title: Expression profiling of CD45- lung tumor cells

summary: Neutrophils are the most abundant circulating leucocytes and constitute an essential component of innate immunity. Although their role in cancer development is still poorly defined, pro- or anti-tumor properties have been attributed to tumor-associated neutrophils (TANs), suggesting major functional diversities. In this study, we focused on the mechanisms involved in neutrophil accumulation within the lung tumor mass. We first identified G-CSF as an inducer of the high-affinity glucose transporter Glut1 in neutrophils, increasing their survival ex vivo. In a genetically engineered mouse model of lung adenocarcinoma, we report that TANs have an increased Glut1 expression and glucose metabolism compared to normal neutrophils. To elucidate the impact of glucose uptake on TANs, we used an in vivo strategy based on two recombinases, Flp to initiate lung tumors, and Cre to delete Glut1 specifically in neutrophils. We demonstrate that the loss of Glut1 decreases the SiglecFhigh TAN subpopulation by accelerating neutrophil turnover in tumors through reduced survival and augmented recruitment. Accelerated TAN turnover led to a decreased tumor growth and synergized with radiotherapy. Altogether, our results demonstrate the importance of Glut1 for neutrophil turnover, which directly affects the pro- versus anti-tumor balance within the tumor. These results also suggest that metabolic vulnerabilities can be exploited to target tumor-supportive neutrophil populations.

overall_design: Expression profile comparisons between CD45- fraction from KPfrt; Ly6gCre/wt; Glut1flox/flox and KPfrt; Ly6gCre/wt; Glut1wt/wt macrodissected independant tumors

first: Pierre-Benoit

last: Ancey

last: Zangger

first: Etienne

last: Meylan


title: RNAseq analysis of B6-Mx1(r) and D2-Mx1(r) mice infected with PR8 virus

status: Public on Jan 23 2021

city: Münster

first: Klaus

last: Schughart

address: Von-Esmarch-Str 56

department: Virology

country: Germany

institute: University Münster

type: Expression profiling by high throughput sequencing

summary:
Gene expression changes in the lung was studied by RNAseq
Results: Strong differences in the immune response between the two mouse strains

overall_design: Lungs were collected from mice, RNA was isolated and sequenced.


title: miR1908-5p

status: Public on Jan 12 2021

city: Ottawa

first: Ruth

last: McPherson

state: Ontario

address: 40 Ruskin

country: Canada

institute: UOHI

summary: Transfection experiments aimed at understanding the role of miR1908-5p in liver.

overall_design: HuH7 cells (hepatocyte-derived carcinoma cell line) were transfected for 72 h with either one of 4 human miRNA analogues: miR1908-5p mimic, miR1908-5p mimic ctl, miR1908-5p inhibitor, miR1908-5p inhibitor ctl. RNA was then isolated and the transcriptome analyzed by transcription array.  The experiment was repeated on 3 distinct occasions (repeat 1-3).

first: Sebastien

last: Soubeyrand


title: Expression profiling of lung neutrophils

status: Public on Mar 18 2021

city: Lausanne

first: Nadine

last: Fournier

state: Vaud

address: Agora Cancer Research Center, Bugon 25A

country: Switzerland

institute: Agora Cancer Research Center & Swiss Institute of Bioinformatics

overall_design: Expression profile comparisons of lung neutrophils from healthy lung or lung tumor. Lung tumor neutrophils are Control (Glut1 proficient) or Glut1 deficient (Glut1KO)


title: Suppression of canonical TGF-β signaling enables GATA4 to interact with H3K27me3 demethylase JMJD3 to promote cardiomyogenesis

status: Public on Feb 13 2021

city: AURORA

last: Danis

state: CO

department: Cancer Center

country: USA

institute: University of Colorado, Denver - Anschutz Medical Campus

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Direct reprogramming of fibroblasts into cardiomyocytes (CMs) represents a promising strategy to regenerate CMs lost after ischemic heart injury. Overexpression of GATA4, HAND2, MEF2C, TBX5, miR-1, and miR-133 (GHMT2m) along with transforming growth factor beta (TGFbeta) inhibition efficiently promotes reprogramming. However, the mechanisms by which TGFbeta; blockade promotes cardiac reprogramming remain unknown. Here, we identify interactions between the histone H3 lysine 27 trimethylation (H3K27me3), demethylase JMJD3, the SWI/SNF remodeling complex subunit BRG1, and cardiac transcription factors. Furthermore, canonical TGFbeta; signaling regulates the interaction between GATA4 and JMJD3. TGF-beta; activation impairs the ability of GATA4 to bind target genes and prevents demethylation of H3K27 at cardiac gene promoters during cardiac reprogramming. Finally, a mutation in GATA4 (V267M) exhibits reduced binding to JMJD3 and impaired cardiomyogenesis. Thus, we have identified an epigenetic mechanism wherein canonical TGFbeta; pathway activation impairs cardiac gene programming by interfering with GATA4-JMJD3 interactions.

overall_design: Examination of H3K27me3 in mouse embryonic fibroblasts following induction of cardiomyocyte differentiation

first: Andrew

middle: S

last: Riching

first: Kunhua

last: Song


title: Neurodegeneration in herpes simplex virus 1 infected human brain organoids

status: Public on Jan 22 2021

city: Berlin

first: Emanuel

last: Wyler

address: Robert Roessle Str 10

department: Berlin Institute for Medical Systems Biology

institute: Max Delbrück Center for Molecular Medicine

summary: We describe the cellular response to HSV-1 infection in human iPSC-derived brain organoids

sample_organism:
Homo sapiens
Human alphaherpesvirus 1

platform_organism:
Homo sapiens
Human alphaherpesvirus 1

overall_design: Single-cell and bulk RNA-sequencing was performed from human brain organoids infected with HSV-1, at different time points post infection

first: Agnieszka

last: Rybak-Wolf

first: Ivano

last: Legnini

first: Petar

last: Glažar

first: Anna

last: Loewa

first: Seung

middle: J

last: Kim

first: Tancredi

middle: M

last: Pentimalli

first: Benjamin

last: Beyersdorf

last: Oliveras Martinez

last: Woehler

first: Markus

last: Landthaler

first: Nikolaus

last: Rajewsky


title: Unbiased genomic analysis of murine lung cancer cell lines

status: Public on Dec 31 2020

city: sunnyvale

first: Chen-Hua

last: Chuang

state: California

address: 963 E homestead rd

institute: amgen

summary: To uncover the gene expression alterations that occur during lung cancer progression, we interrogated the gene expression state of tumor cell lines generated from different stages of malignant progression. We initiated tumors in KrasLSL-G12D/+;p53flox/flox (KP) mice. Cancer cells were isolated from individual primary tumors and metastases

overall_design: This study includes 4 samples: 2 non-metastatic primary tumors, 2 metastasis in lymphnode. total RNA was isolated and prepared for sequencing using the Illumina TruSeq RNA kit to generate 100bp paired end reads. Reads were aligned to mm9.

first: Monte

last: Winslow


title: Distinct exhausted CD8 T cell populations are linked to C-peptide preservation in alefacept-treated, recent onset T1D subjects

status: Public on Dec 10 2020

city: Seattle,

first: Stephanie

last: Osmond

state: WA

address: 1201 9th Ave

institute: Benaroya Research Institute

summary: Bulk RNA-sequencing of sorted CD8 T cells from recent-onset T1D subjects treated with alefacept.

overall_design: 30 recent-onset T1D subjects were enrolled in the T1DAL trial of alefacept. Blood draws were taken at baseline (pre-treatment) and weeks 11, 24, 35, 52, 78 and 104 post-treatment. CD8 T cells were FACS-sorted and analyzed by bulk RNA-sequencing.

first: Kirsten

middle: E

last: Diggins

first: Elisavet

last: Serti

first: Alice

last: Long

first: Peter

last: Linsley


title: Distinct exhausted-like CD8 T cell populations are linked to C-peptide preservation in alefacept-treated, recent onset T1D subjects

status: Public on Dec 23 2020

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

overall_design: Refer to individual Series


title: Characterization of Peripheral Blood Mononuclear Cells Gene Expression Profiles of Pediatric Staphylococcus aureus Persistent and Non-Carriers

status: Public on Dec 07 2020

summary: Defects in innate immunity affect many different physiologic systems and several studies of patients with primary immunodeficiency disorders demonstrated the importance of innate immune system components in disease prevention or colonization of bacterial pathogens. To assess the role of the innate immune system on nasal colonization with Staphylococcus aureus, innate immune responses in pediatric S. aureus nasal persistent carriers (n=3) and non-carriers (n=3) were profiled by RNAseq. We stimulated previously frozen peripheral blood mononuclear cells (PBMCs) from these subjects with i) live S. aureus (a mixture of all carriage isolates), or ii) heat-killed S. aureus.PBMC gene expression profiles differed between persistent and non-S. aureus carriers following stimulation with either live or dead S. aureus. These observations suggest that individuals susceptible to persistent carriage with S. aureus may possess differences in their live/dead bacteria recognition pathway and that innate pathway signaling is different between persistent and non-carriers of S. aureus.

overall_design: Pediatric patients from the Northwest Assistance Ministries Clinic, Houston, TX, were eligible for participation in the current study if they were participants of a prior parent study investigating pediatric S. aureus nasal carriage (n=438). In the parent study (unpublished results), we established nasal carriage phenotypes (persistent, intermittent, or non-carriers) based on the identification of S. aureus on 2 consecutive nasal swabs collected 12-17 days apart. Participants in the parent study testing positive for S. aureus on both swabs were classified as persistent carriers. Non-carriers were S. aureus negative at both time points. PBMCs from pediatric S. aureus nasal persistent carriers (n =3, age 4 (M), 5 (F), and 10 (F) years) and non-carriers (n=3, age 2 (F), 4(M), and 11(F)),

first: Elisabeth

last: Israelsson

first: Damien

last: Chaussabel

first: Rebecca

last: Fischer

first: Heather

middle: C

last: Moore

first: D

middle: A

last: Robinson

first: Jesse

middle: W

last: Dunkle

middle: T

last: Essigmann

first: Eric

middle: L

last: Brown


title: A coregulator shift, rather than the canonical switch, underlies thyroid hormone action in liver

status: Public on Jan 01 2021

city: Philadelphia

first: Mitchell

last: Lazar

state: PA

address: 3400 Civic Center Blvd

department: Perelman School of Medicine

institute: University of Pennsylvania

summary: Thyroid hormones (TH) are powerful regulators of metabolism with major effects on body weight, cholesterol, and liver fat that have been exploited pharmacologically for many years. Activation of gene expression by TH action is canonically ascribed to a hormone- dependent “switch” from corepressor to activator binding to thyroid hormone receptors (TR), while the mechanism of TH-dependent repression is controversial. To address this, we generated a mouse line in which endogenous TRβ1 was epitope-tagged to allow precise chromatin immunoprecipitation at the low physiological levels of TR, and defined high confidence binding sites where TR functioned at enhancers regulated in the same direction as the nearest gene in a TRβ-dependent manner. Remarkably, although positive and negative regulation by TH have been ascribed to different mechanisms, TR binding was highly enriched at canonical DR4 motifs irrespective of the transcriptional direction of the enhancer. The canonical NCoR1/HDAC3 corepressor complex was reduced but not completely dismissed by TH and, surprisingly, similar effects were seen at enhancers associated with negatively as well as positively regulated genes. Conversely, coactivator CBP was found at all TH-regulated enhancers, with transcriptional activity correlating with the ratio of CBP to NCoR rather than their presence or absence. These results demonstrate that, in contrast to the canonical “all or none” coregulator switch model, TH regulates gene expression by orchestrating a shift in the relative binding of corepressors and coactivators.

overall_design: mRNA profile, TR beta cistrome and H3K27ac ChIP-seq of hypo- and hyperthyroid state mice livers.

first: Yehuda

last: Shabtai

first: Kirill

last: Batmanov

first: Nagaswaroop

middle: K

last: Nagaraj

first: Douglas

last: Forrest

first: Young-Wook

last: Cho


title: RNA SEQ NTHi modA2

status: Public on Oct 25 2020

city: Gold Coast

first: John

middle: Michael

last: Atack

state: QLD

address: Gold Coast Campus

department: Institute for Glycomics

country: Australia

institute: Griffith University

summary: Comparison of RNA expression profiles from NTHi strain 723 strains expressing methyltransferase ModA2 (ON) or not (OFF)

overall_design: RNA isolated from triplicate cultures of each strain (modA2 ON and modA2 OFF) grown in sBHI to mid-log; comparison of gene expression profiles by comparing modA2 ON vs modA2 OFF to determine differential gene expression influenced by ModA2 methylation differences

first: Michael

middle: P

last: Jennings


title: Exhausted-like CD8 T cell phenotypes linked to C-peptide preservation in alefacept-treated T1D subjects - sorted CD8 subsets

first: Stephanie

last: Osmond

summary: Bulk RNA-sequencing of sorted CD8 T cell subsets from recent-onset T1D subjects treated with alefacept

overall_design: Frozen PBMC from 4 samples (3 subjects; 1 with sample from both week 52 and 104 post-treatment) were FACS sorted into populations of interest and analyzed by bulk RNA-sequencing. Sorted populations included: PD1+/-, CD57+/-, KLRG1+/TIGIT+ (DP), and KLRG1-/TIGIT- (DN).

first: Kirsten

last: Diggins

first: Elisavet

last: Serti

first: Alice

last: Long

last: Linsley


title: Essential role of ThPOK autoregulatory loop in maintenance of mature CD4 T cell identity and function [RNA-seq]

status: Public on Apr 05 2021

last: Slifker

address: 333 Cottman Ave.

institute: Fox Chase Cancer Center

summary: The transcription factor ThPOK is critical for homeostasis and differentiation of mature T helper cells. Overall goal of this experiment is to show divergent gene expression profiles of  ThPOK-dependent autoregulatory loop disrupted in CD4 T cells.

overall_design: Naïve  T cells (TCRb+CD25-CD44- Cd62l hi) from WT (CD4 and CD8) and OB11/OB11 (CD4hi, CD4+CD8+, CD4lo) mice were sorted based on CD4 and CD8 expression. RNA was extracted using Trizol and were subjected RNA seq  using illumina sequencing.

first: Jayati

last: Basu

first: Yinfei

last: Tan

first: Dietmar

last: Kappes


title: Chromatin accessibility and transcriptomic heterogeneity in human tonsillar B-cell subsets

status: Public on Oct 21 2020

city: Durham

first: Micah

last: Luftig

state: NC

address: 213 Research Dr, DUMC Box 3054

department: Molecular Genetics and Microbiology

institute: Duke University School of Medicine


title: Genome-wide analysis of chromatin accessibility in human tonsillar B cell subsets

summary: We performed ATAC-Seq on three freshly dissociated human tonsils sorted to purity into the following B-cell subsets: CD19+ naïve (IgD+/CD38-), memory (IgD-/CD38-), germinal center light zone (IgD-/CD38mid/CD83+/CXCR4lo), germinal center dark zone (IgD-/CD38mid/CD83lo/CXCR4hi), and plasma cells (IgD-/CD38hi). We used these data to derive differentially accessible sites between B-cell subsets and relative to EBV-immortalized B cell lines (LCLs). These studies revealed similarities between LCLs and both DZ and LZ GC cells at the BCL2A1 (BFL1) locus that formed the impetus for further three-dimensional chromatin studies, which are the focus of the associated manuscript.

overall_design: A comparative study of genome-wide chromatin accessibility and locus specific differences in human tonsillar B-cell subsets and EBV-infected LCLs

first: Joanne

last: Dai

first: Gregory

last: Crawford

first: Lingyun

last: Song


title: Transcriptional response of A. thaliana Root and Shoot tissue to light stress and presence of microbial communities

status: Public on Oct 27 2020

city: Cologne

first: Thorsten

last: Thiergart

address: Carl-con-Linne-Weg 40

institute: MPIPZ cologne

summary: We profiled the root and shoot transcriptomes of BFO-colonized (Bacteria, Fungi, Oomycetes) and germ free A. thaliana exposed to Low PAR (photoactive radiation, LP) and normal condition (NC) in the gnotobiotic FlowPot system five weeks post inoculation.

overall_design: Combination of two different conditions were tested, a) absence/presnece of microbes and b) normal condition and low PAR condition. Respectively in plant Roots (n= 12 samples) and Shoots (n=12 samples)

first: Shiji

last: Hou

first: Stephane

last: Hacquard


title: Potent synergistic interactions between lopinavir and azole antifungal drugs against emerging multidrug-resistant Candida auris

status: Public on Oct 06 2020

city: West Lafayette

first: Nadia

middle: Marie

last: Atallah

state: IN

address: 201 S. University St

department: Center for Cancer Research

institute: Purdue University

summary: The limited number of antifungals and the emergence of multidrug-resistant Candida auris pose a significant challenge to human medicine. Here, we utilized combinatorial drug therapy as an approach to augment the activity of current azole antifungals against C. auris. We evaluated the fluconazole chemosensitization activity of 1547 FDA-approved drugs and clinical molecules against an azole-resistant strain of C. auris. This led to the discovery that lopinavir, an antiviral drug, is a potent agent capable of sensitizing C. auris to the effect of azole antifungals. At a therapeutically achievable concentration (4-8 µg/ml), lopinavir exhibited potent synergistic interactions with azole drugs, particularly with itraconazole, against C. auris (ΣFICI ranged from 0.05-0.50). The lopinavir/itraconazole combination enhanced the survival rate of C. auris-infected Caenorhabditis elegans by 90% and reduced the fungal burden in infected nematodes by 88.5% (p < 0.05). Moreover, lopinavir enhanced the antifungal activity of itraconazole against other medically important Candida species including C. albicans, C. tropicalis, C. glabrata, C. tropicalis, and C. parapsilosis. Comparative transcriptomic profiling revealed that lopinavir interferes with glucose permeation and ATP synthesis. This compromises the function of the efflux pumps presents in C. auris enhancing sensitivity to azole antifungals, as demonstrated by Nile red efflux assays. This study presents lopinavir as a novel, potent and broad-spectrum azole chemosensitizing agent that warrants further investigation against recalcitrant Candida infections.

overall_design: Method: polyA selection of mRNA from Candida auris either treated or untreated, followed by sequenced on a NovaSeq 6000

first: Hassan

last: Eldesouky

first: Ehab

last: Salama

last: Lanman

first: Tony

middle: R

last: Hazbun

first: Mohamed

middle: N

last: Seleem


title: Keystone genes for tooth development [bulk RNA-seq]

status: Public on Nov 01 2020

city: Helsinki

first: Jukka

last: Jernvall

address: P.O. Box 56 (Viikinkaari 9)

country: Finland

institute: University of Helsinki

summary:
The expression levels of developing mouse molar transcriptome were measured.
7 samples each from  embryonic day 13.5,  14.5  and 16.5 of mouse developing molar tooth.

overall_design: Wild type tooth germs were dissected from embryonic stages corresponding to E13, E14 and E16 molar development. For bulk and single-cell RNAseq we used C57BL/6JOlaHsd mice.

first: Outi

last: Hallikas

first: Rishi

last: Das Roy

first: Mona

last: Christensen

first: Elodie

last: Renvoisé

first: Ana-Marija

last: Sulic


title: Effect of Sdc1 knockout in mouse blast crisis chronic myeloid leukemia (bcCML) cells

city: La Jolla

first: Michael

last: Hamilton

state: CA

address: 2880 Torrey Pines Scenic Dr.

institute: UCSD

summary: Here we report the genomic impact on murine bcCML by Sdc1 loss. We compare bcCML derived from a wildtype cell source to bcCML derived from a Sdc1 knockout mouse. This RNA-seq identifies a number of potential critical regulators of growth, migration, and adhesion.

overall_design: RNA-seq comparative analysis of WT and Sdc1 in mouse bcCML cells

first: Kyle

last: Spinler

first: Tannishtha

last: Reya


title: Determination of carbon metabolism response regulators regulons in several Pseudomonas species by DAP-seq

status: Public on Oct 01 2020

city: Berkeley

first: Alexey

last: Kazakov

address: 1 Cyclotron Road

department: Environmental Genomics and Systems Biology Division

institute: Lawrence Berkeley National Laboratory

summary: We identified binding sites genome-wide for carbon metabolism transcriptional response regulators in Pseudomonas stutzeri RCH2, Pseudomonas putida KT2240, Pseudomonas fluorescens FW300-N2E2 and FW300-N2C3 by DNA-affinity-purified (DAP) sequencing.

sample_organism:
Pseudomonas fluorescens
Pseudomonas putida
Stutzerimonas stutzeri

overall_design: The potential response regulator binding sites were acquired by DNA-affinity-purified sequencing (DAP-seq).

first: Megan

last: Garber


title: Pleiotropic effects of trans-regulatory mutations on gene expression and fitness

status: Public on May 25 2021

city: Ann Arbor

first: Patricia

last: Wittkopp

state: MI

address: 1105 N University Ave

department: Department of Molecular, Cellular, and Developmental Biology

institute: University of Michigan

summary: Variation in gene expression arises from cis- and trans-regulatory mutations, which contribute differentially to expression divergence. Here, we compare the impacts on gene expression and fitness for cis- and trans-regulatory mutations affecting expression of the TDH3 gene in Saccharomyces cerevisiae. We use the effects of cis-regulatory mutations to isolate effects of trans-regulatory mutations caused by impacts on TDH3 from pleiotropic impacts on other genes, providing a rare distribution of pleiotropic effects. These pleiotropic effects were often, but not always, deleterious. For cis- and trans-regulatory mutations with similar effects on TDH3, trans-regulatory mutations had more widespread effects on gene expression, with distinct impacts on expression of genes downstream of TDH3. These differences between cis-and trans-regulatory mutations help explain their different contributions to regulatory evolution.

overall_design:
Each of four replicates is one sequencing lane composed of multiplexed library preps for 35 trans-regulatory mutants to TDH3, 3 deletion mutants, 3 alpha wild type controls, 1 a wild type control, 4 cis-regulatory mutants underexpressing TDH3, 1 duplication and cis-regulatory mutant of the TDH3 locus, and one duplication wild type control. A few strains are missing from each replicate due to failed library preps, resulting in 188 samples
Please note that the Strain*txt files are the DESeq2 output for each strain relative to a control and are generated from the data of all four replicate samples for each strain, as indiated in the corresponding sample description field.

first: Petra

last: VandeZande


title: Fibroblasts direct differentiation of human breast epithelial progenitors

status: Public on Sep 30 2020

city: Copenhagen

first: Mikkel

last: Morsing

state: Nørrebro

address: Blegdamsvej 3b

department: Department of Cellular and Molecular Medicine and Danish Stem Cell Center

country: Denmark

institute: University of Copenhagen

summary:
Background: Breast cancer arises within specific regions in the human breast referred to as the terminal duct lobular units (TDLUs). These are relatively dynamic structures characterized by sex hormone driven cyclic epithelial turnover. TDLUs consist of unique parenchymal entities embedded within a fibroblast-rich lobular stroma. Here, we established and characterized a new human breast lobular fibroblast cell line against its interlobular counterpart with a view to assessing the role of region-specific stromal cues in the control of TDLU dynamics.
Methods: Primary lobular and interlobular fibroblasts were transduced to express human telomerase reverse transcriptase (hTERT). Differentiation of the established cell lines along lobular and interlobular pathways was determined by immunocytochemical staining and genome-wide RNA sequencing. Their functional properties were further characterized by analysis of mesenchymal stem cell (MSC) differentiation repertoire in culture and in vivo. The cells’ physiological relevance for parenchymal differentiation was examined in heterotypic co-culture with fluorescence-activated cell sorting (FACS)-purified normal breast primary luminal or myoepithelial progenitors. The co-cultures were immunostained for quantitative assessment of epithelial branching morphogenesis, polarization, growth, and luminal epithelial maturation. In extension, myoepithelial progenitors were tested for luminal differentiation capacity in culture and in mouse xenografts. To unravel the significance of transforming growth factor-beta (TGF-β)-mediated crosstalk in TDLU-like morphogenesis and differentiation, fibroblasts were incubated with the TGF-β signaling inhibitor, SB431542, prior to heterotypic co-culture with luminal cells.
Results: hTERT immortalized fibroblast cell lines retained critical phenotypic traits in culture and linked to primary fibroblasts. Cell culture assays and transplantation to mice showed that the origin of fibroblasts determines TDLU-like and ductal-like differentiation of epithelial progenitors. Whereas lobular fibroblasts supported a high level of branching morphogenesis by luminal cells, interlobular fibroblasts supported ductal-like myoepithelial characteristics. TDLU-like morphogenesis, at least in part, relied on intact TGF-β signaling.
Conclusions: The significance of the most prominent cell type in normal breast stroma, the fibroblast, in directing epithelial differentiation is largely unknown. Through establishment of lobular and interlobular fibroblast cell lines, we here demonstrate that epithelial progenitors are submitted to stromal cues for site-specific differentiation. Our findings lend credence to considering stromal subtleties of crucial importance in the development of normal breast and, in turn, breast cancer.

overall_design: Duplicate primary lobular and interlobular breast fibroblastic cells. Duplicate immortalized lobular and interlobular breast fibroblastic cells.

first: Jiyoung

last: Kim

first: René

last: Villadsen

first: Nadine

last: Goldhammer

first: Abbas

last: Jafari

first: Moustapha

last: Kassem

first: Ole

last: Petersen

first: Lone

last: Rønnov-Jessen


title: Identification and Characterization of N6-Methyladenosine CircRNAs and Methyltransferases in the Lens Epithelium Cells from Age-related Cataract

status: Public on Aug 26 2020

city: Nantong

first: Peng fei

last: li

state: Jiangsu

address: Eye Institute, Affiliated Hospital of Nantong University, 20 Xisi Road, Nantong, Jiangsu, China.

department: Eye Institute

country: China

summary: To explore the involvement of N6-methyladenosine (m6A) modification in circular RNAs (circRNAs) and relevant methyltransferases in the lesion of lens epithelium cells (LECs) under the circumstances of age-related cataract (ARC).

overall_design: LECs were collected from normal subjects and patients with cortical type of ARC (ARCC). M6A-tagged circRNAs expression were analyzed by m6A-modified RNA immunoprecipitation sequencing (m6A-RIP-seq). Our data provided novel evidence regarding the involvement of circRNAs m6A modifications in ARC.

first: Pengfei

last: Li

first: Yong

last: Wang


title: CD109 regulates in vivo tumor invasion in lung adenocarcinoma through TGF-β signaling

city: Nagoya

first: Tetsuro

last: Taki

state: Aichi

address: 65 Tsuromai Showa

department: Pathology

country: Japan

institute: Nagoya University

summary: CD109 is a glycosylphosphatidylinositol-anchored glycoprotein highly expressed in several types of human malignant tumors including lung cancers. We investigated the in vivo functions of CD109 protein in malignant lung tumors.  CD109+/+ and CD109-/- K-ras[LSL-G12D/+];p53[fl/fl] (KP) mice were sacrificed at 20 to 23 weeks of age and total RNA was extracted from the lung tumors. SurePrint G3 Mouse GE Microarray 8×60K Ver.2.0 were performed.

overall_design: We performed microarray analysis of the lung tumors of CD109+/+ and CD109-/- KP mice in order to investigate which gene sets and pathways are significantly associated with CD109 deficiency.

first: Shinji

last: Mii


title: RNAseq analysis of U2OS cell clones made to stably expressing exogenous CIITA

status: Public on Sep 16 2020

first: Stephanie

last: Osmond

summary: Bulk RNAseq analysis was done on single cell clones of U2OS cells either expressing exogeonous CIITA or an empty vector control to identify genes regulated by CIITA expression.

overall_design: RNA was collected from 2.5X106 CIITA expressing cells, empty vector control cells, or wild type U2OS cells. At least four individual clones were used for CIITA expressing cells and three clones for empty vector control cells.

first: Anna

last: Bruchez

first: Adam

last: Lacy-Hulbert


title: Transposon mutagenesis screen identifies MHC class II transactivator (CIITA) as an inducer of cellular resistance to Ebola virus and SARS Coronavirus

summary: U2OS cells were co-transfected with PiggyBac (PB) transposon plasmid pPB-SB-CMV-puro-SD3 and the transposase p-hyPBASE. The modified PiggyBac (PB) transposon, which has a constitutively active CMV promoter which can stimulate or disrupt expression of neighboring genes, depending on insertional orientation. For each library, between 107-108 cells were transfected, cultured with the addition of 2 µg/ml puromycin for one week to select cells that had incorporated the transposon, and then used for screens of virus resistance with minimal further expansion. Mutagenized cells were challenged with replication-competent recombinant EboGP-VSV at MOI of 1 or 10 and propagated for up to 3 weeks with regular media changes to select virus-resistant colonies. After virus resistant cells were selected, genomic DNA (gDNA) was isolated from 5-10 X 106 cells and transposon insertion sites were mapped using high throughput sequencing.

overall_design: Eight individual libraries were created and were combined for common insertion site analysis (CIS).

first: Ky

last: Sha


title: RNA-sequencing of Chronic Myelomonocytic Leukemia and healthy donors bone marrow samples

status: Public on Aug 07 2020

city: Murcia

first: Ana

middle: María

last: Hurtado López

state: Murcia

address: Libertad, n

department: Hematology and Medical Oncology Department

country: Spain

institute: Hospital Morales Meseguer, Instituto Murciano de Investigación Biosanitaria (IMIB)

summary: RNA-seq of bone marrow samples of  chronic myelomonocytic leukemia (CMML) patients and healthy donors  to identify the molecular DNA repair  pathways involved in CMML pathogenic

overall_design: RNA-seq of bone marrow of CMML patients at diagnostic and bone marrow healthy donors

first: Andrés

last: Jerez

last: Hurtado


title: Response of expanded human conventional and regulatory T cells to CD28 or TNFR2 costimulation [Exp2]

status: Public on Jul 10 2020

city: Leiden

first: Mark

last: Mensink

address: Albinusdreef 2

department: Immunology

country: Netherlands

institute: Leiden University Medical Center

summary: Immune responses depend on a dynamic balance between the opposing activities of conventional (Tconv) and regulatory (Treg) CD4+ T cells. While receptor-targeted approaches have been developed based on their modulation of Tconv cells, Tconv and Treg cells share many costimulatory receptors. We aim to determine key differential signaling events downstream of costimulation to find opportunities for selective manipulation of Tconv or Treg cells. This data set includes the transcriptomes of expanded human Tconv and Treg cells that were treated with anti-CD3, anti-CD3/CD28 or anti-CD3/TNFR2 agonistic mAbs for 24 hours.

overall_design: Whole transcriptome analysis of expanded human conventional and regulatory T cells that were treated with anti-CD3, anti-CD3/CD28 or anti-CD3/TNFR2 agonistic mAbs (three healthy blood donors).

first: Sander

last: de Kivit

first: Jannie

last: Borst


title: Synovial Cell Cross-talk with Cartilage Plays a Major Role in the Pathogenesis of Osteoarthritis

status: Public on Jul 06 2020

first: Virginia

middle: Byers

last: Kraus

state: United States

address: 300 N Duke St

department: Duke Molecular Physiology Institute

institute: Duke University

summary: We elucidated the molecular cross-talk between knee articular cartilage and paired synovium in n=3 individuals with knee osteoarthritis using the powerful tool of single-cell RNA-sequencing. Multiple cell types were identified based on profiling of 10,640 synoviocytes and 26,192 chondrocytes (11,579 chondrocytes from the diseased medial vs 14,613 chondrocytes from the relatively non-diseased lateral tibial plateau): 12 distinct synovial cell types and 7 distinct articular chondrocyte phenotypes from matched tissues. Intact cartilage was enriched for homeostatic and hypertrophic chondrocytes, while damaged cartilage was enriched for prefibro- and fibro-, regulatory, reparative and prehypertrophic chondrocytes. A total of 61 cytokines and growth factors were predicted to regulate the 7 chondrocyte cell phenotypes. Based on production by >1% of cells, 55% of the cytokines were produced by synovial cells (39% exclusive to synoviocytes and not expressed by chondrocytes) and their presence in osteoarthritic synovial fluid confirmed. The synoviocytes producing IL-1beta (a classic pathogenic cytokine in osteoarthritis), mainly inflammatory macrophages and dendritic cells, were characterized by co-expression of surface proteins corresponding to HLA-DQA1, HLA-DQA2, OLR1 or TLR2. Strategies to deplete these pathogenic intra-articular cell subpopulations could be a therapeutic option for human osteoarthritis.

overall_design: We performed single-cell transcriptomic analysis on OA knee joint tissues to systematically identify cell types and states within human osteoarthritic (OA) synovium and matched cartilage as well as determined regulators of these articular chondrocyte phenotypes.

first: Ching-Heng

last: Chou

first: Vaibhav

last: Jain

first: Jason

last: Gibson

first: David

last: Attarian

first: Collin

last: Haraden

first: Christopher

middle: B

last: Yohn

first: Remi-Martin

last: Laberge

first: Simon

last: Gregory


title: Response of expanded human conventional and regulatory T cells to CD3/CD28-mediated activation (Exp1) and to CD28 or TNFR2 costimulation (Exp2)

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

overall_design: Refer to individual Series


title: Response of expanded human conventional and regulatory T cells to CD3/CD28-mediated activation [Exp1]

summary: Immune responses depend on a dynamic balance between the opposing activities of conventional (Tconv) and regulatory (Treg) CD4+ T cells. While receptor-targeted approaches have been developed based on their modulation of Tconv cells, Tconv and Treg cells share many costimulatory receptors. We aim to determine key differential signaling events downstream of costimulation to find opportunities for selective manipulation of Tconv or Treg cells. This data set includes the transcriptomes of expanded human Tconv and Treg cells that were either unstimulated or treated with anti-CD3/CD28 agonistic mAbs for 24 hours.

overall_design: Whole transcriptome analysis of expanded human conventional and regulatory T cells that were either unstimulated or treated with anti-CD3/CD28 agonistic mAbs (four healthy blood donors).


title: Characterizing the molecular regulation of inhibitory immune checkpoints with multi-modal single-cell screens.

status: Public on Jun 29 2020

city: New York

first: Efthymia

last: Papalexi

state: NY

address: 101 avenue of americas

institute: New York Genome Center

summary: The expression of inhibitory immune checkpoint molecules such as PD-L1 is frequently observed in human cancers and can lead to the suppression of T-cell mediated immune responses. Here we apply ECCITE-seq, a technology which combines pooled CRISPR screens with single-cell mRNA and surface protein measurements, to explore the molecular networks that regulate PD-L1 expression. We also develop a computational framework, mixscape, that substantially improves the signal-to-noise ratio in single-cell perturbation screens by identifying and removing confounding sources of variation. Applying these tools, we identify and validate regulators of PD-L1, and leverage our multi-modal data to identify both transcriptional and post-transcriptional modes of regulation. In particular, we discover that the kelch-like protein KEAP1 and the transcriptional activator NRF2, mediate levels of PD-L1 upregulation after IFNγ stimulation. Our results identify a novel mechanism for the regulation of immune checkpoints and present a powerful analytical framework for the analysis of multi-modal single-cell perturbation screens.

overall_design:
Identification of novel immune checkpoint regulators using ECCITE-seq.
Please note that the  ECCITE_ADT_Barcodes.csv and ECCITE_Arrayed_ADT_Barcodes.csv files are identical.  To avoid any confusion, duplicated files were generated to link to  each experiment type for users who may try to analyze a specific experiment (ECCITE or ECCITE_Arrayed).


title: The response of A549 transcriptome to RNA Pol II slowdown with alpha-amanitin and DRB

status: Public on Jun 27 2020

city: Moscow

first: Dmitri

middle: D

last: Pervouchine

address: Bolshoy Bul'var, 30, str 1

department: Skoltech Center for Molecular and Cellular Biology

country: Russia

institute: Skolkovo Institute of Science and Technology

summary: According to current estimates, more than 40% of nascent introns are spliced co-transcriptionally and, consequently, the speed of RNA polymerase II elongation must fundamentally affect the rate of RNA folding, spliceosome assembly, and the resulting mature transcript. RNA-Seq data was generated using RNAPII elongation inhibitors DRB and a-amanitin in A549 cells and used to look at genome-wide splicing changes

overall_design: 5x RNA-seq (DMSO, 10 ug/ml DRB, 25 ug/ml DRB, 1 ug/ml a-amanitin, 2 ug/ml a-amanitin) in A549 cells

first: Marina

last: Kalinina

first: Dmintry

last: Skvortsov

first: Maria

last: Logacheva


title: Single cell RNA-seq of CD1 mouse forebrain at embryonic day 15.5

status: Public on Jul 01 2020

city: San Diego

first: Zhen

last: Li

address: 9500 Gilman Dr, CNCB122

summary: Single cell RNA-seq was conducted on embryonic brains from wild-type CD1 mice at embryonic day 15.5 following The Illumina® Bio-Rad® SureCell Single-Cell RNA Sequencing workflow. Four cell captures and sequencing runs were conducted with four to eight fetal brains used for each collection. For each brain, the cerebral cortex was dissected and collected (although ventral forebrain structures, especially the gonglionic eminence may be mixed in due to difficulty in dissection).

overall_design: Single cell RNA-seq analysis of embryonic mouse cerebral cortex at E15.5

first: Tarik

last: Haydar

first: William

last: Tyler


title: RNA-Seq of ischemic stroke in young and aged mice

status: Public on Jun 17 2020

city: Vestec

first: Lukas

last: Valihrach

address: Prumyslova 595

country: Czech Republic

institute: Institute of Biotechnology of the Czech Academy of Sciences

summary: Ischemic stroke is one of the leading causes of mortality and major healthcare and economic burden. It is a well recognized disease of aging, yet it is unclear how the age-dependent vulnerability occurs and what are the underlying mechanisms. To address these issues, we performed a comprehensive RNA-Seq analysis of aging, ischemic stroke and their interaction using a model of permanent middle cerebral artery occlusion (MCAO) in 3 and 18 month old female mice. We assessed differential gene expression across injury status and age, estimated cell type proportion changes, assayed the results against a range of transcriptional signatures from the literature and performed unsupervised co-expression analysis, identifying modules of genes with varying response to injury. We uncovered selective vulnerability of neuronal populations and increased activation of type-I interferon (IFN-I) signaling and several other inflammatory pathways in aged mice. We extended these findings via targeted expression analysis in tissue as well as acutely purified cellular populations to show differential temporal dynamics of IFN-I signaling between age groups and contribution of individual cell types. Together, these results paint a picture of ischemic stroke as a complex age related disease and provide insights into interaction of aging and stroke on cellular and molecular level.

overall_design: 3 and 18 month-old C57Black/6 mice were subjected to permanent middle cerebral artery occlusion (MCAO) to model cerebral ischemia. Parietal cortex tissue was collected 3 days after MCAO and total RNA was isolated using TRIZOL. Intact 3 and 18-month old mice were used as controls. Sequencing libraries were prepared from 400 ng total RNA with Lexogen QuantSeq 3´ mRNA-Seq Library prep kit. 6 animals per treatment were used.

first: Peter

last: Androvič

first: Pavel

last: Abaffy

first: Lukáš


title: Transcriptomic signatures and chromatin structure in adipocytes isolated from premenauposal women subcutaneous fat

status: Public on Jun 08 2020

city: orlando

first: Adeline

last: Divoux

state: FL

address: 301 E. Princeton St

department: Translational Research Institute

institute: Adventhealth


title: RNA Sequencing Analysis of Wild Type and Nell2-KO Caput Epididymal Transcriptomes

status: Public on Jun 05 2020

first: Daiji

last: Kiyozumi

address: Furocho, Chikusa-ku

department: Research Institute of Environmental Medicine

summary:
Purpose: The goal of this study is to detect differentially expressed genes, between Wild type (WT) and Nell2-/- (Nell2-KO) caput epididymis by RNA sequencing
Methods: Caput epidiymal mRNA profiles of 14-week-old wild-type (WT) and NEL-like 2 knockout (Nell2-KO) mice were generated by deep sequencing, in quadplicate, using  Illumina GAIIx.
Results: RNA-seq data identified differentially expressed transcripts; 26 genes showed differential transcript expression between the WT and Nell2-KO caput epididymis, with WT RPKM ≥100, a fold change ≤0.1, and t-test p value <0.05. Altered expression of 26 genes identified.
Conclusions: Our results show that the expression of many genes were downregulated in Nell2-KO caput epididymis compared with WT one.

overall_design: Caput epididymal mRNA profiles of 14-week old wild type (WT) and Nell2-KO mice were generated by deep sequencing, in quadplicate, using Illumina GAIIx.

first: Takashi

last: Kohda

first: Masahito

last: Ikawa


title: Transcriptomic signatures and chromatin structure in adipocytes isolated from premenauposal women subcutaneous fat [ATAC-seq]

summary: Our data showed that lipid and glucose metabolic pathways genes were expressed at higher levels in gluteofemoral adipocyte fraction in pears, while genes associated with inflammation were higher in both abdominal and gluteofemoral apple adipocyte fraction. Gluteofemoral adipocyte chromatin from pear-shaped women contained a significantly higher number of differentially open ATAC-seq peaks relative to chromatin from the apple-shaped gluteofemoral adipocytes. In contrast, abdominal adipocyte chromatin openness showed few differences between apple and pear-shaped women. We revealed a correlation between gene transcription and open chromatin at the proximity of the TSS of some of the differentially expressed genes.

overall_design: Adipocytes isolated from A and GF adipose tissue obtained from premenopausal apple-shaped and pear-shaped women were used to perform transcriptome analysis by RNA-seq and assess accessible chromatin regions by ATAC-seq.

first: S

last: Smith

first: T

middle: F

last: Osborne


title: Comparative transcriptomic and epigenomic analysis identifies key regulators of injury response and neurogenic competence in retinal glia

status: Public on May 01 2020

city: Baltimore

first: Jie

last: Wang

state: Maryland

address: 400 N Broadway

department: Ophthalmology

institute: Johns Hopkins University

summary: Injury induces retinal Muller glia of non-mammalian, but not mammalian, vertebrates to generate neurons.  To identify gene regulatory networks that control neurogenic competence in retinal glia, we used bulk and single-cell RNA-seq and ATAC-seq analysis to comprehensively profile gene expression and chromatin conformation in Muller glia from zebrafish, chick and mice.  This was conducted during glial development, following inner and outer retinal injury, as well as following treatment with extrinsic factors that induce glial reprogramming.   Integration of these data, together with functional analysis of candidate genes, identified evolutionarily conserved and species-specific gene regulatory networks controlling glial quiescence, gliosis, and neurogenic competence.  In zebrafish and chick, transition from quiescence to gliosis is a necessary stage in acquisition of neurogenic competence, while in mice a dedicated network suppresses this transition and rapidly restores quiescence.  These findings may help guide the design of cell-based therapies aimed at restoring retinal neurons lost to disease.

sample_organism:
Danio rerio
Mus musculus

platform_organism:
Danio rerio
Mus musculus

overall_design: In this study, to comprehensively identify transcriptional and epigenetic regulators of neurogenic competence in MG, we profiled mRNA levels using bulk RNA-seq and chromatin accessibility using ATAC (Assay for Transposase-Accessible Chromatin) sequencing technology in zebrafish and mouse in response to multiple neuronal injury models, as well as growth factor treatment. In total, we generated 105 bulk RNA-Seq libraries (including 5 technical replicates) and 40 bulk ATAC-Seq libraries.

first: Thanh

last: Hoang

first: Patrick

last: Boyd

first: Fang

first: David

last: Hyde

first: Jiang

last: Qian

first: Seth

last: Blackshaw


title: IRF5 genetic risk variants drive myeloid-specific IRF5 hyper-activation and pre-symptomatic SLE

status: Public on Apr 07 2020

city: Seattle,

first: Stephanie

last: Osmond

address: 1201 9th Ave

institute: Benaroya Research Institute

summary: The IRF5-SLE risk haplotype is associated with SLE disease severity. We hypothesized that neutrophils from healthy risk donors would carry a pathogenic gene signature compared to non-risk donors. To compare basal neutrophil gene signature between risk and non-risk healthy individuals, fresh whole blood was collected from healthy donors, and granulocytes were extracted from the remaining pellet following Ficoll purification. RNA-seq libraries were prepared from globin-reduced RNA extracted from the granulocytes using the Qiagen RNA preparation kit.

overall_design: Neutrophils from healthy donors with the IRF5-SLE risk haplotype (n=6) and with a non-risk IRF5-SLE haplotype (n=8) were extracted from whole blood for RNA-sequencing. Quality filters were applied to exclude samples with fewer than 4 million total reads, a median CV of coverage greater than 50%, or less than 75% alignment to the reference genome. An additional library was excluded due to outlying quality variance identified by PCA. Following this quality control, 11 of 14 samples (5 risk and 6 non-risk) remained for downstream analysis.

first: Kirsten

last: Diggins

first: Bharati

last: Matta

first: Betsy

middle: J

last: Barnes

middle: S

last: Linsley


title: Successful Preclinical Islet Transplantation in the Subcutaneous Space for Type 1 Diabetes

status: Public on Jun 01 2020

city: Philadelphia

first: Ali

last: Naji

address: 3400 Civic Ctr Blvd

department: Transplant Surgery

institute: University of Pennsylvania

type: Non-coding RNA profiling by high throughput sequencing

summary: The intrahepatic milieu is inhospitable to intraportal islet allografts, limiting their applicability to ameliorate Type 1 Diabetes (T1D). Islet viability in the subcutaneous space represents an unfulfilled paradigm that is crucial to ensure widespread adoption and safety of clinical islet transplantation. Herein we report that human islets transplanted subcutaneously uniformly promote long-term euglycemia when admixed with a device-free Islet Viability Matrix (IVM), through a previously unknown anti-apoptotic mechanism.

overall_design: Human islets transplanted into mouse; 10 samples were used in this analysis. 5 samples were treated with collagen and no collagen for the studied time points .

first: Ming

last: yu

first: Divyansh

last: Agarwal

first: Laxminarayana

last: Korutla

first: Catherine

last: May

first: Negin

last: Griffith

first: Wei

first: Andrei

last: Georgescu

first: Bernard

last: Herring

first: Omaida

middle: C

last: Velazquez

first: Prasanth

last: Vallabhjosyul

first: James

last: Markmann

first: Chengyang

last: Liu


title: A composite immune signature parallels disease progression across T1D subjects (RNA-Seq Cohort 2 Cell Sorted)

status: Public on Feb 03 2020

first: Stephanie

last: Osmond

summary: Peripheral blood mononuculear cells were thawed and sorted into B cell, CD4+ T cells, CD8+ T cells, and monocytes. Samples were obtained from control-arm subjects enrolled in 2 clinical trials evaluating immunotherapies in T1D that were conducted by the Immune Tolerance Network. The trials had similar study timepoints, clinical and metabolic measurements, and data and sample collection plans. Total RNA was isolated from the sorted cell populations and then RNAseq libraries were prepared.

overall_design: Subjects from the placebo arms of 2 new onset T1D trials conducted by the Immune Tolerance Network were aggregated to evaluate the heterogeneity in the rates of insulin secretion decline.  The outcome used for this analysis was the rate of decline in C-peptide as calculated by sequential 2 hour AUC of serum C-peptide after a mixed-meal tolerance test over the 2 years of each trial. We obtained RNA-seq data from the baseline visit (N=28 subjects) of participants in these trials. Of 112 libraries, 106 yielded high-quality RNA-seq data that were used in downstream analyses.

first: Cate

last: Speake

first: Samuel

middle: O

last: Skinner

first: Dror

last: Berel

first: Elizabeth

last: Whalen

first: Matthew

last: Dufort

first: Jared

middle: M

last: Odegard

first: Anne

last: Pesenacker

first: Frans

last: Gorus

first: Eddie

middle: A

last: James

first: Megan

last: Levings

first: Peter

first: Eitan

last: Akirav

first: Alberto

last: Pugliese

first: Martin

last: Hessner

first: Gerald

last: Nepom

first: Raphael

last: Gottardo

first: S

last: Long


title: Analysis of High Molecular Weight RNA-induced silencing complex (HMW-RISC) in CD8+ T cells identifies miR-7 as a modulator of T cell activation

status: Public on Jun 04 2020

city: Edinburgh

first: Amy

last: Buck

address: Ashworth Labs, Charlotte Auerbach Road

department: IIIR

country: United Kingdom

institute: The University of Edinburgh

summary: To identify the miRNAs expected to be potent in suppressing targets, we followed HMW RISC formation upon activation of CD8+ T cells. We show that while most miRNAs distribute between HMW and LMW RISC in activated T cells, several miRNAs were dominant in one complex over the other. Among these, miR-7 is enriched in HMW RISC and inhibition of miR-7 upon T cell activation leads to increased production of IL-2 and expression of IL-2-regulated proteins including the α-subunit of the IL-2 receptor, CD25; transferrin receptor, CD71; and amino acid transporter, CD98, which are direct miR-7 targets. Our data support a model where recruitment of miR-7 to HMW RISC restrains IL-2 signalling and the metabolic processes regulated by IL-2 and thus modulates T cell activation.

overall_design: RNA was isolated from Ago2-IP samples, as well as the IP input and unbound samples. The samples were resuspended in QIAzol lysis reagent and RNA was extracted using the miRNeasy kit (Qiagen) and eluted in 100µl dH2O. The RNA was then ethanol precipitated by adding 2.5 x volume 100% ethanol, 30µl sodium acetate and 1µl GlycoBlue Coprecipitant (Thermo Scientific) to the samples which were then incubated overnight at  -20oC. The following day, the samples were centrifuged at full speed for 30 min, then washed twice with 70% ethanol. The pellets were air-dried on ice then resuspended in 10-15µl dH2O and quantified with Qubit 3.0 fluorometer (Thermo Scientific) using the Qubit RNA HS Assay kit (Thermo Scientific). Small RNA libraries were prepared using the CleanTagTM Small RNA Library Preparation kit (Trilink) using manufacturer’s instructions and 21 cycles. The libraries were size selected to include 145-160 bps and the input, IP-unbound and IP-bound libraries were pooled at a 1:1:2 ratio. The samples were sequenced using NovaSeq 50bp paired end sequencing at Edinburgh Genomics.

first: Matilda

last: Toivakka

first: Katrina

last: Gordon

first: Sujai

last: Kumar

first: Rose

last: Zamoyska


title: A composite immune signature parallels disease progression across T1D subjects

first: Stephanie

last: Osmond


title: A composite immune signature parallels disease progression across T1D subjects (RNA-Seq Cohort 0 WB)

summary: Technical replicate testing was performed to determine the measurement precision of each analyte. Peripheral blood samples were collected in tempus tubes from a healthy control subject with no family history of autoimmunity (n=1) and from subjects with Type 1 diabetes (n=4). Total RNA was isolated independently from 3 replicate aliquots of the same tempus samples and then globin-reduced. RNAseq libraries were prepared from the globin-reduced RNA.

overall_design: The replicate testing cohort was used to determine the technical precision of each analyte by calculating the intra-subject coefficient of variation (CV) for three replicate aliquots (n=3) from the same blood draw from 5 subjects. For each analyte, the mean CV across subjects was then calculated. In order to generate the three replicate aliquots, a tempus tube of 9 mL (3 mL blood plus 6 mL tempus solution) was split into three equally sized samples. These samples were processed separately to RNA, then separate globin reduction and library preparation was conducted.

first: Cate

last: Speake

first: Samuel

last: Skinner

first: Dror

last: Berel

first: Elizabeth

last: Whalen

first: Matthew

last: Dufort

first: Jared

last: Odegard

first: Anne

middle: K

last: Linsley

middle: T


title: Transcriptomic signatures and chromatin structure in adipocytes isolated from premenauposal women subcutaneous fat [RNA-seq]

first: Adeline

last: Divoux

overall_design: Adipocytes isolated from Abdominal (A) and gluteofemoral (GF) adipose tissue obtained from premenopausal apple-shaped and pear-shaped women were used to perform transcriptome analysis by RNA-seq and assess accessible chromatin regions by ATAC-seq.

last: Smith

first: T

last: Osborne


title: Single-cell transcriptome profiling reveals neutrophil heterogeneity in homeostasis and infection

status: Public on May 27 2020

city: Boston

first: Xuemei

last: Xie

state: MA

address: 811 Enders Building, 320 Longwood Ave.

department: Transfusion Medicine

institute: Boston Children's Hospital

summary: The full neutrophil heterogeneity and differentiation landscape remains incompletely characterized. Here we profiled more than 25,000 mouse differentiating and mature neutrophils using single-cell RNA sequencing to provide a comprehensive transcriptional landscape of neutrophil maturation, function, and fate decision in their steady state and during bacterial infection. Eight neutrophil populations (including the GMP population) were defined by distinct molecular signatures, including a new circulating mature neutrophil population highly expressing interferon-stimulated genes. The three mature peripheral blood neutrophil subsets arise from distinct maturing bone marrow neutrophil subsets, a novel mechanism that highlights the complex and precise regulation of neutrophil production. Neutrophil heterogeneity and differentiation are driven by both known and uncharacterized transcription factors. Neutrophils gradually acquire microbicidal capability as cells traverse the transcriptional landscape, representing an evolved mechanism for fine-tuned regulation of an effective but balanced neutrophil response. Bacterial infection reprograms the genetic architecture of neutrophil population, alters dynamic transition between each subpopulation, and primes neutrophils for augmented functionality without affecting overall heterogeneity. Bacterial infection-induced emergency granulopoiesis is mediated by augmented proliferation of early-stage neutrophil progenitors and accelerated post-mitotic maturation. In summary, single-cell transcriptomics enabled the reconstruction of neutrophil differentiation and maturation trajectories and uncovered neutrophil subpopulations, gene pathways, and regulators of neutrophil function and fate decisions. These data establish a reference model and general framework for studying neutrophil-related disease mechanisms, biomarkers, and therapeutic targets at single-cell resolution.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design:
There are 10 mouse samples and 1 human sample. All relative organs consist of bone marrow (BM), peripheral blood (PB), spleen (SP), liver (LV) and peritoneal cavity (PC).
4 mouse control samples: BM sorted by cKit+ Gr1+; BM, PB, SP sorted by Gr1+.
6 mouse E. coli challenged samples: BM sorted by cKit+ Gr1+; BM, PB, SP, LV and PC sorted by Gr1+.
3 human control samples: PB sorted by CD33.

first: Qiang

last: Shi

first: Peng

last: Wu


title: The impact of Prp18p on splicing fidelity and efficiency in budding yeast

status: Public on May 25 2020

city: Los Angeles

first: Kevin

last: Roy

address: 607 Charles E. Young Drive East

department: Chemistry and Biochemistry

institute: UCLA

summary: Fidelity of 3´-splice site (3´SS) selection by the spliceosome is critical for proper gene expression but is a daunting task considering the low complexity of the 3´SS consensus YAG. Here we show that loss of the splicing factor Prp18p in budding yeast activates a diverse array of alternative 3´SS at more than half of all introns. Some alternative sites highly diverge from the YAG consensus, demonstrating a critical role for Prp18p in promoting spliceosome fidelity. Usage of alternative 3´SS is determined by distance from the branchpoint, local RNA secondary structures, upstream poly(U) content, and adenosine enrichment in exons. The 3´SS fidelity function of Prp18p can be genetically uncoupled from its role in splicing efficiency and is promoted by interactions with Slu7p and Prp8p. Taken together, these results provide a comprehensive mechanism into how the spliceosome achieves specificity of 3´SS selection and how it prevents aberrant activation of non-canonical splice sites.

overall_design: RNA sequencing of yeast strains with and without the second step splicing factor Prp18p in a nonsense-mediated decay mutant background


title: Isolated fiber-type transcriptional profiles in human skeletal muscle

status: Public on Feb 01 2020

city: Manhattan

first: Aliza

last: Rubenstein

address: 1 Gustave L. Levy Pl

institute: Icahn School of Medicine at Mount Sinai

summary: Skeletal muscle is a complex heterogeneous tissue comprised of diverse muscle fiber and non-fiber cell types that, in addition to movement, influences other systems such as immunity, metabolism and cognition. We investigated gene expression patterns of resident human skeletal muscle cells using both single-cell RNA-seq and RNA-seq of single muscle fiber dissections from vastus lateralis. We generated transcriptome profiles of the major multinucleated human skeletal muscle fiber-types as well as 11 human skeletal muscle mononuclear cell types, including immune, endothelial, pericyte and satellite cells.  We delineated two fibro-adipogenic progenitor cell subtypes that may contribute to heterotopic ossification and muscular dystrophy fibrosis under pathological conditions. An important application of cell type signatures is for computational deconvolution of cell type specific gene expression changes using data from bulk transcriptome experiments. Analysis of transcriptome data from a 12 week resistance exercise training study using these human skeletal muscle cell-type signatures revealed significant changes in specific mononuclear cell-type proportions related to age, sex, acute exercise and training. This characterization of human skeletal muscle cell types will resolve cell-type specific changes in large-scale physical activity muscle transcriptome studies and can further the understanding of the diverse effects of exercise and the pathophysiology of muscle disease.

overall_design: Muscle fibers were isolated from biopsies, MHC content of each fiber was determined by SDS-PAGE, and fibers of each fiber-type (I vs. IIa) were pooled for sequencing.

first: Gregory

first: Ulrika

last: Raue

first: Frederique

last: Ruf-Zamojski

first: Venugopalan

last: Nair

first: Lan

last: Zhou

first: Elena

last: Zaslavsky

first: Scott

last: Trappe

first: Stuart

last: Sealfon


title: Transcript profiling of putative MTF-1 activating chemicals in wild-type and MTF-1-null MCF-7 cells

city: Research Triangle Park

first: Christopher

last: Corton

address: 109 TW Alexander Dr.

institute: US-EPA

summary: In future high-throughput transcriptomic studies, we propose that the use of cell lines nullizygous for important targets of environmental chemicals will greatly facilitate the interpretation of regulated genes. To provide proof of this concept, we generated transcript profiles from wild-type and MTF-1-null cells exposed to zinc (Zn) and six putative activators of MTF-1 generated using TempO-Seq targeted sequencing of ~3000 human genes. In the absence of chemical exposure, knockout of MTF-1 resulted in expression changes in 44 genes indicating that MTF-1 has some effect on gene expression in the absence of overt activation. Almost all of the genes regulated by Zn and phenoxybenzamine were no longer regulated in the MTF-1-null cells. For the rest of the chemicals, more than half of the genes regulated in the wild-type cells were no longer regulated in the MTF-1-null cells. These studies demonstrate that many of the changes in gene expression after exposure to the putative MTF-1 activators were MTF-1-dependent.

overall_design: An initial concentration-range finding experiment was conducted in MCF-7 cells to determine the optimal concentration of the test chemicals to use in the study. Cells were plated in 12-well plates and incubated 48 hours. The media was replaced with dosing solutions containing DMSO (0.05%) or 1-100 uM of each chemical in DMSO (final concentration 0.05%). After 6 hours of exposure, media was removed and cells were lysed in 0.5 mL Trizol, followed by RNA extraction. Gene expression changes in wild-type and MTF1-null cells were evaluated using the human 1500+ Tempo-Seq platform (BioSpyder, Inc, Carlsbad, CA).

first: J

last: Liu


title: Cryptic developmental events determine medulloblastoma radiosensitivity and cellular heterogeneity without altering transcriptomic profile

status: Public on May 15 2020

city: chapel hill

first: Seth

last: Weir

address: 115 mason farm rd

institute: UNC Chapel Hill

summary: Using scRNA-seq, we show in a murine medulloblastoma model that tumors initiated earlier in development differ from later initiated tumors in their cellular composition and response to radiation therapy.

overall_design: Brains were cut in half sagittally and drop-fixed in 4% paraformaldehyde for IHC, and half cerebellum or tumor collected for Drop-Seq.

first: Daniel

last: Malawsky

first: Taylor

last: Dismuke

first: Jennifer

last: Ocasio

first: Benjamin

last: Babcock

first: Kirk

last: Wilhelmsen

first: Timothy

last: Gershon


title: RNA sequencing of different neutrophil subpopulations from bone marrow

first: Xuemei

last: Xie

summary: The purpose of this study is to learn the dynamic transcriptomic and molecular changes during neutrophil development. We investigated the gene expression profiles of purified morphology-defined neutrophil populations. Promyelocytes (PMs), myelocytes (MCs), metamyelocytes (MMs), band neutrophils and segmented neutrophils (BN/SNs) were isolated by FACS based on differential expression of C-Kit and Ly6g.Their identities were confirmed by morphological examination. Bulk RNA sequencing was then performed to reveal the gene expression profiles and molecular signatures of each cell population. Detailed analysis of gene expression revealed that MBs highly expressed the stem cell marker Cd34 and translation-related genes such as Eef1a1, while the highest expression of primary granule-related genes such as Mpo, Elane, Prtn3, and Cstg was detected in the PM population. MCs and MMs were highly proliferative neutrophils expressing high levels of cell cycle-related genes which were significantly downregulated in mature neutrophils. Finally, this study provides a framework for the detail study of the neutrophil development and for the future application in the modulation of the granulopoiesis under pathological conditions.

overall_design: Five neutrophil populations: promyelocytes (PMs), myelocytes (MCs), metamyelocytes (MMs), band neutrophils and segmented neutrophils (BN/SNs) were isolated by FACS based on differential expression of C-Kit and Ly6g.

first: Xiaoyu

last: Zhang

first: Qiang

last: Shi


title: A composite immune signature parallels disease progression across T1D subjects (Affymetrix Cohort 2)

first: Stephanie

last: Osmond

summary: We used an array-based bioassay whereby serum is used to induce transcription in healthy “reporter” leukocytes.  Serum samples were obtained from control-arm subjects enrolled in 3 clinical trials evaluating immunotherapies in T1D that were conducted by the Immune Tolerance Network. The included trials evaluated immune-modifying therapy in new-onset T1D, with similar trial timecourses, primary outcomes, and data and sample collection. Total RNA was isolated from the reporter cell line for gene expression analysis.

overall_design: Subjects from the placebo arms of three new onset T1D trials conducted by the Immune Tolerance Network were aggregated to evaluate the heterogeneity in the rates of insulin secretion decline. The outcome used for this analysis was the rate of decline in C-peptide as calculated by sequential 2 hour AUC of serum C-peptide after a mixed-meal tolerance test over the 2 years of each trial. UPN727 cells were stimulated with serum collected at baseline visits (N= 40 subjects). Gene expression analysis was performed in order to evaluate the transcriptional signature associated with new onset T1D.

first: Cate

last: Speake

first: Samuel

last: Skinner

first: Dror

last: Berel

first: Elizabeth

last: Whalen

first: Matthew

last: Dufort

first: Jared

last: Odegard

first: Anne

last: Pesenacker

first: Frans

last: Gorus

first: Eddie

last: James

first: Megan

last: Levings

first: Peter

last: Linsley

first: Eitan

last: Akirav

first: Alberto

last: Pugliese

first: Martin

last: Hessner

first: Gerald

last: Nepom

first: Raphael

last: Gottardo

first: S

last: Long


title: Transcriptomic analysis of blood samples collected from SFTS patients

status: Public on Jan 29 2020

city: Wuhan

first: Leike

last: Zhang

address: Xiaohongshan road

institute: The State Key Laboratory of Virology  Wuhan Institute of Virology, CAS

summary: To investigate the mechanisms that contribute to the pathogenesis resulting from SFTSV infection, transcriptomic analysis was performed with blood samples collected from SFTS patients.

overall_design: Patients were divided into three groups based on their clinical status, namely “Recover”: SFTS patients during convalescence who have recovered from acute virus infection (n=19); “Acute. Recover”: SFTS patients during acute virus infection and who eventually recovered from the disease (n=22); and “Acute. Deceased”: SFTS patients during acute virus infection and who eventually died of SFTSV infection (n=17). The group of healthy control is composed by donors who have not experienced SFTSV infection (n=23)

first: Hao

last: Li


title: A composite immune signature parallels disease progression across T1D subjects (RNA-Seq Cohort 0 Cell Sorted)

first: Stephanie

last: Osmond

summary: Technical replicate testing was performed to determine the measurement precision of each analyte. Peripheral blood mononuculear cells were processed and frozen down from blood samples collected from  healthy control subjects (n=3) or subjects with Systemic Lupus Erythematosus (n=3). Three separate PBMC aliquots per subject were then independently thawed and sorted by flow into B cell, CD4+ T cell, CD8+ T cell, and monocyte populations.  Total RNA was isolated from the sorted cell populations and then RNAseq libraries were prepared.

overall_design: The replicate testing cohort was used to determine the technical precision of each analyte by calculating the intra-subject coefficient of variation (CV) for three replicate aliquots (n=3) per blood draw from 6 subjects. For each analyte, the mean CV across subjects was then calculated.  Three independent PBMC aliquots per subject were thawed and sorted by flow cytometry into B cell, CD4+ and CD8+ T cell, and monocyte subsets before RNA sequencing.

first: Cate

last: Speake

first: Samuel

last: Skinner

first: Dror

last: Berel


title: A composite immune signature parallels disease progression across T1D subjects (Affymetrix Cohort 0)

summary: Technical replicate testing was performed to determine the measurement precision of each analyte in the array-based bioassay. Serum from subjects (n=5) with T1D was used to induce transcription in healthy “reporter” leukocytes. Three independent serum aliquots per subject were thawed and used to stimulate separate wells of reporter cells.  Total RNA was isolated from the reporter cell line for gene expression analysis.

overall_design: The replicate testing cohort was used to determine the technical precision of each analyte by calculating the intra-subject coefficient of variation (CV) for three replicate aliquots from the same blood draw from five subjects. For each analyte, the mean CV across subjects was then calculated.  Independent serum aliquots (n=3) per subject were thawed and used to stimulate separate wells of reporter cells (UPN727 cells).  Total RNA was then isolated from the reporter cell line for gene expression analysis using the Affymetrix U137Plus2 chip (54675 probes).

first: Elizabeth

last: Whalen

first: Matthew

last: Dufort

first: Jared

last: Odegard

first: Anne

last: Pesenacker


title: Comparative analysis of mRNA distribution in heavy and light polysomes in wild type and Dicer1 knockout cells

status: Public on May 21 2020

city: Lubbock

first: Degeng Wang

last: Wang

state: TX

address: 1207 Gilbert Drive

department: Environmental Toxicology

institute: Texas Tech University

summary: To gain a global view of translational inhibition by microRNA (miRNA), we isolated polysomes from wild type and Dicer1 knockout HCT116 human cells using sucrose gradient fractionation technique. The polysome fraction was separated into light (9mer or less) and heavy (10mer or more) subfractions. RNA samples were extracted from both subfractions and subjected to RNA-seq analysis. A general shift from light to heavy subfractions was detected for miRNA targted mRNAs.

overall_design: Cytoplasmic extract was subjected to standard sucrose gradient fractionation. Light and heavy polysome fractions were identified based on the OD260 profile.

first: Degeng


title: CRISPR mediated deletion of  RP11-326A19.4/CARMAL

status: Public on Dec 17 2019

city: Ottawa

first: Ruth

last: McPherson

state: Ontario

address: 40 Ruskin

institute: UOHI

summary: Deletion experiment aimed at understanding the role of lncRNA RP11-326A19.4 /CARMAL via its deletion.   The impact on of the deletion on the transcriptome was assessed by array analysis.

overall_design: HEK293T cells were transfected with dCas9-VPR alongside either 2 gRNA  expression constructs spanning exon 1 of RP11-326A19.4/CARMAL  or a tracrRNA only for 48 h.  Single clones were then obtained by cell sorting and genotyped.  RNAs from a deleted clone and 3 control clones, obtained over 4 passages, were isolated and analyzed on  human HuGene-2_0-ST (Affymetrix) arrays. Each  Wtx/KOx pair represents a matched set from passage number x.

first: Sebastien

last: Soubeyrand


title: dCas9 activation of RP11-326A19.4 phase 2

summary: Transfection experiments aimed at understanding the impact of upregulating lncRNA RP11-326A19.4 on the transcriptome; follow-up of GSE132451

overall_design: HEK293T cells were transfected with CRISPRa (dCas9-VPR) alongside either 2 gRNA  expression constructs  targeting the promoter region (CG4:-334 and CG6:-505 relative to TSS) of a lncRNA (RP11-326A19.4) or tracrRNA only for 48 h.  Sample RNAs were then analyzed on  human HuGene-2_0-ST (Affymetrix).


title: Transcriptional profiling of BL/6 mice with alterations in Kras, Stk11, and/or Keap1/Nrf2.

status: Public on Jan 31 2020

city: South San Francisco

first: Anneleen

last: Daemen

address: 1 DNA Way

institute: GENENTECH, INC.

summary:
Transcriptional profiling of BL/6 mice harboring a mutant Kras allele, with or without knockouts of Stk11 and/or Keap1, or with expression of an Nfe2l2 transgene, Nrf2Tg.
In lung adenocarcinoma (LUAD), stabilization of the transcription factor NRF2 through genomic alterations in KEAP1 and NFE2L2 occurs in roughly a quarter of patients, often in the context of STK11 tumor suppressor loss. In this study, we demonstrate that NRF2 activation in the context of concurrent KRAS mutation and STK11 loss promotes aggressive LUAD tumor behavior in both human and mouse preclinical models. This phenotype is associated with metabolic rewiring and rescue by NRF2 of redox stress, high in STK11 null tumors. Applying a novel, pan-lung cancer, diagnostic NRF2 activation gene expression signature that is independent of frequently co-occurring mutations, we dissect the independent contributions of the three most frequent genetic events in human LUAD (NRF2 activation, STK11 loss and KRAS mutations) on patient prognosis and clinical responses in a dataset of second-line LUAD patients treated with immunotherapy or chemotherapy (OAK trial). Our findings underscore that both individual effects and epistatic relationships among oncogenic and tumor suppressor pathways influence tumor biology, immune contexture and patient clinical outcomes. Our work also highlights the value of lung cancer disease sub-classification based on genetic and expression profiling as part of patient clinical management.

overall_design: RNA was isolated from normal tissue of wild-type (WT) BL/6 mice or from tumors in BL/6 mice harboring a mutant Kras allele, with or without knockouts (KO) of Stk11 (LKB) and/or Keap1, or with expression of an Nfe2l2 (Nrf2) transgene (Nrf2Tg). 4 replicates for all conditions, with the exception of Nrf2Tg (3 replicates). The microarray data were generated at the Boston University Microarray and Sequencing Resource lab.

first: Anju

last: Singh

first: Shyam

last: Biswal

first: Georgia

last: Hatzivassiliou


title: A human TH17 population with a tissue-resident signature in healthy and inflamed oral mucosal tissues

status: Public on Dec 08 2019

state: WA

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

overall_design: Refer to individual Series


title: A human TH17 population with a tissue-resident signature in healthy and inflamed oral mucosal tissues [10x]

summary: T cells in mucosal tissues fulfill a complex array of duties to ensure maintenance of barrier immunity. In oral mucosa tissue, we found that increased inflammation altered CD4 T cell subsets in a spatially-dependent manner, although it had a modest effect on the frequency of tissue-resident memory T cells (TRM) and the CD4 T cell transcriptome. In contrast, localization to the tissue profoundly altered the transcriptional profile, emphasizing the importance of studying healthy tissue to understand disease-specific changes. Our data revealed the existence of a TH17 cell population that is predominantly found in the tissue-resident, but not transient, CD4 T cell compartment in mucosal tissue.

overall_design: This project contains bulk RNA-seq data from paired oral mucosa tissue and blood CD4 T cell subsets from 10 subjects and 10X genomics sequencing of CD4 T cell subsets from one individual

first: Amanda

last: Davis

first: Hayley

last: Roozen

first: Hannah

last: DeBerg

first: Martha

last: Delaney

first: Florian

last: Mair

first: Jami

last: Erickson

first: Chloe

last: Slichter

first: Julia

last: Berkson

first: Matthias

last: Mack

first: Yu

last: Lwo

first: Alexander

last: Ko

first: Yu-Wen

last: Chiu

first: Jessica

last: Hsu

first: Peter

last: Linsley

first: Douglas

last: Dixon

first: Martin

last: Prlic


title: Single-cell profiles of retinal neurons differing in resilience to injury reveal neuroprotective genes - Atlas of adult mouse retinal ganglion cells

status: Public on Nov 27 2019

city: Cambridge

first: Wenjun

last: Yan

address: 52 Oxford Street

department: Department of Molecular  and Cellular Biology

institute: Harvard University

summary: Neuronal types in the central nervous system differ dramatically in their resilience to injury or insults. Here we studied selective resilience in mouse retinal ganglion cells (RGCs) following optic nerve crush (ONC), which severs their axons and leads to death of ~80% of RGCs in 2 weeks. To identify expression programs associated with differential resilience, we first used single-cell RNA-seq (scRNA-seq) to generate a comprehensive molecular atlas of 45 RGC types in adult retina. We tracked their survival after ONC, characterized transcriptomic, morphological, and physiological changes that preceded degeneration, and identified genes selectively expressed by each type. Finally, loss- and gain-of-function assays in vivo showed that manipulating some of these genes improved neuronal survival and axon regeneration following ONC. This study provides a systematic framework for parsing type-specific responses to injury, and demonstrates that these responses can be used to reveal molecular targets for intervention.

overall_design: In this experiment, we FACS sorted for retinal ganglion cells using a combination of a transgenic reporter (VGLUT2) and a surface protein CD90 from dissociated cells from retinas of adult (postnatal day 56) mice

first: Irene

middle: E

last: Whitney

first: Nicholas

last: Tran

first: Karthik

last: Shekhar

first: Anne

last: Jacobi

first: Inbal

last: Benhar

first: Zhigang

last: He

first: Aviv

last: Regev

first: Joshua

last: Sanes


title: The conserved WhiA protein influences the fatty acid composition of the Bacillus subtilis cell membrane

status: Public on Oct 18 2019

city: Amsterdam

first: Martijs

last: Jonker

address: Science Park 904

department: Microarray Department

institute: University of Amsterdam

summary: The conserved WhiA protein family is present in most Gram-positive bacteria and plays a role in cell division. WhiA contains a DNA-binding motive and has been identified as a transcription factor in the actinomycetes. In Bacillus subtilis, a deletion of whiA influences cell division and chromosome segregation. However, WhiA does not seem to function as a transcription factor in this organism, and it is still unclear how WhiA influences these processes. The B. subtilis whiA gene is part of an operon and flanked by two metabolic genes, including yvcK required for growth under gluconeogenic conditions, and the gene encoding the catabolite repressor Crh. Therefore, B. subtilis WhiA might play a role in carbon metabolism which could indirectly affect cell division and chromosome maintenance. To study this, we followed a metabolomics approach and measured exo-metabolome flux during growth on different carbon sources, using nuclear magnetic resonance spectroscopy. The carbon utilization of wild type and whiA mutant cells was similar, however, the whiA mutant exhibited depleted pools of branched-chain fatty acid precursors and higher levels of acetate and 2-oxoglutarate. Transcriptome data could not link these effects to gene regulatory differences, but the reduction in branched-chain fatty acid precursors influenced the fatty acid composition of the cell membrane. In conclusion, WhiA does not participate in carbon catabolite regulation, but affects fatty acid composition of the membrane by an as yet unknown mechanism. We speculate that the effect on the membrane might be related to the pleiotropic phenotype of the B. subtilis whiA mutant.

overall_design: wild type and whiA mutant

first: Laura

last: Bohorquez

first: Joana

last: de Sousa

first: Transito

last: Garcia-Garcia

first: Marie-Françoise

last: Noirot

first: Michael

last: Lalk

first: Leendert

middle: W

last: Hamoen


title: SirT7 auto-ADPribosylation regulates glucose starvation response through macroH2A1.1

status: Public on Aug 07 2019

city: Piscataway

last: Thackray

state: NEW JERSEY

address: 145 Bevier Road

department: Human Genetics Institute of New Jersey

institute: Rutgers University


title: Genetic mechanisms regulating the host response during mastitis

status: Public on Oct 22 2019

city: Guelph

first: Angela

last: Cánovas

state: ON

address: 50 Stone Road East

department: Animal Biosciences

institute: University of Guelph

summary: Mastitis is a very costly and common disease in the dairy industry. The study of the transcriptome from healthy and mastitic milk somatic cell samples using RNA-Sequencing technology (RNA-Seq) can provide measurements of transcript levels associated with the immune response to the infection. The objective of this study was to characterize the Holstein milk somatic cells transcriptome from 6 cows to determine host response to intramammary infections. RNA-Sequencing was performed on two samples from each cow from two separate quarters, one classified as healthy (n = 6) and one as mastitic (n = 6). In total, 449 genes were differentially expressed between the healthy and mastitic quarters (P-value < 0.01, FDR < 0.05, FC > ±2). Among the differentially expressed genes, the most expressed genes based on Reads Per Kilo base per Million mapped reads (RPKM) in the healthy group were associated with milk components (CSN2 and CSN3), and in the mastitic group they were associated with immunity (B2M and CD74). In-silico functional analysis was performed using the list of 449 differentially expressed genes, which identified 36 significantly enriched metabolic pathways (FDR < 0.01), some of which were associated with the immune system, such as cytokine-cytokine interaction and cell adhesion molecules. Seven functional candidate genes were selected, based on the criteria of being highly expressed and present in significant pathways that are relevant to the inflammatory process (GLYCAM1, B2M, CD74, BoLA DR-Alpha, FCER1G, SDS and NFKBIA). Lastly, we identified the differentially expressed genes that are located in QTL regions previously known to be associated with mastitis, specifically clinical mastitis, somatic cell count and somatic cell score. It was concluded that there are multiple genes within QTL regions that could potentially impact host response to mastitis causing agents, making some cows more susceptible to intramammary infections.  The identification of key genes with functional, statistical, biological and positional relevance associated with host defense to infection, will contribute to a better understanding of the underlying genetic architecture associated with mastitis. This in turn will improve the sustainability of agricultural practices, by facilitating the selection of cows with improved host defense leading to increased resistance to mastitis.

overall_design: RNA-Sequencing was performed on 6 Holstein dairy cows taking two samples from the same cow, one from a heathy quarter (n = 6) and one from the mastitic quarter (n = 6) to identify differentially expressed genes

first: Victoria

last: Asselstine

first: Filippo

last: Miglior

first: Aroa

last: Suárez-Vega

first: Pablo

last: Fonseca

first: Bonnie

last: Mallard

first: Niel

last: Karrow

first: Alma

last: Islas-Trejo

first: Juan

last: Medrano


title: Single-cell profiles of retinal neurons differing in resilience to injury reveal neuroprotective genes - Time course of transcriptomic changes in single retinal ganglion cells following optic nerve crush

first: Wenjun

last: Yan

overall_design: In these experiments, we FACS sorted for retinal ganglion cells using a combination of a transgenic reporter (VGLUT2) and a surface protein (CD90) from dissociated retinas of adult (6-8 wk old) mice, including uninjured controls, and at 6 time points following optic nerve crush (12h, 1d, 2d, 4d, 1w, 2w).

first: Irene

last: Whitney

first: Inbal

last: Benhar

first: Nicholas

last: Tran

first: Karthik

last: Shekhar

first: Anne

last: Jacobi

first: Zhigang

last: He

first: Aviv

last: Regev

first: Joshua

last: Sanes


title: Global Analysis of the dermatophyte Trichophyton rubrum when shifted from glucose to keratin

status: Public on Sep 19 2019

city: Ribeirão Preto

middle: Rodrigo

last: Sanches

state: SP

address: Av. Bandeirantes, 3900

department: Genetics

country: Brazil

institute: University of São Paulo

summary:
Purpose: We evaluated T. rubrum transcriptome using high-throughput RNA-sequencing (RNA-seq) technology aiming to identify metabolic pathways modulated  the development in keratin.
Methods: T. rubrum strain CBS118892 was inoculated into 100 mL Sabouraud dextrose broth, and incubated under agitation at 28 °C for 96 h. Mycelia were aseptically transferred into 100 mL minimal medium (MM) at pH 5.0 containing 70 mM nitrate and 50 mM glucose (control) or 0.5% (w/v) bovine keratin (treatment) as the carbon source. The cultures were incubated under agitation at 28 °C for 24 h, 48 h, or 96 h. Equal amounts of RNA from three independent biological replicates of keratin or glucose cultures were sequenced with a Hiseq 2000 sequencer. Paired-end reads 150 bp in size were generated. Raw read data obtained were filtered for quality control by FastQC tool and trimmed with Trimmomatic to remove adapters and Illumina-specific sequences. Trimmed paired-end reads from each sample were aligned to the T. rubrum reference genome. Gene-level read counts were quantified with STAR’s ‘-quantModeGeneCounts’ parameter. Differential expression was analyzed with the DESeq2 bioconductor package. A Benjamini-Hochberg correction adjusted the P threshold and was applied to reveal statistically significant changes in gene expression levels. It was set to 0.05, with a log2 fold change ± 1.5 and postulated as a significantly modulated transcript abundance level. Genes surpassing these thresholds are hereinafter referred to as differentially expressed genes (DEG). They were functionally categorized with the Gene Ontology (GO) terms assigned by the Blast2GO algorithm. Highly represented categories were determined by enrichment analysis with the BayGO algorithm.
Results: Our results present adaptive strategies in response to culture media. The global modulation in response to keratin supports fungal virulence and proper adaptation to the environment, in comparison to the availability of glucose as a carbon source.
Conclusions: Our results revealed a profile of the adaptive events to the host environment, crucial to the establishment and maintenance of the infection. The observed metabolic modulation is necessary for fungus survival and perpetuation in the initial steps of the infective process in their host.

overall_design: We examined at the transcriptomic level the effects of carbon source switching from glucose to keratin in Trichophyton rubrum metabolism. We aimed to evaluate the transcriptional pattern during the early stages of the infection in the host.

first: Maíra

middle: P

last: Martins

first: Larissa

middle: G

last: Silva

first: Antonio

last: Rossi

middle: R

last: Souza

first: Nilce

last: Martinez-Rossi


title: Human inflammatory mucosal T cells in the female reproductive tract are poised to participate in immunity to infection

status: Public on Sep 10 2019

city: Seattle,

first: Stephanie

last: Osmond

address: 1201 9th Ave

institute: Benaroya Research Institute

summary: The hypothesis is that HIV affects TRM, by decreasing their numbers and possibly affecting their function. In order to be able to analyse several HIV+ and HIV- samples, we would need to identify some markers for TRM that can be used for flow cytometry-based detection of TRM.

overall_design: To characterize TRM, we will use a RNA seq approach, in order to identify new markers for CD8+ TRM in human tissues. We will compare CD69+, CD69+ CD103+ and CD69- CD103- populations in the tissue and effector memory T cells, as well as CD69+ T cells in the blood.

first: Laura

last: Pattacini

first: Amanda

last: Woodward Davis

first: Julie

last: Czartoski

first: Florian

last: Mair

first: Scott

last: Presnell

first: Sean

last: Hughes

first: Gretchen

last: Lentz

first: Anna

last: Kirby

first: Michael

last: Fialkow

last: Hladik

first: Martin

last: Prlic

first: Jennifer

last: Lund


title: DNA methylation loss coupled with mitotic cell division promotes immune evasion of tumours with high mutation load

status: Public on Jul 31 2019

city: Seoul

first: Sung-Yeon

last: Hwang

address: School of Biological Sciences, Seoul National University

country: South Korea

summary: Mitotic cell division increases tumour mutation burden and copy number load with a positive and inverse correlation, respectively, with the clinical benefit of immunotherapy. Markers of cell division correlate also with genomic demethylation involving methylation loss in late-replicating partial methylation domains. Here we find that immunomodulatory pathway genes are concentrated in these domains and transcriptionally repressed in demethylated tumours with CpG island promoter hypermethylation. Global methylation loss correlated with immune evasion signatures independently of mutation burden and aneuploidy. Methylome data of our cohort (n = 60) and a published cohort (n = 81) in lung cancer and a melanoma cohort (n = 40) consistently demonstrated that genomic methylation alterations counteract the contribution of high mutation burden and increase immunotherapeutic resistance. Higher predictive power was observed for methylation loss than mutation burden. We also found that genomic hypomethylation correlates with the immune escape signatures of aneuploid tumours. Hence, DNA methylation alterations implicate epigenetic modulation as a combination regimens for precision immunotherapy.

overall_design: Bisulphite converted DNA from the 60 samples were hybridised to the Human Infinium MethylationEPIC BeadChip

first: Hong

last: Kim


title: Single-cell profiles of retinal neurons differing in resilience to injury reveal neuroprotective genes

first: Wenjun

last: Yan


title: SirT7 auto-ADPribosylation regulates glucose starvation response through macroH2A1.1 (RNA-seq)

first: Joshua

last: Thackray

summary: Sirtuins are key players in the response to oxidative, metabolic and genotoxic stress, and are involved in genome stability, metabolic homeostasis and aging. Originally described as NAD+-dependent deacetylases, some sirtuins are also characterized by poorly understood mono-ADP-ribosyltransferase (MADPRT) activity. Here we report that the deacetylase SirT7 is a dual sirtuin as it also features auto-MADPRT activity. Molecular and structural evidence suggests that this novel activity occurs at a second previously undefined active site that is physically separated in another domain. Specific abrogation of this activity alters SirT7 chromatin distribution, suggesting a role for this modification in SirT7 chromatin binding specificity and localization. Our studies uncover an epigenetic pathway by which ADP-ribosyl-SirT7 is recognized by the ADP-ribose reader macroH2A1.1, a histone variant involved in chromatin organization, metabolism and differentiation. Glucose starvation (GS) boosts this interaction and promotes SirT7 re-localization intergenic regions in a macroH2A1-dependent manner, which is required for specific up- or downregulation of a subset of nearby genes upon GS in primary cells and in vivo in the livers of calorie-restricted (CR) Wt and SirT7-/- mice. The level of expression of these genes decreases with age in SirT7-deficient mice, reinforcing the link between Sirtuins, CR and aging. Our work provides a novel perspective about sirtuin duality and suggests a key role for SirT7/macroH2A1.1 axis in mammalian glucose homeostasis, calorie restriction signaling and aging.

overall_design: Total RNA from 14.5 day WT and SirT7-/- MEFs (+/- GS) was harvested subjected to high-throughput sequencing.

first: Lourdes

last: Serrano

first: Nicolás

last: Simonet

first: Alejandro

last: Vaquero


title: Transcriptome analyses for salt sensitivity of blood pressure

city: Beijing

first: Han

last: Qi

institute: Beijing An Ding Hospital

type:
Non-coding RNA profiling by array
Expression profiling by array

summary: Salt sensitivity of blood pressure (SSBP) is an independent risk factor for cardiovascular disease. However, the pathogenic mechanisms of SSBP are still uncertain. Thus, ceRNA microarray was applied to identify differentially expressed lncRNAs and mRNAs. The whole blood samples from 10 hypertensives and 10 normotensives were collected by professional nurses in community health centers using EDTA blood tube. The samples were classified according to the salt sensitivity (SS, salt sensitivity; SR, salt resistant) and hypertension (H, hypertensives; N, normotensives). QRT-PCR and cell experiments would be also implemented to validate the reliability of the results.

overall_design: In the ceRNA microarray, 10 salt-sensitive individuals were regarded as cases and 10 salt-resistant individuals were controls. The whole blood samples were collected by professional nurses in the morning after fasting for 8 hours. There were no interventions on participants before collecting the blood samples.

first: Ling

last: Zhang

first: Wenjuan

last: Peng


title: Single-cell RNA-Sequencing data of posteriorized embryonic-like sacs, amniotic ectoderm-like cells, and H9 hESC

status: Public on Jul 26 2019

city: Syracuse

first: Yi

last: Zheng

state: New York

address: 900 S Crouse Ave

department: Biomedical and Chemical Engineering

institute: Syracuse University

summary: The study of early post-implantation human embryo development is limited by the scarcity of embryo samples and ethical restrictions. We developed a human pluripotent stem cell (hPSC)-based microfluidic platform to model several developmental processes during this stage. We applied single-cell mRNA sequencing technique to examine the transcriptomes of posteriorized embryonic-like sacs obtained using the microfluidic platform, as well as of amniotic ectoderm-like cells generated using a Transwell method and human embryonic stem cells (H9 hESCs).

overall_design: Cells retrieved from posteriorized embryonic-like sacs were pooled from 6 microfluidic devices, after 48 hours of exogenous BMP4 stimulation. Amniotic ectoderm-like cells were also obtained by treating H9 hESCs with BMP4 for 48 hours using a Transwell method. H9 hESCs maintained using standard tissue culture dishes were also sequenced for comparison.

first: Y

first: J

last: Fu


title: Experimental evolution identifies adaptive aneuploidy as a mechanism of fluconazole resistance in Candida auris

status: Public on Sep 04 2019

city: Shanghai

first: Jian

last: Bing

address: Songhu Road, Yangpu District

department: School of Life Sciences

institute: Fudan University

summary: In the current study, we set out to explore the developmental mechanism of antifungal resistance in this fungus using an in vitro experimental evolution strategy.

overall_design: total RNA profiles of drug resistant and sensitive strains of Candida auris

first: Guanghua

last: Huang