THE EVI-1 PROTOONCOGENE ENCODES A TRANSCRIPTIONAL REPRESSOR ACTIVITY ASSOCIATED WITH TRANSFORMATION

The myeloid transforming gene Evi-1 encodes a protein with two zinc fi nger domains, designated ZF1 and ZF2, with distinct DNA binding specif icities. For the first time we demonstrate that Evi-1 has transcriptio nal repressor activity which is directly proportional to the amount of Evi-1 protein in cells. Repression has been observed with two distinc t promoters: the minimal HSV-1 tk promoter and a VP16 inducible adenov irus E1b minimal promoter. Optimal repression is DNA binding dependent and is mediated by either ZF1 or a heterologous GAL4 DNA binding doma in (GAL4DBD) but is significantly less efficient through the ZF2 bindi ng site. Both GAL4DBD/Evi-1 fusion and non-fusion proteins have been u sed to map the repressor activity to a proline-rich region located wit hin amino acids 514-724 between the ZF1 and ZF2 domains. Constitutive expression of mutant proteins lacking the repressor domain are defecti ve for transformation of Rat1 fibroblasts demonstrating that this regi on is required for the oncogenic activity of the Evi-1 protein. These studies show that the Evi-1 gene encodes a transcriptional repressor a nd has important implications for the mechanism of action of the Evi-1 protein both in development and in the progression of some myeloid le ukaemias.