NOVEL REPLICATION INHIBITORY FUNCTION OF THE DEVELOPMENTAL REGULATOR TRANSCRIPTION REPRESSOR PROTEIN WT1 ENCODED BY THE WILMS-TUMOR GENE

The tumor suppressor/developmental regulator protein WT1 encoded by th e Wilms' tumor gene is a zinc finger-containing transcription factor w hich binds to the G + C-rich motif 5'-GCGGGGGCG-3' and represses trans cription. Alternatively spliced variants of WT1 (termed + KTS) having an insertion in the zinc finger region are defective for binding to an d hence for repression of transcription from promoters containing this motif. Due to the known interactions of two other tumor suppressor pr oteins with the simian virus 40 (SV40) oncoprotein large tumor antigen (TAg) [which in one case (p53) results in inhibition of the replicati on initiation activity of TAg], and because of the presence of G+C-ric h sequences in the SV40 origin region, we tested the effect of WT1 on TAg- and SV40 origin-dependent DNA replication. WT1 and its alternativ ely spliced variants were found to be potent inhibitors of replication . Inhibition of replication by WT1 required portions of the N-terminal transcription repression domain and the C-terminal DNA binding domain , while other WT1 sequences needed for transcriptional regulation were dispensable. WT1 neither inhibited the synthesis of TAg nor formed a stable complex with it. Studies of the requirement of cis-active origi n sequences in vivo and protein-DNA interactions in vitro indicated th at WT1 and its alternatively spliced variants might inhibit replicatio n by their novel binding to the GC box promoter motifs of the SV40 21 bp repeat replication-auxiliary sequence.