title: Wnt/β-catenin activation epigenetically reprograms Regulatory T cells in IBD and progression to dysplasia

city: Chicago

first: Fotini

last: Gounari

state: IL

address: 924 E 27th Street, JFK

department: Medicine

institute: University of Chicago

summary: Previous work indicated that β-catenin/TCF-1 occupy DNA together with Foxp3. Thus, we anticipated that precise mapping of TCF-1 and Foxp3 co-binding in Tregs, could provide a molecular explanation for our phenotypic observations. To address this, we analyzed TCF-1 DNA binding in Tregs through chromatin immunoprecipitation and deep sequencing (ChIPseq) in Foxp3YFP-Cre WT Tregs. Furthermore, we assessed regions of accessible chromatin in Tregs via transposase-accessible chromatin approach and deep sequencing (ATAC-seq). We also performed RNA-seq expression profiling to investigate how the changes in chromatin accessibility impacted transcription in β-catenin high Tregs.

overall_design: mRNA profiles and ATACseq derived from of 21-day old Foxp3Cre-YFP (WT) and Foxp3Cre-YFP Ctnnb1flex3 (CAT) male mice were generated by deep sequencing, in triplicate, using Illumina

first: Jasmin

last: Quandt


title: A modest change in regulation of NOD1 expression has a major impact on inflammation and gastric cancer

status: Public on Oct 01 2020

city: Seattle

first: Naeha

last: Subramanian

state: Wa

address: 401 Terry Ave N

country: USA

institute: Institute for Systems Biology

summary: Altered protein dosage by defects in single genes leads to haploinsufficiencies and  monogenic disorders, but the impact of small changes in gene expression on  multifactorial disease is unknown. Here we show that a persistent small increase in  expression of NOD1, a key innate sensor of bacterial infection, precipitates a large  physiological effect with dramatically altered cellular function associated with  carcinogenesis. Inhibition of miR-15b and miR-16 microRNA function leads to a ~1.2-1.4 fold increase in NOD1 protein concentration, with even slightly greater increases  leading to ligand-independent, switch-like NOD1 activation. miRNA regulation of NOD1 is impaired in gastric cancer with a small increase being associated with greater early disease mortality. Overall, our data show that tight control of NOD1 expression by  miRNA prevents this sensor from exceeding a physiological switching checkpoint that promotes persistent inflammation and lethal cancer progression, and reveal the impact of a single and modest cellular alteration on cancer.

overall_design:
THP-1 cells were stably transduced with teracycline-inducible NOD1 and NLRP4 coding sequence or vector alone using the pSLIK lentiviral sytem (Shin et al., 2006). Cells were plated at a density of 50,000 cells per well in a 96 well plate in a pre-assigned, non-randomized order provided by the RML Genomics Unit, with four replicates per condition. Expression  of NOD1 and NLRP4 was induced by treatment with 1 μg/ml doxycycline (DOX) for 6 h.
Co-first authors; Leah M. Rommereim, Bhaskar Dutta, Ajay S. Akhade

first: Leah

middle: M

last: Rommereim

first: Bhaskar

last: Dutta

first: Ajay

middle: S

last: Akhade

first: Carolyn

last: Hutcheon

first: Nicolas

middle: W

last: Lounsbury

first: Clifford

middle: C

last: Rostomily

first: Iain

middle: D

last: Fraser

first: Ronald

middle: N

last: Germain


title: Using a novel gene signature for predicting the efficacy of the transarterial chemoembolization in patients with hepatocellular carcinoma.

city: Singapore

first: Kam

last: Hui

address: 11 Hospital Drive

country: Singapore

institute: National Cancer Centre Singapore

summary: Comaprative analysis of gene expression datasets of TACE (transcatheter arterial chemoembolization) responders and non-responders HCC patients.

overall_design: RNA extracted from tumor biopsies of HCC patients to be treated with TACE were analyzed using Affymetrix gene arrays.

first: Ming

last: Shi


title: Transcriptional Analyses of citrus host defense response repression by feeding of Diaphorina citri transmitting Candidatus Liberibacter asiaticus at early stages of infection

status: Public on Sep 30 2020

city: Lake Alffred

first: Xu

last: Wei

state: FL

address: 700 Experiment Station Rd. Lake Alfred, FL

institute: University of Florida

type: Expression profiling by high throughput sequencing

summary:
Purpose: We obtained RNA-seq-based differential expression profile of Valencia sweet orange plants challenged against healthy and CLas-infected psyllid infection at 1 dpi and 5 dpi.  The goals of this study are to reveal the interaction between citrus and psyllid/CaLas during the early phase of infection and understand the molecular mechanisms underlying the host-pathogen interactions and the susceptibility of most citrus varieties.
Methods: leaf mRNA profiles of in vitro cultured Valencia sweet orange (VAL) budwood (WT) and of VAL fed by healthy and CLas-infected psyllid were generated by RNA-seq, in triplicate (one sample is duplicate), using Illumina HiSeq platform. The sequence reads that passed quality filters were used for gene expression and DEG detection analysis by EBseq algorithms. qRT–PCR validation was performed using SYBR Green assays
Results: Using the RNA-seq data analysis workflow, we mapped about 136.80M sequence reads per sample to the reference Citrus clementina v1.0 genome and a total of 32,677 genes were detected. The average total mapping of each library was 71.98%. RNA-seq data were validated with qRT–PCR.
Conclusions: Our study obtained the transcriptional profiles of citrus host by feeding of psyllid transmitting Candidatus Liberibacter asiaticus at early stages of infection, with biologic replicates, generated by RNA-seq technology. The RNA-seq data analysis workflows reported here should provide a framework for comparative investigations of expression profiles. We conclude that RNA-seq based transcriptome characterization would expedite genetic network analyses and permit the dissection of complex biologic functions.

overall_design: Leaf mRNA profiles of in vitro cultured Valencia sweet orange (VAL) budwood (WT) and of VAL fed by healthy and CLas-infected psyllid

first: Amany

last: Mira

first: Qibin

last: Yu

first: Fred

last: Gmitter


title: TCM-stimulated mouse spleen

city: Tokyo

first: Sachie

last: Hiratsuka

address: 8-1 Kawada-cho Shinjyuku-ku

country: Japan

institute: Tokyo Women's Medical University

summary: Comparison of RNA expression in mouse spleens between wild-type and ZC3h12d homozygotes.

overall_design: Wild type and ZC3h12d knockout without replicates.


title: miRNA and gene expression profiling in human thyroid carcinomas and non-neoplastic thyroids

city: Milan

first: Matteo

last: Dugo

address: via Olgettina 60

department: Department of Medical Oncology

country: Italy

institute: IRCCS Ospedale San Raffaele

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

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

overall_design: Refer to individual Series


title: lncRNA and mRNA expression profile in colorectal adenoma and cancer

status: Public on Sep 28 2020

city: Beijing

first: Rui

last: Chen

address: No.10 Xitoutiao, You An Men

country: China

institute: Capital medical university

summary: We evaluated the profile of lncRNA and mRNA expression in 6 colorectal adenoma (CRA), 6 colorectal adenoma (CRC) and 6 matched normal mucosa (NOR) using the Exiqon miRCURY lncRNA and mRNA array,7th generation. We found that global dysregulated lncRNA and mRNAs between colorectal lesions and normal mucosa. Our findings implicates that dysregulation of lncRNA and mRNAs may play important role in the carcinogenesis and present therapeutic targets for CRC.

overall_design: Examination of lncRNA and mRNA expression in CRA, CRC and NOR.

first: Xiaobo

last: Li


title: TFEB in HEK293 cells by TET-OFF system (HEK293-PFL-TFEB).

city: Pozzuoli

first: Rossella

last: De Cegli

address: Via Campi Flegrei 34

institute: Tigem

type: Genome binding/occupancy profiling by high throughput sequencing


title: Ribosome profiling in DDX3X degron cell lines and covering variants

city: San Francisco

first: Srivats

last: Venkataramanan

state: California

address: 513 Parnassus Ave, HSW 740

department: Cell and Tissue Biology

institute: University of California, San Francisco

summary: We describe the subset of transcripts that require DDX3 for efficient translation.

overall_design: Ribosome profiling upon auxin-induced degradation of DDX3X, and covered with either wild-type or mutant DDX3X or wild-type DDX3Y was performed from HCT116 cells.

first: Lorenzo

last: Calviello

first: Emanuel

last: Wyler

first: Karol

last: Rogowski

first: Kevin

last: Wilkins

first: Malvika

last: Tejura

first: Bao

last: Thai

first: Markus

last: Landthaler

first: Stephen

last: Floor


title: Transcriptome profile of a stable clone overexpressing TFEB in HEK293 cells by TET-OFF system (HEK293-PFL-TFEB).

summary: In order to identify the transcriptomic effects of TFEB overexpression, we performed a time series RNASeq experiments

overall_design: Total RNA was extracted at time 0 in presence of tetracycline  to evaluate the basal expression level of TFEB and at each time point (from 6 to 90h) from the removal of tetracyclineto evaluate the effect of TFEB overexpression.

first: R

first: M

last: Moretti


title: miRNA and gene expression profiling in human thyroid carcinomas and non-neoplastic thyroids [miRNA]

summary: We performed miRNA and gene expression profiling in a series of 30 thyroid carcinomas and 6 non-neoplastic thyroids.

overall_design: MiRNA and gene expression profiles were established by microarray analysis in a series of 36 snap-frozen tissues using SurePrint G3 Human miRNA 8x60K microarrays (Agilent Technologies) and HumanHT-12 WG-DASL V4.0 R2 expression beadchip (Illumina), respectively. Tissue samples were obtained from Fondazione IRCCS Istituto Nazionale dei Tumori (Milan) and include various thyroid carcinoma histotypes: 20 papillary carcinomas (PTCs) consisting of different histological variants, 7 poorly differentiated thyroid carcinomas (PDTCs) and 3 lymph node metastases derived from PTC.

middle: G

last: Borrello

first: P

last: Romeo

first: E

last: Minna

first: L

last: De Cecco

first: S

last: Pilotti


title: SARS-CoV-2 infection induces transcriptomic alterations specific to activation of innate immunity and cellular distress.

status: Public on Sep 25 2020

city: Hong Kong

first: Honglin

last: Chen

address: Rm 5-19, 5/F, Lab block, 21 Sassoon Road, Pokfulam

department: Department of Microbiology

country: Hong Kong

institute: The University of Hong Kong

summary: In the grips of the 2019-2020 SARS-CoV-2 pandemic, the pathogenisis of the virus has been shown to induce sharp responses from the immune system of infected hosts and can result in severe/fatal pulmonary distress. To understand how the virus can provoke such responses in infected hosts we infected golden hamsters with validated SARS-CoV-2 clinical isolates from 5 patients to understand the dynamic changes and functionality of induced gene expression.

overall_design: An analysis of the transcriptomic landscape induced by SARS-CoV-2 infection in a suitable live model .Lung tissue from SARS-CoV-2 infected hamsters was extracted and total RNA was collected for sequencing.

first: Bobo

last: Mok

first: Conor

last: Cremin


title: Genome-wide chromatin immunoprecipitation sequencing (ChIP-seq) in a monoclonal population of HEK-293 cells stably expressing a Tet-OFF regulatory system (i.e. HEK293-PFL-TFEB).

first: Rossella

last: De Cegli

summary: In order to identify the direct target of TFEB, we performed a ChIP-seq in HEK293-PFL-TFEB cells where, after removal of tetracycline, samples were collected in duplicates at 18, 36 and 90 hours

overall_design: a ChIP-seq in HEK293-PFL-TFEB cells was performed after removal of tetracycline, samples were collected in duplicates at 18, 36 and 90 hours. Two ChIP samples for each timepoint and one immunoglobulin G (IgG) control samples were subjected to Illumina high-throughput sequencing and mapped to the reference human genome hg19


title: Intraindividual Genetic Variation in Human Fetuses with a Preimplantation Origin

city: Tehran

first: Masood

last: Bazrgar

address: Banihashem

country: Iran

institute: Royan Institute

type: Genome variation profiling by genome tiling array

summary: Intraindividual copy number variations (CNVs) happen post-zygotically, however their origin is largely unknown. They might appear either through aging or may resist following the common chromosome instability at the preimplantation stage. To uncover a part of this question we investigated fetal mosaicism and its origin. According to distribution pattern of frequent CNVs in derivatives of different germ layers, their origin is early development including preimplantation, whereas CNVs with low frequency occur in later stages. Both fetuses share CNVs, some in the same tissues and some other in different tissues. Functional analysis showed that altered genes were involved in embryonic development pathways. Each organ inherits CNVs with an unpredictable pattern due to extensive cell mixing/migration in embryonic development. Since we have found frequent intraindividual reciprocal CNVs as events with preimplantation origin in both fetuses, mosaic embryo transfer should be performed with caution because it may increase susceptibility to develop early/late onset diseases with genetic component even though recent reports seems to encourage IVF clinics for mosaic embryo transfer.

overall_design: Two normal fetuses were achieved following therapeutic abortion due to maternal indications on the 19th week of natural pregnancy.  We studied DNA from 22 tissues of each fetus for copy number changes. The reciprocal aberrations, according to comparison of tissues of each fetus, were subsequently validated using qPCR.

first: Hamid

last: Gourabi

first: Anis

last: Karimpour-Fard

first: Parnaz

middle: B

last: Boroujeni

first: Khadije

last: Anisi

first: Bahar

last: Movaghar

first: Mojtaba

middle: R

last: Valojerdi


title: SARS-CoV-2 infection is effectively treated and prevented by EIDD-2801

status: Public on Sep 23 2020

city: Chapel Hill

first: Paul

middle: O

last: Grant

state: NC

address: 3105 Neuoscience Research Building

department: BARC

institute: UNC SOM

summary: All coronaviruses known to have recently emerged as human pathogens probably originated in bats1. Here we use a single experimental platform based on immunodeficient mice implanted with human lung tissue (hereafter, human lung-only mice (LoM)) to demonstrate the efficient in vivo replication of severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome coronavirus (MERS-CoV) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), as well as two endogenous SARS-like bat coronaviruses that show potential for emergence as human pathogens. Virus replication in this model occurs in bona fide human lung tissue and does not require any type of adaptation of the virus or the host. Our results indicate that bats contain endogenous coronaviruses that are capable of direct transmission to humans. Our detailed analysis of in vivo infection with SARS-CoV-2 in human lung tissue from LoM showed a predominant infection of human lung epithelial cells, including type-2 pneumocytes that are present in alveoli and ciliated airway cells. Acute infection with SARS-CoV-2 was highly cytopathic and induced a robust and sustained type-I interferon and inflammatory cytokine and chemokine response. Finally, we evaluated a therapeutic and pre-exposure prophylaxis strategy for SARS-CoV-2 infection. Our results show that therapeutic and prophylactic administration of EIDD-2801?an oral broad-spectrum antiviral agent that is currently in phase II/III clinical trials?markedly inhibited SARS-CoV-2 replication in vivo, and thus has considerable potential for the prevention and treatment of COVID-19.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Human lung-only mice (LoM) were used as an in vivo model to evaluate infection of lung tissue with recombinant coronaviruses SARS-CoV, MERS-CoV, and SARS-CoV-2 as well as full length bat coronaviruses WIV1 and SHC014 (12, 13, 35, 36). Viruses were directly injected into the human lung tissue of LoMs and lung tissue collected either 2, 6, or 14 days post-exposure for virus titer determination and/or analysis by histology, electron microscopy, or RNA-seq.

first: Angela

last: Wahl

first: Ralph

last: Baric

first: Raymond

middle: J

last: Pickles

first: Edward

middle: P

last: Browne

first: Corbin

last: Jones


title: A senescent liver is rejuvenated in a young systemic environment: A study based on comparison of chronological and biological ages

city: Wuhan

first: Ying

last: He

address: 1095 Jiefan Avenue

institute: Institute of Organ Transplantation

summary: Though the liver plays central roles in metabolism, the pivotal biological events throughout its aging are still unknown, resulting in ambiguity as to whether liver senescence could be reversed. To establish a criterion for liver-aging evaluation, dynamic hepatic biological features, including liver function, histology, transcriptional profile and protein expression, were screened in a cohort of Lewis rats at sequential chronological ages from 3 months to 24 months.

first: Shaofa

last: Wang

first: Yuanyuan

last: Zhao

first: Jing

last: Fang

first: Ji

last: Zhang

first: Hui

last: Guo

first: Hua

last: Zhu

first: Tao

last: Tan

first: Jianjie

last: Ma

first: Zhishui

last: Chen

first: Xuefeng

last: Sun

first: Xiangmei

first: Xiaoping

first: Nianqiao

last: Gong


title: RNA-seq of the E14.5 lacrimal gland

status: Public on Sep 22 2020

city: New york

first: Xin

state: New york

department: Ophthalmology

institute: Columbia University

summary: The goal of this study is to find the transcriptional targets downstream of PI3K signaling during lacrimal gland development.  We performed laser capture microdissection of the lacrimal gland epithelial tissue at embryonic day E14.5 from control embryos and mutant embryos containing lacrimal gland-specific deletion of PI3K subunits. After tissue  harvest, RNA was extracted,  conversion to cDNA and amplification was performed by Clontech SMART-seq v4 Ultra low input RNA kit,and cDNA library construction was performed using Nextera XT DNA library preparation kit by core facility at Columbia university prior to RNA sequencing. After preparation of cDNA library, each sample was sequenced using Illumina platform

overall_design: For this experiment, 3 control samples and 6 mutant samples were analyzed.

first: Qian

first: Abdul

last: Hannan


title: Translated Long Non-Coding Ribonucleic Acid ZFAS1 Promotes Cancer Cell Migration by Elevating Reactive Oxygen Species Production in Hepatocellular Carcinoma

city: Guangzhou

first: zhiwei

state: Guangdong

address: NO.1838 Guangzhou Road

institute: Southern medical university

summary: Micropeptides (≤100 amino acids) are essential regulators of physiological and pathological processes, which can be encoded by small open reading frames (smORFs) derived from long non-coding RNAs (lncRNAs). Recently, lncRNA-encoded micropeptides have been shown to have essential roles in tumorigenesis. Since translated smORF identification remains technically challenging, little is known of their pathological functions in cancer. Therefore, we created classifiers to identify translated smORFs derived from lncRNAs based on ribosome-protected fragment sequencing and machine learning methods. In total, 537 putative translated smORFs were identified and the coding potential of five smORFs was experimentally validated via green fluorescent protein-tagged protein generation and mass spectrometry. After analyzing 11 lncRNA expression profiles of seven cancer types, we identified one validated translated lncRNA, ZFAS1, which was significantly up-regulated in hepatocellular carcinoma (HCC). Functional studies revealed that ZFAS1 can promote cancer cell migration by elevating intracellular reactive oxygen species production by inhibiting nicotinamide adenine dinucleotide dehydrogenase expression, indicating that translated ZFAS1 may be an essential oncogene in the progression of HCC. In this study, we systematically identified translated smORFs derived from lncRNAs and explored their potential pathological functions in cancer to improve our comprehensive understanding of the building blocks of living systems

overall_design: Transcriptomeanalyses of ZFAS1 overexpression and si in SK-Hep1 human hepatoma cell lines by deep sequencing. In total transcriptome data set, include two PCDH control, two ZFAS ORF overexpression, two si GFP, and two si ZFAS1

first: ZhiWei

first: Yu

last: Meng

first: XiangMing

last: Zhai

first: Chen

last: Xie

first: Min

last: Li

first: ChunLian

last: Zhou

first: Kun

first: TianCai

last: Liu

first: XueXi

last: Yang

first: YingSong

last: Wu


title: Differential alteration of plant functions by homologous fungal candidate effectors

status: Public on Sep 24 2020

city: Trois-Rivières

first: KarenCristine

last: GonçalvesdosSantos

state: QC

address: 3351 boulevard des Forges

department: Chemistry, biochemistry and physics

country: Canada

institute: Université du Québec à Trois-Rivières

summary: Rust fungi are plant pathogens that cause epidemics which threatens the production of many important plant species, such as wheat, soy, coffee and poplar. Melampsora larici-populina (Mlp) causes the poplar rust and encodes at least 1 184 candidate effectors (CEs), however their functions are poorly known. In this study, we analysed the transcriptome and metabolome of Arabidopsis constitutively expressing CEs of Mlp to discover processes targeted by these fungal proteins. For this purpose, we sequenced the transcriptome and used mass spectrometry to analyse the metabolome of Arabidopsis plants expressing one of 14 selected CEs and of a control line. We found 2 299 deregulated genes in at least one of the 14 transgenic lines. Among the down-regulated genes, the KEGG pathways MAPK signaling pathway and Plant-pathogen interaction were over-represented in six and five of the 14 transgenic lines, respectively. Moreover, the genes down-regulated across the fourteen transgenic lines are related to hormone response and defense. Regarding the metabolome, there were 680 metabolites deregulated across the 14 transgenic lines, with highly unsaturated and phenolic compounds and peptides enriched among down-regulated and up-regulated metabolites, respectively, in almost all transgenic lines. Interestingly, we found that some transgenic lines expressing CEs with no similarity in amino acid sequence had similar patterns of gene and metabolite deregulation, while plants expressing CEs from the same family deregulated different genes and metabolites. Taken together, our results indicate that the sequence of effectors may not be a good predictor of its impact in the plant.

overall_design: RNA was extracted from pooled aerial tissue of 2-week-old soil-grown plants, using three replicates per genotype, with the Plant Total RNA Mini Kit (Geneaid) using RB buffer following manufacturer's protocol. The samples were treated with DNAse, then RNA quality was assessed using agarose gel electrophoresis. QC was performed using a 2100 Bioanalyzer (Agilent) and only samples having an RNA Integrity Number higher than 7 were kept for library preparation. Libraries were generated with the NeoPrep Library Prep System (Illumina) using the TruSeq Stranded mRNA Library Prep kit (Illumina) and 100 ng of total RNA as per the manufacturer's recommendations. The libraries were then sequenced with Illumina HiSeq 4000 Sequencer with paired-end reads of 100 nt at the Genome Quebec Innovation Centre (McGill University, Montreal, Canada).

first: Karen

last: Goncalves dos Santos

first: Gervais

last: Pelletier

first: Armand

last: Séguin

first: François

last: Guillemette

first: Jeffrey

last: Hawkes

first: Isabel

last: Desgagné-Penix

first: Hugo

last: Germain


title: DREAM-seq, an endonucleases-assisted mapping of R-loops

status: Public on Sep 26 2020

city: Espoo

first: Sini

last: Rautio

address: Konemiehentie 2

country: Finland

institute: Aalto University

summary: Genome-wide identification of R-loops almost uniquely relies on antibody based pull-down, a method that shows sequence and structural biases. Here we describe a novel approach – DREAM-seq – centered around the use of endonuclease digestions. Unstimulated and stimulated B cells showed that R-loops are more widespread than anticipated, mostly covering all actively transcribed genes.

overall_design: 6 stimulated B cell samples, including 3 Mock and 3 RNaseH treated samples. 4 unstimulated B cell samples, with 2 Mock and 2 RNaseH treated samples.

first: Benjamin

last: Delatte

first: Anjana

last: Rao


title: The effects of ETFA knockdown on the gene expression in SMMC-7721 human hepatoma cells

first: Jiyan

last: Zhang

state: China

address: 27 Taiping Road, East Building, Rm0749

institute: Institute of Basic Medical Sciences

summary: SMMC-7721 single clones stably expressing control shRNA or ETFA shRNAs 1# and 2# were generated and subjected to microarray.

overall_design: SMMC-7721 human hepatoma cells were transfected with pGPU6/Hygro vectors carrying the non-targeting control shRNA or hETFA shRNAs (GCATGCAAATGATTCCCTAGC, GGAAATATCAGAGTGGCTTGA). 48 hours later, stable clones were selected in 75 μg/ml hygromycin  for approximately 2 months. Then the stable clones were subjected to microarray.

first: Jian


title: Vascular endothelial cells (VECs) and lymphatic endothelial cells (LECs)  of small intestines

status: Public on Sep 21 2020

city: Sapporo

first: Daigo

last: Hashimoto

state: Hokkaido

address: N15W7 Kita-ku

department: Department of Hematology

institute: Hokkaido University

summary: Analysis of vascular endothrial cells and lympatic endotherial cells purified from small intestines

overall_design: Vascular endotherial cells (n=3), lymphatic endotherial cells (n=3) of small intestines

first: Reiki

last: Ogasawara

first: Takanori

last: Teshima


title: Saturating the Neurospora crassa genome for defective in methylation (dim) mutants

city: Colorado Springs

first: Andrew

middle: David

last: Klocko

state: Colorado

address: 278 Centennial Hall, 1420 Austin Bluffs Pkwy

department: Chemistry and Biochemistry

institute: University of Colorado Colorado Springs

summary: Cytosine methylation, a fundamental form of epigenetic regulation, is found in many eukaryotes and plays a significant role in cancer and other diseases. Using the genetically tractable model organism Neurospora crassa, we have identified genes that when mutated, cause the strains to be defective in methylation (dim). The process of DNA methylation in Neurospora has been shown to be dependent on DCDC, a five-member complex that directs the histone methyltransferase DIM-5 to tri-methylate Lysine 9 on histone H3 (H3K9me3).  This mark is recognized by HP1, which directs DIM-2 to methylate DNA. In contrast, the HCHC complex employs HDA-1, CDP-2, HP1, and CHAP to deacetylate that same residue on H3. While we know a good deal about DNA methylation, it is still unclear whether we have identified all the genes involved in the process. Thus, we continued our search for dim mutants, using a selection for reactivation of silenced drug resistance genes. Interestingly, we predominantly identified known dim genes, including dim-5, dim-7, dim-8, dim-9, hpo, chap, cdp-2, and hda-1. Using a Sanger sequencing-based approach, we identified mutations in these known dim genes, presumably responsible for the Dim- phenotype; many mutations were unique point mutants that could compromise protein activity or structure, or impact protein-protein interactions. For mutants in which the gene was not placed into a complementation group, we employed a bulk segregant analysis and whole genome sequencing approach to identify additional mutations in known DNA methylation genes, including hH3 and dim-1, as well as a novel dim mutant: dim-10, a fungal-specific protein that may work with DCDC for H3K9me3 catalysis. Not only will this dim mutant collection be a useful resource to investigate the roles of these dim genes and their protein products in DNA methylation, but the isolation of a novel dim gene by a forward genetics approach provides an exciting avenue of research into how incipient heterochromatin formation is achieved.

overall_design: ChIP-seq: We analyzed Neurospora crassa strains containing alleles of dim-7 with site directed mutants in the N-terminus for redistribution (Jamieson et al., 2015) of the histone mark H3K27me2me3, as well as Neurospora crassa strains containing alleles of dim-10 for the histone mark H3K9me3 by Chromatin Immunoprecipitation. Strains were grown, crosslinked, lysed, the histone marks were immunopurified, and associated DNA was sequenced.

first: Eric

middle: U

last: Selker


title: Transcriptome analysis of wild-type and drug-resistance-induced melanoma cell lines

status: Public on Sep 18 2020

city: New York

first: Pietro

last: Berico

state: NY

address: 550 1st Ave

department: Pathology

institute: NYU Langone Health

summary: Transcriptional co-acticvators inhibition is proving to be a powerful mean against cancer cells that shown a novel hallmarks named transcriptional addiction. This dependecy on co-activars activity lays on their presence in clusters of enhancer regulatory regions called Super-enhancers (SEs) that establish high level of expression of genes important for cancer phisiology such as oncogenes and master transcription factors of the cellular lineage of origin. Among these co-activators in SEs is the kinase CDK7 of the basal transcription factor TFIIH. Here, using a large panel of melanoma cell lines, we observed an overall dependency of melanoma cells in CDK7 activity by inhibiting its function through the drug THZ1. However, by chronically exposing melanoma cells to THZ1, they univocally become resistant, switching phenotype toward a mesenchymal-like state.

overall_design: Total RNA sequencing of wild-type and drug-resistance-induced melanoma cell lines

first: Frederic

last: Coin

first: Irwin

last: Davidson


title: N6-Methyladenosine mediates alternative intron/exon inclusion in the nascent transcriptome

status: Public on Sep 15 2020

city: Oxford

first: Guifeng

last: Wei

address: South Parks Road

department: Department of Biochemistry

country: United Kingdom

institute: University of Oxford

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

overall_design: Refer to individual Series


title: Genome-wide distribution of CDK7 in human cutaneous melanoma

summary: CDK7 is a cycline-dependent kinase part of the basal transcription factor TFIIH involved in the formation of the pre-inithiation complex of RNApol II dependent-genes. Despite it's well known genomic localization at the level of gene's promoter, it has been recently observed from Kwiatkowski et al., (2014) that CDK7 can be found also in large clusters of gene regulatory elements called Super-enhancers (SEs) of both normal and cancer cells. Here we report that CDK7 is resembling such distribution in a human melanoma context, in particular at the level of genes important in melanoma physiology such as the master transcription factors MITF and SOX10 which indeed, following the inhibition of CDK7 with the molecule THZ1,  are rapidly downregulated at very low nanomolar concentration.

overall_design: Chromatin immunoprecipitation of human CDK7


title: Transcriptome sequencing of human fibroatheromas identifies genetic signatures of plaque formation and suggests roles of TREM1 and PIK3CG in atherosclerosis

status: Public on Sep 20 2020

city: Bethesda

first: Shurjo

middle: Kumar

last: Sen

state: MD

address: 10 Center Drive, Room 7N317

institute: NHGRI

summary:
Purpose: The goals of this study are to use RNA-Seq based transcriptome profiling to compare a very early stage of vascular disease (diffuse intimal thickening) with two advanced states of atherosclerosis (calcified and non-calcified fibroatheromas).methods and to evaluate protocols for optimal high-throughput data analysis
Methods: Total RNA was extracted from sets of thawed plaque cryosections using the Qiagen RNeasy Mini kit (Qiagen). RNA-Seq libraries were constructed using the TruSeq Stranded Total RNA with Ribo-Zero Gold kit (Illumina). The pooled libraries were sequenced on a HiSeq2000 (Illumina) using version 3 chemistry and paired-end 125 bp reads. Raw sequence data were processed using RTA version 1.17.21.3 and CASAVA 1.8.2 (Ilumina). Reads were aligned with the STAR aligner, using the hg19 genome assembly and Ensembl transcript database (release 74), which includes both protein-coding genes and long non-coding RNAs. Quality control metrics were calculated and visualized using the QoRTs software package. Gene-level read counts were generated using the algorithm defined by the HTSeq package, and the differential expression analyses were performed via DESeq2.
Results: We identified 1,052 and 1,959 candidate genes associated with non-calcified and calcified plaque formation, respectively. A smaller set of 42 genes were biomarkers specifically for plaque mineralization. Combining GWAS results for carotid atherosclerosis with our RNA-Seq data, we found that four previous genomic candidates (including PIK3CG, the top locus for carotid plaque) were also strongly perturbed at the transcript level. Functionally, we found overwhelming evidence for upregulation of the TREM1 signaling pathway in plaques, with 34 out of 75 genes differentially expressed. We characterized PIK3CG and TREM1 proteins by immunohistochemical stating of human plaques; both were strongly expressed in macrophage-rich areas of advanced lesions but absent in diffuse intimal thickening.
Conclusions: Using a high-resolution technique, we show pervasive changes in the vascular wall transcriptome linked to formation of advanced atherosclerotic plaques, including genes specifically linked to plaque calcification. Using combined DNA, RNA, and protein characterization, we show that two prior disease candidates (PIK3CG and TREM1) are linked integratively to advanced atherosclerosis.

overall_design: Atherosclerotic plaque transcriptome mRNA profiles in an early disease state (diffuse intimal thickening) and two advanced disease states (calcified and non-calcified fibroatheroma) were generated by deep sequencing.

middle: K

first: Leslie

last: Biesecker


title: Rbm15-irCLIP-seq

summary: we performed infrared crosslinking immunoprecipitation followed by sequencing (irCLIP-seq) (Zarnegar et al., 2016) for Rbm15 to directly map its binding sites on RNA. As a principle of concept, the cells were engineered to simultaneously express emGFP-Rbm15 and Xist RNA together, as Rbm15 strongly interacts with Xist A-repeat to deposit the m6A methylation downstream. Cross-linking induced truncation site (CITS or RT stops) is the main signature occurring at our irCLIP-seq datasets. RNA meta-profile plot against normalized transcripts shows that these CITSs reside across the transcript, with 2 main peaks in transcript starts and near the stop-codon regions, in agreement with the RBM15/15B binding profile in human cells (Patil et al., 2016).Motif analysis against CITSs revealed that Rbm15 binding sites prefer U-rich stretches, namely 3 or 4 consecutive Us. This is also true for crosslinking induced mutations (CIMS).

overall_design: We performed the ChrMeRIP-seq and identified 6-10% of m6A peaks are located in the intronic regions. We would like to know whether these intronic m6A peaks are also installed by Rbm15-Mettl3/14 complex, we thus performed the Rbm15 irCLIP-seq to examine this pattern.

first: G

last: Pintacuda

first: N

last: Brockdorff


title: m6A-seq profiling in Mettl3 Ctrl and dTAG13 mESCs [m6Aseq]

summary: dTAG-13 treated METTL3_FKBP12(F36V) is sufficient to functionally deplete m6A methylation, that catalysed by METTL3/14 complex.

overall_design: We would like to confirm the functional m6A loss by dTAG-13 treatment in the Mettl3_FKBP12(F36V) cells. C3 and H5 are two independent clones correspondingly. The Female mouse embryonic stem cells expressing dox-inducible Xist were employed to generate this dTAG line because Xist is heavily methylated by m6A so that it can be used as a marker to test the presence or absence of m6A modification. Our western blot analysis revealed that Mettl3_FKBP12(F36V) is sensitive to dTAG-13 treatment and 2 hours are sufficient to deplete the protein. Therefore, cells are either treated with dTAG-13 or not for 2 hours, and then added the doxycycline to induce Xist expression for another 24 hours. Total RNAs from these treated cells were isolated and fragmented with independent Drosophila total RNAs in parallel.  All the fragmented RNAs were quantified well by Nanodrop and Qubit, then the equal ms of fragmented mESC RNAs (300ug) were mixed with 30ug of Drosophila total RNAs and subjected to standard m6A-seq. Fragmented mESC total RNA were used input-RNA-seq and used for the conventional m6A-seq analysis.

first: M

last: Almeida


title: N6-Methyladenosine mediates alternative intron/exon inclusion in the nascent transcriptome [4sU-seq]

summary: N6-methyladenosines (m6A) are stoichiometrically deposited on exons of nearly one third of RNA Pol II transcriptome, mainly catalysed by METTL3/14 complex. However, neither the intronic methylation pattern and its functional relevance nor the immediate response upon m6A loss have been fully understood. Here we applied MeRIP-seq on mESC nascent transcriptome and thus revealed that approximately 6-10% of m6A peaks occurred at intronic regions, preferentially the conserved and alternative exon/intron part of longer introns, proximately to 5’-splice sites. These intronic m6A deposition correlates with both Rbm15 binding and H3K36me3. Moreover, coupled 4sU-seq with METTL3 dTAG system in mESC, we found that m6A mediates alternative exon/intron inclusive in nascent transcriptome. Intriguingly, m6A self-regulation including writers and readers are early response and partially contributed by alternative splicing changes. Collectively, our study presents a unified model that m6A mediates alternative splicing, and dTAG METTL3 opens an avenue to interrogate the direct response of functional m6A disruption.

overall_design: dTAG (FKBP12F36V) was inserted into Mettl3 C-terminus. Mettl3_FKBP12F36 is very sensitive to dTAG-13 treatment. With this system, we investigated the immediate response of functional m6A loss on nascent transcriptome by 4sU-seq. Cells (2 clones and 3 independent replicates in total) are either treated with dTAG-13 or not for 3 hours, then 4sU was incorporated into the medium for 30min. The 4sU-contaning RNAs were isolated and subjected to sequencing.


title: m6A-seq on chromatin-assoicated RNA (ChrMeRIP-seq) in mouse embryonic stem cell [ChrMeRIP-seq]

overall_design: ChrRNA enriches substantial intronic fraction. Isolation of ChrRNA from E14 mESC and then subject to the standard MeRIP-seq to identify the m6A peaks, then classification.


title: Time-resolved genome-wide pathogenic gene expression analysis of the plant pathogen Xanthomonas oryzae pv. oryzae (Xoo) via RNA-Seq

status: Public on Sep 14 2020

city: Jeonju

first: Jeong-Gu

last: Kim

address: 370 Nongsaengmoung-ro

country: South Korea

institute: Rural development administration

summary: Host-pathogen interactions, in particular during the early stage of infection, are important for the fate of boththe invading pathogen and host. In pathogens, these interactions activate diverse pahogenic signals to cause disease in the host and evade the inevitably activated immune responses of the infected host. In this study, we used in vitro assay system and RNA-seq to analyze the time-resolved genome-wide gene expressions of Xoo evoked by the interactions with rice leaf extracts. RNA-seq analysis was carried out with samples of 3 different time points within 2 hours

overall_design: The time-dependent, 0, 90, and 120 min, transcriptome analysis of genome-wide Xanthomonas oryzae pv. oryzae genes from the pathogenic interactions with rice leaf extract via RNA-seq

first: Seunghwan

first: Lin-Woo

last: Kang


title: Transcriptional Signaling Centers Regulate Erythroid Gene Expression and are Disrupted in Common Variations of Human Red Blood Cell Traits

city: Boston

first: Leonard

last: Zon

state: MA

address: 1 Blackfan Circle

department: Oncology/Hematology

institute: Boston Children's Hospital

summary: Hematopoietic progenitors respond to developmental and environmental cues to differentiate. The stage-specific steps of differentiation are stereotypic for each cell lineage, and are controlled by transcription. Here we investigate how differential genomic binding of signal-responsive and lineage-restricted transcription factors can specify subsequent stages of erythropoiesis. Using a human erythroid differentiation system, we extensively characterized the co-operation of the BMP signaling transcription factor SMAD1 with the erythroid transcription factors GATA2 and GATA1 in a detailed time-course. BMP signaling enhances erythroid differentiation. SMAD1 is co-recruited with GATA factors at stage-specific genes that are required to have high expression in each stage. Additionally, we observed SMAD1-GATA co-enriched regions are within super-enhancers and span accessible chromatin. Co-bound regions harbor cell-type specific transcription factor motifs that change during commitment, in contrast to GATA-alone regions. Our studies demonstrate that signal-responsive factors together with lineage regulators mark genomic “hotspots” that function to regulate stage-specific gene expression.

overall_design: ChIP-Seq for H3K27ac, GATA1, GATA2, and SMAD1, after a 2hr pulse of hrBMP, on progenitor and differentiating human CD34+ cells towards erythrocytes.

first: Avik

last: Choudhuri

first: Eirini

last: Trompouki

first: Brian

last: Abraham

first: William

last: Mallard

first: Min-Lee

last: Yang

first: Alireza

last: Ghamari

first: Kanen

last: Hoi

first: Sonja

last: Boatman

first: Victoria

last: Chan

first: Barbara

last: Hummel

first: Song

first: Asher

last: Lichtig

first: Yi

last: Zhou

first: Teresa

middle: V

last: Bowman

first: Shinichiro

last: Takahashi

first: Alan

last: Cantor

first: Santhi

last: Ganesh

first: John

middle: L

last: Rinn

first: Richard

middle: A

last: Young

middle: I


title: Differential Gene Expression in MDA-MB231 from transduction of cells with shRNA targeting CDK19 and CDK8

status: Public on Sep 13 2020

city: Stanford

first: Robert

middle: Wenchen

last: Hsieh

state: CA

address: 265 Campus Drive

department: Stem Cell Institute

institute: Stanford University

summary:
When CDK19 was initially discovered, it was assumed that CDK19 played a redundant role to CDK8 because they share 84% amino acid sequence similarity. However, biological clues such as CDK19s different chromosomal location and its more limited tissue distribution suggested a role distinct from CDK8
To investigate whether CDK19 and CDK8 had distinct biological functions in Triple-Negative breast cancer, we knocked down CDK19 and CDK8 in MDA-MB231 and examined differences in the resulting gene expression.

overall_design: MDA-MB231 cells were infected with shRNA targeting CDK19, shRNA targeting CDK8, 'empty vector' control shRNA and grown in cell culture conditions for 96 hours. Infected cells were sorted by FACS (RFP+) and used for microarray studies.

middle: W

first: Michael

middle: F

last: Clarke


title: ChIPseq to profile NELFE occupancy in bone marrow-derived macrophages from C57BL/6 mice

city: Beijing

first: Li

last: Yu

address: Medicial Science Building, Tsinghua University

department: The Institution of Immunology

institute: Tsinghua University

summary: We characterized the NELFE  occupancy on gene loci in conditions of unstimuated and LPS stimualtion in BMDMs

overall_design: WT BMDMs were left untreated or stimulated with LPS  for 1 hour. NELFE was performed and the DNA products were subject to ChIPseq

first: Bin

last: Zhang

first: Yingli

last: Shang


title: FKBP4 knockdown effect on the gene expression of H1975 cells

status: Public on Sep 12 2020

city: Jinan

first: Shuai

last: Zong

address: Jingwu Road 324

institute: Shandong Provincial Hospital Affiliated to Shandong University

summary: H1975 cells were transfected with FKBP4 targeting shRNA and the control shRNA, then mRNA were extracted for high-throughput sequencing.

overall_design: shFKBP4 v.s. shNT, in triplicate

first: Yueran

last: Zhao

first: Yu

last: Xia


title: The neuroanatomical ultrastructure and function of a biological ring attractor

city: Ashburn

first: Vivek

last: Jayaraman

state: VA

address: 19700 Helix Drive

institute: Janelia Research Campus, HHMI

summary: Neural representations of head direction (HD) have been discovered in many species. Theoretical work has proposed that the dynamics associated with these representations are generated, maintained, and updated by recurrent network structures called ring attractors. We evaluated this theorized structure-function relationship by performing electron microscopy-based circuit reconstruction and RNA profiling of identified cell types in the HD system of Drosophila melanogaster. We identified motifs that have been hypothesized to maintain the HD representation in darkness, update it when the animal turns, and tether it to visual cues. Functional studies provided support for the proposed roles of individual excitatory or inhibitory circuit elements in shaping activity. We also discovered recurrent connections between neuronal arbors with mixed pre- and post-synaptic specializations. Our results confirm that the Drosophila HD network contains the core components of a ring attractor while also revealing unpredicted structural features that might enhance the network’s computational power.

overall_design: Three different types of sequencing protocols were performed: bulk sequencing for the EPGs, ∆7s, PEN2s, and PEGs, low-cell sequencing for the PEN2s and R4d neurons, and single cell sequencing for the PEN1 and PEN2 lines. Two biological replicates were run for each genetic line for the bulk sequencing as well as for whole brain samples, which served as a control. Two genetic lines were run for the EPGs and ∆7s while one line was run for the PEGs. For the low-cell sequencing, 14 replicates were run for the PEN2s, 9 for the R4d neurons, and 8 for the whole brain material. One genetic line was used for each. The optic lobes were removed from the brain for the low-cell controls, but not for the bulk sequencing. The former is referred to as the central brain (CB) while the latter is referred to as whole brain (WB). For the single-cell sequencing, one genetic line was used for the PEN1s and one was used for the PEN2s. Six 96 well plates were run for the PEN1s and five 96 well plates were run for the PEN2s. Each plate featured two wells with ERCC RNA and two well with whole brain RNA to act as controls.

first: Daniel

last: Turner-Evans

first: Kristopher

middle: T

last: Jensen

first: Saba

last: Ali

first: Tyler

last: Paterson

first: Arlo

last: Sheridan

last: Ray

first: Tanya

last: Wolff

first: Scott

last: Lauritzen

first: Gerald

middle: M

last: Rubin

first: Davi

last: Bock


title: Cytosine methylation dynamics during spermatogenesis and post-testicular sperm maturation

status: Public on Sep 09 2020

city: Cambridge

first: Felix

last: Krueger

address: Granta Park

department: Bioinformatics

institute: Altos Labs

type: Methylation profiling by high throughput sequencing

summary: We carried out whole genome bisulfite sequencing in seven purified sperm populations, including primary spermatocytes, early and late round spermatids, and mature sperm isolated from the caput, corpus, and cauda epididiymis, and from the vas deferens.

overall_design: Testicular sperm populations were isolated by elutriation. Epididymal and vas deferens sperm were obtained by gently squeezing sperm out of the dissected tissue. Sperm were washed several times prior to genomic DNA extraction, and purified gDNA was subject to WGBS.

first: Oliver

last: Rando

first: Carolina

last: Galan

first: Ryan

last: Serra


title: RNA-Sequencing of FACS sorted neurons and octopaminergic cells

status: Public on Sep 11 2020

city: Lausanne

first: Jean-Yves

last: Roignant

address: genopode building

country: Switzerland

institute: University of Lausanne

summary: Longitudinals lacking (lola) is among the most complex genes in Drosophila melanogaster, encoding up to 20 different protein isoforms and acting as a key transcription factor in axonal pathfinding and neural reprograming. To better characterize Lola function we have generated specific mutations in each isoform using the CRISPR/Cas9 system. Our targeted screen allows us to revisit the previously demonstrated roles for few isoforms, to assign known functions to specific isoforms and to reveal a critical role for a specific variant in the octopaminergic pathway. Thus, our comprehensive study expands the repertoire of Lola functions, and demonstrates that the CRISPR/Cas9 approach is a valuable tool to systematically address the role of complex loci in vivo.

overall_design: For the neuronal control, embryos were collected at 25°C for three hours and subsequently developed for 13 hours. For octopaminergic cells, embryos were collected for three hours and developed for further 20 hours. Cells were isolated as previously described (Salman and Perrin, 2011) and FACS sorted. Per replicate, 10 cells were collected in lysis buffer and subjected to library preparation using the Smart-Seq2 Kit for Illumina.

first: Nadja

last: Dinges

first: Nastasja

last: Kreim

first: Violeta

last: Morin


title: Targeted DamID for Lola-O

overall_design: Embryos were collected for two hours at 25°C and subsequently developed for 20 hours, collected and frozen. Genomic DNA was isolated, RNAseA treated and DpnI digested. DamID adaptors were ligated to DpnI digested DNA followed by DpnII digestion PCR amplification, sonication and removal of DamID adaptors.Libraries were generated using the NebNext Ultra II DNA Library Kit. Libraries were sequenced on a NextSeq 500.

first: Tony

middle: D

last: Southall


title: RNA-Sequencing of Wild Type and lola-O mutant embryos

overall_design: Embryos were collected at 25°C for two hours and subsequently developed for 20 hours. Embryos were subsequently transferred into TRIzol reagent (Thermo Fisher Scientific) and RNA was isolated using the manufacturers protocol. RNA was DNase I (NEB) treated according to the manufacturer's protocol and subjected to library preparation using the NEBNext® Ultra™ Directional RNA Library Prep Kit for Illumina®. 1 µg of total RNA was used as starting material for library preparation.


title: Roles of Lola-O

overall_design: Refer to individual Series


title: Human embryonic stem cell-derived organoidal retinoblastoma reveals cancerous origin and therapeutic target

status: Public on Sep 05 2020

city: Wenzhou

first: Zibing

last: Jin

state: Zhejiang

address: No.270, Xueyuan road

department: Division of Ophthalmic Genetics, The Eye Hospital

institute: Laboratory for Stem Cell & Retinal Regeneration, Institute of Stem Cell Research

summary: Retinoblastoma (Rb) is the most prevalent intraocular malignant tumor in children with less than 30% survival rate globally. Rb genetic predisposition has been well defined for decades, whereas its developmental origin and drug agents remain largely unexplored. Here we developed the first organoidal retinoblastoma model derived from human embryonic stem cells (hESCs) with a biallelic knockout (RB1-/-) or mutagenesis (RB1Mut/Mut) of the RB1 gene, which exhibit extreme consistent properties of Rb tumorigenesis, transcriptome, and genome-wide methylation. Using single-cell RNA-seq and immunostaining of this model, we found that developmentally Rb originated from ARR3+ maturing cone precursors and a new cell type of unfolded protein response occurred in the organoidal retinoblastoma. A key PI3K-Akt pathway was aberrantly regulated, and its strong activator SYK (encoding proto-oncogenic spleen tyrosine kinase) was significantly upregulated in the cancerous organoids. Furthermore, we demonstrated that the SYK inhibitors, A502 and B502, have more significant therapeutic response on the early-stage organoidal retinoblastoma compared to four well-established chemotherapeutic drugs in clinic. Collectively, we established a brand-new organoidal retinoblastoma model derived from human ESCs in-a-dish and discovered its cell-of-origin and potential drug agents, shedding light on development and therapeutics of human cancers.

overall_design:
Whole-Genome Bisulfite Sequencing of 120-day old human Rb organoids (hRBOs) and normal human retinal organoids (hROs).
Transcription profiles of human Rb organoids (hRBOs) and normal human retinal organoids (hROs) at different organoid stages were characterized using RNA-Seq sequencing (RNA-seq).
Transcription profiles of human Rb organoids (hRBOs) and normal human retinal organoids (hROs) at day 90 were characterized using single-cell RNA-Seq sequencing (scRNA-seq).

first: Hui

last: Liu

first: Yan

last: Zhang

first: You-You

first: Yan-Ping

last: Li

first: Zi-Qi

last: Hua

first: Fang

last: Han

first: Si-Qian

first: Chang-Jun

first: Kun-Chao

last: Wu

first: Ji-Neng

last: Lv

first: Fulong

last: Yu

first: Qiang

last: Lin

first: Chong

last: Chen

first: Jianzhong

last: Su

first: Zi-Bing


title: Gene expression profiles of primary human gall bladder epithelial cells with and without infection with S.paratyphi WT and ∆cdtB mutant for 2 and 7 days

status: Public on Sep 04 2020

city: Berlin

first: Thomas

last: Meyer

address: Chariteplatz 1

department: Molecular Biology

country: Germany

institute: Max Planck Institute for Infection Biology

summary: We have successfully established organoid culture model for human and murine gall bladder (GB) epithelial cells allowing long-time culture of primary epithelial cells. This model was used to analyze factors influencing stemness and to study infection with Salmonella paratyphi and its toxin Cdtb.

overall_design:
Primary cells were isolated from healthy tissue of surgery specimen taken from gall bladders of patients undergoing cholecystectomy for reasons not related to diseases of the gall bladder,  and cultured as described in the accompanying publication. 2 Replicates of each condition (2D uninfected, 2D + WT S.paratyphi, 2D + deltaCdtB S.paratyphi, early and differentiated organoids) were generated. Total RNA was extracted from samples and hybridized on Agilent-8x60k Human custom arrays as dual-color hybridizations.

Gall bladder epithelium cells were infected with S.paratyphi WT or ∆cdtB mutant for 2 or 7 days and hybridized on Agilent-4x44k Whole Genome Human arrays as single-color hybridizations.

first: Ludovico

last: Sepe

first: Francesco

last: Boccellato

first: Hilmar

last: Berger

first: Kimberly

last: Hartly


title: Expression data from 4 newly diagnosed multiple myeloma patients and 4 healthy subjects

status: Public on Sep 03 2020

city: Zhengzhou

first: Baijun

last: Fang

address: 127 Dongming Road, Jinshui District

institute: Affiliated Tumor Hospital of Zhengzhou University

summary:
It has been found that miRNA is related to the diagnosis and prognosis of multiple myeloma.
In this project, we We used microarrays to detect the expression of miRNA in bone marrow of newly diagnosed multiple myeloma patients and healthy subjects, screened out differentially expressed miRNAs, and studied its role in multiple myeloma.

overall_design: We compared and analyzed the expression of miRNA in bone marrow of newly diagnosed multiple myeloma patients and healthy subjects, and screened out miRNA with large expression difference, and studied its expression level in plasma of newly diagnosed multiple myeloma patients and healthy subjects, as well as its biological role in multiple myeloma cell lines.

first: Pu

last: Xiang


title: Effects of the novel PI3K δ/γ inhibitor RP6530 on global gene expression in Hodgkin Lymphoma cell lines.

status: Public on Sep 07 2020

city: Rozzano

first: Simone

last: Serio

state: Milano

address: Via Alessandro Manzoni, 56

department: Cardiovascular Medicine

institute: Humanitas Research Hospital

summary: Tumor-associated macrophages (TAMs) are involved in the pathogenesis of Hodgkin lymphoma (HL) and correlate with negative prognosis. The phosphatidylinositol 3-kinase (PI3K) mediates tumor and endothelial cell survival, and macrophage activation. As PI3Kδ and PI3Kγ are constitutively activated in HL, we describe RP6530, a novel PI3Kδ/γ inhibitor, in clinical development for HL and NHL. RP6530 exhibits anti-proliferative and cytotoxic activity both in vitro in HL cell lines and in vivo in HL xenograft mouse models. By inhibiting the metabolic regulator Pyruvate Kinase M2 (PKM2), RP6530 downregulates lactic acid metabolism in HL cells switching the activation of macrophages from an immunosuppressive M2-like phenotype to a more inflammatory M1-like state. RP6530 reshapes the tumor microenvironment (TME), through the re-polarization of TAMs into pro-inflammatory macrophages and the inhibition of tumor vasculature. These data demonstrate the central role of PI3K δ/γ inhibition for targeting HL cells and TME, indicating RP6530 as a novel therapeutic opportunity for HL patients.

overall_design: Profiling of the transcriptome of L-540 and KM-H2 Hodgkin lymphoma cell lines after in vitro treatment with the novel PI3Kδ/γ inhibitor RP6530 (100nM and 10uM) for 6 hours and RP6530 (10uM) for 24 hours and subjected to polyA-RNA sequencing using Illumina platform. Two biological replicates were profiled for each cell type and time point.

first: Silvia

last: Locatelli

first: Carmelo

last: Carlo-Stella


title: Transcriptome Analysis of Wild Type and HMGB1 knock-out iSLK BAC16 cells

status: Public on Sep 02 2020

city: Daejeon

first: Yun Hee

last: Kang

address: 77 Gyeryong-ro 771 beon-gil, Jung-gu

department: Eulji Biomedical Science Research Institute

institute: Eulji University School of Mdicine

summary: Purpose: RNA sequencing (RNA-seq) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare NGS-derived 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

overall_design: HMGB1 WT or KO iSLK BAC16 cells transcriptome were analyzed by RNA seq. cells were or were not induced KSHV lytic replication with doxycycline and sodium butyrate.

first: Sukyung

first: Myung-Shin

last: Lee


title: Digenome for off-target analysis: Start codon disruption with CRISPR/Cas9 prevents murine Fuchs' endothelial corneal dystrophy

status: Public on Sep 01 2020

city: Loma Linda

first: Hironori

last: Uehara

address: 11021 Campus st

institute: Loma Linda University

summary: A missense mutation of collagen type VIII alpha 2 chain (COL8A2) gene leads to early onset Fuchs’ endothelial corneal dystrophy (FECD), which can cause blindness through progressive loss of corneal endothelial cells. We established a novel procedure for achieving structural and functional rescue of the post-mitotic corneal endothelium without surgery, using CRISPR/Cas9-based postnatal gene editing in a mouse model of FECD. A single intraocular injection of an adenovirus encoding both the Cas9 gene and guide RNA (Ad-Cas9-Col8a2gRNA) at a titer below the cytotoxic threshold, efficiently knocked down COL8A2 protein expression in corneal endothelial cells, prevented endothelial cell loss, and rescued corneal endothelium pumping function in adult Col8a2 mutant mice. Here, we evaluated the off-target activity of humanized gRNA targeting COL8A2 gene by a modified digenome analysis (Nat Methods 12, 237-243). Briefly, in vitro SpCas9 digested genomic DNA with/without gRNA are compared by deep sequencing. The target sequence is CGTCCACGGACGCCATGCT. In this study, we identified 13 candidate sites with homology to the gRNA and 8 candidate sites without homology to the gRNA.

overall_design: Deep sequencing comparison between genomic DNAs which were digested by SpCas9 with/without gRNA.

first: Bala

last: Ambati


title: Phenotypic manifestation of alpha-synuclein strains from Parkinson’s disease and multiple system atrophy in human dopaminergic neruons

first: Devika

last: Agarwal

address: JR hospital, Headley way

department: The MRC Weatherall Institute of molecular medicine

summary: Although α-synucleinis implicated in the pathogenesis of Parkinson’s disease and related disorders, it remains unclear whether specific conformations or levels of α-synuclein assemblies are toxic and how they cause progressive loss of human dopaminergic neurons. To address this issue, we used iPSC-derived dopaminergic neurons with a-synuclein triplication or controls where endogenous α-synuclein was imprinted into synthetic or disease-relevant conformations. We used α-synuclein fibrils generated de novo or amplified from homogenates of brains affected with Parkinson’s disease (n=3) .We found that a 2.5-fold increase in α-synuclein levels in α-synuclein gene triplication neurons promoted seeded aggregation in a dose and time-dependent fashion, which was associated with a further increase in α-synuclein gene expression.Transcriptomic analysis and isogenic correction of α-synuclein triplication revealed that intraneuronal α-synuclein levels solely and sufficiently explained vulnerability to cell death.

overall_design: RNASeq experiment to compare the SNCA triplication iPSC lines (n=3 or 3 clones) or healthy control lines (n=3) at baseline and after induction of alpha-synuclein aggregation for 2 weeks. This was to investigate whether there are consistent changes upon synuclein aggregation in Triplication lines (clone 1, clone 3, clone 5) or healthy control lines (AH017, OX3, SFC840) when compared to their corresponding baseline (NSC) using the three different aggregation conditions (WTPFF, PDamp, MSAamp). A total of 24 different  human iPSC samples were sequenced on the Novaseq 6000 platform.

first: Benedict

last: Tanudjojo

first: Samiha

last: Shaikh

first: Alexis

last: Fenyi

first: Luc

last: Bousset

first: Christos

last: Zois

first: Roman

last: Fischer

first: David

last: Sims

first: Ronald

last: Melki

first: George

last: Tofaris


title: Comparative gene expression analysis after exposure to 123I-iododeoxyuridine, γ- and α-irradiation

city: Juelich

first: Marcus

last: Unverricht-Yeboah

address: Wilhelm-Johnen-Strasse

department: Department of Safety and Radiation Protection

institute: Forschungszentrum Juelich

summary: Gene expression analysis was carried out in human T-lymphoma Jurkat cells in order to identify candidate genes showing significant gene expression alterations allowing robust discrimination of the Auger emitter 123I, incorporated into the DNA as 123I-iododeoxyuridine (123IUdR), compared to α- and γ-radiation, which may be useful for biodosimetry purposes. Comparative gene expression analysis was performed employing whole human genome DNA microarrays.  The gene expression of all candidate genes was validated by quantitative real-time PCR. 155, 316 and 982 genes were exclusively regulated after exposure to 123IUdR, α-particles and γ-rays, at equi-effect doses/activity, respectively. Applying the stringent requirements for candidate genes, four , one  and one gene(s) were identified allowing a reliable discrimination between γ- versus 123IUdR exposure, γ- versus α-radiation and α- versus 123IUdR exposure, respectively. Gene expression analysis might be an effective tool for the general discrimination of radiation qualities and might help to elucidate different biological effectiveness on the mechanistic level.

overall_design: The formation of γ-H2AX foci after exposure to different radiation qualities was analyzed to determine equi-effect doses/activities and consequently to compare gene expression on a similar DNA damage effect level. A similar DNA damage effect level was found after exposure to 10 Gy γ-rays, 1 Gy α-particles and after 20 h exposure to 123IUdR, which corresponded roughly to 2600 accumulated 123I decays per cell. Genes with a fold-change > 1.5 and a FDR < 0.15 were used for the functionally categorization  and the venn diagram. A candidate gene allowing the discrimination of different radiation qualities had to show a significant expression fold change of > 1.5 fold  after exposure to a specific radiation quality and display no altered (1-fold ± 0.1) or even opposing (>1.1-fold) regulation in response to the other radiation qualities. Concomitantly, we used P values, which were not adjusted using the FDR, for the analysis of the significance in order to consider more genes. However, this loss of statistical stringency was justified because the gene expression level of the candidate genes were verified and validated by quantitative real-time PCR .

first: Ulrich

last: Giesen

first: Ralf

last: Kriehuber


title: Mapping the landscape of chromatin dynamics during naïve CD4+ T-cell activation.

city: NEDLANDS

last: Martino

state: Western Australia

address: Perth Childrens Hospital, 15 Hospital Avenue

country: Australia

institute: Telethon Kids Institute

summary: T-cell activation induces context-specific gene expression programs that promote energy generation and biosynthesis, progression through the cell cycle and ultimately cell differentiation. The aim of this study was to apply the omni ATAC-seq method to characterize the landscape of chromatin changes induced by T-cell activation in mature naïve CD4+ T-cells. Using a well-established ex vivo protocol of canonical T-cell receptor signaling, we generated genome-wide chromatin maps of naïve T-cells from pediatric donors in quiescent or recently activated states. We identified thousands of individual chromatin accessibility peaks that are associated with T-cell activation. The majority of these were localized to intronic and intergenic enhancer regions, marked by active histone modifications whilst quiescence was maintained by repressive histone marks. Regions of activation-associated gains in chromatin accessibility were enriched for well-known pioneer transcription factor motifs, and super-enhancer regions associated with distinct gene regulatory networks. These cis-regulatory elements together brought about distinct transcriptional signatures in activated cells including TNFa-NFkB signaling, hormone-responsive genes, inflammatory response genes and IL2-STAT5 signaling. Our data provides novel insights into the chromatin dynamics and motif usage of T-cell receptor signaling events in early life. The characterized pathways demonstrate the utility of chromatin profiling techniques applied to bio-banked samples for characterizing gene regulatory elements.

overall_design: ATAC-seq profiles from naïve CD4+T-cells from six healthy 12-month old infant donors


title: A Safety and Efficacy Study of Concomitant Administration of ChAd63/MVA ME-TRAP + RTS,S

city: Oxford

first: Julius

last: Muller

state: Oxfordshire

address: ORCRB, Roosevelt Drive

department: Nuffield Department of Medicine

institute: The Jenner Institute, University of Oxford

summary: This clinical trial (NCT02252640) assesses the safety and immune responses to vaccination with experimental malaria vaccine candidates RTS,S/AS01, ChAd63 ME-TRAP and MVA ME-TRAP in different combinations. All healthy volunteers were recruited in England and administered the experimental malaria vaccines. Furthermore, volunteers were infected with malaria by mosquito bites, 12 weeks after the first vaccination to assess vaccine efficacy.

overall_design: Volunteers in group G1 received vaccination with the malaria vaccine candidate, RTS,S/AS01 at a standard dose (50mg) at Week 0, 4 and 8.  G2 received the same regimen, but only a fractional dose of 10mg RTS,S/AS01 at Week 8. G3 received the same regimen as G1 and in addition ChAd63 ME-TRAP at Week 0 and MVA ME-TRAP at Week 4 and 8. G4 received the same regimen as G2 and in addition ChAd63 ME-TRAP at Week 0 and MVA ME-TRAP at Week 4 and 8.


title: Start codon disruption with CRISPR/Cas9 prevents murine Fuchs' endothelial corneal dystrophy

last: Uehara

state: California

department: Ophthalmology

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


title: A Safety and Efficacy Study of ChAd63/MVA METRAP + RTS,S

summary: The goal of this phase 1/2a sporozoite challenge trial (NCT01883609) was to evaluate novel malaria vaccination regimens of the GSK pre-erythrocytic RTS,S/AS01B vaccine alone and concomitantly same-site administered with the viral vectors ChAd63 & MVA encoding the liver stage antigen construct ME-TRAP. Four vaccine groups were studied and received three vaccinations at a monthly interval. All subjects then underwent controlled human malaria infection (CHMI) 11 weeks after first vaccine administration.

overall_design: Volunteers in group RTSS received vaccination with the malaria vaccine candidate, RTS,S/AS01. The vaccine schedule consisted of 3 doses of RTS,S/AS01 with an interval of 4 weeks between doses (Doses given at 0,4 and 8 week timepoints). The RTSSplus group received a vaccination schedule composed of the same dosage and timing regimen of RTS,S, but they also received vaccination with ChAd63 ME-TRAP, 2 weeks after the first RTS,S followed 8 weeks later by vaccination with MVA ME-TRAP (2 and 10 week timepoints).


title: Reprogramming the endogenous type III-A CRISPR-Cas system for genome editing and RNA interference in Mycobacterium tuberculosis

status: Public on Aug 30 2020

city: Wuhan

first: Weize

last: Xu

state: Hubei

address: shizishan street No.1

institute: Huazhong Agricultural University

summary:
Purpose: Understanding the functional genomics of M.tb (Mycobacterium tuberculosis) and development of novel anti-M.tb drugs and vaccines needs an efficient gene edit tool. The aim of this study was to describe an easy and efficient gene edit tool for functional genomics study of M.tb.
Method: A plasmid was designed containing mini CRISPR array and 400bp up and downside homologues DNA sequences from the target gene interposed by EGFP or BFP. This plasmid was transformed into M.tb that guided the endogenous CRISPR system of M.tb to cut the target gene and insert EGFP or BFP through homologues ends joining. The EGFP/BFP insertion was confirmed and the total genomic was extracted from mutated and wild type strains and subjected to High-throughput DNA sequencing.
Results:  The raw data was filtered by Trimmomatic and the clean reads were mapped to M.tb H37Ra reference genome with bwa 99.94%, and 99.97% genome of wag31 and esxQ deletion strains were covered respectively.

overall_design: DNA library for wild type, H37Ra Δwag and H37Ra ΔesxQ was constructed in triplicate usning Nextera Illumina kit and were subjected to high-throughput sequencing.

first: Muhammad

last: Jamal

first: Khaista

last: Rahman


title: Indel rate analysis:Start codon disruption with CRISPR/Cas9 prevents murine Fuchs' endothelial corneal dystrophy

first: Hironori

summary: A missense mutation of collagen type VIII alpha 2 chain (COL8A2) gene leads to early onset Fuchs’ endothelial corneal dystrophy (FECD), which can cause blindness through progressive loss of corneal endothelial cells. We established a novel procedure for achieving structural and functional rescue of the post-mitotic corneal endothelium without surgery, using CRISPR/Cas9-based postnatal gene editing in a mouse model of FECD. A single intraocular injection of an adenovirus encoding both the Cas9 gene and guide RNA (Ad-Cas9-Col8a2gRNA) at a titer below the cytotoxic threshold, efficiently knocked down COL8A2 protein expression in corneal endothelial cells, prevented endothelial cell loss, and rescued corneal endothelium pumping function in adult Col8a2 mutant mice. Here, to determine the indel rate in mouse corneal endothelium, we performed deep sequencing of PCR products (including the target site) amplified from genomic DNA of corneal endothelium. We found that the indel rate was 23.7 ± 4.5% in mouse corneal endothelium. Most insertions were 1bp insertions (19.8 ± 4.0% in total reads), while 2bp deletions were the most frequent (1.0 ± 0.3% in total reads). We moreover found that A or T insertion was predominant, with the proportion of A:T:G:C being 48.7 : 44.6 : 1.8 : 4.9.

overall_design: gRNA target site was amplified by PCR, and indel rate was determined by deep sequencing

first: Bala

last: Ambati


title: Expression data from human breast cancer tissue

first: Xiaochen

last: Yu

address: Fengtai District, Beijing Fengtai East Street on the 20th

institute: Institute of Biotechnology

summary:
Although cell-in-cell structure (CICs) is often observed in human tumors and is likely to be associated with tumor progression, it is unclear whether CICs  indicate different malignant stages and/or inflammatory conditions. In the case of human breast cancer, the frequency of CICs was significantly different from one patient sample to another, which may indicate different malignant stages and/or inflammatory conditions. Therefore, we used microarrays to identified classes of differential genes and to clarify the correlation of CICs with clinical prognosis in future researches.
Patient samples were divided into two groups, low CICs formation rate (L-CICs) and high CICs formation rate (H-CICs), and used microarrays to detail the global programme of gene expression underlying cellularisation and identified classes of differential genes during this process.

overall_design: Breast tumor tissue samples were collected from patients at the Military 307 Hospital, Beijing and selected according to low and high CICs formation rate for RNA extraction and hybridization on Affymetrix microarrays. We had attained 7 samples (1-7) for L-CICs and 7 samples (1-7) for H-CICs.

first: Qiang

last: Sun

first: Jianqing

last: Liang

first: Zubiao

last: Niu


title: Stable recombinant MCF7 human breast cancer cell lines with inducible expression of CFP (NM_002621.2, RecCFP) or a mutated CFP variant (S27X, RecCFPm) vs. recombinant MCF7 control recombinants expressing an empty expression vector (RecCtrl)

status: Public on Aug 28 2020

city: Odense

first: Ines

last: Block

address: J.B. Winsløws Vej 4

department: Department of Clinical Genetics

country: Denmark

institute: Odense University Hospital

summary: Total RNA expression profiling of recombinant MCF7 breast cancer cell lines with inducible gene expression (pRAPtar-1i expression vector). Profiles after expression induction were analyzed for of a pool of 2 stably transfected RecCFP recombinant cell lines; 2 RecCFPm recombinant cell lines and a pool of 6 control cell lines (RecCtrl).

overall_design: Three-condition experiment (3 Phenotypes): RecCtrl vs. RecCFP recombinant; RecCtrl vs. RecCFPm. Biological replicates: 4 RecCtrl replicates, 4 RecCFP and 4 RecCFPm replicates.

first: Carolin

last: Müller

first: Mads

last: Thomassen

first: Torben

last: Kruse

first: Markus

last: List

first: Jan

last: Mollenhauer


title: Very preterm delivery is associated with microbial signatures in maternal feces and blood and inflammatory activation in the fetus

status: Public on Aug 31 2020

city: Helsinki

first: Sanna

last: Edelman

address: Meilahdentie 2

institute: Helsinki University Central Hospital

summary: Preterm birth is a major cause of infant mortality and morbidity. The rate of preterm birth is 5-9% in most developed countries and 12% in the United States with approximately 15 million children born prematurely each year. Globally, prematurity is the second most common cause of death in children under the age of five years. To characterize the transcriptomic changes between preterm and full term neonates, RNA-sequencing was applied altogether to 14 cord blood samples. The analysis of the RNA-sequencing data revealed that the spontaneous preterm delivery resulted in systemic inflammatory responses at the gene and  pathway level in the fetus.

overall_design: RNAseq gene expression of CD4+ T cells of very preterm, preterm, and full-term neonates compared to fullterm babies

first: Samuli

last: Rautava

first: Erika

last: Isolauri

first: Raakel

last: Luoto

first: Matti

last: Kankainen


title: Regulation of retrotransposon activity and telomeres in primed pluripotent stem cells (ChIP-seq)

status: Public on Aug 22 2020

city: Tainjin

first: Weiyu

last: Zhang

address: No.38 Tongyan Road, Jinnan District

institute: Nankai University

summary: Naïve and primed pluripotent states represent two different states of pluripotency. Mouse naïve pluripotent stem cells (PSCs) exhibit germline competence as determined by chimera production test and can generate all-PSC mice by tetraploid embryo complementation (TEC) test, the most stringent functional test of developmental potency. By contrast, primed PSCs fail in germline chimeras and TEC test. Unfortunately, these tests cannot be applied to characterization of developmental pluripotency of human PSCs due to ethic issue. Extensive studies demonstrated that naïve and primed pluripotent state exhibits clear epigenome differences. Here we uncover surprising differences in telomere maintenance, retrotransposon activity and genomic stability between these two pluripotent states. Although both states express high telomerase activity, naïve PSCs show robust telomere elongation capacity, associated with higher telomere recombination, consistent with sporadic expression of two-cell (2C) genes including Zscan4. Distinctively, primed PSCs only can maintain their telomere length but without elongation, in association with repression of 2C genes, and increased telomere fragility and telomeric DNA damage with passages. RNA-seq revealed that DNA repair and especially recombination repair pathways are down-regulated in primed state compared to naïve state cells, corroborating the robust DNA repair capacity and genome stability in naïve PSCs. These data suggest that vigorous telomere elongation of naïve state may act to minimize DNA damage to the genome. Furthermore, we identified LINE1 family integrant L1Md_T as naïve-specific retrotransposon and ERVK family integrant IAPEz-int to define primed PSCs, distinguishing the two pluripotent states. Heterochromatin modifications and Dnmt3b differentially regulate transcription of the 2C genes and retrotransposons. Notably, aberrant expression of retrotransposons links to increased genomic stability of primed PSCs. Hence, our data reveals that telomere regulation and retrotransposon activity markedly distinguishes naïve from primed pluripotent state. These new criteria may facilitate induction of high quality and additional characterization of developmental pluripotency and scrutinizing the genomic stability of human naïve PSCs for regenerative medicine

overall_design: We cultured naive and primed mESCs for fifteen passages under naïve and primed conditions to test their gene expression, telomere analysis, transposon elements transcription, genomic stability ,etc.

first: Haifeng

last: Fu

first: Niannian

last: Li

first: Lin

last: Liu


title: The role of mesenchymal stromal cells (MSC) in the development of acute myeloid leukemia

city: Essen

first: Ludger

last: Klein-Hitpass

address: Virchowstr. 173

department: Universitaetsklinikum

institute: Institut fuer Zellbiologie

summary: Acute myeloid leukemia (AML) is a clonal disorder of hematopoietic stem cells with a poor prognosis. It has been reported in many tumors that mesenchymal stem/ stromal cells (MSCs) support the growth of the malignant cells. In this study we analyzed the interaction between MSCs and AML cells in patients and in murine models of human AML. We found that MSCs from AML patients or leukemic mice supported the growth of AML cells in vitro better than MSCs originating from healthy mice or control persons. Interestingly, MSCs from AML patients who achieved complete remission didn’t support the growth of leukemia cells  in vitro anymore.   Due to the difficulty to obtain sufficient human MSCs, we developed a novel method to immortalize human MSCs. Immortalized MSC cells derived from AML patients also supported the growth of human AML cells better than MSC cells derived from a healthy donor. Among others, AMSCs increased entering of leukemic cells into cell cycle and at the same time protected the leukemia cells against exogenous toxic events such as chemotherapy or irradiation. In vivo, we observed an increase in the number of MSCs in the bone marrow of leukemic mice compared to healthy mice. Microarray data of AMSCs show upregulation of epithelial mesenchymal trasition genes. The polarization of MSCs towards an AML-supporting state depends on upregulated expression of the transcription factor Growth factor independence 1 (Gfi1). Loss of Gfi1 abrogated the tumor-supporting state of AML-associated MSCs. In summary, we report that MSCs support the growth of AML cells in a Gfi1-dependent manner.

overall_design: 3 mice which are KP8201, KP8203 and KP8966, were transplanted with BM transduced with MLL-AF9 while 3 mice which are KP1820, KP8816 and TU1777, were transplanted with normal BM cells. When the mice became sick, they were killed and the MSCs from the crushed bones were sorted for RNA study.

first: Yahya

middle: S

last: Al-Matary

first: Cyrus

last: Khandanpour

first: Lother

last: Vassan


title: Regulation of retrotransposon activity and telomeres in primed pluripotent stem cells

status: Public on Aug 23 2020

overall_design: Refer to individual Series


title: Genome-wide maps of MBD3 binding sites in HEK393T cell

status: Public on Aug 20 2020

city: shanghai

first: yin

middle: Chuan

address: 320 Yue-yang Road

institute: Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences.

summary: In order to detect MBD3 binding sites in human cells,HEK 293T cells were subjected to ChIP using MBD3 antibody, and the binding genomic DNA were sequenced and peaks were picked.

overall_design: Examination of MBD3 binding sites in HEK293T cell

first: Chuanyin

first: Ronggui

last: Hu


title: The mRNA alteration in HT22 cells induced by the overexpression of long noncoding RNA AK046375

status: Public on Aug 25 2020

city: Chongqing

first: Zhong

last: Jianjun

address: NO.1 Youyi Road, Yuzhong District, Chongqing

department: Neurosurgery Department

institute: The first Affiliated Hospital of Chongqing Medical University

summary: We overexpressed the lncRNA-AK046375 by lentivirus in HT22 cells, which were immortalized from the mice hippocampal neurons. Then we measured the mRNA alteration induced by AK046375 overexpression through the next generation sequencing technique (RNA-seq). A total of 20,319 transcripts were detected in the current sequencing study, in which 1342 transcripts were selected out according to the criteria: |log2(fold change)| > 1, P < 0.05 (717 up-regulated and 625 down-regulated). To explore whether the mRNA change induced by AK046375 overexpression in a stressed culture condition was different from that in normal condition, the cells were dealt with oxygen-glucose-deprived(OGD) and followed by RNA-seq. A total of 22,016 transcripts were detected, in which 87 transcripts were significantly up-regulated and 131 transcripts were significantly down-regulated according to the aforementioned filter rules.

overall_design: Firstly, HT22 cells were infected with lentivirus to overexpress AK046375. Secondly, cells were cultured in either normal or OGD condition. Finally, cells were collected to extract total RNA and subjected to next generation RNA sequencing analysis.

first: Jianjun

last: Zhong

first: Xiaochuan

last: Sun


title: Richter transfromation of chronic lymphocytic leukemia driven by Epstein-Barr virus reactivation during therapy-realted immunosuppression

status: Public on Aug 29 2020

city: Pamplona

first: Victor

last: Segura

state: Navarra

address: Avda. Pío XII, 55

department: Unit of Proteomics, Genomics and Bioinformatics

country: Spain

institute: FIMA

summary: Gene expression analysis in Epstein-Barr virus negative DLBCL samples obtained from patients with Richter syndrome, lymphocytes from CLL patients and epstein-Barr virus positive DLBCL cell lines derived after CLL tumors xenotransplantation

overall_design: Gene expression profiling was conducted in EBV+DLBCL xenograft derived cell lines, in tumor samples from patients with EBV- DLBCL (Richter syndrome) and non-transformed CLL samples using Affymetrix Human Gene ST 1.0 microarrays..

first: Jose

last: Martinez Climent

first: Marta

last: Larrayoz


title: RNA-Seq analysis was performed with total RNA isolated from Col-0 and mee45-1mutant siliques at 4 DAP

city: Tai'an

first: Yingju

address: Daizong Street

institute: Shandong Agricultural University

summary: To establish the regulatory roles of MEE45 in ovule development and seed size control, we identified the regulated genes whose transcriptional activities are associated with relative abundance of MEE45 transcripts by transcriptomic analysis.  Up-regulated genes in mee45-1 were mainly responded to auxin . Down-regulated genes in mee45-1 were predicted to be associated with hormone metabolic process, seed development, fatty acid biosynthetic process and environmental responses

overall_design: Three biological repeats of the Arabidopsis siliques from mee45-1(mee45-1, mee45-2,mee45-3) and Col-0 (1 Col-1,  Col-2,  Col-3) at 4 DAP were used for RNA sequencing

first: Ying

middle: H

last: Su

middle: J

first: Yang

last: Yu


title: Orphan CpG islands boost the regulatory activity of poised enhancers and dictate the responsiveness of their target genes

city: Santander

first: Alvaro

last: Rada-Iglesias

state: Cantabria

address: Albert Einstein 22, PCTCAN

institute: Institute of Biomedicine and Biotechnology of Cantabria (IBBTEC), CSIC/University of Cantabria

summary: CpG islands (CGIs) represent a unique and widespread genetic feature of vertebrate genomes, being associated with ~70% of all annotated gene promoters. CGIs have been proposed to control transcription initiation by conferring nearby promoters with unique chromatin properties. In addition, there are thousands of distal or orphan CGIs (oCGIs) whose functional relevance and mechanism of action are barely known. Here we show that oCGIs are an essential component of poised enhancers (PEs) that boost their long-range regulatory activity and dictate the responsiveness of their target genes. Using a CRISPR/Cas9 knock-in strategy in mESC, we introduced PEs with or without oCGIs within topological associating domains (TADs) harbouring genes with different types of promoters. By evaluating the chromatin, topological and regulatory properties of the engineered PEs, we uncovered that, rather than increasing their local activation, the oCGI boost the physical and functional communication between PEs and distally located developmental genes. Furthermore, we demonstrate that developmental genes with CpG rich promoters are particularly responsive to PEs and that such responsiveness depends on the presence of oCGI. Therefore, our work unveils a novel role for CGIs as genetic determinants of the compatibility between genes and enhancers, which has major implications for the current understanding of how developmental gene expression programs are deployed as well as for our ability to predict the pathological consequences of human structural variation.

overall_design: 4C-seq from mouse embryonic stem cells (mESC) and anterior neural progenitor cells (AntNPC)

first: Tomas

last: Pachano


title: Large-scale tethered function assays identify factors that regulate mRNA stability and translation (eCLIP)

status: Public on Aug 18 2020

city: La Jolla

first: En-Ching

last: Luo

address: 9500 Gilman St

institute: UCSD

summary: Enhanced UV crosslinking and immunoprecipitation (eCLIP) identifies thousands of endogenous mRNA targets for these prioritized RBPs that respond to protein level changes reminiscent of their effects in tethered functional assay.

overall_design: CLIP-seq

first: Gene

last: Yeo


title: Dose-dependent and subset-specific regulation of midbrain dopaminergic neuron differentiation by LEF1-mediated WNT1/b-Catenin signaling

city: Neuherberg

first: Johannes

last: Beckers

address: Ingolstaedter Landstr. 1

department: Institute of Experimental Genetics

institute: Helmholtz Zentrum Muenchen

summary: The development of the mesodiencephalic dopaminergic (mdDA) neurons strongly depends on the WNT1/b-catenin signaling pathway. These neurons include the Substantia nigra pars compacta (SNc) subset that preferentially degenerates in Parkinson’s Disease (PD), and the ventral tegmental area (VTA) subpopulation implicated in a variety of neuropsychiatric disorders. The identity of the cells responding to this signaling pathway in the developing mammalian ventral midbrain (VM) and the precise mechanism of WNT/b-catenin action in these cells, however, are still unknown. Moreover, this signaling pathway has to be accurately balanced during mdDA neuron development: whereas low levels or absence of WNT1/b-catenin signaling abolish their correct specification, constitutive activation of this signaling pathway prevents their proper differentiation in the mouse. We show that the WNT/b-catenin-responsive cells constitute only a fraction of all mdDA progenitors, precursors and neurons in the murine VM. These WNT/b-catenin-responsive cells are mostly located in the Wnt1+, Rspo2+ and Lef1+ lateral floor plate of the medial and caudal midbrain, giving preferentially rise to caudomedial (VTA) mdDA neurons. Strong WNT/b-catenin signaling mediated by RSPO2, a WNT/b-catenin agonist, and LEF1, a nuclear effector of this pathway, inhibits the differentiation of WNT/b-catenin-responsive mdDA progenitors into mature mdDA neurons by repressing the murine Pitx3 gene via conserved LEF1/TCF binding sites in its promoter. Our data indicate that an attenuation of WNT/b-catenin signaling in mdDA progenitors is essential for their correct differentiation into specific mdDA neuron subsets, thus providing a new means for stem cell-based regenerative therapies of PD and in vitro models of neuropsychiatric diseases.

overall_design: We performed gene expression microarray analysis of the laser-microdissected WNT/beta-catenin-responsive mdDA domain from the ventral midbrain of BAT-gal embryos at E12.5 and compared it to a non-WNT/beta-catenin-responsive and non-mdDA domain from the lateral midbrain. In addition, we analysed primary and FACS-sorted GFP-positive Pitx3+/GFP mdDA neurons isolated from the E13.5 and E14.5 ventral midbrain of Pitx3+/GFP embryos

first: Parivash

last: Nouri

first: Sebastian

last: Götz

first: Benedict

last: Rauser

first: Martin

last: Irmler

first: Changgeng

last: Peng

first: Dietrich

last: Trümbach

first: Christian

last: Kempny

first: Carina

last: Nilp

first: Andrea

last: Dlugos

first: Ellen

last: Euchner

first: Rudolf

last: Grosschedl

first: Claude

last: Brodski

first: Wolfgang

last: Wurst

first: Claudia

last: Klümper

first: Nilima

last: Prakash


title: Large-scale tethered function assays identify factors that regulate mRNA stability and translation (HEK293T_RNA-seq)

first: En-Ching

last: Luo

summary: we either depleted RBPs by lentiviral transduction of short-hairpin RNAs (shRNAs) or ectopically expressed the open reading frame of the protein. For each RBP, we performed two independent transductions of two different targeting shRNAs and two independent transductions of a non-targeting shRNA, or performed two independent transfections with a plasmid directing expression of the RBP as a V5-tagged fusion, with the FLAG construct as a control.

overall_design: RNAseq

first: Gene

last: Yeo


title: The expression profiles of lncRNA before and after the initiation of primordial follicle

city: Nanchang

first: su

last: tie

address: XueFu road 999,HongGuTang newly-developed area,NanChang City,JiangXi province

department: Jiangxi Medical College

institute: Nanchang university

type: Non-coding RNA profiling by high throughput sequencing

summary: Primordial follicle accounts for the majority of the ovarian reserve, and recent interest has focused on the mechanisms underlying the follicle initiation. LncRNA is non-coding RNA that has a pivotal role in modulating various bodily activities. Recent studies suggest that lncRNA is able to regulate gene expression at different levels and participate in myriad physiological and pathological processes. For example, lncRNA regulates differentiation of germ cell. However, the regulative effects of lncRNA on follicle initiation have not yet been systemically identified. We compared the expression profile of control group (where ovaries were obtained from three-day-yaer old mice) and experimental group (where samples were cultured in vitro for eight days) by high-throughput sequencing. Our results revealed that 3397 lncRNAs and 1990 mRNAs which showed significantly differential expression (³2.0 fold-change, P < 0.05), and these transcripts are expected to be associated with follicle initiation. Further researches are needed to uncover the mechanisms of follicle initiation based on present study.

overall_design: employing high-throughput sequencing technology to describe lncRNA profiles of ovarian follicle before the initiation (3d) and the follicle after the initiation (3d ovary were cultured in vitro for 8 days)

first: Liping

last: Zheng

first: Tie

last: Su


title: Regulation of retrotransposon activity and telomeres in primed pluripotent stem cells (RNA-seq)

first: Weiyu

last: Zhang

overall_design: We cultured naive and primed mESCs for five passages (P5, relatively early passages) and also ten or fifteen additional passages (P15 or P20, relatively late passages) under naïve and primed conditions to test their gene expression, telomere analysis, transposon elements transcription, genomic stability ,etc.

first: Haifeng

last: Fu


title: Large-scale tethered function assays identify factors that regulate mRNA stability and translation (polysome profiling)

summary: Performed polysome profiling in cell lysates from two independent UBAP2L knockout clonal isolates and from two control samples.


title: Large-scale tethered function assays identify factors that regulate mRNA stability and translation

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


title: Calreticulin Ins5 and Del52 mutations impair unfolded protein and oxidative stress responses in hematopoietic cells

city: Modena

first: Rossella

last: Manfredini

address: Via Gottardi 100

department: Life Sciences

institute: Centre for Regenerative Medicine

summary:
Somatic mutations of calreticulin (CALR) have been described in approximately 30-40% of JAK2 and MPL unmutated essential trombocytemia and primary myelofibrosis patients. CALR is a chaperone that localizes primarily in the endoplasmic reticulum (ER) where it is responsible for the control of proper protein folding and for the calcium retention. Recent data have demonstrated that the TPO receptor (MPL) is essential for the development of CALR mutant-driven myeloproliferative neoplasms (MPNs). However, the precise mechanism of action of CALR mutants haven't been fully unraveled. In this study, we attempted to clarify whether CALR mutations may affect the functions of CALR in the ER under homeostatic conditions. Our results showed that CALR mutants impair the ability to respond to the ER stress and reduce the activation of the pro-apoptotic pathway of the unfolded protein response, therefore allowing the accumulation of unfolded proteins and conferring resistance to ER-stress induced apoptosis. Moreover, our data demonstrated that CALR mutations induce increased sensitivity to oxidative stress, reducing the ability to counteract ROS intracellular accumulation and thus leading to increase oxidative DNA damage. We finally demonstrated that the downmodulation of OXR1 in CALR-mutated cells could be one of the molecular mechanisms responsible for the increased sensitivity to oxidative stress mediated by CALR mutations. Altogether, our data identify a novel MPL-independent mechanism involved in the development of MPNs mediated by CALR mutants: CALR mutations negatively impact on the capability of cells to respond to oxidative stress, and this in turn leads to increase DNA damage and genomic instability.
We used microarrays to detail the change of gene expression caused by calreticulin mutations in K562 cells

overall_design: K562 cells were transduced with CALR wt, CARLIns5 and CALR Del52 respectively and analyzed for gene expression .

first: Simona

last: Salati


title: Gene expression of soybean root tips under water deficits

status: Public on Aug 15 2020

city: Columbia

first: Babu

last: Valliyodan

state: Missouri

address: 1-31 Ag Building

department: Plant Sciences

institute: University of Missouri

summary:
As an important adaptation to drought stress, several agronomic species, such as soybean and maize, can maintain the primary root substantial elongation rates at low water potentials, whereas shoot growth stops completely. In soybean, kinematic characterization of the spatial patterns of cell expansion within the root elongation zone showed that at low water potentials, elongation rates were preferentially maintained toward the root apex but were progressively inhibited at more basal locations, resulting in a shortened growth zone.
To explore the molecular mechanism of root elongation in response to water stress, we set out to examine the expression of soybean genes in different root regions after 5 hours (5h) and 48 hours (48h) water stress treatment using the Affymetrix Soybean GeneChip containing 37,500 G. max probe sets.

overall_design: We examined spatial-temporal expression of soybean  (cv.Magellan) genes in roots under water deficits.  Overall, twenty-seven samples, including three biological replicates for each sample, were analyzed.

first: Henry

last: Nguyen