Cell cycle-coupled variation in topoisomerase II alpha mRNA is regulated by the 3 '-untranslated region - Possible role of redox-sensitive protein binding in mRNA accumulation

Mammalian topoisomerase II alpha (Topo II) is a highly regulated enzyme ess ential for many cellular processes including the G(2) cell cycle checkpoint . Because Topo II gene expression is regulated posttranscriptionally during the cell cycle, we investigated the possible role of the 3'-untranslated r egion (3'-UTR) in controlling Topo II mRNA accumulation. Reporter assays in stably transfected cells demonstrated that, similar to endogenous Topo II mRNA levels, the mRNA levels of reporter genes containing the Topo II 3'-UT R varied during the cell cycle and were maximal in S and G(2)/M relative to G(1). Topo II 3'-UTR sequence analysis and RNA-protein binding assays iden tified a 177-nucleotide (base pairs 4772-4949) region containing an AUUUUUA motif sufficient for protein binding. Multiple proteins (84, 70, 44, and 3 7 kDa) bound this region, and the binding of 84- and 37-kDa (tentatively id entified as the adenosine- or uridine-rich element-binding factor AUF1) pro teins was enhanced in G(1), correlating with decreased Topo II mRNA levels. The binding activity was enhanced in cellular extracts or cells treated wi th thiol-reducing agents, and increased binding correlated with decreased T opo II mRNA levels. These results support the hypothesis that cell cycle-co upled Topo II gene expression is regulated by interaction of the 3'-UTR wit h redox-sensitive protein complexes.