Sp1 mediates cell proliferation-dependent regulation of rat DNA topoisomerase IIα gene promoter

Abstract

DNA topoisomerase IIalpha (topo IIalpha) is an essential nuclear enzyme required for chromosome segregation during mitosis. Consistent with its critical role in cell division is the fact that the expression of the gene for topo IIalpha is strongly regulated by the proliferation state of cells. Using a transient expression system, we determined the contribution of putative cis-acting elements in its promoter region to its basal level and cell proliferation-dependent transcription. Experiments with 5' and/or 3' serial deletion and site-directed mutation revealed that (1) maximal promoter activity resides in the fragment extending to position -663 bp from the ATG initiation codon, (2) minimal promoter activity is harboured at -195 bp, (3) the defined minimal promoter contains only two putative elements, inverted CCAAT box 4 (ICB4) (-166 to -162 bp) and the most proximal GC-rich box in the promoter (GC2) (-149 to -143 bp), and (4) ICB4 is most important in the basal-level transcription of the gene for rat topo IIalpha. The luciferase activities of the mutated reporter plasmids in G(0)-arrested and exponentially growing cells showed that proliferation-specific regulation is controlled mainly by GC2. Electrophoretic mobility-shift assays indicated that Sp1 binds specifically to the GC2 site. The extent of DNA-protein complex formation increases after the stimulation of cells to proliferate. These results indicate that the increased binding activity of Sp1 to GC2 is important in the up-regulation of the gene for topo IIalpha in growing cells.

Down-regulation of human topoisomerase IIα expression correlates with relative amounts of specificity factors Sp1 and Sp3 bound at proximal and distal promoter regions

“…These findings are consistent with other work, which has shown that in the rat topoisomerase IIα promoter, Sp1 is a transcriptional activator at the GC2 element, which is spatially equivalent to the GC1 element of the human topoisomerase IIα promoter [23]. …”

Amplification of the TOP2A gene does not predict high levels of topoisomerase II alpha protein in human breast tumor samples

“…Our results may reflect the fundamental differences between the roles of HER2 and topoisomerase II alpha in the cell. TOP2A expression is regulated through inverted CCAAT box motifs (Adachi et al, 2000), which bind p53 Sandri et al, 1996;Wang et al, 1997;Yoon et al, 1999) and coordinate transcription with the state of cell cycling. The TOP2A promoter responds to increased Ras activity associated with cell proliferation (Woessner et al, 1990;Isaacs et al, 1998;Chen et al, 1999;Stacey et al, 2000) At the mRNA level, topoisomerase II alpha is regulated through untranslated 3Ј sequences (Goswami et al, 2000) that govern the protein's half-life and its location in either the nuclear or cytoplasmic compartments of the cell.…”

Concurrent inhibition of enzymatic activity and NF-Y-mediated transcription of Topoisomerase-IIα by bis-DemethoxyCurcumin in cancer cells

“…9 Among multiple transcription factors controlling TOP2A transcription throughout the cell cycle, Sp and NF-Y have been shown to bind GC consensus sites and four inverted CCAAT boxes (ICB1–ICB4), respectively. 10, 11, 12 …”