title: Wnt signaling regulates neural plate patterning in distinct temporal phases with dynamic transcriptional outputs

city: Houston

first: Arne

last: Lekven

state: TX

address: 3507 Cullen Blvd

department: Biology and Biochemistry

institute: University of Houston

summary: The process that partitions the nascent vertebrate central nervous system into forebrain, midbrain, hindbrain, and spinal cord after neural induction is of fundamental interest in developmental biology, and is known to be dependent on Wnt/beta-catenin signaling at multiple steps. Neural induction specifies neural ectoderm with forebrain character that is subsequently posteriorized by graded Wnt signaling: embryological and mutant analyses have shown that progressively higher levels of Wnt signaling induce progressively more posterior fates.  However, the mechanistic link between Wnt signaling and the molecular subdivision of the neural ectoderm into distinct domains in the anteroposterior (AP) axis is still not clear.  To better understand how Wnt mediates neural AP patterning, we performed a temporal dissection of neural patterning in response to manipulations of Wnt signaling in zebrafish. We show that Wnt-mediated neural patterning in zebrafish can be divided into three phases: (I) a primary AP patterning phase, which occurs during gastrulation, (II) a mes/r1 (mesencephalon-rhombomere 1) specification and refinement phase, which occurs immediately after gastrulation, and (III) a midbrain-hindbrain boundary (MHB) morphogenesis phase, which occurs during segmentation stages. A major outcome of these Wnt signaling phases is the specification of the major compartment divisions of the developing brain: first the MHB, then the diencephalic-mesencephalic boundary (DMB). The specification of these lineage divisions depends upon the dynamic changes of gene transcription in response to Wnt signaling, which we show primarily involves transcriptional repression or indirect activation. We show that otx2b is directly repressed by Wnt signaling during primary AP patterning, but becomes resistant to Wnt-mediated repression during late gastrulation. Also during late gastrulation, Wnt signaling becomes both necessary and sufficient for expression of wnt8b, en2a, and her5 in mes/r1. We suggest that the change in otx2b response to Wnt regulation enables a transition to the mes/r1 phase of Wnt-mediated patterning, as it ensures that Wnts expressed in the midbrain and MHB do not suppress midbrain identity, and consequently reinforce formation of the DMB.  These findings integrate important temporal elements into our spatial understanding of Wnt-mediated neural patterning and may serve as an important basis of a better understanding of neural patterning defects that have implications in human health.

overall_design:
Total RNA seq profiles on zebrafish embroys either wt,hs:dkk1b/+ or hs:wnt8a/+. Samples were run in triplicate except for hs:wnt8a at 5 hpf was run in duplicate. hs:dkk1b/+ and hs:wnt8a/+ transgenic fish were crossed to WT strains. Embryos were collected and heat shocked two time points, 5/5.5 hpf (time 1) or 7/7.5 hpf (time 2) for hs:dkk1 and hs:wnt8a

Pleae note that the samples with the "wt xxx' titles are genotypically wild type sibling embryos to the experimental group and were heat shocked with their transgenic siblings.

first: David

last: Green

first: Amy

middle: E

last: Whitener

first: Saurav

last: Mohanty

first: Brandon

last: Mistretta

first: Preethi

last: Gunaratne

first: Alvin

last: Yeh


title: The effect of oncogenic KrasG12D on the expression of inflammation-related genes in myeloid cells

status: Public on Feb 29 2020

city: Freiburg

first: Geoffroy

last: Andrieux

address: Breisacherstr 153

country: Germany

institute: University clinics Freiburg

summary: Oncogenic Ras mutations occur frequently in hematological malignancies. It was unclear if besides the direct transforming effect via constant RAS/MEK/ERK signaling, an inflammation-related effect of KRAS contributes to the development of the disease. We used microarray-based analysis of murine hematopoietic bone marrow (BM) cells harboring active inducible KrasG12D to reveal the expression of inflammation-related genes.

overall_design: 8 samples each of them from an individual mouse (biological replicate) were analyzed

first: Shaima'a

last: Hamarsheh

first: Robert

last: Zeiser

first: Melanie

last: Boerries


title: Single Cell Transcriptomics Identifies Stem Cell-Derived Graft Composition in a Model of Parkinson’s Disease

status: Public on Mar 01 2020

city: Uppsala

first: Asa

middle: Kristina

last: Bjorklund

address: Box 596

department: Cell and Molecular Biology

country: Sweden

institute: Uppsala University

type: Expression profiling by high throughput sequencing

summary: Cell replacement is a long-standing and realistic goal for the treatment of Parkinson’s disease (PD). Cells for transplantation can be obtained from fetal brain tissue or from stem cells.  However, after transplantation dopamine (DA) neurons are a minor component of grafts and it has remained difficult to determine the identity of other cell types. Here, we report analysis by single cell RNA sequencing (scRNA-seq) combined with comprehensive histological analyses to characterize intracerebral grafts from human embryonic stem cells (hESCs) and fetal tissue after functional maturation in a pre-clinical rat PD model. We show that neurons and astrocytes are major components in both fetal and stem cell-derived grafts. Additionally, we identify a cell type closely resembling a class of newly identified perivascular-like cells in stem cell-derived grafts. Thus, this study uncovers previously unknown cellular diversity in a clinically relevant cell replacement PD model.

overall_design: Single cell RNA sequencing to characterize fetal and clinical grade ventral midbrain human embryonic stem cells after long-term survival in a pre-clinical rat model of PD.

first: Katarina

last: Tiklova

middle: K

first: Sara

last: Nolbrant

first: Agnete

last: Kirkeby

first: Alessandro

last: Fiorenzo

first: Linda

last: Gillberg

first: Andreas

last: Heuer

first: Malin

last: Parmar

first: Thomas

last: Perlmann

first: Yogita

last: Sharma

first: Deidre

middle: B

last: Hoban

first: Tiago

last: Cardoso

first: Andrew

middle: F

last: Adler

first: Marcella

last: Birtele

first: Hilda

last: Lunden-Miguel

first: Nikolaos

last: Volakakis


title: Transcriptional profiles of [ESI+]

city: Stanford

first: Zachary

middle: Hugh

last: Harvey

state: CA

address: 269 Campus Dr

department: Chemical and Systems Biology

country: USA

institute: Stanford University

summary: We profile the transcriptional landscapes associated with acquisition of [ESI+], a prion scaffolded by Snt1, a core component of the Set3C histone deacetylase. We find that acquisition of [ESI+] leads to expression of otherwise silent, heterochromatic loci, proposing a new molecular means by which silent transcriptional states might be reversed.

overall_design: mRNA sequencing of mid-logarithmically grown naïve and [ESI+] diploid cells

middle: H

first: Daniel

last: Jarosz


title: Transcription Profiles of [ESI+] and Set3C loss of function

summary: We report the transcriptional profiles of the [ESI+] prion, as well as snt1∆, set3∆, and hos2∆ diploid cells.

overall_design:
Binding profiles of Snt1, Rap1, and Hos2 in naïve and [ESI+] cells, as well as mRNA-seq and pan-acetyl H4 and H3

This submission corresponds to RNA-seq component of study.


title: Comparison of methylation of D492 before and after EMT in 3D culture

city: Baltimore

first: Winston

last: Timp

state: MD

address: 3400 N. Charles St. Clark 102A

department: Biomedical Engineering

institute: Johns Hopkins University

type: Methylation profiling by genome tiling array

summary: MicroRNAs (miRNAs) regulate developmental events such as branching morphogenesis, epithelial to mesenchymal transition (EMT) and its reverse process mesenchymal to epithelial transition (MET). In this study, we performed small RNA sequencing of a breast epithelial progenitor cell line (D492), and its mesenchymal derivative (D492M) cultured in three-dimensional reconstituted basement membrane (3D-rBM) matrix. Large differences in miRNA expression were seen between D492 and D492M. Among the most downregulated miRNAs in D492M was miR-203a, a miRNA that plays an important role in epithelial differentiation. When analyzed over time (21 days) in 3D culture, increased expression of miR-203a was seen in D492, concomitant with increased complexity of the branching structures. When miR-203a was overexpressed in D492M (D492MmiR-203a), a partial reversion towards epithelial phenotype was seen. Gene expression analysis of D492M and D492MmiR-203a revealed that peroxidasin (PXDN), a collagen IV cross-linker was one of the most significantly downregulated genes in D492MmiR-203a. Bioinformatic analysis revealed that PXDN has three putative binding sites for miR-203a in its 3-prime UTR. When a miR-203a mimic was transfected into D492M, reduced PXDN expression was seen. Conversely, inhibition of miR-203a in D492MmiR-203a with a miR-203a inhibitor increased PXDN expression, demonstrating that miR-203a acts as a repressor of PXDN. To verify the binding of miR-203a to the 3’-UTR of PXDN we set up a luciferase reporter assay that confirmed the binding.    Collectively, we have demonstrated that miR-203a expression temporally correlates with branching morphogenesis in 3D-rBM and is suppressed in the mesenchymal state. Overexpression of miR-203a in D492M induces a partial MET and reduces the expression of PXDN. Furthermore, we demonstrate that miR-203a is a novel repressor of PXDN. MiR-203-PXDN axis may be an important regulator in branching morphogenesis, EMT and basement membrane remodeling.  \

overall_design: 3 vs 3 biological replicates

first: Eirikur

last: Briem

first: Zuzana

last: Budkova

first: Anna

last: Sigurdardottir

first: Bylgja

last: Hilmarsdottir

first: Jennifer

last: Kricker

first: Magnus

last: Magnusson

first: Gunnhildur

last: Traustadottir

first: Thorarinn

last: Gudjonsson


title: Genes regulated by TAZ or YAP in A549 cell

city: Bunkyo-ku

first: Hideaki

last: Isago

state: Tokyo

department: Department of Respiratory Medicine

country: Japan

institute: The University of Tokyo Graduate School of Medicine

summary: To investigate the roles of TAZ and YAP in lung cancer cell proliferation, we compared the expression profiles of A549transfected with control siRNA, siTAZ, and siYAP.

overall_design: We collected RNA from A549 cell lines transfected with control siRNA, siTAZ, and siYAP.

first: Akihisa

last: Mitani

first: Yu

last: Mikami

first: Masafumi

last: Horie

first: Hirokazu

last: Urushiyama

first: Yasuhiro

last: Terasaki

first: Takahide

last: Nagase


title: TAp63-regulated microRNAs suppress cutaneous squamous cell carcinoma [miRNA-seq]

status: Public on Feb 28 2020

city: Tampa

first: Elsa

middle: Renee

last: Flores

state: FL

address: 12902 USF Magnolia Drive

department: Department of Molecular Oncology

institute: Moffit Cancer Center

type: Non-coding RNA profiling by high throughput sequencing

summary: The goal of this study is to compare the differences in the global miRNA expression of WT and TAp63-/- skin and SCC

overall_design: We performed a miRNA-Seq analysis comparing the transcriptomes of WT skin (n=3), TAp63-/- skin (n=3), WT SCC (n=2), and TAp63-/- cSCC (n=4)

first: Andrew

middle: J

last: Davis

middle: R


title: Analysis of the genomic landscape in ALK+ NSCLC patients identifies novel aberrations associated with clinical outcomes

city: Montreal

first: Gerald

last: Batist

state: Québec

address: 3755 Cote Ste-Catherine

country: Canada

institute: Jewish General Hospital

summary: Expression data of primary and metastasic tissues from ALK+ NSCLC patients (Anaplastic lymphoma kinase fusion positive non-small cell lung carcinoma)

overall_design: Total RNA from FFPE slides were sequenced using HiSeq 2500

first: Mathilde

last: Couetoux du Tertre

first: Maud

last: Marques


title: Argonaute2 and LaminB modulate gene expression by controlling chromatin topology (neuRNA-seq, set2)

city: Bethesda

first: Ryan

last: Dale

address: Rm 10D39, 10 Center Drive

department: National Institute of Child Health and Human Development

institute: National Institutes of Health

summary: Drosophila Argonaute2 (AGO2) is known to regulate expression of certain loci in an RNA interference-independent manner, but its genome-wide function on chromatin remains unknown. In order to elucidate the mechanism by which AGO2 regulates transcription, we performed mass spec analysis of factors that interact with AGO2 in nuclear extracts and identified RNA Polymerase II (Pol II) and LaminB among the top-associated proteins. We found that AGO2 and LaminB attenuate transcription genome-wide, including borders between both LaminB-associated domains (LADs) and topologically-associated domains (TADs). Furthermore, we identified a somatic target of AGO2 transcriptional repression, nht, which is immersed in a LAD located within a repressive TAD. Null mutation of AGO2 leads to ectopic expression of spermatogenesis genes dependent on nht. Finally, depletion of either AGO2 or LaminB results in reduced looping interactions within the TAD as well as ectopic inter-TAD interactions, as detected by 4C-seq analysis. Overall, our findings reveal coordination of AGO2 and LaminB to dictate overall genome architecture and thereby regulate gene expression.

overall_design: Nascent RNA-seq (neuRNA-seq) in knockdowns of insulator proteins in Drosophila Kc cells with matched mock knockdown, as well as knockdown of AGO2 with a rescue construct.

first: Ezequiel

last: Nazer

first: Elissa

last: Lei


title: Argonaute2 and LaminB repress transcription by controlling RNA Pol II recruitment and chromatin topology in Drosophila melanogaster (ChIP-Seq)

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Although best known as core components of RNA silencing pathways in the cytoplasm, Argonautes have been shown to regulate gene expression by functioning in the nucleus. Previous work showed that Drosophila AGO2 is preferentially associated with active promoters and can activate gene expression by promoting looping to an enhancer. In order to elucidate the genome-wide role of AGO2 in regulating transcription, we performed mass spec analysis of factors that interact with AGO2 in nuclear extracts and found RNA Pol II and LaminB among the top-associated proteins. Using nascent RNA sequencing, we showed that AGO2 and LaminB co-repress transcription at borders between both LaminB-associated domains (LAD) and topologically-associated domains (TADs). Furthermore, LaminB and AGO2 specifically prevent RNA Pol II recruitment at repressed genes, and AGO2 additionally impairs the ability of RNA Pol II to extend into active elongation. Using mRNA-seq profiling, we identified a somatic target of AGO2 repression, nht, which leads to ectopic expression of spermatogenesis genes in AGO2 null mutants. Interestingly, nht is immersed in a LAD located within a repressive TAD flanked by AGO2 binding sites. 4C-seq analysis showed that depletion of either AGO2 or LaminB results in a significant decrease in frequency of interactions within the TAD as well as ectopic inter-TAD interactions. Overall, our findings reveal a coordinated function for AGO2 and LaminB in dictating the overall architecture of the genome to control the recruitment of RNA Pol II and thereby regulate gene expression.

overall_design: ChIP_seq analysis for a large reportoire of factors in mock, AGO2 and LaminB knockdowns. The experiments have 2 or 3 biological replicates as specified.

middle: P


title: Bivalent promoter hypermethylation in cancer is linked to the H327me3/H3K4me3 ratio in embryonic stem cells

status: Public on Feb 23 2020

city: Edinburgh

first: Donncha

middle: S

last: Dunican

address: Western General Hospital

country: United Kingdom

institute: IGMM / MRC HGU

type: Methylation profiling by high throughput sequencing

summary:
Background
Thousands of mammalian promoters are defined by co-enrichment of the histone tail modifications H3K27me3 (repressive) and H3K4me3 (activating) and are thus termed bivalent. It was previously observed that bivalent genes in human ES cells (hESC) are frequent targets for hypermethylation in human cancers, and depletion of DNA methylation in mouse embryonic stem cells has a marked impact on H3K27me3 distribution at bivalent promoters. However, only a fraction of bivalent genes in stem cells are targets of hypermethylation in cancer, and it is currently unclear whether all bivalent promoters are equally sensitive to DNA hypomethylation and whether H3K4me3 levels play a role in the interplay between DNA methylation and H3K27me3.

Results
We report the sub-classification of bivalent promoters into two groups—promoters with a high H3K27me3:H3K4me3 (hiBiv) ratio or promoters with a low H3K27me3:H3K4me3 ratio (loBiv). HiBiv are enriched in canonical Polycomb components, show a higher degree of local intrachromosomal contacts and are highly sensitive to DNA hypomethylation in terms of H3K27me3 depletion from broad Polycomb domains. In contrast, loBiv promoters are enriched in non-canonical Polycomb components, show lower intrachromosomal contacts and are less sensitive to DNA hypomethylation at the same genomic resolution. Multiple systems reveal that hiBiv promoters are more depleted of Polycomb complexes than loBiv promoters following a reduction in DNA methylation, and we demonstrate that H3K27me3 re-accumulates at promoters when DNA methylation is restored. In human cancer, we show that hiBiv promoters lose H3K27me3 and are more susceptible to DNA hypermethylation than loBiv promoters.

Conclusion
We conclude that bivalency as a general term to describe mammalian promoters is an over-simplification and our sub-classification has revealed novel insights into the interplay between the largely antagonistic presence of DNA methylation and Polycomb systems at bivalent promoters. This approach redefines molecular pathologies underlying disease in which global DNA methylation is aberrant or where Polycomb mutations are present.

overall_design:
17 samples, 1 WTNPC; 2. LSHNPC; 3 WGBS-Ser; 4 WGBS-Dox; 5 WGBS-Rev; 6 K27-Ser;  7 K27-Dox; 8 K27-Rev; 9 K4-Ser;  10 K4-Dox; 11 K4-Rev; 12 WGBS-Cl36; 13 WGBS-Eed; 14 WGBS-R1b; 15 WGBS-DKO; 16 J1_H3K4me3; 17 TKO_H3K4me3; J1_uH2A; J1_Kdm2b

Commercial antibodies used in this study:
H3K27me3 #07-449 Millipore
H3K4me3 #07-473 Millipore

first: Donnchadh

first: Richard

last: Meehan


title: Protein arginine methyltransferase 5 promotes cholesterol biosynthesis mediated Th17 responses and autoimmunity [CD4 T-cells]

status: Public on Feb 27 2020

city: Columbus

first: Mireia

last: Guerau

state: OH

address: 453 W 10th Ave

department: HRS-Medical Laboratory Science

institute: The Ohio State University

summary: Protein Arginine Methyltransferase (PRMT) 5 catalyzes symmetric dimethylation of arginine, a post-translational modification involved in cancer and embryonic development. However, the role and mechanisms by which PRMT5 modulates T helper (Th) cell polarization and autoimmune disease have not yet been elucidated. Here we find that PRMT5 promotes expression of cholesterol biosynthetic pathway enzymes that produce ROR agonists and activate ROR-t, driving Th17 differentiation. Specific loss of PRMT5 in the CD4 Th cell compartment completely protected mice from EAE. We also find that PRMT5 controls thymic and peripheral homeostasis in the CD4 Th cell life cycle, as well as iNK T and CD8 T cell development or maintenance, respectively. This work conclusively demonstrates that PRMT5 expression in recently activated T cells is necessary for expression of a cholesterol biosynthesis metabolic gene expression program that generates ROR-t agonistic activity and promotes Th17 differentiation and EAE. These results point to Th PRMT5 and its downstream cholesterol biosynthesis pathway as promising therapeutic targets in Th17-mediated diseases.

overall_design: Three biological replicates were analyzed for 2 conditions (Two genotypes)

first: Shouvonik

last: Sengupta


title: Protein arginine methyltransferase 5 promotes cholesterol biosynthesis mediated Th17 responses and autoimmunity [Th17]

first: Lindsay

last: Webb

first: Claudia

last: Edell

first: Zayda

middle: L

last: Piedra-Quintero

first: Stephanie

middle: A

last: Amici

first: Janiret

last: Narvaez-Miranda

first: Makenzie

last: Bevins

first: Austin

last: Kennemer

first: Georgios

last: Laliotis

first: Philip

middle: N

last: Tsichlis


title: Pooled CRISPR screens in human intestinal organoids [RNA-seq]

status: Public on Feb 26 2020

city: Zurich

first: giancarlo

last: russo

state: Schweiz

address: winterthurerstrasse 190

department: Functional Genomics Center Zurich

country: Switzerland

institute: ETH/University of Zurich

summary: We performed CRISPR screens on both a sub-library and a genome-wide scale in human intestinal organoids to discover cancer driver genes. We investigated the Wnt and the TGFB pathway and used both WT, APC-mutant and APC-TP53-mutant organoids.

overall_design: To investigate how the identified SWI/SNF screening hits confer TGFB resistance, we assessed changes in gene expression upon ARID1A or SMARCA4 perturbation and TGFB treatment using RNA-seq

first: Till

last: Ringel

first: Nina

last: Frey

first: Femke

last: Ringnalda

first: Sharan

last: Janjuah

first: Sarah

last: Cherkaoui

first: Stefan

last: Butz

first: Sumana

last: Srivatsa

first: Martin

last: Pirkl

first: Giancarlo

last: Russo

first: Lukas

last: Villiger

first: Gerhard

last: Rogler

first: Hans

last: Clevers

first: Niko

last: Beerenwinkel

first: Nicola

last: Zamboni

first: Tuncay

last: Baubec

first: Gerald

last: Schwank


title: Protein arginine methyltransferase 5 promotes cholesterol biosynthesis mediated Th17 responses and autoimmunity

first: Mireia

last: Guerau

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

overall_design: Refer to individual Series


title: Gene expression in crossed cerebellar diaschisis in a transient middle cerebral artery occlusion rat model

city: Okayama

first: Naoya

last: Kidani

state: Okayama

address: 2-5-1 Shikata-cho, Kita-ku

department: Department of Neurological Surgery

institute: Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences

summary: Crossed cerebellar diaschisis (CCD) is hypoperfusion and hypometabolism in the contralateral cerebellar hemisphere caused by supratentorial lesion. We evaluated chronological change of cerebellar blood flow (CbBF) and gene expressions in cerebellum using a rat model of focal ischemic stroke.

overall_design: Gene expression in rat cerebellar cortex was measured at 2 days after transient right middle cerebral artery occlusion(MCAO). Left (contralateral to the infarct) and right (ipsilateral to the infarct) cerebellar cortex of MCAO group and left cerebellar cortex of control group were extracted. To obtain sufficient mass of the experimental material, tissues from four rats in each group (MCAO or control) were combined into one sample.


title: Transcriptome of RABV infected mouse brain

city: Wuhan

first: Zhaochen

last: Luo

state: Hubei

address: No.1 Shi-zi-shan Street

country: China

institute: Huazhong Agricultural University

summary: Here we show the transcriptome of brain from RABV infected B6 mice and TLR7 KO mice. These differential transcripts will provide a reference for studies focus on the relationship between TLR7-dependent signaling and pathogenicity of RABV.

overall_design: B6 mice were infected with RABV and RNA from brain were collected at 6 day p.i. Rabies virus strain B2c is a lab-attenuated rabies virus strain.

first: Ling

last: Zhao


title: A reversible KO model reveals therapeutic potentials of defective Tregs

status: Public on Feb 21 2020

city: Shanghai

first: Zhengyu

last: Jing

address: 393 Middle Huaxia Road, Pudong

department: School of Life Science and Technology

institute: ShanghaiTech University

summary:
To characterize Brg1-reexpressed Tregs, we treated 3-wks-old rKO1 mice with the low dose of TAM and compared gene expression patterns in Brg1-deleted (GFP+) vs. Brg1-reexpressed (GFP- ) Treg subsets isolated 7 days after TAM.
Brg1-deletion in the mosaic females and Brg1-reexpression in rKO1 mice on Day 7 after TAM treatment affected 618 and 1352 genes, respectively, with only 241 genes shared, suggesting divergent roles of Brg1 under the physiological vs. inflammatory conditions. These genes can be divided into two categories:  the “naïve genes” that are predominantly expressed in naïve Tregs  and “activation/effector function genes” preferentially expressed in activated/effector Tregs.
Our data collectively suggest that Brg1 reexpression acted (partly) in conjunction with inflammatory cytokines to convert Brg1-deleted Tregs into hyperactivated Tregs endowed with potent suppressive activity

overall_design: mRNA profiles of 3-4 weeks old wild type (WT) and Nrl-/- mice were generated by deep sequencing, in triplicate, using HiSeq X Ten.  BioProject: PRJNA547476

first: Yongqin

last: Li

first: Yuxing

last: Chen

first: Shaoshuai

last: Mao

first: Ravinder

last: Kaundal

first: Qin

first: Xinxin

last: Wang

first: Jing

last: Xia

first: Jianlong

last: Sun

first: Haopeng

first: Tian

last: Chi


title: LncRNA sequencing of Laoshan dairy goat mammary gland at early, peak and late lactation stages.

status: Public on Feb 22 2020

city: Tai’an

first: Chao

last: Tianle

state: Shandong

address: Daizong street, no. 61, 6/5000 Shandong Agricultural University

institute: 6/5000 Shandong Agricultural University

summary: Five healthy Laoshan dairy goats (four years old, third lactation) from Qingdao Laoshan dairy goat primary farm (Shandong Province, China) were used. The mammary gland samples were collected surgically after general anaesthesia using Xylazine Hydrochloride injection solution (Huamu Animal Health Products Co., Ltd. China) at corresponding lactation stage, including early, peak and late lactations.

overall_design: LncRNA sequencing of Laoshan dairy goat mammary gland at early, peak and late lactation stages, using Illumina HiSeq 2000.

first: Ji

last: Zhibin


title: Massively parallel reporter assay for programmed ribosomal frameshifting on designed sequence libraries

city: Haifa

first: Martin

last: Mikl

address: Abba Khoushy 199

department: Department of Human Biology

country: Israel

institute: University of Haifa

summary: We designed an oligonucleotide libraries containing a potential frameshifting site, cloned the library variants into a dedicated reporter construct, transfected and integrated these constructs in the genome of K562 cells, and performed FACSseq (as well as targeted RNA sequencing) to determine if and to what extent frameshifting occurs.

first: Eran

last: Segal


title: Peyer’s Patch Germinal Centers Select Innate-like BCRs

city: Boston

first: Frederick

middle: W

last: Alt

state: MA

address: 1 Blackfan Circle

department: Program in Cellular and Molecular Medicine (PCMM)

institute: Boston Children's Hospital / Harvard Medical School

summary: Primary B cell receptor (BCR)/antibody variable region exons are generated by V(D)J recombination with junctional diversification creating immensely diverse antigen contact-encoding CDR3s. While most antigen-driven germinal centers (GCs) are transient, gut microbiota-dependent intestinal Peyer’s patch (PP) GCs are chronic. The nature of chronic PP GC BCR repertoires and somatic hyper-mutation (SHM) patterns has remained enigmatic. To elucidate the physiological repertoire of PP GC BCRs, we developed a high throughput antibody repertoire and SHM assay. Remarkably, PP GCs from different mice expanded public clonotypes, each often with identical IgH CDR3. These CDR3s represent innate-like BCRs that appear much more frequently than expected in naïve B cell repertoire by IGoR modeling, but not frequently enough to enter PP GCs at the observed recurrence without cellular selection. Consistently, some public clonotypes are gut microbiota-dependent and encode antibodies reactive to bacteria glycans, while others are not. SPF fecal transfer to germ-free (GF) mice restored two germ-dependent clonotypes, providing direct evidence for BCR selection. In support of this, we identified recurrently selected SHMs in four of the public clonotypes, demonstrating affinity maturation in chronic PP GCs.  Our findings suggest that persistent gut antigens select for innate-like BCR clonotypes to seed chronic PP GCs.

overall_design: We validated our primer design for LAM-HTGTS-Rep-SHM-Seq with mixed JH or JL bait compared with VH or VL bait, with 3 replicates each. We included NP-CGG and SRBCs IP immunized splenic GC and naïve samples as controls for our LAM-HTGTS-Rep-SHM-Seq method to follow BCR selection. We did LAM-HTGTS-Rep-SHM-Seq on PP GC and naïve samples from 18 SPF mice, 10x3 GF mice, 5 GF+SFB mice, 10 GF+SPF mice, 15 AID-deficient SPF mice. We also included single cell RNA sequencing data from 4 of the PP GC samples from SPF mice.

first: Huan

first: Yuxiang

last: Zhang

first: Adam

middle: Y

last: Ye

first: Zhou

last: Du

first: Mo

last: Xu

first: Cheng-Sheng

last: Lee

first: Joyce

last: Hwang

first: Nia

last: Kyritsis

first: Zhaoqing

last: Ba

first: Dan

last: Littman


title: Steroid Hormones Regulate Genome-Wide Epigenetic Programming and Gene Transcription in Human Endometrial Cells with Marked Aberrancies in Endometriosis [methylation]

city: San Francisco

first: Sahar

last: Houshdaran

address: 513 Parnassus Ave HSE1619

department: OB/GYN

institute: UCSF

summary: Genome-wide DNA methylation profiling of hormone treated normal and endometriosis stromal fibroblast cells (eSF). The Illumina Infinium Methylation BeadChip array 450 (HM450)  was used. Samples include 4 normal of no uterine pathology (NUP), 4 endometriosis stage I, 4 endometriosis stage IV samples, each treated with Estrogen (E2), Progesterone (P4), E2+P4, or vehicle (veh), for a total of 48 samples analyzed on HM450.

overall_design: Bisulphite converted DNA from the 48 samples were hybridized to the Illumina Infinium Methylation BeadChip array 450 (HM450).

first: Linda

middle: C

last: Giudice

first: Ashwini

last: Oke

first: Jennifer

last: Fung

first: Kim

last: Vo

first: Camran

last: Nezhat


title: Cortical Neural Stem Cell Lineage Progression Is Regulated by Extrinsic Signaling Molecule Sonic Hedgehog

status: Public on Feb 19 2020

first: Guoping

last: Liu

address: 138 Yi Xue Yuan Road

institute: Fudan University


title: The minimal ProQ protein of Neisseria meningitidis is a global RNA-binding protein that operates in parallel with Hfq

status: Public on Feb 18 2020

city: Würzburg

first: Thorsten

last: Bischler

address: Josef-Schneider-Straße 2 / D15

department: Core Unit SysMed

institute: University of Wuerzburg


title: Transcriptional profiles of skeletal muscle stem cells from mice of different ages, exercise status, and Ccnd1 genotypes

city: Palo Alto

first: Jamie

middle: O.

last: Brett

address: 3801 Miranda Avenue, Building 100, Room E4-216

department: Neurology and Neurological Sciences

institute: Stanford University

summary: Voluntary exercise enhances old skeletal muscle stem cell (MuSC) function in vivo and ex vivo, dependent on upregulation of Ccnd1. To determine the transcriptional changes associated with these phenotypes, RNA-Seq was performed on MuSCs from young and old mice that had exercised or not exercised, and from young mice with MuSC-specific loss-of-function deletions in zero, one, or both alleles of Ccnd1.

overall_design: To study the effects of exercise on MuSCs, 3-month-old (Y) or 18-month-old (O) C57BL6 male mice were given access to a running wheel that was locked (-Ex) or free (+Ex) for three weeks, and hindlimb MuSCs were FACS-isolated as CD31-, CD45-, Sca1-, VCAM+, and viable (DAPI-) cells. To study the effects of Ccnd1 loss in MuSCs, 3-month-old or 18-month-old C57BL6 Pax7_CreERT2; R26R_YFP; Ccnd1_+/+ (WT), Ccnd1_+/flox (HET), or Ccnd1_flox/flox (KO) mice were injected with tamoxifen to induce recombination in MuSCs, which were FACS-isolated six weeks later as YFP+, viable (DAPI-) cells. PolyA-selected RNA libraries were constructed and sequenced on a HiSeq2000, generating paired-end 101-bp reads. Each replicate represents MuSCs from an individual mouse. Y(-Ex) = young, no exercise; O(-Ex) = old, no exercise; Y(+Ex) = young, exercise; O(+Ex) = old, exercise; Y(WT) = young, Pax7_CreERT2; R26R_YFP; Ccnd1_+/+; Y(HET) = young, Pax7_CreERT2; R26R_YFP; Ccnd1_+/flox; Y(KO) = young, Pax7_CreERT2; R26R_YFP; Ccnd1_flox/flox; O(WT) = old, Pax7_CreERT2; R26R_YFP; Ccnd1_+/+.

middle: O

first: Mika

last: Ikeda

first: Thomas

last: Rando


title: The minimal ProQ protein of Neisseria meningitidis is a global RNA-binding protein that operates in parallel with Hfq [CLIP-Seq]

summary: In this study binding sites for the RNA-binding protein ProQ was determined in Neisseria meninigitidis serogroup C strain 8013

overall_design: To detect the binding sites for Neisseria meningitidis (N.meninigitidis) ProQ we collected two biological replicates of N.meninigitidis serogroup C strain 8013 where the C-terminus of the proQ gene was fused to a 3xFLAG tag. Half of both replicate cultures was irradiated with UV light (254 nm, 800 mJ/cm2) (+CL), while the other half was kept untreated (-CL). After lysis of the bacteria, the FLAG-tagged protein was immunoprecipitated using a commercial monoclonal anti-FLAG antibody. The samples were treated with benzonase nuclease, calf intestine phophorylase (CIP), and polynucleotide kinase (PNK) and labeled with radioactive gamma-ATP. After separating the samples by SDS-PAGE, the ProQ-RNA complexes were transfer to nitrocellulose membranes. Radioactively labelled complexes were cut out and eluted from membranes and treated with Proteinase K. Purified RNA was used as input for library preparation using the NEB Next Small RNA Library kit.

first: Saskia

last: Bauriedl

first: Lars

last: Barquist

first: Nadja

last: Heidrich

first: Milan

last: Gerovac

first: Jörg

last: Vogel

first: Christoph

last: Schoen


title: The minimal ProQ protein of Neisseria meningitidis is a global RNA-binding protein that operates in parallel with Hfq [RNA-Seq]

summary: In this study the RNA targetome of the RNA-binding protein ProQ was determined in Neisseria meninigitidis serogroup C strain 8013 by RNA-seq.

overall_design: To uncover the RNA targetome of the RNA-binding protein ProQ of Neisseria meningitidis (N.meninigitidis), bacterial samples were collected of three biological replicates of N.meninigitidis strain 8013 wt and proQ deletion in which the CDS of ProQ was replaced by an antibiotics cassette. Bacterial cultures grown to OD600nm of 2.0 were snapfrozen in liquid nitrogen prior to total RNA-extraction with the hot phenol method and DNase digestion. cDNA libraries of RNA-seq-samples were constructed by Vertis Biotechnology AG, Munich, Germany. To deplete ribosomal transcripts, RNA-seq samples were treated with the Ribo-Zero “Bacteria” kit (Illumina) followed by RNA fragmentation and adapter ligation. cDNA libraries of RNA-seq samples were pooled on an Illumina NextSeq 500 high‐output flow cell and sequenced in single-read mode (1x76 cycles).


title: PIM447 treatment in T-ALL PDX splenocytes

status: Public on Feb 17 2020

city: Gent

first: Renate

last: De Smedt

address: Corneel Heymanslaan 10

department: Department of Biomolecular Medicine

country: Belgium

institute: Ghent University

summary: A subset of T-ALL/T-LBL patients is characterized by overexpression of the oncogenic Serine/Threonine kinase PIM1. Mechanisms for PIM1 overexpression are translocations involving the TCRb and PIM1 loci, JAK/STAT activating mutations and responsiveness to cytokinens in the tumor microenvironment, such as IL7. In this study, we generated a TLX3+ JAKwt/STATwt IL7 responsive T-ALL PDX model using NSG mice. Splenocytes were ex vivo treated with the PIM inhibitor PIM447 and QuantSeq 3’ mRNA sequencing was performed, in order to elucidate the transcriptional effects and molecular mechanisms by which PIM447 induces apoptosis in T-ALL PDX cells.

overall_design: Spleen cells from a TLX3+ T-ALL PDX generated in NSG mice were isolated and red blood cell lysis was performed. Single cells were ex vivo treated with DMSO or 1µM PIM447 for 24 hours in 10% RPMI medium. Cells were washed in PBS after treatment and total RNA was isolated using the miRNeasy Mini Kit (Qiagen).

first: Pieter

last: Van Vlierberghe

first: Wouter

last: Van Loocke


title: Expression data from rat dorsal root ganglion after chronic compression

status: Public on Feb 14 2020

city: Beijing

first: 唐

last: 伟评

address: 2 Anzhen Rd, Chaoyang District,

institute: Beijing Institute of Heart Lung and Blood Vessel Diseases

summary:
Neuropathic pain (NP) is caused by primary or secondary impairment of the peripheral or central nervous systems. Its etiology is complex and involves abnormal patterns of gene expression and pathway activation. Using bioinformatic analysis, we aimed to identify NP-associated changes in genes and pathways in L4 and L5 dorsal root ganglia (DRG) in a rat model of NP induced by chronic compression of the DRG (CCD).
All rats were divided into the sham and CCD groups randomly. Rats in CCD groups were anesthetized, followed by the implantation of two stainless steel rods in the intervertebral foramina between L4 and L5. Rats in the sham-operated group underwent the same procedure without steel rod insertion. DRG were harvested on the 7th day post-surgery.

overall_design: Total RNA was extracted and purified with a RNeasy Mini Kit (Qiagen, Hilden, Germany) according to the manufacturer’s instruction. The quality and amount of RNA were measured using NanoDrop 2000 (Thermo, USA). mRNA expression profiling was performed using Affymetrix Clariom D Rat array (Affymetrix; Thermo Fisher Scientific, Inc), which contains 46,800 gene-level probe sets.

first: Zhanhui

last: Du

first: Yang

last: Zhang


title: Acute Myeloid Leukemia iPSCs reveal a role for RUNX1 in the maintenance of human Leukemia Stem Cells

status: Public on Feb 13 2020

city: New York

first: Franco

last: Izzo

state: NY

address: 1470 Madison Avenue

department: Oncological Sciences

institute: Icahn School of Medicine at Mount Sinai

summary: Leukemia stem cells (LSCs) in acute myeloid leukemia (AML) are believed to possess distinct biological properties than the bulk AML cells, but their rarity and the unavailability of universal immunophenotypic markers for their prospective isolation hampers their study. We report that hematopoietic cells from genetically clonal AML patient-derived induced pluripotent stem cells (iPSCs) contain two morphologically and immunophenotypically distinct subpopulations: a cell fraction with a hematopoietic stem cell (HSC) immunophenotype exhibiting adherent growth which we termed induced leukemai stem cells (iLSCs) and a non-adherent fraction of more differentiated cells, which we termed induced blasts (iBlasts). Through fate-tracking experiments, xenotransplantation and single cell transcriptomics, we show that iLSCs cells reside on the apex of a phenotypic and functional hierarchy and fulfill the hallmark features of leukemia stem cells. Through integrative genomics studies of their transcriptome and chromatin landscape, we derive an LSC 16-gene set that predicts patient survival and identify RUNX1 as a new dependency of AML LSCs.

overall_design: Hematopoietic cells obtained from the AML-4.10 iPSC line were plated in 6 wells of a 6-well tissue culture-treated plate on day 14 of hematopoietic differentiation. After 2 days the iBlasts cells were removed from all 6 wells by washing. The iLSCs cells from 2 wells were collected independently with accutase as the day 16 duplicates. The total cells from two of the remaining wells were independently harvested after 1 and after 2 weeks as day 23 and day 30 duplicates, respectively. All samples were sorted for live (DAPI-) CD45+ cells on a BD FACS Aria II and single cell RNA sequencing was performed using Chromium 10X v2.

first: Josephine

last: Wesely

first: Andriana

middle: G

last: Kotini

first: Hanzhi

last: Luo

first: Han

last: Yuan

first: Maria

last: Georgomanoli

first: Asaf

last: Zviran

first: Neville

last: Dusaj

first: Christina

last: Leslie

first: Dan

last: Landau

first: Michael

last: Kharas

first: Eirini

last: Papapetrou


title: Gene expression profiling of skin adipocyte and adipocyte derived cells in mice 5 days after injury

city: Washington

first: Brett

middle: Adam

last: Shook

state: DC

address: 2300 I St NW

department: Biochemistry & Molecular Medicine

institute: George Washington University

summary: Cells were isolated from day 5wounds from dorsal skin wound beds of 7-9 weeks old using FACS. This experiment describes the gene expression profile associated with mature adipocytes, adipocyte-derived cells and myofibroblasts during tissue repair.

overall_design: Cell were isolated by FACS.  This series includes re-analyzed samples from GSE105790.

first: Valerie

last: Horsley


title: Dynamics of the transcriptional landscape during human gonad development during fetal life

city: Rennes

first: Frédéric

last: Chalmel

state: France

address: 9 avenue du Pr. Léon Bernard

department: Physiology and physiopathology of the urogenital tract

country: France

institute: Inserm U1085-Irset

summary: Fetal life is a crucial period for sexual reproduction when bipotential gonads differentiate into either a testis or an ovary. Gaining insights into the complex molecular events underlying this process is central to a better understanding of disorders of sexual development.  The present work therefore intends to improve the knowledge on molecular pathways at play during gonad development in humans using RNA-Sequencing.

overall_design:
To define the transcriptional landscape of fetal human gonad in greater detail, we extracted and sequenced RNA (library Ribozero, 2x50 pb paired-end, strand specific, on Illumina HiSeq 2500) from testes and ovaries at 6, early 7, late 7, 9, 12, 13-14 and 17 weeks of gestation (n=4 or 2).

The 'Supplementary_table_for_GEO.xlsx' contains all of the information associated with the reconstructed transcripts.

first: Estelle

last: Lecluze

first: Antoine

middle: D

last: Rolland


title: Single cell transcriptome sequencing of CS11 and CS15 human yolk sacs

status: Public on Feb 16 2020

first: Tao

last: Huang

state: Beijing

address: Dong Da Jie No.8

institute: The Fifth Medical Center of Chinese PLA General Hospital

summary: Yolk sac is an important site for early embryonic hematopoiesis. However, our understanding of early hematopoietic development is still very limited. Single cell transcriptome sequencing provides us with a good research method. Here, we performed single cell RNA-seq analysis for  Carnegie stage 11 (CS11) and  Carnegie stage 15 (CS15) human yolk sacs.

overall_design: We performed single cell RNA-seq analysis for  CS11 and  CS15 human yolk sacs.

first: Zhilei

last: Bian

first: Yangdong

last: Gong

first: Christopher

middle: Z

last: Lee

first: Hui

last: Shi


title: Deciphering human macrophage development at single-cell resolution

summary: Macrophages are the earliest emerging cells of the nascent immune system during embryonic development, and as innate immune cells constitutes an important first-line barrier against foreign organisms and pathogens. Rodent macrophages have been shown to infiltrate multiple organs at an early stage, developing symbiotically alongside these organs becoming tissue-resident macrophages (TRM) supporting tissue development and homeostasis. However, knowledge of the development and specialization of macrophages in the early human embryo is still limited. In order to study the spatiotemporal distribution and dynamic process of macrophage development in the early human embryo, we applied single-cell transcriptomic sequencing to generate a map of all CD45+CD235a- hematopoietic cells in human embryos of successive developmental stages from Carnegie stage (CS) 11 to 23. Here, we unravelled for the first time a map of macrophage heterogeneity across multiple anatomical sites and identified multiple macrophage subsets in the early human embryo, including various types of embryonic TRM (head, liver, lung and skin), and traced their developmental trajectories from yolk sac/embryonic liver-derived macrophages to their TRM specification in the head and liver based on core transcriptional factors. Altogether, our analyses provide a comprehensive characterization of the spatial and temporal dynamics of macrophage development in the early human embryo, and provides a reference for future study into human TRM function and embryonic development.

overall_design: We sorted CD45+CD235a- hematopoietic cells using flow cytometry from human embryos at multiple Carnegie stages (CS11, CS12, CS13, CS15, CS17, CS20 and CS23) and sites (yolk sac, head, liver, blood, lung and skin), and then performed a modified single cell tagged reverse transcription (STRT) protocol.

first: Yandong


title: MicroRNA sequencing in ARCaPE and ARCaPM cells

status: Public on Feb 12 2020

city: Oxford

first: Srinivasa

last: Rao

address: Botnar Research Centre, Institute of Musculoskeletal Sciences, Windmill Road, Headington

department: Nuffield Dept. of Surgical Sciences

institute: University of Oxford

summary: ARCaPE and ARCaPM human prostate cancer cell lines are derivatives of the ARCaP cell line, with epithelial and mesenchymal characteristics respectively. Hence, they are a model of epithelial-mesenchymal transition in prostate cancer. The objective of this study was to identify differentially expressed microRNAs in these cell lines, which may be responsible for the EMT phenotype. Unsupervised hierarchical clustering confirmed that microRNA expression profiles are distinctly different between the two cell lines, with 119 miRNAs significantly increased and 81 microRNAs significantly decreased in ARCaPM cells as compared to ARCaPE. Further analysis revealed that many of the differentially expressed microRNAs belong to only a few microRNA families/clusters. Unsurprisingly, microRNAs in these clusters are co-expressed due to common promoters, which are known to be regulated by Wnt signalling and BMP signalling(33) pathways. The microRNAs that were most strikingly overexpressed in ARCaPE cells belong to the miR-372 and miR-302 clusters, which have been previously shown to be key regulators of EMT in embryonic stem cells and induced pluripotent stem cells.

overall_design:
3 biological replicates each of ARCaPE and ARCaPM compared.
Adaptor sequences are as follows:
3' adaptor: AGATCGGAAGAGCACACGTCT
5' adaptor: GTTCAGAGTTCTACAGTCCGACGATC

first: Claire

middle: M

last: Edwards


title: A PKA/PP2A AXIS DRIVES THE GROWTH OF NEUROENDOCRINE SCLC

status: Public on Feb 11 2020

city: Vienna

first: Julia

last: Arand

address: Schwarzspanierstr. 16

country: Austria

institute: Medical University of Vienna

summary: Small cell lung cancer (SCLC) is an aggressive subtype of lung cancer whose biology is still poorly understood. Using a multiplexed inhibitor beads assay, we identified active kinases in SCLC. Among those, we found that PKA is critical for the expansion of SCLC in culture and in vivo. PKA promotes the neuroendocrine epithelial state associated with SCLC tumor-initiating cells. Phosphoproteomics analyses identify ~200 PKA substrates and show that PKA controls multiple facets of SCLC growth. Notably, the PP2A phosphatase counteracts the oncogenic effects of PKA, and PP2A activators inhibit SCLC as single agents and with chemotherapy. Our data uncover key signaling networks in SCLC and indicate that targeting the PKA/PP2A pathway may help inhibit this lethal neuroendocrine cancer.

overall_design: H29 and 82 cells were infected with a lentivirus to express either a dox inducible GFP (control) of active form of PRKACA (aPKA). Puromycin resistant cells were injected into NSG mice and animals were given doxycycline (2mg/ml) in the drinking water.

first: Garry

last: Coles

first: Julien

last: Sage


title: Genome-Wide analysis of gene expression responses to waterlogging stress in roots of barley (Hordeum vulgare L.)

city: Brandon

first: Ana

last: Badea

state: Manitoba

address: 2701 Grand Valley Rd

institute: Agriculture and Agri-Food Canada

summary: Waterlogging is a major abiotic stress causing oxygen depletion and carbon dioxide accumulation in the rhizosphere. Barley is more susceptible to waterlogging stress than other cereals. To gain a better understanding of the effect of waterlogging stress in barley, we carried out a genome-wide gene expression analysis in roots of Yerong and Deder2 barley genotypes under waterlogging and control (well-watered) conditions by RNA-Sequencing, using Illumina HiSeq™ 4000 platform.

overall_design: Two-week-old Yerong and Deder2 seedlings were exposed to waterlogging treatment. Root tissue samples were collected at the time-points 0 and 72 h of treatment for Yerong from both treated and control plants, and at the points 0, 72, and 120 h of treatment for Deder2 from both treated and control plants. Total RNA was prepared and used for the RNA-seq analysis. Four independent biological replicates were used for each plant sample.

last: Borrego-Benjumea

first: Adam

last: Carter

first: James

last: Tucker

first: Zehn

last: Yao

first: Wayne

last: Xu


title: Adapting the 10x Genomics Platform for single-cell RNA-seq in yeast reveals the importance of stochastic gene expression during the lag phase

status: Public on Feb 06 2020

city: Leuven

first: Abbas

last: Jariani

address: Gaston Geenslaan 1, box 2471

department: KU Leuven - VIB Centre for Microbiology

institute: KU Leuven - VIB

summary: Isogenic population of yeast cells was grown in a maltose-glucose-maltose media shift schema. Time point samples were taken throughout the experiment, and were analyzed for single-cell gene expression using 10x Genomics platform.

overall_design: Yeast cells experience a halt in growth after glucose-maltose shift, aka lag phase. Two samples are taken during glucose growth. Two samples are taken during lag phase in maltose. One sample comprises a mix of cells actively grown on glucose and cells actively growing in maltose. This last sample serves as a control to see if we can detect the expected population structure.

first: Lieselotte

last: Vermeersch


title: Single cell RNA-seq of mouse dorsal raphe Pet1 neurons

status: Public on Feb 09 2020

first: Benjamin

last: Okaty

address: 77 Avenue Louis Pasteur

institute: Harvard Medical School

summary: Here we use high-throughput and DR subdomain-targeted single-cell transcriptomics and intersectional genetic tools to map molecular and anatomical diversity of DR-Pet1 neurons. We describe up to fourteen neuron subtypes, many showing biased cell body distributions across the DR.

overall_design: Single cell RNA-seq libraries prepared from genetically labeled Pet1 neurons in two different ways: (1) automated fluorescence-based cell sorting (using the On-Chip Sort) followed by 10X Genomics Chromium Single Cell 3’ v3 protocol (samples 71-72), or (2) DR subdomain-targeted manual cell sorting, followed by SMART-Seq v4 Ultra Low Input cDNA amplification and Nextera XT library preparation (samples 1-70).

first: Nikita

last: Sturrock


title: KDM3A/Ets1/MCAM axis promotes growth and metastatic properties in Rhabdomyosarcoma.

city: Aurora

first: Andrew

last: Goodspeed

state: CO

address: 12801 E 17th Ave, Mail Stop 8303

institute: University of Colorado Anschutz Medical Campus

summary: The goal of this study was to understand the function and mechanisms of action of the chromatin factor KDM3A in the pediatric soft tissue cancer Rhabdomyosarcoma

overall_design: RNA-seq was used to define the effects of KDM3A depletion on gene expression in the patient-derived Rhabdomyosarcoma cell lines RD and Rh30.

first: Paul

last: Jedlicka

first: Lays

last: Sobral

first: Marybeth

last: Secher

first: Janet

middle: K

last: Parrish

first: Tyler

last: McCann

first: Kenneth

last: Jones

first: Joshua

last: Black


title: Elucidating miRNA associated with cisplatin resistance in head and neck cancer cells

status: Public on Feb 01 2020

city: Chuo-ku

first: Yusuke

last: Yamamoto

address: 5-1-1, Tsukiji, Chuo-ku

institute: National cancer center research institute

summary: To investigate the function of the three microRNAs involved in drug resistance in head and neck cancer, we transfected them into oral squamous cancer cell line, SAS. The phenotypes were analyzed and transcriptome of them were investigated.

overall_design: Three microRNAs (miR-210-3p, miR-629-3p and miR-874-3p) were transfected (25nM) into SAS cells that are head and neck cancer cells. And then, we extracted RNA and did microarray.

first: Junichiro

last: Chikuda

first: Takahiro

last: Ochiya


title: The intestinal microbiota programs DNA methylation to control tissue homeostasis and inflammation [ATAC-Seq]

city: Heidelberg

first: Guenter

last: Raddatz

address: Im Neuenheimer Feld 580

institute: German Center for Cancer Research

summary: Although much research has been done on the diversity of gut microbiome, little is known about the way it influences intestinal homeostasis under normal and pathogenic conditions. Epigenetic mechanisms have recently been suggested as operating at the interface between the microbiota and the intestinal epithelium cells (IECs). Using genome-wide analyses, we discovered that exposure to microbiota induced both global DNA hypomethylation and localized changes at regulatory elements, which culminates in activation of a set of “early sentinel” response genes that play a role in maintaining gut homeostasis. Furthermore, we demonstrated that exposure to microbiota in acute inflammation results in profound DNA methylation and chromatin accessibility changes at regulatory elements leading to alterations in the gene expression program in colitis and colon cancer. Our studies add a new dimension to our understanding of the cross talk between the microbiota and IECs, and provide the foundation for how microbiota impact epigenetic programming.

overall_design: Whole genome methylation analysis of M. musculus. Four conditions were sequenced and analyzed, the first is an untreated control, the second corresponds to inflammation, the third mice with a germ-free condition and the fourth to a germ-free condition in combination with inflammation

first: F

last: Lyko

first: Y

last: Bergman


title: RNAseq of renal CD64+CD11b+F4/80hi  from adult mice after cisplatin-induced acute kidney injury

status: Public on Jan 31 2020

city: Planegg

first: Barbara

last: Schraml

address: Großhaderner Strasse 9

institute: LMU Munich

summary: We use bulk RNA sequencing of sorted cells to characterize the gene expression profiles of renal MHCII+/-D11b+F4/80hi cells 72 hours after cisplatin-induced acute kidney injury (AKI). F4/80hi cells are activated after cisplatin induced AKI and start to downregulate MHCII on transcriptional level. Downregulation of MHCII is probably caused by a downregulation/inhibition of its most important transcriptional activator Ciita

overall_design: bulk RNA sequencing of sorted murine renal F4/80hi cells before and after cisplatin induced acute kidney injury

first: S

last: Rambichler

first: N

last: Salei

first: B

middle: U


title: Dynamic induction of drug resistance through a stress-responsive enhancer in acute myeloid leukemia [CHIP-seq]

status: Public on Feb 03 2020

city: Manchester

first: Tim

last: Somervaille

address: Wilmslow Rd

department: Manchester Institute

institute: Cancer Research UK

summary: The drug efflux pump ABCB1 is a key driver of chemoresistance, and high expression predicts for treatment failure in acute myeloid leukemia (AML). We show that both acute and chronic exposure of leukemia cells to daunorubicin activates an integrated stress response-like transcriptional program to induce ABCB1 through remodeling and dynamic activation of an ATF4-bound, stress-responsive enhancer. In primary human AML, stress-responsive ABCB1 enhancers are accessible and acetylated, and exposure of fresh blast cells to daunorubicin induces ABCB1 in a dose-dependent manner. Dynamic induction of ABCB1 by diverse stressors, including chemotherapy, facilitates escape of leukemia cells from targeted third-generation ABCB1 inhibition. Stress-induced up regulation of ABCB1 is mitigated by combined use of pharmacologic inhibitors U0126 and ISRIB, which inhibit stress signaling.

overall_design: TS75 - K562 cells were purchased from DSMZ (Braunschweig, Germany) and cultured in RPMI 1640 medium (Sigma Aldrich) supplemented with 2mM L-Glutamine (Life Technologies) and 10% fetal bovine serum (Sigma Aldrich). Three separate vials of early passage K562 cells were thawed and cultured separately for two weeks. (Figure 1A). Sensitive cell lines were designated K562_S1-3, aliquots were frozen and stored before commencing selection with 10nM daunorubicin (Sigma Aldrich). Whilst under selection, cells were counted and replated twice per week. The daunorubicin concentration was adjusted depending on whether cell numbers were increasing (dose increased by 25-50%), decreasing (dose reduced by 25-50%) or static (dose unchanged). Selection continued until cell lines were capable of expansion in medium containing 500nM daunorubicin. Resistant cell lines were designated K562_R1-3. The time taken to acquire this level of resistance was 106 days (K562_R1 and R3) and 117 days (K562_R2). K562_R1-3 were culture without daunorubicin for 10 days prior to RNA extraction, assessment of daunorubicin IC50 and freezing of aliquots. All cell lines (K562_S1-3, K562_R1-3) were tested for mycoplasma and authenticated by short tandem repeat DNA profiling. H3K27ac ChIP sequencing was performed as detailed below. Data from TS75 was used to identify putative ABCB1 enhancers by comparing resistant (K562_R1-3) with sensitive (K562_S1-3) cells. TS93 - To confirm that putative enhancers were functional, we next performed targeted silencing using a CRISPR-dCas9-KRAB system. We designed multiple sgRNAs for each region and screened them in K562_R3 cells. The most active guides were then selected for use in all resistant cell lines. K562_R1-3 cells were dual infected with pHR-SFFV-dCas9-BFP-KRAB and pLKO5.sgRNA.EFS.tRFP657, the latter expressing an sgRNA targeting an enhancer (E1-4) or the promoter, or a non-targeting control (EV). We used ChIPseq for H3K9 trimethylation to confirm that silencing was discrete and accurate, this was performed as described below. TS98 - To examine enhancer usage in primary AML we performed H3K27ac ChIP sequencing on cryopreserved bulk AML samples, each with their own unique identifier (BBxxx).


title: Dynamic induction of drug resistance through a stress-responsive enhancer in acute myeloid leukemia [RNA-seq]

overall_design: TS60 - K562 cells were purchased from DSMZ (Braunschweig, Germany) and cultured in RPMI 1640 medium (Sigma Aldrich) supplemented with 2mM L-Glutamine (Life Technologies) and 10% fetal bovine serum (Sigma Aldrich). Three separate vials of early passage K562 cells were thawed and cultured separately for two weeks. (Figure 1A). Sensitive cell lines were designated K562_S1-3, aliquots were frozen and stored before commencing selection with 10nM daunorubicin (Sigma Aldrich). Whilst under selection, cells were counted and replated twice per week. The daunorubicin concentration was adjusted depending on whether cell numbers were increasing (dose increased by 25-50%), decreasing (dose reduced by 25-50%) or static (dose unchanged). Selection continued until cell lines were capable of expansion in medium containing 500nM daunorubicin. Resistant cell lines were designated K562_R1-3. The time taken to acquire this level of resistance was 106 days (K562_R1 and R3) and 117 days (K562_R2). K562_R1-3 were culture without daunorubicin for 10 days prior to RNA extraction, assessment of daunorubicin IC50 and freezing of aliquots. All cell lines (K562_S1-3, K562_R1-3) were tested for mycoplasma and authenticated by short tandem repeat DNA profiling. Total RNA was extracted from 5x105 K562_S1-3 and K562_R1-3 using QIAshredder spin columns and an RNeasy® Plus Micro kit (Qiagen, Manchester, UK). RNA sequencing was performed using two replicates for K562_R1-3 and a single replicate for K562_S1-3. Prior to sequencing RNA integrity was checked using a 2100 Bioanalyzer (Agilent Technologies – www.genomics.agilent.com). PolyA libraries were prepared using a SureSelect PolyA kit (Agilent Technologies), samples were then barcoded and pooled. Sequencing was performed using a NextSeq desktop sequencing system (Illumina – www.illumina.com). A single run (400M reads) of 75bp paired-end sequencing produced a mean of 45.7M reads per sample.


title: Dynamic induction of drug resistance through a stress-responsive enhancer in acute myeloid leukemia

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

overall_design: Refer to individual Series


title: RNAseq of renal dendritic cells from adult Clec9a-Cre Rosa-YFP mice

summary: We use bulk RNA sequencing of sorted cells to characterize the gene expression profiles of renal dendritic cell (DC) subsets, cDC1 and cDC2, as well as MHCII+CD64+ F4/80hi and MHCII+CD64+ CD11bhi cells. Splenic DCs and red pulp macrophages serve as reference populations for a macrophage-like or DC-like phenotype. By sorting YFP+/YFP- F4/80hi or YFP+/YFP- CD11bhi cells from Clec9a-Cre Rosa-YFP mice we aim to reveal transcriptional differences between YFP-labelled and unlabelled cells. We showed that F4/80hi cells resemble macrophages on a transcriptional level, despite their DC origin, and that renal CD11bhi cells are a transcriptionally unique subset. However, we were not able to reveal differences between YFP+ and YFP- populations.

overall_design: RNAsequencing of murine dendritic cell and macrophage populations from kidney and spleen


title: Functional genomic consequences of MBD5 knockdown in mouse brain and CRISPR-derived neurons

status: Public on Jan 28 2020

city: Munich

first: Tatsiana

last: Aneichyk

address: Josef-Frankl-Strasse 5A

institute: Independent Data Lab

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus


title: Clonal maintenance of transcriptional and epigenetic states in cancer cells.

status: Public on Jan 29 2020

city: Rehovot

first: Zohar

last: Meir

state: Shfela

address: Herzl 234

department: Mathematics & CS

institute: Weizmann Institute of Science

summary: We performed single-cell transcriptome analysis (using MARS-seq and 10x) of four human cell lines, as well as bulk tracscriptome analysis of hundreds of single-cell clones, derived from the same cells population. Additionally, we performed methylation analysis of HCT116 single cells (using whole-genome PBAT) and of the same clones whose transcriptome was analysed (using capture-PBAT). We also analyse longitudinal single cell transcriptomes of six selected HCT116 clones grown for up to 148 days in our lab, and analysed cells and  clones of HCT116 cells harbouring knock-out mutations in DNMT1 and DNMT3B (DKO).

overall_design: cells from original population were FACS-sorted into single-cell methylation or scRNA profiling. In parallel, single cells were sorted into conditional media for propagation. When single-cell derived clones reached a size of ~500 cells in average (after 9-10 days), they were split into two aliquotes: one was used for bulk methylation analysis, and the other for bulk RNA analysis. For long-term assay of HCT116 cells, low-depth bulk analysis of clones was performed on two time-points (10 and 18 days), and selected clones were chosen fur single-cell RNA sampling in four additional time-points (33, 62, 98 and 148 days), and clonal exome was sequenced at two time-points (d=78, 168).

first: Amos

last: Tanay


title: Functional genomic consequences of MBD5 knockdown in mouse brain and CRISPR-derived neurons [cell lines]

summary:
Background: MBD5, encoding the methyl-CpG-binding domain 5 protein, has been proposed as a necessary and sufficient driver of the 2q23.1 microdeletion syndrome. De novo missense and protein-truncating variants from exome sequencing studies have directly implicated MBD5 in the etiology of autism spectrum disorder (ASD) and related neurodevelopmental disorders (NDDs). However, little is known concerning the specific function(s) of MBD5.
Methods: In an effort to gain insight into the complex interactions associated with genetic alteration of MBD5 in individuals with ASD and related NDDs, we explored the transcriptional landscape of MBD5 haploinsufficiency across multiple mouse brain regions of the heterozygous hypomorphic Mbd5+/GT mouse model and compared these results to CRISPR-mediated mutations of MBD5 in human iPSC-derived neuronal models.
Results: Gene expression analyses of three brain regions from Mbd5+/GT mice showed cortex as the region most affected by the knockdown, indicating context-dependent effects. Gene co-expression network analyses revealed gene clusters that are associated with MBD5 knockdown and enriched for GPCR signaling and terms related to ciliary function. We also identified genes that were downregulated in both models such as EPHA3, OLIG1, related to neuronal development and glial function.
Limitations: These analyses were performed in a limited number of brain regions and neuronal models, and the effects of the gene knockdown are subtle. As such, these results will not reflect the full extent of MBD5 disruption across brain tissues during early brain developmentneurodevelopment in ASD.
Conclusions: Our study points to transcriptional consequences of Mbd5 disruption in the brain and suggests a link between MBD5 and pathways such as ciliary function that are associated with established developmental disorders and syndromes.

overall_design: Human neuronal models of MBD5 disruption from CRISPR/Cas9 gene-edited iPSCs (8330-8) reprogrammed previously from fibroblasts of a healthy male individual using standard retroviral vectors. Three MBD5-edited cell lines with alterations in the conserved MBD domain and three WT cell lines were used in this study

first: Catarina

last: Seabra

first: Serkan

last: Erdin

first: Derek

middle: J

last: Tai

first: Celine

middle: E

last: De Esch

first: Parisa

last: Razaz

first: Yu

last: An

first: Poornima

last: Manavalan

first: Ashok

last: Ragavendran

first: Alexei

last: Stortchevoi

first: Clemer

last: Abad

first: Juan

middle: I

last: Young

first: Patricia

last: Maciel

first: Michael

last: Talkowski

first: James

middle: F

last: Gusella


title: Functional genomic consequences of MBD5 knockdown in mouse brain and CRISPR-derived neurons [mouse]

first: Tatsiana

last: Aneichyk

overall_design: 10 mice with Mbd5 heterozygous deletion, 8 mice with WT genotype from same litters, for each mice three tissues are sequenced: cerebellum(Cb), cortex (Cx) and striatum(St).


title: MLD Project RNAseq

status: Public on Jan 27 2020

city: Amsterdam

first: Evert

last: Bosdriesz

address: De Boelelaan 1111

department: Computer Science

country: Netherlands

institute: VU University Amsterdam

summary: PC9 cells (untreated or treated with 4D combo) were harvested (in trizol) 48h or 72h after 4D combo treatment in replicates.

overall_design: The goal is to look at expression differences between the treated with 4D combo and untreated samples

first: Joao

last: Neto


title: Context-dependent gene regulation by homeodomain transcription factor complexes revealed by shape-readout deficient proteins

city: New York City

first: Harmen

last: Bussemaker

state: New York

address: 1212 Amsterdam Avenue, MC 2441

department: Biological Sciences

institute: Columbia University

summary: Eukaryotic transcription factors (TFs) form complexes with various partner proteins to recognize their genomic target sites. Yet, how the DNA sequence determines which TF complex forms at any given site is poorly understood. Here, we demonstrate that high-throughput in vitro DNA binding assays coupled with unbiased computational analysis provide unprecedented insight into how different DNA sequences select distinct compositions and configurations of homeodomain TF complexes. Using inferred knowledge about minor groove width readout, we design targeted protein mutations that destabilize homeodomain binding both in vitro and in vivo in a complex-specific manner. By performing parallel systematic evolution of ligands by exponential enrichment sequencing (SELEX-seq), chromatin immunoprecipitation sequencing (ChIP-seq), RNA sequencing (RNA-seq), and Hi-C assays, we not only classify the majority of in vivo binding events in terms of complex composition but also infer complex-specific functions by perturbing the gene regulatory network controlled by a single complex.

overall_design: SELEX-seq was performed on different combinations of the three homeodomain transcription factors Deformed, Homothorax and Extradenticle for both wild-type and shape-readout-impaired mutant protein. In addition, ATAC-seq and ChIP-seq was performed in D.melanogaster 3rd instar imaginal wing discs against both wild-type and shape-readout mutant Extradenticle, as well as Homothorax and Antennapedia TFs. Intrinsic DNA Shape (Zhou, PNAS, 2015) was correlated with TF binding affinity to identify position-specific shape readout (minor groove readout). Amino acid mutations abolishing the identified shape readout were subsequently used in vitro and in vivo to probe the degree to which binding mechanism are recapitulated in vivo. Further Hi-C and RNA-seq data was used to dissect the regulatory network of HD TF complexes and identify complex-specific gene function.

first: Richard

middle: S

last: Mann

first: Judith

last: Kribelbauer


title: DNA methylation markers panel can improve prediction of response to neoadjuvant chemotherapy in luminal B breast cancer

status: Public on Jan 25 2020

city: Moscow

first: Alexey

middle: Igorevich

last: Kalinkin

state: Russia

address: Moskvorechie st., 1

department: Epigenetics

country: Russia

institute: RCMG

summary: Despite the advantages of neoadjuvant chemotherapy (NACT), associated toxicity is a serious complication that renders monitoring of the patients response to NACT highly important. Thus, prediction of tumor response to treatment is imperative to avoid exposure of potential non-responders to deleterious complications. We have performed genome-wide analysis of DNA methylation by XmaI-RRBS and selected CpG dinucleotides differential methylation of which discriminates luminal B breast cancer samples with different sensitivity to NACT. With this data, we have developed multiplex methylation sensitive restriction enzyme PCR (MSRE-PCR) protocol for determining the methylation status of 10 genes (SLC9A3, C1QL2, DPYS, IRF4, ADCY8, KCNQ2, TERT, SYNDIG1, SKOR2 and GRIK1) that distinguish BC samples with different NACT response. Analysis of these 10 markers by MSRE-PCR in biopsy samples allowed us to reveal three top informative combinations of markers, (1) IRF4 and C1QL2; (2) IRF4, C1QL2, and ADCY8; (3) IRF4, C1QL2, and DPYS, with the areas under ROC curves (AUCs) of 0.75, 0.78 and 0.74, respectively. A classifier based on IRF4 and C1QL2 better meets the diagnostic panel simplicity requirements, as it consists of only two markers. Diagnostic accuracy of the panel of these two markers is 0.75, with the sensitivity of 75% and specificity of 75%.

overall_design: Reduced representation bisulfite sequencing by XmaI-RRBS (Tanas et al., 2017)

first: Alexander

last: Tanas


title: Integrin αvβ5 internalizes Zika virus during neural stem cells infection and is a target for antiviral therapy

status: Public on Jan 22 2020

city: La jolla

first: Qiong

last: Zhang

state: California

address: 9500 Gilman Dr.

department: Pediatrics

institute: UCSD

summary: Here, we performed the first CRISPR-Cas9 genome-wide screen in glioblastoma stem cells (GSCs) to identify factors determining ZIKV neurotropism. Using a ZIKV-ZsGreen reporter virus, a new strategy was developed to sort infection-resistant cells, expand, and enrich the resistant pool. Importantly, bioinformatics analysis and functional validation revealed integrin αvβ5 as an internalization factor for ZIKV. Expression of αvβ5 is correlated with ZIKV susceptibility in various cell lines and matched the cell tropism in developing human cerebral cortex. Blocking antibody against integrin αvβ5 but not αvβ3 efficiently inhibited ZIKV infection. Mechanistic evaluation showed that ZIKV binds to cells but fails to internalize when treated with integrin αvβ5 blocking antibody. αvβ5 directly binds to ZIKV virions, activates FAK (focal adhesion kinase), which is required for ZIKV infection. Strikingly, pharmaceutical inhibitors of αvβ5, SB273005 and cilengitide suppressed ZIKV infection at nanomolar concentrations. Finally, αvβ5 blocking antibody or inhibitors reduced ZIKV infection and alleviated ZIKV induced pathology in human neural stem cells (hNSC) and in mouse brain. This is the first study, to the best of our knowledge, identifying integrin αvβ5 is an internalization factor for ZIKV providing a new therapeutic target as well as two promising drug candidates that potentially can be used as prophylactics or treatments for ZIKV infections.

overall_design: Examination of sgRNA populations in ZIKV-ZsGreen negative GSCs

first: Shaobo

last: Wang


title: Isolation and analysis of rereplicated DNA by Rerep-seq

first: Joshua

last: Black

address: 12801 East 17th Ave

department: Pharmacology

institute: University of Colorado

summary: Changes in gene copy number contribute to genomic instability, the onset and progression of cancer, developmental abnormalities, and adaptive potential. The origins of gene amplifications have remained elusive; however, one model suggests DNA rereplication could be a source of gene amplifications.  The inability to determine which sequences are rereplicated and under what conditions have made it difficult to determine the validity of these models. Here we present RerepSeq, a technique that selectively fragments and enriches rereplicated DNA in preparation for next generation sequencing. RerepSeq is applicable to any species and can be utilized with low amounts of input DNA with rapid results.  We validated RerepSeq by simulating DNA rereplication in yeast and human cells.  Using RerepSeq, we demonstrate that rereplication induced in S. Cerevisiae by deregulated origin licensing is non-random and defined by broad domains that span multiple replication origins and topological boundaries.

sample_organism:
Saccharomyces cerevisiae
Homo sapiens

platform_organism:
Saccharomyces cerevisiae
Homo sapiens

overall_design: Rerep-seq leverages the semiconservative nature of DNA replication to selectively fragment and enrich rereplicated DNA. Cells are labeled with the thymidine analog BrdU for one cell cycle to allow BrdU incorporation into the newly replicated DNA strand, rereplicated regions incorporate BrdU in both strands. Genomic DNA from cells labeled with BrdU is purified and subjected to biochemical processing to induce ssDNA breaks at the sites of BrdU incorporation. This is performed by subjecting DNA to UVA treatment in the presence of Hoechst 33258 to photolyze the bromine from BrdU leaving deoxyuracil. Deoxyuracil is then removed by treatment with UDG (uracil DNA glycosylase) leaving an abasic site. The abasic site is then converted to a single strand DNA break by deoxyribose excision by APE1 (apurinic/apyrimidinic endodeoxyribonuclease). This results in nicked genomic DNA for normally replicated DNA, but staggered nicks producing dsDNA breaks in rereplicated DNA. The fragmented rereplicated DNA can then be isolated by size fractionation and analyzed by quantitative PCR or next-generation sequencing.

first: Johannes

last: Menzel

first: Philip

last: Tatman


title: Characterization of a new small-molecule inhibitor of HDAC6 in glioblastoma

status: Public on Jan 18 2020

city: Donsotia-San Sebastian

first: charles

last: lawrie

address: Paseo Mikeletegi, 83- planta 3

department: Oncology

country: Spain

institute: Biodonostia Research Institute

summary: Histone deacetylase 6 (HDAC6) is an epigenetic modifier that is an attractive pharmacological target in cancer. In this work, we show that HDAC6 is elevated in glioblastoma, the most malignant and common brain tumor in adults, and in glioma stem cells. Moreover, we identified a new small-molecule inhibitor of HDAC6, called JOC1, which presents strong sensitivity for HDAC6 inhibition and exerts high cytotoxic activity, alone or in combination with temozolomide. Moreover, it is able to significantly reduce tumor growth in vivo. Transcriptomic analysis of patient derived glioma stem cells revealed an increase in cell differentiation and cell death as well as decrease in cell cycle activity and cell division as relevant cellular pathways whose activities are significantly altered by the treatment with JOC1. In conclusion, our data reveal the efficacy of a novel HDAC6 inhibitor in glioblastoma pre-clinical setting

overall_design: Glioblastoma cell line (GNS179)+/- small inhibitor JOC1 and pan-inihibitor SAHA

first: Charles

last: Lawrie


title: Aryl hydrocarbon receptor mediates larval zebrafish fin duplication following exposure to benzofluoranthenes.

status: Public on Jan 21 2020

city: Corvallis

first: Robyn

last: Tanguay

state: OR

address: 28645 East Highway 34

department: Environmental and Molecular Toxicology

institute: Oregon State University

summary: Polycyclic aromatic hydrocarbons (PAHs) can alter gene expression by acting through the aryl hydrocarbon receptor (AHR). In a previous phenotypic screen of over 120 PAHs, we identified four PAHs that induce an ectopic caudal fin (called X-fin) in larval zebrafish: benzo[k]fluoranthene (BkF), dibenzo[b,k]fluoranthene, dibenzo[a,h]anthracene, and benzo[j]fluoranthene. We investigated several plausible mechanisms of X-fin formation using the most potent X-fin inducer, BkF. The X-fin phenotype was dependent on the AHR paralog Ahr2, and we performed RNA sequencing to identify altered gene expression patterns. Transcriptional profiles of distal trunk tissue, where the phenotype was manifest, were generated for animals exposed to 1% DMSO (control) or 12 μM BkF. Four time points of X-fin development were considered for transcriptomics: prior to visual emergence of X-fin (48 hpf), during manifestation of disrupted caudal fin fold development (60 and 72 hpf), and after emergence of X-fin (96 hpf).

overall_design: RNA profiles of larval zebrafish trunk tissue at 48, 60, 72, and 96 hpf, from animals exposed to either 1% DMSO or 12 μM benzo[k]fluoranthene, were generated using Illumina HiSeq 2500 with 100 bp single end reads. There were n = 50 per bioreplicate and four bioreplcates per condition, totaling 32 raw samples. Each processed sample consists of mean normalized counts from four raw data files, totaling eight processed samples.

first: Michael

middle: A

last: Garland

first: Mitra

last: Geier

first: Sean

last: Bugel

first: Prarthana

last: Shankar

first: Cheryl

middle: L

last: Dunham

first: Joseph

last: Brown

first: Susan

last: Tilton


title: Muscle cannabinoid 1 receptor regulates Il-6 and myostatin expression, governing physical performance and whole-body metabolism.

city: Baltimore

first: Supriyo

last: De

state: Maryland

address: 251 Bayview Blvd

department: Laboratory of Genetics and Genomics

institute: NIA-IRP, NIH

summary: Cannabinoid 1 receptor (CB1R) plays a critical role in the regulation of whole-body energy metabolism because of its involvement in controlling food intake thorough a central mechanism, and controlling distribution of fuel and utilization in the periphery, particularly in fat and liver. Additionally, inhibition of CB1R improves insulin secretion from pancreatic beta cells and insulin sensitivity in both pancreatic beta cells and hepatocytes. We now developed a skeletal muscle-specific CB1R-knockout (Skm-CB1R-/-) mouse to study the specific role of CB1R in muscle. Muscle-CB1R ablation prevented diet-induced and age-induced insulin resistance by increasing insulin receptor signaling. Ablation of CB1R in muscle also led to IL-6 secretion and reduced myostatin expression, and enhanced AKT and MAPKs signaling. These changes in myokines induced the expression of myogenic genes, increasing muscle mass and whole-body lean/fat ratio in both obesity and aging, with no change in total body weight. Skm-CB1R-/- mice had increased energy expenditure and their muscle had improved mitochondrial performance, which led to improved physical endurance. These results collectively show that muscle-CB1R is involved in regulating whole-body insulin sensitivity and physical performance.

overall_design: Wild-type (WT) and cannabinoid receptor-1 knock out (CB1KO) male mice were fed a standard chow diet (SD) or high fat-high sugar diet (HFHS) for 15 weeks. Each group is represented by four biological replicates.

first: Isabel

last: González-Mariscal

first: Rodrigo

last: Montoro

first: Marta

last: Gonzalez-Freire

first: Jennifer

last: O'Connell

first: Qing-Rong

last: Liu

first: Irene

last: Alfaras

first: Olga

last: Carlson

first: Elin

last: Lehrmann

first: Yongqing

first: Kevin

last: Becker

first: Josephine

last: Egan


title: Contradictory of gene expression signatures for clinical prognostic prediction caused by update of reference genome and annotation

status: Public on Jan 13 2020

city: harbin

first: Lishuang

last: Qi

address: No. 157, Baojian Road, Nangang District

department: College of Bioinformatics Science and Technology

institute: Harbin Medical University

summary: Gene expression estimates detected by RNA-sequencing technology vary with the updates of reference genome and gene annotation, which might confound existing expression-based prognostic signatures, making them inapplicable to clinical practice. In this study, we proposed a method to decrease these effects and developed a qualitative signature for stage I lung adenocarcinoma, whose classification was based on within-sample relative expression orderings (REOs) of gene pairs. The signature was validated in 471 stage I samples derived from public RNA-sequencing and microarray data (both log-rank p < 0.001). Notably, our signature could effectively predict prognosis for 30 stage I patients with severely degraded FFPE tissues (log-rank p = 0.0177). More important, the risk classification was stable in the latest annotation. In summary, our signature would be a promising signature for clinical individualized application because of its excellent prognostic performance and classification robustness.

overall_design: 30 FFPE tissues with severely RNA degradation from stage I lung adenocarcinoma patients who had surgery with curative intent were sequenced by RNA-Seq.


title: Digital gene expression of directed differenciation protocols

city: Boston

first: Carlos

last: Villacorta-Martin

address: 670 Albany St

institute: Center for Regenerative Medicine at Boston University

summary: Lung and hindgut directed differentiations with multiple protocols and time points

overall_design: Comprehensive characterization at several time points of iPSC directed differenciation protocols

last: Villacorta Martin

first: Aditya

last: Mithal

first: Gustavo

last: Mostoslavsky


title: Calibrated Scc1 ChIP-seq for wild type and chl4Δ strains arrested in metaphase with and without tension

city: Edinburgh

first: Daniel

last: Robertson

state: Midlothian

address: 2.28 Michael Swann Building, Kings Buildings, Mayfield Road

department: Discovery Research Platform for Hidden Cell Biology

institute: University of Edinburgh

summary: Genome binding/occupancy profiling of the cohesin subunit Scc1 by high throughput sequencing. S. pombe is included as the calibration genome.

sample_organism:
Schizosaccharomyces pombe
Saccharomyces cerevisiae

overall_design: Purpose: To determine the requirement for Chl4 in association of cohesin with the centromere and pericentromere borders.

first: Flora

last: Paldi

first: Adele

last: Marston


title: Calibrated Sgo1 and Brn1 ChIP-seq for wild type strains arrested in metaphase without tension

summary: Genome binding/occupancy profiling of Sgo1 and the condensin subunit Brn1 by high throughput sequencing. S. pombe is included as the calibration genome.

platform_organism:
Schizosaccharomyces pombe
Saccharomyces cerevisiae

overall_design: Purpose: to determine the association of Sgo1 and Brn1 (condensin) with pericentromere borders.


title: Calibrated Scc1, Sgo1 and Brn1 ChIP-seq in metaphase with and without tension, in strains where convergent genes at pericentromere borderes are rearranged into a tandem orientation

overall_design: Purpose: Does the localisation of Cohesin, Shugoshin and Condensin change upon the rearrangement of convergent genes into tandem at pericentromere borders?


title: Conformation of the budding yeast pericentromere in metaphase

summary: High-resolution Hi-C in metaphase in wild type cells with and without tension, chl4Δ with tension, and sgo1-3A and sgo1Δ without tension

overall_design: Purpose: To determine how pericentromeric chromosome conformation changes in response to spindle tension and to the absence of pericentromeric cohesin and condensin.


title: Calibrated Scc1 ChIP-seq for wild type and rad61Δ strains arrested in metaphase with and without tension

overall_design: Purpose: To determine the requirement for Rad61 in the tension-dependent removal of cohesin from the pericentromere.


title: Esculetin inhibits cancer cell glycolysis by binding tumor PGK2, GPD2, and GPI

status: Public on Jan 09 2020

city: Wuhan

first: Songtao

last: Wu

address: Faculty of Pharmacy, Hubei University of Chinese Medicine, Wuhan, China

institute: Hubei University of Chinese Medicine

summary:
The target proteins phosphoglycerate kinase 2 (PGK2), glycerol-3-phosphate dehydrogenase (GPD2)GPD2 and glucose-6-phosphate isomerase (GPI) were screened by combining transcriptome, proteomics and reverse docking
We detected the binding constant of the active compound using microscale thermophoresis (MST). It was found that esculetin bound well with three potential target proteins.

overall_design: The HepG2 cells were inoculated into 6-well plates at a concentration of 5 x 105/mL. There are two groups: HepG2 cells without esculetin,  and HepG2 cells containing esculetin

first: Song-Tao

first: Yan-Fang

last: Yang


title: Gene expression analysis of Influenza vaccine response in Young and Old - Year 3 - B and T cells

status: Public on Jan 08 2020

city: New Haven

first: Hailong

last: Meng

state: CT

address: 300 George Str

institute: Yale University

summary: We profiled gene expression of B and T cells from a stratified cohort of subjects to define influenza vaccine response in Young and Old

overall_design: Differential gene expression by human B and T cells from Healthy Adults receiving Influenza Vaccination (Fluvirin, Novartis). Healthy adults (older >65, younger 21-30 years) were recruited at seasonal Influenza Vaccination clinics organized by Yale University Health Services during 2012 - 2013 seasons. With informed consent, healthy individuals were recruited as per a protocol approved by Human Investigations Committee of the Yale University School of Medicine. Each subject was evaluated by a screening questionnaire determining self-reported demographic information, height, weight, medications and comorbid conditions. Participants with acute illness two weeks prior to vaccination were excluded from study. Blood samples were collected into BD Vacutainer Sodium Heparin tube at four different time points, once prior to administration of vaccine and three time points after vaccination on days 4, 7 and 28. Peripheral Blood Mononuclear Cells (PBMC) were isolated from heparinized blood using Histopaque 1077 in gradient centrifugation. About 1.0x10^7 freshly isolated PBMC were lysed in Triso and immediately stored in -800C. Total RNA in aqueous phase of Trisol - Chloroform was isolated in an automated QiaCube instrument using miRNeasy according to manufacturer's instructions. Integrity of RNA samples were assessed by Agilent 2100 BioAnalyser Samples were processed for cRNA generation using Illumina TotalPrep cRNA Amplification Kit and subsequently hybridized to Human HT12-V4.0 BeadChip at Yale Center for Genomic Analysis (YGCA).

first: Albert

middle: C

last: Shaw

first: Subhasis

last: Mohanty

first: Samit

middle: R

last: Joshi

first: Ikuyo

last: Ueda

first: Jean

last: Wilson

first: Juilee

last: Thakar

first: Steven

last: Kleinstein

first: Sui

last: Tsang


title: RNA Sequencing Facilitates Quantitative Analysis of Schwann cells Transcriptomes

status: Public on Jan 01 2020

first: cheng

last: wang

address: No. 1277 Jiefang Avenue

department: Union Hospital of Tongji Medical College

institute: Huazhong University of Science and Technology

summary:
Purpose: Whole-transcriptome sequencing technology and bioinformatics analysis were applied to systematically analyze the differentially expressed mRNAs, lncRNAs and miRNAs in SCs from DPN rats and control rats.
Methods: mRNA, lncRNA and miRNA profiles of Schwann cells from DPN rats and control rats were generated by RNA sequencing, using Illumina Xten. The sequence reads that passed quality filters were analyzed at the transcript isoform level by using sequence analysis programs, including HISAT, Stringtie and DESeq2. qRT–PCR validation was performed using Takara, Vazyme and Clontech kits.
Results: The data showed that 2925 mRNAs, 164 lncRNAs and 49 miRNAs were significantly differently expressed in SCs from DPN rats compared with control rats, with a fold change ≥1.2 or ≤ 0.833 and p value <0.05 . The results of qRT-PCR confirmed 13 mRNAs, 7 lncRNAs and 7 miRNAs which were consistent with the RNA-seq data. Functional and pathway analyses revealed that many enriched biological processes of GO terms and pathways were relevant to the function of SCs and the pathogenesis of DPN.
Our study is the first to detect and analyze mRNAs, lncRNAs and miRNAs changes in Schwann cells from STZ-treated rats with DPN and reveal the novel interactions between dysregulated RNAs and the pathogenesis of DPN. Our data show that dysregulated RNAs have complicated interactions between them and play critical roles in regulating functions of SCs involved in the pathogenesis of DPN

first: Cheng

last: Wang

first: Xu

last: Xiang


title: Expression analysis of genes in human vaginal VK2 E6/E7 cells after treatment with estrogen and challenge with flagellin

city: Newcastle Upon Tyne

first: Judith

last: Hall

state: Tyne & Wear

address: Medical School, Framlington Place, Newcastle University

department: ICaMB

institute: Newcastle University

summary: The female menopause, characterised by reduced estrogen associates with an increased risk of recurrent UTIs caused by uropathogenic Escherichia coli (UPEC). Clinically such infections can be countered by topical vaginal estrogen treatment and the aim of this study was to investigate, in vitro, the effects of topical estrogen treatment on vaginal epithelial responses following challenge with E.coli flagellin used to mimic UPEC. Immortalised vaginal epithelial cells (VK2 E6/E7), modelling the vaginal epithelium were treated with either 4nM 17β-estradiol (E) for seven days, 50ng/ml E.coli flagellin (F) for 12h, or 4nM 17β-estradiol plus 50ng/ml flagellin (E + F(12h)). RNA was analysed by microarray gene profiling using the Illumina HumanHT-12 v 4 Expression Beadchip. Following E + F treatments expression of genes encoding host defence molecules including DEFβ4A, DEFB103A, LCN2 as well as those associated with keratinisation e.g. CNFN and SPRR family genes were significantly enhanced (P<0.05) compared to either E or F treatments alone. Mutation of EREs identified in the DEFβ4 gene promoter abolished the augmented gene expression suggesting estrogen functioned directly through a transcriptional regulatory mechanism involving ESR1/2. Ingenuity pathway analyses also suggested the pro-inflammatory cytokine IL-17A to regulate the vaginal host defences during infection. Pre-treating VK2 E6/E7 cells with estrogen (4nM) and challenging with 1L-17A & F (12h) significantly enhanced DEFβ4, DEF103A and S100A7 expression (P<0.05). Origins of vaginal IL-17 in vivo remain unclear, but vaginal biopsy material suggests gd T cells located within the vaginal epithelium. These data suggest that the vaginal antimicrobial response induced by flagellin activation of TLR5 cell signalling is augmented significantly by topical estrogen treatment.

overall_design: Microarray analysis was performed to identify other potential genes regulated by estrogen and to help determine signalling pathways. VK2 cells that were pretreated with 4nM estrogen for seven days followed by 50ng/ml flagellin challenge were analysed by microarray. The 12 and 24 hour samples of control, estrogen pretreatment, flagellin, and estrogen pretreatment plus flagellin challenged VK2 cells were selected for analysis. Three technical replicates of each treatment were analysed, totalling 24 samples. Samples to be analysed were shipped on dry ice to Service XS, The Netherlands, who performed RNA quality analysis and carried out the microarray experiments.

first: Anna

last: Stanton

first: Catherine

last: Mowbray

first: Marcelo

last: Lanz

first: Karen

last: Brown

first: Paul

last: Hilton

first: Alison

last: Tyson-Capper

first: Robert

last: Pickard

first: Ased

last: Ali


title: Gene expression analysis of Influenza vaccine response in Young and Old - Year 1 - B and T cells

first: Hailong

last: Meng

summary: We profiled gene expression from a stratified cohort of subjects to define influenza vaccine response in Young and Old.

overall_design: Differential gene expression by human B cells, CD4 and CD8 T cells from Healthy Adults receiving Influenza Vaccination (Fluvirin, Novartis). Healthy adults (older >65, younger 21-30 years) were recruited at seasonal Influenza Vaccination clinics organized by Yale University Health Services during October to December of 2010 – 2011 seasons. With informed consent, healthy individuals were recruited as per a protocol approved by Human Investigations Committee of the Yale University School of Medicine. Each subject was evaluated by a screening questionnaire determining self-reported demographic information, height, weight, medications and comorbid conditions. Participants with acute illness two weeks prior to vaccination were excluded from study. Blood samples were collected into BD Vacutainer Sodium Heparin tube at four different time points, once prior to administration of vaccine and three time points after vaccination on days 2/4, 7 and 28. Peripheral Blood Mononuclear Cells (PBMC) were isolated from heparinized blood using Histopaque 1077 in gradient centrifugation. About 1.0x10^7 freshly isolated PBMC were lysed in Triso and immediately stored in -80C. Total RNA in aqueous phase of Trisol - Chloroform was isolated in an automated QiaCube instrument using miRNeasy according to manufacturer’s instructions. Integrity of RNA samples were assessed by Agilent 2100 BioAnalyser Samples were processed for cRNA generation using Illumina TotalPrep cRNA Amplification Kit and subsequently hybridized to Human HT12-V4.0 BeadChip at Yale Center for Genomic Analysis (YGCA).

first: Albert

last: Shaw

first: Subhasis

last: Mohanty

first: Samit

last: Joshi

first: Ikuyo

last: Ueda


title: ChIP_seq data for Rrp6 project analysis in malaria (Plasmodium falciparum)

city: Shanghai

first: Shijun

last: Shen

state: Shanghai

address: 1239 Siping Road, Shanghai, P.R. China

department: Bioninformatics

institute: Tongji University

summary: We carried out chromatin immunoprecipitation followed by deep sequencing (ChIP-seq) to explore the difference of the genome-wide dynamics of histone modifications and HP1 in PfRrp6-DD-1C line.

overall_design: we carried out ChIP-seq assay to detect the signals of H3K9Ac, H3K9me3 and HP1 in each chromosome of PfRrp6-DD-1C clone  with WT 3D7-G7 as control

first: Yanting

last: Fan

first: Guiying

last: Wei

first: Cizhong

last: Jiang

first: Qingfeng

last: Zhang


title: HIC_seq data for Rrp6 project analysis in malaria (Plasmodium falciparum)

summary: We performed Hi-C assay to address the interaction of central var genes in ruf6-var pairs，subtelomeric upsA-subtype var genes and other HP1-associated  genes

overall_design: We performed Hi-C assay with WT 3D7 parasite


title: Gene expression analysis of Influenza vaccine response in Young and Old - Year 2 - B and T cells

overall_design: Differential gene expression by human B cells, CD4 and CD8 T cells from Healthy Adults receiving Influenza Vaccination (Fluvirin, Novartis). Healthy adults (older >65, younger 21-30 years) were recruited at seasonal Influenza Vaccination clinics organized by Yale University Health Services during 2011-2012 seasons. With informed consent, healthy individuals were recruited as per a protocol approved by Human Investigations Committee of the Yale University School of Medicine. Each subject was evaluated by a screening questionnaire determining self-reported demographic information, height, weight, medications and comorbid conditions. Participants with acute illness two weeks prior to vaccination were excluded from study. Blood samples were collected into BD Vacutainer Sodium Heparin tube at four different time points, once prior to administration of vaccine and three time points after vaccination on days 2/4, 7 and 28. Peripheral Blood Mononuclear Cells (PBMC) were isolated from heparinized blood using Histopaque 1077 in gradient centrifugation. About 1.0x10^7 freshly isolated PBMC were lysed in Triso and immediately stored in -80C. Total RNA in aqueous phase of Trisol - Chloroform was isolated in an automated QiaCube instrument using miRNeasy according to manufacturer’s instructions. Integrity of RNA samples were assessed by Agilent 2100 BioAnalyser Samples were processed for cRNA generation using Illumina TotalPrep cRNA Amplification Kit and subsequently hybridized to Human HT12-V4.0 BeadChip at Yale Center for Genomic Analysis (YGCA).

first: Jean

last: Wilson

first: Juilee

last: Thakar

first: Steven

last: Kleinstein

first: Sui

last: Tsang