A mitochondrial nuclease is modified in Drosophila mutants (mus308) that are hypersensitive to DNA crosslinking agents.
The mus308 mutants of Drosophila have previously been demonstrated to be defective in an enzyme that is designated Nuclease 3 (Boyd et al. 1990b). In this study that enzyme is shown to be present in mitochondria of both wild-type flies and embryos. Since the mus308 mutants are hypersensitive to DNA crosslinking agents. Nuclease 3 is potentially required for resistance of the mitochondrial genome to such agents. In support of this hypothesis, electron microscopic studies of mus308 mutant flies that had been exposed to nitrogen mustard revealed an increased frequency of mitochondrial abnormalities. Further investigation of the defect at the enzymological level revealed that the mutants possess a new nuclease activity that is apparently a modified form of the wild-type protein. In the earlier study, enzyme extracts from mus308 mutants were found to lack an enzyme with a pI of approximately 6.2. More precisely defined assay conditions in this study revealed the appearance of a new nuclease activity with a higher pI in extracts from mutants. This observation, together with the finding that only the normal enzyme form is present in heterozygous individuals, supports the hypothesis that the mus308 locus is not the structural gene for the enzyme. Rather, the mus308 gene product is necessary for Nuclease 3 to assume the lower pI. Nuclease 3 has been partially purified and characterized from wild-type embryos. Its activity is stimulated by Mg++ and ATP. Optimum activity is found at a pH of 5.5 and a NaCl concentration of 50-100 mM. Nuclease 3 exhibits a temperature optimum of 42 degrees C and is insensitive to N-ethylmaleimide.(ABSTRACT TRUNCATED AT 250 WORDS)
Sakaguchi
K
K
Department of Genetics, University of California, Davis 95616.
Harris
P V
PV
van Kuyk
R
R
Singson
A
A
Boyd
J B
JB
D000818:Animals; D003432:Cross-Linking Reagents / Q000737:chemistry; D004272:DNA, Mitochondrial / Q000737:chemistry; D003851:Deoxyribonucleases / Q000235:genetics* / Q000378:metabolism; D004331:Drosophila melanogaster / Q000235:genetics*; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D008466:Mechlorethamine / Q000494:pharmacology; D008928:Mitochondria / Q000201:enzymology*
D003432:Cross-Linking Reagents; D004272:DNA, Mitochondrial; D008466:Mechlorethamine; D003851:Deoxyribonucleases