title: AP-1 transcriptional activity drives micrograft-mediated cell migration and promotes the matrix remodeling machinery

city: Leuven

first: Frederic

last: Lluis

state: Leuven

address: Herestraat 49

department: Department of Development and Regeneration

country: Belgium

institute: KU Leuven

summary: Impaired wound healing and tissue regeneration have severe consequences on the patient’s life quality. Micrograft therapies are emerging as promising and affordable alternatives to improve skin regeneration by enhancing the endogenous wound repair processes. However, the molecular mechanisms underpinning the beneficial effects of the micrograft treatments remain largely unknown. In this study, we identified the active protein-1 (AP-1) member Fos-related antigen-1 (Fra-1) to play a central role in the extracellular-signal-regulated kinase (ERK)-mediated enhanced cell migratory capacity of micrograft-treated mouse adult fibroblasts and in the human keratinocyte (HaCaT) cell model. Accordingly, we show that increased micrograft-dependent in vitro cell migration and matrix metalloprotease activity is abolished upon inhibition of AP-1. Furthermore, micrograft treatment leads to increased expression and post-translational phosphorylation of Fra-1 and c-Jun, resulting in the upregulation of wound healing associated genes mainly involved in the regulation of cell migration. Collectively, our work provides insights into the molecular mechanisms behind micrograft treatments, which might contribute to future advances in wound repair therapies.

overall_design: A total of 3 samples (3 biological replicates) have been used to analyse transcriptome changes in wounded murine primary fibroblasts upon 5h of Micrograft (MG) treatment in association with MEK inhibitor PD0325901 (ERK signalling pathway inhibition).

first: Martina

last: Balli

first: Jonathan

middle: S

last: Chui

first: Paraskevi

last: Athanasouli

first: Willy

middle: A

last: Abreu De Oliveira

first: Youssef

last: El Laithy

first: Maurilio

last: Sampaolesi


title: Functional determinants of sensitivity to CDK4/6 inhibition or cell-cycle plasticity

status: Public on Jun 30 2020

city: Buffalo

first: Ram

middle: Mohan

last: Nambiar

state: NY

address: 665 Elm Street

country: USA

institute: Roswell Park

type: Expression profiling by high throughput sequencing

summary:
Intrinsic or acquired resistance to clinically approved CDK4/6 inhibitors have emerged as a major obstacle that hinders their utility beyond ER+ breast cancer.   Palbociclib-sensitive ER+ breast cancer models and pancreatic ductal adenocarcinoma (PDAC) models were employed to identify functional determinants of response.
In all models tested the activation of RB and inhibition of CDK2 activity emerged as determinants of sensitivity.   While depleting CDK4 and 6 was sufficient to limit proliferation in specific resistance setting, RB loss renders cells completely CDK4/6 independent.   The main down-stream target in this context is the activation status of CDK2 which is suppressed with CDK4/6 inhibition in an RB-dependent fashion.  The P27KIP1 protein levels are associated with plasticity/rigidity of the cell cycle and correlates with sensitivity to CDK4/6 inhibiion.   Exogenous overexpression and pharmacological induction of P27KIP1 by targeting the MEK/ERK pathway enhances the cytostatic effect of palbociclib.
Similar to cell-culture data, in vivo experiments revealed that combination treatment of palbociclib and trametinib in mice bearing PDAC PDXs elicited a robust anti-proliferative effect with a corresponding increase in p27 expression. Mice bearing MCF7 xenografts displayed a durable response in the presence of palbociclib; however, over the course of treatment few cells begin to evade the negative cell-cycle regulation.
Based on multispectral staining, the MCF7 tumor cells that underwent RB inactivation in the presence of palbociclib harbored low p27 expression. Finally, we demonstrate that the cell-cycle plasticity that enables tumor models to evade the palbociclib mediated RB activation could be potently targeted using a clinically applicable CDK2 inhibitor.

overall_design: MCF7 wild type and RB knockout cells and the Pancreatic Cells  were exposed to  PF06873600(200 nM). Following 48 hours exposure, RNA was extracted from these cells using RNeasy plus kit (Quiagen) according to manufacturer's protocol and then subjected to RNA sequence analysis.

first: Vishnu

last: Kumarasamy

first: Agnieszka

middle: K

last: Witkiewicz

first: Erik

last: Knudsen


title: Transcriptomic profiling of eggs of pikeperch, Sander lucioperca, obtained from wild and domesticated fish

city: Rennes

first: Aurelie

last: LE CAM

address: Campus de Beaulieu

department: PHASE

country: France

institute: INRA

summary: Domestication of finfishes is a key element toward sustainable expansion of aquaculture sector. However, huge gap in knowledge on the mechanisms of adaptability of fish to the culture environment, being the first step toward successful domestication, exists. This affects the overall production capacity which is hindered, among others, by lowered reproductive capacity of domesticated stocks, including pikeperch – a highly valued commercial freshwater fish species. Transcriptomic profiling of embryos was found to be useful tool significantly contributing into understanding domestication process in aquaculture. Up to now there is no data on transcriptomic profiling of unfertilized eggs in any fish species as well as on pikeperch, in general. Using next generation sequencing the first complex transcriptome of pikeperch have been obtained (with 40,246 unique protein-coding genes). Next, specifically designed, pikeperch-specific microarrays (Agilent technologies) were developed (with 35,343 unique probes) and transcriptomic profiling of eggs coming from wild and domesticated populations (all characterized by high quality) was performed. From among 17,543 genes found to be expressed in eggs of pikeperch 710 were found to be differentially expressed. Functional, enrichment analysis revealed that the most enriched processes represented by the set of genes identified were developmental process, regulation of phosphorylation and nervous system development. The results of the present study provides a new insight into the processes being directly involved in the successful domestication of finfishes. It can be suggested that all the identified processes were pre-determined by the maternally derived set of genes contained already in the unfertilized eggs, what has never been reported to date. In addition to that, it has been shown for the first time that among the most important processes being affected by the domestication the post translation modifications of proteins along with development of the nervous system were identified. This allows to suggest that fish behavior should be considered as a very important trait for selection for aquaculture purposes in pikeperch.

first: Daniel

last: Żarski

last: Le Cam

first: Joanna

last: Nynca

first: Christophe

last: Klopp

first: Sławomir

last: Ciesielski

first: Beata

last: Sarosiek

first: Jerome

last: Montfort

first: Jarosław

last: Król

first: Pascal

last: Fontaine

first: Andrzej

last: Ciereszko

first: Julien

last: Bobe


title: Redox-sensitive bZIP68 plays a role in balancing stress tolerance with growth in Arabidopsis

status: Public on Jul 01 2020

city: HONG KONG

first: Huan

last: ZHONG

state: HONG KONG

address: T1012 CHA CHI-MING SCIENCE TOWER

department: BIOLOGY

country: Hong Kong

institute: HKBU

summary: Perturbation of the cellular redox state by stress conditions is sensed by redox-sensitive proteins so that the cell can physiologically respond to stressors; however, the mechanisms linking sensing to response remain poorly understood in plants. Here we report that the transcription factor bZIP68 underwent in vivo oxidation in Arabidopsis cells under oxidative stress which is dependent on its redox-sensitive Cys320 residue. bZIP68 is primarily localized to the nucleus under normal conditions in Arabidopsis seedlings and oxidative stress reduces its accumulation in the nucleus and increases its cytosolic localization. Chromatin immunoprecipitation followed by sequencing (ChIP-seq) revealed that bZIP68 primarily binds to promoter regions containing the core G-box (CACGTG) or G-box-like motif of the genes involved in abiotic and biotic stress responses, photosynthesis, biosynthetic processes, and transcriptional regulation. The bzip68 mutant displayed slower growth but enhanced tolerance to oxidative stress. The results from the ChIP-seq and phenotypic and transcriptome comparison between the bzip68 mutant and wildtype indicate that bZIP68 normally suppresses expression of stress tolerance genes and promotes expression of growth-related genes, whereas its inactivation enhances stress tolerance but suppresses growth. bZIP68 might balance stress tolerance with growth through the extent of its oxidative inactivation according to the environment.

overall_design: Transcriptome and CHIP-seq of bzip68 mutants

first: Yimin

last: LI

first: Wuzhen

last: LIU

first: Hailei

last: ZHANG

first: Yiji

last: XIA


title: Prominin-1-expressing Hepatic Progenitor Cells Induce Fibrogenesis in Murine Cholestatic Liver Injury

city: Los Angeles

first: Celia

middle: Anne

last: Short

state: CA

address: 4650 Sunset Blvd

department: Pediatric Surgery

institute: Children's Hospital Los Angeles

summary:
Purpose: Cholestatic liver injury is associated with intrahepatic biliary fibrosis, which can progress to cirrhosis. Resident hepatic progenitor cells (HPCs) expressing Prominin-1 (Prom1/CD133) become activated and participate in the expansion of cholangiocytes known as the ductular reaction. Previously, we demonstrated that in biliary atresia, Prom1(+) HPCs are present within developing fibrosis and that null mutation of Prom1 significantly abrogates fibrogenesis. Here, we hypothesized that these activated Prom1-expressing HPCs promote fibrogenesis in cholestatic liver injury.
Methods: Using Prom1CreERT2-nLacZ/+;Rosa26Lsl-GFP/+ mice, we traced the fate of Prom1-expressing HPCs in the growth of the neonatal and adult livers and in biliary fibrosis induced by bile duct ligation (BDL).
Results: Prom1-expressing cell lineage labeling with Green Fluorescent Protein (GFP) on postnatal day 1 exhibited an expanded population as well as bipotent differentiation potential towards both hepatocytes and cholangiocytes at postnatal day 35. However, in the adult liver, they lost hepatocyte differentiation potential. Upon cholestatic liver injury, adult Prom1-expressing HPCs gave rise to both PROM1(+) and PROM1(-) cholangiocytes contributing to ductular reaction without hepatocyte or myofibroblast differentiation. RNA-sequencing analysis of GFP(+) Prom1-expressing HPC lineage revealed a persistent cholangiocyte phenotype and evidence of Transforming Growth Factor-b pathway activation.
Conclusion: Our data indicate that Prom1-expressing HPCs promote biliary fibrosis by activation of myofibroblasts in cholestatic liver injury.

overall_design: Evaluated the mRNA transciptomes of adult hepatic Prom1+ expressing hepatic progenitor cells and their progeny at day 0 (sham), and d3 or d14 after bile duct ligation (cholestatic liver injury model). D0 (n=3), d3 (n=3), d14 (n=4, but only 4 included due to poor quality and <10M bp reads)

first: Michael

last: Fenlon

first: Kasper

last: Wang


title: Circulatory proteins permeate the brain via selective transport impaired with age"

city: Stanford

first: Michelle

middle: Berkeley

last: Chen

address: 318 Campus Drive

institute: Stanford

summary: single cell RNAseq of 3 month mouse brain endothelial cells

overall_design: single cell RNAeq of murine brain endothelial cells

middle: B

first: Andrew

last: Yang


title: Loss of hepatic Mboat7 leads to liver fibrosis

city: Dresden

first: Alexander

last: Herrmann

address: Fetscherstraße 74

department: Clinic of Internal Medicine I

country: Germany

institute: Technical University Dresden, University Hospital Carl Gustav Carus

summary:
Objective: The rs641738C>T variant located near the membrane-bound O-acyltransferase domain containing 7 (MBOAT7) locus is associated with fibrosis in liver diseases, including non-alcoholic fatty liver disease (NAFLD), alcohol-related liver disease, hepatitis B and C. We aim to understand the mechanism by which the rs641738C>T variant contributes to pathogenesis of NAFLD.
Design: Mice with hepatocyte-specific deletion of MBOAT7 (Mboat7 Δhep) were generated and livers were characterised by histology, flow cytometry, qPCR, RNA sequencing and lipidomics. We analysed the association of rs641738C>T genotype with liver inflammation and fibrosis in 846 NAFLD patients and obtained genotype-specific liver lipidomes from 280 human biopsies.
Results: Allelic imbalance analysis of heterozygous human liver samples pointed to lower expression of the MBOAT7 transcript on the rs641738C>T haplotype. Mboat7 Δhep mice showed spontaneous steatosis characterised by increased hepatic cholesterol ester content after 10 weeks. After 6 weeks on a high fat, methionine-low, choline-deficient diet, mice developed increased hepatic fibrosis as measured by picrosirius staining (p<0.05), hydroxyproline content (p<0.05) and transcriptomics, while the inflammatory cell populations and inflammatory mediators were minimally affected. In a human biopsied NAFLD cohort, MBOAT7 rs641738C>T was associated with fibrosis (p=0.004) independent of the presence of histological inflammation. Liver lipidomes of Mboat7 Δhep mice and human rs641738TT carriers with fibrosis showed increased total lysophosphatidylinositol (LPI) levels. The altered LPI and phosphatidylinositol subspecies in MBOAT7 Δhep livers and humans rs641738TT carriers were similar.
Conclusion: Mboat7 deficiency in mice and human points to an inflammation-independent pathway of liver fibrosis that may be mediated by lipid signalling and a potentially targetable treatment option in NAFLD.

overall_design: Identification of deregulated gene expression in Mboat7 deficient mice mice under steady state (CHOW) and under highfat methionine low choline deficient diet.

first: Veera Raghavan

last: Thangapandi

first: Oskar

last: Knittelfelder

first: Mario

last: Brosch

first: Eleonora

last: Patsenker

first: Olga

last: Vvedenskaya

first: Stephan

last: Buch

first: Sebastian

last: Hinz

last: Hendricks

first: Marina

last: Nati

first: Devavrat Ravindra

last: Rekhade

first: Thomas

last: Berg

first: Madlen

last: Matz-Soja

first: Klaus

last: Huse

first: Edda

last: Klipp

first: Josch K

last: Pauling

first: Judith AH

last: Wodke

first: Jacobo Miranda

last: Ackerman

first: Malte

last: von Bonin

first: Elmar

last: Aigner

first: Christian

last: Datz

first: Witigo

last: von Schönfels

first: Sophie

last: Nehring

last: Zeissig

first: Christoph

last: Röcken

first: Andreas

last: Dahl

first: Triantafyllos

last: Chavakis

first: Felix

last: Stickel

first: Andrej

last: Shevchenko

first: Clemens

last: Schafmayer

first: Jochen

last: Hampe

first: Pallavi

last: Subramanian


title: Regulation of liver regeneration by interference with TNF alpha mediated inflammation

city: Bronx

first: Sanjeev

last: Gupta

address: 1300 Morris Park Ave.

department: Medicine

institute: Albert Einstein College of Medicine

summary:
The study examined the role of TNF alpha in regulation of liver regeneration. For this purpose animals were divided in three groups of healthy controls, 2/3 partial hepatectomy alone, and animals pretreated with TNF alpha antagonist followed by 2/3 partial hepatectomy.
Liver regeneration responses were then examined in conjunction with gene expression analysis at the peak of partial hepatectomy induced DNA synthesis. Substantive differences were identified in multiple gene ontology  groups and cellular events and processes to indicate that TNF alpha was deleterious for partial hepatectomy induced liver regeneration.

overall_design: Study groups were adult rats, three in each group of healthy; 2/3 hepatectomy pretreated with saline as vehicle alone control, and experimental group  treated with etanercept for 16-18 hrs before partial hepatectomy. RNA specimens extracted from livers of animals 32 hrs after partial hepatectomy  were pooled in each group and used to generate RNA-seq on Iron Torrent platform

first: Fadi

middle: L

last: Jaber

first: Tatyana

last: Tchaikovskaya

first: David

last: Rhee

first: Wilber

last: Quispe

first: Alexander

middle: Y

last: Maslov

first: Aaron

last: Golden


title: Transcriptional response of Enterococcus faecali to the glycosylated bacteriocin glycocin F

city: Palmerston North

first: Sean

middle: William

last: Bisset

state: Manawatu

address: Tennent Drive

department: School of Fundamental Sciences

country: New Zealand

institute: Massey University

summary: Glycocin F (GccF) is a bacteriocin produced by Lactobacillus plantarum which causes susceptible cells to enter bacteriostasis within minutes of exposure. GccF is diglycosylated with two N-acetylglucosamine (GlcNAc) moieties and is active against strains of Lb. plantarum, as well as potential pathogens such as Enterococcus faecalis and E. faecium. Its mechanism of action, however, is unknown. To better understand how GccF inhibits growth, we carried out RNA sequencing (RNA-seq) on E. faecalis strain JH2-2 exposed to different concentrations of GccF for different lengths of time. Here we demonstrate that free GlcNAc both protects E. faecalis from the effects of GccF, as well as pre-sensitises it to GccF. Transcriptional analysis showed that exposure to free GlcNAc resulted in up-regulation of a GlcNAc-specific phosphotransferase system (PTS) transporter, which is a putative GccF receptor. Time course analysis showed over 100 genes were up- or down-regulated within 10 minutes of exposure, a number that increased to over 300 genes after 40 minutes. A large number of these genes were found to be regulated by the alternative sigma factor 54, which was shown to not be required for susceptibility to GccF. Interestingly, genes encoding the V-type ATPases and sodium:proton antiporters were found to be up-regulated at all times and all concentrations of GccF tested. This suggests an indirect response to GccF, possibly due to low-level permeabilization of the cell membrane.

overall_design: Enterococcus faecalis cells were exposed to the bacteriocin glycocin F at different concentrations and different times, and transcriptional response profiled usind RNA sequncing on an Illumina HiSeq 2000. Samples were carried out in triplicate, with the necessary untreated controls.

middle: W

first: Dave

last: Wheeler


title: Transcriptomic analysis of neuroblastoma cells in response to stable over-expression of myeloid zinc finger 1 antisense RNA 1 (MZF1-AS1)

city: Wuhan

first: Qiangsong

last: Tong

state: Hubei

address: 1277 Jiefang Avenue

department: Department of Surgery

country: China

institute: Union Hospital of Tongji Medical College

summary: Neuroblastoma (NB), a malignant embryonic tumor arising from primitive neural crest cells, accounts for more than 7% of malignancies and around 15% of cancer-related mortality in childhood. Better elucidating the mechanisms of tumorigenesis and aggressiveness is important for improving the therapeutic efficiencies of NB. Through mining of public datasets, we identified myeloid zinc finger 1 antisense RNA 1 (MZF1-AS1) as a novel lncRNA associated with the progression of NB. To investigate the mechanisms underlying the oncogenic functions of MZF1-AS1, we employed the Illumina HiSeq X Ten as a discovery platform to analyze the transcriptome profiling changes of human SH-SY5Y cells in response to stable over-expression of MZF1-AS1. The results showed that stable over-expression of MZF1-AS1 led to altered expression of 2920 human mRNAs, including 1476 up-regulated genes and 1444 down-regulated genes. Then we found the possible roles of these differentially regulated mRNAs in selected pathways including proline synthesis, invasion, and metastasis by Bioinformatic analysis. Furthermore, we validated the RNA-seq results by real-time RT-PCR with high identity. Overall, our results provided fundamental information about the transcriptomic changes in response to MZF1-AS1 over-expression in human NB cells, and these findings will help us understand the pathogenesis of NB.

overall_design: Total RNA of cells stably transfected with empty vector or MZF1-AS1 was extracted using the TRIzol® reagent according to the manufacturer's instructions. RNA concentration was measured using a Qubit® RNA Assay Kit with a Qubit® 2.0 Fluorometer (Life Technologies, Inc.), and integrity was assessed using the RNA Nano 6000 Assay Kit with a Bioanalyzer 2100 system (Agilent Technologies, CA). Library preparation and transcriptome sequencing on an Illumina HiSeq X Ten platform were performed by Novogene Bioinformatics Technology Co., Ltd. (Beijing, China), and 100 bp paired-end reads were generated. HTSeq v0.6.0 was used to count the reads numbers mapped to each gene.

first: Liduan

last: Zheng


title: Epigenetic and Transcriptional Regulation of Supporting Cell to Hair Cell Transdifferentiation

city: Boston

first: Albert

last: Edge

state: Massachusetts

address: 243 Charles St

institute: Harvard Medical School

summary:
Purpose: We have recently extablished a new protocol for the generation of inner ear organoid or Lgr5-positive cochlear progenitors (LCPs). The purpose of this study is to analyze the expression pattern in these organoids as they differentiate and to compare them to native hair cells and supporting cells that were sorted from mouse inner ear at P2.

Methods: Lgr5-cochlear progenitors were harvested from newborn mice, dissociated and plated in Matrigel to generate organoids. The cells were expanded for 10 days and then differentiation for additional 10 days. Samples were harvested for RNA at D0 of differentiation, D2, D4 and D10 and RNA was extracted using RNeasy micro Kit (Qiagen). In parallel, whole cochleae were harvested from P2 litters of Atoh1-GFP, Lgr5-GFP and Sox2-GFP mice, sorted for GFP+ cells and RNA was extracted in the same manner. mRNA libraries were generated from using ThruPLEX DNA-seq kit (Rubicon Genomics) and then sequenced using Illumina NextSeq500 Single-End 75bp. Reads were aligned to mm10 and quality control was assessed using FastQC and Qualimap followed by FeatureCounts, Sailfish v0.10.1 and tximport. Differential expression was evaluated using DESeq2 and subsequent analysis was performed using Ingenuity Pathway Analysis Software (Qiagen) and standard R packages for RNA-Seq analysis. Samples were ran in duplicates and LCP samples were harvest from a single experiment for each of the duplicates.

Results: 40-50 million reads were detected per sample. PCA and Pearson correlation demonstrated a clear segragation of the samples to native cells Vs. cultured cells and along a gradient of differentiation. Multiple pairwise comparisons of differentially expressed genes were conducted to reveal several thousands of DE genes between the different LCP time points and between the native sorted cell types. Additional analysis of expression throughout differentiation segregated the genes into 8 clusters that represent proliferation, early differentiation and late differentiation gene groups in cochlear development. Clear downregulation of supporting cell and cell-cycle markers was detected as the cells differentiation alongside an upregulation of known hair cell markers. A similar distinction was detected between Atoh1-positive native hair cells and Lgr5-positive native supporting cells.

Conclusions: Our results validate the resemblance between our new organoid model system and native cochlear cells. It is evident that the cells at the end of the proliferation phase are similar to supporting cells (and not progenitor cells) and are therefore transdifferentiating into hair cells. The existence of several cochlear markers indicate a separation into outer hair cells and inner hair cells but does not exclude potential differentiation of a portion of the organoids towards a vestibule fate.

overall_design: Samples were generated in duplicates from sorted cochlear tissue at P2 and from expanded and differentiated LCPs. During the analysis process, native hair cells were compared to native supporting cells and LCP samples were compared between their different time points.

first: Danielle

middle: R

last: Lenz

first: Victor

last: Barrera Burgos


title: Gene expression profiling of fibroblasts in a family with LMNA-related cardiomyopathy reveals molecular pathways implicated in disease pathogenesis

city: Irvine

first: Trina

middle: M.

last: Norden-Krichmar

address: School of Medicine

institute: University of California, Irvine

summary: LMNA mutation caused reduced expression of IGFBP5 in patient samples

overall_design: Three groups (Unrelated Control (Donor), Family control (Control) and Patient) with three biological replicates each for a total of 9 samples

middle: M

first: Halida

middle: P

last: Widyastuti

first: Anna

last: Grosberg

first: Michael

middle: V

last: Zaragoza


title: Cell Senescence

city: La Jolla

first: Xue

last: Lei

address: 10901 N Torrey Pines Rd

institute: SBP medical discovery institute

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

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Refer to individual Series


title: RNA-seq and 5hmC-seq in acute myeloid leukemia (AML) cells with and without knockdown of Tet1

city: Monrovia

first: Jianjun

last: Chen

department: Systems Biology

institute: Beckman Research Institute of City of Hope

type: Methylation profiling by high throughput sequencing


title: Whole genome expression profile of human decidual PMN-MDSC in normal pregnancy and unexplained recurrent pregnancy loss

city: Shanghai

first: Congcong

last: Li

state: Shanghai

address: 160 Pujian Road

department: OB & GYN

institute: Ren Ji Hospital, School of Medicine, Shanghai Jiao Tong University

summary: Polymorphonuclear myeloid-derived suppressor cell (PMN-MDSC) accumulate in maternal-fetal interface during pregnancy and play a role in maintenance of immune tolerance. Decreased PMN-MDSC is associated with pregnancy complications such as unexplained recurrent pregnancy loss (URPL). Whether PMN-MDSC function is different between normal pregnancy (NP) and URPL remains unexplored. Here we performed whole genome expression profile of 3 decidual PMN-MDSC from normal early pregnancy and 3 decidual PMN-MDSC from URPL. Total RNA were extracted. Cy5-labeled aRNA was hybridized and scanned on a G2505C Agilent Microarray Scanner with Agilent 0.1 XDR software. The gene expression pattern of the PMN-MDSC was significantly different between the NP group and the URPL group.

overall_design: Total RNA of 3 decidual PMN-MDSC from normal early pregnancy and 3 decidual PMN-MDSC from unexplained recurrent pregnancy loss were analyzed and compared in this study.

first: Aimin

last: Zhao


title: HPV cell lines and HPV tissue MicroRNA SOLiD sequencing

city: Helsinki

first: Petri

last: Auvinen

address: Viikinkaari 5

country: Finland

institute: University of Helsinki

type: Non-coding RNA profiling by high throughput sequencing

summary: Human papillomaviruses (HPV) preferentially infect keratinocytes of mucous membranes or skin and cause numerous benign and malignant lesions at different anatomical locations. We sequenced small RNA (sRNA) libraries of two HPV 16 immortalized cell lines and ten formalin fixed paraffin embedded (FFPE) tissue samples from HPV infected cervical epithelium by SOLiD 4 technology.

overall_design: Small RNA (sRNA) libraries of two HPV 16 immortalized cell lines, HPK IA and HPK II, and ten formalin fixed paraffin embedded (FFPE) tissue samples from HPV infected cervical epithelium, were made according to SOLiD sequencing instructions.

first: Kui

last: Qian

first: Tuuli

last: Pietilä

first: Lars

last: Paulín

first: Eeva


title: Enhancer-promoter association determines cell specificity in mouse pluripotent cells

city: Shenzhen

first: Qing

address: No 7,Pengfei Road,Dapeng District

institute: Agricultural Genomics Institute at Shenzhen, Chinese Academy of Agricultural Sciences,

summary: Enhancer-promoter association is emerging as a common pattern to determine gene function in eukaryotes. However, it remains as an uncultivated land on how its diversity influences gene expression. In this study, we select three defined Sox2 associated enhancers, representing different distal interaction categories, to further explore its biological effects. We construct three enhancer-knock-out cells via CRISPR/Cas9 system in mouse embryonic stem cells. With multiple-level evidences, we realize these associations carried out various biological features. Embryonic cell specific association determines cell proliferation and cell cycle and also involves in cardiac function, which has been validated in vivo. On the other hand, the common or indirect association prefers to regulate cell metabolic process and nerve development in cell function. Our study will enable an alternative way for gene spatiotemporal regulation study and advance our understanding of its diversity since first described decades ago.

overall_design: Analysis of the effect of different promoter-enhancer associations on Sox2 diverse function in mouse embryonic stem cells

first: Lei

last: Huang

first: Qitong

first: Siyuan

last: Kong

first: Xiusheng

last: Zhu

first: Yanling

last: Peng

first: Yubo

last: Zhang


title: ER-associated degradation is required for the maintenance of β cell identity via TGFβ signaling

status: Public on Jun 29 2020

city: Wageningen

first: Guido

last: Hooiveld

address: HELIX, Stippeneng 4

department: Div. Human Nutrition & Health

country: Netherlands

institute: Wageningen University

summary: β cell apoptosis and dedifferentiation are two hotly-debated mechanisms underlying β cell loss in type 2 diabetes (T2D); however, the molecular drivers underlying such events remain largely unclear. Here, by performing a side-by-side comparison of mice carrying β cell-specific deletion of endoplasmic reticulum (ER)-associated degradation (ERAD) and autophagy, we report that while autophagy appears necessary for β cell survival, the highly conserved Sel1L-Hrd1 ERAD protein complex is required for the maintenance of β cell maturation and identity. Notably, SEL1L expression is significantly reduced in human T2D islets compared to healthy human islets. At the single cell level, we demonstrate that Sel1L deficiency is not associated with β cell loss, but rather loss of β cell identity. Mechanistically, we find that Sel1L-Hrd1 ERAD controls β cell identity via TGFβ signaling, in part by mediating the degradation of TGF-β receptor 1 (TGFβRI). Inhibition of TGFβ signaling in Sel1L-deficient β cells augments the expression of β cell maturation markers and increases the total insulin content. Our data reveal profound but distinct pathogenic effects of two major proteolytic pathways in β cells, providing a new framework for therapies targeting distinct mechanisms of protein quality control.

overall_design: Gene expression was profiled in pancreatic islets isolated from 5-week-old wildtype and β cell-specific Sel1L-deficient mice.

first: Neha

last: Shrestha

first: Tongyu

last: Liu

first: Yewei

last: Ji

first: Rachel

last: Reinert

first: Mauricio

last: Torres

first: Maria

first: Chih-Hang

last: Tang

first: Chih-Chi

last: Hu

first: Chengyang

first: Ali

last: Naji

first: Jiandie

last: Lin

first: Sander

last: Kersten

first: Peter

last: Arvan

first: Ling

last: Qi


title: IL-18BP is a secreted immune checkpoint and barrier to effective IL-18 immunotherapy

city: New Haven

first: Meaghan

last: McGeary

state: CT

address: 300 Cedar St.

department: Immunobiology

institute: Yale School of Medicine

summary: Recombinant cytokines were the first modern immunotherapies to produce durable cures in metastatic cancer, but their application has been hampered by only modest efficacy and limited tolerability. Next-generation cytokine therapies are therefore under development to overcome the biological limitations of native cytokines. By analyzing single-cell transcriptomic data from tumor infiltrating lymphocytes (TIL), we found that components of the Interleukin-18 (IL-18) pathway are upregulated on activated and dysfunctional TIL, suggesting that IL-18 therapy could potentially restore anti-tumor immunity by stimulating these key effector cells. However, recombinant IL-18 therapy has consistently failed to demonstrate anti-tumor efficacy in clinical trials. Here we show that the secreted, high-affinity decoy receptor IL-18BP is frequently upregulated in the tumor microenvironment of diverse human cancers and syngeneic murine tumor models. Using directed evolution, we engineered a ‘decoy-resistant’ IL-18 (DR-18), which maintains signaling potential, but is impervious to binding and inhibition by IL-18BP. In contrast to wild-type IL-18, DR-18 exhibits potent anti-tumor efficacy as monotherapy and in combination with anti-PD-1 checkpoint immunotherapy in multiple tumor models. Mechanistically, DR-18 drives the development of poly-functional effector CD8+ T cells, decreases the prevalence of exhausted CD8+ T cells expressing the transcription factor TOX, and expands the pool of TCF1+ precursor CD8+ T cells. DR-18 also enhances NK cell activity and maturation to effectively treat anti-PD-1 resistant tumors that have lost MHC class I surface expression. Together, these results highlight the IL-18 pathway as a powerful target for immunotherapeutic intervention and implicate the secreted immune checkpoint IL-18BP as an obstacle to effective IL-18 immunotherapy.

overall_design: Droplet-based 3' end single cell RNA sequencing was performed on pooled cells from 3 tumors per condition, consisting of 50% CD45+CD3e+, 25% CD45+CD3e-, 25% CD45- after FACS. Droplets and libraries were prepared using Chromium Single Cell 3' Reagent Kits v2 according to manufacturer's protocol (10X genomics). Generated libraries were sequenced on Illumina HiSeq 4000.

first: Ting

last: Zhou

first: William

last: Damsky

first: Orr-El

last: Weizman

first: K

last: Hartmann

first: Connor

middle: E

last: Rosen

first: Ruaidhri

last: Jackson

first: Suzanne

last: Fischer

first: Jun

last: Wang

first: Miguel

middle: F

last: Sanmamed

first: Richard

last: Flavell

first: Marcus

last: Bosenberg

first: Aaron

last: Ring


title: Molecular Pathways of Colon Inflammation Induced by Cancer Immunotherapy

first: Shengbao

last: Suo

state: MA

address: 360 Longwood Ave

institute: Dana-Farber Cancer Institute

summary: Checkpoint blockade with antibodies specific for the PD-1 and CTLA-4 inhibitory receptors can induce durable responses in a wide range of human cancers. However, the immunological mechanisms responsible for severe inflammatory side effects remain poorly understood. Here we report a comprehensive single cell analysis of immune cell populations in colitis, a common and severe side effect of checkpoint blockade. We observed strong enrichment of several T cell clusters in colitis lesions compared to control cases, in particular a striking accumulation of CD8 T cells with signatures reflecting highly cytotoxic and proliferative states. T cell receptor (TCR) sequence analysis demonstrated that a substantial fraction of colitis-associated CD8 T cells originated from tissue-resident populations, explaining the frequently early onset of colitis symptoms following treatment initiation. Our analysis also identified cytokines, chemokines and surface receptors that could serve as therapeutic targets for colitis and potentially other inflammatory side effects of checkpoint blockade.

overall_design: Samples from 22 patients from 3 different cohorts (Normal control, +CPI no colitis, +CPI colitis)

first: Kai

last: Wucherpfennig


title: Molecular profiling of aromatase inhibitor-treated post-menopausal breast tumors identifies immunerelated correlates of resistance

city: London

first: Alice

last: Gao

state: London

address: 237 Fulham Road

department: Breast Cancer Now

country: United Kingdom

institute: The Institute of Cancer Research

summary: To identify in clinical breast cancer those genes and pathways most associated with resistance to aromatase inhibitors by examining the global transcriptional effects of aromatase inhibitor (AI) treatment. Analysis of gene-expression measured in core-cut biopsies at baseline and surgery from patients received 2-week’s presurgical AI revealed: i) the molecular response to AI treatment varies greatly between patients, consistent with the variable clinical benefit from AI treatment; ii) higher baseline expression of an inflammatory signature is associated with poor antiproliferative response,  and should be assessed further as a novel biomarker and potential target for AI-treated patients.

overall_design: Global gene-expression measured in core-cut biopsies, from 112 postmenopausal women with stage I to IIIB ER+ early breast cancer before and after two-weeks’ anastrozole monotherapy in a neoadjuvant trial.

first: Qiong

first: Ricardo

last: Ribas Santos


title: Reciprocal gene editing defines targetable mutant H3.3 oncohistone effectors in pediatric glioma

city: Sacramento

first: Paul

last: Knoepfler

address: 2425 Stockton Blvd, Room 633

department: Cell Biology and Human Anatomy

institute: University of California Davis

type: Genome binding/occupancy profiling by high throughput sequencing

summary: High-grade pediatric gliomas often contain histone H3.3 mutations, but open questions remain about oncogenic mechanisms. To address this gap, we performed ‘reciprocal gene editing’ using CRISPR-Cas9 to introduce H3.3 mutations (K27M, G34R) into H3.3-wildtype brain and glioma cells, while in parallel reverting pre-existing K27M mutations in glioma cells back to wildtype. Analyses of our reciprocally-edited cells indicate that H3.3 mutation leads to specific transcriptomic and epigenetic events, and associated cell biological changes including in xenograft assays. We used these data and the reciprocally-edited cells to screen selected drugs and identify specific putative treatments that are mutant H3.3-specific. Overall, reciprocal gene editing provides new insights into mutant H3.3 oncogenic mechanisms and more broadly may prove useful for studying other cancer-associated mutations.

overall_design: We used CRISPR-Cas9 gene editing of H3F3A to precisely revert preexisting K27M mutations to WT in patient-derived glioma cells to identify consistently functionally important effectors. Two replicates of H3K27me3 and two replicates of H3.3 ChIP-Seq were performed in each of the following cell lines: Line XIII, Line XIII WT, Line XVII, Line XVII WT.

first: Rachel

last: Klein


title: Deep sequencing of human fibroblastic cell line IMR90, either proliferating or senescent,  with or without TSA treatment.

first: Xue

last: Lei

summary:
Gene expression changes were compared in proliferating and senescent human IMR90 cells, with or without TSA treatment.
Method: RNA was extracted, in triplicate, then sequenced with an Illumina NextSeq 500 sequencer. Sequence reads passing the quality control filters were aligned using Tophat2 and then analysed with Cufflinks.

overall_design: polyA mRNA profiles from proliferating or senescent human IMR90 cells, with or without TSA, were generated, in triplicate, using an Illumina NextSeq 500 sequencer.

first: Maria-Grazia

last: Vizioli

first: Peter

middle: D

last: Adams

first: Kathryn

last: Gilroy


title: Array comparative genomic hybridization of canine follicular lymphoma (cFL) and canine marginal zone lymphoma (cMZL)

city: Legnaro

first: Diana

last: Giannuzzi

state: Padova

address: viale dell'Università, 16

department: Dept. of Comparative Biomedicine and Food Science

country: Italy

institute: University of Padova

summary: The aim of this work was to identify the copy number aberrations (CNAs) by high-resolution array comparative genomic hybridization (aCGH) on 7 dogs with newly diagnosed FL and 5 dogs with newly diagnosed MZL.

overall_design: In total, the DNA obtained from 7 cFL and 5 cMZL specimens together with the paired skin biopsies were analyzed using a Agilent-025522 SurePrint G3 Canine CGH Microarray 4x180K (GPL24897). Tumor and reference DNA samples were labelled independently with cyanine 5-deoxyuridine triphosphate (dUTP) and cyanine 3-dUTP, respectively, using SureTag Complete DNA Labeling Kit. Oligo aCGH analysis was performed following the manufacturer’s recommendations, using an Oligo aCGH/ChIP-on-chip Hybridization Kit (Agilent Technologies).

first: Giannuzzi

last: Diana

first: Aresu

last: Luca


title: Atrx deletion in neurons leads to sexually-dimorphic dysregulation of miR-137 and spatial learning and memory deficits

first: Vanessa

last: Dumeaux

state: Ontario

address: Elgin Dr

department: Anatomy and Cell Biology

country: Canada

institute: Western University

summary: Ablation of the ATRX chromatin remodeler specifically in forebrain excitatory neurons of mice causes male-specific deficits in long-term spatial memory associated with miR-137 overexpression, transcriptional changes and structural alterations corresponding to pre- and post-synaptic abnormalities.

overall_design: Hippocampal total RNA was isolated from three pairs of 3-month-old control and Atrx-cKO male and female mice (12 samples total).

first: Renee

middle: J

last: Tamming

first: Luana

last: Langlois

first: Jacob

last: Ellegood

first: Lily

last: Qiu

first: Yan

last: Jiang

first: Jason

last: Lerch

first: Nathalie

middle: G

last: Bérubé


title: Transcriptome analysis of differentially expressed genes in the feronia (fer-4) mutant

first: Chao

last: Li

address: 500 dongchuan Road

institute: East China Normal University

summary: FERONIA (FER) receptor kinase is a multifunctional regulator controlling myriad plant growth and developmental processes and interactions with the environment. We perform a RNA-seq analysis of genes differentially expressed in fer-4 mutant and observed significant changes of groups of genes previously characterized to be involved in AUXIN, ABA, ethylene and RALF1 signaling. The Results provide insight into the molecular mechanisms of FERONIA participation in regulating peptide and hormone signaling.

overall_design: examination of transcript level change

first: Hanqian

last: Feng


title: Deep sequencing of human fibroblastic cell line IMR90, either proliferating or senescent,  expressing either empty vector or Parkin, with or without CCCP treatment

first: Xue

last: Lei

summary:
Gene expression changes were compared in proliferating and senescent human IMR90 cells, with either empty vector, Parkin, or Parkin+CCCP.
Method: RNA was extracted, in triplicate, then sequenced with an Illumina NextSeq 500 sequencer. Sequence reads passing the quality control filters were aligned using Tophat2 and then analysed with Cufflinks.

overall_design: polyA mRNA profiles from proliferating or senescent human IMR90 cells, with either empty vector, parkin, or parkin plus CCCP, were generated, in triplicate, using an Illumina NextSeq 500 sequencer.


title: Genome wide ChIP-seq analysis of human TOP2B occupancy in KG-1 myeloid cell line

city: Newcastle

first: Simon

last: Cockell

address: Framlington Place

institute: Newcastle University

summary: TOP2B is involved in transcriptional initiation in response to nuclear hormone ligands and plays a role in transcriptional elongation. Whole genome TOP2B ChIP-seq was carried out on human KG-1 cells in the presence and absence of the topoisomerase inhibitors mitoxantone and etoposide.  Chromatin was assayed for TOP2B occupancy by peak calling with SISSRs and MACS.

overall_design: Whole genome TOP2B ChIP-seq was carried out on human KG-1 cells in the presence (60 mins exposure) and absence of the topoisomerase inhibitors etoposide and mitoxantrone. "Input" control samples were also sequenced for background detection and comparison.

first: Caroline

last: Austin

first: Zbyslaw

last: Sondka

first: Ian

last: Cowell


title: Expression data of four wheat genotypes viz. HD2967, K307, K9107 and Chotilarma (CL) in response to nitrogen chronic limitation

city: New Delhi

first: Pranab

middle: kumar

last: Mandal

state: Delhi

address: LBS building, Pusa Campus

department: National Research Centre on Plant Biotechnology

country: India

institute: Indian council of Agriculture Research

summary:
The transcriptome profile was examined in four wheat genotypes in roots and shoots under nitrogen stressed condition which indicates genotype specific transcript data-set apart from the common transcripts. Unique genes was identified for nitrogen uptake and utilization process.
We used microarrays to detail the gene expression and identify the candidate genes related to uptake and utilization of nitrogen in root and shoot tissues of wheat genotypes.

overall_design: We collected seedlings from four wheat genotypes viz., HD2967, K9107, K307 and Chotilarma (CL) grown for 15 days in pots containing mixture of vermiculite and perlite in a ratio 2:1. Three sets of pots for each genotype were grown in two different conditions : control including 8mM Ca(NO3)2 and nitrogen starved condition  0.08mM Ca(NO3)2 as source. Total RNA isolated from these tissue samples and amplified RNA were purified and labeled by biotin with GeneChip® IVT labeling kit. The biotinylated cDNAs were fragmented and hybridized to the Affymetrix GeneChip® Wheat Genome Array (Affymetrix, Inc., Carlsbad, CA) for 16 hours. Normalization of the microarray data was conducted using Robust Multichip Average (RMA) algorithm with Affymetrix® GeneChip® Command Console® Software (AGCC) used to analyze the data. Transcriptome profiling was detected in all four different wheat genotypes for N+ and N- plants. Each array dataset was then imported into Agilent GeneSpring software 14.5 for analysis. The signal intensities of the samples were transformed into log2-ratio data. Normalized data with a p-value of ≤ 0.1 that passed a Student’s t- test was used to identify differentially expressed genes.

middle: K

first: Subodh

last: Sinha

first: Akanksha

last: Singh


title: The effects of sulforaphane treatment on the expression of microRNAs in Caco-2 and CCD-841 cells

city: Norwich

first: Tamas

last: Dalmay

address: Earlham Road

department: School of Biological Sciences

institute: University of East Anglia

summary: The effect of sulforaphane on the expression of microRNAs in colorectal adenocarcinoma Caco-2 cells and non-cancerous colorectal CCD-841 cells was investigated, in order to help ascertain the roles of microRNAs in the anti-cancer effects of sulforaphane.

overall_design: Caco-2 and CCD-841 cells were treated for 8 or 24 h with sulforaphane or DMSO alone (control). Final DMSO concentrations were 0.05%. Total RNA was then isolated, and microRNAs were cloned as cDNA-based libraries. The libraries were then subject to deep sequencing.

first: Christopher

last: Dacosta

first: Claudia

last: Paicu

first: Irina

last: Mohorianu

first: Yongping

last: Bao


title: Next Generation Sequencing timecourse of murine hepatocytes with or without paracetamol treatment

summary:
Untreated mice or mice treated with paracetamol, in the presence or absence of TSA, were culled after 14 hrs or 24 hrs of treatment, and RNAseq was performed.
Method: RNA was extracted, in triplicate, then sequenced with an Illumina NextSeq 500 sequencer. Sequence reads passing the quality control filters were aligned using Tophat2 and then analysed with Cufflinks.

overall_design: Quadruplicate polyA mRNA profiles from mouse hepatocytes were generated following various drug treatment protocols, using an Illumina NextSeq 500 sequencer.

first: Maria-Grazia

last: Vizioli

first: Peter

last: Adams

first: Kathryn

last: Gilroy


title: Isolation and molecular characterisation of Dunaliella tertiolecta with truncated light-harvesting antenna for enhanced photosynthetic efficiency.

city: Heidelberg

first: Staffan Andreas

last: Johansson

address: Meyerhofstraße 1

institute: EMBL Heidelberg

summary: Digital gene expression analysis by Ht-SuperSAGE (Matsumura et al. 2011. High-Throughput SuperSAGE. Methods Mol Biol.) was carried out to compare gene expression between wild type and mutant cell lines of the green algae Dunaliella tertiolecta under different light levels.

overall_design:
Novel cell lines were generated using chemical mutagenesis and isolated using a high-throughput selection protocol. The gene expression of two selected lines with reduced chlorophyll content was compared to the wild type in triplicates under low, medium and high light.

The raw data files for this series are multiplexed. They were provided to the submitter in this form. Sample demultiplexing was done by Iwate Biotechnology Research Centre (Kitakami, Iwate, Japan) using custom scripts. The file "Sample_metadata.xls" available at the foot of this record includes the individual sample IDs and metadata. The processed data file includes count data for sequences.

first: S

middle: A

first: P

last: Stephenson

first: R

last: Edwards

first: K

last: Yoshida

first: C

last: Moore

last: Terauchi

first: M

last: Zubkov

last: Terry

first: T

middle: S

last: Bibby


title: 5hmC-seq in human MonoMac6 acute myeloid leukemia (AML) cells with and without knockdown of TET1

first: Jianjun

last: Chen

summary: To dissect the mechanism underlying the regulation of TET1 on its target genes in AML, we performed 5hmC-sequencing for genomic DNA isolated from human MonoMac6 AML cells with and without knockdown of TET1.

overall_design: We lentivirally transduced pLKO.1-based lentiviral shRNA targeting TET1 (i.e., shTET1-1, and shTET1-2) or scramble shRNA (i.e., shNS) into MonoMac6 cells. After puromycin selection (1 µg/mL) for two passages, cells were collected and genomic DNA was extracted using DNeasy blood and tissue kit (Qiagen). DNA was fragmented and labelled with 6-N3-glucose using β-GT enzyme. Biotin tags were added onto the 6-N3-glucose with Huisgen cycloaddition (click) chemistry reaction using DBCO-PEG4-biotin. The biotin-labelled 5hmC containing DNA was enriched with streptavidin-coupled magnetic beads. The enriched DNA was used for library preparation and next-generation sequencing.

first: Hengyou

last: Weng

first: Huilin

last: Huang

first: Chunxiao

last: Song

first: Paulina

last: Siejka-Zielinska


title: Coordinated regulation of cellular identity-associated H3K4me3 breadth by the COMPASS family

status: Public on Jun 27 2020

city: Chicago

first: Ali

last: Shilatifard

address: 303 East Superior Street

institute: Northwestern University

summary: Histone H3 lysine 4 trimethylation (H3K4me3) facilitates recruitment of transcription factors and epigenetic effectors to promote transcriptional activation and ensure cellular identity. This conserved histone mark is implemented by the COMPASS (COMplex of Proteins ASsociated with Set1) family of H3K4 methyltransferases. COMPASS members Set1A and Set1B have been accredited as primary depositors of global H3K4me3 in mammalian cells. Our previous studies in mouse embryonic stem cells (ESCs) demonstrated that deleting the enzymatic SET domain of Set1A does not perturb bulk H3K4me3, indicating possible compensatory roles played by other COMPASS methyltransferases. Here, we generated a series of ESC lines harboring compounding mutations of the COMPASS enzymes. We find that Set1B is functionally redundant to Set1A in implementing H3K4me3 at highly expressed genes, while Mll2 deposits H3K4me3 at less transcriptionally active promoters. Furthermore, Set1A and Set1B are responsible for broad H3K4me3 peaks, whereas Mll2 establishes H3K4me3 with narrow breadth. Most importantly, Mll2 helps preserve global H3K4me3 levels and peak breadth. Our results illustrate the biological flexibility of such enzymes in regulating transcription in a context-dependent manner to maintain stem cell identity, which could assist our understanding of their disease liability.

overall_design: Elucidate roles of various COMPASS methyltransferases on H3K4me3 levels and breadth by investigating a series of generated ESC lines with compounding mutations of the COMPASS enzymes

first: Christie

middle: C

last: Sze

first: Siddhartha

last: Das


title: RNA-seq for  two maize inbred lines

city: Harbin

first: Yu

last: Zhou

state: Heilongjiang

address: Changjiang Road,Xiangfang district

department: College of Agronomy

institute: Northeast Agricultural University

summary: Find differentially expressed genes under low temperature (10℃) and normal conditions (25℃) during maize germination

overall_design: The low temperature resistant lines 55 (named Zao8-3) and one low temperature sensitive line 102 (named Ji853)were used in this study. The seeds were sampled imbibition within the germinating paper under the ‘normal’ condition (25℃)  for CK_102 and CK_55  and low temperature condition (10℃)  for LT_102 and LT_55,biologically repeated three times.Comparison of differentially expressed genes of two maize lines at low temperature by RNA-seq

first: Zhenhua

last: wang

first: Hong

last: Zhang

first: Jiayue


title: Determination of ascorbate-, glutathione- and salicylate-responsive miRNAs in Arabidopsis

first: Tamas

last: Dalmay

summary: miRNA profile of wild type Arabidopsis plants was compared to miRNA profiles in three different lines with altered levels of ascorbate (vtc2-1), glutathione (pad2-1) or salicylate (nahG).

overall_design: 12 libraries were generated (3 repeats for each of the four genotypes) from young leaves of 4-week-old plants grown in standard conditions. Only 2 repeats of vtc2-1 are included here.

first: Zsolt

last: Gulyás

first: Thao

last: Huynh

first: Martina

last: Billmeier

first: Ping

last: Xu

first: Irina

last: Mohorianu

first: Gábor

last: Kocsy


title: H3K9 methylation fine-tunes coordination of transcription by regulating the conformational plasticity of chromosomal folding [RNA-Seq]

city: Stockholm

first: Anita

last: Göndör

address: Nobels väg 16, KI Solna Campus, Karolinska Institutet, Box 280

department: MTC

country: Sweden

institute: Karolinska Institutet

summary: Despite that stochastic gene expression variations underlie lineage specification and adaptive responses, little is known about the plasticity of 3D chromatin folding and its effects on cell-to-cell transcriptional variability. Employing the Chromatin In Situ Proximity (ChrISP) technique we visualize here developmentally acquired chromosome 11-specific chromatin hubs rich in the repressive H3K9me2 mark that project towards the lamina in single cells. Inhibition of the H3Kme2 methyl-transferase G9a/Glp not only antagonizedperipheral chromatin compaction, but strikingly also induced the emergence of dense chromatin clusters in the nuclear interior. Changes in chromatin organization were accompanied by the spatial re-organization of transcriptional coordination without affecting the overall level of transcription. LOCKs thus topologically insulate active genes at the nuclear periphery, while allowing transcriptional coordination in the rest of the chromosome. We conclude that G9a/Glp controls not only the plasticity of chromatin conformations, but also spatial coordination of transcription to affect cellular gene expression variability.

overall_design: 2 Biological replicates and 2 technical replicates where treated with both G9a Inhibitor and siRNA

first: Xingqi

last: Chen

first: Samer

last: Yammine

first: Noriyuki

last: Sumida

first: Alejandro

last: Fernandez-Woodbridge

first: Chengxi

last: Shi

first: Farzaneh

last: Shahin Varnoosfaderani

first: Mariliis

last: Tark-Dame

first: Marta

last: Imreh

first: Rolf

last: Ohlsson


title: Loss of TET2 results in DNA methylation changes in mouse naïve B-cells

city: Farmington

first: Sheng

last: Li

address: 10 Discovery Drive

institute: The Jackson Laboratory

summary: Absence of Tet2 at mouse cells results in gain of DNA methylation

overall_design: Enhanced reduced representation bisulfite sequencing (ERRBS) of mouse TET2 WT and KO GC B-cells

first: Ari

last: Melnick

first: Maria

last: Del Pilar Dominguez

first: Xiaowen

first: Wojciech

last: Rosikiewicz


title: Comprehensive miRNA profiling of human naïve, intermediate and primed pluripotent stem cells

city: Basel

first: Klas

last: Hatje

state: Basel

address: Grenzacherstrasse 124

country: Switzerland

institute: Roche Pharma Research and Early Development

summary: Naïve human pluripotent stem cells (hPSC) represent an earlier time-point in embryogenesis than conventional, ‘primed’ hPSCs. We present a comprehensive miRNA profiling of naïve-to-primed transition in hPSC, a process resembling aspects of early in vivo embryogenesis. We identify miR-143-3p and miR-22-3p as markers of the naïve state and miR-363-5p, several members of the miR-17 family, miR-302 family as primed markers. We uncover that miR-371-373 are highly upregulated in naïve hPSC. MiR-371-373 are the human homologs of the mouse miR-290 family, which are the most highly expressed miRNAs in mPSC. This aligns with the consensus that naïve hPSC resemble mPSC, showing that the absence of miR-371-373 in conventional hPSC is due to cell state rather than a species difference.

overall_design: To investigate miRNAs involved in the differentiation of naïve hPSC to primed hPSC, the effectiveness of the in vitro differentiation process itself was first assessed, using the directly derived naïve HNES1 and HNES2 cell lines. Differentiation was instigated in three separately cultured replicates of each of HNES1 and HNES2 naïve cells, plating them on geltrex and switching medium to E8 after 48 hours, thereby removing potential variability introduced if splitting the cells. The time points were chosen to capture the exit from naïve pluripotency, with a final late time point to ensure complete priming of the cells. Day 42 was chosen for the final time point since the cells had by then regained the ability to differentiate using standard primed protocols. Total RNA samples from naïve cells cultured above (HNES1 and HNES2, each cultured separately in biological triplicates) at day 0, during the transition (day 2) and when fully primed (day 42), were analysed by miRNAseq.

first: Benjamin

middle: T

last: Dodsworth

last: Rostovskaya

first: Rowan

last: Flynn

first: Claas

last: Meyer

first: Sally

last: Cowley


title: An insulin-sensitive circular RNA that regulates lifespan in Drosophila

status: Public on Jun 26 2020

city: Cologne

first: Carina

middle: Marianne

last: Weigelt

address: Joseph-Stelzmann-Str 9b

institute: Max Planck Institute for Biology of Ageing

summary: CircRNAs are a novel, poorly characterized class of non-coding RNAs that accumulate with age in neuronal tissues of diverse species. To characterize the role of circRNAs during ageing, we used RNA sequencing to analyse circRNA expression in Drosophila tissues (brain, thorax, gut and fat body) at different ages (day 10, 30 and 50) in wildtype (wDah) flies and long-lived insulin mutant flies (dilp 2-3,5)

overall_design: Multi-tissue RNA sequencing of Drosophila melanogaster during ageing (Day 10, 30 and 50). Female wildtype flies and dilp 2-3,5 mutant flies. Total RNA or polysome-enriched RNA. Day 10 samples were generated independently.

first: Linda

last: Partridge


title: Expression data from human peripheral B cells in hypoxic and normoxia

status: Public on Jun 23 2020

city: Cambridge

middle: James

last: Stewart

address: Wellcome Genome Campus

department: Cellular Genetics

institute: Wellcome Sanger Institute

summary: Human leukocyte cones were obtained from anonymous blood donors and peripheral blood B cells were isolated from PMBCs following ficoll density centrifugation. Isolated B cells were cultured in normoxia or 1% O2 for 24 h and RNA was extracted using an RNeasy Mini kit (Qiagen). This study was approved by the East of England - Cambridge Central Research Ethics Committee (REC reference 08/H0308/176).

first: N

last: Burrows

first: P

last: Maxwell

first: B

first: M

middle: R

last: Clatworthy


title: Treatment of Bone Marrow derived Macrophages with Immune complex

first: John

middle: Robert

last: Ferdinand

address: Department of Medicine

institute: University of Cambridge

summary: Murine bone marrow derived macrophages were treated with Immne complex for 4 hours after which transcriptional changes were assessed by microarray.

overall_design: BMDM's were derived from 3 separate mice. The cells from each mouse were split, half were stimulated with Ova immune complex for 4 or 14 hours and half were left in normal media for further culture.

first: Rebeccah

last: Mathews

first: Gemma

last: Banham

first: Chenzhi

last: Jing

first: Tomas

last: Castro-Dopico

first: Menna


title: Alternative polyadenylation mediates genetic regulation of gene expression

first: Yoav

last: GiladLi

state: IL

address: 920 e 58th St

department: Section of Genetic Medicine

institute: University of Chicago

summary: Little is known about co-transcriptional or post-transcriptional regulatory mechanisms linking noncoding variation to variation in organismal traits. To begin addressing this gap, we used 3' Seq to study the impact of genetic variation on alternative polyadenylation (APA) in the nuclear and total mRNA fractions of 52 HapMap Yoruba human lymphoblastoid cell lines. We mapped 602 APA quantitative trait loci (apaQTLs) at 10% FDR, of which 152 were nuclear specific. Effect sizes at intronic apaQTLs are negatively correlated with eQTL effect sizes. These observations suggest genetic variants can decrease mRNA expression levels by increasing usage of intronic PAS. We also identified 24 apaQTLs associated with protein levels, but not mRNA expression. Finally, we found that 19% of apaQTLs can be associated with disease. Thus, our work demonstrates that APA links genetic variation to variation in gene expression, protein expression, and disease risk, and reveals uncharted modes of genetic regulation.

overall_design: We extracted whole cell and nuclear mRNA from 52 human LCLs and submitted them for 3' high throughput sequencing.

first: Briana

last: Mittleman

first: Sebastian

last: Pott

first: Shane

last: Warland

first: Tony

last: Zheng

first: Zepeng

last: Mu

first: Mayher

last: Kaur

last: Gilad

first: Yang

middle: I

last: Li


title: Analysis of genome architecture during SCNT reveals a role of cohesin in impeding minor ZGA

city: Beijing

first: Ke

last: Zhang

state: Beijing

address: Haidian District

department: School of Life Sciences

institute: Tsinghua university

summary: Somatic cell nuclear transfer (SCNT) can reprogram a somatic nucleus to a totipotent state. However, the re-organization of three-dimensional chromatin structure in this process remains poorly understood. Using low-input Hi-C, we revealed that during SCNT, the transferred nucleus first enters a mitotic-like state (premature chromatin condensation). Unlike fertilized embryos, SCNT embryos show stronger TADs at the 1-cell stage. TADs become weaker at the 2-cell stage, followed by gradual consolidation. Compartments A/B are markedly weak in 1-cell SCNT embryos and become increasingly strengthened afterward. By the 8-cell stage, somatic chromatin architecture is largely reset to embryonic patterns. Unexpectedly, we found cohesin represses minor zygotic genome activation (ZGA) genes (2-cell specific genes) in pluripotent and differentiated cells, and pre-depleting cohesin in donor cells facilitates minor ZGA and SCNT. These data reveal multi-step reprogramming of 3D chromatin architecture during SCNT and support dual roles of cohesin in TAD formation and minor ZGA repression.

overall_design: Donor MEF/cumulus cells (MEF/CC), the SCNT embryos at the PCC stage (premature chromosome condensation) (1 hour after nuclear transfer) and SCNT embryos after PCC exit through “SCNT activation” using strontium chloride (SrCl2) at the 1-cell(4hours after activation, 6hours after activation and 10hours after activation), late 2-cell (28 hours after activation), and 8-cell (56 hours after activation) stages were obtained. Both MEF and CC are used for donor cells for the collection of SCNT 8-cell samples. HiC and RNA-seq were performed in these cells at various stages during SCNT embryogenesis. For the predepletion of CTCF/cohesin experiment, the short term differentiated cells with/without auxin treatment are collected for HiC experiments, RNA-seq and ChIP-seq experiments.

first: Dan-Ya

last: Wu

first: Hui

first: Yao

last: Wang

first: Qiao-Ran

last: Sun

first: Xin

last: Liu

first: Li-Yan

first: Wen-Jing

last: Xiong

first: Qiujun

first: James D. P.

last: Rhodes

first: Kai

last: Xu

first: Lijia

first: Zili

last: Lin

first: Guang

last: Yu

first: Weikun

last: Xia

first: Bo

last: Huang

first: Zhenhai

last: Du

last: Yao

first: Kim A.

last: Nasmyth

first: Robert J.

last: Klose

first: Yi-Liang

last: Miao

first: Wei

last: Xie


title: MeCP2 links heterochromatin condensates and neurodevelopmental disease

first: Charles

last: Li

address: 455 Main Street

institute: Whitehead Institute for Biomedical Research

summary: MeCP2 (methyl CpG binding protein 2) is a key component of constitutive heterochromatin, which plays important roles in chromosome maintenance and transcriptional silencing. Mutations in MeCP2 cause Rett syndrome (RTT), a postnatal progressive neurodevelopmental disorder associated with severe mental disability and autism-like symptoms that manifests in girls during early childhood. Heterochromatin, long considered a dense and relatively static structure, is now understood to exhibit properties consistent with a liquid-like condensate. Here we report that MeCP2 is a dynamic component of heterochromatin condensates in cells, is stimulated by DNA to form liquid-like condensates, contains multiple domains that contribute to condensate formation, manifests physicochemical properties that selectively concentrate heterochromatin cofactors compared to components of transcriptionally active condensates, and when altered by RTT-causing mutations is disrupted in its ability to form condensates. We propose that MeCP2 enhances heterochromatin/euchromatin separation through its condensate partitioning properties and that condensate disruption may be a common consequence of RTT patient mutations.

overall_design: RNA-seq in MeCP2-GFP WT and R168X mutant murine ESCs and neurons

middle: H


title: Control of gene expression mRNA N6-methyladenosine in pro-B cells

status: Public on Jun 22 2020

first: Chuan

last: He

address: 929 E 57th Street

department: Chemistry

summary: N6-methyladenosine (m6A) is the most abundant internal modification in eukaryotic messenger RNA (mRNA). In mice, loss of METTL14,  the key m6A writer component, severely blocks B cell development. In this group of RNA sequencing data, we 1. conducted RNA-seq on WT and METTL14 KO pro-B cells (sample 1-10); 2. performed m6A-seq in WT pro-B cells (sample 11-14) and 3. conducted YTHDF2 RIP -seq in WT pro-B cells (sample 15-18). Our data supported that m6A repressed a group of genes in pro-B cells via its reader YTHDF2.

overall_design: Pro-B cells were sorted as CD19+B220midCD2-CD43hismallIg(kappa)/Ig(lambda)- cells from mice and subject to sequencing as described in summary.

first: Xianbin

first: Zhong

last: Zheng


title: Control of early B cell development by mRNA N6-methyladenosine

first: Xiaolong

last: Cui

address: 680 N Lake Shore Drive

department: Department of Preventive Medicine

summary: N6-methyladenosine (m6A) is the most abundant internal modification in eukaryotic messenger RNA (mRNA). Recent conceptual and technological advances have sparked extensive research interests in the functions of m6A, which led to the discovery of its writers, erasers and readers; as well as its regulatory roles in a number of biological processes. However, the physiological roles of mRNA m6A in B cells remained unknown. Here, we report that loss of Mettl14, the core component of the m6A writer complex, severely blocked B cell development in mice. In the bone marrow, deletion of Mettl14 in B lineage cells impaired key processes of B lymphopoiesis, including the recombination of immunoglobulin heavy chain (IgH), the IL-7-induced pro-B cell proliferation, and the transition from the large cycling pre-B stage to the small pre-B stage. Associated with these broad biological effects, Mettl14 deficiency in developing B cells dramatically decreased m6A levels in over 40% transcripts of the whole transcriptome. Global gene expression analysis in Mettl14 KO B cells showed extensive abnormalities in gene expression programs important for B cell development. To further characterize the underlying molecular mechanisms, we investigated the role of two m6A reader proteins YTHDF2 and YTHDF1. Loss of Ythdf2 impaired the IgH recombination and IL-7-induced proliferation of pro-B cells but did not affect the large pre-B-to-small pre-B cell transition. In contrast, Ythdf1 deficiency did not cause any significant defects in B cell development. These data highlighted the complexity of reader proteins that may have distinct, overlapping, or synergistic roles.  Altogether, our data demonstrate that mRNA m6A modification represents a novel regulatory mechanism in early B cell development.

overall_design: We used Mettl14 WT and KO mice to study overall immune system changes.

first: Phillip

last: Hsu


title: A yeast-optimized single-cell transcriptomics platform elucidates how mycophenolic acid and guanine alter global mRNA levels

city: West Haven

first: Guste

last: Urbonaite

address: 300 Heffernan Drive, ISTC, Rm 117

department: Molecular and Cellular Biology

institute: Yale University

summary: The ability to measure the number of gene-specific mRNA molecules in individual mammalian cells has transformed the transcriptomics field. Among the key technologies enabling single-cell mRNA sequencing has been Droplet Sequencing (Drop-Seq). While this method works efficiently for mammalian cells, its direct application to yeast cells has been problematic due to cell-type specific differences such as size, doublet formation rate, and cell wall. Here we introduce YeastDropSeq, a single-cell RNA sequencing method for the study of transcriptomics in yeast. We modified and optimized the original Drop-Seq method to address the issues that emerged from smaller cell sizes and the presence of a cell wall in yeast. We also quantified the rate of doublet formation through a species-mixing experiment. As proof-of-principle application of the YeastDropSeq, we investigated the transcriptomic effects of mycophenolic acid (MPA), a lifespan-extending compound that decreases de novo GMP synthesis. We compared transcript levels between cells treated with MPA and cells treated with DMSO and/or guanine, MPA’s epistatic agent. We discovered that isogenic populations of yeast cells contain transcriptionally distinct subpopulations and that the subpopulation structures were maintained despite the different treatment conditions. We found that cells treated with MPA experience an upregulation of genes coding for proteins involved in DNA replication stress-response, antioxidation, pre-RNA processing, and translation initiation. Conversely, a downregulation of mRNA expression was observed for genes encoding translation initiation and elongation factors, the 40S and 60S ribosomal subunits, and for genes involved in metal transport and mitochondrial function. Additionally, we elucidated that expression levels of several genes of unknown function were affected by the MPA treatment. YeastDropSeq will accelerate biological discovery by facilitating droplet-based transcriptomics of yeast cells.

sample_organism:
Saccharomyces bayanus
Saccharomyces cerevisiae

overall_design: There are 4 samples analyzed. One sample, DMSO, is the control. The other samples: Guanine, Guanine and Mycophenolic acid, and Mycophenolic acid are treated conditions

first: Murat

last: Acar

first: Ping

last: Liu


title: RNA-seq of trnascriptome from human embryonic stem cells treated with DMSO or 1 uM of RG7834

status: Public on Jun 28 2020

city: Boulder

first: Siddharth

last: Shukla

state: CO

address: 3415 Colorado Ave

institute: University of Colorado Boulder

summary: We aimed to investigate whether treatment with a small molecule drug RG7834 significantly affected the transcriptome of human embryonic stem cells carrying a pathogenic mutation in the gene DKC1.

overall_design: RNA in triplicate from DMSO or RG7834 treated DKC1 hESCs was used for strand-specific library preparation using Nextflex Rapid Directional RNA-seq kit 2.0. Libraries were multiplexed and seqeunced on Illumina HiSeq using 2x150 bp reads. Demultiplexed reads were checked for quality, followed by adapter trimming, alignment to hg38 p13 version 99 from Ensembl and read counts were obtaiend using HTseq. Read counts were used for differential gene expression analysis using DESeq2. We did not find any significantly affected genes between the two conditions.

first: Roy

last: Parker

first: Hochang

last: Jeong

first: Luis

last: Batista


title: Identification of lncRNAs involved in the PCV2 infection of PK-15 cells

status: Public on Jun 19 2020

city: Hangzhou

first: Jin

address: Yuhangtang Road No.866

institute: Zhejiang University

summary: Porcine circovirus type 2 (PCV2) can cause severe disease in the affected pigs, resulting in massive economic loss for the swine industry. Transcriptomics and proteomics approaches have been widely employed to identify the underlying molecular mechanisms of PCV2 infection. Numerous differentially expressed mRNAs, miRNAs, and proteins, together with their associated signaling pathways, have been identified, paving the way to further validation of their biological functions. Long non-coding RNA (lncRNA) is an important regulator of multiple biological processes. However, rare information regarding its role in the PCV2 infection has been obtained. Hence, in our study, RNA-seq was performed by infecting PK-15 cells with PCV2. Analysis of the differentially expressed genes (DEGs) suggested that cytoskeleton, apoptosis, cell division, and protein phosphorylation were significantly disturbed. Then, using stringent parameters, six lncRNAs were identified. Additionally, the potential targets of the lncRNAs were predicted using both cis- and trans-prediction methods. Interestingly, we found that the HOXB (Homeobox B) gene cluster was probably the target of the lncRNA LOC106505099. Enrichment analysis of the target genes showed that numerous developmental processes were altered during PCV2 infection. Therefore, our study revealed that lncRNAs could affect porcine embryo development through the regulation of the HOXB genes.

overall_design: PK-15 cells were cultured in DMEM (Hyclone) with 10% FBS (Hyclone) in 6-well plates until reaching 80% confluency. Then, 3 wells of one plate were infected with Porcine Circovirus type 2 (PCV2) with MOI of 0.1. After 72 hours, the cells were harvested (3 PCV2-infection, 3 for mock infection) were harvested, subjected to total RNA extraction, and sequencing.

first: Huanan

last: Wang


title: Synaptic Schwann cells: specific labeling reveals unique cellular and molecular features

city: Blacksburg

first: Robert

last: Settlage

state: VA

address: 1 Washington St

department: Advanced Research Computing

institute: Virginia Tech

summary: Perisynaptic Schwann cells (PSCs) are specialized, non-myelinating, synaptic glia of the neuromuscular junction (NMJ), that participate in synapse development, function, maintenance, and repair. The study of PSCs has relied on an anatomy-based approach, as the identities of cell-specific PSC molecular markers have remained elusive. This limited approach has precluded our ability to isolate and genetically manipulate PSCs in a cell specific manner. We have identified neuron-glia antigen 2 (NG2) as a unique molecular marker of S100β+ PSCs in skeletal muscle. NG2 is expressed in Schwann cells already associated with the NMJ, indicating that it is a marker of differentiated PSCs.  Using a newly generated transgenic mouse in which PSCs are specifically labeled, we show that PSCs have a unique molecular signature that includes genes known to play critical roles in PSCs and synapses. These findings will serve as a springboard for revealing drivers of PSC differentiation and function.

overall_design: Number of replicates for each cell type: NG2-dsRed+, 10; S100β-GFP+, 7; NG2-dsRed+;S100β-GFP+, 9

first: Ryan

last: Castro

first: Thomas

last: Taetzsch

first: Sydney

last: Vaughan

first: Kerilyn

last: Godbe

first: John

last: Chappell

first: Gregorio

last: Valdez


title: Reduced Brd1 expression leads to reversible depression-like behaviors and gene-expression changes in female mice [amygdala]

status: Public on Jun 20 2020

city: Aarhus c

first: Per

last: Qvist

state: Denmark

address: Hoegh Guldbergs Gade 10, Building 1116-460

department: Biomedicine

country: Denmark

institute: Aarhus University

summary:
Background: The schizophrenia associated gene, BRD1, encodes an epigenetic regulator which chromatin interactome is enriched with genes implicated in mental health. Alterations in histone modifications and epigenetic regulation contribute to brain transcriptomic changes in affective disorders and preclinical data supports a role for BRD1 in psychopathology. However, the implications of BRD1 on affective pathology remains poorly understood.
Methods: We assessed affective behaviors and associated neurobiology in Brd1+/- mice along with their responses to Fluoxetine and Imipramine. This involves behavioral, neurostructural, and neurochemical characterizations along with regional cerebral gene expression profiling combined with integrative functional genomic analyses.
Results: Here we report behavioral changes in female Brd1+/- mice with translational value to depressive symptomatology that can be alleviated by administration of antidepressant medications. Behavioral changes are accompanied by altered brain morphometry and imbalances in monoaminergic systems. In accordance, gene expression changes across brain tissues reveal altered neurotransmitter signaling and cluster in functional pathways associated with depression  including ‘Adrenergic-, GPCR-, cAMP-, and CREB/CREM-signaling’. Integrative gene expression analysis specifically links changes in amygdaloid intracellular signaling activity to the behavioral treatment response in Brd1+/- mice.
Conclusions: Collectively, our study highlights the importance of BRD1 as a modulator of affective pathology and adds to our understanding of the molecular mechanisms underlying affective disorders and their treatment response.

overall_design: mRNA profiles of right striatum, caudate putamen and anterior cingulate cortex of 8-11 weeks-old female wild type (WT) and Brd1+/- mice, either untreated or treated with antidepressants imipramine or fluoxetine, were generated by RNA-sequencing, using BGISEQ-500, in order to identify diffentially expressed genes between groups.

first: Anto

last: Rajkumar

first: Julie

last: Donskov


title: Reduced Brd1 expression leads to reversible depression-like behaviors and gene-expression changes in female mice

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

overall_design: Refer to individual Series


title: Differential gene expression upon shRNA-mediated silencing of APC in HT-29 colorectal cancer cells

status: Public on Jun 25 2020

city: Munich

first: Peter

last: Jung

address: Thalkirchner Street 36

department: DKTK at LMU Munich

institute: DKFZ Heidelberg

summary: Wnt signaling plays a pivotal role in colorectal cancer. Intrinsic activation of Wnt by mutational events, such as mutations in the tumor suppressor gene APC, represents the most frequent initiating event in this disease background.  Long truncated versions of APC retain partial functionality, which leads to a sub-maximal, “just right” activation state of Wnt signaling supposed to be beneficial for disease initiation. In order to study the transcriptomic alterations of an over-stimulated Wnt signaling pathway, conditional shRNA-mediated silencing of APC was performed in chromosomal instable HT-29 CRC cells which express a 1555 amino acid variant of APC protein able to bind and partially inactivate β-catenin. To achieve this, cells were stably transduced with lentiviral particles encoding for a doxycyline-inducible shRNA directed against APC, or, as a control, a non-silencing shRNA (pTRIPZ inducible shRNA vectors, RHS4740-EG324, Horizon Dharmacon, CO, USA). 72 hours after APC silencing, total RNA was isolated and quality controlled for subsequent RNA-Seq Analysis (DKFZ Heidelberg, Genomic and Proteomic Core Facility) on a HiSeq 2000 instrument (Illumina). Overall, we observed bona-fide Wnt target genes, such as NKD1, AXIN2, PTK7, ASCL2, and SMOC2, and additional putative direct or indirect targets of Wnt signaling up-regulated upon shRNA-mediated APC silencing.

overall_design: HT-29 cells stably transduced to conditionally express a shRNA directed against APC or, as a control, a non-silencing (NonS) shRNA were treated with 500 ng/ml doxycycline for 72 hrs prior to RNA isolation. The experiment was repeated with the same setting, and biological duplicates were analyzed by RNA-Seq. HT-29 cells induced to express the non-silencing RNA were used as the reference sample for analysis and gene expression in these cells was set to “1” in order to examine relative gene expression upon APC silencing.

first: Vanessa

last: Dietinger


title: Characterization of human erythroid chromatin in a two-phase liquid differentiation system

city: Headington

first: Damien

last: Downes

state: Oxfordshire

address: John Radcliffe Hospital

department: MRC Weatherall Institute of Medicine

institute: The University of Oxford

summary: A two-phase liquid differentiation of peripheral blood mononuclear cells (PBMC) can be used to study human erythroid cells. To determine the epigenetic landscape of erythroid cells derived from PBMC we have performs ChIP-seq for histone marks associated with promoters (H3K4me3), enhancers (H3K4me1) and polycomb repression (H3K27me3) as well as for chromatin bound pol2. In addition we characterized the binding of the boundary element CTCF and key erythroid transcription factors NF-E2 and SCL/TAL.

overall_design: Peripheral blood mononuclear cells were isolated and differentiated from a single male donor and a single replicate of ChIP-seq for histone marks (H3K4me1, H3K4me3, H3K27me3) and transcription factors (CTCF, NFE2, SCL/TAL, POL2). NG Capture-C was also performed on the same cells to determine the chromatin conformation of the alpha globin locus.

first: Nigel

last: Roberts

first: James

middle: O

last: Davies

first: Marco

last: de Gobbi

first: Jim

last: Hughes


title: Reduced Brd1 expression leads to reversible depression-like behaviors and gene-expression changes in female mice [anterior cingulate cortex (aCC)]


title: Delineating the molecular pathways regulated by IFN-gamma and IL-17 in the small intestines of humanized mice during staphylococcal superantigen-induced toxic shock syndrome

first: Shiv

middle: D.

last: Kale

address: 1015 Steger Hall, Life Science Circle

department: Biocomplexity Institute

summary: RNASeq identified several pathways that were differentially expressed/regulated in the small intestines that correlated with intestinal failure during toxic shock syndrome caused by staphylococcal superantigen in a humanized mouse model

overall_design: To study the changes in gene expression in the small intestines during staphylococcal superantigen-induced toxic shock syndrome by RNA-Seq

first: Govind

last: Rajagopalan

first: Sheryl

last: Coutermarsh-ott

first: Saikumar

last: Karyala

first: Brittney

last: Mehrkens

first: Molly

last: Stegman


title: Existence of human-virus chimeric proteins generated during infection

city: La Jolla

first: Max

last: Chang

state: CA

address: 9500 Gilman Drive

institute: University of California, San Diego

summary: Influenza A virus (IAV) is a threat to mankind because it generates yearly epidemics and poorly predictable sporadic pandemics with catastrophic potential. Influenza has a small RNA genome (~14 Kb) composed of 8 mini-chromosomes (segments). Segments encode both structural proteins and proteins expressed only during infection. Segments are constituted by a 5’UTR followed by a coding region and a 3’UTR.  Transcription of IAV RNA into mRNA depends on host RNA Polymerase II, as the viral polymerase cleaves 5’ capped cellular nascent transcripts to be used as primers to initiate mRNA synthesis. We hypothesized that host nascent transcripts bearing AUG could generate upstream ORFs in the viral segments, a phenomenon that would depend on the translatability of the viral 5’UTRs. Using orthogonal datasets we report the existence of this mechanism, which generate host-virus chimeric proteins. We show that most segments encode proteins in this manner, expanding the proteome diversity of IAV in infected cells. Host-virus chimeric proteins are conserved across IAV strains, pointing to an evolutionary conservation of function achieved by sampling of the evolutionary space before gene fixation.  Thus, we discover a mechanism that generate human-virus chimeric proteins during infection.

overall_design: (1) Examination of 2 PB1-UFO mutants (2) Examination of PB1-UFO signature and the host transcriptome in the lung tissue of mice

first: Yixuan

last: Ma

first: Matthew

last: Angel

first: Jessica

last: Ho

first: Guojun

last: Wang

last: Zhao

first: Justine

last: Noel

first: Natasha

last: Moshkina

first: Jeffrey

last: Johnson

first: Zuleyma

last: Peralta

first: Nevan

last: Krogan

first: Christopher

last: Benner

first: Harm

last: van Bakel

first: Marta

last: Łuksza

first: Benjamin

middle: D

last: Greenbaum

first: Emily

last: Miraldi

first: Adolfo

last: Garcìa-Sastre

first: Jonathan

middle: W

last: Yewdell

first: Ivan

last: Marazzi


title: Genome-wide maps of ARID1A binding genes in H9 human embryonic stem cells.

city: Indianapolis

first: Lei

last: Yang

state: Indiana

address: 1044 W. Walnut St.

department: Department of Pediatrics

institute: Indiana University School of Medicine

summary: We reported loss of ARID1A promoted neurogenesis and inhibited cardiogenesis. Here we used specific ARID1A antibody to pull down ARID1A binding genomic DNA in human embryonic stem cells, which let us know the potential genes regulated by ARID1A during neurogenesis and inhibited cardiogenesis. 1% Input sample was collected from the same sample after chromatin shearing.

overall_design: Examination of ARID1A binding genes in hESCs.

first: Juli

last: Liu


title: Regenerative reprogramming of the intestinal stem cell state via Hippo signaling suppresses metastatic colorectal cancer

status: Public on Jun 24 2020

city: Boston

first: Priscilla

last: Cheung

address: 1 Blackfan Circle

department: Stem Cell Program

institute: Boston Children's Hospital


title: Single cell RNA-seq revealed different cell types induced by loss of ARID1A in undifferentiated and differentiation (day 10) H9 hESCs.

summary: We reported loss of ARID1A promoted neurogenesis and inhibited cardiogenesis. Under microscopy, we observed that spontaneously differentiated cells were induced in ARID1A KO H9 hESCs cultured in mTesR medium. After cardiac differentiation for 10 days, we also observed the cell types were totally different between WT and ARID1A KO cells. We did not know what cells types were. Here scRNA-seq were used to identify the cell types in WT H9 hESCs and ARID1A KO H9 hESCs.

overall_design: Identification of different cell types induced by loss of ARID1A in undifferentiated and differentiated H9 hESCs.


title: Assay for Transposase Accessible Chromatin with high-throughput sequencing (ATAC-seq) revealed genomic chromatin accessibilities change induced by loss of ARID1A in differentiated (day 4) H9 hESCs

summary: We reported loss of ARID1A promoted neurogenesis and inhibited cardiogenesis. Under microscopy, we observed that spontaneously differentiated cells were induced in ARID1A KO H9 hESCs cultured in mTesR medium. We did not know what cells types were. Here ATAC-seq were used to investigate chromatin accessibilities change in differentiated (day 4) WT H9 hESCs and ARID1A KO hESC cells.

overall_design: Identification of genomic chromatin accessibilities change induced by loss of ARID1A in H9 hESCs.


title: Induced pluripotent stem cells of patients with Tetralogy of Fallot reveal transcriptional alterations in cardiomyocyte differentiation

city: Berlin

first: Silke

last: Sperling

state: Berlin

address: Lindenberger Weg 80

department: Cardiovascular Genetics

institute: Experimental and Clinical Research Center (ECRC), Charité / MDC for Molecular Medicine

summary: We generated induced pluripotent stem cells (iPSCs) of two patients with Tetralogy of Fallot (TOF) and three healthy relatives of two unrelated familes. We furhter performed mRNA sequencing of iPSCs (day 0) and derived cardiomyocytes (CMs) at day 15 and 60 of patients and healthy relatives.

overall_design: Total RNA was isolated using Direct-zol RNA Kits (Zymo Reasearch) and purified with Dynabeads Oligo(dT) (Thermo Fisher Scientific) to get poly(A) RNA. The RNA libraries were prepared with the ScriptSeq v2 RNA-Seq Library Preparation Kit (Illumina). The libraries were evaluated with the Qubit dsDNA HS Assay Kit (Thermo Fisher Scientific) and the Agilent High Sensitivity DNA Kit (Agilent Technologies). Paired-end sequencing was performed on either HiSeq 2500 (2x51 bp) or NextSeq (2x76 bp) device (Illumina). For the latter, the NextSeq 500/550 v2 kits (Illumina) were used. All kits were applied according to the manufacturer's instructions. Overall, mRNA sequencing was performed for 31 samples of 5 individuals for the following three differentiation stages: pluripotent iPSC (day 0), iPSC-CM (day 15) and CM (day 60). Sequencing was separated into two runs, all day 0 samples and three day 15 samples (Mother TOF-02) were sequenced with Illumina’s HiSeq 2500. The remaining samples were sequenced using Illumina’s NextSeq.

last: Rickert-Sperling


title: Essential and Opposite Roles of ARID1A in Coordinating Human Cardiogenesis and Neurogenesis from Pluripotent Stem Cells


title: Transcriptomic Profiling of Fibrosis-Specific Epithelial Cells in Idiopathic Pulmonary Fibrosis

status: Public on Jun 18 2020

city: St. Gallen

first: Florent

last: Baty

address: Rorschacherstrasse 95

department: Lung Center

institute: Cantonal Hospital St. Gallen

summary: We aimed at characterizing disease-specific differences by comparing the transcriptomes of epithelial cells (ECs) from idiopathic pulmonary fibrosis (IPF) and non-IPF sources

overall_design:
Comparison of the transcriptome of ECs from IPF and non-IPF sources
Submitter declares that the raw data are unavailable at this time for confidentiality reasons.

first: Maximilian

last: Boesch

first: Martin

last: Brutsche

first: Michael

last: Tamm

first: Julien

last: Roux

first: Lars

last: Knudsen

first: Amiq

last: Gazdhar

first: Thomas

last: Geiser

first: Petra

last: Khan

first: Katrin

middle: E

last: Hostettler


title: Control of early B cell development by mRNA N6-methyladenosine

city: Chicago

first: Ruitu

last: Lyu

institute: The University of Chicago

summary: N6-methyladenosine (m6A) is the most abundant internal modification in eukaryotic messenger RNA (mRNA). Recent conceptual and technological advances have sparked extensive research interests in the functions of m6A, which led to the discovery of its writers, erasers and readers; as well as its regulatory roles in a number of biological processes. However, the physiological roles of mRNA m6A in B cells remained unknown. Here, we report that loss of Mettl14, the core component of the m6A writer complex, severely blocked B cell development in mice. In the bone marrow, deletion of Mettl14 in B lineage cells impaired key processes of B lymphopoiesis, including the recombination of immunoglobulin heavy chain (IgH), the IL-7-induced pro-B cell proliferation, and the transition from the large cycling pre-B stage to the small pre-B stage. Associated with these broad biological effects, Mettl14 deficiency in developing B cells dramatically decreased m6A levels in over 40% transcripts of the whole transcriptome. Global gene expression analysis in Mettl14 KO B cells showed extensive abnormalities in gene expression programs important for B cell development. To further characterize the underlying molecular mechanisms, we investigated the role of two m6A reader proteins YTHDF2 and YTHDF1. Loss of Ythdf2 impaired the IgH recombination and IL-7-induced proliferation of pro-B cells but did not affect the large pre-B-to-small pre-B cell transition. In contrast, Ythdf1 deficiency did not cause any significant defects in B cell development. These data highlighted the complexity of reader proteins that may have distinct, overlapping, or synergistic roles.  Altogether, our data demonstrate that mRNA m6A modification represents a novel regulatory mechanism in early B cell development.

overall_design: Examination of m6A enriched regions in WT and Mettl14KO mouse proB cells .Examination of the target mRNAs of  YTHDF2 in WT and Mettl14KO mouse proB cells.

first: Xianbin

last: Yu

first: Chuan

last: He

first: Zhong

last: Zheng

first: Linda

last: Zhang


title: Comprehensive structure-function characterization of DNMT3B and DNMT3A reveals distinctive de novo DNA methylation mechanisms

city: Durham

first: Greg

last: Wang

state: NC

address: 3 Genome Court

department: Duke Department of Pharmacology and Cancer Biology  and Duke Cancer Institute

institute: Duke University School of Medicine, and Department of Biochemistry and Biophysics, UNC at Chapel Hill (adjunct)

summary: Mammalian DNA methylation patterns are established by two de novo DNA methyltransferases DNMT3A and DNMT3B, which exhibit both redundant and distinctive methylation activities. However, the related molecular basis remains undetermined. Through comprehensive structural, enzymology and cellular characterizations of DNMT3A and DNMT3B, here we uncovered distinct and interrelated modes-of-action underlying their CpG site and flanking sequence interaction. Strikingly, K777 of DNMT3B makes direct contacts with the DNA base at the +1 flank position of the cytosine methylation site, which contrasts with its counterpart in DNMT3A that forms base-specific contacts with the CpG site. Consequently, there is a divergent substrate and flanking sequence preference between DNMT3A and DNMT3B in vitro and in cells, thus providing an explanation for site-specific epigenomic alterations seen in ICF syndrome patients with DNMT3B mutations. Together, this study reveals crucial, yet complicated interplays of DNMT3s, DNA sequences and resultant methylation.

overall_design: To examine patterns of de novo DNA methylation induced by DNMT3A or DNMT3B in cells, we stably transduced either enzyme into mouse ES cells with compound knockout (TKO) of DNMT1, DNMT3A and DNMT3B, followed by  genome-wide methylation in cells by enhanced reduced representation bisulfite sequencing (eRRBS). eRRBS was carried out side-by-side to generate datasets for across-sample comparisons.

first: Sara

last: Grimm


title: Reduced Brd1 expression leads to reversible depression-like behaviors and gene-expression changes in female mice [Striatum (CPu)]

first: Per

last: Qvist

first: Rajkumar

last: Anto

first: Qvist

last: Per

first: Donskov

last: Julie


title: Developmental Programming: Sheep Granulosa and Theca Cell-Specific Transcriptional Regulation by Prenatal Testosterone

city: Ann Arbor

first: Kelly

last: Bakulski

state: MI

address: 1415 Washington Heights

department: Department of Epidemiology

institute: University of Michigan

summary: Prenatal testosterone treated sheep, similar to PCOS women, manifest reduced cyclicity, functional hyperandrogenism and polycystic ovary (PCO) morphology. The PCO morphology results from increased follicular recruitment and persistence of antral follicles, consequence of reduced follicular growth and atresia, and driven by cell-specific gene expression changes that are poorly understood. Therefore, utilizing RNA sequencing, cell-specific transcriptional changes were assessed in laser capture microdissection isolated antral follicular granulosa and theca cells from 21 months-of- age control and prenatal testosterone treated sheep.

overall_design: 5 control sheep and 5 prenatal testosterone treated sheep, each with a theca cell and granulosa cell sample

first: Muraly

last: Puttabyatappa

first: John

last: Dou

first: Xingzi

last: Guo

first: Daniel

last: Dumesic

middle: M

first: Vasantha

last: Padmanabhan