Role for topoisomerases in the release of DNA into the detergent-soluble fraction of eukaryotic cells.
Detergent-soluble DNA is the fraction (2-4%) of DNA that is released into the supernate upon mild detergent lysis. It is nonmitochondrial in origin. It labels efficiently with deoxy[3H]ribonucleosides and the labeling is prevented by inhibitors of polymerase alpha and ribonucleotide reductase. In previous publications we have characterized detergent-soluble DNA from splenocytes of immunologically activated mice. In this publication we show that incorporation of [3H]thymidine into detergent-soluble DNA is prevented by pretreatment with novobiocin, 4'-(9-acridinylamino)methanesulfon-m-anisidide (m-AMSA), and teniposide (VM26), three inhibitors of type II topoisomerases. Camptothecin, an inhibitor of type I topoisomerases, also reduces incorporation of [3H]thymidine but only to 50% of control levels. In addition to affecting incorporation of [3H]thymidine, preincubation with the topoisomerase II inhibitors m-AMSA and VM26 alters the amount of DNA recovered in the detergent-soluble fraction. At low concentrations of m-AMSA the amount of detergent-soluble DNA increases somewhat, whereas at higher drug concentrations a marked decrease is observed. Treatment with VM26 results in diminished amounts of DNA being released into the detergent-soluble fraction as well. However, maximal inhibition of detergent-soluble DNA release by VM26 requires the presence of camptothecin. Therefore, we suggest that topoisomerases play an important role in making a small part of lymphocyte chromatin detergent labile. Furthermore, these results are consistent with recent studies demonstrating a role for topoisomerases in yeast replication. Thus, the newly synthesized portion of detergent-soluble DNA may arise as DNA replication intermediates not yet stabilized into mature chromatin.
D000609:Aminoacridines / Q000494:pharmacology; D000677:Amsacrine; D000818:Animals; D002166:Camptothecin / Q000494:pharmacology; D004247:DNA / Q000378:metabolism*; D004264:DNA Topoisomerases, Type I / Q000378:metabolism*; D003902:Detergents; D007364:Intercalating Agents / Q000494:pharmacology; D007700:Kinetics; D051379:Mice; D009675:Novobiocin / Q000494:pharmacology; D012995:Solubility; D013154:Spleen / Q000201:enzymology; D013713:Teniposide / Q000494:pharmacology; D013936:Thymidine / Q000378:metabolism
D000609:Aminoacridines; D003902:Detergents; D007364:Intercalating Agents; D000677:Amsacrine; D009675:Novobiocin; D004247:DNA; D013713:Teniposide; D004264:DNA Topoisomerases, Type I; D013936:Thymidine; D002166:Camptothecin