Panopoulou Ekaterini, Gillooly David J, Wrana Jeffrey L, Zerial Marino, Stenmark Harald, Murphy Carol, Fotsis Theodore
Laboratory of Biological Chemistry, University of Ioannina Medical School, 45110 Ioannina, Greece.
J Biol Chem. 2002 May 17;277(20):18046-52. doi: 10.1074/jbc.M107983200. Epub 2002 Mar 4.
Transforming growth factor beta (TGFbeta) receptors require SARA for phosphorylation of the downstream transducing Smad proteins. SARA, a FYVE finger protein, binds to membrane lipids suggesting that activated receptors may interact with downstream signaling molecules at discrete endocytic locations. In the present study, we reveal a critical role for the early endocytic compartment in regulating Smad-dependent signaling. Not only is SARA localized on early endosomes, but also its minimal FYVE finger sequence is sufficient for early endosomal targeting. Expression of a SARA mutant protein lacking the FYVE finger inhibits downstream activin A signaling in endothelial cells. Moreover, a dominant-negative mutant of Rab5, a crucial protein for early endosome dynamics, causes phosphorylation and nuclear translocation of Smads leading to constitutive (i.e. ligand independent) transcriptional activation of a Smad-dependent promoter in endothelial cells. As inhibition of endocytosis using the K44A negative mutant of dynamin and RN-tre did not lead to activation of Smad-dependent transcription, the effects of the dominant-negative Rab5 are likely to be a consequence of altered membrane trafficking of constitutively formed TGFbeta/activin type I/II receptor complexes at the level of early endosomes. The results suggest an important interconnection between early endosomal dynamics and TGFbeta/activin signal transduction pathways.
转化生长因子β（TGFβ）受体需要Smad锚定受体激活蛋白（SARA）来使下游转导性Smad蛋白发生磷酸化。SARA是一种含FYVE结构域的蛋白，可与膜脂结合，这表明活化的受体可能在离散的内吞位点与下游信号分子相互作用。在本研究中，我们揭示了早期内吞区室在调节Smad依赖性信号传导中的关键作用。不仅SARA定位于早期内体，而且其最小的FYVE结构域序列就足以实现早期内体靶向。缺乏FYVE结构域的SARA突变蛋白的表达会抑制内皮细胞中激活素A的下游信号传导。此外，Rab5的显性负性突变体是早期内体动态变化的关键蛋白，它会导致Smad的磷酸化和核转位，从而导致内皮细胞中Smad依赖性启动子的组成型（即不依赖配体）转录激活。由于使用发动蛋白的K44A负性突变体和RN-tre抑制内吞作用不会导致Smad依赖性转录的激活，显性负性Rab5产生的效应可能是早期内体水平上组成型形成的TGFβ/激活素I/II型受体复合物膜转运改变的结果。这些结果表明早期内体动态变化与TGFβ/激活素信号转导途径之间存在重要的相互联系。
