The Ewing’s sarcoma (EWS) proto-oncogene can give rise to a variety of different tumors because of the generation of transforming EWS fusion proteins upon chromosomal translocation. However, the cellular function of the EWS protein itself was hitherto not established. We show that EWS is a nuclear protein, whose nuclear localization is dependent upon its transactivating NH₂ terminus. EWS COOH-terminal amino acids suppress this NH₂-terminal activation domain in the context of a Gal4 fusion protein, which may explain why none of the EWS fusion proteins in cancer cells contains the EWS COOH terminus. Furthermore, EWS expression enhances c-fos, Xvent-2, and ErbB2 promoter activity in a cell-type-dependent manner, indicating that EWS is a transcriptional regulator. Also, the EWS protein stimulates transcription mediated by the COOH-terminal transactivation domain of the cofactor CREB-binding protein (CBP). Coimmunoprecipitation experiments demonstrate that EWS forms a complex with CBP and the homologous p300 protein. A COOH-terminal region of EWS is both required for the physical interaction with CBP/p300 and sufficient to mediate c-fos activation. In addition, suppression of CBP/p300 function by the adenoviral E1A protein abolishes c-fos activation by EWS, indicating that EWS-mediated gene regulation depends on CBP/p300. In conclusion, the nuclear EWS proto-oncoprotein can function as a transcriptional cofactor in conjunction with CBP/p300.