Topoisomerase II binds importin alpha isoforms and exportin/CRM1 but does not shuttle between the nucleus and cytoplasm in proliferating cells

Resistance to anticancer drugs that target DNA topoisomerase II (topo II) isoforms alpha and/or beta is associated with decreased nuclear and increased cytoplasmic topo IIalpha. Earlier studies have confirmed that functional nuclear localization and export signal sequences (NLS and NES) are present in both isoforms. In this study, we show that topo II alpha and beta bind and are imported into the nucleus by importin alpha1, alpha3, and alpha5 in conjunction with importin beta. Topo IIalpha also binds exportin/CRM1 in vitro. However, wild-type topo IIalpha has only been observed in the cytoplasm of cells that are entering plateau phase growth. This suggests that topo IIalpha may shuttle between the nucleus and the cytoplasm with the equilibrium towards the nucleus in proliferating cells but towards the cytoplasm in plateau phase cells. The CRM1 inhibitor Leptomycin B increases the nuclear localization of GFP-tagged topo IIalpha with a mutant NLS, suggesting that its export is being inhibited. However, homokaryon shuttling experiments indicate that fluorescence-tagged wild-type topo II alpha and beta proteins do not shuttle in proliferating Cos-1 or HeLa cells. We conclude that topo II alpha and beta nuclear export is inhibited in proliferating cells so that these proteins do not shuttle.