Interaction of Smads with collagen types I, III, and V

Abstract

Ontogenesis of the mammalian orofacial region is controlled by numerous developmental signals, including those initiated by the transforming growth factors β (TGFβs). Targeted deletion of the genes encoding several of the TGFβs in mice has been shown to result in clefts of the secondary palate. Members of the TGFβ family of growth factors utilize intracellular Smads as signal transducers. Smads 2 and 3 are transcriptional regulators that bind DNA through their conserved MH1 domains and activate/inhibit transcription of TGFβ-responsive genes through their MH2 domains. Using a yeast two-hybrid screen of a cDNA expression library constructed from fetal murine orofacial tissue, we have identified three types of collagens (types I, III, and V) that are capable of binding to the MH2 domain of Smad 3. These interactions were confirmed by glutathione S-transferase (GST) pull-down assays in which the MH2 domain of Smad 3 fused to GST interacted strongly with in vitro translated, 35S-labeled collagen types I, III, and V. Each collagen also bound to the MH2 domains of Smads 4 and 7 and, to a lesser extent, full-length Smads 1, 2, 3, and 4. Binding of Smads to collagen is a novel observation. Moreover, TGFβ is a potent regulator of collagen synthesis and turnover during mammalian orofacial development. These data thus suggest an important means of feedback regulation of the TGFβ signaling cascade.