Guo Xing, Ramirez Alejandro, Waddell David S, Li Zhizhong, Liu Xuedong, Wang Xiao-Fan
Department of Pharmacology and Cancer Biology, Duke University Medical Center, Durham, North Carolina 27710, USA.
Genes Dev. 2008 Jan 1;22(1):106-20. doi: 10.1101/gad.1590908.
The broad range of biological responses elicited by transforming growth factor-beta (TGF-beta) in various types of tissues and cells is mainly determined by the expression level and activity of the effector proteins Smad2 and Smad3. It is not fully understood how the baseline properties of Smad3 are regulated, although this molecule is in complex with many other proteins at the steady state. Here we show that nonactivated Smad3, but not Smad2, undergoes proteasome-dependent degradation due to the concerted action of the scaffolding protein Axin and its associated kinase, glycogen synthase kinase 3-beta (GSK3-beta). Smad3 physically interacts with Axin and GSK3-beta only in the absence of TGF-beta. Reduction in the expression or activity of Axin/GSK3-beta leads to increased Smad3 stability and transcriptional activity without affecting TGF-beta receptors or Smad2, whereas overexpression of these proteins promotes Smad3 basal degradation and desensitizes cells to TGF-beta. Mechanistically, Axin facilitates GSK3-beta-mediated phosphorylation of Smad3 at Thr66, which triggers Smad3 ubiquitination and degradation. Thr66 mutants of Smad3 show altered protein stability and hence transcriptional activity. These results indicate that the steady-state stability of Smad3 is an important determinant of cellular sensitivity to TGF-beta, and suggest a new function of the Axin/GSK3-beta complex in modulating critical TGF-beta/Smad3-regulated processes during development and tumor progression.
转化生长因子-β（TGF-β）在各种组织和细胞中引发的广泛生物学反应主要由效应蛋白Smad2和Smad3的表达水平及活性决定。尽管在稳态下Smad3与许多其他蛋白存在复合体，但目前尚不完全清楚Smad3的基线特性是如何调控的。在此我们发现，未激活的Smad3而非Smad2，由于支架蛋白Axin及其相关激酶糖原合酶激酶3-β（GSK3-β）的协同作用，会经历蛋白酶体依赖性降解。仅在不存在TGF-β时，Smad3才与Axin和GSK3-β发生物理相互作用。Axin/GSK3-β的表达或活性降低会导致Smad3稳定性和转录活性增加，而不影响TGF-β受体或Smad2，而这些蛋白的过表达则会促进Smad3的基础降解并使细胞对TGF-β脱敏。从机制上讲，Axin促进GSK3-β介导的Smad3第66位苏氨酸磷酸化，从而触发Smad3泛素化和降解。Smad3的第66位苏氨酸突变体表现出改变的蛋白稳定性以及转录活性。这些结果表明，Smad3的稳态稳定性是细胞对TGF-β敏感性的重要决定因素，并提示Axin/GSK3-β复合体在发育和肿瘤进展过程中调节关键的TGF-β/Smad3调控过程具有新功能。
