Targeted disruption of Smad3 reveals an essential role in transforming growth factor beta-mediated signal transduction

The Smads are a family of nine related proteins which function as signaling intermediates for the transforming growth factor beta (TGF-beta) superfami ly of ligands. To discern the in vivo functions of one of these Smads, Smad 3, we generated mice harboring a targeted disruption of this gene. Smad3 nu ll mice, although smaller than wild-type littermates, are viable, survive t o adulthood, and exhibit an early phenotype of forelimb malformation. To st udy the cellular functions of Smad3, we generated Smad3 null mouse embryoni c fibroblasts (MEFs) and dermal fibroblasts. We demonstrate that null MEFs have lost the ability to form Smad-containing DNA binding complexes and are unable to induce transcription from the TGF-beta-responsive promoter const ruct, p3TP-lux. Using the primary dermal fibroblasts, we also demonstrate t hat Smad3 is integral for induction of endogenous plasminogen activator inh ibitor 1. We subsequently demonstrate that Smad3 null MEFs are partially re sistant to TGF-beta's antiproliferative effect, thus firmly establishing a role for Smad3 in TGF-beta-mediated growth inhibition, We next examined cel ls in which Smad3 is most highly expressed, specifically cells of immune or igin. Although no specific developmental defect was detected in the immune system of the Smad3 null mice, a functional defect was observed in the abil ity of TGF-beta to inhibit the proliferation of splenocytes activated by sp ecific stimuli, In addition, primary splenocytes display defects in TGF-bet a-mediated repression of cytokine production. These data, taken together, e stablish a role for Smad3 in mediating the antiproliferative effects of TGF -beta and implicate Smad3 as a potential effector for TGF-beta in modulatin g immune system function.