Study on the mechanism of cancer cell death induced by TGFβ1, TGFβ2 downregulation

Abstract

TGF-β signaling has been increasingly recognized as a key driver in cancer. Unlike its tumor suppressor function in normal tissue, TGF-β activation incites tumor progression in cancer tissue and an increase in TGF-β expression often correlates with the malignancy of many cancers. In this study, we tried to unravel the mechanism of TGF-β downregulation-induced by using adenovirus expressing short hairpin RNA against transforming growth factor-β1 or β2 (TGF-β1/2). Notably, we found that TGF-β downregulation could increase the phospho-p38 and phospho-JNK expression, also decrease the survival molecule such as phospho-Akt, phospho-Src, phospho-Stat3 and phospho-p65. Consistent with the increase of phosphor-p38 and p-JNK, the ASK1 phosphorylation (which means ASK1 activation) and reactive oxygen species(ROS) production were also increased in response to TGF-β downregulation, whereas gene expression of Trx and GSTM1 known to be inhibitory binding proteins to ASK1 were decreased. In addition, interactions between GSTM1 and ASK1 or Trx and ASK1 were also decreased. This decrease in Trx and GSTM1 expression was likely to be related to the translocation of Smad complex proteins as a main mediator of canonical signalling pathway of TGF-β playing a tumor-promoting role by transcriptional activation of target genes such as Trx or GSTM1. However, ROS was not directly related to the transcriptional repression of Trx or GSTM1, while inducing dissociation of Trx and GSTM1 from ASK1 activation followed by tumor cell death. Morevoer, ASK1 inhibition with siASK1 or overexpression of a dominant-negative kinase-inactive mutant of ASK1(ASK1-KM) rescued cell death. In addition, p38 MAPK/JNK activation was also inhibited by siASK1 or ASK1-KM, suggesting that ASK1 signaling via p38 MAPK/JNK activation was the main pathway of adenovirus-expressing shTGF- β1 or 2 induced cell death. Taken together, our findings demonstrate that treatment with adenovirus expressing shRNA of TGF-β1 or 2 can cause cell death via ASK1 activation, which was associated with the reduction of Trx and GSTM1 gene expression and dissociation of the Trx/GSTM1 from ASK1–Trx, ASK1-GSTM1 complexes.