Caspase-mediated cleavage of DNA topoisomerase I at unconventional sites during apoptosis

Kumiko Samejima, Phyllis A. Svingen, Guriqbal S. Basi, Timothy Kottke, Peter W. Mesner, Lance Stewart, Françoise Durrieu, Guy G. Poirier, Emad S. Alnemri, James J. Champoux, Scott H. Kaufmann, William C. Earnshaw

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89 Scopus citations

Abstract

Previous studies have demonstrated that topoisomerase I is cleaved late during apoptosis, but have not identified the proteases responsible or examined the functional consequences of this cleavage. Here, we have shown that treatment of purified topoisomerase I with caspase-3 resulted in cleavage at DDVD146 ↓ Y and EEED170 ↓ G, whereas treatment with caspase-6 resulted in cleavage at PEDD123 ↓ G and EEED170 ↓ G. After treatment of Jurkat T lymphocytic leukemia cells with anti-Fas antibody or A549 lung cancer cells with topotecan, etoposide, or paclitaxel, the topoisomerase I fragment comigrated with the product that resulted from caspase-3 cleavage at DDVD146 ↓ Y. In contrast, two discrete topoisomerase I fragments that appeared to result from cleavage at DDVD146 ↓ Y and EEED170 ↓ G were observed after treatment of MDA-MB-468 breast cancer cells with paclitaxel. Topoisomerase I cleavage did not occur in apoptotic MCF-7 cells, which lack caspase-3. Cell fractionation and band depletion studies with the topoisomerase I poison topotecan revealed that the topoisomerase I fragment remains in proximity to the chromatin and retains the ability to bind to and cleave DNA. These observations indicate that topoisomerase I is a substrate of caspase-3 and possibly caspase-6, but is cleaved at sequences that differ from those ordinarily preferred by these enzymes, thereby providing a potential explanation why topoisomerase I cleavage lags behind that of classical caspase substrates such as poly-(ADP- ribose) polymerase and lamin B1.

Original languageEnglish (US)
Pages (from-to)4335-4340
Number of pages6
JournalJournal of Biological Chemistry
Volume274
Issue number7
DOIs
StatePublished - Feb 12 1999

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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    Samejima, K., Svingen, P. A., Basi, G. S., Kottke, T., Mesner, P. W., Stewart, L., Durrieu, F., Poirier, G. G., Alnemri, E. S., Champoux, J. J., Kaufmann, S. H., & Earnshaw, W. C. (1999). Caspase-mediated cleavage of DNA topoisomerase I at unconventional sites during apoptosis. Journal of Biological Chemistry, 274(7), 4335-4340. https://doi.org/10.1074/jbc.274.7.4335