title: During both interphase and mitosis, DNA topoisomerase II interacts with DNA as well as RNA through the protein's C-terminal domain.
abstract: DNA topoisomerase II (topo II) is thought to be a nuclear enzyme; during interphase most was insoluble and could be recovered in the pellet after centrifugation of cell homogenates at 10,000 g (P-10). Upon entry into mitosis, the majority of topo II did not associate with condensed chromosomes but was apparently solubilized and redistributed throughout the cell. Although two non-chromosomal subfractions of mitotic topo II were defined by centrifugation at 130,000 g, the vast majority (>90%) was recovered in the pellet (P-130). In vivo nucleic acid interactions with topo II were monitored by a recently developed approach of UV-photo-crosslinking, immunoprecipitation and (32)P-labeling. P-10 (interphase) topo II was largely associated with DNA. P-130 (mitotic non-chromosomal) topo II was primarily associated with RNA. These nucleic acid interactions with both interphase and mitotic topo II occurred through the catalytically inert and as yet, poorly understood C-terminal domain of the protein. P-10 topo II was highly active enzymatically. Activity, measured by the ability of topo II to decatenate kDNA minicircles, was reduced by treatment with phosphatase. In contrast, P-130 topo II was relatively inactive but activity could be increased by phosphatase treatment. In vivo, P-130 topo II was more heavily phosphorylated than P-10 topo II; in both, only the C-terminal domain of topo II was detectably modified. Our observations suggest that cell cycle-dependent changes in the distribution, nucleic acid interactions and enzymatic activity of topo II are regulated, at least in part, by phosphorylation/dephosphorylation.
authors.affiliation: The Department of Pharmacological Sciences, University Medical Center, The State University of New York at Stony Brook, Stony Brook, NY 11794-8651, USA.
authors.forename: R
authors.initials: R
authors.lastname: Rzepecki
authors.forename: P A
authors.initials: PA
authors.lastname: Fisher
mesh_terms: D000818:Animals; D004247:DNA / Q000378:metabolism*; D004250:DNA Topoisomerases, Type II / Q000737:chemistry / Q000378:metabolism*; D004331:Drosophila melanogaster; D007399:Interphase; D008938:Mitosis / Q000187:drug effects; D010766:Phosphorylation; D012313:RNA / Q000378:metabolism*; D014747:Vinblastine / Q000494:pharmacology
chemical_list: D014747:Vinblastine; D012313:RNA; D004247:DNA; D004250:DNA Topoisomerases, Type II