title: mRNA destabilization by BTG1 and BTG2 maintains T cell quiescence [mTAIL-seq]

first: Jaechul

last: Lim

summary: Quiescence is a cellular state in which cells undergo reversible cell cycle arrest in response to environmental challenges or lack of stimuli. When favourable conditions are engaged, the cells exit quiescence and start to proliferate. For example, quiescent stem cells and oocytes are activated by differentiation signals and fertilization, respectively. The regulation of quiescent state is also crucial for lymphocytes, such as T cells. Inappropriate activation of T cells often leads to autoimmune diseases and thus, the balance between quiescent and activated T cells must be preserved. Despite its importance, however, how T cells maintain the ‘quiescent state’ and exit from it remain largely unknown. Here, we identify BTG1 and BTG2 (BTG1/2) as factors responsible for T cell quiescence. Knockout of Btg1/2 in T cells resulted in a reduced threshold to T cell activation, increased proliferation and a concomitant reduction of the frequency of naive T cells. Loss of BTG1/2 rendered naive T cells to become spontaneously activated even without TCR stimuli. Interestingly, we found a global increase in mRNA abundance in naive T cells that lack BTG1/2. In addition, depletion of BTG1/2 led to an increase in mRNA half-life and poly(A) tail length. Thus, BTG1/2 promotes deadenylation and degradation of mRNA in naive T cells. Our study demonstrates a key mechanism underlying T cell quiescence, and suggests low mRNA abundance as a key feature for maintaining quiescent T cells.

overall_design: Three replicates of naive CD4 T cells (TCRb+CD4+CD25-CD62LhiCD44low) from WT and DKO mice, respectively.

first: Soo-Seok

last: Hwang

first: Richard

last: Flavell


title: Dissecting the regulatory strategies of NFkB RelA target genes in the inflammatory response reveals differential transactivation logics [caRNAseq]

status: Public on Jan 30 2020

city: Los Angeles

first: Kim

last: Ngo

state: CA

address: 611 Charles E Young Dr. East

department: MIMG

country: USA

institute: University of California, Los Angeles

type: Expression profiling by high throughput sequencing

summary: NFkB RelA is the potent transcriptional activator of inflammatory response genes. We stringently defined a list of direct RelA target genes by integrating physical (ChIPseq) and functional (RNAseq in knockouts) datasets. We then dissected each gene's regulatory strategy by testing RelA variants in a novel primary-cell genetic complementation assay.  All endogenous target genes required that RelA makes DNA-base-specific contacts, and none could be activated by the DNA binding domain alone. However, endogenous target genes differed widely in how they employ the two transactivation domains. Through model-aided analysis of the dynamic timecourse data we reveal gene-specific synergy and redundancy of TA1 and TA2. Given that post-translational modifications control TA1 activity and intrinsic affinity for coactivators determines TA2 activity, the differential TA logics suggests context-dependent vs. context-independent control of endogenous RelA-target genes. While some inflammatory initiators appear to require co-stimulatory TA1 activation, inflammatory resolvers are a part of the NFkB RelA core response.

overall_design: chromatin RNA extracted from primary mouse embryonic fibroblasts were subjected to stimulation with TNF.

first: K

middle: A

first: A

last: Hoffmann


title: Dissecting the regulatory strategies of NFkB RelA target genes in the inflammatory response reveals differential transactivation logics [MEFs_2]

overall_design: Total RNA extracted from primary mouse embryonic fibroblasts stimulated with LPS, Poly(I:C), and TNF.


title: Dissecting the regulatory strategies of NFkB RelA target genes in the inflammatory response reveals differential transactivation logics (ChIP-seq.WT.MEFs)

type: Genome binding/occupancy profiling by high throughput sequencing

overall_design: We performed ChIP-seq with antibody specific for NF-kappaB RelA subunit in primary wild-type MEFs treated with TNF.

first: Alexander


title: High-resolution mapping of transcribing RNAPII by CRAC in the presence of different Sen1 variants

status: Public on Jan 31 2020

city: Paris

first: Odil

last: Porrua Fuerte

address: 15 rue Helene Brion

country: France

institute: Institut Jacques Monod-CNRS

summary: Pervasive transcription is a widespread phenomenon leading to the production of a plethora of non-coding RNAs (ncRNAs) without apparent function. Pervasive transcription poses a risk that needs to be controlled to prevent the perturbation of gene expression. In yeast, the highly conserved helicase Sen1 restricts pervasive transcription by inducing termination of non-coding transcription. However, the mechanisms underlying the specific function of Sen1 at ncRNAs are poorly understood. Here we identify a motif in an intrinsically disordered region of Sen1 that mimics the phosphorylated carboxy terminal domain (CTD) of RNA polymerase II  and characterize structurally its recognition by the CTD-interacting domain of Nrd1, an RNA-binding protein that binds specific sequences in ncRNAs. In addition, we show that Sen1-dependent termination strictly requires the recognition of the Ser5-phosphorylated form of the CTD by the N-terminal domain of Sen1. Furthermore, we find that the N-terminal and the C-terminal domains of Sen1 can mediate intra-molecular interactions. Our results shed light onto the network of protein-protein interactions that control termination of non-coding transcription by Sen1.

overall_design: Analysis of the effect of several Sen1 mutations on RNAPII nascent transcription genome-wide using CRAC

last: Porrua


title: Dissecting the regulatory strategies of NFkB RelA target genes in the inflammatory response reveals differential transactivation logics (RNA-seq.WT.p53ko.MEFs_4)

overall_design: Total RNA extracted from two biological replicates of p53-deficient and 1 WT control replicate from primary mouse embryonic fibroblasts were subjected to stimulation with IL-1beta, TNF, and LPS in 8-hour time-course analyis (0, 0.5H, 1H, 3H, 8H). In order to prolong the lifespan of primary MEFs, cells deficient in p53 could be propagated sufficiently longto allow retroviral transduction, selection, and expansion for transcriptional time-course studies. Examining gene expression profiles of inflammatory response genes in p53 MEFs was a control experiment for studying inflammatory response genes. Two biological replicates of p53-deficient MEFs and their WT littermate control MEFs were used for pairwaise comparisons to show reproducibility in inducible gene expression between p53-deficient and WT MEF cells in response to three stimuli known to activate NF-kappaB signaling pathway namely, IL-1beta, TNF, and LPS.


title: Dissecting the regulatory strategies of NFkB RelA target genes in the inflammatory response reveals differential transactivation logics [MEFs_3]

overall_design: Total RNA extracted from retroviral complementation of RelA-deficient primary mouse embryonic fibroblasts with RelA (WT/mutants) stimulated with TNF.


title: Transcriptomic and Functional Studies Reveal Undermined Chemotactic and Angiostimulatory Properties of Aged Microglia During Stroke Recovery

city: Pittsburgh

first: Xiaoming

last: Hu

state: PA

address: 200 lothrop street

department: Neurology

institute: University of Pittsburgh

summary: Age-dependent alterations in microglia behavior have been implicated in neurodegeneration and CNS injuries. Here, we compared the transcriptional profiles of young versus aged microglia during stroke recovery. CD45intermediateCD11b+ microglia were FACS-isolated from the brains of young (10-week-old) and aged (18-month-old) male mice 14 days after distal middle cerebral artery occlusion (dMCAO) or sham operation and subjected to RNA-sequencing analysis. Functional groups enriched in young microglia are indicative of upregulation in cell movement, cell interactions, inflammatory responses and angiogenesis, while aged microglia exhibited a reduction or no change in these features. We confirmed reduced chemoattractive capacities of aged microglia toward ischemic brain tissue in organotypic slide co-cultures, and delayed accumulation of aged microglia around dead neurons injected into the striatum in vivo. In addition, aging is associated with an overall failure to increase the expression of microglial genes involved in cell-cell interactions, such as CXCL10. Finally, impaired upregulation of pro-angiogenic genes in aged microglia was associated with a decline in neovascularization in aged mice compared to young mice after dMCAO. This study provides a new resource to understand the mechanisms underlying microglial alterations in the aged brain milieu and sheds light on new strategies to improve microglial functions in aged stroke victims.

first: Jun

last: Chen


title: Dissecting the regulatory strategies of NFkB RelA target genes in the inflammatory response reveals differential transactivation logics [BMDM]

overall_design: Total RNA extracted from primary mouse bone marrow-derived macrophages (BMDMs) were subjected to stimulation with TNF and LPS.


title: Transcription Factor p73 Regulates Th1 Differentiation

status: Public on Jan 29 2020

city: Bethesda

first: Peng

last: Li

state: MD

address: 9000 Rockville Pike

department: NHLBI

institute: NIH

summary: Inter-individual differences in T helper (Th) cell responses affect susceptibility to infectious, allergic, and autoimmune diseases. To identify factors contributing to these response differences, we analyzed the phenotypic profiles of in vitro-differentiated Th1 cells from 16 inbred mouse strains. Haplotype-based computational genetic analysis indicated that the p53 family protein, p73, affected Th1 differentiation. In cells differentiated under Th1 conditions in vitro, we demonstrated that p73 was a negative regulator of IFNγ production, and this effect required the DNA-binding activity of p73. p73 bound within the gene body or upstream of many genes that affect Th1 differentiation and regulated their expression (e.g., Ifng and Il12rb2). Furthermore, in experimental autoimmune encephalitis, which is a mouse disease model of multiple sclerosis, p73-deficient mice had increased IFNg production and decreased disease severity, whereas in the adoptive transfer model of inflammatory bowel disease, transfer of p73- deficient naïve CD4+ T cells into WT Rag2-/- hosts resulted in increased Th1 responses and enhanced disease severity compared to transfer of WT cells. These studies identify p73 as a regulator of immune responses and indicate that its dysregulation may play a role in susceptibility to autoimmune disease.

overall_design: ChIP-Seq and RNA-Seq using naïve CD4+ T cells that were polarized in Th1 conditions for 3 days

first: Min

last: Ren

first: Majid

last: Kazemian

first: Ming

last: Zheng

first: Jianping

last: He

first: Jangsuk

last: Oh

first: Wei

last: Liao

first: Jessica

first: Jonathan

last: Rajaseelan

first: Brian

middle: L

last: Kelsall

first: Gary

last: Peltz

first: Warren

middle: J

last: Leonard


title: Genome-wide profiling of EDM3 dependent H3K9me2 in Arabidopsis thaliana

city: Riverside

first: Xueqing

middle: Maggie

last: Lu

address: 3401 Watkins Drive, Genomics, Room 2126

department: Cell Biology

institute: University of California, Riverside

summary: We perform genome-wide profiling of H3K9me2 in the Arabidopsis thaliana edm3 mutant. By var-seq, we identified EDM3 as a nuclear-localized protein featuring a single RNA-recognition motif (RRM). Similar to PHD finger-containing histone binding protein EDM2, EDM3 promotes high levels of H3K9me2 at RPP7 and controls transcripts of this NLR gene by suppressing proximal polyadenylation and promoting the synthesis of full-length RPP7-coding mRNAs. Our results showed that EDM3 affects levels of this epigenetic mark at a set of genes and transposons, the vast majority of which also feature EDM2-dependent H3K9me2.

overall_design: H3K9me2 ChIP-seq profiling in edm3 and wild-type Col5 with two replicates respectively, var-seq for edm3 and wild-type Col5

first: Yan

last: Lai

middle: M

first: Neerja

last: Katiyar

first: Karine

last: Le Roche

first: Thomas

last: Eulgem


title: Promoter nucleosome positioning in dendritic cells (DCs) and fibroblasts measured by ChIP-MNase-seq

status: Public on Feb 04 2020

city: Nice

first: Dominic

last: van Essen

address: 28 Avenue Valombrose

institute: Institute for Research on Cancer & Aging, Nice (IRCAN)

summary: We used ChIP-MNase to determine nucleosome positions around promoters in DCs and fibroblasts, after experimental treatment to stimulate inducible gene activation

overall_design: Mouse dendritic cells and 3T3 fibroblasts were treated for 1hr  with LPS or with TNF-alpha, respectively.  Cells were fixed for 10minutes with 4% formaldehyde and used to prepare chromatin.  Chrromatin was fragmented to approximately 5-10kb fragments and used for ChIP.  ChIP-enriched chromatin was then digested using micrococcal nuclease (MNase) to generate approximately 80% mononucleosomal fragments.  DNA was extracted after reversion of fixation and used for sequencing.

first: Simona

last: Saccani


title: MicroRNAs expressed from FSHR and aromatase genes target important ovarian functions.

city: Tallinn

first: Ilmatar

last: Rooda

address: Akadeemia tee 15

department: Chemistry and Biotechnology

country: Estonia

institute: Tallinn University of Technology

summary:
MicroRNAs (miRNAs) have known roles in the post-transcriptional regulation of various biological processes including ovarian follicle development. We have previously identified from human pre-ovulatory granulosa cells miRNAs that are expressed from the intronic regions of two key genes in normal follicular development: FSH receptor (FSHR) and CYP19A1, the latter encoding the aromatase enzyme. In the present study, we aim to identify the targets regulated by those two miRNAs: hsa-miR-548ba and hsa-miR-7973, respectively. The miRNAs of interest were endogenously expressed in KGN cell-line, gene expression changes were analyzed by Affymetrix microarray and confirmed by RT-qPCR. Potential miRNA-regulated sequences were further filtered from the obtained results by bioinformatic target prediction algorithms and validated for direct miRNA:mRNA binding by luciferase reporter assay.
Our results verified Leukemia inhibitory factor receptor (LIFR), Phosphatase and tensin homolog (PTEN), Neogenin 1 (NEO1) and SP110 nuclear body protein (SP110) as target genes for hsa-miR-548ba. Hsa-miR-7973 target genes ADAM metallopeptidase domain 19 (ADAM19), Peroxidasin (PXDN) and Formin like 3 (FMNL3) also passed all verification steps. In conclusion we propose that hsa-miR-548ba may be involved in the regulation of follicle growth and activation via LIFR and PTEN. Hsa-miR-7973 may be implicated in the modulation of extracellular matrix and cell-cell interactions. Taken together, our results suggest that those two miRNAs of interest have important regulatory roles in granulosa cells  and in follicle development in general.

overall_design: For Affymetrix microarray miRNA mimics hsa-miR-548ba, hsa-miR-7973 and control miRNA cel-miR-39-3p were transfected with previously optimized conditions: 12.5nM concentration and 72 hours. Each miRNA was transfected in number of four parallel samples.

first: Birgitta

last: Kaselt

first: Andres

last: Salumets

first: Agne

last: Velthut-Meikas


title: Transcriptomics analysis of lineage-restricted Dendritic-cell (DC) progenitor cells in obesity

status: Public on Jan 28 2020

first: Mehdi

last: Pirooznia

address: 12 SOUTH DR

institute: National Institutes of Health

summary: The overall goal of the project is to understand transcript-level changes in DC-progenitors from bone marrow of diet-induced obese mice by Next Generation Sequencing (NGS; RNASeq)

overall_design:
DC-restricted progenitors (Lin-/c-kit(int)/Sca1(lo)/CD115+/Flt3+; ~50,000 cells) from bone marrow (BM) compartment of high fat (60kcal % fat) & control chow (10 kcal%fat)-fed mic (n=4 per group) were flow-sorted and RNA was isolated.
The whole transcriptome was amplified and library was constructed using NuGen Ovation Solo RNASeq System; and evaluated on high-sensitivity DNAchip (Agilent) and sequenced on a HiSeq 3000 platform

first: Amarjit

last: Mishra

first: Stewart

last: Levine


title: RNA-seq of CRC tissues

status: Public on Jan 27 2020

city: Shanghai

first: Shenglin

last: Huang

address: 270 Dongan

country: China

institute: Fudan University

summary: CRC tissues plus matched normal tissues (Primary colon cancer, liver metastasis colon cancer and matched normal colon tissues from same patient) were subjected to RNA isolation and RNA-seq analyses were generated by deep sequencing using Illumina Hiseq.

overall_design: RNA-seq analyses of CRC tissues plus matched normal tissues by deep sequencing using Illumina Hiseq. Primary colon cancer, liver metastasis colon cancer and matched normal colon tissues from same patient (3 patients, CRC1, CRC2 and CRC3) were subjected to RNA isolation and RNA-seq analyses. RNA libraries were prepared for sequencing using ribosomal RNA depleted RNA-seq protocols. Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to hg38 whole genome using Hisat2. Gene annotated in GencodeV25 were quantified using featureCounts.

first: Guoxiang

last: Cai


title: Dissecting the regulatory strategies of NFkB RelA target genes in the inflammatory response reveals differential transactivation logics [MEF]

first: Kim

last: Ngo

overall_design: Total RNA extracted from primary mouse embryonic fibroblasts stimulated with IL-1b, LPS, and TNF.

first: K

first: A

last: Hoffmann


title: Stagewise keratinocyte differentiation from human embryonic stem cells by defined signal transduction modulators

status: Public on Jan 25 2020

city: Macau

first: Ren

last: Zhili

address: Avenida da Universidade

institute: University of Macau

summary: we use human embryonic stem cells to demonstrate that epidermal ectoderm and subsequent keratinocyte cell fate can be determined step by step under the regulation of defined factors. We show the global gene expression profiles clustered according to the differentiation stages . We also found the human keratinocyte specific gene expression according to the differentiation stages.

overall_design: Total RNA obtained from  Day 0 to Day 8, cells were cultured in differentiation medium with the following treatments: Day 0-6, 10 µM SB431542; Day 1-6, 5 µM CHIR99021; Day 1-8, 10 ng/ml BMP4 (R&D 314-BP-01M); Day 4-8, 5 µM DAPT (Tocris 2634). From Day 9 to Day 11, the medium was changed to low-calcium differentiation medium (containing 0.06 mM calcium) supplemented with BMP4/DAPT/EGF (10 ng/ml). Day 11 to Day 26, the cells in low calcium keratinocyte medium.

first: Guokai

last: Chen


title: RNA sequencing of BT474 cells treated with trastuzumab or trastuzumab + pertuzumab and BT474-derived cells resistant to trastuzumab or trastuzumab + pertuzumab

status: Public on Jan 26 2020

city: New Haven

first: Qin

last: Yan

state: CT

address: Department of Pathology, PO Box 208023

institute: Yale School of Medicine

summary: We sequenced untreated BT474 cells, BT474 cells treated for three days with trastuzumab or trastuzumab + pertuzumab, as well as two BT474-derived trastuzumab-resistant pools and two BT474-derived trastuzumab + pertuzumab resistant pools. Resistant pools were generated by culturing BT474 cells in gradually increasing doses of trastuzumab and trastuzumab + pertuzumab over the course of several months and continually maintained in drug.

overall_design: Examination of mRNA profiles of 1) BT474 cells 2) BT474 cells treated for three days with trastuzumab 3)  BT474 cells treated for three days with trastuzumab + pertuzumab 4) two trastuzumab-resistant cell pools 5) two trastuzumab+pertuzumab resistant cell pools

first: Molly

last: Gale

first: Zongzhi

middle: Z

last: Liu


title: RNA-seq analysis identifies regulators of macrophage reprogramming from M2 to M1-like phenotype

city: Jena

first: Rainer

last: König

address: Am Klinikum 1

country: Germany

institute: University Hospital Jena

summary:
We analyzed expression profiles of thioglyolate elicted peritoneal exudate cells (peritoneal macrophages). Peritoneal macrophages were polarized into M1 and M2 macrophages by activation with IFNgamma+ LPS and Il4, respectively. We predicted a gene-regulatory network, which consists of four transcription factors (E2f1,Myc, Stat6, Pparg) regulating metabolic genes, M1 and M2-associated genes. The predicted regulators were all active in M2 macrophages.
We hypothesized that these transcription factors are essential regulators to maintain M2 phenotype. To further validate our findings, we treated M2-polarized macrophages with siRNA-pools targeting E2f1, Myc, Stat6 and Pparg.
In this study, we observed a switch towards an M1-like phenotype after transfection of siRNA-pools. In addition, Inflammatory pathways were upreguated while fatty acid metabolism was down regulated.

overall_design: RNA was extracted from murine peritoneal macrophages, untreated or treated with LPS+IFNgamma (M1 macrophage) and Il-4 (M2 macrophage). Furthermore, M2-polarized peritoneal macrophages were transfected with siRNA-pools targeting E2f1, Myc, Stat6, Pparg, negative siRNA-pool and mock. Untreated and M1-polarized peritoneal macrophages were treated with mock. The experiments were performed in tripicates.

first: Franziska

last: Hörhold

first: David

last: Eisel

first: Marcus

last: Oswald

first: Amol

last: Kolte

first: Daniela

last: Röll

first: Wolfram

last: Osen

first: Stefan

last: Eichmüller


title: Conditional medium versus MDI on 3T3-L1 differentiation

status: Public on Jan 24 2020

city: Seoul

first: Jae-woo

last: Kim

address: 50 Yonsei-ro, Seodaemun-gu

department: Biochem and Mol Bio

country: South Korea

institute: Yonsei University College of Medicine

summary: RNAseq was performed using 3T3-L1 cells with MDI or conditional medium (CM).

overall_design: To find a factor which is important for 3T3-L1 differentiation

first: Yoonjeong

last: Choi


title: Dissecting the regulatory strategies of NFkB RelA target genes in the inflammatory response reveals differential transactivation logics

first: Kim

last: Ngo

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

overall_design: Refer to individual Series


title: Mammalian RNA decay pathways are highly specialized and widely linked to translation

city: Basel

first: Alex

middle: Charles

last: Tuck

address: Maulbeerstrasse 66

country: Switzerland

institute: Friedrich Miescher Institute for Biomedical Research

summary: RNA decay is crucial for RNA turnover and surveillance, and misregulated in many diseases. This complex system is challenging to study, particularly in mammals, where it remains unclear whether decay pathways perform specialized or redundant roles. Cytoplasmic pathways, and links to translation, are particularly enigmatic. By directly profiling targets of decay factors (XRN1, SKIV2L and MTR4) and normal/aberrant translation events in mouse embryonic stem cells, we uncovered extensive specialization between decay pathways and crosstalk with translation. XRN1 (5’-3’) mediated cytoplasmic bulk mRNA turnover whereas SKIV2L (3’-5’) was universally recruited by ribosomes, tackling aberrant translation and sometimes modulating mRNA abundance. Further exploring translation surveillance, we identified AVEN and FOCAD as SKIV2L interactors. AVEN prevented ribosome stalls at structured regions, which otherwise required SKIV2L for clearance. This pathway was crucial for histone translation, uORF regulation and counteracting spurious non-coding RNA translation. In summary, we identified key targets, components and functions of mammalian RNA decay pathways, and uncovered extensive coupling to translation.

overall_design: mESCs with endogenously 3xFLAG-Avi-tagged decay factors (either MTR4, SKIV2L, XRN1 or AVEN) were used for crosslinking and analysis of cDNAs (CRAC), to identify transcriptome-wide direct RNA binding sites. Various WT or knockout cell lines were also profiled by RNA-seq, and some CRAC experiments were repeated in knockout backgrounds.

first: Aneliya

last: Rankova

first: Alaaddin

middle: B

last: Arpat


title: Retinal endothelial cell phenotypic modifications during experimental autoimmune uveitis: a transcriptomic approach

city: Brussels

first: Frederick

last: Libert

address: 808 route de Lennik

department: IRIBHM

country: Belgium

institute: ULB

summary: Background:  Blood-retinal barrier cells are known to exhibit a massive phenotypic change during experimental autoimmune uveitis (EAU) development. In an attempt to investigate the mechanisms of blood-retinal barrier (BRB) breakdown at a global level, we studied the gene regulation of total retinal cells and retinal endothelial cells during non-infectious uveitis.   Methods:  Retinal endothelial cells were isolated by flow cytometry either in Tie2-GFP mice (CD31+ CD45- GFP+ cells), or in wild type C57BL/6 mice (CD31+ CD45- endoglin+ cells). EAU was induced in C57BL/6 mice by adoptive transfer of IRBP1-20-specific T cells. Total retinal cells and retinal endothelial cells from naïve and EAU mice were sorted and their gene expression compared by RNA-Seq. Protein expression of selected genes was validated by immunofluorescence on retinal wholemounts and cryosections and by flow cytometry.  Results:  Retinal endothelial cell sorting in wild type C57BL/6 mice was validated by comparative transcriptome analysis with retinal endothelial cells sorted from Tie2-GFP mice, which express GFP under the control of the endothelial-specific receptor tyrosine kinase promoter Tie2. RNA-Seq analysis of total retinal cells mainly brought to light upregulation of genes involved in antigen presentation and T cell activation during EAU. Specific transcriptome analysis of retinal endothelial cells allowed us to identify 82 genes modulated in retinal endothelial cells during EAU development. Protein expression of 5 of those genes (serpina3n, lipocalin 2, ackr1, lrg1 and lamc3) was validated at the level of inner BRB cells.  Conclusion:  Those data not only confirm the involvement of known pathogenic molecules but further provide a list of new candidate genes and pathways possibly implicated in inner BRB breakdown during non-infectious posterior uveitis.

overall_design: Regulation of retinal cell and retinal endothelial cell gene expression during experimental autoimmune uveitis

first: Deborah

last: Lipski

first: Vincent

last: Foucart

first: Rémi

last: Dewispelaere

first: Laure

middle: E

last: Caspers

first: Matthieu

last: Defrance

first: Catherine

last: Bruyns

first: François

last: Willermain


title: Vascular Progenitors Generated from Tankyrase Inhibitor-Regulated Naive Diabetic Human iPSC Potentiate Efficient Revascularization of Ischemic Retina

city: Baltimore

first: Yiouli

last: Ktena

address: 1800 Orleans Street

department: SOM Onc Pediatric Oncology

institute: JHU; School of Medicine

summary: We propose that reprogramming of patient donor cells to tankyrase inhibitor-regulated naive hiPSC (N-hiPSC) improves the functionality of differentiated progenitors for subsequent regenerative therapies, and more effectively erases donor epigenetic aberrations sustained from chronic diseases such as diabetes .

overall_design: LIF-3i naïve reversion of conventional, primed hiPSC lines

first: Michael

last: Barbato

last: Koldobskiy

first: Elias

middle: T

last: Zambidis


title: Sexual Dimorphism in Cardiac Transcriptome Associated with a Troponin C Murine Model of Hypertrophic Cardiomyopathy

status: Public on Jan 23 2020

city: Tallahassee

first: Cynthia

last: Vied

state: Florida

address: 1115 W. Call St

department: Translational Science Laboratory

institute: Florida State University

summary: Heart disease remains the number one killer of women in the US. Nonetheless, studies in women and female animal models continue to be underrepresented in cardiac research. Hypertrophic cardiomyopathy (HCM), the most commonly inherited cardiac disorder, has been tied to sarcomeric protein variants. Among the susceptible genes, TNNC1—encoding cardiac troponin C (cTnC)—has contributed to a substantial HCM phenotype in mice. In this study we sought to characterize the sexual dimorphism observed within cardiac physiology and transcriptomics of adult male and female mice bearing the HCM-associated cTnC-A8V point mutation. The HCM mice showed a significant decrease in stroke volume, left ventricular diameter, volume, and relative wall thickness. Importantly, isovolumetric contraction time was significantly higher for female HCM mice. RNA sequencing revealed several altered canonical pathways within the HCM mice vs WT groups including an increase in EIF2 signaling, ILK signaling, actin nucleation by ARP-WASP complex, regulation of actin-based motility by Rho, VDR/RXR activation, and glutathione redox reactions pathways. In contrast, Valine Degradation, TCA Cycle II, Methionine Degradation, and Inositol Phosphate Compound pathways were notably down regulated in HCM mice. HCM male vs. female mice followed similar trends of the canonical pathways altered between the HCM and WT. Interestingly, seven of the genes that were differentially expressed in both the WT and HCM male vs female comparisons changed directions in fold change between the sexes. These data suggest a sexually-dimorphic HCM phenotype and identify several key pathways and genes that could be critical to sex differences seen in disease manifestation.

overall_design: Cardiac left ventricular mRNA from adult male and female mice of KI-cTnC-A8V mice were analyzed by RNA sequencing of 8 biological replicates using Illumina HiSeq 2500

first: Karissa

last: Dieseldorff Jones

first: Isela

middle: c

last: Valera

first: P

last: Chase

first: Michelle

middle: S

last: Parvatiyar

first: Jose

middle: R

last: Pinto


title: Evaluation via Supervised Machine Learning of the Broiler Pectoralis Major and Liver Transcriptome in Association with the Muscle Myopathy Wooden Breast

city: Raleigh

first: Chelsea

middle: Adele

last: Phillips

state: NC

address: 2711 Founders Drive

department: Prestage Department of Poultry Science

institute: NC State University

summary: We use machine learning from PM and liver transcriptome dataset to capture the complex relationships that are not typically revealed by traditional statistical methods.

overall_design: Two separate machine learning algorithms were performed to analyze the data set including the sequential minimal optimization (SMO) of support vector machines (SVMs) and Multilayer Perceptron (MLP) of Artificial Neural Network (ANN). Machine learning algorithms were compared to identify genes within a gene expression data set of approximately 16,000 genes for both liver and PM, which can be correctly classified from birds with or without wooden breast.

first: Christopher

last: Ashwell


title: Identification of Blood Circular RNAs as the Potential Biomarkers for Acute Ischemic Stroke

city: Guangzhou

first: Dan

last: Lu

state: Guangdong

address: No.601, West Huangpu Avenue

department: Neurology and Stroke center

institute: Jinan University

summary: Many hospitals lack facilities for accurate diagnosis of acute ischemic stroke (AIS). Circular RNA (circRNA) is highly expressed in the brain and is closely associated with stroke. In this study, we examined whether the blood-borne circRNAs can be promising candidates as adjunctive diagnostic biomarkers and their pathophysiological roles after stroke. We profiled the blood circRNA expression in mice subjected to experimental focal cerebral ischemia, and validated the selected circRNAs in AIS patients. We demonstrated that 128, 198 and 789 circRNAs were significantly altered at 5 min, 3 h and 24 h after ischemic stroke, respectively.

overall_design: circRNA expression in mouse blood of sham-operated group and of middle cerebral artery occlusion (MCAO)-operated groups at 5 min, 3 hr and 24 hr after MCAO surgery. Three independent experiments were performed in sham-operated group, MCAO-operated groups at 5 min, 3 hr and 24 hr.


title: Neonatal Genistein Exposure Causes Implantation Failure by Disrupting Uterine Developmental Patterning and Endometrial Gland Formation

city: RTP

first: NIEHS

last: Microarray Core

address: 111 T.W. Alexander Drive

department: DIR

institute: NIEHS

summary:
Female mice exposed neonatally to the phytoestrogen genistein (GEN) at doses similar to those in infants consuming soy-based infant formulas are infertile due in part to uterine implantation defects. To determine the mechanisms by which neonatal GEN exposure causes implantation defects, female mice were exposed to vehicle (corn oil) or GEN in corn oil (50 mg/kg/day) on postnatal days (PND) 1-5.
Uterine tissues were collected during early gestation for microarray analysis; gestation day 1.5 (GD1.5), GD3.5, GD4.5 and GD5.5. We conclude that neonatal GEN exposure disrupts expression of genes important for uterine development, causing posteriorization and diminished gland function during pregnancy that together cause implantation failure.

overall_design: CD-1 mice were exposed to corn oil as controls (CON) or genistein (GEN) at a dose of 50 mg/kg/day on postnatal days 1-5. Uteri were collected on gestation day 1.5 (GD1.5), GD3.5, GD4.5 or GD5.5

first: Wendy

middle: N

last: Jefferson

first: Carmen

last: Williams


title: Disruption of DNA damage response by ARID2 knockout in human hepatocellular carcinoma cells

status: Public on Jan 22 2020

city: Bunkyo-ku

first: Shu

last: Shimada

state: Tokyo

address: 1-5-45 Yushima

department: Department of Molecular Oncology

country: Japan

institute: Tokyo Medical and Dental University

summary: Background & Aims: Recent genomic studies have identified frequent mutations of AT-rich interactive domain 2 (ARID2) in hepatocellular carcinoma (HCC), but it is not still understood how ARID2 exhibits tumor suppressor activities. Methods: We established the ARID2 knockout HCC cell lines by using CRISPR/Cas9 system, and investigated the gene expression profiles and biological functions. Results: Bioinformatic analysis indicated that UV-response genes were negatively regulated in the ARID2-KO cells, and they were certainly sensitized to UV irradiation. ARID2 depletion attenuated nucleotide excision repair (NER) of DNA damage sites introduced by exposure to UV as well as chemical compounds known as carcinogens for HCC, benzo[a]pyrene and FeCl3, since XPG could not be accumulated without ARID2. By using large-scale public data sets, we validated that ARID2 knockout could lead to similar molecular changes between in vitro and in vivo, and moreover observed a higher number of somatic mutations in the ARID2-mutated subtypes than that in the ARID2 wild-type across various types of cancers including HCC. Conclusions: We provided evidence that ARID2 knockout could contribute to disruption of NER process through inhibiting the recruitment of XPG, resulting in susceptibility to carcinogens and potential hypermutation. These findings have far-reaching implications for therapeutic targets in cancers harboring ARID2 mutations.

overall_design: We established the ARID2 knockout HCC cell lines (JHH4 and JHH5) by using CRISPR/Cas9 system, and investigated the gene expression profiles and biological functions.

first: Atsushi

last: Oba

first: Yoshimitsu

last: Akiyama

first: Taketo

last: Nishikawaji

first: Kaoru

last: Mogushi

first: Hiromitsu

last: Ito

first: Satoshi

last: Matsumura

first: Arihiro

last: Aihara

first: Yusuke

last: Mitsunori

first: Daisuke

last: Ban

first: Takanori

last: Ochiai

last: Kudo

first: Hiroshi

last: Asahara

last: Kaida

first: Masahiko

last: Miura

first: Minoru

last: Tanabe

first: Shinji

last: Tanaka


title: Classification of Primary Liver Cancer with Immunosuppression Mechanisms and Correlation with Genomic Alterations

status: Public on Jan 21 2020

city: Yokohama

first: Hidewaki

last: Nakagawa

address: 1-7-22 Suehiro-cho, Tsurumi-ku

institute: RIKEN Center for Integrative Medical Sciences

summary: Background: The tumor microenvironment can be classified into immunologically active “inflamed” tumors and inactive “non-inflamed” tumors based on the infiltration of cytotoxic immune cells. Previous studies on liver cancer have reported a superior prognosis for inflamed tumors compared to non-inflamed tumors. However, liver cancer is highly heterogeneous immunologically and genetically, and a finer classification of the liver cancer microenvironment may improve our understanding of its immunological diversity and response to immune therapy.   Methods: We characterized the immune gene signatures of 234 primary liver cancers, mainly virus-related, from a Japanese population using RNA-Seq of tumors and matched non-tumorous hepatitis livers. We then compared them with the somatic alterations detected using the whole-genome sequencing.  Findings: Liver cancers expressed lower levels of immune marker genes than non-tumorous hepatitis livers, indicating immunosuppression in the tumor microenvironment. Several immunosuppression mechanisms functioned actively and mutually exclusively, resulting in four immune subclasses of liver cancer: tumor-associated macrophage (TAM), CTNNB1, cytolytic activity (CYT), and regulatory T cell (Treg). The CYT and Treg subclasses represented inflamed tumors, while the TAM and CTNNB1 subclasses represented non-inflamed tumors. The TAM subclass, which comprised 31% of liver cancers, showed a poor survival, expressed elevated levels of extracellular matrix genes, and was associated with somatic mutations of chromatin regulator ARID2. The results of cell line experiments suggested a functional link between ARID2 and chemokine production by liver cancer cells.  Interpretation: Primary liver cancer was classified into four subclasses based on mutually exclusive mechanisms for immunosuppression. This classification indicate the importance of immunosuppression mechanisms, such as TAM and Treg, as therapeutic targets for liver cancer.

overall_design: mRNA profiles of ARID2 knockout and wildtype JHH4 cells were generated by RNA-Seq in triplicate.


title: miRNA expression profiling in the physiological cardiac hypertrophy of rats

first: Jun

last: Zhang

address: No.100 of Guilin Road

institute: Shanghai Normal University

summary: Using the highly sensitive miRNA array, we screened 220 microRNAs abundant in physiological left ventricular hypertrophy (LVH) and we explored the functions of these miRNAs in the cardiac tissue by Gene Ontology and Kyoto Encyclopedia of Genes annotation. miRNAs showed a high score in the pathway enriched in autophagy. Moreover, the expression levels of miR-26b-5p, miR-204-5p, and miR-497-3p showed an obvious increase in rat hearts. Adenovirus-mediated overexpression of miR-26b-5p, miR-204-5p, and miR-497-3p markedly attenuated IGF-1-induced hypertrophy in H9C2 cells by suppressing autophagy. Furthermore, miR-26b-5p, miR-204-5p, and miR-497-3p attenuated autophagy in H9C2 cells through targeting ULK1, LC3B and Beclin 1, respectively. Taken together, our results demonstrate that swimming exercise induced physiological LVH, at least in part, by modulating the microRNA–autophagy axis, and that miR-26b-5p, miR-204-5p, and miR-497-3p may help distinguish physiological and pathological LVH.

overall_design: In this study, samples from two animals in each group were used to acquire the miRNA expression profiling and the function of the abudant miRNAs in the cardiac tissue were analyzed by bioinformatic methods,finally, our results demonstrate that swimming exercise induced physiological LVH, at least in part, by modulating the microRNA–autophagy axis, mainly inveolved in the miR-26b-5p, miR-204-5p, and miR-497-3p.

first: Jie

last: Qi


title: Transcriptome of Cryptococcus neoformans and Cryptococcus deneoformans

city: Paris

first: Guilhem

last: Janbon

address: 25 rue du Dr Roux

institute: Institut Pasteur

summary: The goal of this study was to position all transcripts extremities in two species of Cryptococcus using TSS-Seq and QuantSeq 3' mRNA-Seq when cells are grown under different conditions. We analysed also the level of expression of each genes in the same condition using the same cell sample. All these data have spiked in using a fixed quantity of S. cereviae cells added just before DNA and RNA extraction.

sample_organism:
Cryptococcus neoformans
Cryptococcus deneoformans

overall_design: total RNA and DNA  were extracted different Cryptococcus  wild type and mutant strains and sequenced. TSS and PAS were positioned and gene expression was evaluated.

first: Corinne

last: Maufrais


title: Analysis of PDCD4 knockdown effects by high throughput sequencing of a human epithelial cell line

status: Public on Jan 20 2020

city: Münster

first: Astrid

last: Haas

department: Biochemistry

institute: WWU Münster

summary: Purpose: examination of gene expression and translation changes in immortalized human epithelial cells (hTERT-RPE-1) induced by PDCD4 siRNA-knockdown.

overall_design: Knockdown induction: hTERT-RPE-1 cells were transfected either with PDCD4-siRNA or control-siRNA (100 nM) and incubated for 24 h. Induced effects were analyzed by ribosome profiling and RNA-seq in triplicate (IlluminaSequencing).

first: Karl-Heinz

last: Klempnauer

first: Benedikt

last: Nilges

first: Sebastian

last: Leidel


title: The epidermis coordinates thermoresponsive growth through the phyB-PIF4-auxin pathway

city: Stanford

first: Eunkyoo

last: Oh

address: 260 Panama Street

department: Plant Biology

institute: Carnegie Institution

summary: In plants, an elevation in ambient temperature induces morphological changes including elongation hypocotyls, considered to be adaptive responses to alleviate the heat-induced damages. The high temperature-induced morphological changes are called thermomorphogenesis, which is predominantly regulated by a bHLH transcription factor PIF4. Although PIF4 is expressed in all aerial tissues including the epidermis, mesophyll, and vascular bundle, its tissue-specific functions in thermomorphogenesis are not known. Here, we found that epidermis-specific expression of PIF4 induced constitutive long hypocotyls, while vasculature-specific expression of PIF4 had no effect on hypocotyl growth. Consistently, RNA-Seq and qRT-PCR analyses revealed that auxin responsive genes and growth-related genes were highly activated by epidermal, but not by vascular, PIF4. The epidermal, but not vascular, inactivation of PIF4 by a PIF4 artificial microRNA or a dominant negative form of PIF4 suppressed thermoresponsive gene expression and hypocotyl growth. Additionally, both the block of epidermal auxin signaling and the epidermal overexpression of a thermosensor phytochrome B (phyB) inhibited thermoresponsive growth, indicating that epidermal PIF4-auxin pathway are essential for the temperature responses. We further show that epidermal PIF4 is increased by high temperatures mainly through the transcriptional activation of PIF4. Taken together, our study demonstrates that the epidermis regulates thermoresponsive growth through the phyB-PIF4-auxin pathway.

overall_design: Seedlings were grown under white light for 5 days at 20 °C before harvesting for total RNA extraction.


title: An Adaptable Soft-mold Embossing Process for Fabricating Optically-accessible, Microfeature-based Culture Systems and Application toward Liver Stage Antimalarial Compound Testing

status: Public on Feb 01 2020

city: Tampa

first: Rays HY

last: Jiang

state: FL

address: 3720 Spectrum Blvd, Suite 304

department: Department of Global Health

institute: University of South Florida

summary: Advanced cell culture methods to model organ function have been demonstrated to better replicate in vivo conditions and produce data with improved translational value.  Because the liver is parituclarly important to human health, several advanced culture methods exist to experiment with liver disease states, including the infectious disease malaria, which can be better controlled with development of new antimalarials effective against the liver parasite population. We developed a novel fabrication method to emboss hepatocyte-confining microfeatures into polysytrene, resulting in microfeature-based hepatocyte confinement (μHEP) slides and plates. RNAseq of hepatocytes in μHEP systems demonstrated microfeatures sustain hepatic differentiation and function, suggesting broader utility for preclinical hepatic assays; while our tailorable embossing process could be repurposed for developing additional organ models.

overall_design: Examination of primary human hepatocyte transcriptome in different culture platforms

first: Steven

middle: P

last: Maher

first: Amy

last: Conway

first: Swamy

last: Adapa

first: Rays

middle: H

first: Dennis

last: Kyle


title: The role of Mediator and Little Elongation Complex in transcription termination

city: Kansas City

first: Shiyuan

middle: (Cynthia)

last: Chen

state: MO

department: Computational Biology

institute: Stowers Institute for Medical Research

summary: Mediator is a coregulatory complex involved in regulating the transcription of Pol II-dependent genes. Metazoan Mediator subunit MED26 functions as a docking site for the ELL/EAF-containing Super Elongation Complex (SEC) and L ittle Elongation Complex (LEC), which regulate the expression of distinct genes. MED26 helps to recruit SEC to protein-coding genes including c-Myc and LEC to small nuclear RNA (snRNA) genes. However, why MED26 takes advantage of SEC or LEC to regulate different claases of genes is unclear. Here, we present evidence that MED26 recruits LEC to support optimal transcription termination at non-polyadenylated genes including snRNA and replication-dependent histone (RDH) genes. Our findings indicate that LEC recruited by MED26 promotes efficient transcription termination by Pol II through interaction with CBC-ARS2 and NELF/DSIF, and then LEC promotes 3’-end processing through the recruitment of Integrator or Heat labile factor to snRNA or RDH genes, respectively.

overall_design: Experiment 1: Rpb1 ChIP-seq HEK293T WT and MED26 mutant cells, using mouse chromatin as spike-in. Experiment 2: MED26 ChIP-seq in HEK293T WT cells. Experiment 3: PRO-seq in HEK293T WT and MED26 mutant cells, using D. melanogastor Kc167 cells as spike-in control. Experiment 4: RNA-seq of polyA-selected libraries prepared from HEK293T cells treated with control, AFF4, KIAA0947/ICE1, or MED26 siRNAs.  Experiment 5: RNA-seq of ribo-depleted libraries prepared from HEK293T cells treated with control or MED26 siRNAs. Experiment 6: RNA-seq of ribo-depleted libraries prepared from HEK293T WT or MED26 mutant cells.

first: Hidehisa

last: Takahashi

first: Amol

last: Ranjan

first: Kai

first: Joan

middle: W

last: Conaway

first: Shigetsugu

last: Hatakeyama


title: An advanced intestinal tissue model reveals host and pathogen  strategies during Salmonella infection

city: Marburg

first: Leon

middle: Nicolas

last: Schulte

state: Hessen

address: Hans-Meerwein-Str. 2

department: Institute for Lung Research

institute: Philipps-Universität Marburg

summary: Here, we introduce an intestinal tissue model to study human enteric infections. Our model comprises epithelial and endothelial layers, a primary intestinal collagen scaffold, and immune cells. We use Dual RNA-seq to chart the communication amongst several different cell types at the intestinal barrier and the pathogen. The results suggest that Salmonella uses its type III secretion systems to manipulate STAT3-dependent inflammatory responses locally in the epithelial compartment. Our approach promises to reveal more human-specific infection strategies employed by Salmonella and other pathogens.

sample_organism: Salmonella enterica subsp. enterica serovar Typhimurium str. SL1344

overall_design: Expression profiles of rRNA depleted total RNA from human intestinal epithelial, endothelial, NK and monocytic cells and Salmonella (+/- S. Typhimurium infection) were recorded in two replicates.

first: Marco

last: Metzger

first: Jörg

last: Vogel


title: GPCR-induced YAP activation sensitizes fibroblasts to profibrotic effects of TGFβ1

city: Allschwil

first: Axel

last: Klenk

address: Hegenheimermattweg 91

department: Research Informatics

institute: Idorsia Pharmaceuticals Ltd.

summary: Fibrosis is an uncontrolled wound healing process resulting from tissue injury, inflammation and fibroblast activation, finally leading to accumulation of extracellular matrix (ECM) components in tissues and to organ failure. The underlying mechanisms of fibrosis have been intensely studied and it has become clear that multiple pathways and their crosstalk regulate fibrotic diseases with a few central players, including TGFβ1. Recently, YAP and TAZ were implicated in fibrosis as one of the essential components of its pathogenesis. YAP/TAZ are known to crosstalk with the TGFβ pathway in the nucleus and regulate its activity on transcriptional level by binding to Smad2/3/4 complexes. We hypothesized that factors, such as GPCR ligands which regulate YAP/TAZ, might modulate the profibrotic responses to TGFβ1. We performed gene expression microarray in NHDF treated with TGFβ1, LPA, the combination of TGFβ1/LPA or vehicle. The aim was to have a broader look at transcriptional signatures induced by the individual treatments and in the combination of TGFβ1 and LPA.

overall_design: Comparison of primary human skin fibroblasts stimulated with TGFβ1, LPA and the combination of TGFβ1 plus LPA; 4 replicates per group

first: Katarina

last: Zmajkovicova

first: Yasmina

last: Bauer

first: Marie

last: Schnoebelen

first: Diego

last: Freti

first: Maxime

last: Boucher

first: Berengere

last: Renault

first: Rolf

last: Studer

first: Magdalena

last: Birker

first: Oliver

last: Nayler

first: John

last: Gatfield


title: Regenerating zebrafish fin epigenome is characterized by stable lineage-specific DNA methylation and dynamic chromatin accessibility

city: St. Louis

first: Hyung Joo

last: Lee

address: 4515 McKinley Avenue

department: Department of Genetics

institute: Washington University School of Medicine

type: Methylation profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Regenerating zebrafish fin epigenome is characterized by stable lineage-specific DNA methylation and dynamic chromatin accessibility (WGBS)

summary:
Background: Zebrafish can faithfully regenerate injured fins through the formation of a blastema, a mass of proliferative cells that can grow and develop into the lost body part. After amputation, various cell types contribute to blastema formation, where each cell type retains fate restriction and exclusively contributes to regeneration of its own lineage. Epigenetic changes that are associated with lineage restriction during regeneration remain underexplored.

Results: We produce epigenome maps, including DNA methylation and chromatin accessibility, as well as transcriptomes, of osteoblasts and other cells in uninjured and regenerating fins. This effort reveals regeneration as a process of highly dynamic and orchestrated transcriptomic and chromatin accessibility changes, coupled with stably maintained lineage-specific DNA methylation. The epigenetic signatures also reveal many novel regeneration-specific enhancers, which are experimentally validated. Regulatory networks important for regeneration are constructed through integrative analysis of the epigenome map, and a knockout of a predicted upstream regulator disrupts normal regeneration, validating our prediction.

Conclusion: Our study shows that lineage-specific DNA methylation signatures are stably maintained during regeneration, and regeneration enhancers are preset as hypomethylated before injury. In contrast, chromatin accessibility is dynamically changed during regeneration. Many enhancers driving regeneration gene expression as well as upstream regulators of regeneration are identified and validated through integrative epigenome analysis.

overall_design: Methylation profiles of the uninjured and regenerating fin by deep sequencing, in duplicate, using Illumina NextSeq and NovaSeq.

first: Ting

last: Wang

first: Stephen

middle: L

last: Johnson


title: Regenerating zebrafish fin epigenome is characterized by stable lineage-specific DNA methylation and dynamic chromatin accessibility (ATAC-seq)

overall_design: Chromatin accessibility profiles of the uninjured and regenerating fin by deep sequencing, in duplicate, using Illumina NextSeq.


title: A hypermethylation strategy utilized by enhancer-bound CARM1 to promote estrogen receptor a-dependent transcriptional activation and breast carcinogenesis

city: Xiamen

first: Jiancheng

last: Ding

state: Fujian

address: School of Pharmaceutical Sciences, Xiamen University Xiang`an South Road

institute: Xiamen University


title: Super-enhancer subregions highly bound by the nuclear receptor ESRRB regulate the exit from naïve pluripotency

status: Public on Jan 19 2020

city: Nijmegen

first: Hendrik

last: Marks

address: Geert Grooteplein 26/28

department: Molecular Biology

country: Netherlands

institute: Radboud University Nijmegen, RIMLS

summary: This study focuses on superenhancers, and includes 1 WGBS profile of EpiSCs.

overall_design: DNA methylation profiling using whole-genome bisulfite sequencing of a pluripotent mouse EpiSC line.

first: Wout

last: Megchelenbrink

first: Emma

last: Bell

first: Ed

last: Curry

first: Luc

last: Jouneau

first: Vincent

last: Brochard

first: Rute

middle: A

last: Tomaz

first: King

last: Mau

first: Henk

middle: G

last: Stunnenberg

first: Alice

first: Véronique

last: Azuara


title: Functional heterogeneity within the developing zebrafish epicardium

city: Oxford

first: Paul

middle: Richard

last: Riley

address: IMS-Tetsuya Nakamura Building, Old Road Campus

country: United Kingdom

institute: University of Oxford

summary: The epicardium is essential during cardiac development, homeostasis and repair and yet fundamental insights into its underlying cell biology, notably epicardium formation, lineage heterogeneity and functional cross-talk with other cell types in the heart, is currently lacking. In this study, we investigated epicardial heterogeneity and the functional diversity of discrete epicardial subpopulations in the developing zebrafish heart. Single-cell RNA-sequencing uncovered three epicardial subpopulations with specific genetic programmes and distinctive spatial distribution within the developing heart. Perturbation of unique gene signatures uncovered distinct functions associated with each subpopulation and established novel epicardial roles in cell adhesion, migration, and chemotaxis as a mechanism for recruitment of leukocytes into the heart. This work elucidates the mutual spatiotemporal relationships between different epicardial subpopulations and assigns unique function to each during cardiac development. Understanding which mechanisms cells employ to establish a functional epicardium and to communicate with other cardiovascular cell types during development will bring us closer to repairing cellular relationships that are disrupted during cardiovascular disease.

overall_design: Single cell RNA sequencing of FACS purified larval cardiac zebrafish cells

first: Michael

last: Weinberger

first: Filipa

middle: C

last: Simões

first: Roger

last: Patient

first: Tatjana

last: Sauka-Spengler


title: Inhibiting WNT and NOTCH in renal cancer stem cells: implications for human patients

city: Berlin

first: Annika

last: Fendler

address: Robert-Roessle-Str. 10

department: Signal Transduction in Development and Cancer

institute: Max Delbrueck Center for Molecular Medicine in the Helmholtz Association

summary: We detected and characterized CXCR4+MET+CD44+ cancer stem cells (CSCs) that were quantitatively correlated with aggressive tumors. Transcriptional profiling and single-cell sequencing revealed a strong activation of WNT and NOTCH signaling in these CSCs. CEL-Seq was performed to study heterogeneity of the CXCR4+MET+CD44+ population.

overall_design: We analyzed freshly isolated single cells from ccRCC tumor tissue of two patients. 90 cells per patient were analyzed (*sc). We included 2 controls: CXCR4+MET+CD44+ cells (10.000 cells, triplicate, *CTC) and control unsorted ccRCC cells (10.000 cells, triplicate, *bulk)

first: Kun

last: Son

first: Wei

last: Chen

first: Walter

last: Birchmeier


title: Chip-seq data of mouse Spermatogonia cells of Tet1-ko and WT

status: Public on Jan 18 2020

city: Tianjin

first: Linlin

last: Liu

address: Weijin Road 97

institute: Nankai university

summary: We analysed 2 samples of mouse spermatogonia cells including  3-month-old WT and 3 month-old Tet1

overall_design: Tet1 chip of mouse spermatogonia cells including 3-month WT and 3-month Tet1 knock out mice.

first: Guian

last: Huang

first: Huasong

last: Wang

first: Lin


title: RNA-seq data of Spermatogonia cells of Tet1-ko and WT

summary: We analyzed 3 repeats of 12 samples of mouse spermatogonia cells including  3-month-old WT and 3 month-old Tet1, 11-month-old WT and 11-month-old Tet1, aimed to find Tet1 function on male fertility.


title: Anti-inflammatory properties of low dose epirubicin on human macrophages

status: Public on Jan 17 2020

city: Jena

first: Sven

last: Stengel

address: Am Klinikum 1

department: Clinic for Internal Medicine IV

institute: Jena University Hospital

summary:
Low-dose epirubicin at non-cytotoxic doses down regulated NLRP3 inflammasome components and reduced the release of proinflammatory cytokines.
These anti-inflammatory effects were a result of global transcriptional suppression of LPS-dependent genes.

overall_design: THP-1 cells were differentiated to macrophages and cells were treated with or without epirubicin (0.25 μg/ml) for 24h and with or without LPS (100 ng/ml) for three hours resulting in four treatment conditions (1. -EPI/- LPS; 2. –EPI/+LPS;  3. +EPI/-LPS; 4. +EPI/+LPS). RNA was isolated and microarray analysis was performed by a commercial transcriptomics provider. Expression analysis was performed using Transcriptome analysis concole (TAC).

first: Tony

last: Bruns


title: Phase separation directs ubiquitination of gene body nucleosomes

city: State College

first: Chitvan

last: Mittal

address: 453 North Frear, University Park

department: BMB

institute: Pennsylvania State University

summary: The conserved yeast E3 ligase Bre1 and its partner E2 Rad6 monoubiquitinate histone H2B across gene bodies during the transcription cycle. While processive ubiquitination might in principle arise from Bre1/Rad6 traveling with RNA polymerase II, we provide a different explanation. Here we implicate liquid-liquid phase separation as the underlying mechanism. Biochemical reconstitution shows that Bre1 binds the scaffold protein Lge1, whose intrinsically disordered region phase separates via dynamic, multivalent interactions. The resulting condensates comprise a core of Lge1 encapsulated by an outer catalytic shell of Bre1. This layered liquid recruits Rad6 and the nucleosomal substrate, accelerating H2B ubiquitination. In vivo, the condensate-forming region of Lge1 is required to ubiquitinate H2B in gene bodies beyond the +1 nucleosome. Our data suggest that layered condensates of histone modifying enzymes generate chromatin-associated reaction chambers with augmented catalytic activity along gene bodies. Equivalent processes may occur in human cells, causing neurological disease when impaired.

overall_design: Enrichment of H2BK123ub and underlying H2B levels in WT and various LGE1 mutants were determined using highthroughput paired end sequencing using ChIP-exo 5.0. In addition, Genome wide binding profile of Lge1 in WT and bre1Δ backgrounds were determined using ChIP-exo 5.0.

first: Laura

middle: D

last: Gallego

first: Maren

last: Schneider

first: Anete

last: Romanauska

first: Ricardo

middle: M

last: Gudino Carrillo

first: Tobias

last: Schubert

first: B

last: Franklin Pugh

first: Alwin

last: Köhler


title: Control of noncoding RNA production and histone levels by a 5’ tRNA fragment

city: Princeton

first: Mike

last: Levine

state: NJ

address: South Dr

department: Lewis Sigler Institute

institute: Princeton University

summary: Small RNAs derived from mature tRNAs, referred to as tRNA fragments or “tRFs”, are an emerging class of regulatory RNAs with poorly understood functions in cellular regulation. We recently identified a role for one specific tRF – 5’ tRF-Gly-GCC, or tRF-GG – in repression of genes associated with the endogenous retroelement MERVL, but the mechanistic basis for this regulation was unknown. Here, we show that tRF-GG plays a role in production of a wide variety of noncoding RNAs normally synthesized in Cajal bodies. Among these noncoding RNAs, tRF-GG regulation of the U7 snRNA modulates heterochromatin-mediated transcriptional repression of MERVL elements by supporting an adequate supply of histone proteins. Importantly, the effects of inhibiting tRF-GG on histone mRNA levels, activity of a histone 3’ UTR reporter, and ultimately on MERVL regulation could all be suppressed by the U7 RNA. We show that the related RNA-binding proteins hnRNPF and H bind directly to tRF-GG, and are required for Cajal body biogenesis. Together, our data reveal a conserved mechanism for 5’ tRNA fragment control of noncoding RNA biogenesis and, consequently, in global chromatin organization.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Mouse embryonic stem cells were transfected with tRNA fragments, LNA sequences against tRNA fragments, snRNAs, or siRNAs. Changes in gene expression were quantified using RNAseq and metabolic labeling. Changes in ribosomal rRNA methylation were quantified using Ribomethseq. Changes in chromatin architecture were probed with ATAC-seq. E14 mESCs  were transfected with LNA-containing oligos antisense to tRF-GG or Mock transfected. iClip libraries were constructed against hnRNPF/H using sc-32310 antibody, with protocol from Zarnegar et al. 2016 Nat Methods.

first: X

middle: Y

last: Bing

first: O

middle: J

last: Rando


title: Transcriptional response of Pseudomonas aeruginosa to the prebiotic Fructooligosaccharide

city: Malaga

first: Carlos

middle: Molina

last: Santiago

state: Malaga

address: Bulevar Louis Pasteur 49

department: MIcrobiology

country: Spain

institute: UMA

summary: The bifidogenic effect of the prebiotic inulin and its hydrolysed form (fructoligosaccharide, FOS) is well established since they promote the growth of specific beneficial (probiotic) gut bacteria such as bifidobacteria. Previous studies of the opportunistic nosocomial pathogen Pseudomonas aeruginosa PAO1 have shown that inulin and FOS reduce growth, biofilm formation, through decrease of its ability to motility and exotoxin secretion. However, the transcriptional basis for these phenotypic alterations remains unclear. To address this question we conducted RNA-sequence analysis.  In most cases changes in the transcript level induced by inulin and FOS were similar.  However, a set of transcripts were increased in expression response to inulin, but reduced in the presence of FOS. In the presence of inulin or FOS 260 and 217 transcript levels, respectively, were altered as compared to the control to which no polysaccharide was added. Importantly, changes in  transcript levels of 57 and 83 genes were found to be specific for either inulin or FOS, respectively, indicating that both compounds trigger different changes. Gene pathway analysis of different exposure genes (DEG) revealed a specific FOS-mediated reduction in transcript levels of genes that participate in several canonical pathways involved in metabolism and growth, motility, biofilm formation, β-lactam resistance and in the modulation of type III and VI secretion system which were supported by real time quantitative PCR measurements. These results may provide solid information to suggest that FOS selectively modulates the P. aeruginosa PAO1 pathogenecity through distinct signaling pathways.

overall_design: Pseudomonas aeruginosa PAO1 was treated with Inulin and FOS

first: A

last: Daddaoua

first: C

last: Molina-Santiago


title: A hypermethylation strategy utilized by enhancer-bound CARM1 to promote estrogen receptor a-dependent transcriptional activation and breast carcinogenesis (RNA-seq)

first: Jiancheng

last: Ding

summary: While protein arginine methyltransferases (PRMTs) and PRMT-catalyzed protein methylation have been well-known to be involved in a myriad of biological processes, their roles in carcinogenesis, particularly in estrogen receptor alpha (ERa)-positive breast cancers, remain incompletely understood. Here we focused on investigating PRMT4 (also called coactivator associated arginine methyltransferase 1, CARM1) due to its high expression and the associated poor prognosis in ERa-positive breast cancers. We first uncovered the chromatin-binding landscape and transcriptional targets of CARM1 in the presence of estrogen in ERa-positive breast cancer cells employing genomic and transcriptomics approaches. CARM1 was found to be predominantly and specifically recruited to ERa-bound active enhancers and essential for the transcriptional activation of cognate estrogen-induced gene transcriptional activation in response to estrogen. Global mapping of CARM1 substrates revealed that CARM1 methylates a large cohort of proteins with diverse biological functions, including regulation of intracellular estrogen receptor signaling, chromatin organization, chromatin remodeling and others. Intriguingly, a number of proteins were hypermethylated exclusively by CARM1 on a cluster of arginine residues. Exemplified by MED12, hypermethylation of these proteins by CARM1 served as a molecular beacon for recruiting coactivator protein, tudor domain-containing 3 (TDRD3), to ensure the full activation of estrogen/ERa target genes. In consistent with its critical role in estrogen-induced gene transcriptional activation, CARM1 was found to promote cell proliferation of ERa-positive breast cancer cells in vitro and tumor growth in mice. Taken together, our study uncovered a “hypermethylation” strategy utilized by CARM1 in gene transcriptional regulation, and suggested that CARM1 can server as a therapeutic target for breast cancer treatment.

overall_design: RNA-seq performed in this study was designed to understand the molecular mechanisms underlying CARM1 regulation of breast cancer

first: Wen

last: Liu

first: Bing-Ling

last: Peng

first: Jian-Cheng


title: Tuft cell formation reflects epithelial plasticity in pancreatic injury; implications for modeling human pancreatitis

status: Public on Jan 16 2020

city: Nashville

first: Kathleen

middle: E

last: DelGiorno

state: TN

address: 465 21st Ave S

department: Cell and Developmental Biology

institute: Vanderbilt University

summary: We performed RNA-seq on pancreatic tuft cells and non-tuft epithelial cells in caerulein-induced pancreatitis

overall_design: The transcriptomes of pancreatic tuft cells and non-tuft epithelial cells were profiled. Five biological replicates for each cell type.

first: Geoffrey

last: Wahl

first: Chi-Yeh

last: Chung


title: A topological cluster of differentially regulated genes in mice lacking PER3

status: Public on Jan 15 2020

city: Guildford

first: Emma

last: Laing

address: Stag Hill

institute: University of Surrey

summary:
Polymorphisms in human circadian clock gene PERIOD3 (PER3) are associated with a wide variety of phenotypes such as diurnal preference, delayed sleep phase disorder, sleep homeostasis, cognitive performance, bipolar disorder, type 2 diabetes, cardiac regulation, cancer, light sensitivity, hormone and cytokine secretion, and addiction. However, the molecular mechanisms underlying these phenotypic associations remain unknown.
Per3 knockout mice (Per3-/-) have phenotypes related to activity, sleep homeostasis, anhedonia, metabolism, and behavioural responses to light. Using a protocol that induces behavioural differences in response to light in wild type and Per3-/- mice, we compared genome-wide expression in the eye and hypothalamus in the two genotypes. Differentially expressed transcripts were related to inflammation, taste, olfactory and melatonin receptors, lipid metabolism, cell cycle, ubiquitination, and hormones, as well as receptors and channels related to sleep regulation. Differentially expressed transcripts in both tissues co-localised with Per3 on an ~8Mbp region of distal chromosome 4. The most down-regulated transcript is Prdm16, which is involved in adipocyte differentiation and may mediate altered body mass accumulation in Per3-/- mice. eQTL analysis with BXD mouse strains showed that the expression of some of these transcripts and also others co-localised at distal chromosome 4, is correlated with brain tissue expression levels of Per3 with a highly significant linkage to genetic variation in that region.
These data identify a cluster of transcripts on mouse distal chromosome 4 that are co-regulated with Per3 and whose expression levels correlate with those of Per3. This locus lies within a topologically associating domain island that contains many genes with functional links to several of the diverse non-circadian phenotypes associated with polymorphisms in human PER3

overall_design: Male, adult WT and Per3-/- mice were exposed to 17 cycles of an LD 3.5:3.5. In the second hour of the last ultradian light cycle, mice (N=8 per genotype) were killed by cervical dislocation and tissue (brain and whole eye) was collected.  mRNA was purified from a frozen block of hypothalamic tissue and whole eye. RNA was used to prepare labelled cRNA, which was hybridised to Agilent Whole Mouse Genome slides (60mer, 4x44K features).

first: Daan

middle: R

last: van der Veen

first: Sung-Eun

last: Bae

first: Jonathan

last: Johnston

first: Derk-Jan

last: Dijk

first: Simon

middle: N

last: Archer


title: ATACseq of kdrl-GFP+ endothelial cells in 28hpf zebrafish embryos

city: Birmingham

first: Rui

last: Monteiro

address: Medical School, Vincent Drive

department: Institute of Cancer and Genomic Sciences

institute: University of Birmingham

summary: The goal of this study is to analyse the open chromatin regions  via ATACseq in zebrafish embryonic endothelial cells at 26-27hpf using Tg(Kdrl:GFP) transgenic zebrafish embryos. 40000-50000 cells were FAC-sorted directly into 100 µl of 1x HBSS/10 mM HEPES/0.25% BSA buffer for Tagmentation (as described by Buenrostro et al. (2013), Nat. Methods 10, 1213-1218), but with 1.5 µl Tn5 transposase (Illumina) in a 50 µl reaction volume. DNA was purified using the QIAquick PCR purification kit (Qiagen). Fragments were amplified for 16 cycles.

overall_design: Genomic control DNA was isolated from 30.000 cells using the DNeasy Blood & Tissue kit (Qiagen) and eluted with EB buffer. Tagmentation was performed as described. Fragments were amplified for 9 cycles. Sequencing was performed on a NextSeq machine. Sequenced reads were checked for base qualities, trimmed where 20% of the bases were below quality score 20, and filtered to exclude adapters using Trimmomatic (Version 0.32).

first: Rossella

last: Rispoli


title: Joint Profiling of Chromatin Accessibility and CAR-T Integration Site Analysis at Population and Single-cell Levels

city: Philadelphia

first: Wenliang

last: Wang

state: PENNSYLVANIA

address: 421 Curie Boulevard

institute: University of Pennsylvania


title: Regenerating zebrafish fin epigenome is characterized by stable lineage-specific DNA methylation and dynamic chromatin accessibility (RNA-seq)

first: Hyung Joo

last: Lee

overall_design: mRNA profiles of the uninjured and regenerating fin by deep sequencing, in duplicate, using Illumina NextSeq.

first: Ting

first: Stephen

last: Johnson


title: Oligosaccharyl transferase that maintains ER homeostasis enhances immunity via up-regulating p38 MAP kinase signaling in C. elegans

status: Public on Jan 30 2020

city: Daejeon

first: Seung-Jae

middle: V

address: 291 Daehak-ro

department: Biological Sciences

institute: KAIST

summary: Oligosaccharyl transferase (OST) protein complex mediates the N-linked glycosylation of substrate proteins in the endoplasmic reticulum (ER), which regulates stability, activity and localization of its substrates. Although many OST substrate proteins have been identified, the physiological role of OST complex remains incompletely understood. Here, we show that OST complex in C. elegans is crucial for ER protein homeostasis and enhances resistance to pathogenic bacteria, Pseudomonas aeruginosa (PA14), via immune-regulatory PMK-1/p38 MAP kinase.

overall_design: The investigation of the changes of gene expression upon knockdown of stt-3 or infection with PA14

first: Dae-Eun

last: Jeong

first: Yujin

first: Seokjin

last: Ham

first: Dongyeop


title: Joint Profiling of Chromatin Accessibility and CAR-T Integration Site Analysis at Population and Single-cell Levels II

summary: We report the development of a novel method called EpiVIA (https://github.com/VahediLab/epiVIA) for the joint profiling of the chromatin accessibility and lentiviral integration site analysis at the population and single-cell levels. We validate our technique in clonal cells with previously defined integration sites and further demonstrate the ability to measure lentiviral integration sites and chromatin accessibility of host and viral genomes at the single-cell resolution in CAR-T cells. We anticipate that EpiVIA will enable the single-cell deconstruction of gene regulation during CAR-T therapy, leading to the discovery of cellular factors associated with durable treatment.

overall_design: And identify the integration sites in two groups of CAR-T cells with single-cell ATAC-seq

first: Golnaz

last: Vahedi


title: Joint Profiling of Chromatin Accessibility and CAR-T Integration Site Analysis at Population and Single-cell Levels I

overall_design: Idenfification of integration sites in two clonal cells with ATAC-seq.


title: Bisphenol A and Bisphenol S Disruptions of the Mouse Placenta and Potential Effects on the Placenta-Brain Axis

city: Ames

first: Geetu

last: Tuteja

state: IA

address: 2106 Molecular Biology Building

department: Genetics, Development and Cell Biology

institute: Iowa State University

summary: C57BL6J mice were fed 200ug/kg body weight BPA or BPS for two weeks and then bred. Placenta tissue was collected at embryonic day 12.5, and dams received checmicals until this day. RNA-Seq was carried out on placenta tissue, and 13 genes were identified to have altered expression to a similar extent in BPA and BPS treated mice.

overall_design: E12.5 Placenta tissue mRNA profiles of wild-type, BPA and BPS treated mice were generated using Illumina NextSeq 500

first: Ashish

last: Jain

first: Jiude

last: Mao

first: R.

last: Roberts

first: Cheryl

last: Rosenfeld


title: mRNA sequencing of human colorectal cancer organoids

status: Public on Jan 14 2020

city: Jinan

first: Yanjun

last: Li

address: 44 Wenhua West Road

institute: Shandong university

type: Non-coding RNA profiling by high throughput sequencing

summary: Given that colorectal cancer stem cells (CCSCs) play key roles in the tumor dormancy, metastasis and relapse, targeting CCSCs is a promising strategy in cancer therapy. Here, we aimed to identify new regulators of CCSCs and found that Cullin 4B (CUL4B), which possesses oncogenic properties in multiple solid tumors, drives the development and metastasis of colon cancer by sustaining cancer stem-like features. Elevated expression of CUL4B was confirmed in colon tumors and was associated with poor overall survival. Inhibition of CUL4B in cancer cell lines and patient-derived tumor organoids led to reduced sphere formation, proliferation and metastasis capacity. Mechanistically, CUL4B coordinates with PRC2 complex to repress miR34a expression,   thus up-regulates oncogenes including MYCN and NOTCH1, which are targeted by miR34a. Mutation of miR34a binding sites in the 3'UTR of the oncogenes rescues the phenotype caused by CUL4B depletion. Significant correlations were found between expression of CUL4B and miR34a (negatively), miR34a target genes (positively) in clinical samples from colon cancer patients. Collectively, our work demonstrates that CUL4B functions to repress miR34a in maintaining cancer stemness in CRC and provides a potential therapeutic target.

overall_design: We generated transciptome data from human colorectal cancer organoids with knocked-down CUL4B or over-expressed CUL4B

first: Huili

last: Hu

first: Yaoqin

last: Gong


title: CRISPR/Cas9 recombineering-mediated deep mutational scanning of essential genes in Escherichia coli

city: Paris

first: Alaksh

last: Choudhury

address: 16 Rue Henri Huchard

department: IAME

institute: INSERM

summary: Deep mutational scanning can provide significant insights into the function of essential genes in bacteria. Here, we developed a high-throughput method for mutating essential genes of Escherichia coli in their native genetic context. We used Cas9-mediated recombineering to introduce a library of mutations, created by error-prone PCR, within a gene fragment on the genome using a single gRNA pre-validated for high efficiency. Tracking mutation frequency through deep sequencing revealed biases in the position and the number of the introduced mutations. We overcame these biases by increasing the homology arm length and blocking mismatch repair to achieve a mutation efficiency of 85% for non-essential genes and 55% for essential genes. These experiments also improved our understanding of poorly characterized recombineering process using dsDNA donors with single nucleotide changes. Finally, we applied our technology to target rpoB, the beta subunit of RNA polymerase, to study resistance against rifampicin. In a single experiment, we validate multiple biochemical and clinical observations made in the previous decades and provide insights into resistance compensation with the study of double mutants.

overall_design: Development and analysis of genomic mutation libraries for 4 genes galK, crp, mreB and rpoB

first: Olivier

last: Tenaillon

first: Ryan

last: T Gill


title: RNA sequencing of S. simiae CCM 7213T, S. capitis SK14 and S. epidermidis RP62A strains

city: Mutilva

first: Alejandro

last: Toledo-Arana

state: Navarra

address: Avenidad de Pamplona 123

department: IDAB

institute: CSIC

summary: The goal of our study was to perform RNA sequencing of whole transcripts to determine and compare the real length of 3’UTRs among closely-related staphylococcal species. We found that most of the mRNAs encoding orthologous genes in staphylococcal species have 3’UTRs with different lengths in addition to sequence variation.

sample_organism:
Staphylococcus epidermidis
Staphylococcus capitis
Staphylococcus simiae

overall_design: Transcriptome mapping of Staphylococcus species by RNA sequencing


title: Acid ceramidase of macrophages traps herpes simplex virus in multivesicular bodies and protects from severe disease

city: Essen

first: Judith

last: Lang

state: NRW

address: Hufelandstr. 55

department: Institute of Immunology

institute: University of Duisburg-Essen

summary: Macrophages have important protective functions during infection with herpes simplex virus type 1 (HSV-1). However, molecular mechanisms that restrict viral propagation and protect from severe disease are unclear. Here we found that macrophages take up HSV-1 via endocytosis and transport the virions into multivesicular bodies (MVBs). In MVBs, acid ceramidase (aCDase) converts ceramide into sphingosine and increases the formation of sphingosine-rich intraluminal vesicles (ILVs). Once HSV-1 particles pass MVBs, sphingosine-rich ILVs bind to HSV-1 particles, which restricts fusion with the limiting endosomal membrane and prevents cellular infection. Lack of aCDase in macrophage cultures or in Asah1–/– mice results in replication of HSV-1 and Asah1–/– mice died soon after systemic or intravaginal inoculation. The treatment of macrophages with sphingosine enhancing compounds blocks HSV-1 propagation, suggesting a therapeutic potential of this pathway. In conclusion, we demonstrate that aCDase loads ILVs with sphingosine, which prevents HSV-1 capsids from penetrating into the cytosol.

overall_design: 3 to 4 independent experiments were performed using different mice for isolation of macrophages and fibroblasts respectively.

first: Patrick

last: Bohn

first: Hilal

last: Bhat

first: Holger

last: Jastrow

first: Bernd

last: Walkenfort

first: Feyza

last: Cansiz

first: Julian

last: Fink

first: Michael

last: Bauer

first: Dominik

last: Olszewski

first: Ana

last: Ramos-Nascimento

first: Vikas

last: Duhan

first: Sarah-Kim

last: Friedrich

first: Katrin

last: Becker

first: Adalbert

last: Krawczyk

last: Edwards

first: Andreas

last: Burchert

first: Magdalena

last: Huber

first: Justa

last: Friebus-Kardash

first: Joachim

last: Göthert

first: Cornelia

last: Hardt

first: Hans

last: Probst

first: Fabian

last: Schumacher

first: Karl

last: Köhrer

first: Burkhard

last: Kleuser

first: Eduard

middle: B

last: Babiychuk

first: Beate

last: Sodeik

first: Jürgen

last: Seibel

first: Urs

middle: F

last: Greber

first: Philipp

first: Erich

last: Gulbins

middle: S


title: Engineering of effector domains for targeted DNA methylation with reduced off-target effects

city: Stuttgart

first: Albert

last: Jeltsch

address: Allmandring 31

department: Institute of Biochemistry and Technical Biochemistry

institute: University Stuttgart

summary: Epigenome editing is a promising technology, potentially allowing the stable reprogramming of gene expression profiles without alteration of the DNA sequence. Targeted DNA methylation has been successfully documented by many groups for silencing of selected genes, but recent publications have raised concerns regarding its specificity. In the current work, we developed new EpiEditors for programmable DNA methylation in cells with a high efficiency and improved specificity. First, we demonstrated that the dCas9-SunTag scaffold, which has been used earlier for signal amplification, can be combined with the DNMT3A-DNMT3L effector domain allowing a strong methylation at the target genomic locus. We demonstrated that off-target activity of this system is mainly due to untargeted freely diffusing DNMT3A-DNMT3L subunits. Therefore, we generated several DNMT3A variants containing mutations, which reduced their endogenous DNA binding strength. We analyzed the genome-wide DNA methylation of selected variants and confirmed a striking reduction of untargeted methylation. For all potential applications of targeted DNA methylation, the efficiency and specificity of the treatment are the key factors. By developing highly active targeted methylation systems with strongly improved specificity, our work contributes to the future applications of this approach.

overall_design: HEK293 cells were transiently transfected with dCas9-SunTag, antibody-fused catalytic domain of DNMT3A/3L (wild type / DNMT3A mutants) and sgRNA targeting the CpG island of the ISG15 gene. After 3 days, the cells were sorted using FACS to obtain only cells carrying each plasmid. MBD2 pulldown was perfomed with sonicated genomic DNA and methylation changes were analysed genome-wide by NGS.

first: Julian

last: Broche

first: Daniel

last: Hofacker

first: Laura

last: Laistner

first: Sabrina

last: Adam

first: Pavel

last: Bashtrykov


title: Stop Codon Context Influences Genome-Wide Stimulation of Termination Codon Readthrough by Aminoglycosides [Dataset 5]

city: Baltimore

first: Rachel

last: Green

state: MD

address: 725 N Wolfe St

department: Molecular Biology and Genetics

institute: Johns Hopkins University School of Medicine

summary: Stop codon readthrough (SCR) occurs when the ribosome miscodes at a stop codon. Such readthrough events can be therapeutically desirable when a premature termination codon (PTC) is found in a critical gene. To study SCR in vivo in a genome-wide manner, we treated mammalian cells with aminoglycosides and performed ribosome profiling. We find that in addition to stimulating readthrough of PTCs, aminoglycosides stimulate readthrough of normal termination codons (NTCs) genome-wide. Stop codon identity, the nucleotide following the stop codon, and the surrounding mRNA sequence context all influence the likelihood of SCR. In comparison to NTCs, downstream stop codons in 3′UTRs are recognized less efficiently by ribosomes, suggesting that targeting of critical stop codons for readthrough may be achievable without general disruption of translation termination. Finally, we find that G418 treatment globally alters gene expression with substantial effects on translation of histone genes, selenoprotein genes, and S-adenosylmethionine decarboxylase (AMD1).

overall_design: 2 ribosome profiling and 4 RNA-seq samples are included from Calu-6 cells

first: Jamie

last: Wangen


title: Stop Codon Context Influences Genome-Wide Stimulation of Termination Codon Readthrough by Aminoglycosides [Dataset 3]

overall_design: 3 ribosome profiling samples are included from HEK293T cells


title: Stop Codon Context Influences Genome-Wide Stimulation of Termination Codon Readthrough by Aminoglycosides [Dataset 1]

overall_design: 18 ribosome profiling and 8 paired-end RNA-seq samples are included from HEK293T cells


title: Analysis of Polycomb Repressive Complex 2 subunits across cell cycle in pluripotent cells

city: Granada

first: Jordi

last: Martorell-Marugan

address: Avenida de la Ilustración 114

department: Bioinformatics Unit

institute: GENyO

summary: We report ChIP-seq of Polycomb proteins and 4sU-seq data using cell-cycle fractions in mouse embryonic stem cells

overall_design: ChIP-seq samples for Ezh2, Jarid2, EPOP, EloB and Ser5-RNAPII obtained from cell-cycle sorted Fucci-mESCs (G1, S, G2-M fractions). 4sU-seq data in Fucci-mESCs of each cell-cycle sample. ChIP-seq of Ser5-RNAPII and 4sU-seq across cell-cycle in Jarid2 -/- Fucci-mESCs.

first: Helena

last: Asenjo

first: Amador

last: Gallardo

first: Pedro

last: Carmona-Saez

first: David

last: Landeira


title: Stop Codon Context Influences Genome-Wide Stimulation of Termination Codon Readthrough by Aminoglycosides [Dataset 4]

overall_design: 2 ribosome profiling samples are included from HEK293T cells


title: Stop Codon Context Influences Genome-Wide Stimulation of Termination Codon Readthrough by Aminoglycosides [Dataset 2]

overall_design: 4 ribosome profiling samples are included from HEK293T cells


title: Profiling astrocyte spatial heterogeneity and region-specific transcription factor networks

status: Public on Jan 13 2020

city: Houston

first: Brittney

last: Lozzi

state: TX

address: 1 Baylor Plaza

department: Cell & Gene Therapy

institute: Baylor College of Medicine

summary: The goals of this study are to profile the molecular signatures of astrocytes from four brain regions (olfactory bulb, hippocampus, cortex, and brainstem) and determine if differential transcription factor enrichment may play a role in shaping astrocyte spatial diversity. We performed RNA-seq on astrocytes from the olfactory bulb, hippocampus, cortex, and brainstem, and determined region-specific molecular signatures. Using transcription factor motif discovery analysis on each region-specific gene signature we uncovered universal and region-specific transcription factor expression profiles.

overall_design: Aldh1l1-GFP positive astrocytes were collected in triplicate from the olfactory bulb, hippocampus, and brainstem and duplicate from the cortex of 16-week old mice and profiled using RNA-seq

first: Benjamin

last: Deneen


title: Comparision of hepatic gene expression between HFHS diet-fed WT and CREBH-Tg mice

status: Public on Jan 11 2020

city: Tsukuba

first: Yoshimi

last: Nakagawa

state: Ibaraki

department: Internal Medicine (Endocrinology and Metabolism)

institute: University of Tsukuba

summary: To determine CREBH-mediated hepatic gene expression changes in hifh-fat high-sucrose (HFHS) dit feeding, we employed the microarray analysis. We collected the livers from male WT and CREBH-Tg mice fed with HFHS diet for 12 weeks from 6 weeks old.

overall_design: Sample mice include WT (C57B6J) and CREBH Tg mice, which were fed with HFHS fro 12 weeks. CREBH Tg mice overexpress the active form of human CREBH cDNA mainly in the liver.


title: Stop Codon Context Influences Genome-Wide Stimulation of Termination Codon Readthrough by Aminoglycosides

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

overall_design: Refer to individual Series


title: Metabolic rearrangements causing elevated proline and polyhydroxybutyrate accumulation during the osmotic adaptation response of Bacillus megaterium

city: Braunschweig

first: Rebekka

last: Biedendieck

address: Rebenring 56

department: Institute of Microbiology

institute: Technische Universtität Braunschweig

summary: First transcriptome analyses of Bacillus megaterium DSM319 grown under mild and severe salt stress (up to 1.8 M NaCl) to analyse the addaptation of the organism on transcriptome level

overall_design: Bacillus megaterium DSM319 was grown in the presence of 0 (control), 0.6, 1.2 and 1.8 M NaCl, respectively, in Minimalmedium. All cultivations were performed as four biological replicates. Control in each case was B. megaterium DSM319 grown in minimal medium without additional NaCl.

first: Thibault

last: Godard

first: Daniela

last: Zühlke

first: Georg

last: Richter

first: Melanie

last: Wall

first: Katharina

last: Riedel

first: Manfred

last: Rohde

first: Ignacio

last: Poblete-Castro

first: Rainer

last: Krull


title: Whole-Transcriptome Profiling in Developing Tubers of the Jerusalem Artichoke

city: Verona

first: Alberto

last: Ferrarini

state: Veneto

address: Strada le Grazie, 15

department: Scientific and Technological Department

country: Italy

institute: University of Verona

summary: Helianthus tuberosus L., known as the Jerusalem Artichoke, is a hexaploid plant species, adapted to low nutrient soils, that accumulates high levels of inulin in its tubers.  Inulin is a fructose-based polysaccharide used either as dietary fiber or for the production of bioethanol.   Key enzymes involved in inulin biosynthesis are well known. However, the gene networks underpinning tuber development and inulin accumulation in H. tuberous remain elusive. To fill this gap, we selected 6,365 ESTs from a H. tuberosus library to set up a microarray platform and record their expression across three tuber developmental stages, when rhizomes start enlarging (T0), at maximum tuber elongation rate (T3) and at tuber physiological maturity (Tm), in “VR” and “K8-HS142”clones. The former was selected as an early tuberizing, the latter as a late-tuberizing clone.   We quantified inulin and starch levels, and q-RT PCR confirmed the expression of key genes accounting for inulin biosynthesis. The microarray analysis revealed that the differences in morphological and physiological traits between tubers of the two clones are genetically determined since T0 and that is relatively low the number of differentially expressed ESTs across the stages shared between the clones (93). The expression of ESTs for sucrose:sucrose 1-fructosyltransferase (1-SST) and fructan:fructan 1-fructosyltransferase (1-FFT), the two critical genes for fructans polymerization, resulted to be temporarily synchronized and mirror the progress of inulin accumulation and stretching.   The expression of ESTs for starch biosynthesis was insignificant throughout the developmental stages of the clones in line with the negligible level of starch into their mature tubers, where inulin was the dominant polysaccharide.  Overall, our study disclosed candidate genes underpinning the development and storage of carbohydrates in the tubers of two H. tuberosus clones. A model according to which the steady state levels of 1-SST and 1-FFT transcripts are developmentally controlled and might represent a limiting factor for inulin accumulation has been provided.  Our finding may have significant repercussions for breeding clones with improved levels of inulin for food and chemical industry.

overall_design: 18 total samples from 3 growth stages of 2 clones of Helianthus tuberosus L. (Violet de Rennes and K8-HS142) in biological triplicate

first: Marco

last: Bizzarri

first: Massimo

last: Delledonne

first: Paola

last: Tononi

first: Elisa

last: Zago

first: Doriano

last: Vittori

first: Francesco

last: Damiani

last: Paolocci


title: Transcriptional Profiling of Lumbar Spinal Cord in a Rat Model of Bone Cancer Pain

city: Changsha

first: Xinran

last: Hou

state: Hunan

address: Xiangya Road 87

institute: Central South University

summary: Bone cancer pain (BCP), in which the majority is caused by metastasis from other cancer sites, is the most common type of chronic cancer pain. Bone cancer pain is developed accompanied by changes of numerous genes expression, from peripheral to central nervous system, which may account for this dysfunctional nociceptive perception. Although the pivotal role of lncRNA in neuropathic pain are well acknowledged, the involvement of lncRNA in bone cancer pain development are remain to be clarified. Thus, in the present study we performed transcriptome sequencing to explore changes in expression profiles of lncRNA and mRNA to provide a landscape of dysregulated lncRNA and mRNA in spinal cord of bone cancer pain.

overall_design: On the fourteenth day after tumor inoculation, total RNA was extracted from the ipsilateral side of spinal cord of bone-cancer-pained and sham-operated Wistar rats, in triplicates, and high throughput sequencing of ncRNA and mRNA was performed by Illumina HiSeq X Ten.

first: Yingqi

last: Weng

first: Jian

last: Wang

first: Jiajia

last: Dai

first: Zhuofeng

last: Ding

first: Zongbin

last: Song

first: Qulian

last: Guo


title: Suppression of oncogenic super enhancers in MYC-dependent AMLs by NR4A Nuclear Receptors

first: Steven

middle: Gregory

last: Call

address: One Baylor Plaza M509

department: Molecular and Cellular Biology

summary: NR4A nuclear receptors are tumor suppressors of acute myeloid leukemia (AML) that are silenced in AML at the level of transcription elongation. Using RNA-Seq, we show that NR4As inhibit leukemic cell growth largely through suppression of MYC pathway genes. MYC is highly expressed in AML and contributes to AML progression and therapeutic resistance. MYC expression in AML is driven by a super enhancer cluster 1.8Mbp downstream of the MYC locus, which is highly sensitive to therapeutic intervention, including treatment with BET inhibitors. We recently identified dihydroergotamine (DHE) as an FDA-approved drug that strongly induces NR4A transcription in AML cells and operates effectively as an antileukemic therapy. RNA-Seq in MOLM-14 identified MYC as the most statistically repressed target of DHE. Using ChIP-Seq for MED1 as a readout for Mediator occupancy, we identified super enhancers in MOLM-14 cells and show that treatment with DHE significantly suppresses MED1 signal across a subset of super enhancers, including the MYC super enhancer. Finally, we demonstrate that NR4As are broadly associated with global super enhancers, including sites where super enhancer activity is retained, reduced, or gained.

overall_design: For this study, we performed total RNA-Seq in MOLM-14 human AML cells electroporated with in vitro-transcribed RNA expression of GFP or NR4A1, followed by 6 hour incubation. Duplicate samples for each treatment condition (ie. GFP1, GFP2, NR4A1-1, NR4A1-2) are included. We also performed RNA-Seq in MOLM-14 treated with vehicle (DMSO) or 10uM dihydroergotamine (DHE), followed by 6 hour incubation. Duplicate samples for each treatment condition (ie. Veh1, Veh2, DHE1, DHE2) are included. Additionally, we performed ChIP-Seq for MED1 in MOLM-14 cells treated with vehicle (DMSO) or 10uM DHE for 4 hours. Duplicate samples for each treatment condition (ie. MED1 Veh1, Veh2, DHE1, DHE2) are included. Finally, we performed ChIP-Seq for NR4A in MOLM-14 cells treated with 10uM DHE for 4 hours, which is included as a single replicate.

first: S

first: O

last: Conneely


title: Comparision of hepatic gene expression between fasted and fed ad lib states in C57B6J (WT) and CREBH KO mice

first: Yoshimi

last: Nakagawa

summary: To determine hepatic gene expression changes in fasted state, we employed the microarray analysis. We collected the livers from 8-weeks-old male WT and CREBH KO mice in both fasted and fed ad lib states.

overall_design: These mice were fasted for 24 h or fed ad lib. CREBH KO mice is CREBH null mice.


title: Snail and Twist expression in HMLE and MCF10A cells

status: Public on Jan 10 2020

city: Lexington

first: Ren

last: Xu

state: KENTUCKY

address: 741 S. Limestone Rd

institute: University of Kentucky

summary:
Snail and Twist are two EMT inducer, expression of Snail or Twist will induce EMT in HMLE and MCF10A cells. By introducing Snail or Twist in HMLE and MCF10A cells, which lack the expression of these two proteins, will identify the genes are induced during EMT.
We used microarray analysis to compare the gene expression profiles between the mammamry epithleial cells and the cells undergone EMT.

overall_design: Expressing Snail or Twist in HMLE and MCF10A cells, stable clones were selected and RNA were prepared for microarray analysis

first: Binhua

middle: P

last: Zhou