The 180-kDa isoform of topoisomerase II is localized in the nucleolus and belongs to the structural elements of the nucleolar remnant

Abstract

Monoclonal antibodies raised against two isoforms (170 and 150 180 kDa) of DNA topoisomerase II showed distinct fluorescence patterns in HeLa cells in different moments of the cell cycle ( C. Negri et al., 1992 , Exp. Cell Res. 200, 452–459). The ultrastructural distribution of the 150 180 -kDa isoform, which in immunofluorescence showed a localization into the nucleolar region, has been analyzed by electron microscopy with a gold-conjugated secondary antibody in HeLa and K562 cells. The results indicate that this isoform of the enzyme is exclusively localized in the nucleolus, mainly in the dense fibrillar component, while the nucleoplasm of interphase cells and the chromosomes of mitotic cells are completely negative. The antibody also reacts with the nucleolus of isolated nuclei and with the nucleolar remnant of purified nuclear matrices. A quantitative evaluation of the label distribution indicates that the percentage of label in the nucleolar remnant of isolated matrix is almost identical to that of the nucleolus in whole cells. The interaction with the insoluble proteins of the isolated nuclear matrix is also demonstrated by quantitative immunoblotting in which the MoAb specifically stains a unique band corresponding to the 150 180 -kDa isoform of topoisomerase II. The localization of the 150 180 -kDa isoform of topoisomerase II in the nucleolar remnant strongly suggests that it represents a structural element for the spatial organization and for the regulation of transcription of the ribosomal genes.