Nuclear distribution of Drosophila DNA topoisomerase II is sensitive to both RNase and DNase.
The nuclear distribution of Drosophila DNA topoisomerase II was determined by immunoblot analysis after nuclease digestion and cell fractionation. About 60% of DNA topoisomerase II could be removed from nuclei by RNase A, about 70% by DNase I, and about 90% by incubation with both enzymes together or with micrococcal nuclease. Nuclease treatment of nuclei did not affect the distribution of lamins Dm1 and Dm2 or other nuclear proteins similarly. Nuclease-mediated solubilization of DNA topoisomerase II from Drosophila nuclei was also dependent on NaCl concentration. Solubilization was not efficient below 100 mM NaCl. Sucrose velocity gradient ultracentrifugation demonstrated that DNA topoisomerase II solubilized from nuclei by either RNase A or DNase I migrated at about 9 S, as expected for the homodimer. Results of chemical crosslinking supported this observation. We conclude that DNA topoisomerase II has both RNA- and DNA-dependent anchorages in Drosophila embryo nuclei.
Meller
V H
VH
Department of Pharmacological Sciences, State University of New York at Stony Brook 11794-8651, USA.
Fisher
P A
PA
D000818:Animals; D002458:Cell Fractionation; D002467:Cell Nucleus / Q000201:enzymology* / Q000648:ultrastructure; D002499:Centrifugation, Density Gradient; D004250:DNA Topoisomerases, Type II / Q000187:drug effects / Q000302:isolation & purification / Q000378:metabolism*; D003720:Densitometry; D003851:Deoxyribonucleases / Q000494:pharmacology*; D004330:Drosophila / Q000196:embryology*; D004591:Electrophoresis, Polyacrylamide Gel; D004625:Embryo, Nonmammalian / Q000201:enzymology*; D015151:Immunoblotting; D012260:Ribonucleases / Q000494:pharmacology*
D003851:Deoxyribonucleases; D012260:Ribonucleases; D004250:DNA Topoisomerases, Type II