title: Disparate anticancer activity of structurally similar multi-kinase inhibitors through cumulative differential effects on individual targets

city: Tampa

first: Steven

last: Eschrich

address: 12902 Magnolia Drive

department: Biostatistics & Bioinformatics

institute: Moffitt Cancer Center

summary: Despite recent improvements many cancer patients do not respond to immune or targeted drugs and require new therapies. Through a systems pharmacology approach including phenotypic screening, chemical and phosphoproteomics and RNA-Seq, we elucidated the mechanism of action underlying the differential anticancer activity of two structurally related multi-kinase inhibitors, foretinib and cabozantinib, in lung cancer cells. Biochemical and cellular target validation using probe molecules and RNA interference revealed a polypharmacology mechanism of foretinib involving MEK1/2, FER and AURKB, which were each more potently inhibited by foretinib than cabozantinib. Based on this, we rationally developed a synergistic combination of foretinib with barasertib, a more potent AURKB inhibitor, for MYC-amplified small cell lung cancer. This systems pharmacology approach showed that small structural changes of drugs can cumulatively through multiple targets result in pronounced anticancer activity differences and that detailed mechanistic understanding can lead to new repurposing opportunities for cancers with unmet medical need.

overall_design: The H1155 cell line was profiled under three conditions: treated with DMSO (control), treated with 1um foretinib or treated with 1um cabozantinib. Each condition was profiled in triplicate.

first: Uwe

last: Rix

first: Natalia

last: Sumi


title: Cue2 and Slh1 define parallel pathways to rescue stalled ribosomes

status: Public on Jun 25 2019

city: Frederick

first: Colin

last: Wu

state: MD

department: RNA Biology Laboratory

country: USA

institute: NCI

summary: Ribosome stalling at problematic sequences in mRNAs leads to collisions that trigger a collection of quality control events including ribosome rescue, targeting the nascent polypeptide for decay (Ribosome-mediated Quality Control or RQC), and targeting of the mRNA for decay (No Go Decay or NGD). Using a reverse genetic screen in yeast, we identify Cue2 as the endonuclease that is recruited to stalled ribosomes to promote NGD. Following Cue2-mediated cleavage, ribosomes upstream of the cleavage site translate to the end of the truncated mRNA and are rescued by the Dom34:Hbs1 complex. We also show that the putative helicase Slh1 (part of the RQC Trigger or RQT complex) removes collided ribosomes behind the lead stalled ribosome and thereby reduces endonucleolytic cleavage by Cue2. The synergistic activities of Cue2 and Slh1 define two parallel pathways that allow cells to recognize and respond to ribosomes trapped on problematic mRNAs.

overall_design: 7 samples for ribosome profiling experiments using various yeast strains and 3 samples for in vitro ribosome footprinting experiments using isloated trisomes from sucrose gradients.

first: Rachel

last: Green


title: Glucose metabolism drives epigenetic landscape transitions that dictate muscle stem cell function

city: PHILADELPHIA

first: Arndt

last: Siekmann

state: Pennsylvania

address: 421 Curie Boulevard, 1114 Biomedical Research Building II/III

department: Perelman School of Medicine

institute: UNIVERSITY OF PENNSYLVANIA

type: Genome binding/occupancy profiling by high throughput sequencing

summary: We report the application of chromatin immunoprecpitation (ChIP) and assay for transposase accessible chromatin (ATAC) followed by sequencing to assay chromatin response to metabolic perturbation.

overall_design: Primary myoblasts were treated with dichloroacetate (DCA) for 24 hours, which increases the contribution of glucose to histone acetylation, as well as overall acetylation. ChIP and ATAC-seq was performed to establish the genomic specificity of this response, as well as the resulting effect on chromatin accesibility.

first: Helen

last: Blau

first: Nora

last: Yucel


title: Genome-wide transcript structure resolution in murine gammaherpesvirus 68: Iso-Seq

city: Gainesville

first: Scott

last: Tibbetts

state: FL

department: Molecular Genetics & Microbiology

institute: University of Florida

type: Expression profiling by high throughput sequencing

summary: Pacific Biosciences Iso-Seq was used in conjunction with Illumina RNA-Seq and deepCAGE to globally resolve transcript structures in lytic murine gammaherpesvirus 68.

overall_design: Pacific Biosciences Iso-Seq of MHV68-infected fibroblasts

first: Tina

last: O'Grady

first: April

last: Feswick

first: Brett

last: Hoffman

first: Yiping

last: Wang

first: Erik

middle: K

last: Flemington

middle: A


title: Systematic evaluation of RNA-Seq preparation protocol performance (RNASeq: SMARTer)

status: Public on Jun 27 2019

city: Houston

first: Yue

last: Lu

state: TX

address: 1881 East Road

department: Epigenetics & Molecular Carcinogenesis

institute: The University of Texas MD Anderson Cancer Center

summary: In this study, we present a comprehensive evaluation of four RNA-Seq library preparation methods. We used three standard input protocols, the Illumina TruSeq Stranded Total RNA and TruSeq Stranded mRNA kits, and a modified NuGEN Ovation v2 kit; and an ultra-low-input RNA protocol, the TaKaRa SMARTer Ultra Low RNA Kit v3. Our evaluation of these kits included quality control measures such as overall reproducibility, 5’ and 3’ end-bias, and the identification of DEGs, lncRNAs, and alternatively spliced transcripts. Overall, we found that the two Illumina kits were most similar in terms of recovering DEGs, and the Illumina, modified NuGEN, and TaKaRa kits allowed identification of a similar set of DEGs. However, we also discovered that the Illumina, NuGEN and TaKaRa kits each enriched for different sets of genes.

overall_design: Two mESC cell lines (biological replicates) from Zbtb24 knockout (1lox/1lox) clones are compared with two wild-type (2lox/+) clones (biological replicates) using the TaKaRa SMARTer Ultra Low RNA protocol directly on cells with no RNA preparation step. Total RNA from 100 mESCs cells and 1000 mESCs cells or approximately 1 and 10 ng RNA were used respectively.

first: Hsueh-Ping

last: Chao

first: Yueping

last: Chen

first: Yoko

last: Takata

first: Mary

middle: W

last: Tomida

first: Kevin

last: Lin

first: Jason

middle: S

last: Kirk

first: Melissa

last: Simper

first: Carol

middle: D

last: Mikulec

first: Joyce

middle: E

last: Rundhaug

first: Susan

middle: M

last: Fischer

first: Taiping

first: Dean

middle: G

last: Tang

first: Jianjun

last: Shen


title: Genome-wide transcript structure resolution reveals novel features of gammaherpesvirus transcription and yields an extensively revised murine gammaherpesvirus 68 genome annotation

first: Scott

last: Tibbetts

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

overall_design: Refer to individual Series


title: High-throughput sequence analysis of peripheral T-cell lymphomas indicates subtype specific viral gene expression patterns and immune cell microenvironments

city: New Orleans

state: LA

address: 1430 Tulane Ave

institute: Tulane University

summary: Certain peripheral T-cell lymphomas (PTCLs) have been associated with viral infection, particularly infection with Epstein-Barr virus (EBV). However, a comprehensive virome analysis across PTCLs has not previously been reported, and viral gene expression profiles have been studied only in certain PTCL subtypes. In this study we utilized published RNA-seq data sets from seven different PTCL studies as well as new RNA-seq data from our laboratory to screen for virus association, to analyze viral gene expression, and to assess B- and T-cell receptor diversity paradigms across these tumor types. In addition to identifying EBV in angioimmunoblastic T-cell lymphoma (AITL) and extranodal NK/T cell lymphoma (ENKTL), two PTCL subtypes with well-established EBV associations, we also detected EBV in several cases of anaplastic large cell lymphoma (ALCL), and we found evidence of infection by the oncogenic viruses KSHV and HTLV-1 in isolated PTCL cases. In AITLs, EBV gene expression analysis showed expression of immediate early, early and late lytic genes, suggesting either abortive lytic replication and/or productive infection in a subset of the EBV infected infiltrating B-lymphocytes. Deconvolution of immune cell subpopulations demonstrated a greater B-cell signal in AITLs than in other PTCL subtypes, consistent with a larger role for B-cell support in the pathogenesis of AITL. T-cell receptor (TCR) and B-cell receptor (BCR) repertoires reconstructed from RNA-seq data demonstrated increased BCR diversity in AITLs, consistent with a possible EBV-driven polyclonal response. These findings indicate potential alternative roles for EBV in PTCLs in addition to the canonical oncogenic mechanisms associated with EBV latent infection. The findings also suggest the involvement of other viruses in T-cell lymphoma pathogenesis and demonstrate immunological alterations associated with these cancers.

overall_design: RNA sequencing of five extranodal NK/T-cell lymphoma (ENKTL)-derived cell lines

first: Hani

last: Nakhoul

first: Zhen

first: Xia

first: Claire

last: Roberts

first: Yan

last: Dong


title: Transcriptome analysis of various TOX+ and TOX- tumor-infiltrating CD8 T cells and in vitro TOX over-expressing T cells.

status: Public on Jun 24 2019

city: New York

first: Andrea

last: Schietinger

state: NY

address: 1275 York Avenue

department: Immunology

institute: Memorial Sloan Kettering Cancer Center

summary: TOX is selectively expressed in tumor-infiltrating CD8 T cells however the role of TOX in peripheral CD8 T cells is not known. The two goals of this study are to elucidate transcriptional changes between TOX-sufficient and TOX-deficient tumor infiltrating CD8 T cells and to elucidate the molecular program induced by TOX overexpression in peripheral CD8 T cells.

overall_design: CD8 T cells were sorted by flow cytometry and RNA-seq was performed.

first: Andrew

middle: C

last: Scott

first: Doron

last: Betel

first: Friederike

last: Dündar

first: Paul

last: Zumbo


title: TOX is a critical regulator of tumour-specific T cell differentiation


title: Metabolic regulation of muscle stem cell epigenetic landscape during regeneration

first: Arndt

last: Siekmann

summary: Muscle is known to be a highly glycolytic tissue, yet the impact of glucose metabolism in muscle stem cell function remains unresolved. Here we use Mass Cytometry (CyTOF) to capture changes in histone acetylation profiles at the single cell level in vivo following injury. We demonstrate that MuSCs transiently increase histone acetylation upon activation, followed by global loss upon commitment. The switch to a committed state is driven by glucose metabolism through pyruvate dehydrogenase (PDH), which fuels histone acetylation via production of acetyl-CoA.  Pharmacological or genetic activation of PDH results in increased histone acetylation and impedes myogenic differentiation.  Moreover, mice lacking PDH inhibitory kinases, PDK2 and PDK4, accumulate uncommitted satellite cells and exhibit impaired muscle regeneration. These studies identify PDH as a previously unrecognized dynamic metabolic mediator of epigenetic state crucial to myogenic commitment and regeneration.

overall_design: Fresh versus 48hr cultured muscle stem cells(MuSCs) were sequenced by RNASeq. 3 replicates (F1-3) of freshly isolated MuSCs were run independently, while 2 replicates (F4-5) were run on the same lane as the cultured (C1-3) replicates.

first: Nora

last: Yucel

first: Helen

last: Blau


title: Genome-wide transcript structure resolution in murine gammaherpesvirus 68: 3T12 deepCAGE

summary: deepCAGE was used in conjunction with Pacific Biosciences Iso-Seq and Illumina RNA-Seq to globally resolve transcript structures in lytic murine gammaherpesvirus 68.

overall_design: deepCAGE of MHV68-infected fibroblasts

first: Tina

last: O'Grady

first: April

last: Feswick

first: Brett

last: Hoffman

first: Yiping

last: Wang

first: Erik

last: Flemington


title: Genome-wide transcript structure resolution in murine gammaherpesvirus 68: Illumina RNA-Seq

first: Scott

last: Tibbetts

summary: Strand-specific Illumina RNA-Seq was used in conjunction with Pacific Biosciences Iso-Seq and deepCAGE to globally resolve transcript structures in lytic murine gammaherpesvirus 68.

overall_design: Strand-specific Illumina RNA-Seq of MHV68-infected fibroblasts


title: ATACseq profiling of various TOX+ and TOX- tumor-infiltrating CD8 T cells

summary: TOX is selectively expressed in tumor-infiltrating CD8 T cells however the role of TOX in peripheral CD8 T cells is not known. The goal of this study is to elucidate changes in chromatin accessibility determined by ATAC-seq between TOX-sufficient and TOX-deficient tumor infiltrating CD8 T cells isolated from murine tumors.

overall_design: CD8 T cells were sorted by flow cytometry and ATAC-seq was performed.


title: A streamlined protocol and analysis pipeline for CUT&RUN chromatin profiling

city: Seattle

first: Jorja

last: Henikoff

state: WA

address: 1100 Fairview AV N, A1-162

department: Basic Sciences

institute: Fred Hutchinson Cancer Research Center

summary: Over the past two decades, the dominant technology for chromatin profiling has been chromatin immunoprecipitation (ChIP), in which cellular components are solubilized to fragment chromatin, and an antibody is added to precipitate a DNA/protein complex of interest. We previously described a novel alternative to ChIP, Cleavage Under Targets & Release Using Nuclease (CUT&RUN), in which the antibody is added to permeabilized cells followed by binding of a Protein A-Micrococcal Nuclease (pA/MNase) fusion protein to the antibody (PMID 29651053). Upon activation of tethered MNase, the bound complex is excised and released into the supernatant for DNA extraction and sequencing. Here we introduce four extensions to CUT&RUN: 1) a hybrid Protein A-Protein G-MNase construct that allows simplified purification using a commercial kit; 2) a modified digestion protocol that prevents release of the enzyme and so minimizes artifactual cleavage of accessible DNA; 3) a calibration strategy based on carryover of E. coli DNA introduced with the fusion protein; and 4) a novel peak-calling strategy that is customized for the low-background profiles obtained using CUT&RUN. These new features, coupled with the previously described low-cost, high efficiency, high reproducibility and high-throughput capability of CUT&RUN make it the method of choice for routine epigenomic profiling.

overall_design: We used Cleavage under targets and Release using nuclease (Cut-and-Run), a chromatin profiling strategy in which antibody-targeted controlled cleavage by micrococcal nuclease releases specific protein-DNA complexes into the supernatant for paired-end DNA sequencing.

first: Michael

middle: P

last: Meers

first: Terri

last: Bryson

first: Steven


title: EZH2 upregulates the PI3K/AKT pathway through IGF1R and MYC in clinically aggressive CLL

city: Gothenburg

first: Subazini

last: TK

address: Medicineragatan 9A

country: Sweden

institute: Gothenburg university

summary: In this study, using ChIP sequencing, we identified EZH2 target genes in two prognostic subgroups of chronic lymphocytic leukemia with distinct outcome, i.e. IGHV-unmutated/subset #1 and IGHV-mutated/subset #4. Our data identified many oncogenic pathways enriched equally by EZH2 target genes in both prognostic subgroups. Importantly, we identified PI3K pathway as differentially enriched pathway by EZH2 between the two prognostic sub groups. Validation of EZH2 target genes for EZH2 occupancy on selected PI3K pathway target genes were performed using independent CLL patient samples and CLL cell lines using siRNA mediated down regulation and ChIP assays. In conclusion, for the first time we characterized EZH2 target genes in CLL to understand the role of EZH2 in regulating the oncogenic pathways for improved therapeutic intervention and designing better drugs for targeting EZH2 in CLL.

overall_design: Investigating the target genes for EZH2 and H3K27me3 enrichment using ChIP seq in chronic lymphocytic leukemia patient samples from age matched healthy individuals.

first: M

last: Kanduri

first: S

middle: T

last: Kosalai


title: Specificity assessment of allele selective Zinc Finger Protein Repressors in hESC-derived HD neurons

status: Public on Jun 30 2019

city: Richmond

first: Bryan

last: Zeitler

state: California

address: 501 Canal Blvd.

institute: Sangamo Therapeutics

summary: Huntington’s disease (HD) is a dominantly inherited neurodegenerative disorder caused by a CAG trinucleotide expansion in the huntingtin gene (HTT), which codes for the pathologic mutant HTT (mHTT) protein. Since normal HTT is thought to be important for brain function, we engineered zinc finger protein transcription factors (ZFP-TFs) to target the pathogenic CAG repeat and selectively lower mHTT as a therapeutic strategy. Using patient-derived fibroblasts and neurons, we demonstrate that ZFP-TFs selectively repress >99% of HD-causing alleles over a wide dose range, while preserving expression of >86% of normal alleles. Other CAG-containing genes are minimally affected, and virally delivered ZFP-TFs are active and well tolerated in HD neurons beyond 100 days in culture and at least 9 months in the mouse brain. Using three HD mouse models, we demonstrate improvements in a range of molecular, histopathological, electrophysiological, and functional endpoints. Our findings support the continued development of an allele-selective ZFP-TF for the treatment of HD.

overall_design: Four treatments (ZFP-A, ZFP-B, ZFP-C, GFP) were tested in neurons derived from HD hESCs (GENEA020). Each ZFP was tested in a different batch with the GFP control. For each batch of treatment and control, 4 replicate transfections were conducted yielding 4 biological replicates. Cells were harvested 24 hours later and processed for total RNA isolation and Affymetrix sample Prep. The same control GFP treatment was used for each batch.

last: Froelich

first: Kimberly

last: Marlen

first: David

last: Shivak

first: Qi

last: Yu

first: Davis

last: Li

first: Jocelynn

middle: R

last: Pearl

first: Jeffrey

last: Miller

first: Lei

last: Zhang

last: Paschon

first: Sarah

middle: J

last: Hinkley

first: Irina

last: Ankoudinova

first: Stephen

last: Lam

first: Dmitry

last: Guschin

first: Lexi

last: Kopan

first: Jennifer

last: Cherone

first: Hoang-Oanh

middle: B

last: Nguyen

first: Guijuan

last: Qiao

first: Yasaman

last: Ataei

first: Matthew

last: Mendel

first: Rainier

last: Amora

first: Richard

last: Surosky

first: Josee

last: Laganiere

first: B

last: Vu

first: Anand

last: Narayanan

first: Yalda

last: Sedaghat

first: Karsten

last: Tillack

first: Christina

last: Thiede

first: Annette

last: Gärtner

first: Seung

last: Kwak

first: Jonathan

last: Bard

first: Ladislav

last: Mrzljak

first: Larry

last: Park

first: Taneli

last: Heikkinen

first: Kimmo

last: Lehtimäki

first: Marie

last: Svedberg

first: Jenny

last: Häggkvist

first: Lenke

last: Tari

first: Miklós

last: Tóth

first: Andrea

last: Varrone

first: Christer


title: Functional diversity of inhibitors tackling the differentiation arrest of MLL-rearranged leukemia

city: Berlin

first: Bernard

last: Haendler

address: Muellerstr. 178

country: Germany

institute: Bayer AG

summary:
Purpose: The chromosomal rearrangements of the mixed-lineage leukemia (MLL) gene have been extensively characterized as a potent oncogenic driver on the molecular and mechanistic level in acute lymphoblastic (ALL) and acute myeloid (AML) leukemia. For its oncogenic function the MLL fusion protein is hijacking the the multi enzyme super elongation complex (SEC) leading to elevated expression of MLL target genes (e.g. HOXA9 and MEIS). High expression of MLL target genes is overwriting the normal hematopoietic differentiation gene expression program, resulting in undifferentiated blasts cells having a more “stem-cell like” cancer-promoting phenotype. Although extensive resources have been devoted to a better understanding of therapeutic targets for the MLL fusion to overcome the de-differentiation, the inter-dependencies of those targets for the pathophysiology of MLL is still barely understood. Here we report a comparative mode of action analysis of different inhibitors potentially interfering with MLL fusion induced differentiation blockade. We used RNA-seq for transcriptomic profiling in 14 AML and ALL cell lines treated with DMSO control or one of the 5 studied inhibitors (EPZ-5676, Brequinar, BAY-155, BAY-1251152 and OTX015).
Methods: We used RNA-seq for transcriptomic profiling in 14 AML and ALL cell lines treated with DMSO control or one of the 5 studied inhibitors (EPZ-5676, Brequinar, BAY-155, BAY-1251152 and OTX015).
Results: We discovered significant differences between compounds in their ability to induce differentiation and interfere with MLL target genes expression. We observed that Menin and DOT1L inhibition act very specifically on MLL fused leukemia cell lines, whereas inhibition of BET, DHODH and P-TEFb have strong effects beyond the MLL fusion .
Conclusions: These results show a substantial diversity in the molecular activities of those inhibitors and provide valuable insights into the further developmental potential as single agents or in combinations in MLL fused leukemias.

overall_design: RNA-sequencing of AML and ALL cell lines treated with inhibitors for 8h (BAY-1251152 - 0.05 µM, OTX015 - 1 µM), 24h (BAY-155 - 2 µM, Brequinar - 2 µM, DMSO – 0.1%) and 96h (EPZ-5676 - 3 µM, DMSO – 0.1%), three biological replicates per condition.

first: Krzysztof

last: Brzezinka

first: Ekaterina

last: Nevedomskaya

first: Ralf

last: Lesche

first: Carlo

last: Stresemann


title: Genomic analyses of primary mouse prostate organoid culture with overexpression of FOXA1

status: Public on Jun 21 2019

first: Hyein

middle: Sophia

last: Cho

address: 430 E 67th St

department: Developmental Biology

summary: Mutations in the FOXA1 transcription factor define a unique subset of prostate cancers but the functional consequences of these mutations and whether they confer gain or loss of function is unknown. By annotating the FOXA1 mutation landscape from 3086 human prostate cancers, we define two hotspots in the forkhead domain: Wing2 (~50% of all mutations) and R219 (~5%), a highly conserved DNA contact residue. Clinically, Wing2 mutations are seen in adenocarcinomas at all stages, whereas R219 mutations are enriched in metastatic tumors with neuroendocrine histology.  Interrogation of the biologic properties of FOXA1WT and 14 FOXA1 mutants revealed gain-of-function in mouse prostate organoid proliferation assays. ATAC-seq of FOXA1WT and representative Wing2 and R219 mutants revealed dramatic, mutant-specific changes in open chromatin at thousands of genomic loci, together with novel sites of FOXA1 binding and associated increases in gene expression. Of note, peaks in R219 mutant expressing cells lack the canonical core FOXA1 binding motifs (GTAAAC/T) but are enriched for a related motif (GTAAAG/A), which is preferentially activated by R219 mutant FOXA1 in reporter assays. Thus, FOXA1 mutations alter its normal pioneering function and promote oncogenesis through perturbation of normal luminal epithelial differentiation programs, providing further support to the role of lineage plasticity in cancer progression.

overall_design: Examination by genotypes and antibodies prepared in 2 replicates

first: Elizabeth

last: Adams

first: Charles

last: Sawyers


title: Gene expression profilings for prostate cancer cells after inhibition of PARP1 or PARP2 using pharmaceutical or siRNA-based approaches

city: Boston

first: Li

last: Jia

state: MA

address: 20 Shattuck Street

department: Surgery

institute: Brigham and Women's Hospital

summary:
Androgen receptor (AR) signaling is a key driver of prostate cancer (PCa) growth and progression. Understanding the factors influencing AR-mediated transcription provides new opportunities for therapeutic intervention. We have previously discovered that a genetic variant in one of the DNA repair genes, PARP2, is associated with aggressive PCa. Here, we show that a high expression level of PARP2 in PCa tumors is associated with high Gleason scores and biochemical recurrence using The Cancer Genome Atlas (TCGA) dataset. Functional studies reveal that PARP2 enhances AR-mediated gene expression through interacting with the pioneer factor FOXA1 and facilitating AR recruitment to the enhancer regions genome-wide in PCa cells. Selective targeting of PARP2, but not PARP1, by genetic or pharmacological means blocks interaction between PARP2 and FOXA1, which in turn attenuates AR-mediated transcription and inhibits AR-positive PCa growth.
SIGNIFICANCE: Current anti-androgens act through blocking ligand binding or inhibiting androgen synthesis. Selective targeting of PARP2 may provide a novel therapeutic approach for AR inhibition by disruption of FOXA1 function, which may be beneficial to patients, irrespective of their DNA repair defect status and, more importantly, when direct AR-targeted therapies fail.

overall_design: Gene expression by RNA-Seq of LNCaP cells treated with PARP inhibitors or after PARP1/PARP2 gene knockdown, with biological duplicates.

first: Bin

last: Gui


title: Genome-wide occupation of AR, FOXA1, and H3K27AC in LNCaP cells treated with selective PARP2 inhibitor UPF-1069

overall_design: Genome-wide occupation of AR, FOXA1, and H3K27AC, by ChIP-seq, in LNCaP cells treated with UPF-1069 or vehicle control, with biological duplicates.


title: Gene expression analysis of primary mouse prostate organoid culture with overexpression of FOXA1

overall_design: Examination by genotypes and days elapsed prepared in 3 replicates


title: TGF-β promotes genomic instability after loss of RUNX3

status: Public on Jun 20 2019

city: Singapore

first: Tuan Zea

last: Tan

address: 14 Medical Drive, #11-01, Singapore

department: Cancer Science Institute Singapore

country: Singapore

institute: National University of Singapore

summary: Studies of genomic instability have historically focused on intrinsic mechanisms rather than extrinsic  mechanisms based in the tumor microenvironment (TME). TGF-β is the most abundantly secreted  cytokine in the TME where it imparts various aggressive characteristics including invasive migration,  drug resistance and epithelial-to-mesenchymal transition (EMT). Here we show that TGF-β also  promotes genomic instability in the form of DNA double strand breaks (DSB) in cancer cells which  lack the tumor suppressor gene RUNX3. Loss of RUNX3 resulted in transcriptional downregulation of  the redox regulator heme oxygenase-1 (HO-1 or HMOX1). Consequently, elevated oxidative DNA  damage disrupted genomic integrity and triggered cellular senescence, which was accompanied by  tumor-promoting inflammatory cytokine expression and acquisition of the senescence-associated  secretory phenotype (SASP). Recapitulating the above findings, tumors harbouring a TGF-β gene  expression signature and RUNX3 loss exhibited higher levels of genomic instability. In summary,  RUNX3 creates an effective barrier against further TGF-β-dependent tumor progression by preventing  genomic instability. These data suggest a novel cooperation between cancer cell-extrinsic TGF-β  signaling and cancer cell-intrinsic RUNX3 inactivation as aggravating factors for genomic instability.

overall_design: A549 cells were seeded into 6-cm dishes at the cell density of 0.2X106 cells/dish and transfected with control siRNA or RUNX3 siRNA. 3 days later, cells were trypsinized and subjected to a second round of siRNA. 24 hours later, cells were exposed to either vehicle control or TGF-β for 48 hours followed by RNA sequencing. C1 and C2 samples are biological replicates of control SiRNA transfection; C-1TGF and C-2TGF are biological replicates following TGF-β for 48 hours. R1 and R2 samples are biological replicates following RUNX3 SiRNA; R-1TGF and R-2TGF are biological replicates following TGF-β for 48 hours of cells transfected with RUNX3 siRNA.

first: Vaidehi

last: Krishnan

first: Yoshiaki

last: Ito


title: Primary mouse prostate organoid culture with overexpression of FOXA1


title: mTOR hyperactivation in Down Syndrome mediates deficits in autophagy induction, autophagosome formation, and mitophagy

status: Public on Jun 22 2019

city: Orangeburg

first: Sandipkumar

last: Darji

address: 140 Old Orangeburg Rd, Bldg 39

department: Center for Dementia Research

institute: Nathan Kline Institute for Psychiatric Research

summary: Down syndrome (DS), a complex genetic disorder caused by chromosome 21 trisomy, is associated with mitochondrial dysfunction leading to the accumulation of damaged mitochondria. Here we report that mitophagy, a form of selective autophagy activated to clear damaged mitochondria is deficient in primary human fibroblasts derived from individuals with DS leading to accumulation of damaged mitochondria with consequent increases in oxidative stress.  We identified two molecular bases for this mitophagy deficiency: PINK1/PARKIN impairment and abnormal suppression of macroautophagy.  First, strongly downregulated PARKIN and the mitophagic adaptor protein SQSTM1/p62 delays PINK1 activation to impair mitophagy induction after mitochondrial depolarization by CCCP or antimycin A plus oligomycin. Secondly, mTOR is strongly hyper-activated, which globally suppresses macroautophagy induction and the transcriptional expression of proteins critical for autophagosome formation such as ATG7, ATG3 and FOXO1. Notably, inhibition of mTOR complex 1 (mTORC1) and complex 2 (mTORC2) using AZD8055 (AZD) restores autophagy flux, PARKIN/PINK initiation of mitophagy, and the clearance of damaged mitochondria by mitophagy. These results recommend mTORC1-mTORC2 inhibition as a promising candidate therapeutic strategy for Down Syndrome.

overall_design: mRNA-Seq profiling of 9 2N and 8 DS human fibroblasts samples of age 5 months (< 1 year) and 2 years. These samples come from 5 unrelated 2N individuals (of which 2 individuals, one each of 5 months and 2 years, have 3 replicates each) and 3 unrelated DS individuals (of which 2 individuals, one each of 5 months and 2 years, have 3 replicates each). Five samples were reanalyzed from GSE55504.

first: Matteo

last: Bordi

first: Yutaka

last: Sato

first: Marian

last: Mellén

first: Martin

last: Berg

first: Asok

last: Kumar

first: Ying

last: Jiang

first: Ralph

last: Nixon


title: Selective Targeting of PARP2 Inhibits Androgen Receptor Signaling and Prostate Cancer Growth Through Disruption of FOXA1 Function

overall_design: Refer to individual Series


title: Gene expression profiling of catfish after treatment with Flavobacterium columnare DnaK protein

city: Auburn

first: Jason

last: Abernathy

state: AL

address: 990 Wire Road

institute: USDA-ARS

summary: Catfish farming remains number one among U.S. aquaculture production. Flavobacterium columnare, the causative agent of columnaris disease, produces substantial mortality during the production of freshwater farmed finfish species. F. columnare is ubiquitous in the aquatic environment, and outbreaks of disease are often triggered during the spring and summer months of the catfish production cycle. As food fish production continues to increase, the frequency of columnaris disease will only continue to rise within the aquaculture industry and thus new preventative measures will be required. Vaccination against different fish pathogens has proven to be an effective disease management practice within the aquaculture industry. To evaluate the immunological effect of bath immunization on catfish with a recombinant F. columnare DNAk protein vaccine, we performed RNA sequencing on skin samples from control and rDNAK immunized catfish at different time intervals. We evaluated the different gene expression patterns between the groups with a focus on identifying differences in innate and adaptive immune function.

overall_design: Channel catfish were spawned and reared at the Stuttgart National Aquaculture Research Center. Catfish averaging 5 g were either sham-bath immunized (control) or immunized in a solution of 200 ug/mL of recombinant Flavobacterium columnare Dnak protein. Skin was sampled from euthanized catfish from each group at different time intervals and skin was placed into RNA later. Skin samples from six catfish from the non-immunized, six from the rDNAK group at week 1, and six from the rDNAk group at week 6 were used to isolate total RNA and construct sequencing libraries.

first: Miles

last: Lange


title: Comparing 3D genome organization in multiple species using Phylo-HMRF

status: Public on Jun 19 2019

city: Pittsburgh

first: Yang

last: Yang

state: PA

address: 5000 Forbes Ave

department: Computational Biology

institute: Carnegie Mellon University

summary: We developed a continuous-trait probabilistic model to identify genome-wide evolutionary patterns of 3D genome structures based on multi-species Hi-C data. We applied the proposed method to analyzing cross-species Hi-C data from the same cell type in multiple primate species. The results showed that the proposed method enables discovery of distinct Hi-C contact frequency evolutionary patterns across species.

sample_organism:
Gorilla gorilla
Pan paniscus
Pan troglodytes

overall_design: Comparing Hi-C contact frequencies among multiple primate species

last: Zhang

first: Bing

last: Ren

first: Jesse

last: Dixon

first: Jian

last: Ma


title: Chromatin Landscape Analysis of primary mouse prostate organoid culture with overexpression of FOXA1

first: Hyein

last: Cho

first: Elizabeth

last: Adams

first: Charles

last: Sawyers


title: Rapid Irreversible Transcriptional Reprogramming in Human Stem Cells Accompanied by Discordance between Replication Timing and Chromatin Compartment [Expression]

city: Tallahassee

first: David

last: Gilbert

address: 319 Stadium Drive

department: Biology

institute: Florida State University

summary: This study demonstrates the relationship between replication timing, transcription and chromatin compartments in human stem cell differenatiation to endoderm lineage.

overall_design: Measuring replication timing, transcription and chromatin organization using Hi-C at different time points during human stem cell differentiation to endoderm lineage.

first: Vishnu

last: Dileep

first: Korey

last: Wilson

first: Claire

last: Marchal

first: Xiaowen

last: Lyu

first: Peiyao

last: Zhao

first: Ben

last: Li

first: Axel

last: Poulet

first: Daniel

last: Bartlett

first: Juan

last: Rivera-Mulia

first: Zhaohui

last: Qin

first: Allan

last: Robins

first: Thomas

last: Schulz

first: Michael

last: Kulik

first: Rachel

last: McCord

first: Job

last: Dekker

first: Stephen

last: Dalton

first: Victor

last: Corces


title: Atlas of Subcellular RNA Localization Revealed by APEX-seq

city: Stanford

first: Furqan

last: Fazal

address: 269 Campus Drive, CCSR 2130

department: Genetics

institute: Stanford University

summary: We introduce APEX-seq, a method for RNA sequencing based on direct proximity labeling of RNA using the peroxidase enzyme APEX2. APEX-seq in nine distinct subcellular locales produced a nanometer-resolution spatial map of the human transcriptome as a resource, revealing extensive patterns of localization for diverse RNA classes and transcript isoforms. We uncover a radial organization of the nuclear transcriptome, which is gated at the inner surface of the nuclear pore for cytoplasmic export of processed transcripts. We identify two distinct pathways of messenger RNA localization to mitochondria, each associated with specific sets of transcripts for building complementary macromolecular machines within the organelle. APEX-seq should be widely applicable to many systems, enabling comprehensive investigations of the spatial transcriptome.

overall_design: APEX-seq libraries for both labeled targets and unlabeled controls were generated for the following constructs: (1) NLS (nucleus), (2) NIK (nucleolus), (3) LMA (nuclear lamina), (4) NucPore (nuclear pore), (5) NES (cytosol), (6) ERM (ER membrane cytosol facing), (7) KDEL (ER lumen), (8) OMM (Outer mitochondrial membrane), and (9) MITO (mitochondrial matrix). 4 libraries were prepared for each construct (2 biological replicates for labeled targets, 2 biological replicates for unlabeled controls). Furthermore, we generated drug-perturbation libraries for OMM and NES using the following treatments: (1) cycloheximide, (2) puromycin and (3) m-chlorophenylhydrazone (CCCP). In addition, nocodazole-perturbation libraries for OMM and NES were generated following drug treatment for: (1) 3 minutes, (2) 6 minutes, (3) 9 minutes, (4) 30 minutes and (5) 2 hours. We also generated nuclear and cytosol fractionation RNA-seq libraries (2 replicates each) following a protocol by Gagnon et al. (Nature Protocols, 2014).

first: Shuo

last: Han

first: Kevin

last: Parker

first: Pornchai

last: Kaewsapsak

first: Jin

last: Xu

first: Alistair

middle: N

last: Boettiger

first: Howard

middle: Y

last: Chang

first: Alice

last: Ting


title: Rapid Irreversible Transcriptional Reprogramming in Human Stem Cells Accompanied by Discordance between Replication Timing and Chromatin Compartment [Repli-Seq]

first: David

last: Gilbert

summary: This study demonstrates the relationship between replication timing, transcription and chromatin compartments in human stem cell differenatiation to endoderm lineage

overall_design: Measuring replication timing, transcription and chromatin organization using Hi-C at different time points during human stem cell differentiation to endoderm lineage

first: Vishnu

last: Dileep

first: Korey

last: Wilson

first: Claire

last: Marchal

first: Xiaowen

last: Lyu

first: Peiyao

last: Zhao

first: Ben

last: Li

first: Axel

last: Poulet

first: Daniel

last: Bartlett

first: Juan

last: Rivera-Mulia


title: Expression data from human lung squamous cell carcinoma tissues and the correponding non-cancerous tissues

city: Shanghai

first: ZHUOAN

last: CHENG

state: Shanghai

address: 2200 Nong, Xietu road

country: China

institute: Shanghai Jiaotong University

summary:
To identifythe the functional roles and the pathophysiological contributions of coding genes and noncoding RNAs in human lung squamous cell carcinoma, we analysed the differenial expression of genes and noncoding RNAs in 5 lung squamous cell carcinoma tissues and the corresponding non-cancerous tissues.
We used microarrays to detail the global programme of gene expression in human lung squamous cell carcinoma and correponding non-cancerous tissues.

overall_design: Total RNA from 5 pairs human lung squamous cell carcinoma tissues and correponding non-cancerous tissues were isolated and purified. Five micrograms of total RNA from each sample were used to synthesize double-stranded complementary DNA (cDNA), and then amplified and labeled with biotin and hybridized to the SBC human ceRNA array V1.0（4x180K, designed by Shanghai Biotechnology corporation, and made by Agilent technologies)

first: Zhuoan

last: Cheng

first: Chengtao

last: Yu

first: Shaohua

last: Cui


title: Rapid Irreversible Transcriptional Reprogramming in Human Stem Cells Accompanied by Discordance between Replication Timing and Chromatin Compartment

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


title: Rapid Irreversible Transcriptional Reprogramming in Human Stem Cells Accompanied by Discordance between Replication Timing and Chromatin Compartment [RNA-Seq]

first: Zhaohui

last: Qin

first: Allan

last: Robins

first: Thomas

last: Schulz

first: Michael

last: Kulik

first: Rachel

last: McCord

first: Job

last: Dekker

first: Stephen

last: Dalton

first: Victor

last: Corces


title: Identification and quantification of modified nucleosides in Saccharomyces cerevisiae mRNAs

city: Ann Arbor

first: mehmet

last: tardu

state: MI

address: 930 N University Ave

department: Chemistry

institute: University of Michigan

summary: Post-transcriptional modifications to messenger RNAs (mRNAs) have the potential to alter the biological function of this important class of biomolecules. The study of mRNA modifications is a rapidly emerging field, and the full complement of chemical modifications in mRNAs is not yet established. We sought to identify and quantify the modifications present in yeast mRNAs using an ultra-high performance liquid chromatography tandem mass spectrometry method to detect 40 nucleoside variations in parallel. We observe six modified nucleosides with high confidence in highly purified mRNA samples (N7-methylguanosine, N6-methyladenosine, 2’-O-methylguanosine, 2’-O-methylcytidine, N4-acetylcytidine and 5-formylcytidine), and identify the yeast protein responsible for N4-acetylcytidine incorporation in mRNAs, Rra1. Additionally, we find that mRNA modification levels change in response to heat shock, glucose starvation and/or oxidative stress. This work expands the repertoire of potential chemical modifications in mRNAs, and highlights the value of integrating mass spectrometry tools in the mRNA modification discovery and characterization pipeline.

overall_design: Total RNA and poly(A) RNA transcriptome profiles of exponentially growing (OD600 = 0.6) Saccharomyces cerevisiae cells were generated by deep sequencing , in two biological replicates, using Illumina HiSeq 4000 platform.

first: Mehmet

last: Tardu

first: Joshua

last: Jones

first: Robert

last: Kennedy

first: Qishan

last: Lin

first: Kristin

last: Koutmou


title: ERBB2 Regulates MED24 during Cancer Progression in Mice with Pten and Smad4 Deletion in the Pulmonary Epithelium

city: Haining

first: Jian

last: Liu

state: Zhejiang

address: 718 East Haizhou Rd.

department: Zhejiang University - University of Edinburgh Institute

institute: Zhejiang University

summary: ERBB2 is an oncogenic driver with frequent gene mutations and amplification in human lung tumors and is an attractive target for lung cancer therapy. However, target therapies can be improved by understanding the in vivo mechanisms regulated by ERBB2 during lung tumor development. Here, we generated genetic mouse models to show that Erbb2 loss inhibited lung tumor development induced by deletion of Pten and Smad4. Transcriptome analysis showed that Erbb2 loss suppressed the significant changes of most of the induced genes by ablation of Pten and Smad4. Overlapping with ERBB2-associated human lung cancer genes further identified those ERBB2 downstream players potentially conserved in human and mouse lung tumors. Furthermore, MED24 was identified as a crucial oncogenic target of ERBB2 in lung tumor development. Taken together, ERBB2 is required for the dysregulation of cancer-related genes, such as MED24, during lung tumor development.

overall_design: We will compare gene expression levels in lung tissues among the following transgenic mice that are with and without tumors:       Ptenf/fSmad4f/fErbb2f/f – 3 lung tissues (Wild type)  CCSPiCrePtenf/fSmad4f/f -- 3 mouse lungs (with tumors)    CCSPiCrePtenf/fSmad4f/fErbb2f/f -- 3 mouse lungs (tumor inhibited)

first: Francesco

last: DeMayo


title: RBPMS knockdown and overexpression in rat PAC1 pulmonary artery smooth muscle cells (SMCs)

status: Public on Jun 18 2019

city: Cambridge

first: Christopher

middle: WJ

last: Smith

address: Tennis Court Road

department: Department of Biochemistry

country: United Kingdom

institute: University of Cambridge

summary: Investigation of RBPMS role in post-transcriptional control of mRNAs in rat PAC1 pulmonary artery smooth muscle cells (SMCs). PolyA mRNA-Seq was carried out after RBPMS knockdown in differentiated PAC1 cells and after inducible RBPMS-A overexpression in dedifferentiated (proliferative) PAC1 cells.

overall_design: mRNA-Seq of RBPMS knockdown: Control siRNA and RBPMS-specific siRNA knockdown; and overexpression: RBPMS-A inducible lentiviral population without and with Dox induction. All experiments were conducted in triplicates.

first: Erick

last: Nakagaki-Silva

first: Clare

last: Gooding

first: Miriam

last: Llorian

first: Chris


title: Choroid plexus tertiary lymphoid structures in lupus: a novel neuro-immune interface

status: Public on Jun 17 2019

city: Bronx

first: Evan

last: Der

address: 1300 Morris Park Avenue

department: Rheumatology

institute: Albert Einstein College of Medicine

summary: The central nervous system manifestations of systemic lupus erythematosus (SLE) remain poorly understood, confounding physician treatment options and leaving patients with little options beyond empirical remedies. We utilized a well-established model of both the systemic and neuropsychiatric manifestations of SLE in an attempt to further understand which immune effectors may be responsible for central nervous system (CNS) disease. We found that a mixed cellular infiltrate in the choroid plexus of these mice represented a tertiary lymphoid structure with in-situ somatic hypermutation and class switch recombination. We further found that this process was driven by cytokine signaling sourced within the CNS that could be abrogated by treatment with biological therapeutics. Therefore, inconsistencies in previous work to determine which neuropathic autoantibodies are involved in the progression of neuropsychiatric lupus may have been lacking, as the source of these autoantibodies are intra-cranial, with limited systemic distribution.

overall_design: Choroid plexus mRNA profiles of 12 week old MRL/lpr (6 samples) and MRL/+ mice (3 samples) and 16 week old MRL/lpr (6 samples) and MRL/+ (3 samples) sequenced with HiSeq2500.

first: Ariel

last: Stock

first: Sivan

last: Gelb

first: Ayal

last: Ben-Zvi

first: Chaim

last: Putterman


title: RNA-seq analysis of rat smooth muscle cells


title: MicroRNA-130a Contributes to Type-2 Classical DC-activation in Sjögren’s Syndrome by Targeting MSK-1

city: Utrecht

first: Ana

middle: Pinheiro

last: Lopes

state: Utrecht

address: Heidelberglaan 100

country: Netherlands

institute: University Medical Center Utrecht

summary: Objectives: Considering the critical role of microRNAs (miRNAs) in regulation of cell activation, we investigated their role in circulating type-2 conventional dendritic cells (cDC2s) of patients with primary Sjögren’s syndrome (pSS) compared to healthy controls (HC).  Methods: CD1c-expressing cDC2s were isolated from peripheral blood. A discovery cohort (15 pSS, 6 HC) was used to screen the expression of 758 miRNAs and a replication cohort (15 pSS, 11 HC) was used to confirm differential expression of 18 identified targets. Novel targets for two replicated miRNAs were identified by SILAC in HEK-293T cells and validated in primary cDC2s. Differences in cytokine production between pSS and HC cDC2s were evaluated by intracellular flow-cytometry. cDC2s were cultured in the presence of MSK1-inhibitors to investigate their effect on cytokine production.  Results: Expression of miR-130a and miR-708 was significantly decreased in cDC2s from pSS patients compared to HC in both cohorts, and both miRNAs were downregulated upon stimulation via endosomal TLRs. Upstream mediator of cytokine production MSK1 was identified as a novel target of miR-130a and overexpression of miR-130a reduced MSK1 expression in cDC2s. pSS cDC2s showed higher MSK1 expression and an increased fraction of IL-12 and TNF-a-producing cells. MSK1-inhibition reduced cDC2 activation and production of IL-12, TNF-a, and IL-6.   Conclusions: The decreased expression of miR-130a and miR-708 in pSS cDC2s seems to reflect cell activation. miR-130a targets MSK1, which regulates pro-inflammatory cytokine production, and we provide proof-of-concept for MSK1-inhibition as a therapeutic avenue to impede cDC2 activity in pSS.

overall_design: RNA was extracted from peripheral blood isolated CD1c-expressing cDC2s from 21 donors; 6 healthy (HC) and 15 primary Sjögren’s syndrome (pSS) patients

first: Joel

last: van Roon

first: Timothy

last: Radstake

first: Maarten

last: Hillen


title: RNAseq analysis of primary differentiated rat aorta tissue compared to proliferative cultured cells

summary: Investigation of alternative splicing changes in rat smooth muscle aorta differentiated tissue compared to dedifferentiated proliferative aorta smooth muscle cells in culture.

overall_design: mRNA-Seq of rat aorta tissue compared to single cell aorta, passage 0 and passage 9.

middle: W

first: Adrian

last: Buckroyd

first: Sushma

last: Nagaraja Grellscheid


title: Effects of cadmium and integrator complex knockdown on gene expression and mRNA splicing

city: Saskatoon

first: Cheng-Wei

last: Wu

state: SK

address: 52 Campus Drive

country: Canada

institute: University of Saskatchewan

summary: RNA undergoes numerous processing steps required for gene expression.  Little is known about RNA homeostasis during environmental stress in metazoan cells.  By studying regulation of a heavy metal-induced gene named numr-1 in C. elegans, we discovered that disruption of RNA processing acts as a signal for environmental stress.  We find that NUMR-1 contains motifs common to RNA splicing factors and regulates RNA splicing in vivo.  A genome-wide screen reveals that numr-1 is strongly and specifically induced by silencing of genes that function in basal RNA metabolism including subunits of the metazoan integrator complex.  Human integrator processes snRNAs for functioning with splicing factors, and we find that silencing of C. elegans integrator subunits disrupts snRNA processing and causes aberrant pre-mRNA splicing.  Cadmium, which also strongly induces numr-1, has the same effects on snRNA processing and pre-mRNA splicing.  Lastly, we find that heat shock factor-1 is required for numr-1 induction by cadmium.  Our results are consistent with a model in which disruption of integrator processing of RNA acts as a molecular damage signal initiating an adaptive stress response mediated by heat shock factor-1.

overall_design:
We report the use of mRNA sequencing to analyze the effects of cadmium exposure and integrator complex gene knockdown have on the transcriptome.
Total RNA was prepared from gravid young adult Caenorhabditis elegans with three biological replicates per group and sent to NovoGene for PE150 sequencing. Sequencing library were constructured with oligo-dT enrichment. Gene expression data were analyzed by DEGSeq with Alternative splicing data analyzed by rMATS.

first: C

first: K

last: Choe

last: Wimberly


title: Molecular comparison of imatinib-naïve and resistant gastrointestinal stromal tumors: differentially expressed microRNAs and mRNAs

status: Public on Jun 12 2019

city: Rotterdam

first: Erik

last: Wiemer

address: Wytemaweg 80

department: Medical Oncology

institute: Erasmus University Medical Center

summary: Despite the success of imatinib in advanced gastrointestinal stromal tumor (GIST) patients, 50% of the patients experience resistance within two years of treatment underscoring the need to get better insight into the mechanisms conferring imatinib resistance. Here the microRNA and mRNA expression profiles in primary (imatinib-naïve) and imatinib-resistant GIST were examined. Fifty-three GIST samples harboring primary KIT mutations (exon 9; n=11/exon 11; n=41/exon 17; n=1) and comprising imatinib-naïve (IM-n) (n=33) and imatinib-resistant (IM-r) (n=20) tumors, were analyzed. The microRNA expression profiles were determined and from a subset (IM-n, n=14; IM-r, n=15) the mRNA expression profile was established. Ingenuity pathway analyses were used to unravel biochemical pathways and gene networks in IM-r GIST. Thirty-five differentially expressed miRNAs between IM-n and IM-r GIST samples were identified. Additionally, miRNAs distinguished IM-r samples with and without secondary KIT mutations. Furthermore 352 aberrantly expressed genes were found in IM-r samples. Pathway and network analyses revealed an association of differentially expressed genes with cell cycle progression and cellular proliferation thereby implicating genes and pathways involved in imatinib resistance in GIST. Differentially expressed miRNAs and mRNAs between IM-n and IM-r GIST were identified. Bioinformatic analyses provided insight into the genes and biochemical pathways involved in imatinib-resistance and highlighted key genes that may be putative treatment targets.

overall_design: The mRNA expression profile was determined using the Affymetrix HG-U133_Plus_2 platform of 14 imatinib-naïve (IM-n) and 15 imatinib-resistant (IM-r) gastrointestinal stromal tumors

first: Azadeh

last: Amirnasr

first: Caroline

last: Gits

first: Patricia

middle: F

last: van Kuijk

first: Marcel

last: Smid

first: Anne

middle: L

last: Vriends

first: Piotr

last: Rutkowski

first: Raf

last: Sciot

first: Patrick

last: Schöffski

first: Maria

last: Debiec-Rychter

first: Stefan

last: Sleijfer

middle: A


title: Epigenetic signature of PD-1+ TCF1+ CD8 T cells thatact as resource cells during chronic viral infectionand respond to PD-1 blockade

status: Public on Jun 15 2019

city: Bethesda

first: Peng

last: Li

address: 9000 Rockville Pike

department: NHLBI

institute: NIH

summary: We have recently defined a novel population of PD-1+TCF1+virus-specific CD8 T cells that function as resource cells during chronic LCMV infection and provide the proliferative burst seen after PD-1 blockade. Such CD8 T cells have been found in other chronic infections and also in cancer in mice and humans. These CD8 T cells exhibit stem-like properties undergoing self-renewal and also giving rise to the terminally differentiated (exhausted) CD8 T cells. Here we compared the epigenetic signature of stem-like CD8 T cells with exhausted CD8 T cells. ATAC-seq analysis showed that stem-like CD8 T cells had a unique signature implicating activity of TCF (HMG family) and NF-κB (RHD family) members. In addition, transcription factor networks including Tcf7, Id3, and Bach2 were enriched in stem-like CD8 T cells. In contrast,exhausted CD8 T cells enriched accessible sites for ETS and Runx motifs in addition to a network encompassing Prdm1 and Il10. We also compared the epigenetic signatures of the two CD8 T-cellsubsets from chronically infected mice with effector and memory CD8 T cells generated after an acute LCMV infection. Both CD8T-cell subsets generated during chronic infection were strikingly different from CD8 T-cell subsets from acute infection. Interestingly, the stem-like CD8 T-cell subset from chronic infection, despite sharing key functional properties with memory CD8 T cells,had a very distinct epigenetic program. These results show that the chronic stem-like CD8 T-cell program represents a specific adaptation of the T-cell response to persistent antigenic stimulation.

overall_design: RNA-seq analysis of CXCR5-Tim-3+ and CXCR5+Tim-3- CD8+ T-cell subsets, PD-1+CXCR5-Tim-3+CD8+ and PD-1+CXCR5+Tim-3-CD8+ cells (> 3x10e5 cells each) were sorted from pooled splenocytes of chronically LCMV infected mice (> day 40 p.i., n=14-15)

first: Rohit

last: Jadhav

first: Se

last: Im

first: Bin

last: Hu

first: Masao

last: Hashimoto

first: Jian-Xin

last: Lin

first: Warren

last: Leonard

first: William

last: Greenleaf

first: Rafi

last: Ahmed

first: Jorg

last: Goronzy


title: Genome-wide maps of H3K79me and H2Bub states

status: Public on Jun 13 2019

city: Amsterdam

first: Fred

last: van Leeuwen

address: Plesmanlaan 121

department: Gene Regulation

institute: Netherlands Cancer Institute

summary: We compare patterns of H3K79me and H2Bub across genes in budding yeast

overall_design: Generate profiles of H3K79me1, H3K79me3 and H2BK123ub1, canonical histones H2B and H3 are used as controls. Strains: wild-type (WT) and rpd3 knock-out (KO).

first: Hanneke

last: Vlaming

first: Tessy

last: Korthout


title: Gene expression of donor monocytes recovered from various tissues of recipient mice one day after transfer

city: Aachen

first: Oliver

last: Pabst

address: Pauwelsstrasse 30

department: Institute of Molecular Medicine

institute: RWTH Aachen University

summary: Monocyte are recruited to almost all peripheral tissues. Micro-environmental stimuli appeared to alter monocyte transcription once they enter the intestine and their subsequent development in the tissue. We therefore investigated whether there is tissue specific monocyte adaption and differentiation in vivo by examining donor monocytes in various tissues i.e. small intestine, liver, spleen and BM.

overall_design: Transferred monocytes recovered and FACS-sorted from the small intenstine, liver, spleen and BM of recipient mice at different time points and used for gene expression analysis. Following sorting, cDNA was made immediately and preamplified with Multiplexed target enrichment (MTE) primers. Hybridization of pre-amplified DNA with target specific oligonucleotide probe pairs were performed using a nanostring technology.

first: Girmay

last: Desalegn


title: Anti-tumor effects of antimicrobial peptides, targets of the innate immune system, against hematopoietic tumors in Drosophila mxc mutants

city: Fukuoka

first: Tetsuya

last: Sato

address: 3-1-1, Maidashi Higashi-ku

country: Japan

institute: Kyushu University

summary: The innate immune response is the first line of defence against microbial infections. In Drosophila, two major immune pathways induce the synthesis of antimicrobial peptides (AMPs) in the fat body. Recently, it has been reported that certain cationic AMPs exhibit selective cytotoxicity against human cancer cells. However, not much is known about their anti-tumour effects. Drosophila mxc(mbn1) mutants exhibit malignant hyperplasia in a larval haematopoietic organ called the lymph gland (LG). Here, using RNA-Seq analysis, we found that many immunoresponsive genes, including AMP genes, were up-regulated in the mutants. Down-regulation of these pathways by either a Toll or an imd mutation enhanced the tumour phenotype of the mxc mutants. Conversely, ectopic expression of each of five different AMPs in the fat body significantly suppressed the LG hyperplasia phenotype in the mutants. Thus, we propose that the Drosophila innate immune system can suppress the progression of haematopoietic tumours by inducing AMP gene expression. Overexpression of any one of these five AMPs resulted in enhanced apoptosis in the mutant LGs, while no apoptotic signals were detected in the controls. We observed that two AMPs, Drosomycin and Defensin, were taken up by circulating haemocyte-like cells, which were associated with the LG regions showing reduced cell-to-cell adhesion in the mutants. On the other hand, another AMP, Diptericin, was directly localised at the tumour site without intermediating haemocytes. These results indicate that AMPs have a specific cytotoxic effect that enhance apoptosis exclusively in the tumour cells.

overall_design: mRNA profiles of 3rd instar larvae wild type  (Canton S), the lymph gland tumor phenotye (mbn1) and a hypomorphic allele not displaying any tumor phenotype (G43) were generated by deep sequencing using illumina sequencer.

first: Mayo

last: Araki

first: Massanori

last: Kurihara

first: Suzuko

last: Kinoshita

first: Rie

last: Awane

first: Yasuyuki

last: Ohkawa

first: Yoshihiro

middle: H

last: Inoue


title: Transcriptomic analysis of isolated zebrafish podocytes and endothelial cells exposed to human APOL1 mRNA

status: Public on Jun 16 2019

city: Durham

first: Allison

last: Ashley-Koch

state: North Carolina

address: Duke Box 104775, 300 N Duke Street

department: Duke Molecular Physiology Institute

institute: Duke University Medical Center

summary: We injected zebrafish embryos with human APOL1 mRNA, dissociated embryos at 4 days of age, and purified podocytes and endothelial cells with fluorescence activated cell-sorting. After sorting, mRNA was extracted from purified cells and used to generate cDNA libraries, which were sequenced on an Illumina HiSeq 2500.

overall_design: Examination of two cell types exposed to one of 3 mRNA treatments (sham, APOL1 G0, APOL1 G2)

first: Joseph

last: Bundy

first: Blair

last: Anderson

first: Lumila

last: Francescatoo

first: Melanie

middle: E

last: Garrett

first: Karen

last: Soldano

first: Marilyn

last: Telen

first: Erica

last: Davis


title: Landscapes of gene translation in hepatocellular carcinoma tumors revealed by ribosome profiling

status: Public on Jun 14 2019

city: Beijing

first: Xuerui

last: Yang

address: Haidian District

department: School of Life Sciences

institute: Tsinghua University

summary: Despite the critical role of translation in the multi-level gene expression regulation program that controls the protein abundance, a high-resolution view on the landscapes of gene translation in cancer has been missing. We used the technique of ribosome profiling to dissect the translatomes of human liver tumors and their adjacent normal tissues. Our results, as a novel resource, revealed genome-wide abnormalities of translation efficiencies in tumors and context-dependent non-canonical ORFs.

overall_design: We profiled the paired tumor and adjacent normal tissue samples from 10 patients of HCC by the procedure of ribosome profiling, which was adapted and optimized for processing of clinical solid tissue samples. Total RNA sequencing was performed in parallel for the same samples.

first: Qin

last: Zou

first: Rongyao

last: Huang

first: Zhengtao

last: Xiao


title: Transcriptional dynamics of pluripotent stem cell derived endothelial cell differentiation revealed by single cell RNA-Seq.

status: Public on Jun 10 2019

city: Edinburgh

first: Ian

last: McCracken

address: 47 Little France Crescent

department: Centre for Cardiovascular Science

institute: University of Edinburgh

summary: Aims: Pluripotent stem cell-derived endothelial cell products possess therapeutic potential in ischemic vascular disease. However, the factors that drive endothelial differentiation from pluripotency and cellular specification are largely unknown. The aims of this study were to use single cell RNA sequencing (scRNA-seq) to map the transcriptional landscape and cellular dynamics of directed differentiation of human embryonic stem cell-derived endothelial cells (hESC-EC), and to compare these cells to mature endothelial cells from diverse vascular beds.Methods and Results: A highly efficient directed 8-day differentiation protocol was used to generate a hESC-derived endothelial cell product (hESC-ECP), in which 66% of cells coexpressed CD31 and CD144. We observed largely homogeneous hESC and mesodermal populations at days 0 and 4 respectively, followed by a rapid emergence of distinct endothelial and mesenchymal populations. Pseudotime trajectory identified transcriptional signatures of endothelial commitment and maturation during the differentiation process. Concordance in transcriptional signatures was verified by scRNA-seq analysis using both a second hESC line RC11, and an alternative hESC-EC differentiation protocol. In total 105,727 cells were subjected to scRNA-seq analysis. Global transcriptional comparison revealed a transcriptional architecture of hESC-EC that differs from freshly isolated and cultured human endothelial cells and from organ-specific endothelial cells. Conclusions: A transcriptional bifurcation into endothelial and mesenchymal lineages was identified, as well as novel transcriptional signatures underpinning commitment and maturation. The transcriptional architecture of hESC-ECP was distinct from mature and foetal human EC.

overall_design: Characterisation of the differentiation of human embryonic stem cell derived endothelial cells using 2 different hESC lines and 2 differentiaton protocols.

first: Richard

middle: S

last: Taylor

first: Fatma

middle: O

last: Kok

first: Fernando

last: De La Cuesta

first: Ross

last: Dobie

first: Beth

last: Henderson

first: Joanne

middle: C

last: Mountford

first: Axelle

last: Caudrillier

first: Neil

first: Chris

last: Ponting

first: Andrew

last: Baker


title: Transcriptional regulation of the glucose-6-phosphate transporter 2

city: East Lansing

first: Kevin

last: Childs

address: 612 Wilson Rd, Room S108, Plant Biology Laboratories

institute: Michigan State University

summary: Arabidopsis Col-0 and WS ecotypes were subjected to a high light treatment and leaf tissues were harvested 0, 15, 30, 60, 120, or 240 after exposure to high light. RNA was isolated and sequenced in order to determine gene expression changes after an increase in light. The transcript for GPT2 transiently increases several hundred-fold within two hours, and the increase in GPT2 transcript is preceded by an increase in expression of many transcription factor genes including Redox Responsive Transcription Factor 1 (RRTF1). It is hypothesized that transient GPT2 expression is adaptive, allowing G6P to move into the chloroplast from the cytosol. The imported G6P can be used for starch synthesis or may flow directly into the Calvin-Benson cycle via an alternative pathway (the G6P shunt),

overall_design: SRP200873; PRJNA548115; Two ecotypes sampled, six time points after start of high light treatment, three biological replicates per sample

first: Sean

last: Weise

first: Tiffany

last: Liu

first: Alyssa

last: Preiser

first: Hailey

middle: M

last: Katulski

first: Christopher

last: Perrin-Porzondek

first: Thomas

middle: D

last: Sharkey


title: Prediction of bacterial infection outcome using single cell RNA-seq analysis of human immune cells [scRNA-seq ind. 2]

status: Public on Jun 07 2019

city: Rehovot

first: Noa

last: Bossel Ben-Moshe

address: Hertzl Street

country: Israel

institute: Weizmann

summary: During host-pathogen encounters, the complex interactions between different immune cell-types can determine the outcome of infection. Advances in single cell RNA-seq (scRNA-seq) allow to probe this complexity of immunity, and afforded the basis for deconvolution algorithms that infer cell-type compositions from bulk RNA-seq measurements. However, immune activation, an important aspect of immune surveillance, is not represented in current algorithms. Here, using scRNA-seq of human peripheral blood cells infected with Salmonella, we developed a novel deconvolution algorithm to infer dynamic immune states from bulk measurements. We applied our dynamic deconvolution algorithm both to cohorts of healthy individuals challenged ex vivo with Salmonella and to cohorts of tuberculosis patients during different stages of disease. We revealed cell-type specific immune responses associated not only with ex vivo infection phenotype but also with clinical disease stage. We propose that our approach provides a predictive power to identify risk for disease, and can be applied to comprehensively study human infection outcome.

overall_design: Frozen PBMCs from healthy individual were defrosted and infectd ex vivo with Salmonella enterica serovar Typhimurium.

first: Shelly

last: Hen-Avivi

first: Natalia

last: Levitin

first: Dror

last: Yehezkel

first: Marije

last: Oosting

first: Leo

last: Joosten

first: Mihai

middle: G

last: Netea

first: Roi

last: Avraham


title: Expression in donor monocyte derived cells

first: Oliver

last: Pabst

summary: Monocyte differentiate into macrophages that display either pro-inflammatory or tolerogenic profile depending on the inflammatory condition of the intestine. We performed adoptive monocyte transfer to dissect the dynamics of monocyte to macropahges differentiation. Gene expression analysis of donor monocyte-derived cells recovered from non-inflamed and inflamed small intestine of recipient mice at day 1, 2 and 5 after transfer for in-depth investigation of monocyte development in the intestine.

overall_design: Transferred monocytes recovered and FACS-sorted from the small intenstine of recipient mice at different time points and used for gene expression analysis. Following sorting, cDNA was made immediately and preamplified with Multiplexed target enrichment (MTE) primers. Hybridization of pre-amplified DNA with target specific oligonucleotide probe pairs were performed using a nanostring technology.

first: Girmay

last: Desalegn


title: Single-cell analysis of adult human ovary using 10X genomics

city: Leiden

first: Susana

last: Chuva de Sousa Lopes

address: Einthovenweg 20

department: Anatomy and Embryology

institute: Leiden University Medical Center

summary: The ovary is perhaps the most dynamic organ in the human body, only rivaled by the uterus. The molecular mechanisms that regulate follicular growth and regression, ensuring ovarian tissue homeostasis, remain elusive. We have performed single-cell RNA-sequencing using human adult ovaries to provide a map of the molecular signature of growing and regressing follicular populations. We have identified different types of granulosa and theca cells and detected local production of components of the complement system by (atretic) theca cells and stromal cells. We also have detected a mixture of adaptive and innate immune cells, as well as several types of endothelial and smooth muscle cells to aid the remodeling process. Our results highlight the relevance of mapping whole adult organs at the single-cell level and reflect ongoing efforts to map the human body. The association between complement system and follicular remodeling may provide key insights in reproductive biology and (in)fertility.

overall_design: 31 tissue samples from 5 ovaries were used for single-cell and cluster analysis

first: S

first: I

last: Moustakas

first: M

last: Bialecka


title: Prediction of bacterial infection outcome using single cell RNA-seq analysis of human immune cells [scRNA-seq ind.1]

overall_design: Frozen PBMCs from healthy individual were defrosted and infected ex vivo with Salmonella enterica serovar Typhimurium.


title: B cell HVEM interaction with T cell BTLA restrains the help delivered to germinal center B cells

city: San Francisco

first: Michelle

middle: Amy

last: Mintz

state: CA

address: 513 Parnassus Ave

department: Microbiology and Immunology

institute: University of California San Francisco

summary: Gene expression of HVEM-deficient versus WT germinal center (GC) B cells.

overall_design: mRNA profiles from HVEM-deficient (CD45.2) and WT (CD45.1/2) mixed bone marrow chimeras analyzed at day 11 after NP-CGG alum immunization were generated using a HiSeq Illumina 4000.

first: Jason

last: Cyster


title: Transcriptomic profile of circulating memory T cells can differentiate between latent tuberculosis individuals and healthy controls

city: La Jolla

first: Bjoern

last: Peters

address: 9420 Athena Cir

institute: La Jolla Institute for Immunology

summary: Tuberculosis (TB) is responsible for the majority of mortality and morbidity associated with infectious diseases worldwide. The characterization of exact molecular components of immune response associated with protection against TB may help design more effective therapeutic interventions. In this study, we aimed to characterize the immune signature of memory T cells associated with latent infection with Mycobacterium tuberculosis. Transcriptomic profiling using RNA sequencing was performed on memory CD4 and CD8 T cells isolated from individuals with latent tuberculosis, as well as from tuberculosis negative healthy controls. Overall, we found specific gene signatures in each cell subset that could successfully discriminate between individuals with latent tuberculosis and healthy controls.

overall_design: RNA-sequencing of sorted memory CD4 and CD8 T cells from cryopreserved PBMC of 10 subjects with latent tuberculosis infection and 10 tuberculosis negative healthy controls

first: Julie

last: Burel

first: Cecilia

last: Lindenstam

first: Gregory

last: Seumois

first: Zheng

last: Fu

last: Greenbaum

first: Alessandro

last: Sette

first: Pandurangan

last: Vijayanand


title: Impact of genome architecture upon the functional activation and repression of Hox regulatory landscapes [4C-seq]

city: Geneva

first: Eddie

last: Rodríguez-Carballo

address: 4, Boulevard d'Yvoy

department: Department of Genetics and Evolution

country: Switzerland

institute: Université de Genève

summary: The spatial organization of the mammalian genome is complex and relies upon the formation of chromatin domains of various scales. At the level of gene regulation in cis, collections of enhancer sequences define large regulatory landscapes that usually match with the presence of topologically associating domains (TADs). These domains are largely determined by bound CTCF molecules and often contain ranges of enhancers displaying similar or related tissue specificity, suggesting that in some cases such domains may act as coherent regulatory units, with a global on or off state. By using the HoxD gene cluster as a paradigm, we investigated the effect of large genomic rearrangements affecting the two TADs flanking this locus, including their fusion into a single chromatin domain. We show that, within a single hybrid TAD, the activation of both proximal and distal limb enhancers initially positioned in either TADs globally occurred as when both TADs are intact. We also show that the timely implementation of distal limb enhancers depends on whether or not target genes had previously responded to proximal enhancers, due to the presence or absence of H3K27me3 marks. From this work, we conclude that antagonistic limb proximal and distal enhancers can exert their specificities when positioned into the same TAD and in the absence of their genuine target genes. We also conclude that removing these target genes reduced the coverage of a regulatory landscape by chromatin marks associated with silencing and thus prolonged its activity in time. Since Polycomb group proteins are mainly recruited at the Hox gene cluster, our results suggest that Polycomb Repressive Complex 2 (PRC2) can extend its coverage to far-cis regulatory sequences as long as confined to the neighboring TAD structure.

overall_design: 4C-seq analysis of Hoxd4 and CS38 3D interactions in proximal and distal limbs from E12.5  HoxD-inverted allele

first: E

last: Rodriguez-Carballo

first: L

last: Lopez-Delisle

first: D

last: Duboule


title: The stringent response inhibits DNA replication initiation in E. coli by modulating supercoiling of oriC

status: Public on Jun 11 2019

city: Cambridge

first: James

last: Kraemer

address: 31 Ames St, Building 68-570

institute: Massachusetts Institute of Technology

summary: The stringent response enables bacteria to respond to a variety of environmental stresses, especially various forms of nutrient limitation. During the stringent response the cell produces large quantities of the nucleotide alarmone ppGpp, which modulates many aspects of cell physiology, including reprogramming transcription, blocking protein translation, and inhibiting new rounds of DNA replication. The mechanism by which ppGpp inhibits DNA replication initiation in Escherichia coli remains unclear. Prior work suggested that ppGpp blocks new rounds of DNA replication by inhibiting transcription of the essential initiation factor dnaA, but we found that replication is still inhibited by ppGpp in cells ectopically producing DnaA. Instead, we provide evidence that a global reduction of transcription by ppGpp prevents replication initiation by modulating the supercoiling state of the origin of replication, oriC. Active transcription normally introduces negative supercoils into oriC to help promote replication initiation, so the accumulation of ppGpp reduces initiation potential at oriC. We find that maintaining transcription near oriC, either by expressing a ppGpp-blind RNA polymerase mutant or by inducing transcription from a ppGpp-insensitive promoter, can strongly bypass the inhibition of replication by ppGpp. Additionally, we show that increasing global negative supercoiling by inhibiting topoisomerase I or by deleting the nucleoid-associated protein seqA, also relieves inhibition. We propose a model, potentially conserved across proteobacteria, in which ppGpp indirectly creates an unfavorable energy landscape for initiation by preventing the introduction of negative supercoils into oriC.

overall_design: RNA sequencing experiments from wild-type and RNAP1-2- cells before and 15 minutes after induction of RelA' to produce ppGpp.

first: Michael

middle: T

last: Laub


title: Prediction of bacterial infection outcome using single cell RNA-seq analysis of human immune cells [WT/TLR10 bulk RNA-seq]

first: Noa

last: Bossel Ben-Moshe

overall_design: PBMCs were isolated from 8 individuals bearing or not TLR10 polymorphism and were infected ex vivo with Salmonella enterica serovar Typhimurium. RNA was extracted before infection, 4 hours post infection and 8 hours post infection.

first: Shelly

last: Hen-Avivi

first: Natalia

last: Levitin

first: Dror

last: Yehezkel

first: Marije

last: Oosting

first: Leo

last: Joosten

first: Mihai

last: Netea

first: Roi

last: Avraham


title: Gain of 20q11.21 in human pluripotent stem cells impairs TGFβ-dependent ectodermal commitment

city: Brussels

first: Claudia

last: Spits

address: Laarbeeklaan 103

department: Research Group of Reproduction and Genetics

country: Belgium

institute: Vrije Universiteit Brussel

summary: Altered TGFb signaling in lines with a 20q11.21 amplification has a dramatic negative effect on ectodermal differentiation

overall_design: Control lines versus lines with a 20q11.21 amplification and Bcl-xL overexpressing line

first: Christina

last: Markouli

first: Edouard

last: Couvreu De Deckersberg


title: Pioneer and repressive functions of p63 during embryonic ectoderm specification

status: Public on Jun 06 2019

city: Sevilla

first: Juan

middle: J.

last: Tena

address: Univ. Pablo Olavide, Ctra. Utrera km1

country: Spain

institute: CABD/CSIC

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

overall_design: Refer to individual Series


title: Impact of genome architecture upon the functional activation and repression of Hox regulatory landscapes [RNA-seq]

first: Eddie

last: Rodríguez-Carballo

overall_design: RNA-seq analysis of wild-type and mutant (delHoxD(attP-Rel5)d9lac and invHoxD(attP-Itga6)) in proximal and distal limbs from E12.5 mouse embryos.

first: E

last: Rodriguez-Carballo


title: Gene expression profile during senescence induced by LSG1 knockdown

middle: Carlos

last: Acosta

address: Crewe Rd South

department: Cancer Research UK - Edinburgh Centre

institute: The University of Edinburgh

summary: In the present study, we analyze the effect of knocking down LSG1 and KRas(V12D) overexpression in MRC5 cells  in the transcriptome using Ampliseq RNA sequencig. We observed that shLSG1 induced a potent senescence response that is characterized by the activation of ER-Stress and cholesterol biosynthetic pathway

overall_design: MRC5  were transfected with siRNA to knockdown the small GTPase LSG1. Total mRNA was extracted and expression profiles were analyzed.

first: Asimina

last: Pantazi

first: Priya

last: Hari

first: Andrew

middle: J

last: Finch


title: Motor dysfunction and neurodegeneration in a C9orf72 mouse line expressing poly-PR

status: Public on Jun 04 2019

city: Suzhou

first: Zongbing

last: Hao

address: 199 Ren-ai Road

institute: Soochow University

summary: GFP-PR28 homozygous mice show decreased survival time, while the heterozygous mice show motor imbalance, decreased brain weight, loss of Purkinje cells and lower motor neurons, and inflammation in the cerebellum and spinal cord. Transcriptional analysis shows that in the cerebellum, GFP-PR28 heterozygous mice show differential expression of genes related to synaptic transmission.

overall_design: The mRNA profiles of cerebellum, cortex and spinal cord of control and heterozygous mice were generated by RNA sequencing, in triplicate, using Illumina HiSeq X Ten

first: Liu

last: Liu

first: Zhouteng

last: Tao

first: Rui

last: Wang

first: Haigang

last: Ren

first: Hongyang

last: Sun

first: Zixuan

last: Lin

first: Zhixiong

last: Zhang

first: Chenchen

last: Mu

first: Jiawei

last: Zhou

first: Guanghui


title: Prediction of bacterial infection outcome using single cell RNA-seq analysis of human immune cells [sorted population Bulk RNA-seq]

first: Noa

last: Bossel Ben-Moshe

overall_design: PBMCs were isolated from a healthy individual and were infected ex vivo with Salmonella enterica serovar Typhimurium or with PBS as control. Monocytes and NKT cells were sorted from naïve and infected PBMCs. RNA was extracted 4 hours post infection.

middle: B

last: Ben-Moshe

first: Shelly

last: Hen-Avivi

first: Natalia

last: Levitin

first: Dror

last: Yehezkel

first: Marije

last: Oosting

first: Leo

last: Joosten

first: Mihai

last: Netea

first: Roi

last: Avraham


title: Prediction of bacterial infection outcome using single cell RNA-seq analysis of human immune cells [WB/PBMCs bulk RNA-seq]

overall_design: Whole-blood (WB) cells and PBMCs were isolated from 4 healthy individuals and were infected ex vivo with Salmonella enterica serovar Typhimurium or with PBS as control. RNA was extracted 4 hours later.


title: CATACOMB: an endogenous inducible gene that antagonizes H3K27 methylation activity of Polycomb Repressive complex 2 via a H3K27M-like mechanism

status: Public on Jun 03 2019

city: Chicago

first: Ali

last: Shilatifard

state: IL

address: 320 E Superior St

department: Department of Biochemistry and Molecular Genetics

institute: Northwestern University Feinberg School of Medicine

type: Methylation profiling by high throughput sequencing

summary: Dysregulation of Polycomb Repressive Complex 2 (PRC2) function is a common feature of many cancer types, including both solid and hematological malignancies. A number of chromosomal translocations involving Polycomb group proteins have been identified, however, the molecular function of these fusion proteins remains largely unexplored. Here we characterize two endometrial stromal sarcoma (ESS) associated fusion proteins: JAZF1-SUZ12 and MBTD1- CXORF67. We identify JAZF1 as a previously unreported subunit of the NuA4 complex, which when fused to SUZ12 creates a bridge between the NuA4 and PRC2 complexes. Intriguingly, the MBTD1-CXORF67 fusion binds to PRC2 and leads to strongly reduced levels of the PRC2 catalytic products H3K27me2/3. We demonstrate that this inhibitory function is mediated by the CXORF67 half of the fusion protein. Because of this striking property, we propose a new gene name: CATACOMB (CATalytic Antagosnist of polyCOMB, official gene name: EZHIP). We map CATACOMB’s inhibitory function to a short highly conserved region and identify a single methionine residue essential for diminution of H3K27me2/3 levels. Remarkably, the amino acid sequence surrounding this critical methionine resembles that of the oncogenic histone H3 lysine-27-to-methionine (H3K27M) mutation found in high grade pediatric gliomas. Finally, we show CATACOMB expression is silenced through DNA methylation and upon treatment with DNA demethylating agents, CATACOMB is expressed, binds to PRC2 and antagonizes its catalytic activity. In conclusion, we have identified an endogenous inducible gene, CATACOMB, that it’s expression can regulate catalytic activity of PRC2 and propose that such mechanism of regulation of histone modifications through the expression of antagonistic subunits may also exist for Trithorax/COMPASS family and other histone modifying enzyme complexes.

overall_design: Bisulfite sequencing and RNA-seq in HEK293T and HCT-116  cells treated with5-azacytidine or 5-aza-2-deoxyazacytidine. RNA-seq of U2OS cells treated with  EPZ-6438.

first: Benjamin

last: Singer

first: Andrea

last: Piunti

first: Edwin

last: Smith


title: Prediction of bacterial infection outcome using single cell RNA-seq analysis of human immune cells


title: Genomic Signatures Reveal DNA Damage Response Deficiency in Brain Metastases of Colorectal Cancer

city: Nanjing

first: Cheng

last: Wang

address: No. 818 East Tianyuan Road

institute: Nanjing Medicial University

summary: We performed RNA-Seq on primary colorectal cancer (CRC) tissues and brain metastatic tissues from two CRC patients. Differential expression analysis was conducted to find genes associated with brain metastasis.

overall_design:
mRNA profiles of primary tissues and metastatic tissues were generated by deep sequencing using Illumina HiSeq platform.
*** Raw sequencing data for all the samples will be available at the European Genome-phenome Archive (http://www.ebi.ac.uk/ega/).

first: Jing

last: Sun

first: Yanhong

last: Gu


title: Scc2 counteracts a Wapl-independent mechanism that releases cohesin from chromosomes during G1

city: Oxford

first: Kim

last: Nasmyth

address: South Parks Road

department: Biochemistry

institute: University of Oxford

summary: Cohesin’s association with chromosomes is determined by loading dependent on the Scc2/4 complex and release due to Wapl. We show here that Scc2 also actively maintains cohesin on chromosomes during G1. It does so by blocking a Wapl-independent release reaction that requires opening the cohesin ring at its Smc3/Scc1 interface as well as the D loop of Smc1’s ATPase. The Wapl-independent release mechanism is switched off as cells activate Cdk1 and enter G2/M and cannot be turned back on without cohesin’s dissociation from chromosomes. The latter phenomenon enabled us to show that in the absence of release mechanisms, cohesin rings that have already captured DNA in a Scc2-dependent manner before replication no longer require Scc2 to capture sister DNAs during S phase.

overall_design: Characterisation of the effect of WT and mutant cohesin complexes and associated factors on loading onto chromosomes in S. cerevisiae, assessed by calibrated ChIP-seq on an Ion Torrent Proton (Life Technologies)

first: Naomi

last: Petela


title: Endodermal maternal transcription factors establish super enhancers during zygotic genome activation

city: Irvine

first: Kitt

middle: D.

last: Paraiso

address: 4410 Nat Sci II, University of California, Irvine

department: Department of Developmental and Cell Biology

institute: University of California, Irvine

summary: We examined the combinatorial function of the Xenopus tropicalis maternal transcription factors Foxh1, Vegt and Otx1. The study involved combinatorial ectopic expression of vegt and otx1 RNA in the putative ectoderm; and combinatorial knock-down of vegt and otx1 in the putative endoderm both followed by transcriptomic analysis (RNA-seq). We performed chromatin immunoprecipitation combined with high throughput sequencing (ChIP-seq) to assess the chromatin association of Vegt and Otx1 prior to zygotic gene activation. In addition, we performed ChIP-seq on H3K4me1 on endoderm tissue to identify active regulatory regions in an unbiased manner.

overall_design:
Screen for animally and vegetally expressed transcription factors: In biological triplicates, the animal and vegetal blastomeres of 8-cell stage embryos were separated. Subsequently, RNA was harvested from both set of blastomeres and whole embryos at the same stage and prepared for RNA sequencing. Combinatorial ectopic expression experiment: In biological duplicates, titrating levels of vegt and otx1 RNA were combinatorially microinjected into 1-cell stage embryos. Animal caps (putative ectoderm) were microdissected at stage 9 (6 hpf) from uninjected and injected samples, and RNA was harvested at stage 10.5 (7 hpf) for RNA sequencing. Double morpholino experiment: In biological duplicates, Vegt and Otx1 morpholinos were injected in 1-cell stage embryos independently or together. Vegetal masses (putative endoderm) were microdissected at stage 9 (6 hpf) and RNA was harvested from uninjected and injected samples at stage 10.5 (7 hpf) for RNA sequencing. Vegt and Otx1 ChIP-seq analysis: Developmentally synchronized embryos were harvested at stage 8 (prior to zygotic gene activation) and fixed with formaldehyde in biological duplicates. Chromatin immunoprecipitation as performed using anti-Vegt or anti-Otx1 antibody and DNA was purified for sequencing. H3K4me1 ChIP-seq analysis: Developmentally synchronized embryos were harvested at stage 10.5 and fixed with formaldehyde in biological duplicates. The vegetal tissue was disssected from the fixed embryos, and ChIP-seq was performed using anti-H3K4me1 antibody.

Previously published Foxh1 ChIP-seq from GSE85273 was re-analyzed along with this study.

GSM2263590 and GSM2263598 used to generate:
GSE118024_foxh1_IDR_optimal_peaks.bed.gz

GSM2263598 used to generate:
GSE118024_otx1_IDR_optimal_peaks.bed.gz, GSE118024_vegt_IDR_optimal_peaks.bed.gz

first: Ken

middle: W

last: Cho


title: Pioneer and repressive functions of p63 during embryonic ectoderm specification [ChIP-Seq]

first: Juan

last: Tena

summary: The transcription factor p63 is a master regulator of ectoderm development essential for epidermal specification. Although previous studies have highlighted the role of p63 triggering the epidermal transcriptomic program, its precise mechanism of target gene regulation in the complex context of a developing embryo remains poorly understood. Here, we used zebrafish embryos to analyze in vivo how p63 regulates the expression of its target genes during development. We generated tp63-knock-out mutants that recapitulate human phenotypes and show down-regulated epidermal gene expression. Following p63-binding dynamics during development, we found two distinct functions clearly separated in space and time. During early development, p63 binds enhancers associated to neural genes, where it limits Sox3 binding and reduces the expression of these neural genes. Indeed, we show that p63 and Sox3 are co-expressed in the neural plate border. Later in development, p63 binds enhancers associated to epidermal genes and promotes their expression, acting as a pioneer factor, as it binds to non-accessible chromatin and is required for its opening. Therefore, our results suggest that p63 is an important regulator of cell fate decisions during ectoderm specification, promoting the epidermal fate and inhibiting the neural program.

overall_design: ChIP-seq (ChIPmentation)  assays in tp63 zebrafish mutants

first: José

last: Santos-Pereira

first: Lourdes

last: Gallardo-Fuentes

first: Ana

last: Neto

first: Rafael

last: Acemel


title: Pioneer and repressive functions of p63 during embryonic ectoderm specification [RNA-seq]

overall_design: RNA-seq assays in tp63 zebrafish mutants


title: Pioneer and repressive functions of p63 during embryonic ectoderm specification [ATAC-seq]

overall_design: ATAC-seq assays in tp63 zebrafish mutants


title: RNA sequencing of Mus musculus thymic lymphomas

city: Paris

first: Ludovic

last: Deriano

address: 25-28 Rue du Docteur Roux

department: Immunology

country: France

institute: Institut Pasteur

overall_design: We performed transcriptome sequencing of T-cell lymphomas originating from p53-deficient mice lacking RAG1/2 activity (Rag2 knock out, Rag2-/- p53-/-) or expressing wild type RAG1/2 (p53-/-). We sequenced five different mice from each genotype and we also sequenced thymus from three wild type mice.

first: Joy

last: Bianchi

first: Valentine

last: Murigneux

first: Marie

last: Bedora-Faure

first: Chloé

last: Lescale


title: OGT knockout S2VP10 cells

status: Public on Jun 01 2019

city: Miami

first: Nikita

middle: Satish

last: Sharma

state: FL

address: 1550 NW 10th Avenue PAP 134

department: Surgery

institute: University of Miami

summary: Pancreatic Cancer cells were transfected with CRISR based OGT knockouts

overall_design: Examination of genes affected by OGT knockout


title: Identification of Host Biomarkers of EBV Latency IIb and Latency III

city: Durham

first: Micah

middle: Alan

last: Luftig

state: NC

address: 213 Research Drive CARL Building Room 0045

department: Molecular Genetics and Microbiology

institute: Duke University

summary: Deciphering the molecular pathogenesis of virally induced cancers is challenging due, in part, to the heterogeneity of both viral and host gene expression. Epstein-Barr Virus (EBV) is a ubiquitous herpesvirus prevalent in B-cell lymphomas of the immune suppressed. EBV infection of primary human B cells leads to their immortalization into lymphoblastoid cell lines (LCLs) serving as a model of these lymphomas. In previous studies, our lab has described a temporal model for immortalization with an initial phase characterized by expression of the Epstein-Barr Nuclear Antigens (EBNAs), high c-Myc activity, and hyper-proliferation in the absence of the Latent Membrane Proteins (LMPs), called latency IIb. This is followed by the long-term outgrowth of LCLs expressing the EBNAs along with the LMPs, particularly the NFkB-activating LMP1, defining latency III. LCLs, however, express a broad distribution of LMP1 such that a subset of these cells expresses LMP1 at levels seen in latency IIb, making it difficult to distinguish these two latency states. In this study, we performed mRNA-Seq on early EBV-infected latency IIb cells and latency III LCLs sorted by NFkB activity. We found that latency IIb transcriptomes clustered independently from latency III independent of NFkB. We identified and validated mRNAs defining these latency states. Indeed, we were able to distinguish latency IIb cells from LCLs expressing low levels of LMP1 using multiplex RNA-FISH targeting EBV EBNA2, LMP1, and human CCR7 or MGST1. This study defines latency IIb as a bona fide latency state independent from latency III and identifies biomarkers for understanding EBV-associated tumor heterogeneity

sample_organism:
Homo sapiens
human gammaherpesvirus 4

platform_organism:
Homo sapiens
human gammaherpesvirus 4

first: Joshua

last: Messinger

first: Joanne

last: Dai

first: Lyla

last: Stanland

first: Alexander

last: Price

middle: A


title: Identifying the transcriptional targets of the Drosophila transcription factor Ets21c via Targeted DamID

city: Cologne

first: Mirka

last: Uhlirova

state: NRW

address: Joseph-Stelzmann Str. 26

department: Institute for Genetics

institute: CECAD Research Center  Cologne

summary: Tissue homeostasis relies on stem cell activity which drives replacement of differentiated cells to maintain tissue integrity and function. The stress-inducible transcription factor Ets21c promotes tissue renewal in the Drosophila adult intestine by controlling intestinal stem cell proliferation and apoptosis of mature enterocytes. To determine how Ets21c fulfills these distinct tasks in the different cell types of the gut, we exploited a genome-wide in vivo Targeted DamID mapping approach to identify its putative direct targets. In young unstressed midguts, Ets21c contacts regulatory loci occupied by as well as devoid of the Polymerase II binding. Follow-up functional genetic studies in intestinal stem cells and enterocytes demonstrate that Ets21c controls cell type-specific sets of target genes that orchestrate cellular behavior via intrinsic and non-autonomous signaling mechanisms. Ets21c thus emerges as a vital determinant of adult health and lifespan.

overall_design: Low-level expression of UAS-LT3-Dam (noise control), UAS-LT3-Dam-PolII (transcribed genes) and UAS-LT3-Dam-ets21c-RA (transcription factor of interest) in Drosophila adults using the temperature sensitive esg-Gal4[TS] (intestinal stem cells and enteroblasts) and Myo1A-Gal4[TS] (enterocytes) TARGET systems. Three biological replicates of dissected midguts for each genotype followed by methylation-specific PCR amplification and single-end sequencing.

first: Juliane

last: Mundorf

first: Colin

last: Donohoe

last: McClure

first: Tony

last: Southall


title: DOT1L inhibition reveals a distinct subset of enhancers dependent on H3K79 methylation (RNA-seq)

status: Public on May 30 2019

first: Thomas

last: Milne

address: John Radcliffe Hospital, Headington

department: MRC Weatherall Institute of Molecular Medicine

summary: Enhancer elements are a key regulatory feature of many important genes. Several general features including the presence of specific histone modifications are used to demarcate potentially active enhancers. Here we reveal that putative enhancers marked with H3 lysine 79 (K79) di or trimethylation (me2/3) (which we name H3K79me2/3 enhancer elements or KEEs) can be found in multiple cell types. Mixed lineage leukemia gene (MLL) rearrangements (MLL-r) such as MLL-AF4 are a major cause of incurable acute lymphoblastic leukemias (ALL). Using the DOT1L inhibitor EPZ-5676 in MLL-AF4 leukemia cells, we show that H3K79me2/3 is required for maintaining chromatin accessibility, histone acetylation and transcription factor binding specifically at KEEs but not non-KEE enhancers. We go on to show that H3K79me2/3 is essential for maintaining enhancer-promoter interactions at a subset of KEEs. Together, these data implicate H3K79me2/3 as having a functional role at a subset of active enhancers in MLL-AF4 leukemia cells.

overall_design: RNA-seq

first: Laura

last: Godfrey

first: Nicholas

last: Crump

first: I-Jun

last: Lau


title: Inhibition of Phosphoinositide-3-kinase signaling promotes the stem cell state of trophoblast

city: Singapore

first: Cheryl

last: Lee

address: 8 College Road

institute: Duke-NUS Medical School

summary: We found that PI3K inhibition increased the expression of stem cell markers in trophoblast stem cells (TSCs). To better understand the PI3K inhibited cells, we compared untreated TSCs with cells treated with PI3K inhibitor ZSTK474 for 3h, 6h and 3 days.

overall_design: Untreated TSCs, TSCs treated with 200nM ZSTK474 for 3h, 6h, and 3 days.

first: Myriam

last: Hemberger

first: Janet

last: Rossant


title: Small RNA-seq of Ago1 and Ago1x immunoprecipitated samples

status: Public on May 31 2019

city: Bangalore

first: Sandeep

last: Eswarappa

state: Karnataka

address: Lab FA08, Dr. Sandeep M Eswarappa, Department of Biochemistry, School of Biological Sciences

country: India

institute: Indian Institite of Science

summary:
Purpose: To identify small RNAs interacting with Ago1x
Methods: FLAG-HA tagged Ago proteins were immunoprecipitated and associated miRNAs were isolated using miRNeasy Mini kit (Qiagen)
Results: Interaction of Ago1x with miRNAs is comparable to that Ago1

overall_design: Small RNA seq was done for RNA isolated from immunoprecipitated samples

first: Anumeha

last: Singh