title: Extracellular Vesicles From Liver Progenitor Cells Downregulates Fibroblast Metabolic Activity and Increase the Expression of Immune-Response Related Molecules

city: Derio

first: Ana

middle: Maria

last: Aransay

state: BIZKAIA

address: Parque tecnologico de Bizkaia, Building 801-A

department: Genome Analysis Platform

institute: CIC bioGUNE

summary: Extracellular vesicles are an important part of cell to cell crosstalk, and help to the acceptor cells to prepare for changes in their environment. MLP29 cells are mouse liver progenitor cells that release EVs loaded with cell fate signaling. In the current work, we have fed 3T3-L1 mouse fibroblast with those EVs, and then analyzed the cellular response by proteomics and transcriptomics. Results showed a downregulation of both proteins and transcripts related to proliferative and metabolic routes dependent on TGFb, and an increase on the ERBB2 inhibitor protein (ERBIN). An increase in the transcription of CXCL2, as well as the increase of ribosome biogenesis and interferon response molecules points toward the activation of immune system signaling, compatible with a response against tissue damage.

overall_design: Approximately 2 million of 3T3-L1 cells were treated with the EVs obtained from 200 million of MLP29 cell line. On average, this represents 20 ug of protein (measured by Bradford) coming from 8.9*E10 particles (measured with Nanosight LM10) for treatment. In total 6 plates were treated, with three different preparations of EVs and 3 preparations of control (the very same procedure of EV isolation over a non conditioned-media, to rule out effects associated to the culture media itself), during 24 hours, in 25 mM HEPES-containing complete DMEM medium (contaminating vesicles were first removed by overnight centrifugation at 110000×g). At the end, plates were washed twice with PBS, raised with Tryple (GIBCO) and then divided equally for proteomic and transcriptomics analysis. An aliquot was employed to measure cell viability, higher than 90% in all the cases. The trypsin was removed after pellet the cells 1000xg 5 min, frozen in dry ice and stored at -80ºC until processing.

first: F

last: Royo

first: M

last: Azkargorta

first: J

last: Lavin

first: A

last: Cortazar

last: Clos

last: Gonzalez-Lopez

first: L

last: Barcena

first: H

last: Del Portillo

last: Yañez-Mo

last: Marcilla

last: Borras

last: Peinado

first: I

last: Guerrero

last: Váles-Gómez

first: U

last: Cereijo

first: T

last: Sardon

last: Elortza

last: Falcon-Perez


title: Single cell RNA sequencing analysis of freshly resected PC9 cell line-derived tumor xenografts

status: Public on Dec 19 2020

city: Chicago

first: Elizaveta

middle: V

last: Benevolenskaya

state: IL

address: 900 S. Ashland Ave.

department: Biochemistry and Molecular Genetics

country: USA

institute: University of Illinois at Chicago

type: Expression profiling by high throughput sequencing

summary: The non-small cell lung carcinoma (NSCLC) PC9 cell line is an established preclinical model for tyrosine kinase inhibitors. Using PC9 cells, we generated EGFR-mutant lung cancer xenografts to study the differences in response between individual cells and cell populations. We performed treatment of PC9 xenograft tumors with the combination of osimertinib and crizotinib as well as single drugs, followed by Drop-seq. The addition of crizotinib was guided by our previous data in PC9 grown in cell culture that identified an erlotinib-resistant drug population sensitive to crizotinib. The results of the xenograft study show that combination treatment targets specific osimertinib-tolerant cell populations but leaves a subset of the population that is tolerant to the combo. Each cell subpopulation is characterized by specific molecular signatures. The results of our study help to address emerging drug resistance that limits clinical usefulness of targeted strategies, particularly in NSCLC.

overall_design: examining drug resistant cell populations using single cell RNA-seq

first: Alexandre

middle: F

last: Aissa

first: Abul

middle: B

last: Islam


title: Profiling non-small cell lung carcinoma cell line PC9 treated with etoposide, erlotinib and its combination with crizotinib

summary: The non-small cell lung carcinoma (NSCLC) PC9 cell line is an established preclinical model for tyrosine kinase inhibitors. To be able to better understand the differences in response between individual cells, we performed treatment of PC9 cells grown in cell culture with etoposide, erlotinib and its combination with crizotinib, followed by Drop-seq. The addition of crizotinib was guided by our previous data that an erlotinib-resistant drug population may be sensitive to crizotinib. To better understand the common events in drug resistance, we compared the resistant cell populations arising from the treatment with etoposide and from the treatment with erlotinib. The results of our study will address emerging drug resistance that limits clinical usefulness of conventional and targeted strategies, particularly in NSCLC.


title: Different forms of KLF5 regulate distinct sets of genes in prostate cancer cells [RNA-seq]

city: Shenzhen

first: Jin-Tang

last: Dong

state: Guangdong

address: 1088 Xueyuan Blvd

department: Human Cell Biology and Genetics

country: China

institute: Southern University of Science and Technology

summary: To identify which genes are regulated by KLF5 and its acetylation, we performed RNA-Seq and bioinformatics analyses in KLF5-null prostate cancer cells expressing KLF5, KLF5KR, and KLF5KQ.

overall_design: RNA-Seq analyses were performed using KLF5 knockout DU 145 and PC-3 cells that ectopically expressed empty vector (EV), wildtype KLF5 (KLF5); acetylation deficient mutant KLF5K369R (KR), and acetylation mimicking mutant KLF5K369Q (KQ).

first: Baotong

last: Zhang

first: Yixiang

last: Li

first: Qiao

last: Wu

first: Benjamin

last: Barwick


title: RNA sequencing of AC16 and AC16-CAR cells

status: Public on Dec 21 2020

city: Charlottesville

first: Kevin

middle: A.

last: Janes

state: VA

address: Box 800759 Health System

department: Department of Biomedical Engineering

institute: University of Virginia

summary: The goal of this study was to determine the differences in gene expression due to ectopic CAR overexpression.

overall_design: Examining transcriptome changes in AC16 cells with ectopic CAR overexpression compared to parental AC16 cells. Both cell lines were run in duplicate.

first: Millie

last: Shah

first: Matthew

middle: D

last: Sutcliffe

first: Andrew

middle: J

last: Sweatt

middle: A


title: Chromatin accessibility to investigate the TF-chromatin information patterns encoded in the accessible genome

city: St Louis

first: Ricardo

last: D'Oliveira Albanus

state: MO

address: 4370 Duncan Avenue, Suite 6306D, Jeffrey T. Fort Neuroscience Bdg.

department: Neurology

institute: Washington University School of Medicine

type: Genome binding/occupancy profiling by high throughput sequencing

summary:
Purpose: The goal of this study is to use chromatin accessibility data to study the information patterns of chromatin accessibility that encode TF-chromatin interactions.

Methods: We generated chromatin accessibility data from the GM12878 lymphoblastoid cell line using a modified protocol where the ATAC-seq fragments are sonicated during the library preparation. The goal of the sonication step is to disrupt the observed fragment size distribution, therefore masking local TF-chromatin interactions encoded in the larger fragment sizes (e.g. nucleosome phasing) without affecting the levels of chromatin accessibility.

overall_design: We generated ATAC-seq libraries of the GM1287 lymphoblastoid cell line under two conditions: with and without a sonication step to disrupt the ATAC-seq fragment size distribution.

first: Yasuhiro

last: Kyono

first: John

last: Hensley

first: Arushi

last: Varshney

first: Peter

last: Orchard

first: Jacob

middle: O

last: Kitzman

first: Stephen

middle: C

last: Parker


title: Chromatin phosphoproteomics unravels a function for AT-hook motif nuclear localized protein AHL13 in PAMP-triggered immunity

status: Public on Dec 20 2020

city: Thuwal

first: Prof. Heribert

last: HIRT

address: 4700 KAUST

country: Saudi Arabia

institute: KAUST

summary: We report the transcriptome composition of ahl13-1 compared to WT (col-0) plant without treatment and after Pst hrcC-application

overall_design: Illumina high-sequencing plateform was used to analyse the transcriptome composition of col0 and ahl13-1 under treated and untreated conditions. col0 samples are in GEO Series GSE118854.

first: Naganand

last: Rayapuram

first: Mai

last: Jarard

first: Hanna

last: Alhoraibi

first: Jean

last: Bigeard

first: Aala

last: Abulfaraj

first: Ronny

last: Voelz

first: Kiruthiga Gayathri

last: Mariappan

first: Mariella

last: Almeida-Trapp

first: Maria

last: Schlöffel

first: Emmanuelle

last: Lastrucci

first: Ludovic

last: Bonhomme

first: Andrea

last: Gust

first: Axel

last: Mithöfer

first: Stefan

last: Arold

first: Delphine

last: Pflieger

first: Heribert

last: Hirt


title: Molecular and Functional Characterization of Disease-Propagating Stem Cells in Juvenile Myelomonocytic Leukemia

city: Oxford

first: Adam

last: Mead

address: Headington

department: WIMM

country: United Kingdom

institute: UNiversity of Oxford

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

overall_design: Refer to individual Series


title: Molecular and Functional Characterization of Disease-Propagating Stem Cells in Juvenile Myelomonocytic Leukemia [FACS sorted populations]

summary: We report on the characterization of  JMML hematopoietic stem and progenitor cells (HSPCs).  In order to molecularly characterize these populations we preformed bulk RNA-seq on purified cord blood and JMML HSCs, GMPs, and JMML double positives (+/+).   We identified a conserved molecular hierarchy in JMML with high stem cell gene expression in HSCs, high progenitor and cell cycle gene expression in GMPs.  We identified enriched gene sets in JMML HSCs vs CB HSCs, and find a number of potential therapeutic targets expressed in JMML.

overall_design: mRNA sequencing of human cord blood and JMML HSPC populations

first: Benjamin

last: Povinelli


title: Molecular and Functional Characterization of Disease-Propagating Stem Cells in Juvenile Myelomonocytic Leukemia [single cell RNA-seq]

summary: We report on the characterization of  JMML hematopoietic stem and progenitor cells (HSPCs).  In order to characterize the single cell heterogeneity in comparison to normal cord blood we preformed droplet based single cell RNA-seq of FACS sorted CD34+ cells using the 10x genomics platform.  We identified distinct clustering of unique JMML HSPC populations in relation to cord blood control.

overall_design: single cell mRNA sequencing of human cord blood and JMML CD34+ HSPCs


title: Alternative splicing and the epigenome in CML remission [WGBS]

city: Sydney

first: Charles

last: Bailey

state: NSW

address: Missenden Rd, Building 93

country: Australia

institute: Centenary Institute

type: Methylation profiling by high throughput sequencing

summary: Transcriptomic and epigenomic profiling of matched CML diagnosis/remission samples revealed a reconfiguration of gene expression and DNA methylation at remission exhibiting patterns similar to those observed in healthy individuals. In contrast, alternative splicing retains chronic phase-like abnormal patterns. Most dramatic dissimilarities between remission and healthy control samples were observed in intron retention. While reduced DNA methylation around splice sites could explain increased intron retention at diagnosis, maintenance of high intron retention levels at remission has other causes, such as reduced splicing factor expression and histone modifications.

overall_design: mRNA sequencing was performed on 16 CML diagnosis and 11 matched remission samples as well as 6 control samples. Whole genome bisulfite sequencing was performed on 10 matched diagnosis/remission samples and 6 healthy controls.

first: Ulf

last: Schmitz

first: Jaynish

middle: S

last: Shah

first: Bijay

last: Dhungel

first: Geoffray

last: Monteuuis

first: Phuc

middle: L

last: Luu

first: Veronika

last: Petrova

first: Cynthia

last: Metierre

first: Shalima

last: Nair

first: William

last: Ritchie

middle: G

first: Ali

last: Turhan

first: Susan

last: Branford

first: Timothy

middle: P

last: Hughes

last: Clark

first: Justin

last: Wong

first: John

middle: E

last: Rasko


title: Profiling non-small cell lung carcinoma cell line PC9 treated with erlotinib

first: Elizaveta

last: Benevolenskaya

summary: The non-small cell lung carcinoma (NSCLC) PC9 cell line is an established preclinical model for tyrosine kinase inhibitors. To be able to better understand the differences in response between individual cells, we performed treatment of PC9 cells grown in cell culture with erlotinib followed by Drop-seq. We were able to clearly distinguish cells that were treated with the drug for five different time points from untreated cells, and we discovered different cell populations within the treated cells, likely reflecting heterogeneity of drug resistant cells. We were able to identify specific biomarkers, as preferentially expressed genes, for each cell population. The results of our study will address preexisting and acquired drug resistance that limits clinical usefulness of targeted strategies, particularly in NSCLC.

overall_design: Examining drug resistant cell populations using single cell RNA-seq

first: Alexandre

last: Aissa

first: Abul

last: Islam


title: Drug holiday expression profiling of non-small cell lung carcinoma cell line PC9 treated with erlotinib

summary: The non-small cell lung carcinoma (NSCLC) PC9 cell line is an established preclinical model for tyrosine kinase inhibitors. To be able to better understand the differences in response between individual cells to drug removal after continuous treatment, we performed treatment of PC9 cells grown in cell culture with erlotinib followed by Drop-seq. We were able to clearly distinguish cells that were treated with the drug, removed from the drug and then re-treated with the same drug, and we discovered different cell populations within the treated cells, likely reflecting heterogeneity of drug resistant cells. We were able to identify specific biomarkers, as preferentially expressed genes, for each cell population. The results of our study will address drug resistance that limits clinical usefulness of targeted strategies, particularly in NSCLC.


title: Biphasic activation of WNT signaling enhances the derivation of midbrain dopamine neurons for translational use.

status: Public on Dec 18 2020

city: New York

first: Doron

last: Betel

state: NY

address: 1305 York Avenue (Box 140)

department: Medicine

institute: Weill Cornell Medical College


title: Drug resistant changes at the level of single cells


title: Different forms of KLF5 regulate distinct sets of genes in prostate cancer cells

first: Jin-Tang

last: Dong


title: Different forms of KLF5 transcriptionally regulate distinct sets of genes in prostate cancer cells [ChIP-Seq]

summary: To identify which genes are directly regulated by KLF5 and its acetylation, we performed ChIP-Seq and bioinformatics analyses in KLF5-null prostate cancer cells expressing KLF5, KLF5KR, and KLF5KQ.

overall_design: ChIP-Seq analyses were performed using KLF5 knockout DU 145 and PC-3 cells that ectopically expressed empty vector (EV), wildtype KLF5 (KLF5); acetylation deficient mutant KLF5K369R (KR), and acetylation mimicking mutant KLF5K369Q (KQ).

first: Baotong

last: Zhang

first: Yixiang

last: Li

first: Qiao

last: Wu

first: Benjamin

last: Barwick


title: Peroxisomal retrograde signaling in C. elegans

city: Ulsan

first: Stéphane

middle: G.

last: Rolland

address: UNIST-Gil 50, Eonyang-eup, Ulju-gun

country: South Korea

institute: IBS Center for Genomic Integrity

summary: Distinct retrograde signaling pathways have been identified for several cellular organelles. These pathways are important to maintain the function of these organelles in response to organelle-specific stress. Using Caenorhabditis elegans, we show for the first time that such a retrograde signaling also exists for peroxisomes. Analysis of the C. elegans transcriptome revealed that peroxisomal import stress caused by the knock-down of the peroxisomal matrix protein import receptor prx-5/PEX5 induces the compensatory up-regulation of genes involved in defense response and lipid metabolic processes, especially peroxisomal beta oxidation. We, therefore, propose that the peroxisomal retrograde signaling participates in the maintenance of peroxisomal function in response to peroxisomal import stress.

overall_design: RNA-seq analysis of tag-208(RNAi) (control) and prx-5(RNAi) treated samples in three biological replicates.

first: Elisabeth

last: Rackles

first: Judith

last: Zacherl


title: MYC Promotes Cancer Progression by Modulating m6A Modification to Suppress Target Gene Translation

city: He Fei

first: Shi-Ting

state: An Hui

address: No.443, Huangshan Road

department: Innovation Center for Cell Biology

institute: University of Science and Technology of China

summary: N6-Methyladenosine (m6A) is the most prevalent internal modification in mammalian mRNAs, and participates in various fundamental bioprocesses as well as cancer. Here we immunoprecipitated m6A methylated poly (A+) RNAs (MeRIP) and then sequenced and profiled mRNA m6A methylation in P493-6 cells.

overall_design: To investigate the effects of MYC on mRNA m6A modification, we used the human P493-6 B cell model of Burkitt’s lymphoma. P493-6 cells carry a MYC tetracycline (Tet)-off system, which enables the generation of cells with low or high MYC expression. P493-6 cells that were treated with Tet for 72 hours or 0 hour with 1 ug/ml Tet.

first: Gongwei

first: Shengqi

last: Shen

first: Yun-gui

last: Yang

first: Ping

last: Gao


title: Profiling non-small cell lung carcinoma cell line PC9 treated with erlotinib using 10x Genomics

first: Elizaveta

summary: The non-small cell lung carcinoma (NSCLC) PC9 cell line is an established preclinical model for tyrosine kinase inhibitors. To be able to better understand the differences in response between individual cells, we performed treatment of PC9 cells grown in cell culture with erlotinib followed by 10x Genomics. We were able to clearly distinguish cells that were treated with the drug from untreated cells, and we discovered different cell populations within the treated cells, likely reflecting heterogeneity of drug resistant cells. We were able to identify specific biomarkers, as preferentially expressed genes, for each cell population. The results of our study will address preexisting and acquired drug resistance that limits clinical usefulness of targeted strategies, particularly in NSCLC.


title: Mobilized multipotent hematopoietic progenitors stabilize and expand regulatory T-cells

city: Vandoeuvre-lès-Nancy

first: Sebastien

last: Hergalant

address: Faculté de Médecine. 9 avenue de la forêt de Haye

department: Bioinformatics

country: France

institute: INSERM

summary:
Objective: Achieving immunoregulation via in vivo expansion of Foxp3+ regulatory CD4+ T cells (Treg) remains challenging. Considerable efforts are being produced aiming at promoting expansion of Tregs in vitro with IL-2, rapamycin, activation with anti-CD3/CD28 mAb-coated beads or with minute foreign antigen doses, for subsequent administration to  patients with autoimmune and alloimmune diseases. We have previously shown that mobilized multipotent hematopoietic progenitors (MPPs)collected in  peripheral blood stem cell grafts, have the capacity to enhance Treg proliferation in vivo.                                                                                Design: To analyze the possible mechanisms involved in mouse CD4+CD25high regulatory T-cells (Treg cells) expansion promoted by mobilized MPPs (MPPs), we sequenced and compared the transcriptome of activated Treg extracted from simple culture or from co-culture with MPPs.
Results: Tregs cultivated with mobilized MPP had a specialized profile, with enhanced expression of sets of genes coding for receptors and transcription factors that stabilize their regulatory function particularly in inflammatory settings including CD4, CD25 (Il2ra), Foxp3, CD127low (Il7ralpha), Ctla4 high, Tnfrsf18 (Gitr) high, Ikzf2 (Helios) high, Tnfrsf4 (Ox40/CD134) high, Itgae (CD103), regulated by Satb1, Runx, Gata3. Moreover, they exhibit enhanced expression of Prdm1, recently reported to prevent methylation of Foxp3 within Tregs in central nervous system inflammation (14), and Nr4a, that stabilizes Tregs against their differentiation into effector T-cells (15). Their survival is improved by a reduced apoptosis controlled by reduced Tnfrsf1b (p75, Tnfr2) signaling pathway and FOXO-mediated transcription. Their cell cycle is arrested at the G1 stage through enhanced Cdkn1a (p21) and regulated by Runx3 and p53.                                                                                conclusion: Tregs interacting with mobilized MPP are shaped, stabilized with high immunosuppressive properties.

overall_design: 6 samples divided in 2 paired groups of 3 biological replicates: Tregs alone, activated by anti-CD3 and anti-CD28 (Treg), and Tregs activated by anti-CD3 and anti-CD28, co-cultured with hematopoietic multipotent progenitor (TH)

first: Maud

last: D'Aveni

first: Nicolas

last: Cagnard

first: Sébastien

first: Flora

last: Zavala


title: p53-induced RNA-binding protein ZMAT3 is splicing regulator that inhibits the splicing of oncogenic CD44 variants in CRC

city: Bethesda

first: Markus

last: Hafner

state: MD

address: 50 South Drive

institute: NIAMS/NIH


title: 19-11-D Gene expression levels in Ninj1 KO BMDMs

city: South San Francisco

first: Rohit

last: Reja

state: CA

address: 1 DNA Way

institute: Genentech

summary: We have identified mouse Ninj1 as a protein responsible for plasma membrane integrity following cell death stimuli. Ninj1 KO BMDMs have a distinct bubble morphology wherein cell cytoplasmic contents fail to be released following cell death. To determine contributing factors to Ninj1 KO morphology, we performed RNAseq in WT and Ninj1 KO murine BMDMs with or without priming.

overall_design: RNA was extracted RNAseq from primary mouse bone marrow derived macrophages (BMDMs) from WT C57BL/6 and Ninj1 KO mice

first: Yue


title: Biphasic activation of WNT signaling enhances the derivation of midbrain dopamine neurons for translational use [ChIP-seq]

first: Doron

last: Betel

summary: We report a novel midbrain dopamine neuron derivation protocol from human pluripotent stem cells (hPSC)s based on a two-step WNT signaling activation strategy. We perform genome-wide chromatin immunoprecipitation (ChIP-seq) for histone modifications including H3K4me3 and H3K27me3 with midbrain dopamine neuron differentiated cells from hPSC in Low-Chir and Boost-Chir (7.5 uM) conditions at day 11. Those results indicate that CHIR-boost promotes a more open chromatin structure at midbrain regions such as an EN1 locus based on increased H3K4me3 and decreased H3K27me3 levels. In contrast, analysis of the contaminating posterior (hindbrain) and anterior (diencephalic) lineage including NKX2-2 and PAX6 loci showed the opposite pattern with decreased H3K4me3 and increased H3K27me3 levels under CHIR-boost conditions. The protocol presented here is the basis for clinical grade dopamine neuron production and preclinical safety and efficacy studies.

overall_design: Examination of 2 different histone modifications at day 11 midbrain dopamine differentiated cells from human pluripotent stem cell in Low-Chir and Boost-Chir (7.5uM) conditions.


title: p53-induced RNA-binding protein ZMAT3 is splicing regulator that inhibits the splicing of oncogenic CD44 variants in CRC [RNA-seq_ZMAT3]

summary:
Purpose: p53 is a well-studied protein in which the activation of its main targets is not enough to explain its tumor suppressor effect. Aiming to search for direct p53 targets that mediate less studied processes within p53 pathway, like post-transcriptional regulation, we selected ZMAT3 as the RNA-binding protein most upregulated by p53 activation.
Methods: We investigated ZMAT3 targets by using PAR-CLIP and RNA-seq in colorectal cancer cell line HCT116.
Results.txt: We found that ZMAT3 binds to ~5000 genes mainly in their introns. It also causes alternative splicing and CD44 was its mostly altered target. ZMAT3 knock-down increased CD44v2-v10 and CD44v3-v10, but decreased CD44s expression. Increased survival and clonogenicity was observed upon knock-down of ZMAT3, which was rescued upon concurrent knock-down of CD44v
Conclusions: We propose that ZMAT3 regulates splicing of CD44 following activation by p53, which upregulates CD44s at the expense of CD44v isoforms and leads to a decrease tumorigenicity.

overall_design: mRNA profiles from HCT116 upon siCTRL, siZMAT3 (48 hours) and treatment with Nutlin-3a or DMSO for 6 hours

first: Bruna

middle: R

last: Muys

first: Dimitrios

last: Anastasakis

first: Duncan

last: Claypool

first: Lorinc

last: Pongor

first: Xiao

first: Ioannis

last: Grammatikakis

first: Minxue

last: Liu

first: Xiantao

last: Wang

first: Kannanganattu

last: Prasanth

first: Mirit

middle: I

last: Aladjem

first: Ashish

last: Lal


title: Comparison of peroxide-inducible gene expression in cells with native or redox-insensitive Stat3

city: Nottingham

first: Fei

last: Sang

address: QMC

department: School of Life Sciences

institute: University of Nottingham

summary: To explore the biological role of Stat3 oxidation we created murine embryonic fibroblasts (MEFs) that express either WT-Stat3 or a redox-insensitive mutant of Stat3 (Stat3-C3S). The Stat3-C3S cells differed from WT-Stat3 cells in morphology, proliferation and resistance to oxidative stress; in response to cytokine stimulation they displayed elevated Stat3 tyrosine phosphorylation and Socs3 expression, implying that Stat3-C3S is insensitive to oxidative inhibition.

overall_design:
Examination of gene expression profiles in control and peroxide-stimulated WT-Stat3 and Stat3-C3S MEFs
Comparative analysis of global gene expression in WT-Stat3 and Stat3-C3S cells revealed differential expression (DE) of genes both under basal conditions and during oxidative stress. Using differential gene regulation pattern analysis, we identified 199 genes clustered into 10 distinct patterns that are selectively responsive to Stat3 oxidation. GO term analysis identified down-regulated genes to be enriched for tissue/organ development and morphogenesis and up-regulated genes to be enriched for cell-cell adhesion, immune responses and transport related processes. Although most DE gene promoters contain consensus Stat3 inducible elements (SIEs), our chromatin immunoprecipitation (ChIP) and ChIP-seq analyses did not detect Stat3 binding at these sites in control or oxidant-stimulated cells, suggesting that oxidised Stat3 regulates these genes indirectly

first: Carolyn

last: Palmer

first: Martin

last: Blythe

first: Rahul

last: Bhosale

first: Peter

last: Shaw


title: Enhanced IL-36R signaling promotes barrier impairment and inflammation in skin and intestine

city: Tarrytown

first: Ruoyu

last: Zhang

address: 777 Old Saw Mill River Road

institute: Regeneron

summary: Deficiency in IL-36R antagonist caused by loss of function mutations in IL-36RN leads to DITRA (1), a rare inflammatory human disease that belongs to a subgroup of Generalized Pustular Psoriasis (GPP). Herein, we report a novel functional genetic mouse model of DITRA with enhanced IL-36R signaling analogous to one observed in DITRA patients which supports and provides new insight in our understanding of IL-36 family of molecules in regulating barrier integrity across multiple tissues. Humanized DITRA-like mice displayed increased skin inflammation in preclinical model of psoriasis, and in vivo blockade of IL-36R pathway using anti-human IL-36R antibody ameliorated IMQ-induced skin pathology at both prophylactic and therapeutic treatments. Deeper characterization of the humanized DITRA-like mice revealed that deregulated IL-36R signaling promoted tissue pathology during intestinal injury and led to impairment in mucosal restoration in the repair phase of chronic DSS-induced colitis. Blockade of IL-36R pathway significantly ameliorated DSS-induced intestinal inflammation and rescued the inability of DITRA-like mice to recover from mucosal damage in vivo. Thus, our results indicate a central role for IL-36 in regulating the pro-inflammatory responses in the skin and epithelial barrier function in the intestine suggesting a new therapeutic potential for IL-36R axis in psoriasis and at the later stages of intestinal pathology.

overall_design: RNA-seq of skin of IMQ-treated DITRA-like mice injected with PBS, anti-human IL-36R and anti-mouse IL-12p40 mAbs. RNA-seq of colon of DSS-treated DITRA-like mice injected with PBS, anti-human IL-36R and anti-mouse IL-12p40 mAbs.

first: Zaruhi

last: Hovhannisyan

first: Nengyin

first: Sara

last: Khalil-Aguero

first: Casandra

last: Panea

first: Jeffrey

last: VanValkenburgh

first: Wei

middle: K

last: Lim

first: Yu

last: Bai

first: Wen

last: Fury

first: Tammy

last: Huang

first: Elena

last: Garnova

first: Jeanette

last: Fairhurst

first: Jee

last: Kim

first: Smita

last: Aryal

first: Dharani

last: Ajithdoss

first: Adelekan

last: Oyejide

first: Maria

last: del Pilar Molina-Portela

first: E

last: Hock

first: William

last: Poueymirou

first: Nicole

last: Stokes Oristian

first: Susannah

last: Brydges

first: Xia

last: Olson

first: George

last: Yancopoulos

first: Andrew

last: Murphy

first: Matthew

last: Sleeman

first: Sokol

last: Haxhinasto


title: Single cell RNA sequencing analysis of freshly resected non-small cell lung carcinoma (NSCLC) tissues

first: Elizaveta

middle: V

last: Benevolenskaya

summary: Lung cancer is the leading cause of cancer-related deaths world-wide. ~85% of lung carcinomas are non–small cell lung carcinoma (NSCLC). Tumor cell heterogeneity is very poorly defined. However, it is known to be important for tumor response to cancer therapy and cancer agressivenes. We subjected three NSCLC tumors resected from different patients to Drop-seq in order to 1) elucidate the capability of scRNA-seq analysis in identifying different tumor cell populations; and 2) ascertain the clinical value of the genes which distinguish cancer cells from other cells in the tissue. As anticipated, the tissue composition of independently collected samples varied. Despite deficient populations in some samples, both donor and patient samples contributed to the majority of cell populations. However, cancer cells were all patient-specific. These findings emphasize the utility of single cell gene expression data in identification of tumor cell populations. The collected data might be further used for predicting of drugs specific to the biology of activated pathways and patient outcome.

overall_design: Examining NSCLC tumor cell populations using single cell RNA-seq

first: Alexandre

middle: F

last: Aissa

first: Abul

last: BIslam


title: p53-induced RNA-binding protein ZMAT3 is splicing regulator that inhibits the splicing of oncogenic CD44 variants in CRC [PAR-CLIP_ZMAT3]

first: Markus

last: Hafner

summary:
Purpose: p53 is a well-studied protein in which the activation of its main targets is not enough to explain its tumor suppressor effect. Aiming to search for direct p53 targets that mediate less studied processes within p53 pathway, like post-transcriptional regulation, we selected ZMAT3 as the RNA-binding protein most upregulated by p53 activation.
Methods: We investigated ZMAT3 targets by using PAR-CLIP and RNA-seq in colorectal cancer cell line HCT116.
Results: We found that ZMAT3 binds to ~5000 genes mainly in their introns. It also causes alternative splicing and CD44 was its mostly altered target. ZMAT3 knock-down increased CD44v2-v10 and CD44v3-v10, but decreased CD44s expression. Increased survival and clonogenicity was observed upon knock-down of ZMAT3, which was rescued upon concurrent knock-down of CD44v
Conclusions: We propose that ZMAT3 regulates splicing of CD44 following activation by p53, which upregulates CD44s at the expense of CD44v isoforms and leads to a decrease tumorigenicity.

first: Bruna

last: Muys

first: Dimitrios

last: Anastasakis

first: Duncan

last: Claypool

first: Lorinc

last: Pongor

first: Xiao

last: Li

first: Ioannis

last: Grammatikakis

first: Minxue

last: Liu

first: Xiantao

last: Wang

first: Kannanganattu

last: Prasanth

first: Mirit

last: Aladjem

first: Ashish

last: Lal


title: Single cell RNA-seq profiling of erlotinib-resistant PC9 cells

summary: The non-small cell lung carcinoma (NSCLC) PC9 cell line is an established preclinical model for tyrosine kinase inhibitors. To be able to better understand the differences in response between individual cells, we performed treatment of PC9 cells grown in cell culture with erlotinib followed by Drop-seq. We were able to clearly distinguish cells that were treated with the drug for 11 days from untreated cells, and we discovered different cell populations within the treated cells, likely reflecting heterogeneity of drug resistant cells. We were able to identify specific biomarkers, as preferentially expressed genes, for each cell population. The results of our study will address preexisting and acquired drug resistance that limits clinical usefulness of targeted strategies, particularly in NSCLC.

middle: B

last: Islam

first: Majd

middle: M

last: Ariss


title: Gene expression profiling of erlotinib-resistant PC9 cells.

last: Benevolenskaya

summary: The non-small cell lung carcinoma (NSCLC) PC9 cell line is an established preclinical model for tyrosine kinase inhibitors. To better understand gene expression changes in cells that survived the inhibitor treatment, we treated the EGFR-mutant PC9 cells with erlotinib, isolated RNA, and performed RNA-seq analysis. We were able to identify genes that are differentially expressed in erlotinib-treated cells compared to untreated. The results of this study will be integrated with single cell RNA-seq to address the utility of bulk RNA versus single cell RNA strategies in identifying biomarkers of drug resistance.

overall_design: examining changes associated with drug resistance using RNA-seq

first: Cammille

last: Go


title: Gene expression of CD4+ T cells

city: Martinsried

first: Tobias

last: Straub

address: Großhadener Str. 9

department: Biomedical Center, Bioinformatics

country: Germany

institute: LMU Munich

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


title: Genome-wide map of H3K4me3 in erlotinib-resistant PC9 cells.

first: Elizaveta

summary: The non-small cell lung carcinoma (NSCLC) PC9 cell line is an established preclinical model for tyrosine kinase inhibitors. Recent evidence suggests that the H3K4 demethylase KDM5A protein is required for emerging drug-resistant cell populations. To better understand the dependence of cell survival on H3K4me3 level, we treated the EGFR-mutant PC9 cells with erlotinib and performed ChIP-seq analysis using H3K4me3 antibodies. We were able to identify genomic regions that exhibit decreased H3K4me3 in drug-treated cells. The results of this study will help to address clinical usefulness of KDM5A-targeted strategies in overcoming drug resistance, particularly in NSCLC.


title: Assessing efficiency of single cell population by Drop-seq using cell mixture of epithelial and myeloid cells.

summary: A lung epithelial cell line was mixed with monocytes/marcophages, and then cells were separated using specific antibodies and/or Drop-seq. PC9 is a non-small cell lung carcinoma (NSCLC) cell line. Myeloid lineage cell line U937 are known to differentiate along monocyte/marcophage lineage by TPA. Using Drop-seq, we were able to clearly distinguish PC9 from U937 cells in the mixture. The separation by Drop-seq was correlated with the separation by cell type-specific antibodies. We were able to identify specific biomarkers, as preferentially expressed genes, for each cell population. The results of our study addresses the quality of cell separation by Drop-seq.

overall_design: efficincy of cell separation using single cell RNA-seq

first: Alexandre

last: Aissa

first: Abul


title: Profiling melanoma cell line M14 treated with vemurafenib

summary: The BRAFV600E mutant melanoma cell line M14 responds to the BRAF inhibitor vemurafenib, however, the differences in response between individual cells are underlying resistance. To highlight the underlying mechanisms, we performed treatment of M14 cells with vemurafenib, followed by Drop-seq. We were able to clearly distinguish cells that were treated with the drug from untreated cells, and we discovered different cell populations within the treated cells, likely reflecting heterogeneity of drug resistant cells. We were able to identify specific biomarkers, as preferentially expressed genes, for each cell population. The results of our study will address preexisting and acquired drug resistance that limits clinical usefulness of targeted strategies.


title: Oscillating and stable promoter-enhancer loops underlie core- and clock-controlled gene expression rhythms in mouse liver

city: Lausanne

first: Felix

last: Naef

address: AAB 040 Station 15

department: Bioengineering

country: Switzerland

institute: École Polytechnique Fédérale de Lausanne

summary: The circadian clock dynamically rewires promoter-enhancer loops in tissues to drive robust daily rhythms in gene transcription and locomoter activity.

overall_design: Total RNA and crosslinked genomic DNA extracted from mouse tissues during the dirunal cycle. Mouse genotype conditions include WT, Bmal1 KO, and Cry1Intron KO.

first: Jerome

last: Mermet

first: Jake

last: Yeung


title: Profiling non-small cell lung carcinoma cell line HCC827 treated with erlotinib

city: Chicago

address: 900 S. Ashland Ave.

institute: University of Illinois at Chicago

summary: The non-small cell lung carcinoma (NSCLC) HCC827 cell line is an established preclinical model for tyrosine kinase inhibitors. To be able to better understand the differences in response between individual cells, we performed treatment of HCC827 cells grown in cell culture with erlotinib followed by Drop-seq. We were able to clearly distinguish cells that were treated with the drug from untreated cells, and we discovered different cell populations within the treated cells, likely reflecting heterogeneity of drug resistant cells. We were able to identify specific biomarkers, as preferentially expressed genes, for each cell population. The results of our study will address preexisting and acquired drug resistance that limits clinical usefulness of targeted strategies, particularly in NSCLC.


title: Gene expression of CD4+ T cells in response to antigen persisting at different doses for different times

summary:
Chronic infections and cancer can induce a dsyfunctional immune response which cannot clear the cause of disease. We asked how antigen persistence alone affects T helper cell reactivity v. exhaustion and compared the gene expression of CD4+ T cell responding to antigen persisting at different levels for different periods of time.
We used microarrays to detail gene expression underlying T helper cell dysfunctionality evolving in response to different levels of antigen at two timepoints.

overall_design: Allogenically marked AND TCR transgenic CD4+ T cells were adoptively transferred into recipients in which the presentation of a moth cytochrome c epitope can be regulated by doxycycline.

first: Anne

last: Trefzer

first: Pallavi

last: Kadam

first: Shu-Hung

first: Stefanie

last: Pennavaria

first: Benedikt

last: Lober

first: Jan

last: Kranich

first: Thomas

last: Brocker

first: Martin

last: Irmler

first: Johannes

last: Beckers

first: Reinhard

last: Obst


title: Drosophila Fezf functions as a transcriptional repressor to direct layer specific synaptic connectivity in the fly visual system

status: Public on Dec 17 2020

city: Boston

first: Jing

last: Peng

state: MA

address: 220 Longwood Ave

department: Neurobiology

institute: Harvard Medical School

overall_design: Refer to individual Series


title: Biphasic activation of WNT signaling enhances the derivation of midbrain dopamine neurons for translational use [RNA-seq]

first: Doron

last: Betel

summary: We report a novel midbrain dopamine neuron derivation protocol from human pluripotent stem cells (hPSC)s based on a two-step WNT signaling activation strategy. By comparing next-generation sequencing (NGS) derived transcriptome profiling (RNA-seq) of dopamine neuron differentiated cells derived from hPSC with different concentration of Chir-boosting (day4 – day9) measured at day 11 including hPSC, we find that CHIR-boosting improves expression of midbrain markers such as EN1 while minimizing expression of contaminating posterior (hindbrain) and anterior (diencephalic) lineage markers. Furthermore, gene expression analysis from RNA-seq study confirmed the progressive differentiation and maturation of Boost CHIR treated precursors into mDA neurons by day 30 of differentiation. The protocol presented here is the basis for clinical grade dopamine neuron production and preclinical safety and efficacy studies.

overall_design: mRNA profiles of human pluripotent stem cells (hPSC)s, midbrain dopamine differentiated cells with different concentration of CHIR-boosting (day 4 -  day 9) at day 11, and CHIR-BOOST (7.5uM) treated mDA neurons at day 20 and day 30 derived from hPSCs.


title: Comparison of Stat3 chromatin distributions in cytokine v peroxide-stimulated MEFs

first: Fei

last: Sang

summary: To explore the biological role of Stat3 oxidation we created murine embryonic fibroblasts (MEFs) that express either WT-Stat3 or a redox-insensitive mutant of Stat3 (Stat3-C3S). The Stat3-C3S cells differed from WT-Stat3 cells in morphology, proliferation and resistance to oxidative stress; in response to cytokine stimulation they displayed elevated Stat3 tyrosine phosphorylation and Socs3 expression, implying that Stat3-C3S is insensitive to oxidative inhibition.  Comparative analysis of global gene expression in WT-Stat3 and Stat3-C3S cells revealed differential expression (DE) of genes both under basal conditions and during oxidative stress. Using differential gene regulation pattern analysis, we identified 199 genes clustered into 10 distinct patterns that are selectively responsive to Stat3 oxidation. GO term analysis identified down-regulated genes to be enriched for tissue/organ development and morphogenesis and up-regulated genes to be enriched for cell-cell adhesion, immune responses and transport related processes. Although most DE gene promoters contain consensus Stat3 inducible elements (SIEs), our chromatin immunoprecipitation (ChIP) and ChIP-seq analyses did not detect Stat3 binding at these sites in control or oxidant-stimulated cells, suggesting that oxidised Stat3 regulates these genes indirectly.

overall_design:
Examination of Stat3 chromatin binding profiles in MEFs stimulated with LIF or peroxide
Please note that each processed data file was generated from both replicates and is linked to the corresponding rep1 sample.

first: Michela

last: Grillo

first: Nadine

last: Holmes

first: Peter

last: Shaw


title: Whole-genome mapping of RUNX1, CBFA2T3, H3K4me1, H3K4me3, H3k27me3 and H3K27ac in pre-B lymphoblast models

city: Rennes

first: Gilles

last: Salbert

address: 263 Av. Gal. Leclerc

department: IGDR UMR6290

institute: University Rennes 1

summary: Genome binding/occupancy profiling of Runt Related Transcription Factor 1 (RUNX1), CBFA2T3 (ETO2, MRG16) and histone marks by high throughput sequencing. RUNX1 and CBFA2T3 are found overexpressed in pediatric t(12;21) ETV6-RUNX1 B cell precursor acute lymphoblastic leukemia. As CBFA2T3 is a transcription regulator, we hypothesized a potential collaboration between the transcription factor RUNX1 and CBFA2T3. Here, we investigated the molecular proximity between RUNX1 and CBFA2T3 proteins on chromatin and identified a regulatory loop between RUNX1 and CBFA2T3.

overall_design: RUNX1, CBFA2T3, H3K4me1, H3K4me3 and H3K27ac genome-wide distributions were determined using ChIP-seq. Cells used in this study are pre-B lymphoblast REH and Nalm6 cell lines and blasts from B-cell precursor acute lymphoblastic leukemia (BCP-ALL) patient. ChIP samples and libraries were done by Sérandour A.A and Debaize L. High-throughput sequencing was conducted on Illumina HiSeq 1500/2500 machines at the Human and Environmental Genomics' platform of Rennes (Biogenouest génomique, Rennes, France).

first: Marie

last: Troadec

first: Lydie

last: Debaize


title: Gene expression of control and anergic CD4+ T cells in the mouse

first: Tobias

last: Straub

summary:
Anergic T cells are unable to respond to antigen though their TCR is specific for it.
Chronic infections and cancer can induce a dsyfunctional immune response which cannot clear the cause of disease. We asked in how far anergic T cells are similar to antigen-exhausted ones.

overall_design: Anergic CD4+ cells carrying the markers FR4 and CD73 were sorted twice from B10.BR spleens and lymph nodes directly into Trizol LS. Control cells were CD4+ and did not express the markers.

first: Anne

last: Trefzer

first: Pallavi

last: Kadam

first: Shu-Hung

last: Wang


title: Nascent chain interactions of proximal ribosomes drive homo-oligomer assembly

city: Heidelberg

first: Kai

last: Fenzl

address: Meyerhofstraße 1

institute: EMBL

summary: Accurate assembly of newly- synthesized proteins into functional oligomers is crucial for cell activity. In this study, we investigated whether direct interaction of two nascent proteins, emerging from nearby ribosomes (co-co assembly), constitutes a general mechanism for oligomer formation. We used a proteome-wide screen to detect nascent chain-connected ribosome pairs and identified hundreds of homomer subunits that co-co assemble in human cells. Interactions are mediated by five major domain classes, among which N-terminal coiled coils are the most prevalent. We were able to reconstitute co-co assembly of nuclear lamin in Escherichia coli, demonstrating that dimer formation is independent of dedicated assembly machineries. Co-co assembly may thus represent an efficient way to limit protein aggregation risks posed by diffusion-driven assembly routes and ensure isoform-specific homomer formation.

sample_organism:
Escherichia coli
Homo sapiens

platform_organism:
Escherichia coli
Homo sapiens

overall_design: Screen of nascent chain dimerization in two human cell lines (HEK293-T and U2OS, 2 replicates each). Total translatomes from HEK293-T cells are also provided in 2 replicates. Similar screen performed with treatment of HEK293-T cell lysates with puromycin (2 replicates) or a titration of Proteinase K (4 conditions, 1 replicate). Similar screen performed in E.coli over-expressing a human gene candidate (5 genes tested, 4 of which are performed in 2 replicates).

first: Matilde

last: Bertolini

first: Florian

last: Wruck

first: Ilia

last: Kats

first: Frank

last: Tippmann

first: Jaro

last: Schmitt

first: Josef

last: Auburger

first: Sander

last: Tans

first: Bernd

last: Bukau

first: Günter

last: Kramer


title: ATAC-seq in NOTCH1-induced mouse T-ALLs

city: New Brunswick

first: Daniel

last: Herranz

state: New Jersey

address: 195 Little Albany Street

institute: Rutgers Cancer Institute of New Jersey

summary: Here we report the genome-wide open chormatin profile (ATACseq) of a NOTCH1-induced mouse T-cell Acute Lymphoblastic Leukemia

overall_design: ATACseq analysis of a mouse NOTCH1-induced leukemia (2 replicates)


title: Drosophila L3 RNA-seq at 24h APF

first: Jing

last: Peng

summary: The layered compartmentalization of functionally-related synaptic connections, a common feature of nervous systems, underlies proper connectivity between neurons and enables parallel processing of neural information. However, the stepwise development of layered neuronal connections is not well understood. The medulla neuropil of the Drosophila melanogaster visual system is comprised of 10 discrete layers (M1-M10), within which neural computations underlying distinct visual features are processed. As such, the Drosophila medulla neuropil serves as a model system for understanding layered compartmentalization of synaptic connectivity. The first step leading to the establishment of neurite layer specificity in the outer medulla (M1-M6) is the innervation of one of two broad, primordial domains that will subsequently expand and transform into the M1-M6 layers. We previously found that the transcription factor dFezf cell-autonomously directs L3 neurons to their proper early broad domain before they innervate and form synapses specifically within the M3 layer of the mature medulla. Here, we examined the transcriptomes of wild type and dFezf mutant L3 neurons by RNA-seq and identified downstream target genes that are regulated by dFezf. We show that dFezf controls L3 layer specificity through temporally precise transcriptional repression of the transcription factor slp1 (sloppy paired 1). During the period of broad domain selection, dFezf limits slp1 expression in L3 neurons. Slp1 is upregulated in dFezf-null L3 neurons, and ablation of slp1 fully rescues the targeting defect observed in dFezf-null L3 growth cones. Surprisingly, L3 innervation of the M3 layer after broad domain selection requires early slp1 expression. DFezf thus functions as a transcriptional repressor to coordinate the temporal dynamics of a transcriptional cascade that orchestrates sequential steps of layer-specific synapse formation. Moreover, we performed ATAC-seq in wild type L3 neurons and identified the accessible regions in the L3 genome, where dFezf may directly bind and regulate gene expression.

overall_design: Fluorescently-labeled wild type and dFezf1 L3 neurons were isolated by FACS at 24h APF, and cDNA libraries were then prepared from total RNA and sequenced using an Illumina platform. Three biological replicates of each genotype were performed, and all the cDNA libraries were prepared and sequenced at the same time to minimize technical variability.


title: Drosophila L3 ATAC-seq at 24h APF

overall_design: Fluorescently-labeled wild type L3 neurons were isolated by FACS at 24h APF. DNA libraries were made from genomic DNA and sequenced using an Illumina platform. Three biological replicates were performed. All three libraries were prepared and sequenced at the same time to minimize technical variability.


title: Combinatorial transcription factor activities on open chromatin induce embryonic heterogeneity in vertebrates

city: Nijmegen

first: Ann

middle: Rose

last: Bright

address: Geert Grooteplein Zuid 30

department: Department of Molecular Biology

country: Netherlands

institute: RIMLS

summary: We applied chromatin accessibility and single cell transcriptome analyses to explore the emergence of heterogeneity and underlying gene-regulatory mechanisms during early gastrulation in Xenopus. TF activity was inferred by integrating chromatin accessibility and single cell transcriptomics. Our results show that combinatorial activity of zygotically expressed transcription factors acts on maternally-regulated accessible chromatin to induce organizer gene expression.

overall_design: ATAC-seq on Xenopus embryos (stage 9, 10.5, 12 and 16) and gastrula stage embryo explants (Animal cap and Dorsal marginal zone). RNA-seq on animal caps overexpressed with Foxb1, Foxb1 + Eomes, Eomes, Irx3, Irx3 + Otx2 and Lhx8 mRNA. ScRNA-seq on Animal cap and Dorsal marginal zone explants. scATAC-seq was performed on stage 10.5 Xenopus embryos.

first: Siebe

last: van Genesen

first: Qingqing

last: Li

first: Alexia

last: Grasso

first: Gert

middle: J

last: Veenstra


title: scRNAseq in Pten enhancer wild-type or deleted thymocytes, enriched for immature thymocyte stages (CD4-CD3-)

summary: Single cell suspensions of total thymocytes were obtained from Pten enhancer (PE) wild-type or knockout mice. This single-cell suspension was enriched in CD4-CD3- immature thymocyte progenitor cells. CD4-CD3- enriched thymocytes were then mixed 1:1 with single-cell suspensions from total unenriched thymocytes and subsequntly loaded in a 10x Chromium instrument for single-cell RNAseq analyses. Our results revealed Pten levels are signifcantly decreased in CD4-CD8- double negative (DN) thymocytes, CD8+ intermediate single positive (ISP) thymocytes and CD4+CD8+ double positive (DP) thymocytes in PE knockout mice, compared to PE wild-type mice.

overall_design: scRNAseq analysis of mouse thymus in two genotypes and one sample per genotype

first: Luca

last: Tottone

first: Olga

last: Lancho

first: Jui

middle: W

last: Loh

first: Amartya

last: Singh

first: Hossein

last: Khiabanian


title: p53-induced RNA-binding protein ZMAT3 is splicing regulator that inhibits the splicing of oncogenic CD44 variants in CRC [RNA-seq_UPF1]

first: Markus

last: Hafner

summary:
Purpose: p53 is a well-studied protein in which the activation of its main targets is not enough to explain its tumor suppressor effect. Aiming to search for direct p53 targets that mediate less studied processes within p53 pathway, like post-transcriptional regulation, we selected ZMAT3 as the RNA-binding protein most upregulated by p53 activation.
Methods: We investigated ZMAT3 targets by using PAR-CLIP and RNA-seq in colorectal cancer cell line HCT116.
Results.txt: We found that ZMAT3 binds to ~5000 genes mainly in their introns. It also causes alternative splicing and CD44 was its mostly altered target. ZMAT3 knock-down increased CD44v2-v10 and CD44v3-v10, but decreased CD44s expression. Increased survival and clonogenicity was observed upon knock-down of ZMAT3, which was rescued upon concurrent knock-down of CD44v
Conclusions: We propose that ZMAT3 regulates splicing of CD44 following activation by p53, which upregulates CD44s at the expense of CD44v isoforms and leads to a decrease tumorigenicity.

overall_design: mRNA profiles from HCT116 upon siCTRL, siZMAT3 (48 hours) and treatment with Nutlin-3a or DMSO for 6 hours

first: Bruna

last: Muys

first: Dimitrios

last: Anastasakis

first: Duncan

last: Claypool

first: Lorinc

last: Pongor

first: Xiao

first: Ioannis

last: Grammatikakis

first: Minxue

last: Liu

first: Xiantao

last: Wang

first: Kannanganattu

last: Prasanth

first: Mirit

last: Aladjem

first: Ashish

last: Lal


title: RNAseq in Pten enhancer wild-type or deleted mouse T-ALLs

first: Daniel

last: Herranz

summary: C57BL6 mice harboring Pten enhancer (PE) conditional knockout NOTCH1-induced leukemias were treated with vehicle (control) or tamoxifen to induce isogenic deletion of PE. Here we report the gene expression profile of leukemic blasts obtained from the spleen from control- or tamoxifen-treated leukemic mice. GSEA analysis show a significant enrichment of Pten signature genes, consistent with a role of PE in regulating Pten expression.

overall_design: RNAseq analysis of leukemic spleens in two genotypes and four or five samples per genotype.

first: Luca


title: A regulatory sub-circuit downstream of Wnt signaling controls developmental transitions in neural crest development

city: Ithaca

first: Marcos

last: Simoes-Costa

state: New York

address: 526 Campus Rd

department: Molecular Biology and Genetics

institute: Cornell University

summary: The process of cell fate commitment requires sequential changes in the gene expression profiles of embryonic progenitors. This is exemplified in the development of the neural crest, a migratory stem cell population derived from the ectoderm of vertebrate embryos. Neural crest formation involves a series of regulatory changes, in which cells adopt distinct transcriptional states in a stepwise manner. The mechanisms underpinning these shifts in cell identity are still poorly understood. Here we employ enhancer analysis to identify a genetic sub-circuit that controls developmental transitions in neural crest development. This sub-circuit links Wnt target genes in an incoherent forward loop that controls the sequential activation of genes in the neural crest lineage. By examining the cis-regulatory apparatus of Wnt effector gene AXUD1, we found that multipotency factor SP5 directly promotes neural plate border identity, while inhibiting premature specification by interacting with tissue specific enhancers.

overall_design: CUT&RUN data for SP5 in chicken neural crest cells

first: Ana

last: Azambuja


title: Androgen signaling in cell fate and cell-cell communication during early prostate development at single-cell resolution

city: Duarte

first: Zijie

last: Sun

address: 1500 E. Duarte Rd.

department: Cancer Biology

institute: City of Hope

summary: Androgens/androgen receptor (AR) mediated signaling pathways are essential for prostate development, morphogenesis, and regeneration.  Here, using newly generated ArIRES-Cre mice with single-cell RNA sequencing (scRNAseq) and other experimental approaches, we profiled the transcriptomes of more than 9000 cells isolated from murine E17.5 UGS tissues and identified a variety of urogenital mesenchyme (UGM) and epithelia (UGE) cell populations.  The expression of AR and other regulators were further assessed in different cell types at single cell resolution to determine their roles and interactions during early prostate development. Psuedotime analyses further explored the differentiation trajectory of UGM and UGE cell populations, and identified a Zeb1-expressing progenitor population.  Using mouse genetic tools, the decisive role of mesenchymal AR action in prostate initiation was further demonstrated.  These data provide fresh, high resolution, comprehensive insight into AR signaling and its roles in initiating dynamic interactions of with distinct signaling pathways between prostatic mesenchymal-epithelial cells.

overall_design: One sequencing dataset was analyzed containing tissue from 4 mice

first: Adam

last: Olson


title: Surviving climate change – the role of acclimatization in reef-building corals [16S]

city: University Park

first: Iliana

last: Baums

state: PA

address: 212 Mueller Laboratory, Penn State University

department: Biology

institute: The Pennsylvania State University

summary: Branching coral species like the Caribbean Acroporids are long lived and reproduce asexually via breakage of branches. Fragmentation is the dominant mode of local population maintenance for these corals across much of their range. Thus, large genets with many member ramets (colonies) are common. Each of the ramets experiences different microenvironments, especially with respect to light and water flow. Here, we investigate whether colonies that are members of the same genet have different epigenomes because of differences in their microenvironments. The Florida Keys experienced a large- scale coral bleaching event in 2014-2015 caused by high water temperatures. During the event, ramets of the same coral genet bleached differently. Previous work had shown that this was unlikely to be due to their eukaryotic algal symbionts (Symbiodinium ‘fitti’) because each genet of this coral species typically harbors a single strain of S. ‘fitti’. Characterization of the microbiome via 16S tag sequencing did not provide evidence for a central role of microbiome variation in determining bleaching response. Instead, epigenetic changes were significantly correlated with the host’s genetic background, the position of the sampled polyps within the colonies (e.g. tip versus base of colony), and differences in the colonies’ condition during the bleaching event. We conclude that microenvironmental differences in growing conditions led to long-term changes in the way the ramets methylated their genomes and thus to a differential bleaching response.

overall_design: 16S tag sequencing of Acropora palmata from Florida with differential bleaching responses were analyzed.

first: Meghann

last: Devlin-Durante

first: Dusty

last: Kemp

first: Dana

last: Williams


title: RNAseq analysis of blood from sepsis patients and healthy controls

city: Münster

first: Klaus

last: Schughart

address: Von-Esmarch-Str 56

department: Virology

institute: University Münster

summary:
Gene expression changes in the blood was studied by RNAseq
Results: Strong differences between patients groups, specific expression changes for heathy control (Hlty), uncomplicated infection (Inf1_P), sepsis (Seps_P), septic shock (Shock_P), follow-up of sepsis (Seps_FU), follow-up of septic shock (Shock_FU) groups.

overall_design: Peripheral blood was collected from patients, RNA was isolated and sequenced.


title: The kinetic landscape of an RNA binding protein in cells

status: Public on Dec 16 2020

city: Cleveland

first: Deepak

last: Sharma

state: Ohio

address: 10900, Euclid Avenue

department: Center for RNA Science and Therapeutics

institute: Case Western Reserve University

summary: The binding and dissociation of RNA Binding Proteins (RBPs) to their cognate sites on cellular RNAs are critical for the regulation of gene expression. Yet, association and dissociation kinetics of RBPs at individual cellular binding sites have not been experimentally accessible. Inaccessibility of binding and dissociation kinetics of RBPs in cells severely limits or even precludes the establishment of quantitative connections between RBP-RNA interactions and cellular RBP function. Here, we describe an approach to measure binding and dissociation kinetics of the RBP Dazl at thousands of individual binding sites in cells. We then show how these kinetic parameters inform a quantitative understanding of the cellular function of Dazl.

overall_design: 16 iCLIP libraries corresponding to 4 time points, 2 in vivo Dazl concentrations and 2 femtosecond, pulsed UV laser powers. Monoclonal cell line with doxycyline inducible Dazl expression (GC-1-spg parental line) was utilized.

first: D

first: L

last: Zagore

first: M

last: Brister

first: X

last: Ye

first: C

last: Crespo-Hernández

middle: D

last: Licatalosi

first: E

last: Jankowsky


title: Brain-infiltrating CD8+ T cell activity after acute infection and during CNS autoimmunity

city: Geneva

first: Sylvain

last: LEMEILLE

address: 1 rue Michel-Servet

department: Department of Pathology and Immunology

institute: University of Geneva Medical School

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

overall_design: Refer to individual Series


title: A Tumor Suppressor Enhancer of PTEN in T-cell development and leukemia

city: Bethesda

first: Pedro

last: Rocha

state: Maryland

address: 6 Center Drive Building 6B 2B220

department: NICHD

institute: National Institutes of Health

summary: Long-range oncogenic enhancers play an important role in cancer. Yet, it remains unclear whether similar regulation of tumor suppressor genes is relevant. In this context, loss of expression of the tumor suppressor PTEN is associated with the pathogenesis of various cancers, including T-cell acute lymphoblastic leukemia (T-ALL). Here, we demonstrate that PTEN is regulated in T-cells by a distal enhancer (PE), which is targeted by recurrent focal deletions in T-ALL. This highly conserved regulatory element interacts with and transactivates the PTEN promoter. Consistently, PE deletion in T-ALL cells leads to reduced PTEN levels and proliferative advantage. Moreover, PE-null mice show reduced PTEN levels in the thymus and develop NOTCH1-induced T-ALL with faster kinetics. Furthermore, secondary loss of PE in established leukemias leads to accelerated progression and a gene expression signature driven by Pten loss. Altogether, our results identify PE as the first long-range tumor suppressor enhancer directly implicated in cancer.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: This 4C-seq analysis of the PTEN locus uses human (DND41, HPBALL and JURKAT) and mouse (N1dE_PKM2 and HDdPEST) cells. In each case 4C-seq was done with a bait on the promoter and putative enhancer loci.


title: Proteinase-mediated macrophage signalling in psoriatic arthritis

status: Public on Dec 15 2020

city: Toronto

first: Vinod

last: Chandran

state: Ontario

address: 399 Bathurst Street

department: Krembil Research Institute

country: Canada

institute: University Health Network

summary: To identify the cell populations in synovial fluid (SF) from psoriatic arthritis (PsA) patients, single cell 3’-RNA-sequencing of cells from PsA SF samples (n=3) was performed using the 10X Genomics platform. Twelve main clusters of cells were predicted and the most abundant clusters had a predominance of monocytes/macrophages gene expression patterns.

overall_design: Single cell RNA-sequencing of synovial fluid samples from psoriatic arthritis patients

first: Fatima

last: Abji

first: Mozhgan

last: Rasti

first: Alejandro

last: Gómez-Aristizábal

first: Carla

last: Muytjen

first: Mahmoud

last: Saifeddine

first: Koichiro

last: Mihara

first: Majid

last: Motahhari

first: Rajiv

last: Gandhi

first: Sowmya

last: Viswanathan

first: Morley

last: Hollenberg

first: Katerina

last: Oikonomopoulou


title: Identification of aberrant circRNAs in peripheral whole blood of patients with Alzheimer's disease

city: Shenyang

first: Ke

last: Zhang

address: No.77 Puhe Road

institute: China Medical University

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

summary: We try to identify the aberrant circRNAs in peripheral whole blood of patients with Alzheimer's disease by circular RNA microarray.

overall_design: Total RNA from fresh blood plasma supernatant was isolated using TRizol reagent. The purified labeled cRNAs were performed the microarray analysis according to the instructors.

first: Li

last: Liu


title: Brain-infiltrating CD8+ T cell chromatin accessibility after acute infection and during CNS autoimmunity [ATAC-Seq]

summary: CD8+ T cells play a pivotal role in eliminating pathogens or tumors, but they equally exert tissue damage in autoimmune or chronic inflammatory diseases. While the mechanisms regulating the differentiation of CD8+ T cells in chronic infections and cancer are just beginning to be unraveled, we still lack knowledge about how this unfolds in autoimmune diseases. Here, we investigated in a model of chronic central nervous system autoimmunity, the transcriptional and epigenetic landscape of autoreactive brain infiltrating CD8+ T cells and how these cells diverge from classical long-lived memory CD8+ T emerging after virus infection.

overall_design: Chromatin accessibility of brain-infiltrating WT and Tox -/- P14 CD8+ T cells adoptively transfered in WT and MOG-GP recipient mice following intracranial infection with LCMV-GP.

first: Page

last: Nicolas

first: Merkler

last: Doron


title: Genomic RNA elements drive phase separation of the SARS-CoV-2 nucleocapsid

city: Chapel Hill

first: Mark

middle: A

last: Boerneke

state: NC

address: Campus Box 3290

department: Chemistry

institute: University of North Carolina

summary: We report that the SARS-CoV-2 nucleocapsid protein (N-protein) undergoes liquid-liquid phase separation (LLPS) with viral RNA. N-protein condenses with specific RNA genomic elements under physiological buffer conditions and condensation is enhanced at human body temperatures (33°C and 37°C) and reduced at room temperature (22°C). RNA sequence and structure in specific genomic regions regulate N-protein condensation while other genomic regions promote condensate dissolution, potentially preventing aggregation of the large genome. At low concentrations, N-protein preferentially crosslinks to specific regions characterized by single-stranded RNA flanked by structured elements and these features specify the location, number, and strength of N-protein binding sites (valency). Liquid-like N-protein condensates form in mammalian cells in a concentration-dependent manner and can be altered by small molecules. Condensation of N-protein is RNA sequence and structure specific, sensitive to human body temperature, and manipulatable with small molecules, and presents a screenable process for identifying antiviral compounds effective against SARS-CoV-2.

sample_organism: Severe acute respiratory syndrome coronavirus 2

overall_design: In vitro transcribed RNA was treated with 5NIA or DMSO (control) for SHAPE-MaP RNA structure probing experiments.  The binding of recombinantly expressed and purified SARS-CoV-2 N-protein at 160x, 80x, 20x, or 0x (control) molar excess concentrations to in vitro transcribed RNA was chemically probed using the bivalent chemical crosslinker SDA for RNP-MaP experiments. Chemically modified RNA samples from SHAPE-MaP and RNP-MaP experiments were subjected to MaP reverse transcription, which encodes the position of chemical adducts as non-templated nucleotides or deletions in the product cDNA. Sequencing libraries were generated using the Nextera XT workflow and sequenced on an Illumin MiSeq instrument. The ShapeMapper 2 software was used to align reads and call modification-induced mutations. Software analysis pipelines were employed to create SHAPE and RNP-MaP profiles, find in-cell reactivity perturbations, locate correlated mutations, and model RNA structures.

first: Kevin

last: Weeks


title: Global RNA stability profiling during selective chemical inhibition of human Dcp2

city: West Haven

first: Sarah

last: Slavoff

state: CT

address: 300 Heffernan Dr

institute: Yale University

summary: Multiple RNA decapping enzymes coexist in mammalian cells to regulate decay of partially overlapping sets of cellular transcripts, but a comprehensive understanding of cellular substrate selectivity of each enzyme is yet to be achieved. Previously we demonstrate the utility of TimeLapse-seq in global profiling of RNA stability changes in human Dcp2 knockout cells. However, secondary transcriptional changes and upregulation of alternative decay pathways have obscured complete mapping of Dcp2 substrates. Here, we present the discovery and first application of a cell-permeable, highly selective Dcp2 ligand in the chemical genetic study of its RNA substrates.

overall_design: RNA isolated from cells pre-treated with respective compounds +/- 2 h s4U feed was subjected to oxidative-nucleophilic-aromatic-substitution chemistry and sequenced. All sequencing for experimental samples was performed in duplicate per condition assessed, while negative controls were performed as single replicates without s4U treatment.

first: Jeremy

last: Schofield

first: Yang

last: Luo

first: Matthew

last: Simon


title: Optimization of CAR T cell signaling by incorporation of null mutations

city: TAMPA

first: Marco

last: Davila

state: Florida

address: 12902 USF MAGNOLIA DRIVE, MRC2

institute: Moffitt Cancer Center

summary: An obstacle with continued clinical development of CAR T cells is the limited understanding of CAR T cell biology and its mechanisms of anti-tumor immunity. We and others have shown that CARs with a CD28 co-stimulatory domain drive high levels of T cell activation that also lead to exhaustion and shortened persistence. This led us to hypothesize that by incorporating null mutations of CD28 subdomains (YMNM, PRRP, or PYAP) we could optimize CAR T cell signaling and reduce exhaustion. In vivo we found that mice given CAR T cells with a PYAP CD28 endodomain had a significant survival advantage. We also observed that mutant CAR T cells had significantly less expression of PD1 and were increased in the bone marrow and spleen. In addition, these mutant CAR T cells had optimized signaling resulting in a reduction of exhaustion related transcription factors and genes.  Our results demonstrate that CAR T cells with a mutant CD28 endodomain have better survival and decreased exhaustion and is the result of decreased CAR dependent NFAT/NR4A1 transcription factors. This work allows for development of enhanced CAR T cell therapies by optimizing CAR T cell signaling.

overall_design: For both ATAC-seq and RNA-seq m1928z, m19hBBz, and mut06 CAR T cells were produced from C57BL6 mice.  CAR T cells were simulated with 3T3-mCD19 target cells at a 10:1 E:T ratio for 24 hours.  After stimulation CAR positive cells were sorted using a BD ARIA FACS sorter.  Samples were pelleted and snap frozen in liquid nitrogen.  All samples were done in duplicate.

first: Justin

middle: C

last: Boucher

middle: L


title: DNA Damage Responses in Mouse Primordial Germ Cells [RNA-Seq]

city: Boston

first: Victor

last: Navarro

address: 221 Longwood Ave

department: Division of Endocrinology

institute: Brigham and Women's Hospital

summary: Germ cells specified during fetal development form the foundation of the mammalian germline. These primordial germ cells (PGCs) undergo rapid proliferation, yet the germline is highly refractory to mutation accumulation compared to somatic cells. Importantly, while the presence of endogenous or exogenous DNA damage has the potential to impact PGCs, there is little known about how these cells respond to stressors. To better understand the DNA damage response (DDR) in these cells, we exposed pregnant mice to ionizing radiation (IR) at specific gestational time points and assessed the DDR in PGCs. Our results show that PGCs prior to sex determination lack a G1 cell cycle checkpoint. Additionally, the response to IR-induced DNA damage differs between female and male PGCs post-sex determination. IR of female PGCs caused uncoupling of germ cell differentiation and meiotic initiation, while male PGCs exhibited repression of piRNA metabolism and transposon de-repression.

overall_design: 17 samples of purified mouse primordial germ cells were collected for RNA-sequencing. There were two conditions (either with or without ionizing radiation).

first: John

last: Schimenti

first: Jordana

last: Bloom


title: DNA Damage Responses in Mouse Primordial Germ Cells

type: Non-coding RNA profiling by high throughput sequencing


title: Competition between PRC2.1 and 2.2 subcomplexes regulates PRC2 chromatin occupancy in human stem cells

city: Boulder

first: Thomas

last: Cech

state: CO

address: 3415 Colorado Ave., Jennie Smoly Caruthers Biotech BLDG

institute: University of Colorado Boulder

summary: Polycomb repressive complex 2 (PRC2) silences expression of developmental transcription factors in pluripotent stem cells by methylating lysine 27 on histone H3. Two mutually exclusive subcomplexes, PRC2.1 and PRC2.2, are defined by the set of accessory proteins bound to the core PRC2 subunits. Here we introduce separation-of-function mutations into the SUZ12 subunit of PRC2 to drive it into a PRC2.1 or 2.2 subcomplex in human induced pluripotent stem cells (iPSCs). We find that PRC2.2 occupies polycomb target genes at low levels and that homeobox transcription factors are upregulated when this complex is exclusively present. In contrast with previous studies, we find that chromatin occupancy of PRC2 increases drastically when it is forced to form PRC2.1. Additionally, several cancer-associated mutations also coerce formation of PRC2.1. We suggest that PRC2 chromatin occupancy can be altered in the context of disease or development by tuning the ratio of PRC2.1 to PRC2.2.

overall_design: ChIP-seq was performed on polyclonal cell lines expressing either WT or mutant SUZ12. RNA-seq was performed on clonal SUZ12(WT) and SUZ12(PEmut) cell lines.

first: Daniel

middle: T

last: Youmans


title: Circadian control of hepatitis B virus replication

city: Oxford

first: Xiaodong

last: Zhuang

state: Oxfordshire

address: Old Road Campus, Roosevelt Drive

institute: University of Oxford

summary: Fah-/-, Rag2-/-, IL2Rg-/-, non-obese diabetic (FRG-NOD) mice transplanted with primary human hepatocytes were infected with 10^9 HBV genome equivalents purified from concentrated supernatant of the HepG2.2.15 cell line (HBV genotype D subtype ayw). Eight weeks later mice were treated twice a day by intraperitoneal injection of either vehicle or 100 mg/kg SR9009 for 2 weeks. At the end of experiment, mouse livers were collected for RNA-seq analysis. The sequence reads were mapped to the human or mouse genome.

overall_design: Liver mRNA profiles of control or SR9009 treated mice.

first: Jane

last: McKeating

first: Robert

last: Maidstone


title: Dkstatin-treated Pseudomonas aeruginosa trascriptome

city: seoul

first: Kyung Bae

last: Min

address: yonsei-ro 50-1

department: college of medicine

institute: Yonsei University

summary: To better understand the effects of Dkstatin treatment, we compared transcriptome landscapes of PAO1 grown without or with either Dkstatin. A global view of the entire transcriptome uncovered the substantial differences between cells treated with Dkstatin-1 or Dkstatin-2.

overall_design: To extract total RNA, P. aeruginosa PAO1 was incubated for 2 h in 5 ml of LB with vigorous shaking at 37°C. Then, additional incubation was conducted for 1 h under the same condition with 150 𝜇M of Dkstatin-1 or Dkstatin-2 supplement. Total RNA samples were extracted using an RNeasy mini kit (Qiagen), and total DNA was eliminated using RNase-free DNase set (Qiagen) following the manufacturer’s instructions. The extraction was performed as an experimental duplicate. Transcriptome profile comparison and processing the expression data were provided by Ebiogen (Korea). RNA quality was assessed by an Agilent 2100 bioanalyzer using the RNA 6000 Nano Chip (Agilent Technologies, Amstelveen, The Netherlands), and RNA quantification was performed using an ND-2000 Spectrophotometer (Thermo Inc., DE, USA). For control and testing RNAs, rRNA was removed using a Ribo-Zero Magnetic kit (Epicentre Inc., USA) from each 5 ug of total RNA. The construction of a library was performed using a SMARTer Stranded RNA-Seq Kit (Clontech Lab Inc., CA, USA) according to the manufacturer’s instructions. T

first: Kyung

first: Sang

middle: S

last: Yoon


title: Alterations in the transcriptome and translatome profiles of Arabidopsis pollen germinated under heat stress

city: Valencia

first: Javier

last: Forment

state: Valencia

address: Ingeniero Fausto Elio, s/n

department: Biotecnología

country: Spain

institute: Instituto de Biología Molecular y Celular de Plantas, IBMCP (UPV-CSIC)

summary: Plant reproduction is one key biological process very sensitive to heat stress and, as a consequence, enhanced global warming imposes serious threats to sustain food safety worldwide. In this work we have focused on the molecular impact that high temperature conditions impose on gene expression of Arabidopsis pollen germinated in vitro. We have used a high-resolution ribosome profiling technology to provide, for the first time, a comprehensive study of how both the transcriptome and the translatome of germinated pollen respond to the increase in temperature. Although heat shock responses operate properly under high temperature conditions, we have uncovered important alterations under elevated temperature regimes down-regulating essential processes linked to cation/proton exchange and to carbohydrate/cation symport transport. These alterations provide molecular explanations to the dramatic alterations of pollen tube growth under heat stress. Overall a high correlation between transcriptional and translational responses to high temperature was found, but specific regulations at the translational level are also present in pollen subjected to temperature challenging conditions.

overall_design: Riboprofiling analysis was performed for Arabidopsis thaliana in vitro germinated pollen at permissive and restrictive temperatures (24°C 35°C respectively).

first: Laetitia

last: Poidevin

first: Dilek

last: Unal

first: Alejandro

last: Ferrando


title: Transcriptional Profiling of Adult Male Rat Liver and Testis following 14 Day Myclobutanil Exposure

city: Indianapolis

first: Kamin

last: Johnson

state: IN

address: 9330 Zionsville Rd

institute: Corteva Agriscience

summary: We report the RNAseq-based transcriptome profiles of adult male rat liver and  testis following 14 day exposure to myclobutanil.  A treatment-related change in mRNA levels were observed in both liver and testis.

overall_design: Adult male rat liver and testis RNAseq profiles following 14 day exposure to myclobutanil

first: Jessica

last: LaRocca

first: Shreedharan

last: Sriram


title: RNA sequencing of naïve, Th1 and Th2 polarised live CD4+ T cells from CD4Cre+ and CD4Cre+DOT1LFL/FL mice

city: Clayton

first: Sebastian

last: Scheer

address: 15 Innovation Walk

institute: Monash University

overall_design: CD4+ T cells were isolated from CD4Cre+ or CD4Cre+DOT1LFL/FL mice and subsequently cultured for 4 days under Th1 or Th2 polarising conditions and sorted for live CD4+ T cells; naïve samples represent the starting population

first: Kirill

last: Tsyganov


title: RNA-seq of B cells treated with recombinant CD52-Fc

status: Public on Dec 14 2020

city: Stanford

first: Kartik

last: Bhamidipati

state: California

address: 269 Campus Drive, Rm 4135

institute: Stanford University School of Medicine

summary: CD52 is a 12 amino acid glycoprotein that is widely expressed on lymphocytes. In T cells, the soluble form of CD52 has been shown to inhibit TCR function by binding Siglec-10. In Monocytes, soluble CD52 has been shown to inhibit TLR signaling and induces apoptosis at high concentrations. To assess the impact of CD52 on B cell function, we left B cells either untreated or exposed to recombinant CD52-Fc for 6 hours prior to lysis and bulk RNA-sequencing. Gene expression analysis revealed that soluble CD52 induced downregulation of immunoglobulin and B cell receptor related genes such as CD19 and CD22. Therefore, soluble CD52 appears to have a suppressive role on B cell function.

overall_design: B cells untreated (n = 6) or CD52-Fc treated (n = 6) for 6 hours at 37C.

first: Bhamidipati

last: Kartik


title: Single-cell RNA sequencing of B cells from healthy donors and SLE patients

summary: Systemic Lupus Erythematosus (SLE) is an autoimmune disease characterized by pathogenic auto-antibodies that cause end organ damage. B cells are thought to play a central role in the immunopathogenesis of SLE and display several abnormalities in patients, including a strong type I interferon signature as well as lower expression of surface markers including TLR9 and FcγRIIB. To characterize differences in the proportion of distinct B cell subsets as well as intrinsic transcriptomic differences between B cells from healthy donors and SLE patients, we performed single-cell RNA sequencing on B cells isolated from PBMCs of donors. Several notable features were observed, including a strong interferon response signature among SLE B cells, and the expansion of CD11c+T-bet+ B cell subsets in these patients. Additionally, several surface molecules including MHC Class II proteins and surface proteins such as CD74 and CD52 were found to be differentially expressed in B cells isolated from SLE patients. The differences in B cell subset proportion as well as expression of various genes might play a role in B cell mediated pathogenesis of SLE.

overall_design: Whole transcriptome data for HC (n = 2) and SLE (n = 3).


title: RNA-Seq of pre-chemo high-grade serous ovarian tumors

first: Zhiyuan

last: Hu

address: John Radcliffe Hospital/Headley Way

department: MRC Weatherall Institute of Molecular Medicine

summary: We performed RNA sequencing on 10 bulk pre-chemo tumor samples collected from 5 patients with HGSOC.

overall_design: Ten bulk pre-chemo tumor samples collected from 5 patients with HGSOC.

first: Mara

last: Artibani

first: Salma

last: El-Sahhar

first: Garry

last: Mallett

first: Christopher

last: Yau

first: Ahmed

last: Ahmed


title: The role of DOT1L in Th cells

summary: ChIP sequencing of naïve Th, bona fide Th1 and bona fide Th2 cells

overall_design: Cells were sorted from the spleens of WT mice for naïve Th cells. Bona fide Th1 or Th2 were generated by culturing enriched Th cells for 4 days under Th1 or Th2 polarising conditions. IL-4 Amcyan - IL-13 dsRed - IFNg-YFP reporter mice were used to sort bona fide Th1 (YFP+) or bona fide Th2 (Amcyan or dsRed+) cells.

first: Adele

last: Burghare


title: Global RNA stability profiling in human NBDY knockout using TimeLapse-seq

first: Sarah

last: Slavoff

summary: We previously identified a conserved human microprotein NoBody/NBDY that can interact with the mRNA decapping complex through EDC4 and affects the expression of an NMD (nonsense-mediated decay) substrate. However, it remains unclear the exact mechanism of NBDY-regulated RNA stability change and whether this is the direct cause/consequence of its modulation of P-body structure/numbers. Here we present a global profile of RNA stability in human NBDY knockout cells via TimeLapse-seq. By comparing this dataset to our previous work done in DCP2 KO cells, we demonstrate that NBDY is a specificity factor of the 5'-3' RNA decay. In addition, these data indicate a role of NBDY in cell cycle and other signaling transduction processes as well.

overall_design: RNA isolated from cells +/- s4U feed was subjected to oxidative-nucleophilic-aromatic-substitution chemistry and sequenced. All sequencing for experimental samples was performed in duplicate per condition assessed, while negative controls were performed as single replicates without s4U treatment.

first: Zhenkun

first: Jeremy

last: Schofield

first: Matthew

last: Simon


title: RNAseq analysis of microglia isolated from treated brain slices or treated mice

city: Cambridge

first: Alex

last: Delbridge

state: MASSACHUSETTS

address: 225 binney street

institute: Biogen

summary: Population based transcriptomic profiling was performed on microglia isolated from TNF or GMCSF treated brain slices or from LPS treated mice

overall_design: For organotypic brain slice microglia samples, brain slices were cultured for 2 weeks and then treated for 24 hours with TNF (50 ng/mL), or GMCSF (20 ng/mL). Following treatment microglia were isolated for analysis, to gain sufficient RNA material for analysis each sample consists of 3 pooled brain slices. For microglia derived from LPS treated mice, mice were treated with LPS (1 mg/kg) injected intraperitoneally 24 hours prior to microglia isolation. In the case of brain tissue analysis samples were flash frozen in liquid nitrogen at the time of collection.

first: Thomas

last: Carlile

first: Michael

last: Mingueneau


title: Organotypic brain slice culture microglia exhibit molecular similarity to acutely-isolated adult microglia and provide a platform to study neuroinflammation

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

overall_design: Refer to individual Series


title: NanoString nCounter with the Oxford Classifier of Carcinoma of the Ovary

summary: Using RNA-seq, we recently developed the 52-gene-based Oxford Classifier of Carcinoma of the Ovary (Oxford Classic, OxC) for molecular stratification of serous ovarian cancers (SOCs) based on the molecular profiles of their cell-of-origin in the fallopian tube epithelium. Here, we developed a 52-gene NanoString panel for the OxC to test the robustness of the classifier. We measured the expression of the 52 genes in an independent cohort of prospectively collected SOC samples (n = 150) from a homogenous cohort who were treated maximal debulking surgery and chemotherapy. We performed data mining of published expression profiles of SOCs and validated the classifier results on tissue arrays comprising 137 SOCs.  We found evidence of profound non-genetic heterogeneity in SOC. ~20% of SOCs were classified as epithelial-mesenchymal-transition-high (EMT-high) tumors, that were associated with poor survival. This was independent of established prognostic factors such as tumor stage, tumor grade and residual disease after surgery (HR = 3.3, p = 0.02). Mining expression data of 593 patients revealed a significant association between the EMT scores of tumors and the estimated fraction of alternatively activated macrophages (M2) (p < 0.0001) suggesting a mechanistic link between immunosuppression and poor prognosis in EMT-high tumors. The OxC-defined EMT-high serous ovarian cancers carry particularly poor prognosis independent of established clinical parameters. These tumors are associated with high frequency of immunosuppressive macrophages suggesting a potential therapeutic target to improve clinical outcome.

overall_design: NanoString expression data of a 52-gene panel (Oxford Classifier of Carcinoma of the Ovary, OxC) in 150 serous ovarian cancer samples (HH-cohort) and ten HGSOC samples.

first: Paula

last: Cunnea

first: Zhe

last: Zhong

first: Christina

last: Fotopoulou


title: Modulation of Aub–Tdrd interactions elucidates piRNA amplification and germ plasm formation

city: Baltimore

first: Manolis

last: Maragkakis

state: MD

address: 251 Bayview Blvd

institute: National Institute on Aging

summary: Aub protein guided by piRNAs ensures genome integrity by cleaving retrotransposons and genome propagation by trapping mRNAs to form the germ plasm that instructs germ cell formation. The amino terminus of Aub (Aub-NT) is rich in arginines (Aub-NTRs), which are symmetrically dimethylated (sDMAs), and interacts with Tudor protein and other Tudor domain containing proteins (Tdrds). Aub-Tdrd interactions play critical roles in suppressing active retrotransposons via piRNA amplification and in germ plasm formation via generation of Aub-Tudor ribonucleoproteins.  Here we show that Aub-NTRs are dispensable for primary piRNA biogenesis but essential for piRNA amplification and transposon control and that sDMAs in Aub-NT are essential for germ plasm formation and germ cell specification, but largely dispensable for piRNA amplification and transposon control.

overall_design: We examine the importance of Aub N-terminal arginine di-methylation in piRNA pathway and germ plasm formation in Drosophila ovaries

first: Nicholas

last: Vrettos

first: Panagiotis

last: Alexiou

first: Daniel

last: Palmieri

first: Paraskevi

last: Sgourdou

first: Fadia

last: Ibrahim

first: Yohei

last: Kirino

first: Zissimos

last: Mourelatos


title: Ethanol exposure drives colon location-specific cell composition changes in a normal colon crypt 3D organoid model

city: Charlottesville

last: Devall

state: Virginia

address: 1335 Lee Street

institute: University of Virginia

summary: Here, we used bulk RNA-seq data derived from healthy colon organoids (un)exposed to ethanol. Through the use of external single cell RNA-seq data, we estimate changes in cell composition that are more pronounced in distal colon organoids following ethanol treatment. We extend this analysis by inferring cell-type-specific effects of ethanol treatment on colon organoids.

overall_design: Analysis of the effects of ethanol treatment on gene expression in colon organoids

first: Sarah

last: Plummer

first: Jennifer

last: Bryant

first: Lucas

last: Jennelle

first: Stephen

last: Eaton

first: Christopher

middle: H

last: Dampier

first: Jeroen

last: Huyghe

first: Ulrike

last: Peters

last: Powell

first: Graham

last: Casey


title: Single cell RNAseq analysis of organotypic brain slice microglia

first: Alex

last: Delbridge

summary: Single cell RNA sequencing analysis was performed on microglia isolated from brain slices  (timepoints: 1 day, 1 week or 3 weeks in culture), 2D in vitro cultured microglia, and acutely isolated adult microglia. Differences in gene expression were examined to determine relative similarity to acutely isolated adult microglia.

overall_design: Microglia were isolated from cultured brain slices at 3 different timepoints, from 2D invitro mixed glial cultures, and adult mouse brain and subjected to scRNAseq analysis using the 10x Chromium system; n = 2 biological replicates per group

middle: R

first: Thomas

last: Carlile

first: Michael

last: Mingueneau


title: Regulation of anti-tumorigenic pathways by the combinatory treatment of calcitriol and TGF-β in PC-3 cells

city: Mexico

first: Sandra

middle: Lorena

last: Romero-Cordoba

state: Mexico

address: Periférico Sur No. 4809, Col. Arenal Tepepan, Delegación Tlalpan. México, D.F.

department: Cancer Genomics

country: Mexico

institute: National Instute of Genomic Medicine

summary: Calcitriol and transforming growth factors beta (TGF-β) are involved in several biological pathways such as cell proliferation, differentiation, migration and invasion. Their cellular effects could be similar or opposite depending on the genetic target, cell type and context. Despite the reported association of calcitriol deficiency and disruption of the TGF-β pathway in prostate cancer and the well-known independent effects of calcitriol and TGF-βs on cancer cells, there is limited information regarding the cellular effects of calcitriol and TGF-β in combination.  In this study, we in vitro analyze the combinatory effects of calcitriol and TGF-β on cell growth and apoptosis using PC-3 and DU145 human prostate cancer cell lines. Using high-throughput microarray profiling of PC-3 cells upon independent and combinatory treatments, we identified distinct transcriptional landscapes of each intervention, with a higher effect established by the combinatorial treatment, following by TGF-β1 and later by calcitriol. A set of genes and enriched pathways converge among the treatments, mainly between the combinatory scheme and TGF-β1, but the majority were treatment-specific. Of note, CYP24A1, IGFBP3, SERPINE1, CDKN1A, NOX4 and UBE2D3 were significantly up-regulated upon the combinatorial treatment whereas CCNA1, members of the CT45A and APOBEC3 family were down-regulated. By public RNA signatures, we were able to confirm the regulation by the co-treatment over cell proliferation and cell cycle. We finally investigated the possible clinical impact of genes modulated by the combinatorial treatment using benchmark prostate cancer data. This comprehensive analysis reveals that the combinatory treatment impairs cell growth without affecting apoptosis and their combinatory actions might synergize and improved their individual effects to reprogram prostate cancer signaling.

overall_design: PC-3 human prostate cancer cells were treated during 24hrs with calcitriol (10 nM) with TGF-β1  (5 ng/mL), alone or in combination. Gene expression profiles were evaluates on Clariom D (Affymetrix) microarrays.

first: Mitzi

last: García-Olivares

last: Romero-Córdoba

first: Elizabeth

last: Ortíz-Sánchez

first: Rocío

last: García-Becerra

first: Mariana

last: Segovia-Mendoza

first: Claudia

last: Rangel-Escareño

first: Ali

last: Halhali

first: Fernando

last: Larrea

first: David

last: Barrera


title: DNA Damage Responses in Mouse Primordial Germ Cells [ncRNA-Seq]

first: Victor

last: Navarro

state: MA

summary: Germ cells specified during fetal development form the foundation of the mammalian germline. These primordial germ cells (PGCs) undergo rapid proliferation, yet the germline is highly refractory to mutation accumulation compared to somatic cells. Importantly, while the presence of endogenous or exogenous DNA damage has the potential to impact PGCs, there is little known about how these cells respond to stressors. To better understand the DNA damage response (DDR) in these cells, we exposed pregnant mice to ionizing radiation (IR) at specific gestational time points and assessed the DDR in PGCs. Our results show that PGCs prior to sex determination lack a G1 cell cycle checkpoint. Additionally, the response to IR-induced DNA damage differs between female and male PGCs post-sex determination. IR of female PGCs caused uncoupling of germ cell differentiation and meiotic initiation, while male PGCs exhibited repression of piRNA metabolism and transposon de-repression.

overall_design: 6 samples of purified E13.5 male primordial germ cells were collected for small RNA-sequencing. There were two conditions (either with or without ionizing radiation). Each of the conditions has 3 replicates

first: John

last: Schimenti

first: Jordana

last: Bloom


title: HMGA2 links promoter-specific H2A.X deposition and active DNA demethylation [ChIP-Seq]

city: Nancy Cedex

first: Guillermo

last: Barreto

address: 9, Avenue de la Forêt de Haye ‐ BP 20199

department: Campus Biologie-Santé. Faculté de Médecine

institute: CNRS Director of Research

summary: During transcriptional activation, chromatin undergoes structural changes with the help of various proteins such as chromatin remodeling factors and histone modifiers in order to make the DNA accessible for the transcription factors and RNA polymerase. We combined ChIP-, MNase- and RNA-sequencing analyzing the Hmga2 knock-out mice and found that HMGA2 is required for promoter specific deposition of the histone variant H2A.X. Consequently, the promoter specific H2A.X deposition resulted in an accumulation of active RNA Polymerase II at these promoters. Stimulation with TGFB1 resulted in increased accumulation of pH2A.X and expression of those “primed” target genes, whereas cells expressing no Hmga2 where not able to respond.

overall_design: ChIP-Seq Data from chromatin extracts after ultracentrifugation from Hmga2+/+ and Hmga2-/- mouse embryonic fibroblasts

first: Stephanie

last: Dobersch

first: Indrabahadur

last: Singh


title: HMGA2 links promoter-specific H2A.X deposition and active DNA demethylation [Mnase-Seq]

overall_design: Mnase-Seq Data from chromatin extracts after ultracentrifugation from Hmga2+/+ and Hmga2-/- mouse embryonic fibroblasts

first: Stephan

last: Günther


title: HMGA2 links promoter-specific H2A.X deposition and active DNA demethylation


title: Day23_exon3lines_Rep2

city: San Francisco

first: Benoit

last: Bruneau

state: CA

address: 1650 Owens St

institute: Gladstone Institutes

summary: Haploinsufficiency of transcriptional regulators causes human congenital heart disease (CHD), predicting gene regulatory network (GRN) imbalances. Here, we define transcriptional responses to reduced transcription factor dosage in human iPSCs heterozygous or homozygous for loss of the CHD gene TBX5.

overall_design:
Single cell RNA sequencing of day 23 cells from exon3 lines (WTC11, TBX5Het and TBX5Hom) - biological replicate 2
Processed data are on the GSE137873 Series record.

first: Irfan

last: Kathiriya

first: Kavitha

last: Rao