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The complete 27,694-bp mitochondrial (mt) DNA sequence of Hansenula wingei, which is a typical budding yeast and contains circular mitochondrial DNA, has been determined. The mt sequence contains genes encoding large and small ribosomal RNAs, 25 tRNAs, three subunits of cytochrome c oxidase (subunits 1, 2 and 3), three subunits of ATPase (subunits 6, 8 and 9), apocytochrome b, seven subunits of NADH dehydrogenase (subunits 1, 2, 3, 4, 4L, 5 and 6), and a ribosomal protein, VAR1. The VAR1 gene is considered to be a typical yeast type. This is consistent with data on DNA and the deduced amino-acid sequence homology comparisons of genes ubiquitous in yeast and fungi. However, we have identified seven genes encoding NADH dehydrogenase subunits, which are not found in other yeast mitochondrial genomes, thus placing the H. wingei mitochondrial genome in a unique position. In addition the H. wingei mitochondrial genome also encodes one tRNA pseudogene and one short unidentified ORF. The genome is compact with only two introns both of which contain an ORF. One intron lies in the large rRNA gene while the other is situated in the cytochrome c oxidase subunit-1 gene. The conserved nonanucleotide motif (A/T)TATAAG (T/A)(A/T), which is a transcription start signal in Saccharomyces cerevisiae mitochondria, has also been found in the H. wingei mitochondrial genome. The codon assignments for ATA and CTN in H. wingei mitochondria are different from those in S. cerevisiae mitochondria. These results indicate a unique and novel structure for the H. wingei mitochondrial genome in terms of characteristics which are typical for both yeast and for filamentous fungi. This is the first complete mt DNA sequence report in yeast.

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We have accomplished the nucleotide sequence of the 1537 bp mitochondrial gene coding for small subunit (SSU) rRNA of yeast Hansenula wingei, and also determined the 5'- and 3'-termini by S1 nuclease mapping. Eight universally conserved (U) elements of the SSU rRNA were identified. Comparison of U regions among five fungal mitochondrial SSU rRNA shows the striking similarity between H. wingei and Saccharomyces cerevisiae. The construction of the secondary structure revealed a core structure similar to the counterpart of Escherichia coli 16S rRNA. The secondary structure also enabled us the specify seven variable (V) regions differing from those of other mitochondrial SSU rRNAs in size, sequence and possible secondary structure. Molecular phylogenetic evaluation based on U regions of five fungi indicates that mitochondria of H. wingei and S. cerevisiae diverged from the same lineage. This suggests that the evolution of mitochondria-encoded genes does not directly correlate with the alteration of mitochondrial genetic system: genome size, gene organization and codon usage.