THE EWS-ATF-1 GENE INVOLVED IN MALIGNANT-MELANOMA OF SOFT PARTS WITH T(12-22) CHROMOSOME-TRANSLOCATION, ENCODES A CONSTITUTIVE TRANSCRIPTIONAL ACTIVATOR

Molecular characterization of malignant melanoma of soft parts or soft tissue clear cell sarcoma which shares t(12;22) chromosome translocat ion revealed fusion of EWS with a transcriptional factor gene ATF-1. T he EWS gene, which encodes an RNA binding protein, was also shown to b e involved in Ewing sarcoma, related primitive neuroectodermal tumors and desmoplastic small round cell tumors. In order to understand the f unctional role of EWS-ATF-1 chimeric protein in human solid tumors, we have cloned the aberrant human ATF-1 (EWS-ATF-1) cDNA and studied its DNA binding, transcriptional activation properties and compared with normal ATF-1 protein. Our results demonstrate that EWS-ATF-1 binds wea kly to DNA in vitro but functions as an efficient constitutive transcr iptional activator unlike the normal ATF-1 which needs to be induced w ith cAMP. Deletion analysis revealed that EWS-fusion domain functions as a regulatory domain for the transcriptional activation properties o f EWS-ATF-1 chimeric protein. Deletion of leucine zipper domain result s in a loss of transcriptional activation of EWS-ATF-1 chimeric protei n suggesting that protein-protein interaction play a role in the trans criptional activation properties of EWS-ATF-1. We demonstrate that EWS -fusion domain negatively regulates the DNA binding activity of EWS-AT F-1 chimeric protein. Therefore replacement of part of the amino-termi nal kinase regulatory domain of ATF-1 protein with EWS regulatory doma in results in an altered DNA binding, protein-protein interactions and transcriptional activation properties of EWS-ATF-1 causing deregulate d gene expression which may be responsible for the genesis of t(12;22) chromosome translocation-bearing human solid tumors. Targeting the tr anscriptional cofactors (CBP, etc) by EWS-fusion proteins could be one of the mechanisms of activation of EWS-fusion proteins in human neopl asia.