Heterogeneous transforming growth factor (TGF)-β unresponsiveness and loss of TGF-β receptor type II expression caused by histone deacetylation in lung cancer cell lines

Transforming growth factor (TGF)-beta strongly inhibits epithelial cell proliferation. Alterations of TGF-beta signaling are thought to play a role in tumorigenesis. We show in the present study that most lung cancer cell lines have lost the growth-inhibitory response to TGF-beta signal, and that those with TGF-beta unresponsiveness can be divided into two major groups, TGF-beta type II receptor (TGF beta RII)(+)/Smad7(+) and TGF beta RII(-)/Smad7(-), suggesting the heterogeneous mechanisms underlying the TGF-beta responsiveness. The mechanism of the loss of TGF beta RII expression of the latter group was further studied, identifying aberrant DNA methylation of the promoter region in a limited fraction of cell lines. Interestingly, we found that the alteration of chromatin structure because of histone deacetylation may also be involved, showing a good correlation with loss of TGF beta RII expression. This notion was supported by the findings of a restriction enzyme accessibility assay, of a chromatin immunoprecipitation assay with anti-acetyl historic antibodies, and of an in vivo induction of TGF beta RII expression by histone deacetylase inhibitors including trichostatin A (TSA) and sodium butyrate. In vitro induction of TGF beta RII promoter reporter activity by TSA was also detected and found to require the CCAAT box within the -127/-75 region. A positive regulatory mechanism for TGF beta RII expression in a TGF-beta -expressing cell line was also investigated, and a TPA-responsive element (TRE)-like motif, TRE2, was detected in addition to the previously reported TRE-like motif Y element in the positive regulatory region. Alterations in two discrete proteins interacting with these two TRE-like motifs were also suspected of being involved in the loss of TGF beta RII expression. This is the first study to demonstrate that, in addition to the TSA-responsive region and TRE2 motif in the TGF beta RII promoter, the alteration of histone deacetylation may be involved in the loss of TGF beta RII expression in lung cancer cell lines.