ArrayExpress Uploader04087noneCytosol_ExtractnoneCytosol_ExtractnoneCytosol_Extractanti-FLAG M2IPanti-FLAG M2IPanti-FLAG M2IPnoneWhole_Cell_Lysateanti-FLAG M2Whole_Cell_Lysate_IParrayexpress_sid4Although EWS/FLI-1 fusion protein is responsible for most Ewing’s sarcoma family tumors (ESFT), the function of native EWS remains...22241085E-GEOD-29313/profile/8773/arrayexpressuploader28cellgenomeprolineproteinsarcomaDec.12, 2014Falserna-immunoprecipitation-rip-chip-analysis-for-ewsE-GEOD-29313_A-AFFY-44RNA immunoprecipitation (RIP)-Chip analysis for EWS-bound mRNASep.16, 2014Although 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).http://www.ebi.ac.uk/arrayexpress/experiments/E-GEOD-29313humanhttp://www.ebi.ac.uk/arrayexpress/experiments/E-GEOD-29313/samples/