title: Inactivated M. tuberculosis and M.tuberculosis Infection remodels the macrophage response

summary:
Background:  Tuberculosis (TB) remains a major public health problem, especially in developing countries, with 1.5 million deaths annually worldwide. Macrophages (Mφs) are central to TB pathogenesis. Mφs play a key role in the outcome of the infection. More importantly, Mφs are the primary cell target of MTB, which has developed different strategies to multiply inside the Mφ phagosome. Here we aim to compare the transcriptome of Mycobacterium tuberculosis (MTB)-infected Mφs with that of cells stimulated with heat-killed MTB.
Results: Here, we used live and heat-inactivated MTB to examine the impact of infection on the human macrophage response. Gene expression profiling has revealed about 3,259 genes differentially expressed in macrophages incubated with heat-inactivated MTB and 4,179  genes differentially expressed with live MTB. We found that the response to live and heat-inactivated MTB was strongly correlated (r>0.78).
Conclusions: Our results show that macrophages respond in a similar manner to live and heat-inactivated MTB.  We also identified some genes differentially expressed only in one condition.

overall_design: Evaluation of host response to live MTB and heat-inactivated MTB in human macrophages.


title: Cerebellar Interpositus Nucleus Transcriptome Following Early-Life Limited Bedding Stress in P70 Male and Female Rats

status: Public on Apr 29 2020

city: Bloomington

first: Alexandra

middle: B

last: Moussa-Tooks

state: Indiana

department: Psychological and Brain Sciences

country: USA

institute: Indiana University

type: Expression profiling by high throughput sequencing

summary: Purpose: This particular experiment is part of a broader study on the effects of early-life stress on the cerebellar endocannabinoid system. Early-life stress is a major developmental risk factor. Though studies are now considering the endogenous cannabinoid system, which is highly intertwied with the stress-response, as a moderator of these effects, studies have been largely focused on cerebral effects. Cerebellum is vulnerable to stress during early development and is dense with cannabinoid receptors, making it an important target for stress effects. Moreover, relatively little is known about sex-specific effects in cerebellum. Methods: Limited bedding,  a naturalistic rodent model of early-life stress, was performed at postnatal days 2-9. Following, rats were reared normally and adult (postnatal day 70) cerebellar interpositus nucleus mRNA was assessed. Results: Transcriptomics for cerebellar interpositus nucleus revealed substantial sex effects, with minimal effects of stress. Specifically, expression of 25% of the genes evaluated significantly differed by sex alone. Interestingly, in the non-stressed groups, there were significant sex differences in Cnr1, cannabinoid receptor 1. Conclusion: We show robust sex-specific transcriptomics effects in cerebellar interpositus, perhaps suggesting a role for this system in sexual differentiation of the brain. Other regions of cerebellum should be evaluated.

overall_design: Cerebellar interpositus nucleus mRNA profiles were obtain for 70 day old Long-Evans rats with or without early-life limited bedding stress. 27 total samples were analyzed including 8 replicates of males having undergone limited bedding stress, 7 replicates of females having undergone limited bedding stress, 7 replicates of control males reared normally, and 6 replicates of control females reared normally.


title: RNA-sequencing identifies factors that contribute to the Dengue-Refractory or –Susceptible phenotype in field-derived strains of Aedes aegypti

status: Public on May 01 2020

city: Burnaby

first: Heather

last: Coatsworth

state: BC

address: 8888 University Drive

department: Biological Sciences

country: Canada

institute: Simon Fraser University

summary: We used RNA-sequencing to identify differentially expressed genes in the midgut of Aedes aegypti that contribute to the field derivied dengue susceptible (Cali-S) and dengue refractory (Cali-R) phenotypes

overall_design: We employed a full factorial design, analyzing differential expression across 1) time (24, 36 and 48 hours post blood meal), 2) feeding condition (blood or blood + DENvDENV2) and 3) strain (Cali-S (=susceptible) and Cali-MIB (=refractory)). Single end reads from 12 biological singetons were analyzed.

middle: G

first: Paola

middle: A

last: Caicedo

first: Geoffrey

middle: L

last: Winsor

first: Fiona

middle: S

last: Brinkman

first: Clara

last: Ocampo

first: Carl

last: Lowenberger


title: Role of cell wall proteins in Saccharomyces cerevisiae’s response to commercial formulations of glyphosate and contribution of Ty elements to adaptation mechanisms

city: Morgantown

first: Apoorva

last: Ravi Shankar

state: WV

address: 53 Campus Drive

department: Biology

institute: West Virginia University

summary: The use of glyphosate-based herbicides is widespread, despite its extensive use its effects are yet to be deciphered completely. The additives in commercial formulations of glyphosate though labelled inert when used individually, has adverse effects when used in combination with other additives and the active ingredient. An RNA seq study was performed to thoroughly investigate the changes that occur in a sensitive and a resistant strain at the transcriptome level on exposure to a commercial formulation of glyphosate. Changes in gene expression of genes contributing to numerous pathways involved in crucial cellular functions such as DNA replication, MAPK signaling, meiosis, cell wall synthesis etc. were found. Common fragile sites were found to play a role in adaptation mechanisms used by cells to attain resistance on long term exposure to a CFG. The cell wall structure acts as a protective barrier in alleviating the stress caused by exposure to Cr41, the thicker the cell wall the more resistant the cell is against CFG.  Hence, a detailed study of the changes occurring at the genome and transcriptome level Is essential to better understand the possible effects of CFG on the cell as a whole.

overall_design: mRNA profiles of GSY147(S288c backgroud) and RM11, in YM or YM supplemented with WYF, treated or not with Cr41, 5 replicates, using Illumina Hiseq.

first: Jennifer

last: Gallagher

last: Ravishankar

first: Amaury

last: Pupo


title: Transcriptomic analysis of endoplasmic reticulum stress (ERS) regulated genes in mouse gastrocnemius muscle

city: Lille

first: Jérôme

last: Eeckhoute

address: Bâtiment J&K, Faculté de Médecine de Lille-Pôle Recherche, Boulevard du Professeur Leclerc

country: France

institute: CNRS

summary: One gastrocnemius muscle was injected with the ERS-drug tunicamycin, the other with control saline solution and muscles were used to prepare RNA.

overall_design: 9 mice were used for injections and RNA was prepared from gastrocnemius muscles and then processed for analysis using MoGene-2_0-st Affymetrix microarrays according to standard procedures.

first: Steve

last: Lancel

first: Hélène

last: Duez

first: Christian

last: Duhem

first: Alexis

last: Boulinguiez


title: Transcriptomic analysis of endoplasmic reticulum stress (ERS) regulated genes in mouse primary hepatocytes.

summary: Mouse primary hepatocytes (MPH) prepared from C57Bl6 male adult mice were treated with the ERS-drug thapsigargin and used to prepare RNA.

overall_design: 3 independent preparations of MPH were performed and used to prepare RNA which was then processed for analysis using MoGene-2_0-st Affymetrix microarrays according to standard procedures.

first: Vanessa

last: Dubois

first: Jerome


title: Transcriptomic and epigenomic analysis of endoplasmic reticulum stress signaling in mouse primary hepatocytes and liver

type: Genome binding/occupancy profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: ChIP-seq analysis of changes in H3K27ac by endoplasmic reticulum stress (ERS) in mouse primary hepatocytes.

summary: Mouse primary hepatocytes (MPH) prepared from C57Bl6 male adult mice were treated with the ERS-drug thapsigargin and ChIP-seq was performed using an H3K27ac antibody (Active Motif ref 39685).

overall_design: 3 independent preparations of MPH were performed and used to perform H3K27ac ChIP-seq.


title: Transcriptomic analysis of endoplasmic reticulum stress (ERS) regulated genes in the liver of NFIL3 KO mice.

summary: Livers from wild-type (WT) or NFIL3 knock-out (NFIL3 KO) C57Bl6 mal adult mice treated with the ERS-drug tunicamycin were used to prepare RNA.

overall_design: Livers from 10 WT and 10 NFIL3 KO adult male mice (5 treated with vehicle and 5 treated with tunicamycin for each genotype) were used to prepare RNA which was then processed for analysis using MoGene-2_0-st Affymetrix microarrays according to standard procedures.


title: Overexpression of the small RNA PA0805.1 modulates the expression of a large set of genes and proteins, resulting in altered motility, cytotoxicity and tobramycin resistance

status: Public on Apr 28 2020

city: Vancouver

first: Shannon

last: Coleman

address: 2259 Lower Mall

institute: University of British Columbia

summary:
Purpose:  Pseudomonas aeruginosa is a motile species that initiates swarming motility in response to specific environmental cues, i.e., a semisolid surface with amino acids as a nitrogen source (relevant to the human lung). Swarming is an intricately regulated process, but to date post-transcriptional regulation has not been extensively investigated. Small non-coding RNAs (sRNA) are hypothesized to play post-transcriptional regulatory roles, largely through suppression of translation, and we previously demonstrated 20 sRNA species that were dysregulated under swarming conditions. Overexpressing these revealed one sRNA, PA0805.1,  that when cloned, transformed into PAO1 WT and overexpressed led to broad phenotypic changes.
Methods: The sRNA PA0805.1 was cloned and overexpressed in PAO1 WT. Phenotypic screens including motility, cytotoxicity and adherence assays were performed. RNA-Seq and proteomics were performed under swarming conditions compared to the empty vector strain to discover which genes were influenced by the overexpression of PA0805.1
Results:  Phenotypic screens revealed that PA0805.1 resulted in reduced swarming, swimming and twitching motility, as well as increased adherence, cytotoxicity, and tobramycin resistance. A deletion mutant ∆PA0805.1 was supersusceptible to tobramycin under swarming conditions. The strain overexpressing PA0805.1 was compared to the empty vector strain by RNA-Seq and proteomics under swarming conditions to determine sRNA targets. Broad transcriptional and proteomic profiles were revealed, encompassing 1121 differentially expressed genes and 258 proteins with significantly different abundance. Importantly, these included 106 transcriptional regulators, two-component regulatory systems, sigma and anti-sigma factors. Downstream of these regulators were found downregulated type IV pilus genes, many upregulated adherence and virulence factors, and two multidrug efflux systems, mexXY and mexGHI-opmD.
Conclusions:  The sRNA PA0805.1 appears to regulate diverse bacterial lifestyles, most likely through a regulatory cascade.

overall_design: Five biological replicates of the wildtype strain overexpressing PA0805.1 were compared to five biological replicates of the wildtype with the empty vector as a control

middle: R

first: Maren

last: Smith

first: Victor

last: Spicer

first: Ying

last: Lao

first: Neeloffer

last: Mookherjee

first: Robert

middle: E

last: Hancock


title: Chromatin isolation by RNA immunoprecipitation (ChIRP) for the long non-coding RNA Pvt1

city: Padova

first: Gerolamo

last: Lanfranchi

address: Via U. Bassi, 58/B

department: CRIBI - Biotechnology Center and Biology Department

country: Italy

institute: University of Padova

summary:
Long non-coding RNAs (lncRNAs) are emerging as important players in the regulation of several aspects of cellular biology. For a better comprehension of their function it is fundamental to determine their expression in single cells, to identify their subcellular localization and eventually DNA interacting regions. In fact, lncRNAs present a cell-type specific expression and different functions depending on their subcellular localization. C2C12 cells are a model to study muscle pathophysiology and differentiation. We used this model to evaluate Pvt1-DNA interacting regions.  We demonstrated that the lncRNA Pvt1, early activated during muscle atrophy, regulate the transcription of DNA and influence mitochondrial respiration and morphology ultimately impinging mito/autophagy and myofiber size in vivo.
We evidenced that the long non-coding RNA Pvt1 is highly expressed during skeletal muscle atrophy. Moreover, it is preferentially expressed in fast contracting myofibers. By using high throughput techniques and Fluorescent In Situ Hybridization we evidenced that Pvt1 localize inside the nucleai of myofibers and C2C12 cells. C2C12 cells are widely used as in vitro model to study different aspects of muscle biology such as development, differentiation and metabolism. Because Pvt1 nuclear localization we checked its capacity to interact with DNA by Chromatin isolation by RNA purification (ChIRP).

overall_design: We performed the Pvt1 precipitation on C2C12 cells, crosslinked with glutaraldehyde, using 18 biotinilated oligonucleotides spanned along the entaire Pvt1 RNA sequence. Before precipitation we fragmented DNA by sonication at the dimension of ~50-130 bp. We subdivided biotinilated oligonucleotides in odd and even probes to perform two different experimental replicates. Replicates were also performed in two different biological samples. Precipitated samples were processed for RNA and DNA extraction to confirm the enrichment in Pvt1 RNA in the precipitated sample and to evaluate DNA associated fragments. Sequencing runs were performed on PGM Ion torrent producing libraries according to Accel-NGS DNA library kit (Swift Bioscience). Libraries were quantized using the Lib Quant Kit Ion/Rox Low (Roches) and used in the Ion One touch (Thermo Fisher Scientific) for the emulsion PCR as described by the manufacturer. Produced sequences were processed by the Ion Torrent server for the adaptors trimming and their alignement against mouse genome (mm10). Alignement was performed using The Torrent Mapping Alignment Program (TMAP) with default parameters. The software MACS with default settings (macs2 version 2.1.0) was used to identify Pvt1-DNA interaction regions. As backgroud for the peacks identification was used unprecipitated DNA. Unprecipitated DNA was fragmented by Bioruptor UCD-200. having the average dimension of 400 bp. The binding effect were evaluated on the genes that have a peak in the region 2000 bp upstream of the TSS.

first: Enrico

last: Alessio

first: Lisa

last: Buson

first: Francesco

last: Chemello

first: Francesca

last: Grespi

first: Caterina

last: Millino

first: Beniamina

last: Pacchioni

first: Paolo

last: Martini

first: Maria

last: Massimino

first: Chiara

last: Romualdi

first: Alessandro

last: Bertoli

first: Luca

last: Scorrano

first: Stefano

last: Cagnin


title: CDK11 is required for transcription of replication-dependent histone genes

city: Brno

first: Dalibor

last: Blazek

address: Studentska 1, Building A35

department: Molecular Medicine

country: Czech Republic

institute: Central European Institute of Technology (CEITEC), Masaryk University

summary: Replication-dependent histones (RDH) are required for packaging of newly synthetized DNA into nucleosomes during the S phase when their expression is highly upregulated. However, the mechanisms of this upregulation in metazoan cells remain poorly understood. Using iCLIP and ChIP–seq, we found that human cyclin-dependent kinase 11 (CDK11) associates with RNA and chromatin of RDH genes primarily in the S phase. Moreover, its amino-terminal region binds FLASH, RDH-specific 3′-end processing factor, which keeps the kinase on the chromatin. CDK11 phosphorylates serine 2 (Ser2) of the carboxy-terminal domain of RNA polymerase II (RNAPII), which is initiated at the middle of RDH genes and is required for further RNAPII elongation and 3′-end processing. CDK11 depletion leads to decreased number of cells in S phase, likely owing to the function of CDK11 in RDH gene expression. Thus, the reliance of RDH expression on CDK11 could explain why CDK11 is essential for the growth of many cancers.

overall_design: Examination of RNA-seq, CDK11 ChIP, CDK11 iCLIP, P-Ser2 ChIP and RNAPolII ChIP in the presence or absence of CDK11 on HCT116 cells

first: Pavla

last: Gajduskova

first: Igor

last: Ruiz de los Mozos

first: Michal

last: Rajecky

first: Milan

last: Hluchy

first: Jernej

last: Ule


title: Histone modifications in livers from wild-type male and female mice, and in livers from mice with a Double Knockout (DKO) of Ezh1 and Ezh2

city: Boston

first: David

middle: J.

last: Waxman

state: MA

address: 5 Cummington Mall

department: Department of Biology and Bioinformatics Program

institute: Boston University

summary: Chromatin immunoprecipitation and sequencing for 3 histone marks (H3K27ac, H3K27me3 and H3K4me1) was performed on livers of male and female mice with a combined loss of Ezh1 and Ezh2. The DKO mouse model used in these analyses is a global deletion of Ezh1 and hepatocyte-specific deletion of Ezh2, and is described in Bae WK et al, FASEB J. 2015 May;29(5):1653-62. doi: 10.1096/fj.14-261537. PMID: 25477280.

overall_design: Chromatin was isolated from frozen livers from 7 week old male and female wild-type and double knockout (DKO; Ezh1-/-, Ezh2-f/f_Alb-Cre) mice and their floxed (f) controls.  Sonicated chromatin from 3-4 individual livers from each group was immunoprecipitated with the following antibodies: H3K27ac (Abcam Cat # ab4729, 3 μg antibody per 15 μg of sonicated chromatin); H3K4me1 (Abcam Cat # ab8895, 1.2 μg antibody per 15 μg of sonicated chromatin); H3K27me3 (Abcam Cat # ab6002, 2 μg antibody per 10 μg of sonicated chromatin); and normal rabbit IgG (Santa Cruz, Cat # sc-2027, 3 μg antibody per 15 μg of sonicated chromatin).

first: Dana

last: Lau-Corona

middle: J


title: Partitioning of cancer therapeutics in nuclear condensates

status: Public on Apr 27 2020

city: Cambridge

first: Richard

last: Young

address: 9 Cambridge Center

institute: Whitehead Institute for Biomedical Research

summary: The nucleus contains diverse phase-separated condensates that compartmentalize and concentrate biomolecules with distinct physicochemical properties. Here we consider whether condensates concentrate small molecule cancer therapeutics such that their pharmacodynamic  properties are altered. We found that antineoplastic drugs become concentrated in specific protein condensates in vitro and that this occurs through physicochemical properties independent of the drug target. This behavior was also observed in tumor cells, where drug partitioning influenced drug activity. Altering the properties of the condensate was found to impact the concentration and activity of drugs. These results suggest that selective partitioning and concentration of small molecules within condensates contributes to drug pharmacodynamics and that further understanding of this phenomenon may facilitate advances in disease therapy.

overall_design: ChIP-seq for MED1 in HCT116 cells treated with 1uM JQ1 for 24 hours + 50uM Cisplatin for 6 hours. The cells were control treated with DMSO for 24 hours + DMF for 6 hours.

first: Isaac

last: Klein

first: Ann

last: Boija

first: Lena

middle: K

last: Afeyan

first: Susana

middle: W

last: Hawken

first: Menyang

last: Fan

first: Alessandra

last: Dall'Agnese

first: Ozgur

last: Oksuz

first: Jonathan

last: Henninger

first: Krishna

last: Shrivas

first: Benjamin

last: Sabari

first: Ido

last: Sagi

first: Victoria

last: Clark

first: Jess

middle: M

last: Platt

first: Mrityunjoy

last: Kar

first: Patrick

last: McCall

first: Alicia

middle: V

last: Zamudio

first: John

middle: C

last: Manteiga

first: Eliot

last: Coffey

first: Charles

middle: H

last: Li

first: Nancy

last: Hannet

first: Yang

last: Guo

first: Tim-Michael

last: Decker

first: Tong

middle: I

last: Lee

first: Tinghu

last: Zhang

first: Jing-ke

last: Weng

first: Dylaan

last: Taatjes

first: Phillip

last: Sharp

first: Young

middle: T

last: Chang

first: Anthony

last: Hyman

first: Nathanael

last: Gray


title: Squamous trans-differentiation of pancreatic cancer cells promotes stromal inflammation

city: Cold Spring Harbor

first: Tim

middle: D D

last: Somerville

state: NY

address: 1 Bungtown Road

department: CSHL

institute: Cold Spring Harbor Laboratory

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus


title: Squamous trans-differentiation of pancreatic cancer cells promotes stromal inflammation (ChIP-seq)

summary: A highly aggressive subset of pancreatic ductal adenocarcinomas undergo trans-differentiation into the squamous lineage during disease progression. While the tumorigenic consequences of this aberrant cell fate transition are poorly understood, recent studies have identified a role for the master regulator TP63 in this process. Here, we investigated whether squamous trans- differentiation of pancreatic cancer cells can influence the phenotype of non-neoplastic cells in the tumor microenvironment. Conditioned media experiments revealed that squamous-subtype pancreatic cancer cells secrete factors that convert quiescent pancreatic stellate cells into a specialized subtype of cancer-associated fibroblasts (CAFs) that express inflammatory genes at high levels. We use gain- and loss-of-function approaches in vivo to show that squamous-subtype pancreatic tumor models become enriched with inflammatory CAFs and neutrophils in a TP63- dependent manner. These non cell-autonomous effects occur, at least in part, through TP63- mediated activation of enhancers at pro-inflammatory cytokine loci, which includes IL1A as a key target. Taken together, our findings reveal enhanced tissue inflammation as a consequence of squamous trans-differentiation in pancreatic cancer, thus highlighting an instructive role of tumor cell lineage in reprogramming the stromal microenvironment.

overall_design: H3K27ac ChIP-seq was performed using antibody recognizing endogenous protein with input genomic DNA as control with/without transduction of sgNEG/sgTP63#2/sgTP63#4 (BxPC3 cells) or MTV/TP63ΔN (SUIT2 cells). TP63 ChIP-seq in BxPC3 cells was performed using antibody recognizing endogenous protein with input genomic DNA as control. FLAG-TP63 ChIP-seq in SUIT2 cells was performed in FLAG-TP63 expressing cells with input genomic DNA as control.

middle: D

first: Christopher

last: Vakoc


title: Squamous trans-differentiation of pancreatic cancer cells promotes stromal inflammation (RNA-seq)

overall_design: For RNA-seq performed in cells harvested from in vivo experiments (flow sorted or xenograft RNA-seq), four biological repeats were used for each experimental group. For RNA-seq performed following dox-inducible knockout of TP63 in T3M4 and KLM1 cells, three technical repeats were performed for each condition. For RNA-seq performed in human PDA cell lines or mouse PSCs following treatment with conditioned media, 1 technical repeat for each cell line was used.


title: Iterative single cell analyses define the transcriptome of the first functional hematopoietic stem cells

city: Shanghai

first: Xiaonan

last: Wang

address: 227 Chongqing South Road

department: School of Public Health

country: China

institute: Shanghai Jiao Tong University School of Medicine

summary: Whereas hundreds of endothelial cells in the aorta-gonad-mesonephros (AGM) region transdifferentiate to hematopoietic cells, only 1-2 of these cells take on hematopoietic stem cell (HSC) identity at a single time point. Gata2 is one of a few transcription factors essential for HSC generation and function. In contrast to cell surface marker-based HSC enrichment approaches, enrichment based on Gata2 expression is directly linked to the internal regulatory network critical for HSC identity.  Here we use index sorting of highly enriched Gata2-expressing intra-aortic hematopoietic cluster (IAHC) cells, iterative single cell transcriptomics and functional analyses to examine the relationship between HSC identity and expression of pivotal factors. Functional HSCs are enriched in Gata2 medium-expressing IAHC cells, which can be further subdivided based on Gata2, cKit and CD27 expression, where each subpopulation is not only associated with a specific molecular profile, but also distinct functionality in both in vitro and in vivo stem/progenitor assays. Immunohistochemical analysis localizes the first putative functional HSCs in vivo to small clusters in the AGM region, thus integrating molecular, functional and spatial information about the first cells in a mammalian embryo that acquire full HSC identity.

overall_design: Single cell RNA sequencing of Gata2-expressing intra-aortic hematopoietic cluster (IAHC) cells

first: C

last: Vink

first: F

last: Calero-Nieto

first: A

last: Maglitto

first: X

first: S

last: A Mariani

first: W

last: Jawaid

first: B

last: Gottgens

first: E

last: Dziezak


title: Single-cell ftATACseq data of mouse thymocytes

city: Hefei

first: Bin

last: Li

address: NO. 96 Jinzhai Road

institute: University of Science and Technology of China

summary: This dataset is related to our recent study "APEC: an accesson-based method for single-cell chromatin accessibility analysis". We adopted a fluorescent tagmentation-based single-cell ATAC-seq technique (ftATAC-seq) to investigated the per cell regulome dynamics of mouse thymocytes. Associated with ftATAC-seq, APEC revealed a detailed epigenomic heterogeneity of thymocytes, characterized the developmental trajectory and predicted the regulators that control the stages of maturation process.

overall_design: To evaluate the performance of this approach in the context of the immune system, we adopted ftATAC-seq technology to investigate the regulome dynamics of the thymic development process. Coordinated with essential cell surface markers, we provided a in-depth classification of thymocytes than the conventional DN, DP, CD4SP and CD8SP stages based on single-cell chromatin status. By reconstructing the developmental pseudotime trajectory, we discovered a transitional stage before thymocytes bifurcate into CD4SP and CD8SP cells and inferred that one of the stages leads to cell apoptosis. Considering that more than 95% of DP cells undergo apoptosis as a programmed cell death process, our data suggested that before DP cells enter the final apoptotic state, there would already be some intracellular changes towards apoptosis at the chromatin level.

first: Kun

last: Qu


title: A complex regulatory landscape involved in the development of external genitals [ChIP-Seq]

status: Public on Apr 25 2020

city: Lausanne

first: Ana Rita

middle: Amandio

last: Lhopitallier

address: SV2842, Station 19

department: SV-ISREC-EPFL

country: Switzerland

institute: EPFL

summary: In vertebrates, developmental genes are often controlled by large regulatory landscapes matching the dimensions of topologically associating domains (TADs). In various ontogenic contexts, the associated constitutive chromatin backbone is modified by fine-tuned specific variations in enhancer-enhancer and enhancer-promoter interaction profiles. In this work, we use a TAD flanking the HoxD gene cluster as a paradigm to address the question of how these complex regulatory architectures are formed and how they are de-constructed once their function has been achieved. We suggest that this TAD can be considered as a coherent functional unit in itself, with several regulatory sequences acting together to elicit a transcriptional response. With one notable exception, the deletion of each of these sequences in isolation did not produce any substantial modification in the global transcriptional outcome of the system, a result at odds with a conventional view of long-range enhancer function. Likewise, both the deletion and inversion of a CTCF site located in a region rich in such sequences did not affect transcription of the target gene. In the latter case, however, slight modifications were observed in interaction profiles in vivo in agreement with the loop extrusion model, despite no apparent functional consequences. We discuss these unexpected results by considering both conventional explanations and an alternative possibility whereby a rather unspecific accumulation of particular factors within the TAD backbone may have a global impact upon transcription.

overall_design: ChIP-seq analysis of H3K27ac and H3K27me3 in the genital tubercle (GT) or the cloaca region (CR) of wildtype mouse embryos. CTCF ChIP-seq analysis of wildtype or del(V) mouse GT.

first: Ana

first: Denis

last: Duboule


title: A complex regulatory landscape involved in the development of external genitals [RNA-Seq]

overall_design: RNA-seq analysis of genital tubercle tissue from E16.5 and E18.5 mouse embryos


title: Gene expression and histone marks in EZH1/EZH2 double knockout mouse liver

first: David

last: Waxman


title: A complex regulatory landscape involved in the development of external genitals [ATAC-Seq]

overall_design: ATAC-seq analysis of the cloaca region of wildtype mouse embryos or the genital tubercle of wildtype or mutant mouse embryos


title: RNA-seq gene expression profiling of livers from wild-type male and female mice, and from mice with a Double Knockout (DKO) of Ezh1 and Ezh2.

summary: PolyA-selected RNA isolated from livers of adult male and female mice with a combined loss of EZH1 and EZH2 in hepatocytes was analyzed by RNA-seq. The DKO mouse model used in these analyses is a global deletion of Ezh1 and hepatocyte-specific deletion of Ezh2, and is described in Bae WK, Kang K, Yu JH, Yoo KH, Factor VM, Kaji K, Matter M, Thorgeirsson S, Hennighausen L. FASEB J. 2015 May;29(5):1653-62. doi: 10.1096/fj.14-261537. PMID: 25477280

overall_design: Liver RNA, polyA-selected, was isolated from 7 week old male and female DKO mice (Ezh1-/-, Ezh2-f/f_Alb-Cre) and their floxed (f) controls.  RNA samples from n=3 mouse livers were pooled to make each of 3 biological replicate libraries per sex and genotype. AC, Albumin-Cre

first: Dana

last: Lau-Corona


title: Single-Cell transcriptomes and immune profiling of IgG+ murine memory B cells from Bone Marrow and Spleen.

status: Public on Apr 24 2020

city: Berlin

first: Pawel

last: Durek

address: Charitéplatz 1

country: Germany

institute: Deutsches Rheuma-Forschungszentrum

summary: Single cell sequencing for analysis gene of gene transcription and Ig repertoires of isotype-switched IgG-expressing memory B cells for paired analysis with Ig repertoires in the same cells. Aim of the study was to explore heterogeneity of isotype-switched memory B cells within and between spleen and bone marrow compartments.

overall_design: Sequencing of RNA and Ig was performed in duplicates of IgG+ B cells isolated from spleen and bone marrow of C57BL/6 mice.

first: René

last: Riedel

first: Richard

last: Addo

first: Gitta

middle: A

last: Heinz

first: Frederik

last: Heinrich

first: Mir-Farzin

last: Mashreghi

first: Andreas

last: Radbruch


title: Clinical significance of gene signature generated from the grade of tumor budding in colorectal cancer

city: Kobe

first: Yuichiro

last: Yoshida

address: 4-4-4 Takatsukadai Nishi-ku

department: central research laboratories

country: Japan

institute: Sysmex corporation

summary:
Tumor budding is a definite prognostic marker, but does not always show favorable reproducibility.
We developed a new molecular signature related to tumor budding levels, a microscopic finding of dedifferentiation, using RNA extracted from the whole tissue section and disclosed its relevance to a high recurrence risk using an inhouse dataset and independent public datasets.

overall_design: mRNA from 85 primary colorectal tumour samples were extracted and hybridized to HG-U133Plus 2.0 expression arrays.  For three samples, the cancer areas at the invasive front of the tumor and  the tumor center were dissected and subjected to DNA microarray measurement Each data was normalized with Microarray Suite (MAS) 5.0.

first: Eiji

last: Shinto

first: Yoshiki

last: Kajiwara

first: Koichi

last: Okamoto

first: Satsuki

last: Mochizuki

first: Masato

last: Yamadera

first: Takehiro

last: Shiraishi

first: Ken

last: Nagata

first: Hitoshi

last: Tsuda

first: Kazuo

last: Hase

first: Hideki

last: Ueno


title: Genome-wide DNA methylation analysis in Williams syndrome (WS patients vs Controls)

city: Kyoto

first: Ryo

last: Kimura

address: Yoshida-Konoe-cho

department: Anatomy and Developmental Biology

institute: Kyoto University Graduate School of Medicine

type: Methylation profiling by genome tiling array

summary: Genome-wide DNA methylation profiling of WS patients and controls using peripheral blood samples.

overall_design: Bisulfite converted DNA was hybridized onto the Illumina Infinium HumanMethylation450 BeadChip array

first: Masatoshi

last: Hagiwara


title: WNT-signaling cells polarize inhibitors to protect their identity and fate

city: New York

first: Irina

last: Matos

address: 1230 York Av

institute: The Rockefeller University

summary: We demonstrated that during development WNT signaling cells produce their own inhibitors polarizing them apically to preserve their own identity

overall_design: Transcriptomes of WNT high cells purified from E14.5 APCcKO and APCHet embryos and from WNT-signaling cells from E17.5 developing placodes. Epidermal keratinocytes were purified from embryonic mouse backskin via FACS sorting RNA, was extracted, cDNA libraries created and RNA sequencing was run on the samples.

first: Amma

last: Asare

first: John

last: Levorse

first: Tamara

last: Ouspenskaia

first: June

last: de la Cruz-Racelis

first: Laura-Nadine

last: Schuhmacher

first: Elaine

last: Fuchs


title: Single-Cell transcriptomes and BCR sequences of IgG+ murine memory B cells from Bone Marrow and Spleen

first: Pawel

last: Durek

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

overall_design: Refer to individual Series


title: Ig repertoire of IgG+ murine memory B cells

summary: BCR repertoire sequencing of isotype-switched memory B cells Aim of the study was to explore the differences of Ig repertoires of isotype-switched memory B cells between spleen and bone marrow compartments.

overall_design: Cells from BM (tibiae, femurae and pelvic bones) and spleen of three times NP-CGG immunized-mice were isolated. Sorted cells of Mouse 1 and 2 were split into two equal aliquots (samples A, B) for BM and spleen samples as cellular (biological) replicates. Sorted cells of Mouse 3 were split into 4 samples (A, B, C, D). Biological replicates were processed independently from this point on.  Isolated RNA was split and library preparation was performed in technical duplicates.

first: René

last: Riedel

first: Richard

last: Addo

first: Gitta

last: Heinz

first: Jannis

last: Kummer

first: Frederik

last: Heinrich

first: Mir-Farzin

last: Mashreghi

first: Andreas

last: Radbruch


title: Next Generation Sequencing Facilitates Quantitative Analysis of Transcriptomics in children with Cryptorchidism

status: Public on Apr 23 2020

city: Nantong

first: Yunzhao

last: Xu

state: China

address: Qixiu Road

department: Prenatal Diagnosis Center

institute: Affiliated Hospital of Nantong University

type: Non-coding RNA profiling by high throughput sequencing

summary: Cryptorchidism is the most common congenital defect in children's genitourinary system. It is one of the risk factors for male infertility and testicular tumors. The occurrence of cryptorchidism is closely related to many factors, and the alteration of genes plays an important role in the occurrence of cryptorchidism. A large number of studies have confirmed that the formation of cryptorchidism is related to the differential expression of genes. Therefore, exploring new differential expression genes of cryptorchidism from the transcriptome level and studying the relationship between susceptibility genes and children with cryptorchidism can provide a new direction for studying the pathogenesis of cryptorchidism.

overall_design: In order to check the difference between the genome-wide transcriptional profile of children with cryptorchidism and in normal control testis, we separately examined testicular tissue including the samples of 6 cases of children with cryptorchidism and 2 cases of testicular trauma children and 1 case of incarcerated inguinal hernias. The transcriptome sequencing of the 9 testicular tissues was performed by RNA-seq technique.

first: Wenliang

last: Ge

first: Hua

last: Xian


title: Transcriptome analysis of control and ERK3-depleted Human Colonic Primary Epithelial cells (HCPECs)

city: Mainz

first: Federico

last: Marini

address: Obere Zahlbacher Strasse 69

department: Institute of Medical Biostatistics, Epidemiology and Informatics (IMBEI)

institute: University Medical Center - Mainz

summary:
RNA sequencing analysis of control and ERK3 knockdown  HCPECs was performed to understand the physiological relevance of ERK3 in maintaining basal as well as LPS-mediated innate immune responses in HCPECs. Results revealed that ERK3 is required for the regulation of several key cytokines and chemokines even at the steady state.
In particular, we discovered that ERK3 is indespansable for the transcriptional regulation of CXCL8/IL-8, CXCL10, MCP-1/CCL2 and GCP-2/CXCL6. Concomitantly, a functional interpretation of the differentially expressed (DE) genes with topGO indicated an important role of ERK3 in regulation of genes involved in, among others, immune responses and leukocyte chemotaxis.

overall_design: Three biological replicates of Control (siCo) and ERK3-depleted (siERK3) HCPECs were challenged with LPS for 24 h. After the stimulation cells were washed with cold PBS and lysed with Trizol (Cat# 15596018, Ambion) according to the manufacturer’s instructions. Total RNA was quantified by a Qubit 2.0 fluorometer (Invitrogen) and quality was assessed using Agilent’s bioanalyzer 2100 and a RNA 6000 Nano chip (Agilent). Samples with RNA integrity number (RIN) > 8 were further subjected for RNA library preparation. Barcoded cDNA libraries were prepared from 300 ng of total RNA using the NEBnext Poly(A) mRNA Magnetic Isolation Module and NEBNext Ultra RNA Library Prep Kit for Illumina (NEB) according to the provided instruction. Library quantity was assessed on a Qubit 2.0 using Invitrogen’s Qubit HS assay kit and library size was determined using Agilent’s Bioanalyzer 2100 and a HS DNA assay chip. Barcoded RNA-Seq libraries were onboard clustered using HiSeq Rapid SR Cluster Kit v2 using 8pM and 59 bps were sequenced on an Illumina HiSeq2500 using a HiSeq Rapid SBS kit v2.

first: Katarzyna

last: Bogucka

first: Krishnaraj

last: Rajalingam


title: A complex regulatory landscape involved in the development of external genitals [4C-seq]

first: Ana Rita

last: Lhopitallier

overall_design: 4C-seq analysis of 3D chromatin interactions in different wild type and mutant mouse embryonic tissues

first: Ana

first: Denis

last: Duboule


title: Sampling time-dependent artifacts in single-cell genomics studies

city: Barcelona

first: Gustavo

last: Rodriguez-Esteban

state: Catalonia

address: C. Baldiri Reixac, 4; Barcelona Science Park

department: Development

country: Spain

institute: Centro Nacional de Análisis Genómico

summary: Robust protocols and automation now enable large-scale single-cell RNA and ATAC sequencing experiments and their application on biobank and clinical cohorts. However, technical biases introduced during sample acquisition can hinder solid, reproducible results and a systematic benchmarking is required before entering large-scale data production. Here, we report the existence and extent of gene expression and chromatin accessibility artifacts introduced during sampling and identify experimental and computational solutions for their prevention.

overall_design: We designed benchmarking experiments to systematically test the effect of varying processing times on single-cell transcriptome and epigenome profiles from healthy and diseased donors, while controlling for technical variability (e.g. batch effects). We isolated peripheral blood mononuclear cells from healthy donors (PBMC) and from patients affected with chronic lymphocytic leukemia (CLL), the most common adult leukemia in the Western world. Samples were either preserved immediately (0 hours) or after 2, 4, 6, 8, 24 and 48 hours; simulating common scenarios in biobank and clinical routines. Single-cell 3`-transcript counting, full-length transcriptome and scATAC-seq were performed to monitor gene expression, RNA integrity and open chromatin variance across preservation time points.

first: Holger

last: Heyn

last: Rodríguez-Esteban


title: Seasonal changes of gene expression of whole brain in Medaka fish (Oryzias latipes) SC

city: Nagoya

first: Takashi

last: Yoshimura

address: Furo-cho, Chikusa-ku

institute: Nagoya University

summary: To examine the seasonal transcriptional landscape of medaka brain, we compared the gene expression of whole brain under SC (short-day and cool temperature conditions: 10 h light/14 h dark and 10 °C) and LW (long-day and warm temperature conditions: 14 h light/10 h dark and 26 °C) conditions.

overall_design: Adult male medaka kept under SC conditions either reimained in SC or were transferred into LW conditions. Two weeks after transfer, whole brains were collected in the middle of the light phase.


title: Gene expression of whole brain between DMSO- and celastrol-treated medaka

summary: We compared the gene expression of whole brain between DMSO- and celastrol-treated medaka.

overall_design: Adult male medaka kept under SC conditions were treated with DMSO or celastrol. Twenty-four hours after the treatment, whole brains were collected in the middle of the light phase.


title: Seasonal changes of gene expression of whole brain in Medaka fish (Oryzias latipes) LW

overall_design: Adult male medaka kept under LW conditions either reimained in LW or were transferred into SC conditions. Two weeks after transfer, whole brains were collected in the middle of the light phase.


title: Transmitting tract transcriptome profile in the quadruple mutant stk ces-4 bee1 bee3 compared to wild type

city: Milan

first: Matteo

last: Chiara

address: via celoria 26

department: Biosciences

institute: University of Milan

summary: This experiment permit to obtain new regulatory genes downstream of STK-CES, CES-BEE1-BEE3 protein complexes, involved in various aspects of transmitting tract development

overall_design: Inflorescence till stage 12 of pistil development were used for RNA extraction in both quadruple and wild type samples

first: Maurizio

last: Di Marzo


title: Seasonal transcriptional landscape of medaka brain


title: Small-Molecule PAPD5 Inhibitors Restore Telomerase Activity in Patient Stem Cells

status: Public on Apr 21 2020

city: Baltimore

first: Emily

last: Lo

state: MD

address: 733 N Broadway

institute: Johns Hopkins

summary: Genetic lesions that reduce telomerase cause a range of incurable diseases including dyskeratosis congenita (DC) and pulmonary fibrosis (PF), and restoring telomere length in these patients would be curative. Ectopic expression of telomerase reverse transcriptase (TERT) risks cellular immortalization, and how to target telomerase in stem cells throughout the body remains unclear. Here we describe a successful screen for small molecules that augment TERC, the non-coding telomerase RNA component, and thereby specifically elongate telomeres in stem cells. PAPD5 is a non-canonical polymerase that oligo-adenylates and destabilizes TERC. Using a high-throughput screen we identify BCH001, a specific PAPD5 inhibitor that decreases TERC 3′- oligo(A) tailing and increases telomerase activity and telomere length by thousands of nucleotides in DC patient induced pluripotent stem cells (iPSCs). BCH001 does not result in immortalization or telomere elongation in somatic cells which lack TERT, establishing a favorable safety profile. When human hematopoietic stem and progenitor cells (HSPCs) engineered by CRISPR-Cas9 to carry PARN mutations that cause DC and PF are xenotransplanted into immunodeficient mice, oral treatment with PAPD5 inhibitors rescues TERC 3′-end maturation and telomere length. Our data demonstrate telomere restoration in human stem cells in vivo using small molecules.

overall_design: Investigation of a small molecule PAPD5 inhibitor in iPSCs from 3 PARN mutant patients and 3 WT controls

first: Neha

last: Nagpal

first: Jianing

last: Wang

first: Jing

last: Zeng

first: Diane

last: Moon

first: Kevin

last: Luk

first: Roman

middle: O

last: Braun

first: Lauri

last: Burroughs

first: Sioban

middle: B

last: Keel

first: Christopher

last: Reilly

first: R

last: Lindsley

first: Scot

last: Wolfe

first: Albert

last: Tai

first: Patrick

last: Cahan

first: Daniel

last: Bauer

first: Yick

last: Fong

first: Suneet

last: Agarwal


title: Infection of monocytes from tuberculosis patients with two virulent clinical isolates of Mycobacterium tuberculosis induces alterations in myeloid effector functions.

city: Medellín

first: LELIA

last: LAVALETT

state: Antioquia

department: Antioquia

country: Colombia

institute: Universidad de antioquia

summary: Monocytes play a critical role during infection with Mycobacterium tuberculosis (Mtb). They are recruited to the lung where they participate in the contention of infection. Alternatively, inflammatory monocytes may help in prolonging inflammation or serve as niches for Mtb infection. Also, monocyte response to infection may vary depending on the particularities of the clinical isolate of Mtb from which they are infected. In this pilot study, using microarrays we have examined the global mRNA profiles of circulating human monocytes from healthy individuals and patients with active tuberculosis (TB).

overall_design: Total RNA obtained from monocytes from TB patients infected with clinical isolates of Mtb to compared to monocytes from  healthy control subjects.

first: Lelia

last: Lavalett

first: Hector

last: Ortega

first: Luis

last: Barrera


title: Transcriptional responses to RTS,S induced vaccination in controlled human malaria infection studies

city: Seattle

last: Zak

state: WA

address: 188 East Blaine Street, Suite 300

institute: Regeneron

summary: Identification of host responses that are induced by RTS,S vaccination in controlled malaria infection

overall_design: In order to examine host responses induced in response to RTS,S-based malaria vaccine, expression profiles from both RNAseq and Microarrays platforms were pulled together from a number of RTS,S-based malaria trials. Mixed effects regression models and various filtering strategies were then applied to identify 6 protection-associated gene pairs that were induced by RTS,S vaccination and common in the RTS,S trials. Lastly, these 6 gene pairs were carefully examined and investigate if they were associated with protection in other RTS,S-related trials.

first: Ying

last: Du

first: Ethan

last: Thompson

middle: E

first: Smitha

last: Shankar


title: Canonical BMP signaling executes epithelial-mesenchymal transition downstream of SNAIL1

status: Public on Apr 22 2020

city: Freiburg

first: Antoine

last: Devisme

address: Breisacherstr. 153

institute: Institute of Medical Bioinformatics and Systems Medicine

summary:
Epithelial-mesenchymal transition (EMT) is a pivotal process in development and disease. In carcinogenesis, various signaling pathways are known to trigger EMT by inducing the expression of EMT transcription factors (EMT-TFs) like SNAIL1, ultimately promoting invasion, metastasis and chemoresistance. However, how EMT is executed downstream of EMT-TFs is incompletely understood. Here, using human colorectal cancer (CRC) and mammary cell line models of EMT, we demonstrate that SNAIL1 critically relies on bone morphogenetic protein (BMP) signaling for EMT execution. This activity requires the transcription factor SMAD4 common to BMP/TGFβ pathways, but is TGFβ signaling-independent. Further, we define a signature of BMP-dependent genes in the EMT-transcriptome which orchestrate EMT-induced invasiveness, and are found to be regulated in human CRC transcriptomes and during EMT in vivo. Collectively, our findings substantially augment the knowledge of mechanistic routes whereby EMT can be effectuated, which is relevant for the conceptual understanding and therapeutic targeting of EMT processes.
To identify BMP-dependent genes regulated during EMT, LS174T cells overexpressing Snail1-HA in a doxycycline (Dox)-inducible manner were treated with Dox and inhibitors of BMP signaling (LDN193189/mNoggin) or DMSO as control for different periods of time.

overall_design: 36 samples of total RNA from LS174T-Snail1-HA cells treated with six different combinations of Dox and BMP inhibitors or DMSO for three different periods of time (6 h, 24 h, 72 h) in two biological replicates.

last: Frey

first: Monika

last: Schrempp

first: Geoffroy

last: Andrieux

first: Melanie

last: Boerries

first: Andreas

last: Hecht


title: Recapitulating early mammalian kidney development in vitro by priming and differentiating mouse embryonic stem cells as a monolayer

status: Public on Apr 20 2020

city: Toronto

first: Theresa

last: Chow

state: Ontario

address: 25 Orde Street

institute: Lunenfeld Tanenbaum Research Institute

summary:
In order to harness the potential of pluripotent stem cells, we need to understand how to differentiate them to our target cell types. Here, we developed a protocol to differentiate mouse embryonic stem cells (ESC) to renal progenitors in a step-wise manner. Microarrays were used to track the transcriptional changes at each stage of differentiation and we observed that genes associated with metanephros, ureteric bud and blood vessel development were significantly upregulated as the cells differentiated towards renal progenitors and kidney organoids
We used microarrays to analyze the gene expression profile of cells and to assess for transcriptional changes that occur during cellular differentiation.

overall_design: Mouse embryonic stem cells were differentiated to mesoderm cells (3 replicates), intermediate mesoderm cells (3 replicates) and renal progenitors (3 replicates). Renal progenitors were then coaxed to form 8D, 14D and 21D organoids (3 replicates each). Control cells/tissues include: embryonic stem cells (3 replicates), embryonic kidneys (3 replicates, and adult mouse kidneys (3 replicates)

first: Frances

last: Wong

first: Ian

last: Rogers


title: Gene responses to RTS,S malaria vaccination in controlled human malaria study

first: Daniel

summary: Identification of gene signatures that are associated with protection in RTS,S malaria vaccination study

overall_design: Forty-six volunteers were randomly selected to receive either three full doses of RTS,S/AS01 vaccines or two full doses following by a delayed fractional third dose. All the volunteers underwent CHMI 3 weeks after the last dose and the protection against CHMI was evaluated accordingly. PBMC samples were collected on day of the 1st vaccination, day of the 3rd vaccination, day 1, 3, 14 post-the 3rd vaccination and day of challenge, and used for RNAseq analysis. Here we are trying to identify protection-associated gene signature induced by RTS,S/AS01 vaccination.


title: Transcriptional responses to RTS,S standard regimen (RRR) and Ad35.CS+R,R vaccination in controlled human malaria infection study

summary: Identification of gene signatures that are associated with RTS,S-mediated protection against controlled malaria infection

overall_design: Forty-six volunteers were randomly selected to receive either three monthly doses of RTS,S/AS01 vaccine according to the standard regiment (RRR) or one injection of Ad35.CS followed by two monthly injections of RTS,S (ARR). All of the volunteers underwent CHMI 3 weeks after the last dose and the protection against CHMI was evaluated accordingly. PBMC samples were collected from all participants prior to vaccination and on days 1, 2, 6, and 14 after the 1st injection and on the days of the 2nd and 3rd injections and on days 1 and 6 after the 2nd injection.  Additionally, PBMC samples were collected on the day of challenge and days 1 and 5 after challenge.  This dataset is suitable for analyses that seek to identify differences between RRR and ARR and to identify transcriptional differences between protected and non-protected recipients of RRR and ARR regimens.


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

city: Beijing

first: Li

last: Yu

address: Medicial Science Building, Tsinghua University

department: The Institution of Immunology

institute: Tsinghua University

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

overall_design: WT BMDMs were left untreated or stimulated with LPS  for indicated time. Anti-NELFE was used to immunoprecipitate interacted DNA and the DNA products were subject to ChIPseq

first: Bin

last: Zhang

first: Maria

last: Sacta


title: PROseq to profile activated transcription in bone marrow-derived macrophages from C57BL/6 mice

address: Zheng Yutong Building room D311

summary: We characterized active transcription in conditions of resting and LPS stimualtion in BMDMs

overall_design: PRO-seq were carried out in murine primary bone marrow-derived macrophages (BMDMs). WT and Nelfb KO BMDMs were stimulated with or without LPS for indicated time.


title: ChIPseq to profile CDK9 occupancy in bone marrow-derived macrophages from C57BL/6 mice

summary: We characterized the CDK9 occupancy on gene loci in conditions of unstimuated and LPS stimualtion in BMDMs

overall_design: WT BMDMs were left untreated or stimulated with LPS  for 1 hour. Anti-CDK9 was used to immunoprecipitate interacted DNA and the DNA products were subject to ChIPseq


title: Differential gene expression analysis in Drosophila adult injected with MR766 strain of ZIKV and PBS at 4 and 8 days post injection

status: Public on Apr 19 2020

city: Washington

first: Ioannis

last: Eleftherianos

state: DC

institute: The George Washington University

summary: Purpose: To gain a comprehensive understanding of the molecular function and biological processes activated upon ZIKV infection in Drosophila adult.

overall_design: Transcriptomic profiles of whole fly injected with PBS (sterile control) and ZIKV (both at 4 and 8 days post injection) were generated by deep sequencing, in triplicate, using Illumina HiSeq 2500

first: Eric

last: Tycksen

first: Sneh

last: Harsh

last: Kenney


title: RNAseq to profile transcriptomes in bone marrow-derived macrophages from Nelfb KO(Nelfbfl/fl, Lyz2-Cre) and WT(Nelfb+/+,Lyz2-Cre) mice

summary: We performed the transcriptomic analysis of RNA-seq of BMDMs generated from WT and Nelfb KO in conditions of unstimuated and LPS stimualtion to identify gene expression changes.

overall_design: BMDMs from WT and KO were left untreated or stimulated with LPS  for indicated time. RNA was extracted, polyA enriched and prepared for sequencing libraries.


title: Expression data from LncRNA TROJAN knockdown breast cancer cells

status: Public on Apr 18 2020

city: Shanghai

first: Xi

last: Jin

state: Shanghai

address: 270 Dong An Road, Build 7, RM301

institute: Fudan Universtiy Shanghai Cancer Canter

summary: Background: Estrogen receptor-positive (ER+) breast cancers represent approximately two-thirds of all breast cancers and have a sustained risk of late disease recurrence. Cyclin-dependent kinase 4 and 6 (CDK4/6) inhibitors have shown significant efficacy in ER+ breast cancer. However, their effects are still limited by drug resistance. In this study, we aim to explore the role of long noncoding RNA TROJAN in ER+ breast cancer.  Methods: The expression level of TROJAN in breast cancer tissue and cell lines was determined by quantitative real-time PCR. In vitro and in vivo assays as well as patient derived organoids were preformed to explore the phenotype of TROJAN in ER+ breast cancer. The TROJAN-NKRF-CDK2 axis were screened and validated by RNA pull-down, mass spectrometry, RNA immunoprecipitation, microarray, dual-luciferase reporter and chromatin immunoprecipitation assays.  Results: Herein, we showed that TROJAN was highly expressed in ER+ breast cancer. TROJAN promoted cell proliferation and resistance to a CDK4/6 inhibitor and was associated with poor survival in ER+ breast cancer. TROJAN can bind to NKRF and inhibit its interaction with RELA, upregulating the expression of CDK2. The inhibition of TROJAN abolished the activity of CDK2, reversing the resistance to CDK4/6 inhibitor. A TROJAN antisense oligonucleotide sensitized breast cancer cells and organoids to the CDK4/6 inhibitor palbociclib both in vitro and in vivo.   Conclusions: TROJAN promotes ER+ breast cancer proliferation and is a potential target for reversing CDK4/6 inhibitor resistance.

overall_design: We used microarrays to detail the transcriptomic profiles of the two TROJAN knockdown cell lines.

first: Xiao-En

last: Xu

first: Yi-Zhou

last: Jiang

first: Li-Ping

last: Ge

first: Da-Qiang

last: Li

first: Zhi-Ming

last: Shao

first: Gen-Hong

last: Di


title: Characterising the Transcriptional and Translational Impact of the Schizophrenia-associated miR-1271-5p in Neuronal Cells

city: Callaghan

first: Dylan

last: Kiltschewskij

state: NSW

address: Room MS616, Level 6, Medical Sciences Building, University Drive

department: School of Biomedical Sciences and Pharmacy

country: Australia

institute: The University of Newcastle

summary: Here, we investigated the transcriptome-wide impact of schizophrenia-associated miR-1271-5p in response to bidirectional modulation. Alteration of miR-1271-5p induced considerable changes to mRNA abundance and translation which spanned a diverse range of cellular functions, including directly targeted genes strongly associated with cytoskeletal dynamics and cellular junctions. Mechanistic analyses additionally revealed that upregulation of miR-1271-5p predominantly repressed mRNAs through destabilisation, wherein 3´UTR and CDS binding sites exhibited similar efficacy. Knockdown, however, produced no discernible trend in target gene expression and strikingly resulted in increased expression of the highly conserved miR-96-5p which shares an identical seed region with miR-1271-5p, suggesting the presence of feedback mechanisms which sense disruptions to miRNA levels. These findings indicate that while bidirectional regulation of miR-1271-5p results in substantial remodelling of the neuronal transcriptome, these effects are not inverse in nature. In addition, we provide further support for the idea that destabilisation of mRNA is the predominant mechanism by which miRNAs regulate complementary mRNAs.

overall_design: Examination of changes in mRNA expression and translation in neuronally differentiated SH-SY5Y cells after overexpression or knockdown of miR-1271-5p

first: Michael

last: Geaghan

first: Murray

last: Cairns


title: Neuronal upregulation of Prospero protein is driven by alternative mRNA polyadenylation and Syncrip-mediated mRNA stabilisation

status: Public on Apr 15 2020

city: Oxford

first: Aino

middle: Inkeri

last: Järvelin

address: South Parks Road

department: Department of Biochemistry

country: United Kingdom

institute: University of Oxford

summary: During Drosophila and vertebrate brain development, the conserved transcription factor Prospero/Prox1 is an important regulator of the transition between proliferation and differentiation. Prospero level is low in neural stem cells and their immediate progeny, but is upregulated in larval neurons and it is unknown how this process is controlled. Here, we use single molecule fluorescent in situ hybridisation to show that larval neurons selectively transcribe a long prospero mRNA isoform containing a 15 kb 3’ untranslated region, which is bound in the brain by the conserved RNA-binding protein Syncrip/hnRNPQ. Syncrip binding increases the mRNA stability of the long prosperoisoform, which allows an upregulation of Prospero protein production. Our findings highlight a regulatory strategy involving alternative polyadenylation followed by differential post-transcriptional regulation.

overall_design: RNA-seq in Drosophila L3 larval brain.

first: Tamsin

last: Samuels

first: Ilan

last: Davis


title: Expression data for Rab27a knockdown in the KPC 4662 pancreatic ductal adenocarcinoma cell line

status: Public on Apr 16 2020

city: Chapel Hill, NC

first: Yuliya

last: Pylayeva-Gupta

state: NC

address: 450 West Drive, CB 7295

department: Lineberger Comprehensive Center

institute: UNC

summary: Pancreatic cancer is an aggressive malignancy, often diagnosed at metastatic stages. Several studies have implicated systemic factors, such as extracellular vesicle release and myeloid cell expansion, in the establishment of pre-metastatic niches in cancer. The Rab27a GTPase is overexpressed in advanced cancers, can regulate vesicle trafficking, and has been previously linked to non-cell autonomous control of tumor growth and metastasis, however, the role of Rab27a itself in the metastatic propensity of pancreatic cancer is not well understood. Here, we have established a model to study how Rab27a directs formation of the pre-metastatic niche. Loss of Rab27a in pancreatic cancer cells did not decrease tumor growth in vivo, but resulted in altered systemic myeloid cell expansion, both in the primary tumors and at the distant organ sites. In metastasis assays, loss of Rab27a expression in tumor cells injected into circulation compromised efficient outgrowth of metastatic lesions. However, Rab27a knockdown cells had an unexpected advantage at initial steps of metastatic seeding, suggesting that Rab27a may alter cell-autonomous invasive properties of the tumor cells. Gene expression analysis of gene expression revealed that downregulation of Rab27a increased expression of genes involved in epithelial-to-mesenchymal transition pathways, consistent with our findings that primary tumors arising from Rab27a knockdown cells were more invasive. Overall, these data reveal that Rab27a can play divergent roles in regulating pro-metastatic propensity of pancreatic cancer cells: by generating pro-metastatic environment at the distant organ sites, and by suppressing invasive properties of the cancer cells.

overall_design: 5 samples were analyzed.  KPC 4662 parental cell line as well was KPC 4662 scramble were analyzed as controls while 3 separate knockdowns of Rab27a were analyzed .

first: Nancy

last: Kren


title: Transcriptome dynamics of hematopoietic stem cell formation revealed using a combinatorial Runx1/Ly6a reporter system

status: Public on Apr 17 2020

first: Yuqi

last: Tan

address: 733 North Broadway, MRB 653

institute: Johns Hopkins University

summary: Single cell RNA-Seq time-course analysis revealed that as pre-HSCs mature into fetal liver-stage HSCs they show signs of interferon exposure, multi-lineage differentiation gene expression signatures, and prolonged cell cycle reminiscent of quiescent adult HSCs.

overall_design: Investigation of transcriptome dynamics of hematopoietic stem cell formation using a combinatorial Runx1/Ly6a reporter system

last: Chen

first: Edroaldo

middle: L

last: da Rocha

first: Patrick

last: Cahan

first: Phoebe

last: Hunter

first: Patricia

last: Sousa

first: Caroline

last: Kubaczka

first: Nate

last: Mullin

first: Yuko

last: Fujiwara

first: Minh

last: Nguyen

first: Samuel

last: Landry

first: Areum

last: Han

first: Song

last: Yang

first: Yi-Fen

last: Lu

first: Deepak

last: Jha

first: Linda

last: Vo

first: Yi

last: Zhou

first: Trista

last: North

first: Leonard

last: Zon

first: George

middle: Q

last: Daley

first: Thorsten

last: Schlaege


title: Telomere dysfunction cooperates with epigenetic alterations to impair murine embryonic stem cell fate commitment II

city: Montreal

first: Lea

last: Harrington

state: Quebec

address: 2950 chemin de Polytechnique, Pavillon Marcelle-Coutu

department: Department of Medicine

institute: University of Montreal

summary: This study demonstrates a specific cooperativity between perturbations in histone methylation and eroded telomeres to destabilize the differentiation of murine embryonic stem cells.

overall_design: Murine embyronic stem cells of two different genotypes (WT or mTert-/-) were subjected to differentiation, and treated with compounds that modulated histone H3K27 tri-methylation to assess their impact upon the differentiation response

first: Mélanie

last: Criqui

first: Aditi

last: Qamra

first: Tsz Wai

last: Chu

first: Monika

last: Sharma

first: Julissa

last: Tsao

first: Danielle

last: Henry

first: Dalia

last: Barsyte-Lovejoy

first: Cheryl

last: Arrowsmith

first: Neil

last: Winegarden

first: Mathieu

last: Lupien


title: RNA-seq analysis of transcriptomic changes in macrophage-lineage cells after acute neural injury in larval zebrafish

city: Edinburgh

first: Leah

last: Herrgen

address: 49 Little France Crescent

department: Centre for Discovery Brain Sciences

institute: University of Edinburgh

summary: Both brain-resident microglia and peripheral macrophages are important cellular effectors of the inflammatory response to neural injury. They respond to neural injury by secreting a wide range of effector molecules including cytokines, chemokines and neurotrophic factors. To identify additional secreted signalling molecules, we used RNA-seq gene expression profiling to detect changes in the transcriptome of macrophage-lineage cells after acute neural injury in larval zebrafish. GO term analysis was then used to analyse the list of differentially expressed genes and identify secreted signalling molecules among them.

overall_design: The transcriptomes of FACS-purified macrophage-lineage cells from mpeg1:GFP transgenic zebrafish larvae with or without neural injury were compared.

first: Chiara

last: Herzog

first: Juan

last: Larraz


title: Integrative Analysis and Machine Learning based Characterization of Single Circulating Tumor Cells

city: Okhla Industrial Estate, Phase III

first: Arvind

last: Iyer

email: arvind16122@iiitd.ac.in, krishang@iiitd.ac.in, naveen.ramalingam@fluidigm.com, debarka@iiitd.ac.in

state: New Delhi

address: IIIT Delhi

department: Computational biology department

country: India

institute: IIIT Delhi

summary: Profiling of 15 circulating tumor cells from three subjects with ER+/PR+/HER- hormone receptor status

first: Debarka

last: Sengupta

first: Naveen

last: Ramalingam


title: Gene expression regulation by the Chromodomain Helicase DNA-binding protein 9 (CHD9) chromatin remodeler is dispensable for murine development

city: Amsterdam

first: Anton

last: Berns

state: Amsterdam

address: Plesmanlann 121

department: Division of Molecular Genetics

country: Netherlands

institute: NKI

summary:
Purpose: Chromodomain helicase DNA-binding (CHD) chromatin remodelers regulate transcription and DNA repair. Chd1-8 show upon germline disruption pronounced, often developmental lethal phenotypes. Here we show that contrary to Chd1-8 disruption, Chd9–/– animals are viable, fertile and display no developmental defects or disease predisposition. Germline deletion of Chd9 only moderately affects gene expression in tissues and derived cells, whereas acute depletion in human cancer cells elicits more robust changes suggesting that CHD9 is a highly context-dependent chromatin regulator that, surprisingly, is dispensable for mouse development.
Methods: The role of CHD9 in gene expression regulation during murine development and lymphomagenesis, as well as in human cancer cell lines was explored by RNA-seq analysis. Embryonic (E15) brain and liver were isolated from littermates and snap-frozen. Mouse embryonic fibroblasts (MEFs) were isolated from the Chd9+/-x Chd9+/- cross at day 13.5 post coitum and cultured using standard methods. Three wild-type control and four Chd9 knockout derived fibroblast lines were used. Embryonic stem cells (ESC) were derived from the Chd9+/-x Chd9+/- cross as described before (Huijbers IJ, et al, 2015). Three wild-type control and three Chd9 knockout derived ESC lines were used. For all the experiments early passage MEFs and ESCs were used. Enlarged lymph nodes from three moribund Emu-Myc control, Chd9+/- and Chd9-/- mice were isolated and snap-frozen. Human SNB19 and K562 cells bearing doxycycline-inducible short hairpin RNAs (shRNA) targeting CHD9 were induced with 1ug/ml doxycycline for 48 hours, harvested, and pellet was snap-frozen. Total RNA was extracted, prepared using TruSeq protocol, and standard sample preparation protocols and RNA-seq was performed on a Hiseq2000 machine (Illumina) at the NKI Genomics Core Facility.
Results: Sequencing data were aligned to mm9 (MEFs, ESCs), mm10 (E15 brain and liver; Emu-Myc lymphomas), hg37 (K562 and SNB19 cells) using TopHat (v2.1.0 using bowtie 1.1.0). and number of reads per gene were measured with HTSeq count (v0.5.3). Statistical analysis of the differential expression of genes were performed using DESeq2. Results were considered statistically significant at an adjusted p < 0.05. Normalized expression values were obtained by correcting for differences in sequencing depth between samples using DESeqs median-of-ratios approach.     Analysis of Chd9 knockout MEFs revealed only downregulation of Chd9 and upregulation of the Hmgb2 and pseudogene Gm13237, (adjusted p–value 0.05, log2FC  ±1). Conversely, in derived Chd9–/– ESCs maintained in serum–free 2i media with leukemia inhibitory factor (LIF), 63 upregulated and 212 downregulated genes were observed (adjusted p–value 0.05, log2FC  ±1).     RNA–seq of Chd9 knockout embryonic (E15) liver and brain revealed only a handful of differentially expressed genes (DEGs) (adjusted p–value 0.05, log2FC  ±1). Both liver and brain show concordant upregulation of an uncharacterized pseudogene Gm4322. Despite high Chd9 expression in the developing brain, only marginal upregulation of the Hoxa (a7), Hoxb (b6, b7, b9) and Hoxc (c5, c6) homeotic gene clusters was observed.    Analysis of lymphomas from Emu-Myc Chd9 wildtype, heterozygous and knockout mice revealed that in heterozygous tumours 20 genes were upregulated, and only Lrnn4 was downregulated. In the knockout tumours 28 genes were up– and 8 down–regulated. The presence of a shared six upregulated genes in both CHD9 heterozygous and knockout lymphomas suggests that they might be direct target genes.    RNA-seq analysis of human cancer cell lines revealed profound gene expression changes 48 hours upon CHD9 depletion. In K562 cells, CHD9 knockdown resulted in 215 down- and 635 upregulated genes, whereas in SNB19 cells, 121 down- and 113 up–regulated genes were observed. Notably, a number of deregulated genes in CHD9-depleted K562 cells – such as PBX1, APLNR, and BIN1– are implicated in neuronal development and pathology. This suggests a more widespread role for CHD9 in regulation of neuronal-specific genes, which even shows up in the context of malignant hematopoietic K562 cells. We did not find any significant overlap in DEGs between both human cancer cell lines.
Conclusions: Our study represents the first analysis of the role of CHD9 in gene expression regulation during murine development and lymphomagenesis, as well as in human cancer cells. The very modest effects on gene expression seen upon germline deletion of Chd9, as compared to the substantial change in gene expression upon acute depletion in two human cell lines, point to compensatory me chanisms mitigating the effects of germline deletion of Chd9. Our gene expression analysis failed to reveal any compensatory transcriptional deregulation of CHD enzymes or other chromatin regulators. Our results reveal that, despite ubiquitous expression, Chd9 is dispensable for embryonic development. Furthermore, knockout animals are viable, fertile, display no overt phenotypes and have a normal lifespan.

overall_design: Gene expression profiles of CHD9-defficient primary murine derived cells, embryonic organs, lymphomas, as well as human cancer cell lines were generated by deep sequencing using TruSeq protocol and Hiseq2000 machine (Illumina).

first: Andrej

last: Alendar

first: Rajith

last: Bhaskaran


title: runx1 controls zebrafish heart regeneration by promoting scar deposition as well as inhibiting myocardial proliferation and survival

first: Xiaonan

last: Wang

address: 227 Chongqing South Road

department: School of Public Health

institute: Shanghai Jiao Tong University School of Medicine

summary: Runx1 is a transcription factor that plays a key role in determining the proliferative and differential state of multiple cell types, during both development and adulthood. Here, we report how runx1 is specifically upregulated at the injury site during zebrafish heart regeneration, but unexpectedly, absence of runx1 results in enhanced regeneration. Using single cell sequencing, we found that the wild-type injury site consists of Runx1-positive endocardial cells and thrombocytes that express smooth muscle and collagen genes without differentiating into myofibroblasts. Both these populations are absent in runx1 mutants, resulting in a less collagenous and fibrinous scar. The reduction in fibrin in the mutant is further explained by reduced myofibroblast formation and by upregulation of components of the fibrin degradation pathway, including plasminogen receptor Annexin 2A as well as downregulation of plasminogen activator inhibitor serpine1 in myocardium and endocardium, resulting in increased levels of Plasminogen. In addition, this we also find enhanced myocardial proliferation as well as increased myocardial survival in the mutant. Our findings suggest that Runx1 controls the regenerative response of multiple cardiac cell-types and that targeting Runx1 is a novel therapeutic strategy to induce endogenous heart repair.

overall_design: To determine the differences between wild type, wild type injured and wild type runx1 KO runx1+ endothelial cells in Zebrafish

first: Jana

last: Koth

first: Abigail

last: Killen

first: William

last: Stockdale

first: Helen

last: Potts

first: Andrew

last: Jefferson

first: Florian

last: Bonkhofer

first: Paul

middle: R

last: Riley

first: Roger

last: Patient

first: Berthold

last: Göttgens

first: Mathilda

last: Mommersteeg


title: Analyzing the impact of Mycobacterium tuberculosis infection on primary human macrophages

city: Leiden

first: Herman

middle: Pieter

last: Spaink

address: Einsteinweg 55

department: Institute of Biology

institute: Leiden University

summary: The success of Mycobacterium tuberculosis (Mtb) as pathogen is tightly linked to its ability to recalibrate host metabolic processes in infected host macrophages. Correspondingly, analysis of RNA-sequencing datasets showed altered gene expression of key metabolic enzymes involved in NAD+, creatine, glucose and glutamine metabolism (e.g NAMPT, IDO1, SLC6A8, CKB, HK2) in Mtb-infected M2 macrophages.

overall_design: Total RNA was extracted from of M1 and M2 macrophages either uninfected or infected with Mtb in triplicate from a single blood bank donor and RNA-seq was performed using an Illumina Hi-Seq 2500.

middle: P

first: Frank

last: Vrieling


title: Telomere dysfunction cooperates with epigenetic alterations to impair murine embryonic stem cell fate commitment

first: Lea

last: Harrington

overall_design: Murine embryonic stem cells of two different genotypes (WT or Tert-/-) were subjected to differentiation, and treated with compounds that modulated histone H3K27 tri-methylation to assess their impact upon the differentiation response

first: Mélanie

last: Criqui

first: Aditi

last: Qamra

first: Tsz Wai

last: Chu

first: Monika

last: Sharma

first: Julissa

last: Tsao

first: Danielle

last: Henry

first: Dalia

last: Barsyte-Lovejoy

first: Cheryl

last: Arrowsmith

first: Neil

last: Winegarden

first: Mathieu

last: Lupien


title: Transcriptomic analysis reveals autophagy flux variation within mouse pancreatic cancer organoids

city: New York

first: Keisuke

last: Yamamoto

address: 550 First Avenue

department: Radiation Oncology

institute: NYU School of Medicine

summary: Murine pancreatic cancer cells (HY15549 cells) established from genetically engineered mouse models (GEMM) of PDAC (p48-Cre+, KrasLSL-G12D/+, Trp53lox/+; KPC mice) were transfected with the GFP-LC3-RFP autophagy flux reporter from Mizushima lab, wherein reduction in the GFP/RFP ratio indicates increase in autophagy flux. Organoids derived from these cells were dissociated into single cells and sorted into autophagy-high (AThi) and autophagy-low (ATlo) populations according to GFP/RFP signal ratio. To validate this reporter, RNA was extracted from these two populations and expression of autophagy/lysosome related genes was examined.

overall_design: RNA was isolated from sorted AThi and ATlo populations (n = 2 and 3 biological replicates), which were used for RNA-sequencing to obtain mRNA expression profiles of these two populations.


title: scRNA-Seq for mouse wounded and non-wounded samples

status: Public on Apr 14 2020

city: New Haven

first: Valerie

last: Horsley

state: CT

address: 225 Propsect St

department: Molecular, Cellular, and Developmental Biology

institute: Yale University

summary: Lineage negative (CD45- & CD31-) cells were isolated from uninjured skin and day 5 dorsal skin wound beds of 7-week-old AdipoqCre; mT/mG mice using FACS. This experiment describes multiple subsets of wound bed myofibroblasts and identifies that Adipoq-traced cells contribute to the myofibroblast pool in wound beds.

overall_design: CD45- and CD31-negative were isolated by FACS and processed for single cell RNAseq.

first: Brett

last: Shook

first: Michael

last: Rutenberg Schoenberg


title: Defining the relative and combined contribution of CTCF and CTCFL to genomic regulation

first: Nishana

last: Mayilaadumveettil

state: New York

address: 31st Street & East River DR, Receiving Department BLDG Smilow Floor 03, RM 307 (Jane Skok Laboratory)

department: Pathology

institute: NYU Langone health

summary: We analyzed the impact of CTCFL and CTCF-CTCFL fusion proteins in the presence and absence of CTCF to obtain insight into the individual functional contributions of the zinc finger and N/C terminal domains

overall_design: ChIP seq, RNA seq, HiC

first: Jane

last: Skok

first: Mayilaadumveettil

last: Nishana

first: Javier

last: Rodriguez-Hernaez


title: YAP/TAZ drives ependymoma-like tumour formation in mice

city: London

first: Noreen

last: Eder

address: 1 Midland Road

institute: The Francis Crick Institute

summary: YAP1 gene fusions have been observed in a subset of paediatric ependymomas.  Here we show that ectopic expression of active nuclear YAP1 (nlsYAP5SA) in ventricular zone neural progenitor cells using conditionally-induced NEX/NeuroD6-Cre is sufficient to drive brain tumour formation in mice.  Neuronal differentiation is inhibited in the hippocampus. Deletion of YAP1’s negative regulators LATS1 and LATS2 kinases in NEX-Cre lineage in double conditional knockout mice also generates similar tumours, which are rescued by deletion of YAP1 and its paralog TAZ. YAP1/TAZ-induced mouse tumours display molecular and ultrastructural characteristics of human ependymoma. RNA sequencing and quantitative proteomics of mouse tumours demonstrate similarities to YAP1-fusion induced supratentorial ependymoma. Finally, we find that transcriptional cofactor HOPX is upregulated in mouse models and in human YAP1-fusion induced ependymoma, supporting their similarity.  Our results show that uncontrolled YAP1/TAZ activity in neuronal precursor cells leads to ependymoma-like tumours in mice.

overall_design: Total RNA sequencing of P11 mouse brains (right hemisphere) investigating changes due to ependymoma-like brain tumour formation in nlsYAP5SA mice using Illumina HiSeq 4000

first: Stuart

last: Horswell


title: A WIZ/Cohesin/CTCF complex anchors DNA loops to define gene expression and cell identity [Hi-C]

city: Chapel Hill

first: Jill

last: Dowen

address: 250 Bell Tower Drive, CB 7100 3360 Genome Sciences Building

institute: UNC Chapel Hill

summary:
Purpose: Chromatin architecture is an important regulator of gene expression, but details of how DNA loops form and function remain unclear. The purpose of this study is to assess the role of Widely Interspaced Zinc-fingers protein (WIZ) in gene regulation and DNA looping.
Method: Genome wide chromatin looping patterns in the presence or absence of WIZ were assessed via Hi-C in both wildtype and CRISPR-Cas9 genome edited WIZ deletion mouse embryonic stem cells (mESCs).

overall_design: Hi-C was performed in wild type and Wiz-deletion mESCs  in duplicate.

middle: M

first: Megan

middle: N

last: Justice

first: Zachary

last: Carico

first: Holden

middle: C

last: Stefan


title: Disrupting mitochondrial copper distribution inhibits leukemic stem cell self-renewal

first: Rashim Pal

last: Singh

address: 101, College Street

institute: Princess Margaret Cancer Centre, UHN

type: Methylation profiling by high throughput sequencing

summary: Leukemic stem cells (LSC) rely on oxidative metabolism and are differentially sensitive to targeting mitochondrial pathways, which spares normal hematopoietic cells. A subset of mitochondrial proteins are folded in the intermembrane space via the mitochondrial intermembrane assembly (MIA) pathway. We found increased mRNA expression of MIA pathway substrates in acute myeloid leukemia (AML) stem cells. Therefore, we evaluated the effects of inhibiting this pathway in AML. Genetic and chemical inhibition of ALR reduces AML growth and viability, disrupts LSC self-renewal and induces their differentiation. ALR inhibition preferentially decreases its substrate COX17, a mitochondrial copper chaperone and knockdown of COX17 phenocopies ALR loss. Inhibiting ALR and COX17 increases mitochondrial copper levels which in turn inhibit S-Adenosylhomocysteine Hydrolase (SAHH) and lower levels of S-adenosylmethionine (SAM), DNA methylation, and chromatin accessibility to lower LSC viability. These results provide insight into mechanisms through which mitochondrial copper controls epigenetic status and viability of LSCs.

overall_design: ATAC seq(3+3+2 replicates), Next Gen RNA seq (3+3+3 replicates) and Cap meth seq (3+3+2 replicates) on OCI-AML2 cells after treatment with ALR (GFER) inhibitor MitoBloCK-6 with or without copper chelator penicillamine.

first: Rashim

first: Veronique

last: Voisin

first: James

last: Hawley

first: Gary

last: Bader

first: Aaron

last: Schimmer


title: bT87Q-globin gene therapy reduces sickle hemoglobin production, allowing for ex vivo anti-sickling activity in human erythroid cells

status: Public on Apr 13 2020

city: Bethesda

first: Mehdi

last: Pirooznia

address: 12 SOUTH DR

institute: National Institutes of Health

summary: We performed RNA-seq analysis on immortalized human erythroid cell line (HUDEP-2), that had been engineered to carry the sickle cell disease (SCD) mutation (sHUDEP-2), and on healthy donor mobilized CD34+ derived cells after transduction with a control GFP lentivirus and lentivirus expressing  βT87Q-globin. Our results show activation of inflammation- and proliferation-related programs, suggesting minimal changes of background gene expression except for βT87Q-globin expression and endogenous β/βs-globin suppression.

overall_design: RNA-seq was performed on non-transduced CD34+ derived human primary cells and on CD34+ derived human primary cells transduced with a control GFP lenitviral vector or βT87Q-globin lentiviral vector at the CD71high/CD235ahigh stage of erythroid development (day 9 of culture) and on non-transduced sHUDEP-2 cells and on sHUDEP-2 cells transduced with a control GFP or βT87Q-globin lentiviral vector at the CD71high/CD235ahigh stage (day 6 of culture).

first: Bjorg

last: Gudmundsdottir

first: John

middle: F

last: Tisdale


title: A WIZ/Cohesin/CTCF complex anchors DNA loops to define gene expression and cell identity [ChIP-Seq]

summary:
Purpose: Chromatin architecture is an important regulator of gene expression, but details of how DNA loops form and function remain unclear. The purpose of this study is to assess the role of Widely Interspaced Zinc-fingers protein (WIZ) in gene regulation and DNA looping.
Method: Genome wide binding patterns of known architectural proteins in the presence or absence of WIZ were assessed via chromatin immunoprecipitation followed by high throughput sequencing (ChIP-seq) in both wildtype and CRISPR-Cas9 genome edited WIZ deletion mouse embryonic stem cells (mESCs).

overall_design: ChIP-seq was performed in wild type mESCs  in duplicate for CTCF, SMC1A, RAD21, and WIZ. ChIP-seq was performed in CRISPR-Cas9 genome edited WIZ deletion mESCs in duplicate for Rad21 and CTCF.


title: A WIZ/Cohesin/CTCF complex anchors DNA loops to define gene expression and cell identity

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

overall_design: Refer to individual Series


title: Darolutamide antagonizes androgen signaling by blocking enhancer and super-enhancer activation

status: Public on Apr 10 2020

city: Berlin

first: Bernard

last: Haendler

address: Muellerstrasse 178

department: Preclinical Research

institute: Bayer AG


title: Mapping DNA interaction landscapes in psoriasis susceptibility loci highlights KLF4 as a target gene in 9q31

city: Manchester

first: Paul

last: Martin

state: Greater Manchester

address: Oxford Road

department: Centre for Genetics and Genomics Versus Arthritis

institute: Univeristy of Manchester

summary: Background: Genome-wide association studies (GWAS) have uncovered many genetic risk loci for psoriasis, yet many remain uncharacterised in terms of the causal gene and their biological mechanism in disease. This is largely a result of the findings that over 90% of GWAS variants map outside of protein-coding DNA, and instead are enriched in cell type and stimulation-specific gene regulatory regions. Results: Here, we use a disease-focused Capture Hi-C (CHi-C) experiment to link psoriasis-associated variants with their target genes in psoriasis-relevant cell lines (HaCaT keratinocytes and My-La CD8+ T cells). We confirm previously assigned genes, suggest novel candidates and provide evidence for complexity at psoriasis GWAS loci. For one locus, uniquely, we combine further epigenomic evidence to demonstrate how a psoriasis-associated region forms a functional interaction with the distant (>500 kb) KLF4 gene. This interaction occurs between the gene and active enhancers in HaCaT cells, but not in My-La cells. We go on to investigate this long-distance interaction further with Cas9 fusion protein-mediated chromatin modification (CRISPR activation) coupled with RNA-seq, demonstrating how activation of the psoriasis associated enhancer upregulates known genes in the KLF4 pathway, specific to skin cells and apoptosis. Conclusions: Taken together, our study design provides a robust pipeline to follow up GWAS disease associated variants and implicate the causal genes, cell types, direction of effect and consequences, which are vital next steps for the functional translation of genetic findings into clinical benefit.

overall_design: Capture Hi-C was performed targeting psoriasis GWAS loci in relevant cell lines (HaCaT keratinocyte - unstimulated or stimulated with IFNg - and My-La CD8+ T cells) in order to uncover gene targets of psoriasis-associated SNPs. 3' mRNA-seq was performed to determine target gene expression in the same cell lines. To further explore an interaction between the psoriasis locus at 9q31 and the KLF4 gene, HiChIP for H3K27ac was performed in the same cell lines and confirmed H3K27ac-mediated interactions bewteen the psoriasis locus and the KLF4 promoter in HaCaT cells, but not in My-La cells. A CRISPRa experiment was conducted in HaCaT dCas9 P300 cells, using a pool of sgRNA targeting a selection of the psoriasis-associated SNPs in 9q31, and further RNA-seq was performed to determine differential gene expression against control cell lines expressing a non-specific sgRNA.

first: Helen

last: Ray-Jones

first: Kate

last: Duffus

first: Amanda

last: McGovern

first: Chenfu

last: Shi

first: Jenny

last: Hankinson

first: Oliver

last: Gough

first: Annie

last: Yarwood

first: Andrew

last: Morris

first: Antony

last: Adamson

first: Christopher

last: Taylor

last: Ding

first: Vasanthi

last: Gaddi

first: Yao

last: Fu

first: Patrick

last: Gaffney

first: Gisela

last: Orozco

first: Richard

last: Warren

first: Steve

last: Eyre


title: A WIZ/Cohesin/CTCF complex anchors DNA loops to define gene expression and cell identity [RNA-Seq]

first: Jill

last: Dowen

summary:
Purpose: Chromatin architecture is an important regulator of gene expression, but details of how DNA loops form and function remain unclear. The purpose of this study is to assess the role of Widely Interspaced Zinc-fingers protein (WIZ) in gene regulation and DNA looping.
Method: The role of WIZ in gene regulation was assessed by performing RNA-seq in both wildtype and CRISPR-Cas9 genome edited WIZ deletion mouse embryonic stem cells (mESCs).

overall_design: RNA-seq was performed in wild type mESCs  and WIZ CRISPR-Cas9 genome edited WIZ deletion mESCs in triplicate.

first: Megan

last: Justice


title: Delineation and modulation of the natural killer cell transcriptome in rhesus macaques during ZIKV and SIV infections

status: Public on Apr 08 2020

city: Atlanta

first: Gregory

middle: K

last: Tharp

state: GA

address: 954 Gatewood Dr

department: Developmental and Cognitive Neuroscience

institute: Yerkes National Primate Research Center

summary: Natural killer (NK) cells are crucial regulators of antiviral and anti-tumor immune responses. Although in humans some NK cell transcriptional programs are relatively well established, NK cell transcriptional networks in non-human primates (NHP) remain poorly delineated. Here we performed RNA-Seq experiments using purified NK cells from experimentally naïve rhesus macaques, providing the first transcriptional characterization of pure NK cells in a NHP species. This novel NK cell transcriptomic signature (NK RMtsig) overlaps with published human NK signatures, allowing us to identify new key signaling and transcription factor networks underlying NK cell function.  Finally, we show that applying NK RMtsig to an unrelated rhesus macaque cohort infected with SIVmac251 or ZIKV can sensitively detect NK cell repertoire perturbations, thus confirming applicability of this approach. In sum, we propose this NHP NK cell signature will serve as a useful resource for future studies involving infection, disease or treatment modalities in NHP

overall_design: Examination of the baseline transcriptome of NK cells from 6 rhesus macaque.

first: Malika

last: Aid

first: Daniel

last: Ram

first: Steven

last: Bosinger

first: Dan

middle: H

last: Barouch

first: R

last: Reeves


title: Myogenic Progenitor Cell Lineage Specification by CRISPR/Cas9-based Transcriptional Activators

city: Durham

first: Charles

last: Gersbach

address: 101 Science Dr

institute: Duke University

summary: Engineered CRISPR/Cas9-based transcriptional activators can potently and specifically activate endogenous fate-determining genes to direct differentiation of pluripotent stem cells. Here, we demonstrate that endogenous activation of the PAX7 transcription factor results in stable epigenetic remodeling and directly reprograms human pluripotent stem cells into skeletal myoblast progenitor cells. Compared to the exogenous overexpression of PAX7 cDNA, we find that endogenous activation results in the generation of more proliferative myogenic progenitors that can maintain PAX7 expression over multiple passages in serum-free conditions while preserving the capacity for terminal myogenic differentiation. Transplantation of human myogenic progenitors derived from endogenous activation of PAX7 into immunodeficient mice resulted in a greater number of human dystrophin+ myofibers compared to exogenous PAX7 overexpression. RNA-seq analysis also revealed transcriptome-wide differences between myogenic progenitors generated via CRISPR-based endogenous activation of PAX7 and exogenous PAX7 cDNA overexpression. These studies demonstrate the utility of CRISPR/Cas9-based transcriptional activators for progenitor cell specification and their potential for regenerative medicine.

overall_design: Transcriptional profiling of differentiated human iPSCs that had been treated with either PAX7-targeting gRNA only, VP64dCas9VP64 with gRNA, cDNA encoding PAX7-A isoform, or cDNA encoding PAX7-B isoform at day 14 of differentiation.

first: Jennifer

last: Kwon


title: Darolutamide antagonizes androgen signaling by blocking enhancer and super-enhancer activation [RNA-seq]

first: Bernard

last: Haendler

summary: The androgen receptor (AR) antagonist darolutamide has very recently been approved for the treatment of non-metastatic castration resistant prostate cancer (PCa). Here we determined the genome-wide effects of darolutamide on cis-acting regulatory elements involved in androgen signaling with a focus on enhancer and super-enhancer (SE) regions. Darolutamide strongly depleted the AR from regulatory elements and abolished the AR transcriptional signaling. Using two different androgen-dependent PCa cell lines we identified genomic-regions with different affinities for the AR in androgen-stimulated, androgen-depleted and darolutamide-antagonized conditions. Altogether, our findings demonstrate that darolutamide is a potent AR antagonist blocking genome-wide AR signaling and AR enhancer activation. Further, we show a dynamic AR cistrome and concomitant adapting chromatin environment to varying conditions and identified regions with high AR affinity in cell lines and tissue samples.

overall_design: Prostate androgen-sensitive cell lines VCaP and LAPC-4 were grown for two days in starvation medium prior to treatment with DMSO control, synthetic androgen R1881 at a concentration of 1 nM or R1881 with darolutamide at a concentration of 500 nM (low) or 2 µM (high). The cells were harvested 8 h or 22 h post-induction for RNA isolation and sequencing.

first: Ekaterina

last: Nevedomskaya

first: Simon

middle: J

last: Baumgart

first: Ralf

last: Lesche


title: RNASeq of cold-sensitive and cold-insensitive neurons from the pre-optic area of hypothalamus of mice

status: Public on Apr 09 2020

first: Viktor

last: Feketa

address: 333 Cedar St SHM BE58

department: Cellular & Molecular Physiology

summary: The preoptic area of the hypothalamus (POA) contains intrinsically warm and cold-sensitive neurons, which are thought to be critically involved in mammalian thermoregulation. However, the precise physiological roles and the molecular markers of the cold-sensitive POA neurons have not been determined yet. Here, we tackle this problem by performing calcium-imaging guided separation and collection of cold-sensitive and cold-insensitive dissociated neurons from the mouse POA, followed by RNASeq and differential transcriptomics of these cell populations.

overall_design: ~100-200 individual cold-sensitive or cold-insensitive neurons from the POA of mouse hypothalamus were collected and pooled together for 1 sample. RNA was isolated from samples and high-throughput deep sequencing of the RNA performed. The study included 3 biological replicates of each type, with a total of 6 samples.

middle: V

first: Elena

last: Gracheva

first: Sviatoslav

last: Bagriantsev


title: Darolutamide antagonizes androgen signaling by blocking enhancer and super-enhancer activation [ChIP-seq]

overall_design: Prostate androgen-sensitive cell lines VCaP and LAPC-4 were grown for two days in starvation medium prior to treatment with DMSO control, synthetic androgen R1881 at a concentration of 1 nM or R1881 with darolutamide at a concentration 2 µM for 22h. The cells were subsequently fixed with 1% formaldehyde and processed for ChIP-seq. For ChIP-seq antibodies against AR, FOXA1, BRD4, MED1, H3K27ac, H3K4me1, H2BK15ac, H3K18ac.


title: Single cell transcriptomics of Ewing Sarcoma cell lines

status: Public on Apr 11 2020

city: San Antonio

first: Henry

last: Miller

state: TX

address: 8403 Floyd Curl Drive

department: Cell Systems and Anatomy

institute: UT Health San Antonio

summary: We report single cell gene expression profiles for Ewing Sarcoma cell lines CHLA9, CHLA10, and TC71. Cell lines were harvested and prepared as single cell suspensions with high viability. Library prep was performed with the 10X Genomics v3.1 3' kit. All samples were sequenced on the illumina HiSeq 3000 with a 28+8+100 configuration. CHLA9 and CHLA10 libraries were re-sequenced on the Illumina NovaSeq with a 28+8+91 configuration. Counts were processed with cellranger v3.1.0. Both the raw data and the cellranger output are provded.

overall_design: 10X Genomics single cell RNA-Seq (scRNA-Seq) of Ewing Sarcoma cell lines CHLA9, CHLA10, and TC71

first: Nicklas

last: Bassani

first: Alexander

last: Bishop