Activation of p21WAF1/Cip1 Transcription through Sp1 Sites by Histone Deacetylase Inhibitor Apicidin

We previously reported that apicidin, a novel histone deacetylase inhibitor, inhibited the proliferation of tumor cells via induction of p21(WAF1/Cip1). In this study, we determined the molecular mechanisms by which apicidin induced the p21(WAF1/Cip1) gene expression in HeLa cells. Apicidin induced p21(WAF1/Cip1) mRNA independent of the de novo protein synthesis and activated the p21(WAF1/Cip1) promoter through Sp1-3 site located at -82 and -77 relative to the transcription start site. This transcriptional activation appears to be mediated by protein kinase C (PKC), because calphostin C, a PKC inhibitor, significantly attenuated the activation of p21(WAF1/Cip1) promoter via Sp1 sites, which was accompanied by a marked suppression of p21(WAF1/Cip1) mRNA and protein expression induced by apicidin. Consistent with the transcriptional activation of p21(WAF1/Cip1) promoter by apicidin, apicidin treatment led to the translocation of PKCepsilon from cytosolic to particulate fraction, which was reversed by pretreatment with calphostin C, indicating the involvement of PKC in the transcriptional activation of p21(WAF1/Cip1) via Sp1 sites by apicidin. However, the PKC-mediated transcriptional activation of p21(WAF1/Cip1) by apicidin appears to be independent of the histone hyperacetylation, because apicidin-induced histone hyperacetylation was not affected by calphostin C. Furthermore, a PKC activator, phorbol 12,13-dibutyrate, alone induced the transcriptional activation of p21(WAF1/Cip1) promoter, p21(WAF1/Cip1) mRNA, and protein expression without induction of the histone hyperacetylation, suggesting that the transcriptional activation of p21(WAF1/Cip1) by apicidin might have been mediated by a mechanism other than chromatin remodeling through the histone hyperacetylation. Taken together, these results suggest that the PKC signaling pathway plays a pivotal role in the transcriptional activation of the p21(WAF1/Cip1) gene by apicidin.