Linear mitochondrial DNAs from yeasts: telomeres with large tandem repetitions

Abstract

The terminal structure of the linear mitochondrial DNA (mtDNA) from the yeast Candida parapsilosis was investigated. This mtDNA, 30 kb long, has symmetrical ends forming inverted terminal repeats. These repeats are made up of a variable number of tandemly repeating units of 738 bp each; the terminal nucleotide corresponds to a precise position within the last repeat unit sequence. The ends had an open structure accessible to enzymes, with a 5' single-stranded extension of about 110 nucleotides. No circular forms were detected in the DNA preparations. Two other unrelated species, Pichia philodendra and Candida salmanticensis also appear to have a linear mtDNA of similar organization. These linear DNAs (which we name Type 2 linear mtDNAs) are distinct from the previously described linear mtDNAs of yeasts whose termini are formed by a closed hairpin loop (Type 1 linear mtDNA). The terminal structure of C. parapsilosis mtDNA is reminiscent of the linear mitochondrial genomes of the ciliate Tetrahymena although, in the latter, the telomeric tandem repeat unit is considerably shorter.

Linear versus circular mitochondrial genomes: intraspecies variability of mitochondrial genome architecture in Candida parapsilosis

“…Restriction fragment patterns of mtDNAs were examined on small-scale preparations (Defontaine et al, 1991) using the restriction endonucleases HindIII, EcoRV, PvuII or KpnI (New England Biolabs) and separated by agarose gel electrophoresis. For physical mapping and cloning purposes mtDNA was prepared and purified by CsCl gradient centrifugation as described previously (Nosek et al, 1995). The HindIII and EcoRV restriction fragments of mtDNA were cloned into the corresponding sites of the pCR-ScriptAmp R -SK(+) plasmid vector (Stratagene) and the sequences at the boundaries of inserts were determined using universal (M13) forward and reverse primers.…”

“…While the mitochondrial telomeres in the strain belonging to group I consist of arrays of tandem repeats of a 738 bp unit (Nosek et al, 1995), the telomeric repeat in the group III strain is remarkably shorter (620 bp). However, the two sequences possess highly conserved regions and overall identity is 72 % (for details see the GenBank/EMBL database entries AY391851, AY391852, X76196 and X76197).…”

“…Previous investigations demonstrated that the mitochondrial genome in C. parapsilosis CBS 7157 (SR23) consists of a population of linear DNA molecules that terminate with a variable number of the 738 bp unit repeated in tandem (Nosek et al, 1995). Southern hybridization and subsequent DNA sequence analysis uncovered a standard set of mitochondrial genes starting with nad3 near the left telomere and terminating with atp6 close to the right telomere ( Fig.…”

“…Southern hybridization and subsequent DNA sequence analysis uncovered a standard set of mitochondrial genes starting with nad3 near the left telomere and terminating with atp6 close to the right telomere ( Fig. 4) (Kovac et al, 1984;Nosek & Fukuhara, 1994a, b;Nosek et al, 1995; J. Nosek, M. Novotna, Z. Hlavatovicova, D. W. Ussery, J. Fajkus & L. Tomaska, unpublished). Results of the restriction enzyme analysis of mtDNA by HindIII, EcoRV, PvuII and KpnI indicated that the restriction fragment patterns seem to be constant among group I strains.…”

“…In addition, several studies have reported that molecular markers derived from mitochondrial DNA (mtDNA) are applicable in molecular diagnostics of C. parapsilosis isolates (Camougrand et al, 1988;Su & Meyer, 1989Yokoyama et al, 2000;. Importantly, studies of genetic organization of mtDNA in C. parapsilosis have uncovered that this yeast, in contrast to the related species C. albicans and C. tropicalis, possesses a linear mitochondrial genome terminating with specific structures termed mitochondrial telomeres (Kovac et al, 1984;Nosek et al, 1995). Due to the uniqueness of mitochondrial telomeres in C. parapsilosis compared to related species, these structures may represent a promising diagnostic or therapeutic target (Nosek et al, 1998.…”