The mitochondrial genome of fission yeast: inability of all introns to splice autocatalytically, and construction and characterization of an intronless genome.

In this paper we report the inability of four group I introns in the gene encoding subunit I of cytochrome c oxidase (cox1) and the group II intron in the apocytochrome b gene (cob) to splice autocatalytically. Furthermore we present the characterization of the first cox1 intron in the mutator strain anar-14 and the construction and characterization of strains with intronless mitochondrial genomes. We provide evidence that removal of introns at the DNA level (termed DNA splicing) is dependent on an active RNA maturase. Finally we demonstrate that the absence of introns does not abolish homologous mitochondrial recombination.

Two distinct mechanisms for deletion in mitochondrial DNA of Schizosaccharomyces pombe mutator strains. Slipped mispairing mediated by direct repeats and erroneous intron splicing.

Mutator strains of the fission yeast Schizosaccharomyces pombe produce mitochondrial respiratory deficient mutants at a high rate, and roughly 20% of these mutants carry deletions in the range of 50 to 1500 base-pairs. To elucidate the mechanism of deletion we have sequenced ten deletion mutants in the mosaic gene encoding apocytochrome b (cob) and three in the split gene coding for the first subunit of cytochrome c oxidase (cox1). Of 13 deletions, ten are correlated with the presence of direct repeats, which could promote deletions by slipped mispairing during DNA replication. In some of these mutants, the termini are located in possible DNA secondary structures. In three independently isolated mutants with identical deletions in the cob gene, the 5' deletion endpoint coincides with the 3' splice point of the intron, whereas the 3' endpoint of the deletion exhibits pronounced homology with the 5' splice point of the intron. This result suggests that these deletions might be initiated by erroneous RNA splicing.

The mitochondrial genome of the fission yeast Schizosaccharomyces pombe : 5. Characterization of mitochondrial deletion mutants.

The three mutator strains ana (r)-8, ana (r)-14, and diu (r)-301 were shown to produce respiratory deficient mutants at different rates. The frequency of respiratory deficient mutants in a culture could be increased by adding ethidium bromide. According to their cytochrome spectra and enzymatic activities they form three classes, namely mutants defective in cytochrome oxidase, in cytochrome b, and in both cytochromes. By restriction enzyme analysis of mitochondrial DNA from about 100 mutants, 22 deletion mutants were identified. The deletions, ranging from 50 to 1,500 base pairs were physically mapped. Deletions were localized in the genes coding for subunit 1 of cytochrome oxidase with its two introns, within the cytochrome b gene and its intron, and within the genes for subunits 2 and 3 of cytochrome oxidase. In several cases, where the physical mapping yielded ambiguous results, pairwise genetic crosses ruled out an overlap between two neighbouring deletions.Using these mitochondrial deletion mutants as tester strains, it was shown that only tetrad analysis and chemical haploidization, but not mitotic segregation analysis, allows a decision between chromosomal and mitochondrial inheritance of respiratory deficiency in Schizosaccharomyces pombe.