title: The genetic architecture of breast papillary lesions as a predictor of progression to carcinoma

city: Melbourne

first: TANJINA

last: KADER

state: VIC

address: 305 Grattan Street

department: Cancer Genomics

institute: The Sir Peter MacCallum Cancer Centre

type:
SNP genotyping by SNP array
Genome variation profiling by SNP array

summary: We analysed a cohort of breast papillary lesions cases with (n=4) or without (n=7) co-existed cancer for genome-wide copy number and loss of heterozygosity using Affymetrix OncoScan® MIP arrays. Cases included pure papilloma (withoout cancer) (n=7), papilloma cases synchronous with cancer (n=4). We have analysed copy number calling from these samples including carcinoma components of synchronous cases in order to understand the precursor relationship as well any biomarker for progression from papillary lesions to carcinoma.

overall_design: 12 cases of papillary breast lesions with or without co-existing carcinoma were microdissected from FFPE tissue and analysed by molecular inversion probe (MIP) copy number arrays. The data in this submission are CN values for the ~300,000 probes common to two batches performed. Raw data is retained by Affymetrix.

first: Kylie

last: Gorringe

first: Tanjina

last: Kader


title: Epithelial gene expression from a novel mouse model (KPFSR) of pancreatic cancer

status: Public on Jan 30 2020

city: Munich

first: Hans

middle: Carlo

last: Maurer

state: Bavaria

address: Ismaninger Str. 22

department: II. Medizinische Klinik

country: Germany

institute: Klinikum rechts der Isar

type: Expression profiling by high throughput sequencing

summary: This study examines the transcriptional changes invoked in pancreatic ductal adenocarcinoma (PDA) epithelium by a conditional deletion of Slc7a11.

overall_design: Gene expression data are presented for tumor epithelia from established KPFSR PDA that were either treated with corn oil (control) or tamoxifen (treatment). RNA was extracted after Laser Capture Microdissection guided collection of tumor epithelium from frozen sections.

first: Michael

middle: A

last: Badgley

middle: C

first: Kenneth

middle: P

last: Olive


title: Expression and transcript isoform profiling of reprogramming cells expressing a Sox2 mutant lacking a putative RNA binding motif [PAR-CLIP]

city: Guangzhou

first: Yiwei

last: Lai

state: Guangdong

address: 190 Kai Yuan Avenue, Science Park

department: South China Institute for Stem Cell Biology and Regenerative Medicine Key Laboratory of Regenerative Biology, CAS

country: China

institute: Guangzhou Institutes of Biomedicine and Health,Chinese Academy of Sciences

overall_design: Perform 2 independent PAR-CLIP sequencing experiments for SOX2 in HEK293T cells.

first: Yuanjie

last: Wei


title: RNA-seq analysis of high fat diet-induced changes in macrophage transcriptome

city: Duarte

middle: Edmund

last: Stapleton

state: CA

address: 1500 Duarte Rd

department: Diabetes Complications and Metabolism

country: USA

institute: City of Hope

summary: Systemic low-grade inflammation associated with obesity and the metabolic syndrome is also a strong risk factor for the development of diabetes. This inflammatory state is primarily caused by the release of proinflammatory cytokines by macrophages. However, the effect of obesity on the macrophage transcriptome, including long non-coding RNAs, is not known. Here, we analyzed global changes in gene expression of elicited peritoneal macrophages from mice fed a high-fat diet (HFD) or control diet for 12 weeks. The HFD-fed mice has significantly increased body weight and impaired glucose tolerance compared to controls. Transcriptome analysis showed that HFD incuced the expression of genes associated with inflammatory response.

overall_design: C57BL/6J mice were fed with standard chow or high-fat diet (HFD, 60%kcal, Research Diets Inc., D12492i) for 12 weeks. Mice were administered with 3% thioglycollate medium via intraperitoneal injection, and four days later, peritoneal macrophages were collected.

first: Rama

last: Natarajan


title: Epigenetic Targeting of the Warburg Effect by Inhibition of Histone-Deacetylases Leads to Activation of Oxidative Metabolism

status: Public on Jan 29 2020

city: New York

first: Markus

last: Siegelin

state: New York

department: Pathology and Cell Biology

institute: Columbia University

type: Genome binding/occupancy profiling by high throughput sequencing

summary: FDA-approved global (panobinostat, vorinostat) and selective (romidepsin) histone-deacetylase (HDAC) inhibitors elicit metabolic reprogramming in concert with disruption of several Warburg-effect related super-enhancers

overall_design: U87 and NCH644 cells were treated with panobinostat or romidepsin for 24h. Thereafter, cells were harvested and subjected to CHIP isolation. DNA was subsequently processed for CHIP sequencing.

first: Trang

last: Nguyen


title: Expression and transcript isoform profiling of reprogramming cells expressing a Sox2 mutant lacking a putative RNA binding motif

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

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Refer to individual Series


title: Tissue-specific factors differentially regulate the expression of antigen-processing enzymes during dendritic cell ontogeny

city: Toulouse

first: Sylvie

last: Guerder

address: CHU Purpan

department: Immunology

country: France

institute: INSERM

summary: Dendritic cells (DC) form a collection of antigen-presenting cells that are distributed throughout the body. Conventional DC (cDC) which include the cDC1 and cDC2 subsets, and plasmacytoid DC (pDC) constitute the two major ontogenically distinct DC populations. The pDC complete their differentiation in bone-marrow (BM) while the cDC subsets derive from pre-committed bone-marrow (BM) precursors, the pre-cDC, that seed lymphoid and non-lymphoid tissues where they further differentiate into mature cDC1 and cDC2. Within different tissues, cDC express distinct phenotype and function. Notably cDC in the thymus are exquisitely efficient at processing and presenting antigens in the class II pathway, while in the spleen they do so only upon maturation induced by danger signals. To appraise this functional heterogeneity, we examined the regulation of the expression of distinct antigen-processing enzymes during DC ontogeny. We analyzed the expression of cathepsin S (CTSS), cathepsin L (CTSL) and thymus specific serine protease (TSSP), three major antigen-processing enzymes regulating class II presentation in cDC, by DC BM precursors and immature and mature cDC from spleen and thymus. We found that pre-cDC in the BM express relatively high levels of these different proteases. Then, their expression is modulated in a tissue-specific and subset–specific manner with immature and mature thymic cDC expressing overall higher levels than immature splenic cDC. On the other hand, TSSP expression level is selectively down-regulated in spleen pDC while CTSS and CTSL are both increased in thymic and splenic pDC. Hence tissue-specific factors program the expression levels of these different proteases during DC differentiation thus conferring tissue-specific function to the different DC subsets.

overall_design: Transcriptomic profiles (mRNA) of immature and mature conventional dendritic cells (cDC) sorted from the Thymus and the Spleen of B6 mice were generated by next generation sequencing using Illumina HiSeq 3000.

first: Chervin

last: Hassel

first: Maeva

last: Girard


title: 3D ATAC-PALM: Super-resolution Imaging of the Accessible Genome

city: Berkeley

first: Robert

last: Tjian

address: LKS

institute: UC Berkeley/HHMI

summary: Spatial genome organization is essential to direct fundamental DNA-templated biological processes (e.g. transcription, replication, and repair), but the 3D in situ nanometer-scale structure of accessible cis-regulatory DNA elements within the crowded nuclear environment remains elusive. Here, we combined the recently developed Assay for Transposase-Accessible Chromatin with visualization (ATAC-see), PALM super-resolution imaging and lattice light-sheet microscope (a method termed 3D ATAC-PALM) to selectively image and quantitatively analyze key features of the 3D accessible genome in single cells. 3D ATAC-PALM reveals that accessible chromatin are non-homogeneously organized into spatially segregated clusters or accessible chromatin domains (ACDs). To directly link imaging and genomic data, we optimized multiplexed imaging of 3D ATAC-PALM with Oligopaint DNA-FISH, RNA-FISH and protein fluorescence. We found that ACDs colocalize with active chromatin and enclose transcribed genes. By applying these methods to analyze genetically purterbed cells, we demonstrated that genome architectural protein CTCF prevents excessive clustering of accessible chromatin and decompacts ACDs. These results highlight the 3D ATAC-PALM as a useful tool to probe the structure and organizing mechanism of the genome.

overall_design: Examine accessible chromatin landscape (bulk ATAC-seq) in mouse embryonic stem cells under a variety of chemical/genetic purturbations. We performed at least two biological replicates for each condition. We also performed Micro-C experiment after acute auxin-induced acute CTCF depeltion for 6 hours.

first: Liangqi

last: Xie


title: RNA-seq of  CDKN2A and EZH2-deficient pro-B and DN3 lymphocyte progenitors

city: Chicago

first: Barbara

middle: L

last: Kee

state: Illinois

address: 924 E57th st

department: Pathology

institute: The University of Chicago

summary: This goal of this study was to identify genes that are deregulated in the absence of EZH2 in early lymphocyte progenitors.  Due to a requirement for EZH2 to repress Cdkn2a in early B and T cell development we generated Cdkn2a-/-Ezh2fl/fl Il7racre/+ mice.  We examined gene expression by RNA-sequencing in sorted pro-B cells (B220+CD19+CD43+), DN3 cells (Lin- CD25+ CD117-),  from Cdkn2a-/- Ezh2fl/fl Il7racre/+  and Cdkn2a-/- Il7racre/+ control mice.  Reads were aligned to the mm10 reference genome by Tophat2.1.0.   Reads were assigned to genes using the htseq-count tool from HTSeq v 0.6.1 and gene annotations from Ensembl release 78.   Differential expression was calculated across 3 independent replicates by EdgeR.  We found that pro-B and DN3 cells remain specified to their respective lineages despite loss of EZH2.  In contrast, loss of EZH2 led to expression of alternate lineage determinants in pro-B but not DN3 cells indicating that EZH2 is required for lineage commitment in B, but not T lymphocyte progenitors.

overall_design: Expression profiling analysis of pro-B and DN3 cells in Cdkn2a-/-Ezh2fl/fl Il7racre/+ mice and Cdkna2a-/- Il7racre/+ controls generated by RNA-sequencing. 3 independent replicates per sample.

first: Jennifer

last: Jacobsen

first: Mikael

last: Sigvardsson

first: Elizabeth

last: Bartom


title: Lineage reprogramming of c-Kit+CD24+ natural killer cells into myeloid cells

status: Public on Jan 28 2020

city: Seoul

first: Chang-Yuil

last: Kang

address: 29-102, Seoul National University, Shillim-dong, Gwanak-gu

country: South Korea

institute: Seoul National University

summary: Myeloid progenitor cells have generally been considered the exclusive source of myeloid cells under steady-state conditions. Here, we show that NK cells contributed to a myeloid cell lineage pool in naïve and tumor-bearing mice. By using fate tracing of NKp46+ cells, we found evidence that myeloid cell populations could be derived from NK-committed NKp46+ cells. Notably, among mature CD11b+CD27+ NK cells, c-Kit+CD24+ NK cells were capable of differentiating into a range of myeloid lineages in vitro and reprogrammed into neutrophils and monocytes in vivo. In addition to the potential for lineage conversion, c-Kit+CD24+ NK cells retained NK cell phenotypes and effector functions. Mechanistically, GATA-2 was essential for the conversion of the c-Kit+CD24+ NK cells into myeloid cells. Therefore, we discovered that GATA-2-mediated conversion of c-Kit+CD24+ NK cells contributes to myeloid cell development and identified a novel pathway for myeloid lineage commitment in physiological conditions.

overall_design: Examination of mRNA profiles of murine NK cells from the spleen/bone marrow by deep sequencing, using HiSeq 2500

first: Boyeong

last: Song


title: Differential response to Lenalidomide from WT and RUNX1-KO MDS-L del(5q) cells

city: Vancouver

first: Aly

last: Karsan

state: BC

address: W10th Ave

department: Genome Sciences Centre

country: Canada

institute: BC Cancer Agency

summary:
We used CRISPR/Cas9 system to generate WT Control and RUNX1-KO MDS-L cells isolated as single clones from the parental MDS-L cell line. We have observed that RUNX1-KO MDS-L cells are resistant to Lenalidomide-induced cell death compared to WT Control MDS-L cells
We have treated WT and RUNX1-KO MDS-L cells with 1µM Lenalidomide for 24h or 72h and performed expression analysis

overall_design: The experiment consists in eight groups in total. Each group was analyzed with three technical replicates. The groups (per timepoint) are as follows: MDS-L WT DMSO; MDS-L WT LEN; MDS-L KO DMSO; MDS-L KO LEN. We then compared the transcriptional response to LEN within each genotype (MDS-L WT or MDS-L RUNX1-KO).Genes with padj<0.05 we identified as differentially expressed.

first: S

last: Martinez-Hoyer

first: A


title: Genetic interaction mapping and exon-resolution functional genomics with a hybrid Cas9-Cas12a platform

city: Toronto

first: Jason

last: Moffat

state: ON

address: 686 Bay Street, Room 13.9705

department: Molecular Genetics

institute: University of Toronto

summary: Systematic mapping of genetic interactions and interrogation of the functions of sizeable genomic segments in mammalian cells represent important goals of biomedical research. To advance these goals, we present a CRISPR-based screening system for combinatorial genetic manipulation that employs co-expression of Cas9 and Cas12a nucleases and machine learning-optimized libraries of hybrid Cas9-Cas12a guide RNAs. This system, named CHyMErA (Cas Hybrid for Multiplexed Editing and Screening Applications), outperforms genetic screens using Cas9 or Cas12a editing alone. Application of CHyMErA to the ablation of mammalian paralog gene pairs reveals extensive genetic interactions and uncovers phenotypes normally masked by functional redundancy. Application of CHyMErA in a chemo-genetic interaction screen identifies genes that impact cell growth in response to mTOR pathway inhibition. Moreover, by systematically targeting thousands of alternative splicing events, CHyMErA identifies exons underlying human cell line fitness. CHyMErA thus represents an effective screening approach for genetic interaction mapping and the functional analysis of sizeable genomic regions, such as alternative exons.

overall_design: Eight CRISPR screens were performed with four different CHyMErA hybird guide (hg)RNA libraries in human and mouse cell lines. All screens were performed with three biologically independent replicates that were derived from the same library infection and were sequenced at multiple time points. (1) A screen were performed using a human optimization hgRNA library in HAP1 cells and that were subjected to 6-thioguanine and thymidine treatment while non-treated cells were used as controls. (2) A screen were performed using a mouse optimization hgRNA library in CGR8 cells. (3) Screens with a human 2nd generation hgRNA library for single- and dual-gene targeting plus paralog pairs were performed in human HAP1 and RPE1 cells. HAP1 cells were also treated with Torin1 and non-treated cells served as control. (4) A screen with a human exon deletion hgRNA library was performed in RPE1 cells.

first: Thomas

last: Gonatopoulos-Pournatzis

first: Michael

last: Aregger

first: Kevin

middle: R

last: Brown

first: Shaghayegh

last: Farhangmehr

first: Ulrich

last: Braunschweig

first: Henry

middle: N

last: Ward

last: Ha

first: Alexander

last: Weiss

first: Maximilian

last: Billmann

first: Tanja

last: Durbic

first: Chad

last: Myers

first: Benjamin

middle: J

last: Blencowe


title: Autism-misregulated eIF4G microexons control synaptic translation and higher-order cognitive functions

status: Public on Jan 27 2020

state: Ontario

address: 160 College Street

department: Donnelly Centre


title: Cyba deficient mice display an increase of Haematopoietic Stem Cells, and a myeloid bias in detriment of B-cell differentiation

city: Salamanca

first: Ángel

last: Hernández-Hernández

state: Salamanca

address: Plaza Doctores de la Reina s/n, Edificio Departamental, Lab 122

department: Biochemistry and Molecular Biology

country: Spain

institute: University of Salamanca (USAL)

summary: The regulation of protein function by reversible oxidation is increasingly recognized as a key mechanism for the control of cellular signalling, modulating crucial biological processes such cell differentiation. In this scenario, the NADPH oxidases must occupy a prominent position. Our results show that haematopoietic stem and progenitor cells (HSPCs) express three p22phox-dependent NADPH oxidases members (Nox1, Nox2 and Nox4). By deleting the p22phox coding gene (Cyba), here we have analysed the importance of this family of enzymes during in vivo haematopoiesis. Cyba-/- mice show a myeloid bias in detriment of B-cells differentiation, and an enrichment of HSPCs populations. By means of haematopoietic transplant experiments we have also tried to dissect the specific role of the NADPH oxidases. While the absence of Nox1 or Nox2 provides an advantage of reconstitution, the lack of Nox4 rendered the opposite result, what suggests a functional specificity among the different NADPH oxidases. Regarding signalling pathways, the activation of AKT and STAT5 is hampered in Cyba-/- cells, the later due to the downregulation of STAT5 protein. This is in line with the diminished response to IL-7 shown by our results, what could explain defective B-cells development observed in Cyba-/- mice, supporting the requirement of NADPH oxidase activity for terminal B-cell differentiation.

overall_design: Total mRNA from bone marrow Lineage negative cells of 2 wild type (WT) and 2 Cyba-/- mice were analized

first: Rodrigo

last: Prieto-Bermejo


title: Differential effect of glucocorticoids on the activation of monocytes and macrophages.

city: RTP

first: NIEHS

last: Microarray Core

state: NC

address: 111 T.W. Alexander Drive

department: DIR

institute: NIEHS

summary:
Glucocorticoids (GCs) reduce the expression of many genes induced in inflammatory conditions in vivo or by pro-inflammatory stimuli in vitro acting on the Glucocorticoid receptor (GR/NR3C1). However, GCs have pleiotropic effects on the immune system. Monocytes and macrophages are important cells of the innate immune system, exhibiting complex properties with enhancing as well as suppressive effects on inflammatory processes depending on their stage of activation and differentiation. Macrophages contribute to host defense, tissue remodeling and wound healing
The mechanisms of GCs actions on monocytes and macrophages and their contribution to systemic anti-inflammatory effects remain unclear. These effects on activation GCs in monocytes and macrophages were investigated using the human monocytic cell line THP-1 and differentiation protocol PMA. Monocytes-THP1 and macrophages derived-monocytes were exposed for 6 hours to 100 nM of dexamethasone (Dex) and analyzed by microarrays. In macrophages-derived monocytes with respect to monocytes-THP-1, the number of genes differentially regulated was almost twice, among these, genes related with pro-inflammatory pathways, like TNFa, CXCL1 and GZMA. Interestingly, one of the genes differentially regulated (decreased) in macrophages exposed to Dex was the GR itself. If this effect modifies the inflammatory function of macrophages derived-monocyte is a question that will be evaluated through this study.

overall_design: For this study, 12 samples were analyzed by NanoString: 2 experimental groups (Monocytes THP-1 and macrophages differentiated from monocytes THP-1; 2x106 cells/well); 2 treatments (vehicle and 100nM Dex); 1 time point (6 h) and x3 replicates. All samples passed nSolver’s initial QA/QC controls.  One of the Mac Dex samples (Mac Dex 2) clustered with the Mac Controls and saw no increased expression of FKBP5. It was therefore excluded from further analysis. All remaining replicates were well correlated (R > 0.98).

first: David

last: Diaz-Jimenez

first: John

last: Cidlowski


title: KDM4A mediates maternal zygotic transition in mammals

city: Copenhagen

first: Aditya

last: Sankar

email: cnj376@ku.dk, aditya.sankar@embl.de

address: Ole Maaloes Vej 5

department: Biotech Research and Innovation Centre

country: Denmark

institute: University of Copenhagen


title: Gene Expression Differences in Host Response to Schistosoma haematobium Infection

status: Public on Jan 25 2020

first: Jennifer

last: Downs

address: 1300 York Ave.

institute: Weill Cornell Medicine

summary: Schistosome worms infect over 200 million people worldwide. They live in the host’s bloodstream and alter host immunity. Epidemiological data suggest  that males and females have different responses to schistosome infection, but the effect of sex on systemic response is undetermined. Our objective was to characterize differences in peripheral blood transcriptional profiles in people with or without active Schistosoma haematobium infection and to determine whether this signature differs between males and females. mRNA was isolated using poly(A) selection and sequenced on an Illumina Hi-Seq4000 platform. Transcripts were aligned to the human hg19 reference genome and counted with the HTSeq package. Genes were compared for differential expression using DESeq2. Ingenuity Pathway Analysis (IPA) was used to identify gene networks altered in the presence of S. haematobium. We  enrolled 33 participants from villages in rural Tanzania where S. haematobium is endemic. After correction for multiple comparisons, we observed 383 differentially expressed genes between those with or without S. haematobium infection when sex was included as a covariate. Heat-mapping of the genes with  1.5-fold differences in gene expression revealed clustering by S. haematobium infection status. The top networks included development, cell death and survival, cell signaling, and immunologic disease pathways. We observed a distinct whole blood transcriptional profile, as well as differences in men and women, with S. haematobium infection. Additional studies are needed to determine the clinical effects of these divergent responses. Attention  to sex-based differences should be included in studies of schistosome infection.

overall_design: Comparisons of men and women with and without S. haematobium infection


title: Differential effect of glucocorticoids on the activation of monocytes and macrophages

summary:
The Glucocorticoid receptor (GR/NR3C1) is expressed in almost all immune cells. Glucocorticoids (GCs) are potent regulators of inflammation with effects mainly immunosuppressive. While, GCs have pleiotropic effects on Macrophages exhibiting complex properties with enhancing as well as suppressive effects on inflammatory processes depending on their stage of differentiation and activation, the mechanisms of GCs actions on Monocytes and Macrophages and their contribution to systemic anti-inflammatory effects remain unclear.
Previously we have demonstrated that NLRP3 expression is induced by GCs in macrophages differentiated from monocytes-THP-1 but not in monocytes.  These findings raise the possibility that the stage of myeloid differentiation and the cellular environment dictates the actions of GR from a pro- or anti-inflammatory perspective. Moreover, our preliminary results using the same cell line THP-1 show that the differentiation process from monocytes to macrophages also is related with the downregulation of GR induced by dex.

overall_design: Human Monocyte cell line THP-1 was used for differentiation into macrophages with PMA during 3 hours and a time of recovery of 24 hours. Monocytes-THP-1 and macrophages-derived monocyte-THP-1 were maintained during 24 hours with charcoal medium before the Dex treatment for 6 hours. RNA samples from all treatments groups (Mono-THP-1 Vehicle and Dex and Macro-THP-1 vehicle and Dex) were extracted using QIAshredder and RNeasy Mini kit with DNase treatment. The RNA concentration will be determined by Nanodrop and the quality using BioAnalyzer. We expect to found significant differences in RNA expression between cells (Mono vs Macrophages) and treatment (Vehicle vs Dex).


title: APOE2 orchestrated differences in transcriptomic and lipidomic profiles of postmortem AD brain.

status: Public on Jan 26 2020

city: Pittsburgh

first: Iliya

last: Lefterov

state: PA

address: 130 De Soto Street

department: Environmental and Occupational Health

institute: University of Pittsburgh

summary: The application of advanced sequencing technologies and improved mass-spectrometry platforms revealed significant changes in gene expression and lipids in Alzheimer’s disease (AD) brain. The results so far have prompted further research using “multi-omics” approaches. These approaches become particularly relevant, considering the inheritance of APOEε4 allele as a major genetic risk factor of AD, disease protective effect of APOEε2 allele, and a major role of APOE in brain lipid metabolism. Significantly affected GO terms and pathways were determined based on the comparisons of APOEε2/c datasets to those of APOEε3/3 and APOEε4/c brain samples. The analysis of lists of genes in highly correlated network modules and of those differentially expressed demonstrated significant enrichment in GO terms associated with genes involved in intracellular proteasomal and lysosomal degradation of proteins, protein aggregates and organelles, ER stress, and response to unfolded protein, as well as mitochondrial function, electron transport, and ATP synthesis. Small nucleolar RNA coding units important for posttranscriptional modification of mRNA and therefore translation and protein synthesis were upregulated in APOEε2/c brain samples compared to both APOEε3/3 and APOEε4/c. The analysis of lipidomics datasets revealed significant changes in ten major lipid classes (exclusively a decrease in APOEε4/c samples), most notably non-bilayer-forming phosphatidylethanolamine and phosphatidic acid, as well as mitochondrial membrane-forming lipids. The results of this study, despite the advanced stage of AD, point to the significant differences in postmortem brain transcriptomes and lipidomes, suggesting APOE allele associated differences in pathogenic mechanisms. Correlations within and between lipidomes and transcriptomes indicate coordinated effects of changes in the proteasomal system and autophagy—canonical and selective, facilitating intracellular degradation, protein entry into ER, response to ER stress, nucleolar modifications of mRNA, and likely myelination in APOEε2/c brains. Additional research and a better knowledge of the molecular mechanisms of proteostasis in the early stages of AD are required to develop more effective diagnostic approaches and eventually efficient therapeutic strategies.

overall_design: Postmortem brain samples from inferior parietal lobule genotyped as APOEε2/c (APOEε2/carriers), APOEε3/3, and APOEε4/c (APOEε4/carriers), age- and gender-matched, were used to reveal APOE allele-associated changes in transcriptomes and lipidomes. Differential gene expression and co-expression network analyses were applied to identify up- and downregulated Gene Ontology (GO) terms and pathways for correlation to lipidomics data.

first: Cody

middle: M

last: Wolfe

first: Nicholas

middle: F

last: Fitz

first: Kyong

last: Nam

first: Florent

last: Letronne

first: Richard

last: Biedrzycki

first: Julia

last: Kofler

first: Xianlin

last: Han

first: Jianing

last: Wang

first: Jonathan

last: Schug

first: Radosveta

last: Koldamova


title: RQT complex dissociates ribosomes collided on endogenous RQC substrate SDD1

city: Tokyo

first: Yoshitaka

last: Matsuo

address: 4-6-1, Shirokanedai, Minato-Ku

department: IMSUT

country: Japan

institute: The University of Tokyo,

summary: Ribosome-associated quality control (RQC) represents a rescue pathway in eukaryotic cells triggered upon translational stalling. Collided ribosomes are recognized for subsequent dissociation followed by degradation of nascent peptides. However, endogenous RQC-inducing sequences and the mechanism underlying the ubiquitin-dependent ribosome dissociation remain poorly understood. Here, we identified the SDD1 mRNA from S. cerevisiae as an endogenous RQC substrate and reveal its mRNA and nascent peptide dependent stalling mechanism by mutational and cryo-EM analyses. In vitro translation of SDD1 mRNA enabled the reconstitution of Hel2-dependent poly-ubiquitination of collided di- and preferentially tri-ribosomes. Distinct trisome architecture was visualized by cryo-EM and provides the structural basis for more efficient recognition by Hel2 over disomes. Subsequently, the Slh1 helicase subunit of the RQC trigger (RQT) complex preferentially dissociates the first stalled poly-ubiquitinated ribosome in an ATP-dependent manner. Together, these findings provide fundamental mechanistic insights into RQC and its physiological role in maintaining cellular protein homeostasis.

overall_design: 2 replicates in 2 different strains

first: Petr

last: Tesina

first: Shizuka

last: Nakajima

first: Masato

last: Mizuno

first: Akinori

last: Endo

first: Robert

last: Buschauer

first: Jingdong

last: Cheng

first: Okuto

last: Shounai

first: Ken

last: Ikeuchi

first: Yasushi

last: Saeki

first: Thomas

last: Becker

first: Roland

last: Beckmann

first: Toshifumi

last: Inada


title: Identification of introduced and stably inherited DNA methylation variants in soybean associated with soybean cyst nematode parasitism [MethylC-Seq]

city: Knoxville

first: Sarbottam

last: Piya

state: Tennessee

address: 2431 Joe Johnson Dr.

department: Plant Science

institute: University of Tennessee

type: Methylation profiling by high throughput sequencing

summary: Methylome changes mediated by the soybean cyst nematode resistance gene GmSHMT08

overall_design: Examining the changes in the methylome of two nearly isogenic lines (TN09-16 and TN09-29) and their parental lines (Fowler and Anand) under soybean cyst nematode infected and non-infected conditions.

first: Tarek

last: Hewezi


title: Sexual dimorphism of gut microbiota dictates therapeutics efficacy of radiation injuries [miRNA-seq]

city: Tianjin

first: Ming

last: Cui

address: 238 Baidi Road

department: Institute of Radiation Medicine,

institute: Chinese Academy of Medical Sciences and Peking Union Medical College

type: Non-coding RNA profiling by high throughput sequencing

summary:
Accidental or iatrogenic ionizing radiation exposure precipitates acute and chronic radiation injuries. Traditional paradigm of mitigating radiotherapy-associated side effects has ignored the gender-specific dimorphism of patients. Here we examined the effects of sexual dimorphism on therapeutic agent efficiencies in murine models. High-throughput sequencing showed that different treatments preserved miRNA expression profile in small intestines of male or female mice, respectively. Collectively, our observations demonstrate that therapeutic strategy efficiencies for radiation toxicity might be dependent on the gender of patients.

We aim to uncover the underlying mechanisms by which simvastatin or high fat diet fights against radiotherapy-induced gastrointestinal tract toxicity for male or female mice. The mice were exposed to 12 Gy total abdominal irradiation (TAI), then treated with simvastatin for male mice or fed with high fat diet for female mice. The small intestine tissues were extracted from male mice without TAI, exposed to TAI only, and exposed to TAI combined with simvastatin treatment as one cohort, and from female mice  without TAI, exposed to TAI only, and exposed to TAI combined with high fat diet feeding as the other cohort.

overall_design: Examination of miRNA profile of small intestine from male mice (without irradiation, total abdominal irradiation or total abdominal irradiation combined with simvastatin treatment) and female mice (without irradiation, total abdominal irradiation or total abdominal irradiation combined with high fat diet).

first: Huiwen

last: Xiao


title: Insights into the involvement of spliceosomal mutations in myelodysplastic disorders from an analysis of SACY-1/DDX41 in Caenorhabditis elegans

city: Minneapolis

first: Micah

last: Gearhart

state: MN

address: 6-160 Jackson Hall, 321 Church St.

department: Genetics, Cell Biology and Development

institute: University of Minnesota

summary: Mutations affecting spliceosomal proteins are frequently found in hematological malignancies, including myelodysplastic syndromes and acute myeloid leukemia. DDX41/Abstrakt is a metazoan-specific spliceosomal DEAD-box RNA helicase found to be recurrently mutated in inherited myelodysplastic syndromes and in relapsing cases of acute myeloid leukemia. The genetic properties and genomic impacts of disease-causing missense mutations in DDX41 and other spliceosomal proteins have been uncertain. Here we conduct a comprehensive molecular genetic analysis of the C. elegans DDX41 ortholog, SACY-1. Our results reveal general essential functions for SACY-1 in both the germline and the soma, as well as specific functions affecting germline sex determination and cell cycle control. Certain sacy-1/DDX41 mutations, including the R525H human oncogenic variant, confer antimorphic activity, suggesting that they compromise the function of the spliceosome. Consistent with these findings, sacy-1 exhibits synthetic lethal interactions with several spliceosomal components, and biochemical analyses suggest that SACY-1 is a component of the C. elegans spliceosome. We used the auxin-inducible degradation system to analyze the impact of SACY-1 on the transcriptome using RNA sequencing. SACY-1 depletion impacts the transcriptome through splicing-independent and splicing-dependent mechanisms. The observed transcriptome changes suggest that disruption of spliceosomal function induces a stress response. Altered 3’ splice site usage represents the predominant splicing defect observed upon SACY-1 depletion, consistent with a role for SACY-1 as a second-step splicing factor. Missplicing events appear more prevalent in the soma than the germline, suggesting that surveillance mechanisms protect the germline from aberrant splicing.

overall_design: Polyadenylated transcripts present in control- or SACY-1-depleted worms following auxin mediated degradation in the soma or germline.

first: Tatsuya

last: Tsukamoto

middle: D

first: David

last: Greenstein


title: RNA-seq of Bifidobacterium longum NCC2705

city: Gifu

first: Tomoya

last: Kozakai

address: Yanagido 1-1

institute: Gifu university

summary:
Bacterial promoters consist of core sequence motifs termed –35 and –10 boxes. The consensus motifs are TTGACA and TATAAT, respectively, which were identified from leading investigations on E. coli. However, the consensus sequences are not likely to fit genetically divergent bacteria. The sigma factor of the genus Bifidobacterium has a characteristic polar domain in the N-terminus, suggesting the possibility of specific promoter recognition. We reevaluated the structure of B. longum NCC2705 promoters and compared them to other bacteria.
Transcriptional start sites (TSS) of the B. longum NCC2705 strain were identified using RNA-Seq analysis to extract promoter regions. Conserved motifs of a bifidobacterial promoter were determined using regions upstream of TSSs and a hidden Markov model. As a result, consensus motifs of the –35 and –10 boxes were TTGTGC and TACAAT, respectively. To assess each base of both motifs, we constructed thirty-seven plasmids based on pKO403-TPCTcon, including the hup promoter connected with a chloramphenicol acetyltransferase as a reporter gene. This reporter assay showed two optimal motifs of the –35 and –10 boxes, TTGNNN and TANNNT, respectively.
We further analyzed spacer-lengths between the –35 and –10 boxes via a bioinformatics approach. The spacer-lengths predominant in bacteria have been generally reported to be approximately 17 bp. In contrast, the predominant spacer-lengths in the genus Bifidobacterium and related species were 11 bp, in addition to 17 bp. A reporter assay to assess the spacer-lengths indicated that the 11 bp spacer-length produced unusually high activity.

overall_design: RNA-Seq libraries were constructed from RNA of Bifidobacterium longum NCC2705 strain. The total RNA were extracted from MRS culture in OD660 0.5, treated with Ribo-Zero rRNA Removal Kit (Illumina) to deplete the ribosomal RNA, and then reverse transcribed. The RNA-Seq was performed using a MiSeq (Illumina). The fragment depth graph was used to assign the transcription start sites (TSSs).

first: Ayako

last: Izumi

last: Horigome

first: Toshitaka

last: Odamaki

first: Jin-zhong

first: Izumi

last: Nomura

first: Tohru

last: Suzuki


title: Oxidative phosphorylation promotes primary melanoma invasion

status: Public on Jan 24 2020

first: Yariv

last: Houvras

state: NY

address: 1300 York Ave

department: Surgery

institute: Weill Cornell Medical College

summary: Dermal invasion is a hallmark of malignant melanoma. The molecular alterations driving the progression of primary melanoma to metastatic disease have been studied extensively, whereas the early progression of non-invasive primary melanoma to an invasive state is not well understood. To elucidate the mechanisms underlying the transition from radial to vertical growth, the first step in melanoma invasion, we developed a zebrafish melanoma model in which constitutive activation of ribosomal protein S6 kinase 1 (RSK1) drives tumor invasion. Transcriptomic analysis of RSK1-activated tumors identified metabolic changes, including upregulation of genes associated with oxidative phosphorylation. Vertical growth phase human melanoma cells show higher oxygen consumption and preferential utilization of glutamine compared to radial growth phase melanoma cells. Peroxisome proliferator-activated receptor gamma coactivator-1α (PGC1a has been proposed as a master regulator of tumor oxidative phosphorylation. In human primary melanoma specimens we show that PGC1a protein expression is positively associated with increased tumor thickness and expression of the proliferative marker Ki-67 and the reactive oxygen species (ROS) scavenger SCARA3. PGC1a depletion modulates cellular processes associated with primary melanoma growth and invasion, including oxidative stress. Our results support a role for PGC1a in mediating glutamine-driven OXPHOS to facilitate the invasive growth of primary melanoma.

overall_design: Melanomas harvested from MiniCoopR-GFP zebrafish (n=6) and MiniCoopR-myrRSK (n=6) zebrafish

first: Iman

last: Osman


title: Gene expression profile of Otx2 KI and Crx KO  retina at P12

city: Suita

first: Takahisa

last: Furukawa

state: Osaka

address: 6-2-4 Furuedai

department: Developmental Biology

institute: Osaka Bioscience Institute

summary: To analyze the expression profile in the Otx2 knock-in (a knock-in mouse line expressing Otx2 from the Crx locus on chromosome 7) and Crx knockout retina, we performed a microarray analysis using wild-type (Crx +/+), Otx2 KI (Crx Otx2/Otx2) and Crx KO (Crx -/-) retina at P12.

overall_design: We performed a microarray analysis using wild-type, Otx2 KI and Crx KO retina at P12, and compared the expression profiles.

first: Haruka

last: Yamamoto


title: Mildew locus O facilitates colonization by arbuscular mycorrhiza in angiosperms

city: Norwich

first: Catherine

last: Jacott

address: Colnet Lane

country: United Kingdom

institute: John Innes Centre

summary: RNA-sequencing of Hordeum vulgare cv. Ingrid wild type and Hvmlo1-5 mutant in non-mycorrhizal (control) and mycorrhizal roots at 17 days post inoculation (dpi) and 26 dpi.

overall_design: H. vulgare cv. Ingrid wild type and Hvmlo1-5 were grown with and without arbuscular mycorrhizal fungi (Rhizophagus irregularis). Roots were harvested at 17 dpi and 26 dpi and RNAs were extracted for sequencing.

first: Burkhard

last: Steuernagel


title: N-myc regulates lens growth and terminal differentiation in vivo

city: Rio de Janeiro

first: Rodrigo

middle: A. P.

last: Martins

state: RJ

address: Av. Pedro Calmon, 550

department: Programa de Biologia Celular e do Desenvolvimento, Instituto de Ciências Biomédicas

country: Brazil

institute: Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ, Brazil

summary: We performed RNA-seq with both wild type and N-myc knock-out E14 mouse lenses and identified the differentially expressed genes affected by N-myc knockout.

overall_design: We have 3 biological replicates for WT controls and 2 biological replicates for N-myc knock-out samples

first: Gabriel

last: Cavalheiro

middle: E

last: Matos-Rodrigues

first: Yilin

last: Zhao

first: Anielle

last: Gomes

first: Deepti

last: Anand

first: Danilo

last: Predes

first: Silmara

last: de Lima

first: José

middle: G

last: Abreu

first: Deyou

last: Zheng

first: Salil

last: Lachke

first: Ales

last: Cvekl


title: Sexual dimorphism of gut microbiota dictates therapeutics efficacy of radiation injuries [RNA-seq]

first: Ming

last: Cui

summary:
Accidental or iatrogenic ionizing radiation exposure precipitates acute and chronic radiation injuries. Traditional paradigm of mitigating radiotherapy-associated side effects has ignored the gender-specific dimorphism of patients. Here we examined the effects of sexual dimorphism on therapeutic agent efficiencies in murine models. High-throughput sequencing of host mRNA showed that different treatments reprogrammed the spectrum of mRNA expression in small intestines of male or female mice, respectively. Collectively, our observations demonstrate that therapeutic strategy efficiencies for radiation toxicity might be dependent on the gender of patients.

We aim to uncover the underlying mechanisms by which simvastatin or high fat diet fights against radiotherapy-induced gastrointestinal tract toxicity for male or female mice. The mice were exposed to 12 Gy total abdominal irradiation (TAI), then treated with simvastatin for male mice or fed with high fat diet for female mice. The small intestine tissues were extracted from male mice without TAI, exposed to TAI only, and exposed to TAI combined with simvastatin treatment as one cohort, and from female mice  without TAI, exposed to TAI only, and exposed to TAI combined with high fat diet feeding as the other cohort.

overall_design: Examination of mRNA profile of small intestine from male mice (without irradiation, total abdominal irradiation or total abdominal irradiation combined with simvastatin treatment) and female mice (without irradiation, total abdominal irradiation or total abdominal irradiation combined with high fat diet).

first: Huiwen

last: Xiao


title: High-throughput screens of PAM-flexible Cas9 variants for gene knock-out and transcriptional modulation

city: New York

first: Neville

last: Sanjana

address: 101 Avenue of the Americas

institute: New York Genome Center

summary: A key limitation of the commonly-used CRISPR enzyme S. pyogenes Cas9 is the strict requirement of an NGG protospacer-adjacent motif (PAM) at the target site, which reduces the number of accessible genomic loci. This constraint can be limiting for genome editing applications that require precise Cas9 positioning. Recently, two Cas9 variants with a relaxed PAM requirement (NG) have been developed (xCas9 and Cas9-NG) but their activity has been measured at only a small number of endogenous sites. Here we devised a high-throughput Cas9 pooled competition screen to compare the performance of both PAM-flexible Cas9 variants and wild-type Cas9 at thousands of genomic loci and across 3 modalities (gene knock-out, transcriptional activation and suppression). We show that PAM flexibility comes at a substantial cost of decreased DNA targeting and cutting. Of the PAM-flexible variants, we found that Cas9-NG outperforms xCas9 regardless of genome engineering modality or PAM. Finally, we combined xCas9 mutations with those of Cas9-NG, creating a stronger transcriptional modulator than existing PAM-flexible Cas9 variants.

overall_design: K562 cells transduced with Cas9 variant and sgRNA library targeting each of three genes separately (CD45, CD46 and CD55) were pooled together prior to antibody staining and sorting based on the expression of the target gene. sgRNA distribution in the cell pool prior to sorting was used as a reference.

first: Mateusz

last: Legut

first: Zharko

last: Daniloski

first: Xinhe

last: Xue

first: Dayna

last: McKenzie

first: Xinyi

last: Guo

first: Hans-Hermann

last: Wessels


title: Diversity of peripheral blood human NK cells identified by single cell RNA sequencing

city: Manchester

first: Paul

last: Martin

state: Greater Manchester

address: Oxford Road

department: Centre for Genetics and Genomics Versus Arthritis

institute: Univeristy of Manchester

summary: Human Natural Killer (NK) cells in peripheral blood perform many functions and classification of specific subsets has been a long-standing goal. Here, we report single cell RNA-sequencing of NK cells, comparing gene expression in unstimulated and IL-2 activated cells, from healthy CMV-negative donors. Three NK cell subsets resembled well-described populations, CD56brightCD16-, CD56dimCD16+CD57- and CD56dimCD16+CD57+. CD56dimCD16+CD57- cells sub-divided to include a population with higher chemokine mRNA and increased frequency of KIR expression. Three novel human blood NK cell populations were identified as: a population of type I interferon responding NK cells which were CD56neg, a population exhibiting a cytokine-induced memory-like phenotype, including increased granzyme B mRNA in response to IL-2, and finally, a small population, with low ribosomal expression, down-regulation of oxidative phosphorylation and high levels of immediate early response genes indicative of cellular activation. Analysis of CMV-positive donors established that CMV altered the proportion of NK cells in each subset, especially an increase in adaptive NK cells, as well as gene regulation within each subset. Together, these data establish an unexpected diversity in blood NK cells and provide a new framework for analysing NK cell responses in health and disease.

overall_design: Single cell transriptional profiling of NK cells isolated from two CMV-negative healthy donors

first: Samantha

last: Smith

first: Philippa

last: Kennedy

first: Kevin

middle: B

last: Stacey

first: Jonathan

last: Worboys

first: Annie

last: Yarwood

first: Wendy

last: Thomson

first: Steve

last: Eyre

first: Daniel

last: Davis


title: Identification of introduced and stably inherited DNA methylation variants in soybean associated with soybean cyst nematode parasitism [RNA-seq]

first: Sarbottam

last: Piya

overall_design: Examining the changes in the transcriptome of two nearly isogenic lines (TN09-16 and TN09-29) under soybean cyst nematode infected and non-infected conditions.

first: Tarek

last: Hewezi


title: Identification of introduced and stably inherited DNA methylation variants in soybean associated with soybean cyst nematode parasitism

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

overall_design: Refer to individual Series


title: Sexual dimorphism of gut microbiota dictates therapeutics efficacy of radiation injuries

first: Ming

last: Cui


title: Knockout of CNOT1 and CNOT10 in primary CD4+ T-cells causes a type-1 interferon response

city: San Francisco

first: David

middle: E.

last: Gordon

state: California

address: 1700 4th Street, Byers Hall 309

institute: UCSF

summary: We report a type 1 interferon response accompanying polyclonal knockout of CNOT1 or CNOT10 in primary human CD4+ T-cells harvested from healthy donors

overall_design: Polyclonal knockouts targeting CNOT1 and CNOT10 in primary human CD4+ T-cells harvested from four healthy donors

first: Nevan

last: Krogan


title: Phosphorylated Groucho delays differentiation in the follicle stem cell lineage by providing a molecular memory of EGFR signaling in the niche

first: Todd

last: Nystul

address: 513 Parnassus Ave, Box 0412

summary: Overexpression of activated EGFR in follicle cells of the drosophila ovary followed by RNA-Seq to assess differential expression

overall_design: Control (UAS-CD8-GFP only) vs EGFR-activated (UAS-EGFR-lambdatop + UAS-CD8-GFP). Two biological replicated for each.

first: Michael

last: Johnston


title: Expression profile of Urothelial carcinoma of the urinary bladder (UCB) or bladder cancer from Blood

city: Braunschweig

first: Ayssar

middle: A.

last: Elamin

state: Nds

address: Salzdahlumer Str. 196

department: Research & Development

institute: LIONEX GmbH

summary:
The main aim of this study was to assess the changes in blood gene expression in UCB patients and to identify genes serving as biomarkers for UCB diagnosis and progression.
Our study characterized the comprehensive expression profile of UCB and highlighted differences in expression patterns between UCB and healthy control

overall_design: The prediction of classes of bladder tumors using a limited set of genes could help in development of a clinical decision model. The main objective of the current study is to predict bladder cancer disease course by identifying markers that will predict the likelihood of progression in patients with high grade tumors. By utilizing gene expression profiling, we identified different genes as a signature biomarkers for UCB and tumor progression in risk and prognostics groups using a multiple group comparison.

first: Saskia

last: Klunkelfuß

first: Susanne

last: Kämpfer

first: Wulf

last: Oehlmann

first: Matthias

last: Stehr

first: Christopher

first: Guy

last: Simpson

first: Richard

last: Morgan

first: Hardev

last: Pandha

first: Mahavir

last: Singh


title: Adaptation to industrial stressors through genomic and transcriptional plasticity in a bioethanol producing fission yeast isolate

status: Public on Jan 23 2020

city: Melbourne

first: Dane

middle: Aaron McKay

last: Vassiliadis

address: Parkville VIC 3000

country: Australia

institute: Peter MacCallum Cancer Centre


title: Silencing Trisomy 21 with XIST in Neural Stem Cells Promotes Neuronal Differentiation (single cells)

city: Worcester

first: Jan

last: Czerminski

state: Massachusetts

address: 55 Lake Ave N, S6-241

department: Neurology

institute: UMass Medical School

summary: Although Down Syndrome (DS) is the leading genetic cause of intellectual disability in children, the developmental pathogenesis remains largely unknown, and better strategies are needed to investigate this. We previously showed that one copy of chromosome 21 can be epigenetically silenced in DS iPSCs by insertion of an XIST transgene, which produces a non-coding RNA that normally silences one X chromosome in female cells. XIST was shown to induce heterochromatin and silence transcription across chromosome 21 in pluripotent stem cells, the natural developmental context of XIST function. Prior literature indicated that initiation of chromosome silencing is only possible within 48 hours of mES cell differentiation, however it would be highly advantageous experimentally if trisomy silencing could be initiated in differentiated cells, and this is critical for any therapeutic potential of XIST. Here we use RNAseq and molecular cytology to investigate the effectiveness of XIST for trisomy silencing in cells undergoing in vitro neural differentiation and examine the potential cell phenotypic effects of chromosomal silencing. Induction of XIST from the onset of differentiation resulted in comprehensive silencing of chromosome 21 genes, providing a powerful approach to examine effects of trisomy on neurogenesis. To determine whether human neural stem cells can initiate XIST-mediated silencing, we induced XIST at several times during neural differentiation. We demonstrate for the first time that differentiated normal human cells can initiate chromosome silencing in response to XIST expression. While this process only takes a few days in pluripotent cells, we show that it takes 2-3 weeks in differentiated cells. Importantly, single-cell RNAseq revealed that cells which express XIST preferentially differentiate into neurons, providing evidence of phenotypic improvement with trisomy silencing, which is seen even when XIST is initiated weeks into differentiation. We are currently investigating whether silenced neurons show other non-chromosome 21 transcriptional changes suggestive of potentially improved function. These studies have important implications for the understanding of XIST biology and DS neurobiology, and also further open the possibility of a chromosomal therapy for DS using a single gene: XIST.

overall_design: Single-cell RNA sequencing of 6 samples from 3 cell lines and 3 dox treatment conditions.

middle: T

first: Jeanne

last: Lawrence


title: Next Generation Sequencing Facilitates Quantitative Analysis of  colorectal cancer cell lines including SW480 and SW620

status: Public on Jan 22 2020

city: Guangzhou

first: Shengyuan

last: Chen

state: Guangdong province

address: No. 1023-1063, shatai South Road, Baiyun District, Guangzhou, Guangdong Province

department: Gastroenterology

institute: Nanfang Hospital, Southern Medical University

summary:
Purpose: The goals of this study are to compare the differentially expressed miRNAs between SW480 and SW620.
Methods:Total RNA of two samples was used to prepare the miRNA sequencing library.The libraries were denatured as single-stranded DNA molecules, captured on Illumina flow cells, amplified in situ as clusters and finally sequenced for 50 cycles on Illumina NextSeq.
Results:We found 72 miRNAs were downregulated on the basis of fold-change less than 0.5. However,total 160 miRNAs were upregulated on the basis of fold-change greater than 2.
Conclusions: Our study represents the detailed analysis of SW480 and SW620, generated by RNA-seq technology. Our results show that NGS offers a comprehensive and more accurate quantitative and qualitative evaluation of miRNA content. We conclude that RNA-seq based transcriptome characterization would expedite genetic network analyses and permit the dissection of complex biologic functions.

overall_design: SW480 and SW620 were generated by deep sequencing, using Illumina NextSeq.

first: Haoxuan

last: Zheng

first: Jiaxuan


title: Single cell RNA sequencing of Hic1 and PDGFRa-labeled cells in the heart

city: Vancouver

first: Fabio

last: Rossi

address: 2222 Health Sciences Mall

department: School of Biomedical Engineering

institute: University of British Columbia

summary: The cardiac stroma contains multipotent mesenchymal progenitors. However, lineage relationships within cardiac stromal cells are poorly defined. Here, we identify heart-resident PDGFRa+ SCA-1+ cells as cardiac Fibro/Adipogenic Progenitors (cFAPs) and show that they respond to ischemic damage by generating SCA-1- fibrogenic cells. Pharmacological blockade of this differentiation step with an anti-fibrotic tyrosine kinase inhibitor decreases post-myocardial infarction (MI) remodeling and leads to improvement in heart function.

overall_design: To understand the heterogeneity of cardiac mesenchymal progenitors in the heart,  hearts were digested and cell suspensions of Hic1 CreERT2 tdTomato+, Hic1-citrine+ and PDGFRa H2BGFP+ cells were subjected to single cell RNA sequencing. Hic1 CreERT2 tdTomato+ and PDGFRa H2BGFP+ cells were collected from undamaged mice as well as 7 days after left anterior descending (LAD) coronary artery ligation. Hic1 citrine+ cells were only collected from undamaged mice. For the Hic1 CreERT2 tdTomato samples (for each time point), 3 mice hearts were pooled together in one sample. For the PDGFRa H2BGFP mice 4 mice hearts were hashed with anti-MHC I antibodies and pooled (2 undamaged and 2 7 days after LAD ligation). For the Hic1 citrine, cells from 3 undamaged hearts were hashed and pooled together and sequenced as 1 sample.

first: Hesham

last: Soliman

first: Ryan

last: Vander Werff


title: Ascorbate oxidation primes rice plants for enhanced defence against root-knot nematode Meloidogyne graminicola through jasmonate/ethylene-activation

city: Ghent

first: Bruno

last: Verstraeten

state: Oost-Vlaanderen

address: Technologiepark 71

country: Belgium

institute: Ghent University

summary: Ascorbic acid (AA) is known to play a vital role in plant growth and detoxification of reactive oxygen species, however little is known about the significance of AA oxidation in plant defence against pathogens. • The role of ascorbate oxidation in rice defence against root-knot nematodes, Meloidogyne graminicola, was tested with application of AA, ascorbate oxidase (AO), dehydroascorbic acid (DHA), biosynthesis inhibitors and use of mutants. Transcriptome analysis was done on AO treated plants, and hormone measurements were executed to confirm the results. Biochemical analyses were used to study oxidative stress markers, including accumulation of H2O2, , malondialdehyde and AA/DHA.

overall_design: The expression profile of ascorbate oxidase treated plants is compared to untreated plants both for plants infected with Meloidigyne graminicola as for uninfected plants. Three replicates for each group. Each sample was multiplexed in four subsamples.

first: Singh

last: Richard

first: Verstraeten

last: Bruno

first: Siddique

last: Shahid

first: Tegene

last: Adelahu

first: Tenhaken

last: Raimund

first: Frei

last: Michael

first: Haeck

last: Ashley

first: Demeestere

last: Kristof

first: Pokhare

last: Somnath

first: Gheysen

last: Lieve

first: Kyndt

last: Tina


title: CDK-Mediator and FBXL19 cooperate in the induction of developmental genes by promoting regulatory interactions [ATAC-seq]

status: Public on Jan 21 2020

city: Oxford

first: Angelika

last: Feldmann

state: England

address: South Parks Road

institute: University of Oxford, Department of Biochemistry

summary: Appropriate developmental gene regulation relies on the capacity of gene promoters to integrate inputs from distal regulatory elements, yet how this is achieved remains poorly understood. In embryonic stem cells (ESCs), a subset of silent developmental gene promoters are primed for activation by FBXL19, a CpG island binding protein, through its capacity to recruit CDK-Mediator. How mechanistically these proteins function together to prime genes for activation during differentiation is unknown. Here we discover that in mouse ESCs FBXL19 and CDK-Mediator support long-range interactions between silent gene promoters that rely on FBXL19 for their induction during differentiation and gene regulatory elements. During gene induction, these distal regulatory elements behave in an atypical manner, in that the majority do not acquire histone H3 lysine 27 acetylation and no longer interact with their target gene promoter following gene activation. Despite these atypical features, we demonstrate by targeted deletions that these distal elements are required for appropriate gene induction during differentiation. Together these discoveries demonstrate that CpG-island associated gene promoters can prime genes for activation by communicating with atypical distal gene regulatory elements to achieve appropriate gene expression.

overall_design: Genome accessibility of E14 mouse embryonic stem cells was profiled using ATAC-seq assay before and after RA-treatment

first: Emilia

last: Dimitrova

first: Robert

last: Klose


title: Adaptation to industrial stressors through genomic and transcriptional plasticity in a bioethanol producing fission yeast isolate [ChIP-seq]

first: Dane

last: Vassiliadis

summary: Schizosaccharomyces pombe is a model unicellular eukaryote with ties to the basic research, oenology and industrial biotechnology sectors. While most investigations into S. pombe cell biology utilize Leupold’s 972h- laboratory strain background, recent studies have described a wealth of genetic and phenotypic diversity within wild populations of S. pombe including stress resistance phenotypes which may be of interest to industry. Here we describe the genomic and transcriptomic characterization of Wilmar-P, an S. pombe isolate used for bioethanol production from sugarcane molasses at industrial scale. Novel sequences present in Wilmar-P but not in the laboratory S. pombe genome included multiple coding sequences with near-perfect nucleotide identity to Schizosaccharomyces octosporus sequences. Wilmar-P also contained a ~100kb duplication in the right arm of chromosome III, a region harboring ght5+, the predominant hexose transporter encoding gene. Transcriptomic analysis of Wilmar-P grown in molasses revealed strong downregulation of core environmental stress response genes and upregulation of hexose transporters and drug efflux pumps compared to laboratory S. pombe. Finally, examination of the regulatory network of Scr1, which is involved in the regulation of several genes differentially expressed on molasses revealed expanded binding of this transcription factor in Wilmar-P compared to laboratory S. pombe in the molasses condition. Together our results point to both genomic plasticity and transcriptomic adaptation as mechanisms driving phenotypic adaptation of Wilmar-P to the molasses environment and therefore adds to our understanding of genetic diversity within industrial fission yeast strains and the capacity of this strain for commercial scale bioethanol production.

overall_design: ChIP-sequencing of Scr1 in Wilmar-P cells grown in rich (YES) medium containing 3% glucose, or in molasses medium. Scr1-TAP ChIP-seq in Laboratory S. pombe grown in molasses medium. RNA Pol II Ser5 ChIP-seq for laboratory S. pombe and Wilmar-P in YES 3% glucose and molasses medium.

first: Brendon

last: Monahan


title: CDK-Mediator and FBXL19 cooperate in the induction of developmental genes by promoting regulatory interactions [Native ChIP-seq]

overall_design: E14 mouse embryonic stem cells were profiled for H3K4me1 using native ChIP-seq before and after RA treatment


title: Transcriptional profile of PDGFRa+ Sca-1+ and Sca-1-  cells at steady state and at 7, 14 and 28 days post LAD ligation

first: Fabio

last: Rossi

state: BC

overall_design: To  assess differences between Pdgfra+ SCA-1+ and SCA-1- cells, we sorted each subset from the stromal fraction (CD45-/CD31-/ Pdgfra  H2BGFP+) of enzymatically dissociated hearts at steady state and 7, 14 and 28 days post left anterior descending (LAD) coronary artery ligation. RNA was extracted using RNeasy (Qiagen) and the transcriptional profile was generated with the Illumina NextSeq 500 using the High Output 150 cycle Kit.

first: Hesham

last: Soliman

first: Ryan

last: Vander Werff


title: Heart-resident PDGFRa+ SCA-1+ cells


title: Polymerase IV Plays a Crucial Role in Pollen Development in Capsella

city: Uppsala

first: Juan

middle: C

last: Santos-González

state: Uppsala

address: Almas Allé 5

department: Department of Plant Biology and Forest Genetics

country: Sweden

institute: SLU

summary: In Arabidopsis thaliana, DNA-dependent RNA polymerase IV (Pol IV) is required for the formation of transposable element (TE)-derived small RNA (sRNA) transcripts. These transcripts are processed by DICER-LIKE3 into 24-nt small interfering RNAs (siRNAs) that guide RNA-directed DNA methylation. In the pollen grain, Pol IV is also required for the accumulation of 21/22-nt epigenetically activated siRNAs (easiRNAs), which likely silence TEs via post-transcriptional mechanisms. Despite this proposed role of Pol IV, its loss of function in Arabidopsis does not cause a discernable pollen defect. Here, we show that the knockout of NRPD1, encoding the largest subunit of Pol IV in the Brassicaceae species Capsella rubella, caused post-meiotic arrest of pollen development at the microspore stage. As in Arabidopsis, all TE-derived siRNAs were 2 depleted in Capsella nrpd1 microspores. In the wild-type background, the same TEs produced 21/22-nt and 24-nt siRNAs; these processes required Pol IV activity. Arrest of Capsella nrpd1 microspores was accompanied by the deregulation of genes targeted by Pol IV-dependent siRNAs. TEs were much closer to genes in Capsella rubella compared to Arabidopsis thaliana, perhaps explaining the essential role of Pol IV in pollen development in Capsella. Our discovery that Pol IV is functionally required in Capsella microspores emphasizes the relevance of investigating different plant models.

sample_organism:
Arabidopsis thaliana
Capsella rubella

platform_organism:
Arabidopsis thaliana
Capsella rubella

overall_design: 12 small RNA sequencing libraries, 8 RNA-seq libraries, 4 genome-wide cytosine methylation libraries and 6 whole genome sequencing libraries. 4 small RNA libraries were generated from Arabidopsis microspore of Col-0 wt and nrpd1a mutants. 4 small RNA libraries were generated from Capsella microspore of wt and nrpd1a mutants. 4 small RNA libraries were generated from Capsella leaves of wt and nrpd1a mutants. 4 RNA libraries were generated from Arabidopsis microspore of Col-0 wt and nrpd1a mutants. 4 RNA libraries were generated from Capsella microspore of wt and nrpd1a mutants. 4 genome-wide cytosine methylation libraries were generated from Capsella leaves of wt and nrpd1a mutants. 6 whole genome sequencing libraries were generated from Capsella leaves of wt and nrpd1a mutants.

first: Zhenxing

last: Wang

first: Nicolas

last: Butel

last: Santos-Gonzalez

first: Filipe

last: Borges

first: Jun

last: Yi

middle: A

last: Martienssen

first: German

last: Martinez

first: Claudia

last: Kohler


title: CDK-Mediator and FBXL19 cooperate in the induction of developmental genes by promoting regulatory interactions [CaptureC]

last: Feldmann

overall_design:
Mouse embryonic stem cells in which either Fbxl19 CxxC domain or Med13/13l can be conditionally ablated, were profiled for interactions using CaptureC

Please note that each processed data file was generated from multiple samples (as indicated in the description field in the corresponding sample records) and is linked to the corresponding rep1 sample records.


title: Fibroblast Growth Factor-21 Regulates PPARγ Activity and the Antidiabetic Actions of Thiazolidinediones

city: La Jolla

first: Ruth

last: Yu

state: CA

address: 10010 N Torrey Pines Rd

department: Gene Expression Lab

institute: Salk Institute

summary: Fibroblast growth factor-21 (FGF21) is a circulating hepatokine that beneficially affects carbohydrate and lipid metabolism.  Here we report that FGF21 is also an inducible, fed-state autocrine factor in adipose tissue that functions in a feed-forward loop to regulate the activity of peroxisome proliferator-activated receptor γ (PPARγ), a master transcriptional regulator of adipogenesis.  FGF21-knockout (KO) mice display defects in PPARγ signaling including decreased body fat and attenuation of PPARγ-dependent gene expression.  Moreover, FGF21-KO mice are refratory to both the beneficial, insulin-sensitizing effects and the detrimental weight gain and edema side effects of the PPARγ agonist rosiglitazone.  This loss of function in FGF21-KO mice is coincident with a marked increase in the sumoylation of PPARγ, which reduces its transcriptional activity.  Adding back FGF21 prevents sumoylation and restores PPARγ activity.  Collectively, these results reveal FGF21 as a key mediator of the physiologic and pharmacologic actions of PPARγ.

overall_design: RNA was prepared from epididymal white adipose tissue of Diet-induced Obesity (DIO) wild-type and FGF21-KO mice treated with rosiglitazone or vehicle for 14 days (2 pools of 2 mice per condition).

first: Paul

last: Dutchak

first: Steven

last: Kliewer


title: CDK-Mediator and FBXL19 cooperate in the induction of developmental genes by promoting regulatory interactions [ChIP-seq]

first: Angelika

sample_organism:
Drosophila melanogaster
Mus musculus

platform_organism:
Drosophila melanogaster
Mus musculus

overall_design:
Mouse embryonic stem cells in which the CxxC domain of Fbxl19  can be conditionally ablated (Dimitrova et al., eLIFE 290018), were profiled for H3K27ac using spike-in calibrated native ChIP-seq before or after retinoic acid-mediated differentiation

Please note that each  *merged_2replicates.bam.bw file was generated from both replicates and is linked to the corresponding rep1 sample records.

first: Emilia

last: Dimitrova

first: Alexander

last: Kenney

last: Klose


title: Chaperone dependent coordination of nutrient sensing and energy metabolism by mTORC2

city: Beijing

first: Sun

last: Fangfang

address: Zhongguancun Street

institute: Tsinghua University

summary: mTORC2 senses nutrients and coordinates substrate metabolism and macromolecule synthesis program with the availability of external nutrient availability. Knockdown of mTORC2 components and its chaperone partners impairs both nutrient sensing and downstrem metabolism/growth programs.

overall_design: adenovirus-mediated knockdown of Dnajb9 and Rictor with their respective lacZ control groups in mouse liver.

first: Fangfang

last: Sun

first: Zhangming

last: Yan

first: Yilie

last: Liao


title: Comparison of MPA regulated gene expression profiles to those regulated by PROG, DHT, DEX

city: ADELAIDE UNIVERSITY

first: Geraldine

last: Laven-Law

state: South Australia

address: Adelaide Health & Med Sci Bldg, Corner George St & North Terrace

institute: The University of Adelaide

summary:
Medroxyprogesterone acetate (MPA) is a progestin that can bind to and activate progesterone, androgen and glucocorticoid receptors. However, it is not known which receptor mediates MPA action in a cellular context where all three of these receptors are co-expressed and functional.
This microarray experiment was performed to compare the transcriptomes induced by MPA and the cognate ligands for these receptors ie progesterone (PROG), 5a-dihydrotestosterone (DHT) and dexamethasone (DEX) in breast cancer cells to determine which was most similar to MPA.

overall_design: Transcriptome analysis by microarray for ZR-75-1 breast cancer cells treated for 6 hours with 10 nM MPA, PROG, DHT, DEX or vehicle.

first: N

middle: L

last: Moore

first: A

last: Hanson

first: T

last: Hickey

first: W

last: Tilley


title: Human plasma-like medium improves T lymphocyte activation

city: Atlanta

first: Arun

last: Boddapati

state: GA

address: 954 Gatewood Road, NE

department: Genomics Core

institute: Yerkes National Primate research center

summary: T lymphocytes are critical effectors of the immune system and the ability to study their behavior in synthetic media in vitro has facilitated major discoveries in their development, function, and fate. Recently, it has been shown that the constituents in human plasma differ markedly from commonly used synthetic media and these differences have important effects on cell physiology. We therefore compared T lymphocyte activation in human plasma-like medium (HPLM) to RPMI conventional medium both with and without serum. We found that HPLM and RPMI induced vastly different transcriptional programs, especially for transcripts encoding proteins involved in activation, proliferation and metabolism, in human peripheral blood T cells after T cell receptor (TCR) stimulation. We also found that in dialyzed fetal bovine serum (dFBS), HPLM supports a stronger response to TCR engagement compared to RPMI. In reconstruction experiments, we demonstrated that this activation difference is due to RPMI being hypocalcemic relative to the in vivo milieu and that addition of calcium chloride to RPMI can improve human T lymphocyte activation. Thus, we conclude that investigators should be cognizant of differences between commonly used media formulations and the in vivo environment since this could profoundly affect their experimental results and that physiologic media is a valuable new way to study immune cells in culture

overall_design: T cells from healthy human volunteers were isolated from PBMCs via magnetic cell sorting (Miltenyi) and cultured in either RPMI (US biologicals) or HPLM (Cantor et al. 2017). Both cultures were supplemented with dialyzed FBS (Thermo Fisher) at 10% by volume. Maxisorp 96 well plates (Thermo Fisher) were coated with 1 mg/ml of anti-CD3 and anti-CD28 antibodies (clone Hit3a, Biolegend) overnight at 4°C. T cells were then stimulated using these plates at a concentration of 0.5 x 10^6 cells/ml for 48 or 120 hours. For the 120 hour cultures, media was changed and cells were re-normalized to a concentration of 0.5 x 10^6 cells/ml at 48 and 96 hours post activation.100 ng – 1 ug of total RNA as the input to an mRNA capture with oligo-dT coated magnetic beads using Illumina TruSeq protocol. The mRNA is fragmented, and then a random-primed cDNA synthesis is performed. The resulting double-strand cDNA is used as the input to a standard Illumina library prep with end-repair, adapter ligation and PCR amplification and then quantitated by qPCR followed by cluster generation and sequencing.

first: Michael

last: Leney-Greene

middle: K

first: Jason

last: Cantor

first: Helen

last: Su

last: Lenardo


title: miR-26 suppresses adipocyte progenitor differentiation and fat production by targeting Fbxl19

city: Dallas

first: Joshua

last: Mendell

state: TX

address: 6000 Harry Hines Blvd., NA6.200A

department: Molecular Biology

institute: UT Southwestern Medical Center

summary: Purpose: Fat storage in adult mammals is a highly regulated process that involves the mobilization of adipocyte progenitor cells (APCs) that differentiate to produce new adipocytes. Here we report an unexpected role for the broadly conserved miR-26 family of microRNAs (miR-26a-1, miR-26a-2, and miR-26b) as key regulators of APC differentiation. Global loss of miR-26 resulted in a dramatic expansion of adipose tissue in adult mice fed normal chow. Conversely, transgenic overexpression of miR-26a protected mice from high fat diet-induced obesity. These effects were attributable to a cell-autonomous function of miR-26 as a potent inhibitor of APC differentiation. miR-26 blocks adipogenesis, at least in part, by repressing expression of Fbxl19, a conserved miR-26 target without a previously known role in adipocyte biology that encodes a component of SCF-type E3 ubiquitin ligase complexes. These findings have therefore revealed a new pathway that plays a critical role in regulating adipose tissue formation in vivo and suggest new potential therapeutic targets for obesity and related disorders.

overall_design: Method: miR-26 targets in adipocyte progenitor cell differentiation were identified by RNA seq analysis of primary stromal vascular fraction cultured in vitro from i) wild-type C57BL/6 and miR-TKO mice, and ii) M2rtTA; eGFP.miR-26a mice with or without 1 mg/mL dox treatment for 4 days.

first: Asha

last: Acharya

first: He

last: Zhang


title: Silencing Trisomy 21 with XIST in Neural Stem Cells Promotes Neuronal Differentiation (bulk)

first: Jan

last: Czerminski

overall_design: Bulk RNA sequencing of a total of 30 samples from two differentiation timepoints and three independent differentiations. Four dox conditions and three cell lines are included.

first: Jeanne

last: Lawrence


title: Loss of bhlha9 impairs thermotaxis and formalin-evoked pain in a sexually dimorphic manner

city: Marseille

first: Andrew

last: Saurin

address: Campus de Luminy

institute: IBDM / CNRS

summary: This RNA-Seq experiment aims at unraveling the sets of genes that may be deregulated in a specific subset of sensory neurons (the C-Low Threshold MechanoReceptors or C-LTMRs), in absence of transcription factor bhlha9.  In this study, we show that the transcription factor bhlha9 is expressed in a subset of adult C-LTMRs. Moreover, we show that bhlha9 KO males but not female displayed impaired thermotaxis behavior in the temperature gradient paradigm and exhibited exacerbated pain hypersensitivity in the formalin test. To address this question, we FACS-sorted C-LTMRs from adult WT and bhlha9 KO males and females and extracted the total RNA. Further RNA-deep sequencing is a major step to pin-point the sets of genes deregulated selectively in these neurons, in absence of bhlha9.

overall_design: FACS-sorted C-LTMRs from adult WT and bhlha9 KO males and females from two biological replicates of each genotype and sex, total RNA extracted and sequenced

first: Manon

last: Bohic

first: Irène

last: Marics

first: Catarina

last: De Pais Paiva Santos

first: Nissim

last: Ben-Arie

first: Malapert

last: Pascale

first: Salio

last: Chiara

first: Ana

last: Reynders

first: Yves

last: Le Feuvre

first: Aziz

last: Moqrich


title: Genomic landscape of young ATLL patients identifies frequent targetable CD28 fusions

city: Boston

first: Samuel

last: Ng

state: MA

address: 450 Brookline Avenue

department: Medical Oncology

institute: Dana-Farber Cancer Institute

summary: We report fusion genes and expression profiles of 8 young adult T-cell leukemia/lymphoma (ATLL) cases for further characterization of the pathophysiology

overall_design: RNA-seq analysis of 8 young ATLL cases <50

middle: Y

first: Noriaki

last: Yoshida

first: David

last: Weinstock


title: p53 activates the long noncoding RNA Pvt1b to inhibit Myc and suppress tumorigenesis

status: Public on Jan 20 2020

city: Orange

first: Matthew

last: Simon

state: CT

address: West Campus, 100 West Campus Drive, Ste MIC312A

department: MBB

institute: Yale University

summary: The tumor suppressor p53 transcriptionally activates target genes to suppress cellular proliferation during stress. p53 has also been implicated in the repression of the proto-oncogene Myc, but the mechanism has remained unclear. Here, we identify Pvt1b, a p53-dependent isoform of the long noncoding RNA (lncRNA) Pvt1, expressed 50 Kb downstream of Myc, which becomes induced by DNA damage or oncogenic signaling and accumulates near its site of transcription. We show that production of the Pvt1b RNA is necessary and sufficient to suppress Myc transcription in cis without altering the chromatin organization of the locus. Inhibition of Pvt1b increases Myc levels and transcriptional activity and promotes cellular proliferation. Furthermore, Pvt1b loss accelerates tumor growth, but not tumor progression, in an autochthonous mouse model of lung cancer. These findings demonstrate that Pvt1b acts at the intersection of the p53 and Myc transcriptional networks to reinforce the anti-proliferative activities of p53.

overall_design: RNA isolated from mouse lung adenocarcinoma (K-rasLA2-G12D/+; p53LSL/LSL; Rosa26-CreERT2+, KPR) cells previously infected with a sgRNA targeting the Pvt1 p53RE (gΔRE) or a control sgRNA (gCon) +/- tamoxifen treatment and +/- s4U feed was subjected to oxidative-nucleophilic-aromatic-substitution chemistry and sequenced. All sequencing for experimental samples was performed in duplicate per condition assessed, while negative controls were performed as single replicates without s4U treatment.

first: Nadya

last: Dimitrova


title: Extracellular vesicle-microRNA transfer controls germinal center reaction and antibody production

city: Madrid

first: Manuel

middle: J

last: Gomez

state: Madrid

address: Melchor Fernández Almagro, 3

institute: CNIC

summary:
Intercellular communication orchestrates effective immune responses against disease-causing agents. Extracellular vesicles (EVs) are potent mediators of cell-cell communication. EVs carry bioactive molecules, including microRNAs, which modulate gene expression and function in the recipient cell. Here, we show that formation of cognate primary T-B lymphocyte immune contacts promotes transfer to the B cell of a very restricted set of T cell EV-microRNAs (mmu-miR20a-5p, mmu-miR-25-3p and mmu-miR-155-3p). Transferred EV-microRNAs target key genes that control B cell function, including the pro-apoptotic BIM and the cell-cycle regulator PTEN. EV-microRNAs transferred in T-B cognate interactions also promote survival, proliferation and antibody class switching. Using mouse chimeras with Rab27KO EV-deficient T cells, we demonstrate that small EV transfer is required for the germinal center reaction and antibody production in vivo, revealing a novel mechanism that controls B cell responses through the transfer of EV-microRNAs of T cell origin. These findings provide mechanistic insight on the Griscelli syndrome, associated with a mutation in the Rab27a gene and may shed light on this pathogenesis and other immune-related and inflammatory disorders.

This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Self-organizing human trunk 3D neuromuscular organoids

city: Berlin

first: Cornelius

last: Fischer

state: Berlin

address: Robert-Rössle-Straße 10

department: Berlin Institute for Medical Systems Biology

institute: Max Delbrück Center for Molecular Medicine (MDC)

summary: Neuromuscular networks assemble during early human embryonic development and are essential for the control of body movement. Previous neuromuscular junction modeling efforts using human pluripotent stem cells (hPSCs) generated either spinal cord neurons or skeletal muscles in monolayer culture. Here, we use hPSC-derived axial stem cells, the building blocks of the posterior body, to simultaneously generate spinal cord neurons and skeletal muscle cells that self-organize to generate human neuromuscular organoids (NMOs) that can be maintained in 3D for several months. Single-cell RNA-sequencing of individual organoids revealed reproducibility across experiments and enabled the tracking of the neural and mesodermal differentiation trajectories as organoids developed and matured. NMOs contain functional neuromuscular junctions supported by terminal Schwann cells. They contract and develop central pattern generator-like neuronal circuits. Finally, we successfully use NMOs to recapitulate key aspects of myasthenia gravis pathology, thus highlighting the significant potential of NMOs for modeling neuromuscular diseases in the future.

overall_design: We have analyzed single cells from 10 organoids at Day 5. The organoids were separated in two samples (5 pooled organoids each) and labelled with different indexes. In total 5,135 cells were analyzed with a median of 3,333 genes per cell. For Day 50 we analyzed 4 organoids. One sample consisted of one organoid analyzed independently, and the other Day 50 sample consisted of 3 organoids tagged with different indexes. In total 17,294 cells were analyzed with a median of 2,343 genes per cell.

first: Jorge-Miguel

last: Martins

first: Alessia

last: Urzi

first: Ramon

last: Vidal

first: Severine

last: Kunz

first: Pierre-Louis

last: Ruffault

first: Loreen

last: Kabuss

first: Iris

last: Hube

first: Elisabetta

last: Gazzero

first: Carmen

last: Birchmeier

first: Simone

last: Spuler

first: Sascha

last: Sauer

first: Mina

last: Gouti


title: DNA methylation analysis in oropharyngeal cancer

city: Chiba

first: Masaki

last: Fukuyo

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

department: Department of Molecular Oncology

institute: Chiba University

type: Methylation profiling by genome tiling array

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


title: Next Generation Sequencing Facilitates Quantitative Analysis of treated and untreated mortical neurons GDE

city: Los Angeles

first: Kailtyn

middle: Yan-Yun

last: Liu

address: 10833 Le Conte Ave

department: Medicine

institute: UCLA/VA

summary: Traumatic Brain Injury (TBI) is associated with disruption of cerebral blood flow leading to localized brain hypoxia. Thyroid hormone (TH) treatment, when administered shortly after injury, has been shown to promote neural protection in rodent TBI models. The mechanism of TH protection, however, is not established. We used mouse primary cortical neurons to investigate the effectiveness and mechanism of T3-promoted cell survival after exposure to hypoxic injury. Cultured primary cortical neurons were exposed to hypoxia (0.2% oxygen) for 7 hours, in either the presence or absence of T3 (5 nM). T3 treatment enhanced DNA 5-hydroxymethylcytosin (5-hmc) levels and attenuated the hypoxia-induced increase in DNA 5-methylcytosin (5-mc). In the presence of T3, mRNA expression of Tet family genes was increased and DNA methyltransferases, (Dnmt) 3a and Dnmt3b, were downregulated, compared to conditions in the absence of T3. These T3-induced changes decreased hypoxia-induced DNA de novo methylation, which reduced hypoxia-induced neuronal damage and apoptosis. We utilized RNA-seq to characterize T3-regulated genes in cortical neurons under hypoxic conditions and identified 22 genes that were upregulated and 15 genes that were downregulated. Krupple-like factor 9 (KLF9), a multifunctional transcription factor that plays a key role in CNS development, was highly upregulated by T3 treatment in hypoxic conditions. Knockdown of the KLF9 gene resulted in early apoptosis and abolished the beneficial role of T3 in neuronal survival. KLF9 mediates, in part, the neuronal protective role of T3. T3 treatment reduces hypoxic damage and acts through pathways that reduce DNA methylation and apoptosis.

overall_design: Comparing GDE in primary cortical neuorons with different treatment (control, hypoxia, TH treatment)

first: Kaitlyn

first: Gregory

last: Brent

first: Jainrong

last: Li


title: CDK-Mediator and FBXL19 cooperate in the induction of developmental genes by promoting regulatory interactions

first: Angelika

last: Feldmann


title: Transcriptomic analysis of neuroblastoma SH-SY5Y cells in response to stable over-expression of neuroblastoma highly expressed 1 (NHEG1)

city: Wuhan

first: Qiangsong

last: Tong

state: Hubei

address: 1277 Jiefang Avenue

department: Department of Surgery

institute: Union Hospital of Tongji Medical College

summary:
Neuroblastoma (NB), a malignant embryonic tumor arising from primitive neural crest cells, accounts for more than 7% of malignancies and around 15% of cancer-related mortality in childhood. Better elucidating the mechanisms of tumorigenesis and aggressiveness is important for improving the therapeutic efficiencies of NB. Through mining public microarray and RNA sequencing datasets, we identified neuroblastoma highly expressed 1 (NHEG1) as a novel 1360-bp lncRNA associated with poor outcome of NB.
To investigate the mechanisms underlying the oncogenic functions of NHEG1, we employed the human whole genome microarray expression profiling as a discovery platform to analyze the transcriptome profiling changes of human SH-SY5Y cells in response to stable over-expression of NHEG1. The results showed that stable over-expression of NHEG1 led to altered expression of 869 human mRNAs, including 499 up-regulated genes and 370 down-regulated genes. Then we found the possible roles of these differentially regulated mRNAs in selected pathways including cell cycle/proliferation, apoptosis, and cytokine/chemokine responses by Bioinformatic analysis. Furthermore, we validated the microarray results by real-time RT-PCR with high identity. Overall, our results provided fundamental information about the transcriptomic changes in response to NHEG1 over-expression in human NB cells, and these findings will help us understand the pathogenesis of NB.

overall_design: Total RNA of cells stably transfected with empty vector or NHEG1 was extracted using the TRIZOL Reagent according to the manufacturer's instructions. Gene expression profiling was performed for each RNA sample separately on the Affymetrix Human Transcriptome Array 2.0 at Shanghai Technology Corporation (Shanghai, China), in which GeneChip microarray service was certificated by Affymetrix.

first: Liduan

last: Zheng


title: DNA methylation analysis in oropharyngeal cancer [EPIC array]

summary: DNA methylation analysis in oropharyngeal squamous carcinoma (OPSCC) samples and oropharyngeal non-cancerous mucosa samples. Infinium MethylationEPIC BeadChip Kit was used to obtain DNA methylation profiles across more than 850,000 CpG sites. Total samples included 89 OPSCC samples and 5 non-cancerous mucosa samples.

overall_design: Bisulphite converted DNA from the 25 OPSCC samples were hybridised to the Infinium MethylationEPIC Beadchip Kit.

first: Takuya

last: Nakagawa

first: Atsushi

last: Kaneda


title: DNA Double Strand Break Repair in E. coli Perturbs Cell Division and Chromosome Dynamics

city: Cambridge

first: Martin

middle: Andrew

last: White

address: Northwest Building room 140, 52 Oxford Street

department: Molecular and Cellular Biology

institute: Harvard University

summary: The effect of a site-specific DNA double-strand break on the abundance of E. coli chromosomal DNA during exponential growth was investigated by marker frequency analysis (MFA).

overall_design: Genomic DNA from 3 independent biological repeats of 4 strains (DL1777, DL2151, DL2859 and DL2874) was sequenced by whole genome sequencing. The genomic DNA from strain DL1777 was isolated from cultures at both stationary and exponential growth phases, whereas the other genomic DNA of the other three strains were isolated from cultures at exponential growth phase only.  Strain DL2859 experiences the site-specific DSB, whereas strains DL1777, DL2151, and DL2874 do not.  Genomic DNA from stationary phase cultures of strain DL1777 was used to control for differences in read number caused by genomic composition (rather than chromosome metabolism).

first: Elise

last: Darmon

first: Manuel

last: Lopez-Vernaza

first: David

last: Leach


title: MiR-29c attenuates pulmonary fibrosis by promoting epithelial renewal and inhibiting epithelial apoptosis

first: ting

last: Xie

address: 127 San Vicente Blvd

department: Pulmonary

institute: Cedars-Sinai Medical Center

summary:
The successful repair and renewal of alveolar epithelial cells are critical steps in prohibiting the accumulation of myofibroblasts and deposition of extracellular matrix in pulmonary fibrogenesis. MicroRNAs (miRNAs) are multi-focal regulators involved in the lung repair process. The contribution of miRNAs to epithelial maintenance and renewal is incompletely understood. We provide evidence that miR-29c on type 2 alveolar epithelial cells (AEC2s) are important for inhibiting AEC2 apoptosis and promoting AEC2 renewal, thus restraining the degree of fibrosis. MiR-29c was lower in AEC2s from lungs of idiopathic pulmonary fibrosis (IPF) individuals than from healthy lungs. Epithelial cells overexpressing miR-29c showed higher proliferative rate and viability. MiR-29c was protective against epithelial apoptosis by targeting Foxo3a. Both overexpression of miR-29c conventionally and AEC2s specifically led to less fibrosis and better recovery. Furthermore, loss of miR-29c in AEC2s resulted in higher apoptosis and reduced epithelial renewal than in control animals. Thus, miR-29c maintains epithelial integrity and promotes recovery from lung injury, attenuating lung fibrosis in mice.
The miR-29c overexpression mouse epithelial cell line was generated based on the strategy described before38. MiR-29c lentiviral construct was generated as follows: a region of 131 nucleotides containing pre-mmu-miR-29c was amplified from mouse genomic DNA by using the following primers: 5’ MIR-29c: GTCGGTTAACATCTCTTACACAGG and 3’ MIR-29c: ACACCTCGAGGATCCTGAGGCTGGT. They were then cloned into the HpaI/XhoI sites of a pSico vector62, named pSico-miR-29c. Viral particles were produced by calcium phosphate–mediated transfection into 293T cells as described63. Lentiviral supernatants were collected 48 hours after transfection, passed through a 0.22-μm filter, and used directly to infect MLE 12 cells. GFP-positive cells were sorted 2-3 days after infection and resulted in MLE 12-SicoGFP and MLE 12-miR-29cGFP cells. Recombinant adenoviral stocks expressing Cre recombinase were purchased from the Gene Transfer Vector Core facility of University of Iowa College of Medicine (Iowa City, IA). Infections of GFP-positive cells were performed by using 100 plaque-forming units of virus per cell. Four days after adenovirus infections, the GPF-negative cells were sorted from GFPpos infected with adeno-Cre as Mlesico and Mle29c cells.

overall_design: Microarray analyses for Mlesico and Mle29c cells

first: Ting

first: Dianhua

last: Jiang


title: DNA methylation analysis in oropharyngeal cancer [450K array]

first: Masaki

summary: DNA methylation analysis in oropharyngeal squamous carcinoma (OPSCC) samples and oropharyngeal non-cancerous mucosa samples. Infinium HumanMethylation450 BeadChip was used to obtain DNA methylation profiles across 485,577 CpG sites. Total samples included 89 OPSCC samples and 5 non-cancerous mucosa samples.

overall_design: Bisulphite converted DNA from the 64 OPSCC samples and 5 non-cancerous mucosa samples were hybridised to the Infinium HumanMethylation450 BeadChip.


title: Real-time quantitative PCR analysis of human EBV-B cells during UPR

status: Public on Jan 19 2020

city: São Paulo

first: Maristela

middle: Martins de

last: Camargo

state: SP

address: Av. Prof. Lineu Prestes 1730

department: Immunology

institute: University of São Paulo

summary: Human B cells were isolated from peripheral blood from one healthy donor and two patients with Common Variable Immunodeficiency (CVID) and were immortalized by infection with Epstein Barr virus (EBV) in vitro. Immortalized EBV-B cells (in biological triplicates) were treated twelve different ways for 4 hours: 1.untreated control, 2. Thapsigargin (Tg) 1 µM, 3. Tunicamycin (Tm) 10 µg/ml, 4. 4-phenylbutyrate (4-PBA) 1 mM, 5. Tg 1 µM plus 4-PBA 1 mM, 6. Tm 10 µg/ml plus 4-PBA 1 mM, 7. Dimethyl sulfoxide (DMSO) 1 mM, 8. Tg 1 µM plus DMSO 1 mM, 9. Tm 10 µg/ml plus DMSO 1 mM, 10. Tauroursodeoxycholic acid (TUDCA) 5 mM, 11. Tg 1 µM plus TUDCA 5 mM, and 12. Tm 10 µg/ml plus TUDCA 5 mM. We used Applied Biosystems TaqMan OpenArray Real-Time PCR System (Thermo Scientific) to quantitate gene expression of relevant genes from the cells (in technical triplicates).

overall_design: qPCR gene expression profiling.  EBV-immortalized B cells from three donors were used and treated separately as indicated in the summary. Equal amount total RNA from each donor was used for cDNA synthesis, and normalized amounts of cDNA were used for gene expression analysis (in technical triplicates). Samples were normalized to Gapdh expression. Fold changes were calculated as a ratio of T/NT (treated / non treated) expressed as log base 2.


title: Combined inhibition of WNT and NOTCH signaling as a therapeutic strategy for human clear cell kidney carcinoma

first: Annika

last: Fendler

address: Robert-Roessle-Str. 10

department: Signal Transduction in Development and Cancer

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

summary: Current therapies for metastasized kidney cancer in humans include treatments with multi-tyrosine kinase inhibitors, monoclonal antibodies against VEGF or inhibitors of mTOR. While many tumors initially respond to these treatments, nearly all resume their progression within two years. Here we characterize CXCR4+MET+CD44+ cancer stem cells in clear cell renal cancer (ccRCC), whose presence represents a major subgroup of the disease that correlates with the aggressiveness of the tumors and poor patient outcomes. Transcriptional profiling revealed that WNT and NOTCH signaling are  activated in ccRCC cancer stem cells.

overall_design: Total RNA was extracted from non-cultured primary clear cell renal cell cancer (ccRCC) cells of three individual patients that were isolated after nephrectomy, sorted CXCR4+MET+CD44+ cells or spheres cultured for 7 days in non-adherent, serum-free condition.

first: Jonas

last: Busch

first: Klaus

last: Jung

last: Wulf-Goldenberg

first: Bettina

last: Ergün

first: Simone

first: Wei

last: Chen

first: Walter

last: Birchmeier


title: Microglial homeostasis requires balanced CSF-1/CSF-2 receptor signaling

status: Public on Jan 17 2020

city: Bronx

first: Deyou

last: Zheng

state: NY

address: 1301 Morris Park Ave

institute: Albert Einstein College of Medicine

summary:
ALSP is a CSF1R-deficiency dementia associated with increased CSF2 expression
Monoallelic deletion of Csf2 rescues most behavioral deficits and histopathological changes in Csf1r+/- mice by preventing microgliosis and eliminating most microglial transcriptomic alterations, including those indicative of oxidative stress and demyelination
Elevation of Csf2 transcripts and of several CSF-2 downstream targets in the brains of ALSP patients demonstrates that the mechanisms identified in the mouse model are functional in man.
Our data provide new insights into the mechanisms underlying ALSP and identify CSF-2 as a therapeutic target.

overall_design: Examination of the effect of single or double Csf1r and Csf2 heterozygosity on behavior, brain histopathology and microglial transcriptomic profiles.

first: Violeta

last: Chitu

first: Ping

last: Wang

first: E

middle: R

last: Stanley