HISTONE DEACETYLASE INHIBITOR ACTIVATES THE WAF1 CIP1 GENE PROMOTER THROUGH THE SP1 SITES/

Treatment of cultured cells with trichostatin A (TSA), a specific hist one deacetylase inhibitor, induces the histone hyperacetylation and mo dulates expression of some mammalian genes. We examined the effects of TSA on cell growth arrest, and its relation to expression of the WAF1 /Cip1 gene, a potent inhibitor of cyclin-dependent kinases, in a p53-m utated human osteosarcoma cell line MG63. TSA at 500 ng/ml induced gro wth arrest at both G1 and G2/M phases, and the expressions of the WAF1 /Cip1 mRNA and protein. We also examined the changes of acetylated iso forms of histone H4. Dose-response and kinetic analysis suggest a clos e correlation between the level of histone acetylation and the inducti on of the WAF1/Cip1 expressions. Using several mutant WAF1/Cip1 promot er fragments, we found that the TSA responsive elements are two Sp1 si tes at -82 and -69 relative to the transcription start site. These fin dings indicate that TSA induces the WAF1/Cip1 promoter through the typ ical Sp1 sites, in a p53-independent fashion. Furthermore, the Sp1-luc plasmid, containing SV40 promoter-derived three consensus Sp1 binding sites, was markedly activated by TSA compared to the mutant Sp1-luc p lasmid. These results demonstrate that transcriptional activation thro ugh the Sp1 sites of the WAF1/Cip1 promoter by TSA coincides with indu ced hyperacetylation of histone H4. (C) 1997 Academic Press.