Transcriptional repression of the insulin-like growth factor I receptor (IGF-I-R) gene by the tumor suppressor WT1 involves binding to sequences both upstream and downstream of the IGF-I-R gene transcription start site

The insulin-like growth factor-I receptor (IGF-I-R) has been implicated in the etiology and/or progression of Wilms' tumor, a pediatric malignancy of the kidney that is often associated with deletion or mutation of the WT1 tumor suppressor gene. The expression of the IGF-I-R gene is increased in Wilms' tumor as compared with normal kidney tissue. Furthermore, the levels of IGF-I-R mRNA in individual tumors have been shown to be inversely correlated to the levels of WT1 mRNA, suggesting that the expression of the IGF-I-R gene is under the negative control of WT1. The activity of an IGF-I-R promoter/luciferase construct in Chinese hamster ovary cells was reduced by cotransfection of a WT1 expression vector. An analysis of various reporter constructs containing different portions of the IGF-I-R 5′-flanking and 5′-untranslated regions suggested that the effect of WT1 depends on the number of WT1 binding sites present, with sites located both upstream and downstream of the IGF-I-R transcription start site involved in mediating this effect. Using the purified zinc finger domain of WT1 in gel retardation and DNase I footprinting assays, we mapped five sites in the 5′-flanking and six sites in the 5′-untranslated regions that were involved in WT1 binding. In addition, the initiator element of the IGF-I-R gene contains a sequence that binds WT1. Thus, the repression of IGF-I-R promoter activity by the WT1 tumor suppressor gene product involves multiple interactions of its zinc finger domain with WT1 binding sites located both 5′ and 3′ of the transcription initiation site.