title: Comparative Analysis Of miRNA Profile In Human Dendritic Cells Infected With Respiratory Syncytial Virus And Human Metapneumovirus

status: Public on Dec 10 2018

city: New Orleans

first: Jovanny

last: Zabaleta

state: LA

address: 1700 Tulane Ave

department: Interdisciplinary Oncology

institute: LSUHSC

summary: Background. Human Metapneumovirus (HMPV) and Respiratory Syncytial Virus (RSV), are responsible for respiratory diseases mostly in children. In spite of the resemblance between these two pneumoviruses, they elicit a different extent of immune response. miRNAs are small non coding RNAs that regulate gene expression and are involved in numerous cellular processes including the immune system.  Methodology. Human monocyte-derived dendritic cells (moDC) were differentiated from peripheral blood mononuclear cells and infected at an MOI of 3 for 24h. RNA was isolated to analyze the miRNAs transcription by high throughput sequencing using illumina technology.  Principal findings. Infection with HMPV up-regulated the expression of hsa-miR-4448, while RSV infection induced significant expression of hsa-miR-30a-5p, hsa-miR-182-5p, hsa-miR-1913, hsa37, miR-4448 and hsa-miR-4634.  Conclusions/Significance. In human monocyte derived dendritic cells (moDC), RSV and HMPV induced different profiles of miRNA expression. Understanding the changes of miRNA expression profiles by RSV and HMPV in immune cells will further our understanding of the differential immune response induced by these respiratory viruses

overall_design: miRNA Profile of non-infected and RSV- or HMPV-infected monocyte derived-dendritic cells (moDC)

first: MD

last: Baños-Lara

first: Jone

last: Garai

first: Melody

last: Baddoo

first: Antonieta

last: Guerrero-Plata


title: Expression data from WT and L-PGDS ko mice aorta

city: Philadelphia

first: Emanuela

last: Ricciotti

state: PA

address: 3400 Civic Center Blvd.

department: Department of Pharmacology/Institute for Translational Medicine and Therapeutics

country: USA

institute: University of Pennsylvania

summary: We used microarray data to look for gene differentially expressed in the aorta of  WT and L-PGDS ko male mice.

overall_design: We profiled mouse aorta harvested from male WT and L-PGDS KO mice.

first: Wenliang

last: Song

first: Garret

middle: A

last: FitzGerald


title: INDETERMINATE-DOMAIN 4 (IDD4) coordinates immune responses with plant-growth in Arabidopsis thaliana.

status: Public on Dec 09 2018

city: Thuwal

first: Prof. Heribert

last: HIRT

address: 4700 KAUST

country: Saudi Arabia

institute: KAUST

type: Expression profiling by high throughput sequencing

type: Genome binding/occupancy profiling by high throughput sequencing

summary: We report the transcriptome composition of idd mutant with and without flg22 treatment compared to WT (ColO). Furthermore, we provide transcriptome data of a IDD4ox line and IDD4-phospho-peptide modified versions without treatment compared to WT (ColO), respectively. In order to identify the genomic regions targeted by IDD4 in vivo, Chromatin-Immunoprecipitation (ChIP) was performed followed by deep sequencing on the Illumina High-Sequencing2000 platform (SEQ).

overall_design: Illumina high-sequencing plateform was used to analyse the transcriptome composition of the different genetic backgrounds (under treated and untreated conditions), and to analyse the chromatin fragments bound on IDD4

first: Ronny

last: Voelz

first: Soon-Kap

last: Kim

first: Jianing

last: Mi

first: Alaguraj

last: Veluchamy

first: Kiruthiga

middle: G

last: Mariappan

first: Naganand

last: Rayapuram

first: Jean-Michel

last: Daviere

first: Patrick

last: Achard

first: Salim

last: Al-Babili

first: Moussa

last: Benhamed

first: Heribert

last: Hirt


title: Genome-wide essential gene identification for Shiga toxin-induced cell death using CRISPR/CAS9 screening system

status: Public on Dec 20 2018

city: Shinjuku-ku

first: Tsuyoshi

last: Sekizuka

state: Tokyo

address: 1-23-1 Toyama

department: Pathogen Genomics Center

country: Japan

institute: National Institute of Infectious Diseases

summary: Isolation of genes whose knockout conferring resistance to STx-Induced cell death

overall_design: A genome-wide CRISPR/CAS9 knockout screen in HeLa cells was performed to identify host factors involved in the regulation of Gb3 biosynthesis by exploiting STx sensitivity. Lentivirus-based GeCKO v2 pooled library, which is delivered as two half-libraries (A and B), was used. Two independent sgRNAs-expressing cell libraries (A-1, A-2, B-1, B-2) were prepared by transducing the lentivirus libraries, and cells were then treated with STx1 to assess toxicity. The sgRNAs integrated into the cellular genomes of the surviving cells were amplified by PCR and analyzed with high-throughput sequencing. For untreated controls, cells in each cell library were cultured for the same period as STx1-treated cells with passages several times. sgRNA profiles of control and STx1-treated HeLa cell were generated by deep sequencing, in biological replicate 2, using Illumina MiSeq.

first: Toshiyuki

last: Yamaji

first: Makoto

last: Kuroda


title: Transcriptional profile of Mycobacterium tuberculosis espL deletion mutant

status: Public on Dec 07 2018

city: Bern

first: Andrej

last: Benjak

address: Murtenstrasse 24-28

department: Department for BioMedical Research

country: Switzerland

institute: University of Bern

summary: The ESX-1, type VII, secretion system represents the major virulence determinant of Mycobacterium tuberculosis. The ESX-1 cluster comprises approximately twenty genes and encodes a specialized secretion apparatus, which releases effectors into the extracellular milieu. The goal of this study is to understand the role of EspL. We show that EspL, a protein of 115 amino acids, is essential for mediating ESX-1-dependent virulence and for stabilization of EspE, EspF and EspH protein levels. Indeed, an espL knock-out mutant was unable to replicate intracellularly, secrete ESX-1 substrates or stimulate innate cytokine production. Moreover, loss of EspL also leads to downregulation in M. tuberculosis of WhiB6, a redox-sensitive transcriptional activator of ESX-1 genes.

overall_design: delta espL vs. wild-type

first: Claudia

last: Sala

first: Stewart

middle: T

last: Cole


title: Comparative RNA-seq data on Selaginella moellendorffii organs

status: Public on Dec 14 2018

city: New York

first: Kenneth

middle: David

last: Birnbaum

state: NY

address: 12 Waverly Place

department: Biology

institute: New York University

summary: RNA-seq expression data was generated for root, shoot, rhizophore, and leaf of S. moellendorffii.

overall_design: Analysis of the meristematic region of the three growth centers was compared.

first: K

first: A

last: Mello

first: E

last: Efroni


title: Transcriptome profiling of a beach-adapted wild legume for dissecting novel mechanisms of salinity tolerance

status: Public on Dec 06 2018

city: CHARLOTTE

first: HENGYOU

last: ZHANG

state: NORTH CAROLINA

address: 9201 University City Blvd

department: Biological Sciences

institute: Univ. of North Carolina at Charlotte

summary: Strophostyles helvola is a close relative to common bean (Phaseolus vulgaris) and inhabits both coastal and non-coastal regions in North America. However, the mechanism of saline adaptation in S. helvola remains unclear. A transcriptome profiling would facilitate dissecting the underlying molecular mechanisms in salinity-adapted S. helvola. In this study, we reported the RNAseq analyses of two genotypes (a salt-tolerant beach genotype and a salt-sensitive inland genotype) of S. helvola stressed with salt. S. helvola plants were grown in pots and treated with half lethal-guided dose of NaCl solution for 3h, 24h, and 7d. The plants supplied with the same amount of water were used as controls. The whole roots sampled from the three time points were equally pooled as one biological replicate, and three replicates were used for library construction and transcriptome sequencing on Illumina Hiseq 2500. The comparative analyses of root transcriptomes presented here provides a valuable resource for discovery of genes and networks involved in salt tolerance in S. helvola.

overall_design: This project has been deposited at NCBI BioProject PRJNA483036. Genome-wide expression profiles of Beach/Inland under salt treatment and control conditions, respectively, were generated by deep sequencing, in triplicate, using Illumina Hiseq 2500.

first: H

last: Zhang

first: B

last: Song


title: Lsd1 as a therapeutic target in Gfi1-activated medulloblastoma [microarray]

city: HEIDELBERG

first: Paul

middle: A.

last: Northcott

address: Im Neuenheimer Feld 280

country: Germany

institute: German Cancer Research Center (DKFZ)

type:
Expression profiling by array
Third-party reanalysis

summary: Drugs that modify the epigenome are powerful tools for treating cancer, but these drugs often have pleiotropic effects, and identifying patients who will benefit from them remains a major clinical challenge. Here we show that medulloblastomas driven by the transcription factor Gfi1 are exquisitely dependent on the enzyme lysine demethylase 1 (Kdm1a/Lsd1). We demonstrate that Lsd1 physically associates with Gfi1, and that these proteins cooperate to inhibit genes involved in neuronal commitment and differentiation. We also show that Lsd1 is essential for Gfi1-mediated transformation: Gfi1 proteins that cannot recruit Lsd1 are unable to drive tumorigenesis and genetic ablation of Lsd1 markedly impairs tumor growth in vivo. Finally, pharmacological inhibitors of Lsd1 potently inhibit growth of Gfi1-driven tumors. These studies provide important insight into the mechanisms by which Gfi1 contributes to tumorigenesis, and identify Lsd1 inhibitors as promising therapeutic agents for Gfi1-driven medulloblastoma.

overall_design:
Affymetrix GeneChip Mouse Genome 430 2.0 Array profiling was conducted on n=7 MG (Myc/Gfi1) murine medulloblastomas. This data was compared to the previously published expression array data generated from n=5 NSC (neural stem cell) samples [GSM842157-GSM842161] deposited in GEO Series GSE34126.
MG medulloblastoma tumors were surgically resected, frozen, and subsequently subjected to RNA extraction for downstream Affymetrix expression array profiling.

The complete dataset representing: (1) the MG tumor Samples and (2) the NSCs Samples [GSM842157-GSM842161] from Series GSE34126 (re-processed using RMA), is linked below as a supplementary file.

first: Robert

middle: J

last: Wechsler-Reya


title: Gene expression in WT and Stat6 KO intestinal stem cells

status: Public on Dec 05 2018

city: Beijing

first: Pingping

last: Zhu

state: Beijing

address: Datun Road 15#

country: China

institute: Institute of Biophysics, Chinese Academy of Sciences

summary:
Purpose: To examine the expression profile of WT and Stat6 KO intestinal stem cells.
Methods: We isolated Lgr5+ ISCs from the intestine tissue, and generated Stat6 KO ISCs through CRISPR/Cas9 approach.
Results: Many genes are changed upon Stat6 KO cells. We pay special attention on Wnt/β-catenin signaling that is the most critical pathway in ISCs, and discovered Stat6 drove the expression of Wnt target genes.
Conclusions: Stat6 crosstalks with Wnt signaling to drive ISC self-renewal.

overall_design: Examination of gene expression in WT ISCs and Stat6 KO ISCs.

first: Jiayi

last: Wu

first: Tianku

last: Lu


title: Determination of Pseudomonas aeruginosa transcriptome induced by oleic acid or its derived oxylipins 10-HOME and 7,10-DiHOME.

city: Birmingham

first: Javier

last: Campos-Gomez

state: Alabama

address: 2000 Ninth Ave South

department: Infectious Diseases

institute: Southern Research

summary: Oxygenated unsaturated fatty acids, known as oxylipins, are signaling molecules commonly used for cell-to-cell communication in eukaryotes. However, a role for oxylipins in mediating communication in prokaryotes has not previously been described. Bacteria mainly communicate via quorum sensing (QS) , which involves the production and detection of diverse small molecules termed autoinducers. We showed that oleic acid-derived oxylipins 10-HOME and 7,10-DiHOME produced by Pseudomonas aeruginosa function as autoinducers of a novel quorum sensing system termed Oxylipin-Dependent QS Sytem (ODS). This experiment was designed to determine the genes whose expression is altered by these P. aeruginosa oxylipins. We found that the ODS system controls the cell density-dependent expression of a P. aeruginosa gene subset through the mediation of 10-HOME and 7,10-DiHOME oxylipins.

overall_design: Pseudomonas aeruginosa mRNA profiles of  strain PAO1 ΔPA2076 (ΔodsR ) in the absence or presence of oleic acid or its derived oxylipins 10-HOME and 7,10-DiHOME.

first: Shiva

middle: K

last: Gadila

first: Eriel

last: Martínez

last: Campos-Gómez


title: Rad53- and Chk1-Dependent DNA Damage Response Pathways Cooperatively Promote Fungal Pathogenesis and Modulate Antifungal Drug Susceptibility

status: Public on Dec 04 2018

city: Jeongeup

first: Kwang-Woo

last: Jung

address: 29 Geumgu-gil

country: South Korea

institute: Korea Atomic Energy Research Institute

summary: To elucidate the signaling circuitry downstream of Rad53, we compared transcriptome profiles between wild type and rad53Δ mutant strains post radiation exposure using RNAseq analysis. Of total 6,962 genes, 5,087 and 4,768 genes showed statistically different expression levels in the wild type and rad53Δ mutant strains, respectively, post radiation exposure (P-value cutoff of < 0.05; false decrease rate (FDR) cutoff of < 0.05). In the wild type strain, 392 genes exhibited ≥ 2-fold expression changes post radiation exposure

overall_design: To investigate downstream genes regulated by Rad53, total RNA was isolated from the wild-type (H99) and rad53∆ mutant. Strains were grown in a 40 ml liquid YPD medium for 16 hr at 30℃. Then, overnight culture was inoculated into 100 ml fresh YPD medium and adjusted to OD600=0.2. The cells were further incubated until OD600 reached approximately 0.6. Fifty ml of cells were pelleted for the zero-time sample and the remaining cells were exposed to γ-radiation (0.5 kGy). After γ-radiation exposure, cells were further incubated at 30℃ for 30 mins.

first: Sangyong

last: Lim

first: Yong-Sun

last: Bahn


title: Immunological properties of neural crest cells derived from human induced pluripotent stem cells (normal vs inflammed condition)

status: Public on Dec 01 2018

city: Kobe

first: Shota

last: Fujii

address: 2-2-3 Minatojima-minamimachi, Chuo-ku

department: The Laboratory for Retinal Regeneration

institute: RIKEN Center for Developmental Biology

summary:
Collecting primary neural crest cells (NCCs) of a sufficient quantity for some experiments is difficult, because NCCs are embryonic transient tissue and basically they do not proliferate. We successfully induced NCCs from human induced pluripotent stem cells (iPSCs) according to the previously described method with some modifications.
iPSCs-derived NCCs (iPSC-NCCs) are reported to have potential for wound healing and expected to be used clinically in the future. However, nobody reported the immunological properties of iPSC-NCCs. It is important to evaluate the immunological properties of iPSC-NCCs before clinical use.
In our previous study, we found that iPSC-NCCs have immunosuppressive properties in resting state. In the next step, we examined gene expression patterns between iPSC-NCCs (normal condition) and iPSC-NCCs (inflamed condition).

overall_design: We induced NCCs from human iPSCs according to the previously described method with some modifications for 3 weeks to complete. We examined how the gene expression of iPSC-NCCs changes in inflamed condition.  iPSC-NCCs were incubated with human peripheral blood mononuclear cells from 5 healthy volunteers in transwell culture plate for 5 days (inflamed condition). We extracted total RNA of iPSC-NCCs (normal condition; n=2) and iPSC-NCCs (inflamed condition; n=2).

first: Satoru

last: Yoshida

first: Emi

last: Inagaki

first: Shin

last: Hatou

first: Kazuo

last: Tsubota

first: Masayo

last: Takahashi

first: Shigeto

last: Shimmura

first: Sunao

last: Sugita


title: Immunological properties of neural crest cells derived from human induced pluripotent stem cells (resting state)

summary: Collecting primary neural crest cells (NCCs) of a sufficient quantity for some experiments is difficult, because NCCs are embryonic transient tissue and basically they do not proliferate. We successfully induced NCCs from human induced pluripotent stem cells (iPSCs) according to the previously described method with some modifications.  iPSCs-derived NCCs (iPSC-NCCs) are reported to have potential for wound healing and expected to be used clinically in the future. However, nobody reported the immunological properties of iPSC-NCCs. It is important to evaluate the immunological properties of iPSC-NCCs before clinical use. We examined gene expression patterns between iPSCs and iPSC-NCCs.

overall_design: We induced NCCs from human iPSCs according to the previously described method with some modifications for 3 weeks to complete.We extracted total RNA of iPSCs (n=2) and iPSC-NCCs (n=2).


title: Bacterial diet and weak cadmium stress affect the age-specific survival rates of Caenorhabditis elegans and its resistance against severe stressors

city: Muenster

first: Ruediger

last: Paul

address: Schlossplatz 8

department: Institute of Zoophysiology

institute: University of Muenster

summary: Stressors may have negative or positive effects in dependence of the dose (hormesis). We studied this phenomenon in Caenorhabditis elegans by applying weak or severe abiotic (cadmium, CdCl2) and/or biotic stress (different bacterial diets) during cultivation/breeding of the worms, and determining developmental speed or survival rates and performing transcriptome profiling and RT-qPCR analyses to explore the genetic basis of the detected phenotypic differences. This study showed that a bacterial diet resulting in higher levels of energy resources in the worms (E. coli OP50 feeding) or weak abiotic and biotic stress especially promote the resistance against severe abiotic or biotic stress and the age-specific survival rate of WT.

overall_design: Five experimental conditions; mostly three replicates per experimental condition; four contrasts between test and control conditions functionally analyzed.

first: Ramona

last: Doelling

first: Martha

middle: N

last: Mendelski


title: Effect of cathepsin inhibitors in signals associated with Th1 and Th2 mediators by Dendritic Cells

city: Würzburg

first: Claus

middle: Juergen

last: Scholz

address: Josef-Schneider-Str. 2

department: Core Unit SysMed

institute: University Hospital Würzburg

summary:
Cathepsin  inhibitors have been associated to induce changes in Th1/Th2 polarization; cathepsin B inhibitor CA074 has been reported to induce a Th1 polarization, whereas cathepsin L inhibitor CLIK148 has been reported to induce a Th2 polarization.
In this study, we pre-treated bone marrow-derived dendritic cells (BMDC) with CA074Me or CLIK148, followed by stimulation with a known pro-Th1 inducer (CpG) and a known pro-Th2 inducer (TNF-alpha) and analyzed by microarrays the expression of genes acting as pro-Th1 or pro-Th2 mediators.

overall_design: A total of 7 experimental conditions were used, and the experiments were performed independently twice; thus, a total of 14 samples were analyzed. We included the following controls: BMDC treated with only DMSO (solvent control) with no stimulation, BMDC pre-treated with either CA074Me or CLIK148 without any stimulation, and BMDC pre-treated with DMSO and further stimulated with either CpG or TNF-alpha.

first: Iris

last: Gonzalez-Leal

first: Uta

last: Schurigt

first: Heidrun

last: Moll


title: Acc1 determines memory potential of individual CD4+ T cells by regulating de novo fatty acid biosynthesis [microarray]

status: Public on Nov 30 2018

city: Chiba-city

first: Yusuke

last: ENDO

state: Chiba

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

department: Immunology

institute: Chiba University

summary: Immunological memory is central to adaptive immunity and protection from disease. Changing metabolic demands as antigen-specific T cells transition from effector to memory have been well documented, but the cell-specific pathways and molecules that govern this transition are poorly defined. ACC1 is a rate-limiting enzym in fatty acid biosynthesis. We show that genetic deletion of ACC1 enhanced the formation of CD4+ T cell memory, and that inhibition of ACC1 function enhanced memory T cell formation during parasite infection in mice. ACC1-deficient effector Th cells had similar metabolic signatures to wild-type memory Th cells, and Acaca expression was inversely correlated with a memory gene signature in individual cells. Single cell analysis allowed us to identify a memory precursor-enriched population (CCR7hiCD137lo) present during early differentiation of effector CD4+ T cells. Thus, fatty acid metabolism directs cell fate determination during the generation of memory CD4+ T cells.

overall_design: Phenotypic characterization of TOFA-treated CD4+ T cell population.

last: Endo

first: Atsushi

last: Onodera

first: Kazushige

last: Obata-Ninomiya

first: Ryo

last: Koyama-Nasu

first: Hikari

last: Asou

first: Toshihiro

last: Ito

first: Takeshi

last: Yamamoto

first: Toshio

last: Kanno

first: Takahiro

last: Nakajima

first: Kenji

last: Ishiwata

first: Hirotaka

last: Kanuka

first: Damon

last: Tumes

first: Toshinori

last: Nakayama


title: Aging alters the epigenetic asymmetry of HSC division [scATAC-Seq]

city: Ulm

first: Medhanie

middle: Assmelash

last: Mulaw

address: Albert-Einstein-Allee 11

institute: University of Ulm

summary: Hematopoietic stem cells (HSCs) balance self-renewal and differentiation to maintain homeostasis. With aging, the  frequency of polar HSCs decreases. Cell polarity in HSCs is controlled by the activity of the small RhoGTPase Cdc42. Here  we demonstrate, using a comprehensive set of paired daughter cell analyses that include single cell 3D-confocal imaging,  single cell transplants, single cell RNA-seq as well as single cell ATAC-seq, that the outcome of HSC divisions is strongly  linked to the polarity status before mitosis, which is in turn determined by the level of the activity Cdc42 in stem cells.  Aged apolar HSCs undergo preferentially self-renewing symmetric divisions, resulting in daughter stem cells with  reduced regenerative capacity and lymphoid potential, while young polar HSCs undergo preferentially asymmetric  divisions. Mathematical modeling in combination with experimental data implies a mechanistic role of the asymmetric  sorting of Cdc42 in determining the potential of daughter cells via epigenetic mechanisms. Therefore, molecules that  control HSC polarity might serve as modulators of the mode of stem cell division regulating the potential of daughter  cells.

overall_design: Sorted single cells were cultured with and without treatment in the presence of cytokines until first cell division (40-44hrs). The daughter cells were manually separated, washed with PBS and collected for ATAC sequencing.


title: Shortening the half-life of Cas9 maintains its gene editing ability and reduces neuronal toxicity

city: Atlanta

first: Gregory

last: Tharp

state: GA

address: 954 Gatewood Dr

department: Developmental and Cognitive Neuroscience

institute: Yerkes National Primate Research Center

summary: Virus-mediated expression of CRISPR/Cas9 has been actively used for genome editing in animal models of genetic disorders, including neurological diseases, but the consequences of overexpressing bacterial Cas9 in the mammalian brain remain unknown. Through RNA-seq analysis, we found that virus-mediated expression of Cas9 caused systematic changes in genes involved in neuronal functions. We also generated a short-lived version of Cas9, which maintains its genome editing capacity, but significantly alleviates neurotoxicity caused by overexpressed Cas9. Thus, modification of Cas9 by shortening its half-life would help develop CRISPR/Cas9-based therapeutic approaches for treating genetic neurological disorders.

overall_design: Wild type mice were injected with adeno-associated virus into the striatum. Three weeks after viral injection, the striatal tissues were collected, and gene expression profiles were analyzed by RNA-seq.

first: Su

last: Yang

first: Shihua

last: Li

first: Xiao-Jiang

first: James

middle: M

last: Billingsley


title: Role of ProQ in Salmonella virulence - pulse expression

status: Public on Nov 28 2018

city: Cologne

first: Konrad

middle: U.

last: Förstner

state: North Rhine-Westphalia

address: Gleueler Str. 60

department: Information Services

institute: ZB MED - Information Centre for Life Sciences

summary: FinO-domain proteins such as ProQ of the model pathogen Salmonella enterica have emerged as a new class of major RNA-binding proteins in bacteria. ProQ has been shown to target hundreds of transcripts including mRNAs from many virulence regions but its role, if any, in bacterial pathogenesis has not been studied. Here, using a Dual RNA-seq approach to profile ProQ-dependent gene expression changes as Salmonella infects human cells, we reveal dysregulation of bacterial motility, chemotaxis and virulence genes which is accompanied by altered mitogen-activated protein kinase signaling in the host. Comparison with Hfq, Salmonella’s other major RNA chaperone, reinforces the notion that these two global RNA binding proteins work in parallel to ensure full virulence. Of newly discovered infection-associated ProQ-bound small noncoding RNAs, we show that STnc540 represses a virulence-related magnesium transporter mRNA in a ProQ-dependent manner. Together, this comprehensive study uncovers the relevance of ProQ for Salmonella infection and highlights the importance of RNA-binding proteins in regulating bacterial virulence programs.

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

overall_design: Bacterial RNA-seq

first: Alexander

last: Westermann

first: Elisa

last: Venturini

first: Mikael

middle: E

last: Sellin

middle: U

first: Wolf-Dietrich

last: Hardt

first: Jörg

last: Vogel


title: Next Generation Sequencing Facilitates Quantitative Analysis of Control and Oct3 Deficient Adipose Transcriptomes

first: wenxin

last: song

address: Hai Dian, Beijing

institute: Tsinghua university

summary: Analysis of Oct3-deficient inguinal and control white adipose tissue from 8-week-old C57BL/6 animals after 4 ºC for one month. Results provide insight into the role of Oct3 in WAT beiging.

overall_design: mRNA profiles of mice were generated by deep sequencing using BGISEQ-500.

first: Wenxin

last: Song

first: Zhilong

last: Ma


title: Molecular basis of mitomycin C enhanced corneal sensory nerve repair

city: Bethesda

first: Maria

middle: Irene

last: Morasso

state: Maryland

address: Bldg 50

institute: NIAMS NIH

summary: We performed RNASeq and confocal imaging analysis to characterize the gene expression changes in the cornea after wounding that modulate sensory axon integrity. We show that by preserving the ability of axons to attach to one another and to corneal epithelial cells, mitomycin C enhances sensory nerve recovery after mechanical injury.

overall_design: Epithelium and stroma from mouse corneas with/without Mitomycin C treatment and wounding were examined, 4 variables were tested (Control, Mitomycin C Control, Wound, Mitomycin C Wound).

middle: I

first: Stephen

middle: R

last: Brooks

first: Mary

last: Stepp


title: HSFA2 functions in the physiological adaptation of undifferentiated plant cells to spaceflight microgravity environment.

city: Gainesville

first: Robert

middle: J.

last: Ferl

state: Florida

address: 1301 Fifield Hall PO Box 110690

department: Horticultural Sciences

institute: University of Florida

summary:
The Heat Shock Factor A2 (HsfA2), as a part of the HSF network, is essential to the plant’s response to almost any environmental stress and to the cellular homeostatic control mechanisms. Plant cell cultures disabled in HsfA2 function were grown aboard the International Space Station (ISS) in order to ascertain whether or not they use the same terrestrially effective systems to adapt to the novel environment of spaceflight. Cultured lines of Arabidopsis thaliana derived from wild type (WT) cultivar Col-0 and from a knock-out line deficient in the gene encoding HSFA2 (HSFA2 KO) were launched to the ISS on SpaceX-2 as part of the Cellular Expression Logic (CEL) experiment of the BRIC17 spaceflight mission and were fixed in-flight after 10 days on orbit. Microarray gene expression data were analyzed using a two-part comparative approach. First, differentially expressed genes were identified between the environments (spaceflight to ground) within cells of the same genotype, which represented physiological adaptation to the spaceflight environment. Second, gene expression profiles were identified between the genotypes (HSFA2 KO to WT) within the same environment, defining genes uniquely required by the two genotypes in the ground and spaceflight adapted states.
The physiological state of the cells as a result of disabling a gene has tremendous control over the mechanisms induced to adapt to the environment of spaceflight.  The HsfA2 demonstrated a role in the physiological adaptation to the spaceflight environment since the cells disabled in the HsfA2 gene used substantially different genes to achieve the spaceflight adapted state than the WT cells. The endoplasmic reticulum (ER) stress and unfolded protein response (UPR) define the HSFA2 KO cells’ physiological state regardless of the environment and likely result from the deficiency in the chaperone-mediated protein folding machinery. HsfA2 seems to have a universal stress response role but also specific roles in the physiological adaptation to spaceflight through cell wall remodeling, signal perception and transduction and starch biosynthesis. Implementation of knock-out cells identified a set of genes with a required expression level in order for a cell to achieve a spaceflight-adapted state. The HSFA2 KO cells helped to unravel the HsfA2-dependent genes of the adaption of wild type cells to the environment of spaceflight.

overall_design: The CEL (Cellular Expression Logic) experiment setup and organization was a modification of a previous Arabidopsis cell culture experiment in BRIC16 (Paul, Zupanska et al. 2012). The CEL BRIC17 experiment was launched on board the Dragon capsule of SpaceX-3 CRS mission to the International Space Station (ISS) on the 1st of March 2013.  The cultured cell lines (both the ground control and the spaceflight samples) were grown within 60mm Petri plates in Petri Dish Fixation Units (PDFUs) that were housed within the Biological Research In Canister (BRIC) hardware. Two BRIC containers (A and B) were assigned to CEL within the BRIC17 payload. Each chamber housed 5 PDFUs, each PDFU holding one 60mm Petri plate. In each BRIC container there were 2 plates with wild type (WT) cells and 3 plates with knock-out cells of two genotypes. The exact same PDFU composition was recapitulated in BRICs on the ground in the ISS Environmental Simulator (ISSES) chamber at Kennedy Space Center (KSC) as Ground Controls. The Ground Controls were initiated with a 48 hrs delay so that the precise temperature environment of the ISS could be recreated for the ground controls in the ISSES chamber. Cells were fixed in-flight in RNAlater on the 10th day on orbit, and the ground controls were fixed 48 hours later. RNAlater fixation was initiated by the crew using an activation tool that moves RNAlater from a storage container in the PDFU into the Petri Plate. Twenty four hours after fixation, the entire BRIC was moved to the MELFI (Minus Eighty-degree Laboratory Freezer for ISS) where it resided until cold stowage transport back to Earth within the Dragon capsule.  After returning to Earth the samples were reclaimed at KSC, and then transported to the University of Florida (UF) laboratories. The total RNA was extracted from spaceflight samples and corresponding Ground Control samples and subjected to microarrays.

first: Agata

last: Zupanska

first: Collin

last: LeFrois

first: Anna-Lisa

last: Paul


title: Transcriptome analysis in hippocampus of DBA/2 mice under different doses of Venlafaxine

first: Claus

last: Scholz

summary: Molecular effectiveness of antidepressants is a still not sufficiently understood process. The hippocampus as a brain region controlling formation of memory and learning processes as well as emotional evaluation during memory retrieval is of special interest in this reagard. We compared gene expression profiles in hippocampi of male DBA/2 mice after one month of different daily doses of the sSNRI antidepressant Venlafaxine to determine genes regulated under the chronic influence of this drug to gain insight in potential molecular changes that might reflect transformation from pathological to physiological condition. Additionally to basic research this might provide new targets of pharmacological interventions for depression.

overall_design: To identify genes that are differentially expressed in the hippocampus under different dosages of the sSNRI Venlafaxine male DBA/2 mice perorally received 0, 30 respectively 100 mg of Venlafaxine per day for 30 days. RNA from hippocampus was extracted and analyzed with the Gene Chip Mouse Genome 430 2.0 (Affymetrix) after pooling of cDNA depending on laterality within dosage groups. Data from ten animals per group was independent of hippocampus laterality pooled within dosage groups.

first: Florian

last: Proft

first: Heike

last: Weber

first: Jürgen

last: Deckert

first: Andreas

last: Reif


title: Induction of muscle stem cell quiescence by the secreted niche factor Oncostatin M

city: San Diego

first: John

last: Walker

state: CA

address: 10675 John Jay Hopkins

institute: Genomics Institute of the Novartis Research Foundation

summary:
Regeneration of skeletal muscle is dependent on the function of tissue-resident muscle stem cells (MuSC), known as satellite cells. MuSC dysfunction is central to muscle pathophysiology, including in age-associated loss of muscle regenerative capacity and congenital disorders such as Duchenne muscular dystrophy. Despite the central role of satellite cells in muscle regeneration, the signals controlling the balance between muscle stem cell quiescence, proliferation, and differentiation remain incompletely understood. Knowledge of the signals that maintain a quiescent state is particularly lacking, yet such cues are crucial to maintaining a stem cell reservoir that can meet the needs of regeneration throughout life. Here we identify Oncostatin M (OSM), a member of the interleukin-6 family of cytokines, as a potent and essential trans-acting regulator of satellite cell quiescence. Key to this discovery is the development of a novel in vivo imaging-based screening strategy allowing identification of proteins that do not induce in vitro proliferation, but instead maintain MuSCs in a non-mitotic state, poised for rapid robust expansion upon transplantation.  We demonstrate that OSM induces reversible exit from the cell cycle and induction of a global transcriptional program significantly enriched within a newly established satellite cell “quiescence signature”.  Genetic ablation of the OSM receptor in mice demonstrates that signaling via OSM/R is essential for maintenance of satellite cell quiescence, and for proper skeletal muscle regeneration in vivo.  Given that aberrant activation and exhaustion of stem cells is seen in a variety of disorders, OSM constitutes an attractive therapeutic target in muscle disease states.
Microarray profiling was used to directly test the hypothesis that OSMR signaling in muscle stem cells promotes their engraftment by inducing a global transcriptional state equivalent to that of quiescent, freshly isolated, non-activated MuSC.

overall_design: Comparison of Oncostatin M-treated cultured muscle stem cells with either fresh or cultured but non-Oncostatin M-treated muscle stem cells

first: Srinath

middle: C

last: Sampath

first: Srihari

first: Andrew

last: Ho

first: Stéphane

middle: Y

last: Corbel

first: Joshua

last: Millstone

last: Lamb

first: Bernd

last: Kinzel

first: Christian

last: Schmedt

first: Helen

last: Blau


title: Microarray data from striatum and prefrontal cortex of ethanol light drinkers and ethanol inflexible drinkers

city: Belo Horizonte

first: Ana Lúcia

last: Brunialti Godard

state: Minas Gerais

address: Avenida Antonio Carlos, 6627

department: Biologia Geral

country: Brazil

institute: UFMG

summary:
Transcriptional study to  investigate differences in prefrontal cortex and striatum of mice that consumed ethanol despite negative consequences, which resembles the compulsive aspect of alcohol addiction.
The transcriptional analysis performed in the striatum and prefrontal cortex revealed genes and biological pathways differentially regulated specifically in animals of the group Inflexible Drinkers that could be involved with the loss of control over voluntary ethanol consumption (da Silva E Silva et al., 2016; de Paiva Lima et al., 2017).

overall_design: In the animal model used, mice were given continuously access to three bottles containing 5 and 10% of ethanol (v/v) and water for 16 weeks. On the last two weeks of the experiment both ethanol solutions were adulterated with the bitter and aversive substance, quinine. Following this procedure, mice were classified in three groups according to their individual ethanol intake and preference as follows: “Light Drinkers” (preference for water at all experimental stages); “Heavy Drinkers” (ethanol preference decreased after adulteration of the ethanol solution with quinine), and “Inflexible Drinkers” (preference for ethanol at all experimental stages, even after addition of quinine to the ethanol solution) (Ribeiro et al., 2012)

first: Daniel

last: Almeida da Silva e Silva

first: Andrea

last: Frozino Ribeiro

first: Samara

last: Damasceno

first: Cristiane

middle: S

last: Rocha

first: Alexandre

middle: H

last: Berenguer de Matos

first: Roseli

last: Boerngen-Lacerda

first: Diego

last: Correia

first: Ana

last: Lúcia Brunialti Godard


title: lncRNA and mRNA Expression Profiling in Early Stage UM CLL

status: Public on Nov 29 2018

city: Rochester

first: Renee

last: Tschumper

state: MN

address: 200 1st St SW

department: Immunology Research

institute: Mayo Clinic

type:
Expression profiling by array
Non-coding RNA profiling by array

summary: Predicting disease progression in chronic lymphocytic leukemia (CLL) remains challenging particularly in patients with Rai Stage 0/I disease that have an unmutated immunoglobulin heavy chain variable region (UM IGHV). Even though patients with UM IGHV have a poor prognosis and generally require earlier treatment, not all UM IGHV patients experience more rapid disease progression with some remaining treatment free for many years. This observation suggests biologic characteristics other than known prognostic factors influence disease progression. Alterations in long non-coding RNA (lncRNA) expression levels have been implicated in diagnosis and prognosis of various cancers, however, their role in disease progression of early Rai stage UM CLL is unknown. Here we use microarray analysis to compare lncRNA and mRNA profiles of Rai 0/I UM IGHV patients who progressed in <2 years relative to patients who had not progressed for >5 years. Over 1300 lncRNAs and 940 mRNAs were differentially expressed (fold change ≥ 2.0; p-value ≤ 0.05). Of interest, the differentially expressed lncRNAs G047155, TCAM1P, and uc.436, have known associated genes that have been linked to CLL. Thus, our study reveals differentially expressed lncRNAs in progressive early stage CLL requiring therapy versus indolent early Rai stage UM CLL. These lncRNAs have the potential to impact relevant biological processes and pathways that influence clinical outcome in CLL.

overall_design: Two-condition experiment, Progressive disease (PD) early Rai (0/I) Stage UM CLL (n=34) and Indolent disease (ID) early Rai (0/I) Stage UM CLL (n=29).  One replicate per array.

first: Tait

middle: D

last: Shanafelt

first: Neil

last: Kay

first: Diane

middle: F

last: Jelinek


title: TraSH analysis of Mycobacterium tuberculosis transposon mutants with reduced bodipy-palmitate uptake on the third day of resting bone marrow-derived macrophage infection

city: Ithaca

first: Evgeniya

last: Nazarova

state: NEW YORK

address: C5 103 VMC Cornell University

department: Microbiology & Immunology

institute: Cornell University

summary: Mycobacterium tuberculosis transposon mutants with reduced bodipy-palmitate intake on the third day of resting bone marrow-derived macrophage infection are identified by TraSH. Discovery of genetic requirements for fatty acid assimilation by the pathogen is essential for understanding its metabolism during host infection.

sample_organism: Mycobacterium tuberculosis str. Erdman = ATCC 35801

overall_design: The screen was performed with transposon (Himar) mutant library generated in M. tuberculosis Erdman strain pDE43-MCH (smyc'::mCherry). Bone marrow-derived macrophages were infected with this transposon library (~2X104) at MOI 4:1, metabolically labeled with Bodipy-palmitate on the third day of infection. After isolation mutants with low Bodipy signal were sorted, plated, and pooled for DNA extraction. This selection was repeated for enrichment. TraSH analysis is performed to compare pools of mutants after 1st and 2nd rounds of selection relative to the input pool (Input). 3 technical replicates were used for each of the rounds of enrichment.

middle: V

first: Thuy

last: La

first: Brian

last: VanderVen

first: David

middle: G

last: Russell


title: circular RNA (circRNA) expression profile in intestinal stem cells

first: Pingping

last: Zhu

address: Datun Road 15#

institute: Institute of Biophysics, Chinese Academy of Sciences

type: Non-coding RNA profiling by high throughput sequencing

summary: Purpose: circRNAs emerge as critical modulators in various biogical processes, but their roles in intestinal stem cells (ISC) reamin unclear. Here we want to identify a functional circRNA in ISCs, and firstly, we explore the expression profile of circRNA in ISCs. Methods: We isolated Lgr5+ ISCs and Lgr5- non-ISCs from the intestine tissue, followed by circRNA seq. Results: Many circRNAs are differently expressed in ISCs and non-ISCs, and we selected circRNAs highly expressed in ISCs for further investigation. Conclusions: circRNAs may play a critical role in ISC self-renewal.

overall_design: Examination of circRNA expression in 2 ISCs and non-ISCs.

first: Zusen

last: Fan


title: Acc1 determines memory potential of individual CD4+ T cells by regulating de novo fatty acid biosynthesis

first: Yusuke

last: ENDO

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

overall_design: Refer to individual Series


title: Cxcr3 expressing leukocytes are necessary for neurofibroma formation in mice

city: Cincinnati

first: Nancy

last: Ratner

state: OH

address: 3333 Burnet Ave.

institute: Cincinnati Children's Hospital Medical Center

summary: Plexiform neurofibroma is a major contributor to morbidity in Neurofibromatosis type I (NF1) patients. Macrophages and mast cells infiltrate neurofibroma, and data from mouse models implicate these leukocytes in neurofibroma development. Anti-inflammatory therapy targeting these cell populations has been suggested as a means to prevent neurofibroma development. Here, we compare gene expression in inflamed nerves from NF1 models which invariably form neurofibroma to those with inflammation driven by EGFR overexpression which rarely progresses to neurofibroma. We find that the chemokine Cxcl10 is uniquely up-regulated in NF1 mice that invariably develop neurofibroma. Global deletion of the Cxcl10 receptor Cxcr3 prevented neurofibroma development in these neurofibroma-prone mice. Cxcr3 expression localized to T cells and dendritic cells (DCs) in both inflamed nerves and neurofibromas. These data support a heretofore unappreciated role for T cells/DCs in neurofibroma initiation.

overall_design: 23 microarrays: Mouse reference samples (4), mouse P30 samples (9), mouse P60 samples (10). Controls: P30 Nf1 fl/fl samples.

first: Nathan

last: Salomonis

first: Walter

middle: J

last: Jessen


title: expression analysis in melanomas and melanocytes from the same patient

first: Susumu

last: Fujiwara

state: Hyogo

address: 7-5-1, Kusunoki-cho, Chuo-ku

department: dermatology

institute: Kobe univ

summary:
DNA methylation is considered the primary epigenetic mechanism underlying the development of malignant melanoma. Since DNA methylation can be influenced by environmental factors, it is preferable to compare cancer and normal cells from the same patient. Here, we employed a novel epidermal sheet cultivation technique to isolate normal melanocytes from unaffected sites of melanoma patients and compared the methylation status with melanoma tissues from the same individuals. We also analyzed primary and metastatic melanoma samples, three commercially available melanocytes, and four melanoma cell lines. Cluster analysis of DNA methylation data classified freshly isolated melanomas and melanocytes into the same group, whereas the four melanoma cell lines were clustered together in a distant clade. Moreover, our analysis discovered methylation at several novel loci (KRTCAP3, AGAP2, ZNF490), in addition to those identified in previous studies (COL1A2, GPX3); however, the latter two were not observed in fresh melanoma samples. Subsequent studies revealed that NPM2 was hypermethylated and downregulated in melanomas, which was consistent with previous reports and indicated that NPM2 immunoreactivity could be used to differentiate melanomas from normal melanocytes or benign disease. Our results highlight the importance of using matched fresh melanoma and melanocyte samples in epigenetic studies.
illumina HumanHT-12_v4 BeadChip

overall_design: total 10 samples, 5 melanocytes and 5 melanomas. Freshly isolated 2 pair of melanoma and melanocyte from the same patient, 3 commercial melanoma cell lines and 3 commercial melanocyte cell lines.

first: Hiroshi

last: Nagai

first: Haruki

last: Jimbo

first: Naoe

first: Tomoyo

last: Tanaka

first: Masukazu

last: Inoie

first: Chikako

last: Nishigori


title: Role of ProQ in Salmonella virulence - in vitro

first: Konrad

last: Förstner

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

first: Alexander

last: Westermann

first: Elisa

last: Venturini

first: Mikael

last: Sellin

first: Wolf-Dietrich

last: Hardt

first: Jörg

last: Vogel


title: Modelling pancreatic beta-cell inflammation in zebrafish identifies a natural product for human beta-cell protection

city: Dresden

first: Luis

middle: Fernando

last: Delgadillo Silva

address: Fetscherstraße 105

institute: CRTD

summary:
To assess the effect of chronic inflammation on the beta-cells transcriptome, we conducted RNA-sequencing, from 3 months old zebrafish transgenic line Tg(ins:IL1B) animals and WT siblings
The total RNA from FACS sorted Beta-cells was isolated using Quick-RNA MicroPrep kit (R1050 Zymo Research) and following the manufacturer instructions. The sequencing was made on llumina HiSeq2500 in 2x75bp paired-end mode.  method
Results:Two replicates for controls and two for IL1B expressing beta cells were sequenced. About 40 million sequence reads per sample were mapped to the Zebrafish genome. We identify around 19 thousand genes per sample. Approximately 6% of the genes were differentially expressed. 1,245 genes with a FDR of 5 showed a fold change ≥0.7 and adjusted p-value <0.05.
Conclusions: Hierarchical clustering by Pearson correlation of differentially expressed genes uncovered the expression of alpha-cell related genes in beta-cells. This provides a possible impairment of beta-cell identity reflected in RNA expression profiles.

overall_design: Beta-cell mRNA profiles of 3 month old wild type (WT) and Tg(ins:IL1B; cryaa:mCherry) Zebrafihs, were generated by deep sequencing, in duplicate, using llumina HiSeq2500.

last: Delgadillo-Silva

first: Nadeem

last: Akhtar

first: Nikolay

last: Ninov


title: Role of ProQ in Salmonella virulence

middle: U.


title: Role of ProQ in Salmonella virulence - dual RNA-Seq

overall_design: Dual RNA-seq or bacterial RNA-seq

middle: E


title: Mammary Tissue: aCGH of primary (IGF-IR induced) and recurrent (IGF-IR independent) mammary tumors from MTB-IGFIR mice

city: Guelph

first: Roger

last: Moorehead

address: 50 Stone Rd E

department: Biomedical Sciences

country: Canada

institute: University of Guelph

type: Genome variation profiling by genome tiling array

summary: Tumor recurrence represents a significant clinical challenge in the treatment and management of breast cancer. To investigate whether copy number aberrations (CNAs) facilitate the re-emergence of tumor growth from residual disease we performed array comparative genomic hybridization (aCGH) on primary and recurrent mammary tumors from an inducible mouse model of type-I insulin-like growth factor receptor (IGF-IR) driven breast cancer. This genome-wide analysis revealed primary and recurrent tumors harbored distinct copy number aberrations (CNAs) with relapsed tumors containing an increased number of gene-level gains and losses. Remarkably, CNAs detected in primary tumors were largely devoid of annotated cancer genes while the vast majority of recurrent tumors harbored at least one CNA containing a known oncogene or tumor suppressor. Specifically, a subset of recurrent tumors carried gains at 6qA2 and 9qA2 which encode the Met and Yap1 oncogenes respectively. The most frequent CNA detected was a focal deletion at 4qC5 involving the Cdkn2a and Cdkn2b tumor suppressor genes. Integrative analysis revealed positive correlations between gene copy number and mRNA expression suggesting Met, Yap1 and Cdkn2a/b may serve as potential driver genes that promote tumor recurrence. Together, these findings indicate that tumor recurrence is facilitated by the acquisition of CNAs with oncogenic potential and provide a framework to dissect the molecular mechanisms that mediate tumor escape from dormancy.

overall_design: Two-color common reference design with 11 primary tumors (doxycycline-dependent) and 8 recurrent tumors (doxycycline-independent) from MTB-IGFIR mice.

first: Robert

last: Jones


title: Expression data from testis in mutant mouse lacking retinoic acid synthesizing enzymes in germ cells

city: ILLKIRCH CEDEX

first: Norbert

middle: B.

last: GHYSELINCK

address: 4 rue Laurent Fries

department: Functional Genomics and Cancer

country: France

institute: Institut de Genetique et Biologie Moleculaire et Cellulaire (IGBMC)

summary:
In mammals, all-trans retinoic acid (ATRA) is instrumental to spermatogenesis. It is synthesized by two retinaldehyde dehydrogenases (RALDH) present notably in germ cells (GC). In order to determine the contributions of the GC-derived source of ATRA, we have generated mice lacking all RALDH activities in GC.
We show that the GC-derived source of ATRA does not perform any specific role despite contributing to two-third of the total amount of ATRA present in the testis.

overall_design: Mice carrying loxP-flanked alleles (L2) of Aldh1a1(tm1Nbg), Aldh1a2 and Aldh1a3(tm1Pcn) were crossed with mice bearing the Tg(Stra8-cre)1Reb transgene. Testis were sampled for analysis at the age of 9 weeks.

first: Manuel

last: Mark

middle: B

last: Ghyselinck


title: Analysis of long non-coding RNA and mRNA expression in bovine macrophages brings up novel aspects of Mycobacterium avium subspecies paratuberculosis infections

status: Public on Nov 27 2018

city: Berlin

first: Jennifer

last: zur Brügge

address: Oertzenweg 19b

department: Veterinary Medicine

institute: Freie Universität Berlin

summary: The alteration in genomic expression profiles of mRNA and long non-coding RNA (lncRNA) in bovine macrophages in response to Mycobacterium avium subspecies paratuberculosis (MAP) infection was investigated by applying an RNA-Seq based approach. A computational pipeline was implemented which identified more than 300 unannotated lncRNAs in macrophages of which 38 were differentially regulated by the infection.

overall_design: Primary bovine macrophages were infected with MAP, total RNA was sequenced and analyzed in terms of mRNA and lncRNA expression

first: Pooja

last: Gupta

last: zurBruegge


title: Mutations in NCAPG2 cause a severe neurodevelopmental syndrome that expands the phenotypic spectrum of condensinopathies

city: Chicago

first: Erica

last: Davis

state: IL

address: 225 E Chicago Avenue

department: Advanced Center for Translational and Genetic Medicine (ACT-GeM)

institute: Ann & Robert H Lurie Children's Hospital of Chicago

summary: The use of whole exome/genome sequencing has been a catalyst for a genotype-first approach to diagnostics. Under this paradigm, we have implemented systematic sequencing of neonates and young children with a suspected genetic disorder. Here we report two families with recessive mutations in NCAPG2 and overlapping clinical phenotypes that include severe neurodevelopmental defects; failure to thrive; ocular abnormalities; and defects in urogenital and limb morphogenesis. NCAPG2 encodes a member of the condensin II complex, necessary for the condensation of chromosomes prior to cell division. Consistent with a causal role for NCAPG2, we found abnormal chromosome condensation, augmented anaphase chromatin bridge formation and micronuclei in daughter cells of proband skin fibroblasts. To test the functional relevance of the discovered variants, we generated an ncapg2 zebrafish model. Morphants displayed clinically-relevant phenotypes, such as microcephaly and renal anomalies with concomitant increases in apoptosis and altered mitotic progression. These could be rescued by wild type but not mutant human NCAPG2 mRNA and were recapitulated in CRISPR/Cas9 F0 mutants. Finally, we noted that the individual with a complex urogenital defect also harbored a heterozygous deletion of NPHP1, a common contributor to nephronophthisis. To test whether sensitization at the NPHP1 locus might contribute to a more severe renal phenotype, we co-suppressed nphp1 and ncapg2, which resulted in significantly more dysplastic renal tubules. Together, our data suggest that impaired function of NCAPG2 results in a severe condensinopathy and highlight the potential utility of examining candidate pathogenic lesions beyond the primary disease locus.

overall_design: To investigate the global impact of NCAPG2 loss in an in vivo developmental context, we performed transcriptome profiling of zebrafish embryo heads isolated at 2 dpf (injected with gRNA1 alone or gRNA1+Cas9; 5 biological replicates)

first: Nicholas

last: Katsanis


title: Novel MVA Vector Expressing Anti-apoptotic gene, B13R, Delays Apoptosis and Enhances Humoral Responses

first: Gregory

middle: K

last: Tharp

address: 954 Gatewood Dr

institute: Yerkes National Primate Research Center

summary: Modified vaccinia virus Ankara (MVA) has been explored as a vaccine vector for use against infectious diseases and cancer. MVA is an immunogenic, attenuated poxvirus capable of eliciting robust cellular and humoral responses in pre-clinical animal models and in patients. However, upon infection with MVA, cells undergo rapid apoptosis leading to faster clearance of recombinant antigens. The fragmentation of the anti-apoptotic gene B13R in MVA could contribute to this effect. Here, we replaced the fragmented B13R with a functional version and observed that MVA-B13R infected HeLa cells and muscle cell lines delayed caspase 3 activation compared to MVA indicating slower progression of apoptosis. For immunogenicity studies, mice were intramuscularly immunized with recombinant MVA or MVA-B13R expressing SIV Gag, Pol and HIV Env (SHIV). We observed higher Env-specific humoral responses from MVA-B13R SHIV compared to MVA SHIV mice. To determine differences in the innate immune response that may have contributed to the augmented humoral response, we performed RNA-Seq analysis on draining lymph node cells after immunization. Gene set enrichment analysis from day 1 after immunization showed that MVA-B13R SHIV immunizations were associated with a negative enrichment for type I and II interferon responses compared to MVA SHIV mice indicating MVA-B13R SHIV induces a delayed anti-viral interferon response that may lead to the enhanced humoral response observed. Taken together, these results demonstrate that restoring B13R functionality in MVA significantly delays MVA-induced apoptosis, augments Env-specific antibody responses, and is associated with reduced interferon-alpha and interferon-gamma responses induced after vaccination.

overall_design: Female BALB/c mice were intramuscularly immunized at 10^7 pfu/dose with MVA expressing SHIV antigens or MVA-B13R expressing SHIV. At days 1, 2, and 6 post immunization, draining inguinal lymph node cells were harvested for RNA extraction. 5 animals per group per time point including 5 naive mice as a control group.

first: Lynette

last: Chea

first: Rama

last: Amara

first: Steven

last: Bosinger

first: Nirav

last: Patel


title: MMP14 empowers tumor-initiating breast cancer cells under hypoxic nutrient-depleted conditions

city: Lübeck

first: Hauke

last: Busch

state: Schleswig-Holstein

address: Ratzeburger Allee 160

department: Lübeck Institute of Experimental Dermatology

institute: University of Lübeck

summary:
Tumor-initiating cells (TICs) exist in breast cancer and are thought to be the cause of tumor initiation, progression, and relapse. In these processes, proteases play an important role. Additionally, conditions of the tumor milieu, including hypoxic nutrient-deprived as well as normoxic nutrient-rich environments, influence the tumor-promoting actions of neoplastic cells. Therefore, we investigated the role of proteases in TICs and their response to different environments by means of a generated immortalized TIC line.
After assessing immortalized TIC tumorigenicity by limiting dilution assays in vivo and their characteristics in different environmental conditions, a transcriptome analysis was performed of iTICS cultured in hypoxic stem cell conditions and 1-7 days in normoxic standard condtitions. A high proteolytic signature was identified and further investigated by inhibitor treatment and shRNA mediated RNA interference of Mmp14.

overall_design: Immortalized tumor-initiating cells (iTICs) were grown in hypoxic stem cell conditions (HSCC) or normoxic standard conditions till confluency of 80% on a 10 cm petridish. Subsequently, total RNA was extracted, analyzed for their quality (RIN), and hybridized to a Mouse WG-6 v2.0 Bead Chip (Illumina). To obtain an overview of expression profile over time iTICs are cultured in NSC, in addition to the HSCC replicates (3x), samples of iTICs cultured in NSC for 1, 2, 3, 4, 5, 6, and 7 days NSC were analyzed. The 7 day sample in NSC was also analyzed in triplicate while all other days in NSC were analyze as one replicate.

first: Larissa

last: Hillebrand

first: Thomas

last: Reinheckel

first: Melanie

last: Boerries


title: Divergent rRNAs as regulators of gene expression at the ribosome level

city: Seoul

first: Seong-il

last: Eyun

address: 84 Heukseok-ro

department: Department of Life Science

institute: Chung-Ang University

summary: It is generally assumed that each organism has evolved to possess a unique ribosomal RNA (rRNA) species optimal for its physiological needs. However, some organisms express divergent rRNAs, the functional roles of which remain unknown. Here, we show that ribosomes containing the most variable rRNAs, encoded by the rrnI operon (herein designated I-ribosomes), direct the preferential translation of a subset of mRNAs in Vibrio vulnificus, enabling the rapid adaptation of bacteria to temperature and nutrient shifts. In addition, genetic and functional analyses of I-ribosomes and target mRNAs suggest that both I-ribosomal subunits are required for the preferential translation of specific mRNAs, the Shine-Dalgarno sequences of which do not play a critical role in I-ribosome binding. This study identifies genome-encoded divergent rRNAs as regulators of gene expression at the ribosome level, providing an additional level of regulation of gene expression in bacteria in response to environmental changes.

overall_design:
Ribosome profiling analysis. Cells were grown at 30 °C in LBS media containing 0.2 μg/ml tetracycline (Tc) to an OD600 of 0.7. After treatment with 100 µg/ml chloramphenicol to stop translation, the cells were incubated at 30 °C for 5 min. The culture was cooled on ice and harvested by centrifugation. The pellet was washed with pre-chilled buffer A (50 mM Tris-HCl, pH 7.6, 10 mM MgCl2, 100 mM NH4Cl, 6 mM 2-mercaptoethanol, 0.5 mM EDTA) containing 1 mM of chloramphenicol and was lysed using a French press. Cellular debris was removed by centrifuging for 30 min at 30,000 × g. The clarified lysate (supplemented with 5 mM CaCl2) was digested with micrococcal nuclease for 1 h at 25 °C and quenched with EGTA to a final concentration of 6 mM. The resulting lysate was loaded onto a 30% sucrose cushion. Crude ribosomes were pelleted by centrifugation at 100,000 × g for 16 h at 4 °C. The pellets were resuspended and loaded onto a 15–40% sucrose gradient and centrifuged at 79,500 × g for 13.5 h at 4 °C. After centrifugation, monosomes were obtained by fractionation and measurement of optical density at 260 nm. Monosomal RNA was obtained by phenol extraction and ethanol precipitation and was resuspended in RNase-free water. To obtain ribosome-protected mRNAs, extracted monosomal RNA was resolved on a 15% polyacrylamide-8 M urea-TBE gel. A band between 28 and 42 bp was excised, gel puriﬁed, precipitated, and resuspended in RNase-free water. For ribosomal RNA depletion the Ribo-Zero rRNA removal kit was used according to the manufacturer’s instructions. Isolated RNAs were used to construct a cDNA library for Illumina sequencing using the TruSeq Small RNA Sample Prep Kit according to the manufacturer’s instructions. RNA sequencing was performed on the Illumina HiSeq 2500 platform using single-end 50 bp sequencing.
Transcriptome analysis. Total cellular RNA was extracted from cells grown at 30 °C in LBS media containing 0.2 μg/ml tetracycline (Tc) to an OD600 of 0.7 using TRIzol according to the manufacturer’s instructions. The mRNA in total RNA was converted into a library of template molecules suitable for subsequent cluster generation using the reagents provided in the Illumina TruSeq RNA Sample Preparation Kit. The first step in the workflow involves removing the rRNA in the total RNA using Ribo-Zero rRNA Removal kit. Following this step, the remaining mRNA is fragmented into small pieces using divalent cations under elevated temperature. The cleaved RNA fragments are copied into first strand cDNA using reverse transcriptase and random primers. This is followed by second strand cDNA synthesis using DNA Polymerase I and RNase H. These cDNA fragments then go through an end repair process, the addition of a single ‘A’ base, and then ligation of the adapters. The products are then purified and enriched with PCR to create the final cDNA library. RNA sequencing was performed on the Illumina HiSeq 2500 platform using paired-end 100 bp sequencing.
Sequencing data analysis. The sequence of the reference genome was retrieved from the NCBI database. RNA-seq reads were aligned to the reference genome sequence using Bowtie2 or BWA. Coordinates of mapped reads were obtained by processing BAM formatted files using BEDOPS tools and were stored in BED formatted files. Coordinates of genome features such as protein coding genes were extracted from the genome annotation data. Boundaries of features such as transcription start sites and gene segments were prepared: a transcription start site was defined as positions from -60 to +60 (the first nucleotide of a start codon was numbered as +1) and a gene segment as positions from -60 to the end of the stop codon. Overlapping genome-mapped RNA-seq reads were assigned to these features. The normalized abundance of reads was measured as number of reads per million mapped reads (RPM).

first: Wooseok

last: Song

first: Minju

last: Joo

first: Ji-Hyun

last: Yeom

first: Eunkyoung

last: Shin

first: Minho

last: Lee

first: Hyung-Kyoon

last: Choi

first: Jihwan

last: Hwang

first: Yong-In

last: Kim

first: Ramin

last: Seo

first: J

first: Christopher

last: Moore

first: Yong-Hak

first: Yoonsoo

last: Hahn

first: Jeehyeon

last: Bae

first: Kangseok


title: Proximity-CLIP provides a snapshot of occupied cis-acting elements on RNA in different subcellular compartments on a transcriptome-wide scale

status: Public on Nov 26 2018

city: Tel Aviv

first: Daniel

last: Benhalevy

address: Tel Aviv University, Green Bldg. Room 205

department: Shmunis School of Biomedicine and Cancer Research

country: Israel

institute: Tel-Aviv University

summary: Many cellular RNAs localize to specific subcellular compartments. Currently, methods to systematically study subcellular RNA localization are limited and lagging behind proteomic approaches. Here, we combined APEX2-mediated proximity biotinylation of proteins with PAR-CLIP to simultaneously profile the proteome and the transcriptome bound by RNA binding proteins in any given subcellular compartment. Our approach is fractionation-independent and does not rely on additional RNA manipulation and labeling steps, thus making it easy to apply. Furthermore, it enables to study the locali-zation of RNA processing intermediates, as well as the identification of regulatory RNA cis-acting elements occupied in different cellular compartments. In a proof-of-concept study we studied RNA and protein localization in the nucleus, cytoplasm and at cell-cell interfaces using Proximity-CLIP. These experiments revealed among other in-sights frequent transcriptional readthrough continuing for several kilobases down-stream of the canonical cleavage and polyadenylation site, a differential binding pat-tern of nuclear and cytoplasmic mRNAs, as well as the localization of mRNAs contain-ing 3’UTR CUG sequence elements at cell-cell interfaces, of which many encode regulatory proteins.

overall_design: Proximity-CLIP takes advantage of the occupancy of cellular RNAs by RBPs throughout their life cycle. An APEX2 fusion protein is targeted to a cellular compartment, and cellular RNAs are labeled with 4SU. Cells are incubated with biotin-phenol (BP) for 30 min, before APEX2-mediated BP oxidation is activated by addition of hydrogen peroxide, followed by reaction quenching and 4SU-dependent protein-RNA crosslinking by UV illumination. BP radicals are created locally and either covalently tag proximate proteins or decay. Compartment-specific RNPs are captured by Streptavidin affinity chromatography. Eluted RNA is analyzed either by RNAse treatment followed by small RNA cDNA library preparation of RBP-protected footprints, analogous to PAR-CLIP, or by standard RNAseq. Data also includes RNAseq analysis of the input, i.e. total cell extract RNA prior to Streptavidin chromatography.

first: Markus

last: Hafner


title: Transcriptomic analysis of MERS-CoV infected Calu-3 cell with or without AM580 treatment

status: Public on Nov 24 2018

city: Hong Kong

first: Jasper

middle: Fuk-Woo

last: Chan

state: Hong Kong

address: Block T, 122 Pokfield Road

department: Microbiology

country: Hong Kong

institute: The University of Hong Kong

summary: We report a broad spectrum antiviral drug AM580, RNA-Seq technology was used to compare the host gene mRNA expression between different groups to indicate the host pathway changes upon MERS-CoV infection and AM580 treatment.

overall_design: Examination of mock-infected cells, MERS-CoV-infected cells and MERS-CoV-infected cells with AM580 treatment, in triplicate

first: Shuofeng

last: Yuan

first: Lei

last: Wen

first: Kwok-Yung

last: Yuen


title: Critical roles of lysine demethylase Kdm3a in craniofacial and neural development during Xenopus embryogenesis

status: Public on Nov 23 2018

city: Ulsan

first: Taejoon

last: Kwon

address: 50 Unist-gil

department: Department of Biomedical Engineering

institute: Ulsan National Institute of Science and Technology

summary: The epigenetic modifier lysine-specific histone demethylase 3a (kdm3a) is specific for demethylation of mono- and di-methylated 9th lysine of histone H3 (H3K9me1/2) and belongs to the Jumonji domain-containing group of demethylases. Kdm3a is known to play significant roles during various biological and pathophysiological processes such as spermatogenesis and metabolism, determination of sex, androgen receptor-mediated transcription, and embryonic carcinoma cell differentiation. In the current study, we evaluated the physiological functions of kdm3a during Xenopus laevis embryogenesis. Spatiotemporal expression pattern indicated maternal nature of kdm3a, exhibiting its expression from early embryonic stages until tadpole stage, however considerable high level of expression were observed during the neurula stage of Xenopus development. Depleting kdm3a using kdm3a antisense morpholino oligonucleotides induced anomalies, such as head deformities and small-sized eyes. Our whole-mount in situ hybridization results demonstrated that kdm3a knockdown is associated with defects in neural crest formation and migration. Further, reverse transcription polymerase chain reaction revealed abnormal expression of neural markers in kdm3a morphants. RNA sequencing of kdm3a morphants indicated that kdm3a is implicated in mesoderm formation, cell adhesion, and metabolic processes of embryonic development. In conclusion, we suggest that kdm3a is critical for neural development during Xenopus embryogenesis and can be targeted for treatment of developmental disorders. The further detailed investigation is required to understand the molecular mechanism involved in neural development regulation by kdm3a.

overall_design: Collect mRNA from whole embryos; two biological replicates were analyzed

first: Hyun-Kyung

first: Tayaba

last: Ismail

first: Chowon

first: Youni

first: Jeen-Woo

last: Park

first: Oh-Shin

first: Beom-Sik

last: Kang

first: Dong-Seok

first: Tae

first: Hyun-Shik


title: Expression data from second oncogene expressing Melanoma Cells.

status: Public on Nov 22 2018

city: Boston

first: Raj

last: Kumar

state: MA

address: 50 Blossom St

department: Dermatology

institute: MGH

summary:
Among the myriad tumors analyzed to date, cutaneous malignant melanomas bear some of the highest mutational burdens. Thus, it is somewhat surprising that oncogene exclusion between is so pronounced in melanomas where there is only a single melanoma out of 366 sequenced specimens which harbors concurrent BRAF(V600E/M) and NRAS(G13R) mutations (TCGA-ES-A2NC Sample; WWW.bioportal.org), In this senario some NRAS and BRAF mutations co-expressing cells show slow growth phenotypes by under study mechanism.
We used Affymetrix GeneChip Expression Arrays to detail the global programme of gene expression underlying oncogenic exclusion and identified distinct classes of up and down-regulated genes during this process.

overall_design: Total mRNA was collected at 5th day from doxycycline induced and non-induced melanoma cell lines (A375(BRAF*)+iNRAS*, GMEL(BRAF*)+iNRAS*,SKMEL119(NRAS*)+iBRAF*) to express second oncogene respectively and subjected  for hybridization on Affymetrix microarrays.

first: Hensin

last: Tsao


title: DAF-16 stabilizes the aging transcriptome and is activated in mid-aged C. elegans to cope with internal stress

city: Beijing

first: Wenhui

last: Li

address: 7 Science Park Road ZGC Life Science Park

institute: National Institute of Biological Sciences

summary: The roles and regulatory mechanisms of transcriptome changes during aging are unclear. It has been proposed that the transcriptome suffers decay during aging owing to age-associated down-regulation of transcription factors. In this study, we characterized the role of a transcription factor DAF-16, which is a highly conserved lifespan regulator, in the normal aging process of Caenorhabditis elegans. We found that DAF-16 translocates into the nucleus in aged wild-type worms and activates the expression of hundreds of genes in response to age-associated cellular stress. Most of the age-dependent DAF-16 targets are different from the canonical DAF-16 targets downstream of insulin signaling. This and other evidence suggest that activation of DAF-16 during aging is distinct from activation of DAF-16 due to reduced signaling from DAF-2. Further analysis showed that it is due in part to a loss of proteostasis during aging, at least in part. We also found that without daf-16, dramatic gene expression changes occur as early as on adult day 2, indicating that DAF-16 acts to stabilize the transcriptome during normal aging. Our results thus reveal that normal aging is not simply a process in which the gene expression program descends into chaos due to loss of regulatory activities; rather, there is active transcriptional regulation during aging.

overall_design: Transcriptome analyses of fer-15(b26ts) and daf-16(mu86);fer-15(b26ts) worms were carried out in three biological replicates. Worms were grown on High Growth (HG) plates supplemented with OP50 bacteria at 25 °C and harvested on adult day 1 through day 7.

first: Shang-Tong

first: Han-Qing

last: Zhao


title: Gene expression profiles in Small Intestine, Colon, iPSCs and hiPS-SIECs

status: Public on Nov 21 2018

city: Suita

first: Ryosuke

last: Negoro

state: Osaka

address: 1-6 Yamadaoka

department: Graduate School of Pharmaceutical Sciences

institute: Osaka University

summary: Efficient Generation of Small Intestinal Epithelial-like Cells from Human iPS Cells for Drug Absorption and Metabolism Studies

overall_design: Small Intestine, Colon, iPSCs and hiPS-SIECs were performed microarray analysis.

first: Kazuo

last: Takayama

first: Hiroyuki

last: Mizuguchi


title: A Cell Proliferation and Inflammatory Signature is Induced by Lawsonia intracellularis Infection in Swine

city: St Paul

first: Fernando

middle: L

last: Leite

address: 1971 Commonwealth Avenue

institute: University of Minnesota

summary: Lawsonia intracellularis causes porcine proliferative enteropathy. This is an enteric disease characterized by thickening of the wall of the ileum that leads to decreased growth and diarrhea of animals. In this study, we investigated the host response to L. intracellularis infection by performing transcriptomic and pathway analysis of intestinal tissue in groups of infected and non-infected animals at 14, 21 and 28 days post challenge.

overall_design: The mucosa of the terminal ileum of two treatment groups, infected and non-infected by Lawsonia intracellularis, were examined at 14, 21 and 28 days post infection.

first: Juan

last: Abrahante

first: Erika

last: Vasquez

first: Fabio

last: Vannucci

first: Connie

last: Gebhart

first: Nathan

last: Winkelman

first: Adam

last: Mueller

first: Jerry

last: Torrison

first: Zachary

last: Rambo

first: Richard

last: Isaacson


title: The Type I interferon response impairs differentiation potential of pluripotent stem cells [RT-qPCR]

city: New York

first: Julie

middle: Anne

last: Eggenberger

state: NY

address: 1 Gustave L. Levy Pl

institute: Icahn School of Medicine at Mount Sinai

summary: Upon virus infection,pluripotent stem cells neither induce nor respond to canonical Type I interferons (IFN-I). To better understand this biology,we characterized induced pluripotent stem cells (iPSCs) as well as their differentiated parental or re-derived counterparts. We confirmed that only iPSCs failed to respond to viral RNA,Type I interferon (IFN-I),or viral infection.  This lack of response could be phenocopied in fibroblasts with the expression of a reprogramming factor which repressed the capacity to induce canonical antiviral pathways. To ascertain the consequences of restoring the antiviral response in the context of pluripotency,we engineered a system to engage these defenses in iPSCs.    Inducible expression of a recombinant virus-activated transcription factor resulted in the successful reconstitution of antiviral defenses through the direct upregulation of IFN-I stimulated genes. Induction of the antiviral signature in iPSCs,even for a short duration,resulted in the dysregulation of genes associated with all three germ layers despite maintaining pluripotency markers. Trilineage differentiation of these same cells showed that engagement of the antiviral defenses compromised ectoderm and endoderm formation and dysregulated the development of mesodermal sublineages. In all,these data suggest that the temporal induction of the antiviral response primes iPSCs away from pluripotency and induces numerous aberrant gene products upon differentiaton. Together these results suggest that the IFN-I system and pluripotency may be incompatible with each other and thus explain why stem cells do not utilize the canonical antiviral system.

overall_design: RT-qPCR

first: Benjamin

last: tenOever


title: Antigen-specific T-cell receptor signatures of cytomegalovirus infection

city: Munich

first: Andreas

last: Moosmann

address: Marchioninistr. 25

department: AGV

institute: Helmholtz Center Munich

summary: We immunosequenced peptide-stimulated PBMCs from 8 CMV-positive donors to identify virus-specific T cell receptors. We also sequenced ex vivo T cell repertoires of multiple CMV-postive and CMV-negative donors to compare the availability of CMV-specific TCRs in virus carriers and uninfected subjects.

overall_design:
Blood samples from healthy anonymous donors were obtained from the "Institut für Transfusionsmedizin Ulm ", Germany. PBMCs were isolated by standard Ficoll gradient centrifugation and mRNA was isolated either directly or after 10-day stimulation with an immunogenic CMV peptide.
For validation of our short-term stimulated TCRs, PBMCs from 3 different donors were stimulated with autologous transformed B cells (mini-LCLs) for 23 or 30 days. These mini-LCLs expressed one of the CMV antigens pp65 or IE1 together with EBV antigens, or EBV antigens alone (control mini-LCLs).
TCR-enriched sequencing libraries were obtained using TCR beta chain-specific primers with Illumina adapter sequence overhangs. After Sequencing on an Illumina HiSeq1500 device, raw data was demultiplexed, quality filtered, and TCR repertoires were generated by alignment and grouping of individual reads.

first: Alina

last: Huth

first: Xiaoling

last: Liang

first: Stefan

last: Krebs

first: Helmut

last: Blum


title: Portal Hypertension Induced Changes in Splenic Gene Expression

city: Kumamoto

first: Junji

last: Saruwatari

state: Select One...

address: 5-1 Oe-honmachi, Chuo-ku

department: Department of Pharmacology and Therapeutics

institute: Kumamoto University

summary: Splenomegaly is caused by several pathological conditions, including portal hypertension, which is most frequently caused by chronic liver disease (e.g., liver cirrhosis).  The detailed mechanisms through which portal pressure induces splenomegaly and the precise pathophysiological conditions in portal hypertension-induced splenomegaly remain to be fully elucidated.  We used microarrays to identify the differential gene expression underlying the portal hypertension-induced splenomegaly.

overall_design: Transcriptomic profile of whole spleen tissues from three male Sprague–Dawley rats with partial portal vein ligation (PPVL) as well as from the three sham rats as controls.

first: Chao

last: Dong

first: Guilin

last: Wang

first: Yasuko

last: Iwakiri


title: Absence of gut microbial colonization attenuates the sympathoadrenal response to hypoglycemia in mice

status: Public on Nov 25 2018

city: Valhalla

first: Bistra

last: Nankova

address: 95 Grassland Rd

institute: New York Medical College

summary: Gut microbiota plays an important role during early development via bidirectional gut- brain signaling. We aimed to explore the potential link between gut microbiota/gut derived metabolites and sympathoadrenal stress responsivity

overall_design: The reponse to insulin induced hypoglycemia was compared in normally colonized male mice (control, C) and sterile male (germ free mice, GF) of the same genetic background (C57Bl6) using whole genome transcriptome profyling of adrenal medullary RNA samples.

first: Priyadarshani

last: Giri

first: Furong

last: Hu

first: Edmund

last: LaGamma


title: Gain of function DNMT3A mutations cause microcephalic dwarfism and hypermethylation of Polycomb-regulated regions (human Illumina methylation)

city: Edinburgh

first: Duncan

last: Sproul

state: Mid Lothian

address: Crewe Road South

department: MRC Human Genetics Unit

country: United Kingdom

institute: University of Edinburgh

type: Methylation profiling by genome tiling array

summary:
DNA methylation and Polycomb are key factors in the establishment of vertebrate cellular identity and fate. Here we report de novo missense mutations in DNMT3A, encoding the DNA methyltransferase DNMT3A, that cause microcephalic dwarfism, a hypocellular disorder of extreme global growth failure. Substitutions in the PWWP domain abrogate binding to the histone modifications H3K36me2/3, and alter DNA methylation in patient cells. Polycomb-associated DNA methylation canyons/valleys, hypomethylated domains encompassing developmental genes, become methylated with concomitant depletion of H3K27me3 and H3K4me3 bivalent marks. Such de novo DNA methylation occurs during differentiation of Dnmt3aW326R pluripotent cells in vitro, and is also evident in Dnmt3aW326R/+ dwarf mice. We therefore propose that the interaction of the DNMT3A PWWP domain with H3K36me2/3 normally limits DNA methylation of polycomb-marked regions. Our findings implicate the interplay between DNA methylation and polycomb at key developmental regulators as a determinant of organism size in mammals.
This series contains Infinium methylation data on human blood and fibroblast samples.

overall_design: Bisulphite converted DNA from 13 samples were hybridized to the Illumina Infinium EPIC Beadchip

first: Patricia

last: Heyn

first: Andrew

middle: P

last: Andrew


title: Microarray analysis with KSHV infection and enrichment for circular RNAs

status: Public on Nov 20 2018

city: Bethesda

first: Joseph

middle: M

last: Ziegelbauer

state: MD

address: 10 Center Dr. 6N110

institute: NIH / National Cancer Institute

summary:
The purpose was to measure expression changes in human circular RNAs.
Total RNA was harvested from KSHV-infected HUVEC and MC116 cells. A portion of the RNA was digested with RNase R to enrich for circular RNAs.

overall_design: Uninfected versus KSHV-infected in two cell types, 3 biological replicates

first: Takanobu

last: Tagawa


title: RNA sequencing for KSHV circular RNAs

summary: The purpose was to detect viral circular RNAs from infected cells.

sample_organism:
Homo sapiens
Human gammaherpesvirus 8

platform_organism:
Homo sapiens
Human gammaherpesvirus 8

overall_design: Total RNA was harvested from KSHV-infected HUVEC cells. A portion of the RNA was digested with RNase R to enrich for circular RNAs.

first: Skyler

last: Kuhn


title: Gain of function DNMT3A mutations cause microcephalic dwarfism and hypermethylation of Polycomb-regulated regions (human RRBS)

type: Methylation profiling by high throughput sequencing

summary: DNA methylation and Polycomb are key factors in the establishment of vertebrate cellular identity and fate. Here we report de novo missense mutations in DNMT3A, encoding the DNA methyltransferase DNMT3A, that cause microcephalic dwarfism, a hypocellular disorder of extreme global growth failure. Substitutions in the PWWP domain abrogate binding to the histone modifications H3K36me2/3, and alter DNA methylation in patient cells. Polycomb-associated DNA methylation canyons/valleys, hypomethylated domains encompassing developmental genes, become methylated with concomitant depletion of H3K27me3 and H3K4me3 bivalent marks. Such de novo DNA methylation occurs during differentiation of Dnmt3aW326R pluripotent cells in vitro, and is also evident in Dnmt3aW326R/+ dwarf mice. We therefore propose that the interaction of the DNMT3A PWWP domain with H3K36me2/3 normally limits DNA methylation of polycomb-marked regions. Our findings implicate the interplay between DNA methylation and polycomb at key developmental regulators as a determinant of organism size in mammals.

overall_design:
This series contains RRBS from human blood samples.
Raw sequencing reads protected as patient material, submitted to EGA. Accession: EGAS00001003232


title: Transcriptomic analyses of the testicular interstitial cells in murine neonates

city: Kurume

first: Yuichi

last: Shima

state: Fukuoka

address: Asahimachi 67

department: Department of Anatomy

institute: Kurume University School of Medicine

summary: Previous studies have established that fetal Leydig cells (FLCs) and adult Leydig cells (ALCs) show distinct functional characteristics. However, the lineage relationship between FLCs and ALCs has not been clarified yet. Here we reveal that a subset of FLCs dedifferentiate at fetal stages to give rise to ALCs at the pubertal stage. Moreover, the dedifferentiated cells contribute to the peritubular myoid cells and vascular pericyte populations in the neonatal testis, and these non-steroidogenic cells serve as potential ALC stem cells. We generated FLC lineage-specific Nr5a1 (Ad4BP/SF-1) gene disrupted mice and mice lacking the fetal Leydig enhancer (FLE) of the Nr5a1 gene. Phenotypes of these mice support the conclusion that most of the ALCs arise from dedifferentiated FLCs, and that the FLE of Nr5a1 gene is essential for both initial FLC differentiation and pubertal ALC redifferentiation.

overall_design: Cells were dispersed from mFLE-Cre;CAG-CAT-EGFP mouse testis at postnatal day 10, and EGFP(weak)/Laminin(negative) population and EGFP(positive)/Laminin(positive) pouplations were collected by cell sorting. Duplicated samples in each population were analyzed by RNA-sequence.

first: Kanako

last: Miyabayashi

first: Yasuyuki

last: Ohkawa

first: Tetsuya

last: Sato

first: Mikita

last: Suyama


title: Identifying lncRNAs as novel modulators of macrophage M2 polarization

city: Hangzhou

first: Rong

last: Dong

state: Zhejiang

address: No.886 Yuhangtang Road

department: College of Pharmaceutical Sciences

institute: Zhejiang University

summary: By utilizing a series of specific modulating M2 macrophages polarization models, 3872 up-regulated and 3091 down-regulated genes were found during the polarization process.

overall_design: To screen for the functional lncRNAs involved in macrophages M2 polarization, we utilized the cellular model of IL13-driven macrophage M2 polarization. The time course samples (0 and 24 hours) of bone marrow-derived macrophages (BMDMs) were harvested after stimulating with IL13 (10 ng/mL).

first: Ji

last: Cao


title: Expression Data from Liver of Rats Dosed with Reference Hepatotoxicants_RNASeq

city: North Chicago

first: Rita

last: Ciurlionis

address: 1 North Waukegan Road

institute: Abbvie

summary: Male Sprague Dawley rats were treated orally for 5 days with reference hepatotoxicants, each at a single dose level, alpha-naphthylisothiocyanate 100mg/kg (vehicle 5ml/kg corn oil); carbon tetrachloride 1582mg/kg (vehicle 5ml/kg corn oil); methylenedianiline 100mg/kg (vehicle 35% ethanol); acetaminophen 1000mg/kg (vehicle 2ml/kg corn oil); diclofenac 10mg/kg (vehicle 10ml/kg water)

overall_design: 3 rats were treated with each hepatotoxicant, 3 rats were treated with corresponding vehicle for each hepatotoxicant, total RNA was isolated form left lateral lobe of liver of all rats, samples run individually on Affymetrix GeneChip Rat Genome 230_2.0 Arrays

first: Mohan

last: Rao


title: Identifying lncRNA-MM2P as a novel modulator of macrophage M2 polarization

summary: By utilizing a series of specific modulating M2 macrophages polarization models, 1556 up-regulated and 953 down-regulated genes were found during the polarization process.

overall_design: To screen for the functional lncRNAs involved in macrophages M2 polarization, we utilized the cellular model of IL13-driven macrophage M2 polarization. The time course samples (0, 24, 48 and 72 h) of RAW264.7 cells were harvested after stimulating with IL13 (10 ng/mL).


title: Gain of function DNMT3A mutations cause microcephalic dwarfism and hypermethylation of Polycomb-regulated regions (mouse RRBS)

overall_design: This series contains RRBS from mouse NPC cells and cortex.


title: Profiling of mouse Fibro/adipogenic progenitors (FAPs) aging and activation upon skeletal muscle injury

city: Lausanne

first: Jerome

last: Feige

address: EPFL Innovation Park, Building H

department: Musculo-Skeletal Health

institute: Nestle Institute Of Health Sciences

summary: Utilizing glycerol intramuscular injections in M. musculus provides a model of skeletal muscle damage followed by skeletal muscle regeneration. In particular, glycerol-induced muscle injury triggers accute activation of muscle Fibro/Adipogenic Progenitors, also called FAPs. However, aging dramatically impairs FAP function. We characterized genome-wide expression profiles of young and old FAPs in the non-proliferative and activated state, freshly isolated to non-injured or damaged muscles, respectively. Our goal was to uncover new regulatory signalings specific to FAP entry into the activation program that are affected with age.

overall_design: Fibro/Adipogenic progenitors (FAPs) were isolated in either quiescent / non-proliferative or activated state from uninjured or 3 days after glycerol-induced injury of tibialis anterior, gastrocnemius and quadriceps, respectively. Young (2-4 months old) and old (20-24 months old) wildtype C57BL/6J male were used, with six biological replicates per group.

first: Laura

last: Lukjanenko

first: Eugenia

last: Migliavacca

first: Sonia

last: Karaz

first: Sylviane

last: Metairon

first: Frederic

last: Raymond

middle: N


title: Response of ECF sigma factor deletion mutants of Cupriavidus metallidurans to mixed metal ions (Co, Cu, Cd, Zn, Ni, 30 µM each)

city: Halle/Saale

first: Cornelia

last: Große

department: Faculty of Biology/Microbiology

summary: Response of Cupriavidus metallidurans CH34 and ECF deletion mutants to mixed metals

overall_design: In this study metal mix-treatment was performed to acquire expression profiles of the total transcriptome of Cupriavidus metallidurans and mutants

last: Grosse

first: Frank

last: Reith

first: Dietrich

middle: H

last: Nies


title: An Optically Decodable Bead Array for Linking Imaging and Sequencing with Single-Cell Resolution

first: Peter

last: Sims

address: 3960 Broadway, 2nd Floor, Room 203AC

institute: Columbia University

summary: Optically decodable beads link the identity of an analyte or sample to a measurement through an optical barcode, enabling libraries of biomolecules to be captured on beads in solution and decoded by fluorescence. This approach has been foundational to microarray, sequencing, and flow-based expression profiling technologies. We have combined microfluidics with optically decodable beads to link phenotypic analysis of living cells to sequencing. As a proof-of-concept, we applied this to demonstrate an accurate and scalable tool for connecting live cell imaging to single-cell RNA-Seq called Single Cell Optical Phenotyping and Expression (SCOPE-Seq).

sample_organism:
Homo sapiens
Mus musculus
synthetic construct

overall_design: Performed SCOPE-Seq on thousands of cells from two cell lines.

first: Jinzhou

last: Yuan

middle: A


title: The effect of type I IFN treatment on global gene expression in monocytes undergoing maturation

status: Public on Nov 19 2018

city: Nanjing

first: Tong

address: Xue Fu Road 12

institute: Nanjing University

summary: To analyze the effect of type I IFN on global gene expression in differentiating monocytes, we employed microarray-based gene expression profiling. The MACS-purified mouse BM monocytes were cultured in M-CSF and treated with or without IFN for 60 h. The cells were harvested and isolated RNAs were subjected to microarray analyses.

overall_design: MACS-purified monocytes from mouse bone marrow were divided into two groups (each in triplicates) and treated with 20 ng/mL M-CSF, together with or without 100 U of mouse IFN-beta for 60 h. The resulting cultures were harvested and the RNA samples were subjected to microarray analyses.

first: Jianghuai

last: Liu

first: Yuanyuan

last: Tong


title: Choroid Plexus Gene Expression in ApoE3 or ApoE4-Knockin Mouse Choroid Plexuses

first: Andreas

middle: Johann Richard

last: Habenicht

state: Bavaria

address: Pettenkoferstraße 9a

department: Medicine

institute: Ludwig Maximilians Universität München

summary: Laser capture microdissected choroid plexuses were obtained and expression arrays were generated to investigate gene expression in ApoE-Knockin choroid plexuses; the choroid plexus forms the cerebrospinal fluid, the cerebrospinal fliod barrier, functions as the major gateway for blood-born leukocytes to enter the brain in degenerative and inflammatory brain diseases, and the principal neuroimmune interface in the brain. We found lipid deposits in the aged choroid plexus of hyperlipidemic ApoE4-Knockin mice but none in normolipidemic ApoE4-Knockin or normolipidemic or hyperlipidemic ApoE3-Knockin control choroid plexuses. Here, we studied the functional impact and gene epressions these choroid plexuses.

overall_design: 6 replicates each of ApoE3 choroid plexuses or of ApoE4-Knockin mice either maintained under normal mouse chow or under high fat-diet (HFD) of aged (78 weeks old) mice were obtained and expression microarrays were generated

first: Changjun

last: Yin


title: Global transcriptome analysis of resistant and susceptible soybean lines following Sclerotinia sclerotiorum infection

city: Madison

first: Mehdi

last: Kabbage

state: Wisconsin

address: Department of Plant Pathology, 583 Russell Labs 1630 Linden Dr

department: Department of Plant Pathology

institute: University of Wisconsin Madison

summary: Sclerotinia sclerotiorum, a necrotrophic fungal pathogen with a broad host range, causes a devastating disease on soybean called Sclerotinia stem rot (SSR), can lead to losses as high as 50-60%. Resistance mechanisms against SSR are poorly understood. We used high throughput RNAseq approach to decipher the molecular mechanisms governing resistance to S. sclerotiorum in soybean. Transcripts of recombinant inbred lines (RILs) of soybean; susceptible (S) and resistant (R) were analyzed in a time course experiment. This study might provide an important step towards understanding resistance responses of soybean to S. sclerotiorum and identified novel mechanisms and targets.

overall_design: Four-week-old soybean plants of both the resistant (R) and the susceptible (S) were inoculated using the cut petiole inoculation method as described by (Ranjan et.al., 2017; MPP., 19(3):700-714). Plant tissue was sampled by cutting horizontally above and below (1.5 cm) the node of the first trifoliate with a clean straight-edge razor. Tissue samples were collected at 24, 48, and 96 h post inoculation (hpi) and immediately frozen in liquid nitrogen prior to RNA extraction. The stem tissue from non-inoculated (0 h) plants were also collected as a control. The experimental design was completely randomized and consisted of three biological replicates for each of the treatments. For each biological replicate, stem segments (~3 cm, first internode) from two different plants were pooled together. Soybean seedlings and plants were maintained in the greenhouse or growth chamber at 24 ± 2◦C with 16 h light/8 h dark photoperiod cycle. Total RNA extracted for each sample was randomly fragmented, and individually indexed libraries were prepared using the TruSeq RNA Sample Preparation v2 kit according to the manufacturer’s instructions and were sequenced using Illumina HiSeq2500. Raw sequence reads were mapped to both genomes using the Subjunc aligner from Subread. Alignments were compared to the gene annotation GFF files for both organisms (Soybean: Gmax_275_Wm82.a2. v1. gene. gff3 (Schmutz, Cannon et al. 2010), S. sclereotiorum: sclerotinia_sclerotiorum_2_transcripts.gtf (Amselem, Cuomo et al. 2011) and raw counts for each gene were generated using the feature Counts tool from subread. The raw counts data were normalized using voom from the R Limma package, then used to generate differential expression (logFC) values.

first: Ashish

last: Ranjan

first: Sachin

last: Jain

first: Craig

middle: R

last: Grau

first: Damon

last: Smith


title: Choroid Plexus Gene Expression in wild-type (wt) and ApoE-deficient Mouse Choroid Plexuses

summary: Laser capture microdissected choroid plexuses were obtained and expression arrays were generated to investigate gene expression in wt and ApoE choroid plexuses; the choroid plexus forms the cerebrospinal fluid, the cerebrospinal fliod barrier, functions as the major gateway for blood-born leukocytes to enter the brain in degenerative and inflammatory brain diseases, and the principal neuroimmune interface in the brain. We found lipid deposits in the aged choroid plexus of hyperlipidemic mice but none in the wt control choroid plexuses. Here, we studied the functional impact and gene epressions in wt and ApoE-deficient choroid plexuses.

overall_design: wt and ApoE mice were maintained on normal mouse chow and allowed to age for 78 weeks; then, choroid plexuses were obtained by laser capture microdissection; total RNA was prepared and whole genome microarrays generated