title: RNA-seq analysis of axolotl embryonic oropharyngeal endoderm explants transcriptome

status: Public on Jun 02 2020

city: New London

first: Deborah

last: Eastman

state: CT

address: 270 Mohegan Ave

institute: Connecticut College

summary: The ability to taste is critical for animal nutrition and toxin deterrence. The majority of research on taste bud development and regeneration is in mice, where taste buds are located within specialized papillae on the tongue.  However, taste buds in fish and amphibians, such as axolotls (Ambystoma mexicanum), are not housed in papillae, rather they are within the pharyngeal epithelium.  This simplified tissue level organization, along with the ability of cultured oropharyngeal explants from early embryos to produce taste buds on the same time-line as embryos, make the axolotl an excellent model to identify molecules specifically involved in taste bud cell differentiation. In this study, we performed de novo transcriptomic analysis on RNA sequences from three different stages of oropharyngeal explants: stages 37/38, 39, and 41.  RNA-seq data from 17 total samples representing these stages were pooled to generate de novo assemblie(s) of the transcriptome using a Trinity pipeline. Raw reads and the assembly were uploaded to Mendeley (doi:10.17632/tvxh3jm83m.1). From 27.9Gb of raw sequences, we identified 21,244 transcripts.  To our knowledge, this study provides the first published assembly of axolotl oropharyngeal endoderm explants. This RNA-seq data and transcriptome assembly provide information on genes expressed in the oropharyngeal endoderm and will be valuable in the identification of taste bud development genes.

overall_design:
Axolotl embryos were obtained from the Ambystoma Genetic Stock Center at the University of Kentucky and incubated in 20% Holtfreter’s solution at 22°C. Oropharyngeal endodermal explants were isolated at stage 15. Explants were harvested at stages 37/38, 39 and 41.  Replicate samples for each stage were analyzed for RNA quantity and quality on a Nanodrop 8000 spectrophotometer. Single-end read RNAseq of RNA isolated from axolotl embryonic oropharyngeal endoderm explants. High-throughput sequencing performed with Illumina NextSeq 500 System.

Please note that the gene isoform counts were collapsed to gene counts for quantification across the samples (as in the 'Trinity.fasta.gene_trans_map')

first: Lauren

last: Marazzi

first: Priya

last: Kohli


title: single-cell RNA-seq in alveolar organoids cultured in MTECplus medium

city: Durham

first: Yoshihiko

last: Kobayashi

state: NC

address: 307 Research Dr

department: Cell Biology

country: USA

institute: Duke University

type: Expression profiling by high throughput sequencing

summary: We performed Drop-seq using alveolar organoids cultured in MTECplus medium to identify cell types, gene expression patterns and signaling pathways in ex vivo condition.

overall_design: single-cell RNA-seq (Drop-seq) in day 10 cultured alveolar organoids.

first: Hiroaki

last: Katsura

first: Purushothama

middle: R

last: Tata


title: Histone modifications in lung epithelial cells

type: Genome binding/occupancy profiling by high throughput sequencing

summary: We report histone modifications in alveolar epithelial type 2 cells in homeostatic mice as well as CTGF-positive alveolar eptihelial cells in mice treated with Bleomycin.

overall_design: We performed Mint-ChIP (ChIP-seq for low cell number samples) using antibodies for H3K4me3, H3K27ac, H3K36me3, H3 and p53)

first: Aleksandra


title: Parallel Bimodal Single-cell Sequencing of Transcriptome and Chromatin Accessibility

status: Public on Jun 09 2020

city: Singapore

first: Chadi

last: El Farran

address: 61 Biopolis Drive

country: Singapore

institute: Institute of Molecular and Cell Biology

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

first: Qiaorui

last: Xing

first: C

last: EL Farran

first: Hu

last: Li

first: Yuin-Han

last: Loh


title: Gene expression profiling of FACS-purified striatal medium spiny neurons and striatum in Huntington's disease model mouse

status: Public on Jun 03 2020

city: Kyotanabe-shi

first: Haruko

last: Miyazaki

state: Kyoto

address: 1-3 Tatara Miyakodani

department: Graduate School of Brain Science

country: Japan

institute: Doshisha University


title: Exome mutations in mouse LMP1 Lymphomas

city: Berlin

first: Thomas

last: Sommermann

address: Robert-Rossle-Str. 10

country: Germany

institute: Max Delbruck Center for Molecular Medicine

summary: 10 LMP1 expressing mouse lymphomas were sorted and sequenced for exome mutations

overall_design: 10 Rag2KOcgKO were reconstituted with  fetal liver hematopoietic stem and progenitor cells (HSPC) from Cd3eKO Cd19-Cre R26LMP1stopf mice. Developing LMP1-driven B-cell lymphomas were transplanted in secondary  Rag2KOcgKO or  Rag2KO mice. Secondary lymphomas were FACSpurified (CD19+huCD2+). gDNA of sorted lymphoma samples was analyzed by exmoe sequencing

first: Klaus

last: Rajewsky

first: Jonathan

last: Ronen


title: Genome-wide mapping of AP sites, binding of repair proteins and G4 structure formation

city: Omaha

first: Shrabasti

last: Roychoudhury

state: NE

address: S 42nd &, Emile St

institute: University of Nebraska Medical Center

summary: We demonstrate that there is a genome-wide correlation between endogenous AP site damages, binding of repair proteins APE1, AcAPE1 and AcOGG1 and G4 structure formation.

overall_design:
Examination of endogenous AP site formation, binding of repair proteins APE1, AcAPE1 and AcOGG1 and formation of G4 structure.

To identify genes that are regulated by APE1 and G4 structures, RNA-Seq analysis was performed with control and Dox-inducible APE1 knock-down (APE1 KD) A549 cells.

first: Bhakat

last: Kishor

first: Roychoudhury

last: Shrabasti


title: RNA-expression profiling of LMP1 expressing mouse B cells before and after transformation in vivo

summary: Non-transformed LMP1 expressing mouse B cells or mouse LMP1 lymphomas were  analysed by RNA-sequencing

overall_design: 14 Rag2KOcgKO were reconstituted with  fetal liver hematopoietic stem and progenitor cells (HSPC) from Cd3eKO Cd19-Cre R26LMP1stopf mice. From 3 mice LMP1 expressing non-transformed B cells were isolated day 18-20 post reconstitution. 11 mice developed LMP1-driven B-cell lymphomas. Thse lymphomas were transplanted into Rag2KOcgKO or Rag2KO mice. Arising secondary lymphomas were FACSpurified (CD19+huCD2+). RNA of sorted lymphoma samples was analysed by RNA sequencing.


title: Tanscriptional profile of primary endothelial cells (HUVEC) in ADA2 depleted setting and effect of Dipyridamole (DPM) treatment

status: Public on Jun 01 2020

city: San Diego

first: Rekha

last: Dhanwani

state: CA

address: 9420 Athena Cir

institute: La Jolla Institute for Immunology

summary: Adenosine deaminase 2 deficiency (DADA2) is an inherited disorder which can cause vasculitis of medium-sized blood vessels. The disease is caused by mutations in the CECR1 gene, which encodes a key metabolic enzyme in the purine pathway. It is not clear how DADA2 leads to vasculitis, and there are currently limited treatment options. We, therefore, as a first step aimed to study the altered gene expression profile of primary endothelial cells in ADA2 depleted setting and then assessed the effect of Dipyridamole (DPM), a pharmacological inhibitor of ENT- transporters, on the differentially expressed genes.  The whole genome transcriptome analysis revealed a robust induction of an IFNβ-stimulated gene signature in unstimulated siADA2-treated cells. Differentially expressed transcripts in this dataset included genes encoding pro-inflammatory cytokines, chemokines, and innate immune response proteins. Pre-treatment with dipyridamole (DPM), reduced expression of the IFNβ-driven gene signature upon depletion of ADA2.

overall_design: Two experiments were performed. In the first experiment 2 conditions in triplicates were considered: 1. HUVEC transfected with non-targeting si RNA referred to as siControl 2. HUVECs transfected with siRNA targeing ADA2 molecule referred to as siADA2. In the second experiment 3 conditions in triplicates were considered: 1. HUVEC transfected with non-targeting si RNA referred to as control 2. HUVECs transfected with siRNA targeing ADA2 molecule referred to as CECR1 3. HUVECs transfected with siRNA targeing ADA2 molecule and pre-treated with Dipyridamole (DPM) refrered as CECR1_DPM

first: Sonia

last: Sharma


title: A simple and highly efficient method for multi-allelic CRISPR-Cas9 editing in primary cell cultures

city: Seattle

first: Sonali

last: Arora

state: WA

address: 1100 Fairview Ave N,

institute: FHCRC

summary:
Background: CRISPR-Cas9-based technologies have revolutionized experimental manipulation of mammalian genomes. None-the-less, limitations of the delivery and efficacy of these technologies restrict their application in primary cells.
Aims: To create an optimized protocol for penetrant, reproducible, and fast targeted genome editing in cell cultures derived from primary cells, using patient-derived glioblastoma (GBM) stem-like cells (GSCs) and human neural stem/progenitor cells (NSCs) for proof-of-concept experiments.
Methods and results: We employed transient nucleofection of Cas9:sgRNA ribonucleoprotein complexes composed of chemically synthesized 2'-O-methyl 3’phosphorothioate-modified sgRNAs and purified Cas9 protein. Insertion-deletion mutation (indel) frequency and size distribution were measured via computational deconvolution of Sanger sequencing trace data. We found that this optimized technique routinely allows for >90% indel formation in only 3 days, without the need to create clonal lines for simple loss-of-function experiments. Using Western blotting, we observed near-total protein loss of target genes in cell pools. Additionally, we found that this approach allows for the creation of targeted genomic deletions. By using RNA-seq in edited NSCs to assess gene expression changes resulting from knockout of tumor suppressors commonly altered in GBM, we also demonstrated the utility of this method for quickly creating a series of gene knockouts that allow for the study of oncogenic activities.
Conclusion: Our data suggest that this relatively simple method can be used for highly efficient and fast gene knockout, as well as for targeted genomic deletions, even in hyperdiploid cells (such as GSCs). This represents an extremely useful tool for the cancer research community when wishing to inactivate not only coding genes, but also non-coding RNAs, UTRs, enhancers, and promoters. This method can be readily applied to diverse cell types by varying the nucleofection conditions.

overall_design: To assess gene expression changes resulting from loss of tumor suppressors commonly altered in glioblastoma, we performed RNA sequencing on hTERT-transduced neural stem cells (NSC-CB660-TERT) harboring various successive knockouts of TP53, CDKN2A, PTEN, and NF1. Knockouts were created by transiently nucleofecting NSC-CB660-TERT cells with Cas9:sgRNA ribonucleoprotein complexes composed of purified Cas9 protein and chemically synthesized 2'-O-methyl 3’phosphorothioate-modified sgRNAs. Three different sgRNAs were used simultaneously for targeting each gene. The pools of various KO cells were sequenced, along with one cell clone harboring knockout of all 4 genes.

first: Pia

last: Hoellerbauer

first: Megan

last: Kufeld

first: Hua-Jun

last: Wu

first: Heather

middle: M

last: Feldman

first: Patrick

middle: J

last: Paddison


title: Transcriptome of the panx1a-knock out zebrafish

city: Toronto

first: Georg

last: Zoidl

state: Ontario

address: 4700 Keele Street

department: Biology

country: Canada

institute: York University

summary: Pannexin 1 (Panx1) forms ATP-permeable membrane channels that play roles in the nervous system. The analysis of roles in both standard and pathological conditions benefits from a model organism with rapid development and early onset of behaviors. Such a model was developed by ablating the zebrafish panx1a gene using TALEN technology. Here, RNA-seq data of 6dpf wild type and knock-out zebrafish is reported which have been generated by gene editing of exon 4 of the panx1a gene.

overall_design: Paired end sequencing of

first: Nickie

last: Safarian


title: RNA-expression profiling of mouse B cells coexpressing LMP1 with EBNA3A, Rel or Ebf1 in vitro

summary: mouse B cells coexpressing LMP1 with EBNA3A, Rel or Ebf1 were cultured for 6 days in vitro and analysed by RNA-sequencing

overall_design: CD43- splenic B cells from R26LMP1stopf mice were isolated and treated with TAT-cre (iLMP1 B cells). After 24h cells were tranduced with MSCV (MIG)-vectors encoding indicated genes. At day 6 cells were sorted for MSCV-reporter (GFP) expression. Alternatively, CD43- splenic B cells from R26LMP1stopf and R26LMP1stopf/R26EBNA3Astopf mice were isolated, treated with TAT-cre (iLMP1 and iLMP1/EBNA3A B cells) and sorted at day 6. RNA from sorted cells was isolated and RNA-sequenced.


title: Genome regulation and gene interaction networks inferred from muscle transcriptome underlying feed efficiency in Pigs

city: Kongens Lyngby

first: Haja

middle: N

last: Kadarmideen

address: Richard Petersens Plads, Building 324

department: Department of Applied Mathematics and Computer Science

country: Denmark

institute: Technical University of Denmark

summary: Feed efficiency (FE) is a key trait in pig production, as it has both high economic and environmental impact. FE is a challenging phenotype to study, as it is complex and affected by many factors, such as metabolism, growth and activity level. Furthermore, testing for FE is expensive, as it requires costly equipment to measure feed intake of individual animals, making FE biomarkers valuable. Therefore, there has been a desire to find single nucleotide polymorphisms (SNPs) as biomarkers, to assist with improved selection and improve our biological understanding of FE. We have done a cis- and trans- eQTL (expressed quantitative trait loci) analysis, in a population of Danbred Durocs (N=11) and Danbred Landrace (N=27) using both a linear and Anova model. We used bootstrapping and enrichment analysis to validate and analyze our detected eQTLs. We identified 15 eQTLs with FDR < 0.01, affecting several genes found in previous studies of commercial pig breeds. Examples include IFI6, PRPF39, TMEM222, CSRNP1,PARK7 and MFF. The bootstrapping results showed statistically significant enrichment of eQTLs with p-value < 0.01 (p-value < 2.2x0-16) in both cis and trans-eQTLs. Based on this, enrichment analysis of top trans-eQTLs revealed high enrichment for gene categories and gene ontologies associated with genomic context and expression regulation. This includes transcription factors (p-value=1.0x10-13), DNA-binding (GO:0003677, p-value=8.9x10-14), DNA-binding transcription factor activity (GO:0003700,) nucleus gene (GO:0005634, p-value<2.2x10-16), positive regulation of expression (GO:0010628), negative regulation of expression (GO:0010629, p-value<2.2x10-16). These results would be useful for future genome assisted breeding of pigs to improve FE, and in the improved understanding of the functional mechanism of trans-eQTLs.

overall_design: RNA-seq was performed on 41 male uncastrated pigs (13 Danbred Duroc and 28 Danbred Landace). Muscle tissue from the psoas major muscle was collected imediatly post-slaughter at 100kg. Feed intake and growth of the pigs was measured during a testing period from ~28kg to ~100kg. Based on these measurements FCR and RFI was calculated for all pigs.

first: Victor

middle: A

last: Carmelo


title: Post-transational regulation of the exon skipping machinery controls aberrant splicing in T cell leukemia [Jurkat_SI_24hr_300bp]

city: Chicago

first: Yalu

last: Zhou

state: IL

address: 303 E. Superior St,

department: Department of Biochemistry and Molecular Genetics

institute: Northwestern University

summary: In this study, we show that pediatric T-cell acute lymphoblastic leukemia (T-ALL) has an alternative mechanism for aberrant splicing that involves post-translational regulation of the splicing machinery via deubiquitination.

overall_design: Whole RNA was extracted from 1-5 million T-ALL (lines) cells or primary cells using the Total RNA Mini Kit (Bio-Rad) according to the manufacturer’s protocol. Poly-A+ (magnetic oligodT-containing beads (Illumina)) or Ribominus RNA was used for library preparation. cDNA preparation and unstranded library construction was performed using the TruSeq RNA Sample Preparation Kit. Libraries were sequenced on the NextSeq 500/HiSeq 2500 using 76bp, 50bp or 151bp paired-end read method. Differential gene expression analysis was performed for primary T cells vs T-ALL patients samples, or each matched treatment vs control pairs, separately in each biological or technical replicate in each of two cell lines (CUTLL1, Jurkat). Seven types of comparisons were tested: (a) T-Cells vs T-ALL, (b) Non high risk T-ALL vs high risk T-ALL, (c) Control vs H3B-8800 treated Jurkat cells, (d) Control vs H3B-8800 treated CUTLL1 cells, (e) Control vs P5091 treated Jurkat cells, (f) Control vs P5091 treated CUTLL1 cells, (g) Control vs SRSF6 knockdown Jurkat cells. Analysis was performed using edgeR package.

first: Panagiotis

last: Ntziachristos


title: Novel Inhibitors of the Histone-Methyltransferase DOT1L Show Potent Antileukemic Activity in Patient-derived Xenografts

city: Hannover

first: Florian

last: Perner

address: Carl-Neuberg-Str. 1

department: Hematology, Hemostasis, Oncology and Stem Cell Transplantation

institute: Hannover Medical School


title: Transcription Analysis of Clustered Essential Genes

city: 天津市

first: peixia

last: wang

address: 天津市津南区海河教育园区雅观路135号天津大学北洋园校区54教203

country: China

institute: 天津大学

summary: All 14 essential genes of chrIII were clustered and reordered, and transcription of the genes was tested. We designed and constructed three types of reorganized essential genes into three different plasmids. For the genomic direction (GD) plasmid, the essential genes were arranged according to their natural chromosomal positions and orientations. For the same direction (SD) plasmid, the essential genes were arrayed in the same positions as GD plasmid, but all genes were assembled in the same direction (SD). For the random direction (RD) plasmid, the essential genes were modularly assembled according to the combined length of three individual genes and thus displayed disorderly. The synIII strains bearing the GD/SD/RD plasmids were phenotypically normal despite of the double expression of 14 essential genes.

overall_design: PCRTags were used to distinguish the transcription of wild-type genes from the transcription of synthetic genes.

first: W

last: Yi


title: Novel Inhibitors of the Histone-Methyltransferase DOT1L Show Potent Antileukemic Activity in Patient-derived Xenografts (ChIP-seq dataset)

summary: using ChIP-seq we examined Chromatin occupancy of the MLL-fusion complex members Menin and DOT1L after treatment with DOT1L methyltransferase inhibitors EPZ5676 and the novel compounds "compound 10" and "compound 11" as well as with the Menin-inhibitor VTP50469 in MOLM13 cells. Furthermore we used ChiP-seq to quantify the loss of H3K79me2 (histone mark deposited by DOT1L) after treatment with the DOT1L inhibitors EPZ5676 and "compound 10" in MOLM13 and RS4,11 cells.

overall_design: Menin and DOT1L occupancy after treatment with DOT1L inhibitors and a Menin-inhibitor was assessed 4d after treatment with the drugs in MOLM13 cells via ChIP sequencing. Deposition of the histone mark H3K79me2 was measured at days 3, 6 and 9 after DOT1L-inhibitor treatment in MOLM13 and RS4,11 cells. For the analysis and normalization of the H3K79me2 ChIPseq we used drosophila spike-in (dm6 spike-in).

first: Charlie

last: Hatton


title: Novel Inhibitors of the Histone-Methyltransferase DOT1L Show Potent Antileukemic Activity in Patient-derived Xenografts (RNA-seq dataset)

summary: Using 3-end RNA-seq followed by digital gene expression analyses we assessed genes that change expression upon treatment with the DOT1L-methyltransferase inhibitors EPZ5676 and the novel drug "compound 10"

overall_design: gene expression was assessed in EPZ5676, compound 10 or DMSO treated MOLM13 cells at 3d, 6d and 9d post-treatment


title: MicroRNA-seq data of mouse during skeletal muscle regeneration after acute injury

status: Public on Jun 04 2020

city: Beijing

first: naixuan

last: cheng

address: Capital Medical University, No.10 Xitoutiao, You An Men, Fengtai District

institute: School of Basic Medical Sciences, Capital Medical University

type: Non-coding RNA profiling by high throughput sequencing

summary: MicroRNAs (miRNAs) are short noncoding RNAs that impact protein expression, regulating a variety of physiological processes. To date, a few miRNAs have been implicated in the process of muscle regeneration after injury; however, our understanding of the miRNAs involved and their regulatory mechanisms remains incomplete.The analysis showed differences of microRNAs in the tibialis anterior muscles at days 0, 1 and 3 post-cardiotoxin injury.

overall_design: Examination of MicroRNA profiles of 10-week old wild type (WT)  tibialis anterior muscle at days 0, 1 and 3 post-cardiotoxin injury

first: Naixuan

last: Cheng

first: Congcong

last: Zhang


title: A negative reciprocal regulatory axis between cyclin D1 and HNF4α modulates cell cycle progression and metabolism in the liver

city: Miinneapolis

first: Jeffrey

middle: H

last: Albrecht

state: MN

address: One Veterans Drive

department: Medicine - Gastroenterology

institute: University of Minnesota

summary: This study examines the regulation of HNF4a by cyclin D1 in hepatic cell lines and mouse liver.

overall_design: Three different cell lines were treated with control of cyclin D1 siRNA, followed by RNA-seq. Mice had transduction of cyclin D1 (or a truncation mutant called cyclin D1-RD) into the liver using recombinant adenovirus, followed by HNF4a ChIP-seq.

first: Heng

first: Yitzhak

last: Reizel

first: Yue

last: Wang

first: Jessica

middle: L

last: Lapiro

first: Betsy

middle: T

last: Kren

first: Jonathan

last: Schug

first: Shilpa

last: Rao

first: Ashleigh

last: Morgan

first: Adam

last: Herman

first: Laurie

last: Shekels

first: Mathew

last: Rassette

first: Andrew

last: Lane

first: Teresa

last: Cassel

last: Fan

first: Juan

middle: C

last: Manivel

first: Sumedha

middle: S

last: Gunewardena

first: Udayan

last: Apte

first: Piotr

last: Sicinski

first: Klaus

last: Kaestner


title: ZipSeq : Barcoding for Real-time Mapping of Single Cell Transcriptomes

city: Houston

first: Kenneth

last: Hu

state: Texas

address: 7455 Fannin Street

department: Immunology

institute: MD Anderson

summary: single cell RNA Sequencing runs for various models linking transcriptomes with regional position using spatially printed barcodes

overall_design:
Using ZipSeq, we mapped gene expression in three settings: in-vitro wound healing, live lymph node sections and in a live tumor microenvironment (TME).
single cell RNA Sequencing data along with zipcodes that determine regional position

first: Matthew

middle: F

last: Krummel


title: Transcriptional Responses to beta-catenin and FoxO with and without oxidative stress

city: St. Louis

first: Leslie

middle: Stuart

last: Gewin

state: MO

address: 660 S Euclid Ave, McDonnell Science Building, Rm 718

department: Medicine (Nephrology)

institute: Washington University in St. Louis

summary: To identify novel transcriptional targets of beta-catenin and FoxO1 and FoxO3 in renal epithelial cells, we used conditionally immortalized murine proximal tubule (PT) cells (these cells, from Tgfbr2floxed mice, are described in detail in manuscript PMID: 23160515 in which Leslie Gewin is first author, JASN 2012). PT cells were either treated with Wnt3a (to activate beta-catenin) or the control diluting buffer, H2O2 to induce oxidative stress, and some were transfected with FoxO1, FoxO3, FoxO1 and 3, or scramble siRNA prior to Wnt and H2O2 treatment.

overall_design:
There were a total of 10 samples, replicates were not done. For publication, we analyzed sample 7 compared to 8 and 7 compared to 9. In other words, we compared the transcripts of PT cells with FoxO1 siRNA plus H2O2 and Wnt3a to PT cells with scramble siRNA and H2O2 and Wnt3a. Also, we compared transcripts of PT cells with FoxO3 siRNA plus H2O2 and Wnt3a to PT cells with scramble siRNA and H2O2 and Wnt3a. Separately, we also did 2 other analyses: 1) compare the genes upregulated by Wnt in regular conditions (PT media) versus oxidative stress (H2O2). This would be comparing the genes that change from sample 1 to 2 and comparing with the genes that change from sample 5 to 6. 2) We also compared the effect of knocking down FoxO1/3 in PT media with those in oxidative stress (comparing the change from sample 3 to 4 with that of sample 7 to 10).
Raw data could not be provided for this submission because the files had been previously deleted

first: Yan

last: Guo


title: Decoding mRNA translatability and stability from 5’ UTR

city: Hangzhou

first: Yuanhui

last: Mao

state: Zhejiang

address: 1369 West Wenyi Road

institute: Zhejiang University

summary: Precise control of protein synthesis by engineering sequence elements in 5’ untranslated region (5’UTR) remains a fundamental challenge. To accelerate our understanding of cis-regulatory code embedded in 5’UTR, we devised massively parallel reporter assays from a synthetic mRNA library composed of over one million 5’UTR variants. A completely randomized 10-nucleotide sequence preceding an upstream open reading frame (uORF) and downstream GFP leads to a broad range of mRNA translatability and stability in mammalian cells. While efficient translation protects mRNA from degradation, uORF translation triggers mRNA decay in a UPF1-dependent manner. We also identified translational inhibitory elements in 5’UTR with G-quadruplex as a mark for mRNA decay in the P-body. Unexpectedly, an unstructured A-rich element in 5’UTR, while enabling cap-independent translation, destabilizes mRNAs in the absence of translation. Our results not only expose diverse sequence features of 5’UTR in controlling mRNA translatability, but also reveal ribosome-dependent and -independent mRNA surveillance pathways.

overall_design: To accelerate our understanding of cis-regulatory code embedded in 5’UTR of mRNA, we devised a high-throughput uORF reporter assay from a synthetic mRNA library composed of over one million 5’UTR variants. This reporter assay is compatible with fluorescence-activated cell sorting and polysome profiling.

first: Longfei

last: Jia

first: Shu-Bing

last: Qian


title: A dynamic pattern of local auxin sources is required for root regeneration

city: Rehovot

first: Idan

last: Efroni

address: POB 12

department: Plant Sciences

country: Israel

institute: Hebrew University

summary: Following removal of its stem cell niche, the root meristem can regenerate by recruitment of remnant cells from the stump. Regeneration is initiated by rapid accumulation of auxin near the injury site but the source of this auxin is unknown. Here, we show that auxin accumulation arises from the activity of multiple auxin biosynthetic sources that are newly specified near the cut site and that their continuous activity is required for the regeneration process. Auxin synthesis is highly localized and PIN-mediate transport is dispensable for auxin accumulation and tip regeneration. Roots lacking the activity of the regeneration competence factor ERF115, or that are dissected at a zone of low-regeneration potential, fail to activate local auxin sources. Remarkably, restoring auxin supply is sufficient to confer regeneration capacity to these recalcitrant tissues. We suggest that regeneration competence relies on the ability to specify new local auxin sources in a precise spatio-temporal pattern.

overall_design: Untreated root meristems control + root meristems treated with 100uM L-Kyn for 3h and 6h. Experiment were done for both uncut and for meristems dissected at 120um above the QC

first: Rotem

last: Matosevich

first: Itay

last: Cohen

first: Naama

last: Gil-Yarom

first: Abelardo

last: Modrego

first: Carla

last: Verna

first: Enrico

last: Scarpella


title: Transcriptome analysis of brain endothelial cells from EC-specific Glut1 knock-out pups (P6) as compared to wild type controls

city: Schwerzenbach

first: Katrien

last: De Bock

state: Zürich

address: Schorenstrasse 16

department: D-HEST

country: Switzerland

institute: ETH Zurich

summary: To explore the mechanisms through which GLUT1 controls EC function, we compared transcriptomic alterations induced by loss of GLUT1 by performing RNA sequencing analysis on freshly isolated ECs from P6 GLUT1EC-/- brains. At P6, 447 genes were differentially expressed between WT and GLUT1-/- ECs. Subsequent gene set enrichment analysis showed that within the most differentially expressed pathways, several ones were related to p53, as well as inflammatory related pathways.

overall_design: Examination of transcriptomic differences in GLUT1EC-/-  vs WT controls

first: Ghobrial

last: Moheb

first: Veys

last: Koen


title: Detection of CNV in LMP1 mouse lymphomas

first: Thomas

last: Sommermann

type: Genome variation profiling by genome tiling array

summary: 12 LMP1 expressing mouse tumors were anayzed by array-CGH to detect chromosomal copy number variations

overall_design: 12 Rag2KOcgKO were reconstituted with  fetal liver hematopoietic stem and progenitor cells (HSPC) from Cd3eKO Cd19-Cre R26LMP1stopf mice. Developing LMP1-driven B-cell lymphomas were transplanted in secondary  Rag2KOcgKO or  Rag2KO mice. Secondary lymphomas were FACSpurified (CD19+huCD2+). gDNA of sorted lymphoma samples was analyzed by array-CGH sequencing.

first: Klaus

last: Rajewsky

first: Jonathan

last: Ronen


title: Low-Dose Silver Nanoparticle Surface Chemistry and Temporal Effects on Gene  Expression in HepG2 Cells

city: RTP

first: John

middle: S.

last: House

address: 111 TW Alexander Drive

department: Biostatistics & Computational Biology Branch

institute: National Instititute of Environmental Health Sciences

summary: We examined effects of surface-coating (BPEI, CIT, PEG) of silver nanoparticles and time (6h, 12h, 24h) on gene expression in human liver cancer cells (HepG2)

overall_design: This study was a two factor design. HepG2 cells were either given Media Only (NoNP) or treated with 1.2 ug/mL of either: CiT(citrate), bPEI(branched polyethylenimine) or PEG (polyethylene glycol). At each of 6h, 12h, and 24h time points post-treatment, treated cells and corresponding NoNP controls were collected in triplicate for targeted TempOseq whole genome sequencing

first: Evangelia

last: Bouzos

first: Kira

last: Fahy

first: Victorino

last: Francisco

first: Dillon

last: Lloyd

first: Fred

last: Wright

first: Alison

last: Motsinger-Reif

first: Prashanth

last: Asuri

first: Korin

middle: E

last: Wheeler


title: Transcriptome analysis of brain endothelial cells from EC-specific Glut1 knock-out mice

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

overall_design: Refer to individual Series


title: Post-translational regulation of the exon skipping machinery controls aberrant splicing in T cell leukemia [Jurkat_shSRSF6]

first: Yalu

last: Zhou

overall_design: Whole RNA was extracted from 1-5 million T-ALL (lines) cells or primary cells using the Total RNA Mini Kit (Bio-Rad) according to the manufacturer’s protocol. Poly-A+ (magnetic oligodT-containing beads (Illumina)) or Ribominus RNA was used for library preparation. cDNA preparation and unstranded library construction was performed using the TruSeq RNA Sample Preparation Kit. Libraries were sequenced on the NextSeq 500/550 using 76bp paired-end read method. Differential gene expression analysis was performed for primary T cells vs T-ALL patients samples, or each matched treatment vs control pairs, separately in each biological or technical replicate in each of two cell lines (CUTLL1, Jurkat). Seven types of comparisons were tested: (a) T-Cells vs T-ALL, (b) Non high risk T-ALL vs high risk T-ALL, (c) Control vs H3B-8800 treated Jurkat cells, (d) Control vs H3B-8800 treated CUTLL1 cells, (e) Control vs P5091 treated Jurkat cells, (f) Control vs P5091 treated CUTLL1 cells, (g) Control vs SRSF6 knockdown Jurkat cells. Analysis was performed using edgeR package.

first: Panagiotis

last: Ntziachristos


title: Post-translational regulation of the exon skipping machinery controls aberrant splicing in T cell leukemia [CUTLL1_P5091]


title: Transcriptome analysis of brain endothelial cells from adult EC-specific Glut1 knock-out mice as compared to wild type controls

first: Katrien

last: De Bock

summary: To explore the mechanisms through which GLUT1 controls EC function, we compared transcriptomic alterations induced by loss of GLUT1 by performing RNA sequencing analysis on freshly isolated ECs from adult (8 week old) GLUT1EC-/- brains. In adult brains ECs, 1494 genes were differentially expressed between WT and GLUT1-/- ECs but we did not observe compensatory upregulation of other GLUTs. Interestingly, pathway analysis revealed that ECs from GLUT1EC-/- mice were characterized by a robust activation of inflammatory and extracellular matrix pathways.


title: Post-translational regulation of the exon skipping machinery controls aberrant splicing in T cell leukemia [Jurkat_P5091]

overall_design: Whole RNA was extracted from 1-5 million T-ALL (lines) cells or primary cells using the Total RNA Mini Kit (Bio-Rad) according to the manufacturer’s protocol. Poly-A+ (magnetic oligodT-containing beads (Illumina)) or Ribominus RNA was used for library preparation. cDNA preparation and unstranded library construction was performed using the TruSeq RNA Sample Preparation Kit. Libraries were sequenced on the NextSeq 500/HiSeq 2500 using 76bp or 50bp paired-end read method. Differential gene expression analysis was performed for primary T cells vs T-ALL patients samples, or each matched treatment vs control pairs, separately in each biological or technical replicate in each of two cell lines (CUTLL1, Jurkat). Seven types of comparisons were tested: (a) T-Cells vs T-ALL, (b) Non high risk T-ALL vs high risk T-ALL, (c) Control vs H3B-8800 treated Jurkat cells, (d) Control vs H3B-8800 treated CUTLL1 cells, (e) Control vs P5091 treated Jurkat cells, (f) Control vs P5091 treated CUTLL1 cells, (g) Control vs SRSF6 knockdown Jurkat cells. Analysis was performed using edgeR package.


title: Post-translational regulation of the exon skipping machinery controls aberrant splicing in T cell leukemia [CUTLL1_H3B-8800]


title: Post-translational regulation of the exon skipping machinery controls aberrant splicing in T cell leukemia [Jurkat_H3B-8800 24h]


title: Systematic identification of factors bound to isolated metaphase ESC chromosomes reveals a role for chromatin repressors in compaction

city: London

first: Ya

last: Guo

address: Faculty of Medicine, Imperial College, Hammersmith Hospital Campus

department: Epigenetics section

country: United Kingdom

institute: MRC London Institute of Medical Sciences

summary: Epigenetic information is transmitted from mother to daughter cells through mitosis. To identify trans-acting factors and cis-acting elements that might be important for conveying epigenetic memory through cell division, we isolated native (unfixed) chromosomes from metaphase-arrested cells using flow cytometry and performed LC-MS/MS to determine the repertoire of chromosome-bound proteins. Quantitative proteomic comparisons between metaphase-arrested cell lysates and chromosome-sorted samples revealed a cohort of proteins that were significantly enriched on mitotic ESC chromosomes. These include pluripotency-associated transcription factors, repressive chromatin-modifiers (such as PRC2 and DNA methyl-transferases) and proteins governing chromosome architecture. We showed that deletion of PRC2, DNMT1/3a/3b or Mecp2 provoked an increase in the size of individual mitotic chromosomes consistent with de-condensation, as did experimental cleavage of cohesin complexes. These data provide a comprehensive inventory of chromosome-bound factors in pluripotent stem cells at mitosis and reveal an unexpected role for chromatin repressor complexes in preserving mitotic chromosome compaction.

overall_design: Hi-C on isolated chromosomes

first: Dounia

last: Djeghloul

first: Matthias

last: Merkenschlager

first: Amanda

middle: G

last: Fisher


title: Whole kidneys at embryonic day 14.5: wildtype (WT) versus Hoxb7Cre transgene mediated conditional Mdm4 knockout kidneys

city: New Orleans

first: Samir

last: El-Dahr

state: Louisiana

address: 1430 Tulane Avenue

department: Pediatric Nephrology

institute: Tulane University

summary: Transcriptional profiling of mouse embryonic kidneys lacking Mdm4 function in the ureteric bud lineage

overall_design: Agilent’s 'Two color Microarray-Based Gene Expression Analysis' (Agilent G4140-90050); 4 wild type replicates and 4 conditional mutant replicates

first: S

last: Hilliard


title: Post-translational regulation of the exon skipping machinery controls aberrant splicing in T cell leukemia [Patients I]

summary: In this study, we show that pediatric T-cell acute lymphoblastic leukemia (T-ALL) has an alternative mechanism for aberrant splicing that involves post-translational regulation of the splicing machinery via deubiquitination.


title: Post-translational regulation of the exon skipping machinery controls aberrant splicing in T cell leukemia


title: The deposition of CENH3 is stringently regulated in Maize

city: Wuhan

first: Handong

last: Su

state: Hubei

address: No.1,Shizishan Street · Hongshan District

institute: Huazhong Agricultural University

summary: The centromere, as an essential element to control chromosome segregation, is epigenetically determined by CENH3-containing nucleosomes as a functional marker, therefore the accurate deposition of CENH3 is crucial to chromosome transmission. We characterized the deposition of CENH3 in maize by over-expression and mutational analysis. Our results revealed that over-expressing CENH3 in callus is lethal while over-expressing GFP-CENH3 and CENH3-YFP in callus and plants is not and can be partly deposited normally. Different mutations of GFP-CENH3 demonstrated that CENH3-Thr4 in the N terminus was needed for the deposition as a positive phosphorylation site and the last five amino acids in the C terminus are necessary for deposition. The C terminal tail of CENH3 is confirmed to be responsible for the interaction of CENH3 and histone H4, which indicates that CENH3 maintains deposition in centromeres via interacting with H4 to form stable nucleosomes. For GFP-CENH3 and CENH3-YFP, the fused tags at the termini probably affect the structure of CENH3 and reduce its interaction with other proteins, which in turn could decrease proper deposition. Taken together, multiple amino acids or motifs were shown to play essential roles in CENH3 deposition, which is suggested to be affected by numerous factors in maize.

overall_design: ChIP-seq was carried out with anti-CENH3 antibodies using material from young leaves with different maize lines.

first: Fangpu

last: Han


title: Expression Data from Cerulein Treated Pancreas

city: Houston.

first: Jason

last: Mills

state: T

address: One Baylor Plaza

department: Gastroenterology, Medicine

institute: Baylor College of Medicine

summary:
IFRD1 is required for reserve stem cell proliferation in multiple tissues. Repeated cerulein treatment causes damage and induces proliferation in otherwise terminally differentiated pancreatic acinar cells.
We used microarrays to detail gene expression changes in IFRD1 null mice treated with cerulein compared to WT mice treated with cerulein.

overall_design: Adult mouse pancreatic tissue was harvested at 5 days post cerulein injection for RNA extraction and hybridization on Affymetrix microarrays. We aimed to isolate tissue during the proliferative phase of the pancreatic injury response.

first: Mark

last: Lewis


title: microRNA Profile in Stage I Clear Cell Renal Cell Carcinoma Predicts Progression to Metastatic Disease

city: Burlington

first: Travis

last: Sullivan

state: MA

address: 41 Mall Rd.

institute: Lahey Hospital & Medical Center


title: microRNA Profile in Stage I Clear Cell Renal Cell Carcinoma Predicts Progression to Metastatic Disease [screen]

summary:
To identify microRNA (miRNA) profiles of small, pathologically confirmed stage 1 clear cell renal cell carcinoma (ccRCC) tumors that could predict progression to metachronous metastatic disease.  : Fifty-five pathologic stage 1 ccRCC tumors ≤5cm from two institutions were examined in an initial miRNA screening, followed by a validation study. This investigation identified miRNA biomarkers that may differentiate between non-progressive ccRCC tumors and those that progress to metastatic disease in this group of stage I tumors.
The data presented in this workbook are for the 20 samples analyzed in the screening analysis.  The data were normalized to the global mean of all targets with a Cq<35.  Prior to normalization each value was scaled by the corresponding internal plate calibrator value.

overall_design: FFPE ccRCC specimens from 20 patients were analyzed for microRNA expression. The 20 candidate microRNA identified in the screening analysis were tested on the remaining 35 samples in an effort to validate microRNA indicative of progressive disease.


title: Post-translational regulation of the exon skipping machinery controls aberrant splicing in T cell leukemia [Jurkat_H3B-8800 6hr]


title: Post-translational regulation of the exon skipping machinery controls aberrant splicing in T cell leukemia [Patients II]


title: microRNA Profile in Stage I Clear Cell Renal Cell Carcinoma Predicts Progression to Metastatic Disease [validation]

summary:
To identify microRNA (miRNA) profiles of small, pathologically confirmed stage 1 clear cell renal cell carcinoma (ccRCC) tumors that could predict progression to metachronous metastatic disease.  : Fifty-five pathologic stage 1 ccRCC tumors ≤5cm from two institutions were examined in an initial miRNA screening, followed by a validation study. This investigation identified miRNA biomarkers that may differentiate between non-progressive ccRCC tumors and those that progress to metastatic disease in this group of stage I tumors.
The data presented in this workbook are for the samples analyzed in the custom array analysis.  The data were normalized to the average of the insignificant control miRNA: miRs-28-5p, -125-5p, and -191-5p, selected from the screening array dataset using NormFinder©.  Prior to normalization each value was scaled by the corresponding internal plate calibrator value (see below).

overall_design: FFPE ccRCC specimens analyzed for microRNA expression. The 20 candidate microRNA identified in the screening analysis were tested on 35 additional samples in an effort to validate microRNA indicative of progressive disease.  In addition, 18 of the 20 samples from the screening analysis were re-tested for the 20 miRNA with the custom array and used along with the 35 validation samples in survival analyses.  Two of the screening samples were excluded from the custom array due to a lack of remaning RNA needed to perform the re-testing.


title: Genome-wide R-loop landscapes during epigenetic programming and reprogramming

status: Public on Jun 17 2020

first: Zunpeng

last: Liu

address: Beichen West Road

department: Institute of Zoology, Chinese Academy of Sciences

institute: State Key Laboratory of Stem Cell and Reproductive Biology

type: Methylation profiling by high throughput sequencing

summary: The role of R-loops, three-stranded nucleic acid structures harboring DNA:RNA hybrids, in regulating cell fate decisions remains elusive. Using an isogenic human stem cell platform, we systematically characterized R-loops, DNA methylation, histone modifications, and chromatin accessibility in pluripotent cells and their lineage-differentiated derivatives. We found that R-loops initially formed co-transcriptionally at pluripotency genes in pluripotent cells. Upon differentiation into neural, mesenchymal, endothelial, and smooth muscle lineages, R-loops and repressive chromatin marks increased synergistically on pluripotency genes and undesired lineage genes, while new R-loops formed co-transcriptionally to activate lineage determination genes. In reprogrammed pluripotent cells, features of epigenetic memory and aberrant gene expression were initially present in regions highly enriched with R-loops, but became resolved with serial passaging. Our analysis defines a multi-faceted role of R-loops in cell fate determination and adds an additional layer of epigenetic control of cell state changes and cell fate memory.

overall_design: Multi-omics sequencing of human neural stem cells (hNSCs), mesenchymal stem cells (hMSCs), vascular endothelial cells (hVECs), as well as vascular smooth muscle cells (hVSMCs), with equal efficiency from human embryonic stem cells (hESCs) or induced pluripotent stem cells (hiPSCs).

first: Guanghui


title: Identification of subcellular gene expression changes due to a Nonsense-Mediated Decay pathway at the Endoplasmic Reticulum

status: Public on Jun 15 2020

city: EDINBURGH

first: Kathryn

middle: Alexandra

last: Jackson-Jones

address: Institute of Genetics and Molecular Medicine, Crewe Road South

department: MRC Human Genetics Unit

institute: University of Edinburgh

summary: Nonsense-mediated decay (NMD) is a translation-dependent RNA quality control mechanism that occurs in the cytoplasm. However, it is unknown how NMD regulates the stability of RNAs translated at the Endoplasmic Reticulum (ER). Here, we identify a localized NMD pathway dedicated to ER-translated mRNAs. We previously identified NBAS, a component of the Syntaxin 18 complex involved in Golgi-to-ER trafficking, as a novel NMD factor. Here, we show that NBAS fulfils an independent function in NMD. This ER-NMD pathway requires the interaction of NBAS with the core NMD factor UPF1, which is partially localized at the ER in the proximity of the translocon. NBAS and UPF1 co-regulate the stability of ER-associated transcripts, in particular those associated with the cellular stress response. We propose a model where NBAS recruits UPF1 to the membrane of the ER and activates an ER-dedicated NMD pathway, thus providing an ER protective function by ensuring quality control of ER-translated mRNAs.

overall_design: Profiling of cytoplasmic and membrane fractions of HeLa cells when NMD factors NBAS, UPF1 or UPF2 depleted, in triplicate, to elucidate NMD targets of each specific factor in thse subcellular locations

first: Magdalena

last: Maslon

first: Robert

last: Young

first: Martin

last: Taylor

first: Dasa

last: Longman

first: Laura

last: Murphy

first: Jack

last: Stoddart

first: Dimitrios

middle: K

last: Papadopoulos

first: Javier

last: Caceres


title: High fat effect on mouse intestinal stem cells

status: Public on May 31 2020

city: Bronx

first: Wenge

last: Li

state: NY

address: 1300 Morris Park Avenue

department: Medicine

institute: Albert Einstein College of Medicine

summary: Examination of transcriptomic profiles of intestinal stem cells fed with different diets

overall_design: 12 samples; 4 diets; triplicates

first: Leonard

last: Augenlicht


title: Endothelial Gene Expression Associated With Early Coronary Atherosclerosis

first: Peng

last: Wei

state: TX

department: Biostatistics

institute: The University of Texas MD Anderson Cancer Center

summary: We examined feasibility of a unique approach towards gaining insight into heritable risk for early atherosclerosis: surveying gene expression by endothelial cells from living subjects. Subjects <50 years old (mean 37, range 22-49) without obstructive coronary artery disease underwent coronary reactivity testing that identified them as having normal (NL) or abnormal (ABNL) coronary endothelial function. Cultures of Blood Outgrowth Endothelial Cells (BOEC) from 6 NLs and 13 ABNLs passed rigorous quality control and were used for microarray assessment of gene expression. Of nine genes differentially expressed at FDR<0.1%, we here focus upon ABNLs having elevated expression of HMGB1 which we unexpectedly found to be linked to low LAMC1 expression. This was corroborated by three of our past studies and confirmed bio-functionally.  Compared to NL BOEC, ABNL BOEC released 13±3 fold more HMGB1 in response to LPS; and they deposited one-tenth as much LAMC1 into collagen subendothelial matrix during culture. Clinical follow-up data are provided for 4 NLs (followed 13.4±0.1 yr) and for 12 ABNLs (followed 9.1±4.5 yr). The known pathogenic effects of high-HMGB1 and low-LAMC1 predict that the combination would biologically converge upon the focal adhesion complex, to the detriment of endothelial shear responsiveness. This gene expression pattern may comprise a heritable risk state that promotes early coronary atherosclerosis. If so, the testing could be applied even in childhood, enabling early intervention. This approach offers a way to bridge the information gap between genetics and clinical phenotype.

overall_design: 6 subjects with normal coronary endothelial function, and 13 subjects with abnormal coronary endothelial function.  All were adults <45 years old.

middle: P

last: Hebbel

first: Amir

last: Lerman


title: Ribosome profiling of eIF3e-KD MCF-10a cells

first: Xuerui

last: Yang

address: Haidian District

department: School of Life Sciences

institute: Tsinghua University

summary: eIF3 is a multi-subunit complex thought to execute numerous functions in canonical translation initiation, including mRNA recruitment to the 40S ribosome, scanning for the start codon, and inhibition of 60S subunit joining 1–3. eIF3 was also found to interact with 40S and 60S ribosomal proteins and translation elongation factors 4, but a direct involvement in translation elongation has never been demonstrated. Using selective ribosome profiling, we made the unexpected observation that eIF3 remains bound to post-initiation 80S ribosomes, followed by release after translation of ~50 codons. Furthermore, eIF3 deficiency reduces early ribosomal elongation speed, particularly on mRNAs encoding proteins associated with membrane-associated functions, resulting in defective synthesis of their encoded proteins and abnormal mitochondrial and lysosomal physiology.  Accordingly, heterozygous eIF3e+/- knockout mice accumulate giant mitochondria in skeletal muscle and show a progressive decline in muscle strength with age. Hence, in addition to its canonical role in translation initiation, eIF3 interacts with 80S ribosomes to enhance, at the level of early elongation, the synthesis of proteins with membrane-associated functions, an activity that is critical for normal muscle health.

overall_design: Using si-RNA to do eIF3e-KD then do the ribosome profiling samples with total RNA, 3 replicates.

first: Yingying

last: Lin

first: Fajin

first: Jianhuo

last: Fang

first: Xudong

last: Xing


title: RNAseq Analysis of the effects of Autophagy depletion  using ATG3KO, GABATKO and HexaKO on differential gene expression

city: ALBUQUERQUE

first: Suresh

last: Kumar

state: NM

address: 3901 Indian School Rd NE, Apt D407

department: MGM

institute: University of new Mexico l

summary: We report that knock out of all mAtg8s (HexaKO) reduced the differential gene expression 451 and upregulated the expression of 294 genes, in addition of several previous known TFEB targets

overall_design: We comapred the differential gene expression in in HeLa wild type and HeLa cells depleted for ATG3, GABARAPs ( GABARAP GABARAPL1 and GABARAPL2) and all six mAtg8s (LC3A, LC3B,LC3C, GABARAP, GABARAPL1, GABARAPL2)

first: Vojo

last: Deretic


title: EZH2 deficient T-cell acute lymphoblastic leukemia is sensitized to CHK1 inhibition through enhanced replication stress

first: Stephen

last: Henderson

address: Paul O'Gorman Building

department: Cancer Institute

institute: UCL

summary: Loss-of-function mutations of EZH2, the enzymatic component of PRC2, have been associated with poor outcome and chemotherapy resistance in T-cell acute lymphoblastic leukemia (T-ALL). Using isogenic T-ALL cells, with and without CRISPR/Cas9-induced EZH2-inactivating mutations, we performed a cell-based synthetic lethal drug screen. EZH2 deficient cells exhibited increased sensitivity to structurally diverse inhibitors of CHK1, an interaction that could be validated genetically. Furthermore, small molecule inhibition of CHK1 had efficacy in delaying tumor progression in isogenic EZH2 deficient, but not EZH2 wild-type T-ALL cells in vivo, as well as in a primary cell model of PRC2 mutant ALL. Mechanistically, EZH2 deficiency resulted in a gene expression signature of immature T-ALL cells, marked transcriptional upregulation of MYCN, increased replication stress, and enhanced dependency on CHK1 for cell survival. Lastly, we demonstrate this phenotype is mediated through de-repression of a distal PRC2-regulated MYCN enhancer. In conclusion, we highlight a novel and clinically exploitable pathway in high-risk EZH2 mutated T-ALL.

overall_design: Jurkat EZH2-WT versus Jurkat CRISPR/Cas9- edited -EZH2-KO1 cell lines were analysed for MYB ChIP-seq

first: Theresa

last: León

first: Tanya

last: Rapoz-D’Silva

first: Cosetta

last: Bertoli

first: Sunniyat

last: Rahman

first: Michael

last: Magnussen

first: Brian

last: Philip

first: Nadine

last: Farah

first: Simon

last: Richardson

first: Sara

last: Ahrabi

first: Jose

last: Guerra-Assunção

first: Rajeev

last: Gupta

first: Elisabeth

last: Nacheva

last: Herrero

first: David

last: Linch

first: Robertus

last: de Bruin

first: Marc

middle: R

last: Mansour


title: Medium-throughput drug screening of patient-derived organoids from colorectal peritoneal metastases can be used to guide therapy

city: Adelaide

first: Susan

last: Woods

state: Select State

address: Level 5 South, SAHMRI building, North Terrace

department: School of Medicine

country: Australia

institute: The University of Adelaide

summary:
Purpose: Colorectal cancer (CRC) patients with peritoneal metastases (CRPM) have limited treatment options and the lowest CRC survival rates. We aimed to determine whether organoid testing could help guide precision treatment for CRPM patients, as the clinical utility of prospective, functional drug screening including non-standard agents is unknown.
Experimental Design: CRPM organoids (peritonoids) isolated from patients underwent parallel next-generation sequencing and medium-throughput drug panel testing ex vivo to identify specific drug sensitivities for each patient. We measured the utility of such a service including: success of peritonoid generation, time to cultivate peritonoids, reproducibility of the medium-throughput drug testing, and documented changes to clinical therapy as a result of the testing.
Results: Peritonoids were successfully generated and validated from 68% (19/28) of patients undergoing standard care. Genomic and drug profiling was completed within 8 weeks and a formal report ranking drug sensitivities was provided to the medical oncology team upon failure of standard care treatment. This resulted in a treatment change for 2 patients, one of whom had a partial response despite previously progressing on multiple rounds of standard care chemotherapy. The barrier to implementing this technology in Australia is the need for drug access and funding for off-label indications.
Conclusions: Our approach is feasible, reproducible and can guide novel therapeutic choices in this poor prognosis cohort, where new treatment options are urgently needed. This platform is relevant to many solid organ malignancies.

overall_design: Fourteen patient derived metastatic CRC peritoneal lesion were sequenced. In addition to this, 3 paired primary CRC resected lesions were sequenced as controls. All samples were subjected to CMS analyses using CMS caller package.

first: Vignesh

last: Narasimhan

first: Josephine

last: Wright

last: Churchill

first: Tongtong

last: Wang

first: Rachele

last: Rosati

first: Tamsin

last: Lannagan

first: Laura

last: Vrbanac

first: Hiroki

last: Kobayashi

first: Annie

first: Timothy

last: Price

first: Gayle

middle: X

last: Tye

first: Julie

last: Marker

first: Peter

last: Hewett

last: Flood

first: Shalini

last: Pereira

first: G

last: Whitney

last: Michael

first: Jeanne

last: Tie

first: Siddhartha

last: Mukherjee

first: Carla

last: Grandori

first: Alexander

last: Heriot

first: Daniel

last: Worthley

first: Robert

last: Ramsay


title: Reduced Type I Interferon response signature detected in dendritic cells from prediabetic NOD mice compared to C57BL/6.g7 and Rag KO [stimulated samples]

city: Bethesda

first: Kristin

middle: V

last: Tarbell

state: MD

address: 10 Center Dr, Bldg 10 5-5832

department: Diabetes, Endocrinology, and Obesity Branch

institute: NIH/NIDDK

summary: Investigating type I interferon in prediabetic stage of DCs from NOD mice compared to C57BL/6.g7 and Rag KO mice.

overall_design: RNAseq was used to study splenic cDC1, cDC2, pDC, moDC, and monocyte populations from 8-10 week old NOD and C57BL/6.g7 and Rag KO mice.


title: Yap1-Scribble polarization is required for hematopoietic stem cell division and fate

status: Public on May 30 2020

city: Cincinnati

first: Mark

middle: Jordan

last: Althoff

state: OH

address: 3333 Burnett Avenue

department: Experimental Hematology and Cancer Biology

institute: Cincinnati Children's Hospital Medical Center

summary: Yap1 and its paralogue Taz largely control epithelial tissue growth. We have identified that hematopoietic stem cell (HSC) fitness response to stress depends on Yap1 and Taz. Deletion of Yap1 and Taz induces a loss of HSC quiescence, symmetric self-renewal ability and renders HSC more vulnerable to serial myeloablative 5-fluorouracil treatment. This effect depends on the predominant cytosolic polarization of Yap1 through its PDZ domain-mediated interaction with the scaffold Scribble. Scribble and Yap1 coordinate to control cytoplasmic Cdc42 activity and HSC fate determination in vivo. Deletion of Scribble disrupts Yap1 co-polarization with Cdc42 and decreases Cdc42 activity, resulting in increased selfrenewing HSC with competitive reconstitution advantages. These data suggest that Scribble/Yap1 co-polarization is indispensable for Cdc42-  dependent activity on HSC asymmetric division and fate. The combined loss of Scribble, Yap1 and Taz results in transcriptional upregulation of Rac-specific guanine nucleotide exchange factors, Rac activation and HSC fitness restoration. Scribble links Cdc42 and the cytosolic functions of the Hippo signaling cascade in HSC fate determination.

overall_design: Four independent biological samples were analyzed (Wild type, Scribble-/-, Taz-/-;Yap1-/- and Scribble-/-;Taz-/-;Yap1-/- hematopoietic stem cells) with experimental replicates. The Wild type sample was used as control/ reference sample

middle: J

first: Ramesh

last: Nayak

first: Shailaja

last: Hegde

first: Ashley

last: Wellendorf

first: Breanna

last: Bohan

first: Marie-Dominique

last: Filippi

first: Mei

last: Xin

first: Richard

last: Lu

first: Hartmut

last: Geiger

first: Yi

last: Zheng

first: Maria

last: Diaz-Meco

first: Jorge

last: Moscat

first: Jose

last: Cancelas


title: RNAseq of 2'3' cGAMP injected Drosophila melanogaster

city: Strasbourg

first: Nelson

middle: Eduardo

last: Martins

address: 2 Allee Konrad Roentgen

country: France

institute: CNRS UPR9022/IBMC

summary: We recently reported that an orthologue of STING regulates infection by picorna-like viruses in drosophila. Here, we show that injection of flies with 2’3’-cGAMP can induce expression of dSTING-regulated genes. Analysis of the transcriptome of 2’3’-cGAMP injected flies reveals a complex pattern of response, with early and late induced genes. Our results reveal that dSTING regulates an NF-κB -dependent antiviral program, which predates the emergence of Interferon Regulatory Factors and interferons in vertebrates.

overall_design: RNAseq of Tris, ci-di-GMP and 2'3' cGAMP injected Drosophila melanogaster

first: Hua

last: Cai

first: Andreas

last: Holleufer

first: Bine

last: Simonsen

first: Juliette

last: Schneider

first: Aurélie

last: Lemoine

first: Hans-Henrik

last: Gad

first: Jingxian

last: Huang

first: Hieqing

first: Di

last: Chen

first: Tao

last: Peng

first: João

last: Marques

first: Rune

last: Hartmann

first: Jean-Luc

last: Imler


title: PRP8A and PRP8B spliceosome subunits act coordinately to  control pollen tube attraction in Arabidopsis thaliana

city: Prague 6

first: Mila

last: Juricek

address: Rozvojova 313

country: Czech Republic

institute: Inst. Exp. Botany, Czech Acad Sci

summary:
Purpose: Alternative splicing is fundamental for post-transcriptional regulation and proteome diversity. The goals of this study are to compare transcriptome and splicing profiling (RNA-seq) between wild type and prp8a prp8b mutant ovules of the spliceosome subunit and define the molecular signature of prp8a prp8b pollen tube attraction phenotype.
Methods: mRNA profiles from mature ovules of 6-weeks-old wild-type (WT) and pre-mRNA processing factor 8 (PRP8Aa prp8bb) Arabidopsis plants were generated by deep sequencing, in triplicates, using Illumina HiSeq4000 100bp paired-end reads. The sequence reads that passed quality filters were were mapped to TAIR10 whole genome and analyzed for differential expression, differential exone usage and intron retention as indicated in DATA PROCESSING PIPELINE section.
Results: Using an optimized data analysis workflow, about 15 million sequence read pairs per sample were mapped to the Arabidopsis genome (TAIR10). Approximately 2.9% of the transcripts showed differential expression between the WT and PRP8Aa prp8bb ovules, with a fold change ≥1.5 and p value <0.05. Analysis for differential gene expression, exon usage and intron retention with DESeq2 v1.22.1 and DEXseq v1.28.0 or IRFinder v1.2.3 respectively, uncovered several as yet uncharacterized genes that may contribute to pollen tube attraction and female gametophyte cell fate specification.
Conclusions: Our work has uncovered a molecular signature through which PRP8A/PRP8B subunits act redundantly to define male-female signaling competence for successful pollen tube attraction in Arabidopsis. Application of DESeq2 algorithms to our ovule RNA-seq data identified downregulation of over 50 different CRP genes with yet unknown function from the synergid and the central cells including all LURE pollen tube attractants. Whereas use of DEXseq workflow, revealed mis-splicing of key genes involved embryo sac specificiation and genes of the secretory pathway. We concluded that 100bp paired-end RNAseq was a sufficient compromise for detection of differential gene expression and splice isoforms, however, our experiment would have benefited with more number of replicates.

overall_design: Mature ovules mRNA profiles of 6-weeks-old wild-type (WT) and pre-mRNA processing factor 8 (PRP8Aa prp8bb) Arabidopsis plants

first: Said

last: Hafidh

first: Katarina

last: Kulikova


title: Gene expression analysis of treatment-naïve hormone receptor-positive metastistic breast cancer

city: San Diego

first: Zhou

last: Zhu

state: CA

address: 10646 Science Center Drive (CB4)

institute: Pfizer Worldwide Research & Development

summary:
A large-panel gene expression analysis was conducted to identify biomarkers associated with the effectiveness of adding palbociclib to letrozole
Cyclin-dependent kinase 4/6 (CDK4/6) inhibitors have demonstrated significant activity in estrogen receptor-positive/human epidermal growth factor receptor 2-negative (ER+/HER2-) advanced breast cancer. Further biomarkers to refine the population beyond ER+/HER2- that benefits remain to be identified. We performed a comprehensive biomarker analysis using patient tissues from PALOMA-2, a randomized phase 3 study of palbociclib-letrozole versus placebo-letrozole, to determine markers of sensitivity and resistance to the combination.

overall_design: Baseline tumor tissue from PALOMA-2 trial were profiled using EdgeSeq Oncology BM Panel, with 455 tumor samples evaluable for analysis

first: Liu

last: Yuan

first: Zhu

last: Zhou


title: Gene expression profile of Arabidopsis thaliana wild type and siz1-2 mutant during in vitro shoot regeneration on callus inducing media (CIM) and shoot inducing media (SIM)

city: Yokohama

first: Duncan

middle: Peter

last: Coleman

state: Kanagawa

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

department: CSRS

institute: Riken

summary: Plants form callus and regenerate new organs when incubated on phytohormone-containing media. While accumulating evidence suggests that these regenerative processes are governed by transcriptional networks orchestrating stress responses and developmental transitions, it remains unknown if post-translational regulatory mechanisms are involved in this process. Here, we find that SIZ1, which encodes an E3 ligase catalyzing attachment of the SMALL UBIQUITIN-LIKE MODIFIER (SUMO) to proteins, regulates wound-induced signal transduction and organ regeneration. We show that loss-of-function mutants for SIZ1 exhibit over-production of shoot meristems under in vitro tissue culture conditions, while this defect is rescued in a complementation line expressing pSIZ1::SIZ1. RNA-sequencing analysis revealed that siz1-2 mutant exhibits enhanced transcriptional responses to wound stress, resulting in the hyper-induction of over 500 genes immediately after wounding. Among them, we show that elevated level of WOUND INDUCED DEDIFFERENTIATION 1 (WIND1) and WIND2 contribute to enhanced shoot regeneration observed in siz1 mutants, as the dominant-negative WIND1-SRDX partly rescues this phenotype in siz1-3. Although compromised SIZ1 function does not modify transcription of genes implicated in auxin-induced callus formation and/or pluripotency acquisition, it does lead to enhanced induction of cytokinin-induced shoot meristem regulators like WUSCHEL (WUS), promoting the formation of WUS-expressing foci in explants. This study thus suggests that SIZ1 negatively regulates shoot regeneration in part by repressing wound-induced cellular reprogramming.

overall_design: Hypocotyls from 7 day old dark grown seedlings were excised  and sampled immediately after cutting, after incubation on CIM for 4 days and after incubation on CIM 4 days plus SIM for 4 and 6 days.

first: Keiko

last: Sugimoto


title: The Arabidopsis epigenetic regulator ICU11 as an accessory protein of Polycomb Repressive Complex 2

city: Norwich

first: Xiaoqi

last: Feng

state: Norfolk

address: Norwich Research Park

department: Cell and Developmental Biology

institute: John Innes Centre

summary: Molecular mechanisms enabling the switching and maintenance of epigenetic states are not fully understood. Distinct histone modifications are often associated with ON/OFF epigenetic states, but how these states are stably maintained through DNA replication, yet in certain situations switch from one to another remains unclear. Here, we address this problem through identification of Arabidopsis INCURVATA11 (ICU11) as a Polycomb Repressive Complex 2 accessory protein. ICU11 robustly immunoprecipitated in vivo with PRC2 core components and the accessory proteins, EMBRYONIC FLOWER 1 (EMF1), LIKE HETEROCHROMATIN PROTEIN1 (LHP1) and TELOMERE_REPEAT_BINDING FACTORS (TRBs). ICU11 encodes a 2-oxoglutarate dioxygenase, an activity associated with histone demethylation in other organisms, and mutant plants show defects in multiple aspects of the Arabidopsis epigenome. To investigate its primary molecular function we identified the Arabidopsis FLOWERING LOCUS C (FLC) as a direct target and found icu11 disrupted the cold-induced, Polycomb-mediated silencing underlying vernalization. icu11 prevented reduction in H3K36me3 levels normally seen during the early cold phase, supporting a role for ICU11 in H3K36me3 demethylation. This was coincident with an attenuation of H3K27me3 at the internal nucleation site in FLC, and reduction in H3K27me3 levels across the body of the gene after plants were returned to the warm. Thus, ICU11 is required for the cold-induced epigenetic switching between the mutually exclusive chromatin states at FLC, from the active H3K36me3 state to the silenced H3K27me3 state. These data support the importance of physical coupling of histone modification activities to promote epigenetic switching between opposing chromatin states.

overall_design: Examination of DNA methylation of WT and icu11-3 seedlings

first: Rebecca

last: Bloomer

first: James

last: Walker

first: Caroline

last: Dean


title: Skeletal muscle DNA methylation profiles in young healthy men before and after acute and chronic HIIT

status: Public on May 29 2020

city: Footscray

first: Sarah

middle: Amandine Caroline

last: Voisin

state: VIC

address: Ballarat Road

department: Institute for Health and Sport

institute: Victoria University

overall_design:
This dataset contains 78 vastus lateralis muscle samples from healthy men who underwent 1) an acute bout of HIIT and 2) four weeks of HIIT. From the 78 samples, 25 were taken at baseline, 25 after one bout of high-intensity interval exercise, and 25 after four weeks of exercise training (25 individuals at 3 time points = 75). The remaining 3 samples were replicates.

Please note that the processed and raw data files have been updated on Aug 10, 2020.

first: Nir

last: Eynon


title: MiRNA analysis in Thoracic Aortic aneurysms

city: Nicosia

first: Areti

last: Moushi

address: Iroon Avenue 6, Agios Dometios

country: Cyprus

institute: Cyprus Insitute of Neurology and Genetics

summary: Thoracic Aortic Aneurysm (TAA) is characterized by the dilation and degradation of the aorta and is fatal if not diagnosed and treated appropriately. There are no specific clinical symptoms, so better knowledge of the physiopathology of TAAs and their underlying genetic mechanisms is needed to improve diagnosis and therapy. MiRNAs regulate gene expression post-transcriptionally and are known to be involved in cerebrovascular disease. The current study aimed to identify differentially expressed miRNAs in patients with TAAs and determine whether their predicted target genes could be associated with this condition.   Nanostring assays identified miRNAs in plasma and tissue samples from four TAA patients. RT-PCR validated the expression levels of these miRNAs in a further 22 plasma samples. Three, hsa-miR140-5p, hsa-miR-191-5p and hsa-miR-214-3p showed significant expression level differences between plasma samples collected pre- and post-surgically from each patient. Analyses of the predicted target gene controlled by these miRNAs revealed nine genes whose expression was investigated in the same 22 plasma samples. The gene expression levels were inversely correlated with the expression of their respective miRNAs. From these, CCND2, CRKL, HEY1, MTMR4, NFIA and PPP1CB, showed fold-change differences >1.5 between the two plasma samples. An in-depth literature search and Cytoscape software three genes; MTMR4, NFIA and PPP1CB, showed a possible association with the TGF-β signalling pathway. It is suggested that the three miRNAs detected together with their target genes could play a role in the TGF-β signalling pathway and thus be involved in TAA pathogenesis.

overall_design: Nanostring assays of miRNAs in plasma and tissue samples from four TAA patients.


title: RNAseq anaysis of entorhinal cortex layer II neurons upon Ptbp1 modulation

city: Boston

first: Jean-Pierre

last: Roussarie

address: 72 E Concord Street

department: Anatomy & Neurobiology

institute: Boston University School of Medicine

summary: Purpose: To identify gene expression changes in entorhinal cortex layer II (ECII) neurons upon Ptbp1 modulation (silencing and overexpression)  Method: bacterial artificial chromosome - Translating Ribosome Affinity Purification (bacTRAP) to isolate actively translated mRNA in ECII neurons, 2 weeks after stereotaxic injection of an AAV1 vector in the EC of ECII-bacTRAP mice; followed by RNAseq. Note: Ptbp1 was significantly overexpressed in the overexpression experiment, but no silencing was achieved with the silencing vector, probably because of tight control of Ptbp1 expression

overall_design: in IP162 (silencing), 5 mice injected with a scrambled-mcherry vector, 5 with a Ptbp1 shRNA-mcherry vector; 1 Ptbp1 shRNA mouse was excluded because no read mapped to mcherry; in IP170 (overexpression), 5 mice injected with control-mcherry vector, 5 mice with a Ptbp1-mcherry vector; 1 control mouse was excluded because no read mapped to mcherry; in both experiments, the two entorhinal cortices were pooled


title: Integrative molecular profiling of autoreactive CD4 T cells in autoimmune hepatitis

city: Marseille

first: Pierre

last: Milpied

address: Parc Scientifique de Luminy, case 906

institute: Centre d'Immunologie de Marseille-Luminy (CIML)

summary: Background & Aims: In most autoimmune disorders, crosstalk of B cells and CD4 T cells results in the accumulation of autoantibodies targeting specific self-antigen like the Soluble Liver Antigen (SLA or SepSecs) in autoimmune hepatitis (AIH). But the associated autoreactive CD4 T cells have not been characterized yet.  Here we isolated and deeply characterized SLA-specific CD4 T cells in AIH patients.  Methods: We used brief ex vivo re-stimulation with overlapping SLA-derived peptides to isolate and phenotype circulating SLA-specific CD4 T cells, and integrative single-cell RNA-seq (scRNA-seq) to characterize their transcriptome and T cell receptor (TCR) repertoire. Identified auto-reactive TCRs were reconstituted cloned and expressed in T cell hybridoma to identify dominant SLA-derived CD4 T cell epitopes. SLA-specific CD4 T cells were tracked in peripheral blood through TCR sequencing, to identify their phenotypic niche. We further characterized disease-associated peripheral blood T cells by high content flow cytometry in an additional cohort of n=42 AIH patients and n=17 non-alcoholic steatohepatitis (NASH) controls.  Results: Frequency of autoreactive SLA-specific CD4 T cells was associated with anti-SLA autoantibodies and had a memory CD45RA-CD27+PD-1+CXCR5-CCR6- phenotype.  ScRNA-seq revealed their pro-inflammatory/B-Helper profile (IL21, IFNG, TIGIT, CTLA4, NR3C1, CD109, KLRB1 and CLEC2D). SLA81-100 and SLA177-204 contain dominant T cell epitopes. Autoreactive TCR clonotypes were restricted to the memory PD-1+CXCR5- CD4 T cells which was significantly increased in the blood of AIH patients and supported B cell differentiation through IL-21. Finally, we identified a specific phenotype (CD45RA-PD-1+CD38+CXCR5-CD127-CD27+) of T cells linked to disease activity and IgG response during AIH.   Conclusions: This work provides for the first time a deep characterization of rare circulating autoreactive CD4 T cells and the identification of their peripheral reservoir in AIH. We also propose a generic phenotype of pathogenic T cells related to AIH disease activity which will be essential to track, delineate and potentially target those pathogenic cells.

overall_design: This dataset contains data from SLA-specific and C.alb-specific CD4 T cells sorted from human peripheral blood of SLA-positive AIH patients after in vitro peptide restimulation. Single-cells were FACS-sorted into several 96-well plates and analyzed by 5'-end single-cell RNAseq following the FB5P-seq protocol. The samples were processed and sequenced in three separate experiments (experiment 1: samples 028-03, 128-01, 018-10; experiment 2: samples 018-13, 004-05, 018-04; experiment 3: sample 051-04).

first: Laurine

last: Gil

first: Chuang

last: Dong

first: Amédée

last: Renand


title: Rational and scalable strategies to identify small “molecular glue” degraders

city: Vienna

first: Hana

last: Imrichova

address: Lazarettgasse 14, AKH BT 25.3

country: Austria

institute: CeMM, Vienna

summary: In our effort to characterize putatite novel molecular glues that we found in a phenotypic  screen, we wanted to gain a more detailed understanding  of the effects of these small-molecules at the trascriptome level.

overall_design: KBM7 cells treated for 5h with DMSO, dCeMM-1[Hit4], dCeMM-2[Hit1], dCeMM-3[Hit2], dCeMM-4[Hit8] (10xEC50) and THZ531 (600nM). Triplicates per condition.

first: Cristina

last: Mayor-Ruiz

first: Sophie

last: Bauer

first: Georg

last: Winter


title: Functional interplay of Epstein-Barr Virus oncoproteins in a mouse model of B cell lymphomagenesis

city: Berlin

first: Thomas

last: Sommermann

address: Robert-Rossle-Str. 10

institute: Max Delbruck Center for Molecular Medicine

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

overall_design: Refer to individual Series


title: Differential gene expression between adipsoe depots in cancer cachexia

city: Stirling

first: Iain

last: Gallagher

address: Room 4B133 Cottrell Building, University of Stirling, Airthrey Road

department: Faculty of Health Sciences and Sport

institute: University of Stirling

summary:
Cancer cachexia (CC) is a poorly understood cause of morbidity and mortality, which has no efficacious treatment or generally-accepted management strategy. The consensus definition for CC identified skeletal muscle loss as a key marker in the diagnosis and classification of cachexia. The importance of fat wasting however, is less understood. During cachexia, different adipose depots demonstrate differential wasting rates. Studies from animal models have suggested adipose tissue may be a key driver of muscle wasting through fat-muscle crosstalk but human studies in this area are lacking. We performed global gene expression profiling of intra-abdominal (IAAT) and subcutaneous (SAT) adipose from weight stable and cachectic cancer patients and healthy controls.
Cachexia was defined as >2% weight loss plus low CT-muscularity. Biopsies of SAT and IAAT were taken from patients undergoing resection for oesophago-gastric cancer, and healthy controls (donor nephrectomy) (n=16 and 8 respectively). RNA was isolated and reverse transcribed. cDNA was hybridised to the Affymetrix Clariom S Microarray and data was analysed using R/Bioconductor. Differential expression of genes was assessed using empirical Bayes and moderated-t-statistic approaches. Category enrichment analysis was used with a tissue-specific background to examine the biological context of differentially expressed genes. Selected differentially regulated genes were validated by qPCR. ELISA for Intelectin-1 was performed on IAAT samples for the corresponding patients. The current cohort plus 12 additional patients from each group also had plasma Intelectin-1 ELISA carried out.
In IAAT versus SAT comparisons there were 2101, 1722 and 1659 significantly regulated genes in the cachectic, weight stable and control groups respectively. There were 2200 significantly regulated genes from IAAT in cachectic patients compared to controls and 1253 significantly regulated genes from IAAT in weight stable cancer patients compared to controls. The gene showing the largest difference in expression was Intelectin-1 (Omentin-1) (FDR corrected p=0.0001); a novel adipocytokine associated with weight loss in other groups. Genes involving inflammation were enriched in cancer and control IAAT versus SAT though different groups of genes contributed. Energy metabolism and fat browning genesets were downregulated in cancer IAAT as were key fat browning genes (e.g. UCP1).
SAT and IAAT have unique gene expression signatures. IAAT is metabolically active in cancer, and maybe a target for therapeutic manipulation. IAAT may play a fundamental role in cachexia, but the downregulation of energy metabolism genes implies a limited role for fat browning in human cachectic patients, in contrast to pre-clinical models.

overall_design: 8 patients from 3 groups (cancer cachexia, cancer weight stable and control) were included. Gene expression in visceral and subcutaneous adipose tissue from each subject was analysed using the Clariom S microarray platform.


title: Phosphorylation of the transcriptional coregulator alphaNAC by DNA-PKcs regulates osteoblastogenesis

status: Public on May 28 2020

city: Montreal

first: Rene

last: St-Arnaud

state: Quebec

address: 1003 Decarie Blvd

department: Human Genetics

institute: McGill University

summary: In order to identify pathways affected by inhibition of the DNA-dependent protein kinase, catalytic subunit in osteoblasts, we performed a whole genome gene expression profiling using the Affymetrix mouse GeneChip system (MoGene-2.0 gene ST arrays). The osteoblastic cell line MC3T3-E1 was grown in osteogenic medium and treated with DMSO (vehicle) or NU7441, an inhibitor of DNA-dependent protein kinase, for 14 days. Six biological samples were used in this experiment, three replicates in the control group and three replicates in the treated group. Isolated RNA was subjected to microarray analysis.

overall_design: In total 6 samples; 3 controls and 3 treatment.

first: Theresa

last: Farhat

first: Amel

last: Dudakovic

first: Caitlin

last: Anderson

first: Jay

middle: H

last: Chung

first: Andre

last: van Wijnen

first: René


title: Enhanced YB1/EphA2 axis signaling promotes acquired resistance to sunitinib and metastatic potential of renal cell carcinoma

city: wuhan

first: Hailong

last: Ruan

state: hubei

address: 1277 Jiefang Avenue, Wuhan, 430022, Hubei Province, China

institute: wuhan union hospital

summary: VHL mutations are the most common tumorigenic lesions in clear cell renal cell carcinoma (ccRCC) and result in continued activation of the HIF/VEGF pathway and uncontrolled cancer progression. Receptor tyrosine kinase (RTK) inhibitors such as sunitinib have been demonstrated to target tumorigenic signaling pathways, delay tumor progression and improve patient prognosis in metastatic renal cell carcinoma (mRCC). Although several mechanisms of sunitinib resistance have been reported, the solutions to overcome this resistance remain still unclear. In our study, we found that increased expression of YB1 (Y box binding protein 1, a multidrug resistance associated protein) and EphA2 (a member of erythropoietin-producing hepatocellular (Eph) receptors, belonging to the RTK family) mediated sunitinib resistance and mRCC exhibited a large phenotypic dependence on YB1 and EphA2. In addition, our findings confirm that YB1 promotes the invasion, metastasis and sunitinib resistance of ccRCC by regulating the EphA2 signaling pathway. Furthermore, pharmacological inhibition of EphA2 through the small molecule inhibitor ALW-II-41-27 reduced the proliferation of sunitinib-resistant tumor cells and suppressed tumor growth in vivo and restored the sensitivity of sunitinib-resistant tumor cells to sunitinib in vitro and in vivo. Mechanistically, YB1 increases the protein levels of EphA2 by maintaining the protein stabilization of EphA2 through inhibiting the proteasomal degradation pathway. Collectively, our findings provide the theoretical rationale that ccRCC metastasis and RTK-directed therapeutic resistance could be prospectively and purposefully targeted.

overall_design: RNA-seq for YB1 and input control. RNA-seq for three independent shRNA against YB1, as well as scrambled control, each performed in triplicate.

first: Xiaoping

last: Zhang


title: Essential role for CD30-Transglutaminase 2 axis in memory Th1 and Th17 cell generation.

city: Chiba

first: Atsushi

last: Onodera

address: Chuo-ku Inohana 1-8-1

department: Immunology

institute: Chiba University

summary: Memory helper T (Th) cells are crucial for secondary immune responses against infectious microorganisms but also drive the pathogenesis of chronic inflammatory diseases. Therefore, it is of fundamental importance to understand how memory T cells are generated. However, the molecular mechanisms governing memory Th cell generation remain incompletely understood. Here, we identified CD30 as a molecule heterogeneously expressed on effector Th1 and Th17 cells, and CD30hi effector Th1 and Th17 cells preferentially generated memory Th1 and Th17 cells. We found that CD30 mediated signal induced Transglutaminase-2 (TG2) expression, and that the TG2 expression in effector Th cells is essential for memory Th cell generation. In fact, Cd30-deficiency resulted in the impaired generation of memory Th1 and Th17 cells, which can be rescued by overexpression of TG2. Furthermore, transglutaminase-2 (Tgm2)-deficient CD4 T cells failed to become memory Th cells. As a result, T cells from Tgm2-deficient mice displayed impaired antigen-specific antibody production and attenuated Th17-mediated allergic responses. Our data indicate that CD30-induced TG2 expression in effector Th cells is essential for the generation of memory Th1 and Th17 cells, and that CD30 can be a marker for precursors of memory Th1 and Th17 cells.

overall_design: CD30high and CD30low effector Th1 and Th17 cells are profiled for mRNA expression

first: Akane

middle: S

last: Suzuki

first: Ryoji

last: Yagi

first: Motoko

middle: Y

last: Kimura

first: Chiaki

last: Iwamura

first: Kenta

last: Shinoda

first: Kiyoshi

last: Hirahara

first: Damon

last: Tumes

first: Ryo

last: Koyama-Nasu

first: Siiri

last: Iismaa

first: Robert

middle: M

last: Graham

first: Shinichiro

last: Motohashi

first: Toshinori

last: Nakayama


title: Gene Expression Profiling of TNBC and OVCA in response to Azacitidine and Talazoparib (RNA-Seq)

city: Baltimore

first: Michael

middle: Jonathan

last: Topper

address: 1650 Orleans St., Suite 530

department: Oncology

institute: Johns Hopkins University

summary: Triple negative breast cancer and ovarian cancer cells were treated with Azacitidine, Talazoparib, or combination for a period of 10 days to assess global transcriptional response. The focus of these transcriptional studies were on the assessment of DNA repair and immune pathway alteration.

overall_design: Therapeutic induced gene expression was assayed in human cell lines after 10 days of continous exposure to doses of  Azacitidine, Talazoparib, or combination treatment.


title: RAW264.7 cells versus H.pylori infected RAW264.7 cells

city: Kuala Lumpur

first: Won

middle: Fen

last: Wong

address: Lembah Pantai

department: Department of Medical Microbiology,  Faculty of Medicine

country: Malaysia

institute: University of Malaya

summary: Comparing gene expression in the RAW264.7 cells before and after H. pylori infection at MOI 10 for 24 hours

overall_design: MOCK vs Hp infected; 2 biological duplicates

middle: F

first: Grace

last: Tan

first: Cheok

last: Ying


title: bacTRAP profiling of seven different types of neurons , vulnerable and resistant to Alzheimer's disease (AD) across the lifetime of the mouse

first: Jean-Pierre

last: Roussarie

summary: We report here the bacTRAP (bacterial artificial chromosome , translating ribosome affinity purification) profiling of 7 different types of neurons in the mouse, at three different ages: two neuron types very vulnerable to AD (principal cells of entorhinal cortex layer II - ECII), pyramidal cells of hippocampus CA1, and 5 types of neurons more resistant to AD (pyramidal cells of hippocampus CA2 and CA3, granule neurons of the dentate gyrus, pyramidal cells from layer IV of primary visual cortex V1, and pyramidal cells from layer II/III and V of primary somatosensory cortex S1). Using these profiles we generated molecular signatures for each of these cell types, proved that the molecular identity of these cell types is very well conserved across mouse and humans, and constructed functional networks for each cell type that allowed us to identify genes and pathways associated with selective neuronal vulnerability in AD. We also report the profiling of ECII neurons in APP/PS1 mice (a mouse model of Abeta accumulation) at 6 months of age.

overall_design: We constructed bacTRAP mice overexpressing eGFP-L10a in a cell-type specific manner. Cell-type specific expression was achieved using bacterial artificial chromosomes (BAC) that contain the regulatory regions of cell-type specific genes. For ECII neurons, we used four different lines: two Rasgrp2-bacTRAP lines (using BACs RP23-199D5 and RP24-344N1), two different founder lines for Sh3bgrl2 (using BAC RP23-307B16); for CA1 neurons, two different lines: one SSTR4-bacTRAP (BAC RP23-126C5), and one Cck-bacTRAP line generated previously by the Heintz lab; for CA2 neurons: one Cacng5-bacTRAP line (BAC RP23- 329L1); for CA3 neurons: one Gprin3-bacTRAP line generated previously by the Heintz lab; for dentate gyrus granule neurons: one SSTR4-bacTRAP line (using BAC RP23-126C5, another founder from the one used for CA1, with ectopic expression in DG granule neurons); for V1: one Calca-bacTRAP line (BAC RP23-181A2); for S1: one Cartpt-bacTRAP line (BAC RP24-68J22). We used these mice to immunoprecipitate polysomes from these neurons at 4-5, 12 and 24 months of age, and profiled the isolated mRNA using RNAseq. Additionally, we profiled APP/PS1 mice crossed to ECII-bacTRAP mice (one of the Sh3bgrl2-bacTRAP lines) at 6 months of age to study the effects of Amyloid accumulation on ECII gene expression. In the end we had 3 to 12 replicates for each neuron type at each age. Each replicate consisted in the pooling of 2 different mice. Male mice were used for the profiling in wild-type mice. For the APP/PS1 profiling we used male and female mice.


title: Dppa2/4 Counteract De Novo Methylation to Establish a Permissive Epigenome for Development [Cut&Run]

city: Monterotondo

first: Kristjan

middle: Holm

last: Gretarsson

state: Roma

address: Via Ercole Ramarini, 32

country: Italy

institute: EMBL Rome

summary: Early mammalian development entails widespread epigenome remodeling, including DNA methylation erasure and reacquisition, which facilitates developmental competence. To uncover the mechanisms that orchestrate global and focal DNA methylation (DNAme) dynamics, we coupled a single-cell ratiometric DNAme reporter with unbiased CRISPR screening. We identify key genes and regulatory pathways that drive developmental DNA hypomethylation, and characterise roles for Cop1 and Dusp6. We also identify Dppa2 and Dppa4 as essential safeguards of focal epigenetic states during (re)programming phases. In their absence, developmental genes and evolutionary young LINE1 elements lose H3K4me3 and gain ectopic de novo DNA methylation. Consequently, lineage-associated genes (and LINE1) acquire a repressive epigenetic memory in pluripotent cells, which renders them incompetent for activation during future lineage-specification. Dppa2/4 thereby sculpt a permissive epigenome for development by targeting H3K4me3 to counteract de novo methylation during (re)programming; a function that has been co-opted by evolutionary young LINE1 to evade epigenetic decommissioning.

overall_design: Examination of Dppa2, H3K4me3, H3K27me3 and H3K9me3 genome localisation in WT,  Dppa2 KO and Dppa4 KO in ESC and EpiLC using Cut and Run

first: Jamie

middle: A

last: Hackett


title: Dppa2/4 Counteract De Novo Methylation to Establish a Permissive Epigenome for Development [CRISPR]

overall_design: Genome wide CRISPR knockout screening coupled with the eRGM cell lines to unbiasedly identify the gene networks that underpin DNA methylation remodeling


title: Dppa2/4 Counteract De Novo Methylation to Establish a Permissive Epigenome for Development [EnzymaticMethyl-seq]

overall_design: Examination of the DNA methylome in WT and Dppa2/4 knock out cell line (ESC and EpiLC)  using EMseq


title: STAT5 is required for lipid breakdown and beta-adrenergic responsiveness of brown adipose tissue

first: Tobias

last: Suske

state: Vienna

address: Veterinärplatz 1

department: Functional Cancer Genomics

institute: University of Veterinary Medicine

summary: Increasing energy expenditure through activation of brown adipose tissue (BAT) thermogenesis is an attractive approach to counteract obesity. Thus, it is essential to understand molecular mechanisms that control BAT functions. Here, we describe signal transducer and activator of transcription (STAT) 5 as key regulator of BAT functionality. We found that STAT5 is necessary for acute cold-induced temperature maintenance and stimulated lipid breakdown in BAT using mice that harbour an adipocyte-specific deletion of Stat5a/b genes. In addition, the mitochondrial respiratory capacity of primary differentiated brown adipocytes from STAT5 deficient mice was diminished. We show that increased sensitivity to cold stress upon STAT5 deficiency was associated with reduced expression of thermogenic key player uncoupling protein 1, while decreased stimulated lipolysis of STAT5-deficient BAT explants was linked to decreased protein kinase A activity. In addition, brown remodeling of white fat was diminished following chronic β-adrenergic stimulation. This impairment was linked to a decrease in mitochondrial functionality. We conclude that STAT5 is essential for the β-adrenergic responsiveness of brown adipose tissue and the physiologic function of thermogenic adipose tissue.

overall_design: Wildtype and Stat5Adipoq, 5 replicates per condition

first: Doris

last: Kaltenecker

first: Katrin

last: Spirk

first: Frank

last: Ruge

first: Florian

last: Grebien

first: Anne

last: Rupprecht

first: Lukas

last: Kenner

first: Elena

last: Pohl

first: Kristina

last: Mueller

first: Richard

last: Moriggl


title: Dppa2/4 Counteract De Novo Methylation to Establish a Permissive Epigenome for Development [RNA-seq]

overall_design: Examination of mRNA levels in WT, Dppa2KO, Dppa4KO, Dusp6KO and Cop1KO in ESC and EpiLC using RNAseq


city: Birmignham

first: Muhan

last: Hu

address: 1900 University BLVD

department: CDIB

country: USA

institute: University of Alabama at Birmingham

summary: total RNA from N2, daf-1(m40), and daf-1(m40);daf-3(mgDf90) adult, gravid hermaphrodites were extracted and sequenced.

overall_design: 3 biological replciates per genotype were harvested

first: Rosa

last: Serra


title: IL-34 actions on FOXP3+ Tregs and CD14+ monocytes control human graft rejection

status: Public on May 27 2020

city: NANTES

first: Céline

last: Sérazin

address: 30 bd Jean monnet

institute: University of Nantes

summary: Overall, we show that IL-34 induced the differentiation of human monocytes with a particular transcriptional profile and these cells favor the development of potent suppressor CD4+ and CD8+ FOXP3+ Tregs.

overall_design: mRNA profils of 5 conditions Monocytes, M1 and M2 macrophages, IL34 induced macrophages and M-CSF induced macrophages, from 3 healthy volunteers


title: Sedentary and Trained Older Adults Have Distinct Circulating Exosomal microRNA Profiles at Baseline and in Response to Acute Exercise

city: New York

first: Yongchao

last: Ge

address: One Gustave L Levy Place #1137

department: Neurology

institute: Icahn School of Medicine at Mount Sinai

summary: Exercise has multi-systemic benefits and attenuates the physiological impairments associated with aging. Emerging evidence suggests that circulating exosomes mediate some of the beneficial effects of exercise via the transfer of microRNAs between tissues. However, the impact of regular exercise and acute exercise on circulating exosomal microRNAs (exomiRs) in older populations remains unknown. In the present study, we analyzed circulating exomiR expression in endurance-trained elderly men and age-matched sedentary males at baseline (Pre), immediately after a forty minute bout of aerobic exercise on a cycle ergometer (Post), and three hours after this acute exercise (3hPost). Plasma exosomes were isolated, characterized, and exomiR levels were determined by sequencing. The effect of regular exercise on circulating exomiRs was assessed using paired t-tests of baseline expression levels in the trained and sedentary groups. The effect of acute exercise was determined by comparing baseline and post-training expression levels in each group. Regular exercise resulted in significantly increased baseline expression of three exomiRs (miR-486-5p, miR-215-5p, miR-941) and decreased expression of one exomiR (miR-151b). Acute exercise altered circulating exomiR expression in both groups. However, exomiRs regulated by acute exercise in the trained group (7 miRNAs at Post and 8 at 3hPost) were distinct from those in the sedentary group (9 at Post and 4 at 3hPost). Pathway analysis prediction and reported target validation experiments revealed that the majority of exercise-regulated exomiRs are targeting genes that are related to IGF-1 signaling, a pathway involved in exercise-induced muscle and cardiac hypertrophy. The immediately post-acute exercise exomiR signature in the trained group correlates with activation of IGF-1 signaling, whereas in the sedentary group it is associated with inhibition of IGF-1 signaling. These results suggest that training status may counteract age-related anabolic resistance by modulating circulating exomiR profiles both at baseline and in response to acute exercise.

overall_design: Sequencing was performed on circulating exosome-derived small RNA samples isolated from 10 elderly individuals (5 endurance-trained and 5 sedentary subjects) at 3 time points: before acute exercise (Pre), immediately after acute exercise (Post), and 3h after acute exercise (3hPost).

first: Venugopalan

middle: D

last: Nair

first: Side

last: Li

first: Hanna

last: Pincas

first: Nimisha

last: Jain

first: Nitish

last: Seenarine

first: Mary-Anne

last: Amper

first: Bret

last: Goodpaster

first: Martin

last: Walsh

first: Paul

last: Coen

first: Stuart

middle: C

last: Sealfon