ABSTRACT
We have recently demonstrated that mammalian spermatozoa have the ability to degrade their DNA by a mechanism that is similar to apoptosis in somatic cells. When this mechanism is activated, the DNA is first degraded into loop-sized fragments by TOP2B (topoisomerase IIB). This degradation, termed sperm chromatin fragmentation, can be reversed by EDTA, which causes TOP2B to religate the double-stranded breaks it originally produced. Under certain conditions, a nuclease then degrades the sperm DNA further, digesting the entire sperm genome. When mouse spermatozoa which have been treated to induce TOP2B-mediated DNA breaks are injected into oocytes, the paternal DNA is specifically and completely degraded. This total digestion of paternal DNA occurs at the time of DNA synthesis initiation. In the present study, we explore the significance of an active TOP2B in the nucleus for mouse sperm function.
Subject(s)
DNA Fragmentation , Spermatozoa/metabolism , Animals , Chromatin/genetics , Chromatin/metabolism , DNA Topoisomerases, Type II/metabolism , DNA-Binding Proteins/metabolism , Genome , Male , Mice , Models, Biological , Poly-ADP-Ribose Binding ProteinsABSTRACT
Nucleolin functions in ribosome biogenesis and contains an acidic N terminus that binds nuclear localization sequences. In previous work we showed that human nucleolin associates with the N-terminal region of human topoisomerase I (Top1). We have now mapped the topoisomerase I interaction domain of nucleolin to the N-terminal 225 amino acids. We also show that the Saccharomyces cerevisiae nucleolin ortholog, Nsr1p, physically interacts with yeast topoisomerase I, yTop1p. Studies of isogenic NSR1(+) and Deltansr1 strains indicate that NSR1 is important in determining the cellular localization of yTop1p. Moreover, deletion of NSR1 reduces sensitivity to camptothecin, an antineoplastic topoisomerase I inhibitor. By contrast, Deltansr1 cells are hypersensitive to the topoisomerase II-targeting drug amsacrine. These findings indicate that nucleolin/Nsr1 is involved in the cellular localization of Top1 and that this localization may be important in determining sensitivity to drugs that target topoisomerases.
