Gunaratne Adrian, Chan Eddie, El-Chabib Tarek H, Carter David, Di Guglielmo Gianni M
J Cell Sci. 2015 Feb 1;128(3):487–98. doi: 10.1242/jcs.155440.
Transforming growth factor b (TGFb) signaling controls many cellular responses including proliferation, epithelial to mesenchymal transition and apoptosis, through the activation of canonical (Smad) as well as non-canonical (e.g., Par6) pathways. Previous studies from our lab have demonstrated that aPKC inhibition regulates TGFb receptor trafficking and signaling. Here, we report that downstream TGFb-dependent transcriptional responses in aPKC-silenced NSCLC cells were reduced compared with those of control cells, despite a temporal extension of Smad2 phosphorylation. We assessed SARA–Smad2–Smad4 association and observed that knockdown of aPKC increased SARA (also known as ZFYVE9) levels and SARA–Smad2 complex formation, increased cytoplasmic retention of Smad2 and reduced Smad2–Smad4 complex formation, which correlated with reduced Smad2 nuclear translocation. Interestingly, we also detected an increase in p38 MAPK phosphorylation and apoptosis in aPKC-silenced cells, which were found to be TRAF6-dependent. Taken together, our results suggest that aPKC isoforms regulate Smad and non-Smad TGFb pathways and that aPKC inhibition sensitizes NSCLC cells to undergo TGFb dependent apoptosis.
转化生长因子β（TGFβ）信号传导通过激活经典（Smad）以及非经典（例如Par6）途径来控制许多细胞反应，包括增殖、上皮-间质转化和细胞凋亡。我们实验室先前的研究表明，非典型蛋白激酶C（aPKC）抑制作用可调节TGFβ受体的运输和信号传导。在此，我们报告，尽管Smad2磷酸化存在时间延长，但与对照细胞相比，aPKC沉默的非小细胞肺癌（NSCLC）细胞中TGFβ依赖性转录反应仍有所降低。我们评估了Smad锚定受体激活蛋白（SARA）-Smad2-Smad4的结合情况，观察到aPKC敲低会增加SARA（也称为锌指FYVE结构域蛋白9）水平和SARA-Smad2复合物的形成，增加Smad2在细胞质中的滞留并减少Smad2-Smad4复合物的形成，这与Smad2核转位减少相关。有趣的是，我们还检测到aPKC沉默细胞中p38丝裂原活化蛋白激酶（MAPK）磷酸化和细胞凋亡增加，且发现这是肿瘤坏死因子受体相关因子6（TRAF6）依赖性的。综上所述，我们的结果表明，aPKC亚型调节Smad和非Smad的TGFβ途径，并且aPKC抑制作用使NSCLC细胞对TGFβ依赖性细胞凋亡敏感。
