Summary
The treatment of yeast cells with high levels of ethidium bromide causes a rapid induction of respiratory deficient mutants followed by a period of recovery to respiratory competence in 60 to 70% of the cells. Prolonged exposure then results in a final irreversible phase of petite formation. Sucrose gradient sedimentation analysis of 3H-adenine labelled mtDNA indicates that limited fragmentation (to about 16-18S) occurs during the initial phase of petite induction followed by a reassembly of the fragments during the period corresponding to the recovery of respiratory competence. The reassembly is associated with an ethidium bromide insensitive incorporation of 3H-adenine into mtDNA at a level consistent with repair synthesis. Genetic analyses, based on the transmission of five markers carried on the mtDNA of “repaired ρ+” clones, suggests that reassembly occurs with a high degree of fidelity, though in two of a total of twenty five clones differences in marker transmission frequency were observed which could possibly reflect an altered gene order. In addition, a description is given of the marked changes in the suppressive nature of the treated cells and the temporary reduction in the capacity for marker transmission seen to accompany the transitory fragmentation of the mtDNA. The final phase of petite induction is an energy dependent degradation of the mtDNA to produce a ρ0 culture.
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References
Bastos, R.N., Mahler, H.R.: A synthesis of labelled ethidium bromide. Arch. Biochem. Biophys. 160, 643–646 (1974a)
Bastos, R.N., Mahler, H.R.: Molecular mechanisms of mitochondrial genetic activity. Effects of ethidium bromide on the deoxyribonucleic acid and energetics of isolated mitochondria. J. biol. Chem. 249, 6617–6627 (1974b)
Bernardi, G., Prunell, A., Kopecka, H.: An analysis of the mitochondrial genome of yeast with restriction enzymes. In: Proceedings of workshop on molecular biology of nucleocytoplasmic relationships during cell morphogenesis in some unicellular organisms. Amsterdam: Elsevier in press 1975
Deutsch, J., Dujon, B., Netter, P., Petrochillo, E., Slonimski, P.P., Bolotin-Fukuhara, M., Coen, D.: Mitochondrial genetics. VI. The petite mutation in Saccharomyces cerevisiae: Interrelationships between the loss of the ρ+ factor and the loss of the drug resistance mitochondrial genetic markers. Genetics 76, 195–219 (1974)
Fauman, M.A., Rabinowitz, M.: DNA-DNA hybridization studies of mitochondrial DNA of ethidium-bromide-induced petite mutants of yeast. Europ. J. Biochem. 42, 67–71 (1974)
Goldring, E.S., Grossman, L.I., Krupnick, D., Cryer, D.R., Marmur, J.: The petite mutation in yeast. Loss of mitochondrial deoxyribonucleic acid during induction of petites with ethidium bromide. J. molec. Biol. 52, 323–335 (1970)
Goldring, E.S., Grossman, L.I., Marmur, J.: Petite mutation in yeast II. Isolation of mutants containing mitochondrial deoxyribonucleic acid of reduced size. J. Bact. 107, 377–381 (1971)
Grossman, L.I., Goldring, E.S., Marmur, J.: Preferential synthesis of yeast mitochondrial DNA in the absence of protein synthesis. J. molec. Biol. 46, 367–376 (1969)
Hall, R.M., Trembath, M.K., Linnane, A.W., Wheelis, L., Criddle, R.S.: Factors affecting petite induction and the recovery of respiratory competence in yeast cells exposed to ethidium bromide. Molec. gen. Genet. 144, 255–264 (1976)
Howell, N., Molloy, P.L., Linnane, A.W., Lukins, H.B.: Biogenesis of mitochondria 34. The synergistic interaction of nuclear and mitochondrial mutations to produce resistance to high levels of mikamycin in Saccharomyces cerevisiae. Molec. gen. Genet. 128, 43–54 (1974)
Linnane, A.W., Howell, N., Lukins, H.B.: Mitochondrial genetics. In: The Biogenesis of mitochondria. Transcriptional, translational and genetic aspects (Kroon, A.M. and Saccone, C. eds.), p. 193–213, New York: Academic Press 1974
Mahler, H.R.: Biogenetic autonomy of mitochondria. C.R.C. Crit. Rev. Biochem. 1, 381–460 (1973a)
Mahler, H.R.: Genetic autonomy of mitochondrial DNA. In: Molecular cytogenetics (Hamkalo, B.A. and Papaconstantinou, J., eds.), p. 181–208. New York: Plenum Press 1973b
Mahler, H.R., Bastos, R.N.: A novel reaction of mitochondrial DNA with ethidium bromide. FEBS Letters 39, 27–34 (1974a)
Mahler, H.R., Bastos, R.N.: Coupling between mitochondrial mutation and energy transduction. Proc. nat. Acad. Sci. (Wash.) 71, 2241–2245 (1974b)
Mahler, H.R., Perlman, P.S.: Mitochondrial membranes and mutagenesis by ethidium bromide. J. Supramol. Structure 1, 105–124 (1972)
Michaelis, G., Douglass, S., Tsai, M.J., Burchiel, K., Criddle, R.S.: In vitro transcription of mitochondrial deoxyribonucleic acid from yeast. Biochemistry 11, 2026–2036 (1972)
Michaelis, G., Douglass, S., Tsai, M., Criddle, R.S.: Mitochondrial DNA and suppressiveness of petite mutants in Saccharomyces cerevisiae. Biochem. Genet. 5, 487–495 (1971)
Molloy, P.L., Linnane, A.W., Lukins, H.B.: Biogenesis of mitochondria 37. Analysis of deletion of mitochondrial antibiotic resistance markers in petite mutants of Saccharomyces cerevisiae. J. Bact. 122, 7–18 (1975)
Molloy, P.L., Linnane, A.W., Lukins, H.B.: Relative retention of mitochondrial markers in petite mutants: Mitochondrially determined differences between rho(+) strains. Genet. Res. (in press 1976)
Nagley, P., Gingold, E.B., Lukins, H.B., Linnane, A.W.: Biogenesis of mitochondria XXV. Studies of the mitochondrial genomes of petite mutants of yeast using ethidium bromide as a probe. J. molec. Biol. 78, 335–350 (1973)
Nagley, P., Linnane, A.W.: Mitochondrial DNA deficient petite mutants of yeast. Biochem. biophys. Res. Commun. 39, 989–996 (1970)
Nagley, P., Linnane, A.W.: Biogenesis of mitochondria XXI. Studies on the nature of the mitochondrial genome in yeast: the degenerative effects of ethidium bromide on mitochondrial genetic information in a respiratory competent strain. J. molec. Biol. 66, 181–193 (1972)
Nagley, P., Mattick, J.S., Hall, R.M., Linnane, A.W.: A comparative study of petite induction and inhibition of mitochondrial DNA replication in yeast by ethidium bromide and berenil. Molec. gen. Genet. 141, 291–304 (1975)
Perlman, P.S., Mahler, H.R.: A premutational state induced in yeast by ethidium bromide. Biochem. biophys. Res. Commun. 44, 261–267 (1971a)
Perlman, P.S., Mahler, H.R>: Molecular consequences of ethidium bromide mutagenesis. Nature (Lond.) New Biol. 231, 12–16 (1971b)
Prunell, A., Bernardi, G.: The mitochondrial genome of wild-type yeast cells. IV. Genes and spacers. J. molec. Biol. 86, 825–842 (1974)
Schachman, H.: Ultracentrifugation in biochemistry. New York: Academic Press 1959
Slonimski, P.P., Perrodin, G., Croft, J.H.: Ethidium bromide induced mutation of yeast mitochondria: Complete transformation of cells into respiratory deficient non-chromosomal petites. Biochem. biophys. Res. Commun. 30, 232–239 (1968)
Sriprakash, K.S., Molloy, P.L., Nagley, P., Lukins, H.B., Linnane, A.W.: Biogenesis of mitochondria 41. Physical mapping of mitochondrial genetic markers in yeast. J. molec. Biol. (in press, 1976)
Wheelis, L., Trembath, M.K., Criddle, R.S.: Petite induction and recovery in the presence of high levels of ethidium bromide. Biochem. biophys. Res. Commun. 65, 838–845 (1975)
Wu, R., Padmanabhan, R., Bambara, R.: Nucleotide sequence analysis of bacteriophage DNA. Methods in Enzymology 29, 231–253 (1974)
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Criddle, R.S., Wheelis, L., Trembath, M.K. et al. Molecular and genetic events accompanying petite induction and recovery of respiratory competence induced by ethidium bromide. Molec. gen. Genet. 144, 263–272 (1976). https://doi.org/10.1007/BF00341724
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DOI: https://doi.org/10.1007/BF00341724