title: Ablation of Smurf2 reveals an inhibition in TGF-β signalling through multiple mono-ubiquitination of Smad3.
abstract: TGF-β signalling is regulated by post-translational modifications of Smad proteins to translate quantitative difference in ligand concentration into proportional transcriptional output. Previous studies in cell culture systems suggested that Smad ubiquitination regulatory factors (Smurfs) act in this regulation by targeting Smads for proteasomal degradation, but whether this mechanism operates under physiological conditions is not clear. Here, we generated mice harbouring a target-disrupted Smurf2 allele. Using primary mouse embryonic fibroblasts and dermal fibroblasts, we show that TGF-β-mediated, Smad-dependent transcriptional responses are elevated in the absence of Smurf2. Instead of promoting poly-ubiquitination and degradation, we show that Smurf2 actually induces multiple mono-ubiquitination of Smad3 in vivo. Phosphorylation of T179, immediately upstream of the Smad3 PY motif, enhances Smurf2 and Smad3 interaction and Smad3 ubiquitination. We have mapped Smurf2-induced Smad3 ubiquitination sites to lysine residues at the MH2 domain, and demonstrate that Smad3 ubiquitination inhibits the formation of Smad3 complexes. Thus, our data support a model in which Smurf2 negatively regulates TGF-β signalling by attenuating the activity of Smad3 rather than promoting its degradation.
authors: Laboratory of Cellular and Molecular Biology, Center for Cancer Research, National Cancer Institute, Bethesda, MD, USA.
authors: Liu-Ya
authors: LY
authors: Tang
authors: Motozo
authors: M
authors: Yamashita
authors: Nathan P
authors: NP
authors: Coussens
authors: Yi
authors: Y
authors: Tang
authors: Xiangchun
authors: X
authors: Wang
authors: Cuiling
authors: C
authors: Li
authors: Chu-Xia
authors: CX
authors: Deng
authors: Steven Y
authors: SY
authors: Cheng
authors: Ying E
authors: YE
authors: Zhang
mesh_terms: D000818:Animals; D015153:Blotting, Western; D005347:Fibroblasts / Q000378:metabolism; D051379:Mice; D008954:Models, Biological; D011485:Protein Binding; D015398:Signal Transduction / Q000502:physiology*; D051900:Smad3 Protein / Q000378:metabolism*; D014158:Transcription, Genetic / Q000502:physiology; D016212:Transforming Growth Factor beta / Q000378:metabolism*; D025801:Ubiquitin / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000235:genetics / Q000378:metabolism; D054875:Ubiquitination / Q000502:physiology*
chemical_list: D051900:Smad3 Protein; D016212:Transforming Growth Factor beta; D025801:Ubiquitin; C533325:Smurf2 protein, mouse; D044767:Ubiquitin-Protein Ligases