Comment[ArrayExpressAccession]	E-MTAB-1662
MAGE-TAB Version	1.1
Investigation Title	Genomic analysis of SRC-2/TIF2 regulated genes in breast cancer cells_
Comment[Submitted Name]	Genomic analysis of SRC-2/TIF2 regulated genes in breast cancer cells_
Experiment Description	We have characterized the global program of transcription in SRC-2-depleted MCF-7 breast cancer cells using short-hairpin RNA technology, and in MCF-7 cells exposed to PKA activating agents. In order to identify genes that may be regulated through PKA-induced downregulation of SRC-2, overlapping transcriptional targets in response to the respective treatments were characterized. MCF-7 human breast cancer cells transduced with shRNA targeting SRC-2 (SRC-2 shRNA) or infected with control shRNA vector (Ctr shRNA)  were seeded in 9,2 cm petri dishes, 2,0 x106 cells in each dishes, in DMEM supplemented with 5% charcoaled stripped FBS and 10 nM 17b-estradiol. After 48 hours a selection of the Ctr shRNA cells were treated with cAMP analog (8-CPT-cAMP, 150 µM) and cAMP elevating agents (Forskolin (10 µM) and IBMX (50 µM)) The cells were then incubated for another 24 hour in the cell incubator. Cells were harvested by washing them twice with 8 ml PBS (37 oC), before adding 1,5 ml 0,25 % trypzine. The trypzine reaction was stopped after 5 minutes by adding 2,5 ml DMEM to the cells.  Trypzine and cell medium was removed by centrifugating the cell suspensions at 1000 rpm for 3 minutes. The cells were then washed with 400 µl PBS, before addition of 350 µl RLT buffer to the cell pellets. The cells were then homogenized by vortexing for 1 minute and then freezed in -80 oC.
Experimental Design	compound_treatment_design	cellular_modification_design	in_vitro_design	co-expression_design
Comment[AEExperimentType]	transcription profiling by array
Comment[AEExperimentDisplayName]	Transcription profiling by array of human SRC-2-depleted MCF-7 human breast cancer cells and MCF-7 cells exposed to PKA activating agents to identify genes that may be regulated through PKA-induced downregulation of SRC-2
Experimental Factor Name	RNA interference
Experimental Factor Type	RNA interference
Quality Control Type	spike-in quality control
Public Release Date	2013-05-15
Person Last Name	Helland	Fenne
Person First Name	Thomas	Ingvild
Person Mid Initials
Person Email	thms.helland@gmail.com	ingvild.fenne@k2.uib.no
Person Phone	4792847793	4797594732
Person Address	Haukeland University Hospital, 5021 Bergen, Norway	Haukeland University Hospital, 5021 Bergen, Norway
Person Affiliation	University of Bergen, Haukeland University Hospital	University of Bergen, Haukeland University Hospital
Person Roles	submitter	First author
PubMed ID
Publication Author List	Ingvild Sveinsgjerd Fenne, Thomas Helland, Marianne Hauglid Flågeng, Simon Nitter Dankel, Gunnar Mellgren, Jørn Vegard Sagen
Publication Title	Downregulation of Steroid Receptor Coactivator-2 Modulates Estrogen-responsive genes and Stimulates Proliferation of MCF-7 Breast Cancer Cells
Publication Status	Submitted
Protocol Name	P-MTAB-32745	P-MTAB-32746	P-MTAB-32747	P-MTAB-32748	P-MTAB-32749	P-MTAB-32750	P-MTAB-32751	P-MTAB-32752	P-MTAB-32753	P-MTAB-32754	P-MTAB-32755
Protocol Type	growth protocol	growth protocol	growth protocol	treatment protocol	treatment protocol	treatment protocol	nucleic acid extraction protocol	nucleic acid labeling protocol	nucleic acid hybridization to array protocol	array scanning protocol	normalization data transformation protocol
Protocol Description	MCF-7 human breast cancer cells transduced with shRNA targeting SRC-2 (SRC-2 shRNA) or infected with control shRNA vector (Ctr shRNA)  were seeded in 9,2 cm petri dishes, 2,0 x106 cells in each dishes, in DMEM supplemented with 5% charcoaled stripped FBS and 10 nM 17b-estradiol.	MCF-7 human breast cancer cells transduced with shRNA targeting SRC-2 (SRC-2 shRNA) were seeded in 9,2 cm petri dishes, 2,0 x106 cells in each dishes, in DMEM supplemented with 5% charcoaled stripped FBS and 10 nM 17b-estradiol.	MCF-7 human breast cancer cells transduced with control shRNA vector (Ctr shRNA)  were seeded in 9,2 cm petri dishes, 2,0 x106 cells in each dishes, in DMEM supplemented with 5% charcoaled stripped FBS and 10 nM 17b-estradiol. 	Cells were treated with 10 nM 17b-estradiol for 3 days	Cells were treated with 10 nM 17b-estradiol for 3 days	The cells were treated with 10 nM 17b-estradiol for 48. After 48 hours the Ctr shRNA cells were treated with cAMP analog (8-CPT-cAMP, 150 µM) and cAMP elevating agents (Forskolin (10 µM) and IBMX (50 µM)) to activate the cAMP/PKA signaling pathway. The cells were then incubated for another 24 hour in the cell incubator.	Total RNA from MCF-7 cells were extracted using RNeasy Mini kit (Qiagen, CA). http://www.genome.duke.edu/cores/microarray/services/rna-qc/documents/RNeasy_Mini_Handbook.pdf	Illumina TotalPrep Amplification Kit http://tools.invitrogen.com/content/sfs/manuals/IL1791ME.pdf	Whole - Genome Gene Expression Direct Hybridization Assay Guide, Illumina inc	iScan system/reader, Illumina	GenomeStudio Data Analysis Software (Illumina, Inc.) J-express 2009 (Molmine AS)
Protocol Term Source REF	EFO	EFO	EFO	EFO	EFO	EFO	EFO	EFO	EFO	EFO	EFO
Term Source Name	EFO
Term Source File	http://www.ebi.ac.uk/efo
SDRF File	E-MTAB-1662.sdrf.txt