Preferential delection of a specific region of mitochondrial DNA in Saccharomyces cerevisiae by ethidium bromide and 3-carbethoxy-psoralen

Abstract

Grande strains of Saccharomyces cerevisiae were mutagenized either by ethidium bromide or by 3-carbethoxy-psoralen (a monofunctional furocoumarin derivative) activated by 365nm light. 973 primary rho- clones induced were randomly collected and analyzed individually for the presence or absence of fifteen mitochondrial genetic markers. 1. Under mild conditions of mutagenesis, 83% of the primary clones showed single-deletion genotypes; a unique order of 14 markers could be deduced from the patterns of the deletion. The gene order confirmed our previous map constructed from the analysis of established non-random petite clones. From the frequencies of disjunction between markers, the distance separating 14 mitochondrial markers were estimated. 2. One region, carrying oxi-3, pho-1 and mit 175 loci, was preferentially lost in rho- mutants: there is a strong constraint in the frequencies of various genotypes found in rho- clones. On each side of this particular region, a bidirectionally oriented pattern of retention of markers is observed.

Influence of the nuclear gene tmp3 on the loss of mitochondrial genes in Saccharomyces cerevisiae.

“…a tmp3 spore segregants were picked from the mother plate (or the first transfer plate, depending on the experiment), suspended in a small amount of water, and spotted onto 11 antibiotic-sensitive strain was added. After overnight growth at 28°C, each plate was replicated onto glucose minimal medium to select diploids; plates were later replicated onto glycerol-containing plates and incubated at both permissive and restrictive temperatures for detection of mit+ alleles (3).…”

“…There is also a preferential deletion of a unique region carrying oxi3, oliII, irrespective of whether the deletions are spontaneous or mutagen induced (11,14,17,20).…”

Spontaneous and induced rho mutants of Saccharomyces cerevisiae: patterns of loss of mitochondrial genetic markers

“…Media. Complete glucose medium, complete glycerol medium, drug-containing media, and minimal medium were as described by Fukuhara et al (12).…”

“…The loss of mtDNA sequences has been studied in primary clones of rho mutants induced by different treatments: ethidium bromide (2,12,13), photoaddition of psoralen derivatives (12,14), UV irradiation (M. Heude and E. Moustacchi, Genetics, in press), mutation of nuclear genes (24), and phosphate starvation (26). The structure of the deletions can be determined genetically by using the numerous mitochondrial genetic markers now.available (9), including mutations which confer drug resistance (17,27) and mit mutations, which are specific defects in mitochondrial genes (1,3,10,15,20,23,25,28).…”

“…In the case of induced rho mutants, detailed studies of deletions, using many genetic markers (11)(12)(13)24; Heude and Moustacchi, in press), have shown that the different segments of mtDNA are not deleted at random. The unique region carrying genes oxi3 and oli2 appears to be preferentially deleted, irrespective of the nature of the mutagen.…”