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E-GEOD-29313 - RNA immunoprecipitation (RIP)-Chip analysis for EWS-bound mRNA

Status
Released on 16 January 2012, last updated on 26 June 2012
Organism
Homo sapiens
Samples (8)
Array (1)
Protocols (8)
Description
Although EWS/FLI-1 fusion protein is responsible for most Ewing’s sarcoma family tumors (ESFT), the function of native EWS remains largely unknown. Here, we first showed that EWS repressed protein expression in a tethering assay. mRNAs bound to EWS were determined by RNA-immunoprecipitation Chip assay, and one of them, proline-rich Akt substrate of 40 kDa (PRAS40) mRNA, directly interacted with EWS. The inhibitor of AKT, API-2, repressed ESFT cell proliferation. We demonstrate that EWS negatively regulated PRAS40 protein expression through binding to PRAS40 3’UTR. Furthermore, PRAS40 knockdown inhibited the proliferation and metastatic potential of ESFT cells. Cytoplasmic lysates or whole cell lysates were prepared from HeLa S3 cells transfected with pFLAG-EWS , and incubated with anti-FLAG M2 Affinity Gel (Sigma) at 4°C for 2 h. RNAs from lysates and immunoprecipitates were analysed using GeneChip Human Genome U133 Plus 2.0 Array (Affymetrix).
Experiment type
transcription profiling by array 
Contacts
Iku Kuwahara, Ken Matsumoto, Lin Huang, Yuji Nakai
Citation
MIAME
PlatformsProtocolsVariablesProcessedRaw
Files
Investigation descriptionE-GEOD-29313.idf.txt
Sample and data relationshipE-GEOD-29313.sdrf.txt
Raw data (1)E-GEOD-29313.raw.1.zip
Processed data (1)E-GEOD-29313.processed.1.zip
Array designA-AFFY-44.adf.txt
R ExpressionSetE-GEOD-29313.eSet.r
Links