aPKC alters the TGFß response in NSCLC cells through both Smad-dependent and Smad-independent pathways.

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 SARA­Smad2­Smad4 association and observed that knockdown of aPKC increased SARA (also known as ZFYVE9) levels and SARA­Smad2 complex formation, increased cytoplasmic retention of Smad2 and reduced Smad2­Smad4 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.