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Temporal and Spatial Distribution of DNA Topoisomerase II Alters
During Proliferation, Differentiation, and Apoptosis in HL-60 Cells
Koichi Sugimoto,
Konagi Yamada,
Motoki Egashira,
Yoshio Yazaki,
Hisamaru Hirai,
Akihiko Kikuchi, and
Kazuo Oshimi
From the Department of Hematology, Juntendo University School of
Medicine, Tokyo, Japan; the Third Department of Internal Medicine,
Faculty of Medicine, University of Tokyo, Tokyo, Japan; and the
Laboratory of Medical Mycology, Research Institute of Disease Mechanism
and Control, Nagoya University School of Medicine, Nagoya, Japan.
We related cellular content of DNA topoisomerase (topo) II and
II with the cell cycle position in proliferating, differentiated, and apoptotic HL-60 cells using two-dimensional flow cytometry. In
logarithmically growing HL-60 cells, topo II increased especially in
late S to G2/M phases, although the topo II level was almost constant throughout the cell cycle. Induction of differentiation by
all-trans retinoic acid dramatically reduced the topo II but not the topo II level. A new G2/M population containing virtually no
topo II appeared during differentiation and was supposed to be alive
and noncycling. Two-dimensional flow cytometry of topo II or II
staining and terminal deoxynucleotidyl transferase-mediated dUTP-biotin
nick end-labeling assay showed that one topo II epitope situated at
the C-terminal end decreased specifically in apoptotic HL-60 cells
treated with Ara-C, etoposide, and vincristine. The amounts of a topo
II epitope and another topo II epitope located at a more central
portion were almost equal between apoptotic and nonapoptotic cells.
Western blot analysis confirmed that topo II protein was completely
degraded into smaller fragments and lost its C-terminal end during
apoptosis. On the contrary, a large portion of topo II remained of
its original size, although both topo II and II left from the
nuclear fraction in apoptotic cells. Confocal laser microscopy showed
nuclear localization of topo II and II in growing HL-60 cells.
Although topo II and II were distributed throughout the cell
during mitosis, only topo II was densely concentrated in the mitotic
chromosomes. Both enzymes were dissociated from the genomic DNA even at
an early phase of apoptosis and completely separated from the propidium iodide signal of DNA in the advanced stage. Chromatin condensation process in apoptosis is therefore completely topo II-independent and
obviously differs from the mitotic one.
Blood, Vol. 91 No. 4 (February 15), 1998:
pp. 1407-1417
© 1998 by The American Society of Hematology.
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