A 2ND TRANSCRIPTIONALLY ACTIVE DNA-BINDING SITE FOR THE WILMS-TUMOR GENE-PRODUCT, WT1

The putative Wilms tumor suppressor gene, wt1, encodes a zinc-finger protein that binds to the DNA sequence 5'-GCGGGGGVG-3'. We previously reported that WT1 has separable domains that function either to activate or suppress transcription. We now have identified a second WT1 binding sequence (5'-TCCTCCTCCTCCTCTCC-3') 3' to the transcription initiation site of the platelet-derived growth factor A-chain gene by DNase I footprinting and gel mobility shift assays. WT1 requires both 5' and 3' binding sites for transcriptional suppression; however, WT1 functions as a transcriptional activator when it binds to either the 5' or 3' site alone. This second WT1 binding sequence functions equally well as the previously identified 5'-GCGGGGGCG-3' sequence when analyzed in transient transfection assays. A core DNA sequence recognized by WT1 was defined by using related synthetic oligonucleotides. We also identified sequences similar to the WT1 binding site within the promoter regions of five other growth-related genes and demonstrated that each of these sequences also binds WT1 in gel mobility shift assays. These results thus identify a second WT1 binding site and suggest that additional growth-related genes may be transcriptionally influenced by WT1.