Doxorubicin and gamma rays increase the level of DNA topoisomerase IIalpha in nuclei of normal and xeroderma pigmentosum fibroblasts.
DNA topoisomerase IIalpha was monitored with the monoclonal antibody Ki-S1 in human fibroblasts after irradiation of cells with gamma rays from a 137Cs source or treatment with the DNA topoisomerase II inhibitor doxorubicin. DNA topoisomerase IIalpha was localized immunohistochemically as bright fluorescent dots in the karyoplasm. The fibroblasts investigated originated from normal human donors and a xeroderma pigmentosum (XP) patient (XP12BE). All cell lines examined showed a time- and dose-dependent increase in DNA topoisomerase IIalpha abundance after irradiation or treatment with doxorubicin. No principal difference in response was seen between normal and XP fibroblasts towards either treatment alone. After irradiation with 9 Gy, the effect was detectable after as little as 30 min and lasted for at least 6 h. After doxorubicin treatment, topoisomerase II overexpression occurred within less than 2 h. It passed through a maximum and began to decrease after approximately 6 h. In principle, the increase in DNA topoisomerase IIalpha may result from (i) architectural changes of interphase chromatin leading to enhanced accessibility of preformed enzyme to the antibody, (ii) enhanced gene expression, or (iii) enhanced stabilization of mRNA or protein molecules. The increase in enzyme levels may be part of the well-known DNA damage responses that operate in cell-protective or DNA-reparative pathways. Thus, the action of DNA topoisomerase II would serve to catalyze preparatory steps in DNA repair. We also found overexpression of the Bax protein and p16 predominantly in treated XP cells, suggesting that the DNA-damaging protocols elicited signals for apoptosis and cell-cycle arrest. From the simultaneous increase in DNA topoisomerase IIalpha and Bax, one may conclude that DNA topoisomerase IIalpha also plays role in apoptosis.
German Cancer Research Center, Division: Interaction of Carcinogens with Biological Macromolecules, Heidelberg.
H W
HW
Thielmann
O
O
Popanda
D000903:Antibiotics, Antineoplastic / Q000494:pharmacology*; D000951:Antigens, Neoplasm; D002460:Cell Line; D002467:Cell Nucleus / Q000187:drug effects / Q000201:enzymology / Q000528:radiation effects; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000096:biosynthesis; D004249:DNA Damage / Q000187:drug effects / Q000528:radiation effects; D004250:DNA Topoisomerases, Type II / Q000096:biosynthesis* / Q000528:radiation effects; D004268:DNA-Binding Proteins; D004305:Dose-Response Relationship, Drug; D004317:Doxorubicin / Q000494:pharmacology*; D005347:Fibroblasts / Q000201:enzymology / Q000528:radiation effects / Q000648:ultrastructure; D005720:Gamma Rays; D006801:Humans; D007150:Immunohistochemistry; D007527:Isoenzymes / Q000096:biosynthesis* / Q000528:radiation effects; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis; D019253:Proto-Oncogene Proteins c-bcl-2; D014983:Xeroderma Pigmentosum / Q000201:enzymology* / Q000648:ultrastructure; D051028:bcl-2-Associated X Protein
D000903:Antibiotics, Antineoplastic; D000951:Antigens, Neoplasm; C494826:BAX protein, human; D019941:Cyclin-Dependent Kinase Inhibitor p16; D004268:DNA-Binding Proteins; D007527:Isoenzymes; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; D004317:Doxorubicin; D004250:DNA Topoisomerases, Type II