first: Xiao

last: Hu

address: 10 Amistad ST

department: Cell Biology

summary: The variability in reprogramming propensity is associated with the activity of the MKL1/SRF transcription factor and concurs with small cell size as well as rapid cell cycle. Reprogramming progressive cells can be prospectively identified by their low activity of a widely used synthetic promoter, CAG. CAG low cells arise and expand during cell cycle acceleration in the early reprogramming culture of both mouse and human fibroblasts. Here, CAG low and CAG high cells from reprogramming culture were FACS-sorted and subject to RNA-sequencing. Data support the notion that Low CAG promoter activity enriches for reprogramming progressive cells. Our work illustrates a molecular scenario underlying the distinct reprogramming propensities and demonstrates a convenient practical approach for their enrichment.

overall_design: For mouse cells, CAG:H2B-GFP transgenic MEFs were transduced with Dox-inducible Yamanaka factors OSKM, and induced for reprogramming. Cells with highest (10%) and lowest (10%) H2B-GFP intensity were FACS-sorted on reprogramming day 4 and subject to RNA-sequencing. For human cells, human secondary fibroblasts (with Dox-inducible Yamanaka factors) were transduced with CAG:GFP virus, and induced for reprogramming. Cells with highest (10%) and lowest (10%) GFP intensity were FACS-sorted on reprogramming day 7 and subject to RNA-sequencing.

first: Qiao

last: Wu

first: Jian

last: Zhang

first: Shangqin

last: Guo


title: RNAseq of splenic dendritic cells from 1 week-old and adult  Clec9a-Cre Rosa-Tomato mice

status: Public on Nov 06 2020

city: Planegg-Martinsried

first: Nikolaos

last: Papaioannou

address: Großhaderner Straße 9

country: Germany

institute: Biomedical Center

type: Expression profiling by high throughput sequencing

summary: We use bulk RNA sequencing of sorted cells to characterize the gene expression profiles of splenic TOM+ cDC2 and TOM- DC2 from 1 week-old mice as well and of TOM+ cDC2 from adult mice. We aim to reveal transcriptional differences based on ontogeny and developmental age. Sequencing results revealed that the developmental age rather than cell origin is a major determinantof the transcriptomic differences observed between young and adult cDC2.

overall_design: RNA sequencing of murine dendritic cell population from the spleen

first: N

middle: E

first: B

middle: U

last: Schraml


title: NAIL: an evolutionarily conserved lncRNA essential for coordinating p38 activation and NFκB mediated chromatin remodelling in inflammation_4

status: Public on Nov 01 2020

city: Singapore

first: Vinay

last: Tergaonkar

address: 61 Biopolis Drive,  Proteos

country: Singapore

institute: Institute of Molecular and Cell Biology

summary: We report the differential gene expression analysis of lncRNAs in wild-type, p65 KO and IKK2 KO MEFs stimulated with TNF alpha for 0min, 45min and 90 min.

overall_design: Immortalized wild-type, p65 KO and IKK2 KO MEFs were stimulated with TNFalpha (10ng/ml) for 0, 45 and 90 mins. Total RNAs were isolated using Trizol. RNA-Seq library was prepared by NEBNext® Ultra™ II Directional RNA library preparation kit according to manufacturer instructions. Paired-end sequencing was performed using Illumina NextSeq, 2x76bp. Raw reads were aligned to mouse genome GRCm38 by using HISAT2 .

first: Semih

middle: C

last: Akincilar


title: RNA-seq of hepatic mRNA from 21 week old PAI-1 +/- and littermate control mice fed standard chow

city: Chicago

first: Douglas

last: Vaughan

state: IL

address: Arkes Pavilion, Suite 2330, 676 N. St. Clair Street

department: Medicine

country: USA

institute: Northwestern University Feinberg School of Medicine

summary: We report RNA-seq data from hepatic mRNA on WT and PAI-1 heterozygous 21 week old mice.

overall_design: RNA-seq of hepatic mRNA

first: Joshua

middle: A

last: Levine


title: Genome-wide analysis of chromatin states of Ectocarpus sporophytes and gametophytes, experiment 1

status: Public on Nov 05 2020

city: Roscoff

first: J. Mark

last: Cock

address: Place Georges Teissier

country: France

institute: Station Biologique de Roscoff

type: Genome binding/occupancy profiling by high throughput sequencing

summary: The aim of this study was to analyze genome-wide patterns of histone post-translational modifications (PTMs) and nucleosome distribution during the sporophyte and gametophyte generations of the life cycle of the brown alga Ectocarpus. Analysis of the results of this experiment along with those generated by a second experiment “Genome-wide analysis of chromatin states of Ectocarpus sporophytes and gametophytes, experiment 2” indicated that the histone PTMs H3K4me2, H3K4me3, H3K9ac, H3K14ac and H3K27ac are associated with the transcriptional start sites (TSSs) of actively expressed genes, H3K36me3 preferentially marks gene bodies and is associated with gene activation, H4K20me3 marks transposons and is associated with gene repression, probably through the silencing of transposons in introns. H3K79me2 occurs in zones corresponding to about a third of the genome. These zones often span several genes and genes within H3K79me2 zones exhibit lower levels of transcript abundance.

overall_design: Analysis of six histone post-translational modifications, along with FAIRE-seq and RNA-seq analyses, during the two generations (sporophyte and gametophyte) of the life cycle of the brown alga Ectocarpus, using duplicate samples for each generation.

first: Simon

last: Bourdareau

first: J

middle: M


title: Coordination between the microbiota and the root diffusion barriers is required for plant mineral nutrient homeostasis

city: Loughborough

first: Gabriel

last: Castrillo

address: Nr Loughborough

department: School of Biosciences

country: United Kingdom

institute: University of Nottingham

summary: RNASeq of roots from two genotypes of Arabidopsis thaliana plants, Col-0 and myb36-2 grown axenically or with a 41 member bacterial Synthetic Community (SynCom) to explore the interaction between the root diffusion barriers and the root microbiome.

overall_design: Explore the interaction of the root diffusion barriers and the microbiome

first: Isai

last: Salas-Gonzalez


title: SO-018 UNH: Influence of omega 3 supplementation over 3 generation of mice on the effect of metabolic stress in different organs

city: clermont ferrand

first: frederic

last: capel

address: 28 place henri dunant

department: alimentation humaine

institute: INRA

type: Non-coding RNA profiling by high throughput sequencing

summary: Study the influence of omega 3 supplementation over 3 generation of mice on the effect of metabolic stress in different organs

overall_design: 3 groups of mice were included n=7-9 per group, a control low fat diet group, a high fat-control lineage group and a high fat-omega 3 lineage group

first: Frederic

last: Capel

first: Karla

middle: F

last: Corral Jara

first: Alexandre

last: Pinel

first: Jean

middle: P

last: Rigaudière

first: Tao

last: Ye


title: Genome-wide analysis of chromatin states of Ectocarpus sporophytes and gametophytes, experiment 2

summary: The aim of this study was to analyze genome-wide patterns of histone post-translational modifications (PTMs) and nucleosome distribution during the sporophyte and gametophyte generations of the life cycle of the brown alga Ectocarpus. Analysis of the results of this experiment along with those generated by a second experiment “Genome-wide analysis of chromatin states of Ectocarpus sporophytes and gametophytes, experiment 1” indicated that the histone PTMs H3K4me2, H3K4me3, H3K9ac, H3K14ac and H3K27ac are associated with the transcriptional start sites (TSSs) of actively expressed genes, H3K36me3 preferentially marks gene bodies and is associated with gene activation, H4K20me3 marks transposons and is associated with gene repression, probably through the silencing of transposons in introns. H3K79me2 occurs in zones corresponding to about a third of the genome. These zones often span several genes and genes within H3K79me2 zones exhibit lower levels of transcript abundance.

overall_design: Analysis of six histone post-translational modifications, along with MNase-seq, H2AZ and RNA-seq analyses, during the two generations (sporophyte and gametophyte) of the life cycle of the brown alga Ectocarpus, using duplicate samples for each generation.


title: RNA-seq of hepatic mRNA in mice fed chow with and without the PAI-1 inhibitor TM5614

summary: We report RNA-seq data from hepatic mRNA in mice on standard chow.  Mice 41-45 were fed standard chow.  Mice 46-50 were fed standard chow + a PAI-1 inhibitor, TM5614 for 10 days.


title: scRNAseq of  sorted splenic MHCII+ cells  from 9 day-old female Clec9a-Cre/Cre Rosa-tdTomato/tdTomato mice after exclusion of CD19+ B cells and autofluorescence+F4/80hi macrophages

first: Nikolaos

summary: We use scRNA sequencing of MHCII+ cells in the neonatal spleen to elucidate the heterogeneity in young cDC populations and their potential ontogenetic relationship with MHCII-expressiong ILCs. Additionally, we aim to uncover the transcriptomic differences of TOM+ cDC2 and TOM- DC2 in the single cell level. Our data reveal that the neonatal spleen consists of the same cDC1 and cDC2 subsets that comprise the adult cDC compartment. Additionallu, ILCs do not seem to relate otogenetically with TOM- DC2.In the cDC2 clusters, TOM+/- cells are evenly distributed indicative of their transciptomic resemblance. A unique and distinct cluster of TOM- DC2 is observed.

overall_design: scRNA sequencing of murine sorted splenic MHCII+ cells  from 9 day-old female mice. 3 mice were pooled to increase the yield of retrieved cells.

first: N


title: SO018 liver gene expression

summary:
Identification of the effect of transgenerationnal supplementation with omega 3 fatty acids on the resistance to a diet induced obesity challenge
Although several lines of evidence suggested that omega 3 fatty acids (FA) may protect from obesity and its related disorders by modulation of the metabolism of insulin-responsive organs, the long term effect remains unknown. Preservation of the liver metabolic homeostasis could be a key mediator of omega 3 FA effects. The impact of transgenerational intake of long chain omega 3 PUFA on hepatic adaptations during a nutritional challenge was investigated in young C57Bl6J male mice.

first: alexandre

last: pinel

first: Claire

last: Naylies

first: Anthony

last: Emile

first: Yannick

last: Lippi


title: NAIL: an evolutionarily conserved lncRNA essential for coordinating p38 activation and NFκB mediated chromatin remodelling in inflammation_2

first: Vinay

last: Tergaonkar

summary: We report the differential gene expression analysis of DSS treated colon tissues obtained from WT and NAIL-ΔNFκB  mouse.

overall_design: Mice were fed with 3% DSS for 4 days and another 4 days with normal water. Total RNA was isolated from the colon tissues (distal) of mice treated with or without DSS.

first: Semih

last: Akincilar

first: Lele

last: Wu


title: Deep sequencing of an AML cell line treated with vehicle control, BETi, MDM2i, or a drug combination

status: Public on Nov 04 2020

city: La Jolla

first: Xue

last: Lei

state: CA

address: 10901 N Torrey Pines Rd

institute: SBP medical discovery institute

summary:
Gene expression changes in human OCI-AML3 cells treated with DMSO (vehicle control), the BET inhibitor CPI-203, the MDM2 inhibitor Nutlin-3a, or a combination of both drugs, were compared.
Method: poly-A mRNA from the OCI-AML3 cell line treated for 24 hrs with either DMSO, CPI-203, Nutlin-3a or a combination of both drugs was extracted, in triplicate, then sequenced with an Illumina NextSeq500 sequencer. Sequence reads passing the quality control filters were aligned using Tophat2 and then analysed with Cufflinks.

overall_design: polyA mRNA profiles from drug-treated human OCI-AML3 cells were generated, in triplicate, using an Illumina GAIIx sequencer.

first: Anne-Louise

last: Latif

first: Peter

middle: D

last: Adams

first: Kathryn

last: Gilroy


title: Deep sequencing of an AML cell line, with and without p53, treated with vehicle control, BETi, MDM2i, or a drug combination

summary:
Gene expression changes were compared in human OCI-AML3 cells, with or without p53 knock down, that were treated with DMSO (vehicle control), the BET inhibitor CPI-203, the MDM2 inhibitor Nutlin-3a, or a combination of both drugs. compared.
Method: OCI-AML3 cells were transduced with either empty vector or vector expressing an shRA targeting p53. Cells were then treated for 24 hrs with either DMSO, CPI-203, Nutlin-3a or a combination of both drugs. PolyA mRNA was extracted, in triplicate, then sequenced with an Illumina NextSeq 500 sequencer. Sequence reads passing the quality control filters were aligned using Tophat2 and then analysed with Cufflinks.

Keywords: Expression profiling by high throughput sequencing

overall_design: polyA mRNA profiles from drug-treated human OCI-AML3 cells with or without p53 were generated, in triplicate, using an Illumina NextSeq500 sequencer.

first: Ashley

last: Newcombe


title: NG Capture-C from human erythroid and embryonic stem cells

status: Public on Nov 02 2020

city: Headington

first: Damien

last: Downes

state: Oxfordshire

address: John Radcliffe Hospital

department: MRC Weatherall Institute of Medicine

institute: The University of Oxford

summary: NG Capture-C provides high resolution chromatin conformation capture (3C) interaction maps. NG Capture-C was carried out in erythroid and hESC cells at the Hba, Hbb, Myc, and Slc25a37 locus to generate sequence libraries for analytical software testing.

overall_design: NG Capture-C was performed in Day 10 differentiating erythroid (Ery) cells from three donors, and human Embryonic Stem Cells (H1 hESC; WiCell) from four independently grown cultures. Capture-C was performed with biotinylated oligonucleotides targeting sequences adjacent to DpnII sites in targeted to the Hba-1, Hba-2, Hbb, Hbd, Myc and Slc25a37 promoters.

middle: J

first: Jim

middle: R

last: Hughes


title: Quantification of transcriptional changes during in vivo mouse erythropoiesis by nascent RNA-sequencing

city: Oxford

first: Robert

last: Beagrie

address: Headley Way

department: Molecular Haematology Unit

institute: MRC Weatherall Institute of Molecular Medicine

summary: We isolated erythroid progenitor cells at various stages of differentiation from mouse fetal liver using flow cytometry. We metabolically labelled RNA from purified progenitor populations with 4-thiouridine (4sU) for 45 minutes in ex vivo culture, then extracted cellular RNA. Nascent RNA was biotinylated by reaction of the incorporated thiol groups with MTS-Biotin, then purified using streptavidin nanobeads and sequenced.

overall_design: Nascent RNA-sequencing of metabolically (4sU) labelled RNA in primary erythroid progenitors from mouse fetal liver


title: Modelling erythropoiesis in congenital dyserythropoietic anaemia type I (CDA-I) for diagnosis and molecular dissection.

summary: Congenital Dyserythropoietic Anaemia type 1 (CDA-I) is an inherited anaemia arising primarily from mutations in C15orf41 and CDAN1, however the molecular mechanisms that cause the disease remain to be fully elucidated. We use an in vitro culture system to study multiple stages of erythropoiesis from CD34+ progenitors of patients with CDA-I. Applying a number of techniques, including ATAC-seq, we show that differentiation of CDA-I patient erythroblasts closely matches that of healthy donors during the early and intermediate stages of erythroid differentiation and maturation. However, a defect in terminal erythropoiesis can be observed in the CDA-I patient derived erythroblasts, resulting in a reduction in the number of enucleated erythroid cells.

overall_design: 32 ATAC-seq samples of cultured erythroblasts from Cogenital DyserythropoieticAnemia Type 1 (CDA-I) CD34 progenitors from 7 Donors, with up to 2 technical replicates for each timepoint during culture (Day7, Day10, Day11, Day13)

first: Caroline

last: Scott


title: Evolutionary Engineering of E. coli MG1655 for Tolerance Against Isoprenol

status: Public on Oct 31 2020

city: Leipzig

first: Jens

middle: Olaf

last: Kroemer

address: Permoserstr. 15

department: Solar Materials

institute: Helmholtz-Zentrum für Umweltforschung GmbH - UFZ

summary: Here we applied adaptive laboratory evolution successfully to evolve E. coli towards high tolerance against isoprenol, increasing growth at the half-maximal inhibitory concentration by 47%. RNA-sequencing showed that the deletion identified upstream of yghB correlated with a strong overexpression of the gene.

sample_organism: Escherichia coli str. K-12 substr. MG1655

overall_design: RNA-Sequencing of 3 different E. coli MG1655 strains (Culture A, Culture B, Culture C) adapted to Isoprenol and wild-type strain in three biological replicates grown with Isoprenol stress.

middle: O

first: Heiko

last: Babel


title: Transcriptomic and epigenetic preservation of genetic sex identity in estrogen-feminized male chicken embryonic gonads

city: Charlestown

first: Toshihiro

last: Shioda

state: MA

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

department: Center for Cancer Research

institute: MGH Cancer Center

type: Methylation profiling by high throughput sequencing

summary: Whereas in ovo exposure of genetically male (ZZ) chicken embryos to exogenous estrogens temporarily feminizes gonads at the time of hatching, the morphologically ovarian ZZ-gonads (FemZZs for feminized ZZ gonads) are masculinized back to testes within one year. To identify the feminization-resistant “memory” of genetic male sex, FemZZs showing varying degrees of feminization were subjected to transcriptomic, DNA methylome, and immunofluorescence analyses. Protein-coding genes were classified based on their relative mRNA expression across normal ZZ-testes, genetically female (ZW) ovaries, and FemZZs. We identified a group of 25 genes that were strongly expressed in both ZZ-testes and FemZZs but dramatically suppressed in ZW-ovaries. Interestingly, 84% (21/25) of these feminization-resistant testicular marker genes, including the DMRT1 master masculinizing gene, were located in chromosome Z. Expression of representative marker genes of germline cells (e.g., DAZL or DDX4/VASA) was stronger in FemZZs than normal ZZ-testes or ZW-ovaries. We also identified 231 repetitive sequences (RSs) that were strongly expressed in both ZZ-testes and FemZZs, but these RSs were not enriched in chromosome Z. Although 94% (165/176) of RSs exclusively expressed in ZW-ovaries were located in chromosome W, no feminization-inducible RS was detected in FemZZs. DNA methylome analysis distinguished FemZZs from normal ZZ- and ZW-gonads. Immunofluorescence analysis of FemZZ gonads revealed expression of DMRT1 protein in medullary SOX9+ somatic cells and apparent germline cell populations in both medulla and cortex. Taken together, our study provides evidence that both somatic and germline cell populations in morphologically feminized FemZZs maintain significant transcriptomic and epigenetic memories of genetic sex.

overall_design: Transcriptomes and DNA methylomes of day-19 chicken embryonic gonads with or without exposure to 17alpha-ethynylestradiol in ovo were examined by deep sequencing methods RNA-seq and MBD-seq, respectively.

first: Keiko

first: Junko

last: Odajima

first: Misato

last: Kobayashi

first: Mutsumi

first: Bianca

last: Cordazzo

first: Kurt

last: Isselbacher

first: Toshi


title: Ambigols from the Cyanobacterium Fischerella ambigua increase Prodigiosin Production in Serratia sp.

city: Halle

first: Tomasz

last: Chilczuk

address: Hoher Weg 8

institute: University of Halle-Wittenberg

summary: Screening a library of 573 cyanobacteria extracts for inhibition of the quorum sensing regulated prodigiosin production of Serratia marcescens, an extract of the cyanobacterium Fischerella ambigua (Näg.) Gomont 108b was found to drastically increase the prodigiosin production. Bioactivity-guided isolation of the active compounds resulted in the two new natural products ambigol D and E along with the known ambigols A and C. Ambigol C treatment increased prodiginine production of Serratia sp. ATCC 39006 (S39006) by a factor of 10, while ambigols A and D were found to have antibiotic activity against this strain. RNA-Seq of S39006 treated with ambigol C and subsequent differential gene expression and functional enrichment analyses indicated a significant downregulation of genes associated with the translation machinery and fatty acid biosynthesis in Serratia, as well as increased expression of genes related to the uptake of l-proline. These results suggest that the ambigols increase the prodiginine production in S39006 not by activating the SmaIR quorum sensing system, but possibly by increasing the precursor supply of l-proline and malonyl-CoA.

overall_design: Six samples were analyzed in this study. Two different treatments were utilized with three biological replicates of each. The two treatments included growth of Serratia sp. ATCC 39006 with 15.6 µM ambigol C and with DMSO (solvent control).

first: Rita

last: Monson

first: Peter

last: Schmieder

first: Vesselin

last: Christov

first: Heike

last: Enke

first: George

last: Salmond

first: Timo

middle: H

last: Niedermeyer


title: LEAD-m6A-seq data of HeLa mRNA

first: Chuan

last: He

state: Illinois

address: 929 E 57th St

department: Chemistry

institute: University of Chicago

summary: We applied LEAD-m6A-seq to 11 sites of HeLa mRNA to probe m6A modification at the target sites.

overall_design: DNA probes were annealed to mRNA extracted from HeLa cells. The annealing product underwent cDNA extension reaction mediated by Bst DNA polymerase and was then subjected to library construction and illumina sequencing.

first: Yuru

last: Wang

first: Zijie

last: Zhang

first: Caraline

last: Sepich-Poore

first: Lisheng

first: Yu

last: Xiao


title: Generation of human induced pluripotent stem cell-derived 3D human striatal spheroids

city: STANFORD

first: Yuki

last: Miura

state: California

address: 318 Campus Drive, W200

department: Department of Psychiatry and Behavioral Sciences

institute: Stanford University

summary: Single cell RNA-seq characterization of stem cell derived organoids resembling the human striatum

overall_design:
Single-cell RNA sequencing of human induced pluripotent stem (hiPS) cell-derived striatal spheroids from three hiPS cell lines
Raw data not submitted due to patient privacy concerns

first: Sergiu

last: Pașca


title: Piwil1 Regulates Glioma Stem Cell Maintenance and Glioblastoma Progression

status: Public on Oct 30 2020

city: Cleveland

first: Haidong

last: Huang

state: Ohio

address: 9500 Euclid Avenue

department: Cancer Biology

institute: Cleveland Clinic Foundation

summary: Piwi proteins are a subfamily of Argonaute proteins that maintain germ cells in eukaryotes. However, the role of their human homologues in cancer stem cells and more broadly in cancer is poorly understood. Here, we report that the Piwi-like family members, including Piwil1 (Hiwi), are overexpressed in glioblastoma (GBM), with Piwil1 levels most frequently elevated.  Piwil1 is enriched in glioma stem cells (GSCs) and helps to maintain their self-renewal. GSCs were transduces with control non-targeting shRNAs (shNT) and shPiwil1 (#1 and #2) and global gene expression was analyzed to identify Piwil1 downscream singalings.

overall_design: 3832 GSCs were transduced with shNT or two shPiwil1 (#1 and #2) lentivirus. Each shRNA had four biological replicates.


title: Quantitative Analysis by Next Generation Sequencing (NGS) of KBM7 and K562 chronic myeloid leukemia (CML) cells transcriptomes upon USP15 siRNAs-mediated knockdown.

city: Amsterdam

first: Elisabetta

last: Citterio

address: Plesmanlaan 121

department: Molecular Genetics

country: Netherlands

institute: Netherlands Cancer Insitute (NKI-AVL)

summary:
Purpose: the goal of this study is to investigate the consequences of ubiquitin specific protease 15 (USP15) knockdown on gene expression in KBM7 and K562 leukemia cells.
Methods: KBM7 or K562 cells were transfected with USP15 ( siGENOME Human USP15 (9958) siRNA-SMART pool M-006066-01) or control (Ctrl) siRNAs (siGENOME Non-Targeting siRNA Control Pool#2 D-001206-14-05, Dharmacon). Cells were transfected using Lipofectamine RNAiMAX Reagent from Life Technologies following the manufacturer’s instructions and assayed at 72 hrs after transfection in Western blotting, RNA-seq and qRT-PCR. Western blotting confirmed knokdown of USP15 protein. mRNA profiles were generated by deep sequencing using Illumina Hiseq2500. Alignments of the sequence reads that passed quality filters were performed using TopHat2.1, Genome build 38 and Ensembl gtf version 77 and genecounts have been generated using Itreecount. https://github.com/NKI-GCF/itreecount.  qRT–PCR validation was performed using SYBR Green assays.The amount of target, normalized to an endogenous reference (HPRT), was calculated by 2-DDCT.
Results: We assigned about 30 million reads per sample uniquely to a gene of the human reference genome (hg38). We identified 26,227 genes in KBM7 and  28,329 genes in K562 cells using TopHat2.1 in combination with Itreecount (https://github.com/NKI-GCF/itreecount) workflow. Comparison of normalized gene expression data resulted in a total of 657 and 330 differential expressed genes between USP15 KD vs Ctrl in KBM7 and K562 cells (in two separate experiments), respectively, with a treshold of logFC_C1 ≥1. and adj.PVal_C1 value <0.05.  Downregulation of USP15 was confirmed by Western blotting and by qRT-PCR. Ingenuity pathway analysis of USP15-dependent genes in the KBM7 and K562 datasets (cut-off logFC > 1; adjusted P value <0.05) uncovered activation of inflammation-related pathways, which involve JAK/STAT and PI3K signal transduction. In K562, we also measured significant down-modulation of TGF-beta signaling. Expression of a set of 17 genes was assessed by RT-qPCR in three independent experiments per cell line and altered expression was validated. Overlap between Top 50 significantly enriched Ingenuity Upstream Regulators in each cell line RNAseq datasets was found. The Upstream Regulators were calculated based on differential gene expression between USP15 and control siRNAs. The nine overlapping terms include: Ifnar, IRF3, STAT1, Interferon alpha, IFNG, CD40LG, IFNB1, TNF and poly rI:rC-RNA.
Conclusions: Our results represent the first detailed analyis of the consequences of USP15 depletion on gene expression in malignant hematopoietic populations such as KBM7 and K562 chronic myeloid leukemia cells by genome wide expression profiling in USP15 KD vs Ctrl cells. We confirmed downregulation of USP15 and validated altered expression of a set of 17 genes by qRT-PCR. Ingenuity pathway analysis identified 657 and 330 differentially regulated genes in KBM7 and K562, respectively.  The changes in gene expression, though not extensive, indicate that RNAi of USP15 led to activation of inflammation-related pathways, which involve JAK/STAT and PI3K signal transduction in both cell lines. In K562, we also measured significant down-modulation of TGF-beta signaling, which is required for USP15 pro-oncogenic role in human glioma cells (Eichhorn et al., 2012). Regulation of inflammatory signals and TGF-beta are relevant both in normal HSC and in malignant development (Blank and Karlsson, 2015).

overall_design: KBM7 or K562 cells were transfected with USP15 or control (Ctrl) siRNAs

first: Gaetano

last: Gargiulo

first: Paul

last: van den Berk

first: Danielle

last: Hulsmann

first: Iris

last: de Rink


title: Prenatal Alcohol Exposure in Rats Diminish Postnatal Cxcl16 Chemokine Ligand Brain Expression

city: Guadalajara

first: Pedro

last: Juárez Rodríguez

state: Jalisco

address: Sierra Mojada #950

department: Biología Molecular y Genómica

country: Mexico

institute: Universidad de Guadalajara

summary: Maternal ethanol consumption during pregnancy is one of the main causes of Neurodevelopmental disorders (NDD). Prenatal alcohol exposure (PAE) produces several adverse manifestations. Even low or moderate intake has been associated with long lasting behavioral and cognitive impairment in offspring. In this study we examined the gene expression profile in the rat nucleus accumbens using microarrays, comparing animals prenatally exposed to ethanol and controls. The microarray gene expression showed an overall downward regulatory effect of PAE. Gene cluster analysis reveals that the main affected gene groups are related to transcription regulation, transcription factors and homeobox genes. We focus on the expression of the C-X-C motif chemokine ligand 16 (Cxcl16) which was differentially expressed. There is a significant reduction in the expression of this chemokine throughout the brain under PAE conditions, evidenced here by qPCR and immunohistochemistry. Chemokines are involved in neuroprotection and implicated in alcohol-induced brain damaged and neuroinflammation in the developing CNS, therefore, the significance of the overall decrease in Cxcl16 expression in the brain as a consequence of PAE, may reflect a reduced ability to neuroprotection against subsequent conditions, such as excitotoxic damage, inflammatory processes or even hypoxic-ischemic insult.

overall_design: Transcriptional profiling of rat nucleus accumbens comparing rats prenatally exposed to ethanol (EtOH) with control isocaloric solution (ISO). Microarray analysis was performed on pools of seven rats for each group (i.e. seven ISO, seven EtOH) to compensate for inter-organism variability in an heterologous hybridization.

last: Juárez-Rodríguez

first: Juliana

last: Godínez-Rubí

first: Carolina

last: Guzmán-Brambila

first: Edgar

last: Padilla-Velarde

first: Arturo

last: Orozco-Barocio

first: Daniel

last: Ortuño-Sahagún

first: Argelia

last: Rojas Mayorquín


title: Dynamic single-cell RNA sequencing identifies immunotherapy persister cells following PD-1 blockade II

city: Boston

first: Cloud

middle: Peter

last: Paweletz

address: 360 Longwood Avenue

department: Medical Oncology

institute: Dana-Farber Cancer Institute

summary: We report results of transcriptional profiling of ex vivo MC38 murine organotypic tumor spheroids following programmed death-1 (PD-1) blockade compared to isotype control IgG treatment by bulk and single-cell RNA-sequencing after 6 days of treatment. Additionally, we report results from single-cell RNA-sequencing of MC38 tumors treated in vivo with PD-1 blockade versus isotype control IgG to validate ex vivo workflow.

overall_design: Transcriptional profiling of residual MC38 tumor cells after PD-1 blockade

first: Kartik

last: Sehgal

first: Andrew

last: Portell

first: Elena

last: Ivanova

first: Patrick

last: Lizotte

first: Juan

last: Miret

last: Pawaletz

first: David

last: Barbie


title: Tracheoesophageal fistula characterisation

city: Rotterdam

first: Erwin

last: Brosens

state: - If in U.S, Select a State -

address: Dr molewaterplein 50

department: Clinical genetics

institute: ErasmusMC

summary:
Esophageal atresia and tracheoesophageal fistula (EA/TEF) are relatively frequently occurring foregut malformations with a largely unknown etiology. EA/TEF is thought to have a strong genetic component and several genes have been proven to be involved in syndromic EA/TEF. However, it is not clear which biological processes or gene networks are disturbed. To gain more insight in the origin of the TEF, we aimed to examine and describe TEF composition using a combination of whole-genome transcription profiling and (immuno-) histochemical stainings. We hypothesized that such characterization of human TEFs provides insight in the molecular and mechanistic etiology of EA/TEF.
Data analysis was carried out using BRB-array tools version 4.6.0 (October 2018) in combination with R version 3.5.1 (July 2018). For each probe set, the geometric mean of the hybridization intensities of all samples was calculated. The level of expression of each probe set was determined relative to this geometric mean and logarithmically transformed (on a base 2 scale) to ascribe equal weight to gene-expression levels with similar relative distances to the geometric mean.

overall_design: We compared the transcriptomes of 21 tracheoesophageal fistulas to 3 esophageal, 3 tracheal and 4 lung sample controls. No replicates are included in the study

first: Robbert

last: Rottier


title: TBX3 promotes proliferation of papillary thyroid carcinoma cells through facilitating PRC2-mediated p57KIP2 repression

city: Tianjin

first: xiaomeng

last: Li

address: No.22 qixiangtai Road,heping district,Tianjin

country: China

institute: Tianjin medical university

summary:
Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare NGS-derived thyroid cancer cell line k1 transcriptome profiling (RNA-seq) to microarray and quantitative reverse transcription polymerase chain reaction (qRT–PCR) methods and to evaluate protocols for optimal high-throughput data analysis
Methods: mRNA profiles of  shCtrl and shTBX3 cells were generated by deep sequencing, in duplicate, using BGISEQ-500. The sequence reads that passed quality filters were analyzed at the transcript isoform level with two methods:Bowtie2 to map clean reads to reference gene and use HISAT to reference genome.Bowtie2 parameters forSE reads:-q--phred64--sensitive--dpad0--gbar and HISTAparameters forSE reads:-p8--phred64--sensitive-I1-X 1000. qRT–PCR validation was performed using SYBR Green assays
Results: Using an optimized data analysis workflow, we mapped about 24 million sequence reads per sample to the human genome (build hg19) and identified 20511 transcripts in K1-shCtrl and K1-shTBX3 cells with RSEM workflow.  Approximately 5% of the transcripts showed differential expression between  K1-shCtrl and K1-shTBX3 cells, with a fold change ≥1.5 and p value <0.01. Altered expression of 16 candidate genes was confirmed with qRT–PCR, demonstrating the high degree of sensitivity of the RNA-seq method. Hierarchical clustering of differentially expressed genes uncovered several as yet uncharacterized genes that may contribute to  thyroid cancer function.
Conclusions: Our study represents the first detailed analysis of  thyroid cancer cell line k1 transcriptomes, with biologic replicates, generated by RNA-seq technology. The optimized data analysis workflows reported here should provide a framework for comparative investigations of expression profiles. Our results show that NGS offers a comprehensive and more accurate quantitative and qualitative evaluation of mRNA content within a cell or tissue. We conclude that RNA-seq based transcriptome characterization would expedite genetic network analyses and permit the dissection of complex biologic functions.

overall_design: Total mRNA profiles of   K1-shCtrl and K1-shTBX3 cells were generated by deep sequencing, in duplicate, using BGISEQ-500

first: Xiaomeng

first: Li

last: Zhao


title: Genome-wide expression changes induced by bisphenol A, F and S in human stem cell derived hepatocyte-like cells

status: Public on Oct 29 2020

city: Dortmund

first: Jan

middle: G.

last: Hengstler

address: Ardeystrasse 67

institute: Leibniz Research Centre for Working Environment and Human Factors at the TU Dortmund University (IfADo)

summary: The debate about possible adverse effects of bisphenol A (BPA) has been ongoing for decades and bisphenol-F (BPF) and -S (BPS) have been suggested as “safer” alternatives. In the present study we used hepatocyte-like cells (HLC) derived from the human embryonic stem cell lines Man12 and H9 to compare the three bisphenol derivatives. Stem cell-derived progenitors were produced using an established system, and, during their transition to HLCs, they were exposed to BPA, BPF and BPS for 8 days. Subsequently, we examined cell viability, inhibition of cytochrome P450 (CYP) activity, and genome-wide RNA profiles. Sub-cytotoxic, inhibitory concentrations (IC50) of CYP3A were 20, 9.5 and 25 µM for BPA, BPF and BPS in Man12 derived HLCs, respectively. The corresponding concentrations for H9-derived HLCs were 19, 29 and 31 µM. These IC50 concentrations were used to study global expression changes in this in vitro study and are higher than unconjugated BPA in serum of the general population. A large overlap of up- as well as down- regulated genes induced by the three bisphenol derivatives was seen. This is at least 28-fold higher compared to randomly expected gene expression changes. Moreover, highly significant correlations of expression changes induced by the three bisphenol derivatives were obtained in pairwise comparisons. Dysregulated genes were associated with reduced metabolic function, cellular differentiation, embryonic development, cell survival and apoptosis. In conclusion, no major differences in cytochrome inhibitory activities of BPA, BPF and BPS were observed and gene expression changes showed a high degree of similarity.

overall_design:
Human embryonic stem cells (hESCs) derived hepatic progenitor cells were mature into HLC in the presence of different concentrations of bisphenol A (BPA), -F (BPF) and -S (BPS) for 8 days to determine the CYP3A IC50 value for each cell line and compound. Analysis of Cyp3A P450 activity upon exposure to BPA revealed that both Man12 and H9 HLCs responded in similar fashion, with IC50 values of 20 μM and 19 μM respectively. However, cell line differences were observed upon exposure to BPF and BPS. Man12 HLCs were more sensitive to these compounds when compared to H9 HLCs. BPF- and BPS-exposed Man12 HLCs displayed IC50 values of ~10 μM and 25 μM respectively, whereas this was increased in H9 HLCs to ~29 μM and 31 μM respectively. Genome-wide gene expression profile and PCR array to examine key genes involved in cell viability and apoptosis were performed in HLCs exposed to the IC50 concentration of the bisphenols for CYP3A function.
Five technical replicates were prepared for each condition. In replicate E of the Man12 cell line untreated control, vehicle control and BPS treated cells died prematurely in culture. Therefore, no microarrays were generated for these experiments. Since the limma analysis was performed paired against the untreated control, Man12 replicate E was excluded from the analysis.

first: Baltasar

last: Lucendo-Villarin

last: Nell

first: Panagiotis

last: Filis

last: Feuerborn

first: Peter

last: O'Shaughnessy

first: Patricio

last: Godoy

first: Birte

last: Hellwig

first: Jörg

last: Rahnenführer

middle: G

first: Anna

last: Cherianidou

first: Agapios

last: Sachinidis

first: Paul

last: Fowler

last: Hay


title: PRRX1 promotes malignant properties in human osteosarcoma

city: okayama

first: Takeshi

last: Takarada

state: okayama

address: 2-5-1 Shikata-cho

department: Graduate School of Medicine, Dentistry and Pharmaceutical Sciences

country: Japan

institute: Okayama university

summary: Compare the gene expression profiles of 143B after PRRX1 knockdown

overall_design: RNA samples extracted from 143B/Ctrl, 143B/ shPRRX1#1 and 143B/shPRRX1#2

first: Daisuke

last: Yamada

first: Masahiro

last: Nakamura


title: Genetic and pharmacological inhibition of the nuclear receptor RORa regulates Th17 driven inflammatory disorders

city: Jupiter

first: Laura

last: Solt

state: FL

address: 130 Scripps Way

department: Immunology and Microbiology

institute: The Scripps Research Institute

summary: Full development of IL-17 producing CD4+ T helper cells (TH17 cells) requires the transcriptional activity of both orphan nuclear receptors RORa and RORgt.  Despite this evidence, RORa is considered functionally redundant to RORgt; thus, the function and therapeutic value of RORa in TH17 cells remains underexplored.  Using mouse models of autoimmune and chronic inflammation, we show that expression of RORa is required for TH17 cell pathogenicity. T-cell specific deletion of RORa reduced the development of experimental autoimmune encephalomyelitis (EAE) and colitis.  Reduced inflammation was associated with decreased TH17 cell development, lower expression of tissue-homing chemokine receptors and integrins, and  increased frequencies of Foxp3+ T regulatory (Treg) cells.  Importantly, inhibition of RORa with a selective small molecule antagonist largely phenocopied our genetic data, potently suppressing the in vivo development of both chronic/progressive and relapsing/remitting EAE but had no effect on overall thymic cellularity.  Furthermore, use of the RORa antagonist effectively inhibited human TH17 cell differentiation and memory cytokine secretion.  Together, these data suggest that RORa acts independently of RORgt in programming TH17 pathogenicity and identifies RORa as a safer and more selective therapeutic target for the treatment of TH17-mediated autoimmunity.

overall_design: 4 samples; 2 replicates per condition


title: Transcriptomic Analysis of Early Stages of Intestinal Regeneration in Holothuria glaberrima

city: San juan

first: David

last: Quispe

state: PR

address: 14, 2534 Av. Universidad Ste. 1401

department: Biology

country: Puerto Rico

institute: Universty of Puerto Rico

summary: We assembled and annotated a de novo transcriptome using RNA-seq data consisting of regenerating and non-regenerating intestinal tissues from the sea cucumber Holothuria glaberrima. Analysis of differential expression revealed a large number of differentially expressed transcripts associated with transcription factors with diverse expression patterns. Activity involving developmental and cancer-related events found in the analysis could be related to the regenerative process. Our study demonstrates the broad and diversified gene expression profile during the early stages of the process using the mesentery as the focal point of intestinal regeneration. It also establishes the genes that are the most important candidates in the cellular processes that underlie regenerative responses.

overall_design: Comparison of non-regenerating mesenteric tissues against 24 hour and 3 days regenerating intestine.

last: Quispe-Parra

first: Joshua

last: Medina-Feliciano

first: Sebastián

last: Cruz-González

first: Humberto

last: Ortiz-Zuazaga

first: José

last: García-Arrarás


title: RNA-seq profiling of malaria pre-exposed but resting effector memory γδ T cells

city: Parkville

first: Waruni

last: Abeysekera

state: Victoria

address: 1G Royal Parade

department: Bioinformatics

country: Australia

institute: Walter and Eliza Hall Institute of Medical Research

summary: Plasmodium chabaudi infected mice were drug-treated on day 14 post-infection with an i.p. injection of CQ (10mg/kg) and pyrimethamine (10mg/kg) followed by CQ (6 mg/ml) and pyrimethamine (70 mg/ml) containing water for 5 days. Spleens were removed 12 weeks after completion of drug treatment. Spleens from drug-treated naive mice were used as controls.

overall_design: Splenocytes were obtained and cells were surface stained ex vivo to identify CD62L-CD44+γδ T cells (EM). These EM γδ T cells were FACS sorted and total RNA was isolated followed by ClonTech RNA sequencing.

first: Emily

last: Eriksson


title: RNA-sequencing of QKI proficient and deficient microglia

city: Montreal

first: Stephane

last: Richard

address: 3755 Côte Ste-Catherine Road

country: Canada

institute: McGill University

summary:
Purpose:RNA_sequencing analysis defined a novel role for the QKI in microglia
Methods:  microglia mRNA profiles of 4-6 weeks-old wild-type (QKIfl/fl) and microglia specific knockout (QKIfl/fl; cx3r1cre-ert2) mice were generated by Illumina HiSeq 4000 in triplicate. The sequence reads that passed quality filters were trimmed with Trimmomatic v0.39. STAR v2.7.1a was then used to align the reads to the mouse genome (mm10/GRCm38). Gene expression was quantified across all samples with HOMER v4.11.1, and the normalization was carried out through the regularized logarithm (rlog) transformation of DESeq2 v1.26.0. Differential expression between the WT and  knockout samples were calculated through DESeq2 v1.26.0, and the gene expression was considered significantly different if the absolute value of the log-fold-change (LFC) was higher than 2, the base means larger than 10 and the false discovery rate (FDR) less than 0.05.
Results:  RNA-seq data revealed the up-regulation of 326 genes and down-regulation of 294 genes in knockout microglia compare to wt (2-fold change at FDR of 0.05, base mean>10). In particular qkICx3cr1-KO microglia exhibited higher expression of genes that are known to encode proteins related to inflammation (Il-6, Apoe, Cxcl-10, Il-1b, Il-12b, and Tnf). In line with this, functional annotation by DAVID, Reactome and, GO analysis showed enrichment of cell cycle and inflammation-related pathways in the up-regulated transcripts in qkICx3cr1-KO mice. Up-regulated transcripts were further validated using qRT-PCR.
Conclusions: Our study represents the first detailed analysis of microglia transcriptomes without QKI expression, with biologic replicates, generated by RNA-seq technology. The data should provide how QKI modulates the microglia gene expression profiles in genome-wide detail.

overall_design: QKI proficient and deficient microglia mRNA profile from adult cx3cr1 mice

first: Jeesan

last: Lee


title: Quantitative Analysis by Next Generation Sequencing of LSK (Lin- Sca1+ cKit+) hematopoietic progenitors transcriptomes from wild type and Usp15-/- mice.

first: Elisabetta

last: Citterio

summary:
Purpose: the goal of this study is to investigate the consequences of ubiquitin specific protease 15 (USP15) deletion on gene expression in mouse LSK hematopoietic progenitors.
Methods: mRNA profiles of 8 weeks-old wild-type (WT) and ubiquitin specific protease 15  knockout (Usp15−/−) mice were generated by deep sequencing using Illumina Hiseq2500. The sequence reads that passed quality filters  Alignments of the sequence reads that passed quality filters were performed using TopHat2.1, Genome build 38 and Ensembl gtf version 77 and genecounts have been generated using Itreecount. https://github.com/NKI-GCF/itreecount
Results: We assigned about 30 million reads per sample uniquely to a gene of the mouse reference genome (mm10) We identified 21,219 genes in the LSKs  of WT and Usp15−/− mice using TopHat2.1 in combination with Itreecount. https://github.com/NKI-GCF/itreecount. Distinct LSK-specific expressed genes (such as Eng, Tek, the MlI receptor and the Kit receptor) are identified.  Comparison of normalized gene expression data for Usp15-/- versus WT LSK confirmed the loss of Usp15. Apart from Usp15, almost no other significantly deregurated genes were detected using a treshold of fold change ≥1.5 and p value <0.05, suggesting the maintenance of an overall stable identity of the cellular compartment in Usp15-/- mice.
Conclusions: Our results represent the first detailed analyis of the consequences of USP15 deletion on gene expression in hematopoietic populations such as LSKs progenitors by genome wide expression profiling in WT and Usp15-/- mice. RNAseq of two freshly isolated biological replicas of sorted LSKs from 8 weeks old Usp15-/- animals confirmed the loss of Usp15, while showing almost no significant other up or down regulated genes among the 21,219 genes identified in Usp15-/- LSKs.  We conclude that young adult hematopoietic stem and progenitor cells (LSKs) perpetuated a stable gene expression program regardless of the homozygous deletion of USP15.

overall_design: mRNA profiles of 8 weeks-old wild type (WT) and Usp15-/- mice were generated by deep sequencing using Illumina Hiseq2500.  WT n=3, Usp15-/- n=2 mice were analized. FACS sorted LSK (Lin- Sca1+ cKit+) cells freshly isolated from the bone marrow of individual animals were subjected to deep sequencing.

first: Gaetano

last: Gargiulo

last: van den Berk

first: Iris

last: de Rink


title: A single master regulator controls asexual cell cycle division patterns in Toxoplasma gondii

city: Lille

first: Mathieu

last: Gissot

address: 1, rue du Pr. calmette

institute: CNRS

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

overall_design: Refer to individual Series


title: A single master regulator controls asexual cell cycle division patterns in Toxoplasma gondii [ChIP-Seq]

summary: ApiAP2 are a family of conserved transcription factors (TF) that play an important role in regulating gene expression in apicomplexan parasites.  ApiAP2 proteins that may control the cell cycle dependent expression program are to be discovered. To better understand how these cell-cycle dependent gene expression profiles are established, we identified ApiAP2 proteins showing a cell cycle dependent expression. Using the Auxin-degradation system, we produced several inducible knock-down (iKD) mutant for these ApiAP2 proteins. Among them, a cell-cycle regulated ApiAP2 is expressed during the late S-phase of the tachyzoite cell-cycle. The iKD mutant parasites for this gene are unable to proliferate. In presence of Auxin, the mutant parasites do not produce daughter cells and are arrested in the early phase of budding. This indicates that this TF may specifically regulate the early steps of the daughter cell formation. Using RNA-Seq, we demonstrate that the level of expression of number of transcripts is affected by the knock-down of this potential TF.

overall_design: Two biological replicates. Chromatin immunoprecipitation profiles  were generated by high throughput sequencing using HiSeq 2500 system (Illumina).

last: GISSOT

first: Cecilia

last: SANCHEZ

first: Kevin

last: LESAGE

first: Ludovic

last: HUOT


title: FoxO maintains a genuine quiescent muscle stem-cell state until geriatric age (RNA-seq)

city: Bethesda

first: Stephen

last: Brooks

state: MD

address: 50 SOUTH DR, RM 1140

department: Biodata Mining and Discovery Section

institute: NIAMS/NIH

summary: We identify two quiescent stem-cell states through relative CD34 expression: CD34High, with stemness properties (genuine state), and CD34Low, more committed to myogenic differentiation (primed state). The genuine-quiescent state is preserved into later life succumbing only in extreme old age due to acquisition of primed-state traits. We identified niche-derived IGF1-dependent Akt activation as detrimental to the genuine stem-cell state by imposing primed-state features via FoxO inhibition. Interventions to neutralize Akt and promote FoxO activity drive primed-to-genuine state conversion, while FoxO inactivation deteriorates the genuine state at young age, causing muscle regenerative failure, as in geriatric mice.

overall_design: Muscles form mice were mechanically disaggregated and dissociated in Ham’s F10 media containing Liberase (0.1mg/g muscle weight) (Roche; 5mg/mL) and Dispase 0.3% at 37 °C for 2 h and then filtered. Cells were then incubated in lysis buffer (BD Pharm Lyse) for 10 min on ice, re-suspended in PBS with 2% goat serum (FACS buffer) and counted. PE-Cy7-conjugated anti-CD45 (Biolegend 103114), anti-Sca-1 (Biolegend 108114) and PE-Cy7-conjugated anti-CD31 (Biolegend 102418) antibodies were used for lineage-negative selection. PE-conjugated anti-α7-integrin (AbLab AB10STMW215) and Alexa Fluor 647-conjugated anti-CD34 (BD Pharmigen 560230)  were used for double positive staining of quiescent satellite cells. Satellite cells were sorted using a FACS Aria II (BD) and processed for RNA-seq and ATAC-seq. This metadata sheet includes data from the 26 ATAC-Seq samples of this project.

first: Laura

last: García-Prat

first: Eusebio

last: Perdiguero

first: Sonia

last: Alonso-Martín

first: Stefania

last: Dell’Orso

first: Srikanth

last: Ravichandran

first: Aster

last: Juan

first: Silvia

last: Campanario

first: Kan

last: Jiang

first: Xiaotong

last: Hong

last: Ortet

first: Vanessa

last: Ruiz-Bonilla

first: Marta

last: Flández

first: Elena

last: Rebollo

first: Mercè

last: Jardí

first: Hong-Wei

last: Sun

first: Antonio

last: Musarò

first: Marco

last: Sandri

last: del Sol

first: Vittorio

last: Sartorelli

first: Pura

last: Muñoz-Cánoves


title: A single master regulator controls asexual cell cycle division patterns in Toxoplasma gondii [RNA-Seq]

first: Mathieu

last: Gissot

overall_design: Three biological replicate of the mutant strain in presence and in abscence of Auxin. mRNA profiles  iKD TgAP2IX-5 parasites was generated by high throughput sequencing using HiSeq 2500 system (Illumina).

last: GISSOT

first: Asma

middle: S

last: KHELIFA

first: Cecilia

last: GUILLEN SANCHEZ

first: Kevin

last: LESAGE


title: CNS-Native Myeloid Cells Drive Immune Suppression in the Brain Metastatic Niche through Cxcl10 [R10_PD-L1+VISTA treatment]

city: Dallas

first: Siyuan

last: Zhang

address: 6001 Forest Park Rd

department: Department of Pathology

institute: UT Southwestern Medical Center

summary: We performed CITE-seq (10x Genomics-based) to profile and compare the transcriptomes and cell surface expression of immune epitopes in the brains of anti-VISTA+anti-PD-L1 treated (Dual TX) and control, non-treated mice. We sequenced a total of eight  samples (4 control, 4 dual TX). We created an antibody pool consisting of 35 different antibodies and stained each sample individually with this antibody pool. Then, we stained each sample with it's own unique hashing antibody so that we could subsequently pool the samples for loading onto the 10x Chromium and later prepare one library consisting of all sight samples and finally separate each sample in silico by it's unique hashing antibody. The samples are as follows: (1) HTO1: Control/Non-treated (2) HTO2: Control/Non-treated ; (3) HTO3: Control/Non-treated ; (4) Control/Non-treated ; (5) HTO5: Dual TX; (6) Dual TX; (7) HTO7: Dual TX; (8) HTO8: Dual TX. The following sample comparisons were made: HTO1 - HTO4 were compared to HTO5 - HTO8.

overall_design: Metastasis-burdened brains were obtained from 2-3 month old female mice with or without anti-VISTA + anti-PD-L1 treatment (Dual TX). These mice were administered Dual TX every other day starting 3 days-post metastasis injection. Brain metastasis was established by carotid injection of E0771 cells. Cell suspensions were prepared by percoll gradient centrifugation enrichment to obtain suspensions enriched for immune cells from the brain. We created an antibody pool consisting of 35 different antibodies and stained each sample individually with this antibody pool. Then, we stained each sample with it's own unique hashing antibody so that we could subsequently pool the samples for loading onto the 10x Chromium and later prepare one library consisting of all eight samples and finally separate each sample in silico by it's unique hashing antibody.

first: Ian

last: Guldner


title: Innate immune function of galectin-1 as a DAMP

city: Farmington

first: Vijay

last: Rathinam

state: Connecticut

address: 263 Farmington Ave

department: Immunology

institute: UConn Health School of Medicine

summary: Galectin-1 is a prototype member of the galectin family of b-galactoside-binding proteins with several immunoregulatory activities (Sundblad et al., 2017). Although this lectin exerts extracellular activities by cross-linking cell surface glycoconjugates, the mechanisms and molecular machinery involved in its secretion are uncertain as it lacks the classical signal sequence required for ER-Golgi release (Camby, 2006; Croci et al., 2014). To assess the global impact of galectin-1 on LPS-induced transcriptomic response, we performed RNAseq analysis of total RNAs from the spleen and the lungs of wild-type and Lgals1-/- mice injected with LPS.

overall_design: Transcriptome profiling of wild-type or Lgals1-/- mice treated with LPS (5 mg/kg for 8 h)

first: Beiyan

last: Zhou


title: CNS-Native Myeloid Cells Drive Immune Suppression in the Brain Metastatic Niche through Cxcl10 [R07_Cx3cr1]

summary: We performed CITE-seq (10x Genomics-based) to profile and compare the transcriptomes and cell surface expression of immune epitopes in the brains of Cx3cr1+/- and Cx3cr1-/- mice during homeostasis or brain metastasis. We sequenced a total of eight different samples. We created an antibody pool consisting of 35 different antibodies and stained each sample individually with this antibody pool. Then, we stained each sample with it's own unique hashing antibody so that we could subsequently pool the samples for loading onto the 10x Chromium and later prepare one library consisting of all sight samples and finally separate each sample in silico by it's unique hashing antibody. The samples are as follows: (1) HTO1: brain of naïve Cx3cr1+/- mouse; (2) HTO2: brain of naïve Cx3cr1+/- mouse; (3) HTO3: brain of metastasis-burdened Cx3cr1+/- mouse; (4) HTO4: brain of metastasis-burdened Cx3cr1+/- mouse; (5) HTO5: brain of naïve Cx3cr1-/- mouse; (6) HTO6: brain of naïve Cx3cr1-/- mouse; (7) HTO7: brain of metastasis-burdened Cx3cr1-/- mouse; (8) HTO8: brain of metastasis-burdened Cx3cr1-/- mouse. The following sample comparisons were made: HTO1 and HTO2 versus HTO5 and HTO6; HTO3 and HTO4 versus HTO7 and HTO8.

overall_design: Naïve brains were obtained from 2-3 month old female Cx3cr1+/- or Cx3cr1-/- mice without brain metastases. Brains from brain metastasis-bearing mice initiated by carotid injection of E0771 cells were obtained from 2-3 month old female mice of the aforementioned genotypes with established brain metastases (~2 weeks post injection). Cell suspensions were prepared by percoll gradient centrifugation enrichment to obtain suspensions enriched for immune cells from the brain. We created an antibody pool consisting of 35 different antibodies and stained each sample individually with this antibody pool. Then, we stained each sample with it's own unique hashing antibody so that we could subsequently pool the samples for loading onto the 10x Chromium and later prepare one library consisting of all eight samples and finally separate each sample in silico by it's unique hashing antibody.


title: SNP array for Cas9 treated human oocytes and embryos

city: North Brunswick Township

first: Jia

last: Xu

state: NJ

address: 675 US Highway One

institute: Genomic Prediction, Inc.

summary: Human eggs were fertilized with sperm carrying a homozygous mutation in the EYS gene at rs758109813 and treated with a Cas9 RNP targeting this mutation at either fertilization or at the 2-cell stage. The purpose of this analysis was to determine loss of heterozygosity  due to CRISPR-induced chromosomal changes. We find that However, in half of the embryos, the break remains unrepaired throughout the first cell cycle, resulting in both segmental, as well as whole chromosome loss. These embryos appear as corrected non-mosaic wild type embryos in an on-target sequence analysis, but are not viable. These results show a surprising tolerance of the human embryo for mitotic entry with unrepaired DNA double strand breaks, and suggest that pericentromeric cleavage by Cas9 may enable the allele-specific removal of chromosomes in trisomic embryos.

first: Dieter

last: Egli

first: Nathan

last: Treff


title: CNS-Native Myeloid Cells Drive Immune Suppression in the Brain Metastatic Niche through Cxcl10 [R08_MgliaDep]

summary: We performed CITE-seq (10x Genomics-based) to profile and compare the transcriptomes and cell surface expression of immune epitopes in the brains and blood of different transgenic mice (Cx3cr1CreERT; Cx3cr1CreERT-ROSA-iDTR) during homeostasis or brain metastasis with or without DT-based microglia depletion. We sequenced a total of eight different samples. We created an antibody pool consisting of 35 different antibodies and stained each sample individually with this antibody pool. Then, we stained each sample with it's own unique hashing antibody so that we could subsequently pool the samples for loading onto the 10x Chromium and later prepare one library consisting of all sight samples and finally separate each sample in silico by it's unique hashing antibody. The samples are as follows: (1) HTO1: brain of naïve Cx3cr1CreERT mouse (non-microglia depleted); (2) HTO2: brain of metastasis-burdened  Cx3cr1CreERT mouse (non-microglia depleted); (3) HTO3: brain of metastasis-burdened  Cx3cr1CreERT mouse (non-microglia depleted); (4) HTO4: brain of naïve Cx3cr1CreERT-ROSA-iDTR mouse (microglia depleted); (5) HTO5: brain of metastasis-burdened  Cx3cr1CreERT-ROSA-iDTR mouse (microglia depleted); (6) HTO6: brain of metastasis-burdened  Cx3cr1CreERT-ROSA-iDTR mouse (microglia depleted); (7) HTO7: blood of metastasis-burdened Cx3cr1CreERT mouse (non-microglia depleted); (8) HTO8: blood of metastasis-burdened Cx3cr1CreERT-ROSA-iDTR mouse (microglia depleted). The following sample comparisons were made: HTO1 versus HTO4; HTO2 and HTO3 versus HTO5 and HTO6; HTO7 versus HTO8.

overall_design: Naïve brains were obtained from 2-3 month old female Cx3cr1CreERT or Cx3cr1CreERT-ROSA-iDTR mice without brain metastases. These mice were administered tamoxifen followed by a one month cessation before being administered diptheria toxin, which induced microglia depletion in Cx3cr1CreERT-ROSA-iDTR, but not control Cx3cr1CreERT mice.  Brains from brain metastasis-bearing mice initiated by carotid injection of E0771 cells were obtained from 2-3 month old female mice of the aforementioned genotypes with established brain metastases (~2 weeks post injection). These mice also underwent the aforementioned tamoxifen and diptheria toxin treatment to deplete microglia during brain metastasis. Cell suspensions were prepared by percoll gradient centrifugation enrichment to obtain suspensions enriched for immune cells from the brain. We created an antibody pool consisting of 35 different antibodies and stained each sample individually with this antibody pool. Then, we stained each sample with it's own unique hashing antibody so that we could subsequently pool the samples for loading onto the 10x Chromium and later prepare one library consisting of all eight samples and finally separate each sample in silico by it's unique hashing antibody. NOTE: Brain immune cells were enriched through percoll gradient before 10X sequencing. Thus, there are remanent microglia cells in the depleted samples.


title: Effect of mast cells on the phenotype of breast cancer cells

city: Milan

first: Matteo

last: Dugo

address: via Olgettina 60

department: Department of Medical Oncology

country: Italy

institute: IRCCS Ospedale San Raffaele

summary: To understand the mechanisms by which mast cells affect the features of breast cancer cells, we collected spontaneous mammary tumors from MMTV-PyMT mice, C57BL/6 wild type or back-crossed with Wsh mice which lack mast cells. Gene expression profiling was performed to select the genes which are differentially expressed in the presence or absence of mast cells.

overall_design: Gene expression profiles were generated for 8 spontaneous mammary tumors from MMTV-PyMT transgenic C57BL/6 mice and 7 tumors from MMTV-PyMT Wsh mice at 20 weeks of age

first: Daniele

last: Lecis

first: Maria

middle: T

last: Majorini

first: Loris

last: De Cecco


title: Targeted high-resolution chromosome conformation capture at genome-wide scale in mouse erythroid cells

status: Public on Oct 28 2020

city: Headington

first: Damien

last: Downes

address: John Radcliffe Hospital

department: MRC Weatherall Institute of Medicine

institute: The University of Oxford

summary: Chromosome conformation capture (3C) provides an adaptable tool through which to study diverse biological questions. Currently, 3C techniques provide either low-resolution interaction profiles across the entire genome, e.g. HiC, or high-resolution interaction profiles at up to several hundred loci, e.g. NG Capture-C and 4C-seq. Generation of high-resolution, genome-wide interaction profiles can feasibly be achieved through efficiency improvements to current high-resolution methods. To this end we systematically tested and removed areas inefficiency in NG Capture-C to develop a new method Nuclear Capture-C, which provides a 300% increase in informative sequencing content. Using Nuclear Capture-C we target 8,026 erythroid promoters in triplicate, showing that this method can achieve high-resolution genome-wide 3C interaction profiles at scale.

overall_design: Nu Capture-C was carried out in triplicate with DpnII for ter119+ mouse erythroid cells from F1 hybrid mice (C57BL/6-cross-CBA/J)

first: Jim

last: Hughes


title: CNS-Native Myeloid Cells Drive Immune Suppression in the Brain Metastatic Niche through Cxcl10

summary: We used CITE-seq (10x Gemoics-based) to profile and compare the transcriptomes and cell surface expression of a set of immune markers of naïve brain myeloid cells from wild type C57Bl/6 mice to brain metastasis-associated myeloid cells (Br.MAM) from C57Bl/6 mice bearing E0771 brain metastases. In each sequencing round, we sequenced a total of four different mouse brains (2 naive and 2 metastasis-burdened). We created an antibody pool consisting of 6 different antibodies (CD45, CD25, CD3, CD4, CD11b, and CD86) and stained each brain individually with this antibody pool. Then, we stained each brain with it's own unique hashing antibody so that we could subsequently pool the samples for loading onto the 10x Chromium and later prepare one library consisting of all six samples and finally separate each sample in silico by it's unique hashing antibody.

overall_design: Naïve brains were obtained from 2-3 month old female C57Bl/6 mice without brain metastases. Brains bearing metastases initiated by carotid injection of E0771 cells were obtained from 2-3 month old female mice (C57Bl/6) with established brain metastases (~2 weeks post injection). Cell suspensions were prepared by percoll gradient centrifugation enrichment to obtain suspensions enriched for immune cells. We created an antibody pool consisting of 6 different antibodies (CD45, CD25, CD3, CD4, CD11b, and CD86) and stained each brain individually with this antibody pool. Then, we stained each brain with it's own unique hashing antibody so that we could subsequently pool the samples for loading onto the 10x Chromium and later prepare one library consisting of all six samples and finally separate each sample in silico by it's unique hashing antibody.


title: Lung transplantation for patients with severe COVID-19

city: Chicago

first: Alexander

middle: V

last: Misharin

address: 303 E Superior

institute: Northwestern University

summary: Lung transplantation can potentially be a life-saving treatment for patients with non-resolving COVID-19-associated respiratory failure. Concerns limiting transplant include recurrence of SARS-CoV-2 infection in the allograft, technical challenges imposed by viral-mediated injury to the native lung, and potential risk for allograft infection by pathogens associated with ventilator-associated pneumonia in the native lung. Most importantly, the native lung might recover, resulting in long-term outcomes preferable to transplant. Here, we report results of the first successful lung transplantation procedures in patients with non-resolving COVID-19-associated respiratory failure in the United States. We performed sm-FISH to detect both positive and negative strands of SARS-CoV-2 RNA in the explanted lung tissue, extracellular matrix imaging using SHIELD tissue clearance, and single cell RNA-Seq on explant and warm post-mortem lung biopsies from patients who died from severe COVID-19 pneumonia. Lungs from patients with prolonged COVID-19 were free of virus but pathology showed extensive evidence of injury and fibrosis which resembled end-stage pulmonary fibrosis. We used a machine learning approach to project single cell RNA-Seq data from patients with late stage COVID-19 onto a single cell atlas of pulmonary fibrosis, revealing similarities across cell lineages. There was no recurrence of SARS-CoV-2 or pathogens associated with pre-transplant ventilator associated pneumonias following transplantation. Our findings suggest that some patients with severe COVID-19 develop fibrotic lung disease for which lung transplantation is the only option for survival.

overall_design:
Distal lung biopsies were obtained from the explanted lung and donor lung from Case 1, and the two post-mortem biopsies. In addition, a biopsy that included the main left bronchus and distal parenchyma from the upper lobe was obtained from another donor lung that was not placed for transplant. Single-cell suspensions from the biopsies were incubated with CD31 beads, and the positive fraction was selected for single-cell. The negative fraction was FACS-sorted for macrophages, epithelial and stromal cells. Single-cell libraries were prepared with 10x Genomics 3' V3 chemistry. Libraries were pooled and sequenced on Illumina HiSeq 4000 or NovaSeq 6000. Gene expression matrices were generated with 10x cellranger v3.1.0 software using custom reference genome of GRCh38.93 and SARS-CoV-2 (NC_045512.2).
****RNA-Seq raw data will be made available through dbGaP (controlled access) due to patient privacy concerns.

first: Ankit

last: Bharat

first: Melissa

last: Querrey

first: Nikolay

last: Markov

first: Samuel

last: Kim

first: Chitaru

last: Kurihara

first: Rafael

last: Garza-Castillon

first: Adwaiy

last: Manerikar

first: Ali

last: Shilatifard

first: Rade

last: Tomic

first: Yuliya

last: Politanska

first: Hiam

last: Abdala-Valencia

first: Anjana

last: Yeldandi

first: Jon

middle: W

last: Lomasney

first: GR

last: Budinger


title: Transcriptome analysis in primary cell models of HIV latency and cells from HIV-infected ART-suppressed individuals

city: San Framcisco

first: Steven

middle: A

last: Yukl

address: 4150 Clement Street

institute: San Francisco VA Medical Center - University of California San Francisco

summary: Comparison of total RNA-seq data from ex vivo unstimulated and stimulated (with anti-CD3/CD28) cells from two primary cell models of HIV latency (resting-cell and wild-type virus models) and peripheral CD4+ T cells from HIV-infected ART-suppressed individuals (ex vivo cells). Two donors were analyzed per model.

overall_design: Total RNA-seq data from ex vivo unstimulated and stimulated (with anti-CD3/CD28) primary CD4+ T cells from the resting-cell model (Lassen et al 2012), the wild-type virus model (Martins et al. 2016) and HIV-infected ART-suppressed individuals were compared (n=2 per condition).


title: CNS-Native Myeloid Cells Drive Immune Suppression in the Brain Metastatic Niche through Cxcl10 [R07_CCR2Cx3cr1B]

first: Siyuan

last: Zhang

summary: We performed CITE-seq (10x Genomics-based) to profile and compare the transcriptomes and cell surface expression of immune epitopes in the brains and blood of different transgenic mice (Cx3cr1+/-; Cx3cr1-/-; CCR2+/-; CCR2-/-) during homeostasis or brain metastasis. We sequenced a total of eight different samples. We created an antibody pool consisting of 35 different antibodies and stained each sample individually with this antibody pool. Then, we stained each sample with it's own unique hashing antibody so that we could subsequently pool the samples for loading onto the 10x Chromium and later prepare one library consisting of all sight samples and finally separate each sample in silico by it's unique hashing antibody. The samples are as follows: (1) HTO1: brain of naïve CCR2+/- mouse; (2) HTO2: brain of naïve  CCR2+/- mouse; (3) HTO3: brain of naïve CCR2-/- mouse; (4) HTO4: brain of naive CCR2-/-; (5) HTO5: blood of naïve Cx3cr1+/- mouse; (6) HTO6: blood of metastasis-burdened Cx3cr1+/- mouse; (7) HTO7: blood of naïve Cx3cr1-/- mouse; (8) HTO8: blood of metastasis-burdened Cx3cr1-/- mouse.The following sample comparisons were made: HTO1 and HTO2 versus HTO3 and HTO4; HTO5 versus HTO7; HTO6 versus HTO8.

overall_design: Naïve brains and blood were obtained from 2-3 month old female Cx3cr1+/-, Cx3cr1-/-, CCR2+/-, or CCR2-/- mice without brain metastases. Blood from brain metastasis-bearing mice initiated by carotid injection of E0771 cells were obtained from 2-3 month old female mice of the aforementioned genotypes with established brain metastases (~2 weeks post injection). Cell suspensions were prepared by percoll gradient centrifugation enrichment to obtain suspensions enriched for immune cells from the brain, and whole blood was RBC-lysed and then pelleted for peripheral blood samples. We created an antibody pool consisting of 35 different antibodies and stained each sample individually with this antibody pool. Then, we stained each sample with it's own unique hashing antibody so that we could subsequently pool the samples for loading onto the 10x Chromium and later prepare one library consisting of all eight samples and finally separate each sample in silico by it's unique hashing antibody.

first: Ian

last: Guldner


title: Ascending colon-derived mucin-type O-glycans form key mucus layers encapsulating microbiota in the colon

city: Los Angeles

first: David

last: Casero

address: 610 Charles Young Drive East

department: Pathology and Laboratory Medicine

institute: UCLA

sample_organism:
Mus musculus
mouse gut metagenome

platform_organism:
Mus musculus
mouse gut metagenome


title: Ascending colon-derived mucin-type O-glycans form key mucus layers encapsulating microbiota in the colon (16S rNA-seq)

summary: The colon contains a dense metabolically potent microbiota. The colonic O-glycan-rich mucus has been recognized as a key barrier to prevent microbial intrusion, but how this system forms and functions remains unclear. Here, we discovered that the colon mucus is mainly forged by microbiota-dependent secretion of O-glycosylated Muc2 by goblet cells in the ascending colon, where it seamlessly encapsulates the fecal materials including the microbiota. Deletion of O-glycans in the ascending colon impaired the segregating function of the mucous coating, leading to altered structure and metabolic output of the microbiota, and transcriptional homeostasis of the entire host mucosa. These findings represent a paradigm change of the prevailing model of the colon mucus system and provides new insights into host and microbiota symbiosis.

overall_design: 16S amplicon sequencing was performed on mouse colon  samples derived from the ascending and descending colon. Differential microbial abundance was compared between 3 Groups of Healthy Control and O-glycan mutant littermates that were all colonized with the same microbiota

first: Kirk

last: Bergstrom

first: Xindi

last: Shan

first: Albert

last: Batushansky

first: Venu

last: Lagishetty

first: Jonathan

last: Jacobs

first: Christopher

last: Hoover

first: J

middle: M

last: McDaniel

last: McGee

first: Courtney

last: Houchen

first: CV

last: Rao

first: Tim

last: Griffin

first: Justin

middle: L

last: Sonnenburg

first: Rodger

last: McEver

last: Braun

first: Lijun

last: Xia


title: The landscape of RNA Pol II binding reveals a step-wise transition during ZGA

city: Beijing

first: Bofeng

last: Liu

address: Haidian District

department: School of Life Science

institute: Tsinghua University

summary: Zygotic genome activation (ZGA) is the first transcription event in life. However, it is unclear how RNA polymerase is engaged in initiating ZGA in mammals. Here, by developing small-scale Tn5-assisted chromatin cleavage with sequencing (Stacc–seq), we investigated the landscapes of RNA polymerase II (Pol II) binding in mouse embryos. We found that Pol II undergoes ‘loading’, ‘pre-configuration’, and ‘production’ during the transition from minor ZGA to major ZGA. After fertilization, Pol II is preferentially loaded to CG-rich promoters and accessible distal regions in one-cell embryos (loading), in part shaped by the inherited parental epigenome. Pol II then initiates relocation to future gene targets before genome activation (pre-configuration), where it later engages in full transcription elongation upon major ZGA (production). Pol II also maintains low poising at inactive promoters after major ZGA until the blastocyst stage, coinciding with the loss of promoter epigenetic silencing factors. Notably, inhibition of minor ZGA impairs the Pol II pre-configuration and embryonic development, accompanied by aberrant retention of Pol II and ectopic expression of one-cell targets upon major ZGA. Hence, step-wise transition of Pol II occurs when mammalian life begins, and minor ZGA has a key role in the pre-configuration of transcription machinery and chromatin for genome activation.

overall_design: Stacc-seq and RNA-seq were applied on mouse oocytes and pre-implantation embryos. All wild-type pre-implantation embryos were collected by crossing C57BL/6N female mice with PWK male mice.

first: Qianhua

last: Xu

first: Qiujun

last: Wang

first: Wei

last: Xie


title: Survival of cells with deregulated MYC requires UVSSA-dependent regulation of RNAPII dynamics

city: NewYork

first: Zhaoqi

state: New York

address: ICRC Building 8th Floor, Office 803B 1130 St. Nicholas Ave

department: Systems Biology

institute: Columbia university

summary: Cancer cells develop strong genetic dependencies enabling survival under oncogenic stress. MYC is a key oncogene activated across most cancers, and identifying associated synthetic lethality can provide important clues about its activity and potential therapeutic strategies. Based on previously conducted genome-wide screenings we identified UVSSA, a gene involved in transcription-coupled repair whose knockdown decreased cell viability when combined with MYC activation. Synthetic lethal interactions between MYC expression and UVSSA loss correlated with ATM/CHK2 activation, suggesting increased genome instability. We show that although the synthetic lethal interaction was diminished by attenuating RNA polymerase II (RNAPII) activity, it becomes independent of UV-induced damage repair, suggesting that UVSSA has a critical function in regulating transcription in the absence of exogenous DNA damage. Supporting this hypothesis, RNAPII-ChIP-seq revealed that MYC-dependent increase in RNAPII promoter occupancy, as well as RNAPII stalling, is reduced or abrogated by UVSSA knockdown, suggesting that UVSSA negatively regulates MYC-dependent transcription. Taken together, our data shows that the UVSSA complex is required to limit MYC-dependent transcription stress and to maintain cell survival. While the role of UVSSA in regulating RNAPII dynamics has only been documented thus far in the context of UV-induced DNA damage repair, we propose that its activity is also required to cope with transcription stress induced by oncogene activation.

overall_design: Examination of chromatin-bound RNAPII levels at actively transcribing genes under 4 conditions: CONTROL, MYC-ER ONLY, shUVSSA ONLY, and MYC+shUVSSA.

first: Mai

last: Sato


title: Pharmacological reversal of synaptic and network pathology in human MECP2-KO neurons and cortical organoids

status: Public on Oct 27 2020

city: La Jolla

first: Alysson

middle: R.

last: Muotri

address: 2880 Torrey Pines Scenic Road MC 0695

institute: UNIVERSITY OF CALIFORNIA, SAN DIEGO

summary: Duplication or deficiency of the X-linked MECP2 gene reliably produces profound neurodevelopmental impairment. MECP2 mutations are almost universally responsible for Rett syndrome (RTT), and particular mutations and cellular mosaicism of MECP2 may underlie the spectrum of RTT symptomatic severity. No clinically approved treatments for RTT are currently available, but human pluripotent stem cell technology offers a platform to identify neuropathology and test candidate therapeutics. Using a strategic series of increasingly complex human stem cell-derived technologies, including human neurons, MECP2-mosaic neurospheres to model RTT female brain mosaicism, and cortical organoids, we identified synaptic dysregulation downstream from knockout of MECP2 and screened select pharmacological compounds for their ability to treat this dysfunction. Two lead compounds, Nefiracetam and PHA 543613, specifically reversed MECP2-knockout cytologic neuropathology. The capacity of these compounds to reverse neuropathologic phenotypes and networks in human models supports clinical studies for neurodevelopmental disorders in which MeCP2 deficiency is the predominant etiology.

overall_design: Examination of the impact of Nefiracetam and PHA 543613 on the recovery of MECP2-knockout cytologic neuropathology

first: Cleber

last: Trujillo

first: Jason

last: Adams

first: Priscilla

last: Negraes

first: Cassiano

last: Carromeu

first: Leon

last: Tejwani

first: Allan

last: Acab

first: Ben

last: Tsuda

first: Charles

last: Thomas

first: Neha

last: Sodhi

first: Katherine

last: Fichter

first: Sarah

last: Romero

first: Fabian

last: Zanella

first: Terrence

last: Sejnowski

first: Henning

last: Ulrich


title: FoxO maintains a genuine quiescent muscle stem-cell state until geriatric age

first: Stephen

last: Brooks

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

overall_design: Refer to individual Series


title: Evolution of advanced chronic lymphoid leukaemia unveiled by single-cell transcriptomics: a case study

city: Vienna

first: Iros

last: Barozzi

address: Borschkegasse 8a

country: Austria

institute: Medical University Vienna

summary: Characterization of the transcriptional changes occurring at the single-cell level during the progression of a chronic lymphoid leukaemia treated with Rituximab, Fludarabine and Cyclophosphamide.

overall_design: Single-cell RNA-seq library from peripheral blood cells of a subject with chronic lymphoid leukaemia; samples at diagnosis and relapse were separatly barcoded and pooled; single-cell captured using 10x Chromium and subjected to high-throughput sequencing.

first: I

first: P

last: Ostasov

first: V

last: Caputo

first: A

last: Datta

last: Piazza

first: M

last: Holubova


title: Enhanced crosslinking and immunoprecipitation of SERBP1 in Hela cells

city: Wuhan

first: Jinglei

last: Xu

state: Hubei

address: Bayi road

department: The Institute of Advanced Studies

institute: Wuhan University

summary:
Purpose:We used eCLIP experiment to analyze binding profile of SERBP1 and its relationship with G4.
Result: we found 505 high-confidence SERBP1 binding sites in Hela cells, and the analysis of sequence revealed transcriptome-wide enrichment of PQS in SERBP1-binding regions

overall_design: We first established a Hela cell line that stably expressed SERBP1,  and then performed eCLIP experiments of SERBP1

first: Haomiao

last: Su

first: Xiang

last: Zhou


title: Widespread transcriptional readthrough caused by Nab2 depletion leads to chimeric transcripts with retained introns

city: New Haven

first: Tara

middle: Diane

last: Alpert

state: CT

address: 333 Cedar St.

department: Molecular Biophysics and Biochemistry

institute: Yale University

summary: Nascent RNA sequencing has recently revealed that pre-mRNA splicing can occur shortly after the intron emerges from RNA polymerase II (Pol II). Differences in co-transcriptional splicing profiles suggest regulation by cis- and/or trans-acting factors. Here we used Single Molecule Intron Tracking (SMIT) in budding yeast to identify a cohort of regulators by machine learning. One candidate, Nab2, displayed reduced co-transcriptional splicing of some pre-mRNAs when depleted. Unexpectedly, these splicing defects were attributable to readthrough transcription, which was revealed by long read sequencing of nascent RNA; individual readthrough transcripts induced by Nab2 depletion sometimes spanned multiple genes. Thus, Nab2 regulation of splicing was indirect. Moreover, unspliced transcripts displayed downstream readthrough in both control and Nab2-depleted cells, highlighting the coupling between splicing and 3′ end formation. We conclude that Nab2 is required for proper 3′ end processing, which ensures both transcription termination as well as proper splicing of downstream genes.

overall_design:
Single Molecule Intron Tracking (SMIT) was performed for 2 replicates each of 6 different yeast deletion strains as well as the wild type control. Additional SMIT libraries were prepared for Anchor-Away strains (Nab2-AA and Control-AA). Finally, long read sequencing libraries were prepared from nascent RNA extracted from the Nab2-AA and Control-AA samples in duplicate.

Please note that the 'Nab2AA_ControlAA_fast5.tar.gz' (linked to the Long read Nab2-AA rep1 sample records) contains raw data for all four "Long read" samples.  As the algorithm for converting and demultiplexing the fast5 into fastq files is proprietary software and not all users will have access to the exact form/version used, both fastq and fast5 data were made available as SRA and GEO records, respectively.

first: Korinna

last: Straube

first: Fernando

last: Carrillo Oesterreich

first: Karla

last: Neugebauer


title: Identification of novel MeCP2 cancer-associated target genes and post-translational modifications

city: Lubbock

first: Kevin

last: Pruitt

state: Texas

address: 3601 4th Street

department: Immunology & Molecular Microbiology

institute: Texas Tech University Health Sciences Center

summary: The goal of this study is to identify novel target genes of MeCP2 in two breast cancer cell lines.

overall_design:
RNA-seq: MDA-MB-468 non-target control (WT) and MecP2 KD (sh1 and sh3).
ChIP-seq: MeCP2 ChIP-Seq in MDA-MB-468 and MCF7 cell lines.


title: Importance of RAD6B in activation of canonical WNT signaling in melanoma development

city: Detroit

first: Malathy

middle: P

last: Shekhar

state: MI

address: 421 East Canfield Avenue

department: Oncology

institute: Wayne State University

summary: We have demonstrated that RAD6B, a ubiquitin conjugating enzyme that is a principal component of translesion synthesis pathway, stabilizes and transcriptionally activates beta-catenin by lysine 63-linked polyubiquitination. RAD6B is also a beta-catenin transcriptional target, triggering a positive feedback loop. Canonical WNT signaling is critical for melanocyte lineage commitment and melanoma development, and RAD6B and beta-catenin are co-expressed in primary and metastatic melanomas. To understand how RAD6B influences melanoma development/progression, we performed RNA-seq of human metastatic M14 melanoma cells, their RAD6B knockout counterpart, and human HEMa-LP normal melanocytes.

overall_design: 9 samples - Control M14 (3 samples each run in duplicates), RAD6BKOM14 (3 samples each in duplicates), HEMa-LP (3 samples)


title: RNAseq analyses of copper effect on Bordetella pertussis and Bordetella bronchiseptica

city: Lille

first: Rudy

last: Antoine

address: 1, rue du Professeur Calmette

institute: CIIL

summary: Copper is both essential and toxic to living beings, which therefore tightly control its intracellular concentration. At the host-pathogen interface, copper is used by phagocytic cells to kill invading microorganisms. We investigated copper homeostasis in the whooping cough agent Bordetella pertussis, which lives in the human respiratory mucosa and has no environmental reservoir. B. pertussis has considerably streamlined copper homeostasis mechanisms relative to other Gram-negative bacteria. Its single remaining defense line against copper intoxication consists in a metallochaperone diverted for copper passivation and two enzymes involved in peroxide detoxification, which together fight two stresses encountered in phagolysosomes. The three proteins are encoded by an original, composite operon assembled in an environmental ancestor and which is under sensitive control by copper. Interestingly, this system appears to play a role in persistent infection in the nasal cavity of B. pertussis-infected mice. Combining responses to co-occurring stresses in a tailored operon reveals a new strategy adopted by a host-restricted pathogen to optimize survival at minimal energy expenditure.

sample_organism:
Bordetella bronchiseptica
Bordetella pertussis

overall_design: NextSeq 500 sequencing of RNA samples obtained in different Bordetella pertussis and Bordetella bronchiseptica culture conditions

first: Alex

last: Rivera-Millot

first: Stéphanie

last: Slupek

first: Jonathan

last: Chatagnon

first: Gauthier

last: Roy

first: Jean-Michel

last: Saliou

first: Gabriel

last: Billon

first: Véronique

last: Alaimo

first: David

last: Hot

first: Sophie

last: Salomé-Denoulez

first: Camille

last: Locht

first: Françoise

last: Jacob-Dubuisson


title: Gene expression changes in atopic dermatitis after treatment with nb-UVB

first: Astrid

middle: Haaskjold

last: Lossius

state: Oslo

address: Sognsvannsveien 20

department: Dermatology/Pathology

country: Norway

institute: University of Oslo

summary:
Background: Atopic dermatitis (AD) is a common inflammatory skin disease with broad impact on quality of life and on the health care system. The pathophysiology is not fully understood, but it is likely multifactorial involving immune dysfunction, altered skin barrier and environmental factors. Narrow band ultraviolet B (nb-UVB) treatment leads to normalization of the tissue and clinical improvement. However, knowledge of early changes in AD skin in response to nb-UVB is lacking and could provide important clues to decipher the disease mechanisms and potential new treatment targets.

Objective: To map the early transcriptional changes in the skin in response to nb-UVB treatment.

Results: When examining the early response after only three local UVB-treatments, gene expression analysis revealed 30 down- and 47 up-regulated transcripts. Among these only a small proportion were related to the inflammatory response. Interestingly, two cytokines of the interleukin (IL)-1 family were differentially expressed: the proinflammatory cytokine IL-36γ was reduced after treatment, while the anti-inflammatory cytokine IL-37 increased in skin after treatment with nb-UVB.

Conclusion: Local nb-UVB induced an early decrease of the pro-inflammatory cytokine IL-36γ and an increase of the anti-inflammatory IL-37. This likely represents one of the first changes in inflammatory signaling induced by nb-UVB in atopic eczema.

overall_design:
Adult patients (n = 16) with mild to moderate AD were included in the study. We performed skin biopsies of patients with AD before and after three treatments of local nb-UVB. The biopsies were analyzed for differences in gene expression with microarrays (Affymetrix, Clariom S).
1 & 4: untreated lesional skin; 2: untreated non-lesional skin; 3: nb-UVB x 3, 5: 6-8 weeks of treatment, lesional skin; 6: 6-8 weeks of treatment, non-lesional skin
The biopsies were analyzed for differences in gene expression with microarrays (Affymetrix, Clariom S).

first: Jan-Øivind

last: Holm

first: Guttorm

last: Haraldsen

first: Teresa

last: Berents

first: Olav

last: Sundnes


title: IN SITU DIFFERENTIATION OF IRIDOPHORE CRYSTALLOTYPES UNDERLIES ZEBRAFISH STRIPING

city: Ashburn

first: Dvir

last: Gur

state: VA

address: 19700 Helix Dr

institute: HHMI Janelia Research Campus

summary: Skin color patterns are ubiquitous in nature, evolve rapidly, and impact social behavior, predator avoidance, and protection from ultraviolet irradiation. A leading model system for vertebrate skin patterning is the zebrafish; its alternating blue stripes and yellow interstripes depend on guanine crystal-containing cells called iridophores that reflect light. It was suggested that the zebrafish’s alternating color pattern arises from a single type of iridophore migrating differentially to stripes and interstripes. When we tracked iridophores, however, we found they did not migrate between stripes and interstripes but instead differentiated and proliferated in place based on their micro-environment. RNA-sequencing analysis further revealed that stripe and interstripe iridophores had different transcriptomic states with many differences in gene expression and pathway enrichment, while cryogenic scanning electron microscopy and micro-X-ray diffraction identified different guanine crystal organizations and responsiveness to norepinephrine, all indicating that stripe and interstripe iridophores are different cell types. Based on these results, we present a new model of skin patterning in zebrafish in which distinct iridophore crystallotypes containing specialized, physiologically responsive, subcellular organelles arise in stripe and interstripe zones by in situ differentiation. In this model, pattern phenotype depends not only on interactions among pigment cells that affect their arrangements, but also on factors that specify subcellular organization and physiological responsiveness of specialized organelles.

overall_design: Unbiased classification of iridophores, based solely on their RNA profiles obtained by single-cell RNA-sequencing

first: Jennifer

last: Lippincott-Schwartz


title: LFY is pioneer transcription factor

city: Philadelphia

first: Doris

last: Wagner

state: PA

address: 103G Carolyn Lynch Laboratory

department: Department of Biology

institute: University of Pennsylvania


title: ChIP-seq identified the regulation of genes by the quorum-sensing regulator OpaR in Vibrio parahaemolyticus

city: Taipei

first: San-Chi

last: Chang

address: 1 Sec. 4, Roosevelt Road

department: Department of Agricultural Chemistry

country: Taiwan

institute: National Taiwan University

summary: We report that ChIP-seq analyses of the quorum-sensing regulator OpaR at late-log and stationary phase in Vibrio parahaemolyticus

overall_design:
We inserted a FLAG-tag before the opaR gene on chromosome. FLAG-OpaR/VP93 was grown to a late-log phase (OD600 3) and stationary phase (OD600 4.2). DNA fragments were sheared by Bioruptor Pico (Diagenode). Anti-FLAG antibody beads (Sigma) were used to enrich the IP DNA fragments. ChIP DNA was then purified using the Agencourt AMPure XP PCR Purification Kit (Beckman Coulter).

DOI: 10.3389/fmicb.2020.534692

first: Chia-Yin

last: Lee


title: Transcriptional dynamics of iPSC differentiation into β cells revealed by single-cell RNA sequencing

city: Milan

first: Giulia

middle: Maria

last: Scotti

state: Milan

address: Viale Rita Levi Montalcini, 1

department: National Facility for Data Handling and Analysis

institute: Fondazione Human Technopole

summary: Induced pluripotent stem cells can be differentiated in vitro into insulin-producing β cells. In this study we unveil the transcriptional dynamics of the differentiation and explain the heterogeneity of the final cell product, in comparison with β cells from organ donors.

overall_design: Single cell RNAseq of hiPSC during differentiation into pancreatic beta cells and of human islets from organ donors

first: Valeria

last: Sordi


title: Regulatory T cells support breast cancer progression by opposing IFN-γ -dependent functional reprogramming of myeloid cells

status: Public on Oct 26 2020

city: Richmond

first: Mikhail

last: Dozmorov

address: 830 E Main St

department: Biostatistics

institute: Virginia Commonwealth University

summary: Regulatory T (Treg) cell infiltration of solid tumors often correlates with poor prognosis, but their tumor suppressive function lacks mechanistic understanding. Through a combination of transgenic mice, cell fate mapping, adoptive transfer and co-injection strategies, we demonstrate that Treg cell ablation-dependent anti-tumor effects in murine breast cancer require intratumoral recruitment of CCR2+ inflammatory monocytes, which primarily differentiate into tumor-associated macrophages (TAMs), and lead to reprogramming of their function in an IFN--dependent manner. Furthermore, transcriptomic signatures from murine TAMs in Treg cell-ablated conditions correlate with increased overall survival in human breast cancer. Our studies highlight the strong myeloid dependency of breast cancer, and provide the basis for the development of therapeutic strategies based on manipulation of the IFN- signaling pathway in monocytes.

overall_design: We mechanistically dissect the tumor-promoting function of Treg cells in poorly immunogenic breast cancer, describing the critical role of TAMs in the anti-tumor phenotype observed upon Treg cell ablation, and the dependency on CD4+ T cell-produced IFN-γ for their functional reprogramming

first: Nicholas

last: Clark

first: Leandro

last: Martínez

first: Steven

last: Murdock

first: James

last: deLigio

first: Amy

last: Olex

first: Comfort

last: Effi

first: Paula

middle: D

last: Bos