This application focuses on the crystallographic studies of the Smad proteins in TGF-beta signaling. TGF-beta signaling from cell membrane to the nucleus is mediated by the Smad family of proteins (Smads), of which Smad2 and Smad4 have been identified as tumor suppressors in humans. Our structural studies on the Smad proteins during the current funding cycle have revealed significant insight into the general mechanisms of TGF-beta signaling. The work proposed here focuses on the fundamental unresolved issues in TGF-beta signaling: (1) Structural and biochemical analyses of a phosphorylated Smad2-Smad4 complex. A phosphorylated Smad heteromeric complex is the ultimate functional unit in TGF-beta signaling. The structure of a phosphorylated Smad2-Smad4 complex will be determined by molecular replacement. Insights will be gained to decipher the mechanism of Smad activation. (2) Structural and biochemical analyses of the Smad2-CBP and Smad2-SnoN complexes. The general transcriptional co-activator CBP/p300 and the proto-oncoprotein SnoN/Ski are among the most important nuclear partners of Smads. Understanding their interactions with Smads will reveal important insights into the mechanisms of Smad-mediated transcriptional regulation. (3) Structural and biochemical analyses of the Smad7-Smurf2 and Smad2-Smurf2 complexes. Smad proteins are routinely degraded by the Smad ubiquitination regulatory factors (Smurfs) and only stabilized in response to TGF-beta signaling. Structural information on the Smad- Smurf complex will help elucidate how Smad proteins are degraded and how Smad recognition by Smurfs is coupled to its ubiquitination. (4) Structural and biochemical analyses of Smad3/Smad4/TFE-3 complexes bound to the plasminogen activator inhibitor-1 (PAI-1) promoter. This structure will ultimately address a pivotal issue in TGF-beta signaling; that is, how Smad proteins cooperate with other nuclear partners to modulate transcription of the TGF-beta-responsive genes. In particular, we will be able to examine the effects of the tumorigenic mutations on the assembly of an active transcriptional complex.
| Lin, Xia; Duan, Xueyan; Liang, Yao-Yun et al. (2006) PPM1A functions as a Smad phosphatase to terminate TGFbeta signaling. Cell 125:915-28 |
| Shi, Yigong; Massague, Joan (2003) Mechanisms of TGF-beta signaling from cell membrane to the nucleus. Cell 113:685-700 |
| Chai, Jijie; Wu, Jia-Wei; Yan, Nieng et al. (2003) Features of a Smad3 MH1-DNA complex. Roles of water and zinc in DNA binding. J Biol Chem 278:20327-31 |
| Wu, Jia Wei; Krawitz, Ariel R; Chai, Jijie et al. (2002) Structural mechanism of Smad4 recognition by the nuclear oncoprotein Ski: insights on Ski-mediated repression of TGF-beta signaling. Cell 111:357-67 |
| Wu, J W; Hu, M; Chai, J et al. (2001) Crystal structure of a phosphorylated Smad2. Recognition of phosphoserine by the MH2 domain and insights on Smad function in TGF-beta signaling. Mol Cell 8:1277-89 |
| Wu, J W; Fairman, R; Penry, J et al. (2001) Formation of a stable heterodimer between Smad2 and Smad4. J Biol Chem 276:20688-94 |
