Inhibition of DNA topoisomerase II with etoposide induces association of DNA topoisomerase IIα, DNA topoisomerase IIβ, and nucleolin with BCR 2 of the ETO gene

Abstract

Chromosomal translocation t(8;21)(q22;q22), which affects AML1 and ETO genes, is often detected in patients with acute myeloid leukemia. The mechanisms that ensure preferential rearrangement at topoisomerase breakpoints remain obscure. It is known that the preferential breakpoints in DNA strands are clustered within the narrow breakpoint-clustering regions (BCRs). We have earlier shown that, in the nuclei of human primary fibroblasts, these two genes are not located close to one another; furthermore, they are located in different chromosome layers. We discovered that the treatment of cells with etoposide (VP-16), an inhibitor of DNA topoisomerase II, changed the preferential nuclear location of the ETO gene so that the AML1 and ETO genes became located in the same chromosome layer. Inhibitor analysis showed that this effect was most likely due to formation of specific stalled cleavable complexes on DNA in the presence of etoposide and that nuclear myosin is apparently involved in the translocation. In this study, we showed that doublestrand breaks are clustered in the immediate vicinity of the nucleolus, which directly indicates that the latter may be involved in illegitimate recombination leading to the occurrence of translocations. We continued studies using chromatin precipitation and real-time TaqMan PCR and found that the BCR2 region of the ETO gene was enriched not only with the DNA topoisomerase II α but also with the nucleolus-specific proteins DNA topoisomerase II β and nucleolin.