Functional Cloning of the Proto-oncogene Brain Factor-1 (BF-1) As a Smad-binding Antagonist of Transforming Growth Factor-β Signaling

Abstract

Using the plasminogen activator inhibitor (PAI) promoter to drive the expression of a reporter gene (mouse CD2), we devised a system to clone negative regulators of the transforming growth factor-β (TGF-β) signaling pathway. We infected a TGF-β-responsive cell line (MvLu1) with a retroviral cDNA library, selecting by fluorescence-activated cell sorter single cells displaying low PAI promoter activity in response to TGF-β. Using this strategy we cloned the proto-oncogene brain factor-1 (BF-1). BF-1 represses the PAI promoter in part by associating with both unphosphorylated Smad3 (in the cytoplasm) and phosphorylated Smad3 (in the nucleus), thus preventing its binding to DNA. BF-1 also associates with Smad1, -2, and -4; the Smad MH2 domain binds to BF-1, and the C-terminal segment of BF-1 is uniquely and solely required for binding to Smads. Further, BF-1 represses another TGF-β-induced promoter (p15), it upregulates a TGF-β-repressed promoter (Cyclin A), and it reverses the growth arrest caused by TGF-β. Our results suggest that BF-1 is a general inhibitor of TGF-β signaling and as such may play a key role during brain development.