title: ELAV/Hu RNA binding proteins determine multiple programs of neural alternative splicing

first: Seungjae

last: Lee

state: NY

address: 430 East 67th St.

department: Department of Developmental Biology

institute: Sloan-Kettering Institute

summary: ELAV/Hu factors are conserved RNA binding proteins that play diverse roles in mRNA processing and regulation. The founding member, Drosophila Elav, was recognized as a vital neural factor 35 years ago. Nevertheless, still little is known about its impacts on the transcriptome, and potential functional overlap with its paralogs. Building on our recent findings that neural-specific lengthened 3' UTR isoforms are co-determined by ELAV/Hu factors, we address their impacts on splicing. While only a few splicing targets of Drosophila are known, we find that ectopic expression of the three family members (Elav, Fne and Rbp9) induces overlapping changes to hundreds of cassette exon and dozens of alternative last exon (ALE) splicing events. Reciprocally, double mutants of elav/fne, but not elav alone, have opposite effects on both classes of regulated mRNA processing events in the larval CNS. While manipulation of Drosophila ELAV/Hu factors induces both exon skipping and inclusion, motif analysis indicates their major direct effects are to suppress cassette exon usage. Moreover, the roles of ELAV/HU factors in global promotion of distal ALE splicing are mechanistically linked to terminal 3' UTR extensions in neurons, since both involve local suppression of proximal polyadenylation signals via ELAV/Hu binding sites downstream of cleavage sites. The phenotypic impact of combined ELAV/Hu activities in neural mRNA processing is overt, since fne loss strongly enhances neuronal differentiation phenotypes in elav mutants. Finally, we provide evidence for conservation in mammalian neurons, which undergo broad programs of distal ALE and APA lengthening, linked to ELAV/Hu motifs downstream of regulated polyadenylation sites. Overall, ELAV/Hu proteins orchestrate multiple conserved programs of neuronal mRNA processing by suppressing alternative exons and polyadenylation sites.

overall_design: RNA-seq profiles from S2R+ cells with the overexpression of wildtype Elav, Rbp9, Fne and RRM mutant versions of Elav, Rbp9, Fne and BrdU-CLIP of wildtype Elav and RRM mutant variant.

first: Lu

last: Wei

first: Binglong

first: Raeann

last: Goering

first: Sonali

last: Majumdar

first: Jiayu

last: Wen

last: Taliaferro

first: Eric

last: Lai


title: Transcriptome profiling to identify a human effector regulatory T cell core signature

status: Public on Feb 23 2021

city: Utrecht

first: Lisanne

last: Lutter

address: Lundlaan 6

country: Netherlands

institute: University Medical Center Utrecht

type: Expression profiling by high throughput sequencing

summary: We investigated the transcriptomic landscape of human Treg in an autoimmune-associated inflammatory environment, i.e. synovial fluid (SF) from inflamed joints of Juvenile Idiopathic Arthritis (JIA) to improve our understanding of human effector Treg programming.

overall_design: Examination of the regulatory and CD4 effector T cell transcriptome in 3-4 biological replicaties of each healthy adult and child peripheral blood, juvenile idiopathic arthritis patient peripheral blood in remission and with active disease as well as synovial fluid.

first: Femke

last: van Wijk

first: Jorg

last: van Loosdregt

first: Michal

last: Mokry


status: Public on Feb 24 2021

city: Stanford

first: Julia

last: Belk

state: California

address: 450 Serra Mall

country: USA

institute: Stanford University

summary: We report the results of CRISPR survival screens in the context of multiple viral infections. Two sgRNA pools were used: one targeting the SARS-CoV-2 RNA interactome proteins, and one targeting mitochondrially-related proteins.

overall_design: Multi-virus CRISPR screening data in VeroE6 cells, sgRNA sequencing results.

middle: A

first: Jin

first: Quanming

last: Shi


title: brpf1 knockdown reduces inhibitory neurotransmission and regulates gene expression of GABAergic interneurons derived from MGE

city: Shanghai

first: Jingli

last: Cao

state: Shanghai

address: 131 Dongan Road, Xuhui District, Shanghai

country: China

institute: Fudan university

summary: Our research is to investigate the role of Brpf1 in the development and function of GABAergic interneurons. To test this, we used mouse primary in vitro cultured GABAergic interneurons derived from medial ganglionic eminence (MGE) and utilized AAV-shBrpf1 virus to knockdown Brpf1. We performed mRNA-seq on the total RNA extracted from the infected neurons .We found that most of the up-regulated genes after Brpf1 knockdown were involved in processes such as protein binding, gene negative transcription, ubiquitination, and inflammation by mRNA-seq. We found that many genes related to neuronal calcium, potassium ion transport and neurotransmitter release changed after Brpf1 was knockdown.Our results demonstrate the key role of Brpf1 in  related gene expression of GABAergic interneurons, and provide new lights on the underlying neurobiological mechanisms of BRPF1 mutations that cause mental retardation in children or adolescents.

overall_design: investigate the role of Brpf1 in the development and function of GABAergic interneurons derived from MGE

first: Weiwei

last: Xian

first: Guomin

last: Zhou

first: Linya

last: You


title: Genome-wide distribution of ASH1L and H3K36me2 in human MV4;11 leukemia cell line

city: Ann Arbor

first: Fengbiao

last: Mao

state: Michigan

address: 1301 Catherine St

department: Department of Pathology

institute: University of Michigan

type: Genome binding/occupancy profiling by high throughput sequencing

summary: The H3K36me2 and ASH1L levels are compared in the presence or absence of the inhibitor of the ASH1L activity.

overall_design: Cut & Run profiling of ASH1L and H3K36me2 in MV4;11 cell line treated with ASH1L inhibitor AS-99 or DMSO.

first: Tomasz

last: Cierpicki

first: Jolanta

last: Grembecka


title: The Role of nuclear protein DEK in hematopoietic stem cells

status: Public on Feb 22 2021

city: Chongqing

first: Zhe

last: Chen

address: 1 Yixueyuan Road, Yuzhong District

institute: Chongqing Medical University

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

overall_design: Refer to individual Series


title: Genome-wide Maps of Chromatin State in Control and DEK-deficient Hematopoietic Stem Cells [CUT&Tag]

summary: Purpose:To identify chromatin state involved in the DEK regulating gene expression of hematopoietic stem cells (HSC) in mice, we performed DEK, H3K27ac, H3K4me3 and H3K9ac CUT&Tag in HSC or LSK cells freshly sorted from mice.

overall_design: Methods: About 20,000 HSC (Lin-Sca-1+c-Kit+CD48-CD150+) or 50,000 LSK cells (Lin-Sca-1+c-Kit+) in bone marrow  were isolated by FACS from Dekfl/fl (floxp) and Dekfl/fl,Tie2-Cre (cKO). Genomic DNA  profiles of HSC or LSK cells treated with DEK, H3K27ac,H3K9ac, and H3K4me3 antibody were generated by deep sequencing, using Illumina HiSeq 2000.

first: Yu

last: Hou


title: RNA-seq of C3H/HeJ AA mice skin biopsies treated with JAK-selective inhibitors

city: New York

first: Zhenpeng

last: Dai

address: 1150 St. Nicholas Ave

department: Dermatology

institute: Columbia University

summary: To define the molecular response to JAKi treatment, we performed RNA-seq analysis on a series of skin biopsies taken before and after systemic treatment with INCB039110 (JAK1i), CEP-33779 (JAK2i), PF-06651600 (JAK3i), ruxolitinib (JAK1/2i), tofacitinib (pan-JAKi) or vehicle control.

overall_design: mRNA profiles to JAKi treatment of C3H/HeJ AA mice.

first: James

first: Yuqian

last: Chang


title: ChIRP-MS RNA-seq during SARS-CoV-2 infection

summary: RNA-seq data of RNAs pulled down with the SARS-CoV-2 viral RNA after formaldehyde crosslinking (ChIRP-MS)

sample_organism:
Homo sapiens
Chlorocebus sabaeus

platform_organism:
Homo sapiens
Chlorocebus sabaeus

overall_design: RNA-RNA interactions during SARS-CoV-2 infection

first: Ryan

last: Flynn

first: Yanyan

last: Qi


title: Next Generation Sequencing Facilitates Quantitative Analysis of the Role of nuclear protein DEK in hematopoietic stem cells [RNA-seq]

summary:
Purpose:To identify genes and the molecular pathways involved in the DEK regulating hematopoietic stem cells (HSC) in mice, we performed RNA-Sequence of HSC freshly sorted from mice.
Methods: mRNA  profiles of HSC in Dekfl/fl (floxp) and Dekfl/fl,Tie2-Cre (cKO)  were generated by deep sequencing, in triplicate, using Illumina HiSeq 2000.
Results:Using an optimized data analysis workflow, we mapped about 50 million sequence reads per sample to the mouse genome (build mm10) and identified 33299 transcripts in HSC (the floxp  and  cKO group) with BWA workflow.
Conclusions: Our results represent the first detailed analysis of HSC transcriptomes in Dek-knockout mice and found that DEK regulated multiple mRNA pathways in HSC.

overall_design: The mRNA  profiles of HSC in Control and Dek-knockout mice were generated by deep sequencing, in triplicate, using Illumina HiSeq 2000.


title: Non-transformed cells respond to fat by inducing glucose metabolism

city: Leuven

first: Sandrine

last: Da Cruz

state: Flemish Brabant

address: ON 5 Herestraat 49

department: KU Leuven Department of Neurosciences

country: Belgium

institute: VIB-KU Leuven Center for Brain and Disease Research

summary:
Purpose: To study the effects of high-fat diet feeding in mouse liver tissues with and without the hepatocellular carcinoma-inducing carcinogen DEN. Here, we used RNA sequencing to identify gene expression patterns associated with high-fat diet feeding.
Methods: C57BL/6N mice were injected at 2-weeks of age with vehicle control (PBS) or DEN. At 6 weeks, mice were randomized to control diet, or 60% high-fat diet. After 8-weeks of diet exposure, mice underwent a 13C6-glucose labelling protocol, and liver tissues were extracted for anlaysis (metabolomic, transcriptomic). RNA was isolated from mouse liver tissue using Triazol extraction, purified, and libraries generated using KAPA Stranded mRNA Sequencing kit. After cDNA synthesis, adapter ligation, and final cDNA library generation, samples were sequenced on a flow cell (1x50bp single-end reads) and HiSeq4000 (Illumina). Data processing was conducted in an NGS pipeline (Snakemake) and quality control was performed with FastQC. Trimmed data was analyzed for differential expression (DEseq2) and gene set enrichment analysis (GSEA) to look for KEGG pathways and gene ontologies (GO) of interest.
Results: Using 13C6-glucose labelling, we determined there was a glycolytic phenotype caused by high-fat diet exposure. Therefore, we focused on a diet effect in our transcriptomcis dataset. By comparing control diet to high-fat diet liver tissue, we found gene sets invloved in peroxisomes and lipid metabolism to be enriched in high-fat diet exposed livers. These findings were in line with in vitro testing of a liver cell line, showing peroxisomal metabolism of palmitate drives ROS production and a glycolytic phenotype.
Conclusions: We found high-fat diet exposed liver exhibits a strong metabolic phenotype towards increased glucose metabolism. At the transcriptome level, we found a lipid-reporgramming signature, and not a glucose metabolism signature. The lipid reprogramming signature was in line with in vitro work using liver cell lines, in which exposure to palmitate stimulated a glycolyitic phenotype that was inhibited by targeting peroxisomal-derived ROS species. In combination with metabolic and lipidomic data after long-term exposure to DEN and subsequent tumor formtion, we discovered that high-fat diet can prime liver tissue for carcinogenesis and tumor development by stimulating a Warburg-like metabolism. These findings suggest that fat can induce similar changes in non-transformed liver cells than found in HCC. In conclusion, we show that normal, non-transformed livers respond to fat by inducing glucose metabolism.

overall_design: 2x2 design in C57BL/6N mice on control diet or 60% high fat diet, with or without DEN injection, with liver tissue RNA sequencing conducted

first: Theo

last: Killian

first: Lindsay

last: Broadfield

first: Sarah-Maria

last: Fendt


title: RNA sequencing of T cells in the genetically encoded mouse model of T cell anergy (W131AOTII)

city: San Francisco, Room S-1024

first: Trang

middle: T. T

last: Nguye

address: 513 Parnassus

department: Department of Medicine

institute: University of California, San Francisco

summary: We found that gene signatures characteristic of unresponsive cells were strongly enriched in peripheral W131AOTII CD4 T cells but not in CD4 single positive (CD4SP) thymocytes. Furthermore, loss of Cbl-b in W131AOTII mice significantly altered gene expression profile associated with improved T cell responsiveness.

overall_design: RNA sequencing of T cells isolated from OTII, Cbl-b-/-OTII, W131AOTII and Cbl-b-/-W131AOTII to investigate the possible downstream signaling pathways that cbl-b controls and rescues T cell responsiveness in T cell anergy.

middle: T

last: Nguyen

first: Andrew

last: Schroeder

first: Walter

last: Eckalbar

first: Arthur

last: Weiss


title: Analysis of MV4;11 cells treated with ASH1L inhibitor AS-99

first: Fengbiao

last: Mao


title: A stem cell population at the anorectal junction maintains homeostasis and participates in tissue regeneration after injury

city: Bron

first: Hector

last: Hernandez-Vargas

state: Rhone

address: 36 bis Rue de la Batterie

country: France

institute: Genomics Consulting

summary: Transition zones are highly plastic junction tissues that connects two distinct epithelia such as stratified and glandular. Here, we used single cell RNA sequencing on normal and wounded mouse anorectal epithelial cells, to study their roles during tissue regeneration in normal and injured state.

overall_design: Single cell RNA sequencing has been performed on the anorectal epithelial cells isolated from normal and 1 week post-EDTA wounded K17CreERT2;R26RGFP mice, injected twice with tamoxifen to mark the anal TZ cells.

first: Géraldine

last: Guasch

first: Louciné

last: Mitoyan


title: Exon Level Expression Profiling: a Novel Unbiased Transcriptome (Human)

city: Houston

first: vanaja

last: konduri

state: Texas

address: One Baylor Plaza

department: Pathology&Immunology

institute: Baylor College of Medicine

summary: Transcriptome analysis of RNA samples from magnetically sorted peripheral blood cells

overall_design: Microarray analysis was performed on peripheral blood derived PBMC from six healthy donors sorted into CD8+CD161+ and CD8+CD161- cells.  harvested, pooled and activated with PDAC antigen-loaded autologous DC overnight.

first: V

last: Konduri

first: Y

last: Baig

first: W

middle: K

last: Decker


title: Interleukin-11-expressing fibroblasts have a unique gene signature correlated with poor prognosis of colorectal cancer

city: Ota-ku

first: Takashi

last: Nishina

state: Tokyo

address: 5-21-16 Omori-nishi

department: Biochemistry

country: Japan

institute: Toho University

summary: Interleukin (IL)-11 is a member of the IL-6 family of cytokines and involved in multiple cellular responses, including tumor development. However, the origin and functions of IL-11-producing (IL-11+) cells are not fully understood. To characterize IL-11+ cells in vivo, we generated Il11 reporter mice. IL-11+ cells appeared in the colon of murine tumor and acute colitis models. Il11ra1 or Il11 deletion attenuated the development of colitis-associated colorectal cancer. IL-11+ cells expressed fibroblast markers, and genes associated with cell proliferation and tissue repair. IL-11 induced the activation of colonic fibroblasts and epithelial cells through phosphorylation of STAT3. Human cancer database analysis revealed that a set of genes enriched in IL-11+ fibroblasts was elevated in human colorectal cancer, and correlated with reduced recurrence-free survival. Together, IL-11+ fibroblasts activate both tumor cells and fibroblasts via secretion of IL-11, thereby constituting a feed forward loop between tumor cells and fibroblasts in the tumor microenvironment.

overall_design: To characterize IL-11+ fibroblasts that appeared in colitis, we used a cell sorter to isolate EGFP+ and EGFP- fibroblasts from the colon of DSS-treated Il11-Egfp reporter mice and compared their gene expression profiles by RNA-seq analysis.

first: Hiroyasu

last: Nakano


title: Transcriptional profiling of microglia in the injured brain reveals distinct molecular features underlying neurodegeneration

city: Beijing

first: Cong

last: LIU

address: 北京市朝阳区北辰西路1号院5号

institute: Chinese Academy Science

summary: We applied whole transcriptome analysis of microglia acutely isolated at different timepoints after a cortical stab wound injury to gain insight into genes that might be dysregulated and transcriptionally different between males and females after cortical injury.

overall_design: Total RNA was extracted from the FACS-sorted microglia that collected from freshly dissociated cortex at the site of cortical stab wound injury or from the equivalent site at the contralateral non-injured hemisphere of adult Cx3cr1CreER mice on days 1, 4, 7 and 14 following stab wound injury, in two biological replicates, using Illumina Hiseq4000.

last: Liu


title: Genome-wide ChIP-seq analysis of H2A.Z and H2A.Zac in HeLa Kyoto cells upon knockdown of JAZF1

city: Giessen

first: Marek

last: Bartkuhn

state: Hessen

address: Aulweg 132

department: Biomedical Informatics and Systems Medicine

country: Germany

institute: Justus-Liebig-University Giessen

summary: The goal was to investigate genome-wide if JAZF1 influences the deposition as well as the acetylation of the histone variant H2A.Z. ChIP-seq analysis revealed that depletion of JAZF1 leads to reduced acetylation of H2A.Z at specific sites.

overall_design: Genome-wide H2A.Z and H2A.Zac profiles of HeLa Kyoto cells upon knockdown of JAZF1 compared to control treatment (with biological replicates) were generated by ChIP-sequencing

first: Tobias

last: Friedrich

first: Tara

last: Procida


title: PRJNA608903 : Genome wide analysis of the transcription factor WIP in Arabidopsis

city: Orsay

first: Maria Victoria

last: Gomez-Roldan

address: bat 630

institute: CNRS

summary: In cucurbits, CmWIP1 is a master gene controlling sex determination. To bring new insight in the function of CmWIP1, we investigated two Arabidopsis WIP proteins, AtWIP1 and AtWIP2. Using an inducible system we showed that WIPs.  are powerful inhibitor of growth and inducer of cell death. Bioproject :

overall_design: ChIP assays were performed as described by Jégu et al. (2017). In total six libraries were sequenced, one for Dex:TT1-GFP and one for its input and two for Dex:NTT-GFP samples and two for its corresponding inputs.

first: Maria

last: Victoria

first: Gomez

last: Roldan

first: Farhaj

last: Izhaq

first: Marion

last: Verdenaud

first: John

last: Eleblu

first: Aimen

last: Haraghi

first: Vivien

last: Sommard

first: Pierre

last: Chambrier

first: David

last: Latrasse

first: Teddy

last: Jegu

first: Moussa

last: Benhamed

first: Judit

last: Szécsi

first: Mohammed

last: Bendahmane

first: Adnane

last: Boualem

first: Abdelhafid


title: Epigenetic and chromatin profiling of wild-type and Dnmt3a-deficient B1 and B2 B cells. (methylation)

city: Cambridge

first: Vinay

middle: S

last: Mahajan

state: MA

address: 65 Landsdowne Street

institute: Brigham and Women's Hospital

type: Methylation profiling by high throughput sequencing

summary: We have comprehensively interrogated wild type B1 and B2 B lineage cells and their progenitors. In mice bearing a conditional deletion of Dnmt3a in the B lineage, we integrated a variety of whole-genome profiling approaches including WGBS, TAB-Seq, RNA-seq, ATAC-Seq and CUT&RUN. The results reveal a stable “foundational methylome” in all B cells that establishes lymphocyte lineage identity. Superimposed on this foundational methylome is a “dynamic methylome” which is differentially modulated  in B1 and B2 B cells by the coincident activity/recruitment of DNMT3A and TET enzymes.

overall_design: We compared the CpG methylomes of wild-type and Dnmt3a-deficient B1 and B2 B cells using WGBS and TABSEQ


title: Single-Cell Transcriptomic Analysis of Somatosensory Neurons Uncovers Temporal Development of Neuropathic Pain

first: Kaikai

last: Wang

address: 393 Middle Huaxia Road

institute: ShanghaiTech University

summary: Peripheral nerve injury could lead to chronic neuropathic pain. Understanding transcriptional changes induced by nerve injury could provide fundamental insights into the complex pathogenesis of neuropathic pain. Gene expression profiles of dorsal root ganglia (DRG) under neuropathic pain condition have been studied. However, little is known about transcriptomic changes in individual DRG neurons after peripheral nerve injury. Here we performed single-cell RNA sequencing on dissociated mouse DRG cells after spared nerve injury (SNI). In addition to DRG neuron types also found under normal conditions, we identified three SNI-induced neuron clusters (SNIICs) characterized by the expression of Atf3/Gfra3/Gal (SNIIC1), Atf3/Mrgprd (SNIIC2) and Atf3/S100b/Gal (SNIIC3). These SNIICs were originated from Cldn9+/Gal+, Mrgprd+ and Trappc3l+ DRG neuron types. Interestingly, SNIIC2 was switched to SNIIC1 by increasing Gal and reducing Mrgprd expression 2 days after nerve injury. Inferring the gene regulatory networks underlying nerve injury, we revealed that activated transcription factor Atf3 and Egr1 in SNIICs could enhance Gal expression while activated Cpeb1 in SNIIC2 might suppress Mrgprd expression within 2 days after SNI. Furthermore, we screened the transcriptomic changes in the development of neuropathic pain to identify the potential analgesic targets. We revealed that the expression of cardiotrophin-like cytokine factor 1, which could activate the astrocytes in the dorsal horn of spinal cord, was increased in SNIIC1 neurons and contributed to SNI-induced mechanical allodynia. Therefore, our results provide a new framework to understand the changes in neuron types and the dynamics of molecular and cellular mechanisms underlying the development of neuropathic pain.

overall_design:
Dorsal root ganglion cells profile of C57 mice in Control, SNI 6h, SNI 24h, SNI 2d, SNI 7d, SNI 14d, and SNI 28d.
Please note that
The "barcode_of_raw_UMI_counts_1.txt" is the barcode list for each cell in "raw_UMI_counts_1.txt"
The   "barcode_of_raw_UMI_counts_2.txt" is the barcode list for each cell in "raw_UMI_counts_2.txt"
A part of processed data in "Smart-seq2_FPKM_metadata.txt" and "Smart-seq2_FPKM.txt" was obtained from GSE63576 and listed in "smart-seq2_data_from GSE63576.txt".

first: Changlin

last: Li

first: Xu

last: Zhang


title: Differential leaf transcriptomes for somatic embryogenesis of Elaeis guineensis

status: Public on Feb 21 2021

city: Bandar Baru Bangi

first: Siew-Eng

last: Ooi

state: Selangor

address: 6 Persiaran Institusi

department: Advanced Biotechnology and Breeding Centre

country: Malaysia

institute: Malaysian Palm Oil Board

summary: Identification of differentially expressed genes from RNA-seq data of non-embryogenic and embryogenic ortets. Selected ortets were previously cloned, thereby somatic embryogenesis rates are known for these ortets. Ortets fitting the study criteria were supplied by two agencies, namely L1 and L2. Principal Component Analysis indicated that variance between agencies were higher than the variance between embryogenesis groups within the agency. Therefore, differential analysis was conducted separately for each agency. Differential expression analysis using DESeq2 package suggested the L2 transcriptomes of zero and low embryogenesis groups were more similar compared to the high embryogenesis group. The L1 transcriptomes consisting of zero and low embryogenesis groups similarly showed overlapping clusters. Differential expression analysis was conducted on the L1  samples (low vs. zero embryogenesis) using DESeq2 R package and the identified differentially expressed genes (DEGs) was used for clustering analysis of the L2 samples. The clustering profiles suggested that expression of these DEGs in L2 samples were able to differentiate high embryogenesis from zero-low embryogenesis L2 groups.

overall_design: Spear leaf (frond F0) was sampled from ortets, supplied by two agencies, L1 (n=32) and L2 (n=20). From the previous somatic embryo production records, ortets were categorized into 3 groups, zero, low and high embryogenesis, with production rates of 0, 1-89 and ≥90 embryoid lines per ortet, respectively. Based on the embryogenesis data, samples from the L2 agency could be categorized into the 3 embryogenesis groups but samples from L1 were categorized into the zero and low groups only. Differential expression analysis was conducted separately for each agency. For the analysis, the zero-low group was designated as reference group for L2 while the zero group was designated as reference group for L1.

first: Ishak

last: Feshah

first: Azimi

last: Nuraziyan

first: Norashikin

last: Sarpan

first: Nabeel

last: Ata

first: Chin-Ching

last: Lim

first: Chin-Nee

last: Choo

first: Wei-Chee

last: Wong

first: Foo-Hin

first: Choo-Kien

first: Meilina

last: Ong-Abdullah


title: Characterization of Interleukin 11-producing fibroblasts that constitute a niche for tumor development

first: Takashi

last: Nishina


title: Identification of the FTO-FOXJ1 axis as a conserved regulatory module of embryonic and homeostatic motile ciliogenesis

status: Public on Feb 20 2021

city: Daejeon

first: Young-suk

last: Lee

address: 1113 CMS(E16), 291 Daehak-ro, Yuseong-gu,

department: Department of Bio and Brain Engineering

country: South Korea

institute: KAIST

summary: Adenosine N6-methylation (m6A) is one of the most pervasive mRNA modifications, and yet the physiological significance of m6A removal (demethylation) remains elusive. Here we report the m6A demethylase FTO functions as a conserved regulator of motile ciliogenesis. Mechanistically, FTO demethylates and thereby stabilizes the mRNA encoding the master ciliary transcription factor FOXJ1. Depletion of Fto in Xenopus laevis embryos caused widespread motile cilia defects, and Foxj1 was identified as one of the major phenocritical targets. In primary human airway epithelium, FTO depletion also led to FOXJ1 mRNA destabilization and a severe loss of ciliated cells with an increase of neighboring goblet cells. Consistently, Fto knockout mice showed strong asthma-like phenotypes upon allergen challenge owing to defective ciliated cells in the airway epithelium. Altogether, our study reveals a conserved role of the FTO-FOXJ1 axis in embryonic and homeostatic motile ciliogenesis.

overall_design: m6A-seq in Xenopus animal cap tissues

first: Hyunjoon

last: Kim

first: V

middle: N


title: ATAC Sequencing Facilitates  the Role of Nuclear Protein DEK in Regulating Chromatin Accessibility of Hematopoietic Stem Cells [ATAC-seq]

first: Zhe

last: Chen

summary: Purpose:To identify genes involved in the DEK regulating chromatin accessibility of hematopoietic stem cells (HSC) in mice, we performed ATAC-Sequence of HSC freshly sorted from mice.

overall_design: Methods: About 20,000 HSC (Lin-Sca-1+c-Kit+CD48-CD150+) were isolated by FACS from Dekfl/fl (floxp) and Dekfl/fl,Tie2-Cre (cKO). Genomic DNA  profiles of HSC treated with Tn5 enzyme were generated by deep sequencing, in duplicate, using Illumina HiSeq 2000.

first: Yu

last: Hou


title: Comparing breast tumor cell-induced gene expression in primary mammary fibroblasts from MEKK1-deficient mice to fibroblasts from MEKK1+/+ mice

city: Vermillion

first: Bruce

last: Cuevas

state: SD

address: 414 E. Clark Street

department: Basic Biomedical Sciences

institute: University of South Dakota

summary:
This study examined the importance of intracellular kinase MEKK1 in breast tumor-induced on gene expression in non-tumor mammary cells. The hypothesis tested in this study was that the kinase MEKK1 regulates breast tumor cells-induced expression of fibroblast genes that change the tumor microenvironment to promote breast tumor progression to metastasis.
Results reveal that MEKK1 regulates expression of chemokines and other genes in normal breast stroma cells that contribute to the development of a tumor microenvironment that promotes metastasis.

overall_design:
Primary mammary fibroblasts extracted from wild type or MEKK1-deficient mice. Total RNA was extracted from fibroblasts that were untreated (control) or treated with breast tumor-conditioned cell culture media for 8 hours. Each time point included 4 independent samples, for a total of 16 samples. Tumor cell-inducible gene expression was determined by comparing samples from the cells exposed to tumor factors to the untreated cells.

>>>Submitter states: Non-normalized data were discarded one year after the experiments were conducted in 2010 and are no longer available<<<


title: Targeting PDGFRa-activated Glioblastoma through Specific Inhibition of SHP-2-mediated Signaling

first: Haizhong

last: Feng

address: Pujian Road 160

department: Ren Ji Hospital

institute: Shanghai Jiao Tong University

summary: SHP-2 is a nonreceptor protein tyrosine phosphatase encoded by the PTPN11 gene and is critical for PDGFR-driven gliomagenesis. SHP099, a novel and potent SHP-2 inhibitor, preferentially attenuated the tumorigenicity of GBM and glioma stem-like cells (GSCs) compared to normal brain cells or neural progenitor cells in vitro. Delivered orally, SHP099 crosses the blood-brain barrier (BBB) and accumulates at efficacious concentrations in the brains, as determined using two different orthotopic xenograft models. SHP099 inhibited tumor growth and extended survival in activated PDGFR-signaling xenografts alone and in combination with first-line chemotherapeutic agent, temozolomide (TMZ). This study highlights SHP-2 inhibitor SHP099 potential for treatment of clinical GBM in combination with TMZ.

overall_design: Examination of effects of SHP-2 inhibitor SHP099 treatment on differential gene expression in glioma cells.


title: Alteration of lipid homeostasis by hydroxypropyl cyclodextrin improves amiodarone-induced alveolar cell disorder

city: Kyoto

first: Shimpei

last: Gotoh

address: 53 Kawaharacho, Shogoin, Sakyo-ku

department: Center for iPS cell Research and Application

institute: Kyoto University

summary: We developed LB cells overexpressing ABCA3, SFTPB, SFTPC, and SFTPD in A549 cells. Using LB cells in high content screening, we identified HPbCD. HPbCD improved  amiodarone-induced LB abnormality in iPS cell-derived alveolar organoids.

overall_design: A549 cells and LB cells were seeded and cultured with DMSO or 10 mg/ml HPbCD. After 24h incubation, the cells were treated with DMSO or 10 μM amiodarone for 24h and harvested. The transcriptomes of these cells were obtained based on paired-end reads (100-bp) with Illumina Novaseq 6000.

first: Shuhei

last: Kanagaki


title: A whole virion vaccine for COVID-19 produced via a novel inactivation method:  results from animal challenge model studies

status: Public on Feb 19 2021

city: Fort Collins

first: Raymond

last: Goodrich

state: Colorado

address: 3185 Rampart Road

department: Microbiology, Immunology, anad Pathology

institute: Colorado State University

summary: The COVID-19 pandemic has generated intense interest in the rapid development and evaluation of vaccine candidates for this disease and other emerging diseases. Several novel methods for preparing vaccine candidates are currently undergoing clinical evaluation in response to the urgent need to prevent the spread of COVID-19. In many cases, these methods rely on new approaches for vaccine production and immune stimulation. We report on the use of a novel method (SolaVAXTM) for production of an inactivated vaccine candidate and the testing of that candidate in a hamster animal model for its ability to prevent infection upon challenge with SARS-CoV-2 virus.  The studies employed in this work included an evaluation of the levels of neutralizing antibody produced post-vaccination, levels of specific antibody sub-types to RBD and spike protein that were generated, evaluation of viral shedding post-challenge, flow cytometric and single cell sequencing data on cellular fractions and histopathological evaluation of tissues post-challenge. The results from this study provide insight into the immunological responses occurring as a result of vaccination with the proposed vaccine candidate and the impact that adjuvant formulations, specifically developed to promote Th1 type immune responses, have on vaccine efficacy and protection against infection following challenge with live SARS-CoV-2. This data may have utility in the development of effective vaccine candidates broadly. Furthermore, the results suggest that preparation of a whole virion vaccine for COVID-19 using this specific photochemical method may have utility in the preparation of one such vaccine candidate.

overall_design: Control (without vaccination) : n=3, SolaVAX vaccinated: n=3, SolaVAX+CpG1018: n=3, SolaVAX+ODN1668: n=3. A total of 12 hamsters were analyzed.

first: Izabela

last: Ragan

first: Lindsay

middle: M

last: Hartson

first: Taru

last: Dutt

first: Andres

last: Obregon-Henao

first: Rachel

last: Maison

first: Paul

last: Gordy

first: Amy

last: Fox

first: Burton

middle: R

last: Karger

first: Shaun

last: Cross

first: Marylee

middle: L

last: Kapuscinski

first: Sarah

last: Cooper

first: Brendan

last: Podell

first: Mark

middle: D

last: Stenglein

first: Richard

last: Bowen

first: Marcela

last: Henao-Tamayo

middle: P


title: Broad transcriptional firing represses bactericidal activity in human airway neutrophils [10hours_transmigrated_CFASN_Neutrophils]

city: Atlanta

first: Diego

last: Moncada

state: Georgia

address: 201 Dowman Dr

department: Pediatrics

institute: Emory University

summary: In this model, naïve blood neutrophils are migrated through a small airway epithelium grown at air-liquid interface toward CF airway fluid supernatant (CFASN, which corresponds to sputum sequentially centrifuged to remove cells and bacteria) or leukotriene B4 (LTB4, transmigration control), placed apically.

overall_design: To investigate a potential causal relationship between de novo transcription and neutrophil fate acquisition, we transmigrated neutrophils in the model for 10 hours toward CFASN or LTB4.

first: Camilla

last: Margaroli

last: Moncada Giraldo

first: Rabindra

last: Tirouvanziam


title: Segment-specific kinetics of mRNA, cRNA and vRNA during influenza infection

city: Minneapolis

first: Wei-Shou

last: Hu

state: MN

address: 421 Washington Ave. SE

institute: University of Minnesota

summary: Influenza A virus (IAV) is a segmented negative-sense RNA virus and is the cause of major global epidemics and pandemics. The replication of IAV is complex, involving both transcription and replication, during which three distinct RNA species, namely mRNA, cRNA, and vRNA are generated for all eight genome segments. While understanding IAV replication kinetics is important for drug development and improving vaccine production, current methods for studying IAV kinetics has been limited by the ability to detect all three different RNA species in a scalable manner. Here were report the development of a novel pipeline using total stranded RNA-Seq, named Influenza Virus Enumerator of RNA Transcripts (InVERT) which allows for the simultaneous quantification of all three RNA species of IAV. Using InVERT provides a full landscape of the IAV replication kinetics, in which we found that different groups of viral genes followed different traits of kinetics. During a cycle of infection, the RNA-dependent RNA Polymerase (RdRP) produced more vRNA than mRNA while some other genes (NP, NS, HA) consistently make more mRNA than vRNA. vRNA expression levels do not correlate with the cRNA expression, suggesting complex regulations of vRNA synthesis. Furthermore, by studying the kinetics of a virus lacking the capacity to generate new polymerase complexes, we found evidence that further supports the model that cRNA synthesis requires newly synthesized RdRP and that incoming RdRP can only generate mRNA.

sample_organism: Influenza A virus (A/Puerto Rico/8/1934(H1N1))

overall_design: Time-series transcriptome profiling of IAV PR/8/34 in MDCK cells and Delta-HA and Delta-PB1 IAV in A549 cells

first: Thu

last: Phan

first: Elizabeth

middle: J

last: Fay

first: Zion

last: Lee

first: Stephanie

last: Aron

first: Ryan

last: Langlois


title: Endometrium on-a-chip reveals insulin- and glucose-induced alterations in the transcriptome

city: Leeds

first: LeedsOmics

last: UoLeeds

state: West Yorkshire

address: Clarendon Way

department: Cross-faculty

country: United Kingdom

institute: University of Leeds

summary: The molecular interactions between the maternal environment and the developing embryo that are key for early pregnancy success are known to be influenced by factors such as maternal metabolic status. Our understanding of the mechanism(s) through which these individual nutritional stressors alter endometrial function and the in utero environment for early pregnancy success is, however, limited. Here we report, for the first time, the use of an endometrium-on-a-chip microfluidics approach to produce a multi-cellular endometrium in vitro, that was exposed to glucose and insulin concentrations associated with maternal metabolic stressors. Following isolation of endometrial cells (epithelial and stromal) from the uteri of non-pregnant cows in the early-luteal phase (Day 4-7 approximately), epithelial cells were seeded in the upper chamber of the device (4-6 104 cells/mL) and stromal cells seeded in the lower chamber (1.5-2 104 cells/mL). Three different concentration of glucose (0.5, 5.0 or 50 mM) or insulin (Vehicle, 1 or 10 ng/mL) were performed in the endometrial cells at a flow rate of 1µL/min for 72 hr to mimic the rate of secretion in vivo. Quantitative differences in the transcriptomic response of the cells and the secreted proteome of in vitro-derived uterine luminal fluid (ULF) were determined by RNA-sequencing and Tandem Mass Tagging Mass Spectrometry (TMT), respectively. Changes in maternal glucose altered 21 and 191 protein-coding genes in epithelial and stromal cells, respectively (p<0.05), with a dose-dependent quantitative change in the protein secretome (1 and 23 proteins in epithelial and stromal cells, respectively). Altering insulin concentrations resulted in limited transcriptional changes including transcripts for insulin-like binding proteins that were cell specific (5, 12, and 20) but altered the quantitative secretion of 196 proteins including those involved in extracellular matrix-receptor interaction and proteoglycan signaling in cancer. Collectively, these findings highlight one potential mechanism by which changes to maternal glucose and insulin associated with metabolic stress may alter uterine function.

overall_design: Two different concentrations of glucose (5 or 50 mM) and three of insulin (Vehicle, 1 or 10 ng/mL) were assessed on two sets of endometrial cells (epithelial or stromal) in three biological replicates, totalizing 30 single-end fastq files (12 glucose experiment + 18 insulin experiment)

first: Tiago

last: DeBem

first: Haidee

last: Tinning

first: Elton

last: Vasconcelos

first: Dapeng

last: Wang

first: Niamh

last: Forde


title: Impaired CHD6 function links misregulation of autophagy and DNA damage response to premature ageing

city: Cologne

first: Yulia

last: Kargapolova

address: Robert-Koch-Straße 21

department: CMMC

institute: Cologne University

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

overall_design: Refer to individual Series


title: Deep sequencing of pre-translational mRNPs reveals hidden flux through evolutionarily conserved AS-NMD pathways

city: Worcester

first: Carrie

last: Kovalak

address: 365 Plantation Street

department: RNA Therapeutics Institute

institute: University of Massachusetts Medical School

summary:
Background:  Alternative splicing (AS), which generates multiple mRNA isoforms from single genes, is crucial for the regulation of eukaryotic gene expression.  The flux through competing AS pathways cannot be determined by traditional RNA-Seq, however, because different mRNA isoforms can have widely differing decay rates.  Indeed, some mRNA isoforms with extremely short half-lives, such as those subject to translation-dependent nonsense-mediated decay (AS-NMD), may be completely overlooked in even the most extensive RNA-Seq analyses.
Results:  RNA immunoprecipitation in tandem (RIPiT) of exon junction complex (EJC) components allows for purification of post-splicing mRNA-protein particles (mRNPs) not yet subject to translation (pre-translational mRNPs) and, therefore, translation-dependent mRNA decay.  Here we compared EJC RIPiT-Seq to whole cell RNA-Seq data from HEK293 cells.  Consistent with expectation, the flux through known AS-NMD pathways is substantially higher than that captured by RNA-Seq.  Our EJC RIPiT-Seq also definitively demonstrates that the splicing machinery itself has no ability to detect reading frame.  We identified thousands of previously unannotated splicing events; while many can be attributed to “splicing noise”, others are evolutionarily-conserved events that produce new AS-NMD isoforms likely involved in maintenance of protein homeostasis.  Several of these occur in genes whose overexpression has been linked to poor cancer prognosis.
Conclusions:  Deep sequencing of RNAs in post-splicing, pre-translational mRNPs provides a means to identify and quantify splicing events without the confounding influence of differential mRNA decay.  For many known AS-NMD targets, the NMD-linked AS pathway predominates.   EJC RIPiT-Seq also enabled identification of numerous conserved but previously unknown AS-NMD events.

overall_design: We created and sequenced rRNA-depleted whole cell RNA-Seq libraries (84-93 million mate pairs each) wherein the captured fragments were of similar length (220-500 nts) to our previously published EJC RIPiT-Seq libraries (GEO: GSE115788).  The new RNA-Seq libraries were generated from cultures (three biological replicates, two technical replicates each) that were (+) or were not (–) subjected to a one hour pre-treatment with harringtonine. Technical replicates were later combined.

first: Scott

last: Donovan

first: Alicia

last: Bicknell

first: Mihir

last: Metkar

first: Melissa

last: Moore


title: Transcriptional profiling of different subsets of PBMCs-CD4+, CD8+, CD14+, CD21+, CD335+ reveals differential response to Sungri/96 PPRV vaccination in Goat and Sheep

city: Karnal

first: Ravi Kumar

last: Gandham

state: Haryana

address: Makrampur

country: India

institute: ICAR-NBAGR

summary: We have used RNA-Seq to examine  expression profiles of different subsets of PBMCs-CD4+, CD8+, CD14+, CD21+, CD335+  from Goat and Sheep under sungri/96 PPRV vaccination.

sample_organism:
Capra hircus
Ovis aries

platform_organism:
Capra hircus
Ovis aries

overall_design: Goat and Sheep were subjected to Sungri/96 PPRV vaccination. RNA isolated from different subsets of PBMCs -CD4+, CD8+, CD14+, CD21+, CD335+ before vaccination (0 day)  and post vaccination (5 day) were subjected to illumina sequencing in both the species.

first: S

last: Wani

first: A

last: Sahu

last: Pandey

first: RIN

last: Khan

first: M

last: Praharaj

first: B

last: Mishra

first: R

last: Singh


title: Impaired CHD6 function links misregulation of autophagy and DNA damage response to premature ageing [ChIP-Seq]

summary: Members of the Chromodomain-Helicase-DNA binding (CHD) protein family are chromatin remodelers critically implicated in human pathologies. CHD6 is the least studied member of this family, and we were motivated to dissect its in-cell roles by discovering a mutated CHD6 allele in a patient suffering from the rare Hallermann-Streiff premature aging syndrome (HSS). We generated isogenic iPSC lines carrying (or not) this single point mutation in the CHD6 SANT/SLIDE domain, which allow studying HSS-relevant cell identities. Using these lines, we show for the first time that CHD6 binds the promoters of a cohort of autophagy and stress response genes across cell types. This CHD6 mutation impairs its ability to recruit co-factors and regulate genes in response to DNA damage and autophagy stimulation, thus leading to accumulation of unresolved DNA damage burden. By combining genomics and functional assays, we describe a molecular mechanism underlying chromatin control of autophagic flux and genotoxic stress surveillance that is broadly applicable to human cell types and can explain HSS establishment.

overall_design: CHD6 ChIP-Seq in three cell lines (iPSCs, derived neural crest cells, derived cardiomyocytes) and TFEB ChIP-Seq in iPSCs.

first: Argyris

last: Papantonis


title: Targeting OCT3 attenuates doxorubicin-induced cardiac injury

city: Columbus

first: Pearlly

last: Yan

state: OH

department: Division of Hematology

institute: The Ohio State University

summary: Doxorubicin is a commonly used anticancer agent that can cause debilitating and irreversible cardiac injury. The initiating mechanisms contributing to this side effect remain unknown, and current preventative strategies only offer modest protection. Using stem cell-derived cardiomyocytes from patients receiving doxorubicin, we probed the transcriptomic landscape of solute carriers and identified OCT3 (SLC22A3) as a critical transporter regulating the cardiac accumulation of doxorubicin. Functional validation studies in mice revealed that targeting OCT3 can attenuate cardiac dysfunction without compromising the anticancer properties of doxorubicin. These findings provide a rationale for the development of targeted approaches to mitigate this debilitating toxicity

overall_design: Evaluating cardiac mRNA expression in untreated or doxorubicin-treated FVB wild-type or OCT3-knockout mice

first: Kevin

last: Huang

first: Megan

middle: Z

last: Thomas

first: Tarek

last: Magdy

first: Eric

last: Eisenmann

first: Muhammad

middle: E

last: Uddin

first: Duncan

middle: F

last: DiGiacomo

first: Alexander

last: Pan

first: Ralf

last: Bundschuh

first: Markus

last: Keiser

last: Otter

first: Sherry

middle: H

last: Xia

first: Yang

last: Li

first: Yan

last: Jin

first: Qiang

last: Fu

first: Alice

last: Gibson

first: Ingrid

last: Bonilla

first: Cynthia

last: Carnes

first: Kara

last: Corps

first: Vincenzo

last: Coppola

first: Sakima

last: Smith

first: Daniel

last: Addison

first: Anne

last: Nies

first: Taosheng

last: Chen

first: Maryam

middle: B

last: Lustberg

first: Joanne

last: Wang

first: Stefan

last: Oswald

first: Sharyn

last: Baker

first: Shuiying

last: Hu

first: Paul

middle: W

last: Burridge

first: Alex

last: Sparreboom


title: The CNOT4 Subunit of the CCR4-NOT Complex is Involved in mRNA Degradation, Efficient DNA Damage Repair, and XY Chromosome Crossover during Male Germ Cell Meiosis

city: Hangzhou

first: Xingxing

last: Dai

address: YuHangTang 866

institute: Life Sciences Institute

summary: The CCR4-NOT complex is a major mRNA deadenylase in eukaryotes and comprise CNOT6/6L and CNOT7/8, both of which are two types of catalytic subunits, as well as CNOT4, which is a regulatory subunit with previously undetermined functions. These subunits are previously hypothesized to play synergistic biochemical functions; however, their in vivo functions have remained poorly investigated. In this study, we constructed viable Cnot6/6l double knockout mice and the males were fertile, whereas Cnot7–/– male mice have been reported to be infertile. These results are against canonical viewpoints and indicate that CNOT7 has CNOT6/6L-independent functions in vivo. We also demonstrate that the mouse testis has an endogenous complex that contains CNOT7 and CNOT4, but not CNOT6/6L. CNOT4 is required for post-implantation embryo development and meiosis progression during spermatogenesis. Selective knockout of Cnot4 in male germ cells leads to defective DNA damage repair and homologous crossover formation between X and Y chromosomes in a cell autonomous manner. CNOT4 functions as a previously unrecognized mRNA adaptor of CCR4-NOT by targeting mRNAs to CNOT7 for deadenylation of the poly(A) tails, thereby mediating the degradation of a subset of transcripts from the zygotene to pachytene stage. Therefore, the mRNA removal promoted by CNOT4-regulated CCR4-NOT complex during the zygotene to pachytene transition is crucial for the selection of DNA repair pathways, subsequent activation of meiosis-related genes, sex chromosome paring, and ultimately, male fertility.

overall_design: Examination of WT and CNOT4-null transcriptome in zygotene and pachytene stage.

first: Xing

middle: X

first: Yu

last: Jiang

first: Jia

last: Gu

first: Zhi

middle: Y


title: Host-virus chimeric events in SARS-CoV2 infected cells are infrequent and artifactual

city: West Lafayette

first: Majid

last: Kazemian

state: Indiana

address: 201 S. University Street

institute: Purdue university

summary: Pathogenic mechanisms underlying severe SARS-CoV2 infection remain largely unelucidated. High-throughput sequencing technologies that capture genome and transcriptome information are key approaches to gain detailed mechanistic insights from infected cells. These techniques readily detect both pathogen and host-derived sequences, providing a means of studying host-pathogen interactions. Recent studies have reported the presence of host-virus chimeric (HVC) RNA in RNA-seq data from SARS-CoV2 infected cells and interpreted these findings as evidence of viral integration in the human genome as a pathogenic mechanism. Since SARS-CoV2 is a positive sense RNA virus that replicates in the cytoplasm it does not have a nuclear phase in its life cycle, so it is biologically unlikely to be in a location where splicing events could result in genome integration. Here, we investigated the biological authenticity of HVC events. In contrast to true biological events, e.g. mRNA splicing and genome rearrangement events, which generate reproducible chimeric sequencing fragments across different biological isolates, we found that HVC events across >100 RNA-seq libraries from patients with COVID-19 and SARS-CoV2 infected cell lines, were highly irreproducible. RNA-seq library preparation is inherently error-prone due to random template switching during reverse transcription of RNA to cDNA. By counting chimeric events observed when constructing an RNA-seq library from human RNA and spike-in RNA from an unrelated species, such as fruit-fly, we estimated that ~1% of RNA-seq reads are artifactually chimeric. In SARS-CoV2 RNA-seq we found that the frequency of HVC events were, in fact, no more frequent than this background “noise”. Finally, we developed a novel experimental approach to enrich SARS-CoV2 sequences from bulk RNA of infected cells. This method enriched viral sequences but did not enrich for HVC events, suggesting that the majority of HVC events are, in all likelihood, artifacts of library construction. In conclusion, our findings indicate that HVC events observed in RNA-sequencing libraries from SARS-CoV2 infected cells are extremely rare and are likely artifacts arising from either random template switching of reverse-transcriptase and/or sequence alignment errors. Therefore, the observed HVC events do not support SARS-CoV2 fusion to cellular genes and/or integration into human genomes.

overall_design: To experimentally enrich for viral RNAs from total RNA of SARS-CoV2 infected cells prior to RNA-seq library preparation, we developed a series of in vitro amplification steps using SARS-CoV2 virus specific primers (VSP) as following. The VSP pool contained ~30 oligonucleotides that span all SARS-CoV2 genes (at least one oligo per gene and nearly 1 oligo per 1kb of the genome). Given our goal to additionally enrich for potential HVC events, we used two approaches, enrichment methods 1 (5’->3’ host-virus chimeras) and enrichment method 2 (5’->3’ virus-host chimeras).

first: Bingyu

last: Yan

first: Srishti

last: Chakravorty

first: Carmen

last: Mirabelli

last: Chauss

first: Dhaneshwar

last: Kumar

first: Christiane

last: Wobus

first: Behdad

last: Afzali


title: RNAseq analysis of mouse CD4+ regulatory Tregs in WT and ∆Foxp3 mice isolated from spleen and lungs.

status: Public on Feb 18 2021

city: Boston

first: CBDM

last: Lab

address: 77 Avenue Louis Pasteur

department: Microbiology and Immunobiology

institute: Harvard Medical School

summary: RNAseq analysis of mouse CD4+ regulatory Tregs in 21-day-old male WT and ∆Foxp3 mice isolated from spleen and lungs.

overall_design: Mouse CD4+ regulatory Tregs  in 21-day-old male WT and ∆Foxp3 mice were isolated from spleen and lungs by flow cytometry, as DAPI–TCRβ+CD4+GFP+CD25+. 1,000 cells were double-sorted directly into lysis buffer for RNA-seq (Smart-seq2)  library preparation following the standard ImmGen low-input protocol (www.immgen.org).

first: David

last: Zemmour

first: Louis

last: Charbonnier

first: Juliette

last: Leon

first: Tala

last: Chatila

first: Diane

last: Mathis

first: Christophe

last: Benoist


title: Co-cultures of enteric neurons and T cells

summary: We reported transcriptional characterization of  Tconvs, Treg and enteric neurons in T cells/neuron co-cultures

overall_design: Naïve T cells were co-cultured with or without enteric neurons for 3 days in iTreg culture conditions, and Tconvs, Treg or enteric neurons were sorted and processed for RNAseq

first: Yiqing


title: MMP (muscularis myenteric plexus) RNAseq profilings in Germ-free (GF) colons with microbe stimulations

summary: We reported transcriptional characterization of MMP with the stimulations of C.ramosum or P.magnus in Germ-free colons

overall_design: Germ-free mice were gavaged with vehicle, P.magnus or C.ramosum for the indicated time, and the whole MMP tissues were prepared and further processed for RNAseq


title: RNAseq analysis of human CD4+ regulatory Tregs and Tconvs in IPEX and healthy donors isolated from peripheral blood.

overall_design: Human Tregs and Tconvs from IPEX and healthy donors were sorted from PBMCs as DAPI– CD4+CD25hiCD127lo and CD4+CD25–CD127hi, respectively. 1,000 cells were double-sorted directly into lysis buffer for RNA-seq (Smart-seq2)  library preparation following the standard ImmGen low-input protocol (www.immgen.org). Samples were collected and sequenced in two cohorts.

first: Marianne

last: Delville

first: Emmanuelle

last: Six

first: Isabelle

last: André


title: The NAC transcription factor FaRIF controls fruit ripening in strawberry

city: Málaga

last: Posé

address: Avenida Louis Pasteur, 49

country: Spain

institute: Instituto de Hortofruticultura Subtropical y Mediterránea

summary: In contrast to climacteric fruits such as tomato, the knowledge on key regulatory genes controlling the ripening of strawberry, a non-climacteric fruit, is still limited. NAC transcription factors mediate different developmental processes in plants. Here, we identified and characterized FaRIF (Ripening Inducing Factor), a NAC transcription factor that is highly expressed and induced in strawberry receptacles during ripening. Functional analyses based on stable transgenic lines aimed at silencing FaRIF by RNA interference, either from a constitutive promoter or the ripe receptacle-specific EXP2 promoter, as well as overexpression lines showed that FaRIF controls critical ripening-related processes such as fruit softening and pigment and sugar accumulation. Physiological, metabolome and transcriptome analyses of receptacles of FaRIF-silenced and overexpression lines point to FaRIF as a key regulator of strawberry fruit ripening from early developmental stages, controlling abscisic acid (ABA) biosynthesis and signaling, cell wall degradation and modification, the phenylpropanoid pathway, volatiles production, and the balance of the aerobic/anaerobic metabolism. FaRIF is therefore a target to be modified/edited to control the quality of strawberry fruits.

overall_design: Transcriptome analysis from strawberry receptacles from control and 35Spro:FaRIF-RNAi #3 and 35Spro:FaRIF-RNAi #11 transgenic lines at the white and red stages or ripening, and from receptacles of control and EXP2pro:FaRIF-RNAi #1, EXP2pro:FaRIF-RNAi #7 and 35Spro:FaRIF #1 at the red stage. Three biological replicates of each sample was analyzed.

last: Martín-Pizarro

first: José

middle: G

last: Vallariino

first: Sonia

last: Osorio

first: Victoriano

last: Meco

first: María

last: Urrrutia

first: Jeremy

last: Pillet

first: Ana

last: Casañal

first: Catharina

last: Merchante

first: Iraida

last: Amaya

first: Lothar

last: Willmitzer

first: Alisdair

last: Fernie

first: James

last: Giovannoni

first: Miguel

middle: A

last: Botella

last: Valpuesta


title: RNAseq profilings of enteric neurons with microbe stimulations

first: CBDM

last: Lab

summary: We reported transcriptional characterization of enteric neurons with the stimulations of C.ramosum or P.magnus in Germ-free colons

overall_design: Germ-free mice were gavaged with vehicle, P.magnus or C.ramosum for the indicated time, and the enteric neurons were sorted, and further processed for RNAseq

last: Yan

first: Christophe

last: Benoist

first: Diane

last: Mathis


title: Characterization of Interleukin 11-producing fibroblasts that constitute a niche for tumor development [Agilent]

city: Ota-ku

first: Takashi

last: Nishina

address: 5-21-16 Omori-nishi

institute: Toho University

summary: Interleukin (IL)-11 belongs to a member of the IL-6 family of cytokines and is involved in a variety of cellular responses. However, the origin and functions of IL-11-producing (IL-11+) cells are not fully understood. To characterize IL-11+ cells in vivo, we generated Il11 reporter mice. IL-11+ cells rapidly appeared in the colon of mice after dextran sulfate sodium treatment. IL-11+ cells expressed fibroblast markers and genes associated with cell proliferation and tissue repair. IL-11+ cells also appeared in the colon of three murine tumor models and human colorectal cancers. Deletion of Il11ra1 or Il11 attenuated the development of colitis-associated colorectal cancers in mice. IL-11 induced STAT3 phosphorylation in colonic fibroblasts, and injection of IL-11 receptor agonist upregulated many genes that were enriched in IL-11+ cells. Together, tumor cells induced IL-11+ fibroblasts, and a feed-forward loop between IL-11 and IL-11+ fibroblasts might constitute a niche for promoting tumor development.

overall_design: To identify target genes by IL-11 in the colon, we intravenously injected 10 mg of the IL-11 receptor agonist, NT-3N into wild-type C57BL/B6 mice (n=2-3 mice per each time point). Then mice were sacrificed, and the colon was removed at 3 hours after injection.

first: Hiroyasu

last: Nakano


title: Functional and conformational impact of cancer-associated SF3B1 mutations depends on the position and the charge of amino acid substitution

city: Paris

first: Alexandre

last: Houy

address: 26 rue d'Ulm

institute: Institut Curie

summary: RNA-sequencing data from HEK293T cell models: Hek293T transfected with expression vectors of SF3B1-WT, SF3B1-R625H, SF3B1-K666T, SF3B1-K700E.

overall_design: In cellulo examination of SF3B1 mutations in HEK293T cell models.

first: Christine

last: Canbezdi

first: Malcy

last: Tarin

first: Dorine

last: Bellanger

first: Tatiana

last: Popova

first: Marc

last: Stern

first: Sergio

last: Roman-Roman

first: Samar

last: Alsafadi


title: Symmetry breaking of tissue mechanics in wound induced hair follicle regeneration [bulk RNA-seq]

city: Los Angeles

first: Hans I-Chen

last: Harn

state: CA

address: 2011 Zonal Ave. HMR304

institute: University of Southern California

summary: Tissue regeneration is a process that recapitulates the molecular and mechanical aspects of development and evolution. We use the wound-induced hair neogenesis (WIHN) model to investigate the mechanical and molecular responses of the laboratory (Mus) and African spiny (Acomys) mice. Laboratory and spiny mice showed an opposite trend of spatiotemporal morphogenetic field for WIHN during wound healing, and wound stiffness gradient across the whole wound bed predicated pattern of hair formation. Using bulk and single-cell RNA-seq analysis and K14-Cre-Twist1 transgenic mice, we identified the central role of the Twist1 pathway as the mediator of epidermal-dermal interaction and the emergence of periodic hair primordia. Lastly, we generated a Turing model with an underlying measure of stiffness to support a two-scale tissue mechanic model to explain the setup of a morphogenetic field from a wound bed (mm scale) or periodically arranged hair primordia from a morphogenetic field (μm scale). Delineating the common and distinct chemo-mechanical events during regenerative wound healing between laboratory and African spiny mice reveal its evo-devo advantages, which provide new perspectives for regenerative medicine.

sample_organism:
Acomys cahirinus
Mus musculus

platform_organism:
Acomys cahirinus
Mus musculus

overall_design: bulk RNA-Seq for epidermal and dermal tissues from lab mouse and spiny mouse

first: Cheng-Ming

last: Chuong

first: Hans

middle: I


title: Next Generation Sequencing Quantitative Analysis of Wild Type and Mki67-/- 3T3 cell transcriptomes

city: Montpellier

first: Daniel

last: Fisher

address: 1919 Route de Mende

department: IGMM

institute: CNRS

summary:
Purpose: The goals of this study were to compare gene expression profiles of mouse NIH/3T3 cells with intact or disrupted Mki67 gene.
Methods: mRNA profiles of wild-type (WT) and two clones of Ki-67 knockout (Mki67−/−) mouse NIH/3T3 cells, all in duplicate, were generated by deep sequencing using Illumina Hiseq2500 in single-end read mode, 50bp. The sequence reads that passed quality filters were aligned to the mouse genome, quantified, and differential gene expression (DGE) analysis was performed using DEseq2.
Results: Using an optimized data analysis workflow, we mapped about 30 million sequence reads per sample to the mouse genome (GRCm38.p6) and identified differentially expressed genes.  This revealed widespread changes in gene expression, with 2558 genes significantly deregulated in independent clones of Mki67-/- cells (q < 0.05)
Conclusions: Our study represents the first detailed analysis of retinal transcriptomes, with biologic replicates, generated by RNA-seq technology. The optimized data analysis workflows reported here should provide a framework for comparative investigations of expression profiles. Our results show that NGS offers a comprehensive and more accurate quantitative and qualitative evaluation of mRNA content within a cell or tissue. We conclude that RNA-seq based transcriptome characterization would expedite genetic network analyses and permit the dissection of complex biologic functions.

overall_design: mRNA profiles of wild-type (WT) and two clones of Ki-67 knockout (Mki67−/−) mouse NIH/3T3 cells, all in duplicate, were generatedby deep sequencing using Illumina Hiseq2500.

first: Priyanka

last: Singh

first: Karim

last: Mrouj

first: Geronimo

last: Dubra

first: Liliana

last: Krasinska


title: Impaired CHD6 function links misregulation of autophagy and DNA damage response to premature ageing [RNA-Seq]

first: Yulia

last: Kargapolova

overall_design:
polyA enriched 3'-end RNA sequencing of iPS cell lines upon etoposide treatment, starvation and culturing normal conditions.
polyA-enriched mRNA profiles of iPS derived neural crest cells and cardiomyocytes with or without mutated CHD6 protein.

first: Argyris

last: Papantonis


title: Irx3 and Irx5 regulate hypothalamic postnatal neurogenesis and leptin sensitivity

city: Toronto

first: Chi-chung

last: Hui

state: ON

address: 686 Bay street

country: Canada

institute: The Hospital for Sick Children

overall_design: Refer to individual Series


title: Ki-67 promotes carcinogenesis by enabling global transcriptional programmes

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus


title: Yap regulates fatty acid oxidation in skeletal muscle

city: Melbourne

first: Enzo

last: Porrello

state: Victoria

address: c/o Royal Children’s Hospital

department: Cell Biology

country: Australia

institute: Murdoch Children's Research Institute


title: The role of queuine as a nutritional regulator of the response of Entamoeba histolyticato oxidative stress

first: Rachel

last: Legendre

address: 28, rue du docteur Roux

department: Research and Resource Center for Scientific Informatics

institute: Institut Pasteur

summary: Amebiasis is an intestinal disease transmitted by the protist parasite, Entamoeba histolytica, following the ingestion of contaminated food and water. Many infected patients (90%) are asymptomatic but for unknown reasons, these trophozoites can become virulent and invasive, cause amebic dysentery, and migrate to the liver, via the portal veins, where they cause hepatocellular damage. We have recently reported that Escherichia coli can modulate E.histolytica 's virulence and resistance to oxidative stress (OS) via the production of oxaloacetate. Queuosine is a naturally occurring modified nucleoside and is found in the first position of the anticodon of the transfer RNAs for Asp, Asn, His and Tyr. Lower and higher eukaryotes are supplied with queuine (the base of queuosine) in the diet or by the intestinal flora. Here, we have tested the hypothesis that queuine being a micronutrient offers an advantage to E.histolytica during OS. Queuine which is efficiently incorporated in E.histolytica’s tRNAs leads to significant modifications of the parasite’s transcriptome, to hypermethylation of C38 in tRNAAspGTC and consequently to OS resistance. While queuine leads to the translation of proteins during OS, we found that that it inhibits protein synthesis otherwise. Localization studies with an antibody against EhTGT in queuine-treated trophozoites revealed the presence of small granular structures. We have biochemically characterized the parasite’s tRNA-guanine transglycosylase (TGT), the enzyme that catalyzes the modification of Q-tRNAs. Trophozoites silenced for TGT expression have a growth defect and they are less resistant to OS compared to control trophozoites.

overall_design: queuine with or without oxidative stress

first: Shruti

last: Nagaraja

first: Maggi

last: Cai

first: Lotem

last: Sarid

first: Yana

last: Shaulov

first: Meirav

last: Trebicz-Geffen

first: Mohit

last: Mazumdar

first: Hugo

last: Varet

first: Jean-Yves

last: Coppée

first: Peter

middle: C

last: Dedon

first: Samudrala

last: Gourinath

first: Nancy

last: Guillen

first: Serge

last: Ankri


title: Irx3 and Irx5 regulate hypothalamic postnatal neurogenesis and leptin sensitivity (DROP-seq)

summary: Drop-Seq single cell RNA sequencing of hypothalamic arcuate nucleus and median eminence from wildtype and Irx3/5 double heterozygous mice

overall_design: Drop-seq was perfomed on WT and Irx3/5dHet adult mouse arcuate nucleus-median eminence. Micro-dissected tissues from 5-7 mice per each group were pooled together for single cell  dissociation, followed by Drop-seq sequencing.

first: Joe

last: Son

first: Zhengchao

last: Dou

first: Troy

last: Ketela

first: Chi-Chung


title: Phosphatidylinositol-3 kinase signaling controls survival and stemness of hematopoietic stem and progenitor cells

status: Public on Feb 17 2021

city: Utrecht

first: Jeroen

last: den Hertog

state: Utrecht

address: Uppsalalaan 8

institute: Hubrecht Institute

summary: Loss of Pten and inhibition of PI3K induced apoptosis of hematopoietic stem/progenitor cells upon endothelial to hematopoietic transition and surviving hematopoietic stem/progenitor cells committed to all blood lineages but displayed reduced stemness

overall_design: AGMs (36hpf) or CHTs (5dpf) were dissected and dissociated, using FACS sorting HSPCs were isolated and single cell RNA seq was performed.

first: Sasja

last: Blokzijl-Franke

first: Bas

last: Ponsioen

first: Stefan

last: Schulte-Merker

first: Philippe

last: Herbomel

first: Karima

last: Kissa

first: Suma

last: Choorapoikayil


title: Next Generation Sequencing Facilitates Quantitative Analysis of the effects of CD200R engagment on sorted mouse pulmonary ILC2 Transcriptomes.

first: Georges Doumet

last: HELOU

state: California

address: 1450 Biggy St, NRT5505, Keck School of Medicine

department: Molecular Microbiology and Immunology

summary: The prevalence of asthma and airway hyperreactivity (AHR) is increasing at an alarming rate. Group 2 innate lymphoid cells (ILC2s) are copious producers of type 2 cytokines, which leads to AHR and lung inflammation. In this study, we demonstrate murine ILC2s express CD200 receptor (CD200R), and this expression is inducible. We ascertain CD200R engagement inhibits activation, proliferation, and type 2 cytokine production, revealing a potent immunoregulatory role for CD200–CD200R axis on ILC2s. Furthermore, we demonstrate that CD200R engagement inhibits both canonical and non-canonical NF-B signaling pathways in activated ILC2s. Importantly, we herein design both preventative and therapeutic approaches utilizing CD200R engagement on ILC2s, which lead to improved airway resistance, dynamic compliance, and eosinophilia. Finally, our results reveal CD200R is expressed on human ILC2s, and its engagement ameliorates AHR in humanized ILC2 models, further emphasizing the translational applications of our findings for treatment of ILC2-related diseases such as allergic asthma.

overall_design: mRNA profiles of sorted mouse pulmonary ILC2s from WT mice treated with CD200 FC or Isotype control were generated by deep sequencing, in triplicate, using a NextSeq 500 (Illumina) system.

first: P

last: Shafiei-Jahani

first: O

last: Akbari


title: Expression data of EGOT-depleted cells and controls

city: Pamplona

first: Puri

last: Fortes

address: Pio XII 55

institute: CIMA/UNAV

summary:
LncRNA EGOT regulates inflammation and growth
We used microarrays to estudy global changes after EGOT depletion and found many inflammation and growth related factors strongly deregulated.

overall_design: HuH7 cells were transfected with a plasmid expressing PKR and poly I:C to induce EGOT levels. In addition, 50 pmols of a gapmer targeting EGOT or a control gapmer were also transfected with lipofectamine 2000. RNA was isolated 36 hours post-transfection and RNA quality was evaluated before samples were hybridized to a Affymetrix Human Gene 32.0 ST array

first: Marina

last: Barriocanal


title: Irx3 and Irx5 regulate hypothalamic postnatal neurogenesis and leptin sensitivity (10X Genomics scRNA-seq)

first: Chi-chung

last: Hui

summary: Single cell sequencing of Ins2-Cre;ptdTomato+ FACS sorted cells from micro-dissected hypothalamic arcuate nucleus and median eminence of wildtype and Irx3/5 double heterozygous mice.

overall_design: Single-cell RNA sequencing (10X Genomics) was performed on tdTomato+ FACS sorted cells from the arcuate nucleus and median eminence of Ins2-Cre;tdTomato and Ins2-Cre;tdTomato;Irx3/5dHet (double-heterozygous) mice at postnatal day 10. For each genotype, micro-dissected tissues from 5 mice were pooled for single-cell dissociation, followed by FACS sorting and 10X single-cell RNA sequencing.

first: Joe-Eun


title: Identification and Functional Analysis of Long Non-coding RNAs in Human Pulmonary Microvascular Endothelial Cells Subjected to Cyclic Stretch

city: Ji'nan

first: wang

last: dong

state: Shandong

address: 44 Wenhua Xi Road

institute: shandong university

summary: To screen the differential expression of mRNAs and lncRNAs in Human pulmonary microvascular endothelial cells (HPMECs) subjected to cyclic stretch, we performed transcriptome profiling using Affymetrix Human Transcriptome Array 2.0.

overall_design: HPMECs were plated on Flexcell plates coated in collagen IV. The cells were subjected to 20% cyclic stretch for 0 or 2 hr (n = 3 per group). Total RNA was isolated and transcriptome profiling was done to identify differentially expressed lncRNAs and mRNAs which occur secondary  to cyclic stretch.

first: Dong

last: Wang

first: Yuelan


title: Yap regulates fatty acid oxidation in skeletal muscle [RNA-seq]

first: Enzo

last: Porrello

summary: The Hippo signalling pathway effector Yap positively regulates adult skeletal muscle mass. However, the biological processes that are modulated by Yap in skeletal muscle remain elusive. Using an integrated transcriptomics and proteomics approach, we define the transcriptional programme regulated by Yap in adult skeletal muscle and demonstrate that Yap is a regulator of metabolic substrate utilisation. Inhibition of Yap in mammalian skeletal muscle results in increased, but incomplete, oxidation of fatty acids and features of lipotoxicity. In line with these findings, we demonstrate that Yap abundance is reduced in the skeletal musculature of obese db/db mice, and in muscle biopsies from obese, insulin-resistant humans where YAP levels positively correlate with whole-body metabolic flexibility. Increasing Yap abundance in the striated muscle of db/db mice attenuated the accumulation of fat mass and development of hepatic steatosis. Our findings demonstrate a vital role for Yap in skeletal muscle as a mediator of metabolic substrate utilisation. Modulating Yap activity in skeletal muscle warrants consideration as part of comprehensive approaches to treat metabolic disease.

overall_design: Transcriptional analysis of cardiomyocytes from mouse strain treated with 2 differently adeno-associated viral vector.

first: Paul

last: Gregorevic

first: Choon

last: Sim


title: Next Generation Sequencing Facilitates Quantitative Analysis of Wild Type and Sirt6-/- mTEC Transcriptomes

city: Beijing

first: Jiayu

last: Zhang

address: Beichen West Road 1-5

department: Institute of Zoology

institute: Chinese Academy of Sciences

summary:
Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare NGS-derived retinal transcriptome profiling (RNA-seq) to microarray and quantitative reverse transcription polymerase chain reaction (qRT–PCR) methods and to evaluate protocols for optimal high-throughput data analysis
Methods:mTECl mRNA profiles of 4 weeks old wild-type (WT) and sirtuin 6 knockout (Sirt6-/-) mice were generated by deep sequencing  using Illumina HiSeq 3000.
Conclusions: Our study revealed that the expression of epigenetic regulator Sirt6 in TECs is crucial for the development and differentiation of mTECs and for the induction of central immune tolerance. Sirt6 deficiency leads to reduced DNA replication and impaired proliferation of mTECs, and specifically accelerates the differentiation of mTECs from CD80-Aire- immature population to CD80+Aire- intermediate mature population by promoting the expression of Spib.

overall_design: mTEC  mRNA profiles of 4 weeks old wild type (WT) and Sirt6-/- mice

first: Qian

first: Zhanfeng

last: Liang


title: MYC-mediated ribosomal gene expression sensitizes enzalutamide-resistant cells to EP300/CREBBP inhibitors

city: Innsbruck

first: Tobias

last: Furlan

state: Tirol

address: Anichstraße 35

department: Urology

country: Austria

institute: Medical University Innsbruck

summary: Various mechanisms have been reported to be responsible for enzalutamide resistance in prostate cancer. In our previous studies, we have demonstrated that the histone acetyltransferase EP300 is highly expressed in castration therapy-resistant prostate cancer. In the present study, we investigated the role of EP300/CREBBP in enzalutamide-resistant prostate cancer. Enzalutamide resistant and control DuCaP cells generatd previously were treated with histone acetyltransferase (C646) and bromodomain (I-CBP112) inhibitors of EP300/CREBBP. Additionally Enzalutamide resistant cells under 5 µM enzalutamide treatment and control LNCaP cells were analyzed. DuCaP and LNCaP cells were seeded in 6-well-plates at 8 x 10^5 and 6 x 10^5 cells per well. The following day, DuCaP cells were treated with 8 µM enzalutamide, 10 µM C646, 10 µM I-CBP112 or DMSO equivalent for 24 hr.

overall_design: Illumina sequencing and examination of differentially expressed genes of parental and resistant cell lines derived from DuCaP and LNCaP. DuCaP cells were also treated with enzalutamide, C646 and I-CBP112.

first: Z

last: Culig

first: T


title: Global analysis of m6A mRNA methylation in human cells

city: Chicago

first: Issam

last: BEN-SAHRA

state: IL

address: 512 North McClurg Ct

department: Biochemistry and Molecular Genetics

institute: Northwestern University

summary: m6A RNA immunoprecipitation was performed according to published procedure (Dominissini et al., 2012). Human m6A-seq data from DTSC2 HeLa cells were aligned to the hg19 transcriptome. To locate m6A peaks, the hg19 transcriptome was divided into 25 nucleotide-wide tiles. The number of reads in the m6A immunoprecipitation (IP) and non-IP (control) sample was counted in each tile, and a p-value was calculated using Fisher’s exact test and adjusted for multiple testing. Tiles with significant enrichment of the m6A signal (adjust-P value < = 0.05) were merged into larger regions. Regions lesser than 100 bp were discarded, and regions over 200 bp were separated into 100 to 200 bp sub-regions; the m6A signal from rapamycin-treated cells compared to control (DMSO) was calculated for each region; and regions with at least a two-fold enrichment in all replicates were identified as m6A peaks. The distributions of m6A peaks and m6A marked genes were identified by overlapping all m6A peaks with the hg19 RefGene annotation (Meng et al., 2014).

overall_design: Examination of m6A mRNA profile in HeLa TSC2 knockout cells treated with vehicle (DMSO) or rapamycin 24h

last: Ben-Sahra

first: Elodie

last: Villa


title: Sensing of mycobacterial arabinogalactan by galectin-9 exacerbates mycobacterial infection by inducing matrix metalloproteinases

city: Shanghai

first: xiangyang

last: wu

address: 507 Zhengmin Road, Shanghai, China

institute: Shanghai Key Lab of Tuberculosis, Shanghai Pulmonary Hospital, Tongji University School of Medicine

summary: Mycobacterial arabinogalactan (AG) is an essential cell wall component of Mycobacteria and a frequent structural and bio-synthetical target for anti-tuberculosis (TB) drug development. Yet, it is unclear whether mycobacterial AG is a pathogen-associated molecular pattern (PAMP) with an elusive pattern recognition receptor (PRR). Here, we report that mycobacterial AG is recognized by galectin-9 and exacerbates mycobacterial infection. Administration of AG-specific aptamers inhibited cellular infiltration caused by Mycobacterium tuberculosis (Mtb) or Mycobacterium bovis BCG, and moderately increased survival of Mtb-infected mice or Mycobacterium marinum-infected zebrafish. AG interacted with carbohydrate recognition domain (CRD) 2 of galectin-9 with high affinity, and galectin-9 associated with transforming growth factor β-activated kinase 1 (TAK1) via CRD2 to trigger subsequent activation of extracellular signal-regulated kinase (ERK) as well as induction of the expression of matrix metalloproteinases (MMPs). Moreover, deletion of galectin-9 or inhibition of MMPs blocked AG-induced pathological impairments in the lung, and the AG-galectin-9 axis aggravated the process of Mtb infection in mice. These results demonstrate that AG is an important virulence factor of mycobacteria and galectin-9 is a novel receptor for Mtb and other mycobacteria, paving the way for the development of novel effective TB immune modulators.

overall_design: Total RNA was isolated and used for RNA-seq analysis. cDNA library construction and sequencing were performed by Beijing Genomics Institute using BGISEQ-500 platform. High-quality reads were aligned to the Mus musculus reference genome (UCSC_mm10) using Bowtie2. The expression levels for each of the genes were normalized to fragments per kilobase of exon model per million mapped reads (FPKM) using RNA-seq by Expectation Maximization (RSEM).

first: Xiangyang

last: Wu


title: QRICH1 Dictates the Outcome of ER Stress through Transcriptional Control of Proteostasis [RNA-seq]

city: Cambridge

first: Kwontae

last: You

state: MASSACHUSETTS

address: 415 main street

institute: Broad Institute

summary: Perturbation of tissue homeostasis accompanies a diversity of inflammatory pathologies and imposes metabolic constraints at the cellular level that elicit ER stress, protein misfolding, and cell death. In response to ER stress, cells initiate the unfolded protein response (UPR), which determines divergent cell fate decisions, either promoting recovery of ER proteostasis and cell survival or triggering programmed cell death. However, the mechanisms by which the UPR transitions from the adaptive to terminal state are not fully understood. Here, we identify QRICH1 as a previously uncharacterized transcriptional regulator of a gene modulethat differentially engages the terminal versus adaptive UPR.

overall_design: To prepare the non-stressed and ER-stressed conditions, we treated HT29 wild type or QRICH1 knockout cells with DMSO or 0.3 ug/ml of Tm for 14 hrs and isolated RNA by TCL buffer (QIAGEN) treatment.  We generated library with three biological replicates.

first: Chih-Hung

last: Chou

first: Ariel

last: Lefkovith

first: Daniel

last: Graham

first: Ramnik

last: Xavier


title: Temporal scRNAseq identifies dynamic transcriptional programs that dictate the outcome of ER stress

summary: Perturbation of tissue homeostasis accompanies a diversity of inflammatory pathologies and imposes metabolic constraints at the cellular level that elicit ER stress, protein misfolding, and cell death. In response to ER stress, cells initiate the unfolded protein response (UPR), which determines divergent cell fate decisions, either promoting recovery of ER proteostasis and cell survival or triggering programmed cell death. However, the mechanisms by which the UPR transitions from the adaptive to terminal state are not fully understood. Here, we leverage single-cell genomics to define dynamic transcriptional states associated with the adaptive versus terminal UPR in the intestinal epithelium. We recovered the terminal-UPR signature, which dictates adaptation versus programmed cell death in response to ER stress.

overall_design: At monolayer differentiation day3 from the intestinal spheroids, treat 0.5ug/ml of Tm (for 25hr sample) or DMSO (for DMSO or 13hr set). 12hrs later, treat Tm on one of the DMSO set to make the 13hr set. Harvest the cells at differentiation day 4 morning. We generated two biological replicates.

first: Lingfei

last: Wang

first: Toru

last: Nakata


title: Genome-wide CRISPR screen for ER stress-UPR pathway

summary: Perturbation of tissue homeostasis accompanies a diversity of inflammatory pathologies and imposes metabolic constraints at the cellular level that elicit ER stress, protein misfolding, and cell death. In response to ER stress, cells initiate the unfolded protein response (UPR), which determines divergent cell fate decisions, either promoting recovery of ER proteostasis and cell survival or triggering programmed cell death. However, the mechanisms by which the UPR transitions from the adaptive to terminal state are not fully understood. To discover novel UPR pathway regulators that influence the outcome of cellular ER stress responses, we performed genome-scale genetic perturbations. Our genome-wide screens revealed that distinct sets of genes regulate UPR activity and maintenance of ER homeostasis. This expansive dataset provides a rich resource that can be leveraged to elucidate signaling crosstalk during ER stress and discover novel UPR regulators.

overall_design:
To measure the UPR activity, we generated an endogenous XBP1s-GFP reporter by CRISPR knock-in. On day 0, we transduced the Genome-wide lentiviral library at an MOI of 0.2 into the 40 million cells of HT29 XBP1s-GFP cells, which infected one virus per cell with over 100 coverage of the library. On day 2, we selected the transduced cells by adding puromycin (Invitrogen) at 5 ug/ml. At day 5, we re-plated the cells with fresh media.   For the XBP1s-GFP screen, we treated DMSO or 0.25 ug/ml of Tm for 13hrs. We filtered (40 μm) the single cells and added DAPI just before the sort to select the live cell fraction. We sorted the cells based on the GFP intensities using FACS, each of high or low bins containing 15% of the cells. We collected about 15 million cells per bin (a representation of over 200 coverage of the library).
There are 3 replicates each for Tm high 15% of bin, Tm low 15% of bin, DMSO high 15% of bin and DMSO low 15% of bin.  All samples were sequenced in two sets of paired-end files. Demultiplexing was performed with poolq. Raw data files are not demultiplexed.  The file sample_index.csv  includes the sample barcodes.  The file count.tsv includes the gene barcode.


title: QRICH1 Dictates the Outcome of ER Stress through Transcriptional Control of Proteostasis

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


title: QRICH1 Dictates the Outcome of ER Stress through Transcriptional Control of Proteostasis [ChIP-seq]

overall_design: To prepare the ER-stressed conditions, we treated HT29 wild type or QRICH1 knockout cells with 0.3 ug/ml of Tm for 14 hrs and performed the chromatin immunoprecipitation. Briefly, we fixed the cells with 1% formaldehyde and quenched with glycine. We prepared the cell lysate and sonicated the lysates using sonifier. We incubated the lysate with anti-QRICH1 antibody, anti-ATF4 antibody, or anti-normal rabbit IgG overnight in the cold room. We generated library with three biological replicates.


title: RNA-seq of different brain tissue areas of Trrap conditional knockout Mus musculus

city: Jena

first: Philipp

last: Koch

address: Beutenbergstraße 11

department: Core Facility Life Science Computing

institute: Leibniz Institute on Aging - Fritz Lipmann Institute

summary: The acetylation levels of histones and other proteins change during aging and have been linked to neurodegeneration. Here we show that deletion of the histone acetyltransferase (HAT) co-factor Trrap specifically impairs the function of the transcription factor Sp1, reduces its stability and causes a decrease in histone acetylation at Sp1 target genes. Modulation of Sp1 function by Trrap acts as a hub regulating multiple processes involved in neuron and neural stem cells function and maintenance including microtubule dynamics and the Wnt signaling pathway. Consistently, Trrap conditional mutants exhibit all hallmarks of neurodegeneration including dendrite retraction and axonal swellings, neuron death, astrogliosis, microglia activation, demyelination and decreased adult neurogenesis. Our results uncovered a novel functional network, essential to prevent neurodegeneration, and involving the specific regulation of Sp1 transcription factor and its downstream targets by Trrap-HAT.

overall_design:
16 samples in total:
8 cortex (4 Trrap Fl/+; Camk2-Cre tg, 4 Trrap Fl/Fl; Camk2-Cre tg),
8 striatum (4 Trrap Fl/+; Camk2-Cre tg, 4 Trrap Fl/Fl; Camk2-Cre tg)

first: Marco

last: Groth

first: David

last: Pellón

first: Alicia

middle: T

last: Soler

first: Zhao-Qi


title: RNA-seq of murine primary adult stem cells of Trrap inducible knockout Mus musculus with and without 4-OHT treatment

overall_design:
15 samples in total:
5 with no treatment (Trrap Fl/+; Rosa26-CreERT2 tg)
5 with 4-OHT treatment (Trrap Fl/+; Rosa26-CreERT2 tg)
5 with 4-OHT treatment (Trrap Fl/Fl; Rosa26-CreERT2 tg)

middle: L


title: Transcriptome of senescent and non-senescent cells within PanIN pancreatic premalignant lesions in a KRas-driven mouse model

city: Jerusalem

first: Ittai

last: Ben-Porath

address: Ein-Kerem

department: Department of Developmental Biology and Cancer Research

country: Israel

institute: The Hebrew University Faculty of Medicine

summary: Cellular senescence is a central barrier to tumorigenesis, acting to block the proliferation of premalignant cells. However, senescent cells residing within tumor lesions can also exert paracrine effects influencing tumor growth and progression. Premalignant pancreatic intraepithelial neoplasia (PanIN) lesions contain senescent cells, yet whether these influence disease progression is unknown. Here we report that senescent cells in PanINs that develop in a Kras-driven mouse model express a pro-inflammatory gene signature, which includes high Cox2 levels. Pharmacologic Cox2 inhibition caused a dramatic reduction in PanIN growth. Senolytic treatment with the Bcl2-family inhibitor ABT-737 reduced the numbers of Cox2-expressing PanIN cells and blocked PanIN formation and progression to carcinoma. These findings indicate that senescent PanIN cells support tumor growth and progression through Cox2 activity, representing crosstalk between interspersed senescent and dividing premalignant cells. Targeted elimination of senescent cells may thus be effective in limiting progression of precancerous lesions.

overall_design: For inducible Kras activation in pancreatic acinar cells, Ptf1a-CreER; Kras+/lsl-G12D; Rosalsl- tdTomato triple transgenic mice, and controls, were injected at 6 weeks of age subcutaneously with two doses of 400 mg/Kg tamoxifen. These mice developed premalignant PanIN lesions and were sacrificed at 3 months of age. We isolated tdTomato+SA-Gal– and tdTomato+SA-Gal high fractions from 3 mice. Total RNA from FACS-sorted cells was isolated by TRIzol (Invitrogen) extraction followed by RNeasy Plus Micro Kit (Qiagen), using >10,000 cells of each sample. Profiling was conducted using an adapted CEL-Seq2 protocol:  3’ cDNA was synthesized and barcoded, followed by RNA synthesis, amplification by in vitro transcription and library generation for paired-end sequencing. Reads were demultiplexed, quality filtered and trimmed for adapters and poly-A tail using Cutadapt and aligned with the human genome (GRCh38) using Tophat2.

first: Dror

last: Kolodkin-Gal


title: Yap regulates fatty acid oxidation in skeletal muscle [ATAC-seq]

first: Enzo

last: Porrello

overall_design: Chomatin landscape analysis of cardiomyocytes from mouse strain treated with 2 differently adeno-associated viral vector.

first: Paul

last: Gregorevic

first: Choon

last: Sim


title: Lipin2 limits macrophage proinflammatory response.

status: Public on Feb 16 2021

city: Sendai

first: Hiroyuki

last: Inuzuka

address: 4-1 Seiryo, Aoba-ku

institute: Tohoku University

summary:
Alterations in Lpin2 encoding the phosphatidate phosphatase Lipin2 cause the autoinflammatory disorder Majeed syndrome.
Lipin2 deficiency enhances lipopolysaccharide (LPS)-induced NLRP3 inflammasome formation in macrophages.

overall_design: To determine the physiological role of Lipin2 at regulating macrophage proinflammatory signaling, we knocked out Lpin2 with CRISPR/Cas9 in murine macrophage RAW-D cells and conducted a microarray expression analysis with LPS-treated or untreated WT and Lpin2 knockout (KO) cells.

first: Asami

last: Watahiki


title: MACF1 promotes osteoblast differentiation by sequestering repressors in cytoplasm

city: Wuhan

first: Dong

last: Chen

state: Hubei

address: 388 GaoXin 2nd Road, East Lake Hi-Tech Development Zone

department: Center for Genome Analysis

institute: ABLife, Inc.

overall_design: Refer to individual Series


title: Histone H3 lysine 4 trimethylation in sperm is transmitted to the embryo and associated with diet-induced phenotypes in the offspring

city: Sainte-Anne-de-Bellevue

first: Sarah

last: Kimmins

address: 21111 Lakeshore Road

institute: McGill University

summary: ChIP-Sequencing for H3K4me3 was performed on the sperm of individual mice fed a folate sufficient or folate deficient diet, and on a wildtype or KDM1A overexpressing transgenic background. Ultra-Low-Input ChIP-Sequencing was performed on 8-cell embryos from male wildtype mice fed a folate sufficient or folate deficient diet. We identified regions of the sperm epigenome that were sensitive to a folate deficient diet and found that multiple epigenetic stressors could cumulatively alter H3K4me3 in sperm. Additionally, we found that alterations in H3K4me3 levels in the sperm of folate deficient males were retained in the pre-implantation embryo. Finally, low-input RNA-Seq on 8-cell embryos from folate sufficient or folate deficient males on a wildtype or KDM1A transgenic background allowed us identify transcripts with deregulated gene expression in the pre-implantation embryo. Regulating promoters with aberrant transcription in the 8-cell embryos overlapped promoters with altered H3K4me3 in sperm.

overall_design: Examination of H3K4me3 in the sperm of wildtype or KDM1A overexpressing transgenic mice fed a folate deficient or folate sufficient diet, examination of H3K4me3 in 8-cell embryos from wildtype males on a folate sufficient or folate deficient diet, and examination of gene expression in 8-cell embryos from folate sufficient or folate deficient males on a wildtype or KDM1A transgenic background.

first: Ariane

last: Lismer


title: IL-22 promotes formation of MUC17 glycocalyx barrier in postnatal small intestine during weaning

city: Gothenburg

first: Thaher

last: Pelaseyed

address: Box 440

department: Medical Biochemistry and Cell Biology

country: Sweden

institute: University of Gothenburg

summary: The intestine is under constant exposure to chemicals, antigens and microorganisms from the external environment. Apical aspects of absorptive epithelial cells form a brush border membrane (BBM), shaped by packed microvilli coated with a dense glycocalyx. We present evidence showing that the glycocalyx forms an epithelial barrier that prevents exogenous molecules and live bacteria from gaining access to BBM. We used a multi-omics approach to investigate function and regulation of membrane mucins exposed on the BBM during postnatal development of the mouse small intestine. Muc17 was identified as a major membrane mucin in the glycocalyx that was specifically upregulated by IL-22 as part of an epithelial defense repertoire during weaning. High levels of IL-22 at time of weaning reprogrammed neonatal postmitotic progenitor enterocytes to differentiate into Muc17-expressing enterocytes, as found in adult intestine during homeostasis. Our findings propose a role for Muc17 in epithelial barrier function in small intestine.

overall_design: This experiment contains 4 biological groups, in 3-4 biological replicates. Samples were whole ileal tissues from mice at postnatal day 14 and 24, and mice treated with recombinant mouse Il22 or PBS via intraperitoneal injection at postnatal days 14 and 15, and sacrificed at postnatal day 16. Ileum samples for mRNA analysis were collected in RNAlater and kept at -80°C until RNA extraction. Total RNA was isolated using RNeasy Plus Mini Kit and DNase Max Kit, followed by storage at -80°C. The quality of isolated RNA was determined using a Bioanalyzer with RIN values greater than 8. cDNA libraries were prepared using the TruSeq Stranded Total RNA Sample Preparation kit with Ribo Zero Gold (Rev. E; Illumina) according to manufacturer´s protocol. Each sample has multiple raw fastq files, corresponding to different sequencing lanes (e.g., L001, L002, etc) and different reads (e.g., R1, R2).

first: Elena

last: Layunta

first: Sofia

last: Jäverfelt

first: Brendan

last: Dolan

first: Liisa

last: Arike


title: PD-1 is imprinted on cytomegalovirus-specific CD4+ T cells and attenuates Th1 cytokine production whilst maintaining cytotoxicity

city: Birmingham

last: Moss

state: West Midlands

address: Vincent Drive

department: Immunology and Immunotherapy

institute: University of Birmingham

summary:
PD-1 is expressed on exhausted T cells in cancer patients but its physiological role remains uncertain. We determined the phenotype, function and transcriptional correlates of PD-1 expression on cytomegalovirus-specific CD4+ T cells during latent infection.
PD-1 expression ranged from 10-85% and remained stable over time within individual donors. This ‘setpoint’ was correlated with viral load at primary infection. PD-1+ CD4+ T cells display strong cytotoxic function but generate low levels of Th1 cytokines which is only partially reversed by PD-1 blockade.
TCR clonotypes showed variable sharing between PD-1+ and PD-1- CMV-specific cells indicating that PD-1 status is defined either during T cell priming or subsequent clonal expansion. Physiological PD-1+ CD4+ T cells therefore display a unique ‘high cytotoxicity-low cytokine’ phenotype and may act to suppress viral reactivation whilst minimizing tissue inflammation.
Improved understanding of the physiological role of PD-1 will help to delineate the mechanisms, and potential reversal, of PD-1+ CD4+ T cell exhaustion in patients with malignant disease.

overall_design: Total RNA profiles of human PD-1 positive and PD-1 negative CMV-specific CD4 T cells

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