Independent interaction of a mad-related protein homologue with rb and spl: a potential signal between TGFβI and TAFs

Stimulation of Spi-mediated transcription through the expression of Rb has been shown to transactivate a variety of genes associated with cell growth and differentiation through Spl binding elements, identified as retinoblastoma control elements (RCE). In addition to the genes encoding the proteins t.los, IGFII, and TGF-21, we show that a gene encoding the 6cyclin-like6 uracil-DNA glycosylase is a target of transactivation directed by Rb expression through Spl-binding elements. The link between Spl-mediated transcription and Rb has previously implicated a protein associated with the TFIII) complex, TAFII250. In an attempt to understand the potential of additional gene products associated with the activation of Spl-mediated transcription by Rb, we have used a modification of the yeast two-hybrid system to identify proteinprotein interactions, common to both Rb and Spl. Interactions with Spl were tested with the two-hybrid system to direct transcription through a RCE by expressing human cDNAs from pancreas fused to a GAL4 yeast expression vector. Results of these experiments identified candidate cDNAs related to the MAD family of gene products. Biochemical studies reveal that an interaction of a known MAD-related gene product, DPC4, with Rb is effected by either TGF31-induction or serine/threonine kinase activity. The interaction of Spl with DPC4 appears to be independent of either TGFβ stimulation, serine/threonine kinase activity, or zinc fingers of Spl. Further studies hope to provide information about the role of MAD-related gene products in directing gene expression.