Comment[ArrayExpressAccession] E-GEOD-68645 MAGE-TAB Version 1.1 Public Release Date 2015-06-24 Investigation Title The DAXX co-repressor is directly recruited to active regulatory elements genome-wide to regulate autophagy programs in a model of human prostate cancer (RNA-seq) Comment[Submitted Name] The DAXX co-repressor is directly recruited to active regulatory elements genome-wide to regulate autophagy programs in a model of human prostate cancer (RNA-seq) Experiment Description This study was aimed at understanding the genome-wide binding and regulatory role of the DAXX transcriptional repressor, recently implicated in PCa. ChIP-Seq analysis of genome-wide distribution of DAXX in PC3 cells revealed over 59,000 DAXX binding sites, found at regulatory enhancers and promoters. ChIP-Seq analysis of DNA methyltransferase 1 (DNMT1), which is a key epigenetic partner for DAXX repression, revealed that DNMT1 binding was restricted to a small number of DAXX sites. DNMT1 and DAXX bound close to transcriptional activator motifs. DNMT1 sites were found to be dependent on DAXX for recruitment by analyzing DNMT1 ChIP-Seq following DAXX knockdown (K/D), corroborating previous findings that DAXX recruits DNMT1 to repress its target genes. Massively parallel RNA sequencing (RNA-Seq) was used to compare the transcriptomes of WT and DAXX K/D PC3 cells. Genes induced by DAXX K/D included those involved in autophagy, and DAXX ChIP-Seq peaks were found close to the transcription start sites (TSS) of autophagy genes, implying they are more likely to be regulated by DAXX. Determine changes in gene expression levels between WT and DAXX K/D prostate cancer cells by RNA-Seq (PC3 Cells). Term Source Name ArrayExpress EFO Term Source File http://www.ebi.ac.uk/arrayexpress/ http://www.ebi.ac.uk/efo/efo.owl Person Last Name Benner Puto Benner Hunter Person First Name Christopher Lorena Christopher Tony Person Mid Initials A Person Email cbenner@salk.edu Person Affiliation Salk Institute Person Address Integrative Genomics and Bioinformatics Core, Salk Institute, 10010 North Torrey Pines Rd, La Jolla, CA, USA Person Roles submitter Protocol Name P-GSE68645-3 P-GSE68645-2 P-GSE68645-1 Protocol Description Illumina Casava v1.8.2 software used for basecalling. Reads were aligned to the human genome (hg19) with STAR (v2.2.0c) with default parameters. Only uniquely alignable reads were used for downstream analysis Reads Per Kilobase of exon per milion mapped reads were calculated using HOMER (http://homer.salk.edu/homer/) across annotated gene exons (RefSeq) Genome_build: hg19 Supplementary_files_format_and_content: Tab delimited text file, Columns: (1) RefSeq acc (2) chr (3) start (4) end (5) strand (6) length (7) copies (8) annotation (9-12) FPKM Total RNA was isolated using NucleoSpin RNA kit (Clontech, cat # 740955). The RNA integrity number (RIN) was determined using The Agilent RNA ScreenTape assay (Agilent 2200 TapeStation, Salk Institute’s Next Generation Sequencing core), and high quality RNA was used for RNA-Seq. The RNA-Seq libraries were prepared using Illumina TruSeq RNA v2 non-stranded sample prep kits and sequenced on an Illumina HiSeq 2500 according to the manufacturer’s instructions. Illumina HiSeq 2500 system and associated protocol was used. Run type: SE50; Run mode: High Output The human prostate cancer cell line PC3 and its DAXX knock-down counterpart were maintained in Roswell Park Memorial Institute-1640 (RPMI-1640) medium, containing 10% fetal bovine serum, FBS (HyClone), and 1% penicillin/streptomycin plus L-glutamine. For the DAXX K/D PC3 cells, the RPMI medium was supplemented with puromycin (Sigma, Cat # P9620; 2 μg/ml). Protocol Type normalization data transformation protocol nucleic acid library construction protocol growth protocol Experimental Factor Name genotype Experimental Factor Type genotype Publication Title The DAXX co-repressor is directly recruited to active regulatory elements genome-wide to regulate autophagy programs in a model of human prostate cancer. Publication Author List Puto LA, Benner C, Hunter T PubMed ID 26097870 Comment[SecondaryAccession] GSE68645 Comment[GEOReleaseDate] 2015-06-24 Comment[ArrayExpressSubmissionDate] 2015-05-07 Comment[GEOLastUpdateDate] 2015-06-24 Comment[AEExperimentType] RNA-seq of coding RNA Comment[AdditionalFile:Data1] GSE68645_fpkm.txt Comment[SecondaryAccession] SRP058098 Comment[SequenceDataURI] http://www.ebi.ac.uk/ena/data/view/SRR2013433-SRR2013436 SDRF File E-GEOD-68645.sdrf.txt