Worm Breeder's Gazette 15(4): 42 (October 1, 1998)
These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.
|1||Department of Biology, Texas Christian University, Fort Worth, TX|
|2||Department of Molecular Life Science, Tokai University School of Medicine, Isehara, Kanagawa 259-11|
|3||Department of Biophysics and Biochemistry, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113|
We have studied mev-1 and rad-8 extensively owing to our interest in free radical damage and its effects on aging.1-3 Mutations in both genes render animals hypersensitive to high concentrations of oxygen as well as cause precocious aging. We now report that oxygen also triggers massive apoptosis in rad-8 and mev-1 embryos. Evidence includes the following: 1) mev-1 and rad-8 embryos contain many cells that appear apoptotic when viewed under Nomarski DIC. The two mutations are different in terms of the kinetics of apoptosis. Specifically, rad-8 early embryos (<8 cell) will arrest- usually before the 16 cell stage--if exposed to high oxygen and not produce apoptotic nuclei. Conversely, apoptosis is readily evident if rad-8 embryos are exposed to high oxygen at the 50-300 cells stage. On the other hand, apoptosis in mev-1 embryos is most prevalent if early embryos exposed immediately to high oxygen. Apoptotic nuclei are not widespread in either mutant until 24 hours after oxygen presentation. The number of apoptotic nuclei per embryo varies and approaches 100% with extended incubation. Such embryos are indistinguishable from ced-9(gf) embryos, which serve as nice positive controls. 2) DAPI stained mev-1 and rad-8 embryos contain nuclei that have a punctate appearance. The kinetics correlate with the Nomarski observations mentioned above. In addition, these embryos appear indistinguishable from DAPI-stained ced-9(gf) embryos, which are known to undergo massive apoptosis. 3) DNA extracted from mev-1 and rad-8 embryos yields a nucleosomal ladder upon gel electrophoresis. As with the other two criteria, the ladder becomes apparent after 24-48 hours of embryo incubation in high oxygen. To elucidate the relationship between this phenotype and that controlled by the ced genes, we have constructed double mutants using lf mutations in ced-3 and ced-9 with rad-8 and mev-1. Under atmospheric oxygen ced-3 suppresses the increased number of apoptotic nuclei caused by the mev-1 mutation; specifically, the number of apoptotic nuclei per embryo were 3.8, 8.4 and 0.1 for unc-50, unc-50 mev-1 and unc-50 mev-1; ced-3 mutants, respectively. In addition, ced-3 renders mev-1 animals slightly less hypersensitive to high oxygen concentrations. These interactions are under continued investigation. 1Ishii et al., (1990) Mutat. Res. 237, 165; 2Ishii et al. (1993) Mech. Ag. Develop. 68, 1; 3Ishii et al. (1998) Nature 394,694.