Loss of cell cycle control by deregulation of cyclin-dependent kinase 2 kinase activity in Evi-1 transformed fibroblasts

The Evi-1 transcriptional repressor protein has two distinct zinc finger DN A binding domains designated ZF1 and ZF2 and is implicated in the progressi on of human and murine leukemias, in which it is abnormally expressed. In t his report, we show that Evi-1-expressing Rat1 fibroblasts are anchorage in dependent, have an abbreviated G(1) phase of the cell cycle, and have a red uced requirement for serum mitogens for S-phase entry, These biological cha nges are accompanied by a moderately increased production of cell cycle-reg ulatory proteins cyclin A and cyclin-dependent kinase (Cdk) 2, a dramatic d eregulation of Cdk2 kinase activity, and a corresponding increase in the le vels of hyperphosphorylated retinoblastoma protein (pRb), We show that the elevated cyclin A-Cdk2 activity is due to the combination of increased accu mulation and stabilization of cyclin A bound to a faster-migrating species of Cdk2 believed to be the active threonine 160 phosphorylated form and a s ubstantial reduction in complexed p27, Cyclin E kinase activity is also ele vated due to a reduction in p27, A significant reduction in total cellular p27 protein levels and a moderate reduction in p27 mRNA are observed, but n o changes in Cdk regulatory kinases and phosphatases occur. The Evi-1 trans criptional repressor domain and the ZF1 DNA binding domain are required for both cell transformation and induction of Cdk2 catalytic activity. We prop ose that one consequence of Evi-1 expression is to repress the transcriptio n of target genes, which may include p27, that deregulate the normal contro l of the G(1) phase of the cell cycle, providing a cellular proliferative a dvantage that contributes to transformation in vitro and leukemogenesis in vivo.