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© The Rockefeller University Press, 0021-9525/1997//775 $5.00
The Journal of Cell Biology, Volume 136, Number 4, , 1997 775-788

Article

Cell Cycle–coupled Relocation of Types I and II Topoisomerases and Modulation of Catalytic Enzyme Activities



Kay N. Meyer*, Eigil Kjeldsen§, Tobias Straub*, Birgitta R. Knudsen||, Ian D. Hickson, Akihiko Kikuchi**, Hans Kreipe{ddagger}, and Fritz Boege*,||

* Medizinische Poliklinik and {ddagger} Institute of Pathology, University of Würzburg, Germany; § Department of Human Genetics and || Department of Molecular and Structural Biology, University of Aarhus, Denmark; Institute of Molecular Medicine, John Radcliffe Hospital, University of Oxford, United Kingdom; and ** Laboratory of Medical Mycology, Institute for Disease Mechanism and Control, Nagoya University, Japan

We visualized DNA topoisomerases in A431 cells and isolated chromosomes by isoenzyme-selective immunofluorescence microscopy. In interphase, topoisomerase I mainly had a homogeneous nuclear distribution. 10–15% of the cells exhibited granular patterns, 30% showed bright intranucleolar patches. Topoisomerase II isoenzymes showed spotted ({alpha}) or reticular (β) nuclear patterns throughout interphase. In contrast to topoisomerase II{alpha}, topoisomerase IIβ was completely excluded from nucleoli. In mitosis, topoisomerase IIβ diffused completely into the cytosol, whereas topoisomerases I and II{alpha} remained chromosome bound. Chromosomal staining of topoisomerase I was homogeneous, whereas topoisomerase II{alpha} accumulated in the long axes of the chromosome arms and in the centriols. Topoisomerase antigens were 2–3-fold higher in mitosis than in interphase, but specific activities of topoisomerase I and II were reduced 5- and 2.4-fold, respectively. These changes were associated with mitotic enzyme hyperphosphorylation. In interphase, topoisomerases could be completely linked to DNA by etoposide or camptothecin, whereas in mitosis, 50% of topoisomerase II{alpha} escaped poisoning. Refractoriness to etoposide could be assigned to the salt-stable scaffold fraction of topoisomerase II{alpha}, which increased from <2% in G1 phase to 48% in mitosis. Topoisomerases I and IIβ remained completely extractable throughout the cell cycle. In summary, expression of topoisomerases increases towards mitosis, but specific activities decrease. Topoisomerase IIβ is released from the heterochromatin, whereas topoisomerase I and II{alpha} remain chromosome bound. Scaffold-associated topoisomerase II{alpha} appears not to be involved in catalytic DNA turnover, though it may play a role in the replicational cycle of centriols, where it accumulates during M phase.

Purified recombinant human topoisomerase IIβ was kindly provided by Dr. Ole Westergaard (Department of Structural and Molecular Biology, University of Aarhus, Denmark). We would also like to thank Dr. Westergaard for helpful discussions. We are grateful to Ms. Berit Hornstrup for helping us prepare the manuscript.

Please address all correspondence to Fritz Boege, Medizinische Poliklinik, University of Wuerzburg, Klinikstr. 6–8, D-97070 Wuerzburg, Germany. Tel.: (49) 931-201-7008; Fax.: (49) 931-201-7120.


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