INHIBITION OF DNA TOPOISOMERASE II-ALPHA GENE-EXPRESSION BY THE P53 TUMOR-SUPPRESSOR

DNA topoisomerase II (topo II) is an essential nuclear enzyme involved in major cellular functions such as DNA replication, transcription, r ecombination, and mitosis. While an elevated level of topo II alpha is associated with cell proliferation, wild-type (wt) p53 inhibits the e xpression of various growth-stimulatory genes. To determine if p53 dow nregulates topo II alpha gene expression, a murine cell line, (10)1val , that expresses a temperature-sensitive p53 was utilized. The (10)1va l cells had significantly lower levels of topo II alpha mRNA and prote in following incubation for 24 h at 32 degrees C (p53 with wt conforma tion) than at 39 degrees C (p53 with mutant conformation). The effect of p53 on the human topo II alpha gene promoter was determined by usin g Luciferase reporter plasmids containing varying lengths of the topo II alpha promoter transiently cotransfected into p53-deficient (10)1 c ells together with wt or mutant p53 expression plasmids. Transcription from the full-length (bp -557 to +90) topo II alpha promoter was decr eased 15-fold by wt p53 in a concentration-dependent manner, whereas m utant p53 exerted much weaker inhibition. Consecutive deletion of the five inverted CCAAT elements (ICEs) from the topo II alpha promoter re duced both the basal promoter activity and wt p53-induced suppression. Transcription of the minimal promoter (-32 to +90), which contains no ICE, was slightly stimulated by wt or mutant p53 expression. When poi nt mutations were introduced into the most proximal ICE (-68), the inh ibitory effect of wt p53 was alleviated and stimulation of topo II alp ha expression resulted. Our study suggests that wt p53 functions as a transcriptional repressor of topo II alpha gene expression, possibly t hrough a functional interaction with specific ICEs. Inactivation of wt p53 may reduce normal regulatory suppression of topo II alpha and con tribute to abortive cell cycle checkpoints, accelerated cell prolifera tion, and alterations in genomic stability associated with neoplasia.