CLONING AND CHARACTERIZATION OF THE 5'-FLANKING SEQUENCE FOR THE HUMAN DNA TOPOISOMERASE-II BETA-GENE

Mammalian cells express two isoforms of type II DNA topoisomerase whic h are the intracellular targets of many structurally diverse antineopl astic agents. The levels of topoisomerase II isozymes are important de terminants for the sensitivity of cells to the cytotoxicity of drugs t hat target topoisomerase II. To investigate whether the expression of topoisomerase II isoforms is coordinated and the mechanisms governing their expression in the context of drug resistance, the 5'-flanking se quence for the gene of human topoisomerase II beta isoform was cloned and characterized. The 5'-flanking region has a very high GC content a nd contains no canonical TATA box element. Two separate transcriptiona l start sites are located to an adenine and a guanine, 193 and 89 nucl eotides, respectively, upstream from the ATG translation initiation co don. Except for a small region immediately upstream of the translation initiation codon, there is no obvious sequence homology between the 5 '-flanking sequences of human topoisomerase II beta gene and the previ ously described or gene. Transient expression assays with different 5' - and 3'-deletions of the 5'-flanking region of the topoisomerase II b eta gene have delineated regions important for transcriptional regulat ion of the gene. Interestingly, sequences within the first intron also contribute to the promoter activity. Gel mobility shift studies demon strate that protein factors from the nuclear extracts can bind specifi cally to the downstream elements and may participate in transcriptiona l regulation. (C) 1997 Elsevier Science B.V.