Crystal Structure of a Smad MH1 Domain Bound to DNA Insights on DNA Binding in TGF-β Signaling

cific cytokine to a pair of specific transmembrane recep-* Department of Molecular Biology tors, leading to the activation of the Ser/Thr kinase in Princeton University the cytoplasmic domain of these receptors (Derynck, Lewis Thomas Laboratory 1994; Massagué and Weis-Garcia, 1996). The signal is Princeton, New Jersey 08544 transferred to the Smad proteins through the receptor † Cell Biology Program kinase–mediated phosphorylation of pathway-specific The signal is then propagated primarily through protein– Summary protein interactions between Smad proteins, which are homooligomeric, and between Smads and transcrip-The Smad family of proteins, which are frequently tar-tion factors. The phosphorylated Smad (1) heterooligo-geted by tumorigenic mutations in cancer, mediate merizes with the ubiquitous Smad4 (Lagna et al., 1996; TGF-␤ signaling from cell membrane to nucleus. The Zhang et al., 1996), (2) translocates into the nucleus crystal structure of a Smad3 MH1 domain bound to an associates with sequence-specific reveals a novel DNA-binding motif. In the crystals, DNA-binding protein(s) (Chen et al., 1996), resulting in base-specific DNA recognition is provided exclusively the activation of agonist-responsive genes. by a conserved 11-residue ␤ hairpin that is embedded The Smad proteins, about 400–500 amino acids each, in the major groove of DNA. A surface loop region, to consist of two conserved domains with pairwise se-which tumorigenic mutations map, has been identi-quence identity of 40%–94% for the N-terminal MH1 fied as a functional surface important for Smad activ-domain and 38%–98% for the C-terminal MH2 domain. ity. This structure establishes a framework for under-The MH2 domain is responsible for transactivation and standing how Smad proteins may act in concert with homo-and heterooligomerization, whereas the MH1 do-other transcription factors in the regulation of TGF-␤-main exhibits sequence-specific DNA binding activity responsive genes. and negatively regulates the functions of MH2 domain Introduction The DNA binding activity of Smad MH1 has been shown to be necessary for proper activation of agonist-respon-The TGF-␤/BMP/activin superfamily of cytokines plays sive genes (Kim et al., 1997; Dennler et al., 1998), and a central role in regulating a broad range of cellular mutation of Smad-binding sequences led to the abolish-responses, including cell growth, differentiation, and ment of TGF-␤ responsiveness in PAI-1 promoter (Denn-specification of developmental fate, in diverse organ-ler et al., 1998). isms from C. elegans to humans (Roberts and Sporn, Investigation of transcriptional activation by Smads 1990; Hogan, 1996). The actions of these cytokines re-has revealed a complex pattern involving multiple pro-sult from their abilities to regulate the …