical Sciences

institute: University of North Dakota

summary: TDP43 is involved in microRNA biogenesis and found in cytoplasmic aggregates in amyotrophic lateral sclerosis (ALS), and microRNAs are important for regulation of gene expression and represent potential biomarkers and therapeutic targets. Therefore, we examined microRNAs that preferentially bind cytoplasmic TDP43 using cellular models expressing TDP43 variants and NanoString miRNA profiling analyses. We identified cytoplasmic TDP43-associated miRNAs and predicted genes and pathways to gain insights into potentially relevant disease pathways, biomarkers, and reversible therapeutic targets for ALS.

overall_design: Stable HeLa cell lines were established that expressed eGFP-His-tagged TDP43 variants (wild-type or ∆NLS) or an empty eGFP-His-tagged control construct. Cells were fractionated into nuclear and cytoplasmic fractions, cytoplasmic TDP43 was pulled down from the cytoplasmic fractions via its His tags, and microRNAs associated with cytoplasmic TDP43 were profiled by NanoString analysis.

first: Eva

last: Feldman


title: Chromatin accessibility and transcriptome changes in injured retinal ganglion cells (RGCs) after Arid1a deletion

status: Public on Apr 20 2020

city: Zibo

first: Dai

middle: Shang

last: Kun

state: Shandong

address: 266 Xincun West Road

department: School of Life Sciences and Medicine

institute: Shandong University of Technology

summary: To gain a deeper insight into roles of Arid1a in injured retinal ganglion cells (RGCs), we performed ATAC-seq and RNA-seq to analyze the genome-wide changes of chromatin accessibility and transcriptome changes before and after Arid1a deletion in RGCs of the adult mouse using Cre-lox system.

overall_design: Total RNA was extracted from WT and Arid1a-cKO RGCs and generated by deep sequencing, in three biological replicates, using Illumina HiSeq X Ten. ATAC-seq was sequenced in two biological replicates using RGCs.

last: Shang-Kun


title: Differential small RNA expression upon induction and establishment of Tuxpeño maize embryogenic callus cultures

city: MEXICO CITY

first: TZVETANKA

middle: DIMITROVA

last: DINKOVA

state: DISTRITO FEDERAL

address: CIRCUITO EXTERIOR, CIUDAD UNIVERSITARIA DELEGACION COYOACAN

department: BIOCHEMESTRY

country: Mexico

institute: UNIVERSIDAD NACIONAL AUTONOMA DE MEXICO, FACULTAD DE QUIMICA

summary:
Purpose: Maize somatic embryogenesis is usually required to achieve genetic transformation and represents an important alternative in plant development. Although many embryogenesis-related genes have been studied in this model, the molecular mechanisms underlying cell dedifferentiation and further plant regeneration are not completely understood.
Methods: Immature embryos smRNA profiles of 15-day-after-pollination (IE) and Embryogenic Callus from one (C1), four (C4), and ten months (C10) were generated by deep sequencing, using Illumina GAIIx. The sequence reads that passed quality filters were analyzed with two methods: Bowtie 1.1.2 and ShortStack 3.4. qRT–PCR validation for selected miRNAs was performed using SYBR Green assays.
Results: We used high throughput sequencing to explore the sRNA populations during maize embryogenic callus induction and established subcultures from the Mexican cultivar VS-535, Tuxpeño landrace. We detected readjustments in 24 nt and 21-22 nt sRNA populations during the embryogenic callus establishment and maintenance. miRNAs related to stress response substantially increased upon callus proliferation establishment, correlating with a reduction in some of their target levels. On the other hand, while 24 nt-long hc-siRNAs derived from transposable retroelements transiently decreased in abundance during the embryogenic callus establishment, a population of 22 nt- hc-siRNAs increased. This was accompanied by reduction in transposon expression in the established callus subcultures.
Conclusions: Stress- and development-related miRNAs are highly expressed upon maize EC callus induction and during maintenance subcultures, while miRNAs involved in hormone response only transiently increase during induction. The establishment of proliferative maize embryogenic callus is accompanied by important readjustments in the length of hc-siRNAs mapping to LTR retrotransposons, and their expression regulation.

first: Tzvetanka

last: Dimitrova

first: Naholi

last: Alejandri-Ramirez


title: Antibody-free enzyme-assisted chemical approach for detection of N6-methyladenosine

city: Los Angeles

first: Ye

state: California

address: 611 Charles E Young Dr E

department: Bioinformatics

institute: UCLA

summary: We applied chemical-assisted, m6A-SEAL method to profile m6A distributions in the transcriptomes of human and plant cells and compared its results against MeRIPm6A-seq.

sample_organism:
Oryza sativa
Homo sapiens

platform_organism:
Oryza sativa
Homo sapiens

overall_design: Examination for RNA methylation states on different species by chemical assisted method (m6A-SEAL) and antibody-based method (MeRIP-seq), then analyzed and compared the performance of those two methods.

first: Guifang

last: Jia


title: Genome-wide methyl-cytosine competition by DNMT and TET

city: Cambridge

first: Jocelyn

last: Charlton

state: MA

address: 7 Divinity Avenue

department: Stem Cell and Regenerative Biology

institute: Harvard University

summary: We define distinct methylated elements that recruit both DNMT3A/B and TETs. We also show the genome-wide activity of DNMTs and TETs.

overall_design: We performed WGBS for a series of HUES64 and HUES8 human embryonic stem cells: wildtype (WT), DNMT3A and DNMT3B double knockout (DKO), TET triple knockout (TKO), TKO plus DNMT3B knockout (QKO) and QKO plus DNMT3A knockout (PKO). We also performed bulk RNA sequencing.

first: Alexander

last: Meissner


title: Gain-of-function genetic alterations of G9a drive oncogenesis

city: Boston

first: David

middle: E.

last: Fisher

state: Massachusetts

address: 55 Fruit Street

department: Dermatology

institute: Massachusetts General Hospital

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

overall_design: Refer to individual Series


title: Gain-of-function genetic alterations of G9a drive oncogenesis II

summary: Epigenetic regulators, when genomically altered, may become driver oncogenes that mediate otherwise unexplained pro-oncogenic changes lacking a clear genetic stimulus, such as activation of the WNT/b-catenin pathway in melanoma. This study identifies previously unrecognized recurrent activating mutations in the G9a histone methyltransferase gene, as well as G9a genomic copy gains in ~26% of human melanomas, which collectively drive tumor growth and an immunologically sterile microenvironment beyond melanoma. Furthermore, the WNT pathway is identified as a key tumorigenic target of G9a gain-of-function, via suppression of the WNT antagonist DKK1. Importantly, genetic or pharmacologic suppression of mutated or amplified G9a using multiple in vitro and in vivo models demonstrate that G9a is a druggable target for therapeutic intervention in melanoma and other cancers harboring G9a genomic aberrations.

overall_design: Whole transcriptome RNA sequencing of total RNA extracted from G9a (EHMT2)-amplified Hs944T melanoma cells with and without shRNA knockdown of G9a

first: Shinichiro

last: Kato

first: Yotam

last: Drier


title: Modeling hypoxia induced neuropathies using a fast and scalable human motor neuron differentiation system

city: Aurora

first: Matthew

last: Taliaferro

state: CO

address: 12801 E 17th Ave, RC1 South Room 10403D

department: Biology

institute: University of Colorado Anschutz Medical Campus

summary: Human pluripotent stem cells carry great potential to provide novel insights into disease mechanisms by enabling generation and study of functional cell types under cell culture conditions, including human motor neurons (MNs). MN disease are a group of conditions where these nerves progressively loose function. Recent findings using animal model systems suggest a previously unappreciated role for mRNA mis-localization in disease development and progression. However, model systems to investigate this novel disease link in the human context are currently absent. Here we describe a scalable suspension-based differentiation system to generate large numbers of hPSCs derived neuronal progenitors that can further differentiate efficiently into functional stem cell derived human motor neurons (sMN) within only 3 weeks. To be able to investigate potential differences in mRNA transcripts between sMN cell soma and neurites we further established a membrane-based culture system that allows efficient fractionation of cell soma and neurites.

overall_design: Quantification of transcript abundance in soma and neurites of hypoxia and control human motor neurons

first: Russ

last: Holger

first: J

middle: M


title: Widespread targeting of nascent transcripts by RsmA in Pseudomonas aeruginosa

city: Iowa City

first: Michael

last: Gebhardt

state: IA

address: 431 Newton Road, 200A EMRB

department: Department of Microbiology and Microbiology

institute: University of Iowa

summary: In the opportunistic pathogen Pseudomonas aeruginosa RsmA is an RNA-binding protein that plays critical roles in the control of virulence, interbacterial interactions and biofilm formation. Although RsmA is thought to exert its regulatory effects by binding full-length transcripts, the extent to which RsmA binds nascent transcripts has not been addressed. Moreover, which transcripts are direct targets of this key post-transcriptional regulator is largely unknown. Using chromatin immunoprecipitation coupled with high-throughput DNA sequencing, with cells grown in the presence and absence of the RNA polymerase inhibitor rifampicin, we identify hundreds of nascent transcripts that RsmA associates with in P. aeruginosa. We also find that the RNA chaperone Hfq targets a subset of the RsmA-associated nascent transcripts and that the two RNA-binding proteins can exert regulatory effects on common targets. Our findings establish that RsmA associates with many transcripts as they are being synthesized in P. aeruginosa, identify the direct targets of RsmA, and suggest that RsmA and Hfq may act in a combinatorial fashion on certain target transcripts. More broadly, our data suggest that the binding of post-transcriptional regulators to nascent transcripts may be commonplace in bacteria where distinct regulators can function alone or in concert to achieve control over the translation of transcripts as soon as they emerge from RNA polymerase.

overall_design:
ChIP-seq: Genome-wide locations of RsmA (with and without rifampicin treatment) Hfq. Additionally, we assess the locations of RsmA in the absence of hfq and the locations of Hfq in the absence of rsmA & rsmF.
ChIP-seq: Please note that each bed file generated from all replicates is linked to the corresponding Sample*A records.

RNA-seq: RNA-sequencing of P. aerugionsa strain PAO1, PAO1 ∆rsmA ∆rsmF, and PAO ∆hfq

first: Tracy

middle: K

last: Kambara

first: Kathryn

last: Ramsey

first: Simon

middle: L

last: Dove


title: Gain-of-function genetic alterations of G9a drive oncogenesis I

overall_design: Whole transcriptome RNA sequencing of total RNA extracted from pMEL/hTERT/CDK4(R24C) cells expressing BRAF(V600E) and either G9a or GFP


title: Transcriptomic changes in response to modulation of long-non-coding RNA, LINC00473

status: Public on Apr 19 2020

city: Worcester

first: Silvia

last: Corvera

address: 373 Plantation St

institute: University of Massachusetts Medical School

summary: Human thermogenic adipose tissue mitigates metabolic disease, raising much interest in understanding its development and function. Here, we show that human thermogenic adipocytes specifically express a primate-specific long non-coding RNA, LINC00473 which is highly correlated with UCP1 expression and decreased in obesity and type-2 diabetes. LINC00473 is detected in progenitor cells, and increases upon differentiation and in response to cAMP. In contrast to other known adipocyte LincRNAs, LINC00473 shuttles out of the nucleus, colocalizes and can be crosslinked to mitochondrial and lipid droplet proteins. Up- or down- regulation of LINC00473 results in reciprocal alterations in lipolysis, respiration and transcription of genes associated with mitochondrial oxidative metabolism. Depletion of PLIN1 results in impaired cAMP-responsive LINC00473 expression and lipolysis, indicating bidirectional interactions between PLIN1, LINC00473 and mitochondrial oxidative functions. Thus, we suggest that LINC00473 is a key regulator of human thermogenic adipocyte function, and reveals a role for a LincRNA in inter-organelle communication and human energy metabolism.

overall_design: To fully understand the functional consequences of LINC00473-mediated modulation of mitochondrial respiration and lipolysis, we analyzed the transcriptome of adipocytes derived from human subcutaneous adipose derived progenitor cells overexpressing LINC00473, and that of cells in which LINC00473 expression was blunted with siRNA.

first: Khanh-Van

last: Tran

first: Erin

last: Brown

first: Tiffany

last: DeSouza

first: Naja

middle: Z

last: Jespersen

first: Cecilie

last: Nandrup-Bus

first: Qin

last: Yang

first: Zinger

first: Anand

last: Desai

first: SoYun

last: Min

first: Raziel

last: Rojas-Rodriguez

first: Morten

last: Lundh

first: Amir

last: Feizi

first: Hanni

last: Willenbrock

first: Therese

last: Larsen

first: Mai

middle: C

last: Severinsen

first: Kimberly

last: Malka

first: Anthony

last: Mozzicato

first: Atul

last: Deshmukh

first: Brice

last: Emanuelli

first: Bente

last: Pedersen

first: Timothy

last: Fitzgibbons

first: Camilla

last: Scheele

first: Søren

last: Nielsen


title: High CD26 and low CD94 expression identifies an IL-23 responsive V(delta)2+ T cell subset with a MAIT cell-like transcriptional profile

city: Melbourne

first: Hyon Xhi

last: Tan

state: Victoria

address: 792 Elizabeth St

department: Department of Microbiology and Immunology

country: Australia

institute: The University of Melbourne, at the Peter Doherty Institute for Infection and Immunity

summary: In this study, we find that CD26 and CD94 co-expression identifies transcriptionally distinct V(delta)2+ T cell subsets that vary in cytokine-responsiveness and cytotoxic potential, with a direct relationship between the loss of CD26 surface expression and upregulation of GzmB. These subsets differ in the expression of key transcriptional regulators including Tbet, Eomes and Hobit, and exist independently of classical memory markers. Analysis of cord blood samples confirms that CD26+ V(delta)2+ T cells predominate at birth, and we find evidence that IL-23 exposure and CD26 ligation may contribute to the generation of CD26- cytotoxic V(delta)2+ T cells in adults.

overall_design: Samples 1-12: Freshly isolated PBMC from 3 healthy donors were stained for CD3, V(delta)2 TCR, CD26 and CD94. Cells were identified as Live/Dead negative, CD3+ V(delta)2+ cells. Four populations (CD26+CD94lo, CD26+CD94hi, CD26-CD94hi, CD26-CD94lo) were sorted from these PBMCs. Transcriptional profiling and differential gene expression was performed using RNA sequencing.    Samples 13-28: For the V(delta)2 expansion RNA sequencing, individual CD26/CD94 V(delta)2 subsets were sorted from a single blood draw and expanded in parallel with (gamma-delta)-depleted PBMC, zoledronate, IL-2 and (if applicable) IL-23. At day 14 post-stimulation, cells were surface stained and live, CD3+ V(delta)2+ cells were sorted. Transcriptional profiling and differential gene expression was performed using RNA sequencing.

first: Kathleen

last: Wragg

first: Hyon-Xhi

first: Anne

middle: B

last: Kristensen

first: Catriona

middle: V

last: Nguyen-Robertson

middle: D

last: Kelleher

first: Matthew

middle: S

last: Parsons

first: Adam

last: Wheatley

first: Stuart

last: Berzins

first: Daniel

middle: G

last: Pellicci

first: Stephen

last: Kent

first: Jennifer

middle: A

last: Juno


title: Zeb2 regulates the output capacity of the neurogenic V-SVZ in the mouse forebrain.

first: Ruben

last: Dries

address: 360 Longwood Ave

department: Computational Biology

institute: DFCI

summary: Using RNA sequencing, this study aims to identify (1) downstream targets and (2) cellular processes depending on the transcription factor Zeb2 in the young postnatal neurogenic V-SVZ niche of the mouse brain

overall_design: RNA sequencing of FACS sorted (GFP+)  Gsh2-Cre; Zeb2fl/ko;RCE versus Gsh2-Cre; Zeb2fl/wt;RCE V-SVZ cells obtained from P2 mouse brains

first: Elke

last: Stappers

first: Eve

last: Seuntjens

first: Danny

last: Huylebroeck


title: miR396e-OsGRF7 module controls rice plant architecture (ChIP-Seq)

city: Wuhan

first: Yunping

last: Chen

state: Hubei

address: No. 299, Ba Yi Road, Wuchang

institute: Wuhan university

summary: Plant-specific growth-regulating factors (GRFs) participate in multiple central developmental processes including root and leaf development, flower and seed formation, plant senescence, and tolerance to stress. While the role of the miRNA-GRFs regulatory module in determining gross morphology, which is one of the most important agronomic traits for crops, have not been comprehensively unraveled yet. Here, we reported that OsGRF7, a target of miR396e and co-activated with OsGIFs, is essential for determining plant architecture in rice. Overexpression of OsGRF7 leads to decreased tiller number, leaf length and leaf angle, reduced plant height and increased grain size, which are mediated by shortened cell length and disordered cell arrangement. Further analyses indicate that OsGRF7 binds the ACRGDA motif in promoters of OsNSP2, OsGASR1 and OsCYP714B1, OsCga1 and OsARF12, which are involved in the synthesis of strigolactones, gibberellins and cytokinins or related to auxin signaling pathway. Our findings establish OsGRF7 as a crucial component in the miR396-OsGRFs/OsGIFs-plant hormone regulatory network that controls rice growth and plant architecture.This dataset records the profile of the binding peaks of OsGRF7 with GFP antibody in 35S:GRF7-GFP overexpression lines.

overall_design: Examination of OsGRF7 regulated genes.

first: Zhiwu

last: Dan


title: miR396e-OsGRF7 module controls rice plant architecture


title: Expression data from wild-type bone marrow-derived macrophages non-treated (NT1 and NT2) or stimulated for 12 h with VSV virus(VSV1 and VSV2)

status: Public on Apr 18 2020

first: zhu

last: Qingchen

address: 320 Yue Yang Rd, Xuhui District

institute: Shanghai Institute of Nutrition and Health, Chinese Academy of Sciences

summary: We conducted gene expression profile analysis by using wild-type bone marrow-derived macrophages. Heatmap revealed the most prominent different expression of genes encoding E3 ligases or DUBs altered in macrophages upon VSV stimulation were associated with typeⅠIFN genes.

overall_design: Genome-wide expression profiling of WT bone marrow-derived macrophages non-treated (NT1 and NT2) or stimulated for 12 h with VSV virus（VSV1 and VSV2) were generated by RNA-sequencing analysis

first: Yichuan

last: Xiao

first: Qingchen

last: Zhu


title: Identification of a Cxcl10+ monocyte subset with a pathogenic signature in the CNS during auto-immune neuroinflammation

city: Rehovot

first: Ido

last: Amit

address: 234 Herzl st.

department: Immunology

country: Israel

institute: Weizmann Institute of Science

summary: Multiple Sclerosis (MS) is characterized by pathological inflammation resulting from recruitment of lymphoid and myeloid immune cells from the blood circulation into the central nervous system (CNS). Due to cellular heterogeneity, defining the functional roles of these subsets in acute and chronic stages of MS has been challenging. Here we used index sorting and transcriptional single-cell sequencing to characterize peripheral mononuclear phagocyte infiltrates in the MS mouse model, experimental autoimmune encephalomyelitis (EAE). Based on their transcriptomes, we identified eight monocyte and three dendritic cell subsets during disease pathology with defined characteristics pointing towards distinct functions. Cell ablation identified two specific monocytic subsets with a pathogenic potential. Congenic monocyte transfer experiments combined with indexed-lineage sorting coupled to scRNA-seq established that these pathogenic cells are not descendants of the canonical Ly6C+ monocytes but derived from early myeloid cell progenitors. These results suggest a potential for targeted therapeutic interventions aimed at blocking specific pathogenic monocytic subsets.

overall_design: Transcriptional profiling of myeloid cells (monocytes, macrophages and dendritic cells) from murine spinal cords, 16 days (acute phase) and 30 days (chronic phase) post induced Experimental Autoimmune Encephalomyelitis (EAE) immunization.

first: Amir

last: Giladi

first: Lisa

last: Wagner

first: Hanjie

last: Li

first: Dorothea

last: Dörr

first: Chiara

last: Medaglia

first: Franziska

last: Paul

first: Anat

last: Shemer

first: Steffen

last: Jung

first: Simon

last: Yona

first: Matthias

last: Mack

first: Achim

last: Leutz

first: Alexander

last: Mildner


title: Single-cell RNA-sequencing of CD4+ T cells from lung and draining lymph node of flu-infected mice

status: Public on Apr 17 2020

city: Basel

first: Carolyn

last: King

address: Hebelstrasse 28

department: Department of Biomedicine

country: Switzerland

institute: University of Basel

summary: We performed a single-cell transcriptome analysis of pooled samples from lung and lymph node of 4-12 mice per time point in days post infection (d9, d14, d30) sorted for I-Ab NP311-325 specific CD4 T cells.  Additionally we include mRNA counts from liver cells from LCMV infected mice day 37 post-infection, which were  sequenced simultaneously with spleen cells from GSE134157.

overall_design: Cells negative for dump and positive for CD4, CD44 and I-Ab NP311-325 were sorted for lung and lymph node at each time point and used for single cell RNA Seq (10x genomics platform). Liver cells were negative for dump and positive for CD4, CD44 and GP66.

first: Nivedya

last: Swarnalekha

first: David

last: Schreiner

first: Julien

last: Roux


title: Epigenetic memory marks determine epiallele stability at loci targeted by de novo DNA methylation

city: West Lafayette

first: Jian-Kang

last: Zhu

state: IN

address: 625 Agriculture Mall Dr.

department: Department of Horticulture and Landscape Architecture

institute: Purdue University

summary: The transgenerational stability of DNA methylation changes is important in setting up genomic DNA methylation patterns and in the formation and transmission of epialleles.  It is generally assumed that DNA methylation changes at genomic regions targeted by the de novo, RNA-directed DNA methylation (RdDM) pathway are unstable. Here, we show that RdDM targets in Arabidopsis can be classified into two groups based on their transgenerational epiallele stability following restoration of NRPD1 function in nrpd1 mutant plants: remethylable loci and non-remethylable loci. Compared to the remethylable loci, non-remethylable ones contain higher levels of the euchromatic marks H3K4me3 and H3K18ac, which interferes with the recruitment of the RdDM molecular machinery and helps to recruit the DNA demethylase ROS1 to antagonize RdDM, respectively. Using targeted de-methylation by CRISPR/dCas9-TET1, we demonstrate that mCG and mCHG are memory marks required for targeting the RdDM machinery to remethylable loci. Our results show that histone and DNA methylation marks are critical in determining the capacity of RdDM target loci to form stable epialleles, and contribute to understanding the formation and transmission of epialleles.

overall_design: MethylC-Seq: 12 samples examined, Col-0, nrpd1-3, nrpe1-11, rdr2-2, two NRPD1 complementation lines,two NRPE1 complementation lines, two RDR2 complementation lines, nrpd1-3xCol-0 F1 and reciprocal F1 hybrids. Small RNA-Seq: 4 samples examined, Col-0, nrpd1-3 and two NRPD1 complementation lines

first: Jingwen

first: Huan

last: Huang


title: Hippo signaling coordinates cellular quiescence with terminal maturation in iNKT cell development and fate decisions [ClariomS]

city: Memphis

first: Hongbo

last: Chi

state: TN

address: 262 Danny Thomas Place

institute: St Jude Children's Research Hospital

summary: Hippo kinases Mst1 and Mst2 act as molecular rheostats for the terminal maturation and effector differentiation programs of iNKT cells. Loss of Mst1 alone or with Mst2 impedes iNKT cell development, associated  with defective IL-15-dependent cell survival

overall_design: Microarray analysis of 6 Samples of WT iNKT cells from stage 1 and stage 2; 4 samples of Mst1 single KO from stage 1 and stage 2; 7 samples of WT iNKT from stage 3; 5 Samples of Mst1KO iNKT from stage 3; 2 samples of Mst1/2 double KO iNKT from stage 1 and stage 2; 1 sample of Mst1/2 double KO from stage 3 (total n = 25)

first: Jana

last: Raynor

first: Yogesh

last: Dhungana


title: High-throughput sequencing analysis of a “hit and run” cell and animal model of KSHV tumorigenesis. [MBD-Seq]

status: Public on Apr 16 2020

city: La Plata

first: Martin

middle: Carlos

last: Abba

state: Buenos Aires

country: Argentina

institute: School of Medical Sciences - UNLP

summary: Kaposi's sarcoma (KS), is an AIDS-associated neoplasm caused by the KS herpesvirus (KSHV/ HHV-8). KSHV-sarcomagenesis is the consequence of oncogenic viral gene expression as well as host genetic and epigenetic alterations. Although KSHV is invariably found in all KS lesions, the percentage of KSHV-infected (LANA+) spindle-cells of the lesion is variable, suggesting the existence of paracrine oncogenic mechanisms and/or spindle cells that have lost KSHV. A mouse model of KSHVBac36-driven tumorigenesis allowed us to induce KSHV-episome loss before and after tumor development. Although cells that lose the KSHV episome prior to tumor formation lose their tumorigenicity, explanted KSHV positive tumor cells that lost the KSHV episome remained tumorigenic. This pointed to the existence of virally-induced irreversible oncogenic alterations occurring during KSHV-tumor formation that support the possibility of hit and run viral sarcomagenesis. RNA sequencing and CpG-methylation analysis were performed on KSHV positive and negative cells, KSHV positive and KSHV negative tumors that developed following KSHV-episome loss from explanted tumor cells. When KSHV positive cells form KSHV-driven tumors, along with viral-gene upregulation there is a tendency for hypo-methylation in genes from oncogenic and differentiation pathways. In contrast, KSHV-negative tumors formed after KSHV-episome loss, show a tendency towards gene hyper-methylation when compared to KSHV-positive tumors. Regarding occurrence of host-mutations, we found the same set of innate-immunity related mutations undetected in KSHV-infected cells but present in all KSHV positive tumors, indicating that pre-existing host mutations that provide an in vivo growth advantage are clonally-selected and contribute to KSHV-tumorigenesis. In addition, KSHV negative tumors display de novo mutations related to cell proliferation that, together with the PDGFRAD842V, were responsible for driving tumorigenesis in absence of the KSHV-episomes. KSHV-induced irreversible genetic and epigenetic oncogenic alterations support the possibility of “hit and run” KSHV-sarcomagenesis consistent with the presence of LANA-negative spindle-cells in KS lesions.

overall_design: KSHV-infected KS lesions are composed of latently-infected cells, as well as cells expressing lytic genes that have been implicated in the development of the KS angioproliferative phenotype. The existence of KS lesions with varying levels of KSHV-infected cells suggests also the existence of virus-independent “hit and run” mechanisms of sarcomagenesis, whereby viral infection irreversibly induce genetic or epigenetic oncogenic alterations in host cells. We used the unique mECK36 animal model of multistep KSHV sarcomagenesis to dissect transcriptional, genetic and epigenetic mechanisms of KSHV dependent tumorigenesis and during tumorigenesis following KSHV-episome loss (“hit and run”) sarcomagenesis in an unbiased high-throughput fashion. These analyses revealed that KSHV in vivo tumorigenesis: A) Occurs predominantly with CpG hypo-methylation of oncogenic and differentiation pathways. B) Selects for pre-existing host mutations that allow the KSHV oncovirus to express the oncogenic lytic program by creating permissive environment for viral-induced innate immunity and inflammation, which provides a selective advantage in vivo conducive to tumorigenesis.

first: E

last: Mesri

first: J

last: Naipauer

first: M

last: Shamay

first: G

last: Journo


title: Development of gene expression signatures for normal and tumor-associated cervix fibroblasts

city: Budapest

first: Alexandra

last: Fullár

address: Üllői út 26

department: 1st Department of Pathology and Experimental Cancer Research

country: Hungary

institute: Semmelweis University

summary: Fresh surgical specimens of Wertheim tumors, not used for for diagnostic purpose, were utilized to establish primary cell cultures. Frozen sections were used to determine the tumor free, and tumorous areas. Small tissue pieces of normal or tumorous regions were let to grow in Amnio-grow medium. Primary cells were positive for vimentin and negative for cytokeratin.

overall_design: To determine the molecules playing critical roles in the function of tumor-associated fibroblasts mRNA expression profile of 3 pairs of fibroblasts isolated from normal and tumorous cervical tissue and additional two normal primary cell cultures were analysed.

first: Ilona

last: Kovalszky


title: Transcriptome analysis upon REXO2 silencing

city: Warsaw

first: Sylwia

last: Czarnomska

state: Mazowieckie

address: Pawinskiego 5a

country: Poland

institute: Institute of Biochemistry and Biophysics Polish Academy of Sciences

summary: Investigation of the role of human REXO2 in mitochondrial RNA metabolism.

overall_design: The dataset corresponds to RNAseq studies and comprises experiment performed in triplicate, with the exception of REXO2 silencing sample for which duplicate was analyzed. The aim of this study was to examine the influence of REXO2 silencing on mitochondrial transcriptome. To this end we engineered two stable cell lines with the use of human HeLa Flp-In T-Rex cells as parental. First cell line inducible expressed miRNAs silencing endogenous copy of REXO2 gene while second one expressed additionally transgenic version of REXO2 which contained silent mutations causing insensitivity to miRNA. We also analyzed RNA isolated from parental HeLa Flp-In T-Rex cells. RNAseq libraries were prepared with the use of strand-specific library preparation procedures. RNAs were random fragmented and reverse transcribed using random oligomers as primers (dUTP-based protocol, see PMID: 29590189, PMID: 22609201; this pipeline enables analysis of RNAs (> ~100 nucleotides)). RNA was isolated from unfractionated cells using TRI-Reagent. Before preparation of the libraries total RNA was subjected to depletion of nuclear-encoded rRNAs (Ribo-Zero rRNA Removal Kit (Human/Mouse/Rat), Epicenter). Libraries were sequenced with the help of Illumina sequencing platform.

first: Maciej

last: Szewczyk

first: Roman

middle: J

last: Szczesny


title: RNA-Sequencing of Murine Stellate Cells and Gli1+ Peribiliary Mesenchymal Cells

city: New Haven

first: Vikas

last: Gupta

address: 1 Gilbert Street

institute: Yale School of Medicine

summary: To determine which genes drive the cellular identity of stellate cells and Gli1+ peribiliary mesenchymal cells, we isolated these two populations from murine livers in the uninjured state and 5 days after bile duct ligation. Gene expression profiles were compared.

first: Robert

last: Ec Schwartz


title: CAR T cells targeting uPAR are effective senolytics

city: New York

first: Yu-Jui

last: Ho

state: New York

address: 417 E 68th St

department: Cancer Biology & Genetics Program

institute: Memorial Sloan Kettering Cancer Center

summary: Cellular senescence is a stress-response program characterized by stable cell cycle arrest and a secretory program that modulates the tissue microenvironment.  Physiologically, senescence serves as a potent tumor suppressive mechanism that prevents the expansion of premalignant cells and plays a beneficial role in certain wound healing responses. Pathologically, the aberrant accumulation of senescent cells in response to chronic tissue damage produces an inflammatory milieu that contributes to diseases such as liver and lung fibrosis, atherosclerosis, diabetes and osteoarthritis.  Accordingly, experimental approaches to eliminate senescent cells from damaged tissues in mice can ameliorate symptoms of these pathologies and even promote longevity. Here we test the therapeutic concept that chimeric antigen receptor (CAR) T cells targeting senescent cells can serve as effective senolytics. We identify the urokinase plasminogen activator receptor (uPAR) as a cell surface protein that is broadly induced during senescence and demonstrate that uPAR-specific CAR T cells efficiently target senescent cells in vitro and in vivo. Administration of uPAR-targeting CAR T cells improves the survival of mice harboring lung adenocarcinoma cells treated with a senescence inducing drug combination, and eliminates fibrosis in mice treated with carbon tetrachloride or on a non-alcoholic steatohepatitis (NASH)-inducing diet. These results establish the potential of senolytic CAR T therapy for a range of senescence-associated diseases.

overall_design: Characterization of the transcriptional profiles of both murine therapy and oncogene induced senescence. For therapy induced senescence murine lung adenocarcinoma cell lines (KRASG12D,Tp53-/-) were cultured for 8 days with either DMSO or combined MEK and Cdk4/6 inhibitors to induce senescence. To study oncogene induced senescence hydrodynamic tail vein injection was performed to overexpress NRASG12V in murine hepatocytes and induce senescence.

first: Corina

last: Amor

first: Xiang

last: Li

first: Scott

middle: W

last: Lowe


title: Global evidence for programmed switching of gene expression from astrocytes  to neurons in response to PTB depletion

city: Shenzhen

first: Zhengyu

last: LIANG

state: Guangzhou

address: 1088 Xueyuan Avenue

department: Department of Biology

institute: Southern University of Science and Technology

summary: RNA-seq data were generated on replicate samples of isolated astrocytes from mouse cortex or midbrain before and after conversion to neurons by depleting PTB for 2 or 4 weeks.

overall_design: To characterize global switch in gene expression from astrocytes to neurons, we performed RNA-seq on isolated astrocytes from mouse cortex or midbrain and their converted neurons. As expected, typical astrocyte-specific genes were suppressed while neuronal-specific genes were induced upon shPTB induced neuronal conversion.

first: Hao

last: Qian

last: Liang

first: Hairi

first: Xiang-Dong

last: Fu


title: Characterization of circulating PDGFRa positive cells in fetus

city: Suita

first: Takashi

last: Shimbo

state: Osaka

address: 2-2 Yamadaoka

country: Japan

institute: Osaka university

summary: Mesenchymal stem/stromal cells (MSCs) are supplied to the damaged tissue via circulation when the injury is massive. MSCs recruited to the damaged tissues support tissue regeneration by differentiating into parenchyma cells and suppressing inflammation. However, most of the analysis of the circulating MSCs have been conducted in adult models, the existence and properties in fetal circulation was not known. Here, we have extensively analyzed the circulating MSCs cells during development.

overall_design: We isolated the circulating PDGFRa positive cells as GFP positive cells using PDGFRa-GFP knockin mice. The isolated cells were cultured and analyzed by single cell RNA-seq to clarify gene expression programs.


title: CARD9-associated Dectin-1 and Dectin-2 are Required for Protective Immunity of a Multivalent Vaccine Against Coccidioides posadasii Infection

city: San Antonio

first: Chiung-Yu

last: Hung

state: Texas

address: One UTSA Circle

country: USA

institute: The University of Texas at San Antonio

summary: Preparation of sequencing libraries and sequencing analysis using Illumina HiSeq 3000 platform (50 bp single-read module) were conducted at the Genomics Core of UT Health San Antonio. Sequencing reads were preprocessed by Trim Sequences tools in CLC Genomics workbench. Remaining reads were then aligned to mouse GRCm38(mm10) reference genome using RNA-Seq Analysis module in CLC Genomics workbench. To determine differentially expressed genes, exact test were performed after filtering lowly expressed gene (counts per million < 1 in more than three samples across the dataset) using edgeR package (Robinson, McCarthy et al. 2010). Gene Ontology (GO) analysis were performed using DAVID (Dennis, Sherman et al. 2003) and then visualized by GOplot (Walter, Sanchez-Cabo et al. 2015).

overall_design: BMMs were incubated with GCP-rCpa1 at a ratio of 50:1 for 12 hours with 5% CO2. BMMs incubated with PBS served as controls. Total RNA was isolated from BMMs cells using the Qiagen RNeasy kit.

last: Zhang

first: Althea

last: Campuzano


title: S-phase enriched non-coding RNAs regulate gene expression and cell cycle progression

city: Boston

first: Manashree

last: Damle

address: 185 Cambridge Street

department: Molecular Biology

institute: Massachusetts General Hospital

summary: We infer that non-coding RNAs can function as key effectors during cell cycle, acting on distinct regulatory networks to ensure maintenance of gene expression programs and cell cycle progression.

overall_design:
We performed nascent RNA-seq in asynchronized RPE-1 cells as well as synchronized RPE-1 cells at five different timepoints of S-phase of cell cycle progression. We also performed strand-specific RNAseq on control and three knockdown cell lines in which a lincRNA of interest was knocked down.
We also performed histone ChIP-seq and Cut-n-run seq on asynchronized RPE1 cells.

first: Ozlem

last: Yildirim

first: Enver

last: Izgu

first: Vladislava

last: Chalei

first: Fei

last: Ji

first: Ruslan

last: Sadrayev

first: Jack

last: Szostak

last: Kingston


title: Global transcriptional analysis of Geobacter sulfurreducens under palladium reducing

city: Mexico

first: Leticia

last: Vega

state: Mexico

address: Circuito Ext s/n

institute: Universidad Nacional Autonoma de Mexico

summary: Geobacter sulfurreducens is capable of reducing Pd(II) to Pd(0) using pyruvate as an electron donor; however, the biochemical and genetic mechanisms involved in this process have not been described. In this work, we carried out transcriptome profiling analyes to identify genes involved in Pd(II) reduction in this bacteria. Our results showed that 254 genes were upregulated while 142 were downregulated during Pd(II) reduction. Among the upregulated genes, 12 were related to energy metabolism and electron transport; 50 were classified as involved in protein synthesis; 42 were associated with regulatory functions and transcription, and 47 have no homologs with known function. RT-qPCR data confirmed upregulation during Pd(II) reduction of genes encoding PilA, the structural protein for the electrically conductive pili, and c-type cytochromes GSU1062, GSU2513, GSU2808, GSU2934, GSU3107, OmcH, OmcM, PpcA, PpcD. ΔpilA and ΔpilR mutant strains showed 20% and 40%, decrease in the Pd(II) reduction respectively, compared to the wild type strain, indicating the central role of the pili in this process. RT-qPCR data also confirmed downregulation during Pd(II) reduction of the omcB, omcC, omcZ, and omcS genes, which have been shown to be involved in Fe(III) reduction and electrodes. Based on these results, we propose mechanisms involved in Pd(II) reduction by G. sulfurreducens.

overall_design: mRNA profiles of Geobacter sulfurreducens in Pd(II) reduction or NBAF medium, in duplicate, using Illumina Genome Analyzer Iix

first: Alberto

last: Hernández-Eligio

first: Aurora

last: Pat-Espadas

last: Vega-Alvarado

first: Manuel

last: Huerta-Amparan

first: Francisco

last: Cervantes

first: Katy

last: Juárez


title: Elevated colonic mucin expression correlates with extended time to surgery for ulcerative colitis patients

city: Hershey

first: Yuka

last: Imamura Kawasawa

state: PA

address: 500 University Dr.

department: College of Medicine, Pharmacology

institute: Penn State University

summary:
Background: Ulcerative colitis (UC) develops from chronic inflammation of the colon. Most patients respond favorably to medical management, however around 25% of UC patients will require surgery to treat their disease. While surgery is the only curative treatment for UC, it can carry its own risks, particularly when the surgery is emergent. Time from diagnosis to surgery varies dramatically amongst UC patients, thus a better understanding of how the colonic microenvironment relates to disease severity is needed to improve patient outcomes.
Methods: We conducted RNA-seq on full-thickness sigmoid colonic tissues from 26 UC patients undergoing colectomy for their disease. Hierarchical clustering of the transcriptomes identified disease subsets. Subsets were characterized using differential gene expression analysis, cell type deconvolution, and network analysis.
Results: We identified two distinct UC subsets in our cohort that were distinguished by 957 differentially expressed genes. Cluster 1 was enriched in genes associated with intestinal epithelial cell differentiation, while cluster 2 was enriched in genes associated with epithelial-to-mesenchymal transition and inflammatory responses. Cluster 1 was associated with an extended time from UC diagnosis to colectomy. Of cluster 1 genes, elevated MUC5B, MUC4, and MUC2 expression displayed the strongest correlation with increased time to surgery.
Conclusion: Our transcriptome analysis indicates that UC can be subclassified into at least two distinct molecular signatures. We find that elevated mucin gene expression correlates with prolonged time to colectomy after diagnosis. These results identified MUC5B, MUC4, and MUC2 as potential prognostic indicators of disease severity, as reflected in time to surgery.

overall_design: Gene expression profiles from the surgically resected, diseased, sigmoid colon tissue of 26 ulcerative colitis patients were generated by deep sequencing.

first: Melanie

last: Eshelman

first: Walter

last: Koltun

first: Gregory

last: Yochum


title: Integrated scRNA-Seq Identifies Human Postnatal Thymus Seeding Progenitors and Regulatory Dynamics of Differentiating Immature Thymocytes

city: Ghent

first: Tom

last: Taghon

address: Corneel Heymanslaan 10

country: Belgium

institute: Ghent University

summary: During postnatal life, thymopoiesis depends on the continuous colonization of the thymus by bone marrow-derived hematopoietic progenitors that migrate through the bloodstream. The current understanding of the nature of thymic immigrants is largely based on data from pre-clinical models. Here, we employed single-cell RNA sequencing (scRNA-seq) to examine the immature postnatal thymocyte population in humans. Integration of bone marrow and peripheral blood precursor datasets identified two putative thymus seeding precursors that varied in expression of CD7; CD10; and the homing receptors CCR7, CCR9, and ITGB7. Whereas both precursors supported T cell development, only one contributed to intrathymic dendritic cell (DC) differentiation, predominantly of plasmacytoid dendritic cells. Trajectory inference delineated the transcriptional dynamics underlying early human T lineage development, enabling prediction of transcription factor (TF) modules that drive stage-specific steps of human T cell development. This comprehensive dataset defines the expression signature of immature human thymocytes and provides a resource for the further study of human thymopoiesis.

overall_design: Single cell transcriptional libraries were generated from CD34+ human post-natal thymocyte samples using the 10X Genomics Chromium 3’ sequencing V2 kit.

first: Marieke

last: Lavaert

first: Niels

last: Vandamme


title: TGF-ß treatment of human melanoma cell lines

city: Zwijnaarde

first: Jordy

last: De Coninck

address: Technologiepark 71

department: Department of Biomedical Molecular Biology

institute: University Gent

summary: To analyse whether the activation of TGF-ß signalling (recombinant hTGF-ß, 5 ng/mL; PeproTech) is able to invoke a full phenotype- switch, we performed RNA sequencing of melanoma  cultures from two independent melanoma cell lines at two different time point after TGF-β1 (5 ng/ml, preprotech).

overall_design: Control and treated replicates were processed using using TRIsure lysis buffer (BioLine, BIO-38032) according to the directives of the manufacturer. RNA quality was assessed using the 2100 Bioanalyzer (Agilent) prior to expression profiling.

first: Kenneth

last: Bruneel

first: Wouter

last: Van Loocke

first: Gillian

last: Blancke

first: Geert

last: Berx


title: BCG_CNRL_wk0vs3_infantMM

city: Seattle

first: Matthew

middle: Price

last: Wood

state: WA

address: 307 Westlake Ave North

department: Global Infectious Diseases

institute: Seattle Children's Research Institute

summary: To assess changes in immune gene expression following BCG vaccination in infants, 6 infant rhesus macaques were vaccinated with BCG at 2 weeks of age. Whole blood was collected at the time of BCG vaccination, and at 3 weeks post-BCG, in EDTA coated tubes. Peripheral blood mononuclear cells were collected by ficoll and RNA was extracted using Macherey Nagel nucleospin columns. RNA from both timepoints was then used for gene expression analysis using the Nanostring nCounter platform.

overall_design: Data from 12 infants was collected at pre-BCG and 3 weeks post-BCG timepoints using the NHP_Immunology_V2.0 770 gene panel. Differential gene expression in was run to measure changes from 0 to 3 weeks post-BCG vaccination.

first: Donald

last: Sodora


title: Ecotype differences in aggression, neural activity and behaviorally relevant gene expression in cichlid fish

city: Atlanta

first: Nicole

middle: M.

last: Baran

state: GA

address: 1510 Clifton Rd NE

department: Department of Psychology

institute: Emory University

summary:
Purpose: The goal of this study is to identify genes that showed a pattern of differential expression between species within activated neurons during mirror-elicited aggression between two species of Lake Malawi cichlid fish which differ in aggressive behaviors, Mchenga conophoros (MC, sand-dwelling) and Petrotilapia chitimba (PC, rock-dwelling). To do this, we use antibodies to phosphorylated ribosomal protein S6 (pS6) to enrich for transcripts bound to phosphorylated ribosomes followed by RNA-seq (PhosphoTRAP). In neurons, this phosphorylation occurs downstream of the binding of neurotransmitters and, thus, pS6 antibodies are increasingly being used as markers of neural activity, similar to immediate early genes (IEGs) like c-fos or egr-1.
Methods: Whole brain mRNA samples were collected from 8 adult male cichlids (4 MC, 4 PC) 90min following either 15min exposure to mirror in home tank (Mirror Test) or a Control condition. RNA-immunoprecipitation using antibodies to pS6 were performed and RNA was extracted from immunoprecipitated (IP) RNA samples. We also collected total RNA (input), prior to immunoprecipitation. For details see extraction protocol below. RNA from both input and IP samples was reverse transcribed using the SMART-Seq v4 Ultra Low Input RNA kit. Libraries were prepped using the NEBNext Ultra II FS DNA library Prep Kit for Illumina and sequenced. Sequence reads that passed quality filters were aligned to the Metriaclima zebra (Malawi cichlid) reference genome UMD2A using HiSat2. Transcripts were quantified using StringTie and differential expressional analyses were performed using DESeq2.
Results: We identified a large number of genes showing differential expression in mirror-elicited aggression, as well as many genes that differ between ecotypes. These genes, which may underly species differences in behavior, include several neuropeptides, genes involved in the synthesis of steroid hormones, and neurotransmitter activity.
Conclusions: This work lays the foundation for future experiments using this emerging genetic model system to investigate the genomic basis of evolved species differences in both brain and behavior.

sample_organism:
Mchenga conophoros
Petrotilapia sp. 'chitimba'

overall_design: Whole brain mRNA samples were collected from 8 adult male cichlids (4 of each species: Mchenga conophoros, MC, and Petrotilapia chitimba, PC), 90min following either 15min exposure to mirror in home tank (Mirror Test) or a Control condition. MC and PC differ in mirror-elicited aggression. Samples were processed in two batches, each containing all four conditions, all collected and processed together on the same day. Immunoprecipitation using antibodies to phosphorylated ribosomal protein S6 (pS6) was used to enrich for transcripts from activated neurons (PhosphoTRAP). We compared input (total RNA) to immunoprecipitated (IP) samples across condition and species to identify behaviorally relevant gene expression and genes exhibit patterns of differential expression across species.

first: J

middle: T

last: Streelman


title: Differential gene expression in mACVR1 NS v wt-ACVR1 NS.

city: Ann Arbor

first: Bioinformatics

last: Core

state: MI

address: BRCF Bioinformatics Core, 2800 Plymouth Rd

institute: University of Michigan

summary: To study the effect of the ACVR1 G328V mutation in tumor neurospheres with a background of NRASV12 overexpression and p53 knockdown. The goal of this study was to identify differentially expressed genes between mACVR1 NS v wt-ACVR1 NS.

overall_design: We had two groups: (1) mACVR1 NS (NPAC) and (2) wt-ACVR1 NS (NP). Each group had 3 biological replicates.

first: Maria

last: Castro

first: Flor

last: Mendez

first: Rebecca

last: Tagett


title: Next Generation Sequencing Facilitates Quantitative Analysis of mock oligodendrocytes (Ols) and tdTomato+ and tdTomato- Ols Transcriptomes.

status: Public on Apr 15 2020

city: Iowa City

first: Stanley

last: Perlman

state: Iowa

address: 3-712 BSB, 51 Newton Rd

institute: University of Iowa

summary:
Purpose: Comparision transcriptomes of isolated tdTomato+ and tdTomato- OLs from the brains and spinal cords of infected mice at 30 dpi to those of OLs isolated from mock-infected central nervous system (CNS) samples by nextgen sequencing
Methods: Brains and spinal cords were harvested at day 30 p.i. and processed as described above. After Live/Dead staining and treatment with Fc block, OLs were identified using APC-anti-O4 (1:50; Miltenyi). RNA was extracted from sorted OLs using an RNeasy Micro Kit (Qiagen). RNA sequence analysis was performed at the University of Minnesota Genomics Center.
Results: We identified 158 and 115 genes, that showed a significant difference (p<0.05) when tdTomato+ and tdTomato- cells were compared to cells from mock-infected mice, respectively, with 61 mRNAs common to both tdTomato- and tdTomato+ cells

overall_design: mRNA profiles of Ols from mock-infected mice, tdTomato+ and tdTomato- Ols from rMHV-infected mice were generated by next-generation sequecing.

first: Ruangang

last: Pan

first: Yu

last: Zhou

first: Qinran

last: Zhang

first: Xiufen

last: Zou