Smads: transcriptional activators of TGF-beta responses.

Based on the observations summarized above, the mechanism of transcriptional activation by Smads is most likely defined by the combined requirements for interactions with other transcription factors and with promoter DNA sequences. Consequently, Smad-responsive promoters have a double DNA sequence requirement. One sequence confers the specificity to bind the transcription factors that cooperate with the Smad complex. Another adjacent sequence is required for direct Smad binding and confers Smad selectivity to the first sequence. Thus, only a subset of promoter sequences that bind these cooperating transcription factors are targets for Smad signaling, and this selectivity is provided by flanking or partially overlapping sequences that allow Smad binding. For example, Smad3/4 cooperates with c-Jun to induce transcription from the collagenase I promoter in response to TGF-β; thus, the AP-1-binding site, which binds c-Jun/c-Fos, and the partially overlapping Smad-binding sequence AGAC mediate responsiveness to TGF-β and Smad3/4 (Yingling et al. 1997xYingling, J, Datto, M, Wong, C, Frederick, J, Liberati, N, and Wang, X.-F. Mol. Cell. Biol. 1997; 17: 7019–7028CrossrefSee all ReferencesYingling et al. 1997; Zhang et al. 1998xZhang, Y, Feng, X.-H, and Derynck, R. Nature. 1998; 394: 909–912Crossref | PubMed | Scopus (567)See all ReferencesZhang et al. 1998). However, the AP-1-binding site without a flanking GAC sequence does not allow inducibility by TGF-β or Smad3/4 (Dennler et al. 1998xDennler, S, Itoh, S, Vivien, D, ten Dijke, P, Huet, S, and Gauthier, J.-M. EMBO J. 1998; 11: 3091–3100Crossref | Scopus (1298)See all ReferencesDennler et al. 1998). Similarly, in the PAI-1 promoter, the TFE3-binding E-box sequence and the flanking sequence that binds the Smad3/4 complex are both required for TGF-β-induced transcription, which is mediated by the cooperation of Smad3/4 and TFE3 (Hua et al. 1998xHua, X, Liu, X, Ansari, D.O, and Lodish, H.F. Genes Dev. 1998; 12: 3084–3095CrossrefSee all ReferencesHua et al. 1998). The dual sequence requirement is also exhibited by an activin-response element, which requires adjacent FAST-1- and Smad-binding sequences for full ligand-inducible transcription (Zhou et al. 1998xZhou, S, Zawel, L, Lengauer, C, Kinzler, K.W, and Vogelstein, B. Mol. Cell. 1998; 2: 121–127Abstract | Full Text | Full Text PDFSee all ReferencesZhou et al. 1998). Therefore, at the Mix.2 promoter, Smad2/4 may not only interact with FAST-1 (Chen et al. 1997xChen, X, Weisberg, E, Fridmacher, V, Watanabe, M, Naco, G, and Whitman, M. Nature. 1997; 389: 85–89Crossref | Scopus (446)See all ReferencesChen et al. 1997), but presumably also with an adjacent sequence, most likely through Smad4 (since Smad2 does not bind DNA). Accordingly, Smad2/4 interacts with FAST-2 at the goosecoid promoter and Smad4 interacts with an adjacent GC-rich bipartite sequence. Again, the FAST-2- and Smad4-binding sequences are both required for full inducibility of transcription by ligand or Smad2/4 (Labbe et al. 1998xLabbe, E, Silvestri, C, Hoodless, P.A, Wrana, J.L, and Attisano, L. Mol. Cell. 1998; 2: 109–120Abstract | Full Text | Full Text PDF | PubMedSee all ReferencesLabbe et al. 1998). Finally, this model may also explain why a CRE sequence and a Mad-binding sequence were independently proposed as sequences required for transcriptional induction of the Ubx promoter by Dpp in Drosophila. Mutational analysis suggests that these overlapping sequences are both required for Dpp-mediated induction (Szuts et al. 1998xSzuts, D, Eresh, S, and Bienz, M. Genes Dev. 1998; 12: 2022–2035CrossrefSee all ReferencesSzuts et al. 1998). The required interactions of Smads with other DNA-binding transcription factors explains in retrospect why no consensus sequences for TGF-β and activin-response elements were found in different promoters, and why previously known transcription factors were implicated as essential factors for ligand-induced transcription.Given this cooperativity with other transcription factors, it may be surprising that the Smad3/4-binding elements themselves allow transcription in response to TGF-β and Smad3/4 (Dennler et al. 1998xDennler, S, Itoh, S, Vivien, D, ten Dijke, P, Huet, S, and Gauthier, J.-M. EMBO J. 1998; 11: 3091–3100Crossref | Scopus (1298)See all ReferencesDennler et al. 1998; Jonk et al. 1998xJonk, L, Itoh, S, Heldin, C.-H, ten Dijke, P, and Kruijer, W. J. Biol. Chem. 1998; 273: 21145–21152Crossref | PubMed | Scopus (444)See all ReferencesJonk et al. 1998; Zawel et al. 1998xZawel, L, Dai, J, Buckhaults, P, Zhou, S, Kinzler, K, Vogelstein, B, and Kern, S. Mol. Cell. 1998; 1: 611–617Abstract | Full Text | Full Text PDF | PubMedSee all ReferencesZawel et al. 1998). However, since a single Smad-binding sequence by itself does not allow transcriptional activation by ligand or Smad3/4, perhaps the multimerization of these sequences in the reporter assays allows recruitment of cooperating transcription factors. Alternatively, multimerization of Smad-binding sites may synergistically amplify the low transcriptional activity of Smad3, which is apparent in yeast or in the absence of cooperating transcription factors (Zhang et al. 1998xZhang, Y, Feng, X.-H, and Derynck, R. Nature. 1998; 394: 909–912Crossref | PubMed | Scopus (567)See all ReferencesZhang et al. 1998).This transcriptional cooperativity model also provides a mechanism for integration of two signaling pathways at the promoter sequence. Thus, transcription by c-Jun/c-Fos is induced by mitogenic stimuli and stress or UV irradiation, whereas Smad3/4 activation is induced by TGF-β. The required cooperation of Smad3/4 with c-Jun/c-Fos at select AP-1-binding sites thus illustrates that a convergence of both signaling pathways is at the basis of Smad3/4-induced transcription from these promoters (Zhang et al. 1998xZhang, Y, Feng, X.-H, and Derynck, R. Nature. 1998; 394: 909–912Crossref | PubMed | Scopus (567)See all ReferencesZhang et al. 1998). A similar type of cross-talk is also required for Dpp-induced transcription from the Ubx promoter. Thus, CREB binding to the promoter results from EGF receptor activation, whereas Mad binding is induced by Dpp receptor activation (Szuts et al. 1998xSzuts, D, Eresh, S, and Bienz, M. Genes Dev. 1998; 12: 2022–2035CrossrefSee all ReferencesSzuts et al. 1998), and Mad is likely to cooperate with CREB for Dpp-induced transcription from this promoter. Based on these examples, a variety of Smad responses may depend on activation of other signaling pathways and modifications in the cooperating transcription factors may regulate Smad responses.