title: Expression data from doxorubicin-resistant hepatoma cells

city: Taoyuan

first: Tse-Ching

last: Chen

address: 5. Fu-Hsing Street

department: Anatomic Pathology

country: Taiwan

institute: Chang Gung Memorial Hospital

summary:
Transarterial chemoembolization or systemic chemotherapy with doxorubicin has been the treatment of choice for unresectable hepatocellular carcinoma (HCC) conferring the best survival benefit. However, HCC is notorious for its predisposition to develop therapeutic tolerance. Thus, Affymetrix microarray analysis was performed on  doxorubicin-resistant hepatoma cells to identify the drug resistance-related genes.
The RNA isolated from primary hepatoma cells and their doxorubicin-resistant counterparts, then subjected to Affymetrix microarray analysis to identify the differential expression profiles of drug resistance-related genes.

overall_design: We used Affymetrix microarrays to detail the global gene expression of primary cells, including 834, 413, and P15 as well as their resistant counterparts to identify the differential expression profiles of drug resistance-related genes.

first: T

first: H

middle: S

last: Wang


title: PLZF targets developmental enhancers for activation during osteogenic differentiation of human mesenchymal stem cells (4C-seq)

status: Public on Jan 17 2019

city: Copenhagen N

first: Mads

last: Lerdrup

address: Blegdamsvej 3C

department: Department of Cellular and Molecular Medicine

country: Denmark

institute: University of Copenhagen

summary: The PLZF transcription factor is essential for osteogenic differentiation of hMSCs, however, its regulation and molecular function during this process is not fully understood. Here we revealed that the ZBTB16 locus encoding PLZF, is repressed by Polycomb (PcG) and H3K27me3 in naïve hMSCs. At the pre-osteoblast stage of differentiation, the locus lost PcG binding and H3K27me3, gained JMJD3 recruitment, and H3K27ac resulting in high expression of PLZF. Subsequently, PLZF was recruited to osteogenic enhancers, influencing H3K27 acetylation and expression of nearby genes important for osteogenic function. Furthermore, we identified a latent enhancer within the ZBTB16/PLZF locus itself that became active, gained PLZF, p300 and Mediator binding and looped to the promoter of the nicotinamide N-methyltransferase (NNMT) gene. The increased expression of NNMT correlated with a decline in SAM levels, which is dependent on PLZF and is required for osteogenic differentiation.

overall_design: Genomic contacts made by EnP element upon osteogenic differenetiation (4C-seq)

first: Shuchi

last: Agrawal Singh

first: Ana-Luisa

middle: R

last: Gomes

first: Harmen

middle: J

last: Van de Werken

first: Jens

last: Vilstrup Johansen

first: Robin

last: Andersson

first: Albin

last: Sandelin

first: Kristian

last: Helin

first: Klaus

last: Hansen


title: RNA sequencing of control and dnm2-knockout mouse sciatic nerves

status: Public on Jan 16 2019

city: Zurich

first: Daniel

last: Gerber

address: Otto-Stern-Weg 7

country: Switzerland

institute: ETH Zurich

type: Expression profiling by high throughput sequencing

summary: In this study, we analyzed the transcriptome profiles of mouse sciatic nerves after developmental (P1 and P5) or  inducible (4 weeks post Tamoxifen) deletion of Dnm2 conditionally in Schwann cells (using a P0Cre-driven or a P0CreERT2-driven recombination of floxed alleles, respectively) as compared to controls (floxed Dnm2 homozygous, Cre-negative).

overall_design: mRNA profiles  from controls and mutants were collected in triplicate from developmental sciatic nerves at P1 and P5, as well as from inducible mice at 4 weeks post tamoxifen

first: Monica

last: Ghidinelli

first: Ueli

last: Suter


title: Conserved Methyltransferase Spb1 Targets mRNAs for Regulated Modification with 2′-O-Methyl Ribose

city: New Haven

first: Wendy

last: Gilbert

state: CT

address: 333 Cedar Street, P.O. Box 208024

country: USA

institute: Yale University

type: Methylation profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Phytochrome-based extracellular matrix with reversibly tunable mechanical properties

city: Freiburg

first: Barbara

last: Hummel

address: Stübeweg 51

country: Germany

institute: Max Planck Institute of Immunobiology and Epigenetics

summary: Interrogation and control of cellular fate and function using optogenetics is providing revolutionary insights into biology. Optogenetic control of cells is achieved by coupling genetically encoded photoreceptors to cellular effectors and enables unprecedented spatiotemporal control of signaling processes. Here, a fast and reversibly switchable photoreceptor is used to tune the mechanical properties of polymer materials in a fully reversible, wavelength-specific, and dose- and space-controlled manner. By integrating engineered cyanobacterial phytochrome 1 into a polyethylene glycol matrix, hydrogel materials responsive to light in the cell-compatible red/far-red spectrum are synthesized. These materials are applied to study in human mesenchymal stem cells how different mechano-signaling pathways respond to changing mechanical environments, and to control the migration of primary immune cells in 3D. This optogenetics-inspired matrix allows addressing fundamental questions of how cells react to dynamic mechanical environments. Further, remote control of such matrices could create new opportunities for tissue engineering or provide a basis for optically stimulated drug depots.

overall_design: Analysis of global gene expression changes due to differences in the mechanical properties of the phytochrome-based hydrogels

first: Maximilian

last: Hörner

first: Wilfried

last: Weber


title: Effect of NFE2L2 silencing on miRNA profile of human aortic endothelial cells (HAECs) in response to GDF-15 and SDF-1

city: Krakow

first: Anna

last: Grochot-Przeczek

address: Gronostajowa 7

country: Poland

institute: Jagiellonian University

type: Non-coding RNA profiling by high throughput sequencing

summary: Nrf2 is a transcription factor playing an essential role in stress response. A knowledge of the significance of Nrf2 in cell function has not, as yet, reached out beyond transactivation of gene expression. This paper shows that GDF-15 and SDF-1-induced angiogenesis strongly depends on Nrf2 presence, but is not related to its transcriptional activity. We propose, that Nrf2 serves as a protein restraining Keap1, its known transcriptional repressor. Deficiency of Nrf2 protein available for Keap1 leads to overabundance of RhoGAP1, the protein regulating Cdc42 activity, and impairs podosome assembly, thereby indisposing actin rearrangements, and preventing angiogenesis. These activities can be rescued by concomitant deletion of RhoGAP1 or Keap1. We suggest that a new Nrf2 function of a Keap1 scavenger implies revising the established murine model of Nrf2 deficiency as a transcriptional knock out (tKO) mouse. The N-terminal fragment of Nrf2, containing Neh2 domain binding Keap1, is present in these animals. Thus, regarding the function of Nrf2 as a protein sequestering Keap1, both published and unpublished data on Nrf2-Keap1 duet may gain new interpretation.

overall_design: miRNA profile of human aortic endothelial cells (HAECs) transfected with scrambled (siMock) or NFE2L2 (siNFE2L2) siRNA, stimulated with PBS, GDF-15 (250 ng/mL) or SDF-1 (100 ng/mL)


title: Overriding FUS autoregulation triggers gain-of-toxic dysfunctions in autophagy-lysosome axis and RNA metabolism

city: Singapore

first: Shuo-Chien

last: Ling

state: Singapore

address: 12 Science Drive 2, MD1, 16-03-H

department: Physiology

country: Singapore

institute: National University of Singapore

summary: Mutations in coding and non-coding regions of FUS cause amyotrophic lateral sclerosis (ALS). The latter mutations may exert toxicity by increasing FUS accumulation. We showhere that broad expression within the nervous system of wild type or either of two ALS-linkedmutants of human FUS produces progressive motor phenotypes accompanied bycharacteristic ALS-like pathology. FUS levels are autoregulated by a mechanism in whichwild type or ALS-linked mutants of human FUS downregulating endogenous FUS at bothmRNA and protein levels. Increasing wild type human FUS expression achieved bysaturating this autoregulatory mechanism produces a rapidly progressive neurologicalphenotype and dose-dependent lethality. Genome-wide expression analysis reveals mis-regulation of genes that are largely distinct from the alterations observed by the acute FUS reduction in the spinal cord. Among these are increased expression of lysosomal proteins,suggestive of disruption in protein homeostasis as a potential gain-of-toxic mechanism.Indeed, increased expression of wild type and ALS-linked mutations in FUS inhibitautophagy with accumulated p62/sequestosome observed in spinal cord motor neurons.Taken together, our data suggests that overriding FUS autoregulation triggers gain-of-toxicproperties in the autophagy-lysosome axis and deregulation of RNA metabolism, highlightinga disruption in protein and RNA homeostasis in FUS-mediated toxicity.

overall_design: Differential gene expression analysis in spinal cord sections of <> mice


title: Paclitaxel plus Cirmtuzumab Achieves Greater Clearance of Patient-derived Xenografts By Targeting ROR1+ Breast Cancer Stem Cells

city: La Jolla

first: suping

last: zhang

state: CA

address: 8955 health science Dr

department: Medical school

institute: UCSD

summary: We applied  high-throughput sequence to identify signaling pathways, stem cell gene signature or target genes of BMI1 that were affected by our newly development humainzed anti-ROR1 antibody (cirmtuzumab) in breast cancer patient-derived xenograft (PDX) mice model

overall_design: Examination of 8 PDX samples isolated from control-treated mice (N=4) versus cirmtuzumab-treated mice (N=4)

first: Suping

last: Zhang

first: Han


title: Epithelial mesenchymal transition using doxycycline-inducible Snail, Zeb1, TGFbeta or Vector in H358 NSCLC cell line

city: Stony Brook

first: John

middle: D

last: Haley

state: NY

address: BST 9-168, SB Medical

institute: Stony Brook University

summary:
Epithelial – mesenchymal transition (EMT) is an important contributor to the invasion and metastasis of epithelial-derived cancers. While considerable effort has focused in the regulators involved in the transition process, we have focused on consequences of EMT to prosurvival signaling. Changes in distinct metastable and ‘epigentically-fixed’ EMT states were measured by correlation of protein, phosphoprotein, phosphopeptide and RNA transcript abundance. The assembly of 1167 modulated components into functional systems or machines simplified biological understanding and increased prediction confidence highlighting four functional groups: cell adhesion and migration, metabolism, transcription nodes and proliferation/survival networks. A coordinate metabolic reduction in a cluster of 17 free-radical stress pathway components was observed and correlated with reduced glycolytic and increased oxidative phosphorylation enzyme capacity, consistent with reduced cell cycling and reduced need for macromolecular biosynthesis in the mesenchymal state. An attenuation of EGFR autophosphorylation and a switch from autocrine to paracrine-competent EGFR signaling was implicated in the enablement of tumor cell chemotaxis. A similar attenuation of IGF1R, MET and RON signaling with EMT was observed. In contrast, EMT increased prosurvival autocrine IL11/IL6-JAK2-STAT signaling, autocrine fibronectin-integrin  activation, autocrine Axl/Tyro3/PDGFR/FGFR RTK signaling and autocrine TGF-R signaling. A relatively uniform loss of polarity and cell-cell junction linkages to actin cytoskeleton and intermediate filaments was measured at a systems level. A more heterogeneous gain of ECM remodeling and associated with invasion and migration was observed. Correlation to stem cell, EMT, invasion and metastasis datasets revealed the greatest similarity with normal and cancerous breast stem cell populations, CD49fhi/EpCAM-/lo and CD44hi/CD24lo respectively.
Thomson, S., Petti, F., Sujka-Kwok, I., Mercado, P., Bean, J., Monaghan, M., Seymour, S.L., Argast, G., Epstein, D. and Haley, J.D. (2011). ‘A systems view of epithelial - mesenchymal transition signaling states’ Clin. Exp. Metastasis 28, 137-55.

overall_design: Affy U133-2 arrays with RT-PCR followup

first: S

last: Thomson

first: J


title: Circulating Long Non-coding RNA ENST00000507296 is a Prognostic Indicator in Patients with Dilated Cardiomyopathy

status: Public on Jan 15 2019

city: wuhan

first: xudong

address: jiefang avenue 1095

country: China

institute: tongji hospital


title: Conserved Methyltransferase Spb1 Targets mRNAs for Regulated Modification with 2′-O-Methyl Ribose [eCLIP]

summary: Non-coding RNAs contain dozens of chemically distinct modifications, of which only a few have been identified in mRNAs. The recent discovery that certain tRNA modifying enzymes also target mRNAs suggests the potential for many additional mRNA modifications. Here, we show that conserved tRNA 2′-O-methyltransferases Trm3, 7,13 and 44, and rRNA 2′-O-methyltransferase Spb1, interact with specific mRNA sites in yeast by crosslinking immunoprecipitation and sequencing (CLIP-seq). We developed sequencing of methylation at two prime hydroxyls (MeTH-seq) for transcriptome-wide mapping of 2′-O-methyl ribose (Nm) with single-nucleotide resolution, and discover thousands of potential Nm sites in mRNAs. Genetic analysis identified hundreds of mRNA targets for the Spb1 methyltransferase, which can target both mRNA and non-coding RNA for environmentally regulated modification. Our work identifies Nm as a prevalent mRNA modification that is likely to be conserved and provides methods to investigate its distribution and regulation.

overall_design: For each protein of interest, we tag it so that it may be pulled down with an antibody, and prepare libraries from the associated RNA. We also prepare a no-pulldown size-matched input control for each protein.

first: Kristen

middle: M

last: Bartoli

first: Cassandra

last: Schaening

first: Thomas

middle: C

last: Carlile

middle: V


title: Etv6-dependent positive and negative gene regulatory network controls vegfa expression in vivo

city: Oxford

first: Lei

last: Li

address: University of Oxford

department: Radcliffe Department of Medicine

country: United Kingdom

institute: WIMM

type: Genome binding/occupancy profiling by high throughput sequencing


title: Checkpoint blockade immunotherapy induces dynamic changes in PD-1-CD8+ tumor-infiltrating T cells

city: Boston

first: Asaf

last: Madi

state: MA

address: 60 Fenwood Road, BTM 10016F

department: Neurology

institute: BWH

summary: An improved understanding of the anti-tumor CD8+ T cell response after checkpoint blockade would enable more informed and effective therapeutic strategies. Here we examined the dynamics of the effector response of CD8+ tumor-infiltrating lymphocytes (TILs) after checkpoint blockade therapy. Bulk and single-cell RNA profiles of CD8+ TILs after combined Tim-3+PD-1 blockade in preclinical models revealed significant changes in the transcriptional profile of PD-1? TILs. These cells could be divided into subsets bearing characterstics of naive-, effector-, and memory-precursor-like cells. Effector- and memory-precursor-like TILs contained tumor-antigen-specific cells, exhibited proliferative and effector capacity, and expanded in response to different checkpoint blockade therapies across different tumor models. The memory-precursor-like subset shared features with CD8+ T cells associated with response to checkpoint blockade in patients and was compromised in the absence of Tcf7. Expression of Tcf7 was requisite for the efficacy of diverse immunotherapies, highlighting the importance of this transcriptional regulator in the development of effective CD8+ T cell responses upon immunotherapy.

overall_design:
(i)  RNAseq of Wild Type Naïve-like, Memory-like and Effector-like subpopulations of PD1-CD8+ Tumor infiltrating lymphocytes isolated from MC38-OVA. CD62LhiSlamf7-CX3CR1-, CD62L-Slamf7hiCX3CR1- and CD62L-Slamf7hiCX3CR1+ subsets within PD-1-CD8+ TILs
(ii)  RNAseq from WT mice, Tim-3+PD-1+ and Tim-3-PD-1- CD8+ TILs were isolated by cell sorting from MC38-OVA tumor-bearing mice that were treated with anti-PD-1 and anti-Tim-3 antibodies or isotype controls.
(iii)  Droplet-based single-cell RNA-Seq of Tim-3-PD-1- CD8+ TILs from MC38-OVA tumor-bearing WT mice that were treated with anti-PD-1 and anti-Tim-3 antibodies or isotype controls.


title: RNAi-dependent epimutation induces broad antifungal resistance in Mucor circinelloides

city: Durham

first: Robert

middle: Blake

last: Billmyre

state: North Carolina

address: 312 CARL Bldg

department: Molecular Genetics and Microbiology

institute: Duke University Medical Center

summary: To elucidate the role of epimutation in antifungal resistance of Mucor, epimutations were isolated that confer resistance to 5-fluoroorotic acid (5-FOA). We identified epimutant strains that exhibit resistance to 5-FOA without mutations in either of the 5-FOA target genes, pyrF or pyrG. Using sRNA sequencing, we demonstrate that these epimutants harbor sRNA against either pyrF or pyrG, and further show that this sRNA is lost after reversion to drug sensitivity.

overall_design: 10 samples were sequenced, including 2 pyrF epimutants and 1 pyrG epimutant before and after passaging.  Before passaging, samples were URA-; after passaging they reverted to the wildtype phenotype  and became URA+. The remaining 4 sequences are derived from the mutant parent strain and the wildtype background for the two pyrF and the single pyrG sequenced strain.

first: Zanetta

last: Chang

first: R

middle: B

first: Soo

last: Lee

first: Joseph

last: Heitman


title: Etv6-dependent positive and negative gene regulatory network controls vegfa expression in vivo [ChIP-seq]

summary: VEGFA signaling is crucial for physiological and pathological angiogenesis and hematopoiesis. Although many context-dependent signaling pathways downstream of  VegfA have been uncovered, vegfa transcriptional regulation in vivo is unclear. Here, we show that ETS transcription factor Etv6 positively regulates vegfa expression during blood stem cell development through multiple transcriptional inputs. In agreement with its established repressive functions, Etv6 directly inhibits expression of vegfa repressor foxo3. Surprisingly, it also directly activates expression of vegfa activator klf4. Finally, it indirectly binds to vegfa promoter by the recruiment of Klf4. Thus, our work uncovers a dual function for Etv6, as both a transcriptional repressor and activator, in controlling a major signaling pathway.

overall_design: Etv6 ChIP-seq was performed on three independent biological samples obtained from the somites of wild-type (WT) stage 22 embryos. Indexed libraries were then generated from immunoprecipited DNA and control input samples, pooled and sequenced using a NextSeq 500 sequencer.

first: Rossella

last: Rispoli

first: Roger

last: Patient

first: Aldo

last: Ciau-Uitz

first: Catherine

last: Porcher


title: LncRNA expression data from human plasma of control donors and chronic HF patients

last: zhang

summary:
There are a lot of non-coding RNAs in human plasma. Some of them could be a biomarker of diseases.
We used microarrys to detect lncRNAs in plasma of 10 control subjects and 10 heart failure (HF) patients diagnosed with dilated cardiomyopathy (DCM) and we identified distinct classes of dysregulated lncRNAs.

overall_design: Blood samples were collected from 10 control donors (victims of accident) and 10 chronic HF patients diagnosed with DCM during heart transplantation surgery. Then, plasma was obtained by centrifugation at 2000g at 4ºC for 10 minutes, and the supernatant was carefully transferred to a new RNAse/DNAse-free 1.5mL microtube and stored at -80ºC until use. Total RNA was extracted from 250ul plasma by TRIzol LS Reagent (Life Technologies, Carlsbad, CA) following the instructions. RNA was pre-amplified and then underwent microarray analysis (Arraystar, Human lncRNA array, version 3.0).

first: Xudong


title: Gene expression analysis in response to hypoxic pathway inhibition

city: Liverpool

first: Sonia

last: Rocha

address: crown street

department: Biochemistry

institute: University of Liverpool

summary: Gene expression analysis of HeLa cells exposed to hypoxia (1% Oxygen), treated with a PHD inhibitor (IOX2) or treated with a VHL inhibitor (VH032) for 16 hours

overall_design: 3 biological replicates of HeLa cells exposed to hypoxia (1% Oxygen), PHD inhibitor (250 μM of IOX2) or  VHL inhibitor (250 μM VH032), comparing to vehicle control (0.05% DMSO). Treatments were performed for 16 hours

first: Julianty

last: Frost

first: Alessio

last: Ciulli


title: ALS implicated protein TDP-43 sustains levels of STMN2 a mediator of motor neuron growth and repair

first: Rory

last: Kirchner

address: 677 Huntington Ave

department: Department of Biostatistics

institute: Harvard T.H. Chan School of Public Health

summary: The discovery that TDP-43 mutations cause familial ALS and that many patients display pathological TDP-43 mislocalization has nominated altered RNA metabolism as a potential disease mechanism. Despite its importance, the identity of RNAs regulated by TDP-43 in motor neurons remains poorly understood. Here, we report transcripts whose abundances in human motor neurons are sensitive to TDP-43 depletion. Notably, we found STMN2, which encodes a microtubule regulator, declined after TDP-43 knockdown, in patient-specific motor neurons, following TDP-43 mislocalization, and in the postmortem patient spinal cords. Loss of STMN2 upon reduced TDP-43 function was due to the emergence of a cryptic exon, which is of substantial functional importance, as we further demonstrate that STMN2 is necessary for both axonal outgrowth and repair. Importantly, post-translational stabilization of STMN2 could rescue neurite outgrowth and axon regeneration deficits induced by TDP-43 depletion.  We propose restoring STMN2 expression warrants future examination as an ALS therapeutic strategy.

overall_design: Transcriptome differential expression analysis in HUES3 cells with and without siRNA knockdown of TDP-43

last: Klim

first: Luis

middle: A

last: Williams

first: Francesco

last: Limone

first: Irune

last: Guerra San Juan

first: Brandi

middle: N

last: Davis-Dusenbery

first: Daniel

last: Mordes

first: Aaron

last: Burberry

first: Michael

last: Steinbaugh

first: Kanchana

middle: K

last: Gamage

first: Rob

last: Moccia

first: Seth

last: Cassel

first: Kuchuan

last: Chen

first: Brian

last: Wainger

first: Clifford

last: Wolff

first: Kevin

last: Eggan


title: Necroptosis inhibition protects from dopaminergic neuronal cell death in OPA1 mutant Parkinson’s disease patient neurons and MPTP treated mice

city: Milano

first: Luca

last: Massimino

state: MI

address: Via Olgettina, 58

department: Gastroenterology

country: Italy

institute: Ospedale San Raffaele

summary: Dysfunctions in mitochondria dynamics and metabolism are common pathological processes associated with Parkinson’s disease (PD). Recently, it was shown that an inherited form of PD and dementia is caused by new mutations in the OPA1 gene, which encodes for a key player of mitochondrial fusion and structure. iPSC-derived neural cells from these patients exhibited severe mitochondrial fragmentation, respiration impairment, ATP deficits and heightened oxidative stress. Reconstitution of normal levels of OPA1 in PD-derived neural cells normalized mitochondria morphology and function. OPA1 mutated neuronal cultures showed reduced survival in vitro. Intriguingly, selective inhibition of necroptosis effectively rescued this survival deficit. Additionally, dampening necroptosis in MPTP treated mice protected from DA neuronal cell loss. This human iPSC-based model captures both the early pathological events in OPA1 mutant neural cells and the beneficial effects of blocking necroptosis, highlighting this cell death process as a promising therapeutic target for PD.

overall_design: 3 replicates for control and 3 replicates for OPA1 F38D mutant cells

first: Vania

last: Broccoli

first: Angelo

last: Iannielli

first: Simone

last: Bido


title: Conserved Methyltransferase Spb1 Targets mRNAs for Regulated Modification with 2′-O-Methyl Ribose [meTH-Seq]

first: Wendy

last: Gilbert

overall_design: Each experiment consists of two libraries, one reverse transcribed in low (restrictive) Mg2+ and one at permissive concentrations. Nm sites are identified through peaks, ie. positions where there are more reverse transcription stops in the restrictive than in the permissive condition. We include data for 41 such experiments across two growth conditions, and in various knockout backgrounds.

first: Kristen

last: Bartoli

first: Cassandra

last: Schaening

first: Thomas

last: Carlile


title: Oligodendrocyte progenitor cells become regionally diverse and heterogeneous with age

city: St Louis

first: Sabine

last: Dietmann

state: MO

address: 4565 McKinley Avenue

institute: Washington University School of Medicine

summary: Oligodendrocyte progenitor cells (OPCs), which differentiate into myelinating oligodendrocytes during central nervous system (CNS) development, are the main proliferative cells in the adult brain. OPCs are conventionally considered a homogeneous population, particularly with respect to their electrophysiological properties, but this has been debated. We show, by using single-cell electrophysiological recordings, that OPCs start out as a homogeneous population, but become functionally heterogeneous, varying both within and between brain regions and with age. These electrophysiological changes in OPCs correlate with the differentiation potential of OPCs; thus, they may underlie the differentiational differences in OPCs between regions and likewise differentiation failure with age.

overall_design: Total RNA profiles of oligodendrocyte precursor cells isolated from 3 to 4 wild-type mice at different time points, using SMARTer® Stranded Total RNA-Seq Kit v2 - Pico Input Mammalian.

first: Kimberley

last: Evans

first: Yasmine

last: Kamen

first: Ragnhildur

middle: T

last: Káradóttir


title: Contribution of SRF and Nkx2-5 to androgen-dependent gene expression in prostate cancer

city: BUFFALO

first: Eduardo

last: Cortes

address: ELM AND CARLTON STREETS

department: Biostatistics and Bioinformatics

institute: ROSWELL PARK CANCER INSTITUTE

summary: To study and compare the effect of siRNA-mediated SRF or Nkx2-5 silencing on androgen-responsive genes in LNCaP cells.

overall_design: Human LNCaP prostate cancer cells were transfected with siRNA SmartPools targeting SRF or Nkx2-5 or a non-targeting siRNA SmartPool.  Forty-two hours after transfection, cells were treated with synthetic androgen R1881 (5nM) or vehicle.  Three biological replicates were generated per treatment group.  Forty-eight hours later, total RNA was isolated and processed for RNA-Seq analysis.

first: Salma

last: Ben-Salem

first: Dhirodatta

last: Senapati

first: Varadha

last: Balaji Venkadakrishnan

first: Yixue

last: Su

first: Qiang

last: Hu

first: Song

last: Liu

first: Hannelore

last: Heemers


title: Insight into novel RNA-binding activities via large-scale analysis of lncRNA-bound proteome and IDH1-bound transcriptome

city: Rochester

first: jinlong

last: lu

address: 213 Hutchinson Hall

institute: University of Rochester

summary: RNA-binding proteins (RBPs) play pivotal roles in directing RNA fate and function. Yet the current annotation of RBPs is largely limited to proteins carrying known RNA-binding domains. To systematically reveal dynamic RNA-protein interactions, we surveyed the human proteome by a protein array-based approach and identified 671 proteins with RNA-binding activity. Among these proteins, 525 lack annotated RNA-binding domains and are enriched in transcriptional and epigenetic regulators, metabolic enzymes, and small GTPases. Using an improved CLIP (crosslinking and immunoprecipitation) method, we performed genome-wide target profiling of isocitrate dehydrogenase 1 (IDH1), a novel RBP. IDH1 binds to thousands of RNA transcripts with enriched functions in transcription and chromatin regulation, cell cycle and RNA processing. Purified IDH1, but not an oncogenic mutant, binds directly to GA- or AU-rich RNA that are also enriched in IDH1 CLIP targets. Our study provides useful resources of unconventional RBPs and IDH1-bound transcriptome, and convincingly illustrates, for the first time, the in vivo and in vitro RNA targets and binding preferences of IDH1, revealing an unanticipated complexity of RNA regulation in diverse cellular processes.

overall_design: Transcriptome-wide profiling of IDH1 RNA targets in mESCs.

first: Lichao

first: Xiaohua

last: Shen


title: Next Generation Sequencing Quantitative Analysis of Wild Type and AML1-ETO Related Fusion Circular RNA (F-CircAE) overexpression NIH3T3 Cells Transcriptomes

city: Tianjin

first: Ying

last: Wang

address: 288 Nanjing Road

institute: State Key Laboratory of Experimental Hematology, Chinese Academy of Medical Sciences & Peking Union Medical College

summary: The goal of this study is to determine the difference in transcriptome expression profils between wild type and AML1-ETO related fusion circular RNA overexpression NIH3T3 cells

overall_design: NIH3T3 cells  mRNA profiles of wild type and  AML1-ETO related fusion circular RNA overexpression were generated by deep sequencing, in triplicate, using Illumina.

first: Yingxi

last: Xu

first: Min

first: Jianxiang


title: Next Generation Sequencing Quantitative Analysis of Wild Type and AML1-ETO Related Fusion Circular RNA (F-CircAE) Knockdown Kasumi-1 Cells Transcriptomes

summary: The goal of this study is to determine the difference in transcriptome expression profils between wild type and AML1-ETO related fusion circular RNA knockdown Kasumi-1 cells

overall_design: Kasumi-1 mRNA profiles of wild type and  AML1-ETO related fusion circular RNA knockdown were generated by deep sequencing, in triplicate, using Illumina.


title: Single cell transcriptomic analysis of the lateral hypothalamic area reveals molecularly distinct populations of inhibitory and excitatory neurons

city: Farmington

first: William

middle: F

last: Flynn

address: 10 Discovery Drive

institute: The Jackson Laboratory

summary: We employed a droplet-based single cell RNA-sequencing (scRNA-seq) approach to develop a comprehensive census of molecularly distinct cell types in the mouse lateral hypothalamic area (LHA). In addition to 13 distinct non-neuronal cell populations, we define 15 distinct populations of glutamatergic and 15 distinct populations of GABAergic neurons, including both known and novel cell types. This comprehensive transcriptomic analysis of LHA cell types lays the groundwork for understanding the circuit-level underpinnings of LHA coordination of innate behavior.

overall_design: Two single cell transcriptomic (Chromium v2 chemistry) data sets of P30-aged mouse LHA, one of 3 pooled male and the other of 2 pooled female animals.

first: Alexander

last: Jackson

first: Paul

last: Robson


title: Expression data from human adipose-derived stem cells (ADSCs)

status: Public on Jan 14 2019

city: Kanazawa

first: Oto

last: Inoue

address: 13-1 Takaramachi

department: System biology

country: Japan

institute: Kanazawa University

summary:
Adipose-derived stem cells are angiogenic and attractive for therapeutic angiogenesis. However, ADSCs contain several subsets according to expression pattern of surface markers.
We identified CD271 (LNGFR) can isolate ADSCs with superior capacity for angiogenesis. Therefore, we aimed to compare gene expression profile between CD271+ and CD271- ADSCs.

overall_design: Human adipose-derived stem cells were obtained from subcutaneous adipose tissue of 4 patients. Total RNA was isolated from sorted CD271+ and CD271- ADSCs.

first: Soichiro

last: Usui


title: Small RNA profiling of Dicer_KO mouse embryonic stem cells.

city: Zurich

first: Constance

last: Ciaudo

email: cciaudo@ethz.ch, ciaudobeyer@gmail.com

address: Otto-Stern-Weg 7

department: D-BIOL

institute: ETH

summary: Purpose:  The goals of this study was to obtain the small RNA profiling of Dicer_KO mESCs.

overall_design: ALL small RNA profiles from total RNA were generated by deep sequencing, in duplicate, using Illumina HiSeq2000.


title: Genetic and Behavioural Requirements for Structural Brain Plasticity

city: Verdun

first: Nicholas

last: O'Toole

state: QC

address: 6875 Boulevard Lasalle

country: Canada

institute: Douglas Mental Health University Institute

summary: Human MRI studies show that experience can lead to changes in the volume of task-specific brain regions; however, the behavioural and molecular processes driving these changes remain poorly understood. Here, we used in-vivo mouse MRI and RNA sequencing to investigate the neuroanatomical and transcriptional changes induced by environmental enrichment, exercise, and social interaction. Additionally, we asked whether the volume changes require CREB, a transcription factor critical for memory formation and neuronal plasticity. Enrichment rapidly increased cortical and hippocampal volume, and these effects were not attributable to exercise or social interaction. Instead, they likely arise from learning and sensorimotor experience. Nevertheless, the volume changes were not attenuated in mice with memory impairments caused by loss of CREB, indicating that these effects are driven by processes distinct from this canonical learning and memory pathway. Finally, within brain regions that underwent volume changes, enrichment increased the expression of genes associated with axonogenesis, dendritic spine development, synapse structural plasticity, and neurogenesis, suggesting these processes underlie the volume changes detected with MRI.

overall_design: RNA-sequencing data from CREB-knockout mice in Dorsal DG, Ventral DG and Visual Cortex in Enriched and Exercise environments

last: O’Toole

first: Tieyuan

last: Zhang


title: LncRNA sequencing in human cardiac myocytes (HCMs), human cardiac fibroblasts-adult ventricular (HCF-av) and human cardiac microvascular endothelial cells (HCMECs) from a single fetal donor

first: xudong

last: zhang

summary: lncRNA-sequencing were performed in three human heart-derived cell types

overall_design: We report the application of sequencing technology for high-throughput profiling of lncRNA expressions in three human heart-derived cell types. In this project, rRNA was removed from the total RNA by ribosomal RNA removal kit, and the way of ion breaking were used to break the RNA into 200-300bp fragments. By using RNA as template, the first strand of cDNA was synthesized with 6 base random primers and reverse transcriptase. The first strand cDNA was used as the template to synthesize the second strand cDNA. When the second strand cDNA is synthesized, the base T is replaced by U to achieve the goal of a chain specific library. All of our RNA-seq projects used strand specific kits to construct the libraries. Finally, we show that lncRNA different expression in three human heart-derived cell types. This study provides abundance of lncRNAs that may predict cardiac diseases.

first: Xudong


title: The mRNA export receptor NXF1 coordinates transcriptional dynamics, alternative polyadenylation and mRNA export

city: Newark

first: Bin

last: Tian

state: NJ

address: 205 South Orange Ave.

department: Department of Microbiology, Biochemistry and Molecular Genetics

institute: Rutgers New Jersey Medical School

summary: Alternative polyadenylation (APA) produces mRNA isoforms with different 3’UTR lengths. Previous studied indicated that 3’ end processing and mRNA nuclear export are intertwined in gene regulation. Here, we show that mRNA export factors generally facilitate usage of distal cleavage and polyadenylation sites (PASs), leading to expression of long 3’UTR isoforms. By focusing on the export receptor NXF1, which exhibits the most potent effect on APA in this study, we reveal a number of gene features that impact NXF1-dependent APA, including 3’UTR size, gene size and AT content. Surprisingly, downregulation of NXF1 results in accumulation of RNAP II at the 3’ end of genes, correlating with its role in APA regulation. Moreover, we show that NXF1 cooperates with CFI-68 to facilitate nuclear export of long 3’UTR isoform with UGUA motifs. Together, our work reveals several important roles of NXF1 in coordinating RNAPII distribution, 3’ end processing, and mRNA export of long 3’UTR transcripts, implicating NXF1 as the nexus of gene expression regulation.

overall_design:
In total 9 3'READS libraries for analysis of APA under KD of export factors using total RNA from Hela cells.
In total 8 3'READS libraries for analysis of APA under siNXF1 and siCFI-68 using cytoplasmic, monosomal fraction, and polysomal fraction from Hela cells.
In total 4 ChIP-seq po2 IP and input libraries from HeLa cells for the analysis of po2 pausing under siNXF1.
In total 2 ChATT-seq libraries from HeLa cells for the analysis of nasent RNA under siNXF1.
In total 2 RNA-seq libraries from HeLa cells for the analysis of total RNA under siNXF1.

first: Suli

last: Chen

first: Ruijia

first: Dinghai

last: Zheng

first: Heng

first: Xingya

last: Chang

first: Wencheng

last: Li

first: Hong

last: Cheng


title: Genome-wide co-localization of RNA-DNA interactions and fusion RNA pairs (RNAseq)

city: La Jolla

first: Sheng

last: Zhong

state: California

address: 9500 Gilman Drive MC 0412

department: Bioengineering

institute: University of California, San Diego

summary: Despite the ever-increasing speed of detecting fusion transcripts in cancer, it remains formidable to predict what unreported RNA pairs can form new fusion transcripts. By systematic mapping of chromatin-associated RNAs (caRNAs) and their respective genomic interaction loci, we obtained genome-wide RNA-DNA interaction maps from two non-cancerous cell types. The gene pairs involved in RNA-DNA interactions in these normal cells exhibited strong overlap with those with cancer-derived fusion transcripts. These data suggest an RNA-poise model, where the spatial proximity of one gene’s transcripts and the other gene’s genomic sequence poises for the creation of fusion transcripts. We validated this model with 96 additional lung cancer samples. One of these additional samples exhibited fusion transcripts without a corresponding fusion gene, suggesting that genome-recombination is not a required step of the RNA-poise model.

overall_design: Targeted RNA-seq on lung cancer patient biopsy and H2228 cell line

first: Zhangming

last: Yan

first: Norman

last: Huang

first: Weixin

last: Wu

first: Weizhong

first: Yiqun

last: Jiang

first: Jingyao

first: Xuerui

first: Xingzhao

last: Wen

first: Kang

first: Zhen

first: Shu

last: Chien


title: The molecular logic of Nanog-induced self-renewal in mouse embryonic stem cells

city: paris

first: Pablo

last: Navarro

address: 25 rue du Docteur Roux

country: France

institute: Institut Pasteur

summary: Identification of Nanog targets by transcriptomic changes of Nanog induction and depletion, identification of Nanog co-binding logic by ChIP-seq of Nanog co-binding factors with and without Nanog depletion. H3K27me3 with Nanog induction in +/- Lif.

overall_design: RNA-seq performed on 44iN +/- Dox, SunTag Nanog & Nanog/Otx2 in +/- Dox +/- Lif. ChIP for Brg1, Esrrb, Sox2, Oct4 in 44iN +/- Dox. ATAC-seq in 44iN +/- Dox. H3K27me3 ChIP-seq in Suntag Nanog +/- Dox +/- Lif.

first: Nick

last: Owens


title: Lin28/let-7 axis regulates the timig of cession of nephrogenesis

first: Areum

last: Han

address: Karp Family Building, 7th Floor. 300 Longwood Ave.

institute: Boston Children's Hospital

summary: To study role of Lin28/let-7 axis on nephrogenesis, we profiled kidney transcriptom of LIN28 OE, let-7 KO, and their wild-type littermate control mice.

overall_design: RNA-seq on kidneys from LIN28B OE, let-7 KO, and their wild-type littermate controls at P3.

first: Alena

last: Yermalovich

first: George

middle: Q

last: Daley


title: SWI/SNF Chromatin Remodeling Factor BRG1 is Synthetic Required for PTEN-deficient Prostate Cancer

city: Shanghai

first: Ni

state: Shanghai

address: 320 Yueyang Road, Shanghai, 200025 P.R. China

institute: Institute of Health Sciences, Shanghai Institute for Biological Sciences, Chinese Academy of Sciences

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

overall_design: Refer to individual Series


title: miRNA expression profiles of uncultured stromal cells of eutopic and ectopic origin

city: Tartu

first: Tõnis

last: Tasa

address: Liivi 2

country: Estonia

institute: University of Tartu

summary: We utilized fluorescence-activated cell sorting to isolate endometrial stromal cells from paired endometrial and endometrioma biopsies and combined it with high-throughput sequencing to determine miRNA alterations in endometriotic stroma. The analysis revealed 149 abnormally expressed miRNAs in endometriotic lesions, including extensive upregulation of miR-139-5p and downregulation of miR-375 compared to eutopic cells. The results of this study provide further insights into the complex molecular mechanisms involved in endometriosis pathogenesis and demonstrate the necessity for cell-type specific analysis of ectopic tissues to understand the interactions between different cell populations in disease onset and progression.

overall_design: miRNA profiles of paired eutopic and ectopic endometrial stromal cells were generated by small-RNA deep sequencing using Illumina NextSeq 500 platform.

first: Kadri

last: Rekker

first: Saare

last: Merli

first: Peters

last: Maire


title: Transcriptome analysis of control and BRG1-depleted 22RV-1 cells with or without PTEN ablation

last: Li

summary: We conduct transcriptome comparison of control and BRG1-depleted 22RV-1 cells with or without PTEN ablation to gain genomic insights on the biological processes that BRG1 is involved in depend on PTEN deletion.  Through unsupervised cluster analysis of differentially expressed genes (DEGs), we found the expressions of 5489 genes were significantly altered in PTEN/BRG1 double knockdown cells, whereas they remained consistent or opposite pattern in control, BRG1-KD and PTEN-KD cells. Differential principal component analysis (PCA) suggested ablation of BRG1 in PTEN-depleted cells forced the cells undergoing a distinct transcriptome as compared to the others. In keeping with these findings, KEGG-DEGs relationship network indicated BRG1 loss in PTEN-deficient cells significantly altered the process related to “Prostate cancer”, “Cell cycle”, “Cell motility” and etc.

overall_design: 22RV-1 cells  were infected with control and BRG1-KD letivirus. After puromycin selection, the two types of cells were infected with control and PTEN-KD letivirus individually.  Total RNA was extracted using TRIzol followed by RNeasy Mini kit (Qiagen) cleanup.

first: Jun

last: Qin

first: Yufeng

last: Ding


title: Calorie restriction regulates translation to regulate metabolism in mouse liver

status: Public on Jan 13 2019

city: CLEVELAND

first: Roman

last: Kondratov

state: Ohio

address: 2399 euclid avenue, bges dept, sr289

department: Biomedical Sciences

institute: CLEVELAND STATE UNIVERSITY

summary: Using next generation sequencing, we report transcripts that are differentially associated in the mouse liver polysomes under calorie restriction.

overall_design: 2 month old animals were put under 30% calorie restricted diet for three months. Control animals were fed ad libitum. Liver tissues were harvested at 6 time points across the day and polysome profiling was performed. Rna extracted from polysome fractions were assayed using next generation sequencing.

first: Kuldeep

last: Makwana

first: Neha

last: Gosai

first: Allan

last: Poe


title: scRNA-seq for lung BASCs stem cells at the bronchioalveolar-duct junction

city: Guangzhou

first: Guangdun

last: Peng

address: 190 Kaiyuan Road

department: Guangzhou Institutes of Biomedicine and Health

institute: Chinese Academy of Sciences

summary: Characterizing the stem cells responsible for lung repair and regeneration is important for the treatment of pulmonary diseases. Recently, a unique cell population located at the bronchioalveolar-duct junctions has been proposed to comprise endogenous stem cells for lung regeneration. However, the role of bronchioalveolar stem cells (BASCs) in vivo remains debated, and the contribution of such cells to lung regeneration is not known. Here we generated a genetic lineage-tracing system that uses dual recombinases (Cre and Dre) to specifically track BASCs in vivo. Fate-mapping and clonal analysis showed that BASCs became activated and responded distinctly to different lung injuries, and differentiated into multiple cell lineages including club cells, ciliated cells, and alveolar type 1 and type 2 cells for lung regeneration. This study provides in vivo genetic evidence that BASCs are bona fide lung epithelial stem cells with deployment of multipotency and self-renewal during lung repair and regeneration.

overall_design: Three groups of tdTomato+ epithelial cells sorted from BASCs-Tracer mouse, Scgb1a1-CreER; R26-tdTomato mouse and Sftpc-DreER; R26-RSR-tdTomato mouse respectively, and subjected to scRNA-seq. Total 480 tdTomato+ epithelial cells were sorted and sequenced (160 per group).

first: Qiaozhen

last: Liu

first: Kuo

first: Guizhong

last: Cui

first: Hongbin

last: Ji

first: Bin

last: Zhou


title: Identification of Transcription Factor Binding Sites using ATAC-seq

city: Aachen

first: Zhijian

state: North Rhine-Westphal

address: Pauwelsstr. 19

institute: RWTH Aachen University

summary: Dendritic cells (DC) are professional antigen presenting cells that comprise different subsets: classical DC type 1 and 2 (cDC1 and cDC2, respectively) and plasmacytoid DC (pDC). In this study cDC1 or pDC were obtained in a two-step in vitro culture system according to Felker et al., J. Immunol. 185, 5326-5335, 2010. Briefly, mouse bone marrow cells were first amplified with a specific cytokine cocktail and then induced to differentiate into DC with Flt3 ligand. cDC1 are CD11c+ CD11b+ XCR1+ and pDC are CD11c+ CD11b- B220+ and thus cDC1 and pDC were obtained by FACS sorting and subjected to Omin-ATAC-seq analysis.  As an example of practical application of HINT-ATAC, we performed Omin-ATAC-seq experiments of cDC and pDC cell populations and used HINT-ATAC to detect footprints within ATAC-seq peaks of each of these two cells. Next, we estimated changes in binding activity for all factors with a motif in JASPAR. Cell-specific TF activity is measured by the depth of footprints and number of reads flanking regions. Two known cDC factors, BATF3 and HES1, are detected. In summary, we demonstrated that HINT-ATAC is capable of identifying relevant factors for dendritic cell specification.

overall_design:
We performed ATAC-seq experiments in mouse classical dendritic cells type1 (cDC1) and plasmacytoid dendritic cells (pDC) in duplicates. Omni-ATAC-seq was performed according to Corces et al., Nature Methods 14, 959-962, 2017 with minor modifications. Prior to transposition dead cells were removed by centrifugation (800 rpm, 4 min, 4°C). The transposition reaction was with 7.5 μl Tagment DNA Enzyme 1 (TDE1) for 60 min at 37°C.
Pre-amplification was with NEBNext Ultra II Q5 Master Mix and Nextera PCR Primers (5 cycles). Quantitative PCR amplification was with NEBNext Ultra II Q5 Master Mix, Nextera PCR Primer, and SYBR Gold to determine the number of additional cycles. PCR amplification of additional cycles was as for pre-amplification. PCR fragments were purified with Qiagen MinElute PCR Purification Kit and library concentration and quality were determined by Agilent High Sensitive DNA Kit and Bioanalyzer, respectively.

first: Marcel

middle: H

last: Schulz

first: Thomas

last: Look

first: Matthias

last: Begemann

first: Martin

last: Zenke

first: Ivan

middle: G

last: Costa


title: Compare chromatin open regions (OCRs) between PTEN-KD and PTEN/BRG1-KD 22RV-1 cells by ATAC-Seq

first: Ni

last: Li

summary: Transposase-accessible chromatin using sequencing (ATAC-seq) assay was performed to compare chromatin open regions (OCRs) between PTEN-KD and PTEN/BRG1-KD 22RV-1 cells. Ablation of BRG1 in PTEN-depleted cells preferentially led to the reductions of OCR at enhancers, but not promoter regions.

overall_design: Lentivirus-mediated RNA interference was used to knockdown PTEN and/or BRG1 expression in prostate cancer cell line 22RV-1. Harvest cells and immediately continue to transposition reaction. Then purify and PCR amplify transposed DNA. library construction protocol is according to the NEB kit.

first: Jun

first: Yufeng


title: BRG1 occupancy profiling by high throughput sequencing from control and PTEN knockdown 22RV-1 cells

summary: To identify the downstream targets and pathways mediated by BRG1, chromatin immunoprecipitation followed by sequencing (ChIP–seq) was performed in control and PTEN-knockdown 22RV-1 cells. Total 6279 genes showed the enhanced BRG1 occupancies in PTEN-knockdown cells as compared to that in control cells. Genome-wide analysis revealed that PTEN loss lead to increase BRG1 intervals specifically localized at enhancers, but not distal promoter regions. We therefore reasoned ablation of BRG1 might impair enhancer configurations in PTEN-deficient cells.

overall_design: Lentivirus-mediated RNA interference was used to knockdown PTEN expression in prostate cancer cell line 22RV-1. The chromatin was prepared and followed by ChIP-Seq analysis by Active Motif, Inc. using the antibody against BRG1 (abcam, ab110641).

last: Qin


title: Next-generation sequencing of murine trophoblast-derived and pregnancy-associated exosome-enriched extracellular vesicle microRNAs

status: Public on Jan 12 2019

city: Stanford

first: Virginia

middle: D.

last: Winn

address: 300 Pasteur Drive, Boswell A364

department: OB-GYN/Reproductive, Perinatal and Stem Cell Biology Research

institute: Stanford University

summary: The role of extracellular vesicles (EVs), specifically exosomes, in intercellular communication likely plays a key role in placental orchestration of pregnancy and maternal immune sensing of the fetus. While murine models are powerful tools to study pregnancy and maternal-fetal immune interactions, in contrast to human placental exosomes, the content of murine placental and pregnancy exosomes remains largely understudied. Using a recently developed in vitro culture technique, murine trophoblast stem cells derived from B6 mice were differentiated into syncytial-like cells. EVs from the conditioned media, as well as from pregnant and non-pregnant sera, were enriched for exosomes. The RNA composition of these murine trophoblast-derived and pregnancy-associated exosome-enriched-EVs (ExoE-EVs) was determined using RNA-sequencing analysis and expression levels confirmed by qRT-PCR. Differentially abundant miRNAs were detected in syncytial differentiated ExoE-EVs, particularly from the X chromosome cluster (mmu-miR-322-3p, mmu-miR-322-5p, mmu-miR-503-5p, mmu-miR-542-3p, and mmu-miR-450a-5p). These were confirmed to be increased in pregnant mouse sera ExoE-EVs by qRT-PCR analysis. Interestingly, fifteen miRNAs were only present within the pregnancy-derived ExoE-EVs compared to non-pregnant controls. Mmu-miR-292-3p and mmu-miR-183-5p were noted to be some of the most abundant miRNAs in syncytial ExoE-EVs and were also present at higher levels in pregnant versus non-pregnant sera ExoE-EVs. The bioinformatics tool, MultiMir, was employed to query publicly available databases of predicted miRNA-target interactions. This analysis reveals that the X-chromosome miRNAs are predicted to target ubiquitin-mediated proteolysis and intracellular signaling pathways. Knowing the cargo of placental and pregnancy-specific ExoE-EVs as well as the predicted biological targets informs studies using murine models to examine not only maternal-fetal immune interactions but also the physiologic consequences of placental-maternal communication.

overall_design: Examination of EVs isolated from 1) conditioned media from in vitro mouse trophoblast cells cultures maintained as "STEM" using Cell Start and FGF4 (n=3), 2) conditioned media from in vitro mouse trophoblast cultures that have undergone syncytial-like differentiation (n=3), 3) non-pregnant B6 mouse sera (n=3); 4) pregnant B6 mouse sera (gestational day 14.5, n=3). EVs were isolated using ExoQuick-TC (1-2) or ExoQuick (3-4).

first: Adrianne

middle: L

last: Stefanski

first: Nadine

last: Martinez

first: Lisa

last: Peterson

first: Tiffany

last: Callahan

first: Eric

last: Treacy

first: Marisa

last: Luck

first: Samantha

last: Friend

first: Amy

last: Hermesch

first: Emin

last: Maltepe

first: Tzu

last: Phang

first: Leonard

last: Dragone


title: small RNA and mRNA sequencing data of human granulosa cells from PCOS women with insulin resistance

city: Hefei

first: Bo

last: Xu

state: Anhui

address: Lujiang road 17

institute: The First Affiliated Hospital of University of Science and Technology of China

summary: Our objective was to identify the miRNA and mRNA expression profiles in PCOS women with or without insulin resistance.

overall_design: In this study, 26 PCOS patients with IR (PCOS-IR) and 24 patients without IR (PCOS-control) were enrolled. We determined the differentially expressed miRNA and mRNA using next-generation sequencing technology

first: Meihong

last: Hu


title: DNA methylation analysis in familial adenomatous polyposis

city: Chiba

first: Masaki

last: Fukuyo

address: 1-8-1 Inohana, Chuo-ku

department: Department of Molecular Oncology

institute: Chiba University

summary: DNA methylation of 23 familial adenomatous polyposis tumors. Infinium HumanMethylation450 BeadChip was used to obtain DNA methylation profiles across 485,577 CpG sites.

overall_design: Bisulphite converted DNA from the 23 samples were hybridised to the Infinium HumanMethylation450 BeadChip.

first: Kiyoko

last: Takane

first: Hisahiro

last: Matsubara

first: Yasuyuki

last: Seto

first: Atsushi

last: Kaneda


title: Cellular collectivity and its effect on early CD4 T cell memory differentiation

city: Rehovot

first: Ido

last: Amit

address: 234 Herzl st.

department: Immunology

country: Israel

institute: Weizmann Institute of Science

summary: Cellular collectivity and its effect on T cell memory formation. Two experiments were performed: 1) 20160314. Gene expression of early differentiated CD62Lpos and CD62Lneg CD4 T cells. 2) 170213. Gene expression of CD4 T cells cultured in high and low initial concentrations over time. And, early CD4 T cell memory differentiation

overall_design: Two experiments were performed: 1) 20160314. naïve CD4 T cells were cultured at an initial concentration of 2.5E5 cells/ml. Cells were activated using antiCD3/antiCD28 coated beads. Cells were cultured in 200ul complete RPMI in 96-well plates. At the indicated times cells were sorted to CD62Lpos and CD62Lneg populations. 2) 20170213. naive CD4 T cells were cultured at either 6.25E4 or 1E6 cells/ml. Cells were activated using antiCD3/antiCD28 coated beads. Cells were cultured in 1ml complete RPMI in 24-well plates. at the indicated times CD4pos cells were sorted. additionally, naive CD4 cells were cultured at an initial concertation of 2.5E5 cells/ml similarly to the first experiment, and sorted at the indicated times to CD4posCD62LposCD44pos or CD4posCD62LnegCD44pos populations.

first: Michal

last: Polonsky

first: Chamutal

last: Bornstein

first: Eyal

last: David

first: Gil

last: Stelzer


title: Quantitative sensing and signalling of single-stranded DNA during the DNA damage response

city: Martinsried/Munich

first: Susanne

middle: Charlotte Sophie

last: Bantele

state: Bavaria

address: Am Klopferspitz 18

department: NWG

institute: Max Planck Institute of Biochemistry

summary: The DNA damage checkpoint senses the presence of DNA lesions and controls the cellular response thereto. A crucial DNA damage signal is single-stranded DNA (ssDNA), which is frequently found at sites of DNA damage and recruits the sensor checkpoint kinase Mec1-Ddc2.  However, how this signal – and therefore the cells’ DNA damage load – is quantified, is poorly understood.  Here, we use genetic manipulation of DNA end resection at a site-specific DNA double-strand break (DSB) in budding yeast to generate quantitatively different DNA damage (ssDNA) signals. Interestingly, two major targets of the Mec1-Ddc2 kinase – Rad53 and γH2A – differ in their response to the ssDNA signal, indicating distinct signalling circuits within the checkpoint. The local checkpoint signalling circuit leading to γH2A phosphorylation is non-quantitative and unresponsive to increased amounts of damage-associated Mec1-Ddc2 kinase. In contrast, the global checkpoint signalling circuit, which triggers Rad53 activation, integrates the ssDNA signal quantitatively.   We find that not only Mec1-Ddc2 association, but also loading of the 9-1-1 co-sensor complex is enhanced during ongoing resection. Moreover, we can uncouple global checkpoint activation from the amount of Mec1-Ddc2 kinase at the lesion by using mutant conditions that hyper-activate the 9-1-1 signalling axis and at the same time show reduced amounts of damage-associated Mec1-Ddc2 kinase. We therefore propose that a key function of the 9-1-1 complex and the downstream checkpoint mediators is to generate a checkpoint response, which is quantitative and proportional to the cellular DNA damage load.  The DNA damage checkpoint senses the presence of DNA lesions and controls the cellular response thereto. A crucial DNA damage signal is single-stranded DNA (ssDNA), which is frequently found at sites of DNA damage and recruits the sensor checkpoint kinase Mec1-Ddc2.  However, how this signal – and therefore the cells’ DNA damage load – is quantified, is poorly understood.  Here, we use genetic manipulation of DNA end resection at a site-specific DNA double-strand break (DSB) in budding yeast to generate quantitatively different DNA damage (ssDNA) signals. Interestingly, two major targets of the Mec1-Ddc2 kinase – Rad53 and γH2A – differ in their response to the ssDNA signal, indicating distinct signalling circuits within the checkpoint. The local checkpoint signalling circuit leading to γH2A phosphorylation is non-quantitative and unresponsive to increased amounts of damage-associated Mec1-Ddc2 kinase. In contrast, the global checkpoint signalling circuit, which triggers Rad53 activation, integrates the ssDNA signal quantitatively.   We find that not only Mec1-Ddc2 association, but also loading of the 9-1-1 co-sensor complex is enhanced during ongoing resection. Moreover, we can uncouple global checkpoint activation from the amount of Mec1-Ddc2 kinase at the lesion by using mutant conditions that hyper-activate the 9-1-1 signalling axis and at the same time show reduced amounts of damage-associated Mec1-Ddc2 kinase. We therefore propose that a key function of the 9-1-1 complex and the downstream checkpoint mediators is to generate a checkpoint response, which is quantitative and proportional to the cellular DNA damage load.

overall_design: In this experiment, RPA and gammaH2A enrichments around two permanent  HO-induced DNA double-strand break are analyzed using ChIP-seq before and after 4 hours of DSB induction in M phase-arrested cells. WT cells were compared to resection-defective exo1∆ sgs1∆ cells.

first: Boris

last: Pfander


title: Long non-coding RNAs discriminate the stages and gene regulatory states of the human humoral immune response (RNA-Seq)

city: New York

first: Cem

last: Meydan

state: New York

address: 1305 York

department: Physiology & Biophysics

institute: Weill Cornell Medical College

summary: Long non-coding RNAs (lncRNAs) make up a vast majority of the human transcriptome and have key regulatory functions. We performed unbiased de novo annotation of transcripts expressed during the humoral immune response in humans. Remarkably, 30% of the total human genome is transcribed during this process, and yet 58% of these transcripts manifest striking differential expression indicating phylogenetic relationships among cell types that is even more robust than coding genes. We provide a regulatory atlas of lncRNAs in naïve B and GC B-cells that indicates their partition into ten functionally distinct categories based on chromatin features including H3K4me1, H3K4me2, H3K4me3, H3K27ac, H3K27me3, DNase hypersensitivity, CTCF, EP300, CREBBP, MED1, BRD4, FOXO1 and FOXP1 localization. This procedure defined lncRNAs classes such as enhancer RNA (eRNA), bivalent lncRNA, and CTCF associated, among others. eRNAs are transcribed in 8.6% of regular enhancers and 36.5% of super enhancers, associated with coding genes that participate in critical immune regulatory pathways. Plasma cells uniquely featured high levels of circular RNAs accounted for by and reflecting the combinatorial clonal state of the Immunoglobulin loci.

overall_design: Detection and funtional study of novel and annotated expressed lncRNAs during human humoral immune response

first: Xabier

last: Agirre

first: Yanwen

last: Jiang

first: Leire

last: Garate

first: Ashley

middle: S

last: Doane

first: Zhuoning

last: Li

first: Akanksha

last: Verma

first: Bruno

last: Paiva

first: José

middle: I

last: Martín-Subero

first: Olivier

last: Elemento

first: Christopher

middle: E

last: Mason

first: Felipe

last: Prosper

first: Ari

last: Melnick


title: Global DNA methylation remodeling during direct reprogramming from fibroblast to neuron [MethylC-seq]

first: Joseph

middle: R

last: Ecker

address: 10010 North Torrey Pines Road

department: Genomic Analysis Laboratory

institute: HHMI-Salk-Institute

summary: Direct reprogramming from fibroblasts to neurons induces widespread cellular and transcriptional reconfigurations. In this study, we characterized global epigenomic changes during direct reprogramming using whole-genome base-resolution DNA methylome (mC) sequencing. We found that the pioneer transcription factor Ascl1 alone is sufficient for inducing robust non-CG methylation (mCH) accumulation in reprogrammed cells, but co-expression of Brn2 and Mytl1 was required to establish a global mCH pattern reminiscent of mature cortical neurons. Ascl1 alone induced strong promoter CG methylation (mCG) of fibroblast specific genes, while BAM overexpression additionally targets a competing myogenic program and directs a more faithful conversion to neuronal cells. Ascl1 induces local demethylation at its binding sites. Surprisingly, co-expression with Brn2 and Mytl1 inhibited the ability of Ascl1 to induce demethylation, suggesting a contextual regulation of transcription factor - epigenome interaction. Finally, we found that de novo methylation by DNMT3A is required for efficient neuronal reprogramming.

overall_design: MethylC-seq of direct reprogramming from fibroblast to neuron

first: Chongyuan

last: Luo

first: Qian

middle: Y

last: Lee

first: Orly

last: Wapinski

first: Rosa

last: Castanon

last: Nery

first: Sean

middle: M

last: Cullen

first: Margaret

last: Goodell

first: Howard

last: Chang

first: Marius

last: Wernig


title: Refinement of BMYB function in response to androgenic stimulation reveals genomic and non-genomic roles involving possible fluid-phase transitions

city: Campinas

first: Rafaela

last: Rosa-Ribeiro

address: Charles Darwin st, s/n

country: Brazil

institute: State University of Campinas

summary: We found a transient increase of BMYB in the prostate gland after castration and down regulation after LNCaP cells were treated to androgen (R1881). The BMYB interacts with chromatin forming large aggregates.

overall_design: Examination BMYB binding site thought ChIP-seq of LNCaP cell treated or not with androgen R1881 for 12 hours.


title: RNA-seq analysis of whole liver derived from endothelial specific TAK1 conditional knockout mice.

status: Public on Jan 11 2019

city: Mitaka

first: Jun-ichi

last: Suehiro

state: Tokyo

address: 6-20-2, Shinkawa

department: Department of Pharmacology and Toxicology

institute: Kyorin University School of Medicine

summary: In this study we show that TGFβ-activated kinase 1 (TAK1), is crucial for ensuring EC survival on TNFα stimulation. EC-specific TAK1-knockout in adult mice resulted in rapid death of the animal due to destruction of intestinal and liver blood vessels. TNFα from myeloid cells responding to intestinal microbiota was responsible for EC death in the intestine and liver. Our results illuminate mechanisms ensuring survival of intestinal and liver EC under physiological conditions

overall_design: Liver mRNA expression profiles derived from VEcadCreERT2/TAK1flox/flox mice injected by tamoxifen were generated by using HiSeq 2500 in triplicate.

first: Naito

last: Hisamichi

first: Suehiro

last: Jun-ichi


title: Next generation sequencing (NGS) facilitates quantitative analysis of miRNA expression

city: Moscow

first: Natalia

last: Baulina

address: Ostrovityanov 1

department: Molecular biology and medical biotechnology

country: Russia

institute: Pirogov Russian National Research Medical University

summary: Using NGS approach we performed the search of multiple sclerosis-related miRNAs. We used PBMC as an informative and easily accessible biological material. To exclude bias in miRNA expression levels caused by disease modifying therapies, miRNA profiling was performed in treatment-naïve RRMS patients. Taking into account hypothetic gender specificity in disease pathogenesis we compared miRNA expression in RRMS patients and HCs separately for men and women. MiRNA profiling in men identified 32 differentially expressed miRNAs, which passed threshold for multiple corrections and may be attributed to MS-related. At the same time we did not find well-defined MS-specific miRNA expression signatures in women using NGS

overall_design: We performed miRNA profiling in peripheral blood mononuclear cells of 16 relapsing-remitting MS (RRMS) patients and 8 healthy controls (HC).

first: Ivan

last: Kiselev

first: German

last: Osmak


title: Circulating Tumor Cells Clustering Triggers DNA Methylation Changes to Enable Metastasis Seeding

city: Zurich

first: Francesc

last: Castro-Giner

address: Otto-Stern-Weg 7

institute: ETH Zurich

summary: The ability of circulating tumor cells (CTCs) to form clusters has been linked to increased metastatic potential. Yet biological features and vulnerabilities of CTC clusters remain largely unknown. Our DNA methylation analysis led us to hypothesize that CTC clusters are characterized by active TF networks that support both stemness and proliferation. To identify whether the stemness- and proliferation-related TF networks are also transcriptionally active in CTC clusters compared to single CTCs, we performed single-cell resolution RNA sequencing analysis of single CTCs and CTC clusters, matched within individual liquid biopsies and isolated from six breast cancer patients with progressive metastatic disease, and of single CTCs and CTC clusters isolated from three xenograft models. In addition, among 2,486 FDA-approved compounds, we identify Na+/K+ ATPase inhibitors that enable the dissociation of CTC clusters into single cells, leading to DNA methylation remodeling at critical sites and metastasis suppression. We performed RNA sequencing analysis of BR16 and Brx50 cells upon treatment to assess the molecular consequences of clusters dissociation.

overall_design: The ability of circulating tumor cells (CTCs) to form clusters is key to accelerate the metastatic process. Yet, biological features and vulnerabilities of CTC-clusters remain largely unknown. Here, we profile DNA methylation and RNA expression landscape of single CTCs and CTC-clusters, matched within individual breast cancer patients and mouse models. We find that clustering of CTC leads to a remarkable hypomethylation of binding sites for stemness-related transcription factors. Through a screen with 2’486 FDA-approved compounds, we identify drugs that are able to dissociate CTC-clusters into single cells, leading to DNA re-methylation at critical sites and to a reduction in metastasis-seeding ability. Thus, clustering of CTCs favors accessibility to binding sites of stemness-related transcription factors, and cluster-dissociating compounds enable to suppress the spread of cancer.

first: Gkountela

last: Sofia

first: Castro-Giner

last: Francesc

first: Szczerba

last: Barbara

first: Vetter

last: Marcus

first: Landin

last: Julia

first: Scherrer

last: Ramona

first: Krol

last: Ilona

first: Beisel

last: Christian

first: Stirnimann

first: Kurzeder

first: Heinzelmann-Schwarz

last: Viola

first: Rochlitz

last: Christoph

first: Weber

last: Walter Paul

first: Aceto

last: Nicola


title: Single Cell Sequencing Reveals Gene Expression Signatures Associated with Bone Marrow Stromal Cell Subpopulations and Time in Culture [NGS_bulk cell RNA-seq]

status: Public on Jan 10 2019

city: BETHESDA

first: Shutong

last: Liu

state: Maryland

address: 9000 Rockville Pike

department: Clinical Center

institute: National Institutes of Health

summary: Background Bone marrow stromal cells (BMSCs) are a heterogeneous population that participates in wound healing, immune modulation and tissue regeneration. Next generation sequencing was used to analyze transcripts from single BMSCs in order to better characterize BMSC subpopulations. Methods Cryopreserved passage 2 BMSCs from one healthy subject were cultured through passage 10. The transcriptomes of bulk BMSCs from designated passages were analyzed with microarrays and RNA sequencing (RNA-Seq). For some passages, single BMSCs were separated using microfluidics and their transcriptomes were analyzed by RNA-Seq. Results Transcriptome analysis microarray and RNA-Seq of unseparated BMSCs from passages 2, 4, 6, 8, 9 and 10 yielded similar results; both data sets grouped passages 4 and 6 and passages 9 and 10 together and genes differentially expressed among these early and late passage BMSCs were similar. 3D Diffusion map visualization of single BMSCs from passages 3, 4, 6, 8 and 9 clustered passage 3 and 9 into two distinct groups, but there was considerable overlap for passage 4, 6 and 8 cells. Markers for early passage, FGFR2, and late passage BMSCs, PLAT, were able to identify three subpopulations within passage 3 BMSCs; one that expressed high levels of FGFR2 and low levels of PLAT; one that expressed low levels of FGFR2 and high levels of PLAT and one that expressed intermediate levels of FGFR2 and low levels of PLAT. Conclusions Single BMSCs can be separated by microfluidics and their transcriptome analyzed by next generation sequencing. Single cell analysis of early passage BMSCs identified a subpopulation of cells expressing high levels of FGFR2 that might include skeletal stem cells.

first: David

last: Stroncek

first: Yingdong

last: Zhao

first: Victoria

last: Chen

first: Rongye

last: Shi

first: Jinguo

first: Jiaqiang

last: Ren

first: Hui

first: Hee

middle: J

last: Bae

first: Steven

last: Highfill

first: Ping

last: Jin


title: Systems genetics identifies a macrophage cholesterol network that associates with physiological wound healing

city: Singapore

first: Aida

last: Moreno-Moral

address: 8 College Road

institute: Duke-NUS Medical School

summary: We generated a genomic and phenotypic resource comprising genetically outbred mice in which we measured (i) quantitative differences in wound healing indicated as ear area (ii) bone marrow derived macrophage (BMDM) mRNA expression by RNA-sequencing (RNA-seq), (iii) genome-wide SNPs genotyping by low-coverage sequencing.We performed gene co-expression network analysis and we identified a network in macrophages enriched in cholesterol genes which is genetically controlled by Runx2 gene. In vivo pharmacological blockage of Fasn with cerulenin showed delayed wound healing in rats and increased macrophage recruitment to the wound.

overall_design: We carried out RNA-sequencing in BMDMs from 146 outbred mice. We performed gene co-expression network analysis and we integrated the data with phenotipic data (wound healing) and with genotype.

first: Marta

last: Bagnati

last: Imprialou

first: Enrico

last: Petretto

first: Jacques

last: Behmoaras


title: ZNF445 is a primary regulator of genomic imprinting

city: Lausanne

first: Julien

last: Duc

state: VAUD

address: Station 19 CH-1015 Lausanne

department: School of Life Science

institute: EPFL

summary: ZFP445/ZNF445 collaborates with ZFP57 to control genomic imprinting in vivo in mouse and in human embryonic stem cells. We determined that ZNF445/ZFP445, a KZFP that can be traced back to marsupials, binds human and murine ICRs, and together with ZFP57 correctly maintains all but one murine ICR in vivo. Furthermore, the early embryonic expression of human ZNF445 and its intolerance to loss of function mutations in the population suggest a critical role in human development.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Study of ZFP445 ChIP-seq in different mouse genomic backgrounds and ZFP57 and KAP1 ChIP-seq in human. All with their respective inputs.

first: Andrea

last: Coluccio


city: Harbin

first: Xu

last: Shouping

address: No.157 Baojian Road

institute: Harbin Medical University

summary: Identification of a novel MYC-TINCR-EGFR signaling pathway loop which initiates receptor tyrosine kinases cascade and confers tumorigenesis in human cancer through examination of transcription factor C-MYC binding in 2 cell types.

overall_design: Examination of C-MYC binding in 2 cell types.

first: Shouping

last: Xu

first: Da

last: Pang


title: Single Cell Sequencing Reveals Gene Expression Signatures Associated with Bone Marrow Stromal Cell Subpopulations and Time in Culture

first: Shutong

last: Liu

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

overall_design: Refer to individual Series


title: Genome-wide chromatin accessibility profile of zebrafish cardiomyocytes throughout heart development

city: Warsaw

first: Michal

last: Pawlak

address: Trojdena 4

institute: International Institute of Molecular and Cell Biology

summary: By using transgenic zebrafish lines Tg(nxk2.5:GFP) (Witzel et al. 2012) and Tg(myl7:EGFP) (D'Amico et al. 2007), we have characterized  chromatin accessibility of FACS-isolated CM (GFP+) from developing zebrafish heart at 24, 48 and 72 hpf, corresponding to heart tube formation, chamber formation and differentiation and heart maturation, respectively. GFP- cells were used as a control. We have identified cardiac regulatory networks playing a crucial role in heart morphogenesis. To validate their importance in heart development, we employed zebrafish mutants of cardiac transciption factors Gata5, Hand2 and Tbx5a, the disruption of which were previously linked to impaired migration of the cardiac primordia to the embryonic midline, reduced number  of  myocardial  precursors and failure of heart looping, respectively (Reiter et al. 1999; Yelon et al. 2000; Garrity et al. 2002). ATAC-seq was performed from homozygous gata5tm236a/tm236a, tbx5am21/ m21, hand2s6/s6 mutant 72 hpf embryos in Tg(myl7:EGFP) genetic background. Homozygous mutant embryos for analyses were selected on the basis of their phenotypes of cardia bifida (gata5tm236a/tm236a, hand2s6/s6) or heart-string (tbx5am21/ m21) .

overall_design: Examination of chromatin accessibility maps of cardiomycytes at 3 stages of heart development

first: Cecilia

last: Winata


title: RNA-seq of MCF7 spheroids after macrophage infiltration.

city: Frankfurt

first: Tobias

last: Schmid

address: Theodor-Stern-Kai 7

department: Institute of Biochemistry I

institute: Goethe University Frankfurt

summary: To determine effects of primary human macrophages on breast tumor cell mRNA levels, breast carcinoma cell line MCF7 were grown as spheroids in the presence or absence of infiltrating macrophages.

overall_design: Breast tumor profiles of MCF7 spheroids +/- infiltrated macrophages were generated by deep sequencing using Illumina TruSeq technology.

first: Sofia

last: Winslow


title: PyMINEr Finds Gene and Autocrine/Paracrine Networks from Human Islet scRNAseq

city: Iowa City

first: John

last: Engelhardt

state: Iowa

address: 51 Newton Rd, 1-111 BSB

department: Developmental and Cellular Biology

institute: University of Iowa

summary: Toolsets available for in-depth analysis of scRNAseq datasets by biologists with little informatics experience is limited. Here we describe an informatics tool (PyMINEr) that fully automates cell type identification, cell type-specific pathway analyses, graph theory-based analysis of gene regulation, and detection of autocrine/paracrine signaling networks in silico. We applied PyMINEr to interrogate human pancreatic islet scRNAseq datasets and discovered several features of co-expression graphs including: concordance of scRNAseq-graph structure with both protein-protein interactions and 3D-genomic architecture; association of high connectivity and low expression genes with cell type-enrichment; and potential for graph-structure to clarify potential etiologies of enigmatic disease-associated variants. We further created a consensus co-expression network and autocrine/paracrine signaling networks within and across islet cell types from 7-datasets. PyMINEr correctly identified changes in BMP/WNT signaling associated with cystic fibrosis pancreatic acinar-cell loss. This proof-of-principle study demonstrates that the PyMINEr framework will be a valuable resource for scRNAseq analyses.

overall_design: Human islets were obtained from the integrated islet distribution program (IIDP), cultured overnight, then prepared for scRNAseq via the Fluidigm C1 platform. RNAseq was perfromed on Illumina HiSeq 2500.

first: Scott

last: Tyler


title: Efficient mRNA processing prevents gene gating-associated replication stress in mammalian cells

city: Utrecht

first: Tuncay

last: Baubec

address: Padualaan 8

department: Department of Biology

country: Netherlands

institute: Utrecht University

summary: Cellular mechanisms that safeguard genome integrity are often subverted in cancer. To identify novel cancer-related genome caretakers, we employed a convergent multi-screening strategy coupled to quantitative image-based cytometry, and ranked candidate genes according to multivariate read-outs reflecting viability, proliferative capacity, replisome integrity and DNA damage signaling. This revealed new regulators of replication stress resilience, including components of the pre-mRNA cleavage and polyadenylation complex. We show that deregulation of pre-mRNA cleavage impairs replication fork speed and leads to excessive origin activity, rendering cells highly dependent on ATR kinase activity. Surprisingly, while excessive formation of RNA:DNA-hybrids under these conditions appears mainly as consequence rather than cause of DNA damage, inhibition of transcription rescued fork speed, origin activation and alleviated replication catastrophe. Importantly, uncoupling pre-mRNA processing from nuclear export by depleting the THO complex also protected cells from DNA damage, suggesting that efficient pre-mRNA cleavage provides a mechanism to prevent gene gating-associated genomic instability.

overall_design: RNA Polymerase II phospho-S2 ChIP in U-2 OS cells treated with control siRNA or siRNA against WDR33

first: Matthias

last: Altmeyer


title: Analysis of gene expression dynamics during iPS cell derivation from mouse embryonic fibroblasts using reprogramming systems with different Klf4 stoichiometry

city: Kyoto

first: Knut

last: Woltjen

address: 53 Kawahara-Cho, Shogoin, Sakyo-ku

department: Reprogramming Science

institute: CiRA, Kyoto University

summary: The forced expression of Yamanaka factors (Oct3/4, Sox2, Klf4, and c-Myc) reprograms cells into induced pluripotent stem cells (iPSCs) through a series of sequential cell fate conversions. The order and robustness of gene expression changes are highly depended on the Yamanaka factor stoichiometry. We specifically focused on two different reprogramming paths induced by high- and low-Klf4 stoichiometry, which were accomplished by introducing OK+9MS or OKMS polycistronic cassettes, respectively, into mouse embryonic fibroblasts. By comparing these reprograming intermediates with embryonic stem cells (ESCs) and primary keratinocytes, we identified high-Klf4 specific, transiently up-regulated epithelial genes. We found that expression of these epithelial genes was enriched in a TROP2-positive cell population. Moreover, we identified a set of transcription factors which are candidates for the regulation of transiently expressed epithelial genes, and revealed their connection to high-Klf4-specific reprogramming hallmarks.

overall_design: 20 samples were analyzed. MEFs were transfected with OKMS (Low-Klf4) or OK+9MS (High-Klf4), followed by FACS sorting for transgene-positive mCherry+ cells (d2, d4, d8, d18) or TROP2+ cells (d8). FACS sorted mCherry+ cells (d6) were submitted previously (GSE65468). A mouse embryonic stem cell, iPS clones derived from each reprogramming methods (OKMS: A7 and A9, OK+9MS: A10 and H10) and primary keratinocyte were used as references for intermediate reprogramming populations.

first: Harunobu

last: Kagawa

first: Shin-Il

last: Kim

first: Takuya

last: Yamamoto


title: Genome-Wide DNA Methylation Profiling of Oesophageal Cancer-Associated Myofibroblasts

city: Liverpool

first: Hanna

last: Najgebauer

address: Crown Street

department: Cellular and Molecular Physiology

institute: University of Liverpool

summary: The Illumina Infinium HumanMethylation450 BeadChip arrays were performed on a collection of primary oesophageal cancer-associated myofibroblasts (CAM) and their patient-matched adjacent tissue myofibroblasts (ATM). CAM and ATM samples were obtained from patients with oesophageal adenocarcinomas undergoing cancer surgery.

overall_design: Bisulphite converted DNA from 3 oesophageal cancer patients, including 3 CAMs and 3 ATMs, were hybridised to the Illumina Infinium HumanMethylation450 BeadChip

first: Olivier

last: Giger


title: DNA IP for 5hmC modified CpGs in DNA of ESC derived hepatocyte like cells exposed to high energy substrates.

city: Edinburgh

first: Marcus

middle: James

last: Lyall

state: Lothian

address: The Queen's Medical Research Institute, 47 Little France Crescent

department: University/British Heart Foundation Centre for Cardiovascular Science

institute: University of Edinburgh

summary: We report the application of antibody affinity DNA immunoprecipitation followed by proton sequencing to map 5'hydroxymethylcytosine profiles in DNA of ESC derived hepatocyte like cells exposed to high energy substrates.

overall_design: Female H9 ESCs were maintained in mTeSR1 (Stem Cell Technologies, Cambridge UK) on matrigel (Corning, Tewksbury, USA) and differentiated into hepatocyte-like cells as previously described (Szkolnicka D, Curr Protoc Stem Cell Biol 2014;30:1G 5 1-12.). At day 20 of differentiation, cells were exposed to Sodium L-Lactate (L7022, Sigma, Gillingham, UK) 10mM, Sodium Pyruvate 1mM (Sigma, Dorset, UK) and Octanoic Acid (C2875, Sigma, Gillingham, UK) 2mM for 48 hours before harvesting.  DNA was extracted using quigen DNAeasy kit.  We carry out 5-hydroxymethylation DNA immunoprecipitation (hmeDIP) prior to sequencing Ion Proton P1 to report locus specific changes in 5hmC induced by LPO treatment.

middle: P

last: Thomson

first: Amanda

last: Drake

first: David

last: Hay


title: Argonaute2 attenuates active transcription by limiting RNA Polymerase II elongation in Drosophila melanogaster [ChIPseq-CDK9]

city: Bethesda

first: Cameron

last: Palmer

state: MD

address: 50 South Drive

department: National Institute of Diabetes and Digestive and Kidney Diseases

summary: Increasing lines of evidence support that Argonaute2 (AGO2) harbors several nuclear functions in metazoa. In particular, Drosophila AGO2 modulates transcription of developmentally regulated genes; however, the molecular mechanisms behind AGO2 recruitment into chromatin and its function in transcription have not been deeply explored. In this study, we show that Drosophila AGO2 chromatin association depends on active transcription. In order to gain insight into how AGO2 controls transcription, we performed differential ChIP-seq analysis for RNA Polymerase II (Pol II) upon depletion of AGO2. Interestingly, we find specific accumulation of the elongating but not initiating form of Pol II after AGO2 knockdown, suggesting that AGO2 impairs transcription elongation. Finally, AGO2 also affects Negative Elongation Factor (CDK9) chromatin association but not the Cyclin Dependent Kinase 9 (CDK9). Altogether, these results provide key insights into the molecular role of AGO2 in attenuating elongation of certain actively transcribed genes.

overall_design: ChIP-seq of CDK9 in Drosophila melanogaster Kc167 cells in mock treatment and Ago2 knockdown. Two replicates per condition, with paired inputs.

first: Ezequiel

last: Nazer

middle: D

first: Elissa

last: Lei


title: Genome wide expression change by RNF168 knocking down in NEC cells

city: Shenzhen

first: Qiangzu

last: Zhang

address: Nan Hai Da Dao Chuang Shi Ji Bin Hai Hua Yuan Building9 9A , Nanshan District, Shenzhen City, Guangdong Province

institute: Lemondata BioTechnology Limited

summary: We aim to the investigate the role of RNF168 in breast cancer progression. NEC cells were used as the model and RNF168 was silenced by siRNA.

overall_design: The NEC cells were treated with 50nM stramble siRNA and siSMURF1. After 48 hours, cells were harvested and the total RNA was extacted by Qiagen kit. The RNA sample was sent to BGI for RNA expression anaylsis.

first: J

last: Zhu


title: Encapsulated miR-200c and Nkx2.1 in a nuclear/mitochondria transcriptional regulatory network of non-metastatic and metastatic lung cancer cells

city: Boston

first: Jean-Bosco

last: Tagne

state: MA

address: 72 East Concord

department: Medicine/Pulmonary

institute: Boston University

summary:
Background: MicroRNAs are noncoding RNA molecules of ~22 nucleotides with diagnostic and therapeutic action, affecting the expression of mRNAs involved in invasion, migration, and development. miR-200c is part of the miR-200c/141 cluster on chromosome 12p13. Its mechanism of action when encapsulated is critical in lung cancer when patients express changes in miRNAs. miR-200c be a potential biomarkers for various lung diseases. As a potential therapy, miR-200c can impact lives as target lung cancer is a leading cause of death with about 234,000 cases annually, high heterogeneity, complex screening, and a 5-year survival rate of 16%. Encapsulated miR-200c efficiently enhances bioavailability, pharmacokinetics of therapeutics and targeting to cells, improves efficacy and provides potential cure.
Methods: The functions of miR-200c were determined in non-metastatic KW-634 and metastatic 821-T4 and 821-LN mouse lung cancer cell lines after various nano vehicle treatments. Viability and cytotoxicity were determined by cell cycle and quantitative real-time PCR analyses were used to quantify levels of miR-200c and its target genes. In situ hybridization was used to visualize patterns of expression of miR-200c and others in the lung and many organs. Next-generation sequencing, invasion and migration assays using transwell chambers, and ActivSignal were used to elucidate the activation and inhibition profiles and perform direct expression measurements and modification of cellular components.

overall_design: 18 samples in triplicates. 3 controls and 3 treatments for each of the 3 cancer cells.

first: Olga

last: D’Almeida

first: Omar

last: Mothar

first: Esther

middle: A

last: Bondzie

first: Yolande

last: Lieumo

first: Laure

first: Sumeet

last: Gupta

first: Thomas

last: Volkert

first: Stuart

last: Levine

first: Jean

middle: B


title: Single Cell Sequencing Reveals Gene Expression Signatures Associated with Bone Marrow Stromal Cell Subpopulations and Time in Culture [scRNA-seq]

first: Shutong

last: Liu

overall_design: Cryopreserved passage 2 BMSCs from one healthy subject were cultured through passage 10. The transcriptomes of bulk BMSCs from designated passages were analyzed with microarrays and RNA sequencing (RNA-Seq). For some passages, single BMSCs were separated using microfluidics and their transcriptomes were analyzed by RNA-Seq.

middle: F

last: Stroncek

first: Yingdong

last: Zhao

first: Victoria

last: Chen

first: Rongye

last: Shi

first: Jinguo

first: Jiaqiang

last: Ren

first: Hui

first: Hee

last: Bae

first: Steven

last: Highfill

first: Ping

last: Jin


title: Genome-Wide DNA Methylation Profiling of Gastric Cancer-Associated Myofibroblasts

first: Hanna

last: Najgebauer

summary: The Illumina Infinium HumanMethylation450 BeadChip arrays were performed on a collection of primary gastric cancer-associated myofibroblasts (CAM) and their patient-matched adjacent tissue myofibroblasts (ATM) as well as unrelated normal tissue myofibroblasts (NTM). CAM and ATM samples were obtained from patients undergoing gastric cancer surgery; whereas NTM samples were generated from deceased transplant donors with normal morphology.

overall_design: Bisulphite converted DNA from the 9 samples, including 3 CAMs, 3 ATMs and 3 NTMs, were hybridised to the Illumina Infinium HumanMethylation450 BeadChip

first: Chris

last: Sanderson


title: Expression data from oral squamous cell carcinomas, 3A and HO1-N-1

status: Public on Jan 09 2019

city: Bunkyo-ku

first: Akane

last: Wada

address: 1-5-45 Yushima

department: Oral Pathology

institute: Tokyo Medical and Dental University

summary:
Co-culture with human oral squamous cell carcinoma cells, 3A or NEM, or culture with each of their conditioned medium induced many osteoclasts from osteoclast precursor cells, which were generated by a 24-h pretreatment of RANKL or TNF-α.  However, HO1-N-1 cells did not induce any osteoclasts. Osteoprotegerin, a decoy RANKL receptor, denosumab, an anti-RANKL antibody drug, and infliximab, an anti-TNF-α antibody drug, did not prevent this tumor-associated osteoclastogenesis.
We compared the expression of molecules associated with osteoclastogenesis between 3A and HO1-N-1 cells.

overall_design: 3A and HO1-N-1 cells were cultured at sub-confluent condition, cells were harvested and extracted total RNA. Gene expression microarray analysis using the Human Gene 2.0 ST Array (Affymetrix, Santa Clara, CA) was consigned to Filgen Inc., Nagoya, Japan. We compared the expression of molecules associated with osteoclastogenesis, IL-1β, IL-6, IL-8, PTHrP, RANKL and TNF-α between 3A and HO1-N-1 cells.

first: Tohru

last: Ikeda

first: Kou

last: Kayamori

first: Kei

last: Sakamoto


title: Genome-wide co-localization of RNA-DNA interactions and fusion RNA pairs (iMARGI)

city: La Jolla

first: Sheng

last: Zhong

address: 9500 Gilman Drive MC 0412

department: Bioengineering

institute: University of California, San Diego

summary: Despite the ever-increasing speed of detecting fusion transcripts in cancer, it remains formidable to predict what unreported RNA pairs can form new fusion transcripts. By systematic mapping of chromatin-associated RNAs (caRNAs) and their respective genomic interaction loci, we obtained genome-wide RNA-DNA interaction maps from two non-cancerous cell types. The gene pairs involved in RNA-DNA interactions in these normal cells exhibited strong overlap with those with cancer-derived fusion transcripts. These data suggest an RNA-poise model, where the spatial proximity of one gene’s transcripts and the other gene’s genomic sequence poises for the creation of fusion transcripts. We validated this model with 96 additional lung cancer samples. One of these additional samples exhibited fusion transcripts without a corresponding fusion gene, suggesting that genome-recombination is not a required step of the RNA-poise model.

overall_design: Map RNA-DNA interactions in two human cell lines, HEK293T and HFFc6 using iMARGI technique.

first: Zhangming

last: Yan

first: Norman

last: Huang

first: Weixin

last: Wu

first: Weizhong

first: Yiqun

last: Jiang

first: Jingyao

first: Xuerui

first: Xingzhao

last: Wen

first: Kang

last: Zhang

first: Zhen

first: Shu

last: Chien


title: Genome-wide co-localization of RNA-DNA interactions and fusion RNA pairs


title: RNA-seq and ChIP-seq analyses of epididymis from ArKI mice

city: Kuopio

first: Einari

last: Niskanen

address: Yliopistonranta 8

department: School of Medicine

country: Finland

institute: University of Eastern Finland


title: Genome-wide maps of SNIP1 in 293T

city: Shanghai

first: Fei

last: Lan

address: No131 Dongan Road, Xuhui District.

institute: Institute of Biochemical Science

summary: We found SNIP1 occupied at promoters and there is an overlap betwween and TET2- and SNIP1-binding events.

overall_design: Examination of genome-wide maps of SNIP1 in 293T overexpressing HA-SNIP1.

first: Leilei

last: Chen

first: Chenxi

last: He


title: ChIP-seq analysis of androgen receptor binding in epididymis of ArKI mice

summary: To analyze how SUMOylation of AR affects its chromatin binding, we generated a mouse model (ArKI) in which the two conserved SUMO acceptor lysines of androgen receptor (AR) were permanently abolished by converting them to arginines (ArK381R, K500R). ChIP-seq was used to analyse how the lack of AR SUMOylation affects the chromatin binding of AR in epididymis.

overall_design: For each sample, caput epididymides (containing initial segments) from three wild type (WT) or ArKI (KI) mice were combined, chromatin binding of AR was analyzed using ChIP and sequenced using Illumina HiSeq 3000 system. We generated two replicates from WT and ArKI mice.

first: Jorma

last: Palvimo

first: Marjo

last: Malinen

first: Petra

last: Sipilä


title: Genome-wide mapping of RTA binding sites in KSHV+ PEL cells during KSHV reactivation.

city: Gainesville

first: Zsolt

last: Toth

state: FL

institute: University of Florida

summary: Chromatin immunoprecipitation coupled with high-throughput sequencing (ChIP-seq) analysis was performed during Kaposi's sarcoma-associated herpesvirus (KSHV) reactivation in KSHV+ recombinant primary effusion B-cell lymphoma cells (PEL). RTA binding sites were identified genome-wide in a recombinant PEL cell line called TRExBCBL1-3xFLAG-RTA cells at 12 hours post-induction (hpi) of RTA expression.

overall_design: The expression of the N-terminally 3xFLAG-tagged RTA was induced by adding 1 μg/ml doxycycline (Dox) to the medium for 12 hours. Cells were cross-linked with formaldehyde and sonicated chromatin prepared.  Chromatin immunoprecipitation was performed using FLAG antibody to detect the FLAG epitope tagged RTA protein in the recombinant PEL cells at 12 hpi. Libraries were generated from the genomic DNA material before immunoprecipitation (Input)  and from the FLAG-RTA ChIP sample in paralel.

first: Bernadett

last: Papp


title: Global host gene expression changes in KSHV+ PEL cells upon KSHV reactivation

summary: Kaposi's sarcoma-associated herpesvirus (KSHV) is a human oncogenic virus, which maintains the persistent infection of the host by intermittently reactivating from latently infected cells to produce viral progenies. Here, we performed a comprehensive time course transcriptome analysis during KSHV reactivation in KSHV+ primary effusion B-cell lymphoma cells (PEL). For this we used a recombinant PEL cell line called TRExBCBL1-3xFLAG-RTA. The expression of the N-terminally 3xFLAG-tagged RTA was induced by adding 1 μg/ml doxycycline (Dox) to the medium. Total gene expression changes were identified in TRExBCBL1-3xFLAG-RTA cells at 0, 6, 12, and 24 hours post-induction (hpi).

overall_design: RNA-seq libraries were generated from recombinant PEL cells (called TRExBCBL1-3xFLAG-RTA). In total, 12 samples were collected following doxycycline treatment to induce RTA expression for 0, 6, 12 or 24 hours, in triplicate.


title: LincK promotes proliferation and epithelial-to-mesenchymal transition and contributes to tumorigenesis and growth in breast cancer I

city: San Diego

first: Yajing

last: Hao

state: CA

address: George Palade Laboratories 9500 Gilman Drive, Room 217

department: Dept. of Cellular and Molecular Medicine

summary:
In this study, we aimed to identify potential lncRNA deregulations associated with breast cancer malignancy instigated by MSCs-MCF7 co-culture. We profiled expression changes of lncRNAs in MCF-7 cells during EMT induced by coculture with hAD-MSCs, and found an intergenic lncRNA with proviouly unknown function (KB-1732A1.1, we termed it LincK), which was significantly elevated. Depletion of LincK decreased the growth, migration, invasion, and EMT in breast cancer cells, while overexpression of LincK exerted the opposite effects. Moreover, knockdown of LincK repressed tumorigenesis, and ectopic expression of LincK promoted tumor growth in MCF-7 xenograft model. LincKs can act as a sponge of mirR-200b, which inhibits its function in proliferation and metastasis. Thus we reported, for the ﬁrst time, the role of LincK in control of EMT and proliferation in breast cancer.
We used microarrays to detail the genes associated with LincK.

overall_design: Transcriptome analysis of LincK siRNA-knockdown in cultured MCF-7 cells.To detect the targeting of LincK, we used microarray to analyze changes in gene expression patterns after siRNA transfection in MCF-7 cells. 3 independent biological replicates were plated, transfected in parallel for each control and siRNA. Two different siRNAs were used.

first: Jing

last: Li

first: Wenzhe

last: Mao

first: Xiaowei

last: Xue

first: Zhao

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