Comment[ArrayExpressAccession]	E-GEOD-42735			
MAGE-TAB Version	1.1			
Public Release Date	2014-04-20			
Investigation Title	"Genome-wide transcriptome profiling of SK-Mel-28, UACC-62 and HCT-116 cells stably expressing scrambled shRNA and RAB7 shRNA"			
Comment[Submitted Name]	"Genome-wide transcriptome profiling of SK-Mel-28, UACC-62 and HCT-116 cells stably expressing scrambled shRNA and RAB7 shRNA"			
Experiment Description	"Purpose: Assess the transcriptional changes induced upon RAB7 knock-down in melanoma (SK-Mel-28 and UACC-62) and in colon cancer (HCT-116) cell lines. Methods: mRNA profiles of tumor cell lines (SK-Mel-28, UACC-62, HCT-116) stably expressing scrambled shRNA or RAB7 shRNA (harvested at day 3 after lentiviral infection) were generated by deep sequencing, using three biological replicates per condition. The sequence reads that passed quality filters were analyzed with TopHat and Cufflinks. Validation of induced / silenced genes was performed by western blot. Results show a differential impact of RAB7 expression in the transcriptomic profile of melanoma vs non-melanoma cell lines, and support a lineage-specific role of this small GTPase in melanoma. Examination of the mRNA profiles RAB7-depleted vs wild type cells, performed in parallel in 3 different tumor cell lines (Melanomas: SK-Mel-28 and UACC-62, Non-melanoma: HCT-116) harvested at day 3 after lentiviral infection."			
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	Soengas	Soengas	Alonso-Curbelo	
Person First Name	Maria	M	D	
Person Mid Initials	S	S		
Person Email	msoengas@cnio.es			
Person Affiliation	Spanish National Cancer Research Institute			
Person Address	"Molecular Pathology, Spanish National Cancer Research Institute, Melchor Fernandez Almagro, 3, Madrid, Spain"			
Person Roles	submitter			
Protocol Name	P-GSE42735-4	P-GSE42735-1	P-GSE42735-3	P-GSE42735-2
Protocol Description	"Read files were quality-checked with FastQC. 40-nt single-end reads were aligned against the human genome (GRCh37/hg19) with TopHat-2.0.4 (using Bowtie 0.12.7 and Samtools 0.1.16), allowing 2 mismatches and 5 multihits. Transcripts quantification and differential expression calculated with Cufflinks 1.3.0 Transcripts annotations used here were Homo_sapiens.GRCh37.65 from Ensembl Genome_build: GRCh37/hg19 Supplementary_files_format_and_content: tables were generated with Cufflinks 1.3.0 and contain transcripts abundance measured in FPKM values"	"Lentivirus-driven gene silencing of RAB7 was performed using the validated shRNA targeting the sequence GATTGACCTCGAAA, purchased from Sigma (St Louis, MO, USA). Parallel infections with the pLKO vector coding for scrambled shRNA (Sigma) served as controls. The lentiviral infections of the 3 different tumor cell lines were performed in parallel."	"RNA was extracted from cell pellets using QIAshredder (Cat No. 79654) and Rneasy Mini Kit (Cat No. 74104) including DNase treatment step. 2 ug of total RNA samples was used as provided by the user. RNA Integrity Numbers were in the range 8.6 to 10 when assayed on an Agilent 2100 Bioanalyzer. PolyA+ RNA fraction was extracted and randomly fragmented, converted to double stranded cDNA and processed through subsequent enzymatic treatments of end-repair, dA-tailing, and ligation to adapters as in Illumina's “TruSeq RNA Sample Preparation v2 Protocol”; (Part # 15026494 Rev. C). Adapter-ligated library was completed by 8 cycles of PCR with Illumina PE primers. The resulting purified cDNA library was applied to an Illumina flow cell for cluster generation (TruSeq cluster generation kit v5) and sequenced on the Genome Analyzer IIx with SBS TruSeq v5 reagents by following manufacturer's protocols."	"Cells were cultured in Dulbecco's modified Eagle's medium (Life Technologies, Rockville, MD, USA) supplemented with 10% fetal bovine serum (Nova-Tech Inc., Grand Island, NY, USA)"
Protocol Type	normalization data transformation protocol	sample treatment protocol	nucleic acid library construction protocol	growth protocol
Experimental Factor Name	cell line	RNA interference		
Experimental Factor Type	cell line	RNA interference		
Publication Author List	Alonso-Curbelo D, Riveiro-Falkenbach E, Pérez-Guijarro E, Cifdaloz M, Karras P, Osterloh L, Megías D, Cañón E, Calvo TG, Olmeda D, Gómez-López G, Graña O, Sánchez-Arévalo Lobo VJ, Pisano DG, Wang HW, Ortiz-Romero P, Tormo D, Hoek K, Rodríguez-Peralto JL, Joyce JA, Soengas MS			
Publication Title	RAB7 controls melanoma progression by exploiting a lineage-specific wiring of the endolysosomal pathway.			
PubMed ID	24981740			
Publication DOI	10.1016/j.ccr.2014.04.030			
Comment[SecondaryAccession]	GSE42735			
Comment[GEOReleaseDate]	2014-04-20			
Comment[ArrayExpressSubmissionDate]	2012-12-04			
Comment[GEOLastUpdateDate]	2014-05-20			
Comment[AEExperimentType]	RNA-seq of coding RNA			
Comment[SecondaryAccession]	SRP017435			
Comment[SequenceDataURI]	http://www.ebi.ac.uk/ena/data/view/SRR628721-SRR628738			
SDRF File	E-GEOD-42735.sdrf.txt