title: Widespread IFN-independent activities of mammalian STING in viral infection and tumor immune evasion

city: Dallas

last: Yan

state: TEXAS

address: 6000 Harry Hines Blvd.

department: Immunology

institute: UT Southwestern Medical Center

summary: Type I interferon (IFN) response is the most recognized signaling activity of STING, and IFN  signaling is commonly believed to be the major (if not the sole) contributor to STING-mediated  anti-viral and anti-cancer responses. Here, we genetically mutated mouse Sting serine 365  residue that is critically required for initiating IFN signaling. Sting-S365A mutation in mice  precisely ablated IFN-dependent while preserving IFN-independent activities of STING. To our  surprise, StingS365A/S365A mice protected against HSV-1 infection remarkably well, similar to wild  type mice, while Sting–/– mice succumbed to infection very quickly. This challenges the current  prevailing view and suggests that STING controls HSV-1 infection through IFN-independent  activities. Transcriptomic analysis of wild type, Sting–/– and StingS365A/S365A macrophages and T  cells stimulated with Sting agonist DMXAA revealed widespread IFN-independent activities of  STING in both cell types and identified many T cell effector functions that are activated by STING  independently of IFN. In mouse tumor models, we found that T cells in the tumor experience  substantial cell death that was in part mediated by STING. Adoptively transferred Sting–/– T cells  were more resistant to cell death in the tumor leading to more effective tumor control compared  to wild type and StingS365A/S365A T cells. Together, our data demonstrate that mammalian STING  possesses widespread IFN-independent activities that are functionally important for restricting  Type I interferon (IFN) response is the most recognized signaling activity of STING, and IFN signaling is commonly believed to be the major (if not the sole) contributor to STING-mediated anti-viral and anti-cancer responses. Here, we genetically mutated mouse Sting serine 365 residue that is critically required for initiating IFN signaling. Sting-S365A mutation in mice precisely ablated IFN-dependent while preserving IFN-independent activities of STING. To our surprise, StingS365A/S365A mice protected against HSV-1 infection remarkably well, similar to wild type mice, while Sting–/– mice succumbed to infection very quickly. This challenges the current prevailing view and suggests that STING controls HSV-1 infection through IFN-independent activities. Transcriptomic analysis of wild type, Sting–/– and StingS365A/S365A macrophages and T cells stimulated with Sting agonist DMXAA revealed widespread IFN-independent activities of STING in both cell types and identified many T cell effector functions that are activated by STING independently of IFN. In mouse tumor models, we found that T cells in the tumor experience substantial cell death that was in part mediated by STING. Adoptively transferred Sting–/– T cells were more resistant to cell death in the tumor leading to more effective tumor control compared to wild type and StingS365A/S365A T cells. Together, our data demonstrate that mammalian STING possesses widespread IFN-independent activities that are functionally important for restricting HSV-1 infection, tumor immune evasion and likely also adaptive T cell immunity.

overall_design: The primary T cells and macrophages were isolated from WT, STING-/- and STING-S365A mice, and then treated with DMSO or DMXAA for 16 hours. The total RNA was extracted and applied to RNA-Seq analysis

first: Jianjun

last: Wu


title: Microarray analysis of transgenic mice specifically overexpressing FOXO1 in skeletal muscle (FOXO1 Tg mice)

status: Public on Jul 07 2020

city: Kyoto

first: Yasutomi

last: Kamei

address: 1-5 Shimogamo-Hangicho, Sakyo-ku

country: Japan

institute: Kyoto Prefectural University

summary: FOXO1, a member of the FOXO forkhead type transcription factors, is markedly up-regulated in skeletal muscle during atrophy. Previously, we created transgenic mice specifically overexpressing FOXO1 in skeletal muscle (FOXO1 Tg mice). These mice weighed less than the wildtype control mice, had a reduced skeletal muscle mass. In this study, to better understand changes in skeletal muscle during atrophy, we performed a microarray analysis of skeletal muscle in wild-type control and FOXO1 Tg mice. The microarray data shows that in the skeletal muscles of FOXO1 Tg mice, gene expression of PGC-1β, a transcriptional regulator whose increased expression activates energy-expenditure-related genes in skeletal muscles, is decreased.

overall_design: For microarray analysis, RNA was isolated from the gastrocnemius skeletal muscle of control mice (25 weeks of age) and FOXO1 Tg mice (25 weeks of age) (Kamei et al., Journal of Biological Chemistry 279:41114-41123, 2004). Samples from wild-type (N=6) and FOXO1 Tg (N=5) mice were pooled before use.


title: Transcriptomic analysis of immortalized oral keratinocytes (n=2) and oral squamous cell carcinoma cells (n=5)

city: Miaoli Conunty

first: Su-Fang

last: Lin

address: No 35, Keyan Road, Zhunan Town

department: NICR

country: Taiwan

institute: National Health Research Institutes, Taiwan

type: Expression profiling by high throughput sequencing

summary: Due to betel nut chewing habit, oral cancer is the fourth leading cause of cancer death among Taiwanese men. In addition, oral cancer is the most common malignancy observed in men with 25-44 years old, alerting an urgent need to curb this disease. It is hypothesized that betel nut ingredients exhibit carcinogenic effects on the oral epithelium and alter local immune system. As a step to identify potential oncogenic gene fusions in Taiwanese oral cancer cells, total RNAs of two immortalized oral keratinocytes (CGHNK2, -K6) and five oral squamous cell carcinoma cell lines (C9, OC3, OEC-M1, TW2.6)  were subjected to RNA-sequencing analysis, followed by an in-house bioinformatic pipeline for fusion gene detection.

overall_design: Total RNA was extracted from specified cell lines before reaching confluency , and the quality of the RNA was assessed with the Agilent Bioanalyzer. Transcriptome libraries from the mRNA fractions were generated following the RNA-Seq protocol (Illumina). Each sample was sequenced in a single lane with the Illumina Genome Analyzer II with an 80-nt read length. Paired-end transcriptome reads passing our filters were mapped to the human reference genome (hg19) using Tophat2. Gene expression profiles were estimated by Cufflinks from SAM/BAM files generated in the previous step. Independently, sequence alignments were processed to nominate gene fusions by using ChimerScan (PMID: 21840877).

first: Sheng-Yao

last: Su

first: Chung-Yen

first: Ann-Joy

last: Cheng

first: Mark

middle: Y

last: Kuo

first: Shu-Chun

first: Yi-Mi

first: Dan

middle: R

last: Robinson


title: Synthetic immunomodulation with a CRISPR super-repressor in vivo

city: Pittsburgh

first: Samira

last: Kiani

state: Pennsylvania

address: A809-B Scaife Hall 3550 Terrace Street Pittsburgh

department: Division of Experimental Pathology

country: USA

institute: University of Pittsburgh

summary: In this study we introduce an enhanced CRISPR-based transcriptional repressor to reprogram immune homeostasis in vivo. By a dual fusion of heterochromatin protein 1 (HP1a) and Krüppel associated box (KRAB) to MS2 coat protein and recruitment to a nuclease competent CRISPR complex, we demonstrate transcriptional repression of the Myeloid differentiation primary response 88 (Myd88) gene in vivo. We report that this strategy can effectively modulate host immune response against adeno-associated virus-mediated gene therapy and influence the course of septicemia.

overall_design:
For the in vitro RNA seq data, Neuro 2A cells received CRISPR components with full length or truncated gRNAs targeting endogenous Myd88 locus. The goal of the experiment was comparing specificity of the gRNAs. Repression levels were assessed 3 days post transfection using RNA-seq and qRT-PCR.
For the in vivo RNA seq data, Cas9 nuclease transgenic mice received AAV1-Myd88-Ms2-HP1aKRAB or AAV1-Myd88-MS2-KRAB through retro-orbital injection. 3 weeks post injection, Myd88 expression levels were assessed in bone marrow by qRT-PCR analysis.

first: Jin

middle: G

last: Park


title: Titanium Tackles the Endolplasmic Reticulum: A First Genomic Study on a Titanium Anticacner Metallodrug

city: Jerusalem

first: Yuval

last: Tabach

address: The Institute For Medical Research-Israel-Canada

department: Department of Developmental Biology and Cancer Research

country: Israel

institute: The Hebrew University of Jerusalem

summary: PhenolaTi is novel non-toxic anticancer chemotherapy; this inert bis(phenolato)bis(alkoxo) Ti(IV) complex demonstrates the intriguing combination of high and wide efficacy with no detected toxicity in animals. Here we unravel the cellular pathways involved in its mechanism of action by a first genome study on Ti(IV)-treated cells, using an attuned RNA-Seq-based available technology. First, phenolaTi induced apoptosis and cell-cycle arrest at the G2/M phase in MCF7 cells. Second, the transcriptome of the treated cells was analyzed, identifying alterations in pathways relating to protein translation, DNA damage, mitochondrial eruption, and p53 regulation. Unlike for common metallodrugs, electrophoresis assay showed no inhibition of DNA polymerase activity. Reduced in vitro cytotoxicity with added endoplasmic reticulum (ER) stress inhibitor supported the ER as a putative cellular target. Altogether, this paper reveals a distinct ER-related mechanism by the Ti(IV) anticancer coordination complex, paving the way for wider applicability of related techniques in mechanistic analyses of metallodrugs.

overall_design: The effect of phenolaTi (drug) on MCF7 cell at 54uM concetration was examined over 6 time points at triplicates. Overall, after 0,3,6,15,24 and 48 hours of incubation. Sample 0 is a control, with untreated cells. Overall 18 samples.

first: Maya

last: Miller

first: Anna

last: Mellul

last: Braun

first: Dana

last: Sheill-Rofe

first: Emiliano

last: Cohen

first: Zohar

last: Shpilt

first: Irene

last: Unterman

first: Ori

last: Braitbard

first: Jacob

last: Hochman

first: Edit

last: Tshuva


title: Aberrantly expressed lncRNAs and mRNAs of osteogenically differentiated mesenchymal stem cells in ossification of the posterior longitudinal ligament

city: Shenzhen

first: Zhaopeng

last: Cai

address: No. 3025, Shennan Middle Road, Futian District

country: China

institute: Sun Yat-sen University

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

summary: Ectopic bone formation is the chief characteristic of ossification of the posterior longitudinal ligament (OPLL). Emerging evidence has revealed that long non-coding RNAs (lncRNAs) can regulate the osteogenic differentiation of mesenchymal stem cells (MSCs), which are the main cells responsible for bone formation. However, the role of lncRNAs in the pathogenesis of OPLL remains unclear. In this study, 725 aberrantly expressed lncRNAs and 664 mRNAs in osteogenically differentiated MSCs from OPLL patients (OPLL MSCs) were identified by microarrays and confirmed by qRT-PCR assays. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses showed that the most enriched pathways included the p53, JAK-STAT and PI3K-Akt signaling pathways. The coexpression network showed the interactions between the aberrantly expressed lncRNAs and mRNAs in OPLL MSCs, and the potential targets and transcription factors of the lncRNAs were predicted. Our research demonstrated the aberrantly expressed lncRNA and mRNA and the potential regulatory networks involved in the ectopic bone formation of OPLL. These findings imply that lncRNAs may play a vital role in OPLL, which provides a new perspective on the pathogenesis of OPLL.

overall_design: The control group including three HDMSC samples was derived from three healthy donors and the experimental group including three OPLLMSC samples was derived from three OPLL patients. The cells used for microarray analysis were cultured in osteogenic differentiation medium for 10 days.

first: Wenjie

last: Liu


title: Stem cell mRNA panel of human pluripotent and adult stem cells

city: Milan

first: Mario

last: Barilani

address: Via F. Sforza 35

country: Italy

institute: Fondazione IRCCS Ca' Granda Ospedale Maggiore Policlinico

summary: Induction of pluripotency in somatic cells needs the full activation of the pluripotency gene regulatory network (PGRN). The core transcription factors of the PGRN establish a  crosstalk with specific micro RNA (miRNA) families to sustain the pluripotent program and govern cell fate decisions. Very recently, circular RNA (circRNA) have been proposed as novel players in the regulation of this molecular circuitry. Herein, we successfully generated human induced pluripotent stem cells (hiPSC) by zero-footprint reprogramming of cord blood mesenchymal stem cells. The hiPSC were extensively characterized for stemness and tri-lineage differentiation potential compared to human embryonic stem cells and parental unreprogrammed cells to assess complete acquisition of the pluripotent identity. High-throughput array-based molecular analyses of messenger RNA (mRNA) (631 genes), miRNA (754 miRNA) and, for the first time, circRNA (13,617 circRNA) were performed to address the role of circRNA in the PGRN. As a result, a circRNA-guided map of miRNA and mRNA associated to naïve and primed pluripotent identity is provided.

overall_design: Seven samples: two primary lines of human cord blood mesenchymal stem cells (reference of unreprogrammed cells, biological replicates); human embryonic stem cells (reference of gold standard pluripotent stem cells); two clonal lines of human induced pluripotent stem cells derived from cord blood mesenchymal stem cells (biological replicates); two lines of human induced pluripotent stem cells derived from fibroblasts (biological replicates).

first: Laura

last: Dioni

first: Alessandro

last: Cherubini


title: Adult mouse hippocampal transcriptome changes associated with long-term behavioral and metabolic effects of gestational air pollution toxicity

city: San Diego

first: Amin

last: Haghani

state: CA

address: 5510 Morehouse Dr

institute: Altos labs

summary: Gestational exposure to air pollution increases the risk of autism spectrum disorder and cognitive impairments with unresolved molecular mechanisms. This study exposed mice throughout gestation to urban derived nanosized particulate matter (nPM). Young adult male and female offspring were studied for behavioral and metabolic changes using forced swim test, fat gain, glucose tolerance, and hippocampal transcriptome. Gestational nPM exposure caused increased depressive behaviors, decreased neurogenesis in the dentate gyrus, and increased glucose tolerance in adult male offspring. Both sexes gained fat and body weight. Gestational nPM exposure induced 29 differentially expressed genes (DEGs) in adult hippocampus related to cytokine production, IL17a signaling, and dopamine degradation in both sexes. Stratification by sex showed 2-fold more DEGs in males than females (69 vs 37), as well as male-specific enrichment of DEGs mediating serotonin signaling, endocytosis, Gai, and cAMP signaling. Gene co-expression analysis (WCGNA) identified a module of 43 genes with divergent responses to nPM between the sexes. Chronic changes in 14 DEGs (e.g., miR9-1) were associated with depressive behaviors, adiposity and glucose intolerance. These genes enriched neuroimmune pathways such as HMGB1 and TLR4. Based on cerebral cortex transcriptome data of neonates, we traced the initial nPM responses of HMGB1 pathway. In vitro, mixed glia responded to 24 h nPM with lower HMGB1 protein and increased proinflammatory cytokines. This response was ameliorated by TLR4 knockdown. In sum, we identified transcriptional changes that could underlie air pollution mediated behavioral and phenotypic changes. These identified genes merit further studies for therapeutic intervention development.

overall_design: Pregnant dams were exposed to nPM during gestational period. The effects were studied in adults.

first: Richard

last: Johnson

first: Nicholas

middle: C

last: Woodward

first: Jason

middle: I

last: Feinberg

first: Kristy

last: Lewis

first: Christine

last: Ladd-Acosta

first: Nikoo

last: Safi

first: Andrew

middle: E

last: Jaffe

first: Constantinos

last: Sioutas

first: Hooman

last: Allayee

first: Daniel

middle: B

last: Campbell

first: Heather

last: Volk

first: Caleb

last: Finch

first: Todd

last: Morgan


title: Expression data from female rat retroperinoneal fat tissue

city: Rennes

first: Frédéric

last: Chalmel

state: France

address: 9 avenue du Pr. Léon Bernard

department: Physiology and physiopathology of the urogenital tract

country: France

institute: Inserm U1085-Irset

summary: Obesity is a complex disease with many causes, including a possible role of environmental chemicals. Perfluorohexane sulfonate (PFHxS) is one of many per- and polyfluoroalkyl substances (PFASs) frequently detected in humans and it is a suspected obesogenic compound. We examined the potential long-term effects of PFHxS on metabolic parameters in rats after developmental exposure to 0.05, 5 or 25 mg/kg bw/day, with or without co-exposure to a background mixture of 12 endocrine disrupting chemicals (EDmix). Both male and female offspring showed signs of lower birth weight following intrauterine exposure. Female offspring exposed to both PFHxS and EDmix showed increased body weight in adulthood. Furthermore, the retroperitoneal fat pad was larger in these female offspring when compared to those exposed to EDmix alone. An attempt to detect putative molecular markers in the fat tissue by performing whole transcriptome profiling resulted in no significant changes between groups and there were no significant effects on plasma leptin levels in exposed females. These results show that early life exposure to endocrine disrupting chemicals can influence body weight later in life, but the effect is not necessarily reflected in changed gene expression in the fat tissue.

overall_design: RNA from the left retroperitoneal fat pad samples from 4 adult female offspring were semi-randomly selected from each group: control, 25-Px, EDmix and 25-Px+ED.

first: Frederic

first: Bertrand

last: Evrard

first: Aurélie

last: Lardenois

first: T

last: Svingen

first: Ramskov

last: Tetzlaff


title: RNA sequencing of silica gel-immobilized and cisplatin-treated ovarian cancer cells

city: Minneapolis

first: Samira

last: Azarin

state: Minnesota

address: 421 Washington Ave SE

department: Chemical Engineering and Materials Science

institute: University of Minnesota Twin Cities

summary: This study aimed to look at the signaling response of ovarian cancer cells to physical confinement in silica gels for 24 and 72 hr relative to standard culture control cells; the study also investigated transcriptomes of ovarian cancer cells surviving physical confinement and later extracted from silica gels and compared signaling to that of ovarian cancer cells surviving cisplatin drug treatment

overall_design: 5 different conditions (Control, Day 1, Day 3, Extracted, and Treated) and 2 biological replicates per condition

first: Tiffany

last: Lam

middle: M


title: miRNome of human pluripotent and adult stem cells

first: Mario

last: Barilani

overall_design: Four samples: two primary lines of human cord blood mesenchymal stem cells (reference of unreprogrammed cells, biological replicates); two clonal lines of human induced pluripotent stem cells derived from cord blood mesenchymal stem cells (biological replicates).

first: Laura

last: Dioni

first: Alessandro

last: Cherubini


title: Copa mutation activates STING and perturbs thymocyte development by increasing type I interferons in the thymus.

city: San Francisco

first: Anthony

last: Shum

address: 555 Mission Bay Blvd South

department: Pulmonary

institute: University of California San Francisco

summary: Pathogenic COPA variants cause a Mendelian syndrome of immune dysregulation with elevated type I interferon signaling. Given the importance of COPA in Golgi-ER transport, we speculated that type I interferon signaling in COPA syndrome involves missorting of STING. Here we show that a defect in COPI transport due to mutant COPA causes ligand-independent activation of STING. Activated STING stimulates type I interferon driven inflammation in CopaE241K/+ mice. Our results demonstrate that activated STING contributes to immune dysregulation in COPA syndrome and may be a new molecular target in treating the disease.

overall_design: Differential gene expression analysis in medullary thymic epithelial cells of CopaE241K/+ mice

middle: K

first: Walter

middle: L

last: Eckalbar

first: Zimu

last: Deng

first: Christopher

middle: S

last: Law


title: Anastrepha ludens early embryogenesis transcriptome

city: Cuernavaca

last: Zurita

state: Morelos

address: Av. Universidad 2001

country: Mexico

institute: UNAM

summary: Anastrepha ludens RNA-Seq experiment of syncytium, cellularization and gastrulation developmental stages.

overall_design: We selected syncytium (0 to 9 h), cellularization (9 to 18 h) and gastrulation (18 to 30 h) embryo developmental stages of A. ludens, with triplicate biological replicas.

first: Ximena

last: Gutierrez-Ramos

middle: A

last: Arteaga-Vazquez


title: Gestational diabetes mellitus alters placental structure, efficiency, and plasticity

city: Kansas City

first: Kaela

middle: Margaret

last: Varberg

state: MO

address: 2401 Gillham Road; CMRI 7902.19

department: Pediatrics

institute: Children's Mercy

summary: The hemochorial placenta provides a critical barrier at the maternal-fetal interface to modulate maternal immune tolerance and enable gas and nutrient exchange between mother and conceptus. Pregnancy outcomes are adversely affected by gestational diabetes mellitus (GDM); however, the effects of GDM on placental formation, and subsequently fetal development, are not fully understood. In this report, streptozotocin was used to induce hyperglycemia in pregnant rats for the purpose of investigating the impact of GDM on placental formation and fetal development. GDM caused placentomegaly and placenta malformation, decreasing placental efficiency and fetal size. Elevated glucose disrupted rat trophoblast stem (TS) cell differentiation in vitro. Evidence of altered trophoblast differentiation was also observed in vivo, as hyperglycemia affected the junctional zone transcriptome and interfered with intrauterine trophoblast invasion and uterine spiral artery remodeling. When exposed to hypoxia, rats with GDM showed decreased proliferation and ectoplacental cone development on gestation day (gd) 9.5 and complete pregnancy loss by gd 13.5.  Furthermore, elevated glucose concentrations inhibited TS cell responses to hypoxia in vitro. Overall, these results indicate that alterations in placental development, efficiency, and plasticity could contribute to the suboptimal fetal outcomes in offspring from pregnancies complicated by GDM.

overall_design: Holtzman Sprague-Dawley rats were housed in an environmentally controlled facility at the University of Kansas Medical Center (KUMC) with lights on from 0600 to 2000 h and allowed free access to food and water. Female rats (8-10 weeks of age) were mated with adult male rats (>3 months of age).  Mating was assessed by inspection of vaginal lavages. The presence of sperm in the vagina was designated gd 0.5. A single dose of either streptozotocin (STZ; 45 mg/kg of body weight) or vehicle control solution (0.1 M citrate buffer, pH 4.2) was administered via tail vein injection on gd 6.5. Placentation sites were dissected on gestation day 13.5 and junctional zones were isolated. Dissected junctional zone tissues were snap-frozen in liquid nitrogen and stored at -80 degrees celsius until processing for RNA isolation.

first: Jackson

last: Nteeba

first: Regan

last: Scott

first: Mikaela

last: Simon

first: Khursheed

last: Iqbal

first: Michael

middle: J

last: Soares


title: Multi-tissue epigenetic analysis of the osteoarthritis susceptibility locus mapping to the plectin gene PLEC

city: NEWCASTLE UPON TYNE

first: John

last: Loughlin

address: Centre For Life

country: United Kingdom

institute: Newcastle University

summary: We report an immortalised CRISPR-Cas9 mesenchymal stem cell line knockdown of Plectin, we initally noted that patients with osteoarthritis association single nucelotide polymorphism rs11780978 had a correlation between genotype at rs11780978 and allelic expression imbalance, with a reduced expression of Plectin from the risk allele. We replicated the effect of the risk allele by deleting 26bp from exon 3 of Plectin to generate multiple knockdown lines to assess the molecular phenotype of Plectin knockdown in the cells. Our data suggest that Plectin knockdown was detectable in the RNA-sequencing data but at a significantly lower level compared to control. The knock-down was very efficient at the protein level, with a near total absence of plectin observed. The knock-down impacted on a range of cellular pathways. Of particular note, both the innate and the acquired immune response showed an upregulation, an effect that has been reported on previously in OA for both cartilage and synovium . Downregulation of Wnt signalling was also of interest, as this pathway is critical to cartilage homeostasis. Furthermore, there is growing evidence of an interplay between Wnt signalling and inflammation in joint tissues.

overall_design: Direct comparison of bulk RNA derived from 4 biological replicates of the same 26bp deletion compared to 3 biological controls (no guide RNA). RNA was collected 10-14 days after treatment (when monolayer confluence was achieved).

first: Antony

last: Sorial

first: Kathleen

last: Cheung


title: Large-scale production of human iPSC-derived macrophages for drug screening

city: Basel

first: Klas

last: Hatje

state: Basel

address: Grenzacherstrasse 124

country: Switzerland

institute: Roche Pharma Research and Early Development

summary: Tissue resident macrophages are key players in inflammatory processes and their activation and functionality is crucial in health and disease. Diseases associated with alterations in homeostasis or dysregulation of the innate immune system are numerous and include allergic reactions, autoimmune diseases as well as cancer. Hence, these cells are of high interest in drug development. Currently, the main sources of macrophages used in drug development are still primary cells isolated from blood or tissue, or immortalized cell lines (e.g. THP-1). Here, we describe an improved method for large-scale production of tissue resident macrophages from induced pluripotent stem cells (iPSC) in unprecedented yields. For this, iPSC-derived macrophages are thoroughly characterized to confirm their cell identity and thus their suitability for screening purposes. We demonstrate that this method to generate macrophages from iPSC overcomes the limitations of using primary cells, i.e. donor variability and limited availability of large cell numbers. Notably, the cells recapitulate key functional characteristics, including cytokine release, phagocytosis and migration. Genetic modifications can be introduced at the macrophage progenitor stage, so this methodology will facilitate the generation of reporters and gain- and loss-of-function mutants in an isogenetic background, essential assets for target validation.

overall_design: Monocytes from primary cells or macrophage progenitors from iPSCs were differentiated into M0 macrophages (MCSF), M1 macrophages (GMCSF, INFG), and M2 macrophages (MCSF, IL4). iPSC-derived monocytes were also differentiated into microglia cells (CD34, GMCSF). 5 replicates were sequenced for each cell type and 5 additional samples were sequenced for primary and iPSC monocytes in a second run.

first: Simon

last: Gutbier

first: Florian

last: Wanke

first: Nadine

last: Dahm

first: Anna

last: Rümmelin

first: Silke

last: Zimmermann

first: Klaus

last: Christensen

first: Fabian

last: Köchl

last: Rautanen

first: Barbara

last: Geering

first: Jitao

middle: D

last: Zhang

first: Markus

last: Britschgi

first: Sally

last: Cowley

first: Christoph

last: Patsch


title: Structural basis of epigenetic protection of mouse lymphoid progenitor cells by Dnmt1 [Mouse RNA-seq]

city: Freiburg im Breisgau

first: Thomas

last: Boehm

address: Stübeweg 51

country: Germany

institute: MPI-IE Freiburg

summary: DNA methylation is a universal epigenetic mechanism involved in regulation of gene expression and genome stability. The DNA maintenance methylase DNMT1 ensures that DNA methylation patterns are faithfully transmitted to daughter cells during cell division. Because DNMT1 is essential for cell viability, little is known about its tissue-specific function. To overcome this impediment, forward genetic screens were conducted in medaka and zebrafish to identify viable dnmt1 alleles. The recovered recessive missense mutations lead to distorted structures of the catalytic pocket and reduced enzymatic activities, resulting in hypomethylation of genomic DNA that specifically impairs lymphoid development. The insights gleaned from fish dnmt1 alleles enabled the structure-guided generation of a hypomorphic variant of mouse Dnmt1, which also causes cell-autonomous defects of lymphoid development. The similar phenotypes observed in three distinct species suggest that the unique sensitivity of lymphocytes to perturbations of DNA methylation patterns is an evolutionarily conserved feature of all vertebrates.

overall_design: Forward genetic screens in zebrafish and medaka identified missense mutations in maintenance DNA methylase Dnmt1 specifically affecting lymphoid development. To examine the molecular consequences of these hypomorphic mutations, DNA methylatio9n levels were assessed in selceted regions of the genomes. Based on strucrual analyses of fish Dnmt1 mutant proteins, a specific Dnmt1 mutation was also generated in mice. This led to hypomethylation of genomic DNA in lineage-negative cells of the bone marrow; additionally, the transcriptomes of several subtypes of hematopoietic progenitor cells (HSC; MPP1; MPP4) were compared by RNA-seq.

first: Norimasa

last: Iwanami

first: Katarzyna

middle: O

last: Sikora


title: Structural basis of epigenetic protection of mouse lymphoid progenitor cells by Dnmt1 (Zebrafish)

type: Methylation profiling by high throughput sequencing


title: Systems-based characterization of specialized metabolites biosynthesis in Cornus officinalis

city: Chiba

first: Mami

last: Yamazaki

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

department: Graduate School of Pharmaceutical Sciences

institute: Chiba University

summary: In this study, we performed LC-QTOFMS-based metabolomics and RNA-seq based transcriptome analysis using seven tissues of C. officinalis.

overall_design: Examination of 7 tissues of Cornus officinalis

first: Kazuki

last: Saito

first: Hideyuki

last: Suzuki

first: Michimi

last: Nakamura

first: Ryo

last: Nakabayashi

first: Tetsuya

last: Mori

first: Hidetaka

last: Kamochi

first: Megha

last: Rai

first: Amit


title: Next Generation Sequencing Facilitates Quantitative Analysis of LAP2α deficient cells and control cells

city: Beijing

first: Yiman

last: Tang

address: Zhongguancun South street

institute: Peking University

summary: Purpose: The goals of this study are to compare the mRNA profiles of LAP2α deficient human adipose-derived stem cells (hASCs) with that of control hASCs by NGS-derived RNA-seq to screen differentially expressed genes and possible involved pathways. Methods: Total RNA was isolated from hASCs transfected with shLAP2α and shNC. RNA sequencing of control and LAP2α deficient cells (each with 2 biological replicates) was performed using BGISEQ-500 sequencing system. Differentially expressed genes (DEGs) between control and LAP2α deficient cells were screened using NOISeq method. KEGG pathway enrichment analysis and Gene Ontology analysis of DEGs were subsequently conducted. Results: Using an optimized data analysis workflow, we mapped about 30 million sequence reads per sample to the cell genome. The scatterplot showed that depletion of LAP2α resulted in the upregulation of 106 genes and downregulation of 91 genes. KEGG pathway analysis suggested that LAP2α knockdown led to the change of several important pathways, including cell cycle, MAPK, and NF-κB signaling pathways that are critically related to cell growth and differentiation. Conclusions: Our results show that NGS offers a comprehensive and more accurate quantitative and qualitative evaluation of mRNA content within a cell or tissue.

overall_design: mRNA profiles of LAP2α deficient human adipose-derived stem cells and control cells were generated by deep sequencing,  in duplicate, using BGISEQ-500.

first: Yiaman

first: Wenshu

last: Ge


title: hPSC-derived Venous Angioblasts Generate Mature Fenestrated LSECs Following Intrahepatic Transplantation

city: Toronto

first: Jeff

last: Liu

state: Ontario

address: 101 College Street, RM 4-601

department: Princess Margaret Cancer Center

country: Canada

institute: University Health Network

summary: The vasculature of the liver is highly specialized and critical to organ function as well as future efforts to engineer new liver tissue.  One key vascular sub type, the liver sinusoidal endothelial cell (LSEC) lines the hepatic sinusoid mediating functions such as passing nutrients to hepatocytes, scavenging blood components, secreting FVIII, and mediating regeneration. To better understand human LSECs and to generate them from human pluripotent stem cells, we differentiated hPSCs to venous endothelial progenitors known as angioblasts, transplanted them intrahepatically in NSG newborn mice, waited 77-days, and isolated the hPSC-derived cells (DAPI-, tdRFP+) for scRNA-seq. This scRNA-seq sample represents a large number of hPSC-derived, matured hepatic- origin fibroblast and endothelial cell types.  Providing a high resolution, large sample population of cells that correlate closely to MacParland et al. (2018, Nat. Commun.) hepatic endothelial clusters providing increased cell population resolution sufficient for human LSEC zonation assessment in a tractable transplantation system.

overall_design: Primary FACS cleaned hPSC-derived hepatic engraftment population from Day 8 venous angioblast population at 77-days post transplant

first: Blair

last: Gage

first: Gary

last: Bader

first: Gordon

last: Keller


title: CTCF and Cohesin (Rad21) ChIP-seq in female mouse liver

city: Boston

first: David

middle: J.

last: Waxman

state: MA

address: 5 Cummington Mall

department: Department of Biology and Bioinformatics Program

institute: Boston University

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Sequencing files provided here include mouse liver ChIP-seq for CTCF and the cohesin subunit Rad21. These files are part of a larger study ("CTCF and Cohesin link sex-biased distal regulatory elements to sex-biased gene expression in mouse liver") where we compare CTCF and cohesin binding in male and female mouse liver as well as differences in chromatin conformation (DNA looping).

first: Bryan

last: Matthews

first: Aram

last: Shin


title: Structural basis of epigenetic protection of mouse lymphoid progenitor cells by Dnmt1 [Mouse WGBS]

first: Thomas

last: Boehm

overall_design: Forward genetic screens in zebrafish and medaka identified missense mutations in maintenance DNA methylase Dnmt1 specifically affecting lymphoid development. To examine the molecular consequences of these hypomorphic mutations, DNA methylatio9n levels were assessed in selected regions of the genomes. Based on structural analyses of fish Dnmt1 mutant proteins, a specific Dnmt1 mutation was also generated in mice. This led to hypomethylation of genomic DNA in lineage-negative cells of the bone marrow; additionally, the transcriptomes of several subtypes of hematopoietic progenitor cells (HSC; MPP1; MPP4) were compared by RNA-seq.

first: Norimasa

last: Iwanami

first: Katarzyna

last: Sikora


title: IGF2BP1 is the first positive marker for anaplastic thyroid carcinoma diagnosis [sWGS]

city: Halle (Saale)

first: Danny

last: Misiak

address: Kurt-Mothes-Str. 3a

department: Molecular Cell Biology

institute: Martin Luther University Halle-Wittenberg

summary: Anaplastic thyroid carcinomas (ATC) are rare, but represent the most lethal malignancy of the thyroid. Selective molecular markers and drivers distinguishing ATC from other thyroid carcinomas of follicular origin remain largely unknown, limiting advances in diagnosis and treatment. In a retrospective study, we analyzed gene expression in 36 ATC, 18 poorly-differentiated, 132 papillary and 55 follicular thyroid carcinoma, as well as 124 paired and un-paired normal thyroid tissues in three independent cohorts by RNA-sequencing and immunohistochemistry. RNA-sequencing data in the test cohort suggested selective ATC protein biomarkers. Evaluation of these revealed that ATCs are characterized by the de novo expression of various testis antigens, including melanoma-associated antigen A3 (MAGEA3), but most importantly the oncofetal IGF2 mRNA binding protein 1 (IGF2BP1). Shallow whole genome-sequencing essentially excluded, that IGF2BP1 upregulation results from gene copy number alterations. Immunohistochemical analyses in all three tumor cohorts confirmed selective de novo expression of IGF2BP1 protein in ATC. In sum, 75 % (27/36) of all tested ATC and 0.5 % (1/204) of poorly and well-differentiated thyroid carcinoma tissue samples were positive for IGF2BP1 protein. This indicates that IGF2BP1 protein expression identifies ATC with a diagnostic odds ratio of 612 (95 % CI: 74.6 to 5021). In addition, we found that MAGEA3 is exclusively, although less consistently upregulated in ATC, presenting with an odds ratio of 411 (95 % CI: 23.8 to 7098.7). Importantly, we provide confirmatory evidence that IGF2BP1 and MAGEA3 expression distinguishes ATC from partially differentiated thyroid carcinomas (PDTCs). IGF2BP1 furthermore identified ATC foci within low-grade follicular thyroid carcinoma. In conclusion, IGF2BP1 represents the most promising single-gene marker available for ATC, followed by MAGEA3, improving on current techniques. Robust markers are essential to help distinguish this high-grade malignancy from other thyroid carcinomas, to guide surgical decision making, therapy and post-resection/therapy monitoring strategies.

overall_design: Shallow Whole Genome sequencing of ATC samples.

first: Jacob

last: Haase

first: Nikolaos

last: Pazaitis

first: Marcus

last: Bauer

first: Juliane

last: Braun

first: Rebecca

last: Pötschke

first: Alexander

last: Mensch

first: Jessica

last: Bell

first: Henning

last: Dralle

first: Udo

last: Siebolts

first: Claudia

last: Wickenhauser

first: Kerstin

last: Lorenz

first: Stefan

last: Hüttelmaier


city: Lexington

first: Jeramiah

middle: James

last: Smith

state: KY

address: 101 Morgan Building

department: Biology

institute: University of Kentucky

summary: 10X sequencing of immune cells and limb regeneratesImmune cells are known to be critical for successful limb regeneration in the axolotl (Ambystoma mexicanum), but many details regarding their identity, behavior, and function are yet to be resolved. We isolated peripheral leukocytes from the blood of adult axolotls and then created two samples for single-cell sequencing: 1) peripheral leukocytes (N = 7,889) and 2) peripheral leukocytes with presumptive macrophages from the intraperitoneal cavity (N = 4,998). Using k-means clustering, we identified 6 cell populations from each sample that presented gene expression patterns indicative of erythrocyte, thrombocyte, neutrophil, B-cell, T-cell, and myeloid cell populations. A seventh, presumptive macrophage cell population was identified uniquely from sample 2. We then isolated cells from amputated axolotl limbs at 1 and 6 days post-amputation (DPA) and performed single cell sequencing (N = 8,272 and 9,906 cells respectively) to identify immune and non-immune cell populations. Using k-means clustering, we identified 8 cell populations overall, with the majority of cells expressing erythrocyte specific genes. Even though erythrocytes predominated, we used an unbiased approach to identify infiltrating neutrophil, macrophage, and lymphocyte populations at both time points. Additionally, populations expressing genes for epidermal cells, fibroblast-like cells, and endothelial cells were also identified. Consistent with results from previous experimental studies, neutrophils were more abundant at 1 DPA than 6 DPA, while macrophages and non-immune cells exhibited inverse abundance patterns. Of note, we identified a small population of fibroblast-like cells at 1 DPA that was represented by considerably more cells at 6 DPA. We hypothesize that these are early progenitor cells that give rise to the blastema. The enriched gene sets from our work will aid future single-cell investigations of immune cell diversity and function during axolotl limb regeneration.

overall_design:
We isolated peripheral leukocytes from the blood of adult axolotls and then created two samples for single-cell sequencing: 1) peripheral leukocytes (N = 7,889) and 2) peripheral  leukocytes with presumptive macrophages from the intraperitoneal cavity (N = 4,998).  A seventh, presumptive macrophage cell population was identified uniquely from sample 2. We then isolated cells from amputated axolotl limbs at 1 and 6 days post-amputation (DPA) and performed single cell sequencing (N = 8,272 and 9,906 cells respectively) to identify immune and non-immune cell populations.
Please note that the reference assembly is not available in any public database.  A copy from (the authors of the referenced PMID  30262634 paper) is included  to be completely clear as to the specific reference that was used for mapping.

first: Katherine

last: Rodgers

first: S

last: Voss


title: Effect of estradiol, zearalenone and apigenin on ER-positive breast cancer cells MCF-7

first: Frédéric

last: Chalmel

summary: To understand the molecular effect of E2 and the dietary compound zearalenone and apigenin on MCF-7 proliferation. Results provide insight into the pathway involved by these dietary compounds on MCF-7 proliferation.

overall_design: Effect on MCF-7 gene expression of E2, zearalenone and apigenin was assessed after 24 h of exposure. Four independent replicates for each treatment were performed.

first: Sylvain

last: Lecomte

first: Florence

last: Demay

first: Thu

middle: H

last: Pham

first: Solenn

last: Moulis

first: Theo

last: Efstathiou

first: Frederic

first: Farzad

last: Pakdel


title: RNA-Seq analysis of Pold3 inducible-knockout embryonic stem cells

city: Tianjin

first: zhongcheng

last: zhou

state: Tianjin

address: 94 Weijin Road, Nankai District

department: Cell biology and Genetics

institute: Nankai Univerisity

summary: To investigate the molecular changes and mechanism during loss of Pold3, Pold3 induble-knockout embryonic stem cells at different passages were subjected to RNA-seq analysis. We found that aberrant cell cycle and defective DNA repair were induced by loss of Pold3.

overall_design: Detection of changes in transcriptome levels at diffrerent passages (P0, P1 and P2)

first: Zhongcheng

last: Zhou

first: Lin

last: Liu


title: Characterization of novel zinc finger protein ZNF414 DNA binding and gene regulatory activity by integrative analysis of ChIP-exo, ATAC-seq and RNA-seq data (ChIP-exo)

status: Public on Jul 06 2020

city: Tampere University

first: Matti

last: Nykter

state: Please select one below:

address: MET / Computational biology

country: Finland

institute: Tampere University

summary: Transcription factor binding to DNA is a central mechanism regulating cellular gene expression. Thus, thorough characterization of this process is essential for understanding cellular biology in both health and disease. In this study, we combined data from three sequencing-based methods to unravel the DNA binding function of the novel zinc finger protein ZNF414 in cells representing two tumor types. ChIP-exo served to map protein binding sites in the genome, ATAC-seq allowed identification of open chromatin, and RNA-seq examined the transcriptome.

overall_design: We characterized ZNF414 DNA binding in human pancreatic tumor cell line Hs700T and in human breast cancer cell line MCF-7 using ChIP-exo with an antibody against V5 tag. Cells were transfected with pcDNA6.2/EmGFP-Bsd/V5-DEST-ZNF414 plasmid for overexpression of V5-tagged ZNF414 48 hours before sample collection and two biological replicates were used per cell line.

first: Alejandra

last: Rodriguez-Martinez

first: Elisa

last: Vuorinen

first: Anastasia

last: Scherban

first: Joonas

last: Tuominen

first: Nina

last: Rajala

first: Anne

last: Kallioniemi


title: Characterization of novel zinc finger protein ZNF414 DNA binding and gene regulatory activity by integrative analysis of ChIP-exo, ATAC-seq and RNA-seq data (ATAC-seq)

overall_design: We characterized accessible chromatin in human pancreatic tumor cell line Hs700T and in human breast cancer cell line MCF-7. Cells were transfected with pcDNA6.2/EmGFP-Bsd/V5-DEST-ZNF414 plasmid for overexpression of V5-tagged ZNF414 48 hours before sample collection. One sample per cell line.


title: Transcriptome analysis of bFGF and WNT3A treated stem cells isolated from human exfoliated deciduous teeth

city: Pathumwan

first: Thanaphum

last: Osathanon

state: Bangkok

address: 34 Henri-dunant Rd. Wongmai

country: Thailand

institute: Chulalongkorn University

summary: bFGF regulated stemness maintenance and cell differentiation in stem cells isolated from human exfoliated deciduous teeth (SHEDs). In addition, WNT3A is also shown to regulate osteogenic differentiation in mesenchymal stem cells. Here, transcriptome analysis of bFGF and WNT3A treated SHEDs was evaluated using a high throughput RNA sequencing technique. Results demonstrated that bFGF regulated genes related to cell cycle control and cell proliferation in SHEDs.

overall_design: Human periodontal ligament cells were treated with the computerized controlled compressive force application apparatus. For static compressive force stimuli, cells were loaded with 1.5 g/cm2 of force continuously while cells were loaded with 1.5 g/cm2 with the loading frequency of 0.23 Hz. All loading was performed in serum free condition and the total RNA was collected at 24 h after loading. Differential gene expression was determined using RNA sequencing technique.

first: Nunthawan

last: Nowwarote

first: Jeeranan

last: Manokawinchoke

first: Waleerat

last: Sukarawan


title: IGF2BP1 is the first positive marker for anaplastic thyroid carcinoma diagnosis

first: Danny

last: Misiak

type: Non-coding RNA profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Distinct transcriptional profiles of the female, male and finasteride-induced feminized male anogenital region in rat fetuses

city: Rennes

first: Frédéric

last: Chalmel

address: 9 avenue du Pr. Léon Bernard

institute: Inserm U1085-Irset

summary: A short anogential distance (AGD) in males is a marker for incomplete masculinization and a predictor of adverse effects on male reproductive health. For this reason, AGD is used to assess the endocrine disrupting potential of chemicals for risk assessment purposes. The molecular mechanisms underpinning this chemically induced shortening of the AGD, however, remains unclear. Although it is clear that AR-mediated signaling is critical, evidence also suggest the involvement of other signaling pathways. This study present the first global transcriptional profile of the anogenital region in male rats with chemically induced short AGD, also including comparison to normal male and female control animals. The anti-androgenic drug Finasteride (10mg/kg bw/day) was used to induce short AGD by exposing pregnant rats at gestation days (GD) 7-21. The AGD was 37% shorter in exposed male offspring compared to control males. Transcriptomics analysis on anogenital tissues at GD21 revealed a sexually dimorphic transcriptional profile. More than 350 genes were found to be differentially expressed between the three groups. The expression pattern of four genes of particular interest (Esr1, Padi2, Wnt2 and Sfrp4) was verified by RT-qPCR analyses. Our transcriptomics profiles provide a stepping-stone for future studies aimed at characterizing the molecular events governing development of the anogenital region, as well as describing the detailed Adverse Outcome Pathways for short AGD; an accepted biomarker of endocrine effects  for chemical regulation.

overall_design: We sought to elucidate the transcriptional profile of the anogenital region of male rats with chemically induced short AGD. We compared this to the transcriptional profile of control males and control females to investigate whether induction of short AGD resulted in a feminization of the anogenital region at the transcriptional level.

first: C

last: Schwartz

first: A

last: Vinggaard

first: S

last: Christiansen

first: T

last: Darde

first: F

last: Svingen


title: 4C-seq analysis of interactions with promoters and enhancers nearby five sex-specific genes, in male and female mouse liver

first: David

last: Waxman

summary: Sequencing files provided here include 4C-seq experiments for a total of 6 viewpoints neighboring 5 highly sex-biased genes in mouse liver. These files are part of a larger study ("CTCF and Cohesin link sex-biased distal regulatory elements to sex-biased gene expression in mouse liver"), where we compare CTCF and cohesin binding in male and female mouse liver as well as differences in chromatin conformation (DNA looping).

overall_design: 4C-seq for viewpoints neighboring highly sex-biased genes

first: Bryan

last: Matthews


title: CD49a+ and CD49a- NK cells from human livers

city: Hefei

first: Cheng

last: Sun

state: Anhui

address: 17, Lujiang Road

department: the First Affiliated Hospital of USTC

institute: University of Science and Technology of China

summary: Purified NK cells from human liver tissues were first enriched by MACS using NK Cell Isolation Kit (Miltenyi Biotec, German) and CD49a+/- hepatic NK cells were isolated by FACS Aria cell sorter (BD Biosciences, United States) to attain a purity greater than 95%.

overall_design: three healthy donors were pooled for each cell type


title: CTCF and Cohesin link sex-biased distal regulatory elements to sex-biased gene expression in mouse liver


title: Genome-wide estrogen receptor-α binding and action in human endometrial stromal cells [ChIP-Seq]

city: Chicago

first: Serdar

last: Bulun

state: Illinois

address: 303 E. Superior, Lurie Research Building

department: Obstetrics and Gynecology

institute: Northwestern University

summary: Endometrial estrogen receptor-α (ESR1) is indispensable for epithelial and stromal proliferation and differentiation during decidualization, yet the gene targets of estradiol (E2) / ESR1 in human stromal cells and associated mechanisms remain unknown. In this study, we characterized global E2-ESR1‒dependent transcriptomic changes and ESR1 recruitment to chromatin. Human endometrial stromal cells were isolated from 4 premenopausal women for primary cell culture. Genome-wide RNA expression by RNA-sequencing was compared in endometrial stromal cells with or without siRNA knockdown of ESR1 in the presence or absence of E2 (n=2). Genome-wide recruitment of ESR1 to chromatin was assessed by chromatin immunoprecipitation sequencing using an antibody against ESR1 was performed to examine binding to target genes (n=1).

overall_design: Genome-wide ESR1 binding profiling in human endometrial stromal cells with estradiol treatment using ChIP-Seq

first: Bahar

last: Yilmaz

first: Christia

last: Sison

first: Ping

last: Yin


title: Transcriptome analysis of male strobilus induction by gibberellic acid treatment in Cryptomeria japonica

city: Koshi

first: Manabu

last: Kurita

institute: Kyushu Regional Breeding Office, Forest Tree Breeding Center, Forestry and Forest Products Research Institute

summary:
Investigation of gene expression dynamics associated with male strobilus induction in Cryptomeria japonica in response to gibberellic acid (GA) treatment.
[SUBMITTER_CITATION]:
Transcriptome Analysis in Male Strobilus Induction by Gibberellin Treatment in Cryptomeria japonica D. Don
Manabu Kurita 1,?, Kentaro Mishima 2,*,? , Miyoko Tsubomura 2, Yuya Takashima 2 , Mine Nose 2, Tomonori Hirao 2 and Makoto Takahashi 2
Forests 2020, 11(6), 633

overall_design: GA-treated and control C. japonica individuals were sampled before at the time of GA treatment (i.e. -1d), and seven times thereafter (i.e. at 0 h, 3 h, 1 d, 3 d, 1 w, 2 w, 4 w and 6 w). Three different plus-trees were used as biological replicates (total 48 samples).

first: Kentaro

last: Mishima


title: Genome-wide estrogen receptor-α binding and action in human endometrial stromal cells


title: Structural basis of epigenetic protection of mouse lymphoid progenitor cells by Dnmt1 (Medaka)

first: Thomas

last: Boehm

summary: DNA methylation is a universal epigenetic mechanism involved in regulation of gene expression and genome stability. The DNA maintenance methylase DNMT1 ensures that DNA methylation patterns are faithfully transmitted to daughter cells during cell division. Because DNMT1 is essential for cell viability, little is known about its tissue-specific function. To overcome this impediment, forward genetic screens were conducted in medaka and zebrafish to identify viable dnmt1 alleles. The recovered recessive missense mutations lead to distorted structures of the catalytic pocket and reduced enzymatic activities, resulting in hypomethylation of genomic DNA that specifically impairs lymphoid development. The insights gleaned from fish dnmt1 alleles enabled the structure-guided generation of a hypomorphic variant of mouse Dnmt1, which also causes cell-autonomous defects of lymphoid development. The similar phenotypes observed in three distinct species suggest that the unique sensitivity of lymphocytes to perturbations of DNA methylation patterns is an evolutionarily conserved feature of all vertebrates.

overall_design: Forward genetic screens in zebrafish amd medaka identified missense mutations in maintenance DNA methylase Dnmt1 specifically affecting lymphoid development. To examine the molecular consequences of these hypomorphic mutations, DNA methylatio9n levels were assessed in selceted regions of the genomes. Based on strucrual analyses of fish Dnmt1 mutant proteins, a specific Dnmt1 mutation was also generated in mice. This led to hypomethylation of genomic DNA in lineage-negative cells of the bone marrow; additionally, the transcriptomes of several subtypes of hematopoietic progenitor cells (HSC; MPP1; MPP4) were compared by RNA-seq.

first: Norimasa

last: Iwanami

first: Kohei

last: Takeshita

first: Divine

middle: F

last: Lawir

first: Isao

last: Suetake

first: Shoji

last: Tajima

first: Katarzyna

last: Sikora

first: Ines

last: Trancoso

first: Connor

last: O'Meara

first: Iliana

last: Siamishi

last: Clapes

first: Eirini

last: Trompouki

first: Yousuke

last: Takahama

first: Makoto

last: Furutani-Seiki

first: Hisato

last: Kondoh

first: Yasushige

last: Yonezawa

first: Michael

last: Schorpp


title: Structural basis of epigenetic protection of mouse lymphoid progenitor cells by Dnmt1

last: Boehm

sample_organism:
Danio rerio
Oryzias latipes
Mus musculus


title: Hematopoietic stem cells acquire survival advantage by loss of RUNX1 methylation identified in familial leukemia

city: Singapore

first: Takayoshi

last: Matsumura

address: 14 Medical Drive #12-01

department: CSI Singapore

country: Singapore

institute: National University of Singapore

summary: RUNX1 is among the most frequently mutated genes in human leukemia, and the loss or dominant-negative suppression of RUNX1 function is found in myelodysplastic syndrome and acute myeloid leukemia (AML). However, how post-translational modifications (PTMs) of RUNX1 affect its in vivo function and whether PTM dysregulation of RUNX1 can cause leukemia are largely unknown. We performed targeted deep sequencing on a family with 3 occurrences of AML and identified a novel RUNX1 mutation R237K. The mutated R237 residue is a methylation site by PRMT1 and loss of methylation has been reported to impair transcriptional activity of RUNX1 in vitro. To explore biological significance of RUNX1 methylation in vivo, we utilized RUNX1 R233K/R237K double mutant mice, in which 2 arginine-to-lysine mutations precluded RUNX1 methylation. Genetic ablation of RUNX1 methylation led to loss of quiescence and expansion of hematopoietic stem cells (HSCs), and changed the genomic and epigenomic signature of phenotypic HSCs to a poised progenitor state. Further, loss of RUNX1 R233/R237 methylation suppressed endoplasmic reticulum stress-induced unfolded protein response genes including Atf4, Ddit3, and Gadd34, radiation-induced p53 downstream genes, Bbc3, Pmaip1, and Cdkn1a, and subsequent apoptosis in HSCs. Mechanistically, ATF4 was identified as a direct transcriptional target of RUNX1. Collectively, defects in RUNX1 methylation in HSCs confer resistance to apoptosis and survival advantage under stress conditions, a hallmark of a pre-leukemic clone which may predispose affected individuals to leukemia. Our study will lead to a better understanding of how dysregulation of PTMs can contribute to leukemogenesis.

overall_design: RNA-seq of control and RUNX1 methylation-deficient CD34-Flt3-Lin-Sca1+cKit+ long-term hematopoietic stem cells (LT-HSCs) in triplicate


title: Hematopoietic stem cells acquire survival advantage by loss of RUNX1 methylation identified in familial leukemia [ATAC-seq]

overall_design: ATAC-seq of control and RUNX1 methylation-deficient CD34-Flt3-Lin-Sca1+cKit+ long-term hematopoietic stem cells (LT-HSCs) in duplicate


title: Gene expression and splicing regulation by hnRNPU in skeletal muscle

first: Ronald

last: Neppl

address: 60 Fenwood Rd.

department: Orthopedic Surgery

institute: Brigham and Women's Hospital, Harvard Medical School

summary: Hnrnpu is the largest member of the heterogenous nuclear ribonucleoprotein family of RNA binding proteins. Hnrnpu is involved in pre-mRNA alternative splicing regulation. We used high throughput sequencing to determine how Hnrnpu regulates skeletal muscle physiology.

overall_design: Gastrocnemius muscles of WT and Hnrnpu muscle specifc knockout mice were harvested for total RNA isolation at 3 months of age.

first: R


title: Differentially expressed genes during pancreatic progenitor cell differentiation and proliferation

city: Kyoto

first: Kenji

last: Osafune

address: Shogoin-Kawahara-cho 53

department: Center for iPS Cell Research and Application (CiRA)

institute: Kyoto University

summary: A small molecule, AT7867, is a proliferation-promoting factor of human iPS cell-derived pancreatic progenitor cells; yet how it promotes cell proliferation is not known. AT7867 induces proliferation of pancreatic progenitor cells, but not that of definitive endoderm cells or primitive gut tube cells suggesting a cell type-specific effect. Our aim is to perform RNA-seq of cells obtained during a three stage differentiation of hiPS cells to the endodermal cell lineage in addition to pancreatic progenitor cells treated with AT7867, and analyze signaling pathways altered by the compound treatment.

overall_design: The overall goal of this study was to examine gene expression changes during pancreatic progenitor cell differentiation and proliferation. Specifically, mRNA profiles of human iPS cells, hiPS cell-derived differentiated cells (definitive endoderm cells, primitive gut tube cells, and pancreatic progenitor cells), and pancreatic progenitor cells treated with a proliferation inducer, AT7867 in two different medium conditions were generated in triplicate.

first: Azuma

last: Kimura

first: Taro

last: Toyoda


title: Characterization of novel zinc finger protein ZNF414 DNA binding and gene regulatory activity by integrative analysis of ChIP-exo, ATAC-seq and RNA-seq data

first: Matti

last: Nykter


title: Characterization of novel zinc finger protein ZNF414 DNA binding and gene regulatory activity by integrative analysis of ChIP-exo, ATAC-seq and RNA-seq data (RNA-seq)

overall_design: We characterized the effect of ZNF414 expression on genomewide gene expression levels in human pancreatic cell line Hs700T and in human breast cancer cell line MCF-7. For this, we silenced ZNF414 gene by siRNA transfection in these cells and performed RNA-seq on transfected and control (Luciferase siRNA-transfected) cells at 12h and 24h timepoints.

first: Alejandra

last: Rodriguez-Martinez

first: Elisa

last: Vuorinen

first: Anastasia

last: Scherban

first: Joonas

last: Tuominen

first: Nina

last: Rajala

first: Anne

last: Kallioniemi


title: Genome-wide estrogen receptor-α binding and action in human endometrial stromal cells [RNA-Seq]

first: Serdar

last: Bulun

overall_design: Genome-wide transcriptome mapping of human endometrial stromal cells with and without ESR1 knockdown in the presense or absence of estradiol treatment

first: Bahar

last: Yilmaz

first: Christia

last: Sison

first: Ping

last: Yin


title: PRC2-mediated H3K27me3 regulates iron homeostasis genes in the shoots of Arabidopsis thaliana

city: Amherst

first: Jeeyon

last: Jeong

address: 25 East Drive

institute: Amherst College

summary: H3K27me3 contributes to the regulation of iron homeostasis genes in shoots

overall_design: RNA-seq analysis of transcripts from wild type and clf shoot tissue grown under iron sufficient and deficient conditions


title: Gene expression classifiers identify bacterial and viral etiologies of acute respiratory illness in Sri Lanka.

city: Durham

first: Sunil

last: Suchindran

state: NC

address: 101 Science Drive

department: Center for Applied Genomics and Precision Medicine

institute: Duke University

summary: Previously developed gene expression signatures had high predictive accuracy at distinguishing viral and bacterial infection in a South Asian cohort with ARI caused by typical and atypical pathogens.

overall_design: Application of transcriptomic signatures in a South Asian cohort with viral and bacterial infections

first: L

middle: G

last: Tillekeratne

first: Emily

middle: R

last: Ko

first: Elizabeth

last: Petzold

first: Champica

last: Bodinayake

first: Ajith

last: Nagahawatte

first: Vasantha

last: Devasiri

first: Ruvini

last: Kurukulasooriya

first: Bradly

middle: P

last: Nicholson

first: Micah

middle: T

last: McClain

first: Thomas

middle: W

last: Burke

first: Ephraim

last: Tsalik

first: Ricardo

last: Henao

first: Geoffrey

last: Ginsburg

first: Megan

last: Reller

first: Christopher

last: Woods


title: Fibronectin deficiency in CCM pathogenesis

city: Hamburg

first: Matthias

last: Rath

address: Am Kaiserkai 1

department: Institute of Molecular Medicine

institute: MSH Medical School Hamburg


title: Expression of ncRNAs on the DLK1-DIO3 locus is associated with basel and mesenchymal phenotype in breast epithelial progenitor cells

city: Reykjavik

first: Zuzana

last: Budkova

address: Vatnsmyrarvegi 14

department: BioMedical Center, School of Health Sciences

country: Iceland

institute: University of Iceland

summary: Epithelial-to-mesenchymal transition (EMT) and its reversed process mesenchymal-to-epithelial transition (MET) play a critical role in epithelial plasticity during development and cancer progression. Among important regulators of these cellular processes are non-coding RNAs (ncRNAs). The imprinted DLK1-DIO3 locus, containing numerous maternally expressed ncRNAs including the lncRNA maternally expressed gene 3 (MEG3) and a cluster of over 50 miRNAs, has been shown to be a modulator of stemness in embryonic stem cells and in cancer progression, potentially through the tumor suppressor role of MEG3. In this study we analyzed the expression pattern and functional role of ncRNAs from the DLK1-DIO3 locus in epithelial plasticity of the breast. We studied their expression in various cell types of breast tissue and revisit the role of the locus in EMT/MET using a breast epithelial progenitor cell line (D492) and its isogenic mesenchymal derivative (D492M). Marked upregulation of ncRNAs from the DLK1-DIO3 locus was seen after EMT induction in two cell line models of EMT. In addition, the expression of MEG3 and the maternally expressed ncRNAs was higher in stromal cells compared to epithelial cell types in primary breast tissue. We also show that expression of MEG3 is concomitant with the expression of the ncRNAs from the DLK1-DIO3 locus and its expression is therefore likely indicative of activation of all ncRNAs at the locus. MEG3 expression is correlated with stromal markers in normal tissue and breast cancer tissue and negatively correlated with the survival of breast cancer patients in two different cohorts. Overexpression of MEG3 using CRISPR activation in a breast epithelial cell line induced partial EMT and enriched for a basal-like phenotype. Conversely, knock down of MEG3 using CRISPR inhibition in a mesenchymal cell line reduced the mesenchymal and basal-like phenotype of the cell line. In summary our study shows that maternally expressed ncRNAs are markers of EMT and suggests that MEG3 is a novel regulator of EMT/MET in breast tissue. Nevertheless, further studies are needed to fully dissect the molecular pathways influenced by non-coding RNAs at the DLK1-DIO3 locus in breast tissue.

overall_design: A total of ten samples were analyzed: Five replicates from D492M KD ctrl and five replicates from D492M KD MEG3. However, sample no. 5 from D492M KD ctrl was omitted due to high deviation from the outher four samples.

first: Snaevar

last: Sigurdsson


title: The “effect” of T staging on resectable metastatic liver colorectal cancer/Early T stage is associated with poor prognosis in patients with metastatic liver colorectal cancer

city: Hangzhou

first: Shu

last: Zheng

state: Zhejiang

address: 88 Jiefang Road,

institute: Zhejiang University

summary: a considerable portion of patients with early T stage who were supposed to have a low distant metastatic probability, were diagnosed with metastatic cancer. Our study aims to evaluate characteristics of T1M1patients via cDNA microarray.

overall_design: Primary colorectal tumors in 19 patients with from resectable CRLM (2 patients with T2N0M1 and 17 patients with T3/4N0M0) were obtained for microarrary

first: Lunpo

last: Wu

first: Jianfei

last: Fu


title: Tumor specific methylome in Chinese high-grade serous ovarian cancer characterized by gene expression profile

status: Public on Jul 04 2020

city: tianjin

first: kexin

last: chen

address: huanhu road, tiyuanbei, hexi district

institute: Tianjin Medical University Cancer Institute and Hospital

summary: Altered methylation patterns in HGSOC were further characterized by gene expression profiles. By integrating DNA methylation and mRNA expression data, we identified genes with negative and positive correlation, aiming to identify molecular drivers and pathways that are HGSOC-related.

overall_design: Differential expression was analyzed in group (tumors vs normals) using the R package PoissonSeq with default parameters and defined by a significance criterion of 0.01 and 1.5 fold changes

first: Fangfang

last: Song

first: Baifeng

last: Zhang


title: Analysis of miR-203 transient over-expression on stemness potential of pluripotent cells

status: Public on Jul 05 2020

city: Madrid

first: Maria

last: Salazar

state: Comunidad de Madrid

address: C/ Melchor Fernández Almagro, 3

department: Molecular Oncology

country: Spain

institute: CNIO

summary: The balance between pluripotency and differentiation is critical during development and regeneration. miR-203 is a microRNA previously involved in differentiation of different tissues as well as in tumor suppression in multiple malignancies. We show here that miR-203 is able to promote differentiation of embryonic stem (ES) and induced pluripotent stem (iPS) cells without decreasing pluripotency. We have observed that transient expression of miR-203 significantly improves the efficiency of ES/iPS cells in the generation of chimeras and tetraploid complementation assays, in addition to inducing complex embryo-like structures when these pluripotent cells are injected in mice. In the present RNA seq, we intend to demonstrate that transient over-expression of miR-203 make pluripotent cells (IPSCs) more similar to ES cells, also in terms of their transcriptomic profile.

overall_design: The general idea was to analyze the transcriptomic proximity between miR-203 transiently-overexpressing IPSCs and ESCs. In this RNA seq, the gene expression of different clones was tested. Three independent pools of clones from three different experiments were represented in ES_WT_C1, C2 and C3. Three independent pools of clones from three independent experiments were represented in samples iPS_WT_C1, C2 and C3. The same applies to KI_DOX_C1, C2, C3, C4 (miR-203 cKI iPS cells transiently treated with DOX to induce miR-203 over-expression). The sample iPS_WT_MIMICS represents a pool of two independent experiments in which WT iPS cells were transiently transfected using 3p/5p miR-203 mimics. The pairs KI-N5 and KI_N5+DOX; KI_N9 and KI_N9+DOX represents two pairs of clones in which N5 and N9 have never been treated with DOX and the very same clones N5+DOX and N9+ DOX have been transiently treated with DOX to over-express miR-203 for 5 days.

first: María

last: Salazar Roa

first: Osvaldo

last: Graña

first: Marcos

last: Malumbres


title: Dnmt1 deletion in PV-positive cortical interneurons of 6 month and 18 month old mice [RNA-seq]

city: Goettingen

first: Gabriela

last: Salinas

state: Lower-Saxony

address: Kreuzbergring 57

department: Department of Pathology

institute: Universitaetsmedizin Goettingen

summary: The emerging role of epigenetic mechanisms like DNA methylation executed by DNA methyltransferases (DNMTs) in synaptic function irrevocably raises the question for the targeted subcellular processes and mechanisms. We here sequenced FAC-sorted PVergic interneurons of 6 month and 18month old PV-Cre/tdTomato/Dnmt1loxp wildtype and knockout mice and performed RNA-Seq.

overall_design: 8 samples in total; WT and KO condition of 6 month and 18 month old mice, each in 2 technical replicates; cells were pooled from 6 individual wild type 6 individual knockout mice. Per mice both hemispheres were FAC-sorted independently and the cells from one hemisphere were used for RNA extraction while the other one were used for DNA extraction.

first: Daniel

last: Pensold

first: Anne

last: Hahn

first: Thomas

last: Lingner

last: Salinas-Riester

first: Judit

last: Symmank

first: Katrin

last: Schubert

first: Tomas

last: Pieler

first: Geraldine

last: Zimmer


title: Wild mice with different social network sizes vary in brain gene expression

city: Orange

first: Patricia

middle: C

last: Lopes

state: CA

address: 450 N Center St

department: Biological Sciences

institute: Chapman University

summary:
Background
Appropriate social interactions influence animal fitness by impacting several processes, such as mating, territory defense, and offspring care. Many studies shedding light on the neurobiological underpinnings of social behavior have focused on nonapeptides (vasopressin, oxytocin, and homologues) and on sexual or parent-offspring interactions. Furthermore, animals have been studied under artificial laboratory conditions, where the consequences of behavioral responses may not be as critical as when expressed under natural environments, therefore obscuring certain physiological responses. We used automated recording of social interactions of wild house mice outside of the breeding season to detect individuals at both tails of a distribution of egocentric network sizes (characterized by number of different partners encountered per day). We then used RNA-seq to perform an unbiased assessment of neural differences in gene expression in the prefrontal cortex, the hippocampus and the hypothalamus between these mice with naturally occurring extreme differences in social network size.
Results
We found that the neurogenomic pathways associated with having extreme social network sizes differed between the sexes. In females, hundreds of genes were differentially expressed between animals with small and large social network sizes, whereas in males very few were. In males, X-chromosome inactivation pathways in the prefrontal cortex were the ones that better differentiated animals with small from those with large social network sizes animals. In females, animals with small network size showed up-regulation of dopaminergic production and transport pathways in the hypothalamus. Additionally, in females, extracellular matrix deposition on hippocampal neurons was higher in individuals with small relative to large social network size.
Conclusions
Studying neural substrates of natural variation in social behavior in traditional model organisms in their habitat can open new targets of research for understanding variation in social behavior in other taxa.

overall_design: RNA-seq dataset from three brain regions (hypothalamus, prefrontal cortex and hippocampus) from wild mice presenting consistently extreme (large/high or small/low) social network sizes. Males and females from each type are included.

first: Barbara

last: König


title: Dnmt1 deletion in PV-positive cortical interneurons of 6 month and 18 month old mice

summary: The emerging role of epigenetic mechanisms like DNA methylation executed by DNA methyltransferases (DNMTs) in synaptic function irrevocably raises the question for the targeted subcellular processes and mechanisms. We here sequenced FAC-sorted PVergic interneurons of 6 month and 18month old PV-Cre/tdTomato/Dnmt1loxp wildtype and knockout mice and performed RNA-Seq and MeDIP-Seq.

overall_design: Refer to individual Series


title: Effect of Dnmt3a/3b transient silencing in WT iPSCs and comparison with miR-203 tKI iPSCs

summary: The balance between pluripotency and differentiation is critical during development and regeneration. miR-203 is a microRNA previously involved in differentiation of different tissues as well as in tumor suppression in multiple malignancies. We have shown that miR-203 is able to promote differentiation of embryonic stem (ES) and induced pluripotent stem (iPS) cells without decreasing pluripotency. We have observed that transient expression of miR-203 significantly improves the efficiency of ES/iPS cells in the generation of quimeras and tetraploid complementation assays, in addition to inducing complex embryo-like structures when these pluripotent cells are injected in mice. Mechanistically, we have shown that miR-203 mediates such effects, at least in part, by modulating the levels of de novo DNA methyltransferases. In the present RNAseq we have transiently silenced the levels of DNMT3a/3b in order to compare their transcriptomic profile with that observed in PSCs transiently exponed to miR-203.

overall_design: The general idea was to analyze the transcriptomic profile of WT iPSCs transiently transfected with siRNA control, Dnmt3a,Dnmt3b or both and compare their transcriptomic signatures with those observed in mir-203 tKI iPSCs. As described previously, miR-203 tKI IPSCs were transiently induced by doxycycline treatment for 5 days, and samples were collected 2 weeks after dox withdrawal.


title: Genome-wide CRISPR screen of rituximab induced cell depletion

city: Aarhus

first: Jacob

middle: GIehm

last: Mikkelsen

address: Høegh-Guldbergs Gade 10

department: Biomedicin

country: Denmark

institute: Aarhus University

summary: We screened the genome of OCI-Ly-7, a Diffuse Large B-cell Lymphoma cell line, for genes mediating the response to rituximab treatment. We utilized the genome-wide library GeCKO v2 (A+B) from Feng Zhang. (Addgene#1000000049)

sample_organism:
Homo sapiens
unidentified plasmid

platform_organism:
Homo sapiens
unidentified plasmid

overall_design: In addition to mock treated sample, we subjected cells to two different rituximab conditions. RTX-SEC (short exposure to CDC conditions, SEC, followed by non-CDC conditions for a total of 14 days) and RTX-REC (repeated exposure to CDC conditions, REC, maintained for 21 days). The rationale was to expose cells in the RTX-SEC group shortly to CDC conditions followed by non-CDC conditions, whereas cells in the RTX-REC group were repeatedly subjected to conditions supporting CDC

first: E

last: Thomsen

first: J


title: Dnmt1 deletion in PV-positive cortical interneurons of 6 month and 18 month old mice [meDIP-seq]

summary: The emerging role of epigenetic mechanisms like DNA methylation executed by DNA methyltransferases (DNMTs) in synaptic function irrevocably raises the question for the targeted subcellular processes and mechanisms. We here sequenced FAC-sorted PVergic interneurons of 6 month and 18month old PV-Cre/tdTomato/Dnmt1loxp wildtype and knockout mice and performed MeDIP-Seq.

overall_design: 4 samples in total; WT and KO condition of 6 month and 18 month old mice; cells were pooled from 6 individual wild type 6 individual knockout mice. Per mice both hemispheres were FAC-sorted independently and the cells from one hemisphere were used for RNA extraction while the other one were used for DNA extraction.


title: mRNA expression changes in CCM3-deficient endothelial cells

first: Matthias

last: Rath

summary: Loss-of-function variants in CCM3/PDCD10 predispose to cerebral cavernous malformations (CCMs) that are vascular lesion of the central nervous system. Using CRISPR/Cas9 genome editing and RNA sequencing, we have shown that long-term inactivation of CCM3 in human endothelial cells dysregulates fibronectin expression and thus impairs the assembly of a functional fibronectin matrix by endothelial cells.

overall_design: Examination of the mRNA expression profile of 3 wild-type CI-huVEC lines and 3 clonal CCM3-knockout CI-huVEC lines.

first: Konrad

last: Schwefel

first: Stefanie

last: Spiegler

first: Bettina

last: Kirchmaier

middle: K

last: Dellweg

first: Christiane

last: Much

first: Jan

last: Pané-Farré

first: Tim

last: Strom

first: Katharina

last: Riedel

first: Ute

last: Felbor


title: The fibronectin mediated endothelial rescue effect in CCM3-deficient cells is independent from acute mRNA changes

summary: Using CRISPR/Cas9 genome editing and RNA sequencing, we demonstrate that fibronectin supplementation restores the impaired spheroid formation and attenuates actin stress fiber assembly in CCM3-/- endothelial cells independently of acute gene expression differences. In particular, there was no significant downregulation of well-known key players in CCM pathogenesis like KLF2 or KLF4.

overall_design: Examination of the mRNA expression profile of 3 wild-type CI-huVEC lines and 3 clonal CCM3-knockout CI-huVEC lines with or without fibronectin treatment.


title: Gene expression signature of bone lesion waldenstrom macroglobulinemia and myeloma

city: Tachikawa

first: NAOHIRO

last: SEKIGUCHI

state: Tokyo

address: 3256, Midori

department: Hematology

country: Japan

institute: Disaster Medical Center

summary:
Bone lesion in waldenstom macroglobulinemia (WM) is less common. In the present study, we compaired gene expresion signature of bone lesion in WM compaired to multiple myeloma (MM).
In WM, APRIL and HBGB1 were up-regulated compaired to MM.

overall_design: retrospective study, 7 cases were utilized

first: Naohiro

last: Sekiguchi


title: Effect of miR-203 on the transcriptomic profile of IPSCs and IPSC-derived teratomas

first: Maria

last: Salazar

summary: The balance between pluripotency and differentiation is critical during development and regeneration. miR-203 is a microRNA previously involved in differentiation of different tissues as well as in tumor suppression in multiple malignancies. We have shown that miR-203 is able to promote differentiation of embryonic stem (ES) and induced pluripotent stem (iPS) cells without decreasing pluripotency. We have observed that transient expression of miR-203 significantly improves the efficiency of ES/iPS cells in the generation of quimeras and tetraploid complementation assays, in addition to inducing complex embryo-like structures when these pluripotent cells are injected in mice. In the present RNA seq, we intend to analyze the trascriptomic profile of WT IPSCs, compared to miR-203 KO IPSCs and miR-203 tKI IPSCs (in which we have induced a transient over-expression of miR-203). Moreover, we analyze the mRNA profiles of the teratomas derived from those IPSCs.

overall_design: The general idea was to analyze the transcriptomic profile of miR-203 transiently over-expressing IPSCs, and compare it with WT IPSCs and miR-203 KO IPSCs profiles. Also, we have included mRNA samples extracted from teratomas generated by inoculation of those IPSCs in mice. In the present RNA seq, the gene expression of different clones was tested. Two independent pools of clones from two independent experiments were represented in iPS KI_C1 and C2 (miR-203 cKI iPS cells transiently-treated with DOX to induce miR-203 over-expression). And the same applies to the samples iPS KO_C3 and C4 (miR-203 KO iPS cells) and the samples iPS WT_C1 and C6. The teratomas generated subcutaneously in nude cells and derived from those WT, miR-203 cKI and miR-203 KO iPS cells were tested in this experiment as samples TERAT_WT, TERAT_KO and TERAT_KI 1 and 2. Each of the samples named as TERAT_KI_1, TERAT_KI_2, TERAT_KO and TERAT_WT represent a pool of two teratomas derived from the respective IPSCs.

first: María

last: Salazar Roa

first: Osvaldo

last: Graña

first: Marcos

last: Malumbres


title: single cell RNA sequencing for the paired TNF-free and -treated intestinal organoids

city: Seoul

first: Sung Noh

last: Hong

address: 81 Irwon-Ro, Gangnam-gu

department: Gastroenterolgy

country: South Korea

institute: Samsung Medical Center

summary: Organoids grown from intestinal crypts of controls and patients with Crohn's disease were sub-cultured at least 6 passages. Single cell RNA sequencing was performed for the paired TNF-free and -treated control and Crohn's organoids.

overall_design:
Examination of RNA sample of human intestinal organoids (TNF-free intestinal organoids: n=1 each for control and Crohn’s organoids; TNF(30ng/mL)-treated intestinal organoids: n=1 each for control and Crohn’s organoids)

***Please note that the data was derived from material obtained from patients in Samsung Medical Center and raw data are not provided due to patient privacy concerns.

first: Sung

middle: N


title: Enhanced Pluripotent Cell Function through miR-203-mediated Control of DNA Methylation

summary: Maintaining a full differentiation potential along self-renewal ability is a major property of stem cells during development and regeneration. miR-203 is a microRNA previously involved in skin differentiation and tumor suppression. We show here that transient expression of miR-203 enhances the potential of embryonic (ESC) and induced pluripotent stem cells (iPSC) in contributing to multiple lineages without decreasing their self-renewal properties. In fact, miR-203 significantly improves the efficiency of ES/iPS cells in the generation of quimeras and tetraploid complementation assays, in addition to improving the generation of complex teratomas and embryo-like structures in vivo. These effects are mediated by the direct miR-203-dependent repression of de novo DNA methyltransferases Dnmt3a and Dnmt3b, leading to genome-wide demethylation of pluripotent cells. Transient exposure to miR-203 improves functional differentiation and maturation of pluripotent cells into cardiomyocytes in a Dnmt3a/b-dependent manner, suggesting the possible therapeutic uses of this microRNA in regenerative medicine.

overall_design: The general idea was to analyze the transcriptomic profile of miR-203 transiently over-expressing IPSCs, and compare different time points, from t=0 (before DOX treatment) to embryoid bodies formation. miR-203 tKI IPSCs were transiently induced by doxycycline treatment for 5 days, and then the following time points were analyzed: t=10 (5 days after DOX treatment); t=25 (20 days after DOX treatment) and t=32 (7 days after embryoid body differentiation).

last: Salazar-Roa

first: Fátima

last: Valdés-Mora

first: Timothy

last: Peters

first: Susan

last: Clark


title: Silencing of TRIM28 in human iPSC

status: Public on Jul 03 2020

city: Poznań

first: Maciej

last: Wiznerowicz

address: Fredry 10

country: Poland

institute: Poznań University of Medical Sciences

summary: TRIM28, a multi-domain protein, is crucial in the development of mouse embryos and the maintenance of embryonic stem cells (ESC) self-renewal potential. As the epigenetic factor modulating the structure of chromatin, TRIM28 regulates the expression of numerous genes and is associated with progression and poor prognosis in many types of cancer. IPSC served as a model of the cells with stem cell-like phenotype, e.g., cancer stem cells. We evaluated the role of TRIM28 in pluripotency maintenance in iPSC by silencing endogenous TRIM28 expression with siRNA (iPSC-siTRIM28). IPSC treated with siRNA with no target sequence served as a control (siCTRL) of the experiment. Cells lacking TRIM28 lose the expression of pluripotency markers, as well as the ability to self-renew, and they start to differentiate. Pathway enrichment analysis using Gene Ontology datasets showed significant upregulation of pathways related to apoptosis, differentiation, cellular response to DNA damage stimulus and cell cycle regulation in iPSC-siTRIM28, relative to reference iPSC (iPSC-WT and iPSC-siCTRL).

overall_design: Analysis was performed on 1 biological replicate, 3 technical replicates, 14 days upon siRNA treatment. Samples were compared to control iPSC (treated with control siRNA, with no target), and to wild-type iPSC (non-treated).

first: Joanna

last: Wróblewska


title: Time course of the miR-203 effect on the transcriptomic profile of IPSCs

summary: The balance between pluripotency and differentiation is critical during development and regeneration. miR-203 is a microRNA previously involved in differentiation of different tissues as well as in tumor suppression in multiple malignancies. We have shown that miR-203 is able to promote differentiation of embryonic stem (ES) and induced pluripotent stem (iPS) cells without decreasing pluripotency. We have observed that transient expression of miR-203 significantly improves the efficiency of ES/iPS cells in the generation of quimeras and tetraploid complementation assays, in addition to inducing complex embryo-like structures when these pluripotent cells are injected in mice. In the present RNA seq, we intend to analyze the trascriptomic profile of WT IPSCs transiently treated with DOX to induce miR-203. We analyzed the transcriptomic profile at different time points before and after the induction.

middle: S

last: Roa


title: Real-time quantitative PCR analysis of human collecting lymphatic vessels

city: London

first: Sarah

last: Johnson

state: London

address: Prince Consort Road

department: Bioengineering

institute: Imperial College London

summary: The aim of this feasibility study was to assess whether inflammatory status of the lymphatic vessels (LVs) and/or changes in the miRNA profile of the LVs have potential prognostic and predictive value for overall outcome and risk of relapse. Tissue specimens were obtained from the external iliac vessels draining the pelvic region of patients undergoing debulking surgery (n=10). The tissue was transported directly from surgical theatre to the laboratory in ice-cold sterile Phosphate Buffered Saline. LVs were identified and isolated along with a small amount of surrounding tissue under a dissection microscope within a sterile laminar flow cabinet. Each vessel was divided into two sections. One half of each vessel was further cleaned of surrounding fatty tissue, placed in RNA stabilising solution, frozen on dry ice and stored at -20°C. The miRNAs were extracted with an miRNA easy kit and reverse-transcribed to cDNA using a miScript II RT Kit . Subsequent qPCR was carried out using a miScriptSybrGreen real-time PCR kit and pre-defined miScript miRNA PCR Array.  Quantification of the inflammatory state (low, medium and high) and presence of cancer-infiltration of each LV was carried out using immunohistochemistry with the remaining half of each vessel. The LV miRNA expression profiling was then analysed in the context of high versus low inflammation, and cancer-infiltrated versus non-cancer-infiltrated. Results were correlated with clinical outcome data including relapse with an average follow-up time of 13.3 months.

overall_design: qPCR miRNA expression profiling.  Each sample contains miRNA extracted from the LVs of a single patient (n=10). RNA concentration per sample was quantified using absorbance ratios and an estimated equal amount of total RNA was reverse transcribed per sample. Well G06 was used as a negative control. Housekeeping gene at H06 was used to normalise the data after analysis to identify the most stable housekeeping gene with NormFinder and GeNorm software.


title: RNA sequencing for the paired TNF-free and -treated intestinal organoids

summary: Organoids grown from intestinal crypts of controls and patients with Crohn's disease were sub-cultured at least 6 passages. RNA sequencing was performed for the 12 paired TNF-free and -treated control and Crohn's organoids.

overall_design:
Examination of 24 RNA samples of human intestinal organids (TNF-free intestinal organoids: n=3 each for jejunal control, ileal control, jejunal Crohn’s, and ileal Crohn’s organoids; TNF-treated intestinal organoids: n=3 each for jejunal control, ileal control, jejunal Crohn’s, and ileal Crohn’s organoids; ),
Note from submitter: Our data was derived from material obtained from patients in Samsung Medical Center. Therefore, based on the hospital policy and Korean raw, it is difficult to upload raw data because they contained the patients information.


title: Differential analysis of testosterone induced genes by peritubular cells

city: New Delhi

first: Subeer

last: Majumdar

state: Delhi

address: Aruna Asaf Ali Marg, JNU Complex

country: India

institute: National Institute of Immunology

summary: Agilent's whole genome microarray platform was used to profile differentially expressed genes induced upon testosterone treatment of testicular myoid peritubular cells isolated from adult rat testes.

overall_design: Organism : Rattus Norvegicus, Agilent Catalogue Rat Gene Expression  4x44k Array (AMADID: 014879)

first: Mukesh

last: Gautam


title: Genome-wide RNA sequencing on myeloid derived suppressor cells from mice treated with lymphodepleting chemotherapy reveals distinct IL-6 driven regulatory patterns

city: Tampa

first: Dongliang

last: Du

state: FL

address: 12902 USF Magnolia Drive

country: USA

institute: Moffitt Cancer Center

summary: Myeloid derived suppressor cells (MDSCs) were sorted from B16 tumor-bearing mice, WT and IL-6KO, that were untreated or given lymphodepleting cyclophosphamide and fludarbine.  We report that monocytic and polymorphonuclear MDSC subsets (M-MDSCs and PMN-MDSCs respectively) have a unique gene expression profile in comparison to MDSCs collected from untreated tumor-bearing animals.

overall_design: M-MDSCs and PMN-MDSCs were sorted from WT untreated  mice (n=3) and lymphodepleted (LD) mice (n=3); IL-6KO untreated mice (n=3) and lymphodepleted (LD) mice. Lymphodepleting treatment was initated in mice with established tumors and MDSCs were collected from the spleens of mice 7 days after treatment.

first: Patrick

last: Innamarato

first: Shari

last: Pilon-Thomas


title: Genome-wide RNA sequencing on myeloid derived suppressor cells from mice treated with lymphodepleting chemotherapy

summary: Myeloid derived suppressor cells (MDSCs) were sorted from B16 tumor-bearing mice that were untreated or given lymphodepleting cyclophosphamide and fludarbine.  We report that monocytic and polymorphonuclear MDSC subsets (M-MDSCs and PMN-MDSCs respectively) have a unique gene expression profile in comparison to MDSCs collected from untreated tumor-bearing animals.

overall_design: M-MDSCs and PMN-MDSCs were sorted from untreated (Ctrl) mice (n=3) and lymphodepleted (LD) mice (n=3). Lymphodepleting treatment was initated in mice with established tumors and MDSCs were collected from the spleens of mice 7 days after treatment.


title: Post-transcriptional regulation of MRTF-A by miRNAs during myogenic differentiation of myoblasts

city: Halle (Saale)

first: Danny

last: Misiak

address: Kurt-Mothes-Str. 3a

department: Molecular Cell Biology

institute: Martin Luther University Halle-Wittenberg

summary: The differentiation and regeneration of skeletal muscle from myoblasts to myotubes involves myogenic transcription factors, such as myocardin-related transcription factor A (MRTF-A) and serum response factor (SRF). In addition, post-transcriptional regulation by miRNAs is required during myogenesis. Here we provide evidence for novel mechanisms regulating MRTF-A during myogenic differentiation. Endogenous MRTF-A protein abundance and activity decreased during C2C12 differentiation, which was attributable to miRNA-directed inhibition. Conversely, overexpression of MRTF-A impaired differentiation and myosin expression. Applying miRNA trapping by RNA affinity purification (miTRAP), we identified miRNAs which directly regulate MRTF-A via its 3’UTR, including miR-1a-3p, miR-206-3p, miR-24-3p and miR-486-5p. These miRNAs were upregulated during differentiation and specifically recruited to the 3’UTR of MRTF-A. Concomitantly, Ago2 recruitment to the MRTF-A 3’UTR was considerably increased, whereas Dicer1 depletion or 3’UTR deletion elevated MRTF-A and inhibited differentiation. MRTF-A protein expression was inhibited by ectopic miRNA expression in murine C2C12 and primary human myoblasts. 3'UTR reporter activity diminished upon differentiation or miRNA expression, whereas deletion of the predicted binding sites reversed these effects. Furthermore, TGF-β abolished MRTF-A reduction and decreased miR-486-5p expression. Our findings implicate miR-24-3p and miR-486-5p in the repression of MRTF-A and suggest a complex network of transcriptional and post-transcriptional mechanisms regulating myogenesis.

overall_design: microRNA sequencing of murine C2C12 cell lysates and eluates of miRNA trapping by RNA affinity purification (miTRAP) in undifferentiated and differentiated conditions

first: Ingo

last: Holstein

first: Anurag

last: Singh

first: Falk

last: Pohl

first: Juliane

last: Braun

first: Laura

last: Leitner

first: Stefan

last: Hüttelmaier

first: Guido

last: Posern


title: Analysis of miRNA Expression in Tri-o-cresyl Phosphate-Induced Delayed Neurotoxicity

city: beijing

first: yuyuan

last: li

address: 1 Beichen West Road, Chaoyang District

country: China

institute: Institute of Zoology, Chinese Academy of Sciences

summary: miRNAs have received much attention in the past several years. They have been proven to play important role in the neurodegenerative disease. To identify miRNAs specifically expressed in OPIDN, the expression of lncRNAs in spinal cord obtained from hens with OPIDN were compared with spinal cord obtained from control using NGS.

overall_design: In this study, six samples from hens with OPIDN and healthy hens in control were used to acquire the miRNA expression profiling and the function of the abundant miRNAs in the spinal cord were analyzed by bioinformatic methods. Finally, gga-miR-144-3p, gga-miR-6586-5p, gga-miR-6609-3p, gga-miR-1456-5p, gga-miR-6582-3p, gga-miR-3538, gga-miR-1434 and gga-miR-214 were experimentally validated to play important roles in OPIDN.

first: Yuyuan

last: Li

first: Yijun

last: Wu


title: Rapid activation of distinct members of multigene families in Plasmodium spp.

city: Singapore

first: Han Ping

last: Loh

address: 60 Nanyang Drive

institute: NTU

summary: Variant antigens that are encoded by large multigene families play an important role in the adaptation and immune evasion of a wide range of pathogens. However, the study of their biological function is significantly hampered by the difficulty in controlling their expression in its cellular setting. The genomes of Plasmodium spp. encode a number of different multigene families that are thought to play a critical role for their survival. However, with the exception of the P. falciparum var genes very little is known about the biological roles of any of the other multigene families. Here we report a highly efficient genetic system to study variant antigens in Plasmodium spp. using the Selection Linked Integration method; we are able to activate the expression of a single member of a multigene of our choice using its endogenous promoter.

overall_design: 2 clones of the yir transfectants and 1 P. yoelii YM wildtype RNA, each with 3 replicates were extracted and sent for RNA sequencing. Differential Gene expression analysis was performed to compare the gene expression between the wildtype and the transfected parasites.

first: Peter

last: Preiser

first: Loh

last: Ping


title: Selective time-dependent changes in activity- and cell-specific gene expression in human postmortem brain

status: Public on Jul 02 2020

city: CHICAGO

first: Fabien

last: Dachet

state: IL

address: 912 S. Wood Street

department: Department of NEurology

institute: UIC

summary: RNA seq data at each time point

overall_design: A human brain part is left to spoil for 24h on workbench at room temperature. Time course of 7 time points is performed. The initial sampling withdraw immediately '0H' used a reference until the last one withdraw after waiting 24 hours '24H'. The 7 time points are: '0H', '1H', '2H', '4H', '8H', '12H' and '24H' corresponding respectively to no wait (), a wait of 1 hour, 2 hours, 4 hours, 8 hours, 12 hours and 24 hours

first: James

middle: B

last: Brown

first: Tibor

last: Valyi-nagy

first: Kunwar

last: Narayan

first: Anna

last: Serafini

first: Nathan

last: Boley

first: Thomas

last: Gingeras

first: Susan

last: Celniker

first: Gayatry

last: Mohapatra

first: Jeffrey

last: Loeb