TGF-β downregulates PTEN via activation of NF-κB in pancreatic cancer cells

TGF-β utilizes receptor-activated SMAD signaling to mediate growth suppression; however, non-SMAD signaling that modulates the TGF-β response in epithelial cells become apparent when the SMAD signaling is abrogated, a common occurrence in pancreatic cancers. Here, we examined whether TGF-β utilized NF-κB to downregulate PTEN, a gene that is rarely mutated in pancreatic cancers. SMAD4-null BxPc3 and CAPAN-1 pancreatic cancer cells were treated with TGF-β (10 ng/ml) and lysed, and cellular proteins were analyzed by Western blots using p-IκB, p65, and PTEN antibodies. PTEN promoter and NF-κB activities were assessed by PTEN-luc and p-NF-luc constructs, respectively. Dominant negative p-IκB-α-M (NF-κB superrepressor) was used to block activation of NF-κB. Cell motility was assessed by Boyden chamber migration assay. TGF-β induced IκB-α phosphorylation followed by NF-κB p65 subunit nuclear translocation and increased NF-κB activity. IκB-α-M blocked TGF-β-induced NF-κB activity, reversed downregulated PTEN promoter activity and PTEN expression, and prevented augmentation of cell motility induced by TGF-β. SMAD4 restoration, but not knockdown of SMAD2 and/or 3, reversed TGF-β-induced NF-κB activity. Thus TGF-β suppresses PTEN in pancreatic cancer cells through NF-κB activation and enhances cell motility and invasiveness in a SMAD4-independent manner that can be counteracted when TGF-β-SMAD signaling is restored. The TGF-β/NF-κB/PTEN cascade may be a critical pathway for pancreatic cancer cells to proliferate and metastasize.