Skip to main content
NCBI home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1986 Aug;83(16):5871–5874. doi: 10.1073/pnas.83.16.5871

Role for topoisomerases in the release of DNA into the detergent-soluble fraction of eukaryotic cells.

L H Zhang, S C Mui, J C Todt, P R Strauss
PMCID: PMC386398  PMID: 3016725

Abstract

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.

Full text

PDF
5871

Images in this article

5872Image
on p.5872
5872Image
on p.5872

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Colwill R. W., Sheinin R. ts A1S9 locus in mouse L cells may encode a novobiocin binding protein that is required for DNA topoisomerase II activity. Proc Natl Acad Sci U S A. 1983 Aug;80(15):4644–4648. doi: 10.1073/pnas.80.15.4644. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Cozzarelli N. R. DNA topoisomerases. Cell. 1980 Nov;22(2 Pt 2):327–328. doi: 10.1016/0092-8674(80)90341-4. [DOI] [PubMed] [Google Scholar]
  3. Drlica K., Snyder M. Superhelical Escherichia coli DNA: relaxation by coumermycin. J Mol Biol. 1978 Apr 5;120(2):145–154. doi: 10.1016/0022-2836(78)90061-x. [DOI] [PubMed] [Google Scholar]
  4. Edenberg H. J. Novobiocin inhibition of simian virus 40 DNA replication. Nature. 1980 Jul 31;286(5772):529–531. doi: 10.1038/286529a0. [DOI] [PubMed] [Google Scholar]
  5. Gellert M. DNA topoisomerases. Annu Rev Biochem. 1981;50:879–910. doi: 10.1146/annurev.bi.50.070181.004311. [DOI] [PubMed] [Google Scholar]
  6. Holm C., Goto T., Wang J. C., Botstein D. DNA topoisomerase II is required at the time of mitosis in yeast. Cell. 1985 Jun;41(2):553–563. doi: 10.1016/s0092-8674(85)80028-3. [DOI] [PubMed] [Google Scholar]
  7. Hsiang Y. H., Hertzberg R., Hecht S., Liu L. F. Camptothecin induces protein-linked DNA breaks via mammalian DNA topoisomerase I. J Biol Chem. 1985 Nov 25;260(27):14873–14878. [PubMed] [Google Scholar]
  8. Liu L. F. DNA topoisomerases--enzymes that catalyse the breaking and rejoining of DNA. CRC Crit Rev Biochem. 1983;15(1):1–24. doi: 10.3109/10409238309102799. [DOI] [PubMed] [Google Scholar]
  9. Long B. H., Musial S. T., Brattain M. G. Comparison of cytotoxicity and DNA breakage activity of congeners of podophyllotoxin including VP16-213 and VM26: a quantitative structure-activity relationship. Biochemistry. 1984 Mar 13;23(6):1183–1188. doi: 10.1021/bi00301a024. [DOI] [PubMed] [Google Scholar]
  10. Mattern M. R., Painter R. B. Dependence of mammalian DNA replication on DNA supercoiling. II. Effects of novobiocin on DNA synthesis in Chinese hamster ovary cells. Biochim Biophys Acta. 1979 Jul 26;563(2):306–312. doi: 10.1016/0005-2787(79)90049-2. [DOI] [PubMed] [Google Scholar]
  11. Meechan P. J., Killpack S., Cleaver J. E. Novobiocin-mediated inhibition of polymerization and ligation of DNA in vitro. Mutat Res. 1984 Oct;141(2):69–73. doi: 10.1016/0165-7992(84)90013-7. [DOI] [PubMed] [Google Scholar]
  12. Meinke W., Hall M. R., Goldstein D. A., Kohne D. E., Lerner R. A. Physical properties of cytoplasmic membrane-associated DNA. J Mol Biol. 1973 Jun 25;78(1):43–56. doi: 10.1016/0022-2836(73)90427-0. [DOI] [PubMed] [Google Scholar]
  13. Nelson E. M., Tewey K. M., Liu L. F. Mechanism of antitumor drug action: poisoning of mammalian DNA topoisomerase II on DNA by 4'-(9-acridinylamino)-methanesulfon-m-anisidide. Proc Natl Acad Sci U S A. 1984 Mar;81(5):1361–1365. doi: 10.1073/pnas.81.5.1361. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Rogers J. C. Characterization of DNA excreted from phytohemagglutinin-stimulated lymphocytes. J Exp Med. 1976 May 1;143(5):1249–1264. doi: 10.1084/jem.143.5.1249. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Ross W., Rowe T., Glisson B., Yalowich J., Liu L. Role of topoisomerase II in mediating epipodophyllotoxin-induced DNA cleavage. Cancer Res. 1984 Dec;44(12 Pt 1):5857–5860. [PubMed] [Google Scholar]
  16. Schneider W. C., Kuff E. L. The association of deoxyribonucleic acid with liver microsomes. J Biol Chem. 1969 Sep 25;244(18):4843–4851. [PubMed] [Google Scholar]
  17. Strauss P. R., Andrutis A. T., Leong S. R., Nickeson S., Supple E. Characterization of rapidly labeled detergent-soluble DNA in murine splenocytes. Biochemistry. 1984 Feb 28;23(5):915–921. doi: 10.1021/bi00300a019. [DOI] [PubMed] [Google Scholar]
  18. Strauss P. R., Banerjee P. T., LaGree K., Mui S. C. Kinetics of thymidine incorporation into detergent-soluble DNA of mouse lymphocytes. Proc Natl Acad Sci U S A. 1984 Nov;81(22):7056–7060. doi: 10.1073/pnas.81.22.7056. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Strauss P. R. Incorporation of 3H-labeled nucleosides and 3H-labeled deoxynucleosides into detergent soluble DNA. Proc Soc Exp Biol Med. 1985 Sep;179(4):487–491. doi: 10.3181/00379727-179-42127. [DOI] [PubMed] [Google Scholar]
  20. Strauss P. R., Sheehan J. M., Kashket E. R. Membrane transport by murine lymphocytes. I. A rapid sampling technique as applied to the adenosine and thymidine systems. J Exp Med. 1976 Oct 1;144(4):1009–1021. doi: 10.1084/jem.144.4.1009. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Wang J. C. DNA topoisomerases. Annu Rev Biochem. 1985;54:665–697. doi: 10.1146/annurev.bi.54.070185.003313. [DOI] [PubMed] [Google Scholar]
  22. Zwelling L. A., Michaels S., Erickson L. C., Ungerleider R. S., Nichols M., Kohn K. W. Protein-associated deoxyribonucleic acid strand breaks in L1210 cells treated with the deoxyribonucleic acid intercalating agents 4'-(9-acridinylamino) methanesulfon-m-anisidide and adriamycin. Biochemistry. 1981 Nov 10;20(23):6553–6563. doi: 10.1021/bi00526a006. [DOI] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES