Ca super(2+), Mg super(2+)-dependent DNase Involvement in Apoptotic Effects in Spermatozoa of Sea Urchin Strongylocentrotus intermedius Induced by Two-Headed Sphingolipid Rhizochalin

• Published in: Marine biotechnology (New York, N.Y.) N.Y.), 2011-06, Vol.13 (3), p.536-543
• Main Authors: Sibirtsev, Juriy T, Shastina, Valeria V, Menzorova, Natalia I, Makarieva, Tatyana N, Rasskazov, Valeriy A
• Format: Article
• Language: English
• Subjects: Echinoidea; Marine; Strongylocentrotus intermedius
• ISSN: 1436-2228; 1436-2236
• DOI: 10.1007/s10126-010-9324-9

Previously, we have purified three distinct DNases from spermatozoa of sea urchin Strongylocentrotus intermedius and we suppose the role of Ca super(2+), Mg super(2+)-dependent DNase (Ca, Mg-DNase) in apoptosis of spermatozoa. Two-headed sphingolipid rhizochalin (Rhz) induced characteristic apoptotic nuclear chromatin changes, internucleosomal DNA cleavage, and activation of caspase-9, caspase-8, and caspase-3 in spermatozoa as was shown by fluorescence Hoechst 33342/PI/FDA analysis, DNA fragmentation assay, and fluorescence caspase inhibitors FAM-LEHD-fmk, FAM-IETD-fmk, and FAM-DEVD-fmk, respectively. Inhibitor of caspase-3 z-DEVD-fmk subdued Rhz-induced internucleosomal ladder formation, which confirmed the major role of caspase-3 in apoptotic DNA cleavage probably through Ca, Mg-DNase activation. Participation of sea urchin Ca, Mg-DNase in apoptosis of spermatozoa was demonstrated by ions Zn super(2+) blocking of Rhz-induced DNA fragmentation due to direct inhibition of the Ca, Mg-DNase and internucleosomal cleavage of HeLa S and Vero E6 cell nuclei chromatin by highly purified Ca, Mg-DNase.